Getachew Aderaye

Sputum concentration improves diagnosis of tuberculosis in a setting with a high prevalence of HIV

Sputum microscopy for acid-fast bacilli (AFB), although relatively insensitive, is still the cornerstone of tuberculosis (TB) diagnosis in the developing world. Its diagnostic value has been eroded owing to the increasing number of HIV-related smear-negative pulmonary TB cases. Concentration of sputum by centrifugation after liquefaction with sodium hypochlorite is a possible means of increasing the sensitivity of direct microscopy. This procedure has been studied recently in developing countries although with conflicting results. The aim of our study, performed in 1996 in Addis Ababa, Ethiopia, was to evaluate the sensitivity of the concentration method in a large cohort of consecutive patients with suspected pulmonary TB. We show that the overall sensitivity increased from 54.2% using conventional direct microscopy to 63.1% after concentration (P < 0x0015). In HIV-positive patients, sensitivity increased from 38.5% before to 50.0% after concentration (P < 0x0034). The significant increase in yield of AFB in HIV-positive patients suggests that this method has a place in routine diagnosis of pulmonary TB in countries with a high prevalence of HIV.

Pharmacogenetic & Pharmacokinetic Biomarker for Efavirenz Based ARV and Rifampicin Based Anti-TB Drug Induced Liver Injury in TB-HIV Infected Patients

Background Implication of pharmacogenetic variations and efavirenz pharmacokinetics in concomitant efavirenz based antiviral therapy and anti-tubercular drug induced liver injury (DILI) has not been yet studied. We performed a prospective case-control association study to identify the incidence, pharmacogenetic, pharmacokinetic and biochemical predictors for anti-tubercular and antiretroviral drugs induced liver injury (DILI) in HIV and tuberculosis (TB) co-infected patients. Methods and Findings Newly diagnosed treatment naïve TB-HIV co-infected patients (n = 353) were enrolled to receive efavirenz based ART and rifampicin based anti-TB therapy, and assessed clinically and biochemically for DILI up to 56 weeks. Quantification of plasma efavirenz and 8-hydroxyefaviernz levels and genotyping for NAT2, CYP2B6, CYP3A5, ABCB1, UGT2B7 and SLCO1B1 genes were done. The incidence of DILI and identification of predictors was evaluated using survival analysis and the Cox Proportional Hazards Model. The incidence of DILI was 30.0%, or 14.5 per 1000 person-week, and that of severe was 18.4%, or 7.49 per 1000 person-week. A statistically significant association of DILI with being of the female sex (p = 0.001), higher plasma efavirenz level (p = 0.009), efavirenz/8-hydroxyefavirenz ratio (p = 0.036), baseline AST (p = 0.022), ALT (p = 0.014), lower hemoglobin (p = 0.008), and serum albumin (p = 0.007), NAT2 slow-acetylator genotype (p = 0.039) and ABCB1 3435TT genotype (p = 0.001). Conclusion We report high incidence of anti-tubercular and antiretroviral DILI in Ethiopian patients. Between patient variability in systemic efavirenz exposure and pharmacogenetic variations in NAT2, CYP2B6 and ABCB1 genes determines susceptibility to DILI in TB-HIV co-infected patients. Close monitoring of plasma efavirenz level and liver enzymes during early therapy and/or genotyping practice in HIV clinics is recommended for early identification of patients at risk of DILI.

Evaluation of Outpatients with Suspected Pulmonary Tuberculosis in a High HIV Prevalence Setting in Ethiopia: Clinical, Diagnostic and Epidemiological Characteristics

In a setting with a high prevalence of HIV we studied (i) the prevalence of pulmonary tuberculosis (PTB) and HIV; (ii) clinical and epidemiological characteristics of PTB; and (iii) the usefulness of standard procedures for diagnosing PTB. Of 509 consecutive outpatients evaluated on clinical suspicion of PTB in Addis Ababa, 33.0% were culture-verified as having PTB. PTB patients, non-TB patients and controls were HIV-1-positive in 57.1%, 38.5% and 8.3% of cases, respectively. Predictors for culture-verified PTB were age < 25 y, male gender and the presence of HIV and fever, whereas profound weight loss indicated HIV infection. Diagnosis of PTB based on clinical symptoms, sputum microscopy for acid-fast bacilli and chest radiography was sensitive (86.7%) but unspecific (64.1%). In HIV-positive patients both sensitivity and specificity were significantly lower (p < 0.05). HIV-related pulmonary infections are often misinterpreted as smear-negative PTB. HIV screening is therefore warranted not only in cases of verified TB but also as part of the diagnostic work-up in patients with respiratory symptoms suggestive of PTB. Also, increased awareness of, and improved diagnostic tools for, HIV-related pulmonary infections other than PTB are required, together with algorithms for patients with suspected PTB.

Importance of ethnicity, CYP2B6 and ABCB1 genotype for efavirenz pharmacokinetics and treatment outcomes: a parallel-group prospective cohort study in two sub-Saharan Africa populations.

Objectives We evaluated the importance of ethnicity and pharmacogenetic variations in determining efavirenz pharmacokinetics, auto-induction and immunological outcomes in two African populations. Methods ART naïve HIV patients from Ethiopia (n = 285) and Tanzania (n = 209) were prospectively enrolled in parallel to start efavirenz based HAART. CD4+ cell counts were determined at baseline, 12, 24 and 48 weeks. Plasma and intracellular efavirenz and 8-hydroxyefvairenz concentrations were determined at week 4 and 16. Genotyping for common functional CYP2B6, CYP3A5, ABCB1, UGT2B7 and SLCO1B1 variant alleles were done. Result Patient country, CYP2B6*6 and ABCB1 c.4036A>G (rs3842A>G) genotype were significant predictors of plasma and intracellular efavirenz concentration. CYP2B6*6 and ABCB1 c.4036A>G (rs3842) genotype were significantly associated with higher plasma efavirenz concentration and their allele frequencies were significantly higher in Tanzanians than Ethiopians. Tanzanians displayed significantly higher efavirenz plasma concentration at week 4 (p<0.0002) and week 16 (p = 0.006) compared to Ethiopians. Efavirenz plasma concentrations remained significantly higher in Tanzanians even after controlling for the effect of CYP2B6*6 and ABCB1 c.4036A>G genotype. Within country analyses indicated a significant decrease in the mean plasma efavirenz concentration by week 16 compared to week 4 in Tanzanians (p = 0.006), whereas no significant differences in plasma concentration over time was observed in Ethiopians (p = 0.84). Intracellular efavirenz concentration and patient country were significant predictors of CD4 gain during HAART. Conclusion We report substantial differences in efavirenz pharmacokinetics, extent of auto-induction and immunologic recovery between Ethiopian and Tanzanian HIV patients, partly but not solely, due to pharmacogenetic variations. The observed inter-ethnic variations in efavirenz plasma exposure may possibly result in varying clinical treatment outcome or adverse event profiles between populations.

High plasma efavirenz level and CYP2B6*6 are associated with efavirenz-based HAART-induced liver injury in the treatment of naïve HIV patients from Ethiopia: a prospective cohort study.

The objective of this study was to assess the incidence, timing and identify pharmacogenetic, efavirenz (EFV) pharmacokinetic and biochemical predictors of EFV-based antiretroviral therapy (ART) drug-induced liver injury (DILI). ART-naïve HIV patients (n=285) were prospectively enrolled. Pretreatment laboratory evaluations included hepatitis B surface antigen and C antibody, CD4 count and viral load. Liver tests were done at baseline, 1st, 2nd, 4th, 8th, 12th, 24th and 48th weeks during ART. Plasma EFV and 8-hydroxyefvairenz concentration was determined at week 4 using liquid chromatography–mass spectrometry. CYP2B6, CYP3A5, ABCB1 3435C/T and UGT2B7*2 genotyping was done using Taqman genotyping assay. Data were analyzed using survival analysis and Cox proportional hazards model. The incidence of DILI was 15.7% or 27.9 per 100 person-years and that of severe injury was 3.4% or 6.13 per 100 person-years. The median time for the development of DILI and severe injury was 2 and 4 weeks after initiation of ART, respectively. There was significant association of DILI with lower baseline platelet, albumin, log plasma viral load and CD4 count (P=0.031, 0.037, 0.06 and 0.019, respectively). Elevated baseline alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, plasma EFV level and CYP2B6*6 were good predictors for the development of DILI (P=0.03, 0.01, 0.016, 0.017 and 0.04, respectively). We report for the first time CYP2B6*6 as a putative genetic marker and high plasma EFV concentration as intermediate biomarker for vulnerability to EFV-induced liver injury in HIV patients. CYP2B6 genotyping and/or regular monitoring of EFV and lever enzymes level during early therapy is advised for early diagnosis and management of DILI.

Molecular Epidemiology and Drug Resistance of Mycobacterium tuberculosis Isolates from Ethiopian Pulmonary Tuberculosis Patients with and without Human Immunodeficiency Virus Infection

ABSTRACT We have analyzed the molecular epidemiology and drug resistance of 121 Mycobacterium tuberculosis isolates from consecutive patients with culture-positive pulmonary tuberculosis attending a university hospital outpatient department in Addis Ababa, Ethiopia. Restriction fragment length polymorphism analysis and spoligotyping were used to analyze the DNA fingerprinting patterns. Fifty-one (41.2%) of the isolates were found in 13 clusters with two or more identical DNA patterns. Two such clusters contained 49.0% of all clustered isolates. In a multivariate logistic regression model, human immunodeficiency virus (HIV)-positive serostatus was significantly associated with clustering of isolates for patients of both sexes (odds ratio [OR], 2.55; 95% confidence interval [CI], 1.17 to 5.80). There was a trend toward increased clustering of isolates from tuberculous women residing in Addis Ababa (OR, 2.10; 95% CI, 0.85 to 5.25). In total, 17 of 121 isolates (14.0%) were resistant to one or more of the antituberculosis drugs isoniazid (8.3%), streptomycin (7.4%), rifampin (2.5%), and ethambutol (1.7%). The high rate of drug-resistant isolates (29.6%) coincided with the peak prevalence of HIV infection (77.8%) in patients 35 to 44 years old. The majority (62.5%) of resistant isolates in this group were found within clusters. The simultaneous accumulation of certain bacterial clones in a patient population likely reflects recent transmission. Hence, we conclude that tuberculosis is commonly caused by recent infection with M. tuberculosis in HIV-positive Ethiopian patients. Furthermore, with the rapidly increasing prevalence of HIV infection in Ethiopia, the burden of tuberculosis, including drug-resistant tuberculosis, is likely to increase. Strengthening of classical tuberculosis control measures by promoting active case finding among HIV-positive adults with tuberculosis is warranted to reduce rates of transmission.

Keratin-18 and microRNA-122 complement alanine aminotransferase as novel safety biomarkers for drug-induced liver injury in two human cohorts

There is a demand for more sensitive, specific and predictive biomarkers for drug‐induced liver injury (DILI) than the gold standard used today, alanine aminotransferase (ALT). The aim of this study was to qualify novel DILI biomarkers (keratin‐18 markers M65/M30, microRNA‐122, glutamate dehydrogenase and alpha‐foetoprotein) in human DILI.

The relationship between disease pattern and disease burden by chest radiography, M. tuberculosis Load, and HIV status in patients with pulmonary tuberculosis in Addis Ababa.

Background:We evaluated the impact of HIV coinfection on the chest radiographic pattern and extent of disease and its relation to the load of Mycobacterium tuberculosis in Ethiopian out-patients with pulmonary tuberculosis.Patients and Methods:A total of 168 patients with cultureverified pulmonary tuberculosis had their chest X-rays (CXR) reviewed for the site, pattern, and extent of disease and the findings were correlated to (a) the mycobacterial culture count and bacillus load after sputum concentration and (b) the HIV status of the patients.Results:HIV-positive patients were less likely to have cavitary disease (p < 0.001) and more likely to have pleural effusion (p = 0.08), miliary (p < 0.05), and interstitial (p < 0.01) patterns. A total of 15 (9.2%) patients had normal chest X-rays. HIV-infected patients had a CXR classified as normal or with minimal involvement (p = 0.059) and a reduced mycobacterial colony count (p = 0.002) compared to HIV-negative patients. Middle and lower lung involvement were more common in HIV-positive patients.Conclusion:CXR findings in the setting of an underlying HIV infection tend to be more atypical and could present as either normal or with minimal involvement. In general, HIV-positive patients had lower colony count of M. tuberculosis than HIV-negative patients. Of particular interest is the finding of a large number of normal chest X-rays in HIV-infected patients. With the rising incidence of both tuberculosis and HIV infection in Ethiopia, the finding of a normal chest X-ray and a negative smear poses a challenge for the diagnosis of pulmonary tuberculosis.

Anti-tuberculosis therapy-induced hepatotoxicity among Ethiopian HIV-positive and negative patients.

Background To assess and compare the prevalence, severity and prognosis of anti-TB drug induced hepatotoxicity (DIH) in HIV positive and HIV negative tuberculosis (TB) patients in Ethiopia. Methodology/Principal Findings In this study, 103 HIV positive and 94 HIV negative TB patients were enrolled. All patients were evaluated for different risk factors and monitored biochemically and clinically for development of DIH. Sub-clinical hepatotoxicity was observed in 17.3% of the patients and 8 out of the 197 (4.1%) developed clinical hepatotoxicity. Seven of the 8 were HIV positive and 2 were positive for HBsAg. Conclusions/Significance Sub-clinical hepatotoxicity was significantly associated with HIV co-infection (p = 0.002), concomitant drug intake (p = 0.008), and decrease in CD4 count (p = 0.001). Stepwise restarting of anti TB treatment was also successful in almost all the patients who developed clinical DIH. We therefore conclude that anti-TB DIH is a major problem in HIV-associated TB with a decline in immune status and that there is a need for a regular biochemical and clinical follow up for those patients who are at risk.

Long-term effect of efavirenz autoinduction on plasma/peripheral blood mononuclear cell drug exposure and CD4 count is influenced by UGT2B7 and CYP2B6 genotypes among HIV patients

OBJECTIVES We investigated the long-term effect of efavirenz autoinduction on its plasma/peripheral blood mononuclear cell (PBMC) exposure and the CD4 count, and the importance of sex and pharmacogenetic variations. METHODS Treatment-naive HIV patients (n = 163) received efavirenz-based antiretroviral therapy. Plasma and intracellular (PBMC) concentrations of efavirenz and 8-hydroxyefavirenz were determined at weeks 4 and 16 of antiretroviral therapy. CD4 count was determined at baseline, and at weeks 12, 24 and 48. Genotyping for CYP2B6*6, CYP3A5*3, CYP3A5*6, CYP3A5*7, ABCB1 3435C/T and UGT2B7 (-327G→A, *2) was done. RESULTS There was a significant increase in the median plasma (32%) and intracellular (53%) 8-hydroxyefavirenz concentrations with a decrease in the efavirenz metabolic ratio (MR) (calculated by dividing the concentration of efavirenz by that of 8-hydroxyefavirenz) (20% and 5%, respectively) by week 16 compared with at week 4. While the CYP2B6 genotype significantly influenced efavirenz pharmacokinetics at weeks 4 and 16, the effect of the UGT2B7 genotype and sex was significant only at week 16. The Wilcoxon matched pairs test indicated that the change in 8-hydroxyefavirenz concentration and efavirenz MR over time was significant in females and in CYP2B6*1 and UGT2B7*1 carriers. The intracellular 8-hydroxyefavirenz level at week 16 was a negative predictor of the CD4 count at week 24 (P = 0.03) and at week 48 (P = 0.007). CYP2B6 (P = 0.02) and UGT2B7 (P = 0.05) genotypes predicted the CD4 count at week 48. Among CYP2B6*1/*1 and UGT2B7*1/*1 carriers there was no significant change in the mean CD4 count after week 24, while it continuously increased until week 48 in CYP2B6*6 and UGT2B7*2 carriers. CONCLUSIONS The effects of long-term efavirenz autoinduction on its plasma/PBMC exposure and the CD4 count over time display wide interpatient variability, partly due to sex and CYP2B6 and UGT2B7 genetic variation. Patients with the CYP2B6*1/*1 and UGT2B7*1/*1 genotypes are at risk of suboptimal immune recovery due to pronounced long-term autoinduction.