Background

Cryptococcal disease is estimated to be responsible for significant mortality in Sub-Saharan Africa; however, only scarce epidemiology data exists. We sought to evaluate the prevalence of and risk factors for cryptococcal antigenemia in Ethiopia.

Methods

Consecutive adult HIV-infected patients from two public HIV clinics in Addis Ababa, Ethiopia were enrolled into the study. A CD4 count ≤200 cells/μl was required for study participation. Patients receiving anti-retroviral therapy (ART) were not excluded. A cryptococcal antigen test was performed for all patients along with an interview, physical exam, and medical chart abstraction. Logistic regression analysis was used to assess risk factors for cryptococcal antigenemia.

Results

369 HIV-infected patients were enrolled; mean CD4 123 cells/μl and 74% receiving ART. The overall prevalence of cryptococcal antigenemia was 8.4%; 11% in patients with a CD4 count <100 cells/μl, 8.9% with CD4 100 to 150 cells/μl and 5.7% with CD4150-200 cell/μl. 84% of patients with cryptococcal antigenemia were receiving ART. In multivariable analysis, increasing age, self reported fever, CD4 count <100 cells/μl, and site of screening were associated with an increased risk of cryptococcal antigenemia. No individual or combination of clinical symptoms had optimal sensitivity or specificity for cryptococcal antigenemia.

Conclusion

Cryptococcal antigenemia is high in Ethiopia and rapid scale up of screening programs is needed. Screening should be implemented for HIV-infected patients with low CD4 counts regardless of symptoms or receipt of ART. Further study into the effect of location and environment on cryptococcal disease is warranted.

Background

Ethiopia has the largest cattle population in Africa. The vast majority of the national herd is of indigenous zebu cattle maintained in rural areas under extensive husbandry systems. However, in response to the increasing demand for milk products and the Ethiopian government's efforts to improve productivity in the livestock sector, recent years have seen increased intensive husbandry settings holding exotic and cross breeds. This drive for increased productivity is however threatened by animal diseases that thrive under intensive settings, such as bovine tuberculosis (BTB), a disease that is already endemic in Ethiopia.

Methodology/Principal Findings

An extensive study was conducted to: estimate the prevalence of BTB in intensive dairy farms in central Ethiopia; identify associated risk factors; and characterize circulating strains of the causative agent, Mycobacterium bovis. The comparative intradermal tuberculin test (CIDT), questionnaire survey, post-mortem examination, bacteriology, and molecular typing were used to get a better understanding of the BTB prevalence among dairy farms in the study area. Based on the CIDT, our findings showed that around 30% of 2956 tested dairy cattle from 88 herds were positive for BTB while the herd prevalence was over 50%. Post-mortem examination revealed gross tuberculous lesions in 34/36 CIDT positive cattle and acid-fast bacilli were recovered from 31 animals. Molecular typing identified all isolates as M. bovis and further characterization by spoligotyping and MIRU-VNTR typing indicated low strain diversity within the study area.

Conclusions/Significance

This study showed an overall BTB herd prevalence of 50% in intensive dairy farms in Addis Ababa and surroundings, signalling an urgent need for intervention to control the disease and prevent zoonotic transmission of M. bovis to human populations consuming dairy products coming from these farms. It is suggested that government and policy makers should work together with stakeholders to design methods for the control of BTB in intensive farms in Ethiopia.

Understanding the genetic diversity of Mycobacterium tuberculosis is needed for a better understanding of the epidemiology of TB and could have implications for the development of new diagnostics, drugs, and vaccines. M. tuberculosis isolates were characterized using spoligotyping and were compared with the SpoIDB4 database of the Pasteur Institute of Guadeloupe. A total of 53 different patterns were identified among 192 isolates examined. 169 of the isolates were classified into one of the 33 shared SITs, whereas the remaining 23 corresponded to 20 orphan patterns. 54% of the isolates were ascribed to the T family, a family which has not been well defined to date. Other prominent families were CAS, Haarlem, LAM, Beijing, and Unknown comprising 26%, 13%, 2.6%, 0.5%, and 2.1%, respectively. Among HIV-positive patients, 10 patterns were observed among 25 isolates. The T (38.5%), H (26.9%), and CAS (23.1%) families were the most common among HIV-positive individuals. The diversity of the M. tuberculosis strains found in this study is very high, and there was no difference in the distribution of families in HIV-positive and HIV-negative TB patients except the H family. Tuberculosis transmission in Addis Ababa is due to only the modern M. tuberculosis families (CAS, LAM, T, Beijing, Haarlem, and U).

Background

Neglected tropical diseases (NTDs) are a group of chronic parasitic diseases and related conditions that are the most common diseases among the 2·7 billion people globally living on less than US$2 per day. In response to the growing challenge of NTDs, Ethiopia is preparing to launch a NTD Master Plan. The purpose of this review is to underscore the burden of NTDs in Ethiopia, highlight the state of current interventions, and suggest ways forward.

Results

This review indicates that NTDs are significant public health problems in Ethiopia. From the analysis reported here, Ethiopia stands out for having the largest number of NTD cases following Nigeria and the Democratic Republic of Congo. Ethiopia is estimated to have the highest burden of trachoma, podoconiosis and cutaneous leishmaniasis in sub-Saharan Africa (SSA), the second highest burden in terms of ascariasis, leprosy and visceral leishmaniasis, and the third highest burden of hookworm. Infections such as schistosomiasis, trichuriasis, lymphatic filariasis and rabies are also common. A third of Ethiopians are infected with ascariasis, one quarter is infected with trichuriasis and one in eight Ethiopians lives with hookworm or is infected with trachoma. However, despite these high burdens of infection, the control of most NTDs in Ethiopia is in its infancy. In terms of NTD control achievements, Ethiopia reached the leprosy elimination target of 1 case/10,000 population in 1999. No cases of human African trypanosomiasis have been reported since 1984. Guinea worm eradication is in its final phase. The Onchocerciasis Control Program has been making steady progress since 2001. A national blindness survey was conducted in 2006 and the trachoma program has kicked off in some regions. Lymphatic Filariasis, podoconiosis and rabies mapping are underway.

Conclusion

Ethiopia bears a significant burden of NTDs compared to other SSA countries. To achieve success in integrated control of NTDs, integrated mapping, rapid scale up of interventions and operational research into co implementation of intervention packages will be crucial.

BACKGROUND

Podoconiosis is a tropical lymphedema resulting from long-term barefoot exposure to red-clay soil derived from volcanic rock. The World Health Organization recently designated it as a neglected tropical disease. Podoconiosis develops in only a subgroup of exposed people, and studies have shown familial clustering with high heritability (63%).

METHODS

We conducted a genomewide association study of 194 case patients and 203 controls from southern Ethiopia. Findings were validated by means of family-based association testing in 202 family trios and HLA typing in 94 case patients and 94 controls.

RESULTS

We found a genomewide significant association of podoconiosis with the single-nucleotide polymorphism (SNP) rs17612858, located 5.8 kb from the HLA-DQA1 locus (in the allelic model: odds ratio, 2.44; 95% confidence interval [CI], 1.82 to 3.26; P = 1.42×10−9; and in the additive model: odds ratio, 2.19; 95% CI, 1.66 to 2.90; P = 3.44×10−8), and suggestive associations (P<1.0×10−5) with seven other SNPs in or near HLA-DQB1, HLA-DQA1, and HLA-DRB1. We confirmed these associations using family-based association testing. HLA typing showed the alleles HLA-DRB1*0701 (odds ratio, 2.00), DQA1*0201 (odds ratio, 1.91), and DQB1*0202 (odds ratio, 1.79) and the HLA-DRB1*0701–DQB1*0202 haplotype (odds ratio, 1.92) were risk variants for podoconiosis.

CONCLUSIONS

Association between variants in HLA class II loci with podoconiosis (a noncommuni-cable disease) suggests that the condition may be a T-cell–mediated inflammatory disease and is a model for gene–environment interactions that may be relevant to other complex genetic disorders. (Funded by the Wellcome Trust and others.)

Background

In northern Ethiopia the prevalence of visceral leishmaniasis is steadily rising posing an increasing public health concern. In order to develop effective control strategies on the transmission of the disease it is important to generate knowledge on the epidemiological determinants of the infection.

Methodology/Principal Findings

We conducted a cross-sectional survey on children 4–15 years of age using a multi staged stratified cluster sampling on high incidence sub-districts of Amhara regional state, Ethiopia. The survey included a socio-demographic, health and dietary questionnaire, and anthropometric measurements. We performed rK39-ICT and DAT serological tests in order to detect anti-Leishmania antibodies and carried out Leishmanin Skin Test (LST) using L.major antigen. Logistic regression models were used. Of the 565 children surveyed 56 children were positive to infection (9.9%). The individual variables that showed a positive association with infection were increasing age, being male and sleeping outside [adjusted odds ratios (95% CI): 1.15 (1.03, 1.29), 2.56 (1.19, 5.48) and 2.21 (1.03, 4.71) respectively] and in relation to the household: past history of VL in the family, living in a straw roofed house and if the family owned sheep [adjusted OR (95% CI): 2.92 (1.25, 6.81), 2.71 (1.21, 6.07) and 4.16 (1.41, 12.31) respectively].

Conclusions/Significance

A behavioural pattern like sleeping outside is determinant in the transmission of the infection in this area. Protective measures should be implemented against this identified risk activity. Results also suggest a geographical clustering and a household focalization of the infection. The behaviour of the vector in the area needs to be clarified in order to establish the role of domestic animals and house materials in the transmission of the infection.

Author Summary

Visceral leishmaniasis is a vector borne disease that can be fatal if left untreated. Its prevalence is steadily rising in northern Ethiopia posing a public health challenge in the region. We conducted a study on the factors associated to asymptomatic infection in Libo Kemkem and Fogera, Amhara regional state, where little is known about Leishmania transmission. Sleeping outside was identified as a risk activity so measures towards it are recommended. Our results also showed a geographical clustering and a household focalization of the infection although the reasons behind it are not clearly understood. More entomological studies are needed in order to clarify the vecto's behaviour in the area. Individuals living in houses that owned sheep were more likely to be infected but no association was found with other domestic animals like cattle chicken or dogs. These results add up to the debate found in the literature regarding the role of domestic animals in the transmission of Leishmania in different regions of the world. No specific recommendation should be given until the exact role of the domestic animal in the transmission cycle is clearly understood.

Epidemiological evidence from tuberculosis outbreaks revealed that some genotypes of M. tuberculosis are more transmissible and capable of causing disease than others. We analysed the plasma cytokine levels of pulmonary tuberculosis patients infected with different strains of M. tuberculosis to test the hypothesis that immune responses would be linked to the bacterial genotype. Spoligotyping was carried out for genotyping, and we used Luminex technology to measure 17 cytokines (EGF, fractalkine, GM-CSF, IFN-γ, IL-1, IL-10, IL-12, IL-17, IL-4, IL-7, IL-9, IP-10, MCP-1, MCP-3, MIP-1β, TNF, and VEGF) from plasma samples of tuberculosis patients. The levels of IL-12 (p40), IL-4, IL-7, and MIP-1beta were higher in patients infected with lineage 3, however, it was only IL-4 which showed statistically significant difference (P < 0.05) between lineage 3 and lineage 4. We further grouped the lineages into families (CAS, H and T families), and we found that the plasma level of IL-4 was significantly higher in patients infected with the CAS family (P < 0.05) in comparison with T and H families. However, there was no difference between T and H families. Therefore, the higher level of IL-4 in lineage 3 families might indicate that possible differences in the response elicited from host depend on strain lineages in the studied population.

Background

Areas endemic of helminth infection, tuberculosis (TB) and HIV are to a large extent overlapping. The aim of this study was to assess the impact of asymptomatic helminth infection on the immunological response among TB patients with and without HIV, their house hold contacts and community controls.

Methodology

Consecutive smear positive TB patients (n = 112), their household contacts (n = 71) and community controls (n = 112) were recruited in Gondar town, Ethiopia. Stool microscopy, HIV serology, serum IgE level, eosinophil and CD4 counts were performed and tuberculosis patients were followed up for 3 months after initiation of anti-TB treatment.

Results

Helminth co-infection rate was 29% in TB patients and 21% in both community control and household contacts (p = 0.3) where Ascaris lumbricoides was the most prevalent parasite. In TB patients the seroprevalence of HIV was 47% (53/112). Eosinophilia and elevated IgE level were significantly associated with asymptomatic helminth infection. During TB treatment, the worm infection rate of HIV+/TB patients declined from 31% (10/32) at week 0 to 9% (3/32) at week 2 of TB treatment, whereas HIV−/TB patients showed no change from baseline to week 2, 29% (13/45) vs. 22.2% (10/45). This trend was stable at week 8 and 12 as well.

Conclusion

One third of smear positive TB patients were infected with helminths. Eosinophilia and elevated IgE level correlated with asymptomatic worm infection, indicating an effect on host immunity. The rate of worm infection declined during TB treatment in HIV+/TB co-infected patients whereas no decline was seen in HIV−/TB group.

Background

One third of the world’s population is thought to have latent tuberculosis infection (LTBI) with the potential for subsequent reactivation of disease. To better characterize this important population, studies comparing Tuberculin Skin Test (TST) and the new interferon-γ release assays including QuantiFERON®-TB Gold In-Tube (QFT-GIT) have been conducted in different parts of the world, but most of these have been in countries with a low incidence of tuberculosis (TB). The aim of this study was therefore to evaluate the use of QFT-GIT assay as compared with TST in the diagnosis of LTBI in Ethiopia, a country with a high burden of TB and routine BCG vaccination at birth.

Methods

Healthy medical and paramedical male students at the Faculty of Medicine, Addis Ababa University, Ethiopia were enrolled into the study from December 2008 to February 2009. The TST and QFTG-IT assay were performed using standard methods.

Results

The mean age of the study participants was 20.9 years. From a total of 107 study participants, 46.7% (95%CI: 37.0% to 56.6%) had a positive TST result (TST≥10 mm), 43.9% (95%CI: 34.3% to 53.9%) had a positive QFT-GIT assay result and 44.9% (95%CI: 35.2% to 54.8%) had BCG scar. There was strong agreement between TST (TST ≥10mm) and QFT-GIT assay (Kappa = 0.83, p value = 0.000).

Conclusion

The TST and QFT-GIT assay show similar efficacy for the diagnosis of LTBI in healthy young adults residing in Ethiopia, a country with high TB incidence.

Background

Monitoring the outcome of tuberculosis treatment and understanding the specific reasons for unsuccessful treatment outcome are important in evaluating the effectiveness of tuberculosis control program. This study investigated tuberculosis treatment outcomes and predictors for unsuccessful treatment outcome in the Tigray region of Ethiopia.

Methods

Medical records of smear-positive pulmonary tuberculosis (PTB) patients registered from September 2009 to June 2011 in 15 districts of Tigray region, Northern Ethiopia, were reviewed. Additional data were collected using a structured questionnaire administered through house-to-house visits by trained nurses. Tuberculosis treatment outcomes were assessed according to WHO guidelines. The association of unsuccessful treatment outcome with socio-demographic and clinical factors was analyzed using logistic regression model.

Results

Out of the 407 PTB patients (221 males and 186 females) aged 15 years and above, 89.2% had successful and 10.8% had unsuccessful treatment outcome. In the final multivariate logistic model, the odds of unsuccessful treatment outcome was higher among patients older than 40 years of age (adj. OR = 2.50, 95% CI: 1.12-5.59), family size greater than 5 persons (adj. OR = 3.26, 95% CI: 1.43-7.44), unemployed (adj. OR = 3.10, 95% CI: 1.33-7.24) and among retreatment cases (adj. OR = 2.00, 95% CI: 1.37-2.92) as compared to their respective comparison groups.

Conclusions

Treatment outcome among smear-positive PTB patients was satisfactory in the Tigray region of Ethiopia. Nonetheless, those patients at high risk of an unfavorable treatment outcome should be identified early and given additional follow-up and social support.

Background

In Ethiopia where there is no strong surveillance system and diagnostic facilities are limited, the real burden of tuberculosis (TB) lymphadenitis is not well known. Therefore, we conducted a study to estimate the prevalence of TB lymphadenitis in Southwest Ethiopia.

Methods

A community based cross-sectional study was conducted from February to March 2009 in the Gilgel Gibe field research area. A total of 30,040 individuals 15 years or older in 10,882 households were screened for TB lymphadenitis. Any individual 15 years or older with lumps in the neck, armpits or groin up on interview were considered TB lymphadenitis suspect. The diagnosis of TB lymphadenitis was established when acid fast bacilli (AFB) smear microscopy of fine needle aspiration (FNA) sample, culture or cytology suggested TB. HIV counseling and testing was offered to all TB lymphadenitis suspects. Descriptive and bivariate analysis was done using SPSS version 15.

Results

Complete data were available for 27,597 individuals. A total of 87 TB lymphadenitis suspects were identified. Most of the TB lymphadenitis suspects were females (72.4%). Sixteen cases of TB lymphadenitis were confirmed. The prevalence of TB lymphadenitis was thus 58.0 per 100,000 people (16/27,597) (95% CI 35.7-94.2). Individuals who had a contact history with chronic coughers (OR 5.58, 95% CI 1.23-25.43) were more likely to have TB lymphadenitis. Lymph nodes with caseous FNA were more likely to be positive for TB lymphadenitis (OR 5.46, 95% CI 1.69-17.61).

Conclusion

The prevalence of TB lymphadenitis in Gilgel Gibe is similar with the WHO estimates for Ethiopia. Screening of TB lymphadenitis particularly for family members who have contact with chronic coughers is recommended. Health extension workers could be trained to screen and refer TB lymphadenitis suspects using simple methods.

Background and Objective

The relative contribution of nitric oxide (NO) to the killing of Mycobacterium tuberculosis in human tuberculosis (TB) is controversial, although this has been firmly established in rodents. Studies have demonstrated that clinical strains of M. tuberculosis differ in susceptibility to NO, but how this correlates to drug susceptibility and clinical outcome is not known.

Methods

In this study, 50 sputum smear- and culture-positive patients with pulmonary TB in Gondar, Ethiopia were included. Clinical parameters were recorded and drug susceptibility profile and spoligotyping patterns were investigated. NO susceptibility was studied by exposing the strains to the NO donor DETA/NO.

Results

Clinical isolates of M. tuberculosis showed a dose- and time-dependent response when exposed to NO. The most frequent spoligotypes found were CAS1-Delhi and T3_ETH in a total of nine known spoligotypes and four orphan patterns. There was a significant association between reduced susceptibility to NO (>10% survival after exposure to 1 mM DETA/NO) and resistance against first-line anti-TB drugs, in particular isoniazid (INH). Patients infected with strains of M. tuberculosis with reduced susceptibility to NO showed no difference in cure rate or other clinical parameters but a tendency towards lower rate of weight gain after two months of treatment, independent of antibiotic resistance.

Conclusion:

There is a correlation between resistance to first-line anti-TB drugs and reduced NO susceptibility in clinical strains of M. tuberculosis. Further studies including the mechanisms of reduced NO susceptibility are warranted and could identify targets for new therapeutic interventions.

Background

Despite the importance of the human leukocyte antigen (HLA) gene locus in research and clinical practice, direct HLA typing is laborious and expensive. Furthermore, the analysis requires specialized software and expertise which are unavailable in most developing country settings. Recently, in silico methods have been developed for predicting HLA alleles using single nucleotide polymorphisms (SNPs). However, the utility of these methods in African populations has not been systematically evaluated.

Methodology/Principal Findings

In the present study, we investigate prediction of HLA class II (HLA-DRB1 and HLA-DQB1) alleles using SNPs in the Wolaita population, southern Ethiopia. The subjects comprised 297 Ethiopians with genome-wide SNP data, of whom 188 had also been HLA typed and were used for training and testing the model. The 109 subjects with SNP data alone were used for empirical prediction using the multi-allelic gene prediction method. We evaluated accuracy of the prediction, agreement between predicted and HLA typed alleles, and discriminative ability of the prediction probability supplied by the model. We found that the model predicted intermediate (two-digit) resolution for HLA-DRB1 and HLA-DQB1 alleles at accuracy levels of 96% and 87%, respectively. All measures of performance showed high accuracy and reliability for prediction. The distribution of the majority of HLA alleles in the study was similar to that previously reported for the Oromo and Amhara ethnic groups from Ethiopia.

Conclusions/Significance

We demonstrate that HLA class II alleles can be predicted from SNP genotype data with a high level of accuracy at intermediate (two-digit) resolution in an African population. This finding offers new opportunities for HLA studies of disease epidemiology and population genetics in developing countries.

Bovine tuberculosis in domestic livestock such as cattle is an economically important disease with zoonotic potential, particularly in countries with emerging economies. We discuss the findings of recent epidemiological and immunological studies conducted in Ethiopia on host susceptibility differences between native zebu and the exotic Holstein–Friesian cattle that are increasingly part of the Ethiopian National herd, due to the drive to increase milk yields. These findings support the hypothesis that native Zebu cattle are more resistant to bovine tuberculosis. We also summarise the results of experimental infections that support the epidemiological data, and of laboratory experiments that suggest a role for the innate immune response, and in particular interleukin-6, in the outcome of bovine tuberculosis infection.

Despite huge global efforts in tuberculosis (TB) control, pastoral areas remain under-investigated. During two years sputum and fine needle aspirate (FNA) specimens were collected from 260 Ethiopian pastoralists of Oromia and Somali Regional States with suspected pulmonary TB and from 32 cases with suspected TB lymphadenitis. In parallel, 207 suspected tuberculous lesions were collected from cattle, camels and goats at abattoirs. All specimens were processed and cultured for mycobacteria; samples with acid-fast stained bacilli (AFB) were further characterized by molecular methods including genus and deletion typing as well as spoligotyping. Non-tuberculous mycobacteria (NTM) were sequenced at the 16S rDNA locus. Culturing of AFB from human sputum and FNA samples gave a yield of 174 (67%) and 9 (28%) isolates, respectively. Molecular typing was performed on 173 of these isolates and 160 were confirmed as Mycobacterium tuberculosis, three as M. bovis, and the remaining 10 were typed as NTMs. Similarly, 48 AFB isolates (23%) yielded from tuberculous lesions of livestock, of which 39 were molecular typed, including 24 M. bovis and 4 NTMs from cattle, 1 M. tuberculosis and 1 NTM from camels and 9 NTMs from goats. Isolation of M. bovis from humans and M. tuberculosis from livestock suggests transmission between livestock and humans in the pastoral areas of South-East Ethiopia

TB remains a major public health problem despite all the efforts that have been made since it was declared a global emergency in 1993. Different strategies have been implemented to curb the spread of the epidemic. Early case detection and treatment is one of the pillars of the TB control program. In 1991, WHO set targets for increasing case detection and treatment success rates to 70% and 85% respectively. Although the target of treatment success rate has been achieved, the case detection rate remains far below target at currently less than 50%. It is high time that control programs move from simple passive to a more systematic active case finding in order to accelerate TB control.

Background

In Ethiopia where there is no strong surveillance system and state of the art diagnostic facilities are limited, the real burden of tuberculosis (TB) is not well known. We conducted a community based survey to estimate the prevalence of pulmonary TB and spoligotype pattern of the Mycobacterium tuberculosis isolates in Southwest Ethiopia.

Methods

A total of 30040 adults in 10882 households were screened for pulmonary TB in Gilgel Gibe field research centre in Southwest Ethiopia. A total of 482 TB suspects were identified and smear microscopy and culture was done for 428 TB suspects. Counseling and testing for HIV/AIDS was done for all TB suspects. Spoligotyping was done to characterize the Mycobacterium tuberculosis isolates.

Results

Majority of the TB suspects were females (60.7%) and non-literates (83.6%). Using smear microscopy, a total of 5 new and 4 old cases of pulmonary TB cases were identified making the prevalence of TB 30 per 100,000. However, using the culture method, we identified 17 new cases with a prevalence of 76.1 per 100,000. There were 4.3 undiagnosed pulmonary TB cases for every TB case who was diagnosed through the passive case detection mechanism in the health facility. Eleven isolates (64.7%) belonged to the six previously known spoligotypes: T, Haarlem and Central-Asian (CAS). Six new spoligotype patterns of Mycobacterium tuberculosis, not present in the international database (SpolDB4) were identified. None of the rural residents was HIV infected and only 5 (5.5%) of the urban TB suspects were positive for HIV.

Conclusion

The prevalence of TB in the rural community of Southwest Ethiopia is low. There are large numbers of undiagnosed TB cases in the community. However, the number of sputum smear-positive cases was very low and therefore the risk of transmitting the infection to others may be limited. Active case finding through health extension workers in the community can improve the low case detection rate in Ethiopia. A large scale study on the genotyping of Mycobacterium tuberculosis in Ethiopia is crucial to understand transmission dynamics, identification of drug resistant strains and design preventive strategies.

The stable incidence of new leprosy cases suggests that transmission of infection continues despite worldwide implementation of MDT. Thus, specific tools are needed to diagnose early stage Mycobacterium leprae infection, the likely sources of transmission. M. leprae antigens that induce T-cell responses in M. leprae exposed and/or infected individuals thus are major targets for new diagnostic tools. Previously, we showed that ML1601c was immunogenic in patients and healthy household contacts (HHC). However, some endemic controls (EC) also recognized this protein. To improve the diagnostic potential, IFN-γ responses to ML1601c peptides were assessed using PBMC from Brazilian leprosy patients and EC. Five ML1601c peptides only induced IFN-γ in patients and HHC. Moreover, 24-hour whole-blood assay (WBA), two ML1601c peptides could assess the level of M. leprae exposure in Ethiopian EC. Beside IFN-γ, also IP-10, IL-6, IL-1β, TNF-α, and MCP-1 were increased in EC from areas with high leprosy prevalence in response to these ML1601c peptides. Thus, ML1601c peptides may be useful for differentiating M. leprae exposed or infected individuals and can also be used to indicate the magnitude of M. leprae transmission even in the context of various HLA alleles as present in these different genetic backgrounds.

A cross-sectional study of bovine tuberculosis (BTB) detected by the comparative intradermal tuberculin test (CIDT) was conducted in livestock of the Somali region in southeast Ethiopia—in four pastoral associations from January to August 2009. In 94 herds, each of 15 cattle, camels, and goats was tested per herd leading to a total of 1,418 CIDT tested animals, with 421 cattle, 479 camels, and 518 goats. A herd was considered positive if it had at least one reactor. Prevalence per animal species was calculated using a xtgee model for each species. The individual animal prevalence was 2.0% [95% confidence interval (CI), 0.5–8.4], 0.4% (95% CI, 0.1–3%), and 0.2% (95% CI, 0.03–1.3) in cattle, camels, and goats, respectively. Prevalence of avian mycobacterium purified protein derivative (PPD) reactors in cattle, camels, and goats was 0.7% (95% CI, 0.2–2.0%), 10.0% (95% CI, 7.0–14.0%), and 1.9 (95% CI, 0.9–4.0%), respectively, whereby camels had an odds ratio of 16.5 (95% CI, 5.0–55.0) when compared to cattle. There was no significant difference between livestock species in BTB positivity. In the present study, the prevalence of bovine tuberculosis was low in Somali pastoral livestock in general and in camels and goats in particular. The high proportion of camel reactors to avian PPD needs further investigation of its impact on camel production.

Background

Ethiopia has had mechanisms for TB/HIV collaborative activities since 2002. However, no published account has defined the role of these collaborative efforts in strengthening linkages between HIV and TB management units at the point-of-care level. Our objective was to assess the extent of linkages between the two programs at the patient management level at Zewditu Memorial Hospital in Addis Ababa, Ethiopia. Between January and December 2008, the registers of 241 TB patients were reviewed to determine the HIV testing rate, the treatment charts of 238 randomly selected patients were reviewed for providers' compliance with evaluation criteria, and exit interviews were conducted with 309 TB/HIV co-infected clients to validate providers' compliance.

Results

From register review, it was determined that the HIV testing acceptance rate was 95%, and that 70% of patients received post-test counseling. A review of the patient chart revealed that of 51 patients with a complaint of cough, duration for cough was recorded in 35 (68.6%) cases and cough > 2 weeks was recorded in 25 (49.0%) cases. Seventy two percent (18 of 25) were linked for sputum microscopy. Linkage to cotrimoxazole prophylactic treatment was 81%, but only 47% of eligible patients were linked to isoniazid preventive therapy (IPT). Correct diagnosis was accomplished at a rate of 100% for smear positive pulmonary TB, 23% for smear negative pulmonary TB and 88% for extra pulmonary TB patients. Both chart review and exit interviews indicated that history of TB contact and cough > 2 weeks predicted TB disease.

Conclusion

The rates of HIV testing and linkage to cotrimoxazole prophylactic therapy were high. Improvement is needed in the areas of recording patient information, screening HIV positives for TB, initiation of IPT, referral, linkages, and TB diagnostic capacity.

Background

Bovine tuberculosis (BTB) is endemic in Ethiopian cattle. The aim of this study was to assess BTB prevalence at an intensive contact interface in Meskan Woreda (district) in cattle, small ruminants and suspected TB-lymphadenitis (TBLN) human patients.

Methods

The comparative intradermal test (CIDT) was carried out for all animals involved in the cross-sectional study and results interpreted using a > 4 mm and a > 2 mm cut-off. One PPD positive goat was slaughtered and lymph nodes subjected to culture and molecular typing. In the same villages, people with lymphadenitis were subjected to clinical examination. Fine needle aspirates (FNA) were taken from suspected TBLN and analyzed by smear microscopy and molecular typing.

Results

A total of 1214 cattle and 406 small ruminants were tested for BTB. In cattle, overall individual prevalence (> 2 mm cut-off) was 6.8% (CI: 5.4-8.5%) with 100% herd prevalence. Only three small ruminants (2 sheep and 1 goat) were reactors. The overall individual prevalence in small ruminants (> 2 mm cut-off) was 0.4% (CI: 0.03-5.1%) with 25% herd prevalence. Cattle from owners with PPD positive small ruminants were all PPD negative. 83% of the owners kept their sheep and goats inside their house at night and 5% drank regularly goat milk.

FNAs were taken from 33 TBLN suspected cases out of a total of 127 screened individuals with lymph node swellings. Based on cytology results, 12 were confirmed TBLN cases. Nine out of 33 cultures were AFB positive. Culture positive samples were subjected to molecular typing and they all yielded M. tuberculosis. M. tuberculosis was also isolated from the goat that was slaughtered.

Conclusions

This study highlighted a low BTB prevalence in sheep and goats despite intensive contact with cattle reactors. TBLN in humans was caused entirely by M. tuberculosis, the human pathogen. M. tuberculosis seems to circulate also in livestock but their role at the interface is unknown.

We have identified a clonal complex of Mycobacterium bovis isolated at high frequency from cattle in Uganda, Burundi, Tanzania, and Ethiopia. We have named this related group of M. bovis strains the African 2 (Af2) clonal complex of M. bovis. Af2 strains are defined by a specific chromosomal deletion (RDAf2) and can be identified by the absence of spacers 3 to 7 in their spoligotype patterns. Deletion analysis of M. bovis isolates from Algeria, Mali, Chad, Nigeria, Cameroon, South Africa, and Mozambique did not identify any strains of the Af2 clonal complex, suggesting that this clonal complex of M. bovis is localized in East Africa. The specific spoligotype pattern of the Af2 clonal complex was rarely identified among isolates from outside Africa, and the few isolates that were found and tested were intact at the RDAf2 locus. We conclude that the Af2 clonal complex is localized to cattle in East Africa. We found that strains of the Af2 clonal complex of M. bovis have, in general, four or more copies of the insertion sequence IS6110, in contrast to the majority of M. bovis strains isolated from cattle, which are thought to carry only one or a few copies.

This study, nested within a clinical trial, by Luis Cuevas and colleagues finds that LED-FM microscopy has higher sensitivity but lower specificity than Zn microscopy for detecting tuberculosis in sputum samples.

Background

The diagnosis of tuberculosis (TB) in resource-limited settings relies on Ziehl-Neelsen (ZN) smear microscopy. LED fluorescence microscopy (LED-FM) has many potential advantages over ZN smear microscopy, but requires evaluation in the field. The aim of this study was to assess the sensitivity/specificity of LED-FM for the diagnosis of pulmonary TB and whether its performance varies with the timing of specimen collection.

Methods and Findings

Adults with cough ≥2 wk were enrolled consecutively in Ethiopia, Nepal, Nigeria, and Yemen. Sputum specimens were examined by ZN smear microscopy and LED-FM and compared with culture as the reference standard. Specimens were collected using a spot-morning-spot (SMS) or spot-spot-morning (SSM) scheme to explore whether the collection of the first two smears at the health care facility (i.e., “on the spot”) the first day of consultation followed by a morning sample the next day (SSM) would identify similar numbers of smear-positive patients as smears collected via the SMS scheme (i.e., one on-the-spot-smear the first day, followed by a morning specimen collected at home and a second on-the-spot sample the second day). In total, 529 (21.6%) culture-positive and 1,826 (74.6%) culture-negative patients were enrolled, of which 1,156 (49%) submitted SSM specimens and 1,199 (51%) submitted SMS specimens. Single LED-FM smears had higher sensitivity but lower specificity than single ZN smears. Using two LED-FM or two ZN smears per patient was 72.8% (385/529, 95% CI 68.8%–76.5%) and 65.8% (348/529, 95% CI 61.6%–69.8%) sensitive (p<0.001) and 90.9% (1,660/1,826, 95% CI 89.5%–92.2%) and 98% (1,790/1,826, 95% CI 97.3%–98.6%) specific (p<0.001). Using three LED-FM or three ZN smears per patient was 77% (408/529, 95% CI 73.3%–80.6%) and 70.5% (373/529, 95% CI 66.4%–74.4%, p<0.001) sensitive and 88.1% (95% CI 86.5%–89.6%) and 96.5% (95% CI 96.8%–98.2%, p<0.001) specific. The sensitivity/specificity of ZN smear microscopy and LED-FM did not vary between SMS and SSM.

Conclusions

LED-FM had higher sensitivity but, in this study, lower specificity than ZN smear microscopy for diagnosis of pulmonary TB. Performance was independent of the scheme used for collecting specimens. The introduction of LED-FM needs to be accompanied by appropriate training, quality management, and monitoring of performance in the field.

Trial Registration

Current Controlled Trials ISRCTN53339491

Please see later in the article for the Editors' Summary

Editors' Summary

Background

Tuberculosis is a global public health problem. Every year, about 1.7 million people die from this contagious bacterial infection, and about 9 million new cases occur, mainly in low- and middle-income countries. Mycobacterium tuberculosis, which causes tuberculosis, is spread in airborne droplets when people with the disease cough or sneeze, and usually infects the lungs (pulmonary tuberculosis). Symptoms of tuberculosis include a persistent cough, weight loss, and night sweats. Because tuberculosis is easily transmitted and potentially deadly, it is important that it is diagnosed quickly and accurately and immediately treated. The “gold standard” diagnostic test for tuberculosis is mycobacterial culture (in liquid or solid medium), in which laboratory technicians try to grow M. tuberculosis from sputum (mucus brought up from the lungs by coughing). However, this test is expensive, so most patients suspected of having pulmonary tuberculosis in resource-limited countries are investigated using sputum smear microscopy. In this cheaper but less sensitive test, sputum samples are “smeared” onto microscope slides, stained with Ziehl-Neelsen (ZN) dye, and then examined with a microscope for the presence of M. tuberculosis.

Why Was This Study Done?

With smear microscopy, multiple samples have to be examined to increase the test's sensitivity (the proportion of patients with culture-positive tuberculosis that the test detects). Because each smear examination takes up to 10 minutes, tuberculosis diagnosis with ZN smear microscopy creates a large laboratory workload. A variant form of smear microscopy—light-emitting-diode fluorescence microscopy (LED-FM)—could reduce this workload. With LED-FM, smears stained with a fluorescent dye can be examined in a quarter of the time it takes to examine ZN smears. In this study, the researchers evaluate the sensitivity and specificity (the proportion of people with a negative smear among people without tuberculosis; a high specificity indicates a low false-positive rate) of LED-FM using samples collected in a trial undertaken in four resource-limited countries (Ethiopia, Nepal, Nigeria, and Yemen) to investigate two schemes for sputum sample collection. In the spot-morning-spot (SMS) scheme, patients provide an on-the-spot specimen at their initial consultation, a specimen collected at home the next morning, and a second on-the-spot sample when they deliver their morning specimen. In the spot-spot-morning (SSM) scheme, patients provide two on-the-spot samples during their first clinic visit and a sample collected at home the next morning.

What Did the Researchers Do and Find?

In the main trial, the researchers collected sputum samples using the SMS or SSM scheme from 6,627 patients with a cough lasting more than two weeks. For their investigation of LED-FM, they examined nearly 2,400 samples (half SSM and half SMS specimens, about a quarter of which were tuberculosis culture-positive) with both ZN smear microscopy and LED-FM and determined the sensitivity and specificity of both tests—with one, two, or three sputum samples per patient—relative to mycobacterial solid culture. Single LED-FM smears had higher sensitivity but lower specificity than single ZN smears. The sensitivities of two LED-FM and two ZN smears were 72.8% and 65.8%, respectively; the specificities of these tests were 90.9% and 98.0%. The sensitivities of three LED-FM and three ZN smears were 77% and 70.5%, respectively; the specificities of these tests were 88.1% and 96.5%. The sensitivity and specificity of both tests was similar for samples collected using the SMS and the SSM schemes.

What Do These Findings Mean?

These findings show that in the resource-limited countries included in this trial, LED-FM has a higher sensitivity but lower specificity than ZN smear microscopy. The researchers calculate that in this study the accuracy of three LED-FM examinations was 85% (2,017 out of 2,355 patients were correctly classified as infected or uninfected), whereas the accuracy of three ZN smears was 91.8%. Thus, although LED-FM should identify more people with tuberculosis than ZN smear microscopy, because of its lower specificity, its use might also lead to more people without tuberculosis being needlessly treated for the disease. Nevertheless, provided that the introduction of LED-FM is accompanied by appropriate training and performance monitoring, LED-FM is an attractive potential tool for the laboratory diagnosis of tuberculosis that, together with a move towards the collection of two on-the-spot smears in a single clinic visit, could ensure that poor patients have access to timely tuberculosis diagnosis and prompt treatment.

Additional Information

Please access these Web sites via the online version of this summary at http://dx.doi.org/10.1371/journal.pmed.1001057.

Details of the parent trial in which the samples used in this study were collected are available in a PLoS Medicine Research Article by Cuevas et al.

The World Health Organization provides information on all aspects of tuberculosis, including information on tuberculosis diagnostics; recent WHO policy statements on diagnosis of tuberculosis are available; the Stop TB Partnership provides information on global tuberculosis control (some information in several languages)

The US Centers for Disease Control and Prevention has information about tuberculosis, including information on the diagnosis of tuberculosis disease

The US National Institute of Allergy and Infectious Diseases also has detailed information on all aspects of tuberculosis

MedlinePlus has links to further information about tuberculosis (in English and Spanish)

A new Web site dedicated to the discussion and optimization of smear microscopy has recently been launched