Distribution of prophages in the oenococcus oeni species

Oenococcus oeni is the most exploited lactic acid bacterium in the wine industry and drives the malolactic fermentation of wines. Although prophage-like sequences have been identified in the species, many are not characterized, and a global view of their integration and distribution amongst strains is currently lacking. In this work, we analyzed the complete genomes of 231 strains for the occurrence of prophages, and analyzed their size and positions of insertion. Our data show the limited variation in the number of prophages in O. oeni genomes, and that six sites of insertion within the bacterial genome are being used for site-specific recombination. Prophage diversity patterns varied significantly for different host lineages, and environmental niches. Overall, the findings highlight the pervasive presence of prophages in the O. oeni species, their role as a major source of within-species bacterial diversity and drivers of horizontal gene transfer. Our data also have implications for enhanced understanding of the prophage recombination events which occurred during evolution of O. oeni, as well as the potential of prophages in influencing the fitness of these bacteria in their distinct niches.Evolution expérimentale en mileu extrême de la bactérie lactique Oenococcus oeni et applications à la sélection de levains malo-lactiques plus performant

Bioversity of oenococci in fermented beverages et deciphering of their interactions with cognate bacteriophages.

Les bactéries à Gram positif de l’espèce Oenococcus oeni sont des coques fermentaires présents à la surface des baies de raisin et sur les équipements des chais. Ces bactéries lactiques sont connues pour leur rôle dans la fermentation malolactique des vins. L’intérêt croissant pour les boissons fermentées a encouragé les inventaires moléculaires de leur microbiote. Ces travaux ont révélé l’existence de niches supplémentaires pour O. oeni comprenant le cidre, ainsi que d’autres boissons à teneur plus réduite en alcool comme le kombucha et le kéfir. Ils ont également mis en lumière l’existence de trois nouvelles espèces dans le genre Oenococcus. Toutefois, malgré la multiplication des études consacrées aux écosystèmes alimentaires microbiens ces dernières années, un nombre limité de souches issues de niches autres que le vin a été isolé et caractérisé dans le genre Oenococcus. Les relations entre les espèces qui sont proches d’un point de vue phylogénétique, mais exploitent des niches différentes, restent donc peu caractérisées. Dans ce contexte, ce projet de thèse a eu pour objectif de mieux comprendre la contribution des phages dans l’évolution des oenocoques et leur biologie. Dans ce travail, nous avons (i) isolé des oenocoques à partir de kéfirs d’eau et de kombucha et décrypté leur génome (ii) évalué si les oenocoques affiliés à des espèces différentes sont connectés via leur phages en explorant les loci CRISPR bactériens et enfin (iii) caractérisé les modalités d’interaction entre le phage lytique X27 et ses bactéries hôtes. L’ensemble de nos résultats suggèrent des traces d’infection entre les phages d’O. oeni et O. sicerae. Par ailleurs, nous avons amélioré nos connaissances sur les modalités de recombinaison entre génomes phagiques et précisé la structure de la partie caudale responsable de la reconnaissance de l’enveloppe de l’hôte. Ces résultats ouvrent la voie à l’élucidation des interactomes entre les oenocoques et leurs phages.Gram positive strains of the Oenococcus oeni species are fermentative cocci found at the surface of berries and equiments in wineries. Such lactic acid bacteria play a prominent role during the malolactic fermentation of wines. The growing interest for fermented beverages has led to molecular inventories of their microbiota. These approaches have revealed additional niches for O. oeni, of which cider, and other low alcohol beverages such as kombucha and kefir. They also highlighted three novel species in the Oenococcus genus. Yet, despite the number of studies dedicated to food microbial ecosystems during past years, a limited number of strains that did not originate from wine have been isolated and further characterized. The relationships between phylogenetically related species, yet exploiting distinct niches, have so far received little attention. In such context, the objectives of this thesis project were to better understand the contribution of phages in the evolution and biology of oenococci. In this work, we have (i) isolated oenococci from water kefirs and kombucha and deciphered their genomic data (ii) assessed whether oenococci from distinct species are connected through their phages through the exploration of the bacterial CRISPR loci and last (iii) characterized the interactions between the lytic phage X27 and its hosts. Our results suggest traces of infection between phages of O. oeni and O. sicerae. In addition, we improved our knowledge of the mechanisms responsible for recombinaison between phage genomes and detailed the structure of the tail tip which recognizes and bind to the host. Such data open the way to the deciphering of the interactome between the cell wall of oenococci and their cognate phages

Bioversity of oenococci in fermented beverages et deciphering of their interactions with cognate bacteriophages.

Les bactéries à Gram positif de l’espèce Oenococcus oeni sont des coques fermentaires présents à la surface des baies de raisin et sur les équipements des chais. Ces bactéries lactiques sont connues pour leur rôle dans la fermentation malolactique des vins. L’intérêt croissant pour les boissons fermentées a encouragé les inventaires moléculaires de leur microbiote. Ces travaux ont révélé l’existence de niches supplémentaires pour O. oeni comprenant le cidre, ainsi que d’autres boissons à teneur plus réduite en alcool comme le kombucha et le kéfir. Ils ont également mis en lumière l’existence de trois nouvelles espèces dans le genre Oenococcus. Toutefois, malgré la multiplication des études consacrées aux écosystèmes alimentaires microbiens ces dernières années, un nombre limité de souches issues de niches autres que le vin a été isolé et caractérisé dans le genre Oenococcus. Les relations entre les espèces qui sont proches d’un point de vue phylogénétique, mais exploitent des niches différentes, restent donc peu caractérisées. Dans ce contexte, ce projet de thèse a eu pour objectif de mieux comprendre la contribution des phages dans l’évolution des oenocoques et leur biologie. Dans ce travail, nous avons (i) isolé des oenocoques à partir de kéfirs d’eau et de kombucha et décrypté leur génome (ii) évalué si les oenocoques affiliés à des espèces différentes sont connectés via leur phages en explorant les loci CRISPR bactériens et enfin (iii) caractérisé les modalités d’interaction entre le phage lytique X27 et ses bactéries hôtes. L’ensemble de nos résultats suggèrent des traces d’infection entre les phages d’O. oeni et O. sicerae. Par ailleurs, nous avons amélioré nos connaissances sur les modalités de recombinaison entre génomes phagiques et précisé la structure de la partie caudale responsable de la reconnaissance de l’enveloppe de l’hôte. Ces résultats ouvrent la voie à l’élucidation des interactomes entre les oenocoques et leurs phages.Gram positive strains of the Oenococcus oeni species are fermentative cocci found at the surface of berries and equiments in wineries. Such lactic acid bacteria play a prominent role during the malolactic fermentation of wines. The growing interest for fermented beverages has led to molecular inventories of their microbiota. These approaches have revealed additional niches for O. oeni, of which cider, and other low alcohol beverages such as kombucha and kefir. They also highlighted three novel species in the Oenococcus genus. Yet, despite the number of studies dedicated to food microbial ecosystems during past years, a limited number of strains that did not originate from wine have been isolated and further characterized. The relationships between phylogenetically related species, yet exploiting distinct niches, have so far received little attention. In such context, the objectives of this thesis project were to better understand the contribution of phages in the evolution and biology of oenococci. In this work, we have (i) isolated oenococci from water kefirs and kombucha and deciphered their genomic data (ii) assessed whether oenococci from distinct species are connected through their phages through the exploration of the bacterial CRISPR loci and last (iii) characterized the interactions between the lytic phage X27 and its hosts. Our results suggest traces of infection between phages of O. oeni and O. sicerae. In addition, we improved our knowledge of the mechanisms responsible for recombinaison between phage genomes and detailed the structure of the tail tip which recognizes and bind to the host. Such data open the way to the deciphering of the interactome between the cell wall of oenococci and their cognate phages

Phage Encounters Recorded in CRISPR Arrays in the Genus <i>Oenococcus</i>

The Oenococcus genus comprises four recognized species, and members have been found in different types of beverages, including wine, kefir, cider and kombucha. In this work, we implemented two complementary strategies to assess whether oenococcal hosts of different species and habitats were connected through their bacteriophages. First, we investigated the diversity of CRISPR-Cas systems using a genome-mining approach, and CRISPR-endowed strains were identified in three species. A census of the spacers from the four identified CRISPR-Cas loci showed that each spacer space was mostly dominated by species-specific sequences. Yet, we characterized a limited records of potentially recent and also ancient infections between O. kitaharae and O. sicerae and phages of O. oeni, suggesting that some related phages have interacted in diverse ways with their Oenococcus hosts over evolutionary time. Second, phage-host interaction analyses were performed experimentally with a diversified panel of phages and strains. None of the tested phages could infect strains across the species barrier. Yet, some infections occurred between phages and hosts from distinct beverages in the O. oeni species

Draft Genome Sequence of Oenococcus kitaharae CRBO2176, Isolated from Homemade Water Kefir

Here, we announce the draft genome sequence of an Oenococcus kitaharae strain isolated from homemade water kefir in Bordeaux, France. O. kitaharae CRBO2176 is deposited at the Biological Resources Center Oenology (CRBO) of the Institute of Vine and Wine Science (ISVV; Villenave d’Ornon, France).Evolution expérimentale en mileu extrême de la bactérie lactique Oenococcus oeni et applications à la sélection de levains malo-lactiques plus performant

Distribution of Prophages in the Oenococcus oeni Species

Oenococcus oeni is the most exploited lactic acid bacterium in the wine industry and drives the malolactic fermentation of wines. Although prophage-like sequences have been identified in the species, many are not characterized, and a global view of their integration and distribution amongst strains is currently lacking. In this work, we analyzed the complete genomes of 231 strains for the occurrence of prophages, and analyzed their size and positions of insertion. Our data show the limited variation in the number of prophages in O. oeni genomes, and that six sites of insertion within the bacterial genome are being used for site-specific recombination. Prophage diversity patterns varied significantly for different host lineages, and environmental niches. Overall, the findings highlight the pervasive presence of prophages in the O. oeni species, their role as a major source of within-species bacterial diversity and drivers of horizontal gene transfer. Our data also have implications for enhanced understanding of the prophage recombination events which occurred during evolution of O. oeni, as well as the potential of prophages in influencing the fitness of these bacteria in their distinct niches