Browsing by Author "Derendinger, Brigitta"
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- ItemBedaquiline microheteroresistance after cessation of tuberculosis treatment(Massachusetts Medical Society, 2019-05-30) De Vos, Margaretha; Wiggins, Kristin B.; Derendinger, Brigitta; Reuter, Anja; Dolby, Tania; Burns, Scott; Schito, Marco; Engelthaler, David M.; Metcalfe, John; Theron, Grant; Van Rie, Annelies; Posey, James; Warren, Rob; Cox, HelenENGLISH ABSTRACT: Bedaquiline improves survival among persons with multidrug-resistant tuberculosis (MDR-TB).1 We report the case of a 65-year-old South African man who was negative for human immunodeficiency virus and in whom MDR-TB was diagnosed in 2013 (resistant to rifampin and isoniazid; phenotypically susceptible to a fluoroquinolone and amikacin). A baseline radiograph showed changes consistent with bilateral tuberculosis with left apex cavitation. He started standardized treatment that included moxifloxacin, pyrazinamide, kanamycin, ethionamide, isoniazid, and terizidone. After initial sputum culture conversion (at month 3) and clinical improvement, the patient again became culture-positive, and bilateral cavitation developed. After detection of phenotypic resistance to fluoroquinolones (at month 6), his treatment was revised (at month 8) to include high-dose isoniazid, ethambutol, pyrazinamide, terizidone, linezolid, paraaminosalicylic acid, and kanamycin (Figure 1 and the Supplementary Appendix, available with the full text of this letter at NEJM.org). Bedaquiline was added 22 days later and was administered for 6 months.2 The patient remained culture-positive (treatment failure), and treatment was stopped 15 months after revision of the regimen. The patient died 7 months later.
- ItemExtract from used Xpert MTB/ RIF Ultra cartridges is useful for accurate second-line drug-resistant tuberculosis diagnosis with minimal rpoB-amplicon cross-contamination risk(Nature Research (part of Springer Nature), 2020) Venter, Rouxjeane; Minnies, Stephanie; Derendinger, Brigitta; Tshivhula, Happy; De Vos, Margaretha; Dolby, Tania; Ruiters, Ashley; Warren, Robin M.; Theron, GrantXpert MTB/RIF Ultra (Ultra) detects Mycobacterium tuberculosis and rifampicin resistance. Follow-on drug susceptibility testing (DST) requires additional sputum. Extract from the diamond-shaped chamber of the cartridge (dCE) of Ultra’s predecessor, Xpert MTB/RIF (Xpert), is useful for MTBDRsl-based DST but this is unexplored with Ultra. Furthermore, whether CE from non-diamond compartments is useful, the performance of FluoroType MTBDR (FT) on CE, and rpoB cross-contamination risk associated with the extraction procedure are unknown. We tested MTBDRsl, MTBDRplus, and FT on CEs from chambers from cartridges (Ultra, Xpert) tested on bacilli dilution series. MTBDRsl on Ultra dCE on TB-positive sputa (n = 40) was also evaluated and, separately, rpoB amplicon cross-contamination risk . MTBDRsl on Ultra dCE from dilutions ≥103 CFU/ml (CTmin <25, >“low semi-quantitation”) detected fluoroquinolone (FQ) and second-line injectable (SLID) susceptibility and resistance correctly (some SLIDs-indeterminate). At the same threshold (at which ~85% of Ultra-positives in our setting would be eligible), 35/35 (100%) FQ and 34/35 (97%) SLID results from Ultra dCE were concordant with sputa results. Tests on other chambers were unfeasible. No tubes open during 20 batched extractions had FT-detected rpoB cross-contamination. False-positive Ultra rpoB results was observed when dCE dilutions ≤10−3 were re-tested. MTBDRsl on Ultra dCE is concordant with isolate results. rpoB amplicon cross-contamination is unlikely. These data mitigate additional specimen collection for second-line DST and cross-contamination concerns.
- ItemMycobacterial genomic DNA from used Xpert MTB/RIF cartridges can be utilised for accurate second-line genotypic drug susceptibility testing and spoligotyping(Nature, 2017) Venter, Rouxjeane; Derendinger, Brigitta; De Vos, Margaretha; Pillay, Samantha; Dolby, Tanya; Simpson, John; Kitchin, Natasha; Ruiters, Ashley; Van Helden, Paul D.; Warren, Robin M.; Theron, GrantXpert MTB/RIF (Xpert) is a widely-used test for tuberculosis (TB) and rifampicin-resistance. Second-line drug susceptibility testing (DST), which is recommended by policymakers, typically requires additional specimen collection that delays effective treatment initiation. We examined whether cartridge extract (CE) from used Xpert TB-positive cartridges was, without downstream DNA extraction or purification, suitable for both genotypic DST (MTBDRplus, MTBDRsl), which may permit patients to rapidly receive a XDR-TB diagnosis from a single specimen, and spoligotyping, which could facilitate routine genotyping. To determine the limit-of-detection and diagnostic accuracy, CEs from dilution series of drug-susceptible and -resistant bacilli were tested (MTBDRplus, MTBDRsl). Xpert TB-positive patient sputa CEs (n = 85) were tested (56 Xpert-rifampicin-susceptible, MTBDRplus and MTBDRsl; 29 Xpert-rifampicin-resistant, MTBDRsl). Spoligotyping was done on CEs from dilution series and patient sputa (n = 10). MTBDRplus had high non-valid result rates. MTBDRsl on CEs from dilutions ≥103CFU/ml (CT ≤ 24, >“low” Xpert semiquantitation category) was accurate, had low indeterminate rates and, on CE from sputa, highly concordant with MTBDRsl isolate results. CE spoligotyping results from dilutions ≥103CFU/ml and sputa were correct. MTBDRsl and spoligotyping on CE are thus highly feasible. These findings reduce the need for additional specimen collection and culture, for which capacity is limited in high-burden countries, and have implications for diagnostic laboratories and TB molecular epidemiology.
- ItemMycobacterial genomic DNA from used Xpert MTB/RIF cartridges can be utilised for accurate secondline genotypic drug susceptibility testing and spoligotyping(Springer Nature, 2017-11-01) Venter, Rouxjeane; Derendinger, Brigitta; De Vos, Margaretha; Pillay, Samantha; Dolby, Tanya; Simpson, John; Kitchin, Natasha; Ruiters, Ashley; Van Helden, Paul D.; Warren, Robin M.; Theron, GrantENGLISH ABSTRACT: Xpert MTB/RIF (Xpert) is a widely-used test for tuberculosis (TB) and rifampicin-resistance. Second-line drug susceptibility testing (DST), which is recommended by policymakers, typically requires additional specimen collection that delays effective treatment initiation. We examined whether cartridge extract (CE) from used Xpert TB-positive cartridges was, without downstream DNA extraction or purification, suitable for both genotypic DST (MTBDRplus, MTBDRsl), which may permit patients to rapidly receive a XDR-TB diagnosis from a single specimen, and spoligotyping, which could facilitate routine genotyping. To determine the limit-of-detection and diagnostic accuracy, CEs from dilution series of drug-susceptible and -resistant bacilli were tested (MTBDRplus, MTBDRsl). Xpert TB-positive patient sputa CEs (n = 85) were tested (56 Xpert-rifampicin-susceptible, MTBDRplus and MTBDRsl; 29 Xpert-rifampicin-resistant, MTBDRsl). Spoligotyping was done on CEs from dilution series and patient sputa (n = 10). MTBDRplus had high non-valid result rates. MTBDRsl on CEs from dilutions ≥103CFU/ml (CT ≤ 24, >“low” Xpert semiquantitation category) was accurate, had low indeterminate rates and, on CE from sputa, highly concordant with MTBDRsl isolate results. CE spoligotyping results from dilutions ≥103CFU/ml and sputa were correct. MTBDRsl and spoligotyping on CE are thus highly feasible. These findings reduce the need for additional specimen collection and culture, for which capacity is limited in high-burden countries, and have implications for diagnostic laboratories and TB molecular epidemiology.
- ItemNew and improved methods for the diagnosis of susceptibility to tuberculosis drugs(Stellenbosch : Stellenbosch University, 2023-01) Derendinger, Brigitta; Theron, Grant; De Vos, Margaretha; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Molecular Biology and Human Genetics.ENGLISH ABSTRACT: Drug-resistant tuberculosis (DR-TB) is a global threat. Diagnosing patients with DR-TB and initiating them onto appropriate treatment, in the shortest possible time, is of utmost importance. The optimisation of existing rapid molecular assays and the implementation of drug susceptibility testing (DST) for new drugs to monitor patients on treatment is crucial in curbing transmission. Firstly (chapter 2), showed that Mycobacterium tuberculosis (Mtb) DNA recoverable from used Xpert MTB/RIF (Xpert) cartridges ‒ cartridge extract (CE) – that would otherwise be discarded, can be used for downstream second-line molecular DST. No additional DNA extraction or sample purification was needed. We defined a threshold of Xpert semiquantitative category “low” to ensure that no MTBDRsl assays are wasted on CE likely to give invalid results. This alleviated the need for collection of a second sputum specimen, thereby reducing diagnostic delays, and enabling patients to be placed on treatment sooner. This approach is being developed into a cartridge extraction device and will be evaluated by TB programmes. Secondly and thirdly (chapter 3 and 4), MTBDRplus and MTBDRsl focussed on WHOendorsed rapid molecular assays that have reported suboptimal sensitivities and high indeterminate rates especially in smear-negative specimens. We hypothesised that ramp rate (speed of temperature change between PCR cycles) could impact assay performance. We showed that correcting thermocycler ramp rate (manufacturer-recommended ramp rate ≤2.2°C/s) likely improved the yield of rapid diagnoses for first-and second-line DST, done with MTBDRplus and MTBDRsl respectively, especially in paucibacillary specimens. Our survey showed that suboptimal ramp rate is a common problem but is easily fixable. This manuscript informed WHO course training material (https://openwho.org/courses/multi-drug-resistant-tb). Finally (chapter 5), bedaquiline (BDQ), a lifesaving TB drug, is undergoing rapid scale-up but largely in the absence of DST. In a group of programmatic patients still culture-positive after ≥4 months of BDQ-based treatment, more than half had isolates that gained BDQ resistance (mostly acquisition, some transmission). Several Rv0678 and pepQ variants were associated with phenotypic resistance, many previously undescribed. Patients with baseline fluoroquinolone-resistance, clofazimine exposure, and ≤4 effective drugs were more likely to be BDQ-resistant. This thesis has resulted in four first author manuscripts (three published, one submitted). Additionally, two 2nd author manuscripts and seven manuscripts as a middle co-author. These nine total are briefly discussed in chapter 6 and can be found in the appendices (and included as ancillary publications). The candidate presented three times at international and twice at national peer-reviewed conferences. In summary, this work shows that second-line DR time-to-treatment initiation could be reduced by doing second-line DST on CE from used cartridges. The number of smear-negative patients in whom DST is possible will improve substantially after ramp rate correction for MTBDRplus and MTBDRsl. Finally, we show the existence of a potentially infectious pool of BDQ resistant strains created under programmatic conditions, as well as the challenges and risks associated with starting patients with complex TB-treatment histories on a regimen containing a novel drug without routinely available DST. Our findings also inform on how and in whom new TB drugs are prioritised for use.