In a Policy Forum, Teodora Wi and colleagues discuss the challenges of antimicrobial resistance in gonococci.

SUMMARY: Gonorrhea, which may become untreatable due to multiple resistance to available antibiotics, remains a public health problem worldwide. Precise methods for typing Neisseria gonorrhoeae , together with epidemiological information, are crucial for an enhanced understanding regarding issues involving epidemiology, test of cure and contact tracing, identifying core groups and risk behaviors, and recommending effective antimicrobial treatment, control, and preventive measures. This review evaluates methods for typing N. gonorrhoeae isolates and recommends various methods for different situations. Phenotypic typing methods, as well as some now-outdated DNA-based methods, have limited usefulness in differentiating between strains of N. gonorrhoeae. Genotypic methods based on DNA sequencing are preferred, and the selection of the appropriate genotypic method should be guided by its performance characteristics and whether short-term epidemiology (microepidemiology) or long-term and/or global epidemiology (macroepidemiology) matters are being investigated. Currently, for microepidemiological questions, the best methods for fast, objective, portable, highly discriminatory, reproducible, typeable, and high-throughput characterization are N. gonorrhoeae multiantigen sequence typing (NG-MAST) or full- or extended-length porB gene sequencing. However, pulsed-field gel electrophoresis (PFGE) and Opa typing can be valuable in specific situations, i.e., extreme microepidemiology, despite their limitations. For macroepidemiological studies and phylogenetic studies, DNA sequencing of chromosomal housekeeping genes, such as multilocus sequence typing (MLST), provides a more nuanced understanding.

Antimicrobial resistance (AMR) in Neisseria gonorrhoeae is a major concern worldwide. It has been recently feared that the bla TEM-1 gene is, via bla TEM-135, evolving into an extended-spectrum β-lactamase (ESBL), which could degrade all cephalosporins including ceftriaxone. The aims of the present study were to characterize the bla TEM genes, types of β-lactamase plasmids, the degradation of ampicillin by TEM-135 compared to TEM-1, and to perform molecular epidemiological typing of β-lactamase-producing N. gonorrhoeae strains internationally. β-lactamase producing N. gonorrhoeae isolates (n = 139) cultured from 2000 to 2011 in 15 countries were examined using antibiograms, bla TEM gene sequencing, β-lactamase plasmid typing, and N. gonorrhoeae multiantigen sequence typing (NG-MAST). Furthermore, the bla TEM gene was sequenced in the first described Toronto plasmid (pJD7), one of the first Asian plasmids (pJD4) and African plasmids (pJD5) isolated in Canada. The degradation of ampicillin by TEM-135 compared to TEM-1 was examined using a MALDI-TOF MS hydrolysis assay. Six different bla TEM sequences were identified (among isolates with 125 different NG-MAST STs), i.e. bla TEM-1 (in 104 isolates), bla TEM-135 (in 30 isolates), and four novel bla TEM sequences (in 5 isolates). The bla TEM-1 allele was only found in the African and Asian plasmids, while all Rio/Toronto plasmids possessed the bla TEM-135 allele. Most interesting, the first described gonococcal Toronto plasmid (pJD7), identified in 1984, also possessed the highly conserved bla TEM-135 allele. The degradation of ampicillin by TEM-135 compared to TEM-1 was indistinguishable in the MALDI-TOF MS hydrolysis assay. bla TEM-135, encoding TEM-135, is predominantly and originally associated with the Rio/Toronto plasmid and prevalent among the β-lactamase producing gonococcal strains circulating globally. bla TEM-135 does not appear, as previously hypothesized, to have recently evolved due to some evolutionary selective pressure, for example, by the extensive use of extended-spectrum cephalosporins worldwide. On the contrary, the present study shows that bla TEM-135 existed in the Toronto plasmid from its discovery and that bla TEM-135 is highly conserved (not further evolved in the past >30 years). Nevertheless, international studies for monitoring the presence of different bla TEM alleles, the possible evolution of the bla TEM-135 allele, and the types of β-lactamase producing plasmids, remain imperative.

To elucidate loci in Neisseria gonorrhoeae implicated in reduced susceptibility to ceftriaxone. N. gonorrhoeae isolates were collected in Shanghai, China, in 2005 and 2008. Twenty-eight isolates with reduced susceptibility to ceftriaxone (CRO^Red; MIC = 0.125–0.25 mg/L) were studied for mutations in PorB (porB), MtrR (mtrR), PBP2 (penA) and PBP1 (ponA). The mutation profiles of the 28 CRO^Red isolates were compared with those of 32 ceftriaxone-susceptible isolates (CRO^S; MIC = 0.004–0.016 mg/L). porB-based DNA sequence typing and N. gonorrhoeae multi-antigen sequence typing (NG-MAST) analyses were performed. Significantly more CRO^Red isolates (89.3%) exhibited a PIB phenotype as compared with the CRO^S isolates (59.4%) (P = 0.02). Double mutations (G45D/H105Y or A39T/H105Y) in MtrR were associated with CRO^Red phenotypes. A ‘wild-type’ MtrR protein characterized CRO^Red isolates (50.0%, 14/28), while a single H105Y mutation was observed only in CRO^S isolates (43.8%, 14/32). Both CRO^Red and CRO^S isolates carried an ‘−A’ deletion in the mtrR promoter. Six of 15 mutation patterns observed in PBP2 were new. Mutation patterns XIII (17.9% of CRO^Red isolates) and XVII or XVIII (25.0% of CRO^Red isolates) of PBP2 comprised A501V/G542S or A501V/P551S double mutations and were associated with a CRO^Red phenotype. The mosaic PBP2 (pattern X) was not observed. The L421P mutation in PBP1 was observed in all CRO^Red and in 97.0% of CRO^S isolates. CRO^Red isolates were non-clonal. Reduced susceptibility to ceftriaxone in N. gonorrhoeae is mediated by porB1b alleles and is associated with specific mutations in PBP2 and in the DNA binding and dimerization domains of MtrR.

Background: The successful treatment of Neisseria gonorrhoeae (NG) infections is increasingly problematic because of the resistance of this pathogen to multiple antimicrobial agents. This development underscores the need for new antimicrobial sources. In the current study, 21 crude methanol extracts, from 19 plants used in Colombian traditional medicine for cutaneous infections, were screened for antimicrobial activity against NG. Methods: Extracts were screened by disc susceptibility assay. In addition, the minimum inhibitory concentrations of active compounds from P. lanceaefolium were assayed using a panel of 26 NG strains comprising 12 antibiotic-resistant phenotypes. Results: In all, 71% of the crude extracts exhibited antibacterial activity against the antibiotic susceptible NG strain WHO V, whereas 10% of the extracts inhibited penicillinase-producing NG strain GC1–182. The crude extract of Piper lanceaefolium was the only extract to show significant activity without ultraviolet (UV) light activation. Preliminary screening identified 3 compounds in this plant possessing antimicrobial activity: the flavonoids 5,7-dihydroxyflavanone (pinocembrin), 2′,4′,6′-trihydroxychalcone (pinocembrin chalcone), and the prenylated benzoic acid derivative cyclolanceaefolic acid methyl ester. Pinocembrin and pinocembrin chalcone inhibited 100% of the NG panel at 64 μg/mL and 128 μg/mL, respectively, whereas cyclolanceaefolic acid methyl ester inhibited 44% of the strains at 128 μg/mL. Conclusions: This is the first report of the antibacterial activity of Columbian plants against NG. The activity of the 2 flavonoids, pinocembrin, and pinocembrin chalcone, toward both susceptible and resistant NG strains makes them promising candidates for further research.