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Abstract Background The mollicute Mycoplasma conjunctivae is the etiological agent leading to infectious keratoconjunctivitis (IKC) in domestic sheep and wild caprinae. Although this pathogen is relatively benign for domestic animals treated by antibiotics, it can lead wild animals to blindness and death. This is a major cause of death in the protected species in the Alps (e.g., Capra ibex, Rupicapra rupicapra). Methods The genome was sequenced using a combined technique of GS-FLX (454) and Sanger sequencing, and annotated by an automatic pipeline that we designed using several tools interconnected via PERL scripts. The resulting annotations are stored in a MySQL database. Results The annotated sequence is deposited in the EMBL database (FM864216) and uploaded into the mollicutes database MolliGen http://cbi.labri.fr/outils/molligen/ allowing for comparative genomics. Conclusion We show that our automatic pipeline allows for annotating a complete mycoplasma genome and present several examples of analysis in search for biological targets (e.g., pathogenic proteins).

Abstract Background Fourmidable is an infrastructure to curate and share the emerging genetic, molecular, and functional genomic data and protocols for ants. Description The Fourmidable assembly pipeline groups nucleotide sequences into clusters before independently assembling each cluster. Subsequently, assembled sequences are annotated via Interproscan and BLAST against general and insect-specific databases. Gene-specific information can be retrieved using gene identifiers, searching for similar sequences or browsing through inferred Gene Ontology annotations. The database will readily scale as ultra-high throughput sequence data and sequences from additional species become available. Conclusion Fourmidable currently houses EST data from two ant species and microarray gene expression data for one of these. Fourmidable is publicly available at http://fourmidable.unil.ch

Background: Building Metagenome-Assembled Genomes (MAGs) from highly complex metagenomics datasets encompasses a series of steps covering from cleaning the sequences, assembling them to finally group them into bins. Along the process, multiple tools aimed to assess the quality and integrity of each MAG are implemented. Nonetheless, even when incorporated within end-to-end pipelines, the outputs of these pieces of software must be visualized and analyzed manually lacking integration in a complete framework., Methods: We developed a Nextflow pipeline (MAGFlow) for estimating the quality of MAGs through a wide variety of approaches (BUSCO, CheckM2, GUNC and QUAST), as well as for annotating taxonomically the metagenomes using GTDB-Tk2. MAGFlow is coupled to a Python-Dash application (BIgMAG) that displays the concatenated outcomes from the tools included by MAGFlow, highlighting the most important metrics in a single interactive environment along with a comparison/clustering of the input data., Results: By using MAGFlow/BIgMAG, the user will be able to benchmark the MAGs obtained through different workflows or establish the quality of the MAGs belonging to different samples following the divide and rule methodology., Conclusions: MAGFlow/BIgMAG represents a unique tool that integrates state-of-the-art tools to study different quality metrics and extract visually as much information as possible from a wide range of genome features., Competing Interests: No competing interests were disclosed., (Copyright: © 2024 Yepes-García J and Falquet L.)

Botrytis cinerea is a necrotrophic phytopathogen able to attack more than 200 different plant species causing strong yield losses worldwide. Many synthetic fungicides have been developed to control this disease, resulting in the rise of fungicide-resistance B. cinerea strains. The aim of this study was to identify Streptomyces strains showing antagonistic activity against B. cinerea to contribute to plant protection in an environmentally friendly way. We isolated 15 Actinomycete strains from 9 different Swiss soils. The culture filtrates of three isolates showing antifungal activity inhibited spore germination and delayed mycelial growth of B. cinerea . Infection experiments showed that Arabidopsis thaliana plants were more resistant to this pathogen after leaf treatment with the Streptomyces filtrates. Bioassay-guided isolation of the active compounds revealed the presence of germicidins A and B as well as of oligomycins A, B, and E. While germicidins were mostly inactive, oligomycin B reduced the mycelial growth of B. cinerea significantly. Moreover, all three oligomycins inhibited this fungus' spore germination, suggesting that these molecules might contribute to the Streptomyces 's ability to protect plants against infection by the broad host-pathogen Botrytis cinerea ., Importance: This study reports the isolation of new Streptomyces strains with strong plant-protective potential mediated by their production of specialized metabolites. Using the broad host range pathogenic fungus Botrytis cinerea , we demonstrate that the cell-free filtrate of selected Streptomyces isolates efficiently inhibits different developmental stages of the fungus, including mycelial growth and the epidemiologically relevant spore germination. Beyond in vitro experiments, the strains and their metabolites also efficiently protected plants against the disease caused by this pathogen. This work further identifies oligomycins as active compounds involved in the observed antifungal activity of the strains. This work shows that we can harness the natural ability of soil-borne microbes and of their metabolites to efficiently fight other microbes responsible for significant crop losses. This opens the way to the development of environmentally friendly health protection measures for crops of agronomical relevance, based on these newly isolated strains or their metabolic extracts containing oligomycins., Competing Interests: The authors declare no conflict of interest.

Introduction: The early-life intestinal microbiome plays an important role in the development and regulation of the immune system. It is unknown whether the administration of vaccines influences the composition of the intestinal microbiome., Objective: To investigate whether Bacille Calmette-Guérin (BCG) vaccine given in the first few days of life influences the abundance of bacterial taxa and metabolic pathways in the intestinal microbiome at 1 week of age., Methods: Healthy, term-born neonates were randomized at birth to receive BCG or no vaccine within the first few days of life. Stool samples were collected at 1 week of age from 335 neonates and analyzed using shotgun metagenomic sequencing and functional analyses., Results: The composition of the intestinal microbiome was different between neonates born by cesarean section (CS) and those born vaginally. Differences in the composition between BCG-vaccinated and BCG-naïve neonates were only minimal. CS-born BCG-vaccinated neonates had a higher abundance of Staphylococcus lugdunensis compared with CS-born BCG-naïve neonates. The latter had a higher abundance of Streptococcus infantis and Trabulsiella guamensis . Vaginally-born BCG-vaccinated neonates had a higher abundance of Clostridiaceae and Streptococcus parasanguinis compared with vaginally-born BCG-naïve neonates, and a lower abundance of Veillonella atypica and Butyricimonas faecalis. Metabolic pathways that were differently abundant between BCG-vaccinated and BCG-naïve neonates were mainly those involved in sugar degradation and nucleotide/nucleoside biosynthesis., Conclusion: BCG given in the first few days of life has little effect on the composition of the intestinal microbiome at 1 week of age but does influence the abundance of certain metabolic pathways., Competing Interests: The authors have no funding or conflicts of interest to disclose., (Copyright © 2023 Wolters Kluwer Health, Inc. All rights reserved.)

Cassava (Manihot esculenta Crantz) is a vital tropical root crop providing essential dietary energy to over 800 million people in tropical and subtropical regions. As a climate-resilient crop, its significance grows as the human population expands. However, yield improvement faces challenges from biotic and abiotic stress and limited breeding. Advanced sequencing and assembly techniques enabled the generation of a highly accurate, nearly complete, haplotype-resolved genome of the African cassava cultivar TMEB117. It is the most accurate cassava genome sequence to date with a base-level accuracy of QV > 64, N50 > 35 Mbp, and 98.9% BUSCO completeness. Over 60% of the genome comprises repetitive elements. We predicted over 45,000 gene models for both haplotypes. This achievement offers valuable insights into the heterozygosity genome organization of the cassava genome, with improved accuracy, completeness, and phased genomes. Due to its high susceptibility to African Cassava Mosaic Virus (ACMV) infections compared to other cassava varieties, TMEB117 provides an ideal reference for studying virus resistance mechanisms, including epigenetic variations and smallRNA expressions., (© 2023. The Author(s).)

The rixosome defined in Schizosaccharomyces pombe and humans performs diverse roles in pre-ribosomal RNA processing and gene silencing. Here, we isolate and describe the conserved rixosome from Chaetomium thermophilum, which consists of two sub-modules, the sphere-like Rix1-Ipi3-Ipi1 and the butterfly-like Las1-Grc3 complex, connected by a flexible linker. The Rix1 complex of the rixosome utilizes Sda1 as landing platform on nucleoplasmic pre-60S particles to wedge between the 5S rRNA tip and L1-stalk, thereby facilitating the 180° rotation of the immature 5S RNP towards its mature conformation. Upon rixosome positioning, the other sub-module with Las1 endonuclease and Grc3 polynucleotide-kinase can reach a strategic position at the pre-60S foot to cleave and 5' phosphorylate the nearby ITS2 pre-rRNA. Finally, inward movement of the L1 stalk permits the flexible Nop53 N-terminus with its AIM motif to become positioned at the base of the L1-stalk to facilitate Mtr4 helicase-exosome participation for completing ITS2 removal. Thus, the rixosome structure elucidates the coordination of two central ribosome biogenesis events, but its role in gene silencing may adapt similar strategies., (© 2023 The Authors. Published under the terms of the CC BY 4.0 license.)

The Streptomyces genus is known to produce many specialized metabolites of value for medicine, but the potential of these metabolites in agronomy remains largely unexplored. In this study, we investigated three phylogenetically closely related Streptomyces strains (B5, B91, and B135) isolated from three distinct soil samples in Sudan. Despite belonging to the same species, these strains exhibited different ranges of Phytophthora infestans inhibition. The objective of this work was to identify the active compound(s) responsible for the inhibition of P. infestans and of other plant pathogens by comparing the genomes and metabolomes of the three strains which showed distinct activity patterns: B5 was the strongest inhibitor of oomycetes, B5 and B91 both inhibited most fungi and B135 was the only strain showing antibacterial activity. Our comparative genomic and metabolomic analysis identified borrelidin as the bioactive compound underlying B5's strong anti-oomycete activity and highlighted a few other metabolites as putative candidates underlying the strains' antifungal and antibacterial activities. This study illustrates the power of comparative genomics and metabolomics on phylogenetically closely related strains of differing activities to highlight bioactive compounds that could contribute to new sustainable crop protection strategies., (© 2023 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.)

Importance: Bacteria communicate by exchanging chemical signals, some of which are volatile and can remotely reach other organisms. HCN was one of the first volatiles discovered to severely impact exposed organisms by inhibiting their respiration. Using HCN-deficient mutants in two Pseudomonas strains, we demonstrate that HCN's impact goes beyond the sole inhibition of respiration and affects both emitting and receiving bacteria in a global way, modulating their motility, biofilm formation, and production of antimicrobial compounds. Our data suggest that bacteria could use HCN not only to control their own cellular functions, but also to remotely influence the behavior of other bacteria sharing the same environment. Since HCN emission occurs in both clinically and environmentally relevant Pseudomonas , these findings are important to better understand or even modulate the expression of bacterial traits involved in both virulence of opportunistic pathogens and in biocontrol efficacy of plant-beneficial strains., Competing Interests: The authors declare no conflict of interest.

Mycoplasma feriruminatoris is a fast-growing Mycoplasma species isolated from wild Caprinae and first described in 2013. M. feriruminatoris isolates have been associated with arthritis, kerato conjunctivitis, pneumonia and septicemia, but were also recovered from apparently healthy animals. To better understand what defines this species, we performed a genomic survey on 14 strains collected from free-ranging or zoo-housed animals between 1987 and 2017, mostly in Europe. The average chromosome size of the M. feriruminatoris strains was 1,040±0,024 kbp, with 24 % G+C and 852±31 CDS. The core genome and pan-genome of the M. feriruminatoris species contained 628 and 1312 protein families, respectively. The M. feriruminatoris strains displayed a relatively closed pan-genome, with many features and putative virulence factors shared with species from the M. mycoides cluster, including the MIB-MIP Ig cleavage system, a repertoire of DUF285 surface proteins and a complete biosynthetic pathway for galactan. M. feriruminatoris genomes were found to be mostly syntenic, although repertoires of mobile genetic elements, including Mycoplasma Integrative and Conjugative Elements, insertion sequences, and a single plasmid varied. Phylogenetic- and gene content analyses confirmed that M. feriruminatoris was closer to the M. mycoides cluster than to the ruminant species M. yeatsii and M. putrefaciens . Ancestral genome reconstruction showed that the emergence of the M. feriruminatoris species was associated with the gain of 17 gene families, some of which encode defence enzymes and surface proteins, and the loss of 25 others, some of which are involved in sugar transport and metabolism. This comparative study suggests that the M. mycoides cluster could be extended to include M. feriruminatoris . We also find evidence that the specific organization and structure of the DnaA boxes around the oriC of M. feriruminatoris may contribute to drive the remarkable fast growth of this minimal bacterium.

The rapid development of machine learning (ML) techniques has opened up the data-dense field of microbiome research for novel therapeutic, diagnostic, and prognostic applications targeting a wide range of disorders, which could substantially improve healthcare practices in the era of precision medicine. However, several challenges must be addressed to exploit the benefits of ML in this field fully. In particular, there is a need to establish "gold standard" protocols for conducting ML analysis experiments and improve interactions between microbiome researchers and ML experts. The Machine Learning Techniques in Human Microbiome Studies (ML4Microbiome) COST Action CA18131 is a European network established in 2019 to promote collaboration between discovery-oriented microbiome researchers and data-driven ML experts to optimize and standardize ML approaches for microbiome analysis. This perspective paper presents the key achievements of ML4Microbiome, which include identifying predictive and discriminatory 'omics' features, improving repeatability and comparability, developing automation procedures, and defining priority areas for the novel development of ML methods targeting the microbiome. The insights gained from ML4Microbiome will help to maximize the potential of ML in microbiome research and pave the way for new and improved healthcare practices., Competing Interests: CJ is employed by Biome diagnostics GmbH. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2023 D’Elia, Truu, Lahti, Berland, Papoutsoglou, Ceci, Zomer, Lopes, Ibrahimi, Gruca, Nechyporenko, Frohme, Klammsteiner, Pau, Marcos-Zambrano, Hron, Pio, Simeon, Suharoschi, Moreno-Indias, Temko, Nedyalkova, Apostol, Truică, Shigdel, Telalović, Bongcam-Rudloff, Przymus, Jordamović, Falquet, Tarazona, Sampri, Isola, Pérez-Serrano, Trajkovik, Klucar, Loncar-Turukalo, Havulinna, Jansen, Bertelsen and Claesson.)

Introduction: The intestinal microbiome forms a major reservoir for antibiotic resistance genes (ARGs). Little is known about the neonatal intestinal resistome., Objective: The objective of this study was to investigate the intestinal resistome and factors that influence the abundance of ARGs in a large cohort of neonates., Methods: Shotgun metagenomics was used to analyse the resistome in stool samples collected at 1 week of age from 390 healthy, term-born neonates who did not receive antibiotics., Results: Overall, 913 ARGs belonging to 27 classes were identified. The most abundant ARGs were those conferring resistance to tetracyclines, quaternary ammonium compounds, and macrolide-lincosamide-streptogramin-B. Phylogenetic composition was strongly associated with the resistome composition. Other factors that were associated with the abundance of ARGs were delivery mode, gestational age, birth weight, feeding method, and antibiotics in the last trimester of pregnancy. Sex, ethnicity, probiotic use during pregnancy, and intrapartum antibiotics had little effect on the abundance of ARGs., Conclusion: Even in the absence of direct antibiotic exposure, the neonatal intestine harbours a high abundance and a variety of ARGs., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2023 Leo, Cetiner, Pittet, Messina, Jakob, Falquet, Curtis and Zimmermann.)

A growing body of evidence links gut microbiota changes with inflammatory bowel disease (IBD), raising the potential benefit of exploiting metagenomics data for non-invasive IBD diagnostics. The sbv IMPROVER metagenomics diagnosis for inflammatory bowel disease challenge investigated computational metagenomics methods for discriminating IBD and nonIBD subjects. Participants in this challenge were given independent training and test metagenomics data from IBD and nonIBD subjects, which could be wither either raw read data (sub-challenge 1, SC1) or processed Taxonomy- and Function-based profiles (sub-challenge 2, SC2). A total of 81 anonymized submissions were received between September 2019 and March 2020. Most participants' predictions performed better than random predictions in classifying IBD versus nonIBD, Ulcerative Colitis (UC) versus nonIBD, and Crohn's Disease (CD) versus nonIBD. However, discrimination between UC and CD remains challenging, with the classification quality similar to the set of random predictions. We analyzed the class prediction accuracy, the metagenomics features by the teams, and computational methods used. These results will be openly shared with the scientific community to help advance IBD research and illustrate the application of a range of computational methodologies for effective metagenomic classification., (© 2023. The Author(s).)

Soil tillage or herbicide applications are commonly used in agriculture for weed control. These measures may also represent a disturbance for soil microbial communities and their functions. However, the generality of response patterns of microbial communities and functions to disturbance have rarely been studied at large geographical scales. We investigated how a soil disturbance gradient (low, intermediate, high), realized by either tillage or herbicide application, affects diversity and composition of soil bacterial and fungal communities as well as soil functions in vineyards across five European countries. Microbial alpha-diversity metrics responded to soil disturbance sporadically, but inconsistently across countries. Increasing soil disturbance changed soil microbial community composition at the European level. However, the effects of soil disturbance on the variation of microbial communities were smaller compared to the effects of location and soil covariates. Microbial respiration was consistently impaired by soil disturbance, while effects on decomposition of organic substrates were inconsistent and showed positive and negative responses depending on the respective country. Therefore, we conclude that it is difficult to extrapolate results from one locality to others because microbial communities and environmental conditions vary strongly over larger geographical scales., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Steiner et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: In eukaryotes, cell-to-cell communication relies on the activity of small signaling peptides. In plant genomes, many hundreds of genes encode for such short peptide signals. However, only few of them are functionally characterized and due to the small gene size and high sequence variability, the comprehensive identification of such peptide-encoded genes is challenging. The CLAVATA3 (CLV3)/EMBRYO SURROUNDING REGION-RELATED (CLE) gene family encodes for short peptides that have a role in plant meristem maintenance, vascular patterning and responses to environment. The full repertoire of CLE genes and the role of CLE signaling in tomato (Solanum lycopersicum)- one of the most important crop plants- has not yet been fully studied., Results: By using a combined approach, we performed a genome-wide identification of CLE genes using the current tomato genome version SL 4.0. We identified 52 SlCLE genes, including 37 new non annotated before. By analyzing publicly available RNAseq datasets we could confirm the expression of 28 new SlCLE genes. We found that SlCLEs are often expressed in a tissue-, organ- or condition-specific manner. Our analysis shows an interesting gene diversification within the SlCLE family that seems to be a result of gene duplication events. Finally, we could show a biological activity of selected SlCLE peptides in the root growth arrest that was SlCLV2-dependent., Conclusions: Our improved combined approach revealed 37 new SlCLE genes. These findings are crucial for better understanding of the CLE signaling in tomato. Our phylogenetic analysis pinpoints the closest homologs of Arabidopsis CLE genes in tomato genome and can give a hint about the function of newly identified SlCLEs. The strategy described here can be used to identify more precisely additional short genes in plant genomes. Finally, our work suggests that the mechanism of root-active CLE peptide perception is conserved between Arabidopsis and tomato. In conclusion, our work paves the way to further research on the CLE-dependent circuits modulating tomato development and physiological responses., (© 2022. The Author(s).)

Members of the Staphylococcaceae family, particularly those of the genus Staphylococcus, encompass important human and animal pathogens. We collected and characterized Staphylococcaceae strains from apparently healthy and diseased camels ( n = 84) and cattle ( n = 7) in Somalia and Kenya. We phenotypically characterized the strains, including their antimicrobial inhibitory concentrations. Then, we sequenced their genomes using long-read sequencing, closed their genomes, and subsequently compared and mapped their virulence- and resistance-associated gene pools. Genome-based phylogenetics revealed 13 known Staphylococcaceae and at least two novel species. East African strains of different species encompassed novel sequence types and phylogenetically distant clades. About one-third of the strains had non-wild-type MICs. They were resistant to at least one of the following antimicrobials: tetracycline, benzylpenicillin, oxacillin, erythromycin, clindamycin, trimethoprim, gentamicin, or streptomycin, encoded by tet (K), blaZ / bla ARL , mecA / mecA1 , msrA / mphC , salA , dfrG , aacA-aphD , and str , respectively. We identified the first methicillin- and multidrug-resistant camel S. epidermidis strain of sequence type (ST) 1136 in East Africa. The pool of virulence-encoding genes was largest in the S. aureus strains, as expected, although other rather commensal strains contained distinct virulence-encoding genes. We identified toxin-antitoxin (TA) systems such as the hicA/hicB and abiEii/abiEi families, reported here for the first time for certain species of Staphylococcaceae . All strains contained at least one intact prophage sequence, mainly belonging to the Siphoviridae family. We pinpointed potential horizontal gene transfers between camel and cattle strains and also across distinct Staphylococcaceae clades and species. IMPORTANCE Camels are a high value and crucial livestock species in arid and semiarid regions of Africa and gain importance giving the impact of climate change on traditional livestock species. Our current knowledge with respect to Staphylococcaceae infecting camels is very limited compared to that for other livestock species. Better knowledge will foster the development of specific diagnostic assays, guide promising antimicrobial treatment options, and inform about potential zoonotic risks. We characterized 84 Staphylococcaceae strains isolated from camels with respect to their antimicrobial resistance and virulence traits. We detected potentially novel Staphylococcus species, resistances to different classes of antimicrobials, and the first camel multidrug-resistant S. epidermidis strain of sequence type 1136.

The kidney is strongly dependent on a continuous oxygen supply, and is conversely highly sensitive to hypoxia. Controlled oxygen gradients are essential for renal control of solutes and urine-concentrating mechanisms, which also depend on various hormones including aldosterone. The cortical collecting duct (CCD) is part of the aldosterone-sensitive distal nephron and possesses a key function in fine-tuned distal salt handling. It is well known that aldosterone is consistently decreased upon hypoxia. Furthermore, a recent study reported a hypoxia-dependent down-regulation of sodium currents within CCD cells. We thus investigated the possibility that cells from the cortical collecting duct are responsive to hypoxia, using the mouse cortical collecting duct cell line mCCDcl1 as a model. By analyzing the hypoxia-dependent transcriptome of mCCDcl1 cells, we found a large number of differentially-expressed genes (3086 in total logFC< −1 or >1) following 24 h of hypoxic conditions (0.2% O2). A gene ontology analysis of the differentially-regulated pathways revealed a strong decrease in oxygen-linked processes such as ATP metabolic functions, oxidative phosphorylation, and cellular and aerobic respiration, while pathways associated with hypoxic responses were robustly increased. The most pronounced regulated genes were confirmed by RT-qPCR. The low expression levels of Epas1 under both normoxic and hypoxic conditions suggest that Hif-1α, rather than Hif-2α, mediates the hypoxic response in mCCDcl1 cells. Accordingly, we generated shRNA-mediated Hif-1α knockdown cells and found Hif-1α to be responsible for the hypoxic induction of established hypoxically-induced genes. Interestingly, we could show that following shRNA-mediated knockdown of Esrra, Hif-1α protein levels were unaffected, but the gene expression levels of Egln3 and Serpine1 were significantly reduced, indicating that Esrra might contribute to the hypoxia-mediated expression of these and possibly other genes. Collectively, mCCDcl1 cells display a broad response to hypoxia and represent an adequate cellular model to study additional factors regulating the response to hypoxia.

Toxins of toxin-antitoxin systems use diverse mechanisms to control bacterial growth. Here, we focus on the deleterious toxin of the atypical tripartite toxin-antitoxin-chaperone (TAC) system of Mycobacterium tuberculosis, whose inhibition requires the concerted action of the antitoxin and its dedicated SecB-like chaperone. We show that the TAC toxin is a bona fide ribonuclease and identify exact cleavage sites in mRNA targets on a transcriptome-wide scale in vivo. mRNA cleavage by the toxin occurs after the second nucleotide of the ribosomal A-site codon during translation, with a strong preference for CCA codons in vivo. Finally, we report the cryo-EM structure of the ribosome-bound TAC toxin in the presence of native M. tuberculosis cspA mRNA, revealing the specific mechanism by which the TAC toxin interacts with the ribosome and the tRNA in the P-site to cleave its mRNA target., (© 2022. The Author(s).)

The biogenesis of eukaryotic ribosomes involves the ordered assembly of around 80 ribosomal proteins. Supplying equimolar amounts of assembly-competent ribosomal proteins is complicated by their aggregation propensity and the spatial separation of their location of synthesis and pre-ribosome incorporation. Recent evidence has highlighted that dedicated chaperones protect individual, unassembled ribosomal proteins on their path to the pre-ribosomal assembly site. Here, we show that the co-translational recognition of Rpl3 and Rpl4 by their respective dedicated chaperone, Rrb1 or Acl4, reduces the degradation of the encoding RPL3 and RPL4 mRNAs in the yeast Saccharomyces cerevisiae . In both cases, negative regulation of mRNA levels occurs when the availability of the dedicated chaperone is limited and the nascent ribosomal protein is instead accessible to a regulatory machinery consisting of the nascent-polypeptide-associated complex and the Caf130-associated Ccr4-Not complex. Notably, deregulated expression of Rpl3 and Rpl4 leads to their massive aggregation and a perturbation of overall proteostasis in cells lacking the E3 ubiquitin ligase Tom1. Taken together, we have uncovered an unprecedented regulatory mechanism that adjusts the de novo synthesis of Rpl3 and Rpl4 to their actual consumption during ribosome assembly and, thereby, protects cells from the potentially detrimental effects of their surplus production., Competing Interests: BP, AM, GM, SF, MS, LF, DK No competing interests declared, (© 2022, Pillet et al.)

In order to thrive in constantly changing environments, animals must adaptively respond to threatening events. Noxious stimuli are not only processed according to their absolute intensity, but also to their context. Adaptation processes can cause animals to habituate at different rates and degrees in response to permanent or repeated stimuli. Here, we used a forward genetic approach in Caenorhabditis elegans to identify a neuropeptidergic pathway, essential to prevent fast habituation and maintain robust withdrawal responses to repeated noxious stimuli. This pathway involves the FRPR-19A and FRPR-19B G-protein coupled receptor isoforms produced from the frpr-19 gene by alternative splicing. Loss or overexpression of each or both isoforms can impair withdrawal responses caused by the optogenetic activation of the polymodal FLP nociceptor neuron. Furthermore, we identified FLP-8 and FLP-14 as FRPR-19 ligands in vitro. flp-14, but not flp-8, was essential to promote withdrawal response and is part of the same genetic pathway as frpr-19 in vivo. Expression and cell-specific rescue analyses suggest that FRPR-19 acts both in the FLP nociceptive neurons and downstream interneurons, whereas FLP-14 acts from interneurons. Importantly, genetic impairment of the FLP-14/FRPR-19 pathway accelerated the habituation to repeated FLP-specific optogenetic activation, as well as to repeated noxious heat and harsh touch stimuli. Collectively, our data suggest that well-adjusted neuromodulation via the FLP-14/FRPR-19 pathway contributes to promote nociceptive signals in C. elegans and counteracts habituation processes that otherwise tend to rapidly reduce aversive responses to repeated noxious stimuli., Competing Interests: The authors have declared that no competing interests exist.

Mycoplasmas are minute bacteria controlled by very small genomes ranging from 0.6 to 1.4 Mbp. They encompass several important medical and veterinary pathogens that are often associated with a wide range of chronic diseases. The long persistence of mycoplasma cells in their hosts can exacerbate the spread of antimicrobial resistance observed for many species. However, the nature of the virulence factors driving this phenomenon in mycoplasmas is still unclear. Toxin-antitoxin systems (TA systems) are genetic elements widespread in many bacteria that were historically associated with bacterial persistence. Their presence on mycoplasma genomes has never been carefully assessed, especially for pathogenic species. Here we investigated three candidate TA systems in M. mycoides subsp. capri encoding a (i) novel AAA-ATPase/subtilisin-like serine protease module, (ii) a putative AbiEii/AbiEi pair and (iii) a putative Fic/RelB pair. We sequence analyzed fourteen genomes of M. mycoides subsp. capri and confirmed the presence of at least one TA module in each of them. Interestingly, horizontal gene transfer signatures were also found in several genomic loci containing TA systems for several mycoplasma species. Transcriptomic and proteomic data confirmed differential expression profiles of these TA systems during mycoplasma growth in vitro. While the use of heterologous expression systems based on E. coli and B. subtilis showed clear limitations, the functionality and neutralization capacities of all three candidate TA systems were successfully confirmed using M. capricolum subsp. capricolum as a host. Additionally, M. capricolum subsp. capricolum was used to confirm the presence of functional TA system homologs in mycoplasmas of the Hominis and Pneumoniae phylogenetic groups. Finally, we showed that several of these M. mycoides subsp. capri toxins tested in this study, and particularly the subtilisin-like serine protease, could be used to establish a kill switch in mycoplasmas for industrial applications., Competing Interests: The authors have declared that no competing interests exist.

The ability of gut bacterial pathogens to escape immunity by antigenic variation-particularly via changes to surface-exposed antigens-is a major barrier to immune clearance 1 . However, not all variants are equally fit in all environments 2,3 . It should therefore be possible to exploit such immune escape mechanisms to direct an evolutionary trade-off. Here, we demonstrate this phenomenon using Salmonella enterica subspecies enterica serovar Typhimurium (S.Tm). A dominant surface antigen of S.Tm is its O-antigen: a long, repetitive glycan that can be rapidly varied by mutations in biosynthetic pathways or by phase variation 4,5 . We quantified the selective advantage of O-antigen variants in the presence and absence of O-antigen-specific immunoglobulin A and identified a set of evolutionary trajectories allowing immune escape without an associated fitness cost in naive mice. Through the use of rationally designed oral vaccines, we induced immunoglobulin A responses blocking all of these trajectories. This selected for Salmonella mutants carrying deletions of the O-antigen polymerase gene wzyB. Due to their short O-antigen, these evolved mutants were more susceptible to environmental stressors (detergents or complement) and predation (bacteriophages) and were impaired in gut colonization and virulence in mice. Therefore, a rationally induced cocktail of intestinal antibodies can direct an evolutionary trade-off in S.Tm. This lays the foundations for the exploration of mucosal vaccines capable of setting evolutionary traps as a prophylactic strategy.

The human microbiome has emerged as a central research topic in human biology and biomedicine. Current microbiome studies generate high-throughput omics data across different body sites, populations, and life stages. Many of the challenges in microbiome research are similar to other high-throughput studies, the quantitative analyses need to address the heterogeneity of data, specific statistical properties, and the remarkable variation in microbiome composition across individuals and body sites. This has led to a broad spectrum of statistical and machine learning challenges that range from study design, data processing, and standardization to analysis, modeling, cross-study comparison, prediction, data science ecosystems, and reproducible reporting. Nevertheless, although many statistics and machine learning approaches and tools have been developed, new techniques are needed to deal with emerging applications and the vast heterogeneity of microbiome data. We review and discuss emerging applications of statistical and machine learning techniques in human microbiome studies and introduce the COST Action CA18131 "ML4Microbiome" that brings together microbiome researchers and machine learning experts to address current challenges such as standardization of analysis pipelines for reproducibility of data analysis results, benchmarking, improvement, or development of existing and new tools and ontologies., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Moreno-Indias, Lahti, Nedyalkova, Elbere, Roshchupkin, Adilovic, Aydemir, Bakir-Gungor, Santa Pau, D’Elia, Desai, Falquet, Gundogdu, Hron, Klammsteiner, Lopes, Marcos-Zambrano, Marques, Mason, May, Pašić, Pio, Pongor, Promponas, Przymus, Saez-Rodriguez, Sampri, Shigdel, Stres, Suharoschi, Truu, Truică, Vilne, Vlachakis, Yilmaz, Zeller, Zomer, Gómez-Cabrero and Claesson.)

Whole genome sequencing (WGS) enables high resolution typing of bacteria up to the single nucleotide polymorphism (SNP) level. WGS is used in clinical microbiology laboratories for infection control, molecular surveillance and outbreak analyses. Given the large palette of WGS reagents and bioinformatics tools, the Swiss clinical bacteriology community decided to conduct a ring trial (RT) to foster harmonization of NGS-based bacterial typing. The RT aimed at assessing methicillin-susceptible Staphylococcus aureus strain relatedness from WGS and epidemiological data. The RT was designed to disentangle the variability arising from differences in sample preparation, SNP calling and phylogenetic methods. Nine laboratories participated. The resulting phylogenetic tree and cluster identification were highly reproducible across the laboratories. Cluster interpretation was, however, more laboratory dependent, suggesting that an increased sharing of expertise across laboratories would contribute to further harmonization of practices. More detailed bioinformatic analyses unveiled that while similar clusters were found across laboratories, these were actually based on different sets of SNPs, differentially retained after sample preparation and SNP calling procedures. Despite this, the observed number of SNP differences between pairs of strains, an important criterion to determine strain relatedness given epidemiological information, was similar across pipelines for closely related strains when restricting SNP calls to a common core genome defined by S. aureus cgMLST schema. The lessons learned from this pilot study will serve the implementation of larger-scale RT, as a mean to have regular external quality assessments for laboratories performing WGS analyses in a clinical setting., (Copyright © 2020 Dylus, Pillonel, Opota, Wüthrich, Seth-Smith, Egli, Leo, Lazarevic, Schrenzel, Laurent, Bertelli, Blanc, Neuenschwander, Ramette, Falquet, Imkamp, Keller, Kahles, Oberhaensli, Barbié, Dessimoz, Greub and Lebrand.)

Microbially-mediated methylation of arsenic (As) plays an important role in the As biogeochemical cycle, particularly in rice paddy soils where methylated As, generated microbially, is translocated into rice grains. The presence of the arsenite (As(III)) methyltransferase gene ( arsM ) in soil microbes has been used as an indication of their capacity for As methylation. Here, we evaluate the ability of seven microorganisms encoding active ArsM enzymes to methylate As. Amongst those, only the aerobic species were efficient methylators. The anaerobic microorganisms presented high resistance to As exposure, presumably through their efficient As(III) efflux, but methylated As poorly. The only exception were methanogens, for which efficient As methylation was seemingly an artifact of membrane disruption. Deletion of an efflux pump gene ( acr3 ) in one of the anaerobes, Clostridium pasteurianum , rendered the strain sensitive to As and capable of more efficiently methylating As. Our results led to the following conclusions: (i) encoding a functional ArsM enzyme does not guarantee that a microorganism will actively drive As methylation in the presence of the metalloid and (ii) there is an inverse relationship between efficient microbial As efflux and its methylation, because the former prevents the intracellular accumulation of As.

Background: Providencia rettgeri is a nosocomial pathogen associated with urinary tract infections and related to Healthcare-Associated Infection (HAI). In recent years isolates producing New Delhi Metallo-β-lactamase (NDM) and other β-lactamases have been reported that reduce the efficiency of clinical antimicrobial treatments. In this study, we analyzed antibiotic resistance, the presence of resistance genes and the clonal relationship of two P. rettgeri isolates obtained from male patients admitted to the same hospital in Bogotá - Colombia, 2015., Results: Antibiotic susceptibility profile evaluated by the Kirby-Bauer method revealed that both isolates were resistant to third-generation carbapenems and cephalosporins. Whole-genome sequencing (Illumina HiSeq) followed by SPAdes assembling, Prokka annotation in combination with an in-house Python program and resistance gene detection by ResFinder identified the same six β-lactamase genes in both isolates: bla NDM-1 , bla VIM-2 , bla CTX-M-15 , bla OXA-10 , bla CMY-2 and bla TEM-1 . Additionally, various resistance genes associated with antibiotic target alteration (arnA, PmrE, PmrF, LpxA, LpxC, gyrB, folP, murA, rpoB, rpsL, tet34) were found and four efflux pumps (RosAB, EmrD, mdtH and cmlA). The additional resistance to gentamicin in one of the two isolates could be explained by a detected SNP in CpxA (Cys191Arg) which is involved in the stress response of the bacterial envelope. Genome BLAST comparison using CGView, the ANI value (99.99%) and the pangenome (using Roary) phylogenetic tree (same clade, small distance) showed high similarity between the isolates. The rMLST analysis indicated that both isolates were typed as rST-61,696, same as the RB151 isolate previously isolated in Bucaramanga, Colombia, 2013, and the FDAARGOS&#95;330 isolate isolated in the USA, 2015., Conclusions: We report the coexistence of the carbapenemase genes bla NDM-1 , and bla VIM-2 , together with the β-lactamase genes bla CTX-M-15 , bla OXA-10 , bla CMY-2 and bla TEM-1 , in P. rettgeri isolates from two patients in Colombia. Whole-genome sequence analysis indicated a circulation of P. rettgeri rST-61,696 strains in America that needs to be investigated further.