SGM Journals
Research Article

Escherichia coli carrying the blaCTX-M-15 gene of ST648

Zhiyong Zong, Rujia Yu

  • Center of Infectious Diseases, West China Hospital, Sichuan University, Chengdu, PR China
  • Correspondence
    : Zhiyong Zong
    (zongzhiyong{at}gmail.com)

    • Journal of Medical Microbiology 2010; 59(12):1536–1537 · https://doi.org/10.1099/jmm.0.022459-0

    View at publisher PubMed

    Abstract

    blaCTX-M genes have emerged as the dominant genes encoding extended-spectrum β-lactamases in the world. Among more than 90 blaCTX-M genes that have been identified so far, blaCTX-M-15 is the most widespread variant. Although Escherichia coli of O antigen type 25 (O25) and sequence type 131 (ST131) is largely responsible for the intercontinental spread of blaCTX-M-15, some other sequence types have also been found associated with this gene. During a local surveillance of blaCTX-M genes, an isolate of ST648 carrying blaCTX-M-15 was identified and is reported here. As an ST648 isolate carrying blaCTX-M-15 was found in the USA (Sidjabat et al., 2009) recently, the findings here together with the USA report suggest a possible intercontinental spread of this lineage.

    E. coli clinical isolate WCE227 was collected from a urine sample and was resistant to cefotaxime (MIC >32 μg ml−1) and ceftazidime (MIC >16 μg ml−1) as determined by the Phoenix automated microbiology system (BD). The WCE227 was hospital acquired as it was recovered from a patient hospitalized for more than 48 h, although the patient did not receive antimicrobial agents prior to sample collection. The blaCTX-M gene was detected and was subsequently identified as blaCTX-M-15 by PCR and sequencing as described elsewhere (Zong et al., 2008).

    WCE227 was identified as being of the O25b subtype by O25b allele PCR. Phylogenetic group typing (Clermont et al., 2000) revealed that WCE227 belonged to group D (subgroup D1, having chuA but lacking yjaA and Tspe4.C2). WCE227 was of ST648 as determined by multilocus sequence typing (MLST) following an established scheme based on seven housekeeping genes (adk, fumC, gyrB, icd, mdh, purA and recA) available through the University College Cork MLST database ().

    Since blaCTX-M-15 usually co-exists with a few other resistance determinants, aac(3)-II, blaTEM, blaOXA-1 (also called blaOXA-30) and plasmid-encoded quinolone resistance determinants including qnrA, qnrB, qnrS and qepA, were screened by PCR, and aac(6′)-Ib-cr was identified by sequencing. WCE227 had blaOXA-1 and aac(6′)-Ib-cr but no others. WCE227 were resistant to trimethoprim (TMP)/sulfamethoxazole and had a class 1 integron with the dfrA17–aadA5 cassette array that was determined by PCR and sequencing. Class 1 integrons contain sul1, a sulfonamide-resistance gene, in the 3′ conserved segment. dfrA17 encodes TMP-insensitive dihydrofolate reductases conferring resistance to TMP. As WCE227 was resistant to ciprofloxacin, the gyrA allele was partially sequenced. This revealed Ser83Tyr and Asp87Asn substitutions that have been seen in fluoroquinolone-resistant isolates before (Cagnacci et al., 2008).

    Mating was carried out in brain heart infusion broth (Oxoid) with E. coli DH5αRf, a spontaneous rifampicin-resistant mutant of DH5α (ΔlacZ) as the recipient strain. WCE227 was sensitive to rifampicin and transconjugants were selected on 4 μg cefotaxime ml−1 plus 250 μg rifampicin ml−1. In WCE227, blaCTX-M-15 was carried by a conjugative plasmid, for which the incompatibility (Inc) group could not be assigned by PCR-based replicon typing (Carattoli et al., 2005). Surprisingly, aac(6′)-Ib-cr and blaOXA-1 were not co-transferred with blaCTX-M-15 in WCE227, in contrast to the findings of reports from elsewhere. The dfrA17–aadA5 cassette array was also not located on the plasmid carrying blaCTX-M-15 in WCE227.

    ST648 isolates carrying blaCTX-M have been seen before, including two isolates carrying blaCTX-M-15 from the USA (Sidjabat et al., 2009) and two carrying blaCTX-M-1 or blaCTX-M-32 from Spain (Blanco et al., 2009). All of those ST648 isolates like WCE227 belonged to phylogenetic group D. Interestingly, ST648 without blaCTX-M-15 but carrying aac(6′)-Ib-cr had been found in London, UK, prior to the epidemic of blaCTX-M-15 (Jones et al., 2008). The findings from the UK together with the fact that aac(6′)-Ib-cr was not located on the plasmid carrying blaCTX-M-15 in WCE227 suggest that the ST648 lineage acquired the two genes independently. This is in contrast to the ST131 lineage, for which blaCTX-M-15 and aac(6′)-Ib-cr were usually co-transferred on individual plasmids, most of which were FII-like.

    This work was partially supported by a grant from the National Natural Science Foundation of China (project no. 30900052).

    blaCTX-M genes have emerged as the dominant genes encoding extended-spectrum β-lactamases in the world. Among more than 90 blaCTX-M genes that have been identified so far, blaCTX-M-15 is the most widespread variant. Although Escherichia coli of O antigen type 25 (O25) and sequence type 131 (ST131) is largely responsible for the intercontinental spread of blaCTX-M-15, some other sequence types have also been found associated with this gene. During a local surveillance of blaCTX-M genes, an isolate of ST648 carrying blaCTX-M-15 was identified and is reported here. As an ST648 isolate carrying blaCTX-M-15 was found in the USA (Sidjabat et al., 2009) recently, the findings here together with the USA report suggest a possible intercontinental spread of this lineage.

    E. coli clinical isolate WCE227 was collected from a urine sample and was resistant to cefotaxime (MIC >32 μg ml−1) and ceftazidime (MIC >16 μg ml−1) as determined by the Phoenix automated microbiology system (BD). The WCE227 was hospital acquired as it was recovered from a patient hospitalized for more than 48 h, although the patient did not receive antimicrobial agents prior to sample collection. The blaCTX-M gene was detected and was subsequently identified as blaCTX-M-15 by PCR and sequencing as described elsewhere (Zong et al., 2008).

    WCE227 was identified as being of the O25b subtype by O25b allele PCR. Phylogenetic group typing (Clermont et al., 2000) revealed that WCE227 belonged to group D (subgroup D1, having chuA but lacking yjaA and Tspe4.C2). WCE227 was of ST648 as determined by multilocus sequence typing (MLST) following an established scheme based on seven housekeeping genes (adk, fumC, gyrB, icd, mdh, purA and recA) available through the University College Cork MLST database ().

    Since blaCTX-M-15 usually co-exists with a few other resistance determinants, aac(3)-II, blaTEM, blaOXA-1 (also called blaOXA-30) and plasmid-encoded quinolone resistance determinants including qnrA, qnrB, qnrS and qepA, were screened by PCR, and aac(6′)-Ib-cr was identified by sequencing. WCE227 had blaOXA-1 and aac(6′)-Ib-cr but no others. WCE227 were resistant to trimethoprim (TMP)/sulfamethoxazole and had a class 1 integron with the dfrA17–aadA5 cassette array that was determined by PCR and sequencing. Class 1 integrons contain sul1, a sulfonamide-resistance gene, in the 3′ conserved segment. dfrA17 encodes TMP-insensitive dihydrofolate reductases conferring resistance to TMP. As WCE227 was resistant to ciprofloxacin, the gyrA allele was partially sequenced. This revealed Ser83Tyr and Asp87Asn substitutions that have been seen in fluoroquinolone-resistant isolates before (Cagnacci et al., 2008).

    Mating was carried out in brain heart infusion broth (Oxoid) with E. coli DH5αRf, a spontaneous rifampicin-resistant mutant of DH5α (ΔlacZ) as the recipient strain. WCE227 was sensitive to rifampicin and transconjugants were selected on 4 μg cefotaxime ml−1 plus 250 μg rifampicin ml−1. In WCE227, blaCTX-M-15 was carried by a conjugative plasmid, for which the incompatibility (Inc) group could not be assigned by PCR-based replicon typing (Carattoli et al., 2005). Surprisingly, aac(6′)-Ib-cr and blaOXA-1 were not co-transferred with blaCTX-M-15 in WCE227, in contrast to the findings of reports from elsewhere. The dfrA17–aadA5 cassette array was also not located on the plasmid carrying blaCTX-M-15 in WCE227.

    ST648 isolates carrying blaCTX-M have been seen before, including two isolates carrying blaCTX-M-15 from the USA (Sidjabat et al., 2009) and two carrying blaCTX-M-1 or blaCTX-M-32 from Spain (Blanco et al., 2009). All of those ST648 isolates like WCE227 belonged to phylogenetic group D. Interestingly, ST648 without blaCTX-M-15 but carrying aac(6′)-Ib-cr had been found in London, UK, prior to the epidemic of blaCTX-M-15 (Jones et al., 2008). The findings from the UK together with the fact that aac(6′)-Ib-cr was not located on the plasmid carrying blaCTX-M-15 in WCE227 suggest that the ST648 lineage acquired the two genes independently. This is in contrast to the ST131 lineage, for which blaCTX-M-15 and aac(6′)-Ib-cr were usually co-transferred on individual plasmids, most of which were FII-like.

    Acknowledgments

    This work was partially supported by a grant from the National Natural Science Foundation of China (project no. 30900052).

    References

    1. Blanco, M., Alonso, M. P., Nicolas-Chanoine, M. H., Dahbi, G., Mora, A., Blanco, J. E., López, C., Cortés, P., Llagostera, M. & other authors (2009). Molecular epidemiology of Escherichia coli producing extended-spectrum β-lactamases in Lugo (Spain): dissemination of clone O25b : H4–ST131 producing CTX-M-15. J Antimicrob Chemother 63, 1135–1141.
    2. Cagnacci, S., Gualco, L., Debbia, E., Schito, G. C. & Marchese, A. (2008). European emergence of ciprofloxacin-resistant Escherichia coli clonal groups O25 : H4-ST 131 and O15 : K52:H1 causing community-acquired uncomplicated cystitis. J Clin Microbiol 46, 2605–2612.
    3. Carattoli, A., Bertini, A., Villa, L., Falbo, V., Hopkins, K. L. & Threlfall, E. J. (2005). Identification of plasmids by PCR-based replicon typing. J Microbiol Methods 63, 219–228.
    4. Clermont, O., Bonacorsi, S. & Bingen, E. (2000). Rapid and simple determination of the Escherichia coli phylogenetic group. Appl Environ Microbiol 66, 4555–4558.
    5. Jones, G. L., Warren, R. E., Skidmore, S. J., Davies, V. A., Gibreel, T. & Upton, M. (2008). Prevalence and distribution of plasmid-mediated quinolone resistance genes in clinical isolates of Escherichia coli lacking extended-spectrum β-lactamases. J Antimicrob Chemother 62, 1245–1251.
    6. Sidjabat, H. E., Paterson, D. L., Adams-Haduch, J. M., Ewan, L., Pasculle, A. W., Muto, C. A., Tian, G. B. & Doi, Y. (2009). Molecular epidemiology of CTX-M-producing Escherichia coli isolates at a tertiary medical center in western Pennsylvania. Antimicrob Agents Chemother 53, 4733–4739.
    7. Zong, Z., Partridge, S. R., Thomas, L. & Iredell, J. R. (2008). Dominance of blaCTX-M within an Australian extended-spectrum β-lactamase gene pool. Antimicrob Agents Chemother 52, 4198–4202.