Molecular characterisation of class 1 integrons in Salmonella enterica serovar Choleraesuis isolates from southern Taiwan

Abstract

Integron-mediated multidrug resistance in Salmonellaenterica serovar Choleraesuis poses a major concern in human salmonellosis in Taiwan. In this study, 71 strains of S. Choleraesuis harbouring a class 1 integron from humans and animals were characterised for detection of integrons, gene cassettes and the spvC virulence gene using molecular genetic techniques. All 71 strains tested were negative for class 2 and 3 integrons. Fifty-eight (81.7%) of the isolates harboured a class 1 integron with different gene cassettes, including dfr12-orfF-aadA2 in 50 strains, dfrA1-UN in 7 strains and aadA1 in 1 strain. Among the 71 isolates, 1 isolate of S. Choleraesuis H40 harboured two class 1 integrons, the first integron with the dfr12-orfF-aadA2 gene cassette and the second integron with a novel single gene cassette aadA22, a new variant of the aadA gene family. Moreover, 13 of the 71 isolates were negatively amplified by 5′-CS and 3′-CS specific primer pairs and represented a novel gene cassette array of sat-psp-aadA2-cmlA1-aadA1-like-qacH-tnp-sul3 in class 1 integrons. In mating experiments, the transferability of integron-mediated drug resistance in S. Choleraesuis isolates was demonstrated. The class 1 integron-borne gene cassette dfr12-orfF-aadA2 was characterised as becoming predominant in multidrug-resistant S. Choleraesuis isolates. Furthermore, both the dfrA1-UN gene cassette and the sul3-associated novel gene cassette were also identified in the S. Choleraesuis isolates. To date, this is the first report describing the dissemination of a sul3-associated integron in S. Choleraesuis isolates in Taiwan.

Introduction

Salmonellosis is still a major infectious disease worldwide, including in developed, industrialised countries [1]. In particular, Salmonellaenterica serovar Choleraesuis, a host-adapted, facultative intracellular pathogen of swine, most often results in severe invasive human infections [2].

In recent years, molecular mechanisms for acquisition and dissemination of resistance determinants among clinical isolates of bacterial populations have been studied intensively [3], [4]. Integrons are the most important resistance mechanism for Gram-negative pathogens, carrying resistance gene cassettes captured via site-specific recombination catalysed by specific integrase genes [4]. Three classes of resistance integrons are known to be associated with antibiotic resistance phenotypes in clinical isolates of Gram-negative bacteria [3]. Class 1 integrons are the most frequently detected integrons associated with sul1 genes that are part of the 3′ conserved segment (3′-CS). In recent years, class 1 integrons with variable 3′ ends have been described [5]. Moreover, dissemination of sul3, associated with plasmid-borne class 1 integrons containing an unusual 3′-CS site, has also been reported [6].

In Taiwan, owing to the abuse of antimicrobials [7], multidrug-resistant (MDR) S. Choleraesuis has emerged as a major concern in human salmonellosis, which always leads to difficulties in antibiotic therapy and a higher morbidity and mortality rate [2], [8]. In Taiwan, the first quinolone-resistant S. Choleraesuis isolated from swine and humans were identified in 1993 and 1996, respectively [9]. Moreover, the prevalence of human infections by ciprofloxacin-resistant S. Choleraesuis increased [10]. Notably, >60% of S. Choleraesuis isolates were resistant to ciprofloxacin and pose a significant threat to the population of Taiwan [11].

However, few reports have elucidated the characteristics of gene cassettes in integrons and the resistance phenotypes in S. Choleraesuis among human and animal isolates. The aims of this study were to investigate the molecular characterisation of class 1 integrons and gene cassettes in MDR S. Choleraesuis isolated in southern Taiwan. Antimicrobial resistance phenotypes, class 1 integron-associated resistance profiles, and localisation of resistance determinants were also analysed. Furthermore, a novel aadA22 gene, presenting as a single-gene cassette in a class 1 integron, together with a novel gene cassette array of sat-psp-aadA2-cmlA1-aadA1-like-qacH- tnp-sul3, were also characterised.