Frequency of conjugative transfer of plasmid-encoded ISEcp1 - blaCTX-M-15 and aac(6')-lb-cr genes in Enterobacteriaceae at a tertiary care center in Lebanon - role of transferases

<p>Abstract</p> <p>Background</p> <p>The frequency of transfer of genes encoding resistance to antimicrobial agents was determined by conjugation in ESBL-producing and/or fluoroquinolone or aminoglycoside resistant Enterobacteriaceae clinical isolates at a tertiary care center in Lebanon. In addition, the role of <it>tra </it>genes encoding transferases in mediating conjugation was assessed.</p> <p>Methods</p> <p>Conjugation experiments were done on 53 ESBL-producing and/or fluoroquinolone resistant <it>E. coli </it>and <it>K. pneumoniae </it>and ESBL-producing <it>S. sonnei </it>isolates. Antimicrobial susceptibility testing on parent and transconjugant isolates, and PCR amplifications on plasmid extracts of the resistance-encoding genes: <it>bla</it>_CTX-M-15with the <it>ISEcp1 </it>insertion sequence, the <it>aac(6')-lb-cr </it>and <it>qnr</it>S genes, as well as <it>tra </it>encoding transferases genes were done. Random amplified polymorphic DNA (RAPD) analysis was performed to demonstrate whether conjugative isolates are clonal and whether they are linked epidemiologically to a particular source.</p> <p>Results</p> <p>Antimicrobial susceptibility testing on transconjugants revealed that 26 out of 53 (49%) ESBL-producing <it>Enterobacteriaceae </it>were able to transfer antimicrobial resistance to the recipients. Transfer of high-level resistance to the transconjugants encoded by the <it>bla</it>_CTX-M-15gene downstream the <it>ISEcp1 </it>insertion sequence against 3rd generation cephalosporins, and of low-level resistance against ciprofloxacin, and variable levels of resistance against aminoglycosides encoded by <it>aac(6')-lb-cr </it>gene, were observed in transconjugants. <it>tra </it>encoding transferase genes were detected exclusively in conjugative isolates.</p> <p>Conclusion</p> <p>In conclusion, the frequency of transfer of antimicrobial resistance in non clonal <it>Enterobacteriaceae </it>at the tertiary care center by conjugation was 49%. Conjugation occurred in isolates expressing the <it>tra </it>encoding transferase genes. Multiple conjugative strains harboring the plasmid encoded antimicrobial resistant genes were circulating in the medical center. Molecular epidemiology analysis showed that conjugative isolates are neither clonal nor linked to a particular site and transfer of antimicrobial resistance is by horizontal transfer of plasmids.</p

Waste water effluent contributes to the dissemination of CTX-M-15 in the natural environment.

OBJECTIVES Multidrug-resistant Enterobacteriaceae pose a significant threat to public health. We aimed to study the impact of sewage treatment effluent on antibiotic resistance reservoirs in a river. METHODS River sediment samples were taken from downstream and upstream of a waste water treatment plant (WWTP) in 2009 and 2011. Third-generation cephalosporin (3GC)-resistant Enterobacteriaceae were enumerated. PCR-based techniques were used to elucidate mechanisms of resistance, with a new two-step PCR-based assay developed to investigate bla(CTX-M-15) mobilization. Conjugation experiments and incompatibility replicon typing were used to investigate plasmid ecology. RESULTS We report the first examples of bla(CTX-M-15) in UK river sediment; the prevalence of bla(CTX-M-15) was dramatically increased downstream of the WWTP. Ten novel genetic contexts for this gene were identified, carried in pathogens such as Escherichia coli ST131 as well as indigenous aquatic bacteria such as Aeromonas media. The bla(CTX-M-15) -gene was readily transferable to other Gram-negative bacteria. We also report the first finding of an imipenem-resistant E. coli in a UK river. CONCLUSIONS The high diversity and host range of novel genetic contexts proves that evolution of novel combinations of resistance genes is occurring at high frequency and has to date been significantly underestimated. We have identified a worrying reservoir of highly resistant enteric bacteria in the environment that poses a threat to human and animal health

In vivo reprogramming of wound-resident cells generates skin epithelial tissue

Large cutaneous ulcers are, in severe cases, life threatening(1,2). As the global population ages, non-healing ulcers are becoming increasingly common(1,2). Treatment currently requires the transplantation of pre-existing epithelial components, such as skin grafts, or therapy using cultured cells(2). Here we develop alternative supplies of epidermal coverage for the treatment of these kinds of wounds. We generated expandable epithelial tissues using in vivo reprogramming of wound-resident mesenchymal cells. Transduction of four transcription factors that specify the skin-cell lineage enabled efficient and rapid de novo epithelialization from the surface of cutaneous ulcers in mice. Our findings may provide a new therapeutic avenue for treating skin wounds and could be extended to other disease situations in which tissue homeostasis and repair are impaired