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Influenza A(H1N1)pdm09 pandemic virus causing the 2009 global outbreak moved into the post-pandemic period, but its variants continued to be the prevailing subtype in the 2015-2016 influenza season in Europe and Asia. To determine the molecular characteristics of influenza A(H1N1)pdm09 isolates circulating during the 2015-2016 season in Turkey, we identified mutations in the hemagglutinin (HA) genes and investigated the presence of H275Y alteration in the neuraminidase genes in the randomly selected isolates. The comparison of the HA nucleotide sequences revealed a very high homology (>99.5%) among the studied influenza A(H1N1)pdm09 isolates, while a relatively low homology (96.6%-97.2%), was observed between Turkish isolates and the A/California/07/2009 vaccine virus. Overall 14 common mutations were detected in HA sequences of all 2015-2016 influenza A(H1N1)pdm09 isolates with respect to the A/California/07/2009 virus, four of which located in three different antigenic sites. Eleven rare mutations in 12 HA sequences were also detected. Phylogenetic analysis revealed that all characterized influenza A(H1N1)pdm09 isolates formed a single genetic cluster, belonging to the genetic subclade 6B.1, defined by HA amino acid substitutions S84N, S162N, and I216T. Furthermore, all isolates showed an oseltamivir-sensitive genotype, suggesting that Tamiflu (Oseltamivir) could still be the drug of choice in Turkey.

This study aimed to determine the presence of vancomycin resistance (vanA and vanB) and virulence genes (esp, asa1, gelE, ace, hyl, cylA, cpd and ebpA) in vancomycin-resistant Enterococcus faecium (VREfm) strains and to analyse the clonal relationships among the strains. E. faecium strains were identified from rectal and clinical specimens by biochemical tests and the API-20 Strep kit. Susceptibility testing was performed using disc-diffusion and broth-dilution methods. PFGE was used for molecular typing of the VREfm strains. The vancomycin resistance and virulence genes were amplified by two-step multiplex PCR. All 55 VREfm isolates were resistant to penicillin G, ampicillin and high-level gentamicin but were susceptible to quinupristin/dalfopristin and linezolid. Multiplex PCR analysis indicated that all isolates harboured vanA and that 41 (75 %) were positive for virulence genes. The esp gene was the most common virulence factor and was detected in nine (41 %) invasive and 32 (96.7 %) non-invasive isolates. Multiple virulence genes were observed only in two non-invasive isolates; one harboured esp and ebpA and the other harboured esp, ebpA, asa1, gelE and cpd. PFGE typing yielded 16 different types, seven of which were clusters with two to 14 strains each. The clustering rates of the rectal swab, blood and urine isolates were 72.7 %, 61.5 % and 87.5 %, respectively. The genetic similarity observed among the VREfm isolates indicated cross-transmission in the hospital. Further studies on the virulence factors present in the strains might provide insight into the acquisition of these traits and their contribution to increased prevalence of VREfm.

Arrest of DNA replication in Escherichia coli is mediated by specific interactions between the Tus protein and terminator (Ter) sequences. Binding of Tus to a Ter site forms a asymmetric protein-DNA complex that arrests DNA replication in an orientation-dependent fashion. In this study, mutant Ter sites carrying single base pair substitutions at 16 different positions were examined for their ability to bind purified Tus protein and arrest DNA replication. In vitro competition assays demonstrated that base pair substitutions at positions 8-19 had significant effects on the free energy of Tus binding (DeltaDeltaG0 of 1.5 to4. 0 kcal/mol). Concomitant with loss of binding affinity, mutations at these positions also showed significantly lower or undetectable replication arrest activities in vivo. Substitutions at positions 6, 20, and 21 had moderate effects on Tus-Ter interactions, suggesting that these base pairs contribute to, but are not absolutely critical for, Tus binding. Even though the effects on binding were minimal, these Ter mutants were not as efficient as wild type Tus-TerB complexes at arresting replication forks. Three new potential Ter sites, referred to as TerH, TerI, and TerJ, were identified by searching the E. coli genome for sequence similarity to a consensus Ter site sequence.

ari, filiz/0000-0002-8107-732X WOS: 000312108500016 PubMed: 23226519 The life-threatening illnesses caused by Streptococcus pneumoniae have been declined significantly after the use of pneumococcal conjugate vaccines. Continuous monitoring of the vaccine serogroups/types is necessary to follow the changing epidemiology of invasive pneumococcal diseases. Recently, the sequential multiplex PCR approach, which uses several different sets of reactions, has been commonly adopted for determining capsular serogroups/types of S. pneumoniae isolates. In our study, we focused on development of a one-step multiplex PCR assay detecting all 1, 3, 4, 5, 6A/B, 7F, 9V, 14, 18C, 19A, 19F and 23F serogroups/types targeted by PCV13. The content of multiplex PCR mix and the cycling conditions were optimized in a manner that allowed rapid and accurate serotyping of a pneumococcal isolate by performing only a single amplification reaction. In our study of 182 clinical isolates, the one-step multiplex PCR assay exhibited 100% sensitivity and specificity, suggesting that its utilization can significantly reduce the use of traditional antiserum method requiring expensive reagents. Citation: Coskun-Ari FF, Guldemir D, Durmaz R (2012) One-Step Multiplex PCR Assay for Detecting Streptococcus pneumoniae Serogroups/Types Covered by 13-Valent Pneumococcal Conjugate Vaccine (PCV13). PLoS ONE 7(12): e50406. doi:10.1371/journal.pone.0050406