Background The microbiome of the oral cavity is the second-largest and diverse microbiota after the gut, harboring over 700 species of bacteria and including also fungi, viruses, and protozoa. With its diverse niches, the oral cavity is a very complex environment, where different microbes preferentially colonize different habitats. Recent data indicate that the oral microbiome has essential functions in maintaining oral and systemic health, and the emergence of 16S rRNA gene next-generation sequencing (NGS) has greatly contributed to revealing the complexity of its bacterial component. However, a detailed site-specific map of oral microorganisms (including also eukaryotes and viruses) and their relative abundance is still missing. Here, we aimed to obtain a comprehensive view of the healthy oral microbiome (HOM), including its drug-resistance features. Results The oral microbiome of twenty healthy subjects was analyzed by whole-genome sequencing (WGS) and real-time quantitative PCR microarray. Sampled oral micro-habitat included tongue dorsum, hard palate, buccal mucosa, keratinized gingiva, supragingival and subgingival plaque, and saliva with or without rinsing. Each sampled oral niche evidenced a different microbial community, including bacteria, fungi, and viruses. Alpha-diversity evidenced significant differences among the different sampled sites (p < 0.0001) but not among the enrolled subjects (p = 0.876), strengthening the notion of a recognizable HOM. Of note, oral rinse microbiome was more representative of the whole site-specific microbiomes, compared with that of saliva. Interestingly, HOM resistome included highly prevalent genes conferring resistance to macrolide, lincosamides, streptogramin, and tetracycline. Conclusions The data obtained in 20 subjects by WGS and microarray analysis provide for the first time a comprehensive view of HOM and its resistome, contributing to a deeper understanding of the composition of oral microbiome in the healthy subject, and providing an important reference for future studies, allowing to identify microbial signatures related to functional and metabolic alterations associated with diseases, potentially useful for targeted therapies and precision medicine.

The human oral microbiome (HOM) is the second largest microbial community after the gut and can impact the onset and progression of several localized and systemic diseases, including those of viral origin, especially for viruses entering the body via the oropharynx. However, this important aspect has not been clarified for the new pandemic human coronavirus SARS-CoV-2, causing COVID-19 disease, despite it being one of the many respiratory viruses having the oropharynx as the primary site of replication. In particular, no data are available about the non-bacterial components of the HOM (fungi, viruses), which instead has been shown to be crucial for other diseases. Consistent with this, this study aimed to define the HOM in COVID-19 patients, to evidence any association between its profile and the clinical disease. Seventy-five oral rinse samples were analyzed by Whole Genome Sequencing (WGS) to simultaneously identify oral bacteria, fungi, and viruses. To correlate the HOM profile with local virus replication, the SARS-CoV-2 amount in the oral cavity was quantified by digital droplet PCR. Moreover, local inflammation and secretory immune response were also assessed, respectively by measuring the local release of pro-inflammatory cytokines (L-6, IL-17, TNFα, and GM-CSF) and the production of secretory immunoglobulins A (sIgA). The results showed the presence of oral dysbiosis in COVID-19 patients compared to matched controls, with significantly decreased alpha-diversity value and lower species richness in COVID-19 subjects. Notably, oral dysbiosis correlated with symptom severity (p = 0.006), and increased local inflammation (p &lt; 0.01). In parallel, a decreased mucosal sIgA response was observed in more severely symptomatic patients (p = 0.02), suggesting that local immune response is important in the early control of virus infection and that its correct development is influenced by the HOM profile. In conclusion, the data presented here suggest that the HOM profile may be important in defining the individual susceptibility to SARS-CoV-2 infection, facilitating inflammation and virus replication, or rather, inducing a protective IgA response. Although it is not possible to determine whether the alteration in the microbial community is the cause or effect of the SARS-CoV-2 replication, these parameters may be considered as markers for personalized therapy and vaccine development.

Purpose Antimicrobial resistance (AMR) is one of the major threats to human health, and the high frequency of resistant pathogens in the hospital environment can contribute to the transmission of difficult-to-treat health care-associated infections (HAIs). We recently reported that, compared with conventional chemical cleaning, the use of a microbial-based sanitation strategy (Probiotic Cleaning Hygiene System [PCHS]) was associated with remodulation of hospital microbiota and reduction of HAI incidence. Here, we aimed to analyze the impact of PCHS on AMR and related effects, such as HAI-associated antimicrobial drug consumption and costs. Patients and methods Five Italian hospitals, enrolled in a multicenter study where conventional sanitation methods were replaced with PCHS, were included in the analysis. The study period included a 6-month observation for each sanitation type. Surface microbiota AMR was analyzed using microarray, nested PCR, antibiogram, and microdilution tests. Drug consumption data and related costs were obtained from the medical records of all hospitalized patients affected by HAIs. Results PCHS use was associated with up to 99% decrease of the AMR genes harbored by surface hospital microbiota, independently of the resistance types originally present in each individual setting ( P c <0.01). Functional assays confirmed the molecular data, demonstrating a 33%–100% decrease of resistant strains depending on the antibiotic type. Antimicrobial drug consumption associated with HAI onset showed a global 60.3% decrease, with a 75.4% decrease of the associated costs. Conclusion The spread of AMR in the hospital environment can be limited by the use of sanitation methods to remodulate the hospital microbiota, leading to lower antimicrobial consumption and costs. This approach might be considered as part of broader infection prevention and control strategies.

Healthcare-associated infections (HAIs) affect up to 15% of all hospitalized patients, representing a global concern. Major causes include the persistent microbial contamination of hospital environment, and the growing antimicrobial-resistance (AMR) of HAI-associated microbes. The hospital environment represents in fact a reservoir of potential pathogens, continuously spread by healthcare personnel, visiting persons and hospitalized patients. The control of contamination has been so far addressed by the use of chemical-based sanitation procedures, which however have limitations, as testified by the persistence of contamination itself and by the growing AMR of hospital microbes. Here we review the results collected by a microbial-based sanitation system, inspired by the microbiome balance principles, in obtaining more effective control of microbial contamination and AMR. Whatever the sanitation system used, an important aspect of controlling AMR and HAIs relates to the ability to check any variation of a microbial population rapidly and effectively, thus effective monitoring procedures are also described.

PurposeMany hospital-acquired infections (HAIs) can be transmitted by pathogens contaminating hospital surfaces, not efficiently controlled by conventional sanitation, which can indeed contribute to the selection of MDR strains. Bacteriophages have been suggested as decontaminating agents, based on their selective ability to kill specific bacteria. However, there are no data on their stability in detergents and their potential use in routine sanitation. On the other hand, a probiotic-based sanitation system (Probiotic Cleaning Hygiene System, PCHS) was recently shown to stably reduce pathogens on treated surfaces. However, its action is not specific and slow, being based on competitive antagonism. This work aimed to assess the effectiveness of a combined use of phages and PCHS in removing HAI-associated pathogens from different hard surfaces.Materials and methodsThe decontamination ability of phages in PCHS was tested in vitro and in situ, against drug-susceptible or resistant Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa strains, and using bacterial densities similar to those detected on hospital surfaces.ResultsPhages targeted efficiently all tested bacteria, maintaining their full activity when added to the PCHS detergent. Notably, the combined use of phages and PCHS not only resulted in a rapid reduction (up to >90%) of the targeted pathogens, but also, due to the stabilizing effect of probiotics, the pathogens were maintained at low levels (>99%) at later times too, when instead the effect of phages tends to diminish.ConclusionThese results suggest that a combined biological system might be successfully used in hospital sanitation protocols, potentially leading to effective and safe elimination of MDR pathogens from the hospital environment.