Molecular genotyping of bacillus anthracis strains from Georgia and northeastern part of Turkey

Bacillus anthracis is the causal agent of anthrax and has a history of use as a biological weapon. Anthrax cases occur worldwide and the disease is endemic in certain regions. Here we describe a study of the genetic diversity of B. anthracis strains in two endemic areas: The country of Georgia and the Kars region of Turkey. Thirty Turkish isolates and thirty Georgian isolates were subjected to Single Nucleotide Polymorphism (SNP) sub typing, followed by higher-resolution genotyping using 25-loci variable-number tandem repeat analysis (MLVA-25). Canonical SNP typing indicated that Turkish strains belonged to both the A.Br.003 linage and the Australian 94 lineage. In light of a recent analysis that placed the majority of Georgian B. anthracis isolates in one phylogenetic group, we screened the Turkish strains using a previously developed Georgian SNP panel. Minimal diversity was observed among the Kars strains within the Georgian SNP lineage: all 30 of these strains grouped with A.Br.026, ten strains were derived from A.Br.028, and only two isolates belonged to A.Br.029. According to the results of MLVA-25 genotyping, all 30 Turkish strains belong to two clusters. Cluster A is more diverse than cluster B. Our results suggest that B. anthracis strains originating from Georgia and the northeastern part of Turkey are genetically interrelated, which could be explained by the geographic proximity of the countries

Molecular genotyping of bacillus anthracis strains from Georgia and northeastern part of Turkey

Bacillus anthracis is the causal agent of anthrax and has a history of use as a biological weapon. Anthrax cases occur worldwide and the disease is endemic in certain regions. Here we describe a study of the genetic diversity of B. anthracis strains in two endemic areas: The country of Georgia and the Kars region of Turkey. Thirty Turkish isolates and thirty Georgian isolates were subjected to Single Nucleotide Polymorphism (SNP) sub typing, followed by higher-resolution genotyping using 25-loci variable-number tandem repeat analysis (MLVA-25). Canonical SNP typing indicated that Turkish strains belonged to both the A.Br.003 linage and the Australian 94 lineage. In light of a recent analysis that placed the majority of Georgian B. anthracis isolates in one phylogenetic group, we screened the Turkish strains using a previously developed Georgian SNP panel. Minimal diversity was observed among the Kars strains within the Georgian SNP lineage: all 30 of these strains grouped with A.Br.026, ten strains were derived from A.Br.028, and only two isolates belonged to A.Br.029. According to the results of MLVA-25 genotyping, all 30 Turkish strains belong to two clusters. Cluster A is more diverse than cluster B. Our results suggest that B. anthracis strains originating from Georgia and the northeastern part of Turkey are genetically interrelated, which could be explained by the geographic proximity of the countries

Respiratory distress and perinatal lethality in Nedd4-2-deficient mice

The epithelial sodium channel (ENaC) is essential for sodium homoeostasis in many epithelia. ENaC activity is required for lung fluid clearance in newborn animals and for maintenance of blood volume and blood pressure in adults. In vitro studies show that the ubiquitin ligase Nedd4-2 ubiquitinates ENaC to regulate its cell surface expression. Here we show that knockout of Nedd4-2 in mice leads to increased ENaC expression and activity in embryonic lung. This increased ENaC activity is the likely reason for premature fetal lung fluid clearance in Nedd4-2−/− animals, resulting in a failure to inflate lungs and perinatal lethality. A small percentage of Nedd4-2−/− animals survive up to 22 days, and these animals also show increased ENaC expression and develop lethal sterile inflammation of the lung. Thus, we provide critical in vivo evidence that Nedd4-2 is essential for correct regulation of ENaC expression, fetal and postnatal lung function and animal survival

Effective DNA/RNA Co-Extraction for Analysis of MicroRNAs, mRNAs, and Genomic DNA from Formalin-Fixed Paraffin-Embedded Specimens

Background: Retrospective studies of archived human specimens, with known clinical follow-up, are used to identify predictive and prognostic molecular markers of disease. Due to biochemical differences, however, formalin-fixed paraffinembedded (FFPE) DNA and RNA have generally been extracted separately from either different tissue sections or from the same section by dividing the digested tissue. The former limits accurate correlation whilst the latter is impractical when utilizing rare or limited archived specimens. Principal Findings: For effective recovery of genomic DNA and total RNA from a single FFPE specimen, without splitting the proteinase-K digested tissue solution, we optimized a co-extraction method by using TRIzol and purifying DNA from the lower aqueous and RNA from the upper organic phases. Using a series of seven different archived specimens, we evaluated the total amounts of genomic DNA and total RNA recovered by our TRIzol-based co-extraction method and compared our results with those from two commercial kits, the Qiagen AllPrep DNA/RNA FFPE kit, for co-extraction, and the Ambion RecoverAll TM Total Nucleic Acid Isolation kit, for separate extraction of FFPE-DNA and-RNA. Then, to accurately assess the quality of DNA and RNA co-extracted from a single FFPE specimen, we used qRT-PCR, gene expression profiling and methylation assays to analyze microRNAs, mRNAs, and genomic DNA recovered from matched fresh and FFPE MCF10A cells. These experiments show that the TRIzol-based co-extraction method provides larger amounts of FFPE-DNA and –RNA tha

Predictions for the future of kallikrein-related peptidases in molecular diagnostics

Kallikrein-related peptidases (KLKs) form a cancer-related ensemble of serine proteases. This multigene family hosts the most widely used cancer biomarker that is PSA-KLK3, with millions of tests performed annually worldwide. The present report provides an overview of the biomarker potential of the extended KLK family (KLK1-KLK15) in various disease settings and envisages approaches that could lead to additional KLK-driven applications in future molecular diagnostics. Particular focus is given on the inclusion of KLKs into multifaceted cancer biomarker panels that provide enhanced diagnostic, prognostic and/or predictive accuracy in several human malignancies. Such panels have been described so far for prostate, ovarian, lung and colorectal cancers. The role of KLKs as biomarkers in non-malignant disease settings, such as Alzheimer’s disease and multiple sclerosis, is also commented upon. Predictions are given on the challenges and future directions regarding clinically oriented KLK research

Complete plastid genomes of South Caucasian, European and Mediterranean Basin wild grapevines (<em>Vitis vinifera</em> subsp. <em>sylvestris</em>)

International audienceThe South Caucasus region is widely believed to be the area in which grape domestication began, therefore the study of genetic diversity of wild grape samples in this region is viewed as a key to understanding grape domestication in general. The main goal of the presented research was assessment of genetic diversity of wild grape samples from different places of Europe and Mediterranean basin by using complete chloroplast DNA Illumina sequencing. The analyzed sequences were compared with the plastid genomes of Georgian wild grape samples from our previous research. The presented work is a first attempt of studying of wide range of the genus Vitis L., in particular wild grape samples from Europe and Mediterranean basin with the next-generation technologies and adopting this application for the tracing of grape ancestry. The obtained results will help to understand the genetic relationships between wild and cultivated grapes from different geographical locations and explain the molecular bases of grape origin and evolution. Phylogenetic trees representing evolutionary relationship between analyzed grape samples are presented

Correction to: Genomic characterization of three novel Basilisk-like phages infecting Bacillus anthracis

Following the publication of this article [1], the authors noted two typographical errors: one in Table 1 with regard to the location of the Basilisk Phage, which was incorrectly captured as “Kutaisis, country of Georgia Utah, USA” but should be “Utah, USA”

Genomic characterization of three novel Basilisk-like phages infecting Bacillus anthracis

Abstract Background In the present study, we sequenced the complete genomes of three novel bacteriophages v_B-Bak1, v_B-Bak6, v_B-Bak10 previously isolated from historical anthrax burial sites in the South Caucasus country of Georgia. We report here major trends in the molecular evolution of these phages, which we designate as “Basilisk-Like-Phages” (BLPs), and illustrate patterns in their evolution, genomic plasticity and core genome architecture. Results Comparative whole genome sequence analysis revealed a close evolutionary relationship between our phages and two unclassified Bacillus cereus group phages, phage Basilisk, a broad host range phage (Grose JH et al., J Vir. 2014;88(20):11846-11860) and phage PBC4, a highly host-restricted phage and close relative of Basilisk (Na H. et al. FEMS Microbiol. letters. 2016;363(12)). Genome comparisons of phages v_B-Bak1, v_B-Bak6, and v_B-Bak10 revealed significant similarity in sequence, gene content, and synteny with both Basilisk and PBC4. Transmission electron microscopy (TEM) confirmed the three phages belong to the Siphoviridae family. In contrast to the broad host range of phage Basilisk and the single-strain specificity of PBC4, our three phages displayed host specificity for Bacillus anthracis. Bacillus species including Bacillus cereus, Bacillus subtilis, Bacillus anthracoides, and Bacillus megaterium were refractory to infection. Conclusions Data reported here provide further insight into the shared genomic architecture, host range specificity, and molecular evolution of these rare B. cereus group phages. To date, the three phages represent the only known close relatives of the Basilisk and PBC4 phages and their shared genetic attributes and unique host specificity for B. anthracis provides additional insight into candidate host range determinants