Canary in the Coal Mine: How Resistance Surveillance in Commensals Could Help Curb the Spread of AMR in Pathogenic Neisseria

Abstract

Antimicrobial resistance (AMR) is widespread within Neisseria gonorrhoeae populations. Recent work has highlighted the importance of commensal Neisseria (cN) as a source of AMR for their pathogenic relatives through horizontal gene transfer (HGT) of AMR alleles, such as mosaic penicillin binding protein 2 (penA), multiple transferable efflux pump (mtr), and DNA gyrase subunit A (gyrA) which impact beta-lactam, azithromycin, and ciprofloxacin susceptibility, respectively. However, nonpathogenic commensal species are rarely characterized. Here, we propose that surveillance of the universally carried commensal Neisseria may play the role of the "canary in the coal mine," and reveal circulating known and novel antimicrobial resistance determinants transferable to pathogenic Neisseria. We summarize the current understanding of commensal Neisseria as an AMR reservoir, and call to increase research on commensal Neisseria species, through expanding established gonococcal surveillance programs to include the collection, isolation, antimicrobial resistance phenotyping, and whole-genome sequencing (WGS) of commensal isolates. This will help combat AMR in the pathogenic Neisseria by: (i) determining the contemporary AMR profile of commensal Neisseria, (ii) correlating AMR phenotypes with known and novel genetic determinants, (iii) qualifying and quantifying horizontal gene transfer (HGT) for AMR determinants, and (iv) expanding commensal Neisseria genomic databases, perhaps leading to the identification of new drug and vaccine targets. The proposed modification to established Neisseria collection protocols could transform our ability to address AMR N. gonorrhoeae, while requiring minor modifications to current surveillance practices. IMPORTANCE Contemporary increases in the prevalence of antimicrobial resistance (AMR) in Neisseria gonorrhoeae populations is a direct threat to global public health and the effective treatment of gonorrhea. Substantial effort and financial support are being spent on identifying resistance mechanisms circulating within the gonococcal population. However, these surveys often overlook a known source of resistance for gonococci-the commensal Neisseria. Commensal Neisseria and pathogenic Neisseria frequently share DNA through horizontal gene transfer, which has played a large role in rendering antibiotic therapies ineffective in pathogenic Neisseria populations. Here, we propose the expansion of established gonococcal surveillance programs to integrate a collection, AMR profiling, and genomic sequencing pipeline for commensal species. This proposed expansion will enhance the field's ability to identify resistance in and from nonpathogenic reservoirs and anticipate AMR trends in pathogenic Neisseria.

Keywords: N. gonorrhoeae; Neisseria; Neisseria gonorrhoeae; antibiotic resistance; commensal bacteria; horizontal gene transfer; microbiome.

FIG 1

(A) Schematic representation of some of the known mechanisms of resistance in N. gonorrhoeae, including those that have been acquired from (bold text and *) or have been documented to be present in the commensal Neisseria (bold text). Known resistance encompasses multiple mechanisms, including: (A1) mutations enhancing the affinity of antibiotics for the Mtr efflux pump, increasing their efflux from the cell; (A2) mutations altering the structures of the targets of antibiotics reducing binding affinity (e.g., β-lactams [PEN] and PBP2, azithromycin [AZM] and ribosomal proteins, and fluoroquinolones [FQ] and DNA Gyrase Subunit A); (A3) genes encoding enzymes that modify antibiotics through degradation or blocking their target site; (A4) altered promoter motifs which increase or decrease the expression of genes; (A5) changes in cell surface permeability through mechanisms such as the decoration of Lipid A with phosphoethanolamine (PEA) (not yet described in commensal Neisseria); and (A6) development of alternative pathways to antibiotic targets (not yet described in commensal Neisseria). (B) Of the six described mechanisms, four have been shared from or documented to be present within Neisseria commensal communities and thus available for horizontal gene transfer (HGT) to pathogenic Neisseria through their natural competence and pilus-mediated DNA uptake machinery; the other two (A5 and A6) are yet to be described in commensal Neisseria. (C) Of great concern is the possibility that commensal Neisseria could acquire extended spectrum β-lactamase (ESBL) genes from other bacteria with which they live proximally to in the oral and nasopharynx, rendering them resistant to ceftriaxone, our last line of defense against gonorrhea. To note, chromosomal mutations encoding ceftriaxone resistance are already present in gonococcal populations (164); however, additional acquired β-lactam resistance mechanisms could hasten the wide-spread failure of this drug as an effective anti-gonococcal therapy. cN, commensal Neisseria; Nci, N. cinerea; Ngo, N. gonorrhoae; Nla, N. lactamica; Nme, N. meningitidis; Nmu, N. mucosa; Nsu, N. subflava; CAT, chloramphenicol acetyltransferase; CAMP, cationic antimicrobial peptides; SFA, sulfonamides; TMP, trimethoprim; TET, tetracycline; Chl, chloramphenicol; FDH, folate dehydrogenase; PM, polymyxins.

FIG 2

Flowchart of the current protocol for the Gonococcal Isolate Surveillance Project (GISP, run by the CDC). For each clinical collection site, the current GISP (orange) and eGISP (blue) programs sample 25 individuals from genital and 25 from nongenital sites per month, respectively. Currently, all samples are submitted for morphological testing and nucleic acid amplification testing (NAAT) and retained if bacterial isolates are Gram-negative, diplococci, oxidase-positive, have a tan/transparent colony morphology on modified Thayer-Martin media, and NAAT-positive for N. gonorrhoeae (Ngo). ^aSentinel sites not participating in eGISP discard their non-Ngo samples. Retained colonies are then subjected to AMR profiling and WGS. We propose a minor modification to the eGISP program (bold and red), in which all pharyngeal samples collected via oral rinse-and-gargle (138) are conserved if they display features inconsistent with N. gonorrhoeae, yet consistent with other members of the Neisseria genus (Gram−, diplococci or rods, and oxidase+) (bold and red). We also suggest that pharyngeal samples are inoculated onto LBVT.SNR media, which is selective for commensal Neisseria. Subsequent development of a pipeline for complete analysis of each resultant non-N. gonorrhoeae strain (AMR profiling, WGS, etc.) will allow for detailed characterization of Neisseria species and strain diversity, allowing for a comprehensive evaluation of the Neisseria resistome and other comparative genomics analyses of interest.