Rolling the evolutionary dice:
Neisseria
commensals as proxies for elucidating the underpinnings of antibiotic resistance mechanisms and evolution in human pathogens
Authors: Frost Kelly M. , Charron-Smith Sierra L. , Cotsonas Terence C. , Dimartino Daniel C. , Eisenhart Rachel C. , Everingham Eric T. , Holland Elle C. , Imtiaz Kainat , Kornowicz Cory J. , Lenhard Lydia E. , Lynch Liz H. , Moore Nadia P. , Phadke Kavya , Reed Makayla L. , Smith Samantha R. , Ward Liza L. , Wadsworth Crista B. 
Published: January 2024

Species within the genus
Neisseria
are adept at sharing adaptive allelic variation, with commensal species repeatedly transferring resistance to their pathogenic relative
Neisseria gonorrhoeae
. However, resistance in commensals is infrequently characterized, limiting our ability to predict novel and potentially transferable resistance mechanisms that ultimately may become important clinically. Unique evolutionary starting places of each
Neisseria
species will have distinct genomic backgrounds, which may ultimately control the fate of evolving populations in response to selection as epistatic and additive interactions coerce lineages along divergent evolutionary trajectories. Alternatively, similar genetic content present across species due to shared ancestry may constrain existing adaptive solutions. Thus, identifying the paths to resistance across commensals may aid in characterizing the
Neisseria
resistome—or the reservoir of alleles within the genus as well as its depth. Here, we use
in vitro
evolution of four commensal species to investigate the potential and repeatability of resistance evolution to two antimicrobials, the macrolide azithromycin and the β-lactam penicillin. After 20 days of selection, commensals evolved resistance to penicillin and azithromycin in 11/16 and 12/16 cases, respectively. Almost all cases of resistance emergence converged on mutations within ribosomal components or the
mtrRCDE
efflux pump for azithromycin-based selection and
mtrRCDE
,
penA
, and
rpoB
for penicillin selection, thus supporting constrained adaptive solutions despite divergent evolutionary starting points across the genus for these particular drugs. Though drug-selected loci were limited, we do identify novel resistance-imparting mutations. Continuing to explore paths to resistance across different experimental conditions and genomic backgrounds, which could shunt evolution down alternative evolutionary trajectories, will ultimately flesh out the full
Neisseria
resistome.

IMPORTANCE

Neisseria gonorrhoeae
is a global threat to public health due to its rapid acquisition of antibiotic resistance to all first-line treatments. Recent work has documented that alleles acquired from close commensal relatives have played a large role in the emergence of resistance to macrolides and beta-lactams within gonococcal populations. However, commensals have been relatively underexplored for the resistance genotypes they may harbor. This leaves a gap in our understanding of resistance that could be rapidly acquired by the gonococcus through a known highway of horizontal gene exchange. Here, we characterize resistance mechanisms that can emerge in commensal
Neisseria
populations via
in vitro
selection to multiple antimicrobials and begin to define the number of paths to resistance. This study, and other similar works, may ultimately aid both surveillance efforts and clinical diagnostic development by nominating novel and conserved resistance mechanisms that may be at risk of rapid dissemination to pathogen populations.