Phylogenetic and Complementation Analysis of a Single-Stranded DNA Binding Protein Family from Lactococcal Phages Indicates a Non-Bacterial Origin

Background: The single-stranded-nucleic acid binding (SSB) protein superfamily includes proteins encoded by different organisms from Bacteria and their phages to Eukaryotes. SSB proteins share common structural characteristics and have been suggested to descend from an ancestor polypeptide. However, as other proteins involved in DNA replication, bacterial SSB proteins are clearly different from those found in Archaea and Eukaryotes. It was proposed that the corresponding genes in the phage genomes were transferred from the bacterial hosts. Recently new SSB proteins encoded by the virulent lactococcal bacteriophages (Orf14bIL67-like proteins) have been identified and characterized structurally and biochemically. Methodology/Principal Findings: This study focused on the determination of phylogenetic relationships between Orf14bIL67-like proteins and other SSBs. We have performed a large scale phylogenetic analysis and pairwise sequence comparisons of SSB proteins from different phyla. The results show that, in remarkable contrast to other phage SSBs, the Orf14bIL67–like proteins form a distinct, self-contained and well supported phylogenetic group connected to the archaeal SSBs. Functional studies demonstrated that, despite the structural and amino acid sequence differences from bacterial SSBs, Orf14bIL67 protein complements the conditional lethal ssb-1 mutation of Escherichia coli. Conclusions/Significance: Here we identified for the first time a group of phages encoded SSBs which are clearly distinct from their bacterial counterparts. All methods supported the recognition of these phage proteins as a new family within the SSB superfamily. Our findings suggest that unlike other phages, the virulent lactococcal phages carry ssb genes that were not acquired from their hosts, but transferred from an archaeal genome. This represents a unique example of a horizontal gene transfer between Archaea and bacterial phages

Expanding Diversity of Firmicutes Single-Strand Annealing Proteins: a Putative Role of Bacteriophage-Host Arms Race

Bacteriophage-encoded single strand annealing proteins (SSAPs) are recombinases which can substitute the classical, bacterial RecA and manage the DNA metabolism at different steps of phage propagation. SSAPs have been shown to efficiently promote recombination between short and rather divergent DNA sequences and were exploited for in vivo genetic engineering mainly in Gram-negative bacteria. In opposition to the conserved and almost universal bacterial RecA protein, SSAPs display great sequence diversity. The importance for SSAPs in phage biology and phage-bacteria evolution is underlined by their role as key players in events of horizontal gene transfer (HGT). All of the above provoke a constant interest for the identification and study of new phage recombinase proteins in vivo, in vitro as well as in silico. Despite this, a huge body of putative ssap genes escapes conventional classification, as they are not properly annotated. In this work, we performed a wide-scale identification, classification and analysis of SSAPs encoded by the Firmicutes bacteria and their phages. By using sequence similarity network and gene context analyses, we created a new high quality dataset of phage-related SSAPs, substantially increasing the number of annotated SSAPs. We classified the identified SSAPs into seven distinct families, namely RecA, Gp2.5, RecT/Redb, Erf, Rad52/22, Sak3, and Sak4, organized into three superfamilies. Analysis of the relationships between the revealed protein clusters led us to recognize Sak3-like proteins as a new distinct SSAP family. Our analysis showed an irregular phylogenetic distribution of ssap genes among different bacterial phyla and specific phages, which can be explained by the high rates of ssap HGT. We propose that the evolution of phage recombinases could be tightly linked to the dissemination of bacterial phage-resistance mechanisms (e.g., abortive infection and CRISPR/Cas systems) targeting ssap genes and be a part of the constant phage-bacteria arms race

An examination of the bacteriophages and bacteria of the Namib desert.

International audienceBacteria and their viruses (called bacteriophages, or phages), have been found in virtually every ecological niche on Earth. Arid regions, including their most extreme form called deserts, represent the single largest ecosystem type on the Earth's terrestrial surface. The Namib desert is believed to be the oldest (80 million years) desert. We report here an initial analysis of bacteriophages isolated from the Namib desert using a combination of electron microscopy and genomic approaches. The virus-like particles observed by electron microscopy revealed 20 seemingly different phage-like morphologies and sizes belonging to the Myoviridae and Siphoviridae families of tailed phages. Pulsed-field gel electrophoresis revealed a majority of phage genomes of 55-65 kb in length, with genomes of approximately 200, 300, and 350 kb also observable. Sample sequencing of cloned phage DNA fragments revealed that approximately 50% appeared to be of bacterial origin. Of the remaining DNA sequences, approximately 50% displayed no significant match to any sequence in the databases. The majority of the 16S rDNA sequences amplified from DNA extracted from the sand displayed considerable (94-98%) homology to members of the Firmicutes, and in particular to members of the genus Bacillus, though members of the Bacteroidetes, Planctomycetes, Chloroflexi, and delta-Proteobacteria groups were also observed

A novel bacteriophage morphotype with a ribbon-like structure at the tail extremity

7 pagesInternational audienceWe have isolated a novel Siphoviridae phage (named Sol-P11) morphotype from the surface sands of the Sahara Desert with a ribbon-like structure at the tail extremity. Sol-P11 was found to grow on a Bacillus subtilis strain isolated from the same environment and to contain a double stranded DNA genome of approximately 120 kb in length incapable of being hydrolysed by a wide variety of restriction endonucleases. The major constituent proteins of CsCl-purified Sol-P11 virions were 65, 50, 30, and 24 kDa in size, with the 30 kDa polypeptide being the major protein of the 85 nm diameter icosahedral capsid, and the other three proteins comprising the major polypeptides of the tail (320 nm in length) and ribbon-like structure. Moreover, different sized phages displaying a Sol-P11 morphology were observed in phage preparations from the Death Valley and Namib deserts. Sol-P11-like phage morphotypes have been previously described, including PBPI, a flagellum-specific phage that infects B. pumilis and phage BcP15 infecting the marine bacterium, Burkholderia cepacia DR11. We thus propose that Sol-P11 represents a member of a novel morphotype of Siphoviridae phages that use a ribbon-like structure, instead of caudal fibers, to attach to their host cell

Variations of bacterial 16S rDNA phylotypes prior to and after chlorination for drinking water production from two surface water treatment plants.

International audienceWe examined the variations of bacterial populations in treated drinking water prior to and after the final chlorine disinfection step at two different surface water treatment plants. For this purpose, the bacterial communities present in treated water were sampled after granular activated carbon (GAC) filtration and chlorine disinfection from two drinking water treatment plants supplying the city of Paris (France). Samples were analyzed after genomic DNA extraction, polymerase chain reaction (PCR) amplification, cloning, and sequencing of a number of 16S ribosomal RNA (rRNA) genes. The 16S rDNA sequences were clustered into operational taxonomic units (OTUs) and the OTU abundance patterns were obtained for each sample. The observed differences suggest that the chlorine disinfection step markedly affects the bacterial community structure and composition present in GAC water. Members of the Alphaproteobacteria and Betaproteobacteria were found to be predominant in the GAC water samples after phylogenetic analyses of the OTUs. Following the chlorine disinfection step, numerous changes were observed, including decreased representation of Proteobacteria phylotypes. Our results indicate that the use of molecular methods to investigate changes in the abundance of certain bacterial groups following chlorine-based disinfection will aid in further understanding the bacterial ecology of drinking water treatment plants (DWTPs), particularly the disinfection step, as it constitutes the final barrier before drinking water distribution to the consumer's tap

Assessment of phylogenetic diversity of bacterial microflora in drinking water using serial analysis of ribosomal sequence tags.

International audienceWe examined chlorinated drinking water samples from three different surface water treatment plants for bacterial 16S rDNA diversity using the serial analysis of V6 ribosomal sequence tag (SARST-V6) method. A considerable degree of diversity was observed in each sample, with an estimated richness ranging from 173 to 333 phylotypes. The community structure shows that there are differences in bacterial evenness between sampled sites. The taxonomic composition of the microbial communities was found to be dominated by members of the Proteobacteria (57.2-77.4%), broadly distributed among the classes Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, and Deltaproteobacteria. Additionally, a large proportion of sequences (6.3-36.5%) were found to be distantly related to database sequences of unknown phylogenetic affiliation. Given the apparent significance of this bacterial group in drinking water, a 16S rDNA analysis was performed and confirmed their presence and phylogeny. Notwithstanding the potential under-representation of certain bacterial phyla using the SARST-V6 primer pairs, as revealed by a refined computer algorithm, our results suggest that 16S rDNA corresponding to a variety of eubacterial groups can be detected in finished drinking water, suggesting that this water may contain a higher level of bacterial diversity than previously observed

Quantitative prediction of genome-wide resource allocation in bacteria

Predicting resource allocation between cell processes is the primary step towards decoding the evolutionary constraints governing bacterial growth under various conditions. Quantitative prediction at genome-scale remains a computational challenge as current methods are limited by the tractability of the problem or by simplifying hypotheses. Here, we show that the constraint-based modeling method Resource Balance Analysis (RBA), calibrated using genome-wide absolute protein quantification data, accurately predicts resource allocation in the model bacterium Bacillus subtilis for a wide range of growth conditions. The regulation of most cellular processes is consistent with the objective of growth rate maximization except for a few suboptimal processes which likely integrate more complex objectives such as coping with stressful conditions and survival. As a proof of principle by using simulations, we illustrated how calibrated RBA could aid rational design of strains for maximizing protein production, offering new opportunities to investigate design principles in prokaryotes and to exploit them for biotechnological applications

Service sanitaire des étudiants en santé : ancrer l’éducation pour la santé dans le cursus des étudiants

National audienceIntroduction : Instauré au niveau national depuis 2018, le Service sanitaire des étudiants en santé (SSES) vise à les former à mener des actions d’éducation pour la santé.Méthode et résultats : Un dispositif pédagogique d’une durée équivalente à six semaines à temps plein a été mis en place en troisième année de médecine à Rennes (35). L’objectif est que les étudiants développent les compétences nécessaires pour mener des interventions fondées sur une démarche projet, auprès de publics variés, sur des thèmes prioritaires de santé publique.De nouvelles approches pédagogiques ont été développées pour intégrer au cursus médical l’apprentissage de la promotion et de l’éducation pour la santé. Des innovations ont été mises en place : travail sur la posture éducative, tutorat des étudiants de troisième année par des internes en médecine, forum de simulation des actions concrètes sous la supervision d’une double expertise thématique et populationnelle. Au-delà de l’acquisition de compétences en éducation pour la santé, la formation vise à susciter une démarche réflexive et s’appuie sur l’éducation par les pairs.Les 240 étudiants de la faculté préparent en trinôme leur projet tout au long de l’année universitaire. Leurs actions se déroulent sur une plage de dix demi-journées auprès d’une centaine d’établissements, dans la diversité des territoires de la subdivision. Elles permettent d’intervenir auprès d’environ 10 000 personnes par an.Discussion : L’éducation et la promotion de la santé occupent désormais une place centrale dans la formation des étudiants de troisième année, condition indispensable pour amorcer l’acquisition durable de ce champ de compétences par les futurs professionnels de santé

The bacteria and bacteriophages from a Mesquite Flats site of the Death Valley desert

International audienceArid zones cover over 30 % of the Earth's continental surface. In order to better understand the role of microbes in this type of harsh environment, we isolated and characterized the bacteriophages from samples of the surface sand of the Mesquite Flats region via electron microscopy and DNA sequencing of a select number of cloned phage DNAs. An electron microscopic analysis of the recovered virus-like particles revealed at least 11 apparently different morphotypes sharing structural characteristics of the Caudoviridae family of tailed phages. We found that 36 % of the sequences contained no significant identity (e-value >10(-3)) with sequences in the databases. Pilot sequencing of cloned 16S rRNA genes identified Bacteroidetes and Proteobacteria as the major bacterial groups present in this severe environment. The majority of the 16S rDNA sequences from the total (uncultured) bacterial population displayed ≤96 % identity to 16S rRNA genes in the database, suggesting an unexplored bacterial population likely adapted to a desert environment. In addition, we also isolated and identified 38 cultivable bacterial strains, the majority of which belonged to the genus Bacillus. Mitomycin-C treatment of the cultivable bacteria demonstrated that the vast majority (84 %) contained at least one SOS-inducible prophage