Mycobacteriology

Risk factors associated with unsuppressed viral load in HIV-1 infected patients at the first antiretroviral therapy in Morocco Titou Hicham, Elkassimi Ilyas, Hanafi Tarik, Baba Noureddine, Boudi Omar, Frikh Rachid, Hjira Naoufal, Boui Mohammed International Journal of Mycobacteriology 2019 8(2):113-117 Background: Nonsuppression of viral load (VL) in HIV-infected patients on antiretroviral therapy (ART) is associated with increased HIV transmission and poor survival. The objective of this work was to evaluate the factors associated with the unsuppressed VL (VL >400 copies/ml) in HIV-1-infected patients after 6 months of the first-line ART. Methods: This was a retrospective cohort study of 181 patients living with HIV-1 on ART in Dermatology and Venereology Department of Mohamed V Military Teaching Hospital of Rabat, during the period between January 1, 2007, and January 1, 2017. Associated factors were identified through a logistic regression model. Results: Of the 181 patients, 76% were men. At inclusion, the median age was 35 years. Five variables (male sex, initial fasting glucose >1.1 g/l, alcoholism, smoking, and initial VL >10,000 copies/ml) were significantly associated (P < 0.05) with unsuppressed VL. In multivariate analysis, smoking (relative risk [RR]: 4.27, 95% confidence interval [CI]: 1.67–10.89) and initial VL >10,000 (RR: 9.78, 95% CI: 2.40–39.73) were associated independently with unsuppressed VL. Conclusion: Approximately 83% of the patients in the cohort had been able to suppress VL after 6 months of the first-line ART. Smoking and high initial VL were independent risk factors of unsuppressed VL. This work highlights the importance of developing and evaluating targeted interventions for patients at risk of unsuppressed VL on ART.

Determinants of pulmonary tuberculosis in public health facilities of Dire Dawa City, Eastern Ethiopia: Unmatched Case–control study Jafer Kezali Hassen, Andamlak Gizaw, Shikur Mohamed International Journal of Mycobacteriology 2019 8(2):118-123 Background: The Federal Ministry of Health of Ethiopia Annual performance report in 2014–2015 showed that the highest prevalence of tuberculosis (TB) case in Ethiopia was reported from Dire Dawa city which was 400/100,000 population. The aim of this study was to identify the determinants of pulmonary TB (PTB) in public health facilities of Dire Dawa city, Eastern Ethiopia. Methods: A case–control study was conducted from October to December 2017 among 95 cases and 190 controls in Dire Dawa city. Dire Dawa is one of the two chartered cities in Ethiopia like the capital city Addis Ababa. Cases and controls were identified and selected randomly from the health facilities. Data were collected using a pretested and structured questionnaire by trained data collectors. We used logistic regression to model the associations of independent variables with PTB infection. Results: PTB was associated with patients' education (no formal education vs. formal education) (adjusted odds ratio [AQR] [95% confidence interval [CI]: 3.0, [1.3, 7.1]), human immunodeficiency virus (HIV) positive status (AOR [95% CI]: 3.1: [1.1,9.1]), previous contact history with TB patient (AOR [95% CI]: 9.9 [4.3,23.0]), body mass index (BMI) of ≤18 (AOR [95% CI]: 14.9 [6.4,35.1]), and cigarette smoking history (ever vs. never) (AOR [95% CI]: 6.7 [2.3,19.5]). Conclusion: This study showed that patients' educational status, HIV status, cigarette smoking, contact history with PTB patient, and BMI were independently associated with being infected with PTB. To reduce PTB transmission, peoples should be educated on TB prevention and consequences of risky behaviors.

A rapid viability and drug-susceptibility assay utilizing mycobacteriophage as an indicator of drug susceptibilities of Anti-TB drugs against Mycobacterium smegmatis mc2 155 Gillian Catherine Crowley, Jim O'Mahony, Aidan Coffey, Riona Sayers, Paul Cotter International Journal of Mycobacteriology 2019 8(2):124-131 Background: A rapid in-house TM4 mycobacteriophage-based assay, to identify multidrug resistance against various anti-tuberculosis drugs, using the fast-growing Mycobacterium smegmatis mc2 155 in a microtiter plate format was evaluated, based on phage viability assays. Methods: A variety of parameters were optimized before the study including the minimum incubation time for the drugs, phage and M. smegmatis mc2 155 to be in contact. An increase in phage numbers over 2 h was indicative that M. smegmatis mc2 155 is resistant to the drugs under investigation, however when phage numbers remained static, M. smegmatis mc2 155 found to be sensitive to the drug. Results: The study confirmed that the data are statistically significant and that M. smegmatis mc2 155 is, in fact, sensitive to isonazid, iifampicin, pyranzaimide, and ethambutol as phage numbers doubled over 2 h (P = 0.015, 0.018, 0.014, and 0.020). The study also confirmed that M. smegmatis mc2 155 is resistant to the drugs ampicillin, erythromycin, amoxicillin streptomycin as phage numbers remain static over the same 2 h period (P = 0.028, 0.052, 0.049, and 0.04). This drug-susceptibility profiling of eight different drugs against M. smegmatis mc2 155 was detected in as little as 1½ days with a cost of ~ one euro and fifteen cent to test four drugs. Conclusion: This test is rapid to perform and will have widespread applications in drug-susceptibility testing of other members of the mycobacterial genus. In addition, the platform could also be used as a tool for high-throughput screening of novel antimycobacterial drugs. The main assets of this assay include its relatively cheap cost, versatility, and quick turnaround time.

Evaluation of GeneXpert Mycobacterium tuberculosis/Rifampin for the detection of Mycobacterium tuberculosis complex and rifampicin resistance in nonrespiratory clinical specimens Maya Habous, Maimona Ahmed E. Elimam, Rajesh Kumar, Zulfa A L. Deesi International Journal of Mycobacteriology 2019 8(2):132-137 Background: The objective of this study is to assess the performance of Xpert Mycobacterium tuberculosis (MTB)/rifampin (RIF), an automated molecular test for MTB and resistance to RIF, against smear microscopy and culture method for the diagnosis of MTB infection. Methods: This is a retrospective analysis of 168 nonrespiratory patient specimens suspected of tuberculosis (TB) at TB Laboratory of Dubai Health Authority in the United Arab Emirates between September 2016 and November 2018. Each sample underwent smear microscopy, mycobacterial culture, and GeneXpert MTB/RIF test. Results: Of 168 nonrespiratory samples, 52 samples were positive by both culture and Xpert MTB/RIF, 9 samples were detected positive only by culture. Sensitivity, specificity, positive predictive value, and negative value of the Xpert MTB/RIF test were 82.69%, 100%, 100%, and 92.80%, respectively. No false positive was yielded by the Xpert MTB/RIF, and all 116 samples were true negative by Xpert MTB/RIF. The sensitivity of the Xpert MTB/RIF was 76.92% in lymph node tissue and aspirates, 66.67% in cerebrospinal fluid, 100% in gastric lavage and aspirate, 81.25% in other body fluids, 100% in pus, 85.71% in urine, and 66.67% in other tissue samples. Of 168 strains, five strains were rifampicin resistant by phenotypic and Xpert MTB/RIF and 163 were susceptible to rifampicin with culture and Xpert MTB/RIF. Conclusion: The performance of Xpert MTB/RIF assay was comparable to the gold standard culture method for identification of MTB in nonrespiratory clinical specimens. It does not replace the gold standard culture method, but it helps to achieve better sensitivity and obtain rapid results within 2 h.

Prevalence and species diversity of nontuberculous mycobacteria in drinking water supply system of Bahía Blanca City, Argentina Alejandra Soledad Oriani, María Jimena Marfil, Martín Jose Zumárraga, Mónica Diana Baldini International Journal of Mycobacteriology 2019 8(2):138-145 Background: There is evidence that tap water is the vehicle through which nontuberculous mycobacteria (NTM) infect or colonize the human body. The objective of this study was to determine the presence and diversity of NTM in the water distribution system of Bahía Blanca city, Argentina (sites S2/S3) and in the dike that supplies water to it (S1). Methods: Culture-dependent method, biochemical tests, and molecular method (16S rRNA sequencing gene) were combined to detect and identify NTM. Results: NTM were isolated in 51.6% (64/124) of all the samples analyzed. Mycobacterium gordonae was the most frequently isolated organism (15/64) in all samples analyzed, followed by Mycobacterium peregrinum and Mycobacterium frederiksbergense. Significant differences were found in the residual chlorine values between sampling S2 and S3. In both sites, maximum counts were recorded but they did not correlate with low chlorine values. A concentration higher than 500 colony-forming unit/L of NTM was never found, which can be attributed to the negative effect caused by decontamination methods being a point to consider for the recovery of NTM. In 46.9% (30/64) of samples, both methods coincided in the identification, and the obtained sequences presented ≥99% identity. Identification at the species level was achieved in 50% (32/64) of the isolates. Nearly 17.2% (11/64) of the isolates showed a similarity <99%. Conclusions: It should be taken into account that sequencing of the 16S rRNA gene and biochemical tests are useful for the identification of several species, but it is necessary to incorporate other genes (hsp 65 and rpo B) to obtain accurate identification.

Evaluation of apoptotic protease-activating Factor-1 and cluster of Differentiation-4+ T-Cell counts in patients-infected with mycobacterium tuberculosis in Bauchi, Nigeria Muhammad Sagir Shehu, Juliana Unoifa OKpapi, Bolanle Olufunke Priscilla Musa, Idris Nasir Abdullahi, Abdurrahman Elfulaty Ahmad, Yahaya Usman International Journal of Mycobacteriology 2019 8(2):146-152 Background: This cross-sectional study evaluated Apoptotic Protease Activating Factor and cluster of differentiation-4+ (CD4+) T-cell counts in patients infected with Mycobacterium tuberculosis in Bauchi, Nigeria. Methods: This involved 180 blood samples from 90 tuberculosis (TB)-infected patients and 90 of their close contacts at home or attending Federal Medical Center Azare and Infectious Disease Hospital Bayara, Bauchi, Nigeria. The blood samples were analyzed for Apoptotic Protease Activating Factor (Apaf-1) expression using ELISA and CD4+ T cells using cyflow counter. Structured questionnaires were also used to collect the sociodemographic and clinical data of the study participants. Results: Eighty-six of the TB-infected patients had pulmonary TB (PTB), two had spine TB, and two had pleural TB. No statistically significant difference was recorded in CD4+ T-cell counts (P = 0.2935) between participants with PTB (mean ± standard deviation [SD]: 680.4 ± 235 cells/mm3) and those with extra-PTB (mean ± SD: 553.0 ± 130.5 cells/mm3). Similarly, there was no significant difference in Apaf-1 concentration (P = 0.1432) between participants with PTB (mean ± standard error of the mean [SEM]: 320.3 ± 35.4 pg/ml), and participants with extra-PTB (mean ± SEM: 143.7 ± 7.8 pg/ml). No significant difference was recorded in CD4+ T-cell counts (P = 0.4299) between the participants on treatment (mean ± SD: 758.6 ± 358.6 cells/mm3) and those who are treatment naïve (mean ± SD: 637.7 ± 208.4 cells/mm3). Similarly, there was no significant difference in Apaf-1 concentration (P = 0.6829) between the study participants on treatment (mean ± SEM: 336.3 ± 34.7 pg/ml) and those who are not on treatment (mean ± SEM: 381.2 ± 176.8 pg/ml). The CD4+ T-cells count was significantly higher in the controls (866.7 ± 288.4 cells/mm3) compared to the TB (675.0 ± 232.7 cells/mm3) patients (P < 0.0001). However, there was no significant difference in Apaf-1 expression between the control (312.4 ± 34.6 pg/ml) and the TB patients (329.1 ± 44.0 pg/ml) (P = 0.7658). Conclusion: Findings from this study showed a lower T-cell immune function during TB infection. However, Apaf-1 has no relevance on TB progression and control.

Evaluation of thin-layered agar for Mycobacterium tuberculosis isolation and drug susceptibility testing Divya Tharmalingam, Sridharan Sathyamoorthy Kopula, Kennedy Kumar Palraj International Journal of Mycobacteriology 2019 8(2):153-156 Background: Tuberculosis (TB) is India's major public health problem. According to the WHO, India harbors the largest number of cases, and TB control remains a challenge in diagnosis, drug resistance, and treatment. We undertook this study to compare the isolation rates of Mycobacterium tuberculosis (MTB) in agar, egg-based media, incidence of multidrug-resistant (MDR), and extended drug resistance (XDR) in MTB. This study aimed to compare and evaluate thin-layered agar (TLA) for the cultivation and drug susceptibility pattern of MTB with the conventional egg-based Lowenstein–Jensen's (LJ) medium and to differentiate atypical Mycobacterium by incorporating para-nitrobenzoic acid (PNB) in the TLA medium. This cross-sectional study was conducted in Sri Ramachandra Medical College and Research Institute, Porur, Chennai. Methods: A total of 68 smear-positive samples were inoculated into TLA (Middle Brook 7H 11) and LJ media with and without antibiotics (rifampicin, isoniazid, and ofloxacin) simultaneously after decontamination by the modified Petroff's method. TLA with PNB was also used to differentiate the growth of nontuberculous mycobacterium (NTM). Incubation was done at 37°C, and reading was taken every 3rd day for 6 weeks in case of TLA and for 8 weeks in case of LJ medium. Results: Out of the 68 samples, 64 (94.1%) grew in LJ, and the growth observed at the end of the 1st, 2nd, 3rd, 4th, 5th, and 6–10th weeks was 0, 12 (18.8%), 10 (15.6%), 14 (21.9%), 15 (23.4%), and 13 (20.3%), respectively. Similarly, in TLA, 65 (95.5%) samples were grown, among which 22 (33.8%) grew in the 1st week and the rest (43 [66.2%]) in the 2nd week. MDR and XDR were observed in 4 (5.8%) and 3 (4.4%) samples, respectively. Seven of them were NTM. Conclusions: TLA is a better medium, with time to positivity ranging from 1 to 2 weeks with drug susceptibility and the pattern is also comparable with LJ medium. Incorporation of PNB in TLA helps in differentiating NTM.

Laboratory diagnosis of nontuberculous mycobacteria in a Belgium Hospital Anandi Martin, Alexandre Colmant, Alexia Verroken, Hector Rodriguez-Villalobos International Journal of Mycobacteriology 2019 8(2):157-161 Background: Nontuberculous mycobacteria (NTM) have been identified in human pulmonary and extrapulmonary infections and are increasing globally, which makes it challenging to identify them. This article reports our experience with the laboratory identification of NTM in clinical practice among pulmonary and extrapulmonary samples received in our routine work. Methods: The study was conducted at the Université Catholique de Louvain at the Cliniques Universitaires Saint-Luc, Brussels, Belgium, from 2015 to 2018. A total of 386 clinical samples were collected from patients suspected of having pulmonary or extrapulmonary mycobacterial infections. Routine laboratory methods phenotypic and molecular tests were performed. Results: The majority of NTM species were isolated from pulmonary samples (68%). The most prevalent species identified were Mycobacterium chimaera_intracellulare group (32%), followed by Mycobacterium avium complex (21%), Mycobacterium abscessus complex (18%), Mycobacterium gordonae (9%), and Mycobacterium chelonae (4%). In extrapulmonary samples, M. avium and M. chimaera_intracellulare were the most frequently isolated. Conclusion: The species diversity of NTM found in our setting suggests the importance of the use of new modern methods for accurate identification of NTM at species level and in some case at subspecies level for the proper treatment and management of patients.

HbA1c levels at presentation do not impact the clinical presentation or outcomes in abdominal tuberculosis Jimil Shah, Harshal S Mandavdhare, Naresh Sachdeva, Kaushal K Prasad, Harjeet Singh, Usha Dutta, Vishal Sharma International Journal of Mycobacteriology 2019 8(2):162-165 Background: The relationship between diabetes mellitus and tuberculosis (TB) has received increasing attention, and diabetes may be associated with poor outcomes in pulmonary TB. Clinical relevance of HbA1c measurement in abdominal TB is unknown. Methods: We did a retrospective study of consecutive patients diagnosed with abdominal TB and treated with antitubercular therapy for 6 months. Patients were categorized as those with normal HbA1c (<5.7%) or those having elevated HbA1c (≥5.7%). The patients' baseline characteristics, their mode of presentation, type of abdominal TB, and their outcomes after treatment were retrieved and compared. Results: Of 84 patients included in the final analysis, the mean age was 35.52 years and 50% were male. While 19 patients had a confirmed diagnosis, 65 patients were diagnosed as probable abdominal TB. Of the 84 patients, 60 patients (71.43%) had a normal HbA1c and 24 patients (28.57%) had an elevated HbA1c. Those with elevated HbA1c were older in age (47.00 ± 13.98 vs. 30.93 ± 13.91; P < 0.001), and there were more males (75.0% vs. 40.0%; P = 0.004) as compared to the normal HbA1c group. However, other parameters such as their presenting features, pattern of abdominal TB, extra-abdominal involvement, outcomes after treatment, and need of intervention (surgery and dilatation) were comparable between both the groups. Conclusion: Presence of prediabetes or diabetes does not seem to impact the clinical presentation or outcomes in patients with abdominal TB.

Detection of Anti-Phenolic Glycolipid-I antibody in sera from tuberculosis patients in Bandung, West Java, Indonesia Hendra Gunawan, Nina Roslina, Jono Hadi Agusni, Iceu Dimas Kulsum, Kristina Makarti, Reti Hindritiani, Oki Suwarsa International Journal of Mycobacteriology 2019 8(2):166-169 Background: Mycobacterium tuberculosis (M. tuberculosis) and Mycobacterium leprae (M. leprae) are morphologically, immunologically, and pathologically similar. The incidence of simultaneous tuberculosis (TB) and leprosy is still controversial. The aim of this study was to detect anti-phenolic glycolipid-I (anti-PGL-I) antibody in sera from TB patients at Dr. Hasan Sadikin Hospital Bandung, West Java, Indonesia. The aim of this study is to detect anti-phenolic glycolipid-I (anti-PGL-I) antibody in sera from TB patients at Dr. Hasan Sadikin Hospital Bandung, West Java, Indonesia. Methods: We performed a cross-sectional descriptive study with consecutive sampling from 112 TB patients clinically diagnosed by internist from the Internal Medicine Department and confirmed through bacteriological, histological, and chest radiograph examinations. The specimens were taken from the blood serum of the patient. Furthermore, the anti-PGL-I immunoglobulin (Ig) M and IgG serum level were evaluated using the enzyme-linked immunosorbent assay. Results: The mean of anti-PGL-I IgM and IgG serum levels in TB patients of this study was 34.17 ± 21.94 pg/ml and 41.44 ± 18.93 pg/ml with the mean of optical density values was 0.18 ± 0.05 and 0.26 ± 0.07. The seropositivity of anti-PGL-I in TB patients was 27.68% for IgM and 41.96% for IgG. The seropositivity of anti-PGL-I IgM and IgG level based on clinical manifestation of TB in this study from the highest to the lowest were as follows: extrapulmonary TB patients (61.29% and 59.57%), pulmonary TB patients (29.03% and 36.17%), and pulmonary with extrapulmonary TB patients (9.68% and 4.26%), respectively. Conclusion: The seropositivity of anti-PGL-I antibody in sera from TB patients in Bandung, West Java, Indonesia was 27.68% for IgM and 41.96% for IgG. Furthermore, periodic observations are needed to determine the likelihood of clinical manifestation of leprosy in TB patients.