Differences in genotype and virulence among four multidrug-resistant <i>Streptococcus pneumoniae</i> isolates belonging to the PMEN1 clone

We report on the comparative genomics and characterization of the virulence phenotypes of four &lt;i&gt;S. pneumoniae&lt;/i&gt; strains that belong to the multidrug resistant clone PMEN1 (Spain&lt;sup&gt;23F&lt;/sup&gt; ST81). Strains SV35-T23 and SV36-T3 were recovered in 1996 from the nasopharynx of patients at an AIDS hospice in New York. Strain SV36-T3 expressed capsule type 3 which is unusual for this clone and represents the product of an in vivo capsular switch event. A third PMEN1 isolate - PN4595-T23 - was recovered in 1996 from the nasopharynx of a child attending day care in Portugal, and a fourth strain - ATCC700669 - was originally isolated from a patient with pneumococcal disease in Spain in 1984. We compared the genomes among four PMEN1 strains and 47 previously sequenced pneumococcal isolates for gene possession differences and allelic variations within core genes. In contrast to the 47 strains - representing a variety of clonal types - the four PMEN1 strains grouped closely together, demonstrating high genomic conservation within this lineage relative to the rest of the species. In the four PMEN1 strains allelic and gene possession differences were clustered into 18 genomic regions including the capsule, the blp bacteriocins, erythromycin resistance, the MM1-2008 prophage and multiple cell wall anchored proteins. In spite of their genomic similarity, the high resolution chinchilla model was able to detect variations in virulence properties of the PMEN1 strains highlighting how small genic or allelic variation can lead to significant changes in pathogenicity and making this set of strains ideal for the identification of novel virulence determinant

Development Of An Engineered Bioluminescent Reporter Phage For Detection Of Bacterial Blight Of Crucifers

Bacterial blight, caused by the phytopathogen Pseudomonas cannabina pv. alisalensis, is an emerging disease afflicting important members of the Brassicaceae family. The disease is often misdiagnosed as pepper spot, a much less severe disease caused by the related pathogen Pseudomonas syringae pv. maculicola. We have developed a phage-based diagnostic that can both identify and detect the causative agent of bacterial blight and differentiate the two pathogens. A recombinant >light>-tagged reporter phage was generated by integrating bacterial luxAB genes encoding luciferase into the genome of P. cannabina pv. alisalensis phage PBSPCA1. The PBSPCA1::luxAB reporter phage is viable and stable and retains properties similar to those of the wildtype phage. PBSPCA1::luxAB rapidly and sensitively detects P. cannabina pv. alisalensis by conferring a bioluminescent signal response to cultured cells. Detection is dependent on cell viability. Other bacterial pathogens of Brassica species such as P. syringae pv. maculicola, Pseudomonas marginalis, Pectobacterium carotovorum, Xanthomonas campestris pv. campestris, and X. campestris pv. raphani either do not produce a response or produce significantly attenuated signals with the reporter phage. Importantly, the reporter phage detects P. cannabina pv. alisalensis on diseased plant specimens, indicating its potential for disease diagnosis.National Science Foundation Small Business Innovative Research 1012059U.S. Department of EducationU.S. Department of AgricultureCellular and Molecular Biolog

Genome-wide high-resolution mapping of UV-induced mitotic recombination events in Saccharomyces cerevisiae.

In the yeast Saccharomyces cerevisiae and most other eukaryotes, mitotic recombination is important for the repair of double-stranded DNA breaks (DSBs). Mitotic recombination between homologous chromosomes can result in loss of heterozygosity (LOH). In this study, LOH events induced by ultraviolet (UV) light are mapped throughout the genome to a resolution of about 1 kb using single-nucleotide polymorphism (SNP) microarrays. UV doses that have little effect on the viability of diploid cells stimulate crossovers more than 1000-fold in wild-type cells. In addition, UV stimulates recombination in G1-synchronized cells about 10-fold more efficiently than in G2-synchronized cells. Importantly, at high doses of UV, most conversion events reflect the repair of two sister chromatids that are broken at approximately the same position whereas at low doses, most conversion events reflect the repair of a single broken chromatid. Genome-wide mapping of about 380 unselected crossovers, break-induced replication (BIR) events, and gene conversions shows that UV-induced recombination events occur throughout the genome without pronounced hotspots, although the ribosomal RNA gene cluster has a significantly lower frequency of crossovers

Tuberculosis vaccine strain _Mycobacterium bovis_ BCG Russia is a natural _recA_ mutant

The current tuberculosis vaccine is a live vaccine derived from _Mycobacterium bovis_ and attenuated by serial _in vitro_ passaging. All vaccine substrains in use stem from one source, strain Bacille Calmette-Gu&#xe9;rin. However, they differ in regions of genomic deletions, antigen expression levels, immunogenicity, and protective efficacy. As a RecA phenotype increases genetic stability and may contribute restricting the ongoing evolution of the various BCG substrains, we aimed to inactivate _recA_ by allelic replacement in BCG vaccine strains representing different phylogenetic lineages (Pasteur, Frappier, Denmark, Russia). Homologous gene replacement was successful in three out of four strains. However, only illegitimate recombination was observed in BCG substrain Russia. Sequence analyses of _recA_ revealed that a single nucleotide insertion in the 5&#x27; part of _recA_ led to a translational frameshift with an early stop codon making BCG Russia a natural _recA_ mutant. At the protein level BCG Russia failed to express RecA. According to phylogenetic analyses BCG Russia is an ancient vaccine strain most closely related to the parental _M. bovis_. Our data suggest that _recA_ inactivation in BCG Russia occurred early and is in part responsible for its high degree of genomic stability, resulting in a substrain that has less genetic alterations than other vaccine substrains with respect to _M. bovis_ AF2122/97 wild type

Statistical analysis on detecting recombination sites in DNA-beta satellites associated with the old world geminiviruses

Although an exchange of genetic information by recombination plays an important role in the evolution of viruses, it is not clear how it generates diversity. {\it Geminiviruses} are plant viruses which have ambisense single-stranded circular DNA genomes and one of the most economically important plant viruses in agricultural production. Small circular single-stranded DNA satellites, termed DNA-$\beta$, have recently been found associated with some geminivirus infections. In this paper we analyze a satellite molecule DNA-$\beta$ of geminiviruses for recombination events using phylogenetic and statistical analysis and we find that one strain from ToLCMaB has a recombination pattern and is possibly recombinant molecule between two strains from two species, PaLCuB-[IN:Chi:05] (major parent) and ToLCB-[IN:CP:04] (minor parent).Comment: 8 figures and 2 tables. To appear in Frontiers in Systems Biolog

Genome characterization and population genetic structure of the zoonotic pathogen, streptococcus canis

Background - Streptococcus canis is an important opportunistic pathogen of dogs and cats that can also infect a wide range of additional mammals including cows where it can cause mastitis. It is also an emerging human pathogen. Results - Here we provide characterization of the first genome sequence for this species, strain FSL S3-227 (milk isolate from a cow with an intra-mammary infection). A diverse array of putative virulence factors was encoded by the S. canis FSL S3-227 genome. Approximately 75% of these gene sequences were homologous to known Streptococcal virulence factors involved in invasion, evasion, and colonization. Present in the genome are multiple potentially mobile genetic elements (MGEs) [plasmid, phage, integrative conjugative element (ICE)] and comparison to other species provided convincing evidence for lateral gene transfer (LGT) between S. canis and two additional bovine mastitis causing pathogens (Streptococcus agalactiae, and Streptococcus dysgalactiae subsp. dysgalactiae), with this transfer possibly contributing to host adaptation. Population structure among isolates obtained from Europe and USA [bovine = 56, canine = 26, and feline = 1] was explored. Ribotyping of all isolates and multi locus sequence typing (MLST) of a subset of the isolates (n = 45) detected significant differentiation between bovine and canine isolates (Fisher exact test: P = 0.0000 [ribotypes], P = 0.0030 [sequence types]), suggesting possible host adaptation of some genotypes. Concurrently, the ancestral clonal complex (54% of isolates) occurred in many tissue types, all hosts, and all geographic locations suggesting the possibility of a wide and diverse niche. Conclusion - This study provides evidence highlighting the importance of LGT in the evolution of the bacteria S. canis, specifically, its possible role in host adaptation and acquisition of virulence factors. Furthermore, recent LGT detected between S. canis and human bacteria (Streptococcus urinalis) is cause for concern, as it highlights the possibility for continued acquisition of human virulence factors for this emerging zoonotic pathogen

The Landscape of Realized Homologous Recombination in Pathogenic Bacteria

Recombination enhances the adaptive potential of organisms by allowing genetic variants to be tested on multiple genomic backgrounds. Its distribution in the genome can provide insight into the evolutionary forces that underlie traits, such as the emergence of pathogenicity. Here, we examined landscapes of realized homologous recombination of 500 genomes from ten bacterial species and found all species have “hot” regions with elevated rates relative to the genome average. We examined the size, gene content, and chromosomal features associated with these regions and the correlations between closely related species. The recombination landscape is variable and evolves rapidly. For example in Salmonella, only short regions of around 1 kb in length are hot whereas in the closely related species Escherichia coli, some hot regions exceed 100 kb, spanning many genes. Only Streptococcus pyogenes shows evidence for the positive correlation between GC content and recombination that has been reported for several eukaryotes. Genes with function related to the cell surface/membrane are often found in recombination hot regions but E. coli is the only species where genes annotated as “virulence associated” are consistently hotter. There is also evidence that some genes with “housekeeping” functions tend to be overrepresented in cold regions. For example, ribosomal proteins showed low recombination in all of the species. Among specific genes, transferrin-binding proteins are recombination hot in all three of the species in which they were found, and are subject to interspecies recombination