High prevalence of TEM-135 expression from the Asian plasmid in penicillinase-producing Neisseria gonorrhoeae from Hangzhou, China.

Penicillinase-producing Neisseria gonorrhoeae (PPNG) expressing the TEM β-lactamase variant TEM-135 are a global public-health concern as this variant requires only a single amino acid substitution to develop into an extended-spectrum β-lactamase. The aim of this study was to investigate the epidemiology of blaTEM-135 in 505 N. gonorrhoeae isolates from Hangzhou, China, during the periods 2011-2012 and 2015-2017. Investigation by nitrocefin test and mismatch amplification PCR showed that 41.0% (207/505) of the isolates were PPNG, of which 37.2% (77/207) contained the blaTEM-135 gene. Further PCR-based plasmid typing showed that blaTEM-135 was predominantly expressed from the Asian plasmid (94%). PPNG isolates consisted of three major clusters, namely Asian plasmid/blaTEM-135 (34.8%), Asian plasmid/blaTEM-1 (32.4%) and African plasmid/blaTEM-1 (28.0%), which showed significant differences in penicillin minimum inhibitory concentrations (MICs) determined by the agar dilution method. Representative isolates were investigated by quantitative real-time PCR (plasmid copy number and blaTEM gene expression), western blot analysis (TEM levels and TEM stability) and in vivo β-lactamase activity assays to elucidate the cause of the observed differences in penicillin MIC. Overall, isolates of the Asian plasmid/blaTEM-135 cluster showed the highest β-lactamase activity, which was explained by higher blaTEM gene expression (Asian versus African plasmid) and higher TEM stability (TEM-135 versus TEM-1). In conclusion, the blaTEM-135 gene is commonly present on the Asian plasmid in PPNG isolates from Hangzhou. The PPNG isolate cluster containing the Asian plasmid and blaTEM-135 showed the highest penicillin MICs, which might explain its abundance in the Hangzhou population.