Plasmid-Mediated Transference of Multiple-Antibiotic Resistance between Escherichia coli Isolates in the Western Montana Region

Abstract

This thesis investigates the multiple drug resistant (MDR) characteristic within enteric Escherichia coli (E. coli) isolates and their ability to transfer this phenotype via resistant-plasmid (R-plasmid) conjugation. E. coli were isolated from bovine fecal samples in the western Montana region. These isolates were subjected to antibiotic resistance testing, then sorted and stored based on general resistance to two broad-spectrum antibiotics, ampicillin and chlorotetracycline. All isolates underwent molecular differentiation using RAPD PCR, then plasmid extraction using a Qiagen Maxi kit. These results were compared to determine if genetically similar isolates, as determined by RAPD PCR, were more likely to contain a plasmid. Isolates containing a plasmid were subjected to conjugation assays to determine transferability of plasmids. In the conjugation assay, each MDR donor isolate was mated with azide-resistant strain, J53AZr. The following hypotheses were addressed: 1) a strain of E. coli will demonstrate a MDR pattern with resistance to more antibiotics if a plasmid is present and 2) the ability to transfer the MDR characteristic to other strains lacking the plasmid is specific to the donor strain of E. coli and its MDR pattern designated by a plasmid. Plasmid presence in E. coli isolates increased with level of MDR. Results also demonstrated plasmid conjugation between E. coli isolates was more likely to occur as level of MDR increased; however the conjugation assay may require larger sample size to confirm the trend. The RAPD PCR method should also be altered to illustrate more accurate banding patterns and verify molecular differences between strains containing plasmids.