Exploration of the Neisseria Resistome Reveals Resistance Mechanisms in Commensals That May Be Acquired by N. gonorrhoeae through Horizontal Gene Transfer

Abstract

Nonpathogenic Neisseria transfer mutations encoding antibiotic resistance to their pathogenic relative Neisseria gonorrhoeae. However, the resistance genotypes and subsequent phenotypes of nonpathogens within the genus have been described infrequently. Here, we characterize the minimum inhibitory concentrations (MICs) of a panel of Neisseria (n = 26)-including several commensal species-to a suite of diverse antibiotics. We furthermore use whole genome sequencing and the Comprehensive Antibiotic Resistance Database Resistance Gene Identifier (RGI) platform to predict putative resistance-encoding mutations. Resistant isolates to all tested antimicrobials including penicillin (n = 5/26), ceftriaxone (n = 2/26), cefixime (n = 3/26), tetracycline (n = 10/26), azithromycin (n = 11/26), and ciprofloxacin (n = 4/26) were found. In total, 63 distinct mutations were predicted by RGI to be involved in resistance. The presence of several mutations had clear associations with increased MIC such as DNA gyrase subunit A (gyrA) (S91F) and ciprofloxacin, tetracycline resistance protein (tetM) and 30S ribosomal protein S10 (rpsJ) (V57M) and tetracycline, and TEM-type β-lactamases and penicillin. However, mutations with strong associations to macrolide and cephalosporin resistance were not conclusive. This work serves as an initial exploration into the resistance-encoding mutations harbored by nonpathogenic Neisseria, which will ultimately aid in prospective surveillance for novel resistance mechanisms that may be rapidly acquired by N. gonorrhoeae.

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

Figure 1

Maximum likelihood phylogenetic tree of 38 Neisseria isolates based on the 16S rRNA gene: species are coded by unique colors. The scale bar represents 0.08 substitutions per nucleotide site. Antibiotic Resistance (AR) Bank # 0961, one of the Moraxella catarrhalis in the Neisseria species MALDI-TOF verification panel, was used as an outgroup.

Figure 2

Heatmap showing the distribution of Comprehensive Antibiotic Resistance Database (CARD) Resistance Gene Identifier (RGI) predicted resistance-encoding mutations for all sequenced isolates (n = 24). Upper panel: Neisseria species’ susceptibility phenotypes as quantified by MIC. Black denotes loss of susceptibility (CLSI standards), grey denotes one dilution below the reduced susceptibility breakpoint, and white denotes susceptibility to the antibiotics listed on the right (PEN, penicillin; CRO, ceftriaxone; CFX, cefixime; TET, tetracycline; AZI, azithromycin; and CIP, ciprofloxacin). Lower panel: presence of each recovered mutation is indicated by blue fill. CARD-predicted resistance phenotypes for sets of mutations are denoted for beta-lactam resistance (red shading), beta-lactam and tetracycline resistance (purple), tetracycline (blue), macrolides (orange), fluoroquinolones (green), and resistance to other antibiotics (grey).