Staphylococcus aureus temperate bacteriophage: carriage and horizontal gene transfer is lineage associated.

Staphylococcus aureus is a major cause of human and animal infections. Bacteriophage are a class of mobile genetic element (MGE) that carry virulence genes and disseminate them horizontally, including Panton-Valentine leukocidin (PVL), the immune evasion cluster (IEC) associated with human specificity, and enterotoxin A the major toxin associated with food poisoning. S. aureus isolates group into major clonal complex (CC) lineages that largely evolve independently due to possession of different restriction-modification (RM) systems. We aimed to better understand the horizontal and vertical transmission dynamics of virulence and resistance genes by bacteriophage by using (i) bioinformatic approaches to analyze bacteriophage genomes from the first 79 sequenced S. aureus isolates and (ii) S. aureus microarrays to analyze the distribution of bacteriophage and virulence genes in S. aureus isolates from a broader range of lineages. The distribution of eight bacteriophage families was highly variable but lineage associated. Nevertheless, there was evidence of frequent acquisition and loss and not just vertical transmission. Most bacteriophage genes were dispensable, and extensive mosaicism was seen. Surprisingly, virulence genes were tightly associated with specific phage families. This data suggests S. aureus bacteriophage evolve rapidly, and the horizontal gene transfer (HGT) of virulence genes encoded by bacteriophage is restricted by bacteriophage family and the lineage of the host bacterium, delaying the evolution of fully resistant and virulent strains

Investigating bacteriophages targeting the opportunistic pathogen Acinetobacter baumannii

The multi-drug resistance of the opportunistic pathogen Acinetobacter baumannii is of growing concern, with many clinical isolates proving to be resistant to last resort as well as front line antibiotic treatments. The use of bacteriophages is an attractive alternative to controlling and treating this emerging nosocomial pathogen. In this study, we have investigated bacteriophages collected from hospital wastewater in Thailand and we have explored their activity against clinical isolates of A. baumannii. Bacteriophage vB_AbaM_PhT2 showed 28% host range against 150 multidrug resistant (MDR) isolates and whole genome sequencing did not detect any known virulence factors or antibiotic resistance genes. Purified vB_AbaM_PhT2 samples had endotoxin levels below those recommended for preclinical trials and were not shown to be directly cytotoxic to human cell lines in vitro. The treatment of human brain and bladder cell lines grown in the presence of A. baumannii with this bacteriophage released significantly less lactate dehydrogenase compared to samples with no bacteriophage treatment, indicating that vB_AbaM_PhT2 can protect from A. baumannii induced cellular damage. Our results have also indicated that there is synergy between this bacteriophage and the end line antibiotic colistin. We therefore propose bacteriophage vB_AbaM_PhT2 as a good candidate for future research and for its potential development into a surface antimicrobial for use in hospitals. View Full-Tex

Structural modification of polysaccharides: A biochemical-genetic approach

Polysaccharides have a wide range of industrial and biomedical applications. An industry trend is underway towards the increased use of bacteria to produce polysaccharides. Long term goals of this work are the adaptation and enhancement of saccharide properties for electronic and optic applications. In this report we illustrate the application of enzyme-bearing bacteriophage on strains of the enteric bacterium Klebsiella pneumoniae, which produces a polysaccharide with the relatively rare rheological property of drag-reduction. This has resulted in the production of new polysaccharides with enhanced rheological properties. Our laboratory is developing techniques for processing and structurally modifying bacterial polysaccharides and oligosaccharides which comprise their basic polymeric repeat units. Our research has focused on bacteriophage which produce specific polysaccharide degrading enzymes. This has lead to the development of enzymes generated by bacteriophage as tools for polysaccharide modification and purification. These enzymes were used to efficiently convert the native material to uniform-sized high molecular weight polymers, or alternatively into high-purity oligosaccharides. Enzyme-bearing bacteriophage also serve as genetic selection tools for bacteria that produce new families of polysaccharides with modified structures

The Xp10 bacteriophage protein P7 inhibits transcription by the major and major variant forms of the host RNA polymerase via a common mechanism

The σ factor is a functionally obligatory subunit of the bacterial transcription machinery, the RNA polymerase. Bacteriophage-encoded small proteins that either modulate or inhibit the bacterial RNAP to allow the temporal regulation of bacteriophage gene expression often target the activity of the major bacterial σ factor, σ70. Previously, we showed that during Xanthomonas oryzae phage Xp10 infection, the phage protein P7 inhibits the host RNAP by preventing the productive engagement with the promoter and simultaneously displaces the σ70 factor from the RNAP. In this study, we demonstrate that P7 also inhibits the productive engagement of the bacterial RNAP containing the major variant bacterial σ factor, σ54, with its cognate promoter. The results suggest for the first time that the major variant form of the host RNAP can also be targeted by bacteriophage-encoded transcription regulatory proteins. Since the major and major variant σ factor interacting surfaces in the RNAP substantially overlap, but different regions of σ70 and σ54 are used for binding to the RNAP, our results further underscore the importance of the σ–RNAP interface in bacterial RNAP function and regulation and potentially for intervention by antibacterials

Photocatalytic antimicrobial activity of thin surface films of TiO2, CuO and TiO2 /CuO dual layers on Escherichia coli and bacteriophage T4

TiO2 coated surfaces are increasingly studied for their ability to inactivate microorganisms. The activity of glass coated with thin films of TiO2, CuO and hybrid CuO/TiO2 prepared by atmospheric Chemical Vapour Deposition (Ap-CVD) and TiO2 prepared by a sol-gel process was investigated using the inactivation of bacteriophage T4 as a model for inactivation of viruses. The chemical oxidising activity was also determined by measuring stearic acid oxidation. The results showed that the rate of inactivation of bacteriophage T4 increased with increasing chemical oxidising activity with the maximum rate obtained on highly active sol-gel preparations. However these were delicate and easily damaged unlike the Ap-CVD coatings. Inactivation rates were highest on CuO and CuO/TiO2 which had the lowest chemical oxidising activities. The inactivation of T4 was higher than that of Escherichia coli on low activity surfaces. The combination of photocatalysis and toxicity of copper acted synergistically to inactivate bacteriophage T4 and retained some selfcleaning activity. The presence of phosphate ions slowed inactivation but NaCl had no effect. The results show that TiO2/CuO coated surfaces are highly antiviral and may have applications in the food and healthcare industries

Draft Genome Sequence of an Enterococcus faecalis ATCC 19433 Siphovirus Isolated from Raw Domestic Sewage.

We previously isolated and characterized an Enterococcus faecalis ATCC 19433 siphovirus from raw domestic sewage as a viral indicator of human fecal pollution. Here, we report the draft genome sequence of this bacteriophage

Bacteriophages and their structural organisation

Viruses are extremely small infectious particles that are not visible in a light microscope, and are able to pass through fine porcelain filters. They exist in a huge variety of forms and infect practically all living systems: animals, plants, insects and bacteria. All viruses have a genome, typically only one type of nucleic acid, but it could be one or several molecules of DNA or RNA, which is surrounded by a protective stable coat (capsid) and sometimes by additional layers which may be very complex and contain carbohydrates, lipids, and additional proteins. The viruses that have only a protein coat are named “naked”, or non- enveloped viruses. Many viruses have an envelope (enveloped viruses) that wraps around the protein capsid. This envelope is formed from a lipid membrane of the host cell during the release of a virus out of the cell

Genome signatures, self-organizing maps and higher order phylogenies: a parametric analysis

Genome signatures are data vectors derived from the compositional statistics of DNA. The self-organizing map (SOM) is a neural network method for the conceptualisation of relationships within complex data, such as genome signatures. The various parameters of the SOM training phase are investigated for their effect on the accuracy of the resulting output map. It is concluded that larger SOMs, as well as taking longer to train, are less sensitive in phylogenetic classification of unknown DNA sequences. However, where a classification can be made, a larger SOM is more accurate. Increasing the number of iterations in the training phase of the SOM only slightly increases accuracy, without improving sensitivity. The optimal length of the DNA sequence k-mer from which the genome signature should be derived is 4 or 5, but shorter values are almost as effective. In general, these results indicate that small, rapidly trained SOMs are generally as good as larger, longer trained ones for the analysis of genome signatures. These results may also be more generally applicable to the use of SOMs for other complex data sets, such as microarray data

Comparative genomics of Shiga toxin encoding bacteriophages

Background Stx bacteriophages are responsible for driving the dissemination of Stx toxin genes (stx) across their bacterial host range. Lysogens carrying Stx phages can cause severe, lifethreatening disease and Stx toxin is an integral virulence factor. The Stx-bacteriophage vB_EcoP-24B, commonly referred to as 24B, is capable of multiply infecting a single bacterial host cell at a high frequency, with secondary infection increasing the rate at which subsequent bacteriophage infections can occur. This is biologically unusual, therefore determining the genomic content and context of 24B compared to other lambdoid Stx phages is important to understanding the factors controlling this phenomenon and determining whether they occur in other Stx phages. Results The genome of the Stx2 encoding phage, 24B was sequenced and annotated. The genomic organisation and general features are similar to other sequenced Stx bacteriophages induced from Enterohaemorrhagic Escherichia coli (EHEC), however 24B possesses significant regions of heterogeneity, with implications for phage biology and behaviour. The 24B genome was compared to other sequenced Stx phages and the archetypal lambdoid phage, lambda, using the Circos genome comparison tool and a PCR-based multi-loci comparison system. Conclusions The data support the hypothesis that Stx phages are mosaic, and recombination events between the host, phages and their remnants within the same infected bacterial cell will continue to drive the evolution of Stx phage variants and the subsequent dissemination of shigatoxigenic potentia