Genome Sequencing and Analysis of a Type A Clostridium perfringens Isolate from a Case of Bovine Clostridial Abomasitis

Clostridium perfringens is a common inhabitant of the avian and mammalian gastrointestinal tracts and can behave commensally or pathogenically. Some enteric diseases caused by type A C. perfringens, including bovine clostridial abomasitis, remain poorly understood. To investigate the potential basis of virulence in strains causing this disease, we sequenced the genome of a type A C. perfringens isolate (strain F262) from a case of bovine clostridial abomasitis. The ∼3.34 Mbp chromosome of C. perfringens F262 is predicted to contain 3163 protein-coding genes, 76 tRNA genes, and an integrated plasmid sequence, Cfrag (∼18 kb). In addition, sequences of two complete circular plasmids, pF262C (4.8 kb) and pF262D (9.1 kb), and two incomplete plasmid fragments, pF262A (48.5 kb) and pF262B (50.0 kb), were identified. Comparison of the chromosome sequence of C. perfringens F262 to complete C. perfringens chromosomes, plasmids and phages revealed 261 unique genes. No novel toxin genes related to previously described clostridial toxins were identified: 60% of the 261 unique genes were hypothetical proteins. There was a two base pair deletion in virS, a gene reported to encode the main sensor kinase involved in virulence gene activation. Despite this frameshift mutation, C. perfringens F262 expressed perfringolysin O, alpha-toxin and the beta2-toxin, suggesting that another regulation system might contribute to the pathogenicity of this strain. Two complete plasmids, pF262C (4.8 kb) and pF262D (9.1 kb), unique to this strain of C. perfringens were identified

The search for therapeutic bacteriophages uncovers one new subfamily and two new genera of Pseudomonas-infecting Myoviridae.

In a previous study, six virulent bacteriophages PAK_P1, PAK_P2, PAK_P3, PAK_P4, PAK_P5 and CHA_P1 were evaluated for their in vivo efficacy in treating Pseudomonas aeruginosa infections using a mouse model of lung infection. Here, we show that their genomes are closely related to five other Pseudomonas phages and allow a subdivision into two clades, PAK_P1-like and KPP10-like viruses, based on differences in genome size, %GC and genomic contents, as well as number of tRNAs. These two clades are well delineated, with a mean of 86% and 92% of proteins considered homologous within individual clades, and 25% proteins considered homologous between the two clades. By ESI-MS/MS analysis we determined that their virions are composed of at least 25 different proteins and electron microscopy revealed a morphology identical to the hallmark Salmonella phage Felix O1. A search for additional bacteriophage homologs, using profiles of protein families defined from the analysis of the 11 genomes, identified 10 additional candidates infecting hosts from different species. By carrying out a phylogenetic analysis using these 21 genomes we were able to define a new subfamily of viruses, the Felixounavirinae within the Myoviridae family. The new Felixounavirinae subfamily includes three genera: Felixounalikevirus, PAK_P1likevirus and KPP10likevirus. Sequencing genomes of bacteriophages with therapeutic potential increases the quantity of genomic data on closely related bacteriophages, leading to establishment of new taxonomic clades and the development of strategies for analyzing viral genomes as presented in this article

Efficacy and safety assessment of two enterococci phages in an in vitro biofilm wound model

Chronic wounds affect thousands of people worldwide, causing pain and discomfort to patients and represent significant economical burdens to health care systems. The treatment of chronic wounds is very difficult and complex, particularly when wounds are colonized by bacterial biofilms which are highly tolerant to antibiotics. Enterococcus faecium and Enterococcus faecalis are within the most frequent bacteria present in chronic wounds. Bacteriophages (phages) have been proposed as an efficient and alternative against antibiotic-resistant infections, as those found in chronic wounds. We have isolated and characterized two novel enterococci phages, the siphovirus vB_EfaS-Zip (Zip) and the podovirus vB_EfaP-Max (Max) to be applied during wound treatment. Both phages demonstrated lytic behavior against E. faecalis and E. faecium. Genome analysis of both phages suggests the absence of genes associated with lysogeny. A phage cocktail containing both phages was tested against biofilms formed in wound simulated conditions at a multiplicity of infection of 1.0 and a 2.5log CFU.mL1 reduction in the bacterial load after at 3h of treatment was observed. Phages were also tested in epithelial cells colonized by these bacterial species and a 3log CFU.mL1 reduction was observed using both phages. The high efficacy of these new isolated phages against multi-species biofilms, their stability at different temperatures and pH ranges, short latent periods and non-cytotoxicity to epithelial cells suggest their therapeutic use to control infectious biofilms present in chronic wounds.This study was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469/2013 unit, COMPETE 2020 (POCI-01-0145-FEDER-006684) and the Project PTDC/BBB-BSS/6471/2014 (POCI-01-0145-FEDER-016678). This work was also supported by BioTecNorte operation (NORTE-01-0145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte. A.R.C. and H.O. acknowledge FCT for grant SFRH/BPD/94648/2013 and SFRH/BPD/111653/2015, respectively. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.info:eu-repo/semantics/publishedVersio