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Free-ranging and feral dogs represent a group of unattended companion animals. They impact wild animal populations by predating native species, displacing predators and introducing exotic pathogens. The aim of this work was to describe the molecular occurrence of Rickettsia, Ehrlichia, Anaplasma, Mycoplasma and Bartonella in feral dogs. The study was carried out in the last relict of a protected area in Mexico City. Blood clots samples from 19 dogs were obtained and analyzed for detection of specific fragments of the 16S-rRNA gene for Anaplasma, Ehrlichia and Mycoplasma and citrate synthase (gltA) for Bartonella and Rickettsia. Our results showed that DNA from three bacteria species (Bartonella vinsonii subsp. berkhoffii, Ehrlichia canis and Mycoplasma haemocanis) was present with frequencies ranging from 5.3 to 15.8%. This is the first record of B. vinsonii subsp. berkhoffii and M. haemocanis in dogs from México, and also the first finding of Ehrlichia canis in Mexico City. It is important to perform surveillance of feral dog populations in order to identify the impact of these pathogens on wild animal populations and Public Health in order to establish prevention and protection programs.

SUMMARY Mechanistic insights into the role of the human microbiome in the predisposition to and treatment of disease are limited by the lack of methods to precisely add or remove microbial strains or genes from complex communities. Here, we demonstrate that engineered bacteriophage M13 can be used to deliver DNA to Escherichia coli within the mouse gastrointestinal (GI) tract. Delivery of a programmable exogenous CRISPR-Cas9 system enables the strain-specific depletion of fluorescently marked isogenic strains during competitive colonization and genomic deletions that encompass the target gene in mice colonized with a single strain. Multiple mechanisms allow E. coli to escape targeting, including loss of the CRISPR array or even the entire CRISPR-Cas9 system. These results provide a robust and experimentally tractable platform for microbiome editing, a foundation for the refinement of this approach to increase targeting efficiency, and a proof of concept for the extension to other phage-bacterial pairs of interest., Graphical Abstract, In brief Lam et al. show that filamentous bacteriophage can be harnessed as agents of gene delivery to bacteria colonizing the gastrointestinal tract. Using M13 to deliver CRISPR-Cas9, they demonstrate sequence-specific targeting of GFP-marked E. coli in the gut and show that CRISPR-Cas9 can induce genomic deletions at the target site.

Despite the remarkable microbial diversity found within humans, our ability to link genes to phenotypes is based upon a handful of model microorganisms. We report a comparative genomics platform for Eggerthella lenta and other Coriobacteriia, a neglected taxon broadly relevant to human health and disease. We uncover extensive genetic and metabolic diversity and validate a tool for mapping phenotypes to genes and sequence variants. We also present a tool for the quantification of strains from metagenomic sequencing data, enabling the identification of genes that predict bacterial fitness. Competitive growth is reproducible under laboratory conditions and attributable to intrinsic growth rates and resource utilization. Unique signatures of invivo competition ingnotobiotic mice include an adhesin enriched in poor colonizers. Together, these computational and experimental resources represent a strong foundation for the continued mechanistic dissection of the Coriobacteriia and a template that can be applied to study other genetically intractable taxa.

Bacteriophages are the most abundant entities found within the human gastrointestinal tract, yettheir interaction with bacterial hosts remain poorly understood, particularly with respect to CRISPRCas immunity. Here, we show that the Type IC CRISPRCas system in the prevalent gut Actinobacterium Eggerthella lenta i s active both transcriptionally and catalytically, cleaving foreign and self DNA. Comparative analyses of E. lenta CRISPRCas systems across genomes and metagenomes revealed two distinct clades according to their shared cas gene sequence similarity and spacer content. To identify putative CRISPR targets, we assembled a human virome database (HuVirDB) encompassing 1,831 samples, revealing matches for the majority of the identified spacers, including “hypertargeted” phage with as many as 96 protospacers. Modeling observed spacer/protospacer pairs, we mapped positional mismatch tolerance in crRNAs. This work emphasizes the utility of merging computational and experimental approaches for determining the function and targets of human gut bacterial CRISPRCas systems.

Despite the remarkable evolutionary and metabolic diversity found within the human microbiome, the vast majority of mechanistic studies focus on two phyla: the Bacteroidetes and the Proteobacteria. Generalizable tools for studying the other phyla are urgently needed in order to transition microbiome research from a descriptive to a mechanistic discipline. Here, we focus on the Coriobacteriia class within the Actinobacteria phylum, detected in the distal gut of 90% of adult individuals around the world, which have been associated with both chronic and infectious disease, and play a key role in the metabolism of pharmaceutical, dietary, and endogenous compounds. We established, sequenced, and annotated a strain collection spanning 14 genera, 8 decades, and 3 continents, with a focus on Eggerthella lenta. Genome-wide alignments revealed inconsistencies in the taxonomy of the Coriobacteriia for which amendments have been proposed. Re-sequencing of the E. lenta type strain from multiple culture collections and our laboratory stock allowed us to identify errors in the finished genome and to identify point mutations associated with antibiotic resistance. Analysis of 24 E. lenta genomes revealed an “open” pan-genome suggesting we still have not fully sampled the genetic and metabolic diversity within this bacterial species. Consistent with the requirement for arginine during in vitro growth, the core E. lenta genome included the arginine dihydrolase pathway. Surprisingly, glycolysis and the citric acid cycle was also conserved in E. lenta despite the lack of evidence for carbohydrate utilization. We identified a species-specific marker gene and validated a multiplexed quantitative PCR assay for simultaneous detection of E. lenta and specific genes of interest from stool samples. Finally, we demonstrated the utility of comparative genomics for linking variable genes to strain-specific phenotypes, including antibiotic resistance and drug metabolism. To facilitate the continued functional genomic analysis of the Coriobacteriia, we have deposited the full collection of strains in DSMZ and have written a general software tool (ElenMatchR) that can be readily applied to novel phenotypic traits of interest. Together, these tools provide a first step towards a molecular understanding of the many neglected but clinically-relevant members of the human gut microbiome.

Bacteriophages are abundant within the human gastrointestinal tract, yet their interactions with gut bacteria remain poorly understood, particularly with respect to CRISPR-Cas immunity. Here, we show that the type I-C CRISPR-Cas system in the prevalent gut Actinobacterium Eggerthella lenta is transcribed and sufficient for specific targeting of foreign and chromosomal DNA. Comparative analyses of E.lenta CRISPR-Cas systems across (meta)genomes revealed 2 distinct clades according to cas sequence similarity and spacer content. We assembled a human virome database (HuVirDB), encompassing 1,831 samples enriched for viral DNA, to identify protospacers. This revealed matches for a majority of spacers, a marked increase over other databases, and uncovered "hyper-targeted" phage sequences containing multiple protospacers targeted by several E.lenta strains. Finally, we determined the positional mismatch tolerance of observed spacer-protospacer pairs. This work emphasizes the utility of merging computational and experimental approaches for determining the function and targets of CRISPR-Cas systems.

Currently, there is a growing demand to determine the protective status of vaccinated fish in order to prevent diseases outbreaks. A set of different parameters that include the infectious and immunological status of vaccinated salmonids from 622 Chilean farms were analyzed during 2011–2014. The aim of this study was to optimize the vaccination program of these centers through the determination of the protective state of vaccinated fish using oral immunizations. This state was determined from the association of the concentration of the immunoglobulin M (IgM) in the serum and the mortality rate of vaccinated fish. Salmonids were vaccinated with different commercial mono- or polyvalent vaccines against salmonid rickettsial septicemia (SRS) and infectious salmon anemia (ISA), first by the intraperitoneal injection of oil-adjuvanted antigens and then by the stimulation of mucosal immunity using oral vaccines as a booster vaccination. The results showed that high levels of specific IgM antibodies were observed after injectable vaccination, reaching a maximum concentration at 600–800 degree-days. Similar levels of antibodies were observed when oral immunizations were administrated. The high concentration of antibodies [above 2750 ng/mL for ISA virus (ISAv) and 3500 ng/mL for SRS] was maintained for a period of 800 degree-days after each vaccination procedure. In this regard, oral immunizations maintained a long-term high concentration of anti-SRS and anti-ISAv specific IgM antibodies. When the concentration of antibodies decreased below 2000 pg/mL, a window of susceptibility to SRS infection was observed in the farm, suggesting a close association between antibody levels and fish protective status. These results demonstrated that, in the field, several oral immunizations are essential to uphold a high level of specific anti-pathogens antibodies and, therefore, the protective status during the whole productive cycle.

Janthinobacterium lividum is a Gram-negative bacterium able to produce violacein, a pigment with antimicrobial and antitumor properties. Janthinobacterium lividum colonizes the skin of some amphibians and confers protection against fungal pathogens. The mechanisms underlying this association are not well understood. In order to identify the advantages for the bacterium to colonize amphibian skin we sequenced Janthinobacterium lividum strain MTR, a strain isolated from Cajón del Maipo, Chile. The strain has capnophilic behavior, with growth favored by high concentrations (5 %) of carbon dioxide. Its genome is 6,535,606 bp in size, with 5,362 coding sequences and a G + C content of 62.37 %. The presence of genes encoding for products that participate in the carbon fixation pathways (dark CAM pathways), and the entire set of genes encoding for the enzymes of the glyoxylate cycle may explain the capnophilic behavior and allow us to propose that the CO2 secreted by the skin of amphibians is the signal molecule that guides colonization by Janthinobacterium lividum. Electronic supplementary material The online version of this article (doi:10.1186/s40793-015-0104-z) contains supplementary material, which is available to authorized users.