Examination of factors promoting progressive emergence of antibiotic resistance among microbiota strains of gastrointestinal tract

Abstract

The benefits of antibiotic usage in treatment of human infection, which we have all enjoyed for decades, are being rapidly eroded by the gradual emergence of antibiotic resistance among bacterial pathogens, posing not only a huge economic cost to the society, but also threatening the lives of millions of patients worldwide. To devise a solution to this problem or at least slow down the rate of antibiotic resistance development, more studies need to be performed to investigate the nature and scope of the factors that promote antibiotic resistance development in bacteria, so as to devise novel approaches to enhance the effectiveness of antibiotic usage and prolong the shelf life of the currently used antimicrobial agents. The objective of this study is therefore to better understand the cellular mechanisms that govern expression of antibiotic resistance phenotypes among bacterial pathogens during the treatment process, especially among microbiota strains in the gastrointestinal tract (GI tract). In order to study the mechanisms underlying the rapid increase in emergence of the antibiotic resistant strains, the first phase of this study involved investigation of various aspects of antibiotic resistance in China, so as to accumulate sufficient material and identify potential targets for subsequent experiments. Currently, antibiotic resistant strains may be divided into two categories: the hospital-acquired pathogens and the community-acquired resistant organisms. The first phase of my work therefore focuses on investigating the current status of the problem, with a particular emphasis on the nosocomial pathogens of Carbapenem-resistant enterobacteriaceae (CRE), and multiple drug resistant (MDR) isolates collected from meat samples. The first chapter of this thesis comprises five separate sections, beginning with studies on clinical CRE isolates, the results of which demonstrate the seriousness of the situation in hospitals. This is followed by three sections on antibiotic resistant Salmonella which is a food borne pathogen, in which we revealed not only a high isolation rates of the pathogen in meat products, but also a high resistance rate among those strains. Analysis on isolates recovered from meat samples hints on a close linkage between community acquired strains and the nosocomial strains, suggesting that a significant proportion of drug resistant clinical strains originated from food animals. In order to delineate mechanisms behind the rapid development of resistance, we then tested the range of mutations that may occur in selected antibiotic target genes, and factors promoting acquisition of extra antibiotic resistance genes. The second chapter describes the range of antibiotic target mutations detectable among gut microbiota strains subjected to antibiotic selection pressure. The first section of the chapter is on the effect of in vivo mutation induction by sub-inhibitory concentrations of antibiotics; we showed that low doses of antibiotic were sufficient to induce mutational changes, often producing identical target mutations. That finding allows us to conclude that a specific cellular mechanism in bacteria is involved in active generation of target mutations. Following this finding, the second section of this chapter demonstrated the role of biofilm formation in resistance development upon in vivo ciprofloxacin treatment. Our results indicate that production of a biofilm is essential or a prerequisite for mutational changes. The third chapter comprises three sections. The first reports result of investigation of CRE strains in food animals China, which allow us to conclude that the carbapenemase genes could readily be transferred without carbapenem selection pressure. The second section reports on analysis of the response of GI tract strains to antibiotic treatment, with results showing that dissemination of the antibiotic resistance genes occur even during the parenteral administration. Based on this finding, we were eventually able to identify factors which play a role in promoting transmission of antibiotic resistance genes among the gut microbiota strains.