Full Text Available
Note: Clicking the button above will open the full text document at the original institutional repository in a new window.
Resistance to the novel nitroimidazole drug class in Mycobacterium tuberculosis
Astrid Gail, Paulse. 2023. Resistance to the novel nitroimidazole drug class in Mycobacterium tuberculosis. Unpublished master's dissertation. Stellenbosch : Stellenbosch University [online]. Available at: https://scholar.sun.ac.za/items/52eb99f9-0718-432e-85d9-9d723fa425f0
Saved in:
| Main Author: | |
|---|---|
| Other Authors: | |
| Format: | Thesis |
| Language: | English |
| Published: |
Stellenbosch : Stellenbosch University
2023
|
| Subjects: | |
| Tags: |
No Tags, Be the first to tag this record!
|
| _version_ | 1867613881517473792 |
|---|---|
| access_status_str | Open Access |
| author | Astrid Gail, Paulse |
| author2 | Grobbelaar, Melanie |
| author_browse | Astrid Gail, Paulse Grobbelaar, Melanie |
| author_facet | Grobbelaar, Melanie Astrid Gail, Paulse |
| author_sort | Astrid Gail, Paulse |
| collection | Thesis |
| dc_rights_str_mv | Stellenbosch University |
| description | Astrid Gail, Paulse. 2023. Resistance to the novel nitroimidazole drug class in Mycobacterium tuberculosis. Unpublished master's dissertation. Stellenbosch : Stellenbosch University [online]. Available at: https://scholar.sun.ac.za/items/52eb99f9-0718-432e-85d9-9d723fa425f0 |
| format | Thesis |
| id | oai:scholar.sun.ac.za:10019.1/129406 |
| institution | Stellenbosch University (South Africa) |
| language | English |
| last_indexed | 2026-06-10T12:43:11.727Z |
| license_str | Other — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from SUNScholar — Stellenbosch University Repository |
| publishDate | 2023 |
| publishDateRange | 2023 |
| publishDateSort | 2023 |
| publisher | Stellenbosch : Stellenbosch University |
| publisherStr | Stellenbosch : Stellenbosch University |
| record_format | dspace |
| source_str | SUNScholar — Stellenbosch University Repository |
| spelling | oai:scholar.sun.ac.za:10019.1/129406 Resistance to the novel nitroimidazole drug class in Mycobacterium tuberculosis Astrid Gail, Paulse Grobbelaar, Melanie Warren, Robin Ismail, Nabila Dippenaar, Anzaan Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Molecular Biology and Human Genetics. Multidrug-resistant tuberculosis Mycobacterium tuberculosis -- Treatment Microbial mutation Anti-infective agents UCTD Astrid Gail, Paulse. 2023. Resistance to the novel nitroimidazole drug class in Mycobacterium tuberculosis. Unpublished master's dissertation. Stellenbosch : Stellenbosch University [online]. Available at: https://scholar.sun.ac.za/items/52eb99f9-0718-432e-85d9-9d723fa425f0 Thesis (MSc)--Stellenbosch University, 2023. ENGLISH ABSTRACT: Given the low treatment success of drug-resistant tuberculosis (DR-TB) with a limited number of available drugs, the development of new tuberculosis (TB) drugs is expected to enhance the treatment success rates. Various concerns have been raised regarding the effectiveness of the novel drug pretomanid (PMD) since its implementation in the new short-oral treatment regimen, BPaL/M, for the treatment of DR-TB in South Africa (SA). PMD belongs to the same class of drugs (nitroimidazoles) as delamanid (DLM) and shares a similar mechanism of action, target genes and molecular structure. These similarities underpin potential cross-resistance between the two drugs, which has been previously reported. Furthermore, mutations within the six target genes (ddn, fgd1, fbiA, fbiB, fbiC and fbiD) involved with the activation of DLM and PMD have been reported to confer resistance to the two drugs. However, limited information is available on the genotypic and phenotypic correlation of mutations within these target genes. Additionally, the specific impact of mutations linked to DLM and PMD in Mycobacterium tuberculosis drug-resistant strains is poorly characterized. In this study, we determined the prevalence of resistance-associated variants (RAV) to PMD and DLM in SA using the whole-genome sequencing (WGS) databank created by the TB Genomics group. We identified n=40 M. tuberculosis clinical isolates with mutations within the six target genes, of which 23 were novel mutations. We assessed the phenotypic effect of these mutations on PMD and DLM susceptibility using phenotypic drug susceptibility testing (pDST) and minimum inhibitory concentration (MIC) assays. Furthermore, two in vitro mutant generation techniques, Luria-Delbrück (spontaneous approach) and the serial passaging assay (induction approach) were used to generate mutants to identify cross-resistance between the two drugs. Of the 23 novel mutations, only one mutation within the ddn (p.Gly70Val) gene conferred resistance to PMD (MIC ≤ 4 μg/mL). Four out of the forty clinical isolates, belonging to Lineage 1 had an increase in PMD MIC levels in comparison to Lineage 2 and 4, while the DLM MIC was not affected by strain genetic background. In addition to the effect of Lineage 1, the mutations within the ddn (p.Gly70Val) and fbiA (p.Val160Leu). In vitro and in vivo assessments of the bactericidal activity of both drugs showed that DLM exhibited lower MIC values and higher mycobacterial load reduction potential than PMD. Moreover, high MICs (DLM ≥4 μg/mL, PMD ≥8 μg/mL) were observed in vitro for the two mutant generation assays. Spontaneously generated PMD mutants that contained RAVs within ddn (p.Arg151Lys, p.Arg90Arg) and fbiA (p.Lys421Gln) were all associated with high MIC values and were found to be resistant to PMD or DLM, of which fbiA (p.Lys421Gln) and ddn (p.Arg151Lys) were found to cause cross-resistance to PMD and DLM. Furthermore, the induced method produced mutants with high DLM MIC values, but causal mutations were not found within the three amplified target genes (fbiA, ddn and fgd1). In conclusion, our data shows that PMD and DLM resistance-related mutations are diverse and distributed across the entire region of each gene targeted by these drugs. The findings of this study are of critical importance for the development of comprehensive molecular diagnostic tools for the prediction of PMD and DLM resistance. The in vitro mutation generation assays present an excellent approach to studying the acquisition of RAV and cross-resistance mechanisms to novel antimicrobial drugs for TB treatment. The methods used in this study will further benefit in monitoring the emergence of resistance to PMD and DLM, especially as these drugs are rolled out in South Africa. AFRIKAANSE OPSOMMING: Gegewe die lae behandelingsukses van middelweerstandige tuberkulose (DR-TB) met ’n beperkte aantal beskikbare middels, word verwag dat die ontwikkeling van nuwe tuberkulose (TB) middels die sukseskoers sal verhoog. Kommer oor die veiligheid en doeltreffendheid van die nuwe nitroimidasool-middel, pretomanied (PMD), is uitgespreek sedert die implementering van die middel in die nuwe kort-orale behandelingsregime, BPaL/M, vir die behandeling van DR-TB in Suid-Afrika (SA). PMD behoort aan dieselfde klas geneesmiddels (nitroimidazole) as delamanied (DLM) en het ’n soortgelyke meganisme van werking, teikengene en struktuur. Hierdie ooreenkomste ondersteun potensiële kruisweerstandigheid tussen die twee middels wat voorheen gerapporteer is. Verder is daar berig dat mutasies binne die ses teikengene (ddn, fgd1, fbiA, fbiB, fbiC en fbiD) betrokke by die aktivering van DLM en PMD weerstand teen die twee middels verleen. Daarbenewens is die spesifieke impak van mutasies gekoppel aan DLM en PMD in *Mycobacterium tuberculosis*-middelweerstandige stamme nie goed gekarakteriseer nie.,In hierdie studie het ons die voorkoms van weerstand-geassosieerde variante (RAV) vir PMD en DLM binne SA vasgestel deur gebruik te maak van die heelgenoom (WGS) databank wat deur die TB Genomics-groep geskep is. Ons het n=40 *M. tuberculosis* kliniese isolate geïdentifiseer wat mutasies binne die ses teikengene gehad het, waarvan 23 nuwe mutasies is. Om die fenotipiese effek van hierdie mutasies in *M. tuberculosis* op PMD en DLM vatbaarheid te evalueer, is fenotipiese geneesmiddelvatbaarheidstoetse (pDST) en minimum inhiberende konsentrasie (MIC) toetse uitgevoer om hul weerstandsprofiele te karakteriseer. Verder is twee *in vitro* mutantgenerasie-tegnieke, Luria-Delbrück en die inokuleringsreeks-toets (induksiebenadering), gebruik om mutante te skep om die mate van kruisweerstandigheid tussen die twee middels te bepaal. Slegs een van die 23 nuwe mutasies het weerstandigheid verleen teen PMD (MIC ≤ 4 µg/mL), maar hierdie observasie is gekompliseer deur die teenwoordigheid van ’n tweede mutasie wat nie alleen weerstandigheid in ’n ander isolaat veroorsaak het nie. Vier uit die veertig kliniese isolate, wat aan Lineage 1 behoort, het ’n toename in PMD MIC-vlakke gehad in vergelyking met Lineage 2 en 4, terwyl die DLM MIC nie deur stamgenetiese agtergrond beïnvloed is nie. Benewens die effek van Lineage 1 is die mutasies binne die *ddn* (p.Gly70Val) en *fbiA* (p.Val160Leu) ondersoek. *In vitro* en *in vivo* assesserings van die bakteriosidiese aktiwiteit van beide middels toon dat DLM laer MIC-waardes en hoër mikobakteriese ladingsverminderingvermoë het in vergelyking met PMD. Verder is hoë MIC’s *in vitro* waargeneem vir die twee mutantgenerasie-toetse (DLM ≥4 μg/mL, PMD ≥8 μg/mL). Spontane PMD-mutante wat RAV’s bevat het in *ddn* (p.Arg151Lys, p.Arg90Arg) en *fbiA* (p.Lys421Gln), is almal geassosieer met hoë MIC-waardes en daar is gevind dat dit weerstandbiedend is teen PMD of DLM. Verder het geïnduseerde mutante hoë MIC-waardes vir DLM tot gevolg gehad, maar is nie gevind binne die drie versterkte teikengene nie (*fbiA, ddn* en *fgd1*). Ten slotte toon ons data dat beide PMD- en DLM-weerstandsverwante mutasies uiteenlopend is en versprei word oor die hele streek van elke geen wat deur hierdie middels geteiken word. Die bevindinge van hierdie studie is van kritieke belang vir die ontwikkeling van omvattende molekulêre diagnostiese toetse vir die bepaling van PMD- en DLM-weerstand. Die *in vitro* mutasiegenerasie-toetse bied ’n goeie benadering om verworwe kruisweerstandigheidsmeganismes sowel as RAV’s vir nuwe antimikrobiese middels vir TB-behandeling te bestudeer. Verder sal dit die monitering van toekomstige implementering van nuwe middels baat. Dit is egter nodig om meganismes van PMD- of DLM-weerstand in *M. tuberculosis* verder te ondersoek. Masters 2023-11-16T09:57:43Z 2024-02-20T10:28:06Z 2023-11-16T09:57:43Z 2024-02-20T10:28:06Z 2023-08 Thesis https://scholar.sun.ac.za/handle/10019.1/129406 en Stellenbosch University 122 pages : illustrations application/pdf Stellenbosch : Stellenbosch University |
| spellingShingle | Multidrug-resistant tuberculosis Mycobacterium tuberculosis -- Treatment Microbial mutation Anti-infective agents UCTD Astrid Gail, Paulse Resistance to the novel nitroimidazole drug class in Mycobacterium tuberculosis |
| title | Resistance to the novel nitroimidazole drug class in Mycobacterium tuberculosis |
| title_full | Resistance to the novel nitroimidazole drug class in Mycobacterium tuberculosis |
| title_fullStr | Resistance to the novel nitroimidazole drug class in Mycobacterium tuberculosis |
| title_full_unstemmed | Resistance to the novel nitroimidazole drug class in Mycobacterium tuberculosis |
| title_short | Resistance to the novel nitroimidazole drug class in Mycobacterium tuberculosis |
| title_sort | resistance to the novel nitroimidazole drug class in mycobacterium tuberculosis |
| topic | Multidrug-resistant tuberculosis Mycobacterium tuberculosis -- Treatment Microbial mutation Anti-infective agents UCTD |
| url | https://scholar.sun.ac.za/handle/10019.1/129406 |
| work_keys_str_mv | AT astridgailpaulse resistancetothenovelnitroimidazoledrugclassinmycobacteriumtuberculosis |