NOVEL DNA SEQUENCE OF DHA-1 GENE ON AN INC F PLASMID FROM AN ENVIRONMENTAL K. PNEUMONIAE

The rapid spread of antibiotic resistance represents a severe problem in healthcare; therefore, understanding the dissemination of antibiotic resistant genes through mobile elements is vital. The DHA-1 gene is a plasmid-mediated inducible AmpC beta-lactamase that confers resistance to cephamycin and extended-spectrum cephalosporin antibiotics. DHA-1 gene was found in 1997. As part of a large survey of antibiotic resistant bacteria, we identified an environmental isolate with a plasmid-mediated DHA-1 gene. The objective of this study is to obtain the complete sequence of the plasmid and compare the mobile element harboring the DHA-1 gene from our isolate with clinical isolates containing the DHA-1 gene found in the NCBI GenBank. The environmental K. pneumoniae was collected from Eisenhower Park in California and designated SW4037. Polymerase chain reaction identified a DHA gene in SW4037. A conjugation experiment was performed successfully between SW4037 and a J53 E. coli recipient strain. The transconjugant was designated SW4037-J53-TMP-1. An 86 kb plasmid was isolated and sequenced from SW4037-J53-TMP-1 and then assembled to produce an 86,430 bp plasmid designated pSW4037-86430. This plasmid belongs to incF Incompatibility group and harbors six antibiotic resistant genes located in a multiresistance region (MRR) of 21kb conferring resistance to six groups of antibiotics: aminoglycoside (aac(3)-IIa), sulphonamide (sul1), macrolide (mphA), quinolone (QnrB4), trimethoprim (dfrA17) and beta-lactam (DHA-1). Our analysis of plasmid pSW4037-86430 shows a novel IS26 transposase harboring a DHA-1 gene with a chrA/mphA ending (characteristics of class 1 integrons). This region (99% identical coverage and 99% query coverage) has only been found in an ISCr1 class 1 integron of a K. pneumoniae incR plasmid from a clinical sample in France in the year 2008. Furthermore, a total of 10 mobile elements were found on pSW4037-86430: IS1 (2), IS3 (1), IS26 (4), IS6100 (1), IS150 (1) and Tn3 (1). All clinical isolates gathered from GenBank show an ISCr1 class 1 integron harboring DHA-1. The mobile elements as well as the Incompatibility types of the plasmids harboring the DHA-1 gene from NCBI clinical isolates are unknown since only partial sequencing was performed. This study indicates that an IS26 transposon truncated an ISCr1 class 1 integron with a chrA/mphA ending--which suggests that new configurations of mobile elements harboring antibiotic resistant genes can be found in environmental bacteria derived from clinical bacteria. This is the first report of a novel genetic configuration of a DHA-1 gene found in an IS26 transposase carried by an incF Incompatibility group plasmid of an environmental K. pneumonia.