Review Reports

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The authors have reported a randomized, double-blind, placebo-controlled clinical trial evaluating the efficacy of Lacticaseibacillus rhamnosus CRL1505 for prevention of upper respiratory tract infections (URTIs) in healthy adults.

The study included 140 randomized participants, with 122 completing sufficient follow-up for analysis. Daily supplementation with the probiotic for 12 weeks was associated with reductions in recurrent URTI episodes, cumulative URTI duration, symptomatic medication use, and increased URTI-free time.

The manuscript addresses an important and clinically relevant topic. The randomized, double-blind, placebo-controlled design is a clear strength, and the study is generally well conducted and clearly written. The findings are likely to be of interest to the readership of Microorganisms and will contribute additional clinical data regarding probiotic approaches for respiratory infection prevention in adults.

Some aspects however may benefit from clarification or a more cautious interpretation before publication.

1. Interpretation of the Primary Endpoint. The primary endpoint was the proportion of participants experiencing at least 1, 2, or 3 URTI episodes. However, significant differences were observed only for the ≥3 episode category at 16 weeks.

Although these findings are encouraging, some portions of the Discussion appear to overstate their clinical significance. The study did not demonstrate statistically significant reductions for:

• ≥1 URTI episodes
• time to first URTI
• symptom severity (WURSS-21)

In view of this perhaps the findings should be expressed more cautiously as reduction in recurrent URTI burden rather than broader prevention of URTI overall.

2.     Attrition and Analysis Population. Although 140 participants were randomized, only 122 were included in the final analysis.

It is stated that an intention-to-treat approach was used, but it appears that participants without follow-up data were excluded from efficacy analyses.

Please clarify:

• whether a modified intention-to-treat analysis was actually performed,
• how missing data were handled,
• and whether sensitivity analyses were conducted.

This point is important because the observed treatment effects are quite modest.

3. Statistical Considerations

The statistical methods appear generally to be appropriate. However, several endpoints reached only borderline statistical significance (e.g., P=0.05, P=0.06, P=0.07). Given the large number of secondary outcomes analyzed, the manuscript would benefit from:

• clarification regarding multiplicity adjustment,
• or some explicit acknowledgement that secondary endpoint analyses were exploratory.

In addition, some variables (e.g., URTI episodes per participant) may not follow normal distributions. Please clarify whether distributional assumptions for t-tests were assessed.

4. Reliance on Self-Reported URTI Outcomes

The authors appropriately acknowledge the limitations of the Jackson scale and symptom-based URTI diagnosis.

However, the manuscript should further emphasize that:

• no virological confirmation was performed,
• diagnoses relied heavily on self-reported symptom questionnaires,
• and common cold definitions remain inherently subjective.

Although this does not invalidate the study, it should moderate any mechanistic and clinical interpretations of the findings.

5. Mechanistic Interpretations

The lack of salivary IgA modulation is interesting. Since no other immune biomarkers other than salivary IgA were assessed, the authors should probably avoid implying specific immunological mechanisms underlying the clinical effects in the discussion.

6 Minor Comments

It would be helpful to provide:

• absolute numbers of total URTI episodes in each group,
• and adherence details for randomized versus analyzed participants.

Please clarify whether adverse events were analyzed statistically between groups.

 

The term “more vulnerable populations” in the Discussion should be defined more precisely.

Some Discussion statements regarding labour productivity and absenteeism are speculative, as these outcomes were not directly measured.

Author Response

Thanks for the comments. Please see the attachment.

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

This manuscript presents a randomized, double-blind, placebo-controlled clinical trial evaluating the efficacy of Lacticaseibacillus rhamnosus CRL1505 in reducing URTI burden in healthy adults. The topic is clinically relevant and the study is generally well designed. The manuscript is clearly written, and the findings may contribute to the growing evidence regarding probiotic-mediated modulation of respiratory health.

However, several points should be addressed before acceptance.

Major Comments

1. Although the study is described as intention-to-treat (ITT), only participants who completed at least two visits were included in the analysis. The authors should clarify how missing data and dropouts were handled statistically and explain whether a true ITT analysis was performed.
2. More details regarding adherence assessment should be provided. The manuscript states that adherence exceeded 95%, but the methodology used to determine compliance is insufficiently described.
3. The diagnosis of URTI relied primarily on symptom-based criteria and the Jackson scale. Since no virological confirmation was performed, the limitations of symptom-based diagnosis should be discussed more critically.
4. The manuscript would benefit from a more detailed explanation of the proposed immunological mechanisms underlying the observed effects of L. rhamnosus CRL1505, especially considering that salivary IgA levels did not significantly differ between groups.
5. Since several authors are affiliated with companies related to the probiotic strain, the role of the sponsor in study design, data analysis, manuscript preparation, and publication decision should be clarified further to avoid concerns regarding potential bias.

Minor Comments

1. Please carefully revise the manuscript for typographical and formatting inconsistencies.
2. Some references require formatting corrections according to journal style.
3. The CONSORT flow diagram resolution could be improved.
4. Please define more clearly whether symptomatic medication use was self-reported or physician-confirmed.
5. The discussion section could better compare the findings with previous adult probiotic URTI trials.

Author Response

Thanks for your comments. Please see the attachment.

Author Response File: Author Response.pdf