Trichomonas vaginalis metronidazole resistance is associated with single nucleotide polymorphisms in the nitroreductase genes ntr4Tv and ntr6Tv

Abstract

Metronidazole resistance in the sexually transmitted parasite Trichomonas vaginalis is a problematic public health issue. We have identified single nucleotide polymorphisms (SNPs) in two nitroreductase genes (ntr4Tv and ntr6Tv) associated with resistance. These SNPs were associated with one of two distinct T. vaginalis populations identified by multilocus sequence typing, yet one SNP (ntr6Tv A238T), which results in a premature stop codon, was associated with resistance independent of population structure and may be of diagnostic value.

FIG 1

Metronidazole susceptibility in 197 isolates stratified by ntr_Tv genotype. Intact open reading frames (ORFs) are represented by +, and the presence of stop codons is indicated by −. Horizontal bars represent median values. Isolates with a range of MLC values (e.g., 100 to 200 μg/ml) are plotted using their mean MLC values. Isolates with an MLC value of >400 μg/ml are assigned a value of 800 μg/ml for display purposes. *, P < 0.05; **, P < 0.01; and ***, P < 0.001.

FIG 2

Determination of T. vaginalis population using Structure 2.3 to analyze SNP data from 100 isolates. (a) Mean values ± standard error of the means (SEM) (n = 5) for log probability of assumed population structure. (b) Second-order rate of change in likelihood (ΔK) between the number of assumed populations.

FIG 3

T. vaginalis population structure. Red represents ancestry from population 1, and green represents ancestry from population 2. (a) Total population sorted by Q (ancestry vector) (100 isolates), (b) population sorted by ascending MLC values (100 isolates), and (c) population sorted by ntr_Tv genotype (99 isolates, 1 excluded due to undetermined ntr6_Tv genotype).

FIG 4

Distribution of metronidazole MLC values within 50 population 2 isolates by presence or absence of ntr6_Tv A238T (K80STOP). Horizontal bars represent median values. Isolates with range of MLC values (e.g., 100 to 200 μg/ml) are plotted using their mean MLC values. Isolates with an MLC value of >400 μg/ml are assigned a value of 800 μg/ml for display purposes. P value determined by Mann-Whitney U test.