Genetic Analysis of Drug Resistance in Neisseria gonorrhoeae : Identification and Linkage Relationships of Loci Controlling Drug Resistance

Abstract

The genetic basis of multiple drug resistance of Neisseria gonorrhoeae was investigated by the technique of transformation. Six different genetic loci were characterized by the type and amount of antibiotic resistance they controlled, and also by the degree of linkage to other resistance markers. A streptomycin resistance locus is linked to separate loci determining resistance to tetracycline, chloramphenicol, and erythromycin. A multiple resistance locus was identified. This genetic locus determines resistance to a variety of antibacterial agents. Lastly, a locus determining resistance to the penicillins was found which is unlinked to any other resistance locus.The emergence of gonococci with increased drug resistance has rendered many antibiotics ineffective or barely effective in the treatment of gonorrheae (17,20). With growing frequency, multiply drug-resistant strains are encountered which exhibit increased resistance to penicillin, tetracycline, and streptomycin, as well as to other antimicrobial agents (10,11,15).We have undertaken studies to investigate the genetic basis of multiple drug resistance in naturally occurring strains of Neisseria gonorrhoeae. It was possible to analyze antibiotic resistance using transformation, since the gonococcus has been shown previously to be a competent species (16 Media and cultivation of gonococci. GC agar consisting of GC medium base (Difco), and 1.0% chemically defined supplements 1 and 2 (17) were used for routine culturing of gonococci and as the base medium when antibiotics were incorporated. The GC broth used in these experiments was prepared following the formula of GC medium base with the omission of starch and agar. Gonococci were incubated at 36 C under 5% CO2 in a CO2 incubator.MIC technique. The minimal inhibitory concentration (MIC) of a gonococcal isolate was determined by an agar dilution technique. Gonococci to be tested were streaked twice for isolation on GC agar. Colonies (18 to 24 h) were suspended in GC broth, vortexed vigorously, and diluted to contain approximately 5 x 107 colony-forming units (CFU) per ml, as judged against a borium sulfate turbidity standard. The desired concentrations of antibiotic were incorporated into 20 ml of GC agar in a 100-mm Petri dish.

A Gonococcal Efflux Pump System Enhances Bacterial Survival in a Female Mouse Model of Genital Tract Infection

“…The mtr locus was initially identified in studies on the genetic basis of resistance to multiple antibiotics in N. gonorrhoeae (10,25,39,43). Further definition of the substrate repertoire of this efflux system was facilitated by the subsequent cloning of the Here we present data that support the role of the MtrCDE, but not the FarAB-MtrE, efflux system in survival of N. gonorrhoeae in the lower genital tract of female mice.…”

Taxonomy of the Neisseriae: Deoxyribonucleic Acid Base Composition, Interspecific Transformation, and Deoxyribonucleic Acid Hybridization

“…Transformation procedures were performed in a manner slightly modified from that described by Maier, Zubrzycki, and Coyle (23). Eighteen-hour-old cultures of the recipient organisms were suspended at a concentration of approximately 3 x 10' colony-forming units per ml in GC broth containing 2 mM MgC12 and were exposed to the action of a Vortex mixer to minimize cell clumping.…”

Genetic analysis of spontaneous resistance to ampicillin in Neisseria gonorrhoeae

“…These results indicate that ampicillin resistance in N. gonorrhoeae arises by a series of mutations in multiple genes distinct from those previously mapped (6,8,11,15,16). Mutation frequencies from initial selection were 10-9 to 10-1o, although the first-step resistance was shown to derive from two mutation sites.…”