Perfluoroalkyl substances (PFAS) have been linked to immunotoxic effects in children, and the consequences of immune suppression could be particularly severe for children in low-income settings. We thus aimed to investigate associations between infant and maternal PFAS exposure and infant vaccine antibody concentrations in Burkina Faso. Infants (n = 200) were vaccinated against measles at baseline (age 4-7 months) and/or at 9 months. Associations between maternal and infant PFAS concentrations (quantified in serum collected at baseline) and infant measles antibody concentrations before and after vaccination were assessed in confounder-adjusted linear regression models. Influential observations were excluded in sensitivity analyses. Infant PFAS exposure was positively associated with pre-vaccination antibody concentrations, but associations were largely driven by a few influential observations. Maternal and infant PFAS concentrations were not consistently associated with infant measles antibody concentrations after one or two vaccinations. Thus, these results from a low-exposed population did not confirm findings from previous studies.

BACKGROUND:It has been suggested that non-live vaccines may increase the risk of non-targeted infections, especially in girls. We examined whether the introduction of non-live human papillomavirus (HPV) vaccine was associated with an increased rate of infectious disease hospitalization among girls in Denmark, Norway, and Sweden. METHODS:Using data from national patient registries, we performed controlled interrupted time series analyses including an intervention group (13-year-old girls in Denmark, 8th-grade girls in Norway, 7th grade girls in Sweden) and a control group not offered HPV vaccine (boys of the same age/grade). Infectious disease hospitalization rates were modelled using negative binomial regression, and the level change, i.e. the incidence rate ratio corresponding to the relative change to the rate's underlying trend following introduction of HPV vaccine, was estimated. RESULTS:For infectious disease hospitalizations of any duration, the level change among girls and boys, respectively, was 1.21 (95% confidence interval (CI) 0.94-1.57) and 0.73 (95% CI 0.56-0.96) in Denmark; 1.21 (95% CI 0.94-1.56) and 1.14 (95% CI 0.88-1.49) in Norway; and 0.83 (95% CI 0.65-1.07) and 0.72 (95% CI 0.56-0.92) in Sweden. For hospitalizations with ≥2 overnight stays, the level change was 1.05 (95% CI 0.71-1.56) and 0.81 (95% CI 0.54-1.20) among Danish girls and boys, respectively, whereas the results in Norway and Sweden were similar to the results for hospitalizations of any duration. CONCLUSION:Changes to the rate following introduction of HPV vaccine were similar among girls and boys in Norway and Sweden, in line with there being no association between HPV vaccination and infectious disease hospitalizations. The difference between Danish girls and boys seems unlikely to be attributable to HPV vaccine, given the Norwegian and Swedish results. Overall, our study does not support that HPV vaccination increases risk of infectious disease hospitalization.

The Bacillus Calmette-Guérin (BCG) vaccine, primarily designed for tuberculosis, exerts non-specific immunological effects that vary considerably among children. Limited knowledge exists regarding genetic variants that can impact immune responses post-vaccination. Understanding the genetic factors influencing the immune response in children following BCG vaccination is crucial for optimizing vaccine strategies. Genome-wide pQTL mapping of 65 circulating inflammatory proteins profiled at Month 13 uncovered 11 independent genome-wide significant loci. Following pQTL mapping in BCG-unvaccinated children, whose mean age was not significantly different from that of BCG-vaccinated children, a P-value lookup complemented with statistical colocalization and interaction analyses identified four pQTLs (rs10217747: CXCL11, rs13187850: CD6, rs56283092: LAP-TGF-beta 1, and rs11708321: CDCP1) showing evidence of genotype-by-BCG interactions and larger effect estimates in the BCG group. These variants have previously been associated with inflammatory and autoimmune traits. Genetic analysis of in vitro cytokine production capacity after BCG vaccination identified nominal associations mainly at Month 13 rather than Day 4 or Month 3. These findings enhance our understanding of immune responses to vaccination in children but, given the exploratory genome-wide approach, should be considered hypothesis-generating and require further validation.

OBJECTIVES:The Bacillus Calmette-Guerin (BCG) vaccine has beneficial effects on the immune system, which may lead to non-specific protection against non-tuberculous infections and increase the response to subsequent vaccinations. Seasonal influenza vaccination is used to protect senior citizens against influenza, but the serological response to the inactivated influenza vaccine (IIV) decreases in the elderly due to immunosenescence. The aim of this study was to test the capacity of BCG to boost the specific immune response to IIV and explore the effect of BCG on the innate immune system and health of senior citizens. METHODS:A randomised controlled trial with a nested immunological study including 273 Danish citizens >65 years. Participants were randomised into four equally sized groups combining BCG with IIV in different sequences compared with IIV alone. The primary outcome was change in influenza antibody hemagglutination inhibition (HI) titre four weeks after vaccination. Secondary outcomes were infection rate during six months follow-up, and association between influenza antibody titre and infection rate. In subgroup analyses, we explored the effect of BCG on lymphocyte populations seven days after IIV and on cytokine production after stimulation of mononuclear cells in vitro. RESULTS:Four weeks after influenza vaccination, the mean fold change in HI titre over all serotypes was 2.3-2.5 with no significant differences between the treatment groups. Seroconversion rate was comparable between treatment groups, and in strata of age and sex. There was no difference in the rate of infection between the groups and there was no association with influenza antibody level. We found no difference in distribution of lymphocytes. Combining BCG with IIV had modest impact on in vitro cytokine production compared with IIV alone. CONCLUSION:BCG vaccination did not increase serological response to seasonal influenza vaccination or reduce the incidence of infection in this population of Danish senior citizens. TRIAL REGISTRATION:EU Clinical Trials Register (EudraCT number 2019-002781-12). SUMMARY:In this randomised clinical trial of Danish senior citizens, BCG vaccination did not influence serological response to influenza vaccination or reduce the risk of infection compared with placebo. There was no clear indication of induction of trained immunity.

Introduction: Healthcare workers (HCWs) faced a heightened risk of SARS-CoV-2 infection compared to the general population. The Bacillus Calmette-Guérin (BCG) vaccine, used for over a century, is known to modulate immune responses to various pathogens, including the SARS-CoV-2. This study examines the association between lifetime BCG vaccination and the risk of SARS-CoV-2 infection among HCWs in Cape Verde. Methods: We conducted a case-control study among HCWs in Cape Verde in 2021. Cases were HCWs, 18 years or older who tested positive for SARS-CoV-2 via PCR or Rapid Antigen Test, identified by Public Health Authorities, while controls were HCWs, 18 years or older with no reported or suspected SARS-CoV-2 infection, confirmed by POC Rapid Antigen Test. Both were selected from HCWs of three hospitals on Santiago and São Vicente islands in Cape Verde. The primary outcome measured was the status of SARS-CoV-2 infection (yes/no), and the primary exposure examined was BCG vaccination status (yes/no). The study also considered sociodemographic, epidemiological, and employment variables. Data collection was conducted using an electronic questionnaire applied to all participants, and analysis employed a maximum likelihood forward selection logistic regression model to assess the impact of BCG vaccination on the odds of SARS-CoV-2 infection. Results: The study included 215 cases and 250 controls. Most cases of SARS-CoV-2 infection that occurred before completion of COVID-19 vaccination were identified through epidemiological surveillance or among people who reported being quarantined and/or absent from work after diagnosis. Hospitalization for COVID-19 was required for 7.7% of the cases. Persistent symptoms following COVID-19 were reported in fewer than 1% of cases. Having at least one child in the house, not having double employment, and being of normal weight were more likely to be observed among cases than controls. BCG vaccination showed no association with SARS-CoV-2 infection (aOR of BCG-vaccinated vs. not BCG-vaccinated = 0.76 [0.48; 1.20]). Completing the COVID-19 vaccination regimen was associated with reduced odds of SARS-CoV-2 infection, but there was likely some reversed causation, as vaccination was postponed for 6 months post-COVID-19 infection for cases. Conclusions: Lifetime BCG vaccination did not reduce the likelihood of SARS-CoV-2 infection among HCWs.

BACKGROUND:Within a randomised controlled trial providing Bacillus Calmette-Guérin, measles vaccine, or placebo to women of fertile age, we aimed to study the risk of adverse events following vaccination. This study addresses the lack of data on adverse reactions in adults, especially women of childbearing age, to inform broader vaccination policies. METHODS:The trial included HIV-negative, non-pregnant women aged 15 to 35 years, who had given birth to at least one child, and residing in the Bandim Health Project study area in the capital Bissau of Guinea-Bissau. Excluded were individuals who were frail, had serious illnesses like active tuberculosis, or had no prior births. Participants were randomised 1:1:1 to bacillus calmette-guérin, measles vaccine, or placebo. A subset of participants was enrolled into the adverse events study. Adverse events were registered immediately 15 min post-vaccination and at home visits 7, 14, 28 and 90 days after enrolment to register symptoms that could be related to the trial interventions. We compared symptom distributions by randomisation using chi-squared tests. RESULTS:Of the total 366 women included in the study, 99% (363) were eligible to follow up for adverse events (Placebo, 121; measles vaccine, 121; Bacillus Calmette-Guérin, 121) and 97% (1414/1452) of the planned study contacts were successful, 1.3% (18/1414) of which were mobile phone interviews. The baseline characteristics did not differ between the randomisation groups. There were no immediate reactions within the first 15 min post-vaccination. Seven days after vaccination, there was a tendency for more episodes of headache reported among both the Bacillus Calmette-Guérin and measles vaccine vaccinated participants compared to placebo. We registered 11 medical consultations (placebo: 4, measles vaccine: 5, Bacillus Calmette-Guérin: 2) and no deaths or hospitalisations during 90 days of post-enrolment follow-up. CONCLUSION:Bacillus Calmette-Guérin and measles vaccine were safe and tolerable for adult women of childbearing age, with few short- and medium-term adverse events.

Trained immunity, a de-facto innate immune memory, has been extensively studied in response to live-attenuated vaccines, but its presence following the new COVID-19 vaccines has not yet been fully elucidated. In this study, we investigate markers of trained immunity in individuals vaccinated with mRNA-1273 or BNT162b2. As part of the vaccine roll-out in Denmark and recruited for a comparative study. Our primary objective was to determine whether these vaccines elicit lasting changes in innate immune responses, particularly in monocyte populations and cytokine production following stimulation with a panel of agonists. The study was conducted at four time points: Day-0 (pre-vaccination), Day-28, Day-90, and Day-180 post-vaccination. We observed no significant differences in monocyte subpopulations between vaccine groups; however, cytokine and chemokine analysis revealed distinct immune signatures. While IL-6 and TNFα production remained unchanged after ex-vivo restimulation in the BNT162b2 group, individuals vaccinated with mRNA-1273 exhibited a sustained increase in the production of these cytokines, persisting for up to 180 days post-vaccination. Additionally, CCL2, a key chemokine involved in monocyte recruitment, was upregulated following mRNA-1273 vaccination but decreased in the BNT162b2 group, further supporting the finding of differential innate immune responses between the two vaccines. In conclusion, our study provides evidence that mRNA-1273, but not BNT162b2, induces immune responses consistent with the concept of trained immunity. These results highlight the potential for mRNA vaccine platforms to shape innate immunity, with implications for future vaccine design aimed at enhancing non-specific and specific protection against infectious diseases.

The use of Bacillus Calmette-Guérin (BCG) vaccine against tuberculosis (TB) spans more than a century. Besides protection against severe paediatric TB, randomized trials and novel advances within innate immunology documented that BCG has beneficial non-specific effects, providing protection against non-TB infections. Since paediatric intradermal BCG vaccination has proved unable to contain adult pulmonary tuberculosis, several novel TB vaccines are under development, most of which build upon BCG. BCG's status as an essential remedy against TB will therefore be maintained, but despite many decades of near-universal intradermal use, the local responses to BCG in the skin have not been thoroughly elucidated. We therefore developed appropriate methods to capture the localised skin events at the cellular and molecular level after intradermal BCG vaccination. This work informs future studies to identify the immunological events induced following administration of BCG to accelerate development of improved or new vaccines against Mycobacterium tuberculosis (Mtb). We employed advanced technologies such as spatial transcriptomics on skin tissue punch biopsies and cell-free plasma transcriptomics (liquid biopsies) to characterize both the local (in situ) skin and systemic (peripheral blood) response to BCG via:•Whole blood transcriptomics and epigenetic analysis•Blood immune cell characterization•Plasma proteomic and metabolomic analysis.

Objectives: Bacillus Calmette-Guerin (BCG), similar to some other live vaccines, may offer partial protection against unrelated infections. During the COVID-19 pandemic, several trials tested whether BCG provides protection against COVID-19. Design: Meta-analysis of randomised placebo-controlled trials which provided mortality data. Setting: High-income settings as well as low- and middle-income settings (LMICs). Participants: Trial populations were healthcare workers (HCW) or individuals of older age, followed for 6-12 months. Interventions: BCG vaccine versus placebo. Main outcome measure: Overall mortality and COVID-19 mortality. Results Nine of the 15 RCTs reported at least one death. Trial populations were healthcare workers (4 RCTs) or individuals of older age (5 RCTs), in Europe (5 RCTs) or LMICs (4 RCTs) with a total of 8,169 participants randomly allocated to BCG (18 deaths) and 8,176 to placebo (37 deaths). In a meta-analysis, receiving BCG vs placebo was associated with a 51% (95% CI: 15 to 72%) reduction in all-cause mortality. The all-cause mortality RRR was 78% (-3 to 95%) in HCWs and 43% (95% CI: -6 to 69%) in the elderly. It was 37% (95 CI: -15 to 66%) in Europe and 90% (95% CI: 22 to 99%) in LMICs. The relative reduction of deaths (6 BCG; 12 placebo) from COVID-19 was 50% (95 CI: -33 to 81). Conclusions: BCG may have reduced adult all-cause mortality in the context of the COVID-19 pandemic. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement This study did not receive any funding ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: This meta-analysis used only data extracted from previously published studies. No new or individual participant data were generated. All underlying source data are available in the original trial reports and their supplementary materials. All data received was de-identified. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes Data sharing: This meta-analysis used only data extracted from previously published studies. No new or individual participant data were generated. All underlying source data are available in the original trial reports and their supplementary materials. The complete dataset compiled for this meta-analysis, including all extracted variables used in the analyses, is submitted with the manuscript and will be deposited in a public repository and made openly available prior to publication. Code sharing: The full Stata code used for the meta-analysis is submitted as a supplementary file and will be deposited in a public repository and made openly available prior to publication.

AimEstimate the incidence of asthma among children aged 0 to 15 years in Denmark, Finland, Norway and Sweden during 2000-2017.MethodsCases of preschool asthma (up to 6 years) and school-age asthma (from 6 years) were identified through national registers using an algorithm including hospital diagnoses and prescription medicines. The respective cumulative incidence (CI) was estimated in 1-year age intervals for each country and birth year.ResultsThe CI of algorithm-based preschool asthma peaked for the birth cohorts 2008 or 2009 at 14.8% in Denmark, 11.0% in Finland, 15.1% in Norway and 13.7% in Sweden. For later birth cohorts, a slight decrease was observed. The CI of school-age asthma was 7.1% in Denmark, 10.5% in Finland, 9.7% in Norway and 10.2% in Sweden (children born in 2002). A slight decline over time was seen in Denmark and Norway, and a slight increase in Sweden and Finland.ConclusionFinland had a markedly lower CI of preschool asthma and Denmark lower CI of school-age asthma as estimated by prescriptions and hospital diagnoses. Preschool asthma may have plateaued in the Nordic countries. For school-age asthma trends over time varied by country. Differences in diagnostic and prescription practices may have influenced the results.

ABSTRACT Background In low-income settings, WHO recommended a booster-dose of DTP (DTP4) and OPV (OPV4) at 18 months of age. Previous studies found DTP after MV to be associated with higher female mortality. We conducted a randomised trial (RCTs) in Guinea-Bissau, in which children from 18 months of age were randomised to receive DTP4+OPV4 vs OPV4-only to assess the impact on survival and hospitalisations, overall and by sex. Methods The trial was conducted from 2005-2012. Children were randomised and followed from 18 months to 4 years of age. Hospitalisations were captured through a surveillance system at the main paediatric ward, and deaths were detected through a demographic surveillance system in the study area and through yearly home visits to the study participants. Data was analysed in Cox proportional hazards models. The RCT aimed to enrol 6,000 children based on an expected annual mortality rate of 3%. During the trial period many new interventions, including many national health campaigns, were carried out. Results The trial enrolled 5,918 children of which 96% (5,673) were included in the analysis. Annual non-accidental mortality rate was 0.56%, 81% lower than the expected rate. There was no difference in mortality between the randomisation groups. With 30 and 35 deaths, the hazard ratio (HR) for DTP4+OPV4 vs OPV4-only was 0.84 (95% CI=0.52-1.37); 0.86 (0.42-1.77) in males and 0.82 (0.42-1.60) in females. The HR for hospitalisations was 0.90 (0.77-1.06); 0.87 (0.70-1.09) in males; 0.94 (0.74-1.20) in females). Conclusion The study found no negative effect of DTP4 and no sex-differential effect of DTP4, but it was strongly underpowered. Furthermore, due to the large number of health interventions, not envisioned at the initiation of the trial, a limited part of the follow-up was a comparison between DTP4+OPV4 vs OPV4 as the most recent vaccinations. ### Competing Interest Statement The authors have declared no competing interest. ### Clinical Trial https://clinicaltrials.gov/study/[NCT00244673][1] ### Funding Statement Funding: Aarhus Universitetshospitals Forskningsinitiativ, Lundbeckfonden, Aase og Ejnar Danielsens Fond, Det danske Pasteur Selskab, Jakob og Olga Madsens Fond, Laegernes Forsikringsforening, Dagmar Marshalls Fond, Scandinavian Society for Antimicrobial Chemotherapy Foundation, Forskerskolen for International Sundhed KU, Faculty of Health Aarhus Universitet contributed to the study. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: The National Ethical Committee in Guinea-Bissau approved the protocol, and the Danish Central Ethical Committee gave consultative approval. The trial was registered on October 25th 2005 at clinicaltrials.gov: https://clinicaltrials.gov/study/[NCT00244673][1]. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes Data availability statement: By contact to the corresponding author. [1]: /lookup/external-ref?link_type=CLINTRIALGOV&access_num=NCT00244673&atom=%2Fmedrxiv%2Fearly%2F2025%2F10%2F31%2F2025.10.29.25339081.atom

Tuberculosis (TB) is caused by Mycobacterium tuberculosis (Mtb) and is a leading cause of death. BCG is the only licensed TB vaccine. Preclinical studies have shown that in adults, intravenous administration of BCG improves protection against TB. We hypothesize that intradermal administration of BCG to the human newborn leads to low-grade BCG bacteremia and that this systemic dissemination improves protection against Mtb infection. This hypothesis is based on supporting observations including animal and human studies. It is a testable hypothesis and offers to deliver immediately actionable insight to advance the global efforts against TB.

Objectives: To investigate if receipt of measles-mumps-rubella (MMR) vaccine following the third dose of diphtheria-tetanus-acellular pertussis (DTaP3) is associated with reduced rates of non-targeted infectious disease hospitalisations. Methods: Register based cohort study following 1,397,027 children born in Denmark, Finland, Norway, and Sweden until 2 years of age. Rates of infectious disease hospitalisations with minimum one overnight stay according to time-varying vaccination status were compared using Cox proportional hazards regression analysis with age as the underlying timescale and including multiple covariates. Summary estimates were calculated using random-effects meta-analysis. Results: Compared with DTaP3 and no MMR vaccine, MMR after DTaP3 was associated with reduced rates of infectious disease hospitalisations: aHR was 0.86 (0.83-0.89) in Denmark, 0.70 (0.64-0.75) in Finland, 0.71 (0.68-0.74) in Norway, and 0.71 (0.65-0.77) in Sweden: summary estimate was 0.75 (0.65 to 0.84). A beneficial association was also seen in a negative control exposure analysis (3 vs. 2 DTaP doses): summary estimate aHR was 0.81 (0.75-0.87). Conclusions: Having MMR as the most recent vaccine was consistently associated with reduced rates of infectious disease hospitalisation. However, bias may account for at least some of the observed association. Randomised controlled trials are warranted to inform the optimal timing of MMR for both its specific and potential non-specific effects. (c) 2024 The Author(s). Published by Elsevier Ltd on behalf of The British Infection Association. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

BACKGROUND:The decline in global child mortality is an important public health achievement, yet child mortality remains disproportionally high in many low-income countries like Guinea-Bissau. The persisting high mortality rates necessitate targeted research to identify vulnerable subgroups of children and formulate effective interventions. OBJECTIVE:This study aimed to discover subgroups of children at an elevated risk of mortality in the urban setting of Bissau, Guinea-Bissau, West Africa. By identifying these groups, we intend to provide a foundation for developing targeted health interventions and inform public health policy. METHODS:We used data from the health and demographic surveillance site, Bandim Health Project, covering 2003 to 2019. We identified baseline variables recorded before children reached the age of 6 weeks. The focus was on determining factors consistently linked with increased mortality up to the age of 3 years. Our multifaceted methodological approach incorporated spatial analysis for visualizing geographical variations in mortality risk, causally adjusted regression analysis to single out specific risk factors, and machine learning techniques for identifying clusters of multifactorial risk factors. To ensure robustness and validity, we divided the data set temporally, assessing the persistence of identified subgroups over different periods. The reassessment of mortality risk used the targeted maximum likelihood estimation (TMLE) method to achieve more robust causal modeling. RESULTS:We analyzed data from 21,005 children. The mortality risk (6 weeks to 3 years of age) was 5.2% (95% CI 4.8%-5.6%) for children born between 2003 and 2011, and 2.9% (95% CI 2.5%-3.3%) for children born between 2012 and 2016. Our findings revealed 3 distinct high-risk subgroups with notably higher mortality rates, children residing in a specific urban area (adjusted mortality risk difference of 3.4%, 95% CI 0.3%-6.5%), children born to mothers with no prenatal consultations (adjusted mortality risk difference of 5.8%, 95% CI 2.6%-8.9%), and children from polygamous families born during the dry season (adjusted mortality risk difference of 1.7%, 95% CI 0.4%-2.9%). These subgroups, though small, showed a consistent pattern of higher mortality risk over time. Common social and economic factors were linked to a larger share of the total child deaths. CONCLUSIONS:The study's results underscore the need for targeted interventions to address the specific risks faced by these identified high-risk subgroups. These interventions should be designed to work to complement broader public health strategies, creating a comprehensive approach to reducing child mortality. We suggest future research that focuses on developing, testing, and comparing targeted intervention strategies unraveling the proposed hypotheses found in this study. The ultimate aim is to optimize health outcomes for all children in high-mortality settings, leveraging a strategic mix of targeted and general health interventions to address the varied needs of different child subgroups.

The Bacille Calmette-Guerin (BCG) vaccine is a well-established inducer of innate immune memory (also termed trained immunity), causing increased cytokine production upon heterologous secondary stimulation. Innate immune responses are known to be influenced by season, but whether seasons impact induction of trained immunity is not known. To explore the influence of season on innate immune memory induced by the BCG vaccine, we vaccinated healthy volunteers with BCG either during winter or spring. Three months later, we measured the ex vivo cytokine responses against heterologous stimuli, analyzed gene expressions and epigenetic signatures of the immune cells, and compared these with the baseline before vaccination. BCG vaccination during winter induced a stronger increase in the production of pro-inflammatory cytokines by peripheral blood mononuclear cells (PBMCs) upon stimulation with different bacterial and fungal stimuli, compared to BCG vaccination in spring. In contrast, winter BCG vaccination resulted in lower IFNγ release in PBMCs compared to spring BCG vaccination. Furthermore, NK cells of the winter-vaccinated people had a greater pro-inflammatory cytokine and IFNγ production capacity upon heterologous stimulation. BCG had only minor effects on the transcriptome of monocytes 3 months later. In contrast, we identified season-dependent epigenetic changes in monocytes and NK cells induced by vaccination, partly explaining the higher immune cell reactivity in the winter BCG vaccination group. These results suggest that BCG vaccination during winter is more prone to induce a robust trained immunity response by activating and reprogramming the immune cells, especially NK cells. (Dutch clinical trial registry no. NL58219.091.16)