Novel Chewable Pleuran-Based Supplement Decreases Respiratory Tract Infections in Children: A Randomised Controlled Trial

Abstract

Introduction

Recurrent respiratory tract infections (RRTIs) are common in childhood and impose substantial socioeconomic burden. β-Glucans, particularly pleuran from Pleurotus ostreatus, demonstrate immunomodulatory properties through the pathogen-associated molecular pattern receptor interactions. This study evaluated a novel chewable pleuran-based supplement with vitamin D and zinc for preventing respiratory infections in children with RRTIs.

Methods

This international, multicentre, randomised, double-blind, placebo-controlled trial enrolled 249 children with RRTIs from Slovakia, Czech Republic, and Serbia. Participants received either pleuran-based supplement (IMG® with vitamin D and zinc) or active placebo (vitamin D and zinc) for 3 months during respiratory infection season (October–March). Primary endpoint was total respiratory tract infections (RTIs). Secondary endpoints included RTI subtypes, RTI duration, missed school days, and safety evaluations.

Results

In total, 217 children completed the study (104 active, 113 placebo). The active group experienced significantly fewer total RTIs versus placebo (2.35 ± 1.25 vs. 2.77 ± 1.78; P = 0.042), representing 15.2% reduction over 3 months. Common cold episodes were reduced by 18.6% (1.53 ± 1.22 vs. 1.88 ± 1.25; P = 0.040). Effects were pronounced in children over 6 years (P = 0.026 for total RTIs; P = 0.005 for common cold) and evident after the first month (P = 0.037). Tonsillopharyngitis showed significant reductions in frequency (P = 0.003) and duration (P = 0.009). Post hoc analysis of children enrolled at respiratory season onset confirmed significant reductions in common cold frequency (P = 0.004) and duration (P = 0.023). The supplement demonstrated excellent tolerability with 98.7% compliance and only mild adverse events.

Conclusion

Chewable pleuran-based supplement with vitamin D and zinc significantly reduced respiratory tract infections in children with RRTIs, with rapid onset and favourable safety profile, demonstrating therapeutic potential in this vulnerable population.

Trial Registration

ClinicalTrials.gov ID NCT06974747.

Key Summary Points

Why carry out this study?

Recurrent respiratory tract infections (RRTIs) are very common in childhood, associated with mortality and morbidity, and impose substantial socioeconomic burden.

While β-glucans like pleuran have shown promise as natural immunomodulators in previous studies, there was a need to evaluate a newly developed chewable formulation combining pleuran with vitamin D and zinc specifically designed for paediatric use.

The study aimed to assess whether a chewable pleuran-based supplement (containing pleuran, vitamin D, and zinc) could reduce the incidence of respiratory infections in children with RRTIs during a 3-month administration period compared to an active placebo (vitamin D and zinc only).

What was learned from the study?

Children receiving the pleuran-based supplement had significantly fewer total respiratory tract infection episodes compared to the active placebo group.

The study demonstrated that pleuran provides additional protective benefits beyond vitamin D and zinc alone, with particularly pronounced effects in children aged 6 years and older and for common cold episodes.

This first-of-its-kind study establishes the chewable pleuran formulation as a promising therapeutic option for children with RRTIs, with excellent safety and tolerability profiles.

Introduction

Recurrent respiratory tract infections (RRTIs) are very common in childhood, may be associated with both mortality and morbidity, and impose a substantial socioeconomic burden [1]. The definition of RRTIs depends on paediatric age [2]. According to intersociety consensus, six or more respiratory tract infections (RTIs) per year are required to define RRTIs in children under 3 years of age, five or more in those aged 3 to 6 years, and three or more in those over 6 years of age [2]. Most RRTIs occurring in children are of viral origin and therefore bacterial vaccines and antibiotics have neither preventive nor curative effects [3]. Several consensus reports advocate the use of immunomodulators to boost natural defence mechanisms against RTIs and to support the maturation of the immune system [4, 5]. β-Glucans—well-known biologically active polysaccharides—are among the most widely studied natural immunomodulators, with a range of potential indications and clinical applications across different age groups. They have been investigated as therapeutic agents for the prevention of RTIs [4]. In addition to their beneficial interactions with the immune system, their favourable safety profile is another notable advantage. Pleuran (a β-glucan isolated from the oyster mushroom Pleurotus ostreatus) has shown both preventive and acute treatment effects on RTIs in several clinical trials and observational studies in both children and adults, and in both healthy individuals as well as in high-risk populations [6–11]. A recent study of children with RRTIs demonstrated beneficial effects of short-term administration of a pleuran-based chewable supplement containing vitamin D and zinc, notably increasing salivary secretory IgA levels and supporting mucosal immunity [12].

New solid formulations of pleuran-based products have recently been developed for paediatric use, including orally disintegrating and chewable tablets [13]. Topical (mucosal) application of immunomodulatory agents may act directly within the oral cavity and respiratory tract, promoting protective immunity in the most exposed parts of the body.

The aim of this randomised, double-blind, active placebo-controlled, international study was to assess the preventive potential of a newly developed chewable form of highly purified natural immunomodulator pleuran, in combination with vitamin D and zinc, on the incidence of respiratory infections in children with RRTIs during 3 months of administration.

Methods

Study Population

The study involved children of both sexes aged 3 to 18 years with a documented history of RRTIs. Participants were recruited from hospital paediatric outpatient departments and paediatric private practices in Slovakia (16 settings), the Czech Republic (10 settings), and Serbia (10 settings). Screening took place during August and September 2023, involving physical examination and a thorough review of medical history to confirm eligibility and ensure compliance with the study protocol. Enrolment was based on the definition of RRTIs adapted from De Martino and Ballotti [14]: for younger children (< 6 years), ≥ 6 (upper or lower) RTIs, and for older children (≥ 6 years), ≥ 3 RTIs during the October–March period preceding enrolment.

Exclusion criteria were primary or severe secondary immunodeficiency syndromes, bronchopulmonary dysplasia, cystic fibrosis, chronic kidney or liver diseases, endocrinological or metabolic disorders, chronic diarrhoea, oncological conditions, congenital cardiovascular malformations, malnutrition, and intolerance to any of the study product ingredients. Children who were receiving, or had received, any preventive immunomodulatory therapies (including β-glucans or medicinal mushroom extracts, bacterial lysates, or inosine pranobex) within 14 days prior to enrolment were also excluded. Additionally, children whose parents or legal guardians declined to provide written consent or were unwilling to adhere to the study procedures were not included.

Ethics Statement

The study was conducted in accordance with legal and regulatory requirements and the ethical standards of the 1964 Declaration of Helsinki and its later amendments. The study protocol was reviewed and approved by local ethics committees in accordance with national regulations: the Bratislava Self-Governing Region (8658/2023/HF), the Trencin Self-Governing Region (TSC/2023/07875-2), the Banska Bystrica Self-Governing Region (14230/0DDZ/37796/2023), the Kosice Self-Governing Region (6396/2023/ODDZ-29243), Hospital Agel Ostrava-Vítkovice (EK/107/2023), and the Clinical Hospital Centre Dr Dragisa Mišović-Dedinje, Belgrade (20899/3-2023). Written informed consent was obtained from the participants’ parents or legal guardians prior to any study-related procedures. They were fully informed about the purpose, risks and benefits of the study, understood their voluntary role, and agreed to both participation of their children in the study and to the use and publication of their anonymised data. The study has been registered in ClinicalTrials.gov, ID NCT06974747.

Study Design

This prospective, multicentre, randomised, double-blind, placebo-controlled study was conducted between September 2023 and March 2024. Children meeting the inclusion criteria were enrolled and randomised 1:1 into either the active or placebo group. Randomisation, allocation concealment, and implementation (patient enrolment and intervention assignment) were carried out independently by separate parties, each responsible for their respective assignment. A qualified third-party administrator used Microsoft Excel and permuted block randomisation with a block size of four to assign children to one of the two treatment groups (two active, two placebo per block), ensuring balance and using a table of random digits to select the allocation sequence for each block. This procedure was concealed from investigators and monitors to prevent selection bias. Both the active treatment and placebo had an identical appearance and taste; thus, it was not possible for investigators or patients to distinguish between them at any time during the study. Study products were packaged identically in labelled, opaque, sealed bottles, distinguishable only by a unique three-digit kit number and were stored at the trial sites in securely locked cabinets with restricted access. Distribution to allocated patients was confined to trained site investigators or delegated qualified staff. Allocation concealment was ensured using opaque, sealed envelopes with pre-assigned kit and randomisation numbers. Study coordinators, investigators, data managers, statisticians and all participants and their legal guardians remained blinded to treatment allocation throughout the study. Randomised participants received the study medication daily for 3 months.

During this period, four onsite study visits were scheduled (V0–V3; Fig. 1). Prior to inclusion (V0), investigators collected demographic and anthropometric data from medical records, recorded vital signs and physical examination results including comorbidities, and the incidence of RTIs during respiratory infection season (October to March) of the previous year. To track new RTI episodes following enrolment, parents were instructed to maintain a diary documenting the incidence and duration of RTI episodes and their subtypes, number of emergency department and physician visits, absence from nursery/school, use of antibiotics, adherence to study medication and any adverse events (AEs). Parents were also instructed to contact and/or visit their investigator upon occurrence of any acute respiratory symptoms for a physician’s objective physical examination and potentially diagnostic tests to confirm the diagnosis. Onsite follow-up visits were scheduled at 1 (V1), 2 (V2), and 3 months (V3) after enrolment.

Fig. 1.

Study design. AEs, adverse events; RTIs, respiratory tract infections; V0, initial visit; V1, visit at 1 month post-enrollment; V2, visit at 2 months post-enrollment; V3, visit at 3 months post-enrollment

The primary endpoint of the study was to assess the efficacy of the pleuran-based supplement (Imunoglukan P4H® chewable tablets containing 50 mg of IMG®, 5 mg of zinc, and 10 μg of vitamin D₃ per tablet) compared with an active placebo (zinc 5 mg and vitamin D₃ 10 μg per tablet) in reducing the total number of new RTI episodes in children with RRTIs over a 3-month period.

The secondary endpoints included the total duration of RTIs, the number and duration of RTI subtypes (common cold, otitis media, influenza, COVID-19, tonsillopharyngitis, bronchitis, pneumonia), and the evaluation of product tolerability and the occurrence of AEs. RTI subtypes were classified on the basis of clinical signs and/or symptoms and confirmed by the treating paediatrician. Parents were additionally asked to report the child’s absence from daycare or school, the number of parental working days lost as a result of child illness, emergency department visits, and the use of antibiotics. Compliance with the study procedures and medication intake was verified by investigators through the review of parental diaries and inspection of returned medication bottles. Adherence to study medication was assessed by counting the remaining pills in a patient’s medication container during each study visit and at the study end. Children who took less than 75% of the prescribed study medication were excluded from the analysis. AEs were documented both in the patient’s medical records and in the AE form, including the date of onset, type of event, severity (mild, moderate, or severe), causality (possible, unlikely, unrelated, or cannot be evaluated), actions taken (if any: interventions by medications, permanent or temporary discontinuation, or other), and outcome (resolved, resolved without consequences, ongoing, or death).

Study Medication

The main active substance, IMG®, of the study medication—previously identified and chemically characterised by Karacsonyi and Kuniak [15]—contains biologically active polysaccharides consisting of beta-1,3/1,6-d-glucan pleuran, isolated from the oyster mushroom Pleurotus ostreatus using a previously patented technology. In this study, the active product was administered in a novel galenic formulation of chewable tablets, designed to facilitate direct interaction of the active ingredients with the mucosa-associated lymphatic tissue within the oral cavity.

The dosing regimen was based on body weight: children weighing up to 25 kg received one tablet of the study medication once daily, while children over 25 kg received two tablets once daily, between meals, in the morning, for 3 months.

Statistical Analysis

To detect a clinically meaningful reduction of at least 15% in the number of RTI episodes between the active treatment and placebo over a 3-month period, a sample size calculation was conducted prior to study initiation. The calculation assumed a two-sided significance level of 0.05 and a statistical power of 80%. The standard deviation used in the calculation was based on data reported in a previous study investigating a similar immunomodulatory intervention in a paediatric population [7]. On the basis of these parameters, the minimum required sample size was estimated to be approximately 100 children per group. To account for potential dropouts during the study, the planned enrolment was increased by 20%, resulting in a target of approximately 120 participants per group. This approach ensured adequate power to detect statistically significant differences in RTI incidence attributable to the immunomodulatory treatment.

Quantitative data are presented using frequency tables displaying absolute and relative numbers including percentages and were analysed using chi-square tests. Continuous data are shown as mean values with standard deviation and were analysed using a two-sided Student’s t test if normally distributed, or a two-sided Wilcoxon test if non-normally distributed. Multivariate analysis was performed to evaluate the impact of risk factors for RRTIs within the study population. All analyses were two-tailed, and P values of < 0.05 were considered statistically significant. Statistical analyses were performed using IBM SPSS Statistics for Windows, version 20 (IBM Corp., Armonk, NY, USA).

Results

In total, 249 children were recruited and randomised into the study. Thirty-two of these 249 patients were excluded from data analysis (17 from the active group and 15 from the placebo group). In most cases (19 of 32; 59.4%), data inconsistency was the reason for withdrawal from both the study and the analysis. In the remaining cases, participation was discontinued prematurely because of intolerance to the taste of the study medication (n = 5; 15.6%), non-compliance with the study protocol (n = 3; 9.4%), or voluntary withdrawal by the parents (n = 4; 12.5%). One patient withdrew early because of a non-serious AE (nausea; 3.1%) following administration of the active treatment. A total of 217 patients (92 from Slovakia, 58 from the Czech Republic, and 67 from Serbia) completed the study as per protocol and were included into the analysis. Of these, 104 were assigned to the active group and 113 to the placebo group, as shown in Fig. 2. Compliance with the study treatment was high and comparable between the active and placebo groups (98.7% ± 3.5% vs. 97.9% ± 4.3%, respectively).

Fig. 2.

Patient recruitment and enrolment

Table 1 presents the demographic and clinical characteristics of the participants included in the per protocol analysis. The active and placebo groups were comparable in terms of country of residence, sex (male: n = 61; 58.7% vs. n = 53; 46.9%; P = 0.083), nursery or school attendance (n = 104; 100.0% vs. n = 113; 100.0%; not statistically significant), number of siblings (≥ 1 sibling: n = 81; 77.9% vs. n = 93; 82.3%; P = 0.415), passive smoking (n = 22; 21.2% vs. n = 20; 17.7%; P = 0.520), and prevalence of comorbidities (n = 52; 50.0% vs. n = 49; 43.4%; P = 0.327). Allergy was the most common comorbidity, reported in 35.6% of patients in the active group and 31.0% in the placebo group (P = 0.472). The mean number of RTIs reported during the respiratory infection season (October–March) prior to study enrolment was similar between the active and placebo groups (5.73 ± 1.60 vs. 5.49 ± 1.52; P = 0.250), as were subgroup values by age: children aged 3–5.99 years (6.62 ± 0.88 vs. 6.61 ± 0.88; P = 0.945) and children ≥ 6 years (4.61 ± 1.60 vs. 4.72 ± 1.38; P = 0.703).

Table 1.

Baseline demographic and clinical characteristics of children included in the per protocol analysis (N = 217) and in the post hoc analysis (N = 91)

	Per protocol analysis			Post hoc analysis
	Active group (N = 104)	Placebo group (N = 113)	P value	Active group (N = 44)	Placebo group (N = 47)	P value
Country [N; %]
Slovakia	45; 43.3	47; 41.6	0.945	27; 61.4	26; 55.3	0.700
Czech Republic	28; 26.9	30; 26.5		15; 34.1	17; 36.2
Serbia	31; 29.8	36; 31.9		2; 4.5	4; 8.5
Gender [N; %]
Male	61; 58.7	53; 46.9	0.083	27; 61.4	21; 44.7	0.111
Female	43; 41.3	60; 53.1		17; 38.6	26; 55.3
Age [N; %]
3–5.99	58; 55.8	46; 40.7	0.027*	19; 43.2	14; 29.8	0.184
≥ 6 years	46; 44.2	67; 59.3		25; 56.8	33; 70.2
Age [years ± SD]	5.97 ± 2.90	6.92 ± 3.29	0.025*	6.46 ± 2.37	7.48 ± 3.13	0.014*
3–5.99 years	4.17 ± 0.82	3.96 ± 0.82	0.184	5.72 ± 2.45	5.74 ± 2.75	0.090
≥ 6 years	8.24 ± 2.99	8.96 ± 2.76	0.193	7.47 ± 2.25	8.54 ± 2.78	0.017*
Body weight [mean ± SD]	23.82 ± 11.20	28.33 ± 13.73	0.009*	24.73 ± 8.22	30.08 ± 13.54	0.006*
< 25 kg	18.60 ± 3.00	18.96 ± 3.16	0.488	22.16 ± 6.70	22.70 ± 7.74	0.391
≥ 25 kg	38.33 ± 12.84	39.33 ± 13.20	0.748	30.95 ± 8.29	35.65 ± 13.27	0.177
Body weight/dosing [N; %]
< 25 kg (1 tablet/day)	77; 74.0	61; 54.0	0.002*	31; 70.5	20; 42.6	0.007*
≥ 25 kg (2 tablets/day)	27; 26.0	52; 46.0		13; 29.5	27; 57.4
Attending nursery/school [N; %]	104; 100.0	113; 100.0	–	44; 100.0	47; 100.0	-
Number of siblings [N; %]
0	23; 22.1	20; 17.7	0.415	6; 13.6	8; 17.0	0.655
1+	81; 77.9	93; 82.3		38; 86.4	39; 83.0
Passive smoking from parents [N; %]
Yes (at least one parent)	22; 21.2	20; 17.7	0.520	11; 25.0	9; 19.1	0.501
Comorbidities [N; %]
Yes	52; 50.0	49; 43.4	0.327	20; 45.5	26; 55.3	0.347
Allergy	37; 35.6	35; 31.0	0.472	12; 27.3	22; 46.8	0.054
Allergic rhinitis	21; 20.2	19; 16.8	0.521	6; 13.6	12; 25.5	0.155
Asthma	10; 9.6	13; 11.5	0.652	5; 11.4	9; 19.1	0.304
Atopic dermatitis	11; 10.6	8; 7.1	0.363	2; 4.5	5; 10.6	0.276
Number of RTIs between October and March prior enrolment [mean ± SD]
Whole study population	5.73 ± 1.60	5.49 ± 1.52	0.250	5.73 ± 1.60	5.49 ± 1.52	0.250
3–5.99 years	6.62 ± 0.88	6.61 ± 0.88	0.945	6.62 ± 0.88	6.61 ± 0.88	0.945
≥ 6 years	4.61 ± 1.60	4.72 ± 1.38	0.703	4.61 ± 1.60	4.72 ± 1.38	0.703

RTIs respiratory tract infections

*Significant p value

As shown in Table 1, after unblinding the study, statistically significant differences were observed between the active and placebo groups with respect to baseline age (5.97 ± 2.90 vs. 6.92 ± 3.29; P = 0.025). There was a significantly higher proportion of younger children (3–5.99 years: n = 58; 55.8% vs. n = 46; 40.7%) and a lower proportion of older children (≥ 6 years: n = 46; 44.2% vs. n = 67; 59.3%; P = 0.027) in the active group compared to the placebo group. This age distribution was reflected in the dosing pattern: a significantly higher percentage of children weighing < 25 kg (and thus receiving one tablet per day) were in the active group (n = 77; 74.0% vs. n = 61; 54.0%), whereas a higher proportion of children weighing ≥ 25 kg (receiving two tablets per day) were in the placebo group (n = 52; 46.0% vs. n = 27; 26.0%; P = 0.002).

Almost half of the study participants (n = 91; 41.9%) were enrolled in October 2023, prior to the onset of the autumn and winter respiratory infection season. Consequently, a post hoc sub-analysis of the study product’s effect in this subgroup was performed (Table 1). The remaining participants were enrolled during November and December 2023. Baseline demographic and clinical characteristics of this early-enrolment subgroup were comparable to those of the overall study population.

Primary Endpoint: Total Number of RTIs

There was a gradual decrease in the number of RTIs in both study groups, including their respective age subgroups, after 3 months of treatment. The total number of RTI episodes was significantly lower among children receiving the pleuran-based supplement compared to those receiving the active placebo (2.35 ± 1.25 vs. 2.77 ± 1.78; P = 0.042), with the most notable difference observed during the first month of treatment (0.97 ± 0.74 vs. 1.22 ± 1.00; P = 0.037). Overall, a 15.2% reduction in total RTIs over 3 months was observed in the active group compared with the placebo group (Fig. 3). A similarly significant decline in RTIs was seen in the ≥ 6 years age subgroup (2.15 ± 1.05 vs. 2.79 ± 1.93; P = 0.026). Among children younger than 6 years, no statistically significant difference was observed between the active and placebo groups (2.50 ± 1.38 vs. 2.74 ± 1.56; P = 0.409).

Fig. 3.

Total number of respiratory tract infections (RTIs) during the study period (*P < 0.05). V1, visit at 1 month post enrolment; V2, visit at 2 months post enrolment; V3, visit at 3 months post enrolment

Secondary Endpoints

Per protocol analysis of each RTI subtype revealed a significant reduction in the number of common cold episodes, with a mean decrease of 18.6% in children who received the pleuran-based supplement compared with those given the active placebo (1.53 ± 1.22 vs. 1.88 ± 1.25; P = 0.040). Subgroup analysis only showed a statistically significant decline in children aged ≥ 6 years (1.39 ± 0.93 vs. 2.03 ± 1.41; P = 0.005) in the active group compared with the placebo group. The incidences of other RTI subtypes (COVID-19, influenza, otitis, tonsillopharyngitis, bronchitis, and pneumonia) were similar across study groups and age subgroups and did not differ significantly (Table 2). After the first month of treatment, we observed a statistically significant reduction in both the mean number (0.08 ± 0.27 vs. 0.22 ± 0.42; P = 0.003) and mean duration (0.5 ± 1.9 vs. 1.4 ± 2.9; P = 0.009) of tonsillopharyngitis in the active group compared with the placebo group.

Table 2.

Number and duration of total respiratory tract infections (RTIs) and RTI subtypes in the per protocol analysis after 3 months of treatment

	Active group (N = 104)	Placebo group (N = 113)	P value	Younger children 3–5.99 years		P value	Older children ≥ 6 years		P value
				Active group (N = 58)	Placebo group (N = 46)		Active group (N = 46)	Placebo group (N = 67)
Number of RTIs (mean ± SD)
All infections	2.35 ± 1.25	2.77 ± 1.78	0.042*	2.50 ± 1.38	2.74 ± 1.56	0.409	2.15 ± 1.05	2.79 ± 1.93	0.026*
Common cold	1.53 ± 1.22	1.88 ± 1.25	0.040*	1.64 ± 1.41	1.65 ± 0.95	0.951	1.39 ± 0.93	2.03 ± 1.41	0.005*
COVID-19	0.03 ± 0.17	0.04 ± 0.21	0.550	–	–	–	0.07 ± 0.25	0.07 ± 0.26	0.850
Influenza	0.13 ± 0.37	0.07 ± 0.26	0.145	0.14 ± 0.40	0.11 ± 0.31	0.683	0.13 ± 0.34	0.04 ± 0.21	0.133
Tonsillopharyngitis	0.28 ± 0.57	0.41 ± 0.70	0.139	0.24 ± 0.54	0.41 ± 0.78	0.206	0.33 ± 0.60	0.40 ± 0.65	0.526
Otitis	0.14 ± 0.38	0.14 ± 0.60	0.969	0.19 ± 0.44	0.20 ± 0.65	0.956	0.09 ± 0.28	0.10 ± 0.55	0.844
Bronchitis	0.19 ± 0.46	0.21 ± 0.56	0.774	0.28 ± 0.56	0.33 ± 0.76	0.698	0.09 ± 0.28	0.13 ± 0.34	0.443
Pneumonia	0.04 ± 0.19	0.02 ± 0.13	0.354	0.02 ± 0.13	0.04 ± 0.21	0.432	0.07 ± 0.25	–	0.083
Duration of RTIs (mean ± SD)
All infections	15.17 ± 10.13	16.60 ± 11.88	0.343	15.70 ± 10.40	17.26 ± 11.63	0.484	14.46 ± 9.83	16.15 ± 12.10	0.433
Common cold	8.88 ± 8.79	10.32 ± 8.57	0.225	9.30 ± 9.70	9.41 ± 6.94	0.933	8.39 ± 7.56	10.94 ± 9.54	0.133
COVID-19	0.17 ± 1.05	0.27 ± 1.30	0.531	–	–	–	0.39 ± 1.56	0.46 ± 1.67	0.819
Influenza	0.97 ± 2.81	0.37 ± 1.49	0.054	0.80 ± 2.30	0.57 ± 1.86	0.561	1.17 ± 3.34	0.24 ± 1.17	0.074
Tonsillopharyngitis	1.88 ± 4.12	2.67 ± 4.74	0.186	1.60 ± 3.50	2.78 ± 5.41	0.192	2.26 ± 4.77	2.60 ± 4.25	0.695
Otitis	0.93 ± 2.47	1.06 ± 4.35	0.790	1.20 ± 2.80	1.59 ± 5.50	0.679	0.54 ± 1.89	0.70 ± 3.34	0.772
Bronchitis	1.81 ± 4.62	1.58 ± 3.77	0.684	2.70 ± 5.70	2.11 ± 4.27	0.546	0.65 ± 2.20	1.21 ± 3.38	0.291
Pneumonia	0.53 ± 2.80	0.33 ± 2.70	0.590	0.10 ± 0.90	0.80 ± 4.21	0.285	1.04 ± 4.04	–	0.087

RTIs respiratory tract infections

*Significant p value

Although the mean duration of total RTIs was shorter in children given the pleuran-based supplement compared to those receiving the active placebo, the difference did not reach statistical significance (15.17 ± 10.13 vs. 16.60 ± 11.88; P = 0.343). Analysis of individual RTI subtypes showed a trend toward shorter duration of common cold (8.88 ± 8.79 vs. 10.32 ± 8.57; P = 0.225) and tonsillopharyngitis (1.88 ± 4.12 vs. 2.67 ± 4.74; P = 0.186) in the active group (Table 2).

The prevalence of allergy, exposure to cigarette smoke, and number of siblings did not influence the number or duration of RTIs.

No significant differences were observed in other secondary parameters (missed school or workdays, number of emergency department visits, or use of antibiotic therapy) between the study groups or within age subgroups at the end of the 3-month study period.

Post Hoc Subgroup Analysis of Patients Enrolled in October 2023

A post hoc subgroup analysis was performed on patients whose initial visit (V0) occurred during October, a month generally considered the beginning of the respiratory infection season. Of the 217 patients eligible for per protocol statistical analysis, 91 (41.9%) were included in this analysis, of whom 44 were assigned to the active group and 47 to the placebo group (Fig. 2). A comparable, gradual decrease in the number of RTIs was observed in both groups over the 3-month study period. However, already after the first month of treatment, the number of total RTIs was significantly lower in the active group compared to the placebo group (0.86 ± 0.73 vs. 1.34 ± 1.07; P = 0.016). This difference remained statistically significant at the end of the treatment period, with fewer RTIs in the active group versus the placebo group (2.14 ± 1.05 vs. 2.85 ± 1.78; P = 0.021). Common cold was the most frequently occurring RTI subtype, and a statistically significant reduction was observed in both its mean number (1.39 ± 1.19 vs. 2.17 ± 1.34; P = 0.004) and mean duration (8.64 ± 9.23 vs. 13.06 ± 9.00; P = 0.023) among children receiving the pleuran-based supplement compared with those given the active placebo. Differences in the number and duration of other RTI subtypes did not reach statistical significance within the observation period (Table 3).

Table 3.

Number and duration of respiratory tract infections (RTIs) and RTI subtypes in patients enrolled in October 2023

	Active group (N = 44)	Placebo group (N = 47)	P value	Active group (N = 44)	Placebo group (N = 47)	P value
	Number of RTIs after 1 month (mean ± SD)			Number of RTIs after 3 months (mean ± SD)
All infections	0.86 ± 0.73	1.34 ± 1.07	0.016*	2.14 ± 1.05	2.85 ± 1.78	0.021*
Common cold	0.61 ± 0.72	1.02 ± 0.82	0.014*	1.39 ± 1.19	2.17 ± 1.34	0.004*
COVID-19	–	–	–	0.07 ± 0.25	0.09 ± 0.28	0.765
Influenza	0.02 ± 0.15	–	0.323	0.09 ± 0.29	0.02 ± 0.15	0.158
Tonsillopharyngitis	0.09 ± 0.29	0.13 ± 0.34	0.580	0.30 ± 0.55	0.19 ± 0.40	0.309
Otitis	0.02 ± 0.15	0.09 ± 0.35	0.269	0.07 ± 0.25	0.19 ± 0.68	0.251
Bronchitis	0.09 ± 0.29	0.09 ± 0.46	0.943	0.16 ± 0.43	0.17 ± 0.64	0.923
Pneumonia	0.02 ± 0.15	0.02 ± 0.15	0.963	0.07 ± 0.25	0.02 ± 0.15	0.289

	Duration of RTIs after 1 month (mean ± SD)		P value	Duration of RTIs after 3 months (mean ± SD)		P value
All infections	6.25 ± 6.30	8.68 ± 7.27	0.093	15.16 ± 9.65	18.04 ± 12.03	0.212
Common cold	3.41 ± 4.49	6.45 ± 5.86	0.007*	8.64 ± 9.23	13.06 ± 9.00	0.023*
COVID-19	–	–	–	0.41 ± 1.59	0.55 ± 1.86	0.693
Influenza	0.30 ± 1.96	–	0.323	0.77 ± 2.60	0.11 ± 0.73	0.108
Tonsillopharyngitis	0.61 ± 2.06	0.94 ± 2.62	0.517	1.98 ± 4.07	1.53 ± 3.38	0.571
Otitis	0.23 ± 1.51	0.72 ± 3.27	0.360	0.50 ± 1.95	1.70 ± 5.92	0.193
Bronchitis	1.27 ± 4.28	0.36 ± 1.76	0.195	1.91 ± 5.20	0.87 ± 2.81	0.246
Pneumonia	0.43 ± 2.86	0.21 ± 1.46	0.644	0.95 ± 3.82	0.21 ± 1.46	0.232

RTIs respiratory tract infections

*Significant p value

Tolerability and Safety

The pleuran-based supplement in chewable tablet form, at the administered doses, was generally well tolerated and considered safe. Five of the 247 enrolled children withdrew prematurely because of intolerance to the taste of the study product—four in the active group and one in the placebo group—during the first month of treatment. Five mild, self-limiting, non-serious AEs were reported by parents to investigators over the course of the study. These included three episodes of nausea, one episode of acute gastritis, and one episode of vomiting. All AEs were assessed as possibly related to the study medications. Only one patient, who reported nausea during the first month following enrolment, chose to withdraw from the study early. The incidence of AEs was low and similar across both groups (n = 1 (0.96%) in the active group and n = 4 (3.5%) in the placebo group; P > 0.05).

Discussion

β-Glucans, a heterogeneous group of biologically active polysaccharides, possess a wide range of complex immunomodulatory properties intricately linked to their structural characteristics, natural sources, and extraction methods [15]. Pleuran, an insoluble β-1,3/1,6-d-glucan isolated from the oyster mushroom Pleurotus ostreatus, exemplifies this complexity, demonstrating multifaceted interactions with both the innate and adaptive immune systems. Its branched molecular structure, stabilised by unique glycosidic bonds, enables recognition by pathogen-associated molecular pattern receptors such as dectin-1, Toll-like receptors (TLRs), and complement receptor 3, initiating downstream immunomodulatory cascades [8, 16]. Innate immune modulation by pleuran is evidenced by its capacity to enhance phagocytic activity of macrophages and neutrophils, prime natural killer cell cytotoxicity, and to support mucosal defences. Clinical trials involving children with RRTIs previously demonstrated significant increases in natural killer cell counts and stabilisation of CD8⁺ T lymphocytes, associated with a reduced respiratory morbidity [8, 17]. Furthermore, recent research highlights pleuran’s potential for topical (mucosal) application in strengthening mucosal immunity, showing a significant increase in salivary IgA levels in children with RRTIs after just 8–10 days of supplementation. This rapid enhancement of the mucosal barrier function underscores pleuran’s protective potential against acute respiratory infections [12].

The immunomodulatory mechanisms of insoluble beta-glucans are orchestrated through selective engagement with immune cells within intestinal Peyer’s patches, operating via local immunological activation without systemic absorption, thereby adhering to established principles of targeted immunomodulation [17]. Notably, the dose–response kinetics for insoluble beta-glucans, including pleuran, demonstrate a non-linear, quality-dependent profile rather than conventional quantity-dependent relationships, suggesting sophisticated receptor-mediated recognition mechanisms [18]. Furthermore, insoluble beta-glucans demonstrate superior immunobiological potency compared to their soluble counterparts, orchestrating comprehensive activation of both innate and adaptive immune cascades through specialised dectin-1 receptor-mediated signalling pathways [19].

This international, randomised, double-blind, placebo-controlled study was intentionally conducted from September to March, which corresponds to the peak respiratory infection season in Central Europe. This period is characterised by higher viral circulation, reduced vitamin D synthesis due to limited sun exposure, and increased exposure of children to indoor environments. Conducting the study during this season, therefore, maximised the likelihood of capturing a clinically meaningful number of endpoints. Importantly, both the active and placebo groups were exposed to the same seasonal conditions.

This study showed that an innovative formulation of chewable pleuran-based supplement enriched with vitamin D and zinc significantly reduced respiratory morbidity in children with RRTIs during the respiratory infection season, with a clinically meaningful effect already present in the first month of intervention. This effect was particularly pronounced in relation to common cold episodes and in the subgroup of children aged 6 years and older. While the overall duration of RTIs showed a favourable trend, a statistically significant decrease was observed in both the frequency (P = 0.003) and duration (P = 0.009) of tonsillopharyngitis during the initial month of supplementation, likely reflecting the benefit of oral administration. Furthermore, a post hoc analysis focusing on children enrolled at the start of the typical respiratory infection season reinforced these findings, showing statistically significant reductions in both the number (P = 0.004) and duration (P = 0.023) of common cold episodes over the 3-month period. Importantly, the supplement demonstrated a favourable safety and tolerability profile throughout the study. This was reflected by high treatment compliance and a low incidence of mild, self-limiting, non-serious AEs, primarily minor gastrointestinal complaints such as nausea, gastritis, or vomiting, which occurred at similar, statistically insignificant rates in both study groups. Comparable results have been reported in previous studies conducted in recent years [5–10].

Vitamin D plays a crucial role in modulating and regulating multiple immune responses, particularly within the respiratory tract. In the context of mucosal immunity, it enhances host defences by stimulating the synthesis of key antimicrobial peptides, including defensins and cathelicidin, thereby strengthening intrinsic mechanisms against pathogenic microorganisms [20]. The high prevalence of vitamin D deficiency among children with RRTIs highlights the potential value of supplementation in this population. Supporting this, clinical evidence suggests that vitamin D supplementation can reduce the risk of RTIs, demonstrating efficacy in lowering both the incidence and severity of infections in individuals with baseline vitamin D deficiency [21, 22]. Alongside vitamin D, the trace element zinc is also fundamental to the proper functioning of both innate and adaptive immunity. It contributes by modulating cytokine production, regulating signalling pathways and enzyme activities, and supporting the activity of various immune cell populations involved in defending against respiratory pathogens. Moreover, evidence indicates that zinc supplementation may exert a protective effect against RTIs, although the effectiveness of such interventions appears to depend on factors such as dosage and baseline zinc status—particularly relevant in paediatric populations [23]. In our study, the active placebo group, which received zinc and vitamin D alone, showed a trend toward gradually reduced respiratory infection rates during the 3-month study period, consistent with the known effects of these nutrients. However, the group receiving the pleuran-based product (containing IMG®, zinc, and vitamin D) demonstrated a significantly greater reduction in respiratory infection rates. This suggests that pleuran contributes additional protective benefits beyond those provided by zinc and vitamin D alone and may serve as a valuable supplement for the prevention of respiratory infections. The combination of pleuran, vitamin D, and zinc may offer synergistic effects among these active compounds, thereby achieving enhanced clinical effectiveness.

Allergic diseases accounted for the most frequently observed comorbidities in our study population, affecting more than 30% of the children. Allergic inflammation is known to impair antiviral host defence, predisposing individuals to increased respiratory morbidity. The T2-dominant inflammatory environment, characteristic of allergy, disrupts key components of the innate immune system. Within the affected tissues, recruited and activated eosinophils suppress the expression of TLR7 and TLR8, thereby dampening the interferon signalling pathway [24, 25]. Plasmacytoid dendritic cells (pDCs), which are crucial for tolerance induction and immune homeostasis through the production of type I interferons in response to viral infections, exhibit a markedly reduced capacity for interferon synthesis within a T2-skewed inflammatory milieu. Elevated IgE levels further contribute to suppressed production of interferons by enabling FcεRI receptor cross-linking on pDCs, which then competitively inhibits the TLR7/8 signalling pathways that are crucial for initiating interferon production in response to viral infections [26]. In children with allergic conditions, multiple pathognomonic mechanisms contribute to a compromised structural and functional integrity of the respiratory epithelium. This may include increased expression of adhesion molecules such as intercellular adhesion molecule 1, which serve as binding receptors for rhinoviruses. Such epithelial dysfunction may further exacerbate susceptibility to viral respiratory infections in this population [27]. Given that pleuran exhibits both anti-allergic and immunomodulatory effects, its use in this population may help mitigate the increased susceptibility to respiratory infections associated with allergic comorbidities [7].

The European Medicines Agency highlights the importance of age-appropriate formulations for paediatric medicines, with particular emphasis on acceptability and safety (Regulation (EC) No. 1901/2006 on medicinal products for paediatric use). Orally dissolvable and chewable tablets are recognised as suitable dosage forms, especially for school-age children and adolescents, because of their ease of administration and palatability [28]. This form of application is particularly relevant as it facilitates direct interaction of the product’s active ingredients with the mucosa-associated lymphatic tissue within the oral cavity. Specifically, local mucosal application of immunomodulators can act directly in the oral cavity and upper respiratory tract, promoting protective immunity in the most exposed regions of the body [29]. Therefore, formulations like the pleuran-based supplement used in this study offer advantages including improved taste masking and ease of swallowing, which may enhance medication adherence and treatment outcomes in children, while simultaneously supporting local mucosal immunity.

Although our clinical study was conducted in Europe, pleuran’s immunomodulatory mechanisms appear not to be region-specific and are therefore likely applicable across populations. Moreover, previous clinical studies using liquid formulations of pleuran in different paediatric populations conducted in different countries and geographic regions have consistently demonstrated comparable protective effects against RRTIs [8, 10, 12, 17]. These findings support the assumption that our results with the novel chewable formulation may also be generalizable. Nevertheless, we recognize that environmental, nutritional, and epidemiological factors may influence outcomes, and future international trials specifically evaluating this new formulation will be valuable to further confirm global applicability.

Among the strengths of this study are its large cohort allowing evaluation of the novel chewable formulation of pleuran in the paediatric population known for their high burden of RRTIs. While this study provides relevant evidence for the efficacy of pleuran-based supplementation in reducing respiratory morbidity, several limitations warrant consideration. The relatively short intervention period of 3 months potentially limits the assessment of sustained benefits, while the combined vitamin D/zinc formulations in both groups make it difficult to isolate pleuran-specific effects. Imbalances in age distribution and reliance on caregiver-reported outcomes—despite methodological safeguards—introduce potential confounding and recall biases. Furthermore, although the number of siblings did not differ between study groups, the study did not account for other potential confounding variables, including household income, socioeconomic status, parental educational level, and healthcare access—factors that have been demonstrated to significantly influence research outcomes and may compromise the validity of the findings. Similarly, no epidemiological data were systematically collected or documented throughout the clinical trial. In addition, other significant methodological limitations of our study are the absence of (more objective) virological diagnosis and the lack of systematic monitoring of infectious, allergic, and immunological biomarkers that could have added more objective insights into treatment efficacy and safety profiles. Future research should include longer trial durations in more balanced and larger age groups and employ factorial designs to disentangle the specific contribution of pleuran from that of co-administered micronutrients. Studies involving more diverse populations, as well as investigations into pleuran’s impact on microbiota modulation, vaccine responsiveness, and cost-effectiveness compared with standard therapeutic options, will be essential to support optimal clinical translation for children with RRTIs.

Conclusions

This study demonstrates, for the first time, the beneficial effects of a novel chewable supplement containing pleuran, vitamin D, and zinc on the incidence of RTIs in children who are frequently affected during the respiratory infection season. Pleuran’s rapid onset of action—combining immunomodulatory and anti-allergic effects—together with its synergistic interaction with micronutrients vitamin D and zinc, positions the new formulation as a promising therapeutic option for children with RRTIs. The supplement was well tolerated and showed a favourable safety profile throughout the 3-month intervention period.

Author Contributions

Milos Jesenak: Conceptualisation, Writing – original draft, Writing – review & editing. Elena Prokopova: Conceptualisation, Investigation, Methodology, Project administration, Supervision, Writing – original draft, Writing – review & editing. Jan Bozensky: Conceptualisation, Investigation, Methodology, Project administration, Supervision, Writing – review & editing. Branka Bonaci-Nikolic: Conceptualisation, Investigation, Methodology, Project administration, Supervision, Writing – review & editing. Katarina Milosevic: Investigation, Project administration, Writing – review & editing. Katarina Stankovic: Investigation, Project administration, Writing – review & editing. Olivera Ostojic: Investigation, Project administration, Writing – review & editing. Zorica Zivkovic: Investigation, Project administration, Supervision, Writing – review & editing. Zuzana Diamant: Writing – critical review & editing. Peter Kunc: Data curation, Writing – review & editing. Kamil Janeczek: Data curation, Writing – review & editing. Juraj Majtan: Data curation, Formal analysis, Writing – original draft, Writing – review & editing.

Funding

Open access funding provided by The Ministry of Education, Science, Research and Sport of the Slovak Republic in cooperation with Centre for Scientific and Technical Information of the Slovak Republic. This research was funded by PLEURAN, s.r.o. The journal’s Rapid Service Fee was funded by PLEURAN, s.r.o.

Data Availability

The raw data will be made available by the authors on reasonable request. For privacy reasons the data are not public.

Declarations

Conflict of Interest

The authors (Milos Jesenak, Elena Prokopova, Jan Bozensky, Branka Bonaci-Nikolic, Katarina Milosevic, Katarina Stankovic, Olivera Ostojic, Zorica Zivkovic, Zuzana Diamant, Peter Kunc, Kamil Janeczek, Juraj Majtan) declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. The funder had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.

Ethical Approval

The study was conducted in accordance with legal and regulatory requirements and the ethical standards of the 1964 Declaration of Helsinki and its later amendments. The study protocol was reviewed and approved by local ethics committees in accordance with national regulations: the Bratislava Self-Governing Region (8658/2023/HF), the Trencin Self-Governing Region (TSC/2023/07875-2), the Banska Bystrica Self-Governing Region (14230/0DDZ/37796/2023), the Kosice Self-Governing Region (6396/2023/ODDZ-29243), Hospital Agel Ostrava-Vítkovice (EK/107/2023), and the Clinical Hospital Centre Dr Dragisa Mišović-Dedinje, Belgrade (20899/3-2023). Written informed consent was obtained from the participants’ parents or legal guardians prior to any study-related procedures. They were fully informed about the purpose, risks and benefits of the study, understood their voluntary role, and agreed to both participation of their children in the study and to the use and publication of their anonymised data. The study has been registered in ClinicalTrials.gov, ID NCT06974747.