Plasmid mediated antibiotic resistance : with focus on extended spectrum β-lactamases (ESBL)

Abstract: Enterobacteriaceae is a family of bacteria including Escherichia coli and Klebsiella pneumoniae that are common colonizers of the gastrointestinal tract. Extended spectrum β- lactamases (ESBL) are bacterial enzymes that degrade β-lactam antibiotics and thus make the bacteria resistant to β-lactam antibiotics. ESBL-producing Enterobacteriaceae (EPE) is an increasing problem constituting a burden on health care systems conferring excess morbidity, prolonged hospital stay and mortality. There are many hundreds of different ESBL-genes described. The most common are the blaCTX-M group. ESBL-genes are often situated on plasmids. This enables a fast dissemination since they can be spread both vertically and horizontally. Horizontal dissemination also enables them to spread between bacteria of different species. Outbreaks of resistant bacteria, such as EPE, must be rapidly detected and contained. Today the main focus during an outbreak is to conduct epidemiological typing of the bacterial strain. However, a horizontal outbreak of resistance genes in the bacterial population can also occur and go undetected if only the strain is subjected to epidemiological typing. There is a need for better understanding of plasmid dissemination and their role in transferring antibiotic resistance genes. In this thesis I have evaluated different methods for epidemiological typing of both strains and plasmids by analyzing different nationwide collections of Swedish EPEs from 2007 to 2011. In Sweden EPE became notifiable according to the communicable disease act in 2007. The number of reported cases has since increased from ~2000 to ~7000 cases per year. In paper I and II we found that both bacterial strain types and ESBL-genotypes were stable over the five- year period. However, a decline in median age for contracting an EPE infection in the Swedish population was seen. This suggests that EPE-carriage is increasing among healthy people in the community. The decline of EPE amongst elderly people might also be an effect of increased awareness and improved infection control in hospitals limiting outbreaks amongst elderly and multi-ill individuals. Paper III describes plasmids of EPE in detail and presents a new plasmid typing approach using next generation sequencing (NGS). The Swedish EPE were found to often carry several plasmids as well as multi-replicon plasmids of IncF-type. Small cryptic plasmids were common in the EPE isolates which raises the question of their role in the bacterial cell. Paper IV describes the strain-plasmid-interplay in the gut flora of three EPE carriers over time. The three patients presented different examples of this interplay. In one of the patients isolates with identical epidemiological strain types were found to also have identical plasmids. In a second patient plasmid variation was observed between identical epidemiological strain types. A third patient had a mixed EPE flora with different epidemiological strain types and different species where similar plasmids were seen in divergent strains as well as the opposite. These results most likely reflect the effect of antibiotic usage and EPE risk factors, such as travel abroad, on the gut flora in terms of bacterial communication and dissemination of ESBL-genes. This thesis has contributed to increased knowledge of the Swedish epidemiology of EPE and increased awareness of the impact of plasmids in the dissemination of common resistance genes.

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