CTX-M-123, a Novel Hybrid of the CTX-M-1 and CTX-M-9 Group β-Lactamases Recovered from Escherichia coli Isolates in China

Abstract

T he rapid and extensive dissemination of CTX-M-type extended-spectrum ␤-lactamases (ESBLs) in humans, animals, and the environment is one of the most successful stories of microbial drug resistance (1). CTX-M ␤-lactamases were originally named for their substrate preference for cefotaxime over ceftazidime (2), but subsequent microevolution means that many of the 140 variants currently listed (as of June 2013) at http://www.lahey.org /studies/other.asp#table1 confer resistance to both cefotaxime and ceftazidime (3, 4). CTX-M variants can be divided into six clusters (CTX-M-1, CTX-M-2, CTX-M-8, CTX-M-9, CTX-M-25, and KLUC, with Ͻ90% identity between clusters and Ͼ95% identity within clusters) (1, 5, 6). CTX-M-1 and CTX-M-9 group enzymes are the most frequently reported groups worldwide, and CTX-M-15 and CTX-M-14, respectively, are the most common variants within these groups (4, 6). In 2010, CTX-M-64, a chimera of the CTX-M-1 (N-and C-terminal domains) and CTX-M-9 (central domain) group enzymes, was identified in Japan (7) and then China (8), and a similar chimeric protein, CTX-M-132, was also recently reported in Escherichia coli from a chicken carcass in China (GenBank accession no. JX313020) (Fig. 1). Here, we have identified bla CTX-M-123 , a third hybrid of bla CTX-M-1 and bla CTX-M-9 group genes.Two E. coli isolates were obtained from the feces of diseased animals, FKP358 (pig) in Guangdong Province in December 2010, and AHC4 (chicken) in Anhui Province (ϳ1,200 km from Guangdong Province) in June 2011. Multilocus sequence typing (MLST) (http://mlst.ucc.ie/mlst/dbs/Ecoli) identified AHC4 as sequence type 746 (ST746) and FKP358 as ST165. Both isolates were resistant to most ␤-lactams, including ceftazidime and cefquinome (Ͼ64 g/ml), but were susceptible to imipenem and cefoxitin (Table 1). PCR screening for selected bla genes (bla TEM , bla SHV , bla CTX-M , and bla CMY-2 ) (8, 9), plasmid-mediated quinolone resistance (PMQR) genes, including oqxAB (10), and fosfomycin resistance (fos) genes (11) revealed bla CTX-M , bla TEM , fosA3, and oqxAB in FKP358 and bla CTX-M in AHC4.Broth mating with E. coli C600 (streptomycin resistant; MIC, Ͼ2,000 g/ml) and selection on cefotaxime (2 g/ml) and streptomycin (2,000 g/ml) gave transconjugants from AHC4 only, carrying two plasmids of ϳ90 kb (pHNAH4-1) and Ͻ5 kb (pHNAH4-3). Hybridization with a probe generated by digoxigenin (DIG) High Prime DNA labeling (Roche, Germany) of the amplicon generated with primers M123-F and M123-R (see Table S1 in the supplemental material) indicated that the bla CTX-M gene is on pHNAH4-1. Transformation into E. coli DH5␣ with selection on cefotaxime (2 g/ml) gave transformants carrying a single ϳ60-kb plasmid with bla CTX-M from AHC4 (pHNAH4-2) and a single ϳ90-kb plasmid with bla CTX-M , bla TEM , and fosA3 from FKP358 (pHNP358).Complete sequencing of bla CTX-M genes amplified using primers designed from the typical context of bla CTX-M-1 group genes (see Table S1 in the supplemental material) revealed bla CTX-M-55 in pHNAH4-2 and a nove...

Residues Distal to the Active Site Contribute to Enhanced Catalytic Activity of Variant and Hybrid β-Lactamases Derived from CTX-M-14 and CTX-M-15

“…This suggests that recombination may take place between bla CTX-M- 14 and bla CTX-M-55 , producing various hybrid enzymes. Interestingly, all three published hybrids, CTX-M-64, CTX-M-123, and CTX-M-137, confer relatively high ceftazidime MICs compared to the majority of other CTX-M-type ESBLs (3)(4)(5). Conversely, the hydrolytic activities of CTX-M-55, CTX-M-123, and CTX-M-132 enzymes have not been described.…”

“…More than 95% identity is often observed within the same cluster, whereas less than 90% identity is detectable between clusters (2). Several hybrids of the CTX-M-1 and -9 groups, namely, CTX-M-64, CTX-M-123, CTX-M-137, and CTX-M-132, have also been identified recently (3)(4)(5). Formation of these hybrid CTX-M enzymes was suggested to be the result of recombination between bla CTX-M- 15 and bla CTX-M-14 , the two most dominant variants detectable worldwide (1,6).…”

bla CTX-M-1/9/1 Hybrid Genes May Have Been Generated from bla CTX-M-15 on an IncI2 Plasmid

“…CTX-M-137 is a simple hybrid, with the N terminus matching CTX-M-14 and the C terminus matching CTX-M-15 (6). The other three hybrids match CTX-M-1 group enzymes at the start and end but CTX-M-9 group enzymes in the middle, suggesting different double crossover events (4). Interestingly, these four CTX-M hybrids have all been found in China, where three of them were first reported, and have been detected in animals and in animal food (4,(6)(7)(8)(9)(10)(11).…”

“…Four CTX-M enzymes (CTX-M-64, CTX-M-123, CTX-M-132, and CTX-M-137) that are hybrids of members of the CTX-M-1 and CTX-M-9 groups have emerged in recent years (4)(5)(6). CTX-M-137 is a simple hybrid, with the N terminus matching CTX-M-14 and the C terminus matching CTX-M-15 (6).…”

Antimicrobial resistance and integrons of ESBL-producing thermotolerant coliforms from a water reservoir in Tai'an, China

“…The blaCTX-M-3 gene was frequently detected in Chinese hospitals [30,35]. The blaCTX-M-123 gene, a hybrid of blaCTX-M-1 and blaCTX-M-9 group genes, was first isolated from diseased animals in Guangdong province of China in 2010 [36]. SHV-12 was first reported in China, while a subsequent largescale study of isolates from six provinces, collected between 1998 and 2002, reported SHV-12 as the most common SHV type (ten isolates), with much smaller number of SHV-5, -2 and -9 [28].…”