Contribution of Streptococcus pseudopneumoniae and Streptococcus salivarius to vocal fold mucosal integrity and function.

Structural changes to the vocal fold (VF) epithelium, namely, loosened intercellular junctions, have been reported in VF benign lesions. The potential mechanisms responsible for the disruption of cell junctions do not address the contribution of resident microbial communities to this pathological phenomenon. In this study, we focused on determining the relationship between Streptococcus pseudopneumoniae (SP), a dominant bacterial species associated with benign lesions, and Streptococcus salivarius (SS), a commensal bacterium, with human VF epithelial cells in our three-dimensional model of the human VF mucosa. This experimental system enabled direct deposition of bacteria onto constructs at the air/liquid interface, allowing for the assessment of bacterium-host interactions at the cellular, molecular and ultrastructural levels. Our findings demonstrate that SP disrupts VF epithelial integrity and initiates inflammation via the exported products HtrA1 and pneumolysin. In contrast, SS attaches to the VF epithelium, reduces inflammation and induces Mmp2-mediated apical desquamation of infected cells to mitigate the impact of pathogens. In conclusion, this study highlights the complexity of microbial involvement in VF pathology and potential VF mucosal restoration in the presence of laryngeal commensals.

Probiotic Streptococcus salivarius K12 Alleviates Radiation-Induced Oral Mucositis in Mice.

Background: Oral mucositis is the most common oral complication of cancer patients receiving radiotherapy and/or chemotherapy, leading to poor quality of life. Limitations of the current interventions on radiation-induced oral mucositis (RIOM) urge the development of novel therapeutics. Here, we evaluated the treatment outcome of probiotic Streptococcus salivarius K12 on RIOM mice, and oral microbiota that is associated with the progress of RIOM was further investigated. Methods: An experimental RIOM mouse model was established, and S. salivarius K12 was applied to the mouse oral cavity daily. Histological analyses were performed to evaluate the severity of oral mucositis and the treatment outcome of S. salivarius K12. The oral microbiota of mice was further analyzed by 16S rRNA sequencing, microbial culture and qPCR. Results: Irradiation induced conspicuous mucositis in the oral cavity of mice. S. salivarius K12 treatment was beneficial for the healing of RIOM, as reflected by reduced ulcer size, increased basal layer epithelial cellularity and mucosal thickness, and elevated epithelial proliferation and attenuated apoptosis. RIOM mice presented significant oral microbial dysbiosis, with an overgrowth of oral anaerobes. S. salivarius K12 treatment reconstituted the oral microbiota and decreased the abundance of oral anaerobes of RIOM mice. In addition, S. salivarius K12 treatment inhibited NI1060 in Pasteurella genus and downregulated the expression of nitrate reductase. Conclusions: S. salivarius K12 treatment can alleviate RIOM and reconstituted the dysbiotic oral microbiota in mice. S. salivarius K12 may represent a promising adjuvant treatment to improve the quality of life of cancer patients receiving radiotherapy.

Local Bacteriotherapy - a promising preventive tool in recurrent respiratory infections.

INTRODUCTION: Children with recurrent respiratory infections (RRI) represent a social issue for the economic burden and the negative family impact. Local Bacteriotherapy is an attractive therapeutic strategy that could be potentially effective in preventing infections. The current article remarks on the existing evidence of preventing RRI by Local Bacteriotherapy. AREAS COVERED: The literature search methodology was based on the articles cited by PubMed from 1980 to 2020. Respiratory infections include rhino-pharyngitis, otitis media, rhinosinusitis, pharyngo-tracheitis, bronchitis, and pneumonia. Several studies were performed to investigate the effects of Local Bacteriotherapy in children with RRI. Both intranasal and oral Local Bacteriotherapy were evaluated. The findings showed that Local Bacteriotherapy significantly reduced the number of RI episodes, their severity, the use of antibiotics, and school absences. EXPERT OPINION: Local Bacteriotherapy is a promising approach to RRI prevention and could be a profitable strategy to contrast infections in the future.

Sex effects in the association between airway microbiome and asthma.

BACKGROUND: Sex differences exist in asthma susceptibility and severity. Accumulating evidence has linked airway microbiome dysbiosis to asthma, and airway microbial communities have been found to differ by sex. However, whether sex modifies the link between airway microbiome and asthma has not been investigated. OBJECTIVE: To evaluate sex effects in the association between airway microbiome and asthma. METHODS: We analyzed induced sputum samples from 47 subjects (n = 23 patients with asthma and n = 24 normal controls) using 16S ribosomal RNA gene sequencing methods. The bacterial composition was analyzed for sex differences. Bacterial associations with asthma were assessed for each sex at the core taxa and genus levels. RESULTS: The microbiome in induced sputum differed in women vs men at the community level. A total of 5 core bacterial taxa were found in all samples. No sex-specific core taxa were detected. The most abundant core taxon, Streptococcus salivarius, was significantly enriched in women than in men (P = .02). Within each sex, individuals with relatively lower abundance of S salivarius were more likely to have asthma (P = .006). For both sexes, increased Lactobacillus species were found in sputum samples of patients with patients compared with normal controls (adjusted P = .01). Haemophilus species were associated with asthma in men and not in women. CONCLUSION: The airway microbiome differed by sex, and sex effects exist in the association of airway microbial markers and asthma. Future airway microbiome studies may yield better resolution if the context of specific sex is considered. The airway microbiome is a potential mechanism driving sex differences in asthma.

Randomised, double-blind, placebo-controlled trial of oral probiotic Streptococcus salivarius M18 on head and neck cancer patients post-radiotherapy: a pilot study.

Xerostomia detrimentally affects the oral health of many head and neck cancer patients who undergo radiotherapy. Its sequelae become an ongoing burden for patients that often manifest as periodontal disease and dental decay. Bacteria play a major role in the pathogenesis of these conditions and here we explore the use of an oral probiotic to beneficially modulate the oral bacterial community post-radiotherapy. In this pilot study, a four-week intervention with oral probiotic lozenges containing Streptococcus salivarius M18 was trialled in seven patients. Post-intervention changes in oral health and in the composition of the plaque and saliva bacterial communities were compared with six patients in a placebo group. An improvement in periodontal screening and plaque index scores was observed in both groups after the intervention period. The oral probiotic lozenges did not significantly impact bacterial community composition or diversity, nor did the probiotic lozenges increase the relative sequence abundance of ZOTU_1 (the probiotic-associated sequence assigned to S. salivarius) detected in the samples. Network analyses suggest negative interactions occurred between ZOTU_1 and species from the periopathogenic genera Campylobacter, Fretibacterium, Selenomonas and Treponema but further investigation is required to more fully understand the beneficial properties of this oral probiotic.

Abundance, Diversity and Role of ICEs and IMEs in the Adaptation of Streptococcus salivarius to the Environment.

Streptococcus salivarius is a significant contributor to the human oral, pharyngeal and gut microbiomes that contribute to the maintenance of health. The high genomic diversity observed in this species is mainly caused by horizontal gene transfer. This work aimed to evaluate the contribution of integrative and conjugative elements (ICEs) and integrative and mobilizable elements (IMEs) in S. salivarius genome diversity. For this purpose, we performed an in-depth analysis of 75 genomes of S. salivarius and searched for signature genes of conjugative and mobilizable elements. This analysis led to the retrieval of 69 ICEs, 165 IMEs and many decayed elements showing their high prevalence in S. salivarius genomes. The identification of almost all ICE and IME boundaries allowed the identification of the genes in which these elements are inserted. Furthermore, the exhaustive analysis of the adaptation genes carried by these elements showed that they encode numerous functions such as resistance to stress, to antibiotics or to toxic compounds, and numerous enzymes involved in diverse cellular metabolic pathways. These data support the idea that not only ICEs but also IMEs and decayed elements play an important role in S. salivarius adaptation to the environment.

Alteration of Streptococcus salivarius in Buccal Mucosa of Oral Lichen Planus and Controlled Clinical Trial in OLP Treatment.

Oral lichen planus (OLP) is a T cell-mediated common chronic inflammatory mucosal disease, with limited therapies available for long-term use. Previous study showed that ratio of genus Streptococcus decreased significantly in OLP patients when compared with controls. Buccal cotton swab samples of 43 OLP patients and 48 healthy individuals were collected for real-time quantitative polymerase chain reaction (RT-PCR) to investigate relative abundance alteration of Streptococcus salivarius in OLP lesions. Bacterial supernatants of S. salivarius ATCC® BAA-2593™ were collected by centrifugation and added to HSC-3 cells, and quantitative analysis of expression of IL-1ß, IL-6, IL-8, and TNF-α in the HSC-3 cells was determined by RT-PCR. Then, a randomized, non-blinded, controlled study was conducted. Forty patients with symptomatic OLP were randomly allocated into two groups and received topical treatment of 0.1% triamcinolone acetonide dental paste (group A) and S. salivarius K12 lozenge (group B), respectively, for 4 weeks. Sign scores, visual analogue scale (VAS), and adverse reactions were recorded. Relative abundance of S. salivarius in the OLP group was lower than that of control group (P < 0.05). After treated with 0.1% supernatants of S. salivarius ATCC® BAA-2593™, the expression level of IL-6 in the HSC-3 cells significantly reduced (P < 0.001), while IL-1ß, IL-8, and TNF- α showed a decreasing tendency (P > 0.05). There was significant reduction in sign scores and VAS scores in both groups after the 4-week treatment, with no significant difference between two groups. No adverse reaction was observed. S. salivarius might maintain local immune balance by inhibiting the NF-κB pathway. Topical application of Streptococcus salivarius K12 seemed to be effective in treatment of symptomatic OLP, especially with promising potential in long-term use. More detailed clinical studies with long follow-up period and standardized usage/dosage are expected to acquire definite conclusions.

The Effect of Streptococcus salivarius K12 on Halitosis: a Double-Blind, Randomized, Placebo-Controlled Trial.

This study was to evaluate the effect of Streptococcus salivarius K12 on tongue coating-associated halitosis. Twenty-eight subjects having tongue coating-associated halitosis were randomly divided into either a test or control group. For each of the 30 days, the test subjects sucked S. salivarius K12 tablet while the control subjects sucked placebo tablets. All the subjects did not take physical (tongue scraping) and chemical (antiseptic mouth-rinse) oral cavity pretreatment prior to use of the tablets. At baseline, and on the 1st, 7th, and 14th day after completing the course of tablets, the subjects were assessed for their organoleptic test (OLT) scores, volatile sulfur compound (VSC) levels, and tongue coating scores (TCS). During the course, all subjects kept their routine oral care habits without scraping their tongue coating. Plaque index, probing depth, and bleeding index were recorded at baseline and at the completion of the trial. On the 1st day following the end of tablet use, the OLT scores and VSC levels had significantly decreased in the test group when compared with the baseline values (P = 0.001 and P = 0.012). The TCS in the test group were also significantly decreased (P = 0.05). At days 7 and 14, the OLT scores in the test group were still significantly lower than the baseline levels (P = 0.006 and P = 0.039 respectively). However, there were no statistical differences with OLT, VSC, and TCS between the test group and the placebo group by analysis of multi-level regression model. The use of S. salivarius K12 did not have significant effect on halitosis with tongue coating cause when the tongue coating was not physically or chemically pre-treated, which implies removing tongue coating is required before Streptococcus salivarius K12 use.

Strain-level analysis of gut-resident pro-inflammatory viridans group Streptococci suppressed by long-term cotrimoxazole prophylaxis among HIV-positive children in Zimbabwe.

Antimicrobials have become a mainstay of healthcare in the past century due to their activity against pathogens. More recently, it has become clear that they can also affect health via their impact on the microbiota and inflammation. This may explain some of their clinical benefits despite global increases in antimicrobial resistance (AMR) and reduced antimicrobial effectiveness. We showed in a randomized controlled trial of stopping versus continuing cotrimoxazole prophylaxis among HIV-positive Zimbabwean children taking antiretroviral therapy (ART), that continuation of cotrimoxazole persistently suppressed gut-resident viridans group streptococcal species (VGS) that were associated with intestinal inflammation. In this addendum, we provide a broader overview of how antibiotics can shape the microbiota and use high read-depth whole metagenome sequencing data from our published study to investigate whether (i) the impact of cotrimoxazole on gut VGS and (ii) VGS associated inflammation, is attributable to strain-level variability. We focus on S. salivarius, the VGS species that was most prevalent in the cohort and for which there was sufficient genome coverage to differentiate strains. We demonstrate that suppression of S. salivarius by cotrimoxazole is not strain specific, nor did stool concentration of the pro-inflammatory mediator myeloperoxidase vary by S. salivarius strain. We also show that gut-resident S. salivarius strains present in this study population are distinct from common oral strains. This is the first analysis of how cotrimoxazole prophylaxis used according to international treatment guidelines for children living with HIV influences the gut microbiome at the strain-level. We also provide a detailed review of the literature on the mechanisms by which suppression of VGS may act synergistically with cotrimoxazole's anti-inflammatory effects to reduce gut inflammation. A greater understanding of the sub-clinical effects of antibiotics offers new insights into their responsible clinical use.

A single-centre investigator-blinded randomised parallel group clinical trial to investigate the effect of probiotic strains Streptococcus salivarius M18 and Lactobacillus acidophilus on gingival health of paediatric patients undergoing treatment with fixed orthodontic appliances: study protocol.

BACKGROUND: There is limited data on the beneficial effects of probiotics on the gingival health of patients undergoing treatment with fixed orthodontic appliances. This study aims to compare the effect of probiotic tablets combined with regular oral hygiene versus regular oral hygiene alone on gingival status in these patients. The effect of probiotic intake on plaque formation and salivary microbiome composition will be also assessed. METHODS AND ANALYSIS: This is a 3 month single-centre, single blind (clinical and laboratory examiners), parallel group randomised controlled two arm superiority trial. Fifty paediatric patients attending the Postgraduate Orthodontic Clinic at the Hamdan Bin Mohammed College of Dental Medicine (HBMCDM), Mohammed Bin Rashid University of Medicine and Health Sciences (MBRU), Dubai, United Arab Emirates, who meet the eligibility criteria will be recruited. Block randomisation with 1:1 allocation and concealment of allocation will be carried out. The treatment group will receive probiotic tablets containing Streptococcus salivarius M18 and Lactobacillus acidophilus together with regular oral hygiene versus the control group on regular oral hygiene alone. Clinical examination and collection of saliva for microbiome assay will be carried out at baseline and end of study. Self-reporting by patients will be used to document acceptability and adverse effects. Statistically significant decrease in gingival bleeding on probing in the treatment group will be classified as primary outcome of treatment success. Statistically significant reduction in Plaque Index, Gingival Index and shift in the composition of the oral microbiome in favour of beneficial bacteria are secondary outcomes indicative of efficacy of probiotic intake. ETHICS AND DISSEMINATION: Ethical approval for the study has been granted by the HBMCDM, MBRU, Institutional Review Board (Reference #: MBRU-IRB-2018-015). Study findings will be disseminated via publication in peer-reviewed journal. TRIAL REGISTRATION NUMBER: ISRCTN95085398.

Assessment of Efficacy of BLIS-Producing Probiotic K12 for the Prevention of Group A Streptococcus Pharyngitis: a Short Communication.

The neutral outcome of the recently reported school-based trial of probiotic K12 (The effect of the oral probiotic Streptococcus salivarius K12 on group A streptococcus pharyngitis: a pragmatic trial in schools) can be attributed at least partially to several readily identifiable confounding factors. Mainly, the execution and outcome were negatively impacted by (a) the suboptimal efficacy and frequency of K12 administration, (b) the failure both clinically and microbiologically to adequately diagnose and distinguish active group A streptococci (GAS) pharyngitis from harmless GAS carriage, and (c) the exceptionally low occurrence of GAS in this population at the time of the probiotic intervention due to recent high-intensity antibiotic exposure.

On the influence of Streptococcus salivarius on the wear response of dental implants: An in vitro study.

This study aims to understand microorganisms' effect and relevance of aseptic conditions on wear performance of dental materials. Tribocouple zirconia/titanium was submitted to tribological tests in the presence of Streptococcus salivarius biofilm, the most abundant bacteria in the oral cavity. Reciprocating ball-on-plate tests were carried out at 37°C, pH 7, 2 N, and 1 Hz, during 2 h, using Ti6Al4V balls as pins and Y-TZP plates. Simultaneous OCP readings assessed corrosion tendency. Tested lubricants were artificial saliva, artificial saliva plus glucose, and artificial saliva plus glucose and S. salivarius. Wear volume and worn surfaces were analyzed after test. S. salivarius proliferation was enhanced in artificial saliva plus glucose. When a highly populated biofilm was present at the zirconia/titanium interface, titanium wear decreased by ~60% compared to artificial saliva and artificial saliva plus with glucose, respectively, with 0.0060 and 0.0053 mm3 loss. Wear was not observed in zirconia under any tested condition. Presence of S. salivarius also decreased corrosion activity during wear. Lowest free OCP value reached during sliding was -260 mV in artificial saliva, -246 mV in artificial saliva plus glucose, and -196 mV in artificial saliva plus glucose and bacteria. This decreased to -147 mV when a highly-populated S. salivarius biofilm was present at the interface. Overall, obtained results show that the presence of S. salivarius clearly changes the corrosion-wear performance of the tested dental materials, confirming that it must be recognized, and that asepsis level and conditions during wear test of dental materials must be clearly established and controlled to ensure reproducible results and supported conclusions. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 1393-1399, 2019.

Putative Microbial Population Shifts Attributable to Nasal Administration of Streptococcus salivarius 24SMBc and Streptococcus oralis 89a.

Changes in bacterial composition of nasal microbiota may alter the host's susceptibility to several infectious and allergic diseases such as chronic rhinosinusitis and allergic rhinitis. The aim of this study was to evaluate the effects of 1-week administration of a probiotic product, composed by a combination of Streptococcus salivarius 24SMBc and Streptococcus oralis 89a, on the nostril microbiota. Differences in the nasal microbiota composition were investigated by using a next-generation sequencing approach. A strong and significant decrease in Staphylococcus aureus abundance was detected immediately after the bacterial administration. Moreover, comparing the microbial networks of nostril microbiota before and 1 month after the end of treatment, we detected an increase in the total number of both bacterial nodes and microbial correlations, with particular regard to the beneficial ones. Furthermore, a less abundance of microbial genera commonly associated to potential harmful bacteria has been observed. These results suggest a potential ability of S. salivarius 24SMBc and S. oralis 89a to regulate and reorganize the nasal microbiota composition, possibly favoring those microorganisms that may be able to limit the overgrowth of potential pathogens.

Probiotics Streptococcus salivarius 24SMB and Streptococcus oralis 89a interfere with biofilm formation of pathogens of the upper respiratory tract.

BACKGROUND: Infections of the ears, paranasal sinuses, nose and throat are very common and represent a serious issue for the healthcare system. Bacterial biofilms have been linked to upper respiratory tract infections and antibiotic resistance, raising serious concerns regarding the therapeutic management of such infections. In this context, novel strategies able to fight biofilms may be therapeutically beneficial and offer a valid alternative to conventional antimicrobials. Biofilms consist of mixed microbial communities, which interact with other species in the surroundings and communicate through signaling molecules. These interactions may result in antagonistic effects, which can be exploited in the fight against infections in a sort of "bacteria therapy". Streptococcus salivarius and Streptococcus oralis are α-hemolytic streptococci isolated from the human pharynx of healthy individuals. Several studies on otitis-prone children demonstrated that their intranasal administration is safe and well tolerated and is able to reduce the risk of acute otitis media. The aim of this research is to assess S. salivarius 24SMB and S. oralis 89a for the ability to interfere with biofilm of typical upper respiratory tract pathogens. METHODS: To investigate if soluble substances secreted by the two streptococci could inhibit biofilm development of the selected pathogenic strains, co-cultures were performed with the use of transwell inserts. Mixed-species biofilms were also produced, in order to evaluate if the inhibition of biofilm formation might require direct contact. Biofilm production was investigated by means of a spectrophotometric assay and by confocal laser scanning microscopy. RESULTS: We observed that S. salivarius 24SMB and S. oralis 89a are able to inhibit the biofilm formation capacity of selected pathogens and even to disperse their pre-formed biofilms. Diffusible molecules secreted by the two streptococci and lowered pH of the medium revealed to be implied in the mechanisms of anti-biofilm activity. CONCLUSIONS: S. salivarius 24SMB and S. oralis 89a possess desirable characteristics as probiotic for the treatment and prevention of infections of the upper airways. However, the nature of the inhibition appear to be multifactorial and additional studies are required to get further insights.

Surface proteins involved in the adhesion of Streptococcus salivarius to human intestinal epithelial cells.

The adhesion properties of 14 Streptococcus salivarius strains to mucus (HT29-MTX) and non-mucus secreting (Caco-2/TC7) human intestinal epithelial cells were investigated. Ability to adhere to these two eukaryotic cell lines greatly differs between strains. The presence of mucus played a major factor in adhesion, likely due to high adhesiveness to mucins present in the native human mucus layer covering the whole cell surface. Only one S. salivarius strain (F6-1), isolated from the feces of a healthy baby, was found to strongly adhere to HT-29 MTX cells at a level comparable to that of Lactobacillus rhamnosus GG, a probiotic strain considered to be highly adherent. By sequencing the genome of F6-1, we were able to identify 36 genes encoding putative surface proteins. Deletion mutants were constructed for six of them and their adhesion abilities on HT-29 MTX cells were checked. Our study confirmed that four of these genes encode adhesins involved in the adhesion of S. salivarius to host cells. Such adhesins were also identified in other S. salivarius strains.

Circuitry Rewiring Directly Couples Competence to Predation in the Gut Dweller Streptococcus salivarius.

Small distortions in transcriptional networks might lead to drastic phenotypical changes, especially in cellular developmental programs such as competence for natural transformation. Here, we report a pervasive circuitry rewiring for competence and predation interplay in commensal streptococci. Canonically, in streptococci paradigms such as Streptococcus pneumoniae and Streptococcus mutans, the pheromone-based two-component system BlpRH is a central node that orchestrates the production of antimicrobial compounds (bacteriocins) and incorporates signal from the competence activation cascade. However, the human commensal Streptococcus salivarius does not contain a functional BlpRH pair, while the competence signaling system ComRS directly couples bacteriocin production and competence commitment. This network shortcut might underlie an optimal adaptation against microbial competitors and explain the high prevalence of S. salivarius in the human digestive tract. Moreover, the broad spectrum of bacteriocin activity against pathogenic bacteria showcases the commensal and genetically tractable S. salivarius species as a user-friendly model for competence and bacterial predation.

Streptococcus salivarius MS-oral-D6 promotes gingival re-epithelialization in vitro through a secreted serine protease.

Gingival re-epithelialization represents an essential phase of oral wound healing in which epithelial integrity is re-establish. We developed an automated high-throughput re-epithelialization kinetic model, using the gingival epithelial cell line Ca9-22. The model was employed to screen 39 lactic acid bacteria, predominantly including oral isolates, for their capacity to accelerate gingival re-epithelialization. This screen identified several strains of Streptococcus salivarius that stimulated re-epithelialization. Further analysis revealed that S. salivarius strain MS-oral-D6 significantly promoted re-epithelialization through a secreted proteinaceous compound and subsequent experiments identified a secreted serine protease as the most likely candidate to be involved in re-epithelialization stimulation. The identification of bacteria or their products that stimulate gingival wound repair may inspire novel strategies for the maintenance of oral health.

Influence of Oral Probiotic Streptococcus salivarius K12 on Ear and Oral Cavity Health in Humans: Systematic Review.

Traditionally, probiotics are linked to the good health of the intestine and most clinical studies focus on that field. Evidence of oral probiotic use for ear and oral cavity disease prevention with impact on human health is limited. This work reviews existing studies and literature on Streptococcus salivarius K12 as an oral probiotic and effects of S. salivarius K12 on human ear and oral cavity human health. The studies were accessed via database searches: MEDLINE, PubMed, and Elsevier. The search included/focused on/encompassed publications from 2003 to 2016 with keywords related to K12 Streptococcus salivarius, bacteriocin-like inhibitory substances (BLIS) K12, probiotic K12 salivarius, and K12 probiotic health effects. Only a small amount of studies was identified: the total of 68 studies was identified, 35 of which were relevant after screening, and 9 were included in the final analysis. Very little literature is available about the association/correlation between/connection/interrelation of S. salivarius K12 with/and human ear and oral cavity health. S. salivarius K12 may have a role in reducing the occurrence and/or severity of secretory otitis media (SOM) and also in prevention of streptococcal and viral pharyngotonsillitis in children. Research highlights that S. salivarius K12 has shown promising results in treatment of halitosis, but data are still deficient. Further studies need to be initiated to improve understanding of the association of oral probiotic S. salivarius K12 with human ear and oral cavity health.

Oral cavities of healthy infants harbour high proportions of Streptococcus salivarius strains with phenotypic and genotypic resistance to multiple classes of antibiotics.

Emerging antibiotic resistance in the oropharyngeal microbiota, of which Streptococcus salivarius is a prominent species, represents a challenge for treating paediatric populations. In this study, we investigated the role of Streptococcussalivarius as a reservoir for antibiotic resistance genes (ARG) in the oral microbiota by analysing 95 Streptococcussalivarius isolates from 22 healthy infants (2-16 months of age). MICs of penicillin G, amoxicillin, erythromycin, tetracycline, doxycycline and streptomycin were determined. ARG profiles were assessed in a subset of 21 strains by next-generation sequencing of genomes, followed by searches of assembled reads against the Comprehensive Antibiotic Resistance Database. Strains resistant to erythromycin, penicillins and tetracyclines were isolated from 83.3, 33.3 and 16.6 %, respectively, of infants aged 2 to 8 months with no prior antibiotic treatment. These percentages were100.0, 66.6 and 50.0 %, by 13 to 16 months of age. ARG or polymorphisms associated with antibiotic resistance were the most prevalent and involved genes for macrolide efflux (mel, mefA/E and macB), ribosomal protection [erm(B), tet(M) and tet(O)] and ß-lactamase-like proteins. Phylogenetically related strains showing multidrug-resistant phenotypes harboured multidrug efflux ARG. Polymorphic genes associated with antibiotic resistance to drugs affecting DNA replication, folate synthesis, RNA/protein synthesis and regulators of antibiotic stress responses were detected. These data imply that Streptococcussalivarius strains established during maturation of the oral microbiota harbour a diverse array of functional ARG, even in the absence of antibiotic selective pressures, highlighting a potential role for this species in shaping antibiotic susceptibility profiles of oropharyngeal communities.