PhageLeads: Rapid Assessment of Phage Therapeutic Suitability Using an Ensemble Machine Learning Approach.

The characterization of therapeutic phage genomes plays a crucial role in the success rate of phage therapies. There are three checkpoints that need to be examined for the selection of phage candidates, namely, the presence of temperate markers, antimicrobial resistance (AMR) genes, and virulence genes. However, currently, no single-step tools are available for this purpose. Hence, we have developed a tool capable of checking all three conditions required for the selection of suitable therapeutic phage candidates. This tool consists of an ensemble of machine-learning-based predictors for determining the presence of temperate markers (integrase, Cro/CI repressor, immunity repressor, DNA partitioning protein A, and antirepressor) along with the integration of the ABRicate tool to determine the presence of antibiotic resistance genes and virulence genes. Using the biological features of the temperate markers, we were able to predict the presence of the temperate markers with high MCC scores (>0.70), corresponding to the lifestyle of the phages with an accuracy of 96.5%. Additionally, the screening of 183 lytic phage genomes revealed that six phages were found to contain AMR or virulence genes, showing that not all lytic phages are suitable to be used for therapy. The suite of predictors, PhageLeads, along with the integrated ABRicate tool, can be accessed online for in silico selection of suitable therapeutic phage candidates from single genome or metagenomic contigs.

Biogeographic study of human gut-associated crAssphage suggests impacts from industrialization and recent expansion.

CrAssphage (cross-assembly phage) is a bacteriophage that was first discovered in human gut metagenomic data. CrAssphage belongs to a diverse family of crAss-like bacteriophages thought to infect gut commensal bacteria belonging to Bacteroides species. However, not much is known about the biogeography of crAssphage and whether certain strains are associated with specific human populations. In this study, we screened publicly available human gut metagenomic data from 3,341 samples for the presence of crAssphage sensu stricto (NC_024711.1). We found that crAssphage prevalence is low in traditional, hunter-gatherer populations, such as the Hadza from Tanzania and Matses from Peru, as compared to industrialized, urban populations. Statistical comparisons showed no association of crAssphage prevalence with variables such as age, sex, body mass index, and health status of individuals. Phylogenetic analyses show that crAssphage strains reconstructed from the same individual over multiple time-points, cluster together. CrAssphage strains from individuals from the same study population do not always cluster together. Some evidence of clustering is seen at the level of broadly defined geographic regions, however, the relative positions of these clusters within the crAssphage phylogeny are not well-supported. We hypothesize that this lack of strong biogeographic structuring is suggestive of an expansion event within crAssphage. Using a Bayesian dating approach, we estimate that this expansion has occurred fairly recently. Overall, we determine that crAssphage presence is associated with an industrialized lifestyle and the absence of strong biogeographic structuring within global crAssphage strains is likely due to a recent population expansion within this bacteriophage.

Filamentous phages: masters of a microbial sharing economy.

Bacteriophage ("bacteria eaters") or phage is the collective term for viruses that infect bacteria. While most phages are pathogens that kill their bacterial hosts, the filamentous phages of the sub-class Inoviridae live in cooperative relationships with their bacterial hosts, akin to the principal behaviours found in the modern-day sharing economy: peer-to-peer support, to offset any burden. Filamentous phages impose very little burden on bacteria and offset this by providing service to help build better biofilms, or provision of toxins and other factors that increase virulence, or modified behaviours that provide novel motile activity to their bacterial hosts. Past, present and future biotechnology applications have been built on this phage-host cooperativity, including DNA sequencing technology, tools for genetic engineering and molecular analysis of gene expression and protein production, and phage-display technologies for screening protein-ligand and protein-protein interactions. With the explosion of genome and metagenome sequencing surveys around the world, we are coming to realize that our knowledge of filamentous phage diversity remains at a tip-of-the-iceberg stage, promising that new biology and biotechnology are soon to come.

Classifying the Unclassified: A Phage Classification Method.

This work reports the method ClassiPhage to classify phage genomes using sequence derived taxonomic features. ClassiPhage uses a set of phage specific Hidden Markov Models (HMMs) generated from clusters of related proteins. The method was validated on all publicly available genomes of phages that are known to infect Vibrionaceae. The phages belong to the well-described phage families of Myoviridae, Podoviridae, Siphoviridae, and Inoviridae. The achieved classification is consistent with the assignments of the International Committee on Taxonomy of Viruses (ICTV), all tested phages were assigned to the corresponding group of the ICTV-database. In addition, 44 out of 58 genomes of Vibrio phages not yet classified could be assigned to a phage family. The remaining 14 genomes may represent phages of new families or subfamilies. Comparative genomics indicates that the ability of the approach to identify and classify phages is correlated to the conserved genomic organization. ClassiPhage classifies phages exclusively based on genome sequence data and can be applied on distinct phage genomes as well as on prophage regions within host genomes. Possible applications include (a) classifying phages from assembled metagenomes; and (b) the identification and classification of integrated prophages and the splitting of phage families into subfamilies.

Numbers of coliforms, Escherichia coli, F-RNA phage, rotavirus, bovine enteric calicivirus and presence of non-O157 STEC on commercial vacuum packaged beef.

The numbers of coliforms, Escherichia coli, F-RNA coliphages, bovine enteric calicivirus (BEC) and rotavirus (RV) and presence of non-O157 shiga toxigenic E. coli (STEC) were determined on commercial vacuum packaged beef subprimals at the retail level from swabs obtained from the entire surfaces of 150 cuts that originated from federally and provincially registered plants. The prevalence and log mean numbers of E. coli were higher in provincially registered plants than in federally registered plants; 64% vs 20%, respectively, and -0.3 vs -1.22 log cfu/100 cm(2), respectively. In contrast, the prevalence and mean log numbers of F-RNA coliphages were lower for the provincially registered plants than for the federally registered plants; 31% vs 68% and -0.86 vs -0.13 log cfu/100 cm(2), respectively. One E. coli sample tested positive for stx2 and eae. F-RNA coliphages associated with human origin (GII/GIII) were detected in 12% and 30% of samples that originated from provincially and federally registered plants, respectively. RV RNA was detected in 4% of samples while BEC RNA was not detected. Although the infectivity of RV is unknown, the presence of viable F-RNA coliphages suggests that consumers could potentially be at risk when consuming undercooked meat that is contaminated with RV.

Phage therapy.

There is a renaissance of interest in the antimicrobial potential of phages as more pathogens become multiply antibiotic resistant. Phage therapy is not a new concept, and it is important to ask why it is not part of the current repertoire of western medicine despite the fact that it has been continuously and extensively used in Eastern Europe for almost a century. Answering this question successfully will, largely, determine whether phage therapy can gain the credibility needed to overcome the scientific, financial and regulatory hurdles facing its adoption in mainstream clinical practice. Despite a paucity of such information from human studies, pharmacokinetic data and clinical outcomes from animal studies are currently providing convincing evidence for the safety and efficacy of phage therapy.

Is there a cost of virus resistance in marine cyanobacteria?

Owing to their abundance and diversity, it is generally perceived that viruses are important for structuring microbial communities and regulating biogeochemical cycles. The ecological impact of viruses on microbial food webs, however, may be influenced by evolutionary processes, including the ability of bacteria to evolve resistance to viruses and the theoretical prediction that this resistance should be accompanied by a fitness cost. We conducted experiments using phylogenetically distinct strains of marine Synechococcus (Cyanobacteria) to test for a cost of resistance (COR) to viral isolates collected from Mount Hope Bay, Rhode Island. In addition, we examined whether fitness costs (1) increased proportionally with 'total resistance', the number of viruses for which a strain had evolved resistance, or (2) were determined more by 'compositional resistance', the identity of the viruses to which it evolved resistance. A COR was only found in half of our experiments, which may be attributed to compensatory mutations or the inability to detect a small COR. When detected, the COR resulted in a approximately 20% reduction in relative fitness compared to ancestral strains. The COR was unaffected by total resistance, suggesting a pleiotropic fitness response. Under competitive conditions, however, the COR was dependent on compositional resistance, suggesting that fitness costs were associated with the identity of a few particular viruses. Our study provides the first evidence for a COR in marine bacteria, and suggests that Synechococcus production may be influenced by the composition of co-occurring viruses.

Isolation and characterization of five Erwinia amylovora bacteriophages and assessment of phage resistance in strains of Erwinia amylovora.

Phages able to infect the fire blight pathogen Erwinia amylovora were isolated from apple, pear, and raspberry tissues and from soil samples collected at sites displaying fire blight symptoms. Among a collection of 50 phage isolates, 5 distinct phages, including relatives of the previously described phages phiEa1 and phiEa7 and 3 novel phages named phiEa100, phiEa125, and phiEa116C, were identified based on differences in genome size and restriction fragment pattern. phiEa1, the phage distributed most widely, had an approximately 46-kb genome which exhibited some restriction site variability between isolates. Phages phiEa100, phiEa7, and phiEa125 each had genomes of approximately 35 kb and could be distinguished by their EcoRI restriction fragment patterns. phiEa116C contained an approximately 75-kb genome. phiEa1, phiEa7, phiEa100, phiEa125, and phiEa116C were able to infect 39, 36, 16, 20, and 40, respectively, of 40 E. amylovora strains isolated from apple orchards in Michigan and 8, 12, 10, 10, and 12, respectively, of 12 E. amylovora strains isolated from raspberry fields (Rubus spp.) in Michigan. Only 22 of 52 strains were sensitive to all five phages, and 23 strains exhibited resistance to more than one phage. phiEa116C was more effective than the other phages at lysing E. amylovora strain Ea110 in liquid culture, reducing the final titer of Ea110 by >95% when added at a ratio of 1 PFU per 10 CFU and by 58 to 90% at 1 PFU per 10(5) CFU.