Mycobacterium tuberculosis spoligotypes and drug susceptibility pattern of isolates from tuberculosis patients in South-Western Uganda

BACKGROUND: Determination of the prevalence and drug susceptibility of the M. tuberculosis strains is important in tuberculosis control. We determined the genetic diversity and susceptibility profiles of mycobacteria isolated from tuberculosis patients in Mbarara, South Western Uganda. METHODS: We enrolled, consecutively; all newly diagnosed and previously treated smear-positive TB patients aged ≥ 18 years. The isolates were characterized using regions of difference (RD) analysis and spoligotyping. Drug resistance against rifampicin and isoniazid were tested using the Genotype(® )MDRTBplus assay and the indirect proportion method on Lowenstein-Jensen media. HIV-1 testing was performed using two rapid HIV tests. RESULTS: A total of 125 isolates from 167 TB suspects (60% males) with a mean age 33.7 years and HIV prevalence of 67.9% (55/81) were analyzed. Majority (92.8%) were new cases while only 7.2% were retreatment cases. All the 125 isolates were identified as M. tuberculosis strict sense with the majority (92.8%) of the isolates being modern strains while seven (7.2%) isolates were ancestral strains. Spoligotyping revealed 79 spoligotype patterns, with an overall diversity of 63.2%. Sixty two (49.6%) of the isolates formed 16 clusters consisting of 2-15 isolates each. A majority (59.2%) of the isolates belong to the Uganda genotype group of strains. The major shared spoligotypes in our sample were SIT 135 (T2-Uganda) with 15 isolates and SIT 128 (T2) with 3 isolates. Sixty nine (87%) of the 79 patterns had not yet been defined in the SpolDB4.0.database. Resistance mutations to either RIF or INH were detected in 6.4% of the isolates. Multidrug resistance, INH and RIF resistance was 1.6%, 3.2% and 4.8%, respectively. The rpoβ gene mutations seen in the sample were D516V, S531L, H526Y H526D and D516V, while one strain had a Δ1 mutation in the wild type probes. There were three strains with katG (codon 315) gene mutations only while one strain showed the inhA promoter gene mutation. CONCLUSION: The present study shows that the TB epidemic in Mbarara is caused by modern M. tuberculosis strains mainly belonging to the Uganda genotype and anti-TB drug resistance rate in the region is low

Vaginal colonisation of women in labour with potentially pathogenic bacteria: A cross sectional study at three primary health care facilities in Central Uganda

Background: Potentially pathogenic bacteria that colonise the lower genital tract of women in labour can be passed to the baby during birth. While many babies become colonised with these bacteria after delivery, a few develop neonatal infections. The lower genital tract is a reservoir for potential pathogens and a source of infection for neonates. We determined the prevalence of vaginal colonisation of potentially pathogenic bacteria among women in labour in Central Uganda and identified potential risk factors associated with this colonisation. Methods: We conducted a cross sectional study at three primary health care facilities and collected vaginal swabs from HIV-1 negative women in labour. Specimens were cultured on different selective microbiological media, and biochemical tests were used to classify bacterial isolates on the species level. Multivariable logistic regression analyses were used to estimate the association between relevant exposures and colonisation with potentially pathogenic bacteria. Results: We recruited 1472 women in labour whose mean age was 24.6 years (standard deviation [SD] 4.9). Of these, 955 (64.9%; 95% Confidence Interval [CI] 62.4, 67%) were vaginally colonised with at least one potentially pathogenic bacterial species. The most commonly isolated species were Escherichia coli (n = 508; 34.5%), Klebsiella pneumoniae (n = 144; 9.8%) and Staphylococcus aureus (n = 121; 8.2%). Results from exploratory multivariable regression analyses indicated that having had ≥5 previous pregnancies (adjusted odds ratio [aOR] 0.59; 95% CI 0.35, 0.97) or being ≥30 years old (aOR 1.52; 95% CI 1.03, 2.23) could be associated with vaginal colonisation with any potentially pathogenic bacteria, as well as with vaginal colonisation with S. aureus (aOR 0.33; 95% CI 0.12, 0.88, and aOR 2.17; 95% CI 1.17, 4.00, respectively). Possession of domestic animals in a household (aOR 0.57; 95% CI 0.35, 0.92) could be associated with vaginal colonisation with E. coli. Conclusions: Two-thirds of HIV-1 negative women in labour were vaginally colonised by potentially pathogenic bacteria, mainly E. coli, K. pneumoniae, and S. aureus.publishedVersio

Sputum quality and diagnostic performance of GeneXpert MTB/RIF among smear-negative adults with presumed tuberculosis in Uganda.

BackgroundIntroduction of GeneXpert MTB/RIF (Xpert) assay has constituted a major breakthrough for tuberculosis (TB) diagnostics. Several patient factors may influence diagnostic performance of Xpert including sputum quality.ObjectiveWe carried out a prospective, observational, cross-sectional study to determine the effect of sputum quality on diagnostic performance of Xpert among presumed TB patients in Uganda.MethodsWe collected clinical and demographic information and two sputum samples from participants. Staff recorded sputum quality and performed LED fluorescence microscopy and mycobacterial culture on each sample. If both smear examinations were negative, Xpert testing was performed. We calculated diagnostic yield, sensitivity, specificity, and other indicators for Xpert for each stratum of sputum quality in reference to a standard of mycobacterial culture.ResultsPatients with salivary sputum showed a trend towards a substantially higher proportion of samples that were Xpert-positive (54/286, 19%, 95% CI 15-24) compared with those with all other sputum sample types (221/1496, 15%, 95% CI 13-17). Blood-stained sputum produced the lowest sensitivity (28%; 95% CI 12-49) and salivary sputum the highest (66%; 95% CI 53-77). Specificity didn't vary meaningfully by sample types. Salivary sputum was significantly more sensitive than mucoid sputum (+13%, 95% CI +1 to +26), while blood-stained sputum was significantly less sensitive (-24%, 95% CI -42 to -5).ConclusionsOur findings demonstrate the need to exercise caution in collecting sputum for Xpert and in interpreting results because sputum quality may impact test yield and sensitivity. In particular, it may be wise to pursue additional testing should blood-stained sputum test negative while salivary sputum should be readily accepted for Xpert testing given its higher sensitivity and potentially higher yield than other sample types. These findings challenge conventional recommendations against collecting salivary sputum for TB diagnosis and could inform new standards for sputum quality

Umbilical Cord Stump Infections in Central Uganda: Incidence, Bacteriological Profile, and Risk Factors

Umbilical cord stump infection (omphalitis) is a risk factor for neonatal sepsis and death. We assessed the incidence of omphalitis, described the bacteriological and antibiotic-resistance profile of potentially pathogenic bacteria isolated from the umbilical cord stump of omphalitis cases, and evaluated whether bacteria present in the birth canal during birth predicted omphalitis. We enrolled 769 neonates at birth at three primary healthcare facilities and followed them for 28 days with scheduled visits on days 3, 7, 14, and 28. Cox regression models were used to estimate the rates of omphalitis associated with potential risk factors. Sixty-five (8.5%) neonates developed omphalitis, with an estimated incidence of 0.095 cases per 28 child-days (95% CI 0.073, 0.12). Potentially pathogenic bacteria were isolated from the cord stump area of 41 (63.1%) of the 65 neonates with omphalitis, and the most commonly isolated species were Escherichia coli (n = 18), Klebsiella pneumoniae (n = 10), Citrobacter freundii (n = 5), and Enterobacter spp. (n = 4). The Enterobacteriaceace isolates were resistant to gentamicin (10.5%, 4/38), ampicillin (86.8%, 33/38), and ceftriaxone (13.2%, 5/38). Delayed initiation of breastfeeding was associated with an increased risk of omphalitis (aHR 3.1; 95% CI 1.3, 7.3); however, vaginal colonization with potentially pathogenic bacteria did not predict omphalitis.publishedVersio

Accuracy of GenoQuick MTB test in detection of Mycobacterium tuberculosis in sputum from TB presumptive patients in Uganda

The author(s) disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: MIND study was funded by National Institutes of Health (NIH) (NIH K24 HL087713, R01 HL090335, and R01 HL128156). MBN Clinical Laboratories provided PCR reagents, other laboratory consumables, and workspace. Sylvia Kaswabuli received a partial scholarship from the Pulmonary Complications of AIDS Research Training Program (NIH D43TW009607).Objective: The objective of the study was to determine the diagnostic performance of the GenoQuick MTB test on heated sputum against the conventional Lowenstein–Jensen Mycobacterium tuberculosis culture as the reference method for tuberculosis diagnosis. Introduction: Fast, reliable, and easy-to-use tests for tuberculosis diagnosis are essential to achieving the Sustainable Development Goal of diagnosing and treating 90% of tuberculosis patients by 2030. We evaluated the diagnostic performance of the GenoQuick MTB, a polymerase chain reaction–lateral flow test, in Uganda, a resource-constrained, high tuberculosis- and HIV-burden setting. Methods: Fresh sputum samples from presumptive tuberculosis patients at Mulago Hospital were tested for M. tuberculosis using smear microscopy, GenoQuick MTB test, and Lowenstein–Jensen culture. For the GenoQuick MTB test, mycobacterial DNA was extracted by heating sputum at 95°C for 30 min while DNA amplification and detection were done following the manufacturer’s protocol (Hain Lifescience, Nehren, Germany). Sensitivity, specificity, and kappa agreements were calculated against Lowenstein–Jensen M. tuberculosis culture as a reference test using STATA V12. Results: Of the 86 tested samples, 30.2% had culture-confirmed pulmonary tuberculosis. Overall, sensitivity was higher for GenoQuick MTB (81%, 95% confidence interval: 60%−93%) than for smear microscopy (69%, 95% confidence interval: 48%−86%). Among people living with HIV, sensitivity was identical for GenoQuick MTB and smear tests (75%, 95% confidence interval: 42%−95%). Contrastingly, smear had a higher overall specificity (98%, 95% confidence interval: 91%−100%) than for GenoQuick MTB (92%, 95% confidence interval: 81%−97%). A similar trend of specificity was observed among the people living with HIV for smear microscopy (100%, 95% CI: 87%−100%) and for GenoQuick MTB (96%, 95% confidence interval: 81%−100%). Conclusion: The GenoQuick MTB test could be a potential tuberculosis diagnostic test given its higher sensitivity. Evaluation of this test in larger studies is recommended.Publisher PDFPeer reviewe

Vaginal colonization with antimicrobial-resistant bacteria among women in labor in central Uganda: prevalence and associated factors

Background According to WHO ( CISMAC. Centre for Intervention Science in Maternal and Child health), the antimicrobial resistant bacteria considered to be clinically most important for human health and earmarked for surveillance include extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae, carbapenem-resistant bacteria, methicillin-resistant (MRSA) and, macrolide-lincosamide-streptogramin B -resistant vancomycin-resistant (VRSA) Staphylococcus aureus and vancomycin-resistant Enterococcus (VRE). If these bacteria are carried in the female genital tract, they may be transmitted to the neonate causing local or systemic neonatal infections that can be difficult to treat with conventionally available antimicrobials. In order to develop effective treatment strategies, there is need for updated information about the prevalence of colonization with important antimicrobial-resistant pathogens. Objective We sought to estimate the prevalence of vaginal colonization with potentially pathogenic and clinically important AMR bacteria among women in labour in Uganda and to identify factors associated with colonization. Methods We conducted a cross-sectional study among HIV-1 and HIV-2 negative women in labour at three primary health care facilities in Uganda. Drug susceptibility testing was done using the disk diffusion method on bacterial isolates cultured from vaginal swabs. We calculated the prevalence of colonization with potentially pathogenic and clinically important AMR bacteria, in addition to multidrug-resistant (MDR) bacteria, defined as bacteria resistant to antibiotics from ≥ 3 antibiotic classes. Results We found that 57 of the 1472 enrolled women (3.9% prevalence; 95% Confidence interval [CI] 3.0%, 5.1%) were colonized with ESBL-producing Enterobacteriaceace, 27 (1.8%; 95% CI 1.2%, 2.6%) were colonized with carbapenem-resistant Enterobacteriaceae, and 85 (5.8%; 95% CI 4.6%, 7.1%) were colonized with MRSA. The prevalence of colonization with MDR bacteria was high (750/1472; 50.9%; 95% CI 48.4%, 53.5%). Women who were ≥ 30 years of age had higher odds of being colonized with MDR bacteria compared to women aged 20–24 years (OR 1.6; 95% CI 1.1, 2.2). Conclusion Most of the women included in our study were vaginally colonized with potentially pathogenic MDR and other clinically important AMR bacteria. The high prevalence of colonization with these bacteria is likely to further increase the incidence of difficult-to-treat neonatal sepsis.publishedVersio

Direct Nitrate Reductase Assay versus Microscopic Observation Drug Susceptibility Test for Rapid Detection of MDR-TB in Uganda

The most common method for detection of drug resistant (DR) TB in resource-limited settings (RLSs) is indirect susceptibility testing on Lowenstein-Jensen medium (LJ) which is very time consuming with results available only after 2–3 months. Effective therapy of DR TB is therefore markedly delayed and patients can transmit resistant strains. Rapid and accurate tests suitable for RLSs in the diagnosis of DR TB are thus highly needed. In this study we compared two direct techniques - Nitrate Reductase Assay (NRA) and Microscopic Observation Drug Susceptibility (MODS) for rapid detection of MDR-TB in a high burden RLS. The sensitivity, specificity, and proportion of interpretable results were studied. Smear positive sputum was collected from 245 consecutive re-treatment TB patients attending a TB clinic in Kampala, Uganda. Samples were processed at the national reference laboratory and tested for susceptibility to rifampicin and isoniazid with direct NRA, direct MODS and the indirect LJ proportion method as reference. A total of 229 specimens were confirmed as M. tuberculosis, of these interpretable results were obtained in 217 (95%) with either the NRA or MODS. Sensitivity, specificity and kappa agreement for MDR-TB diagnosis was 97%, 98% and 0.93 with the NRA; and 87%, 95% and 0.78 with the MODS, respectively. The median time to results was 10, 7 and 64 days with NRA, MODS and the reference technique, respectively. The cost of laboratory supplies per sample was low, around 5 USD, for the rapid tests. The direct NRA and MODS offered rapid detection of resistance almost eight weeks earlier than with the reference method. In the study settings, the direct NRA was highly sensitive and specific. We consider it to have a strong potential for timely detection of MDR-TB in RLS

Evaluation of an Aspergillus IgG/IgM lateral flow assay for serodiagnosis of fungal asthma in Uganda

Background: Diagnosis of fungal allergies in asthma remains problematic in low-and middle-income countries due to non-availability of point-of-care testing. In this study, we aimed to evaluate the performance of an Aspergillus immunochromatographic technology (ICT) IgG/M lateral flow device (LFD) for the serological diagnosis of allergic bronchopulmonary aspergillosis (ABPA) and severe asthma with fungal sensitisation (SAFS) among Ugandan adult asthmatics. Methods: 374 adult (aged ≥18years) asthmatics in the African Severe Asthma Program study, Ugandan site constituted the study population. ABPA and SAFS were diagnosed according to standard criteria. Asthmatics who did not meet the above criteria constituted a control group. The LFD tests were performed and read according to manufacturer’s instructions. Results: ABPA was found in 12/374 (3.2%) and SAFS in 60/374 (16%) participants. The sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) for the Aspergillus ICT for the diagnosis of ABPA were 0.0%, 96.4%, 0.0% and 96.7% respectively, and for SAFS 6.7%, 97.1%, 30.8% and 84.5% respectively. False positive and negative rates were 3.5% and 3.2% for ABPA and 2.4% and 14.9% for SAFS, respectively. Patients with a positive LFD significantly had higher median Aspergillus fumigatus-specific IgE levels compared to those with negative LFD (median: 0.06 kUA/l VS 0.03 kUA/L, P = 0.011). Conclusion: The Aspergillus ICT IgG/M LFD had a poor diagnostic performance for the diagnosis of both ABPA and SAFS. Its greatest value may be in distinguishing chronic and allergic aspergillosis in Africa

Rapid screening of MDR-TB using molecular Line Probe Assay is feasible in Uganda

<p>Abstract</p> <p>Background</p> <p>About 500 new smear-positive Multidrug-resistant tuberculosis (MDR-TB) cases are estimated to occur per year in Uganda. In 2008 in Kampala, MDR-TB prevalence was reported as 1.0% and 12.3% in new and previously treated TB cases respectively. Line probe assays (LPAs) have been recently approved for use in low income settings and can be used to screen smear-positive sputum specimens for resistance to rifampicin and isoniazid in 1-2 days.</p> <p>Methods</p> <p>We assessed the performance of a commercial line probe assay (Genotype MTBDR<it>plus</it>) for rapid detection of rifampicin and isoniazid resistance directly on smear-positive sputum specimens from 118 previously treated TB patients in a reference laboratory in Kampala, Uganda. Results were compared with MGIT 960 liquid culture and drug susceptibility testing (DST). LPA testing was also performed in parallel in a University laboratory to assess the reproducibility of results.</p> <p>Results</p> <p>Overall, 95.8% of smear-positive specimens gave interpretable results within 1-2 days using LPA. Sensitivity, specificity, positive and negative predictive values were 100.0%, 96.1%, 83.3% and 100.0% for detection of rifampicin resistance; 80.8%, 100.0%, 100.0% and 93.0% for detection of isoniazid resistance; and 92.3%, 96.2%, 80.0% and 98.7% for detection of multidrug-resistance compared with conventional results. Reproducibility of LPA results was very high with 98.1% concordance of results between the two laboratories.</p> <p>Conclusions</p> <p>LPA is an appropriate tool for rapid screening for MDR-TB in Uganda and has the potential to substantially reduce the turnaround time of DST results. Careful attention must be paid to training, supervision and adherence to stringent laboratory protocols to ensure high quality results during routine implementation.</p