Evaluation of in-house PCR for diagnosis of smear-negative pulmonary tuberculosis in Kampala, Uganda

ABSTRACT: BACKGROUND: Nucleic acid amplification tests (NAATs) have offered hope for rapid diagnosis of tuberculosis (TB). However, their efficiency with smear-negative samples has not been widely studied in low income settings. Here, we evaluated in-house PCR assay for diagnosis of smear-negative TB using Lowenstein-Jensen (LJ) culture as the baseline test. Two hundred and five pulmonary TB (PTB) suspects with smear-negative sputum samples, admitted on a short stay emergency ward at Mulago Hospital in Kampala, Uganda, were enrolled. Two smear-negative sputum samples were obtained from each PTB suspect and processed simultaneously for identification of MTBC using in-house PCR and LJ culture. RESULTS: Seventy two PTB suspects (35%, 72/205) were LJ culture positive while 128 (62.4%, 128/205) were PCR-positive. The sensitivity and specificity of in-house PCR for diagnosis of smear-negative PTB were 75% (95% CI 62.6-85.0) and 35.9% (95% CI 27.2-45.3), respectively. The positive and negative predictive values were 39% (95% CI 30.4-48.2) and 72.4% (95% CI 59.1-83.3), respectively, while the positive and negative likelihood ratios were 1.17 (95% CI 0.96-1.42) and 0.70 (95% CI 0.43-1.14), respectively.One hundred and seventeen LJ culture-negative suspects (75 PCR-positive and 42 PCR-negative) were enrolled for follow-up at 2 months. Of the PCR-positive suspects, 45 (60%, 45/75) were still alive, of whom 29 (64.4%, 29/45) returned for the follow-up visit; 15 (20%, 15/75) suspects died while another 15 (20%, 15/75) were lost to follow-up. Of the 42 PCR-negative suspects, 22 (52.4%, 22/42) were still alive, of whom 16 (72.7%, 16/22) returned for follow-up; 11 (26.2%, 11/42) died while nine (21.4%, 9/42) were lost to follow-up.Overall, more PCR-positive suspects were diagnosed with PTB during follow-up visits but the difference was not statistically significant (27.6%, 8/29 vs. 25%, 4/16, p = 0.9239). Furthermore, mortality was higher for the PCR-negative suspects but the difference was also not statistically significant (26.2% vs. 20% p = 0.7094). CONCLUSION: In-house PCR correlates poorly with LJ culture for diagnosis of smear-negative PTB. Therefore, in-house PCR may not be adopted as an alternative to LJ culture.     

Long delays and missed opportunities in diagnosing smear-positive pulmonary tuberculosis in Kampala, Uganda: a cross-sectional study

Background

Early detection and treatment of tuberculosis cases are the hallmark of successful tuberculosis control. We conducted a cross-sectional study at public primary health facilities in Kampala city, Uganda to quantify diagnostic delay among pulmonary tuberculosis (PTB) patients, assess associated factors, and describe trajectories of patients'' health care seeking.

Methodology/Principal Findings

Semi-structured interviews with new smear-positive PTB patients (≥15 years) registered for treatment. Between April 2007 and April 2008, 253 patients were studied. The median total delay was 8 weeks (IQR 4–12), median patient delay was 4 weeks (inter-quartile range [IQR] 1–8) and median health service delay was 4 weeks (IQR 2–8). Long total delay (>14 weeks) was observed for 61/253 (24.1%) of patients, long health service delay (>6 weeks) for 71/242 (29.3%) and long patient delay (>8 weeks) for 47/242 (19.4%). Patients who knew that TB was curable were less likely to have long total delay (adjusted Odds Ratio [aOR] 0.28; 95%CI 0.11–0.73) and long patient delay (aOR 0.36; 95%CI 0.13–0.97). Being female (aOR 1.98; 95%CI 1.06–3.71), staying for more than 5 years at current residence (aOR 2.24 95%CI 1.18–4.27) and having been tested for HIV before (aOR 3.72; 95%CI 1.42–9.75) was associated with long health service delay. Health service delay contributed 50% of the total delay. Ninety-one percent (231) of patients had visited one or more health care providers before they were diagnosed, for an average (median) of 4 visits (range 1–30). All but four patients had systemic symptoms by the time the diagnosis of TB was made.

Conclusions/Significance

Diagnostic delay among tuberculosis patients in Kampala is common and long. This reflects patients waiting too long before seeking care and health services waiting until systemic symptoms are present before examining sputum smears; this results in missed opportunities for diagnosis.     

Potential Sources and Transmission of Salmonella and Antimicrobial Resistance in Kampala,Uganda

In sub‒Saharan Africa, non‒typhoidal Salmonellae (NTS) cause invasive disease particularly in children and HIV infected adults, but the disease epidemiology is poorly understood. Between 2012 and 2013, we investigated NTS sources and transmission in Kampala. We detected Salmonella in 60% of the influent and 60% of the effluent samples from a wastewater treatment plant and 53.3% of the influent and 10% of the effluent samples from waste stabilization ponds that serve the human population; 40.9% of flush‒water samples from ruminant slaughterhouses, 6.6% of the poultry fecal samples from live bird markets and 4% of the fecal samples from swine at slaughter; and in 54.2% of the water samples from a channel that drains storm–water and effluents from the city. We obtained 775 Salmonella isolates, identified 32 serovars, and determined resistance to 15 antimicrobials. We genotyped common serovars using multiple‒locus variable number tandem repeats analysis or pulsed‒field gel electrophoresis. In addition, we analyzed 49 archived NTS isolates from asymptomatic livestock and human clinical cases. Salmonella from ruminant and swine sources were mostly pan‒susceptible (95%) while poultry isolates were generally more resistant. Salmonella Kentucky isolated from poultry exhibited extensive drug resistance characterized by resistance to 10 antimicrobials. Interestingly, similar genotypes of S. Kentucky but with less antimicrobial resistance (AMR) were found in poultry, human and environmental sources. The observed AMR patterns could be attributed to host or management factors associated with production. Alternatively, S. Kentucky may be prone to acquiring AMR. The factors driving AMR remain poorly understood and should be elucidated. Overall, shared genotypes and AMR phenotypes were found in NTS from human, livestock and environmental sources, suggesting zoonotic and environmental transmissions most likely occur. Information from this study could be used to control NTS transmission.     

Effectiveness of the standard WHO recommended retreatment regimen (category II) for tuberculosis in Kampala, Uganda: a prospective cohort study

Background

Each year, 10%–20% of patients with tuberculosis (TB) in low- and middle-income countries present with previously treated TB and are empirically started on a World Health Organization (WHO)-recommended standardized retreatment regimen. The effectiveness of this retreatment regimen has not been systematically evaluated.

Methods and Findings

From July 2003 to January 2007, we enrolled smear-positive, pulmonary TB patients into a prospective cohort to study treatment outcomes and mortality during and after treatment with the standardized retreatment regimen. Median time of follow-up was 21 months (interquartile range 12–33 months). A total of 29/148 (20%) HIV-uninfected and 37/140 (26%) HIV-infected patients had an unsuccessful treatment outcome. In a multiple logistic regression analysis to adjust for confounding, factors associated with an unsuccessful treatment outcome were poor adherence (adjusted odds ratio [aOR] associated with missing half or more of scheduled doses 2.39; 95% confidence interval (CI) 1.10–5.22), HIV infection (2.16; 1.01–4.61), age (aOR for 10-year increase 1.59; 1.13–2.25), and duration of TB symptoms (aOR for 1-month increase 1.12; 1.04–1.20). All patients with multidrug-resistant TB had an unsuccessful treatment outcome. HIV-infected individuals were more likely to die than HIV-uninfected individuals (p<0.0001). Multidrug-resistant TB at enrolment was the only common risk factor for death during follow-up for both HIV-infected (adjusted hazard ratio [aHR] 17.9; 6.0–53.4) and HIV-uninfected (14.7; 4.1–52.2) individuals. Other risk factors for death during follow-up among HIV-infected patients were CD4<50 cells/ml and no antiretroviral treatment (aHR 7.4, compared to patients with CD4≥200; 3.0–18.8) and Karnofsky score <70 (2.1; 1.1–4.1); and among HIV-uninfected patients were poor adherence (missing half or more of doses) (3.5; 1.1–10.6) and duration of TB symptoms (aHR for a 1-month increase 1.9; 1.0–3.5).

Conclusions

The recommended regimen for retreatment TB in Uganda yields an unacceptable proportion of unsuccessful outcomes. There is a need to evaluate new treatment strategies in these patients. Please see later in the article for the Editors'' Summary     

Participatory planning, management and alternative livelihoods for poor wetland-dependent communities in Kampala, Uganda

The paper is based on an on-going 3-year study in the wetland communities of Kampala. The study uses participatory methods and aims to contribute to (i) the development of low-income wetland communities, (ii) to prepare these communities to become less dependent on wetlands without receding into poverty, and (iii) the better management of the wetlands. The communities in direct dependence and intimate interaction with Nakivubo wetlands are mainly poor, live and work under hazardous conditions, and their activities pose a threat to the ecological function of the wetlands. Yet these wetlands are important for filtering the city's waste and storm water before it flows into Lake Victoria's Murchison Bay, which is Kampala's source of piped water. Government approaches to the problem of wetland encroachment have largely failed because they are confrontational, and are not consistent or participatory. The study has in the first year conducted a series of activities including stakeholder analysis, resource analysis, livelihood analysis, a questionnaire survey and action planning. Preliminary data show that wetland dependency is very high among the poor nearby communities. They practice cultivation, brick-making and harvesting of wetland vegetation. However, these activities are under threat because wetland resources are dwindling due to increasing population and over-use. Livelihoods are threatened not only by the decreasing productivity of the wetland, but also by the ever-present government threat to evict wetland encroachers to restore its ecology. The study therefore works with communities to prepare for less dependence on wetlands so that they do not suddenly recede into worse poverty if they are evicted. They formulate strategies to enhance alternative livelihood, and for management of the wetland. Action plans have been formulated to address the situation through a newly created association.     

Four years of IVF/ICSI experience in Kampala (Uganda)

⁴Correspondence address. Zonlaan 49, 1700 Dilbeek, Belgium. E-mail: peterplatteau{at}telenet.be We all know that starting and running an ART clinic is not soeasy as some people might perceive from outside. Doing the samething in the middle of Africa is even more challenging as someevidences in the Western world are not so obvious in this partof the world. We started our clinic in Kampala in 2004. Theclinic was a converted apartment from a four flat building.In the beginning, we had difficulties with importing drugs,culture media and consumables; we had the feeling everybodywas against us. We overcame multiple power failures, night intrudersand a 20% masturbation failure, but once the first IVF/ICSIbabies were born, people started to believe in the project.At present, 250 IVF/ICSI cycles a year are done in batches,we have a successful embryo freezing programme, offer IUI/ICSIfor sero discordant HIV couples and have the first babies afterIVF, ICSI, testicular biopsy, embryo freezing, oocyte donationand surrogacy in Central Africa. The results are comparableto the ones in the Western world.     

Massive structural and compositional changes over two decades in forest fragments near Kampala,Uganda

Private forests harbor considerable biodiversity, however, they are under greater threat than reserved areas, particularly from urbanization, agriculture, and intense exploitation for timber and fuel wood. The extent to which they may act as habitats for biodiversity and how level of protection impacts trends in biodiversity and forest structure over time remain underresearched. We contribute to filling this research gap by resampling a unique data set, a detailed survey from 1990 of 22 forests fragments of different ownership status and level of protection near Kampala, Uganda. Eleven of the 22 fragments were lost over 20 years, and six of the remnants reduced in size. Forest structure and composition also showed dramatic changes, with six of the remnant fragments showing high temporal species turnover. Species richness increased in four of the remaining forests over the resample period. Forest ownership affected the fate of the forests, with higher loss in privately owned forests. Our study demonstrates that ownership affects the fate of forest fragments, with private forests having both higher rates of area loss, and of structural and compositional change within the remaining fragments. Still, the private forests contribute to the total forest area, and they harbor biodiversity including IUCN “vulnerable” and “endangered” species. This indicates the conservation value of the fragments and suggests that they should be taken into account in forest conservation and restoration.     

An autopsy study describing causes of death and comparing clinico-pathological findings among hospitalized patients in Kampala, Uganda

Background

Information on causes of death in HIV-infected patients in Sub-Saharan Africa is mainly derived from observational cohort and verbal autopsy studies. Autopsy is the gold standard to ascertain cause of death. We conducted an autopsy study to describe and compare the clinical and autopsy causes of death and contributory findings in hospitalized HIV-infected and HIV-uninfected patients in Uganda.

Methods

Between May and September 2009 a complete autopsy was performed on patients that died on a combined infectious diseases gastroenterology ward in Mulago Hospital in Kampala, Uganda. Autopsy cause of death and contributing findings were based on the macro- and microscopic post-mortem findings combined with clinical information. Clinical diagnoses were reported by the ward doctor and classified as confirmed, highly suspected, considered or not considered, based on information derived from the medical chart. Results are reported according to HIV serostatus.

Results

Fifty-three complete autopsies were performed in 66% HIV-positive, 21% HIV-negative and 13% patients with an unknown HIV serological status. Infectious diseases caused death in 83% of HIV-positive patients, with disseminated TB as the main diagnosis causing 37% of deaths. The spectrum of illness and causes of death were substantially different between HIV-positive and HIV-negative patients. In HIV-positive patients 12% of postmortem diagnoses were clinically confirmed, 27% highly suspected, 16% considered and 45% not considered. In HIV-negative patients 17% of postmortem diagnoses were clinically highly suspected, 42% considered and 42% not considered.

Conclusion

Autopsy examination remains an important tool to ascertain causes of death particularly in settings with limited access to diagnostic testing during life. HIV-positive patients continue to die from treatable and clinically undiagnosed infectious diseases. Until rapid-point of care testing is available to confirm common infections, empiric treatment should be further investigated.     

Molecular Characterization of Staphylococcus aureus from Patients with Surgical Site Infections at Mulago Hospital in Kampala,Uganda

Background

The prevalence of Methicillin resistant Staphylococcus aureus (MRSA) is progressively increasing globally with significant regional variation. Understanding the Staphylococcus aureus lineages is crucial in controlling nosocomial infections. Recent studies on S. aureus in Uganda have revealed an escalating burden of MRSA. However, the S. aureus genotypes circulating among patients are not known. Here, we report S. aureus lineages circulating in patients with surgical site infections (SSI) at Mulago National hospital, Kampala, Uganda.

Methods

A cross-sectional study involving 314 patients with SSI at Mulago National Hospital was conducted from September 2011 to April 2012. Pus swabs from the patients’ SSI were processed using standard microbiological procedures. Methicillin sensitive Staphylococcus aureus (MSSA) and MRSA were identified using phenotypic tests and confirmed by PCR-detection of the nuc and mecA genes, respectively. SCCmec genotypes were determined among MRSA isolates using multiplex PCR. Furthermore, to determine lineages, spa sequence based-genotyping was performed on all S. aureus isolates.

Results

Of the 314 patients with SSI, S. aureus accounted for 20.4% (64/314), of which 37.5% (24/64) were MRSA. The predominant SCCmec types were type V (33.3%, 8/24) and type I (16.7%, 4/24). The predominant spa lineages were t645 (17.2%, 11/64) and t4353 (15.6%, 10/64), and these were found to be clonally circulating in all the surgical wards. On the other hand, lineages t064, t355, and t4609 were confined to the obstetrics and gynecology wards. A new spa type (t10277) was identified from MSSA isolate. On multivariate logistic regression analysis, cancer and inducible clindamycin resistance remained as independent predictors of MRSA-SSI.

Conclusion

SCCmec types I and V are the most prevalent MRSA mecA types from the patients’ SSI. The predominant spa lineages (t645 and t4353) are clonally circulating in all the surgical wards, calling for strengthening of infection control practices at Mulago National Hospital.     

北京城市化进程中家庭食物碳消费动态

不可持续的家庭消费是造成全球环境问题的主要原因,食物碳消费研究是促进可持续家庭消费的重要内容。基于政府宏观统计数据,分析北京城市化进程中城市家庭食物碳消费的变化趋势和影响因素。结果表明,与1979年相比,1999年北京城市家庭人均及户均食物消费量分别减少了15.2%和38.6%,而食物碳消费总量增加了28.5%,食物碳消费结构由“以粮食为主”转变成“以粮食和肉类为主”。城市化进程中,以1993年为界,家庭食物人均及户均碳消费量均由明显减少趋势转变为明显增长趋势,变化的主要原因由“食物消费结构变化引起的人均谷物类碳消费量的减少”转变为“人均食物消费量增加引起的人均肉类碳消费量的增加”。北京城市家庭已基本完成食物消费结构的转变,人均食物消费量仍继续增加。GDP指数是影响人均食物碳消费量的主要经济因素,经济的继续增长可能带来人均食物碳消费量的增加,北京城市家庭食物碳消费尚未达到稳定状态。     

Aetiology of Pulmonary Symptoms in HIV-Infected Smear Negative Recurrent PTB Suspects in Kampala,Uganda: A Cross-Sectional Study

Introduction

Previously treated TB patients with pulmonary symptoms are often considered recurrent TB suspects in the resource-limited settings, where investigations are limited to microscopy and chest x-ray. Category II anti-TB drugs may be inappropriate and may expose patients to pill burden, drug toxicities and drug-drug interactions.

Objective

To determine the causes of pulmonary symptoms in HIV-infected smear negative recurrent pulmonary tuberculosis suspects at Mulago Hospital, Kampala.

Methods

Between March 2008 and December 2011, induced sputum samples of 178 consented HIV-infected smear negative recurrent TB suspects in Kampala were subjected to MGIT and LJ cultures for mycobacteria at TB Reference Laboratory, Kampala. Processed sputum samples were also tested by PCR to detect 18S rRNA gene of P.jirovecii and cultured for other bacteria.

Results

Bacteria, M. tuberculosis and Pneumocystis jirovecii were detected in 27%, 18% and 6.7% of patients respectively and 53.4% of the specimens had no microorganisms. S. pneumoniae, M. catarrhalis and H. influenzae were 100% susceptible to chloramphenicol and erythromycin but co-trimoxazole resistant.

Conclusion

At least 81.5% of participants had no microbiologically-confirmed TB. However our findings call for thorough investigation of HIV-infected smear negative recurrent TB suspects to guide cost effective treatment.     

Transmission and dynamics of tuberculosis on generalized households

Tuberculosis (TB) transmission is enhanced by systematic exposure to an infectious individual. This enhancement usually takes place at either the home, workplace, and/or school (generalized household). Typical epidemiological models do not incorporate the impact of generalized households on the study of disease dynamics. Models that incorporate cluster (generalized household) effects and focus on their impact on TB's transmission dynamics are developed. Detailed models that consider the effect of casual infections, that is, those generated outside a cluster, are also presented. We find expressions for the Basic Reproductive Number as a function of cluster size. The formula for R0 separates the contributions of cluster and casual infections in the generation of secondary TB infections. Relationships between cluster and classical epidemic models are discussed as well as the concept of critical cluster size.     

Associations between Antibodies to a Panel of Plasmodium falciparum Specific Antigens and Response to Sub-Optimal Antimalarial Therapy in Kampala,Uganda

Background

Antibodies are important in the control of blood stage Plasmodium falciparum infection. It is unclear which antibody responses are responsible for, or even associated with protection, partly due to confounding by heterogeneous exposure. Assessment of response to partially effective antimalarial therapy, which requires the host to assist in clearing parasites, offers an opportunity to measure protection independent of exposure.

Methods

A cohort of children aged 1–10 years in Kampala, Uganda were treated with amodiaquine+sulfadoxine-pyrimethamine for uncomplicated malaria. Serum samples from the time of malaria diagnosis and 14 days later were analyzed for total IgG to 8 P. falciparum antigens using a quantitative indirect ELISA. Associations between antibody levels and risk of treatment failure were estimated using Cox proportional hazard regression.

Results

Higher levels of antibodies to apical membrane antigen 1 (AMA-1), but to none of the other 7 antigens were significantly associated with protection against treatment failure (HR 0.57 per 10-fold increase in antibody level, CI 0.41–0.79, p = 0.001). Protection increased consistently across the entire range of antibody levels.

Conclusions

Measurement of antibody levels to AMA-1 at the time of malaria may offer a quantitative biomarker of blood stage immunity to P. falciparum, a tool which is currently lacking.     

Quantifying greenhouse gases from the production,transportation and utilization of charcoal in developing countries: a case study of Kampala,Uganda

Purpose

This study aims to quantify greenhouse gases (GHGs) from the production, transportation and utilization of charcoal and to assess the possibilities of decreasing greenhouse gases (GHGs) from the charcoal industry in general in Uganda. It also aims to assess the emission intensity of the Ugandan “charcoal production” sector compared to that of some other major charcoal producing nations.

Methods

This work was done in accordance with ISO 14040 methodology for life-cycle assessment (LCA), using GABi 4.0—a software for life-cycle assessment. A cradle-to-grave study was conducted, excluding emissions arising from machinery use during biomass cultivation and harvesting. The distance from charcoal production locations to Kampala was estimated using ArcGIS 10.0 software and a GPS tool. Emission data from a modern charcoal production process (PYREG methane-free charcoal production equipment), which complies with the German air quality standards (TA-Luft), was compared with emissions from a traditional charcoal production process. Four coupled scenarios were modelled to account for differences in the quantity of greenhouse gases emitted from the “traditional charcoal production phase”, “improved charcoal production phase (biomass feedstock sourced sustainably and unsustainably)”, “transportation phase” and “utilization phase”. Data for this study was obtained via literature review and onsite measurements.

Results and discussion

The results showed that greenhouse gases emitted due to charcoal supply and use of traditional production technique in Kampala was 1,554,699 tCO₂eq, with the transportation phase accounting for approximately 0.15 % of total greenhouse gases emitted. The utilization phase (charcoal cookstoves) emitted 723,985 tCO₂eq (46.6 %), while the charcoal production phase emitted 828,316 tCO₂eq (53.3 %). Changing the charcoal production technology from a traditional method to an improved production method (PYREG charcoal process) resulted in greenhouse gases reductions for the city of 230,747 tCO₂eq; however, by using sustainably sourced biomass, this resulted in reductions of 801,817 tCO₂eq.

Conclusions

This study showcased and quantified possible GHG emission reduction scenarios for the charcoal industry in Uganda. The result of 3 tCO₂eq emitted per tonne of charcoal produced, using earth mound method, can be applied to other countries in Eastern Africa where similar charcoal production methods are used; this will allow for somewhat better regional estimates of the inventory of greenhouse gas emissions from the production of charcoal. The results of this study also suggests that the primary use of charcoal for cooking will lead to increases in GHG emissions and increases in deforestation on the long term, if legal frameworks are not made to ensure that biomass used for charcoal production is obtained via sustainable sources or if alternative cheap energy-generating technologies for cooking are not developed and deployed to the masses.     

First Things First: Effectiveness and Scalability of a Basic Prehospital Trauma Care Program for Lay First-Responders in Kampala,Uganda

Background

We previously showed that in the absence of a formal emergency system, lay people face a heavy burden of injuries in Kampala, Uganda, and we demonstrated the feasibility of a basic prehospital trauma course for lay people. This study tests the effectiveness of this course and estimates the costs and cost-effectiveness of scaling up this training.

Methods and Findings

For six months, we prospectively followed 307 trainees (police, taxi drivers, and community leaders) who completed a one-day basic prehospital trauma care program in 2008. Cross-sectional surveys and fund of knowledge tests were used to measure their frequency of skill and supply use, reasons for not providing aid, perceived utility of the course and kit, confidence in using skills, and knowledge of first-aid. We then estimated the cost-effectiveness of scaling up the program.At six months, 188 (62%) of the trainees were followed up. Their knowledge retention remained high or increased. The mean correct score on a basic fund of knowledge test was 92%, up from 86% after initial training (n = 146 pairs, p = 0.0016). 97% of participants had used at least one skill from the course: most commonly haemorrhage control, recovery position and lifting/moving and 96% had used at least one first-aid item. Lack of knowledge was less of a barrier and trainees were significantly more confident in providing first-aid. Based on cost estimates from the World Health Organization, local injury data, and modelling from previous studies, the projected cost of scaling up this program was $0.12 per capita or $25–75 per life year saved. Key limitations of the study include small sample size, possible reporter bias, preliminary local validation of study instruments, and an indirect estimate of mortality reduction.

Conclusions

Lay first-responders effectively retained knowledge on prehospital trauma care and confidently used their first-aid skills and supplies for at least six months. The costs of scaling up this intervention to cover Kampala are very modest. This may be a cost-effective first step toward developing formal emergency services in Uganda other resource-constrained settings. Further research is needed in this critical area of trauma care in low-income countries.