Uganda's experience in Ebola virus disease outbreak preparedness, 2018-2019.

BACKGROUND: Since the declaration of the 10th Ebola Virus Disease (EVD) outbreak in DRC on 1st Aug 2018, several neighboring countries have been developing and implementing preparedness efforts to prevent EVD cross-border transmission to enable timely detection, investigation, and response in the event of a confirmed EVD outbreak in the country. We describe Uganda's experience in EVD preparedness. RESULTS: On 4 August 2018, the Uganda Ministry of Health (MoH) activated the Public Health Emergency Operations Centre (PHEOC) and the National Task Force (NTF) for public health emergencies to plan, guide, and coordinate EVD preparedness in the country. The NTF selected an Incident Management Team (IMT), constituting a National Rapid Response Team (NRRT) that supported activation of the District Task Forces (DTFs) and District Rapid Response Teams (DRRTs) that jointly assessed levels of preparedness in 30 designated high-risk districts representing category 1 (20 districts) and category 2 (10 districts). The MoH, with technical guidance from the World Health Organisation (WHO), led EVD preparedness activities and worked together with other ministries and partner organisations to enhance community-based surveillance systems, develop and disseminate risk communication messages, engage communities, reinforce EVD screening and infection prevention measures at Points of Entry (PoEs) and in high-risk health facilities, construct and equip EVD isolation and treatment units, and establish coordination and procurement mechanisms. CONCLUSION: As of 31 May 2019, there was no confirmed case of EVD as Uganda has continued to make significant and verifiable progress in EVD preparedness. There is a need to sustain these efforts, not only in EVD preparedness but also across the entire spectrum of a multi-hazard framework. These efforts strengthen country capacity and compel the country to avail resources for preparedness and management of incidents at the source while effectively cutting costs of using a "fire-fighting" approach during public health emergencies

Whole genome sequencing to complement tuberculosis drug resistance surveys in Uganda

YesUnderstanding the circulating Mycobacterium tuberculosis resistance mutations is vital for better TB control strategies, especially to inform a new MDR-TB treatment programme. We complemented the phenotypic drug susceptibility testing (DST) based drug resistance surveys (DRSs) conducted in Uganda between 2008 and 2011 with Whole Genome Sequencing (WGS) of 90 Mycobacterium tuberculosis isolates phenotypically resistant to rifampicin and/or isoniazid to better understand the extent of drug resistance. A total of 31 (34.4 %) patients had MDR-TB, 5 (5.6 %) mono-rifampicin resistance and 54 (60.0 %) mono-isoniazid resistance by phenotypic DST. Pyrazinamide resistance mutations were identified in 32.3% of the MDR-TB patients. Resistance to injectable agents was detected in 4/90 (4.4%), and none to fluoroquinolones or novel drugs. Compensatory mutations in rpoC were identified in two patients. The sensitivity and specificity of drug resistance mutations compared to phenotypic DST were for rpoB 88.6% and 98.1%, katG 60.0% and 100%, fabG1 16.5% and 100%, katG and/or fabG1 71.8% and 100%, embCAB 63.0% and 82.5%, rrs 11.4% and 100%, rpsL 20.5% and 95.7% and rrs and/or rpsL 31.8% and 95.7%. Phylogenetic analysis showed dispersed MDR-TB isolate, with only one cluster of three Beijing family from South West Uganda. Among tuberculosis patients in Uganda, resistance beyond first-line drugs as well as compensatory mutations remain low, and MDR-TB isolates did not arise from a dominant clone. Our findings show the potential use of sequencing for complementing DRSs or surveillance in this setting, with good specificity compared to phenotypic DST. The reported high confidence mutations can be included in molecular assays, and population-based studies can track transmission of MDR-TB including the Beijing family strains in the South West of the country.Erasmus Mundus Joint Doctorate Program of the European Union through a training grant to WS and the European Research Council-INTERRUPTB starting grant (nr.311725) to BdJ

The 2021 WHO catalogue of Mycobacterium tuberculosis complex mutations associated with drug resistance: a genotypic analysis.

Background: Molecular diagnostics are considered the most promising route to achievement of rapid, universal drug susceptibility testing for Mycobacterium tuberculosis complex (MTBC). We aimed to generate a WHO-endorsed catalogue of mutations to serve as a global standard for interpreting molecular information for drug resistance prediction. Methods: In this systematic analysis, we used a candidate gene approach to identify mutations associated with resistance or consistent with susceptibility for 13 WHO-endorsed antituberculosis drugs. We collected existing worldwide MTBC whole-genome sequencing data and phenotypic data from academic groups and consortia, reference laboratories, public health organisations, and published literature. We categorised phenotypes as follows: methods and critical concentrations currently endorsed by WHO (category 1); critical concentrations previously endorsed by WHO for those methods (category 2); methods or critical concentrations not currently endorsed by WHO (category 3). For each mutation, we used a contingency table of binary phenotypes and presence or absence of the mutation to compute positive predictive value, and we used Fisher's exact tests to generate odds ratios and Benjamini-Hochberg corrected p values. Mutations were graded as associated with resistance if present in at least five isolates, if the odds ratio was more than 1 with a statistically significant corrected p value, and if the lower bound of the 95% CI on the positive predictive value for phenotypic resistance was greater than 25%. A series of expert rules were applied for final confidence grading of each mutation. Findings: We analysed 41 137 MTBC isolates with phenotypic and whole-genome sequencing data from 45 countries. 38 215 MTBC isolates passed quality control steps and were included in the final analysis. 15 667 associations were computed for 13 211 unique mutations linked to one or more drugs. 1149 (7·3%) of 15 667 mutations were classified as associated with phenotypic resistance and 107 (0·7%) were deemed consistent with susceptibility. For rifampicin, isoniazid, ethambutol, fluoroquinolones, and streptomycin, the mutations' pooled sensitivity was more than 80%. Specificity was over 95% for all drugs except ethionamide (91·4%), moxifloxacin (91·6%) and ethambutol (93·3%). Only two resistance mutations were identified for bedaquiline, delamanid, clofazimine, and linezolid as prevalence of phenotypic resistance was low for these drugs. Interpretation: We present the first WHO-endorsed catalogue of molecular targets for MTBC drug susceptibility testing, which is intended to provide a global standard for resistance interpretation. The existence of this catalogue should encourage the implementation of molecular diagnostics by national tuberculosis programmes. Funding: Unitaid, Wellcome Trust, UK Medical Research Council, and Bill and Melinda Gates Foundation

Implementation of GeneXpert MTB/Rif proficiency testing program: A Case of the Uganda national tuberculosis reference laboratory/supranational reference laboratory.

BackgroundFollowing the WHO's endorsement of GeneXpert MTB/RIF assay for tuberculosis diagnosis in 2010, Uganda's ministry of health introduced the assay in its laboratory network in 2012. However, assessing the quality of the result produced from this technique is one of its major implementation challenges. To bridge this gap, the National tuberculosis reference laboratory (NTRL) introduced the GeneXpert MTB/RIF proficiency testing (PT) Scheme in 2015.MethodsA descriptive cross-sectional study on the GeneXpert PT scheme in Uganda was conducted between 2015 and 2018. Sets of panels each comprising four 1ml cryovial liquid samples were sent out to enrolled participants at preset testing periods. The laboratories' testing accuracies were assessed by comparing their reported results to the expected and participants' consensus results. Percentage scores were assigned and feedback reports were sent back to laboratories. Follow up of sites with unsatisfactory results was done through "on and off-site support". Concurrently, standardization of standard operating procedures (SOPs) and practices to the requirements of the International Organization for Standardization (ISO) 17043:2010 was pursued.ResultsParticipants gradually increased during the program from 56 in the pilot study to 148 in Round 4 (2018). Continual participation of a particular laboratory yielded an odd of 2.5 [95% confidence interval (CI), 1.22 to 4.34] times greater for achieving a score of above 80% with each new round it participated. The "on and off-site" support supervision documented improved performance of failing laboratories. Records of GeneXpert MTB/RIF PT were used to achieve accreditation to ISO 17043:2010 in 2018.ConclusionContinued participation in GeneXpert MTB/RIF PT improves testing accuracy of laboratories. Effective implementation of this scheme requires competent human resources, facility and equipment, functional quality management system, and adherence to ISO 17043:2010

Anti-tuberculosis drug resistance among new and previously treated sputum smear-positive tuberculosis patients in Uganda: results of the first national survey.

BACKGROUND: Multidrug resistant and extensively drug resistant tuberculosis (TB) have become major threats to control of tuberculosis globally. The rates of anti-TB drug resistance in Uganda are not known. We conducted a national drug resistance survey to investigate the levels and patterns of resistance to first and second line anti-TB drugs among new and previously treated sputum smear-positive TB cases. METHODS: Sputum samples were collected from a nationally representative sample of new and previously treated sputum smear-positive TB patients registered at TB diagnostic centers during December 2009 to February 2011 using a weighted cluster sampling method. Culture and drug susceptibility testing was performed at the national TB reference laboratory. RESULTS: A total of 1537 patients (1397 new and 140 previously treated) were enrolled in the survey from 44 health facilities. HIV test result and complete drug susceptibility testing (DST) results were available for 1524 (96.8%) and 1325 (85.9%) patients, respectively. Of the 1209 isolates from new cases, resistance to any anti-TB drug was 10.3%, 5% were resistant to isoniazid, 1.9% to rifampicin, and 1.4% were multi drug resistant. Among the 116 isolates from previously treated cases, the prevalence of resistance was 25.9%, 23.3%, 12.1% and 12.1% respectively. Of the 1524 patients who had HIV testing 469 (30.7%) tested positive. There was no association between anti-TB drug resistance (including MDR) and HIV infection. CONCLUSION: The prevalence of anti-TB drug resistance among new patients in Uganda is low relative to WHO estimates. The higher levels of MDR-TB (12.1%) and resistance to any drug (25.3%) among previously treated patients raises concerns about the quality of directly observed therapy (DOT) and adherence to treatment. This calls for strengthening existing TB control measures, especially DOT, routine DST among the previously treated TB patients or periodic drug resistance surveys, to prevent and monitor development and transmission of drug resistant TB

Advancing proficiency testing for ultra in resource-limited settings using dried tube specimen: A study by SRL-Uganda.

BackgroundProficiency testing (PT) has been hard to set up due to cost limitations and technical capacity. Conventional Xpert MTB/RIF PT programs use liquid and culture spots which require stringent storage and transportation conditions with cross-contamination chances prevalent. These setbacks prompted the use of dried tube specimens (DTS) for Ultra assay PT. For continuity of PT provision, stability of DTS and compatibility with testing protocols when kept for a long period needs to be established.MethodsDTS were prepared from known isolates inactivated using a hot air oven at 85°C. 100μl of bacterial suspensions were aliquoted and dried inside a Biosafety cabinet. Panel validation was done to establish the baseline Deoxyribonucleic acid (DNA) concentration in terms of cycle threshold (Ct) value. DTS aliquots were shipped to participants to test and report within six weeks. The remaining DTS were kept at 2-8°C and room temperature for one year with testing at six months. Twenty (20) DTS samples per set remaining at one year were heated at 55°C for two weeks before testing. The means of the different samples were compared to validation data using paired t-tests. Boxplots were designed to visualize the differences in the medians of the DTS.ResultsOverall mean Ct value increased by 4.4 from the validation to testing after one year at the different storage conditions. Samples heated at 55°C showed a 6.4 Ct difference from validation data. Testing done at six months on 2-8°C stored items showed no statistical difference. At all the remaining testing times and conditions, P-values were less than 0.008 although the absolute mean Ct when compared showed slight increments and accommodated differences for the detection of MTB and rifampicin resistance. Median values for samples stored at 2-8°C were lower compared to those at room temperature.ConclusionDTS stored at 2-8°C remain more stable for one year compared to higher temperatures and can be consistently used as PT materials in more than one PT round for biannual PT providers

Diagnostic accuracy of Xpert^® MTB/XDR test using phenotypic drug susceptibility test as a reference comparator.

Diagnostic accuracy of Xpert® MTB/XDR test using phenotypic drug susceptibility test as a reference comparator.</p

Evaluation of Xpert MTB/XDR test in Uganda.

BackgroundDrug-Resistant Tuberculosis (DR-TB) is one of the major challenges to TB control.Design and methodsThis was a blinded, laboratory-based cross-sectional study using sputum samples or culture isolates. Samples were from patients with rifampicin-resistant—TB and/or with high risk for isoniazid (INH) resistance and/or 2nd line fluoroquinolones (FQ) and injectable agents (IAs). The diagnostic accuracy of the Xpert® MTB/XDR test was compared to MGIT960 and the Hain Genotype® MTBDRplus and MDRsl assays (LPA) as reference DST methods. Factors for laboratory uptake of the Xpert® MTB/XDR test were also evaluated.ResultsOf the 100 stored sputum samples included in this study, 65/99 (65.6%) were resistant to INH, 5/100 (5.0%) were resistant to FQ and none were resistant to IAs using MGIT960. The sensitivity and specificity, n (%; 95% Confidence Interval, CI) of Xpert® MTB/XDR test for; INH was 58 (89.2; 79.1–95.5) and 30 (88.2; 72.5–96.6) and for FQ; 4 (80.0; 28.3–99.4) and 95 (100; 96.2–100), respectively. Using LPA as a reference standard, a total of 52/98 (53.1%) were resistant to INH, 3/100 (3.0%) to FQ, and none to IA. The sensitivity and specificity, n (%; 95%CI) of Xpert® MTB/XDR test compared to LPA for; INH was 50 (96.1; 86.7–99.5) and 34 (74.0; 58.8–85.7) for FQ 3 (100; 29.2–100) and 96 (99.0; 94.3–99.9) respectively. The factors for laboratory uptake and roll-out of the Xpert® MTB/XDR test included: no training needed for technicians with, and one day for those without, previous Xpert-ultra experience, recording and reporting needs were not different from those of Xpert-ultra, the error rate was 4/100 (4%), one (1%) indeterminate rate and test turn-around-time were 1hr/45 minutes.ConclusionThere is high sensitivity and specificity of Xpert® MTB/XDR test for isoniazid and fluoroquinolones. There are acceptable Xpert® MTB/XDR test attributes for the test uptake and roll-out.</div

Diagnostic accuracy of Xpert^® MTB/XDR test using hain MDR<i>plus</i> and MTBDR<i>sl</i> as a reference comparator.

Diagnostic accuracy of Xpert® MTB/XDR test using hain MDRplus and MTBDRsl as a reference comparator.</p

Flow Chart of patient enrollment in the National Anti-TB drug resistance survey in Uganda; December 2009–February 2011.

<p>Figure legend: **NTM = Non Tuberculous Mycobacteria.</p