École Nationale Vétérinaire de Toulouse

Transient infection of eukaryotic cells with commensal and extraintestinal pathogenic Escherichia coli of phylogenetic group B2 blocks mitosis and induces megalocytosis. This trait is linked to a widely spread genomic island that encodes giant modular nonribosomal peptide and polyketide synthases. Contact with E. coli expressing this gene cluster causes DNA double-strand breaks and activation of the DNA damage checkpoint pathway, leading to cell cycle arrest and eventually to cell death. Discovery of hybrid peptide-polyketide genotoxins in E. coli will change our view on pathogenesis and commensalism and open new biotechnological applications.

The List of Bacterial Names with Standing in Nomenclature includes, alphabetically and chronologically, the official names of bacteria as published or validated in the International Journal of Systematic Bacteriology. It encompasses 5,569 taxa (as of 31 December 1996) and is available on the Internet (URL: ftp:@ftp.cict.fr/pub/ bacterio/).

Motivation: Metagenomics leads to major advances in microbial ecology and biologists need user friendly tools to analyze their data on their own. Results: This Galaxy-supported pipeline, called FROGS, is designed to analyze large sets of amplicon sequences and produce abundance tables of Operational Taxonomic Units (OTUs) and their taxonomic affiliation. The clustering uses Swarm. The chimera removal uses VSEARCH, combined with original cross-sample validation. The taxonomic affiliation returns an innovative multi-affiliation output to highlight databases conflicts and uncertainties. Statistical results and numerous graphical illustrations are produced along the way to monitor the pipeline. FROGS was tested for the detection and quantification of OTUs on real and in silico datasets and proved to be rapid, robust and highly sensitive. It compares favorably with the widespread mothur, UPARSE and QIIME. Availability and implementation: Source code and instructions for installation: https://github.com/geraldinepascal/FROGS.git. A companion website: http://frogs.toulouse.inra.fr. Contact: geraldine.pascal@inra.fr. Supplementary information: Supplementary data are available at Bioinformatics online.

Escherichia coli is a normal inhabitant of the human gut. However, E. coli strains of phylogenetic group B2 harbor a genomic island called "pks" that codes for the production of a polyketide-peptide genotoxin, Colibactin. Here we report that in vivo infection with E. coli harboring the pks island, but not with a pks isogenic mutant, induced the formation of phosphorylated H2AX foci in mouse enterocytes. We show that a single, short exposure of cultured mammalian epithelial cells to live pks(+) E. coli at low infectious doses induced a transient DNA damage response followed by cell division with signs of incomplete DNA repair, leading to anaphase bridges and chromosome aberrations. Micronuclei, aneuploidy, ring chromosomes, and anaphase bridges persisted in dividing cells up to 21 d after infection, indicating occurrence of breakage-fusion-bridge cycles and chromosomal instability. Exposed cells exhibited a significant increase in gene mutation frequency and anchorage-independent colony formation, demonstrating the infection mutagenic and transforming potential. Therefore, colon colonization with these E. coli strains harboring the pks island could contribute to the development of sporadic colorectal cancer.

Enterohaemorrhagic Escherichia coli (EHEC) constitute a subset of serotypes (E. coli O157 and some other serogroups) of Shiga toxin (Stx)-producing E. coli (STEC) firmly associated with severe human illnesses like bloody diarrhoea and haemolytic uraemic syndrome. Stx production is essential but not sufficient for EHEC virulence. Most strains are capable of colonising the intestinal mucosa of the host with the "attaching and effacing" mechanism, genetically governed by a large pathogenicity island (PAI) defined as the Locus of Enterocyte Effacement. Other virulence factors carried by mobile genetic elements like PAI and plasmids have been recently described, and their role in the pathogenic process has not been fully elucidated. EHEC are zoonotic pathogens. They rarely cause disease in animals, and ruminants are recognised as their main natural reservoir. Cattle are considered to be the most important source of human infections with EHEC O157, and the ecology of the organism in cattle farming has been extensively studied. The organism has also been reported in sheep, goats, water buffalos, and deer. Pigs and poultry are not considered to be a source of EHEC and the sporadic reports may derive from accidental exposure to ruminant dejections. The epidemiology of EHEC infections has remarkably changed during the past ten years and an increasing number of unusual food vehicles have been associated with human infections. New routes of transmission have emerged, like contact with animals during farm visits and a wide variety of environment-related exposures. As for other zoonotic agents, having animals and raw products that are free from EHEC is not possible in practice. However, their occurrence can be minimised by applying high standards of hygiene in all the steps of the food production chain.

Staphylococci are the main aetiological agents of small ruminants intramammary infections (IMI), the more frequent isolates being S. aureus in clinical cases and coagulase negative species in subclinical IMI. The clinical IMI, whose annual incidence is usually lower than 5%, mainly occur at the beginning of machine milking and during the first third of lactation. These features constitute small ruminant peculiarities compared to dairy cattle. Small ruminant mastitis is generally a chronic and contagious infection: the primary sources are mammary and cutaneous carriages, and spreading mainly occurs during milking. Somatic cell counts (SCC) represent a valuable tool for prevalence assessment and screening, but predictive values are better in ewes than in goats. Prevention is most often based on milking machine management, sanitation and annual control, and milking technique optimisation. Elimination mainly relies on culling animals exhibiting clinical, chronic and recurrent IMI, and on drying-off intramammary antibiotherapy; this treatment allows a good efficacy and may be used selectively by targeting infected udders only. Heritability values for lactation mean SCC scores are between 0.11 and 0.15. Effective inclusion of ewe's mastitis resistance in the breeding goal has recently been implemented in France following experimental and large scale estimations of genetic parameters for SCC scores.

International recommendations for determination of reference intervals have been recently updated, especially for small reference sample groups, and use of the robust method and Box-Cox transformation is now recommended. Unfortunately, these methods are not included in most software programs used for data analysis by clinical laboratories. We have created a set of macroinstructions, named Reference Value Advisor, for use in Microsoft Excel to calculate reference limits applying different methods. For any series of data, Reference Value Advisor calculates reference limits (with 90% confidence intervals [CI]) using a nonparametric method when n≥40 and by parametric and robust methods from native and Box-Cox transformed values; tests normality of distributions using the Anderson-Darling test and outliers using Tukey and Dixon-Reed tests; displays the distribution of values in dot plots and histograms and constructs Q-Q plots for visual inspection of normality; and provides minimal guidelines in the form of comments based on international recommendations. The critical steps in determination of reference intervals are correct selection of as many reference individuals as possible and analysis of specimens in controlled preanalytical and analytical conditions. Computing tools cannot compensate for flaws in selection and size of the reference sample group and handling and analysis of samples. However, if those steps are performed properly, Reference Value Advisor, available as freeware at http://www.biostat.envt.fr/spip/spip.php?article63, permits rapid assessment and comparison of results calculated using different methods, including currently unavailable methods. This allows for selection of the most appropriate method, especially as the program provides the CI of limits. It should be useful in veterinary clinical pathology when only small reference sample groups are available.

Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that induces in humans a disease characterized by fever, rash, and pain in muscles and joints. The recent emergence or reemergence of CHIKV in the Indian Ocean Islands and India has stressed the need to better understand the pathogenesis of this disease. Previous CHIKV disease models have used young or immunodeficient mice, but these do not recapitulate human disease patterns and are unsuitable for testing immune-based therapies. Herein, we describe what we believe to be a new model for CHIKV infection in adult, immunocompetent cynomolgus macaques. CHIKV infection in these animals recapitulated the viral, clinical, and pathological features observed in human disease. In the macaques, long-term CHIKV infection was observed in joints, muscles, lymphoid organs, and liver, which could explain the long-lasting CHIKV disease symptoms observed in humans. In addition, the study identified macrophages as the main cellular reservoirs during the late stages of CHIKV infection in vivo. This model of CHIKV physiopathology should allow the development of new therapeutic and/or prophylactic strategies.

Human toxocariasis is a helminthozoonosis due to the migration of Toxocara species larvae through human organism. Humans become infected by ingesting either embryonated eggs from soil (geophagia, pica), dirty hands or raw vegetables, or larvae from undercooked giblets. The diagnosis relies upon sensitive immunological methods (ELISA or western-blot) which use Toxocara excretory-secretory antigens. Seroprevalence is high in developed countries, especially in rural areas, and also in some tropical islands. The clinical spectrum of the disease comprises four syndromes, namely visceral larva migrans, ocular larva migrans, and the more recently recognized "common" (in adults) and "covert" (in children) pictures. Therapy of ocular toxocariasis is primarily based upon corticosteroids use, when visceral larva migrans and few cases of common or covert toxocariasis can be treated by anthelmintics whose the most efficient appeared to be diethylcarbamazine. When diagnosed, all of these syndromes require thorough prevention of recontamination (especially by deworming pets) and sanitary education.

Whole-genome sequencing projects are increasingly populating the tree of life and characterizing biodiversity1–4. Sparse taxon sampling has previously been proposed to confound phylogenetic inference5, and captures only a fraction of the genomic diversity. Here we report a substantial step towards the dense representation of avian phylogenetic and molecular diversity, by analysing 363 genomes from 92.4% of bird families—including 267 newly sequenced genomes produced for phase II of the Bird 10,000 Genomes (B10K) Project. We use this comparative genome dataset in combination with a pipeline that leverages a reference-free whole-genome alignment to identify orthologous regions in greater numbers than has previously been possible and to recognize genomic novelties in particular bird lineages. The densely sampled alignment provides a single-base-pair map of selection, has more than doubled the fraction of bases that are confidently predicted to be under conservation and reveals extensive patterns of weak selection in predominantly non-coding DNA. Our results demonstrate that increasing the diversity of genomes used in comparative studies can reveal more shared and lineage-specific variation, and improve the investigation of genomic characteristics. We anticipate that this genomic resource will offer new perspectives on evolutionary processes in cross-species comparative analyses and assist in efforts to conserve species. A dataset of the genomes of 363 species from the Bird 10,000 Genomes Project shows increased power to detect shared and lineage-specific variation, demonstrating the importance of phylogenetically diverse taxon sampling in whole-genome sequencing.

Processed meat intake may be involved in the etiology of colorectal cancer, a major cause of death in affluent countries. The epidemiologic studies published to date conclude that the excess risk in the highest category of processed meat-eaters is comprised between 20% and 50% compared with non-eaters. In addition, the excess risk per gram of intake is clearly higher than that of fresh red meat. Several hypotheses, which are mainly based on studies carried out on red meat, may explain why processed meat intake is linked to cancer risk. Those that have been tested experimentally are (i) that high-fat diets could promote carcinogenesis via insulin resistance or fecal bile acids; (ii) that cooking meat at a high temperature forms carcinogenic heterocyclic amines and polycyclic aromatic hydrocarbons; (iii) that carcinogenic N-nitroso compounds are formed in meat and endogenously; (iv) that heme iron in red meat can promote carcinogenesis because it increases cell proliferation in the mucosa, through lipoperoxidation and/or cytotoxicity of fecal water. Nitrosation might increase the toxicity of heme in cured products. Solving this puzzle is a challenge that would permit to reduce cancer load by changing the processes rather than by banning processed meat.

Abstract Food-grade titanium dioxide (TiO 2 ) containing a nanoscale particle fraction (TiO 2 -NPs) is approved as a white pigment (E171 in Europe) in common foodstuffs, including confectionary. There are growing concerns that daily oral TiO 2 -NP intake is associated with an increased risk of chronic intestinal inflammation and carcinogenesis. In rats orally exposed for one week to E171 at human relevant levels, titanium was detected in the immune cells of Peyer’s patches (PP) as observed with the TiO 2 -NP model NM-105. Dendritic cell frequency increased in PP regardless of the TiO 2 treatment, while regulatory T cells involved in dampening inflammatory responses decreased with E171 only, an effect still observed after 100 days of treatment. In all TiO 2 -treated rats, stimulation of immune cells isolated from PP showed a decrease in Thelper (Th)-1 IFN-γ secretion, while splenic Th1/Th17 inflammatory responses sharply increased. E171 or NM-105 for one week did not initiate intestinal inflammation, while a 100-day E171 treatment promoted colon microinflammation and initiated preneoplastic lesions while also fostering the growth of aberrant crypt foci in a chemically induced carcinogenesis model. These data should be considered for risk assessments of the susceptibility to Th17-driven autoimmune diseases and to colorectal cancer in humans exposed to TiO 2 from dietary sources.

Enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic E. coli (EHEC) produce the characteristic "attaching and effacing" (A/E) lesion of the brush border. Intimin, an outer membrane protein encoded by eae, is responsible for the tight association of both pathogens with the host cell. Several eae have been cloned from different EPEC and EHEC strains isolated from humans and animals. These sequences are conserved in the N-terminal region but highly variable in the last C-terminal 280 amino acids (aa), where the cell binding activity is localized. Based on these considerations, we developed a panel of specific primers to investigate the eae heterogeneity of the variable 3' region by using PCR amplification. We then investigated the distribution of the known intimin types in a large collection of EPEC and EHEC strains isolated from humans and different animal species. The existence of a yet-unknown family of intimin was suspected because several EHEC strains, isolated from human and cattle, did not react with any of the specific primer pairs, although these strains were eae positive when primers amplifying the conserved 5' end were used. We then cloned and sequenced the eae present in one of these strains (EHEC of serotype O103:H2) and subsequently designed a PCR primer that recognizes in a specific manner the variable 3' region of this new intimin type. This intimin, referred to as "epsilon," was present in human and bovine EHEC strains of serogroups O8, O11, O45, O103, O121, and O165. Intimin epsilon is the largest intimin cloned to date (948 aa) and shares the greatest overall sequence identity with intimin beta, although analysis of the last C-terminal 280 aa suggests a greater similarity with intimins alpha and gamma.

The immune system is known to be involved in the early phase of scrapie pathogenesis. However, the infection route of naturally occurring scrapie and its spread within the host are not entirely known. In this study, the pathogenesis of scrapie was investigated in sheep of three PrP genotypes, from 2 to 9 months of age, which were born and raised together in a naturally scrapie-affected Romanov flock. The kinetics of PrP(Sc) accumulation in sheep organs were determined by immunohistochemistry. PrP(Sc) was detected only in susceptible VRQ/VRQ sheep, from 2 months of age, with an apparent entry site at the ileal Peyer's patch as well as its draining mesenteric lymph node. At the cellular level, PrP(Sc) deposits were associated with CD68-positive cells of the dome area and B follicles before being detected in follicular dendritic cells. In 3- to 6-month-old sheep, PrP(Sc) was detected in most of the gut-associated lymphoid tissues (GALT) and to a lesser extent in more systemic lymphoid formations such as the spleen or the mediastinal lymph node. All secondary lymphoid organs showed a similar intensity of PrP(Sc)-immunolabelling at 9 months of age. At this time-point, PrP(Sc) was also detected in the autonomic myenteric nervous plexus and in the nucleus parasympathicus nervi X of the brain stem. These data suggest that natural scrapie infection occurs by the oral route via infection of the Peyer's patches followed by replication in the GALT. It may then spread to the central nervous system through the autonomic nervous fibres innervating the digestive tract.

Prediction of genetic merit or disease risk using genetic marker information is becoming a common practice for selection of livestock and plant species. For the successful application of genome-wide marker-assisted selection (GWMAS), genomic predictions should be accurate and unbiased. The effect of selection on bias and accuracy of genomic predictions was studied in two simulated animal populations under weak or strong selection and with several heritabilities. Prediction of genetic values was by best-linear unbiased prediction (BLUP) using data either from relatives summarized in pseudodata for genotyped individuals (multiple-step method) or using all available data jointly (single-step method). The single-step method combined genomic- and pedigree-based relationship matrices. Predictions by the multiple-step method were biased. Predictions by a single-step method were less biased and more accurate but under strong selection were less accurate. When genomic relationships were shifted by a constant, the single-step method was unbiased and the most accurate. The value of that constant, which adjusts for non-random selection of genotyped individuals, can be derived analytically.

There are huge numbers of wild animals distributed throughout the world and the diversity of wildlife species is immense. Each landscape and habitat has a kaleidoscope of niches supporting an enormous variety of vertebrate and invertebrate species, and each species or taxon supports an even more impressive array of macro- and micro-parasites. Infectious pathogens that originate in wild animals have become increasingly important throughout the world in recent decades, as they have had substantial impacts on human health, agricultural production, wildlife-based economies and wildlife conservation. The emergence of these pathogens as significant health issues is associated with a range of causal factors, most of them linked to the sharp and exponential rise of global human activity. Among these causal factors are the burgeoning human population, the increased frequency and speed of local and international travel, the increase in human-assisted movement of animals and animal products, changing agricultural practices that favour the transfer of pathogens between wild and domestic animals, and a range of environmental changes that alter the distribution of wild hosts and vectors and thus facilitate the transmission of infectious agents. Two different patterns of transmission of pathogens from wild animals to humans are evident among these emerging zoonotic diseases. In one pattern, actual transmission of the pathogen to humans is a rare event but, once it has occurred, human-to-human transmission maintains the infection for some period of time or permanently. Some examples of pathogens with this pattern of transmission are human immunodeficiency virus/acquired immune deficiency syndrome, influenza A, Ebola virus and severe acute respiratory syndrome. In the second pattern, direct or vector-mediated animal-to-human transmission is the usual source of human infection. Wild animal populations are the principal reservoirs of the pathogen and human-to-human disease transmission is rare. Examples of pathogens with this pattern of transmission include rabies and other lyssaviruses, Nipah virus, West Nile virus, Hantavirus, and the agents of Lyme borreliosis, plague, tularemia, leptospirosis and ehrlichiosis. These zoonotic diseases from wild animal sources all have trends that are rising sharply upwards. In this paper, the authors discuss the causal factors associated with the emergence or re-emergence of these zoonoses, and highlight a selection to provide a composite view of their range, variety and origins. However, most of these diseases are covered in more detail in dedicated papers elsewhere in this Review.

Objective Campylobacter jejuni produces a genotoxin, cytolethal distending toxin (CDT), which has DNAse activity and causes DNA double-strand breaks. Although C. jejuni infection has been shown to promote intestinal inflammation, the impact of this bacterium on carcinogenesis has never been examined. Design Germ-free (GF) Apc Min/+ mice, fed with 1% dextran sulfate sodium, were used to test tumorigenesis potential of CDT-producing C. jejuni . Cells and enteroids were exposed to bacterial lysates to determine DNA damage capacity via γH2AX immunofluorescence, comet assay and cell cycle assay. To examine the interplay of CDT-producing C. jejuni , gut microbiome and host in tumorigenesis, colonic RNA-sequencing and faecal 16S rDNA sequencing were performed. Rapamycin was administrated to investigate the prevention of CDT-producing C. jejuni -induced tumorigenesis. Results GF Apc Min/+ mice colonised with human clinical isolate C. jejuni 81–176 developed significantly more and larger tumours when compared with uninfected mice. C. jejuni with a mutated cdtB subunit, mut cdtB , attenuated C. jejuni -induced tumorigenesis in vivo and decreased DNA damage response in cells and enteroids. C. jejuni infection induced expression of hundreds of colonic genes, with 22 genes dependent on the presence of c dtB. The C. jejuni -infected group had a significantly different microbial gene expression profile compared with the mut cdtB group as shown by metatranscriptomic data, and different microbial communities as measured by 16S rDNA sequencing. Finally, rapamycin could diminish the tumorigenic capability of C. jejuni . Conclusion Human clinical isolate C. jejuni 81–176 promotes colorectal cancer and induces changes in microbial composition and transcriptomic responses, a process dependent on CDT production.

Red meat and processed meat intake is associated with a risk of colorectal cancer, a major cause of death in affluent countries. Epidemiological and experimental evidence supports the hypothesis that heme iron present in meat promotes colorectal cancer. This meta-analysis of prospective cohort studies of colon cancer reporting heme intake included 566,607 individuals and 4,734 cases of colon cancer. The relative risk of colon cancer was 1.18 (95% CI: 1.06-1.32) for subjects in the highest category of heme iron intake compared with those in the lowest category. Epidemiological data thus show a suggestive association between dietary heme and risk of colon cancer. The analysis of experimental studies in rats with chemically-induced colon cancer showed that dietary hemoglobin and red meat consistently promote aberrant crypt foci, a putative precancer lesion. The mechanism is not known, but heme iron has a catalytic effect on (i) the endogenous formation of carcinogenic N-nitroso compounds and (ii) the formation of cytotoxic and genotoxic aldehydes by lipoperoxidation. A review of evidence supporting these hypotheses suggests that both pathways are involved in heme iron toxicity.

Terminal plasma half-life is the time required to divide the plasma concentration by two after reaching pseudo-equilibrium, and not the time required to eliminate half the administered dose. When the process of absorption is not a limiting factor, half-life is a hybrid parameter controlled by plasma clearance and extent of distribution. In contrast, when the process of absorption is a limiting factor, the terminal half-life reflects rate and extent of absorption and not the elimination process (flip-flop pharmacokinetics). The terminal half-life is especially relevant to multiple dosing regimens, because it controls the degree of drug accumulation, concentration fluctuations and the time taken to reach equilibrium.

The evolutionary basis of domestication has been a longstanding question and its genetic architecture is becoming more tractable as more domestic species become genome-enabled. Before becoming established worldwide, sheep and goats were domesticated in the fertile crescent 10,500 years before present (YBP) where their wild relatives remain. Here we sequence the genomes of wild Asiatic mouflon and Bezoar ibex in the sheep and goat domestication center and compare their genomes with that of domestics from local, traditional, and improved breeds. Among the genomic regions carrying selective sweeps differentiating domestic breeds from wild populations, which are associated among others to genes involved in nervous system, immunity and productivity traits, 20 are common to Capra and Ovis. The patterns of selection vary between species, suggesting that while common targets of selection related to domestication and improvement exist, different solutions have arisen to achieve similar phenotypic end-points within these closely related livestock species.