The defensome of complex bacterial communities.

Bacteria have developed various defense mechanisms to avoid infection and killing in response to the fast evolution and turnover of viruses and other genetic parasites. Such pan-immune system (defensome) encompasses a growing number of defense lines that include well-studied innate and adaptive systems such as restriction-modification, CRISPR-Cas and abortive infection, but also newly found ones whose mechanisms are still poorly understood. While the abundance and distribution of defense systems is well-known in complete and culturable genomes, there is a void in our understanding of their diversity and richness in complex microbial communities. Here we performed a large-scale in-depth analysis of the defensomes of 7759 high-quality bacterial population genomes reconstructed from soil, marine, and human gut environments. We observed a wide variation in the frequency and nature of the defensome among large phyla, which correlated with lifestyle, genome size, habitat, and geographic background. The defensome's genetic mobility, its clustering in defense islands, and genetic variability was found to be system-specific and shaped by the bacterial environment. Hence, our results provide a detailed picture of the multiple immune barriers present in environmentally distinct bacterial communities and set the stage for subsequent identification of novel and ingenious strategies of diversification among uncultivated microbes.

Tepidibacter aestuarii sp. nov., isolated from the Bach Dang Estuary, Haiphong, Vietnam.

A mesophilic, anaerobic, endospore-forming, fermentative bacterium designated strain 8C15bT was isolated from bank sediment of the Bach Dang Estuary, Haiphong, Vietnam. The Bach Dang Estuary, where Haiphong harbour is located, is subject to strong anthropogenic influence, resulting in high concentrations of black carbon and heavy metals. Strain 8C15bT grew optimally at 30 °C, pH 7.5 and with 2.5 % (w/v) NaCl. The main cellular fatty acids consisted of iso-C15 : 0 (51 %), iso-C15:1 ω7c (32 %) and iso-C13 : 0 (5 %). Genomic considerations of strain 8C15bT and comparisons with the phylogenetically closest strains of the genus Tepidibacter provide evidence that Tepidibacter thalassicus SC562T (=DSM 15285T), Tepidibacter formicigenes DV1184T (=DSM 15518T), Tepidibacter mesophilus B1T (=JCM 16806T) and strain 8C15bT could be differentiated at the species level. We propose the name Tepidibacter aestuarii sp. nov. for the type strain 8C15bT (=JCM 35983T=KCTC 25692T). Finally, the nickel-tolerance properties of strain 8C15bT are highlighted in this study.

Synthesis of non-anomeric C-glycosyl pyrazolidinone derivatives via visible-light photoredox catalysis.

C-Glycosyl compounds have gained considerable attention over the last few decades due to their high chemical stability and promising applications in drug discovery. Herein we disclose an operationally simple, metal-free, photocatalytic approach for the glycosylation of azomethine imines using 4-glycosyl-1,4-dihydropyridines (DHPs) as radical precursors. The protocol features mild reaction conditions, scalability, broad substrate scope, and good functional group tolerance. Moreover, the resulting pyrazolidinone moiety can be easily deprotected, acylated or reduced into a glycosyl ß-alanine analog.

Diversity of the Pacific Ocean coral reef microbiome.

Coral reefs are among the most diverse ecosystems on Earth. They support high biodiversity of multicellular organisms that strongly rely on associated microorganisms for health and nutrition. However, the extent of the coral reef microbiome diversity and its distribution at the oceanic basin-scale remains to be explored. Here, we systematically sampled 3 coral morphotypes, 2 fish species, and planktonic communities in 99 reefs from 32 islands across the Pacific Ocean, to assess reef microbiome composition and biogeography. We show a very large richness of reef microorganisms compared to other environments, which extrapolated to all fishes and corals of the Pacific, approximates the current estimated total prokaryotic diversity for the entire Earth. Microbial communities vary among and within the 3 animal biomes (coral, fish, plankton), and geographically. For corals, the cross-ocean patterns of diversity are different from those known for other multicellular organisms. Within each coral morphotype, community composition is always determined by geographic distance first, both at the island and across ocean scale, and then by environment. Our unprecedented sampling effort of coral reef microbiomes, as part of the Tara Pacific expedition, provides new insight into the global microbial diversity, the factors driving their distribution, and the biocomplexity of reef ecosystems.

Integrative omics framework for characterization of coral reef ecosystems from the Tara Pacific expedition.

Coral reef science is a fast-growing field propelled by the need to better understand coral health and resilience to devise strategies to slow reef loss resulting from environmental stresses. Key to coral resilience are the symbiotic interactions established within a complex holobiont, i.e. the multipartite assemblages comprising the coral host organism, endosymbiotic dinoflagellates, bacteria, archaea, fungi, and viruses. Tara Pacific is an ambitious project built upon the experience of previous Tara Oceans expeditions, and leveraging state-of-the-art sequencing technologies and analyses to dissect the biodiversity and biocomplexity of the coral holobiont screened across most archipelagos spread throughout the entire Pacific Ocean. Here we detail the Tara Pacific workflow for multi-omics data generation, from sample handling to nucleotide sequence data generation and deposition. This unique multidimensional framework also includes a large amount of concomitant metadata collected side-by-side that provide new assessments of coral reef biodiversity including micro-biodiversity and shape future investigations of coral reef dynamics and their fate in the Anthropocene.

Visible-Light Mediated Carbamoylation of Nitrones under a Continuous Flow Regime.

Herein, we report a rapid and scalable continuous-flow photocatalytic approach for the carbamoylation of nitrones. This protocol makes use of readily available 4-amido-1,4 dihydropyridines as carbamoyl radical precursors. The scope of this transformation exhibits high compatibility with complex structures containing amino acids, peptides, and glycosides. Importantly, the developed method allows a photocatalytic synthetic strategy in combination with flow conditions, maximizing the potential and efficiency for the synthesis of valuable α-(N-hydroxy)amino amides.

Photoinduced carbamoylation reactions: unlocking new reactivities towards amide synthesis.

The preparation of amide-containing compounds is among the most interesting and challenging topics for the synthetic community. Such relevance is given by their reactive aspects explored in the context of organic synthesis and by the direct application of these compounds as pharmaceuticals and useful materials, and their key roles in biological structures. A simple and straightforward strategy for the amide moiety installation is the use of carbamoyl radicals - this nucleophilic one-electron intermediate is prone to undergo a series of transformations, providing a range of structurally relevant derivatives. In this review, we summarize the latest advances in the field from the perspective of photoinduced protocols. To this end, their synthetic applications are organized accordingly to the nature of the radical precursor (formamides through HAT, 4-substituted-1,4-dihydropyridines, oxamic acids, and N-hydroxyphthalimido esters), the mechanistic aspects also being highlighted. The discussion also includes a recent approach proceeding via photolytic C-S cleavage of dithiocarbamate-carbamoyl intermediates. By exploring fundamental concepts, this material aims to offer an understanding of the topic, which will encourage and facilitate the design of new synthetic strategies applying the carbamoyl radical.

Complete Genome Sequence of Tepidibacter sp. Strain 8C15b, Isolated from Bank Sediments of Haiphong Bay, Vietnam.

We report the complete genome sequence of Tepidibacter sp. strain 8C15b, isolated from bank sediments of Haiphong Bay, Vietnam. The genome includes a 3,628,320-bp circular chromosome and a plasmid of 38,213 bp.

Bacterial Epigenomics: Coming of Age.

Epigenetic DNA methylation in bacteria has been traditionally studied in the context of antiparasitic defense and as part of the innate immune discrimination between self and nonself DNA. However, sequencing advances that allow genome-wide analysis of DNA methylation at the single-base resolution are nowadays expanding and have propelled a modern epigenomic revolution in our understanding of the extent, evolution, and physiological relevance of methylation. Indeed, as the number of mapped bacterial methylomes recently surpassed 4,000, increasing evidence supports roles for methylation in gene expression regulation, virulence, and host colonization, among others. In this paper, I summarize lessons taken from high-dimensional methylome data analyses and recent efforts that we and others are developing to leverage such findings into meaningful biological insights and overarching frameworks. Ultimately, I highlight anticipated research avenues and technological developments likely to unfold in the coming years.

Carbamoylation of Azomethine Imines via Visible-Light Photoredox Catalysis.

A versatile and robust photocatalytic methodology to install the amide functional group into azomethine imine ions is described. This protocol is distinguished by its broad scope and mild reaction conditions, which are well suited for the preparation of structurally complex compounds in the form of amino acids, peptides, and small drug-like molecules. Moreover, the generated pyrazolidinone core could be easily converted into ß-alanine analogues.

Conserved DNA Methyltransferases: A Window into Fundamental Mechanisms of Epigenetic Regulation in Bacteria.

An increasing number of studies have reported that bacterial DNA methylation has important functions beyond the roles in restriction-modification systems, including the ability of affecting clinically relevant phenotypes such as virulence, host colonization, sporulation, biofilm formation, among others. Although insightful, such studies have a largely ad hoc nature and would benefit from a systematic strategy enabling a joint functional characterization of bacterial methylomes by the microbiology community. In this opinion article, we propose that highly conserved DNA methyltransferases (MTases) represent a unique opportunity for bacterial epigenomic studies. These MTases are rather common in bacteria, span various taxonomic scales, and are present in multiple human pathogens. Apart from well-characterized core DNA MTases, like those from Vibrio cholerae, Salmonella enterica, Clostridioides difficile, or Streptococcus pyogenes, multiple highly conserved DNA MTases are also found in numerous human pathogens, including those belonging to the genera Burkholderia and Acinetobacter. We discuss why and how these MTases can be prioritized to enable a community-wide, integrative approach for functional epigenomic studies. Ultimately, we discuss how some highly conserved DNA MTases may emerge as promising targets for the development of novel epigenetic inhibitors for biomedical applications.

Author Correction: The chromosomal organization of horizontal gene transfer in bacteria.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Epigenomic characterization of Clostridioides difficile finds a conserved DNA methyltransferase that mediates sporulation and pathogenesis.

Clostridioides (formerly Clostridium) difficile is a leading cause of healthcare-associated infections. Although considerable progress has been made in the understanding of its genome, the epigenome of C. difficile and its functional impact has not been systematically explored. Here, we perform a comprehensive DNA methylome analysis of C. difficile using 36 human isolates and observe a high level of epigenomic diversity. We discovered an orphan DNA methyltransferase with a well-defined specificity, the corresponding gene of which is highly conserved across our dataset and in all of the approximately 300 global C. difficile genomes examined. Inactivation of the methyltransferase gene negatively impacts sporulation, a key step in C. difficile disease transmission, and these results are consistently supported by multiomics data, genetic experiments and a mouse colonization model. Further experimental and transcriptomic analyses suggest that epigenetic regulation is associated with cell length, biofilm formation and host colonization. These findings provide a unique epigenetic dimension to characterize medically relevant biological processes in this important pathogen. This study also provides a set of methods for comparative epigenomics and integrative analysis, which we expect to be broadly applicable to bacterial epigenomic studies.

Host Range and Genetic Plasticity Explain the Coexistence of Integrative and Extrachromosomal Mobile Genetic Elements.

Self-transmissible mobile genetic elements drive horizontal gene transfer between prokaryotes. Some of these elements integrate in the chromosome, whereas others replicate autonomously as plasmids. Recent works showed the existence of few differences, and occasional interconversion, between the two types of elements. Here, we enquired on why evolutionary processes have maintained the two types of mobile genetic elements by comparing integrative and conjugative elements (ICE) with extrachromosomal ones (conjugative plasmids) of the highly abundant MPFT conjugative type. We observed that plasmids encode more replicases, partition systems, and antibiotic resistance genes, whereas ICEs encode more integrases and metabolism-associated genes. ICEs and plasmids have similar average sizes, but plasmids are much more variable, have more DNA repeats, and exchange genes more frequently. On the other hand, we found that ICEs are more frequently transferred between distant taxa. We propose a model where the different genetic plasticity and amplitude of host range between elements explain the co-occurrence of integrative and extrachromosomal elements in microbial populations. In particular, the conversion from ICE to plasmid allows ICE to be more plastic, while the conversion from plasmid to ICE allows the expansion of the element's host range.

The chromosomal organization of horizontal gene transfer in bacteria.

Bacterial adaptation is accelerated by the acquisition of novel traits through horizontal gene transfer, but the integration of these genes affects genome organization. We found that transferred genes are concentrated in only ~1% of the chromosomal regions (hotspots) in 80 bacterial species. This concentration increases with genome size and with the rate of transfer. Hotspots diversify by rapid gene turnover; their chromosomal distribution depends on local contexts (neighboring core genes), and content in mobile genetic elements. Hotspots concentrate most changes in gene repertoires, reduce the trade-off between genome diversification and organization, and should be treasure troves of strain-specific adaptive genes. Most mobile genetic elements and antibiotic resistance genes are in hotspots, but many hotspots lack recognizable mobile genetic elements and exhibit frequent homologous recombination at flanking core genes. Overrepresentation of hotspots with fewer mobile genetic elements in naturally transformable bacteria suggests that homologous recombination and horizontal gene transfer are tightly linked in genome evolution.Horizontal gene transfer (HGT) is an important mechanism for genome evolution and adaptation in bacteria. Here, Oliveira and colleagues find HGT hotspots comprising ~ 1% of the chromosomal regions in 80 bacterial species.

Bond Strength of Abraded and Non-Abraded Bleached Enamel to Resin After Er,Cr:YSGG Laser Irradiation.

OBJECTIVE: The objective of the study was to evaluate the microtensile bond strength (µTBS) of a composite resin to abraded or non-abraded bleached enamel after Er,Cr:YSGG laser irradiation and to observe the fracture patterns of the tested interfaces. MATERIALS AND METHODS: Two hundred twenty-eight bovine incisors were sectioned, resulting in 228 enamel blocks (7 × 4 × 4 mm3) that were divided into 12 groups (n = 19) according to the factors "adhesion" after bleaching (immediate adhesion; after 14 days; and a control group with adhesion on unbleached teeth); enamel "abrasion" (with or without abrasion simulating cavity preparation); and "laser" (with or without Er,Cr:YSGG laser irradiation). Bleached enamel groups were treated with 20% carbamide peroxide, 8 h/day for 21 days. Abrasion was performed with silicon carbide sandpaper. Specimens were restored with adhesive system and a composite resin (Adper Single Bond 2 and Z250; 3M ESPE). After 7 days, specimens were prepared by cutting into 1 mm beans to µTBS test performed in a universal testing machine. Fracture mode analysis was performed by using a stereoscopic loupe. The µTBS data were statistically analyzed by three-way analysis of variance with 95% confidence level and compared by running a Tukey post hoc test (α = 0.05). RESULTS: There was no statistically significant difference between triple interaction and double interactions among factors. There was no significant difference between the factors "adhesion," "abrasion," and "laser." Laser irradiation produced significantly lower bond strength values in irradiated groups compared with the non-irradiated ones. All groups had a high percentage of adhesive failures. CONCLUSIONS: Abrasion provided no benefit to bond strength of composite resins to bleached enamel. Er,Cr:YSGG (20 Hz, 0.5 W, 3.97 J/cm2) treatment reduced the bond strength of composite resins to enamel.

Regulation of genetic flux between bacteria by restriction-modification systems.

Restriction-modification (R-M) systems are often regarded as bacteria's innate immune systems, protecting cells from infection by mobile genetic elements (MGEs). Their diversification has been recently associated with the emergence of particularly virulent lineages. However, we have previously found more R-M systems in genomes carrying more MGEs. Furthermore, it has been suggested that R-M systems might favor genetic transfer by producing recombinogenic double-stranded DNA ends. To test whether R-M systems favor or disfavor genetic exchanges, we analyzed their frequency with respect to the inferred events of homologous recombination and horizontal gene transfer within 79 bacterial species. Genetic exchanges were more frequent in bacteria with larger genomes and in those encoding more R-M systems. We created a recognition target motif predictor for Type II R-M systems that identifies genomes encoding systems with similar restriction sites. We found more genetic exchanges between these genomes, independently of their evolutionary distance. Our results reconcile previous studies by showing that R-M systems are more abundant in promiscuous species, wherein they establish preferential paths of genetic exchange within and between lineages with cognate R-M systems. Because the repertoire and/or specificity of R-M systems in bacterial lineages vary quickly, the preferential fluxes of genetic transfer within species are expected to constantly change, producing time-dependent networks of gene transfer.