Introduction: Upper respiratory tract infections (URTI) are the most common illnesses affecting athletes, causing absences from training and competition. Salivary immunoglobulin A (sIgA) is the main immune factor in saliva, and a consistent association between low concentrations of sIgA and an increased incidence of URTIs has been reported. The oral probiotic Streptococcus salivarius K12 has been suggested to have the potential to improve oral diseases and mucosal barrier function. However, the effects of this probiotic on active young subjects performing a high-intensity training (HIT) program have not been investigated.

Methods: Active young students were randomised into a treated group (S. salivarius K12) and a control (placebo) group and asked to take the product daily for 30 days. After this period, participants performed a graded exercise test and five HIT sessions, all within 3 days. They were also asked to complete the Wisconsin Upper Respiratory Symptom Survey daily to monitor URTI’s presence. Before and after the 30 days, and at 0h, 24h and 72h after the last training session, saliva samples were collected to quantify sIgA level, secretion rate, and flow. The effect of S. salivarius K12 intake on these parameters was tested using an ANOVA for repeated measures.

Results: Twenty (M = 14, F = 6) young subjects (23.5 ± 2.3 years old) participated in the study. The total accumulated training load (sRPE) in the supplementation period was similar in the two groups (treated: 4345 ± 3441 AU; control: 4969 ± 4165 AU; p > 0.05). Considering both sIgA level and secretion rate, significant time (F_(4,15) = 3.38; p = 0.037; F_(4,15) = 6.00; p = 0.004) and time×group interactions (F_(4,15) = 2.49; p = 0.049; F_(4,15) = 5.01; p = 0.009) were reported, with the treated group showing higher sIgA levels at 72h post-exercise and increased secretion rate both at 0h and 72h. The number of URTI episodes was similar in the treated and control groups (χ² = 1.83; p > 0.05).

Conclusion: This study demonstrates that relatively short-term S. salivarius K12 supplementation increased sIgA level and secretion in healthy subjects performing a demanding exercise-training programme composed of HIT sessions.

Introduction: Low microbial biomass and high human DNA content in nasopharyngeal aspirate samples hinder comprehensive characterization of microbiota and resistome. We obtained samples from premature infants, a group with increased risk of developing respiratory disorders and infections, and consequently frequent exposure to antibiotics. Our aim was to devise an optimal protocol for handling nasopharyngeal aspirate samples from premature infants, focusing on host DNA depletion and microbiome and resistome characterization.

Methods: Three depletion and three DNA extraction protocols were compared, using RT-PCR and whole metagenome sequencing to determine the efficiency of human DNA removal, taxonomic profiling and assignment of antibiotic resistance genes. Protocols were tested using mock communities, as well as pooled and individual patient samples.

Results: The only extraction protocol to retrieve the expected DNA yield from mock community samples was based on a lytic method to improve Gram positive recovery (MasterPure™). Host DNA content in non-depleted aliquots from pooled patient samples was 99%. Only samples depleted with MolYsis™ showed satisfactory, but varied reduction in host DNA content, in both pooled and individual patient samples, allowing for microbiome and resistome characterisation (host DNA content from 15% to 98%). Other depletion protocols either retrieved too low total DNA yields, preventing further analysis, or failed to reduce host DNA content. By using Mol_MasterPure protocol on aliquots from pooled patient samples, we increased the number of bacterial reads by 7.6 to 1,725.8-fold compared to non-depleted reference samples. PCR results were indicative of achieved microbial enrichment. Individual patient samples processed with Mol_MasterPure protocol varied greatly in total DNA yield, host DNA content (from 40% to 98%), species and antibiotic resistance gene richness.

Discussion: Despite high human DNA and low microbial biomass content in nasopharynx aspirates of preterm infants, we were able to reduce host DNA content to levels compatible with downstream shotgun metagenomic analysis, including bacterial species identification and coverage of antibiotic resistance genes. Whole metagenomic sequencing of microbes colonizing the nasopharynx may contribute to explaining the possible role of airway microbiota in respiratory conditions and reveal carriage of antibiotic resistance genes.