Amino Acid Substitutions in Mosaic Penicillin-Binding Protein 2 Associated with Reduced Susceptibility to Cefixime in Clinical Isolates of Neisseria gonorrhoeae

Abstract

The molecular mechanisms of reduced susceptibility to cefixime in clinical isolates of Neisseria gonorrhoeae, particularly amino acid substitutions in mosaic penicillin-binding protein 2 (PBP2), were examined. The complete sequence of ponA, penA, and por genes, encoding, respectively, PBP1, PBP2, and porin, were determined for 58 strains isolated in 2002 from Japan. Replacement of leucine 421 by proline in PBP1 and the mosaic-like structure of PBP2 were detected in 48 strains (82.8%) and 28 strains (48.3%), respectively. The presence of mosaic PBP2 was the main cause of the elevated cefixime MIC (4-to 64-fold). In order to identify the mutations responsible for the reduced susceptibility to cefixime in isolates with mosaic PBP2, penA genes with various mutations were transferred to a susceptible strain by genetic transformation. The susceptibility of partial recombinants and site-directed mutants revealed that the replacement of glycine 545 by serine (G545S) was the primary mutation, which led to a two-to fourfold increase in resistance to cephems. Replacement of isoleucine 312 by methionine (I312M) and valine 316 by threonine (V316T), in the presence of the G545S mutation, reduced susceptibility to cefixime, ceftibuten, and cefpodoxime by an additional fourfold. Therefore, three mutations (G545S, I312M, and V316T) in mosaic PBP2 were identified as the amino acid substitutions responsible for reduced susceptibility to cefixime in N. gonorrhoeae.

Whole-Genome Phylogenomic Heterogeneity of Neisseria gonorrhoeae Isolates with Decreased Cephalosporin Susceptibility Collected in Canada between 1989 and 2013

“…The molecular markers previously described as contributing to cephalosporin resistance, such as penA, mtrR, porB, ponA, and pilQ mutations, do not adequately account for the observed phenotypes (4,9,10,15,20,(24)(25)(26)51). Of these markers identified in silico using the WGS data, the presence of a penA mosaic allele, mtrR Ϫ35A deletion and/or a porB G120K mutation were the best indicators of decreased susceptibility to cephalosporins (Table 2).…”

Gonorrhea in Men Sex Men and Heterosexual Men

“…They hypothesized that the L421P substitution in the gene encoding PBP1 ponA may be important in conferring resistance to ceftriaxone. The possible importance of ponA L421P was subsequently supported by data Takahata in which strains with the substitution L421P was associated with increased cephalosporin MICs, compared with only transformants that had the mosaic PBP2 (Takahata, 2006). However, Nicholas found neither the presence or absence of ponA affected the MICs of cephalosporins (Nicholas, 2008).…”

A Study Of The Antimicrobial Susceptibility Pattern Of Neisseria Gonorrhoeae

“…QRNG is also widely distributed in other WHO SEAR and WPR countries [12, 13, 14, 15, and 16]. Recently and most disquieting, the emergence of N. gonorrhoeae isolates with reduced susceptibility or resistance in vitro to broad-spectrum cephalosporins such as cefixime [7,17,18,19,20,21,22] and ceftriaxone [8] have been reported. In occasional cases, treatment failures when using cefixime have been reported [22].…”