In the shadow of COVID-19: changes in the seroprevalence of pediatric Mycoplasma pneumoniae

Abstract

Background

Mycoplasma pneumoniae (MP) is a leading cause of respiratory tract infections in children. The coronavirus disease 2019 (COVID-19) pandemic has significantly heightened global attention to respiratory infections, including MP. This retrospective study analyzed the MP seroprevalence among children with acute respiratory tract infections (ARTIs) in Nantong, China, across different phases of the COVID-19 pandemic to assess the changes in the epidemiological landscape of MP in children.

Methods

This study retrospectively analyzed MP-IgM antibody test data from 56,411 children aged 0–14 years with ARTIs at the Affiliated Maternity and Child Health Care Hospital of Nantong University between April 2018 and September 2024. The study period was divided into three phases: pre-pandemic (April 2018–January 2020), pandemic (January 2020–January 2023), and post-pandemic (January 2023–September 2024). Chemiluminescent immunoassay (CLIA) was used to detect serum MP-IgM antibodies, and comparisons were made across the three phases regarding the sex distribution, age distribution, positivity rates, antibody titers, and temporal distribution patterns.

Results

The overall MP-IgM positivity rate was 33.39% (18,833/56,411). Positivity rates exhibited a “V”-shaped fluctuation across the three phases, with rates of 39.70% (6,038/15,209) pre-pandemic, 30.12% (6,553/21,758) during the pandemic, and 32.10% (6,242/19,444) post-pandemic (χ²=391.564, P < 0.001). Significant differences in positivity rate distribution occurred among age groups, with the highest positivity rates in the 3 ~ < 6 years group. In the post-pandemic phase, there was a marked increase in infection rates in the ≥ 6 years group (P < 0.001). There were significant differences in the monthly MP positivity rates every year but 2022 (P < 0.05). Significant fluctuations in MP-IgM antibody titers were observed across phases, with the median titer decreasing from 1.80 (IQR: 1.22–3.18) COI pre-pandemic to 1.65(IQR: 1.18–2.67) COI during the pandemic, and subsequently rebounding to 2.16(IQR: 1.31–4.30) COI post-pandemic (H = 368.022, P < 0.001).

Conclusion

The epidemiological pattern of MP was significantly altered by the COVID-19 pandemic. The rebound in MP infection rates and the marked increase in antibody titers post-pandemic suggest the need to adjust control strategies for pediatric respiratory infections to address shifts in the patterns of MP epidemics. Moreover, the dynamic changes in MP-IgM titers provide novel insights into the evolution of immune responses.

Introduction

In recent years, ARTIs have emerged as one of the leading causes of pediatric outpatient visits. MP is an important pathogen of community-acquired pneumonia (CAP), and is the most common cause in children [1]. Statistics show that MP is responsible for approximately 40% of CAP cases in children over 5 years of age [2, 3]. This high prevalence makes MP a significant public health concern, especially in pediatric populations.

MP is primarily transmitted via respiratory droplets and is particularly likely to cause respiratory infections in confined settings, such as households and schools [4]. This transmission pattern can lead to outbreaks in these settings, especially in preschool and school-aged children. The infection rate of MP varies significantly across different regions, seasons, and pediatric age groups, often demonstrating seasonal trends [5]. Previous studies have shown that MP has an epidemic cycle of 3–7 years, with each outbreak typically lasting 1–2 years [6]. Prior to 2018, MP outbreaks were reported in numerous regions globally [7]. However, since the COVID-19 pandemic emerged in late 2019, the epidemiological patterns of several respiratory infectious diseases, including MP, have been altered, presenting new challenges for epidemiological research [8].

During the COVID-19 pandemic, non-pharmaceutical interventions (NPIs), such as mask-wearing, social distancing, and home quarantine, were implemented globally. These measures not only significantly reduced the transmission of SARS-CoV-2 and curbed the spread of COVID-19, but also likely influenced the transmission dynamics of other respiratory pathogens [9]. In support of this, studies have demonstrated that during the COVID-19 pandemic, the detection rates of other respiratory pathogens, including MP, decreased [10]. However, following the COVID-19 pandemic, as NPIs were relaxed, some regions experienced a resurgence in MP infections [11].

MP infections present with a wide range of clinical manifestations, from mild upper respiratory tract symptoms to severe pneumonia, and can even cause extrapulmonary complications, posing a serious threat to children’s health [12]. IgM antibodies are usually produced 3–7 days after MP infection and peak at 3 weeks of infection when patients start to present clinical symptoms [13, 14]; thus, IgM positivity can serve as a diagnostic indicator of acute-phase infection.

This study retrospectively analyzed MP detection data from the Nantong Maternity and Child Health Hospital from April 2018 to September 2024. To our knowledge, this study is the first to examine the longitudinal changes in MP serum antibody positivity rates and titer values in children with ARTIs across different phases of the COVID-19 pandemic, providing novel insights into the epidemiological shifts of this important respiratory pathogen.

Materials and methods

General data

This single-center retrospective study included 56,411 children with ARTIs who underwent MP-IgM testing at Nantong Maternal and Child Health Hospital between April 2018 and September 2024. All data were extracted from the hospital’s Laboratory Information System (LIS), including age, sex, date of visit, clinical diagnosis, and MP-IgM titer values. A standardized extraction form was used by one researcher and independently verified by a second.

The study was approved by the Ethics Commission of the Affiliated Maternity and Child Health Care Hospital of Nantong University (approval No. Y2025043). As this study was retrospective in nature, the need for individual informed consent was waived.

Inclusion and exclusion criteria

The inclusion criteria were: (1) children aged 0–14 years; (2) who met the diagnostic criteria for ARTIs [15], defined as: ① body temperature > 37.5 °C; and ② at least one of cough, sore throat, rhinorrhoea or expectoration; (3) a whole-blood sample was collected; (4) a diagnosis of MP infection (or no evidence of infection) was based on positive serum MP-IgM (where ≥ 0.9 COI was considered positive).

The exclusion criteria were: (1) infection combined with serious underlying diseases (e.g., congenital immunodeficiency, severe cardiopulmonary disease, etc.); (2) patients treated with immunosuppressants or systemic antibiotics; (3) clinical data deemed incomplete, specifically when records were missing age, sex, or date of visit.

Sample collection and method used to detect MP-IgM

Venous blood (2 mL) or capillary blood (120 µL) was collected from each child and immediately sent to the laboratory for MP-IgM testing. Samples that could not be tested immediately were stored at 4 °C for no more than 48 h.

Quantitative detection of MP-IgM antibodies was performed using chemiluminescent immunoassay (CLIA) platforms (iFlash 3000 series; YHLO Biotech Co., Ltd., China) commencing in April 2018. Sample types were serum and/or plasma; the injection volume was 5 µL; and the reaction temperature was room temperature (20–25 °C). The instrument automatically reports results with a cut-off index (COI) ≥ 0.9 as positive and COI < 0.9 as negative. Calibration was performed strictly according to the manufacturer’s instructions. Identical NMPA-approved reagents (Registration ID: Guoxie Zhun 20173400758) and standardized protocols were utilized throughout the study period.

Daily internal quality control using kit-supplied controls is performed, and instrument calibration is carried out strictly according to the manufacturer’s instructions. Because the National Center for Clinical Laboratories (NCCL) of China has not yet established an external quality assessment (EQA) programme for MP serology, we organise a regional inter-laboratory proficiency exercise every six months. Five serum samples (covering high, medium and low positive levels plus one negative) are tested in parallel with other local laboratories that use the same detection system and are accredited to ISO 15189. Agreement on ≥ 4 samples (≥ 80%) is defined as an acceptable result. Our laboratory has passed every proficiency round, thereby confirming the analytical reliability of our data.

Grouping based on different phases of the pandemic

Based on the timeline of the COVID-19 pandemic and the announcements issued by the National Health Commission of the People’s Republic of China (NHC) [16, 17], the study period was divided into three phases: (1) Pre-pandemic phase: April 1, 2018 to January 20, 2020 (Pre-pandemic group); (2) Pandemic phase: January 21, 2020 to January 7, 2023 (Pandemic group); (3) Post-pandemic phase: January 8, 2023 to September 30, 2024 (Post-pandemic group).

Statistical methods

Data were analyzed using SPSS version 29.0. Categorical data, including sex and positivity rates, were expressed as counts (n) and percentages (%), with intergroup comparisons performed using the chi-squared test (χ²). Non-normally distributed titer values were summarized as medians with interquartile ranges (P25, P75). Pairwise comparisons were conducted using the Bonferroni-adjusted Kruskal-Wallis H test, with the significance level set at P < 0.05.

Results

Sex and age distribution of patients with ARTIs before, during, and after the pandemic

From April 2018 to September 2024, a total of 56,411 children (30,204 males and 26,207 females) with ARTIs were included in the study. These children were divided into three groups: 15,209 from the pre-pandemic period, 21,758 during the pandemic, and 19,444 from the post-pandemic period. No significant sex differences were found in children with ARTIs across the pre-pandemic, pandemic, or post-pandemic periods (P = 0.473). However, there were significant differences in the age distribution of children with ARTIs before, during, and after the pandemic (P < 0.001). Among different age groups, the 3 ~ < 6-year-old group consistently accounted for the highest proportion of ARTIs cases across all pandemic stages (Table 1).

Table 1.

Demographic characteristics of children with ARTIs across pandemic phases

Characteristic	Pre-pandemic (n = 15209)	Pandemic (n = 21758)	Post-pandemic (n = 19444)	χ2	P
Sex				1.496	0.473
Male	8206(53.95)	11,603(53.33)	10,395(53.46)
Female	7003(46.05)	10,155(46.67)	9049(46.54)
Age (Years)				1417.088	< 0.001
0 ~ < 1	1894(12.45)	2086(9.59)	1559(8.02)
1 ~ < 3	3862(25.39)	4638(21.32)	3670(18.87)
3 ~ < 6	7330(48.20)	10,729(49.31)	8491(43.67)
≥ 6	2123(13.96)	4305(19.78)	5724(29.44)

Data are presented as the n (%)

MP-IgM positivity rates before, during, and after the pandemic

The overall positivity rate of MP infection in children was 33.39% (18,833/56,411). MP-IgM seropositivity declined significantly from 39.70% in the pre-pandemic period to 30.12% during the pandemic, followed by a rebound to 32.10% in the post-pandemic period (P < 0.001) (Table 2). This decline and subsequent rebound suggest that the COVID-19 containment measures had a significant impact on the transmission dynamics of MP.

Table 2.

MP-IgM positivity rates in children with ARTIs before, during, and after the COVID-19 pandemic

Total (n = 56411)	Pre-pandemic (n = 15209)	Pandemic (n = 21758)	Post-pandemic (n = 19444)	χ2	༰
18,833(33.39)	6038(39.70)	6553(30.12)	6242(32.10)	391.564	< 0.001

Data are presented as the n (%)

Sex and age distribution of MP-positive children during different phases of the pandemic

A total of 18,833 children (8,793 males and 10,040 females) tested positive for MP during the study period. Significant differences in the sex and age distribution were observed among the pre-pandemic, pandemic, and post-pandemic groups (P < 0.001). The proportion of female children among MP-positive patients was significantly higher than that of male children throughout all pandemic phases. For children aged 0 ~ < 1 years, 1 ~ < 3 years, and 3 ~ < 6 years, the MP-IgM seropositivity rate decreased from the pre-pandemic to the post-pandemic period. In contrast, for children aged ≥ 6 years, the proportion of children exhibiting MP-IgM positivity consistently increased from the pre-pandemic to the post-pandemic period (Table 3).

Table 3.

Characteristics of MP-positive children during different phases of the COVID-19 pandemic

Characteristic	Pre-pandemic (n = 6038)	Pandemic (n = 6553)	Post-pandemic (n = 6242)	χ2	P
Sex				15.299	< 0.001
Male	2883(47.75)	2932(44.74)	2978(47.71)
Female	3155(52.25)	3621(55.26)	3264(52.29)
Age (Years)				1417.088	< 0.001
0 ~ < 1	453(7.50)	371(5.66)	305(4.89)
1 ~ < 3	1701(28.17)	1704(26.00)	1265(20.27)
3 ~ < 6	3013(49.90)	3159(48.21)	2570(41.17)
≥ 6	871(14.43)	1319(20.13)	2102(33.68)

Data are presented as the n (%)

Distribution of MP positivity rates across different years and months from 2018 to 2024

Significant differences in the MP positivity rates were observed across different years and months from 2018 to 2024 (P < 0.05). The MP positivity rates varied significantly across different years from 2018 to 2024 (P < 0.001). Significant differences in monthly positivity rates were observed for all years except 2022 (P < 0.05); month-to-month variation in 2022 was not significant (P = 0.073). Specifically, higher positivity rates were observed in April (51.94%) and October (53.40%) of 2018; August to December (41.01%-43.08%) of 2019; January to September (34.57%-71.91%) of 2020; March to April (30.75%-30.92%) and August to September (31.01%-32.41%) of 2021; December (33.13%) of 2022 (not significantly higher than other months); October to December (32.16%-36.87%) of 2023; and April (42.50%) and August (47.24%) of 2024 (Table 4; Fig. 1).

Table 4.

Distribution of MP positivity rates across different months from 2018 to 2024

Fig. 1.

Monthly distribution of MP-IgM positivity (April 2018-September 2024)

Differences in MP-Positive sample titers across different phases of the pandemic from 2018 to 2024

We compared the titer values of MP-IgM-positive samples across the three pandemic phases. As detailed in Table 5, titers differed significantly among the pre-pandemic, pandemic and post-pandemic periods (H = 368.022, P < 0.001); pairwise comparisons with Bonferroni correction showed statistically significant between-phase differences (P < 0.001).

Table 5.

Comparison of median MP-IgM titers in positive samples across pandemic phases (COI; median [IQR])

Group	Median (P25, P75)	Kruskal-Wallis H test
		H	P
Pre-pandemic	1.80(1.22, 3.18)	368.022	< 0.001
Pandemic	1.65(1.18, 2.67)
Post-pandemic	2.16(1.31, 4.30)

Discussion

This study analyzed the detection of MP IgM antibodies in children with ARTIs admitted to the Affiliated Maternity and Child Health Care Hospital of Nantong University from 2018 to 2024, helping to elucidate the impact of the COVID-19 pandemic on MP infections in children. The results showed that the MP positivity rate initially decreased during the COVID-19 pandemic and then increased post-pandemic. The age distribution of MP infections, seasonal epidemiological characteristics, and titer values of positive samples changed significantly over the different periods. Our data suggest that the NPIs implemented during the COVID-19 pandemic significantly curtailed the transmission of MP. These measures, including mask-wearing, social distancing, home quarantine, and school closures, etc., effectively curtailed the spread of SARS-CoV-2 and likely altered the transmission dynamics of other respiratory pathogens [9]. Our study found that the MP positivity rate decreased from 39.70% pre-pandemic to 30.12% during the COVID-19 pandemic, representing a nearly 10% decline, aligning with findings from other regions [7, 10, 18, 19]. However, following the relaxation of NPIs post-pandemic, the MP infection rate rebounded, likely due to increased exposure and potentially due to fluctuations in population immunity to MP [11].

Our results indicated that the proportion of MP infections was highest in the 3 ~ < 6-year-old group across all pandemic phases, suggesting that preschool children are particularly susceptible to MP, consistent with the findings reported by Zhang et al. [10]. MP infections are primarily transmitted via droplets and have the highest transmission rates in closed or semi-closed settings, such as schools and institutions [20, 21]. Compared to infants aged 0 ~ < 1 year and toddlers aged 1 ~ < 3 years, children aged 3 ~ < 6 years spend more time in kindergartens or daycare centers, increasing their opportunities for contact with peers. Moreover, compared to school-aged children (≥ 6 years), this age group has poorer compliance with hand hygiene and mask-wearing, making them more susceptible to MP infections.

Notably, our study found that the proportion of MP infections in the ≥ 6 years group increased significantly post-pandemic compared to pre-pandemic levels, possibly related to the “immune theft” hypothesis [22]. Proponents of this hypothesis suggest that SARS-CoV-2 may impair the human immune system through various mechanisms, potentially leading to an increased incidence of other respiratory infections post-pandemic [23–25]. However, further research is needed to elucidate the mechanisms underlying these findings.

It is also possible that these children were simply not exposed to MP during the 3-6-year-old period due to the pandemic, and thus were not already immune when they were exposed at an older age [19, 22, 26]. Pre-pandemic (April 1, 2018 to January 20, 2020), the MP positivity rate remained relatively stable, exhibiting seasonal peaks during autumn and winter. During the pandemic (January 21, 2020 to January 7, 2023 [16, 17]), strict pandemic prevention and control measures likely suppressed the spread of most respiratory pathogens [27], leading to an absence of significant MP epidemic peaks. The notable rise in MP positivity rates from April to September 2020 may be linked to the stringent home-quarantine measures in place during that period, wherein only patients with severe symptoms sought medical attention, leading to a higher proportion of positive results. In the post-pandemic period (January 8, 2023 to September 30, 2024), the gradual lifting of control measures saw the MP positivity rate rebound and sustain an epidemic peak from August 2023 (30.23% positivity rate). This may have been due to the decreased population immunity after the pandemic, increasing susceptibility to MP, and is also likely closely related to the resumption of school and social activities to pre-pandemic levels, which facilitated the re-emergence of MP transmission (Table 4; Fig. 1). However, the post-pandemic data in this study collection ended in September 2024, and long-term trends require further monitoring.

Our study employed a chemiluminescent immunoassay (CLIA) for the serological diagnosis of MP infections [28, 29]. Compared with enzyme-linked immunosorbent assay (ELISA) and particle agglutination assay (PA), CLIA demonstrates significant advantages. First, CLIA provides greater sensitivity and specificity [30]. This allows for the early identification of low-concentration MP-IgM antibodies in acute infection samples, thereby decreasing the likelihood of false-negative results [29, 30]. Second, CLIA employs an automated platform that utilizes magnetic bead solid-phase carriers and a chemiluminescent signal amplification system, standardizing the detection process. This approach reduces the within-batch coefficient of variation (CV) and minimizes analytical deviations that may arise from manual operations. In terms of quantitative analysis, CLIA’s ability to quantitatively detect IgM antibody titers offers a more reliable foundation for efficacy evaluation and disease monitoring compared to the semi-quantitative nature of ELISA and PA [30, 31].

Our study is the first to reveal the dynamic changes and significant (P < 0.001) differences in the MP-IgM titer across different phases of the COVID-19 pandemic (Table 5). Specifically, the MP-IgM titer values decreased significantly during the pandemic but rebounded sharply in the post-pandemic period. This finding suggests that the protective effects of NPIs during the pandemic may have reduced the population’s natural immunity to MP, leading to higher pathogen loads during infections in the post-pandemic period. Given the near-universal community exposure to SARS-CoV-2, it is also possible that these infections heightened host susceptibility, as has been suggested by several groups showing reduced immunity after COVID-19 infection [32–34]. Moreover, the generation of antibodies is a complex process, and differences may also be related to infection with different strains of bacteria (including MP) circulating at different times [35–38]. Further basic research is needed to elucidate the specific mechanisms underlying the apparently stronger infections with MP post-pandemic, and whether the findings also apply to other organisms.

The limitations of this study include its retrospective nature, which may introduce selection bias because data were extracted from past records, and its single-center design, which may limit the external validity of the conclusions. To mitigate the risk of incomplete data, we applied strict exclusion criteria and double-verified all extracted records. These findings need to be further validated through multicenter studies in diverse populations with larger sample sizes. IgM antibody positivity only indicates recent infection and cannot distinguish the current state of infection. Moreover, reliance on IgM serology alone may lack the specificity of PCR-based detection methods. Future studies incorporating molecular methods could enhance diagnostic accuracy.

Conclusion

In conclusion, this study provides valuable insights into the impact of the COVID-19 pandemic on MP infections in children. There were significant increases in both the positivity rate and IgM titer, suggesting both more frequent and more severe infections with MP, especially in older children. Future research could further explore the long-term trends in MP infection rates after the pandemic and the potential impacts of these changes on children’s health.

Abbreviations

MP

Mycoplasma pneumoniae

ARTIs

Acute respiratory tract infections

COVID-19

Coronavirus disease 2019

CAP

Community-acquired pneumonia

NPIs

Non-pharmacological interventions

SARS-CoV-2

Severe acute respiratory syndrome coronavirus 2

CLIA

Chemiluminescent immunoassay

Author contributions

HG and QW collected and analyzed the data. HG drafted the manuscript. ER and XW revised the manuscript. XW and XB conducted data analysis. RH and HL designed and supervised the project. All authors read and approved the final manuscript.

Funding

This study was supported by the Scientific Research Project of Nantong Municipal Health Commission (No. QN2025043); the Nantong University Clinical Medicine Specialized Research Fund Project (No. 2024JQ041) and the National Natural Science Foundation of China (No. 82202063).

Data availability

The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.

Declarations

Ethics approval and consent to participate

The procedures followed in this study were in accordance with principles of the Declaration of Helsinki (1964, amended most recently in 2008) of the World Medical Association. The protocol was approved by the Ethics Commission of Affiliated Maternity and Child Health Care Hospital of Nantong University (approval No. Y2025043), who also waived the written informed consent due to the retrospective design of the study.

Consent for publication

Not applicable. Patient consent was waived due to the retrospective nature of the study and the use of anonymous clinical data.

Competing interests

The authors declare no competing interests.