Studies of beta-lactamase mediated resistance in Salmonella enterica

Abstract

This thesis is divided into three sections. Each section focuses on occurrences of beta-lactamase/ ESBL mediated resistance in Salmonella and the role that plasmids play in the dissemination of same. Section 1 of this thesis investigated the genetic basis of a novel ESBL phenotype (cefepimase). This phenotype was observed in 19 isolates of S. Typhimurium from Kenya, Ireland and Malawi. All isolates harboured a c. 39kb plasmid termed pFEP39. Plasmid pFEP39-1dr confered resistance to cefepime with significant inhibition by CA. Sequencing results identified blaOXA-1 in conjunction with an unusual promoter combination directly upstream of blaOXA-1. The P2 promoter was unusual in that there was a GGG triplet upstream of the -10 signal. The cefepimase phenotype is transferred with a plasmid construct in which blaOXA-1 was the only beta-lactamase gene present. Section 2 of this thesis investigated ESBL production in seven S. Kentucky isolates from poultry samples in Ireland. S. Kentucky is a common serovar from poultry in Ireland. Four isolates harboured blaSHV-12 and 3 harboured blaCMY-2. All 7 isolates harboured 2 plasmids. All isolates were similar but distinguishable by PFGE. This thesis reported the emergence of plasmid-mediated broad-spectrum cephalosporin resistance in S. Kentucky in poultry. Section 3 of this thesis investigated the comparison of S1 nuclease PFGE and alkaline lysis for analysis of plasmid DNA. The aim was to compare the reproducibility, accuracy and convenience of both the AL and S1-PFGE methods. S1-PFGE was concluded to be more accurate and reproducible than AL. Plasmids were more clearly defined and better separated using S1-PFGE. The S1-PFGE procedure is more convenient to perform than AL and the availability of agarose plugs for other PFGE analysis is an advantage. The final section of this thesis- the discussion, reflects on the individual chapters.