Citrobacter spp. simultaneously harboring blaCTX-M, blaTEM, blaSHV, blaampC, and insertion sequences IS26 and orf513: an evolutionary phenomenon of recent concern for antibiotic resistance.

A collection of 40 confirmed isolates of Citrobacter spp., obtained from the culture of 5,732 clinical samples during 2006 to 2008, was studied for the presence of blaCTX-M, blaTEM, blaSHV, and blaampC by monoplex and multiplex PCRs. These isolates were also looked at for the presence of insertion elements IS26 and orf513, which play an important role in the resistance gene pool. blaCTX-M, blaTEM, blaSHV, and blaampC were noticed in 67.5%, 40%, 25%, and 40% of isolates, respectively. Sequencing for the specific CTX-M type revealed the presence of CTX-M-15-type extended-spectrum beta-lactamases (ESBL). sul1-type integrons were detected in 32.5% (13/40) of isolates, and a single plasmid of ca. approximately 22 kb was also noticed in all the isolates harboring bla genes. A total of 48.2% (13/27) of isolates harboring blaCTX-M alleles were found to carry IS26 elements and 53.4% (7/13) of isolates harboring the sul1-type integrons were found to carry orf513. Alarmingly, a few isolates simultaneously carried IS26 and orf513, reflecting the occurrence of complex evolution. All the Citrobacter isolates were noticed to produce an amplicon of 400 bp in an IS26 PCR, similar to that of the predominant UK CTX-M-15-producing Escherichia coli clone (clone A) and suggesting a probable genetic relatedness between the two, which could not be proved with British and Indian E. coli isolates in our earlier studies. Here, a first report is being presented describing the occurrence of blaCTX-M-15 in Indian Citrobacter spp. This is also the first report describing the simultaneous occurrence of blaTEM, blaSHV, and blaampC along with blaCTX-M in class 1 integrons. The recent findings indicate that the genetic environment of blaCTX-M-15 has changed through the insertion of IS26 and orf513, although the consequences of these events remain uncertain. It perhaps also suggests a "turnover" of these mobile elements in the population over time.