Search results for "PLASMID"

Extraintestinal pathogenic Escherichia coli sequence type 131 H30-R and H30-Rx subclones in retail chicken meat, Italy

2016

Extraintestinal pathogenic Escherichia coli sequence type 131 (ST131), typically fluoroquinolone-resistant (FQ-R) and/or extended-spectrum β-lactamase (ESBL)-producing, has emerged globally. Among clinical isolates, ST131, primarily its H30-R and H30-Rx subclones, accounts for most antimicrobial-resistant E. coli and is the dominant E. coli strain worldwide. We assessed its prevalence and characteristics among raw chicken meat samples on sale in Palermo, Italy. A collection of 237 fluoroquinolone resistant and ESBL/AmpC producing E. coli isolates, which had been isolated from processed retail chicken meat in the period May 2013-April 2015, was analyzed. Established polymerase chain reaction…

Conjugative ESBL plasmids differ in their potential to rescue susceptible bacteria via horizontal gene transfer in lethal antibiotic concentrations.

2017

Conjugative ESBL plasmids differ in their potential to rescue susceptible bacteria via horizontal gene transfer in lethal antibiotic concentrations

New ΦBT1 site-specific integrative vectors with neutral phenotype in Streptomyces.

2016

Integrative plasmids are one of the best options to introduce genes in low copy and in a stable form into bacteria. The ΦC31-derived plasmids constitute the most common integrative vectors used in Streptomyces. They integrate at different positions (attB and pseudo-attB sites) generating different mutations. The less common ΦBT1-derived vectors integrate at the unique attB site localized in the SCO4848 gene (S. coelicolor genome) or their orthologues in other streptomycetes. This work demonstrates that disruption of SCO4848 generates a delay in spore germination. SCO4848 is co-transcribed with SCO4849, and the spore germination phenotype is complemented by SCO4849. Plasmids pNG1-4 were crea…

2017

Despite rapid progress, many problems and limitations persist and limit the applicability of gene-editing techniques. Making use of meganucleases, TALENs, or CRISPR/Cas9-based tools requires an initial step of pre-screening to determine the efficiency and specificity of the designed tools. This step remains time consuming and material consuming. Here we propose a simple, cheap, reliable, time-saving, and highly sensitive method to evaluate a given gene-editing tool based on its capacity to induce chromosomal translocations when combined with a reference engineered nuclease. In the proposed technique, designated engineered nuclease-induced translocations (ENIT), a plasmid coding for the DNA-…

Efficacy of interleukin 10 gene hydrofection in pig liver vascular isolated ‘in vivo’ by surgical procedure with interest in liver transplantation

2019

AIM Liver transplantation is the only curative strategy for final stage liver diseases. Despite the great advances achieved during the last 20 years, the recipient immune response after transplantation is not entirely controlled. This results in high rates of acute cell rejection and, approximately, 10% of early mortality. Therapeutic treatment could be improved by efficiently transfecting genes that encode natural immunosuppressant proteins, employing safe procedures that could be transferred to clinical setting. In this sense, interleukin 10 plays a central role in immune tolerance response by acting at different levels. METHODS hIL10 gene was hydrofected by retrograde hydrodynamic inject…

Establishment of two quantitative nested qPCR assays targeting the human EPO transgene.

2016

International audience; For ethical and safety reasons it is critical to develop easily implemented assays with high sensitivity and specificity for gene doping surveillance. Two nested quantitative real-time PCR (qPCR) assays were developed that target the human EPO (hEPO) cDNA sequence in a circular form, representative of recombinant adeno-associated viral (rAAV) vector genomes found in vivo. Through an interlaboratory evaluation, the assays were validated and utilized in an in vitro blinded study. These assays are specific and extremely sensitive with a limit of detection (LOD) of 1 copy of circular plasmid DNA and a limit of quantification (LOQ) of 10 to 20 copies in the presence of 50…

Corrigendum: Phylogeny of Vibrio vulnificus From the Analysis of the Core-Genome: Implications for Intra-Species Taxonomy

2018

Vibrio vulnificus (Vv) is a multi-host pathogenic species currently subdivided into three biotypes (Bts). The three Bts are human-pathogens, but only Bt2 is also a fish-pathogen, an ability that is conferred by a transferable virulence-plasmid (pVvbt2). Here we present a phylogenomic analysis from the core genome of 80 Vv strains belonging to the three Bts recovered from a wide range of geographical and ecological sources. We have identified five well-supported phylogenetic groups or lineages (L). LI comprises a mixture of clinical and environmental Bt1 strains, most of them involved in human clinical cases related to raw seafood ingestion. LII is linked to the aquaculture industry and incl…

Convergent Evolution in Intracellular Elements: Plasmids as Model Endosymbionts

2018

Endosymbionts are organisms that live inside the cells of other species. This lifestyle is ubiquitous across the tree of life and is featured by unicellular eukaryotes, prokaryotes, and by extrachromosomal genetic elements such as plasmids. Given that all of these elements dwell in the cytoplasm of their host cell, they should be subject to similar selection pressures. Here we show that strikingly similar features have evolved in both bacterial endosymbionts and plasmids. Since host and endosymbiont are often metabolically tightly intertwined, they are difficult to disentangle experimentally. We propose that using plasmids as tractable model systems can help to solve this problem, thus allo…

2019

The possibility to modify gut bacterial flora has become an important goal, and various approaches are used to achieve desirable communities. However, the genetic engineering of existing microbes in the gut, which are already compatible with the rest of the community and host immune system, has not received much attention. Here, we discuss and experimentally evaluate the possibility to use modified and mobilizable CRISPR-Cas9-endocing plasmid as a tool to induce changes in bacterial communities. This plasmid system (briefly midbiotic) is delivered from bacterial vector into target bacteria via conjugation. Compared to, for example, bacteriophage-based applications, the benefits of conjugati…

Molecular epidemiology and drug-resistance mechanisms in carbapenem-resistant Klebsiella pneumoniae isolated in patients from a tertiary hospital in …

2020

Abstract Objectives The aim of this study has been to characterize carbapenem-resistant Klebsiella pneumoniae isolates and to determine the resistance mechanisms involved, the clonal relationship between strains and clinical and demographical data of the infected patients. Methods Clinical and demographical data from patients were collected and statistically analysed. Antimicrobial susceptibility testing was performed and resistance genes were detected both phenotypically and genotypically. Conjugation assays were performed to show horizontal transferability of resistance genes. Clonal relationship was also studied. Next-generation sequencing (NGS) was performed to obtain information regard…