MOBILIZATION OF SIDEROPHORE GENES IN ANTIBIOTIC RESISTANT ENVIRONMENTAL ESCHERICHIA COLI

Siderophores are iron chelating compounds secreted by bacteria to acquire iron in low-iron environments. Some siderophores play an important role in virulence of extra-intestinal pathogenic E. coli (ExPEC) by allowing them to acquire iron in their host. In ExPEC, genes encoding for virulence-associated siderophores can be found in plasmids or chromosomal pathogenicity islands. They are mobilized through genetic elements such as conjugative plasmids, which are self-replicating DNA molecules that are transferred from one cell to another by a process called conjugation. Other mobile genetic elements called Transposases can move genes by extracting DNA sequences and inserting them in other regions of the genome. These processes facilitate the existence of an antibiotic-resistance gene pool in the environment. We hypothesize we can find siderophore genes in plasmids that are typically associated with the antibiotic-resistance gene pool in the environment. Our objectives were: (1) to describe an environmental strain that has a plasmid-mediated siderophore and antibiotic resistance genes, and (2) to study their conjugative plasmids encoding for siderophores and transposases that promote the mobilization of these genes. As part of a larger sample, an isolate was collected in Buckhall Branch creek in Mannassas, Virginia that was identified as E. coli using MALDI-TOF. Disk diffusion tests showed that this isolate is resistant to Ampicillin, Cephalothin, Sulfisoxaszole, Streptomycin, Tetracycline, Ticarcillin, and Trimethoprim. A conjugation experiment was conducted to confirm that a conjugative plasmid is responsible for the aforementioned resistances to antibiotics. Chrome Azurol Assay Test confirmed an increased siderophore production in a transconjugant carrying this plasmid. DNA was extracted and sequenced. The plasmid obtained was of 169,397 base pairs with three regions: (1) conjugative IncF type region followed by (2) an antibiotic resistance region with genes encoding for resistance to Tetracycline (tetA), Aminoglycosides (strAB), Beta-lactams (blaTEM-1b) and Sulfanomides (sul2). These genes are flanked by transposases Tn3 and IS6. Next is (3) a region of virulence-associated genes with the siderophore Salmochelin iroBCDEN flanked by transposases IS3, IS66, and Aerobactin iucABCD flanked by IS110, IS481 –suggesting transposon-mediated acquisition of virulence factors. Manganese ABC transporter sitABCD, temperature-sensitive hemagglutinin tsh and colicin V cvaB were also found in this gene cluster. This virulent configuration is similar to conjugative plasmids of avian pathogenic strains of E. coli like pAPEC-1, which contains the genes sitABCD, cvaB, tsh, siderophores Aerobactin and Salmochelin. A conjugative plasmid of this particular siderophore configuration and a wide array of antibiotic resistance genes found in the environment suggests a potential a reservoir of siderophore genes that parallels the antibiotic resistance gene pool present in the environment.