Effects of incentive spirometer training on dyspnea and functional status in patients with long COVID

Yao-Hsiang Chen; Chia-Huei Lin; Ju-Han Liu; Hsin-An Lin; Yu-Shan Hsieh

doi:10.1371/journal.pone.0351553

. 2026 Jun 22;21(6):e0351553. doi: 10.1371/journal.pone.0351553

Effects of incentive spirometer training on dyspnea and functional status in patients with long COVID

Yao-Hsiang Chen ^1,², Chia-Huei Lin ^3,⁴, Ju-Han Liu ^1,⁵, Hsin-An Lin ⁶, Yu-Shan Hsieh ^1,^7,^*

Editor: Davor Plavec⁸

PMCID: PMC13286201 PMID: 42330008

Abstract

Background

Since the emergence of Coronavirus Disease 2019 (COVID-19), it has become a global pandemic, profoundly affecting public health and daily life. Many recovering individuals report persistent or recurrent symptoms—fatigue, palpitations, cognitive impairment, shortness of breath, anxiety, and chest discomfort. These lingering effects impair work, daily function, and social interaction, placing a significant burden on individual quality of life and society.

Objective

This study aims to evaluate the effectiveness of using an induced Incentive Spirometer as a respiratory training tool to relieve long COVID symptoms.

Methods

This study, conducted from July 1, 2023, to May 11, 2024, at a regional teaching hospital in northern Taiwan, involved participants who had recovered from COVID-19 within the past year and had at least one long COVID respiratory symptom. Participants were assigned to one waiting control group and four experimental groups based on recovery time: within 3 months (Experimental Group 1), 3–6 months (Experimental Group 2), 6–9 months (Experimental Group 3), and 9–12 months (Experimental Group 4). The waiting control group received no interventions, while the experimental groups underwent inspiratory training using an induced Incentive Spirometer three times a week for 6 weeks (30 repetitions per session). Assessments were conducted before and after the intervention. Primary outcomes were the Dyspnoea-12 scale and Post-COVID-19 Functional Status scale. Secondary outcomes included the 6-minute walk distance and CaO₂.

Results

Ninety participants were enrolled, with five withdrawing, leaving 85 for final analysis. After 6 weeks of intervention, the waiting control group showed no significant changes in dyspnea (p = 0.463) or post-COVID-19 functional status (p = 0.343). In contrast, all experimental groups showed significant improvements. Dyspnoea-12 scale scores improved in Experimental Groups 1 (p < 0.001), 2 (p = 0.008), 3 (p = 0.011), and 4 (p = 0.001). The Post-COVID-19 Functional Status scale also showed improvements in all Experimental Groups (Group 1: p < 0.001, Group 2: p = 0.003, Group 3: p = 0.002, and Group 4: p = 0.011). Significant improvements in 6-min walk distance were observed in some experimental groups, improvements were seen in Experimental Groups 1 (p < 0.001), 2 (p = 0.027), 3 (p = 0.68), and 4 (p = 0.172). No significant changes in CaO2 were observed (pre-test, p = 0.872 and post-test, p = 0.585).

Conclusion

Respiratory training using an induced Incentive Spirometer may help alleviate dyspnea and improve post-COVID-19 functional status in individuals with Long COVID. Earlier intervention appeared to yield better outcomes, although improvements were also observed even 9–12 months after infection. However, further studies with comprehensive pulmonary assessments are needed to confirm these findings.

Clinical trial number

NCT06165835, registered on 9 December 2023.

Introduction

The Coronavirus Disease 2019 (COVID-19) has emerged as one of the most significant global public health crises of the 21st century. While acute-phase mortality has declined substantially—owing to viral evolution, medical advances, and widespread vaccination—many patients continue to experience persistent symptoms following infection. This condition, referred to as “Long COVID,” is defined as the persistence or onset of new symptoms 3 months after a SARS-CoV-2 infection, lasting for at least 2 months without an alternative diagnosis [1]. Long COVID affects approximately 80% of patients and commonly presents with fatigue, dyspnea, chest tightness, and cognitive or emotional disturbances, all of which significantly reduce the quality of life [2–7]. Currently, there is a lack of definitive treatment for Long COVID. Several systematic reviews and meta-analyses have investigated the effects of pulmonary rehabilitation in patients with long COVID, consistently suggesting improvements in post-COVID-19 symptoms [8–10]. However, the current evidence remains limited and heterogeneous. [11–14], and no standardized or definitive therapeutic approach has been established. Therefore, evaluating the effectiveness of respiratory training for Long COVID has emerged as an urgent public health priority in the post-pandemic era.

Although aerobic exercise is known to improve cardiopulmonary function, it often poses challenges for individuals with impaired lung function, potentially leading to hypoxemia or hypercapnia [15]. In contrast, pulmonary rehabilitation exercises present a lower risk and are easier to implement, making them a more viable alternative for patients experiencing dyspnea following COVID-19 recovery [16]. An incentive spirometer (IS) is a simple and safe handheld device that provides real-time visual feedback on inspiratory strength and volume, thereby promoting lung expansion and enhancing lung capacity [17–19]. Previous studies have shown that IS can improve maximum inspiratory volume and oxygenation levels and reduce anxiety in patients with COVID-19 [20–22]. However, most available evidence pertains primarily to acute or postoperative populations [23,24], with limited systematic investigation into the effects of IS on Long COVID symptoms.

Although evidence-based medicine supports the effectiveness of IS in improving pulmonary function, its role in managing Long COVID symptoms—especially concerning the optimal timing of intervention during recovery—remains unclear. Therefore, an important aim of the present study is to determine whether, although IS cannot replace pulmonary rehabilitation, it may serve as a low-cost, self-administered intervention option, and to evaluate the efficacy of IS as a respiratory training intervention for improving dyspnea and functional status in post-COVID-19 patients. Furthermore, it seeks to examine the differential outcomes based on varying recovery intervals, with the objective of developing a more specific, feasible, and evidence-based clinical rehabilitation strategy to address existing gaps in the study.

Methods

Study design

The flow diagram of participant enrollment and allocation is shown in Fig 1. Due to the natural tendency for Long COVID symptoms to improve over time, strict randomization could lead to imbalance in disease severity at the time of intervention and introduce systematic bias resulting from spontaneous recovery. Moreover, it is not feasible in clinical practice to require patients who actively seek intervention to be assigned to a control group. To ensure ethical considerations for the control group, participants in the control group were offered the intervention after the study period if they wished; however, data collected after the intervention were not included in the analysis(Fig 1).

Accordingly, the study adopted an open-label randomized controlled trial design. Eligible participants were randomly assigned to the experimental group (EG) or waitlist control group (CG) using a computer-generated random number. Ten participants were randomly assigned to the CG, while the other participants were placed in the EG, which was further stratified into four subgroups based on the time elapsed since COVID-19 recovery. Only participants in the EG received a 6-week IS intervention, whereas those in the CG continued with routine care and received no additional pulmonary rehabilitation. Participants who dropped out were excluded from the final analysis (Fig 2).

Participants in the EG received inspiratory training using an IS (Taiwan FDA Device Approval No. 003427). They were instructed to complete three training sessions per week, each consisting of 30 sustained inhalations lasting at least 3 s. Training intensity was adjusted based on individual tolerance and physical condition. To support adherence and evaluate the feasibility of the intervention, the research team conducted regular telephone follow-ups at the third weeks. All participants completed pre- and post-intervention assessments over 6 weeks. Primary outcome measures included the D-12 scale, the PCFs scale and blood biomarkers (CaO₂). The secondary outcome is 6-min walk distance (6MWD). These assessments were conducted to evaluate the effects of the IS intervention on dyspnea and physical function among post-COVID-19 individuals

Participants and group allocation criteria

Individuals who had recovered from COVID-19 within the past year but continued to experience at least one respiratory-related symptom associated with Long COVID were recruited in this study. Participants were assigned into five groups: one waiting control group and four experimental groups, defined a priori based on the study objective of comparing intervention effects across different recovery durations and following the commonly adopted stratification of Long COVID symptom persistence (within 3, 3–6, 6–9, and 9–12 months post-recovery).

Ten participants were randomly assigned to the CG, while the other participants were placed in the EG, which was further stratified into four subgroups based on the time elapsed since COVID-19 recovery. (Fig 3).

Inclusion criteria:

Individuals who had recovered from COVID-19 within the past year (confirmed by a negative rapid antigen test). The corresponding ICD-10 codes included U07.1 (COVID-19; virus identified) and U09.0 (Post COVID-19 condition; unspecified).
Presence of at least one respiratory-related symptom related to Long COVID (such as exertional dyspnea, shortness of breath, chest tightness, cough, or dyspnea).
Aged between 20 and 90 years.
Intact consciousness with normal cognitive function and behavioral stability.
Ability to communicate, verbally or non-verbally, and understand Mandarin Chinese or a Taiwanese dialect.
Willingness to participate in the study and accept random group assignments.

Exclusion criteria:

Individuals with severe disabilities or those who are permanently bedridden.
Diagnosed with dementia (such as Alzheimer’s disease and Parkinson’s disease)
Presence of acute psychiatric symptoms that impair communication.
Individuals with a high risk of litigation.
Diagnosed with chronic obstructive pulmonary disease (COPD) or any other respiratory disorders.
Diagnosed with moderate to severe heart disease.

Quality control

The initial IS training session was conducted under the supervision of a nurse, who provided instructions and guidance to ensure participants fully understood the standardized training protocol. Written informed consent was obtained from all participants before study enrollment.

Statistical analysis and sample size calculation

The required sample size was calculated using G*Power software (version 3.1.2, Germany), employing repeated measures of one-way analysis of variance (ANOVA) (within-between interaction) as the analysis method. The following parameters were used: statistical power of 0.95, effect size of 0.25, and significance level (α) of 0.05. With five groups and two measurement points, the estimated minimum sample size was 80. Considering a potential dropout rate of approximately 10%, the final target sample size was adjusted to 90 participants.

All data were organized using Excel 2019 (Microsoft, USA) and analyzed with SPSS version 18.0 (SPSS Inc., Chicago, IL). Descriptive statistics were reported as means, standard deviations, and percentages. Inferential analyses included independent and paired t-tests, ANOVA, chi-square tests, and both linear and logistic regression. Generalized Estimating Equations (GEEs) were used to evaluate group-by-time interaction effects and assess the efficacy of the intervention. A p-value of < 0.05 was considered statistically significant.

Research instruments and measurements

Based on the study objectives, the research instruments were categorized as follows:

1
Demographic and Clinical Characteristics

The collected variables included age, sex, medical history, oxygen requirement, frequency of regular physical activity, number of previous COVID-19 infections, inspiratory volume (measured using an incentive spirometer), and medication use during the infection period.

2
Chinese Version of the Post-COVID-19 Functional Status Scale (PCFS Scale).

Klok et al. developed the Post-COVID-19 Functional Status Scale to assess the functional status of participants following COVID-19 recovery. The scale includes five levels, ranging from 0 (no functional limitations) to 4 (severe limitations), with level 5 indicating death. The scale has been validated and shown to effectively reflect the influence of Long COVID symptoms on daily functioning [25–27].

3
Dyspnoea-12 scale (D-12)

The Dyspnoea-12 scale was developed by Yorke et al. It was used to assess the subjective experience of breathlessness of the participants, encompassing physical and emotional dimensions across 12 items. Higher scores indicate more severe dyspnea. The Chinese version has demonstrated strong reliability and validity [28–30].

4
Blood Biomarkers

Hemoglobin (Hb) and hematocrit (Hct) levels were measured using the HemoSmart GOLD hemoglobin analyzer (Taiwan FDA Registration No. 004460/004455). Peripheral oxygen saturation (SpO₂), measured with a fingertip pulse oximeter, was used as a surrogate for arterial oxygen saturation (SaO₂), supported by prior validation studies [31–33]. Arterial oxygen content (CaO₂) was calculated using the formula: CaO₂ = Hb × SaO₂ × 1.34/ 100 (mL/dL). This calculation served as a reference for assessing pulmonary oxygenation function [34].

5
Six-Minute Walk Test

The 6MWT was used to assess the exercise tolerance and cardiopulmonary function of the participants. The main outcome measure was the 6-min walk distance (6MWD). Heart rate and SpO₂ were recorded before and after the test to assess physiological responses [35–37].

Results

Participant demographics and baseline characteristics

Overall, 90 eligible participants were enrolled in this study at a regional teaching hospital in Taipei, Taiwan, from July 1, 2023, to May 11, 2024. Participants were randomly assigned to the experimental and control groups. The experimental group was further stratified based on recovery duration. Five participants (5.6%) were lost to follow-up owing to reinfection, fractures, or voluntary withdrawal. Among the remaining 85 participants, 3 participants completed only the questionnaire assessments but did not undergo the physiological measurements. Therefore, their physiological data were excluded from the analysis, whereas their questionnaire data were retained for relevant statistical analyses. Ultimately, 85 participants were included in the final analysis. Group allocation was as follows: Control Group (CG): n = 10; Experimental Group 1 (EG1; recovered within 3 months): n = 43; Experimental Group 2 (EG2; recovered 3–6 months): n = 10; Experimental Group 3 (EG3; recovered 6–9 months): n = 12; Experimental Group 4 (EG4; recovered 9–12 months): n = 10. Table 1 provides a summary of the group-specific data (Table 1).

Table 1. Respiratory-related long COVID symptoms by group.

	CG	EG1	EG2	EG3	EG4
Cough	4(40%)	31(72%)	5(50%)	6(50%)	3(30%)
Chest tightness	0(0%)	3(7%)	0(0%)	0(0%)	0(0%)
Short of breath	7(70%)	17(40%)	8(80%)	9(75%)	8(80%)
Excessive sputum	0(0%)	15(35%)	2(20%)	2(17%)	0(0%)

Open in a new tab

Of the 85 participants included in the final analysis, 66% were female, with a mean age of 46.3 years. The most prevalent comorbidities were hypertension and cardiovascular disease. Most participants were non-smokers, and approximately 40% reported not engaging in regular physical activity. Regarding their COVID-19 history, 54% had been infected once, while 40% had experienced two infections. Treatments during infection included oral antiviral medications and the traditional Chinese medicine formula NRICM101; a minority of participants received no treatment. Table 2 presents a summary of the detailed demographic and clinical characteristics.

Table 2. Baseline demographic and clinical information of participants.

	All	CG	EG1	EG 2	EG 3	EG 4	p value
	(n = 85)	(n = 10)	(n = 43)	(n = 10)	(n = 12)	(n = 10)	p value
Gender [N(%)]
Male	29(34%)	2(20%)	15(35%)	3(30%)	6(50%)	3(30%)	–
Female	56(66%)	8(80%)	28(65%)	7(70%)	6(50%)	7(70%)	–
Age [Mean (Median)]	46.29(45)	38.1(33)	49.4(52)	41.2(40.5)	49.1(53)	43.1(34.5)	0.28
Height [Mean (SD)]	164.38(8.25)	161.9(6.05)	163.7(9.05)	165.3(10.25)	167.6(7.1)	164.9(5.17)	0.541
Weight[Mean (SD)]	66.96(15.06)	61.6(8.13)	65.4(14.64)	73.9(17.69)	74.3(18.29)	63.9(12.32)	0.123
Duration Post-Recovery [Mean (SD)]	4.29(3.88)	6.2(4.5)	1.2(0.63)	5(0.74)	7.5(0.8)	11.1(1.13)	p < 0.001***
Hypertension [N(%)]
Yes	14(16%)	2(20%)	9(21%)	2(20%)	1(8%)	0(0%)	–
No	71(84%)	8(80%)	34(79%)	8(80%)	11(92%)	10(100%)	–
Cardiovascular Disease [N(%)]
Yes	13(15%)	0(0%)	10(23%)	1(10%)	1(8%)	1(10%)	–
No	72(85%)	10(100%)	33(77%)	9(90%)	11(92%)	9(90%)	–
Diabetes [N(%)]
Yes	4(5%)	0(0%)	2(5%)	0(0%)	0(0%)	1(10%)	–
No	81(95%)	10(100%)	41(95%)	10(100%)	12(100%)	9(90%)	–
Smoking [N(%)]
Yes	10(12%)	1(10%)	6(14%)	1(10%)	0(0%)	2(20%)	–
No	74(87%)	9(90%)	36(84%)	9(90%)	12(100%)	8(80%)	–
Quit smoking	1(1%)	0(0%)	1(2%)	0(0%)	0(0%)	0(0%)	–
Medication [N(%)]
No	28(33%)	5(50%)	18(42%)	1(10%)	2(17%)	2(20%)	–
Injectable Antivirals	3(4%)	1(10%)	1(2%)	1(10%)	0(0%)	0(0%)	–
Oral Antivirals	23(27%)	0(0%)	14(33%)	4(40%)	2(17%)	3(30%)	–
NRICM101	23(27%)	4(40%)	6(14%)	3(30%)	7(58%)	3(30%)	–
Other Folk Remedies	7(8%)	0(0%)	3(7%)	1(10%)	1(8%)	2(20%)	–
Injectable Antivirals and NRICM101	1(1%)	0(0%)	1(2%)	0(0%)	0(0%)	0(0%)	–
Exercise [N(%)]
No	36(42%)	2(20%)	19(44%)	5(50%)	5(42%)	5(50%)	–
1 ~ 3 times/week	32(38%)	7(70%)	14(33%)	3(30%)	4(33%)	4(40%)	–
4 ~ 5 times/week	9(11%)	1(10%)	6(14%)	1(10%)	1(8%)	0(0%)	–
6 ~ 7 times/week	8(9%)	0(0%)	4(9%)	1(10%)	2(17%)	1(10%)	–
Number of Infections [N(%)]	1.52(0.61)	1.6(0.52)	1.6(0.66)	1.5(0.7)	1.3(0.45)	1.4(0.52)	0.448
1times	46(54%)	4(40%)	21(49%)	6(60%)	9(75%)	6(60%)	–
2 times	34(40%)	6(60%)	18(42%)	3(30%)	3(25%)	4(40%)	–
3 times	5(6%)	0(0%)	4(9%)	1(10%)	0(0%)	0(0%)	–
Hb (g/dL) [Mean (SD)]	11.61(1.98)	11.3(1.6)	11.6(2.12)	12.2(1.88)	11.5(1.82)	11.6(2.19)	0.88
Hct (%) [Mean (SD)]	34.89(5.86)	33.9(4.78)	34.8(6.25)	36.7(5.63)	34.7(5.42)	34.9(6.58)	0.874
PCFS [Mean (SD)]	1.56(0.57)	1.2(0.42)	1.6(0.54)	1.9(0.57)	1.3(0.49)	1.7(0.68)	0.029*
D-12 [Mean (SD)]	8.72(7.55)	8.2(7.47)	8.3(7.54)	10.2(9.58)	6(5.43)	12.8(7.13)	0.285
D-12 of physiological items	5.09(3.87)	4.1(3.76)	5.1(4.02)	5.9(4.86)	3.4(2.19)	7.1(3.32)	0.196
D-12 of psychological items	3.62(4.21)	4.1(3.99)	3.2(4.05)	4.3(5.12)	2.6(3.9)	5.7(4.48)	0.403
pre HR (times/minute) [Mean (SD)]	84.46(12.82)	88.1(13.26)	83.7(12.59)	85.4(12.69)	77.9(10.26)	91.4(14.17)	0.129
preSpO₂ (%) [Mean (SD)]	98.68(0.68)	98.7(0.95)	98.7(0.63)	98.6(0.84)	98.8(0.39)	98.4(0.7)	0.63
6MWD (meter) [Mean (SD)]	346.7(79.59)	378.1(61.57)	340.1(71.9)	294.5(82.05)	374.1(53.85)	363.1(123.7)	0.088
post HR (times/minute) [Mean (SD)]	99.84(16.01)	104.1(25.31)	99.6(14.62)	97.7(15.3)	93.1(7.62)	107(17.46)	0.286
postSpO ₂ (%) [Mean (SD)]	98.35(1.34)	98.6(0.97)	98.6(0.76)	98.3(1.16)	97.5(2.81)	98.2(0.92)	0.159
CaO2 (ml/dL) [Mean (SD)]	15.3(2.56)	14.9(2.02)	15.3(2.78)	16.1(2.45)	15(2.37)	15.3(2.86)	0.872
Ball [Mean (SD)]	N/A	N/A	2(0.87)	2(0.82)	1.8(0.87)	1.9(1)	0.834

Open in a new tab

Note 2: ***=p<0.001

At baseline, most physiological indicators (Hb, Hct, heart rate, SpO₂, and CaO₂) were within normal ranges. The mean 6MWD was 346.7 meters. The PCFS scale scores showed statistically significant differences across groups (p = 0.029), whereas no significant differences were found in other baseline indicators (p > 0.05) (Table 2).

Within-group analysis

1
Comparison of Post-COVID-19 Functional Status Scale Within Each Group

Post-intervention comparisons of the PCFS scale demonstrated significant improvements across all EGs. EG1 experienced the most improvement, with scores decreasing from 1.6 to 0.58 (p < 0.001). The other experimental groups also exhibited statistically significant changes (p < 0.05). In contrast, the CG showed no significant change, with scores decreasing from 1.2 to 1.0 (p = 0.343). The trend indicates that earlier intervention is associated with better improvement in functional status, as demonstrated by more significant reductions in the PCFS scale. This emphasizes the importance of timely intervention in maximizing rehabilitation outcomes (Table 3).

Table 3. Results of within and between analysis compare of Groups.

	CG	EG1	EG2	EG3	EG4	P value
*Primary outcomes*
Pre-test PCFS scale [mean (SD)]	1.2(0.42)	1.6(0.54)	1.9(0.57)	1.3(0.49)	1.7(0.68)	0.029*
Post-test PCFS scale [mean (SD)]	1(0.82)	0.6(0.63)	1.1(0.74)	0.5(0.52)	0.9(0.74)	0.099
△	−0.2	−1	−0.8	−0.8	−0.8	–
*p value*	0.343	<0.001***	0.003**	0.002**	0.011*	–
Pre-test D-12[mean (SD)]	8.2(7.47)	8.3(7.54)	10.2(9.58)	6(5.43)	12.8(7.13)	0.285
Post-test D-12[mean (SD)]	6.7(8.67)	1.8(3.04)	4.2(7.27)	2.3(3.65)	5.5(4.93)	0.028*
△	−1.5	−6.5	−6	−3.7	−7.3	–
*p value*	0.463	<0.001***	0.008**	0.011*	0.001**	–
Pre-test D-12 of physiological items [mean (SD)]	4.1(3.76)	5.1(4.01)	5.9(4.86)	3.4(2.19)	7.1(3.32)	0.196
Post-test D-12 of physiological items [mean (SD)]	3.5(5.02)	1.3(1.78)	2.8(3.88)	1.3(1.92)	3(2.31)	0.044*
△	−0.6	−3.8	−3.1	−2.1	−4.1	–
*p value*	0.546	<0.001***	0.007**	0.014*	<0.001***	–
Pre-test D-12 psychological items [mean (SD)]	4.1(3.99)	3.2(4.05)	4.3(5.12)	2.6(3.9)	5.7(4.48)	0.403
Post-test D-12 psychological items [mean (SD)]	3.2(3.97)	0.6(1.53)	1.4(3.44)	1(1.76)	2.5(3.06)	0.017*
△	−0.9	−2.6	−2.9	−1.6	−3.2	–
*p value*	0.434	<0.001***	0.028*	0.086	0.016*	–
Pre-test CaO2[mean (SD)]	14.9(2.02)	15.3(2.78)	16.1(2.45)	15(2.37)	15.3(2.86)	0.872
Post-test CaO2[mean (SD)]	15.4(2.52)	15.5(2.75)	14.7(2.25)	16.3(1.27)	16.1(1.81)	0.585
△	0.5	0.2	−1.4	1.3	0.8	–
*p value*	0.299	0.514	0.046*	0.061	0.104	–
*Secondary outcomes*
Pre-test 6MWD [mean (SD)]	378.1(61.57)	340.1(71.9)	294.5(82.05)	374.1(53.85)	363.1(123.7)	0.088
Post-test 6MWD [mean (SD)]	359.6(113.8)	391.1(81.24)	354.2(84.9)	393.3(49.28)	375.5(110.83)	0.664
△	−18.5	51	59.7	19.2	12.4	–
*p value*	0.573	<0.001***	0.027*	0.068	0.172	–
Pre-test Hb [mean g/dL(SD)]	11.3 (1.6)	11.5(2.12)	12.2(1.88)	11.5(1.82)	11.6(2.19)	0.88
Post-test Hb [mean g/dL (SD)]	11.6(1.94)	11.7(2.06)	11.2(1.68)	12.3(1.06)	12.2(1.42)	0.571
△	0.3	0.2	1	0.8	0.6	–
*p value*	0.327	0.513	0.037*	0.082	0.111	–
Pre-test Hct [mean% (SD)]	33.9(4.78)	34.8(6.25)	36.7(5.63)	34.7(5.42)	34.9(6.58)	0.874
Post-test Hct [mean% (SD)]	34.8(5.56)	35.2(6.25)	33.7(5.05)	37(3.22)	36.8(4.21)	0.582
△	0.9	0.6	3	2.3	1.9	–
*p value*	0.42	0.463	0.036*	0.085	0.111	–
Pre-test Hb [mean g/dL(SD)]	11.3 (1.6)	11.5(2.12)	12.2(1.88)	11.5(1.82)	11.6(2.19)	0.88
Post-test Hb [mean g/dL (SD)]	11.6(1.94)	11.7(2.06)	11.2(1.68)	12.3(1.06)	12.2(1.42)	0.571
△	0.3	0.2	1	0.8	0.6	–
*p value*	0.327	0.513	0.037*	0.082	0.111	–
Pre-HR (before 6MWT) [mean (SD)]	88.1(13.26)	83.7(12.59)	85.4(12.69)	77.9(10.26)	91.4(14.17)	0.129
Post-HR (before 6MWT) mean (SD)]	83.5(13.13)	79.7(11.44)	84.8(14.25)	78.3(11.15)	91.9(17.92)	0.077
△	−4.6	−4	−0.6	0.4	0.5	–
*p value*	0.284	0.019*	0.917	0.867	0.905	–
Pre-SpO2 (before 6MWT) [mean% (SD)]	98.7(0.95)	98.7(0.63)	98.6(0.84)	98.8(0.39)	98.4(0.7)	0.63
Post-SpO2 (before 6MWT) [mean% (SD)]	97.7(2.54)	98.7(0.72)	98.9(0.32)	98.9(0.29)	98.9(0.32)	0.037*
△	−1	0	0.3	0.1	0.5	–
*p value*	0.128	1	0.343	0.339	0.096	–
Pre-HR (after 6MWT) [mean (SD)]	104.1(25.31)	99.6(14.62)	97.7(15.3)	93.1(7.62)	107(17.46)	0.286
Post-HR (after 6MWT) [mean (SD)]	99.9 ± 17.83	98.1(14.42)	105.9(22.56)	93.3 ± 13.61	107.5(18.88)	0.22
△	−4.2	−1.5	8.2	0.2	0.5	–
*p value*	0.464	0.462	0.238	0.926	0.884	–
Pre-SpO2 (after 6MWT) [mean% (SD)]	98.6(0.97)	98.6(0.76)	98.3(1.16)	97.5(2.81)	98.2(0.92)	0.159
Post-SpO2 (after 6MWT) [mean% (SD)]	98.8(0.63)	98.6(1.48)	98.4(1.08)	98.5(1.45)	98.4(1.08)	0.747
△	0.2	0	0.1	1	0.2	–
*p value*	0.168	0.78	0.868	0.197	0.509	–
Pre-test ball [mean (SD)]	N/A	2(0.87)	2(0.82)	1.8(0.87)	1.9(1)	0.834
Post-test ball [mean (SD)]	N/A	2.8(0.45)	2.7(0.68)	2.8(0.62)	2.9(0.32)	0.906
△	N/A	0.8	0.7	1	1	–
*p value*	N/A	<0.001***	0.025*	0.002**	0.008**	–

Open in a new tab

Note 1: Hb = Hemoglobin; Hct = Hematocrit; PCFS = Post-COVID-19 Functional Status total score; D-12 = Dyspnea-12 total score; 6MWD = Six-Minute Walk Distance; CaO₂ was calculated using post-exercise SpO₂; ball = Incentive spirometer ball-lift count; △ = Difference between pre- and post-test values. P value = The statistical significance of between-group differences; p value=Within-group pre–post differences

Note 2: *=p<0.05； **=p<0.01；***=p<0.001

2
Comparison of Dyspnea-12 Scores Within Each Group

The pre- and post-intervention comparisons of D-12 scores showed significant reductions across all EGs, thereby indicating improvements in perceived dyspnea symptoms. EG1 exhibited the most significant reduction, with scores decreasing from 8.3 to 1.84 (p < 0.001). The other experimental groups also experienced statistically significant improvements (p < 0.05). In contrast, the CG did not exhibit a statistically significant change in D-12 scores, with values decreasing from 8.2 to 6.7 (p = 0.463).

These findings indicate that earlier intervention is associated with more significant improvements in dyspnea symptoms. The post-test mean score of 1.84 for EG1 approached the threshold, indicating the absence of symptoms, highlighting the potential benefits of early rehabilitation in relieving symptoms.

3
Comparison of Cardiopulmonary Function

Pre- and post-intervention comparisons were conducted within each of the five groups. No statistically significant changes were observed in the CG. For example, the 6MWD decreased from 378.1 to 359.6 meters (p = 0.573), and the resting heart rate declined from 88.1 to 83.5 beats per minute (bpm) (p = 0.284), indicating no meaningful variation (p > 0.05). In contrast, EG1 demonstrated the most prominent improvements. Resting heart rate significantly decreased from 83.7 to 79.7 bpm (p = 0.019), 6MWD increased significantly from 340.1 to 391.1 meters (p < 0.001), and inspiratory ball counts improved from 2.0 to 2.8 (p < 0.001). EG2 also demonstrated significant improvements in resting heart rate and 6MWD (p < 0.05) but concurrently experienced slight reductions in hematological indices such as Hb, Hct, and CaO₂. EG3 and EG4 showed significant improvements only in inspiratory volume: EG3 increased from 1.8 to 2.8 (p = 0.002), and EG4 from 1.9 to 2.9 (p = 0.008). No other physiological or functional indicators in these groups showed statistically significant changes.

Overall, the respiratory training intervention yielded the most significant improvements in participants who had recovered from COVID-19 within 3 months. These participants demonstrated significant enhancements in physical endurance and inspiratory capacity, along with a trend toward reduced heart rate. In contrast, the improvements in the other EGs were more limited, suggesting that the timing of the intervention plays a critical role in determining its therapeutic efficacy.

Between-group comparisons

1
Between-Group Comparisons of PCFS Scores

At baseline, the PCFS scale differed significantly among all groups (p = 0.029), and the CG showed better functional status than that of EG1, EG2, and EG4 (mean score: CG = 1.2, EG1 = 1.6, EG2 = 1.9, and EG4 = 1.7). However, after 6 weeks of intervention, these differences were no longer statistically significant (p = 0.099).

2
Between-Group Comparisons of D-12

No significant differences were observed in D-12 scores among all groups at baseline (p = 0.285). However, post-intervention scores showed a statistically significant difference (p = 0.028). Further analysis revealed significant differences between the CG and EG1 (p = 0.006), CG and EG3 (p = 0.040), and between EG1 and EG4 (p = 0.036).

3
Between-Group Comparisons of Oxygenation-Related Parameters

No significant between-group differences were observed in blood parameters (Hb and Hct), physiological indices (pre-activity and post-activity heart rate), or oxygenation indicators (CaO₂ and SpO₂) at pre-intervention and post-intervention (p > 0.05), except for pre-activity SpO₂ at the post-test, which showed a significant difference among groups (p = 0.037). Further analysis revealed that the CG exhibited lower SpO₂ than that of all EGs at the post-test.

4
Between-Group Comparisons of 6MWD

No significant differences were observed in 6MWD at either pre-test (p = 0.088) or post-test (p = 0.664) in between-groups. Nonetheless, the within-group analysis revealed significant improvements in EG1 (p < 0.001) and EG2 (p = 0.027), with a trend toward improvement also observed in other EGs. Conversely, CG showed a decline in 6MWD. The initial difference between CG and EG2 at baseline (p = 0.018) was no longer present post-intervention (p = 0.890), suggesting that the intervention timing may influence physical endurance recovery (Table 3).

Generalized Estimating Equations (GEEs) Analysis of Group-by-Time Interactions

1
Interaction Between Group and Time on the PCFS Scale

GEE analysis of the group-by-time interaction effects on the PCFS scale revealed statistically significant differences in change between the EGs and the CG following the intervention. Compared with the CG, significant reductions in PCFS scores were observed in EG1 (p < 0.001, B = −3.487), EG2 (p = 0.026, B = −2.754), EG3 (p = 0.012, B = −2.892), and EG4 (p = 0.047, B = −2.387), indicating greater functional recovery in the intervention groups (Table 4).

Table 4. Interaction effects between group and time.

	B	95% Wald CI
	B	Lower	Upper
*PCFS Scale*
EG4	−0.600	−1.194	−0.006
EG3	−0.633	−1.171	−0.095
EG2	−0.600	−1.126	−0.074
EG1	−0.823	−1.255	−0.392
CG	–
*D-12*
EG4	−5.800	−10.760	−0.840
EG3	−2.167	−6.916	2.583
EG2	−4.500	−9.460	0.460
EG1	−4.965	−8.859	−1.071
CG	–
*Oxygenation Parameters*
EG4	0.337	−1.321	1.995
EG3	0.719	−0.868	2.307
EG2	−1.832	−3.490	−0.173
EG1	−0.278	−1.588	1.031
CG	–
*6MWD*
EG4	30.990	−7.620	69.600
EG3	37.718	0.752	74.685
EG2	78.260	39.650	116.870
EG1	68.498	37.989	99.007
CG	–

Open in a new tab

*=p< 0.05; ***=p<0.001

Note 1: The β coefficients represent the interaction effects (group × time) estimated from the GEE model. The model included group, time, and group × time as independent variables. The interaction terms indicate the differences in the magnitude of change before and after the intervention between each experimental group and the control group.

2
Interaction Between Group and Time on D-12

GEE analysis of the group-by-time interaction effects on D-12 scores demonstrated statistically significant differences in change between the EGs and the CG following the intervention. Compared with the CG, significant reductions in D-12 scores were observed in EG1 (p = 0.012, B = −4.965) and EG4 (p = 0.022, B = −5.800), indicating greater improvements in perceived dyspnea in these intervention groups. EG2 showed a reduction relative to the CG (B = −4.500), although this difference did not reach statistical significance (p = 0.075). No statistically significant difference in change was observed between EG3 and the CG (p = 0.371, B = −2.167) (Table 4).

3
Interaction Effects on Oxygenation Parameters

GEE analysis of the group-by-time interaction effects on oxygenation-related parameters demonstrated that only EG2 showed a statistically significant difference in change compared with the CG following the intervention (p = 0.030, B = −1.832). No significant differences in change relative to the CG were observed in EG1 (p = 0.677, B = −0.278), EG3 (p = 0.374, B = 0.719), or EG4 (p = 0.690, B = 0.337). Although EG2 exhibited a relative decline in oxygenation compared with the CG, participants in this group still demonstrated improvements in functional status and physical performance. The clinical significance of this seemingly paradoxical finding warrants further investigation (Table 4).

4
Interaction Between Group and Time on 6MWD

GEE analysis of the group-by-time interaction effects on 6MWD demonstrated that, relative to the CG, EG1 (p < 0.0001, B = 68.498), EG2 (p < 0.0001, B = 78.260), and EG3 (p = 0.046, B = 37.718) showed statistically significant greater improvements in 6MWD following the intervention. In contrast, the change observed in EG4 was not statistically different from that of the CG (p = 0.116, B = 30.990). These findings indicate that the intervention was associated with greater improvements in physical performance in most experimental groups compared with the CG (Table 4).

Item-level analysis of D-12 responses across groups

A detailed item-level analysis of the D-12 revealed no statistically significant changes in total or individual item scores in the CG (p > 0.05). In EG1, all items showed significant improvement except for Item 6 (“My breathing is uncomfortable”), which was not statistically significant. This item demonstrated significant improvement in the other EGs, suggesting that symptom responsiveness may vary depending on the timing of the intervention.

EG1 exhibited the most pronounced pre-to-post differences across most items (p < 0.0001). As the intervention was administered later in EG2 and EG3, the magnitude of improvement declined; however, a significant resurgence was observed in EG4, forming a bimodal trend. This pattern warrants further investigation into the potential roles of recovery motivation and intervention timing.

Additionally, an analysis of score composition revealed that, both before and after the intervention, the scores of physiological items (Items 1–6) were consistently higher than those of psychological items (Items 7–12) across all groups. This indicates that individuals with Long COVID experience greater distress from the physiological aspects of dyspnea than from the psychological components (supplementary table 1)

Discussion

The findings indicate that IS is an effective, safe, and low-cost respiratory training modality for improving dyspnea and functional status in individuals with Long COVID. IS significantly alleviated subjective dyspnea and enhanced overall functional capacity in post-COVID-19 participants. In terms of functional status (PCFS scale) and dyspnea severity (D-12), the EGs demonstrated significantly greater improvements after the intervention compared with the CG, reaching statistical significance; in contrast, the CG did not show statistically significant changes.

A study reports that the PCFS scale is highly correlated with other indicators, such as quality of life and mental health [25]. In the present study, all EGs demonstrated statistically significant improvements in functional status following the intervention, with notable reductions in limitations to daily activities. These findings align with those of prior research, which supports the efficacy of respiratory training in enhancing overall function and quality of life for post-COVID-19 [38–42]. Collectively, this study confirms that IS is a practical, low-risk respiratory training modality that effectively reduces subjective dyspnea and improves functional limitations among COVID-19 survivors. The therapeutic benefits were most pronounced with early implementation, underscoring the clinical significance of timely intervention.

Based on D-12 scale assessments, all EGs showed improvements in perceived dyspnea post-intervention. In addition, the generalized estimating equation (GEE) analysis examining the time × group interaction demonstrated that improvements in dyspnea severity were statistically significant in both the EG1 and EG4 groups. Moreover, during the telephone follow-up conducted in week 3, many participants reported noticeable relief from dyspnea within three weeks of initiating IS training, along with enhanced subjective comfort and adherence. These findings align with those of Altmann et al. [43], who report that earlier initiation of respiratory rehabilitation leads to superior pulmonary recovery and overall clinical outcomes. Similarly, studies by Kusumawardani et al [44] and Abo Elyazed et al. [45] support the superiority of IS over other respiratory training modalities in improving pulmonary function.

Dyspnea and functional limitations are closely linked to psychological well-being, as persistent symptoms can significantly impair quality of life and contribute to emotional distress. Studies suggest that prolonged dyspnea in post-COVID-19 patients may stem from direct viral effects on the central nervous system or from a bidirectional interaction between physiological dysfunction and psychological stressors [46–49].

Harenwall et al. [50] and Abelson et al. [51] propose that chronic dyspnea may result in overstimulation of the hypothalamic-pituitary-adrenal axis, leading to dysregulation that exacerbates respiratory symptoms. Conversely, Gudivada et al. [21] report that respiratory training could alleviate psychological disturbances such as anxiety and depression commonly observed during post-COVID-19 recovery. Furthermore, Bai et al. [52] and Malesevic et al. [53] show that gender-related differences in coping strategies significantly influence respiratory and psychological outcomes. Female patients are more likely to report pronounced symptoms and higher levels of psychological distress [54,55]. Huang et al. [56] further attribute these effects to a combination of factors, including delayed physical recovery, social isolation, and financial strain following infection.

Regarding the timing of IS-based respiratory training and its influence on the improvement of Long COVID symptoms, a bimodal trend was observed in the D-12 analysis. Although the earliest intervention group (EG1) exhibited the greatest improvement (p < 0.001), the therapeutic effect declined modestly in EG2 and EG3 before resurging significantly in EG4 (9–12 months post-recovery). Further analysis revealed that EG4 participants exhibited higher baseline levels of perceived dyspnea and psychological distress—particularly in the item “My breathing is distressing,” which averaged 1.2 points—suggesting that persistent symptoms may exacerbate anxiety, thus enhancing motivation to actively engage in respiratory training.

McGregor et al. [57] also report a similar bimodal pattern in the effectiveness of interventions for Long COVID, with the most significant improvements observed at 3 and 12 months post-intervention, while outcomes around 6 months were less pronounced. Studies also confirm the persistence of anxiety and depression among COVID-19 survivors, with a higher prevalence in female patients [58–60]. Hanania et al. [50] propose that anxiety and dyspnea may form a vicious cycle wherein activity avoidance behaviors exacerbate respiratory symptoms. Beyond its physiological benefits, respiratory training may also alleviate psychological symptoms by modulating emotional stress pathways via vagal nerve activation [61]. Moreover, psychological expectation and placebo effects have been suggested as potential contributors to symptom improvement [62].

In contrast, the improvement observed in EG3 (6–9 months post-recovery) was relatively modest. Participant data revealed that this group exhibited milder initial symptoms, and 58% had taken NRICM101 (Traditional Chinese Medicine, Qing-guan Yi-hau) during their COVID-19 infection.

Developed by the National Research Institute of Chinese Medicine in Taiwan, NRICM101 is widely used across the region. Previous studies report its antiviral properties, ability to alleviate pulmonary damage in patients with mild COVID-19 [63,64], and potential to enhance cardiopulmonary function and reduce the risk of severe illness [65,66]. These factors may explain the limited post-intervention gains observed in EG3. This study confirms that early intervention with IS yields the greatest improvement in dyspnea. Moreover, a bimodal response pattern was observed, with significant effects also present when intervention occurred 9–12 months post-recovery. Future research is warranted to clarify the psychological drivers and physiological mechanisms underlying this phenomenon, with the goal of optimizing clinical intervention strategies.

However, regarding oxygenation-related parameters, no significant changes were observed in Hb, Hct, SpO₂, or CaO₂ across groups, except for a significant decline in Hb and Hct levels in EG2. These findings suggest that the effect of IS on pulmonary oxygenation capacity may be limited. This observation is consistent with that of Hockele et al. [39], who report that despite no improvement in SpO₂, patients still exhibited significant reductions in dyspnea following inspiratory muscle training.

This outcome may indicate that nonphysiological factors contribute to dyspnea regulation. Hanania et al. [50] propose that anxiety elevates respiratory rate and promotes gas trapping, thereby exacerbating perceived dyspnea. Beyond enhancing lung expansion and ventilation, IS training may also modulate autonomic nervous system activity, potentially alleviating anxiety and stress perception. Furthermore, Lladós et al. [67] report that vagal and phrenic nerve dysfunction is common among COVID-19 survivors, which may contribute to inspiratory muscle weakness and perceived breathlessness. Thus, IS training may mitigate symptoms indirectly by enhancing respiratory muscle function.

After 6 weeks of IS training, EG1 and EG2 exhibited significant improvements in 6MWD, while EG3 and EG4 demonstrated upward trends that were not statistically significant. These outcomes may be influenced by baseline values, intervention timing, and initial symptom severity. These findings align with those of previous studies. Andrea et al. [68] show that IS enhances 6MWD in patients with COPD without prolonging hospitalization. Vallier et al. [69] also report that pulmonary rehabilitation improves 6MWD and quality of life in patients recovering from COVID-19 [70]. Aljazeeri et al. [58] further show that although pulmonary training significantly improves 6MWD, its effects on perceived dyspnea remain limited.

Additionally, in this study, the inspiratory ball count significantly increased across all EGs following the intervention, indicating a significant improvement in inspiratory capacity. Srinivasan et al. [24] report that IS training leads to significant increases in forced expiratory volume in one second and forced vital capacity. Gudivada et al. [21] also show that 50% of participants who underwent IS training achieved restored pulmonary function, outperforming those in nonintervention groups. Although direct evidence linking inspiratory ball count to perceived dyspnea is currently lacking, the present findings suggest promising clinical potential. Further research incorporating comprehensive pulmonary function assessments and long-term follow-up is warranted to clarify its physiological relevance and practical applicability.

Given that this study was conducted during the transitional period following the COVID-19 pandemic, when uncertainties regarding recurrent outbreaks and infection control remained substantial, several practical and methodological limitations should be acknowledged. To minimize unnecessary hospital exposure and maintain infection-control precautions, the study was designed primarily as a community- and home-based intervention, which inevitably influenced certain aspects of participant assessment and follow-up. First, recruitment challenges resulted in unequal group distribution, particularly among participants with prolonged Long COVID symptoms, who were less likely to seek intervention. In addition, the control group had a relatively small sample size and was not stratified according to recovery duration, which may have affected between-group comparisons. Participants who withdrew were excluded from the final analysis, and per-protocol rather than intention-to-treat analysis was applied, potentially introducing attrition bias. Furthermore, the use of NRICM101 during acute infection was not controlled for and may have influenced post-COVID recovery trajectories.

Second, several clinical and environmental variables could not be fully controlled. Variability in SARS-CoV-2 variants, intercurrent respiratory symptoms, repeated infections, antiviral treatment exposure, and residual differences in baseline cardiopulmonary status may have influenced intervention responsiveness [71]. Environmental exposures and age-related differences in exercise tolerance were also not specifically adjusted for in the analyses.

Third, the quality of intervention execution may have been influenced by the self-administered nature of IS training and reliance on self-reported adherence. Although standardized instruction and telephone follow-up were provided, variations in technique and compliance may still have introduced performance-related variability. In addition, because of the open-label design and absence of a sham intervention group, placebo and expectation-related effects on subjective symptom outcomes could not be completely excluded.

Another important limitation is the absence of diffusion capacity of the lung for carbon monoxide assessment (DLco). As the present study primarily focused on a simple home-based intervention among non-hospitalized individuals recovering in community settings, comprehensive pulmonary function testing was difficult to perform routinely during the study period. Future studies incorporating DLco and additional pulmonary function parameters may help further clarify the physiological effects of IS intervention.

Finally, most participants had mild-to-moderate Long COVID with relatively preserved functional status. Therefore, the findings may not be generalizable to individuals with severe pulmonary sequelae or advanced functional impairment.

Despite these limitations, this study provides preliminary evidence supporting IS as a safe, simple, and accessible respiratory training strategy for improving dyspnea and functional status in individuals with Long COVID. Future studies should incorporate larger multicenter cohorts, comprehensive pulmonary assessments, and longer follow-up periods to further clarify the optimal timing and mechanisms of IS intervention.

Conclusion

This study evaluated the effects of a six-week IS training program on dyspnea and functional status in individuals with respiratory-related Long COVID symptoms. The findings suggest that IS is a feasible and low-intensity intervention that may help alleviate persistent respiratory symptoms associated with Long COVID. Participants who initiated IS training within 3 months of recovery demonstrated the greatest improvements, although benefits were also observed in those who began the intervention at later recovery stages.

In conclusion, IS may represent a practical adjunctive strategy for improving dyspnea and functional limitations in individuals with Long COVID. Further large-scale studies incorporating comprehensive pulmonary function assessments are warranted to confirm these findings and clarify the optimal timing of intervention.

Supporting information

S1 Table. Item-Level Analysis of D-12 scale.

(DOCX)

pone.0351553.s001.docx^{(34.9KB, docx)}

S1 File. CONSORT checklist and Protocol documents.

(PDF)

pone.0351553.s002.pdf^{(1.8MB, pdf)}

Data Availability

All relevant data are within the manuscript and its Supporting Information files.

Funding Statement

This work was support by Tri-service General Hospital Songshan Branch (Grant number is TSGH-SS_D_113004 and TSGH-SS_E_114015). Initials of the authors who received: Yao-Hsiang Chen. Initials of the authors who received award: Conceptualization, Data collection, Formal analysis, Visualization, Investigation. There was no additional external funding received for this study.

References

1.Koc HC, Xiao J, Liu W, Li Y, Chen G. Long COVID and its Management. Int J Biol Sci. 2022;18(12):4768–80. [DOI] [PMC free article] [PubMed] [Google Scholar]
2.Cabrera Martimbianco AL, Pacheco RL, Bagattini ÂM, Riera R. Frequency, signs and symptoms, and criteria adopted for long COVID-19: A systematic review. Int J Clin Pract. 2021;75(10):e14357. doi: 10.1111/ijcp.14357 [DOI] [PMC free article] [PubMed] [Google Scholar]
3.Davis HE, Assaf GS, McCorkell L, Wei H, Low RJ, Re’em Y, et al. Characterizing long COVID in an international cohort: 7 months of symptoms and their impact. EClinicalMedicine. 2021;38:101019. doi: 10.1016/j.eclinm.2021.101019 [DOI] [PMC free article] [PubMed] [Google Scholar]
4.O’ Mahony L, Buwalda T, Blair M, Forde B, Lunjani N, Ambikan A, et al. Impact of Long COVID on health and quality of life. HRB Open Res. 2022;5:31. doi: 10.12688/hrbopenres.13516.1 [DOI] [PMC free article] [PubMed] [Google Scholar]
5.Torres-Castro R, Vasconcello-Castillo L, Alsina-Restoy X, Solis-Navarro L, Burgos F, Puppo H, et al. Respiratory function in patients post-infection by COVID-19: a systematic review and meta-analysis. Pulmonology. 2021;27(4):328–37. doi: 10.1016/j.pulmoe.2020.10.013 [DOI] [PMC free article] [PubMed] [Google Scholar]
6.van Kessel SAM, Olde Hartman TC, Lucassen PLBJ, van Jaarsveld CHM. Post-acute and long-COVID-19 symptoms in patients with mild diseases: a systematic review. Fam Pract. 2022;39(1):159–67. doi: 10.1093/fampra/cmab076 [DOI] [PMC free article] [PubMed] [Google Scholar]
7.Visco V, Vitale C, Rispoli A, Izzo C, Virtuoso N, Ferruzzi GJ, et al. Post-COVID-19 Syndrome: Involvement and Interactions between Respiratory, Cardiovascular and Nervous Systems. J Clin Med. 2022;11(3):524. doi: 10.3390/jcm11030524 [DOI] [PMC free article] [PubMed] [Google Scholar]
8.Li S, Dai B, Hou Y, Zhang L, Liu J, Hou H, et al. Effect of pulmonary rehabilitation for patients with long COVID-19: a systematic review and meta-analysis of randomized controlled trials. Ther Adv Respir Dis. 2025;19. doi: 10.1177/17534666251323482 [DOI] [PMC free article] [PubMed] [Google Scholar]
9.Martínez-Pozas O, Meléndez-Oliva E, Rolando LM, Rico JAQ, Corbellini C, Sánchez Romero EA. The pulmonary rehabilitation effect on long covid-19 syndrome: A systematic review and meta-analysis. Physiother Res Int. 2024;29(2):e2077. doi: 10.1002/pri.2077 [DOI] [PubMed] [Google Scholar]
10.Singh SJ, Baldwin MM, Daynes E, Evans RA, Greening NJ, Jenkins RG, et al. Respiratory sequelae of COVID-19: pulmonary and extrapulmonary origins, and approaches to clinical care and rehabilitation. Lancet Respir Med. 2023;11(8):709–25. doi: 10.1016/S2213-2600(23)00159-5 [DOI] [PMC free article] [PubMed] [Google Scholar]
11.Ahmed I, Mustafaoglu R, Yeldan I, Yasaci Z, Erhan B. Effect of Pulmonary Rehabilitation Approaches on Dyspnea, Exercise Capacity, Fatigue, Lung Functions, and Quality of Life in Patients With COVID-19: A Systematic Review and Meta-analysis. Arch Phys Med Rehabil. 2022;103(10):2051–62. doi: 10.1016/j.apmr.2022.06.007 [DOI] [PMC free article] [PubMed] [Google Scholar]
12.Bargahi M, Alavi-Moghaddam M, Karimi M, Azizan Z, Jafarzadeh F, Javaherian M, et al. Safety and Efficacy of Incentive Spirometer in Covid-19 Pneumonia; a Randomized Clinical Trial. Arch Acad Emerg Med. 2024;12(1):e51. doi: 10.22037/aaem.v12i1.2244 [DOI] [PMC free article] [PubMed] [Google Scholar]
13.Jimeno-Almazán A, Pallarés JG, Buendía-Romero Á, Martínez-Cava A, Franco-López F, Sánchez-Alcaraz Martínez BJ, et al. Post-COVID-19 Syndrome and the Potential Benefits of Exercise. International Journal of Environmental Research and Public Health. 2021;18(10). [DOI] [PMC free article] [PubMed] [Google Scholar]
14.Liu K, Zhang W, Yang Y, Zhang J, Li Y, Chen Y. Respiratory rehabilitation in elderly patients with COVID-19: A randomized controlled study. Complement Ther Clin Pract. 2020;39:101166. doi: 10.1016/j.ctcp.2020.101166 [DOI] [PMC free article] [PubMed] [Google Scholar]
15.Troosters T, Janssens W, Demeyer H, Rabinovich RA. Pulmonary rehabilitation and physical interventions. Eur Respir Rev. 2023;32(168):220222. doi: 10.1183/16000617.0222-2022 [DOI] [PMC free article] [PubMed] [Google Scholar]
16.Chen Y-H, Hsieh Y-S. A Narrative Review of Impact of Incentive Spirometer Respiratory Training in Long COVID. Int J Gen Med. 2024;17:5233–46. doi: 10.2147/IJGM.S492772 [DOI] [PMC free article] [PubMed] [Google Scholar]
17.Eltorai AEM, Baird GL, Eltorai AS, Pangborn J, Antoci V Jr, Cullen HA, et al. Perspectives on Incentive Spirometry Utility and Patient Protocols. Respir Care. 2018;63(5):519–31. doi: 10.4187/respcare.05872 [DOI] [PubMed] [Google Scholar]
18.Eltorai AEM, Szabo AL, Antoci V Jr, Ventetuolo CE, Elias JA, Daniels AH, et al. Clinical Effectiveness of Incentive Spirometry for the Prevention of Postoperative Pulmonary Complications. Respir Care. 2018;63(3):347–52. doi: 10.4187/respcare.05679 [DOI] [PubMed] [Google Scholar]
19.Sum S-K, Peng Y-C, Yin S-Y, Huang P-F, Wang Y-C, Chen T-P, et al. Using an incentive spirometer reduces pulmonary complications in patients with traumatic rib fractures: a randomized controlled trial. Trials. 2019;20(1):797. doi: 10.1186/s13063-019-3943-x [DOI] [PMC free article] [PubMed] [Google Scholar]
20.Curci C, Negrini F, Ferrillo M, Bergonzi R, Bonacci E, Camozzi DM, et al. Functional outcome after inpatient rehabilitation in postintensive care unit COVID-19 patients: findings and clinical implications from a real-practice retrospective study. Eur J Phys Rehabil Med. 2021;57(3):443–50. doi: 10.23736/S1973-9087.20.06660-5 [DOI] [PubMed] [Google Scholar]
21.Gudivada KK, Tikka SK, Bonthala Y, Miryala SK, Ch RP, Garre S. Association Between Post-COVID Pulmonary Dysfunction and New-Onset Anxiety and Depression and the Impact of Incentive Spirometer-Based Exercises. Prim Care Companion CNS Disord. 2023;25(5):23m03490. doi: 10.4088/PCC.23m03490 [DOI] [PubMed] [Google Scholar]
22.Toor H, Kashyap S, Yau A, Simoni M, Farr S, Savla P, et al. Efficacy of Incentive Spirometer in Increasing Maximum Inspiratory Volume in an Out-Patient Setting. Cureus. 2021;13(10):e18483. doi: 10.7759/cureus.18483 [DOI] [PMC free article] [PubMed] [Google Scholar]
23.Garcia A, Sarenas J. Blowing in the Wind: Use of Incentive Spirometry in Preventing Acute Respiratory Distress Syndrome for Patients with Moderate COVID. European Respiratory Society. 2022. [Google Scholar]
24.Srinivasan V, Kandakurti PK, Alagesan J, Suganthirababu P, Kishore J, Jenifer A, et al. Efficacy of pursed lip breathing with bhastrika pranayama vs incentive spirometry in rehabilitating post Covid 19 follow up-a randomized control study. Turkish Journal of Physiotherapy and Rehabilitation. 2021;32(3):402–7. [Google Scholar]
25.Benkalfate N, Eschapasse E, Georges T, Leblanc C, Dirou S, Melscoet L, et al. Evaluation of the Post-COVID-19 Functional Status (PCFS) Scale in a cohort of patients recovering from hypoxemic SARS-CoV-2 pneumonia. BMJ Open Respir Res. 2022;9(1):e001136. doi: 10.1136/bmjresp-2021-001136 [DOI] [PMC free article] [PubMed] [Google Scholar]
26.Klok FA, Boon GJAM, Barco S, Endres M, Geelhoed JJM, Knauss S, et al. The Post-COVID-19 Functional Status scale: a tool to measure functional status over time after COVID-19. Eur Respir J. 2020;56(1):2001494. doi: 10.1183/13993003.01494-2020 [DOI] [PMC free article] [PubMed] [Google Scholar]
27.Machado FVC, Meys R, Delbressine JM, Vaes AW, Goërtz YMJ, van Herck M, et al. Construct validity of the Post-COVID-19 Functional Status Scale in adult subjects with COVID-19. Health Qual Life Outcomes. 2021;19(1):40. doi: 10.1186/s12955-021-01691-2 [DOI] [PMC free article] [PubMed] [Google Scholar]
28.Amado Diago CA, Puente Maestu L, Abascal Bolado B, Agüero Calvo J, Hernando Hernando M, Puente Bats I, et al. Translation and validation of the multidimensional dyspnea-12 questionnaire. Arch Bronconeumol. 2018;54(2):74–8. [DOI] [PubMed] [Google Scholar]
29.Choi TCM, Chan LLY, Tsang HC, Vong YP, Cheng YK, To YL, et al. Adaptation and validation of the Chinese version of Dyspnoea-12 scale in individuals with chronic obstructive pulmonary disease. Clin Respir J. 2021;15(10):1081–7. doi: 10.1111/crj.13411 [DOI] [PMC free article] [PubMed] [Google Scholar]
30.Williams MT, Lewthwaite H, Paquet C, Johnston K, Olsson M, Belo LF, et al. Dyspnoea-12 and Multidimensional Dyspnea Profile: Systematic Review of Use and Properties. J Pain Symptom Manage. 2022;63(1):e75–87. doi: 10.1016/j.jpainsymman.2021.06.023 [DOI] [PubMed] [Google Scholar]
31.Dubowitz G, Breyer K, Lipnick M, Sall JW, Feiner J, Ikeda K, et al. Accuracy of the Lifebox pulse oximeter during hypoxia in healthy volunteers. Anaesthesia. 2013;68(12):1220–3. doi: 10.1111/anae.12382 [DOI] [PubMed] [Google Scholar]
32.Nitzan M, Romem A, Koppel R. Pulse oximetry: fundamentals and technology update. Med Devices (Auckl). 2014;7:231–9. doi: 10.2147/MDER.S47319 [DOI] [PMC free article] [PubMed] [Google Scholar]
33.Silverston P, Ferrari M, Quaresima V. Pulse oximetry and the pandemic. BMJ. 2022;378:e071474. doi: 10.1136/bmj-2022-071474 [DOI] [PubMed] [Google Scholar]
34.Rizvi A, Macedo P, Babawale L, Tighe HC, Hughes JMB, Jackson JE, et al. Hemoglobin Is a Vital Determinant of Arterial Oxygen Content in Hypoxemic Patients with Pulmonary Arteriovenous Malformations. Ann Am Thorac Soc. 2017;14(6):903–11. doi: 10.1513/AnnalsATS.201611-872OC [DOI] [PubMed] [Google Scholar]
35.Agarwala P, Salzman SH: Six-Minute Walk Test: Clinical Role, Technique, Coding, and Reimbursement. Chest. 2020, 157(3):603–11. [DOI] [PMC free article] [PubMed] [Google Scholar]
36.Bellet RN, Adams L, Morris NR. The 6-minute walk test in outpatient cardiac rehabilitation: validity, reliability and responsiveness--a systematic review. Physiotherapy. 2012;98(4):277–86. doi: 10.1016/j.physio.2011.11.003 [DOI] [PubMed] [Google Scholar]
37.Pires IM, Denysyuk HV, Villasana MV, Sá J, Marques DL, Morgado JF, et al. Development Technologies for the Monitoring of Six-Minute Walk Test: A Systematic Review. Sensors (Basel). 2022;22(2):581. doi: 10.3390/s22020581 [DOI] [PMC free article] [PubMed] [Google Scholar]
38.Del Corral T, Menor-Rodríguez N, Fernández-Vega S, Díaz-Ramos C, Aguilar-Zafra S, López-de-Uralde-Villanueva I. Longitudinal study of changes observed in quality of life, psychological state cognition and pulmonary and functional capacity after COVID-19 infection: A six- to seven-month prospective cohort. J Clin Nurs. 2024;33(1):89–102. doi: 10.1111/jocn.16352 [DOI] [PMC free article] [PubMed] [Google Scholar]
39.Hockele LF, Sachet Affonso JV, Rossi D, Eibel B. Pulmonary and Functional Rehabilitation Improves Functional Capacity, Pulmonary Function and Respiratory Muscle Strength in Post COVID-19 Patients: Pilot Clinical Trial. Int J Environ Res Public Health. 2022;19(22):14899. doi: 10.3390/ijerph192214899 [DOI] [PMC free article] [PubMed] [Google Scholar]
40.Nopp S, Moik F, Klok FA, Gattinger D, Petrovic M, Vonbank K, et al. Outpatient pulmonary rehabilitation in patients with long COVID improves exercise capacity, functional status, dyspnea, fatigue, and quality of life. Respiration. 2022;101(6):593–601. [DOI] [PMC free article] [PubMed] [Google Scholar]
41.Omar A, Ferreira AS, Hegazy FA, Alaparthi GK. Cardiorespiratory response to six-minute step test in post COVID-19 patients - a cross sectional study. Healthcare (Basel). 2023;11(10). [DOI] [PMC free article] [PubMed] [Google Scholar]
42.Szarvas Z, Fekete M, Horvath R, Shimizu M, Tsuhiya F, Choi HE, et al. Cardiopulmonary rehabilitation programme improves physical health and quality of life in post-COVID syndrome. Ann Palliat Med. 2023;12(3):548–60. doi: 10.21037/apm-22-1143 [DOI] [PubMed] [Google Scholar]
43.Altmann CH, Zvonova E, Richter L, Schüller PO. Pulmonary recovery directly after COVID-19 and in Long-COVID. Respir Physiol Neurobiol. 2023;315:104112. doi: 10.1016/j.resp.2023.104112 [DOI] [PubMed] [Google Scholar]
44.Kusumawardani RI, Tinduh D, Andriati, Poerwandari D, Marhana IA, Melaniani S. The effectiveness of incentive spirometry exercise on pulmonary function in COVID-19 survivors: a randomized controlled trial study. Bali Med J. 2023;12(1):539–44. doi: 10.15562/bmj.v12i1.3956 [DOI] [Google Scholar]
45.Abo Elyazed TI, Abd El-Hakim AAE-M, Saleh OI, Sonbol MMF, Eid HA, Moazen E, et al. Diaphragmatic strengthening exercises for patients with post COVID-19 condition after mild-to-moderate acute COVID-19 infection: a randomized controlled study. J Rehabil Med. 2024;56:jrm25491. doi: 10.2340/jrm.v56.25491 [DOI] [PMC free article] [PubMed] [Google Scholar]
46.Bellan M, Soddu D, Balbo PE, Baricich A, Zeppegno P, Avanzi GC, et al. Respiratory and Psychophysical Sequelae Among Patients With COVID-19 Four Months After Hospital Discharge. JAMA Netw Open. 2021;4(1):e2036142. doi: 10.1001/jamanetworkopen.2020.36142 [DOI] [PMC free article] [PubMed] [Google Scholar]
47.Gutiérrez-Canales LG, Muñoz-Corona C, Barrera-Chávez I, Viloria-Álvarez C, Macías AE, Martínez-Navarro LJ, et al. Persistence of COVID-19 Symptoms and Quality of Life at Three and Twelve Months after Hospital Discharge. Medicina (Kaunas). 2024;60(6):944. doi: 10.3390/medicina60060944 [DOI] [PMC free article] [PubMed] [Google Scholar]
48.Lippi G, Sanchis-Gomar F, Henry BM. COVID-19 and its long-term sequelae: what do we know in 2023? Pol Arch Intern Med. 2023;133(4):16402. doi: 10.20452/pamw.16402 [DOI] [PubMed] [Google Scholar]
49.Nasrullah A, Virk S, Javed A, Shah A, Ramanujam D, Sharma A, et al. Effects of pulmonary rehabilitation on functional and psychological parameters in post-acute sequelae of SARS-CoV-2 infection (PASC) patients. BMC Pulm Med. 2024;24(1):231. doi: 10.1186/s12890-024-03047-0 [DOI] [PMC free article] [PubMed] [Google Scholar]
50.Hanania NA, O’Donnell DE. Activity-related dyspnea in chronic obstructive pulmonary disease: physical and psychological consequences, unmet needs, and future directions. Int J Chron Obstruct Pulmon Dis. 2019;14:1127–38. doi: 10.2147/COPD.S188141 [DOI] [PMC free article] [PubMed] [Google Scholar]
51.Abelson JL, Khan S, Giardino N. HPA axis, respiration and the airways in stress--a review in search of intersections. Biol Psychol. 2010;84(1):57–65. doi: 10.1016/j.biopsycho.2010.01.021 [DOI] [PubMed] [Google Scholar]
52.Bai F, Tomasoni D, Falcinella C, Barbanotti D, Castoldi R, Mulè G, et al. Female gender is associated with long COVID syndrome: a prospective cohort study. Clin Microbiol Infect. 2022;28(4):611.e9–611.e16. doi: 10.1016/j.cmi.2021.11.002 [DOI] [PMC free article] [PubMed] [Google Scholar]
53.Malesevic S, Sievi NA, Baumgartner P, Roser K, Sommer G, Schmidt D, et al. Impaired health-related quality of life in long-COVID syndrome after mild to moderate COVID-19. Sci Rep. 2023;13(1):7717. doi: 10.1038/s41598-023-34678-8 [DOI] [PMC free article] [PubMed] [Google Scholar]
54.Xiong Q, Xu M, Li J, Liu Y, Zhang J, Xu Y, et al. Clinical sequelae of COVID-19 survivors in Wuhan, China: a single-centre longitudinal study. Clin Microbiol Infect. 2021;27(1):89–95. doi: 10.1016/j.cmi.2020.09.023 [DOI] [PMC free article] [PubMed] [Google Scholar]
55.Zhao Y, Shi L, Jiang Z, Zeng N, Mei H, Lu Y, et al. The phenotype and prediction of long-term physical, mental and cognitive COVID-19 sequelae 20 months after recovery, a community-based cohort study in China. Mol Psychiatry. 2023;28(4):1793–801. doi: 10.1038/s41380-023-01951-1 [DOI] [PMC free article] [PubMed] [Google Scholar]
56.Huang L, Li X, Gu X, Zhang H, Ren L, Guo L, et al. Health outcomes in people 2 years after surviving hospitalisation with COVID-19: a longitudinal cohort study. Lancet Respir Med. 2022;10(9):863–76. doi: 10.1016/S2213-2600(22)00126-6 [DOI] [PMC free article] [PubMed] [Google Scholar]
57.McGregor G, Sandhu H, Bruce J, Sheehan B, McWilliams D, Yeung J, et al. Clinical effectiveness of an online supervised group physical and mental health rehabilitation programme for adults with post-covid-19 condition (REGAIN study): multicentre randomised controlled trial. BMJ. 2024;384:e076506. doi: 10.1136/bmj-2023-076506 [DOI] [PMC free article] [PubMed] [Google Scholar]
58.Aljazeeri J, Almusally R, Wert Y, Abdelhalim M, Klinger C, Ramesh N, et al. Pulmonary Rehabilitation for Post-COVID-19. J Cardiopulm Rehabil Prev. 2023;43(6):438–43. doi: 10.1097/HCR.0000000000000813 [DOI] [PubMed] [Google Scholar]
59.Natarajan A, Shetty A, Delanerolle G, Zeng Y, Zhang Y, Raymont V, et al. A systematic review and meta-analysis of long COVID symptoms. Syst Rev. 2023;12(1):88. doi: 10.1186/s13643-023-02250-0 [DOI] [PMC free article] [PubMed] [Google Scholar]
60.Zeng N, Zhao Y-M, Yan W, Li C, Lu Q-D, Liu L, et al. A systematic review and meta-analysis of long term physical and mental sequelae of COVID-19 pandemic: call for research priority and action. Mol Psychiatry. 2023;28(1):423–33. doi: 10.1038/s41380-022-01614-7 [DOI] [PMC free article] [PubMed] [Google Scholar]
61.Fincham GW, Strauss C, Montero-Marin J, Cavanagh K. Effect of breathwork on stress and mental health: A meta-analysis of randomised-controlled trials. Sci Rep. 2023;13(1):432. doi: 10.1038/s41598-022-27247-y [DOI] [PMC free article] [PubMed] [Google Scholar]
62.Benedetti F, Mayberg HS, Wager TD, Stohler CS, Zubieta J-K. Neurobiological mechanisms of the placebo effect. J Neurosci. 2005;25(45):10390–402. doi: 10.1523/JNEUROSCI.3458-05.2005 [DOI] [PMC free article] [PubMed] [Google Scholar]
63.Tsai K-C, Huang Y-C, Liaw C-C, Tsai C-I, Chiou C-T, Lin C-J, et al. A traditional Chinese medicine formula NRICM101 to target COVID-19 through multiple pathways: A bedside-to-bench study. Biomed Pharmacother. 2021;133:111037. doi: 10.1016/j.biopha.2020.111037 [DOI] [PMC free article] [PubMed] [Google Scholar]
64.Lin C-H, Chen Y-J, Lin M-W, Chang H-J, Yang X-R, Lin C-S. ACE2 and a Traditional Chinese Medicine Formula NRICM101 Could Alleviate the Inflammation and Pathogenic Process of Acute Lung Injury. Medicina (Kaunas). 2023;59(9):1554. doi: 10.3390/medicina59091554 [DOI] [PMC free article] [PubMed] [Google Scholar]
65.Huynh DTN, Nguyen HT, Hsieh C-M. Taiwan Chingguan Yihau may improve post-COVID-19 respiratory complications through PI3K/AKT, HIF-1, and TNF signaling pathways revealed by network pharmacology analysis. Mol Divers. 2025;29(3):2305–21. doi: 10.1007/s11030-024-10993-8 [DOI] [PubMed] [Google Scholar]
66.Wei W-C, Tsai K-C, Liaw C-C, Chiou C-T, Tseng Y-H, Liao G-Y, et al. NRICM101 ameliorates SARS-CoV-2-S1-induced pulmonary injury in K18-hACE2 mice model. Front Pharmacol. 2023;14:1125414. doi: 10.3389/fphar.2023.1125414 [DOI] [PMC free article] [PubMed] [Google Scholar]
67.Lladós G, Massanella M, Coll-Fernández R, Rodríguez R, Hernández E, Lucente G, et al. Vagus nerve dysfunction in the post-COVID-19 condition: a pilot cross-sectional study. Clin Microbiol Infect. 2024;30(4):515–21. doi: 10.1016/j.cmi.2023.11.007 [DOI] [PubMed] [Google Scholar]
68.Andrea Ban YL, Siti Istiana AS, Nik Nuratiqah NA, Ng BH, Rose Azzlinda O, Hasni J, et al. The benefits of early pulmonary rehabilitation with incentive spirometer among chronic obstructive pulmonary disease patients with exacerbation of chronic obstructive pulmonary disease. Med J Malaysia. 2024;79(5):561–8. [PubMed] [Google Scholar]
69.Vallier J-M, Simon C, Bronstein A, Dumont M, Jobic A, Paleiron N, et al. Randomized controlled trial of home-based vs. hospital-based pulmonary rehabilitation in post COVID-19 patients. Eur J Phys Rehabil Med. 2023;59(1):103–10. doi: 10.23736/S1973-9087.22.07702-4 [DOI] [PMC free article] [PubMed] [Google Scholar]
70.Eksombatchai D, Wongsinin T, Phongnarudech T, Thammavaranucupt K, Amornputtisathaporn N, Sungkanuparph S. Pulmonary function and six-minute-walk test in patients after recovery from COVID-19: A prospective cohort study. PLoS One. 2021;16(9):e0257040. doi: 10.1371/journal.pone.0257040 [DOI] [PMC free article] [PubMed] [Google Scholar]
71.de Bruijn S, van Hoek AJ, Mutubuki EN, Knoop H, Slootweg J, Tulen AD, et al. Lower prevalence of post-Covid-19 Condition following Omicron SARS-CoV-2 infection. Heliyon. 2024;10(7):e28941. doi: 10.1016/j.heliyon.2024.e28941 [DOI] [PMC free article] [PubMed] [Google Scholar]

PLoS One. doi: 10.1371/journal.pone.0351553.r001

Decision Letter 0

Toufic Ahmad Chaaban

20 Nov 2025

PONE-D-25-41209Study on Post-Acute COVID-19 Syndrome in Improvement of COVID-19 Rehabilitated Patients by Respiratory TrainingPLOS ONE

Dear Dr. Hsieh,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

Please submit your revised manuscript by Jan 02 2026 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.
A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.
An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

We look forward to receiving your revised manuscript.

Kind regards,

Toufic Ahmad Chaaban, M.D.

Academic Editor

PLOS ONE

Journal Requirements:

When submitting your revision, we need you to address these additional requirements.

1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at

https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and

https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf

2. Thank you for stating in your Funding Statement:

“Initials of the authors who received : Yao-Hsiang Chen

Enter a statement with the following details:

Initials of the authors who received award: Conceptualization, Data collection, Formal analysis, Visualization, Investigation

This work was support by Tri-service General Hospital Songshan Branch (Grant number is TSGH-SS_D_113004).”

Please provide an amended statement that declares *all* the funding or sources of support (whether external or internal to your organization) received during this study, as detailed online in our guide for authors at http://journals.plos.org/plosone/s/submit-now. Please also include the statement “There was no additional external funding received for this study.” in your updated Funding Statement.

Please include your amended Funding Statement within your cover letter. We will change the online submission form on your behalf.

3. Thank you for stating the following financial disclosure:

“Initials of the authors who received : Yao-Hsiang Chen

Enter a statement with the following details:

Initials of the authors who received award: Conceptualization, Data collection, Formal analysis, Visualization, Investigation

This work was support by Tri-service General Hospital Songshan Branch (Grant number is TSGH-SS_D_113004).”

Please state what role the funders took in the study. If the funders had no role, please state: "The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript."

If this statement is not correct you must amend it as needed.

Please include this amended Role of Funder statement in your cover letter; we will change the online submission form on your behalf.

4. Your ethics statement should only appear in the Methods section of your manuscript. If your ethics statement is written in any section besides the Methods, please move it to the Methods section and delete it from any other section. Please ensure that your ethics statement is included in your manuscript, as the ethics statement entered into the online submission form will not be published alongside your manuscript.

5. We are unable to open your Supporting Information file table 1-8.rar. Please kindly revise as necessary and re-upload.

6. Please include captions for your Supporting Information files at the end of your manuscript, and update any in-text citations to match accordingly. Please see our Supporting Information guidelines for more information: http://journals.plos.org/plosone/s/supporting-information.

7. If the reviewer comments include a recommendation to cite specific previously published works, please review and evaluate these publications to determine whether they are relevant and should be cited. There is no requirement to cite these works unless the editor has indicated otherwise.

Additional Editor Comments:

This manuscript require significant edits and revisions before deciding on publication. Please address reviewers comments and consider sharing the protocol prior to the study and if it was published

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Partly

**********

2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: No

Reviewer #2: Yes

Reviewer #3: Yes

**********

3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: No

Reviewer #2: Yes

Reviewer #3: Yes

**********

4. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

**********

5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: This reports the results of an RCT that compared the efficacy of IS for improving dyspnea and functional status in post COVID 19 patients. Unfortunately, this paper is impossible to read and understand.

-A ton of outcomes are presented - with no distinction about which are the primary outcomes and which are secondary.

-It is unclear how the randomization was carried out.

-The experimental group is stratified by time since infection, but it is unclear whether this was defined a priori or not.

-The analysis compares within changes from baseline to post intervention in each group. What is important is if these changes are different in the experimental arms vs. the control arm. See suggestions below.

1. How exactly was the randomization carried out? Please see the consort checklist and ensure that all the details are given, in one section. Right now these are scattered throughout the manuscript.

2. Why was such a high power used for the sample size calculation? What outcome was the sample size based on? What detectable difference was used? What SD was used? Please justify each choice.

3. Please be specific about the primary outcome. Was this 6MT walk at follow up or was it the change from baseline? OR was it dyspnea?

4. Were the groups defined a priori?

5. Please remove the p value from Table 2. Also standardize the number of decimal places. For the continuous variables present mean (SD). Also - please fix the format - it is impossible to read as is. What does a mean 1.8 for gender mean??

6. The primary analysis should compare the change in 6 minute walk between the intervention groups vs the control group overall. And then, the same thing for dyspnea. Then, use a regression model that includes arm and the group, and the interaction between arm and group - the crucial thing is if the effect of the intervention differs by group.

7. How exactly were the GEE fit? What correlation structure was used? Were robust standard errors used? Table 4 presents group by time interaction - this is a little better - now we can see that some of the groups have different changes than the control group.

8. Table 8 is extraneous, and not part of the objectives. Remove all these results.

9. the discussion says that "Among these, EG1, who underwent the intervention within 3 months of recovery, showed the greatest improvement, with a 79% reduction in D-12 scores." Is this statistically different than the changes observed in the other EGs?

Reviewer #2: This is an excellent article focusing on a highly relevant and common issue currently encountered in pulmonary and medicine clinics.

The study effectively evaluates the use of incentive spirometry as a component of respiratory training to relieve symptoms associated with long COVID. While there are currently no established treatment alternatives proven to resolve long COVID, respiratory training—including techniques like incentive spirometry—has long been utilized in the management of other chronic lung diseases. This approach aims to improve lung function, reduce dyspnea, and enhance overall functional status, suggesting that such interventions may play a valuable role in the multidisciplinary management of post-COVID patients

The study features an innovative approach by employing multiple experimental groups alongside a control group. These groups are evaluated longitudinally, according to the duration elapsed since recovery from COVID-19. This design allows for assessment of intervention effectiveness over time, providing valuable insights into how respiratory training may impact functional recovery and symptom resolution in the post-COVID population.

A key strength of the study lies in the intervention's simplicity, which is easy to implement and associated with limited to no immediate perceived harm to the patient. Moreover, the fact that incentive spirometry can be self-performed by patients adds to its practicality and feasibility, making it an attractive option for managing long COVID symptoms in the outpatient setting. The sample size for this study was appropriately calculated using statistical methods, including repeated measures one-way ANOVA. The investigators also accounted for a potential 10% dropout rate by enrolling an adequate number of additional participants. This approach enhances the robustness of the findings and ensures that sufficient data would be available for meaningful analysis despite potential attrition.

The diagnosis of long COVID is a complex and nuanced topic that warrants discussion on its own. Unlike many other conditions, long COVID remains primarily a diagnosis of exclusion, as there is currently no definitive imaging or blood-based test to confirm its presence. Clinicians must therefore rely on clinical evaluation, history, and the persistence of characteristic symptoms beyond the acute phase of infection, while actively ruling out alternative explanations. This diagnostic uncertainty presents a significant challenge in both research and clinical practice.

A significant limitation of the study is its lack of consideration for the inherent severity or risk posed by pre-existing cardiopulmonary disease. For example, the study does not account for whether participants’ initial COVID-19 illness was complicated by respiratory failure, myocarditis, hypoxemic respiratory failure, or pulmonary embolism. Instead, it includes individuals who have recovered from COVID-19 within the past year, without stratifying them according to the severity or type of their prior cardiopulmonary involvement. A mean 6-minute walk distance of 347 meters indicates that the study group consisted of a relatively healthy population with moderate baseline functional capacity. This may limit the applicability of the findings to individuals with more severe impairment or lower exercise tolerance. Subjective performance in interventions such as respiratory training may vary significantly with age, as younger patients often tolerate these exercises better than older individuals. There is no age-based stratified analysis to address these potential differences. Furthermore, the study does not fully account for underlying health risk factors such as pre-existing cardiopulmonary disease and environmental exposures, including urban air pollution. Although hypertension and cardiovascular disease were the most reported underlying medical conditions among participants, it remains uncertain whether individuals with chronic lung diseases were excluded. These factors could substantially influence both baseline functional status and response to intervention, and their omission may limit the generalizability of the study’s findings. The study comments on baseline exercise tolerance, noting that 40% of participants did not engage in regular physical activity. However, it is unclear whether this lack of activity reflects participants’ habits prior to their COVID-19 episode or is a consequence of their illness and recovery. This ambiguity makes it difficult to interpret the impact of the intervention on overall exercise capacity and recovery trajectories. Although participants of the study have at least 1 or 2 episodes of covid infection, the study does not address whether outcomes differed between individuals with repeated COVID-19 infections compared to those who experienced a single episode. This information is not provided, leaving uncertainty about whether recurrent infection may impact functional recovery or response to respiratory training in long COVID patients. Injectable antivirals were only used in 3 out of 85 participants (4% of study population). so, I would expect the participants to have a lower acuity of covid infection at base line. What is the difference among those who were treated with the medications vs no medication group (28 of 85)

Despite robust sample calculation several subgroups had a limited number of patients; detailed interpretation in some of the subgroup analyses was extremely limited.

The effect of the Chinese medicine formula on the results is not clearly addressed in the study. It is unclear whether a separate analysis was conducted for patients who did not receive the Chinese medicine formula, which raises concerns about the generalizability of the findings—particularly to populations that do not use traditional Chinese medicine. Additionally, the possible interaction between the Chinese medicine formula and the intervention being studied (such as respiratory training) remains uncertain. Without subgroup analysis, it is difficult to determine whether the observed improvements in recovery are attributable to the respiratory intervention itself, the Chinese medicine formula, or a combination of both. This limitation should be considered when interpreting the applicability of the study to broader patient populations

Quality control in the study was addressed by having a nurse perform the initial training for participants. However, subsequent performance of the intervention relied on patient self-reporting, which introduces the potential for variation in both execution and documentation of the exercise. While telephone follow-up calls were used to assess compliance, a video call could have provided a more objective assessment, allowing direct observation of patient technique without the confounding factor of patients talking during incentive spirometry sessions. Employing video-based monitoring might enhance the accuracy and consistency of performance evaluation in future studies.

The study mentions regular telephone follow-ups; however, it does not specify the exact frequency of these calls. Further clarification regarding the timing and intervals of follow-up contacts would be useful to assess the consistency of monitoring and its potential impact on patient compliance and outcomes.

There appears to be some lack of clarity regarding the final number of participants included in the analysis. The study notes that five patients were lost to follow-up, and an additional three participants, who only completed the questionnaire assessments, were excluded from the final analysis. Based on these exclusions, the total analysis group was stated as 85; however, by calculation, one might expect the final number to be 82. Clarifying the exact criteria for inclusion and ensuring consistency in reporting participant numbers is important for accurate interpretation of study results.

Smoking prevalence in Taiwan, where this study was conducted, is low at around 13% overall. This aligns with the study population, in which female smoking prevalence is reported at only 2% and male smoking at 23%. These rates reflect national averages and provide important context for interpreting study outcomes, especially when considering risk factors commonly associated with cardiopulmonary health.

In table 2

When we look at the average duration of post recovery. For experimental Group 2 it is mentioned that the average duration of post recovery is only 1.2 months, I am not sure. If this is represented adequately as experimental, group 2 should be Three to six months. It is unclear why there are numbers in the row beside Hypertension in the mean and SD, I do not think we need this is necessary for categorical variables

Based on the PCFS score, there are significant differences that are noted in the Experimental Groups. It indicates that the study population is a healthier population, and the underlying COVID infection was less severe. Without any long-term Needs for supplemental oxygen. So, I would expect that the patients would not have any diseases like COVID-induced ILD.

There is a paragraph repetition between lines 110 and 120 and this could be corrected.

Reviewer #3: I appreciate the authors' commitment to rigorously evaluating a low-cost strategy for managing Long COVID syndrome, especially given the crucial need for accessible interventions in low and middle-income countries.

However, I have a significant methodological concern regarding the composition of the standard care arm and its relationship to the intervention being tested.

My concern is two-fold:

1. Exclusion of Pulmonary Rehabilitation:

Pulmonary rehabilitation is widely recognized and increasingly accepted as a cornerstone of management for persistent respiratory and functional impairment in Long COVID. Excluding it as a component of standard care for both the control and intervention groups may create a potential ethical and scientific deficit by not providing the currently accepted optimal care to the participants.

2. Role of Incentive Spirometry:

I believe that both groups should have received a structured pulmonary rehabilitation program. The authors' chosen low-cost intervention, such as incentive spirometry, should have been evaluated as an additive or specific, targeted component in addition to comprehensive pulmonary rehabilitation, rather than a potential replacement for it.

I would appreciate a strong justification from the authors for this trial design choice, particularly on why they elected to avoid including pulmonary rehabilitation in the standard care protocol, which is critical for fully assessing the incremental benefit of their low-cost strategy.

**********

6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: Yes: Madhu Kalyan Pendurthi

Reviewer #3: Yes: Dapanaduwage Amila Vindana Rathnapala

**********

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

To ensure your figures meet our technical requirements, please review our figure guidelines: https://journals.plos.org/plosone/s/figures

You may also use PLOS’s free figure tool, NAAS, to help you prepare publication quality figures: https://journals.plos.org/plosone/s/figures#loc-tools-for-figure-preparation.

NAAS will assess whether your figures meet our technical requirements by comparing each figure against our figure specifications.

Attachment

Submitted filename: PLOS Reviews 30.10.2025.docx

pone.0351553.s003.docx^{(13.6KB, docx)}

PLoS One. 2026 Jun 22;21(6):e0351553. doi: 10.1371/journal.pone.0351553.r002

Author response to Decision Letter 1

26 Dec 2025

We sincerely thank the reviewers for their extensive and valuable comments and suggestions, which have greatly contributed to improving the quality and clarity of our manuscript. In response to these constructive critiques, we have carefully revised the manuscript and provided detailed, point-by-point responses in the accompanying Response to Reviewers document.

-A ton of outcomes are presented - with no distinction about which are the primary outcomes and which are secondary.

-It is unclear how the randomization was carried out.

-The experimental group is stratified by time since infection, but it is unclear whether this was defined a priori or not.

1. How exactly was the randomization carried out? Please see the consort checklist and ensure that all the details are given, in one section. Right now these are scattered throughout the manuscript.

Reply: Thank you for the reviewer’s suggestion. Because Long COVID symptoms tend to improve naturally over time, strict randomization could result in imbalance in disease status at the time of intervention and introduce systematic bias related to spontaneous recovery. In addition, it is not feasible in clinical practice to require patients who actively seek intervention to be randomly assigned to a control group. Therefore, this study adopted a predefined stratification based on recovery duration rather than random allocation. We have added this design consideration and its rationale to the Study Design section. (line124-132)

2. Why was such a high power used for the sample size calculation? What outcome was the sample size based on? What detectable difference was used? What SD was used? Please justify each choice.

Reply: Thank you for the reviewer’s insightful comment. We adopted a power of 0.95 based on the following considerations. First, the final analysis of this trial was performed using generalized estimating equations (GEE) with five study groups and a group × time interaction term. Given the nature of Long COVID, patients’ symptoms may improve spontaneously over time, and we anticipated heterogeneous recovery trajectories across the four experimental groups (0–3, 3–6, 6–9, and 9–12 months post-recovery). Such heterogeneity can disperse the overall effect across groups and reduce the effective power for detecting the interaction effect, which is the key parameter of interest in this design. To mitigate this potential dilution of statistical power, we chose a more conservative power level (0.95) to ensure adequate sensitivity for detecting clinically meaningful differences. Second, randomized controlled trials in Long COVID populations remain limited, and there is insufficient preliminary evidence to precisely estimate the true effect size of incentive spirometry in this population. Under circumstances in which the expected effect size is uncertain, using a higher power helps reduce the risk of type II error, preventing the study from overlooking a potentially meaningful intervention effect. In addition, several prior studies in similar rehabilitation contexts have also adopted a high statistical power (e.g., Power = 0.95) to ensure that clinically meaningful intervention effects can be reliably detected.( Çetin, Ş., Çetin, M., Kaplan, A., Kaplan, Ö., & Çelik, İ. (2024). The Effect of Respiratory Exercises on People with Ongoing Dyspnea and Recovered from COVID-19. EJMA, 4(1), 13-21. https://doi.org/10.14744/ejma.2023.29491.)

For these reasons, adopting a power of 0.95 was deemed an appropriate and conservative strategy for a multi-group design requiring interaction analyses.

The sample size calculation in this study was based on the primary outcome, the Dyspnea-12 (D-12) score. In studies involving Long COVID–related dyspnea, the variability of the D-12 measure generally falls within a moderate range, with standard deviations commonly reported between 5 and 7 points, which represents a typical variation observed in the literature. (Ekström, M. P., Bornefalk, H., Sköld, C. M., Janson, C., Blomberg, A., Bornefalk-Hermansson, A., Igelström, H., Sandberg, J., & Sundh, J. (2020). Minimal Clinically Important Differences and Feasibility of Dyspnea-12 and the Multidimensional Dyspnea Profile in Cardiorespiratory Disease. Journal of pain and symptom management, 60(5), 968–975.e1. https://doi.org/10.1016/j.jpainsymman.2020.05.028).The G*Power parameters adopted in this study are consistent with those later reported in similar respiratory training intervention research. For example, Çetin et al. (2024) employed sample size estimation settings comparable to those used in our study.

3. Please be specific about the primary outcome. Was this 6MT walk at follow up or was it the change from baseline? OR was it dyspnea?

Reply: Thank you for the reviewer’s insightful comment. The primary outcomes of this study were D-12 (Dyspnea-12), PCFS (Post-COVID Functional Status), and CaO₂ (arterial oxygen content). These measures were selected as the primary endpoints because Long COVID is characterized mainly by dyspnea, post-exertional discomfort, and functional limitations. D-12 and PCFS directly assess symptom burden and functional impairment, while CaO₂ reflects physiological oxygenation status. Together, these indicators more accurately capture the direct effects of the respiratory intervention on patients’ respiratory symptoms and functional recovery.

In contrast, the 6-minute walk distance (6MWD) was designated as a secondary outcome. Although 6MWD is widely used to evaluate overall cardiopulmonary endurance, it is an indirect functional measure and can be influenced by multiple non-respiratory factors. For instance, during the early phases of the pandemic in Taiwan, confirmed COVID-19 cases were required to undergo home isolation or stay in designated quarantine facilities. This markedly reduced physical activity levels and led to muscle deconditioning in many individuals. Consequently, lower 6MWD values in some participants may reflect reduced muscle strength or deconditioning rather than impaired cardiopulmonary function alone. Therefore, 6MWD was considered more suitable as a complementary functional measure rather than a primary endpoint.

Accordingly, the primary outcomes in this study were D-12, PCFS, and CaO₂, while 6MWD served as a secondary outcome assessing overall exercise tolerance. (line143-153)

4. Were the groups defined a priori?

Reply: Thank you for the reviewer’s comment. The grouping strategy in this study was defined a priori, prior to the initiation of data collection. Because one of the objectives of the study was to compare whether the effectiveness of incentive spirometry differed across varying recovery durations, the groups were predetermined during the study design phase based on the principle of Long COVID symptom duration stratification. Accordingly, the recovery periods were divided into four stages, rather than being created post hoc after data collection. We have clarified the grouping rationale and criteria in the Methods section. (line 158-167)

Reply: Thank you for the reviewer’s suggestion. Table 2 has been revised accordingly.

Reply: Thank you for the reviewer’s suggestion. We have revised the footnotes of Table 3 to clearly explain the meaning of the p-values and P-values, allowing readers to directly interpret the pre–post changes in 6MWD and dyspnea measures for each group, as well as the statistical significance of these changes.

In addition, we thank the reviewer for this valuable suggestion. While our study was not originally designed to test a predictive hypothesis (e.g., that a longer duration of triflow training would result in greater improvement), we understand the rationale for exploring such an analysis. Accordingly, we conducted an exploratory regression model including arm, group, and their interaction. However, the results did not show a clear or robust interaction effect.

Reply: Thanks to the reviewer for the insightful comments. The GEE model in this study was constructed using a repeated-measures framework to evaluate changes before and after the intervention across the five study groups, as well as the group × time interaction. An autoregressive correlation structure of order 1 (AR(1)) was applied, and robust standard errors were used to ensure stable parameter estimation even if the working correlation structure was not perfectly specified.

8. Table 8 is extraneous, and not part of the objectives. Remove all these results.

Reply: Thank you for the reviewer’s suggestion. Table 8 has been removed from the main text and placed in the Appendix as supplementary table 1. (line 409)

Reply: Thank you for the reviewer’s comment. The improvement in D-12 was calculated as (post − pre) / pre × 100%, and the corresponding values are presented in Table 3. EG1 showed a 79% reduction, whereas EG2, EG3, and EG4 showed reductions of 59%, 62%, and 57%, respectively, indicating that EG1 demonstrated the greatest relative improvement among the experimental groups. We have referenced Table 3 in the main text for readers’ convenience. (line 418)

Reviewer #2: This is an excellent article focusing on a highly relevant and common issue currently encountered in pulmonary and medicine clinics.

Reply: Thank you for the reviewer’s valuable comments. This study has indeed presented the major cardiopulmonary comorbidities in Table 1, and chronic obstructive pulmonary disease (COPD) was excluded based on the predefined inclusion and exclusion criteria. However, certain comorbidities that could not

Attachment

Submitted filename: Response to Reviewers.docx

pone.0351553.s004.docx^{(35.2KB, docx)}

PLoS One. doi: 10.1371/journal.pone.0351553.r003

Decision Letter 1

Toufic Ahmad Chaaban

28 Jan 2026

PONE-D-25-41209R1Study on Post-Acute COVID-19 Syndrome in Improvement of COVID-19 Rehabilitated Patients by Respiratory TrainingPLOS One

Dear Dr. Hsieh,

Please address some residual minor comments related to structuring and results presentation

Please submit your revised manuscript by Mar 14 2026 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

A letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.
A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.
An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

We look forward to receiving your revised manuscript.

Kind regards,

Toufic Ahmad Chaaban, M.D.

Academic Editor

PLOS One

Journal Requirements:

If the reviewer comments include a recommendation to cite specific previously published works, please review and evaluate these publications to determine whether they are relevant and should be cited. There is no requirement to cite these works unless the editor has indicated otherwise.

Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: (No Response)

Reviewer #2: All comments have been addressed

Reviewer #3: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

Reviewer #1: No

Reviewer #2: Yes

Reviewer #3: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: No

Reviewer #2: Yes

Reviewer #3: N/A

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

Reviewer #1: No

Reviewer #2: Yes

Reviewer #3: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

**********

6. Review Comments to the Author

Reviewer #1: The paper is more understandable now.

1. Why wasn't the control group stratified on the basis of long covid? How long had the control group been suffering from long covid? How could this have affected the results?

2. The tables are now in separate documents - this is hard to review.

4. Table 4-7 - these tables are very hard to understand, and interpret. But I am happy to see the p for interaction. Can you please instead integrate the p for interaction into Table 3.

5. Also - re-arrange table 3 into Primary and Secondary outcomes.

6. Please integrate the sample size information into the main manuscript. I cannot replicate the sample size calculation - thus more details are needed. With two groups you would need a much larger sample size - I am not clear how you get to 90 total subjects.

7. the discussion says that "Among these, EG1, who underwent the intervention within 3 months of recovery, showed the greatest improvement, with a 79% reduction in D-12 scores." Is this statistically different than the changes observed in the other EGs? Your reply did not address my question.

8. The discussion emphasizes the wrong thing. The first paragraph should talk about whether D-12 change from baseline to follow up was different in the control group to the intervention group. Within group différences could have occurrence due to usual improvement of symptoms over time.

9. The second paragraph of discussion: " Moreover, many participants reported noticeable relief from dyspnea within 3 weeks of initiating IS training, along with enhanced subjective comfort and adherence." Where are these results presented? Similarly for results pertaining to the inspiratory ball count?

10. The interpretations of the effects in lines 357-389 should emphasize the difference of differences vs the control group.

11. Everywhere - the order of results should first describe the primary results.

12. For the between group differences - should these be adjusted for relevant confounders since the EG groups are not randomized?

13. lines 339-40 "indicating that participants in EGs improved their functional status to a comparable functional level as the CG." This is an incorrect interpretation. An equivalence test would need to be conducted to make this statement.

Reviewer #2: Replies from the authors suffice the questions raised in the review. I appreciate your effort in revising several key areas of the draft.

Reviewer #3: I have reviewed the revised manuscript and the authors' response to my previous concerns regarding the study design and the standard of care provided to the control group. I appreciate the authors' thorough engagement with these points.

1. Regarding the Standard Care Arm and Pulmonary Rehabilitation: I accept the authors' explanation that at the time of the study, a standardized pulmonary rehabilitation program for Long COVID was not routinely available in clinical practice in Taiwan. This context satisfactorily justifies why pulmonary rehabilitation could not be included as the "standard of care" for the control group. It resolves the ethical and scientific concern regarding the withholding of optimal care, as the study design reflected the real-world clinical reality of that specific period and region.

2. Regarding the Role of Incentive Spirometry (IS): I commend the authors for revising the Introduction to clarify the positioning of the intervention. The added text explicitly states that "although IS cannot replace pulmonary rehabilitation, it may serve as a low-cost, self-administered intervention option... to address existing gaps". This successfully shifts the framing of the study from proposing IS as a substitute for comprehensive rehab to evaluating it as a feasible, accessible alternative for resource-limited settings.

General Observations: The inclusion of Generalized Estimating Equations (GEE) analysis and the expanded Limitations section—addressing comorbidities and pre-existing conditions —have significantly strengthened the rigor of the manuscript. The revisions have adequately addressed my methodological concerns.

I have no further comments and recommend the manuscript for publication.

**********

7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: Yes: Madhu Kalyan Pendurthi

Reviewer #3: Yes: Dapanaduwage Amila Vindana Rathnapala

**********

To ensure your figures meet our technical requirements, please review our figure guidelines: https://journals.plos.org/plosone/s/figures

You may also use PLOS’s free figure tool, NAAS, to help you prepare publication quality figures: https://journals.plos.org/plosone/s/figures#loc-tools-for-figure-preparation.

NAAS will assess whether your figures meet our technical requirements by comparing each figure against our figure specifications.

PLoS One. 2026 Jun 22;21(6):e0351553. doi: 10.1371/journal.pone.0351553.r004

Author response to Decision Letter 2

21 Feb 2026

Reviewer #1: The paper is more understandable now.

1. Why wasn't the control group stratified on the basis of long covid? How long had the control group been suffering from long covid? How could this have affected the results?

Reply:

Thank you for the reviewer’s suggestion. This study employed a quasi-experimental design rather than a fully randomized controlled trial. The primary objective was to compare the effectiveness of incentive spirometry initiated at different recovery time points; therefore, stratification based on recovery duration from Long COVID was applied only within the experimental groups. The control group was designed to observe the natural course of symptom changes over time without intervention, and thus was not further stratified according to the duration of Long COVID.

The duration from COVID-19 infection to enrollment for participants in the control group has been presented in Table 2 for reference.

We acknowledge that the absence of stratification in the control group may have introduced a degree of within-group heterogeneity. However, as recruitment occurred during the later phase of the pandemic, the distribution of Long COVID cases was inherently uneven. Imposing additional stratification criteria might have further reduced the sample size and compromised the feasibility of statistical analyses. Nevertheless, baseline comparisons revealed no significant differences among groups. Therefore, we believe that this design limitation is unlikely to have materially affected the main conclusions. Future larger-scale studies may consider stratified designs to further enhance internal validity. We appreciate the reviewer’s insightful suggestion.

2. The tables are now in separate documents - this is hard to review.

Reply:Thank you for the reviewer’s suggestion. We have now reintegrated all tables into the main manuscript to facilitate review and improve readability.

4. Table 4-7 - these tables are very hard to understand, and interpret. But I am happy to see the p for interaction. Can you please instead integrate the p for interaction into Table 3.

Reply:

Thank you for the reviewer’s valuable suggestion. Original Table 3 presents the between-group comparisons as well as the within-group pre–post differences and their corresponding p-values. In contrast, Tables 4–7 report the results of the generalized estimating equations (GEE) analysis examining the time × group interaction. As these analyses are based on different statistical methods and analytical frameworks, they were not combined into a single table.

However, we acknowledge that presenting Tables 4–7 separately may have affected the readability and clarity of the results. After considering the reviewer’s suggestion, we have consolidated the GEE results from Tables 4–7 into a single table and reorganized them according to primary and secondary outcomes to improve overall clarity and interpretability.

5. Also - re-arrange table 3 into Primary and Secondary outcomes.

Reply:

Thank you for the reviewer’s suggestion. The order of presentation in Table 3 has been revised to distinguish between primary and secondary outcomes.

Reply:

Thank you for the reviewer’s comment. The sample size estimation in this study was not based on a two-group comparison. Instead, it was calculated according to a five-group design (four experimental groups and one control group) using a repeated-measures within–between interaction framework, with all groups analyzed concurrently. The control group received no intervention. We have clearly specified this design in the Methods section (Line 202-208) to avoid misunderstanding that the study was structured as a two-group comparison trial.

Reply:Thank you for the reviewer’s further clarification. Although EG1 demonstrated the highest percentage reduction in D-12 scores, EG4 showed a relatively larger absolute mean difference between pre- and post-intervention measurements. This may be attributable to more severe baseline symptoms in EG4, resulting in greater room for improvement. However, to avoid overinterpretation, we have revised the wording in the Discussion. The original statement emphasizing that EG1 showed the “greatest” improvement has been removed and replaced with a statement indicating that both EG1 and EG4 demonstrated statistically significant improvements in the GEE analysis (Line 451-453), in order to more accurately reflect the statistical findings.

Reply:

1. Thank you for the reviewer’s important comment. We have revised the first paragraph of the Discussion to explicitly emphasize the difference in change between the EGs and the CG, rather than focusing solely on within-group improvements. (line 436-439)

2. We fully agree that within-group improvement may partly reflect the natural resolution of symptoms over time. Therefore, this potential effect was considered at both the study design and statistical analysis stages. In this study, GEE were used to examine the time × group interaction, allowing us to assess whether the intervention effect was significantly greater than the natural change observed in the CG, thereby minimizing the influence of time effects alone.

Reply:

Thank you for the reviewer’s comment. Regarding the statement that “many participants reported noticeable relief from dyspnea within 3 weeks of initiating IS training, along with enhanced subjective comfort and adherence,” we have clarified in the revised manuscript that this information was obtained during the mid-intervention telephone follow-up conducted at week 3. During this follow-up, participants provided verbal reports regarding changes in their symptoms and training experiences. (line454-456)

As for the inspiratory ball count, its primary purpose was to serve as a training reference during the intervention, helping participants monitor their inspiratory effort. The corresponding data are presented in Table 3. However, because this measure represents only a semi-quantitative visual feedback provided by the incentive spirometer and does not precisely reflect actual inspiratory volume, and because it was not predefined as a primary or secondary outcome in the study design, it was not described as a formal efficacy outcome in the Results section. Instead, it is presented as supplementary information for reference.

10. The interpretations of the effects in lines 357-389 should emphasize the difference of differences vs the control group.

Reply:

Thank you for the reviewer’s comment. We have revised the interpretation accordingly, shifting the emphasis from within-group pre–post changes to highlighting the difference-in-differences between the EGs and the CG.

11. Everywhere - the order of results should first describe the primary results.

Reply:T

hank you for the reviewer’s comment. We have reorganized the Results section to present the primary outcomes first, followed by the secondary outcomes, in order to improve clarity and better align with the study design.

12. For the between group differences - should these be adjusted for relevant confounders since the EG groups are not randomized?

Reply:Thank you for the reviewer’s comment. We agree with the reviewer’s perspective. Although randomization was not performed, we made efforts to ensure comparability at baseline, and no significant differences were observed among groups in baseline characteristics (as presented in Table 2). In addition, the GEE approach accounts for within-subject correlations and time effects, which partially addresses potential confounding related to repeated measurements. Therefore, no additional covariate adjustments were included in the primary analysis. We appreciate this suggestion and will take it into consideration in future larger-scale studies.

Reply:

Thank you for the reviewer’s correction. We agree that this statement may constitute an overinterpretation, and that the term “comparable” should not be used without conducting an equivalence test. Therefore, we have removed this sentence from the main text to avoid potential misunderstanding.

Reviewer #2: Replies from the authors suffice the questions raised in the review. I appreciate your effort in revising several key areas of the draft.

Reply:

Thank you very much for the reviewer’s recognition. We sincerely appreciate your thoughtful comments and encouragement. We will continue to refine our work in this field and consider conducting larger-scale studies in the future to further strengthen the evidence base.

I have no further comments and recommend the manuscript for publication.

Reply:

Thank you very much for the reviewer’s thoughtful comments and encouragement. We truly appreciate your support, and will continue to refine our work and aim to conduct larger-scale studies in the future to further strengthen the evidence in this field.

Attachment

Submitted filename: Response to Reviewers-0217.pdf

pone.0351553.s005.pdf^{(194.4KB, pdf)}

PLoS One. doi: 10.1371/journal.pone.0351553.r005

Decision Letter 2

Toufic Ahmad Chaaban

10 Apr 2026

PONE-D-25-41209R2Study on Post-Acute COVID-19 Syndrome in Improvement of COVID-19 Rehabilitated Patients by Respiratory TrainingPLOS One

Dear Dr. Hsieh,

==============================

ACADEMIC EDITOR: please make corrections according to reviewers' comments or write a detailed rebuttal on a point-by-point basis.

==============================

Please submit your revised manuscript by May 25 2026 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

A letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.
A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.
An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

As the corresponding author, your ORCID iD is verified in the submission system and will appear in the published article. PLOS supports the use of ORCID, and we encourage all coauthors to register for an ORCID iD and use it as well. Please encourage your coauthors to verify their ORCID iD within the submission system before final acceptance, as unverified ORCID iDs will not appear in the published article. Only the individual author can complete the verification step; PLOS staff cannot verify ORCID iDs on behalf of authors.

We look forward to receiving your revised manuscript.

Kind regards,

Davor Plavec, MD, MSc, PhD, Prof.

Academic Editor

PLOS One

Journal Requirements:

Additional Editor Comments:

Dear Authors,

please make corrections according to reviewers' comments or write a detailed rebuttal on a point-by-point basis.

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

Reviewer #1: (No Response)

Reviewer #2: All comments have been addressed

Reviewer #3: All comments have been addressed

Reviewer #4: (No Response)

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

Reviewer #1: Partly

Reviewer #2: Yes

Reviewer #3: Yes

Reviewer #4: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: N/A

Reviewer #4: I Don't Know

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

Reviewer #1: No

Reviewer #2: Yes

Reviewer #3: Yes

Reviewer #4: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

Reviewer #4: Yes

**********

6. Review Comments to the Author

Reviewer #1: 1. Table 3 remove the t-value from the table.

2. Table 4; Present only the beta and a 95% CI. Remove Se, Wald chi square, df and p value.

3. table 4 is described as interaction effects between group and time. Are you sure that is what you are presenting here? Please write out the model and show which parameters are included in the table.

4. How were the subjects who dropped out handled? If these are excluded then this is not an intention to treat analysis and this should be acknowledged as a potential source of bias.

5. Please also acknowledge as a limitation the very small size of the control group, which was not stratified by length of long COVID.

Reviewer #2: (No Response)

Reviewer #3: The authors have thoroughly and satisfactorily addressed my previous concerns regarding the study design and the intervention's clinical role.

Specifically, the authors provided crucial context regarding the lack of standardized pulmonary rehabilitation programs in the study's region during the data collection period. This adequately justifies the composition of the standard care control arm and resolves my ethical and methodological concerns. Furthermore, the revised Introduction accurately positions Incentive Spirometry not as a replacement for comprehensive pulmonary rehabilitation, but as a practical, low-cost, and accessible adjunct for resource-limited settings.

These clarifications significantly strengthen the manuscript's clinical relevance and scientific integrity. I have no further concerns and recommend the manuscript for publication.

Reviewer #4: Dear colleagues, thank you for giving me the opportunity to review your article. Basically, the article is of interest in the sense of post-covid rehabilitation possibilities. I had not the opportunity to review previous versions, but this version is much improved. I would have some minor suggestions to even more improve your article:

1. “Study on Post-Acute COVID-19 Syndrome in Improvement of 2 COVID-19 Rehabilitated Patients by Respiratory Training.”

The article title can be misleading and not informative enough what the article is about so I would suggest you to rephrase it.

2. “Currently, no definitive treatment for Long COVID.”

This is not a valid sentence, correct it.

3. “Respiratory training—commonly used to manage symptoms in chronic pulmonary disease—is considered a potentially beneficial intervention for Long COVID; however, supporting evidence remains limited.”

This is not truth, evidence exist so please rephrase this sentence and add new references. That should be corrected throughout the article.

4. “provides patients with an alternative choice”

No truth because it was not compared with pulmonary rehabilitation. You should omit this sentence.

5. “Participants were randomly assigned to groups based on recovery duration.”

If they were assigned by recovery duration what was random? The later explanation in the article is correct, but this sentence is not.

6. “The flow diagram of participant enrollment and allocation in figure 1.”

Incorrect sentence, please correct it.

Best regards

**********

7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: Yes: Madhu Kalyan Pendurthi

Reviewer #3: Yes: Dapanaduwage Amila Vindana Rathnapala

Reviewer #4: No

**********

To ensure your figures meet our technical requirements, please review our figure guidelines: https://journals.plos.org/plosone/s/figures

You may also use PLOS’s free figure tool, NAAS, to help you prepare publication quality figures: https://journals.plos.org/plosone/s/figures#loc-tools-for-figure-preparation.

NAAS will assess whether your figures meet our technical requirements by comparing each figure against our figure specifications.

PLoS One. 2026 Jun 22;21(6):e0351553. doi: 10.1371/journal.pone.0351553.r006

Author response to Decision Letter 3

24 Apr 2026

Reviewer #1:

1. Table 3 remove the t-value from the table.

Reply: Thank you for the reviewer’s comment. The t-values have been removed from Table 3. (line367)

2. Table 4; Present only the beta and a 95% CI. Remove Se, Wald chi square, df and p value.

Reply: Thank you for the reviewer’s suggestion. We have revised Table 4 to present only the β coefficients and 95% confidence intervals, and have removed the SE, Wald chi-square, degrees of freedom, and p-values accordingly. (line 414)

Reply: Thank you for the reviewer’s comment. We confirm that the results presented in Table 4 represent the interaction effects (group × time) in the GEE analysis.

In our study, the GEE model included group, time, and the group × time interaction term as the main independent variables. The β coefficients presented in Table 4 correspond to the interaction terms, reflecting the differences in the magnitude of change before and after the intervention between each EG and the CG. A clarification has also been added below Table 4. (line 416-419)

4. How were the subjects who dropped out handled? If these are excluded then this is not an intention to treat analysis and this should be acknowledged as a potential source of bias.

Reply: Thank you for the reviewer’s comment. In this study, participants who dropped out were excluded from the final analysis; therefore, a per-protocol analysis was conducted rather than an intention-to-treat (ITT) analysis. We acknowledge that this approach may introduce attrition bias, particularly if the characteristics of participants who dropped out differ from those who completed the study, which may affect the internal validity of the results. This potential limitation has been addressed in both the Methods and Limitations sections to provide a more comprehensive explanation. (line 142 and line 564-566)

5. Please also acknowledge as a limitation the very small size of the control group, which was not stratified by length of long COVID.

Reply: Thank you for the reviewer’s comment. This study adopted a quasi-experimental design, with the primary aim of comparing intervention effects across different recovery durations; therefore, stratification based on recovery time was applied only within the EGs. The CG was used as a reference for the natural progression of symptoms without intervention and was not further stratified. We agree that the small sample size of the CG and the lack of stratification by duration of long COVID may affect the stability of between-group comparisons and the internal validity of the study, thereby influencing the interpretation of the results. This limitation has been acknowledged in the revised manuscript. (line 560-564)

Reviewer #2: (No Response)

Reviewer #3: The authors have thoroughly and satisfactorily addressed my previous concerns regarding the study design and the intervention's clinical role.

These clarifications significantly strengthen the manuscript's clinical relevance and scientific integrity. I have no further concerns and recommend the manuscript for publication.

Reply: Thank you for the reviewer’s comments. We will continue to refine our work and contribute further evidence in this field in future studies.

1. “Study on Post-Acute COVID-19 Syndrome in Improvement of 2 COVID-19 Rehabilitated Patients by Respiratory Training.”

The article title can be misleading and not informative enough what the article is about so I would suggest you to rephrase it.

Reply: Thank you for the reviewer’s suggestion. We agree that the original title was not sufficiently precise and did not fully reflect the focus of the study. Therefore, we have revised the title to “Effects of Incentive Spirometer Training on Dyspnea and Functional Status in Patients with Long COVID,” emphasizing the main intervention and key outcome measures (dyspnea and functional status) to improve clarity and informativeness.

2. “Currently, no definitive treatment for Long COVID.”

This is not a valid sentence, correct it.

Reply: Thank you for the reviewer’s suggestion. We have corrected the grammatical error and revised the sentence to avoid an overly absolute statement. (line 86)

This is not truth, evidence exist so please rephrase this sentence and add new references. That should be corrected throughout the article.

Reply: Thank you for the reviewer’s comment. We agree that existing studies have supported the potential benefits of respiratory training for long COVID. Therefore, we have revised the statement to emphasize the heterogeneity of the current evidence and the lack of consistent conclusions, and have added relevant references to provide a more comprehensive and accurate description. (line87-91)

4. “provides patients with an alternative choice”

No truth because it was not compared with pulmonary rehabilitation. You should omit this sentence.

Reply: Thank you for the reviewer’s suggestion. We agree with this concern and have removed the statement accordingly. (line 111)

5. “Participants were randomly assigned to groups based on recovery duration.”

If they were assigned by recovery duration what was random? The later explanation in the article is correct, but this sentence is not.

Reply: Thank you for the reviewer’s comment. We agree that the original statement was not sufficiently precise and may have caused misunderstanding regarding the group allocation process. In this study, participants were first randomly assigned to the experimental and control groups, and the experimental group was subsequently stratified based on recovery duration. Therefore, the grouping based on recovery duration was not part of the randomization process. The relevant sentence has been revised to more clearly describe the study design. (line120 and line 258-260)

6. “The flow diagram of participant enrollment and allocation in figure 1.”

Incorrect sentence, please correct it.

Reply: Thank you for the reviewer’s comment. We have revised the sentence for clarity and accuracy. (line 126)

Attachment

Submitted filename: Response to Reviewers 0420.docx

pone.0351553.s006.docx^{(22KB, docx)}

PLoS One. doi: 10.1371/journal.pone.0351553.r007

Decision Letter 3

Toufic Ahmad Chaaban

18 May 2026

PONE-D-25-41209R3Effects of Incentive Spirometer Training on Dyspnea and Functional Status in Patients with Long COVIDPLOS One

Dear Dr. Hsieh,

==============================

ACADEMIC EDITOR: Dear Authors, please make suggested revisions according to reviewers' comments or write a detailed rebuttal on a point-by-point basis.

==============================

Please submit your revised manuscript by Jul 02 2026 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

A letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.
A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.
An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

We look forward to receiving your revised manuscript.

Kind regards,

Davor Plavec, MD, MSc, PhD, Prof.

Academic Editor

PLOS One

Journal Requirements:

Additional Editor Comments:

Dear Authors,

please make suggested revisions according to reviewers' comments or write a detailed rebuttal on a point-by-point basis.

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

Reviewer #1: (No Response)

Reviewer #3: All comments have been addressed

Reviewer #4: (No Response)

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

Reviewer #1: Yes

Reviewer #3: Partly

Reviewer #4: No

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: I Don't Know

Reviewer #3: N/A

Reviewer #4: I Don't Know

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

Reviewer #1: No

Reviewer #3: Yes

Reviewer #4: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

Reviewer #1: Yes

Reviewer #3: Yes

Reviewer #4: Yes

**********

6. Review Comments to the Author

Reviewer #1: Thank you for the revisions.

I think there is a problem with the values for the PCFS interaction terms presented in Table 4. These do not seem to line up with what would be expected given the results presented in Table3. Please verify.

The other values presented are close to what is presented in Table 3.

Reviewer #3: The authors have successfully maintained the crucial clarifications requested in the previous review rounds, effectively resolving my primary methodological and ethical concerns.

Specifically, the manuscript correctly contextualizes the control arm by acknowledging the absence of standardized pulmonary rehabilitation programs during the study period. This provides a solid ethical and scientific justification for the study design based on the clinical reality of the time.

Furthermore, the authors have appropriately defined the role of Incentive Spirometry. By framing it as a practical, self-administered adjunct rather than a replacement for comprehensive pulmonary rehabilitation, the study offers a highly relevant, low-cost option. This distinction is particularly valuable for optimizing respiratory care in resource-limited settings where formal, multidisciplinary rehabilitation programs may be inaccessible to patients.

The revisions have significantly strengthened the manuscript's clinical applicability and scientific integrity. The authors have adequately addressed all of my feedback, and I have no further concerns. I recommend this article for publication.

Reviewer #4: Dear colleagues,

Thank you for giving me the opportunity to review your article. This is an interesting idea but unfortunately, I don’t believe that you have scientifically proof your conclusion: “In conclusion, IS represents an effective and practical strategy for alleviating dyspnea and improving functional limitations in individuals with Long COVID.“

The design of your study has many limitations as you have correctly stated in the limitations section but there is also one big limitation and you have not mentioned it. The lung function analysis of the patients with CO diffusion capacity which is known to be the important parameter considering the exercise limitation in post-COVID patients. From my point of view, no studies of post-COVID respiratory investigations should be done or at least published without the DLco data. Also, the intervention itself could have important placebo effect that was not possible to exclude by the study design.

If you can correct for previously stated problems, I would be glad to re-review your article.

Best regards

**********

7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #3: Yes: Dapanaduwage Amila Vindana Rathnapala

Reviewer #4: No

**********

To ensure your figures meet our technical requirements, please review our figure guidelines: https://journals.plos.org/plosone/s/figures

You may also use PLOS’s free figure tool, NAAS, to help you prepare publication quality figures: https://journals.plos.org/plosone/s/figures#loc-tools-for-figure-preparation.

NAAS will assess whether your figures meet our technical requirements by comparing each figure against our figure specifications.

PLoS One. 2026 Jun 22;21(6):e0351553. doi: 10.1371/journal.pone.0351553.r008

Author response to Decision Letter 4

23 May 2026

Reviewer #1: Thank you for the revisions.

The other values presented are close to what is presented in Table 3.

Reply: Thank you for your careful review and for identifying this issue. After re-examining the analysis, we found that the previous GEE model for the PCFS scale had been conducted using an ordinal-variable approach, whereas Table 3 presented the PCFS results as continuous mean score changes. This difference in modeling strategy may have contributed to inconsistency in the interpretation of the interaction coefficients presented in Table 4.

To improve the consistency and interpretability of the results, we reanalyzed the PCFS outcomes using a continuous-variable GEE model and updated the corresponding interaction coefficients in Table 4 accordingly. We sincerely appreciate the reviewer’s careful observation, which helped us further improve the clarity of the manuscript. (P.19 Table4)

Reviewer #3: The authors have successfully maintained the crucial clarifications requested in the previous review rounds, effectively resolving my primary methodological and ethical concerns.

Reply: We sincerely thank the reviewer for the thoughtful comments and positive evaluation of our manuscript. Your feedback has been highly valuable in strengthening the clinical applicability and scientific rigor of this study. We are grateful for your encouragement and will continue striving to improve the quality and clinical relevance of our future research.

Reviewer #4: Dear colleagues,

Thank you for giving me the opportunity to review your article. This is an interesting idea but unfortunately, I don’t believe that you have scientifically proof your conclusion: „In conclusion, IS represents an effective and practical strategy for alleviating dyspnea and improving functional limitations in individuals with Long COVID.“

If you can correct for previously stated problems, I would be glad to re-review your article.

Best regards

Reply: We sincerely thank the reviewer for the thoughtful and constructive comments. We agree that the conclusions of our study should be interpreted cautiously given the limitations of the study design. Accordingly, we have revised the limitation and conclusions throughout the manuscript to avoid overly definitive statements regarding the effectiveness of IS intervention.

Regarding the absence of DLco assessment, we fully acknowledge that diffusion capacity is an important parameter when evaluating pulmonary impairment and exercise limitation in patients with post-COVID conditions. However, the primary aim of this study was to evaluate the feasibility and potential benefits of a simple, low-cost, home-based respiratory training intervention that could be implemented in non-hospitalized individuals recovering from COVID-19.

During the study period (2023–2024), most participants had already returned to community or home settings and were not hospitalized. Performing DLco measurements would have required participants to repeatedly return to the hospital pulmonary function laboratory, which posed practical difficulties and potentially increased unnecessary exposure risks during the post-pandemic period.

In addition, at the time the study protocol was initially designed, there was still limited evidence regarding standardized respiratory assessment strategies for Long COVID. Therefore, DLco measurement was not incorporated into the original study design. We appreciate the reviewer for highlighting this important issue, and we agree that future studies should incorporate comprehensive pulmonary function assessments, including DLco analysis, to further clarify the physiological mechanisms underlying symptom improvement. (P.25 line580-585)

We also agree that placebo effects and expectation-related responses could not be fully excluded because of the open-label design and the absence of a sham intervention group. Therefore, we have further emphasized this issue in the Limitations section and revised the interpretation of the findings accordingly. (P.26 line 577-579)

Attachment

Submitted filename: Response to Reviewer 0520.pdf

pone.0351553.s007.pdf^{(172.1KB, pdf)}

PLoS One. doi: 10.1371/journal.pone.0351553.r009

Decision Letter 4

Toufic Ahmad Chaaban

28 May 2026

Effects of Incentive Spirometer Training on Dyspnea and Functional Status in Patients with Long COVID

PONE-D-25-41209R4

Dear Dr. Hsieh,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

An invoice will be generated when your article is formally accepted. Please note, if your institution has a publishing partnership with PLOS and your article meets the relevant criteria, all or part of your publication costs will be covered. Please make sure your user information is up-to-date by logging into Editorial Manager at Editorial Manager® and clicking the ‘Update My Information' link at the top of the page. For questions related to billing, please contact billing support.

If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

Kind regards,

Davor Plavec, MD, MSc, PhD, Prof.

Academic Editor

PLOS One

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

Reviewer #1: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

Reviewer #1: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

Reviewer #1: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

Reviewer #1: Yes

**********

6. Review Comments to the Author

Reviewer #1: (No Response)

**********

7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

**********

PLoS One. doi: 10.1371/journal.pone.0351553.r010

Acceptance letter

Toufic Ahmad Chaaban

PONE-D-25-41209R4

PLOS One

Dear Dr. Hsieh,

I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS One. Congratulations! Your manuscript is now being handed over to our production team.

At this stage, our production department will prepare your paper for publication. This includes ensuring the following:

* All references, tables, and figures are properly cited

* All relevant supporting information is included in the manuscript submission,

* There are no issues that prevent the paper from being properly typeset

You will receive further instructions from the production team, including instructions on how to review your proof when it is ready. Please keep in mind that we are working through a large volume of accepted articles, so please give us a few days to review your paper and let you know the next and final steps.

Lastly, if your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

You will receive an invoice from PLOS for your publication fee after your manuscript has reached the completed accept phase. If you receive an email requesting payment before acceptance or for any other service, this may be a phishing scheme. Learn how to identify phishing emails and protect your accounts at https://explore.plos.org/phishing.

If we can help with anything else, please email us at customercare@plos.org.

Thank you for submitting your work to PLOS ONE and supporting open access.

Kind regards,

PLOS ONE Editorial Office Staff

on behalf of

Dr. Davor Plavec

Academic Editor

PLOS One

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

S1 Table. Item-Level Analysis of D-12 scale.

(DOCX)

pone.0351553.s001.docx^{(34.9KB, docx)}

S1 File. CONSORT checklist and Protocol documents.

(PDF)

pone.0351553.s002.pdf^{(1.8MB, pdf)}

Attachment

Submitted filename: PLOS Reviews 30.10.2025.docx

pone.0351553.s003.docx^{(13.6KB, docx)}

Attachment

Submitted filename: Response to Reviewers.docx

pone.0351553.s004.docx^{(35.2KB, docx)}

Attachment

Submitted filename: Response to Reviewers-0217.pdf

pone.0351553.s005.pdf^{(194.4KB, pdf)}

Attachment

Submitted filename: Response to Reviewers 0420.docx

pone.0351553.s006.docx^{(22KB, docx)}

Attachment

Submitted filename: Response to Reviewer 0520.pdf

pone.0351553.s007.pdf^{(172.1KB, pdf)}

Data Availability Statement

All relevant data are within the manuscript and its Supporting Information files.

[pone.0351553.ref001] 1.Koc HC, Xiao J, Liu W, Li Y, Chen G. Long COVID and its Management. Int J Biol Sci. 2022;18(12):4768–80. [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref002] 2.Cabrera Martimbianco AL, Pacheco RL, Bagattini ÂM, Riera R. Frequency, signs and symptoms, and criteria adopted for long COVID-19: A systematic review. Int J Clin Pract. 2021;75(10):e14357. doi: 10.1111/ijcp.14357 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref003] 3.Davis HE, Assaf GS, McCorkell L, Wei H, Low RJ, Re’em Y, et al. Characterizing long COVID in an international cohort: 7 months of symptoms and their impact. EClinicalMedicine. 2021;38:101019. doi: 10.1016/j.eclinm.2021.101019 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref004] 4.O’ Mahony L, Buwalda T, Blair M, Forde B, Lunjani N, Ambikan A, et al. Impact of Long COVID on health and quality of life. HRB Open Res. 2022;5:31. doi: 10.12688/hrbopenres.13516.1 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref005] 5.Torres-Castro R, Vasconcello-Castillo L, Alsina-Restoy X, Solis-Navarro L, Burgos F, Puppo H, et al. Respiratory function in patients post-infection by COVID-19: a systematic review and meta-analysis. Pulmonology. 2021;27(4):328–37. doi: 10.1016/j.pulmoe.2020.10.013 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref006] 6.van Kessel SAM, Olde Hartman TC, Lucassen PLBJ, van Jaarsveld CHM. Post-acute and long-COVID-19 symptoms in patients with mild diseases: a systematic review. Fam Pract. 2022;39(1):159–67. doi: 10.1093/fampra/cmab076 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref007] 7.Visco V, Vitale C, Rispoli A, Izzo C, Virtuoso N, Ferruzzi GJ, et al. Post-COVID-19 Syndrome: Involvement and Interactions between Respiratory, Cardiovascular and Nervous Systems. J Clin Med. 2022;11(3):524. doi: 10.3390/jcm11030524 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref008] 8.Li S, Dai B, Hou Y, Zhang L, Liu J, Hou H, et al. Effect of pulmonary rehabilitation for patients with long COVID-19: a systematic review and meta-analysis of randomized controlled trials. Ther Adv Respir Dis. 2025;19. doi: 10.1177/17534666251323482 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref009] 9.Martínez-Pozas O, Meléndez-Oliva E, Rolando LM, Rico JAQ, Corbellini C, Sánchez Romero EA. The pulmonary rehabilitation effect on long covid-19 syndrome: A systematic review and meta-analysis. Physiother Res Int. 2024;29(2):e2077. doi: 10.1002/pri.2077 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref010] 10.Singh SJ, Baldwin MM, Daynes E, Evans RA, Greening NJ, Jenkins RG, et al. Respiratory sequelae of COVID-19: pulmonary and extrapulmonary origins, and approaches to clinical care and rehabilitation. Lancet Respir Med. 2023;11(8):709–25. doi: 10.1016/S2213-2600(23)00159-5 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref011] 11.Ahmed I, Mustafaoglu R, Yeldan I, Yasaci Z, Erhan B. Effect of Pulmonary Rehabilitation Approaches on Dyspnea, Exercise Capacity, Fatigue, Lung Functions, and Quality of Life in Patients With COVID-19: A Systematic Review and Meta-analysis. Arch Phys Med Rehabil. 2022;103(10):2051–62. doi: 10.1016/j.apmr.2022.06.007 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref012] 12.Bargahi M, Alavi-Moghaddam M, Karimi M, Azizan Z, Jafarzadeh F, Javaherian M, et al. Safety and Efficacy of Incentive Spirometer in Covid-19 Pneumonia; a Randomized Clinical Trial. Arch Acad Emerg Med. 2024;12(1):e51. doi: 10.22037/aaem.v12i1.2244 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref013] 13.Jimeno-Almazán A, Pallarés JG, Buendía-Romero Á, Martínez-Cava A, Franco-López F, Sánchez-Alcaraz Martínez BJ, et al. Post-COVID-19 Syndrome and the Potential Benefits of Exercise. International Journal of Environmental Research and Public Health. 2021;18(10). [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref014] 14.Liu K, Zhang W, Yang Y, Zhang J, Li Y, Chen Y. Respiratory rehabilitation in elderly patients with COVID-19: A randomized controlled study. Complement Ther Clin Pract. 2020;39:101166. doi: 10.1016/j.ctcp.2020.101166 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref015] 15.Troosters T, Janssens W, Demeyer H, Rabinovich RA. Pulmonary rehabilitation and physical interventions. Eur Respir Rev. 2023;32(168):220222. doi: 10.1183/16000617.0222-2022 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref016] 16.Chen Y-H, Hsieh Y-S. A Narrative Review of Impact of Incentive Spirometer Respiratory Training in Long COVID. Int J Gen Med. 2024;17:5233–46. doi: 10.2147/IJGM.S492772 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref017] 17.Eltorai AEM, Baird GL, Eltorai AS, Pangborn J, Antoci V Jr, Cullen HA, et al. Perspectives on Incentive Spirometry Utility and Patient Protocols. Respir Care. 2018;63(5):519–31. doi: 10.4187/respcare.05872 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref018] 18.Eltorai AEM, Szabo AL, Antoci V Jr, Ventetuolo CE, Elias JA, Daniels AH, et al. Clinical Effectiveness of Incentive Spirometry for the Prevention of Postoperative Pulmonary Complications. Respir Care. 2018;63(3):347–52. doi: 10.4187/respcare.05679 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref019] 19.Sum S-K, Peng Y-C, Yin S-Y, Huang P-F, Wang Y-C, Chen T-P, et al. Using an incentive spirometer reduces pulmonary complications in patients with traumatic rib fractures: a randomized controlled trial. Trials. 2019;20(1):797. doi: 10.1186/s13063-019-3943-x [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref020] 20.Curci C, Negrini F, Ferrillo M, Bergonzi R, Bonacci E, Camozzi DM, et al. Functional outcome after inpatient rehabilitation in postintensive care unit COVID-19 patients: findings and clinical implications from a real-practice retrospective study. Eur J Phys Rehabil Med. 2021;57(3):443–50. doi: 10.23736/S1973-9087.20.06660-5 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref021] 21.Gudivada KK, Tikka SK, Bonthala Y, Miryala SK, Ch RP, Garre S. Association Between Post-COVID Pulmonary Dysfunction and New-Onset Anxiety and Depression and the Impact of Incentive Spirometer-Based Exercises. Prim Care Companion CNS Disord. 2023;25(5):23m03490. doi: 10.4088/PCC.23m03490 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref022] 22.Toor H, Kashyap S, Yau A, Simoni M, Farr S, Savla P, et al. Efficacy of Incentive Spirometer in Increasing Maximum Inspiratory Volume in an Out-Patient Setting. Cureus. 2021;13(10):e18483. doi: 10.7759/cureus.18483 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref023] 23.Garcia A, Sarenas J. Blowing in the Wind: Use of Incentive Spirometry in Preventing Acute Respiratory Distress Syndrome for Patients with Moderate COVID. European Respiratory Society. 2022. [Google Scholar]

[pone.0351553.ref024] 24.Srinivasan V, Kandakurti PK, Alagesan J, Suganthirababu P, Kishore J, Jenifer A, et al. Efficacy of pursed lip breathing with bhastrika pranayama vs incentive spirometry in rehabilitating post Covid 19 follow up-a randomized control study. Turkish Journal of Physiotherapy and Rehabilitation. 2021;32(3):402–7. [Google Scholar]

[pone.0351553.ref025] 25.Benkalfate N, Eschapasse E, Georges T, Leblanc C, Dirou S, Melscoet L, et al. Evaluation of the Post-COVID-19 Functional Status (PCFS) Scale in a cohort of patients recovering from hypoxemic SARS-CoV-2 pneumonia. BMJ Open Respir Res. 2022;9(1):e001136. doi: 10.1136/bmjresp-2021-001136 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref026] 26.Klok FA, Boon GJAM, Barco S, Endres M, Geelhoed JJM, Knauss S, et al. The Post-COVID-19 Functional Status scale: a tool to measure functional status over time after COVID-19. Eur Respir J. 2020;56(1):2001494. doi: 10.1183/13993003.01494-2020 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref027] 27.Machado FVC, Meys R, Delbressine JM, Vaes AW, Goërtz YMJ, van Herck M, et al. Construct validity of the Post-COVID-19 Functional Status Scale in adult subjects with COVID-19. Health Qual Life Outcomes. 2021;19(1):40. doi: 10.1186/s12955-021-01691-2 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref028] 28.Amado Diago CA, Puente Maestu L, Abascal Bolado B, Agüero Calvo J, Hernando Hernando M, Puente Bats I, et al. Translation and validation of the multidimensional dyspnea-12 questionnaire. Arch Bronconeumol. 2018;54(2):74–8. [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref029] 29.Choi TCM, Chan LLY, Tsang HC, Vong YP, Cheng YK, To YL, et al. Adaptation and validation of the Chinese version of Dyspnoea-12 scale in individuals with chronic obstructive pulmonary disease. Clin Respir J. 2021;15(10):1081–7. doi: 10.1111/crj.13411 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref030] 30.Williams MT, Lewthwaite H, Paquet C, Johnston K, Olsson M, Belo LF, et al. Dyspnoea-12 and Multidimensional Dyspnea Profile: Systematic Review of Use and Properties. J Pain Symptom Manage. 2022;63(1):e75–87. doi: 10.1016/j.jpainsymman.2021.06.023 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref031] 31.Dubowitz G, Breyer K, Lipnick M, Sall JW, Feiner J, Ikeda K, et al. Accuracy of the Lifebox pulse oximeter during hypoxia in healthy volunteers. Anaesthesia. 2013;68(12):1220–3. doi: 10.1111/anae.12382 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref032] 32.Nitzan M, Romem A, Koppel R. Pulse oximetry: fundamentals and technology update. Med Devices (Auckl). 2014;7:231–9. doi: 10.2147/MDER.S47319 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref033] 33.Silverston P, Ferrari M, Quaresima V. Pulse oximetry and the pandemic. BMJ. 2022;378:e071474. doi: 10.1136/bmj-2022-071474 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref034] 34.Rizvi A, Macedo P, Babawale L, Tighe HC, Hughes JMB, Jackson JE, et al. Hemoglobin Is a Vital Determinant of Arterial Oxygen Content in Hypoxemic Patients with Pulmonary Arteriovenous Malformations. Ann Am Thorac Soc. 2017;14(6):903–11. doi: 10.1513/AnnalsATS.201611-872OC [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref035] 35.Agarwala P, Salzman SH: Six-Minute Walk Test: Clinical Role, Technique, Coding, and Reimbursement. Chest. 2020, 157(3):603–11. [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref036] 36.Bellet RN, Adams L, Morris NR. The 6-minute walk test in outpatient cardiac rehabilitation: validity, reliability and responsiveness--a systematic review. Physiotherapy. 2012;98(4):277–86. doi: 10.1016/j.physio.2011.11.003 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref037] 37.Pires IM, Denysyuk HV, Villasana MV, Sá J, Marques DL, Morgado JF, et al. Development Technologies for the Monitoring of Six-Minute Walk Test: A Systematic Review. Sensors (Basel). 2022;22(2):581. doi: 10.3390/s22020581 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref038] 38.Del Corral T, Menor-Rodríguez N, Fernández-Vega S, Díaz-Ramos C, Aguilar-Zafra S, López-de-Uralde-Villanueva I. Longitudinal study of changes observed in quality of life, psychological state cognition and pulmonary and functional capacity after COVID-19 infection: A six- to seven-month prospective cohort. J Clin Nurs. 2024;33(1):89–102. doi: 10.1111/jocn.16352 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref039] 39.Hockele LF, Sachet Affonso JV, Rossi D, Eibel B. Pulmonary and Functional Rehabilitation Improves Functional Capacity, Pulmonary Function and Respiratory Muscle Strength in Post COVID-19 Patients: Pilot Clinical Trial. Int J Environ Res Public Health. 2022;19(22):14899. doi: 10.3390/ijerph192214899 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref040] 40.Nopp S, Moik F, Klok FA, Gattinger D, Petrovic M, Vonbank K, et al. Outpatient pulmonary rehabilitation in patients with long COVID improves exercise capacity, functional status, dyspnea, fatigue, and quality of life. Respiration. 2022;101(6):593–601. [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref041] 41.Omar A, Ferreira AS, Hegazy FA, Alaparthi GK. Cardiorespiratory response to six-minute step test in post COVID-19 patients - a cross sectional study. Healthcare (Basel). 2023;11(10). [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref042] 42.Szarvas Z, Fekete M, Horvath R, Shimizu M, Tsuhiya F, Choi HE, et al. Cardiopulmonary rehabilitation programme improves physical health and quality of life in post-COVID syndrome. Ann Palliat Med. 2023;12(3):548–60. doi: 10.21037/apm-22-1143 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref043] 43.Altmann CH, Zvonova E, Richter L, Schüller PO. Pulmonary recovery directly after COVID-19 and in Long-COVID. Respir Physiol Neurobiol. 2023;315:104112. doi: 10.1016/j.resp.2023.104112 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref044] 44.Kusumawardani RI, Tinduh D, Andriati, Poerwandari D, Marhana IA, Melaniani S. The effectiveness of incentive spirometry exercise on pulmonary function in COVID-19 survivors: a randomized controlled trial study. Bali Med J. 2023;12(1):539–44. doi: 10.15562/bmj.v12i1.3956 [DOI] [Google Scholar]

[pone.0351553.ref045] 45.Abo Elyazed TI, Abd El-Hakim AAE-M, Saleh OI, Sonbol MMF, Eid HA, Moazen E, et al. Diaphragmatic strengthening exercises for patients with post COVID-19 condition after mild-to-moderate acute COVID-19 infection: a randomized controlled study. J Rehabil Med. 2024;56:jrm25491. doi: 10.2340/jrm.v56.25491 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref046] 46.Bellan M, Soddu D, Balbo PE, Baricich A, Zeppegno P, Avanzi GC, et al. Respiratory and Psychophysical Sequelae Among Patients With COVID-19 Four Months After Hospital Discharge. JAMA Netw Open. 2021;4(1):e2036142. doi: 10.1001/jamanetworkopen.2020.36142 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref047] 47.Gutiérrez-Canales LG, Muñoz-Corona C, Barrera-Chávez I, Viloria-Álvarez C, Macías AE, Martínez-Navarro LJ, et al. Persistence of COVID-19 Symptoms and Quality of Life at Three and Twelve Months after Hospital Discharge. Medicina (Kaunas). 2024;60(6):944. doi: 10.3390/medicina60060944 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref048] 48.Lippi G, Sanchis-Gomar F, Henry BM. COVID-19 and its long-term sequelae: what do we know in 2023? Pol Arch Intern Med. 2023;133(4):16402. doi: 10.20452/pamw.16402 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref049] 49.Nasrullah A, Virk S, Javed A, Shah A, Ramanujam D, Sharma A, et al. Effects of pulmonary rehabilitation on functional and psychological parameters in post-acute sequelae of SARS-CoV-2 infection (PASC) patients. BMC Pulm Med. 2024;24(1):231. doi: 10.1186/s12890-024-03047-0 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref050] 50.Hanania NA, O’Donnell DE. Activity-related dyspnea in chronic obstructive pulmonary disease: physical and psychological consequences, unmet needs, and future directions. Int J Chron Obstruct Pulmon Dis. 2019;14:1127–38. doi: 10.2147/COPD.S188141 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref051] 51.Abelson JL, Khan S, Giardino N. HPA axis, respiration and the airways in stress--a review in search of intersections. Biol Psychol. 2010;84(1):57–65. doi: 10.1016/j.biopsycho.2010.01.021 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref052] 52.Bai F, Tomasoni D, Falcinella C, Barbanotti D, Castoldi R, Mulè G, et al. Female gender is associated with long COVID syndrome: a prospective cohort study. Clin Microbiol Infect. 2022;28(4):611.e9–611.e16. doi: 10.1016/j.cmi.2021.11.002 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref053] 53.Malesevic S, Sievi NA, Baumgartner P, Roser K, Sommer G, Schmidt D, et al. Impaired health-related quality of life in long-COVID syndrome after mild to moderate COVID-19. Sci Rep. 2023;13(1):7717. doi: 10.1038/s41598-023-34678-8 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref054] 54.Xiong Q, Xu M, Li J, Liu Y, Zhang J, Xu Y, et al. Clinical sequelae of COVID-19 survivors in Wuhan, China: a single-centre longitudinal study. Clin Microbiol Infect. 2021;27(1):89–95. doi: 10.1016/j.cmi.2020.09.023 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref055] 55.Zhao Y, Shi L, Jiang Z, Zeng N, Mei H, Lu Y, et al. The phenotype and prediction of long-term physical, mental and cognitive COVID-19 sequelae 20 months after recovery, a community-based cohort study in China. Mol Psychiatry. 2023;28(4):1793–801. doi: 10.1038/s41380-023-01951-1 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref056] 56.Huang L, Li X, Gu X, Zhang H, Ren L, Guo L, et al. Health outcomes in people 2 years after surviving hospitalisation with COVID-19: a longitudinal cohort study. Lancet Respir Med. 2022;10(9):863–76. doi: 10.1016/S2213-2600(22)00126-6 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref057] 57.McGregor G, Sandhu H, Bruce J, Sheehan B, McWilliams D, Yeung J, et al. Clinical effectiveness of an online supervised group physical and mental health rehabilitation programme for adults with post-covid-19 condition (REGAIN study): multicentre randomised controlled trial. BMJ. 2024;384:e076506. doi: 10.1136/bmj-2023-076506 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref058] 58.Aljazeeri J, Almusally R, Wert Y, Abdelhalim M, Klinger C, Ramesh N, et al. Pulmonary Rehabilitation for Post-COVID-19. J Cardiopulm Rehabil Prev. 2023;43(6):438–43. doi: 10.1097/HCR.0000000000000813 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref059] 59.Natarajan A, Shetty A, Delanerolle G, Zeng Y, Zhang Y, Raymont V, et al. A systematic review and meta-analysis of long COVID symptoms. Syst Rev. 2023;12(1):88. doi: 10.1186/s13643-023-02250-0 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref060] 60.Zeng N, Zhao Y-M, Yan W, Li C, Lu Q-D, Liu L, et al. A systematic review and meta-analysis of long term physical and mental sequelae of COVID-19 pandemic: call for research priority and action. Mol Psychiatry. 2023;28(1):423–33. doi: 10.1038/s41380-022-01614-7 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref061] 61.Fincham GW, Strauss C, Montero-Marin J, Cavanagh K. Effect of breathwork on stress and mental health: A meta-analysis of randomised-controlled trials. Sci Rep. 2023;13(1):432. doi: 10.1038/s41598-022-27247-y [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref062] 62.Benedetti F, Mayberg HS, Wager TD, Stohler CS, Zubieta J-K. Neurobiological mechanisms of the placebo effect. J Neurosci. 2005;25(45):10390–402. doi: 10.1523/JNEUROSCI.3458-05.2005 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref063] 63.Tsai K-C, Huang Y-C, Liaw C-C, Tsai C-I, Chiou C-T, Lin C-J, et al. A traditional Chinese medicine formula NRICM101 to target COVID-19 through multiple pathways: A bedside-to-bench study. Biomed Pharmacother. 2021;133:111037. doi: 10.1016/j.biopha.2020.111037 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref064] 64.Lin C-H, Chen Y-J, Lin M-W, Chang H-J, Yang X-R, Lin C-S. ACE2 and a Traditional Chinese Medicine Formula NRICM101 Could Alleviate the Inflammation and Pathogenic Process of Acute Lung Injury. Medicina (Kaunas). 2023;59(9):1554. doi: 10.3390/medicina59091554 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref065] 65.Huynh DTN, Nguyen HT, Hsieh C-M. Taiwan Chingguan Yihau may improve post-COVID-19 respiratory complications through PI3K/AKT, HIF-1, and TNF signaling pathways revealed by network pharmacology analysis. Mol Divers. 2025;29(3):2305–21. doi: 10.1007/s11030-024-10993-8 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref066] 66.Wei W-C, Tsai K-C, Liaw C-C, Chiou C-T, Tseng Y-H, Liao G-Y, et al. NRICM101 ameliorates SARS-CoV-2-S1-induced pulmonary injury in K18-hACE2 mice model. Front Pharmacol. 2023;14:1125414. doi: 10.3389/fphar.2023.1125414 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref067] 67.Lladós G, Massanella M, Coll-Fernández R, Rodríguez R, Hernández E, Lucente G, et al. Vagus nerve dysfunction in the post-COVID-19 condition: a pilot cross-sectional study. Clin Microbiol Infect. 2024;30(4):515–21. doi: 10.1016/j.cmi.2023.11.007 [DOI] [PubMed] [Google Scholar]

[pone.0351553.ref068] 68.Andrea Ban YL, Siti Istiana AS, Nik Nuratiqah NA, Ng BH, Rose Azzlinda O, Hasni J, et al. The benefits of early pulmonary rehabilitation with incentive spirometer among chronic obstructive pulmonary disease patients with exacerbation of chronic obstructive pulmonary disease. Med J Malaysia. 2024;79(5):561–8. [PubMed] [Google Scholar]

[pone.0351553.ref069] 69.Vallier J-M, Simon C, Bronstein A, Dumont M, Jobic A, Paleiron N, et al. Randomized controlled trial of home-based vs. hospital-based pulmonary rehabilitation in post COVID-19 patients. Eur J Phys Rehabil Med. 2023;59(1):103–10. doi: 10.23736/S1973-9087.22.07702-4 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref070] 70.Eksombatchai D, Wongsinin T, Phongnarudech T, Thammavaranucupt K, Amornputtisathaporn N, Sungkanuparph S. Pulmonary function and six-minute-walk test in patients after recovery from COVID-19: A prospective cohort study. PLoS One. 2021;16(9):e0257040. doi: 10.1371/journal.pone.0257040 [DOI] [PMC free article] [PubMed] [Google Scholar]

[pone.0351553.ref071] 71.de Bruijn S, van Hoek AJ, Mutubuki EN, Knoop H, Slootweg J, Tulen AD, et al. Lower prevalence of post-Covid-19 Condition following Omicron SARS-CoV-2 infection. Heliyon. 2024;10(7):e28941. doi: 10.1016/j.heliyon.2024.e28941 [DOI] [PMC free article] [PubMed] [Google Scholar]

PERMALINK

Effects of incentive spirometer training on dyspnea and functional status in patients with long COVID

Yao-Hsiang Chen

Chia-Huei Lin

Ju-Han Liu

Hsin-An Lin

Yu-Shan Hsieh

Roles

Abstract

Background

Objective

Methods

Results

Conclusion

Clinical trial number

Introduction

Methods

Study design

Fig 1. Flow diagram of participant enrollment and allocation.

Fig 2. Study framework.

Participants and group allocation criteria

Fig 3. Participant recruitment flowchart.

Quality control

Statistical analysis and sample size calculation

Research instruments and measurements

Results

Participant demographics and baseline characteristics

Table 1. Respiratory-related long COVID symptoms by group.

Table 2. Baseline demographic and clinical information of participants.

Within-group analysis

Table 3. Results of within and between analysis compare of Groups.

Between-group comparisons

Generalized Estimating Equations (GEEs) Analysis of Group-by-Time Interactions

Table 4. Interaction effects between group and time.

Item-level analysis of D-12 responses across groups

Discussion

Conclusion

Supporting information

Data Availability

Funding Statement

References

Decision Letter 0

Toufic Ahmad Chaaban

Roles

Author response to Decision Letter 1

Decision Letter 1

Toufic Ahmad Chaaban

Roles

Author response to Decision Letter 2

Decision Letter 2

Toufic Ahmad Chaaban

Roles

Author response to Decision Letter 3

Decision Letter 3

Toufic Ahmad Chaaban

Roles

Author response to Decision Letter 4

Decision Letter 4

Toufic Ahmad Chaaban

Roles

Acceptance letter

Toufic Ahmad Chaaban

Roles

Associated Data

Supplementary Materials

Data Availability Statement

ACTIONS

PERMALINK

RESOURCES

Similar articles

Cited by other articles

Links to NCBI Databases