Genetic indicators of drug resistance in the highly repetitive genomes of<i>Trichomonas vaginalis</i>and other trichomonads

Bradić, Martina; Warring, SD; Tooley, GE; Scheid, Paul; Secor, W. Evan; Land, KM; Huang, Po-Jung; Chen, Tai‐Di; Lee, C; Tang, Petrus; Sullivan, SA; Carlton, JM

doi:10.1101/076729

Cited by 1 publication

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References 68 publications

(128 reference statements)

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“…Of course it must be noted that predicted functions are just that, and that the true nature of the many hypothetical proteins in Giardia awaits experimental determination. Only two reports to date have investigated transcriptional correlates of Mtz resistance using genome-wide techniques (Müller et al, 2008 ; Bradic et al, 2016 ). In this study, in addition to sensitive genetically-controlled analyses of transcriptional changes between Mtz-resistant isotypes of G. duodenalis , we compared our results to these independently generated data sets to infer key transcriptional themes in Mtz resistant parasites more broadly.…”

Section: Discussionmentioning

confidence: 99%

“…Selected SNPs were visually confirmed using Integrative Genomics Viewer (Robinson et al, 2011 ), and amino acid substitutions (products of non-synonymous SNPs), were predicted using getORF (Rice et al, 2000 ) and custom scripts. For comparative transcriptomics between species, published RNA transcript count data for Mtz-susceptible and –resistant T. vaginalis (Bradic et al, 2016 ) were submitted to edgeR in a two-sample comparison of transcript abundance. Count data for resistant and susceptible G. duodenalis lines generated in this study were compared using the same design, and orthologous genes (defined in OrthoMCL-DB; Chen, 2006 ) that were differentially transcribed in the same direction in both species, were retained for further analysis.…”

Section: Methodsmentioning

confidence: 99%

“…Further, as we have argued previously (Ansell et al, 2015b ), enzymes that produce and consume the substrates and cofactors of oxidoreductase enzymes may thereby modulate their function, which potentially implicates a host of metabolic enzymes in Mtz metabolism and resistance. Recent whole genome and transcriptome sequencing has refined and expanded the complement of genes associated with Mtz resistance in H. pylori (Binh et al, 2015 ) and T. vaginalis (Bradic et al, 2016 ), among which hitherto under-characterized flavin reductases, ribosomal proteins, and canonical nitroreductases feature prominently.…”

Section: Introductionmentioning

confidence: 99%

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Transcriptomics Indicates Active and Passive Metronidazole Resistance Mechanisms in Three Seminal Giardia Lines

et al. 2017

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Giardia duodenalis is an intestinal parasite that causes 200–300 million episodes of diarrhoea annually. Metronidazole (Mtz) is a front-line anti-giardial, but treatment failure is common and clinical resistance has been demonstrated. Mtz is thought to be activated within the parasite by oxidoreductase enzymes, and to kill by causing oxidative damage. In G. duodenalis, Mtz resistance involves active and passive mechanisms. Relatively low activity of iron-sulfur binding proteins, namely pyruvate:ferredoxin oxidoreductase (PFOR), ferredoxins, and nitroreductase-1, enable resistant cells to passively avoid Mtz activation. Additionally, low expression of oxygen-detoxification enzymes can allow passive (non-enzymatic) Mtz detoxification via futile redox cycling. In contrast, active resistance mechanisms include complete enzymatic detoxification of the pro-drug by nitroreductase-2 and enhanced repair of oxidized biomolecules via thioredoxin-dependent antioxidant enzymes. Molecular resistance mechanisms may be largely founded on reversible transcriptional changes, as some resistant lines revert to drug sensitivity during drug-free culture in vitro, or passage through the life cycle. To comprehensively characterize these changes, we undertook strand-specific RNA sequencing of three laboratory-derived Mtz-resistant lines, 106-2ID10, 713-M3, and WB-M3, and compared transcription relative to their susceptible parents. Common up-regulated genes encoded variant-specific surface proteins (VSPs), a high cysteine membrane protein, calcium and zinc channels, a Mad-2 cell cycle regulator and a putative fatty acid α-oxidase. Down-regulated genes included nitroreductase-1, putative chromate and quinone reductases, and numerous genes that act proximal to PFOR. Transcriptional changes in 106-2ID10 diverged from those in 713-r and WB-r (r ≤ 0.2), which were more similar to each other (r = 0.47). In 106-2ID10, a nonsense mutation in nitroreductase-1 transcripts could enhance passive resistance whereas increased transcription of nitroreductase-2, and a MATE transmembrane pump system, suggest active drug detoxification and efflux, respectively. By contrast, transcriptional changes in 713-M3 and WB-M3 indicated a higher oxidative stress load, attributed to Mtz- and oxygen-derived radicals, respectively. Quantitative comparisons of orthologous gene transcription between Mtz-resistant G. duodenalis and Trichomonas vaginalis, a closely related parasite, revealed changes in transcripts encoding peroxidases, heat shock proteins, and FMN-binding oxidoreductases, as prominent correlates of resistance. This work provides deep insight into Mtz-resistant G. duodenalis, and illuminates resistance-associated features across parasitic species.

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Section: Discussionmentioning

confidence: 99%