Patrick Orikiriza

Diagnostic accuracy of Xpert MTB/RIF Ultra for tuberculous meningitis in HIV-infected adults: a prospective cohort study

Summary Background WHO recommends Xpert MTB/RIF as initial diagnostic testing for tuberculous meningitis. However, diagnosis remains difficult, with Xpert sensitivity of about 50–70% and culture sensitivity of about 60%. We evaluated the diagnostic performance of the new Xpert MTB/RIF Ultra (Xpert Ultra) for tuberculous meningitis. Methods We prospectively obtained diagnostic cerebrospinal fluid (CSF) specimens during screening for a trial on the treatment of HIV-associated cryptococcal meningitis in Mbarara, Uganda. HIV-infected adults with suspected meningitis (eg, headache, nuchal rigidity, altered mental status) were screened consecutively at Mbarara Regional Referral Hospital. We centrifuged CSF, resuspended the pellet in 2 mL of CSF, and tested 0·5 mL with mycobacteria growth indicator tube culture, 1 mL with Xpert, and cryopreserved 0·5 mL, later tested with Xpert Ultra. We assessed diagnostic performance against uniform clinical case definition or a composite reference standard of any positive CSF tuberculous test. Findings From Feb 27, 2015, to Nov 7, 2016, we prospectively evaluated 129 HIV-infected adults with suspected meningitis for tuberculosis. 23 participants were classified as probable or definite tuberculous meningitis by uniform case definition, excluding Xpert Ultra results. Xpert Ultra sensitivity was 70% (95% CI 47–87; 16 of 23 cases) for probable or definite tuberculous meningitis compared with 43% (23–66; 10/23) for Xpert and 43% (23–66; 10/23) for culture. With composite standard, we detected tuberculous meningitis in 22 (17%) of 129 participants. Xpert Ultra had 95% sensitivity (95% CI 77–99; 21 of 22 cases) for tuberculous meningitis, which was higher than either Xpert (45% [24–68]; 10/22; p=0·0010) or culture (45% [24–68]; 10/22; p=0·0034). Of 21 participants positive by Xpert Ultra, 13 were positive by culture, Xpert, or both, and eight were only positive by Xpert Ultra. Of those eight, three were categorised as probable tuberculous meningitis, three as possible tuberculous meningitis, and two as not tuberculous meningitis. Testing 6 mL or more of CSF was associated with more frequent detection of tuberculosis than with less than 6 mL (26% vs 7%; p=0·014). Interpretation Xpert Ultra detected significantly more tuberculous meningitis than did either Xpert or culture. WHO now recommends the use of Xpert Ultra as the initial diagnostic test for suspected tuberculous meningitis. Funding National Institute of Neurologic Diseases and Stroke, Fogarty International Center, National Institute of Allergy and Infectious Disease, UK Medical Research Council/DfID/Wellcome Trust Global Health Trials, Doris Duke Charitable Foundation.

Use of Colorimetric Culture Methods for Detection of Mycobacterium tuberculosis Complex Isolates from Sputum Samples in Resource-Limited Settings

ABSTRACT Despite recent advances, tuberculosis (TB) diagnosis remains imperfect in resource-limited settings due to its complexity and costs, poor sensitivity of available tests, or long times to reporting. We present a report on the use of colorimetric methods, based on the detection of mycobacterial growth using colorimetric indicators, for the detection of Mycobacterium tuberculosis in sputum specimens. We evaluated the nitrate reductase assay (NRA), a modified NRA using para-nitrobenzoic acid (PNB) (NRAp), and the resazurin tube assay using PNB (RETAp) to differentiate tuberculous and nontuberculous mycobacteria. The performances were assessed at days 18 and 28 using mycobacterium growth indicator tube (MGIT) and Löwenstein-Jensen (LJ) medium culture methods as the reference standards. We enrolled 690 adults with suspected pulmonary tuberculosis from a regional referral hospital in Uganda between March 2010 and June 2011. At day 18, the sensitivities and specificities were 84.6% and 90.0% for the NRA, 84.1% and 92.6% for the NRAp, and 71.2% and 99.3% for the RETAp, respectively. At day 28, the sensitivity of the RETAp increased to 82.6%. Among smear-negative patients with suspected TB, sensitivities at day 28 were 64.7% for the NRA, 61.3% for the NRAp, and 50% for the RETAp. Contamination rates were found to be 5.4% for the NRA and 6.7% for the RETAp, compared with 22.1% for LJ medium culture and 20.4% for MGIT culture. The median times to positivity were 10, 7, and 25 days for colorimetric methods, MGIT culture, and LJ medium culture,respectively. Whereas the low specificity of the NRA/NRAp precludes it from being used for TB diagnosis, the RETAp might provide an alternative to LJ medium culture to decrease the time to culture results in resource-poor settings.

Asymptomatic Plasmodium Infections in Children in Low Malaria Transmission Setting, Southwestern Uganda

A survey of asymptomatic children in Uganda showed Plasmodium malariae and P. falciparum parasites in 45% and 55% of microscopy-positive samples, respectively. Although 36% of microscopy-positive samples were negative by rapid diagnostic test, 75% showed P. malariae or P. ovale parasites by PCR, indicating that routine diagnostic testing misses many non–P. falciparum malarial infections.

Evaluation of Capilia TB assay for rapid identification of Mycobacterium tuberculosis complex in BACTEC MGIT 960 and BACTEC 9120 blood cultures

BackgroundCapilia TB is a simple immunochromatographic assay based on the detection of MPB64 antigen specifically secreted by the Mycobacterium tuberculosis complex (MTC). Capilia TB was evaluated for rapid identification of MTC from BACTEC MGIT 960 and BACTEC 9120 systems in Kampala, Uganda. Since most studies have mainly dealt with respiratory samples, the performance of Capilia TB on blood culture samples was also evaluated.MethodsOne thousand samples from pulmonary and disseminated tuberculosis (TB) suspects admitted to the JCRC clinic and the TB wards at Old Mulago hospital in Kampala, Uganda, were cultured in automated BACTEC MGIT 960 and BACTEC 9120 blood culture systems. BACTEC-positive samples were screened for purity by sub-culturing on blood agar plates. Two hundred and fifty three (253) samples with Acid fast bacilli (AFB, 174 BACTEC MGIT 960 and 79 BACTEC 9120 blood cultures) were analyzed for presence of MTC using Capilia TB and in-house PCR assays.ResultsThe overall Sensitivity, Specificity, Positive and Negative Predictive values, and Kappa statistic for Capilia TB assay for identification of MTC were 98.4%, 97.6%, 97.7%, 98.4% and 0.96, respectively. Initially, the performance of in-house PCR on BACTEC 9120 blood cultures was poor (Sensitivity, Specificity, PPV, NPV and Kappa statistic of 100%, 29.3%,7%, 100% and 0.04, respectively) but improved upon sub-culturing on solid medium (Middlebrook 7H10) to 100%, 95.6%, 98.2%, 100% and 0.98, respectively. In contrast, the Sensitivity and Specificity of Capilia TB assay was 98.4% and 97.9%, respectively, both with BACTEC blood cultures and Middlebrook 7H10 cultured samples, revealing that Capilia was better than in-house PCR for identification of MTC in blood cultures. Additionally, Capilia TB was cheaper than in-house PCR for individual samples (

Male Gender is independently associated with pulmonary tuberculosis among sputum and non-sputum producers people with presumptive tuberculosis in Southwestern Uganda

2.03 vs.

Lowenstein-Jensen Selective Medium for Reducing Contamination in Mycobacterium tuberculosis Culture

12.59, respectively), and was easier to perform with a shorter turnaround time (20 min vs. 480 min, respectively).ConclusionCapilia TB assay is faster and cheaper than in-house PCR for rapid identification of MTC from BACTEC MGIT 960 and BACTEC 9120 culture systems in real-time testing of AFB positive cultures.

A review of antimicrobial resistance in East Africa

BackgroundLittle is known about the association between gender and risk of TB infection. We sought to assess the impact of gender on TB prevalence among people with presumptive tuberculosis at a regional referral hospital in a high TB and HIV prevalence setting.MethodsWe analyzed data from two diagnostic TB studies conducted in rural, southwestern Uganda. People with presumptive tuberculosis were evaluated by chest X-ray, fluorescence microscopy, TB culture, and HIV testing. Our primary outcome of interest was TB infection, as defined by a positive TB culture. Our primary explanatory variable of interest was gender. We fit univariable and multivariable logistic regression models to investigate associations between TB infection and gender, before and after adjusting or possible confounding factors, including ability to produce sputum, age and residence.ResultsBetween April 2010 and September 2012, 863 people with presumptive tuberculosis (PWPTB) were enrolled in the two studies at Mbarara Regional Referral Hospital (MRRH) in Uganda. Among them 664 (76.9%) were able to produce sputum. X-ray was suggestive of TB for 258 (66.5%) of males and 175 (44.8%) of female (p < 0.001). using microscopy 84 (20%) of males and 48 (10.9%) of females were diagnosed with TB (p < 0.001) while 122 (30.3%) of males and 76 (18.4%) of females were diagnosed with TB (p < 0.001) using TB culture.In multivariable logistic regression models, the odds of having TB was higher in males than females (AOR 2.2 (1.56-3.18 95% CI°, P < 0.001), after adjustment for age, HIV status, ability to produce sputum, and residence.ConclusionIn Southwestern Uganda, TB prevalence is higher among male than female people with presumptive TB. The increased risk of TB among males is independent of other TB risk factors. These findings emphasize the need for gender-focused interventions aimed at reducing TB transmission.

Low Resistance to First and Second Line Anti- Tuberculosis Drugs among Treatment Naive Pulmonary Tuberculosis Patients in Southwestern Uganda

ABSTRACT We compared Mycobacterium tuberculosis sputum culture recovery and contamination rates between Lowenstein-Jensen medium (LJ) containing the following decontaminants and LJ alone: (i) PANTA (n = 299), (ii) Selectatab-MB (n = 299), and (iii) penicillin G (n = 234). The contamination rate for LJ alone was approximately 31%, versus 5.0% for PANTA-containing, 2% for Selectatab-containing, and 9% for penicillin-containing media (P < 0.001). M. tuberculosis isolation rates were 9.8%, 17%, 18%, and 12% for standard LJ, PANTA, Selectatab, and penicillin cultures, respectively.

Detection of Plasmodium species by high resolution melt analysis of DNA from blood smears acquired in Southwestern Uganda

Background and objectives Knowledge of local and regional antimicrobial resistance (AMR) is important for clinical decision making. However, surveillance capacity for AMR is lacking throughout East Africa, and current AMR data are sparse. We sought to address this gap by summarising all available high-quality data on AMR in the East Africa region. Method We searched the PubMed database and African Journals Online archives in April and May 2015 using the search term ‘antimicrobial resistance AND sub-Saharan Africa’ to find articles published from 2005 onwards. Only full-text articles in English were included. Results We included 12 published articles in our analysis. Most articles were on bloodstream infections, hospital-based and cross-sectional in design; a majority described either community- or hospital-acquired infections. High levels of AMR to commonly-used antibiotics were reported, including 50% – 100% resistance to ampicillin and cotrimoxazole infections, emerging resistance to gentamicin (20% – 47%) and relatively high levels of resistance to ceftriaxone (46% – 69%) among Gram-negative infections. Much of the resistance was reported to be in Klebsiella species and Escherichia coli. Among Gram-positive infections, extensive resistance was reported to ampicillin (100%), gentamicin and ceftriaxone (50% – 100%), with methicillin-resistant Staphylococcus aureus prevalence ranging from 2.6% – 4.0%. Conclusion Overall, bacterial resistance was reported among commonly-used antibiotics (ampicillin, gentamicin and ceftriaxone), raising concern that these antibiotics may no longer be useful for treating moderate or severe bacterial infections in East Africa. Thus, empirical treatment of bacterial infections needs to be reconsidered and guided by local assessment of AMR. Improvements in the limited amount of quality data and lack of harmonisation in assessing the burden of AMR are also needed.

Detection and Quantification of Mycobacterium tuberculosis in the Sputum of Culture-Negative HIV-infected Pulmonary Tuberculosis Suspects: A Proof-of-Concept Study

Background There are limited data on region-specific drug susceptibility of tuberculosis (TB) in Uganda. We performed resistance testing on specimens collected from treatment-naive patients with pulmonary TB in Southwestern Uganda for first and second line anti-TB drugs. We sought to provide data to guide regional recommendations for empiric TB therapy. Methods Archived isolates, obtained from patients at Mbarara Regional Referral Hospital from February 2009 to February 2013, were tested for resistance to isoniazid and rifampicin using the MTBDRplus and Xpert MTB/RIF assays. A subset of randomly selected isolates was tested for second line agents, including fluoroquinolones (FQs), aminoglycosides, cyclic peptides, and ethambutol using the MTBDRsl assay. We performed confirmatory testing for FQ resistance using repeated MTBDRsl, the Mycobacteria growth indicator tube (MGIT) assay, and sequencing of the gyrA and gyrB genes. Results We tested isolates from 190 patients. The cohort had a median age of 33 years (IQR 26-43), 69% (131/190) were male, and the HIV prevalence was 42% (80/190). No isolates (0/190) were rifampicin-resistant and only 1/190 (0.5%) was isoniazid-resistant. Among 92 isolates tested for second-line drug resistance, 71 (77%) had interpretable results, of which none were resistant to aminoglycosides, cyclic peptides or ethambutol. Although 7 (10%) initially tested as resistant to FQs by the MTBDRsl assay, they were confirmed as susceptible by repeat MTBDRsl testing as well as by MGIT and gyrase gene sequencing Conclusion We found no MDR-TB and no resistance to ethambutol, FQs, or injectable anti-TB drugs in treatment naïve patients with pulmonary TB in Southwestern Uganda. Standard treatment guidelines for susceptible TB should be adequate for most patients with TB in this population. Where possible, molecular susceptibility testing methods should be routinely validated by culture methods.