Laura Martín

Acquisition of Resistance to Extended-Spectrum Cephalosporins by Salmonella enterica subsp. enterica Serovar Newport and Escherichia coli in the Turkey Poult Intestinal Tract

ABSTRACT Salmonella enterica subsp. enterica serovar Newport resistant to the extended-spectrum cephalosporins (ESCs) and other antimicrobials causes septicemic salmonellosis in humans and animals and is increasingly isolated from humans, animals, foods, and environmental sources. Mechanisms whereby serovar Newport bacteria become resistant to ESCs and other classes of antimicrobials while inhabiting the intestinal tract are not well understood. The present study shows that 25.3% of serovar Newport strains isolated from the turkey poult intestinal tract after the animals were dosed with Escherichia coli harboring a large conjugative plasmid encoding the CMY-2 β-lactamase and other drug resistance determinants acquired the plasmid and its associated drug resistance genes. The conjugative plasmid containing the cmy-2 gene was transferred not only from the donor E. coli to Salmonella serovar Newport but also to another E. coli serotype present in the intestinal tract. Laboratory studies showed that the plasmid could be readily transferred between serovar Newport and E. coli intestinal isolates. Administration of a single dose of ceftiofur, used to prevent septicemic colibacillosis, to 1-day-old turkeys did not result in the isolation of ceftiofur-resistant E. coli or Salmonella serovar Newport. There was a remarkable association between serotype, drug resistance, and plasmid profile among the E. coli strains isolated from the poults. This study shows that Salmonella serovar Newport can become resistant to ESCs and other antibiotics by acquiring a conjugative drug resistance plasmid from E. coli in the intestines.

Subinhibitory concentrations of tetracycline affect virulence gene expression in a multi-resistant Salmonella enterica subsp. enterica serovar Typhimurium DT104

Treatment of salmonellosis with antibiotics is controversial and may prolong carriage and shedding. Therefore, this study sought to investigate if exposure to antimicrobials influences the expression of factors involved in virulence and host colonization. The effect of subinhibitory tetracycline treatment (16 microg/ml, 30 min) on a multi-drug resistant Salmonella Typhimurium DT104 strain was investigated using a targeted microarray. Real-time reverse transcriptase PCR was used to confirm and further assess transcription of 10 selected genes. An in vitro cell invasion assay was performed to assess the invasiveness of the tetracycline-treated isolate. Out of 323 genes, 11 were significantly up-regulated and four were down-regulated in the microarray assays. The hilD and hilA genes, both regulators of Salmonella Pathogenicity Island 1, were up-regulated. Other up-regulated genes included the fliC, fliD, motA and motB genes, involved in motility, the fur gene, an important regulator of iron acquisition systems and of acid tolerance. The drug-exposed replicates showed a 2.5-fold increase in intracellular bacteria over the non-exposed control in cell cultures. These findings suggest a drug-induced expression profile consistent with the early stages of Salmonella infection and invasion concomitant with an increased ability to invade epithelial cells in vitro.

Characterization of blaCMY-2 Plasmids in Salmonella and Escherichia coli Isolates from Food Animals in Canada

ABSTRACT One hundred thirty-four bla CMY-2 plasmids from Salmonella and Escherichia coli strains from animals and food in Canada were characterized. Five plasmid groups were identified based on replicon type and restriction profiles. Three groups contained E. coli plasmids only. IncA/C plasmids included most multiresistant plasmids and all those of bovine origin.

Real-Time PCR Detection of Phaeomoniella chlamydospora and Phaeoacremonium aleophilum

ABSTRACT Phaeomoniella chlamydospora and Phaeoacremonium aleophilum are the two main fungal causal agents of Petri disease and esca. Both diseases cause significant economic losses to viticulturalists. Since no curative control measures are known, proactive defensive measures must be taken. An important aspect of current research is the development of sensitive and time-saving protocols for the detection and identification of these pathogens. Real-time PCR based on the amplification of specific sequences is now being used for the identification and quantification of many infective agents. The present work reports real-time PCR protocols for identification of P. chlamydospora and P. aleophilum. Specificity was demonstrated against purified DNA from 60 P. chlamydospora isolates or 61 P. aleophilum isolates, and no amplification was obtained with 54 nontarget DNAs. The limits of detection (i.e., DNA detectable in 95% of reactions) were around 100 fg for P. chlamydospora and 50 fg for P. aleophilum. Detection was specific and sensitive for P. chlamydospora and P. aleophilum. Spores of P. chlamydospora and P. aleophilum were detected without the need for DNA purification. The established protocols detected these fungi in wood samples after DNA purification. P. chlamydospora was detectable without DNA purification and isolation in 67% of reactions. The detection of these pathogens in wood samples has great potential for use in pathogen-free certification schemes.