The identification of relevant virus-host pairs that globally account for a large pool of carbon and nutrients in the ocean is paramount to build accurate ecological models. A previous work using single-virus genomics led to the discovery of the uncultured single-virus vSAG 37-F6, originally sorted from the Mediterranean Sea (Blanes Bay Microbial Observatory), that represents one of the most abundant dsDNA viral population in the marine surface virosphere. Here, from same sampling site, we report that a Pelagibacter single-cell contained a viral member of vSAG 37-F6 population, by means of PCR screening of sorted, genome-amplified single cells with vSAG 37-F6-specific primers and whole-genome sequencing. Furthermore, viruses from this population were also found in three other Pelagibacter single cells from the South Pacific and Atlantic oceans. These new uncultured pelagiphages were genetically different from the previously characterized pelagiphage isolates. Data showed that the uncultured vSAG 37-F6 population represents the Pelagibacter phages that inhabit the sunlit ocean better, and contains a vast unrecognized microdiversity.

Summary Wastewater treatment plants effluents are considered as hotspots for the dispersion of antibiotic resistance genes (ARGs) into natural ecosystems. The bacterial resistome (ARG collection in a metagenome) analyses have provided clues on antibacterial resistance dynamics. However, viruses and vesicles are frequently ignored. Here, we addressed the bacterial, viral and vesicle resistomes from a representative wastewater effluent in natural conditions and amended with polymyxin, which is used as a last resort antibiotic. Metagenomics showed that the natural prokaryotic resistome was vast (9000 ARG hits/Gb metagenome) and diverse, while viral resistome was two orders of magnitude lower (50 ARG hits/Gb metagenome) suggesting that viruses rarely encoded ARGs. After polymyxin amendment, data showed no ARG enrichment – including to polymyxin – in the microbiome. Remarkably, microbiomes responded to polymyxin with a vast release of putative vesicles (threefold increase compared with the control), which might be used as 'traps' to decrease the antibiotic concentration. Intriguingly, although polymyxin resistance genes (PRGs) were rare in the microbiome (0.018% of total ARG found), in the viral and vesicle fractions, PRGs were more abundant (0.5%–0.8% of total ARG found). Our data suggest that vesicles could have a more active role in the context of transmission of antibiotic resistances.

Absolute abundances of prokaryotes are typically determined by FISH. Due to the lack of a universal conserved gene among all viruses, metagenomic fragment recruitment is commonly used to estimate the relative viral abundance. However, the paucity of absolute virus abundance data hinders our ability to fully understand how viruses drive global microbial populations. The cosmopolitan marine Pelagibacter ubique is host for the highly widespread HTVC010P pelagiphage isolate and the extremely abundant uncultured virus vSAG 37-F6 recently discovered by single-virus genomics. Here we applied droplet digital PCR (ddPCR) to calculate the absolute abundance of these pelagiphage genotypes in the Mediterranean Sea and the Gulf of Maine. Abundances were between 360 and 8,510 virus mL-1 and 1,270–14,400 virus mL-1 for vSAG 37-F6 and HTVC010P, respectively. Illumina PCR-amplicon sequencing corroborated the absence of ddPCR non-specific amplifications for vSAG 37-F6, but showed an overestimation of 6% for HTVC010P from off-targets, genetically unrelated viruses. Absolute abundances of both pelagiphages, two of the most abundance marine viruses, suggest a large viral pelagiphage diversity in marine environments, and show the efficiency and power of ddPCR to disentangle the structure of marine viral communities. Results also highlight the need for a standardized workflow to obtain accurate quantification that allows cross data comparison.

A high-fat diet (HFD) can induce hyperglycemia and metabolic syndromes that, in turn, can trigger visual impairment. To evaluate the acute effects of HFD feeding on retinal degeneration, we assessed retinal function and morphology, inflammatory state, oxidative stress, and gut microbiome in dystrophic retinal degeneration 10 (rd10) mice, a model of retinitis pigmentosa, fed an HFD for 2 to 3 wk. Short-term HFD feeding impaired retinal responsiveness and visual acuity and enhanced photoreceptor degeneration, microglial cell activation, and Müller cell gliosis. HFD consumption also triggered the expression of inflammatory and oxidative markers in rd10 retinas. Finally, an HFD caused gut microbiome dysbiosis, increasing the abundance of potentially proinflammatory bacteria. Thus, HFD feeding drives the pathological processes of retinal degeneration by promoting oxidative stress and activating inflammatory-related pathways. Our findings suggest that consumption of an HFD could accelerate the progression of the disease in patients with retinal degenerative disorders.

The gut microbiome is known to influence the pathogenesis and progression of neurodegenerative diseases. However, there has been relatively little focus upon the implications of the gut microbiome in retinal diseases such as retinitis pigmentosa (RP). Here, we investigated changes in gut microbiome composition linked to RP, by assessing both retinal degeneration and gut microbiome in the rd10 mouse model of RP as compared to control C57BL/6J mice. In rd10 mice, retinal responsiveness to flashlight stimuli and visual acuity were deteriorated with respect to observed in age-matched control mice. This functional decline in dystrophic animals was accompanied by photoreceptor loss, morphologic anomalies in photoreceptor cells and retinal reactive gliosis. Furthermore, 16S rRNA gene amplicon sequencing data showed a microbial gut dysbiosis with differences in alpha and beta diversity at the genera, species and amplicon sequence variants (ASV) levels between dystrophic and control mice. Remarkably, four fairly common ASV in healthy gut microbiome belonging to Rikenella spp., Muribaculaceace spp., Prevotellaceae UCG-001 spp., and Bacilli spp. were absent in the gut microbiome of retinal disease mice, while Bacteroides caecimuris was significantly enriched in mice with RP. The results indicate that retinal degenerative changes in RP are linked to relevant gut microbiome changes. The findings suggest that microbiome shifting could be considered as potential biomarker and therapeutic target for retinal degenerative diseases.

Technologies to detect the entire bacterial diversity spectra and foodborne pathogens in food represent a fundamental advantage in the control of foodborne illness. Here, we applied high-throughput 16S rRNA sequencing of amplicons obtained by PCR and RT-PCR from extracted DNA and RNA targeting the entire bacterial community and the active bacterial fraction present in some of the most consumed and distributed ready-to-eat (RTE) salad brands in Europe. Customer demands for RTE food are increasing worldwide along with the number of associated foodborne illness and outbreaks. The total aerobic bacterial count in the analyzed samples was in the range of 2–4 × 106 CFU/g (SD ± 1.54 × 106). Culture validated methods did not detect Salmonella spp., Escherichia coli, and other fecal coliforms. 16S rRNA gene Illumina next-generation sequencing (NGS) data were congruent with these culture-based results and confirmed that these and other well-known foodborne bacterial pathogens, such as Listeria, were not detected. However, the fine-resolution of the NGS method unveiled the presence of the opportunistic pathogens Aeromonas hydrophyla and Rahnella aquatilis (relative frequency of 1.33–7.33%) that were metabolically active in addition to non-pathogenic, active members of Yersinia spp. (relative frequency of 0.0015–0.003%). The common ail and foxA marker genes of Yersinia enterocolitica were not detected by qPCR. Finally, our NGS data identified to non-pathogenic Pseudomonas spp. as the most abundant and metabolically active bacteria in the analyzed RTE salads (53–75% of bacterial abundance). Our data demonstrate the power of sequencing, in parallel, both 16S rRNA and rDNA to identify and discriminate those potentially and metabolically active bacteria and pathogens to provide a more complete view that facilitates the control of foodborne diseases, although further work should be conducted to determine the sensitivity of this method for targeting bacteria

Background: The gut microbiome is known to influence the pathogenesis and progression of neurodegenerative diseases. However, there has been relatively little focus upon the implications of the gut microbiome in retinal diseases such as retinitis pigmentosa (RP) that leads to photoreceptor degeneration and is the main worldwide cause of complete blindness in adulthood. Here, we investigated changes in gut microbiome composition linked to RP, by assessing both retinal degeneration and gut microbiome in the rd10 mouse model of RP as compared to control C57BL/6J mice.Results: In rd10 mice, retinal responsiveness to flashlight stimuli was deteriorated with respect to observed in age-matched control mice, with decreased amplitudes in the a- and b-wave responses of the electroretinogram and concomitant reduction of the visual acuity. This functional decline in dystrophic animals was accompanied by photoreceptor loss, evidenced by decreased outer nuclear layer thickness, and morphologic anomalies in photoreceptor cells. Changes in retinal neurons were paralleled to induction of reactive gliosis in retina, with increased microglial cell numbers and higher Müller cell reactivity in rd10 mice. Furthermore, 16S rRNA gene amplicon sequencing data showed a microbial gut dysbiosis with differences in alpha and beta diversity at the genera, species and amplicon sequence variants (ASV) levels between dystrophic and control mice. A taxonomic partitioning of ASV was observed since unique ASVs present in only one group had a cumulative relative microbial abundance between 17.6% (rd10 mice) and 26.7% (C57BL/6J mice). Remarkably, four fairly common ASV in healthy gut microbiome belonging to -Rikenella spp., Muribaculaceace spp., Prevotellaceae UCG-001 spp., and Bacilli spp.- were absent in the gut microbiome of retinal disease mice, while Bacteroides caecimuris was significantly enriched in mice with retinitis pigmentosa. Conclusions: Our results indicate that retinal degenerative changes in retinitis pigmentosa are linked to relevant gut microbiome changes. The findings suggest that microbiome shifting could be considered as potential biomarker and therapeutic target for retinal degenerative diseases.

A comprehensive characterization of the human body resistome [sets of antibiotic resistance genes (ARGs)] is yet to be done and paramount for addressing the antibiotic microbial resistance threat. Here, we study the resistome of 771 samples from five major body parts (skin, nares, vagina, gut, and oral cavity) of healthy subjects from the Human Microbiome Project (HMP) and addressed the potential dispersion of ARGs in pristine environments. A total of 28,714 ARGs belonging to 235 different ARG types were found in the HMP proteome dataset (n = 9.1 × 107 proteins analyzed). Our study reveals a distinct resistome profile (ARG type and abundance) between body sites and high interindividual variability. Nares had the highest ARG load (≈5.4 genes/genome) followed by the oral cavity, whereas the gut showed one of the highest ARG richness (shared with nares) but the lowest abundance (≈1.3 genes/genome). The fluroquinolone resistance genes were the most abundant in the human body, followed by macrolide–lincosamide–streptogramin (MLS) or tetracycline. Most ARGs belonged to common bacterial commensals and multidrug resistance trait were predominant in the nares and vagina. Many ARGs detected here were considered as low risk for human health, whereas only a few of them, such as BlaZ, dfrA14, dfrA17, or tetM, were classified as high-risk ARG. Our data also provide hope, since the spread of common ARG from the human body to pristine environments (n = 271 samples; 77 Gb of sequencing data and 2.1 × 108 proteins analyzed) thus far remains very unlikely (only one case found in an autochthonous bacterium from a pristine environment). These findings broaden our understanding of ARG in the context of the human microbiome and the One-Health Initiative of WHO uniting human host–microbes and environments as a whole.