| Summary: | Nonpathogenic <i>Neisseria</i> transfer mutations encoding antibiotic resistance to their pathogenic relative <i>Neisseria gonorrhoeae</i>. 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 <i>Neisseria</i> (<i>n</i> = 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 (<i>n</i> = 5/26), ceftriaxone (<i>n</i> = 2/26), cefixime (<i>n</i> = 3/26), tetracycline (<i>n</i> = 10/26), azithromycin (<i>n</i> = 11/26), and ciprofloxacin (<i>n</i> = 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 (<i>gyrA</i>) (S91F) and ciprofloxacin, tetracycline resistance protein (<i>tetM</i>) and 30S ribosomal protein S10 (<i>rpsJ</i>) (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 <i>Neisseria</i>, which will ultimately aid in prospective surveillance for novel resistance mechanisms that may be rapidly acquired by <i>N. gonorrhoeae</i>.
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