JAC Advance Access published online on September 25, 2007
Journal of Antimicrobial Chemotherapy, doi:10.1093/jac/dkm326
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Oligonucleotide microarray for molecular characterization and genotyping of Salmonella spp. strains
1 Department of Genomics and Biotechnology, Institute of Molecular Biology SAS, Dubravska cesta 21, 84551 Bratislava 45, Slovakia 2 Department of Microbiology, Slovak Medical University, Limbova 14, 83303 Bratislava 37, Slovakia
Received 1 May 2007; returned 3 June 2007; revised 17 July 2007; accepted 31 July 2007
* Correspondence address. University of Colorado at Denver and Health Sciences Center, 12800 E 19th Avenue, Aurora, CO 80045, USA. Tel: +1-303-724-3813; Fax: +1-303-724-3838; E-mail: tomas.majtan{at}uchsc.edu or Tel: +421-2-59307429; Fax: +421-2-59307416; E-mail: tomas.majtan{at}savba.sk
Objectives: To characterize and subtype multidrug-resistant Salmonella isolates by determining the virulence factors, prophage sequences and antimicrobial resistance genes using a novel Salmonella-specific oligonucleotide microarray.
Methods: Preliminary screening of 24 Salmonella clinical isolates was carried out by using susceptibility testing, plasmid profiling and class 1 integron PCR. Subsequently, oligonucleotide microarray was involved in genotypic characterization and localization of monitored genetic markers. The presence of antimicrobial resistance genes was also detected and confirmed by PCR and subsequent sequencing. The potential spread of emerging blaSHV-2 was investigated by bacterial conjugation.
Results: All Salmonella strains revealed resistance to two or more (up to nine) antibiotics. Nineteen of them carried class 1 integrons including dfrA1, dfrA12, aadA1, aadA2, blaPSE-1 and blaTEM-1 gene cassettes, respectively. Twenty-three out of 24 Salmonella isolates possessed one or more plasmids. Oligonucleotide microarray characterization and typing revealed the conserved character of Salmonella pathogenicity island virulence factors among three Salmonella enterica serovars, significant variability in prophage sequences and many different antimicrobial resistance gene patterns. Differential labelling of genomic and plasmid DNA, respectively, and hybridization to the microarray made it possible to localize important resistance determinants. Microarray results were successfully confirmed and verified by using PCR. The emerging blaSHV-2 gene from Salmonella Kentucky SK10944 conferring resistance to ceftriaxone and cefotaxime was transferred via bacterial conjugation to Escherichia coli K-12 3110.
Conclusions: Salmonella isolates were quickly and thoroughly characterized by a novel oligonucleotide microarray, which could become a useful tool for detection of virulence and resistance genes and monitoring of their dissemination among salmonellae and closely related bacteria.
Key Words: multidrug resistance , virulence , conjugation
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