Isolation of a PRD1-like phage uncovers the carriage of three putative conjugative plasmids in clinical Burkholderia contaminans.

The Burkholderia cepacia complex (Bcc) is a group of increasingly multi-drug resistant opportunistic bacteria. This resistance is driven through a combination of intrinsic factors and the carriage of a broad range of conjugative plasmids harbouring virulence determinants. Therefore, novel treatments are required to treat and prevent further spread of these virulence determinants. In the search for phages infective for clinical Bcc isolates, CSP1 phage, a PRD1-like phage was isolated. CSP1 phage was found to require pilus machinery commonly encoded on conjugative plasmids to facilitate infection of Gram-negative bacteria genera including Escherichia and Pseudomonas. Whole genome sequencing and characterisation of one of the clinical Burkholderia isolates revealed it to be Burkholderia contaminans. B. contaminans 5080 was found to contain a genome of over 8 Mbp encoding multiple intrinsic resistance factors, such as efflux pump systems, but more interestingly, carried three novel plasmids encoding multiple putative virulence factors for increased host fitness, including antimicrobial resistance. Even though PRD1-like phages are broad host range, their use in novel antimicrobial treatments shouldn't be dismissed, as the dissemination potential of conjugative plasmids is extensive. Continued survey of clinical bacterial strains is also key to understanding the spread of antimicrobial resistance determinants and plasmid evolution.

Burkholderia contaminans Bacteriophage CSP3 Requires O-Antigen Polysaccharides for Infection.

The Burkholderia cepacia complex is a group of opportunistic pathogens that cause both severe acute and chronic respiratory infections. Due to their large genomes containing multiple intrinsic and acquired antimicrobial resistance mechanisms, treatment is often difficult and prolonged. One alternative to traditional antibiotics for treatment of bacterial infections is bacteriophages. Therefore, the characterization of bacteriophages infective for the Burkholderia cepacia complex is critical to determine their suitability for any future use. Here, we describe the isolation and characterization of novel phage, CSP3, infective against a clinical isolate of Burkholderia contaminans. CSP3 is a new member of the Lessievirus genus that targets various Burkholderia cepacia complex organisms. Single nucleotide polymorphism (SNP) analysis of CSP3-resistant B. contaminans showed that mutations to the O-antigen ligase gene, waaL, consequently inhibited CSP3 infection. This mutant phenotype is predicted to result in the loss of cell surface O-antigen, contrary to a related phage that requires the inner core of the lipopolysaccharide for infection. Additionally, liquid infection assays showed that CSP3 provides suppression of B. contaminans growth for up to 14 h. Despite the inclusion of genes that are typical of the phage lysogenic life cycle, we saw no evidence of CSP3's ability to lysogenize. Continuation of phage isolation and characterization is crucial in developing large and diverse phage banks for global usage in cases of antibiotic-resistant bacterial infections. IMPORTANCE Amid the global antibiotic resistance crisis, novel antimicrobials are needed to treat problematic bacterial infections, including those from the Burkholderia cepacia complex. One such alternative is the use of bacteriophages; however, a lot is still unknown about their biology. Bacteriophage characterization studies are of high importance for building phage banks, as future work in developing treatments such as phage cocktails should require well-characterized phages. Here, we report the isolation and characterization of a novel Burkholderia contaminans phage that requires the O-antigen for infection, a distinct phenotype seen among other related phages. Our findings presented in this article expand on the ever-evolving phage biology field, uncovering unique phage-host relationships and mechanisms of infection.

Tyroviruses are a new group of temperate phages that infect Bacillus species in soil environments worldwide.

BACKGROUND: Bacteriophages are widely considered to be highly abundant and genetically diverse, with their role in the evolution and virulence of many pathogens becoming increasingly clear. Less attention has been paid on phages preying on Bacillus, despite the potential for some of its members, such as Bacillus anthracis, to cause serious human disease. RESULTS: We have isolated five phages infecting the causative agent of anthrax, Bacillus anthracis. Using modern phylogenetic approaches we place these five new Bacillus phages, as well as 21 similar phage genomes retrieved from publicly available databases and metagenomic datasets into the Tyrovirus group, a newly proposed group named so due to the conservation of three distinct tyrosine recombinases. Genomic analysis of these large phages (~ 160-170 kb) reveals their DNA packaging mechanism and genomic features contributing to virion morphogenesis, host cell lysis and phage DNA replication processes. Analysis of the three tyrosine recombinases suggest Tyroviruses undergo a prophage lifecycle that may involve both host integration and plasmid stages. Further we show that Tyroviruses rely on divergent invasion mechanisms, with a subset requiring host S-layer for infection. CONCLUSIONS: Ultimately, we expand upon our understanding on the classification, phylogeny, and genomic organisation of a new and substantial phage group that prey on critically relevant Bacillus species. In an era characterised by a rapidly evolving landscape of phage genomics the deposition of future Tyroviruses will allow the further unravelling of the global spread and evolutionary history of these Bacillus phages.

Isolation and Characterisation of the Bundooravirus Genus and Phylogenetic Investigation of the Salasmaviridae Bacteriophages.

Bacillus is a highly diverse genus containing over 200 species that can be problematic in both industrial and medical settings. This is mainly attributed to Bacillus sp. being intrinsically resistant to an array of antimicrobial compounds, hence alternative treatment options are needed. In this study, two bacteriophages, PumA1 and PumA2 were isolated and characterized. Genome nucleotide analysis identified the two phages as novel at the DNA sequence level but contained proteins similar to phi29 and other related phages. Whole genome phylogenetic investigation of 34 phi29-like phages resulted in the formation of seven clusters that aligned with recent ICTV classifications. PumA1 and PumA2 share high genetic mosaicism and form a genus with another phage named WhyPhy, more recently isolated from the United States of America. The three phages within this cluster are the only candidates to infect B. pumilus. Sequence analysis of B. pumilus phage resistant mutants revealed that PumA1 and PumA2 require polymerized and peptidoglycan bound wall teichoic acid (WTA) for their infection. Bacteriophage classification is continuously evolving with the increasing phages' sequences in public databases. Understanding phage evolution by utilizing a combination of phylogenetic approaches provides invaluable information as phages become legitimate alternatives in both human health and industrial processes.

Bacteriophages in Natural and Artificial Environments.

Bacteriophages (phages) are biological entities that have attracted a great deal of attention in recent years. They have been reported as the most abundant biological entities on the planet and their ability to impact the composition of bacterial communities is of great interest. In this review, we aim to explore where phages exist in natural and artificial environments and how they impact communities. The natural environment in this review will focus on the human body, soils, and the marine environment. In these naturally occurring environments there is an abundance of phages suggesting a role in the maintenance of bacterial community homeostasis. The artificial environment focuses on wastewater treatment plants, industrial processes, followed by pharmaceutical formulations. As in natural environments, the existence of bacteria in manmade wastewater treatment plants and industrial processes inevitably attracts phages. The presence of phages in these environments can inhibit the bacteria required for efficient water treatment or food production. Alternatively, they can have a positive impact by eliminating recalcitrant organisms. Finally, we conclude by describing how phages can be manipulated or formulated into pharmaceutical products in the laboratory for use in natural or artificial environments.