RESUMEN
The prevalence of plasmid-mediated quinolone resistance (PMQR) determinants (qnrA, qnrB, qnrS, aac(6')-Ib-cr, and qepA) was investigated in a collection of 47 extended-spectrum ß-lactamase (ESBL) producing enterobacterial isolates with reduced susceptibility to fluoroquinolones, recovered at Nantes University hospital, in 2006. qnr, aac(6')-Ib-cr, and qepA genes were screened by PCR, and positive results were subsequently confirmed by sequencing. The epidemiological relationship between positive isolates was studied by pulsed-field gel electrophoresis (PFGE). qnr-positive isolates were analyzed for antimicrobial susceptibility and presence of mutations in the quinolone resistance-determining region (QRDR) of gyrA and parC genes. ESBL genes were characterized by PCR and sequencing. Conjugation experiments were performed to determine whether the qnr-carrying plasmids were self-transferable. Two Klebsiella pneumoniae isolates (4.3%), not clonally related, harboured a qnrS1 gene, whereas no qnrA- or qnrB-positive isolate was detected. The aac(6')-Ib-cr gene was detected in 11 Escherichia coli and one K. pneumoniae isolates. None of the 47 isolates carried the qepA gene. ESBLs associated with QnrS1 were CTX-M-14 and CTX-M-15. The CTX-M-15 producing isolate was highly resistant to fluoroquinolones and harboured three mutations in the QRDR and two PMQR determinants (qnrS1 and aac(6')-Ib-cr). The CTX-M-14-producing isolate exhibited reduced susceptibility or resistance to fluoroquinolones without resistance to nalidixic acid. This strain harboured only a qnr gene on a single 170 kb transferable plasmid, without any mutation in the QRDR. In conclusion, our study showed that aac(6')-Ib-cr gene had occurred in multiclonal ESBL-producing enterobacterial isolates collected at Nantes University hospital in 2006, with a higher prevalence than qnr genes.
Asunto(s)
Antibacterianos/farmacología , Proteínas Bacterianas/genética , Farmacorresistencia Bacteriana Múltiple , Infecciones por Enterobacteriaceae/microbiología , Enterobacteriaceae/efectos de los fármacos , Quinolonas/farmacología , Factores R/aislamiento & purificación , Conjugación Genética , Girasa de ADN/genética , Topoisomerasa de ADN IV/genética , ADN Bacteriano/genética , Farmacorresistencia Bacteriana Múltiple/genética , Electroforesis en Gel de Campo Pulsado , Enterobacteriaceae/enzimología , Enterobacteriaceae/aislamiento & purificación , Infecciones por Enterobacteriaceae/epidemiología , Francia/epidemiología , Hospitales Universitarios , Humanos , Reacción en Cadena de la Polimerasa , Prevalencia , Factores R/genética , Análisis de Secuencia de ADN , Especificidad de la Especie , Resistencia betalactámica/genética , beta-Lactamasas/genéticaAsunto(s)
Proteínas Bacterianas/biosíntesis , Infecciones por Escherichia coli/microbiología , Escherichia coli/clasificación , Escherichia coli/genética , beta-Lactamasas/biosíntesis , Animales , Bovinos , Análisis por Conglomerados , Dermatoglifia del ADN , Escherichia coli/enzimología , Escherichia coli/aislamiento & purificación , Infecciones por Escherichia coli/veterinaria , Genotipo , Humanos , Epidemiología Molecular , Aves de Corral , Análisis de Secuencia de ADN , PorcinosRESUMEN
The search for new protein phosphatase inhibitors in shellfish contaminated by toxin-producing dinoflagellates generally relies on preliminary separation techniques followed by biological tests. To detect such substances without purifying them initially, we developed an approach based on a correlation of the results of two different analytical techniques applied to toxic extracts: high-performance liquid chromatography after derivation of the toxins and the cell morphology transformation assay on KB cells. Application of this protocol to stored frozen mussels showed a decrease in okadaic acid concentration during storage, with formation of degradation derivatives, some of which possessed notable protein phosphatase inhibition activity.