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Streptococcus dentisani has been proposed as a promising probiotic against tooth decay, due to its ability to buffer acidic pH and to inhibit the growth of oral pathogens. However, it is unknown if this bacterial species has a global distribution. The current study aimed to establish the presence of S. dentisani in oral samples from different geographic locations by identifying the sequence of its 16S rRNA gene in available datasets from across the globe. In addition, an analytical and cross-sectional study was carried out to determine if the levels of this probiotic strain are higher in caries-free individuals compared to those with dental caries. Samples from various geographical sources demonstrated that S. dentisani is present in saliva and dental plaque from individuals of different continents. Typical S. dentisani levels in saliva ranged from 10(4) to 10(5) cells/ml and a total of 10(6)-10(7) cells in dental plaque. Using real-time qPCR, S. dentisani was quantified from supragingival dental plaque of 25 caries-free and 29 caries-active individuals from a Mexican children population, where significantly higher proportions of S. dentisani were found in the caries-free group (p = 0.002). Finally, a negative correlation was found between caries levels (as measured by the dmft caries index) and the percentage of S. dentisani (p < 0.001). Thus, the current manuscript indicates that this species has a global distribution, can be found in saliva and dental plaque, and appears to be present in higher numbers in plaque samples from caries-free children.

Background Tooth decay is one of the most prevalent diseases worldwide, and efficient tooth brushing with a fluoride-containing dentifrice is considered fundamental to caries prevention. Fluoride-containing dentifrices have been extensively studied in relation to enamel resistance to demineralization. Arginine (Arg) has also been proposed as a promising prebiotic to promote pH buffering through ammonia production. Here, we present the first metagenomic (DNA sequencing of the whole microbial community) and metatranscriptomic (RNAseq of the same community) analyses of human dental plaque to evaluate the effect of brushing with fluoride (Fl) and a Fl+Arg containing dentifrices on oral microbial composition and activity. Fifty-three patients were enrolled in a longitudinal clinical intervention study with two arms, including 26 caries-active and 27 caries-free adults. After a minimum 1-week washout period, dental plaque samples were collected at this post-washout baseline, 3 months after the use of a 1450-ppm fluoride dentifrice, and after 6 months of using a 1450-ppm fluoride with 1.5% arginine dentifrice. Results There was a shift in both the composition and activity of the plaque microbiome after 3 months of brushing with the fluoride-containing toothpaste compared to the samples collected at the 1-week post-washout period, both for caries-active and caries-free sites. Although several caries-associated bacteria were reduced, there was also an increase in several health- and periodontitis-associated bacteria. Over 400 genes changed proportion in the metagenome, and between 180 and 300 genes changed their expression level depending on whether caries-free or caries-active sites were analyzed. The metagenome and metatranscriptome also changed after the subjects brushed with the Fl+Arg dentifrice. There was a further decrease of both caries- and periodontitis-associated organisms. In both caries-free and caries-active sites, a decrease of genes from the arginine biosynthesis pathway was also observed, in addition to an increase in the expression of genes associated with the arginine deiminase pathway, which catabolizes arginine into ammonia, thereby buffering acidic pH. Bacterial richness and diversity were not affected by either of the two treatments in the two arms of the study. Conclusions Our data demonstrate that long-term use of both assayed dentifrices changes the bacterial composition and functional profiles of human dental plaque towards a healthier microbial community, both in caries-free and caries-active sites. This observation was especially apparent for the Fl+Arg dentifrice. Thus, we conclude that the preventive benefits of tooth brushing go beyond the physical removal of dental plaque and that the active ingredients formulated within dentifrices have a positive effect not only on enamel chemistry but also on the metabolism of oral microbial populations.

Bacterial colonization in the oral cavity is critical for efficient action of probiotics. However, limited colonization rates have been reported in many clinical trials. The aim of this pilot clinical study was to evaluate the colonization efficiency of Streptococcus dentisani under different dosing schedules and pre-treatment conditions. Eleven adult volunteers enrolled in the study. A professional ultrasound cleaning was performed in quadrants 1 and 4. The probiotic was applied in all four quadrants at a total dose of 1010 CFUs, administered in a buccoadhesive gel for 5 min, either in a single dose (n = 5) or daily for a week (n = 6). Dental plaque and saliva samples were collected at baseline and after 14 and 28 days of first application. Amounts of S. dentisani and the cariogenic organism Streptococcus mutans were measured by qPCR and salivary pH was measured by reflectometry. There was a significant increase in S. dentisani cells at day 14 but not at day 28 under both dosing schedules. A non-significant higher colonization was found in the half-mouth with previous professional cleaning as compared to the intact half. There was a significant increase in salivary pH at day 14 (p = 0.024) and day 28 (p = 0.014), which was stronger in multi-dose patients, and a significant decrease in S. mutans at day 28 (p

Human milk represents a source of bacteria for the initial establishment of the oral (and gut) microbiomes in the breastfed infant, however, the origin of bacteria in human milk remains largely unknown. While some evidence points towards a possible endogenous enteromammary route, other authors have suggested that bacteria in human milk are contaminants from the skin or the breastfed infant mouth. In this work 16S rRNA sequencing and bacterial culturing and isolation was performed to analyze the microbiota on maternal precolostrum samples, collected from pregnant women before delivery, and on oral samples collected from the corresponding infants. The structure of both ecosystems demonstrated a high proportion of taxa consistently shared among ecosystems, Streptococcus spp. and Staphylococcus spp. being the most abundant. Whole genome sequencing on those isolates that, belonging to the same species, were isolated from both the maternal and infant samples in the same mother-infant pair, evidenced that in 8 out of 10 pairs both isolates were >99.9% identical at nucleotide level. The presence of typical oral bacteria in precolostrum before contact with the newborn indicates that they are not a contamination from the infant, and suggests that at least some oral bacteria reach the infant’s mouth through breastfeeding., This research was supported by grant AGL2016-75476-R. LR was funded by grant 624773 (FP-7-PEOPLE-2013IEF, European Commission) and is currently supported by the Juan de la Cierva Postdoctoral Trainee Program of the Spanish Ministry of Economy and Competitiveness (MINECO; IJCI-2015-23196).

Human breastmilk contains a variety of bacteria that are transmitted to the infant and have been suggested to contribute to gut microbiota development and immune maturation. However, the characterization of fungal organisms in milk from healthy mothers is currently unknown although their presence has been reported in the infant gut and also in milk from other mammals. Breastmilk samples from healthy lactating mothers (n = 65) within 1 month after birth were analyzed. Fungal presence was assessed by different techniques, including microscopy, growth and identification of cultured isolates, fungal load estimation by qPCR, and fungal composition using 28S rRNA gene high-throughput sequencing. In addition, milk macronutrients and human somatic cells were quantified by spectrophotometry and cytometry. qPCR data showed that 89% of samples had detectable levels of fungal DNA, at an estimated median load of 3,5 × 10

Epidemiological studies demonstrate a link between gastrointestinal cancers and environmental factors such as diet. It has been suggested that environmental cancer risk is determined by the interaction between diet and microbes. Thus, the purpose of this study was to examine the hypothesis that microbiota composition during colorectal cancer (CRC) progression might differ depending on the stage of the disease. A total of 28 age-matched and sex-matched subjects, seven with CRC adenocarcinoma, 11 with tubular adenomas and ten healthy subjects with intact colon, were included into the study. Microbiomes from mucosal and fecal samples were analyzed with 16S ribosomal RNA gene pyrosequencing, together with quantitative PCR of specific bacteria and archaea. The principal coordinates analysis clearly separated healthy tissue samples from polyps and tumors, supporting the presence of specific bacterial consortia that are associated with affected sites and that can serve as potential biomarkers of CRC progression. A higher presence of Fusobacterium nucleatum and Enterobacteriaceae was found by qPCR in samples from CRC compared to healthy controls. We observed a correlation between CRC process development and levels of Methanobacteriales (R = 0.537, P = 0.007) and Methanobrevibacterium (R = 0.574, P = 0.03) in fecal samples. Differences in microbial and archaeal composition between mucosal samples from healthy and disease tissues were observed in tubular adenoma and adenocarcinoma. In addition, microbiota from mucosal samples represented the underlying dysbiosis, whereas fecal samples seem not to be appropriate to detect shifts in microbial composition. CRC risk is influenced by microbial composition, showing differences according to disease progression step and tumor severity.

The oral cavity of humans is inhabited by hundreds of bacterial species and some of them have a key role in the development of oral diseases, mainly dental caries and periodontitis. We describe for the first time the metagenome of the human oral cavity under health and diseased conditions, with a focus on supragingival dental plaque and cavities. Direct pyrosequencing of eight samples with different oral-health status produced 1 Gbp of sequence without the biases imposed by PCR or cloning. These data show that cavities are not dominated by Streptococcus mutans (the species originally identified as the ethiological agent of dental caries) but are in fact a complex community formed by tens of bacterial species, in agreement with the view that caries is a polymicrobial disease. The analysis of the reads indicated that the oral cavity is functionally a different environment from the gut, with many functional categories enriched in one of the two environments and depleted in the other. Individuals who had never suffered from dental caries showed an over-representation of several functional categories, like genes for antimicrobial peptides and quorum sensing. In addition, they did not have mutans streptococci but displayed high recruitment of other species. Several isolates belonging to these dominant bacteria in healthy individuals were cultured and shown to inhibit the growth of cariogenic bacteria, suggesting the use of these commensal bacterial strains as probiotics to promote oral health and prevent dental caries.

The deep chlorophyll maximum (DCM) is a zone of maximal photosynthetic activity, generally located toward the base of the photic zone in lakes and oceans. In the tropical waters, this is a permanent feature, but in the Mediterranean and other temperate waters, the DCM is a seasonal phenomenon. The metagenome from a single sample of a mature Mediterranean DCM community has been 454 pyrosequenced both directly and after cloning in fosmids. This study is the first to be carried out at this sequencing depth (ca. 600 Mb combining direct and fosmid sequencing) at any DCM. Our results indicate a microbial community massively dominated by the high-light-adapted Prochlorococcus marinus subsp. pastoris, Synechococcus sp., and the heterotroph Candidatus Pelagibacter. The sequences retrieved were remarkably similar to the existing genome of P. marinus subsp. pastoris with a nucleotide identity over 98%. Besides, we found a large number of cyanophages that could prey on this microbe, although sequence conservation was much lower. The high abundance of phage sequences in the cellular size fraction indicated a remarkably high proportion of cells suffering phage lytic attack. In addition, several fosmids clearly belonging to Group II Euryarchaeota were retrieved and recruited many fragments from the total direct DNA sequencing suggesting that this group might be quite abundant in this habitat. The comparison between the direct and fosmids sequencing revealed a bias in the fosmid libraries against low-GC DNA and specifically against the two most dominant members of the community, Candidatus Pelagibacter and P. marinus subsp. pastoris, thus unexpectedly providing a feasible method to obtain large genomic fragments from other less prevalent members of this community.

Not all isolates of a species contain the same set of genes. In this Opinion article, Rodriguez-Valera and colleagues propose the constant-diversity model to account for these differences. In this model, predation by phages promotes bacterial diversity and allows more efficient use of the nutrients in the environment. The remarkable differences that have been detected by metagenomics in the genomes of strains of the same bacterial species are difficult to reconcile with the widely accepted paradigm that periodic selection within bacterial populations will regularly purge genomic diversity by clonal replacement. We have found that many of the genes that differ between strains affect regions that are potential phage recognition targets. We therefore propose the constant-diversity dynamics model, in which the diversity of prokaryotic populations is preserved by phage predation. We provide supporting evidence for this model from metagenomics, mathematical analysis and computer simulations. Periodic selection and phage predation dynamics are not mutually exclusive; we compare their predictions to shed light on the ecological circumstances under which each type of dynamics could predominate.

Background The phylogenetic position and evolutionary relationships of Fusobacteria remain uncertain. Especially intriguing is their relatedness to low G+C Gram positive bacteria (Firmicutes) by ribosomal molecular phylogenies, but their possession of a typical gram negative outer membrane. Taking advantage of the recent completion of the Fusobacterium nucleatum genome sequence we have examined the evolutionary relationships of Fusobacterium genes by phylogenetic analysis and comparative genomics tools.

Background: The wealth of genomic data in bacteria is helping microbiologists understand the factors involved in gene innovation. Among these, the expansion and reduction of gene families appears to have a fundamental role in this, but the factors influencing gene family size are unclear. Results: The relative content of paralogous genes in bacterial genomes increases with genome size, largely due to the expansion of gene family size in large genomes. Bacteria undergoing genome reduction display a parallel process of redundancy elimination, by which gene families are reduced to one or a few members. Gene family size is also influenced by sequence divergence and physiological function. Large gene families show wider sequence divergence, suggesting they are probably older, and certain functions (such as metabolite transport mechanisms) are overrepresented in large families. The size of a given gene family is remarkably similar in strains of the same species and in closely related species, suggesting that homologous gene families are vertically transmitted and depend little on horizontal gene transfer (HGT). Conclusions: The remarkable preservation of copy numbers in widely different ecotypes indicates a functional role for the different copies rather than simply a back-up role. When different genera are compared, the increase in phylogenetic distance and/or ecological specialization disrupts this preservation, albeit in a gradual manner and maintaining an overall similarity, which also supports this view. HGT can have an important role, however, in nonhomologous gene families, as exemplified by a comparison between saprophytic and enterohemorrhagic strains of Escherichia coli.

Very small genomes have evolved repeatedly in eubacterial lineages that have adopted obligate associations with eukaryotic hosts. Complete genome sequences have revealed that small genomes retain very different gene sets, raising the question of how final genome content is determined. To examine the process of genome reduction, the tiny genome of the endosymbiont Buchnera aphidicola was compared to the larger ancestral genome, reconstructed on the basis of the phylogenetic distribution of gene orthologs among fully sequenced relatives of Escherichia coli and Buchnera. The reconstructed ancestral genome contained 2,425 open reading frames (ORFs). The Buchnera genome, containing 564 ORFs, consists of 153 fragments of 1-34 genes that are syntenic with reconstructed ancestral regions. On the basis of this reconstruction, 503 genes were eliminated within syntenic fragments, and 1,403 genes were lost from the gaps between syntenic fragments, probably in connection with genome rearrangements. Lost regions are sometimes large, and often span functionally unrelated genes. In addition, individual genes and regulatory regions have been lost or eroded. For the categories of DNA repair genes and rRNA genes, most lost loci fall in regions between syntenic fragments. This history of gene loss is reflected in the sequences of intergenic spacers at positions where genes were once present. The most plausible interpretation of this reconstruction is that Buchnera lost many genes through the fixation of large deletions soon after the acquisition of an obligate endosymbiotic lifestyle. An implication is that final genome composition may be partly the chance outcome of initial deletions and that neighboring genes influence the likelihood of loss of particular genes and pathways.