High prevalence of genotype B0/W148 of Mycobacterium tuberculosis among HIV-TB patients in Perm Krai and Irkutsk Region

Background. The population with HIV-infection plays significant role in ongoing tuberculosis pandemic. Immunosuppression due to HIV-infection is one of the causes of TB disseminated forms in this group of people. Having low immune status is also often associated with a polyclonal M. tuberculosis infection. Aim of the research: comparative assessment of epidemic genotypes of M. tuberculosis prevalence and mixed genotypes identification within HlV-TB co-infected patients in two Russian regions. Materials and methods. The DNAs of 78 clinical isolates from Irkutsk Region (IR) and 64 strains from Perm Krai (PK) have been genotyped by MIRU VNTR 24 and RD105/RD207. Strains were obtained from patients who did not have significant age and sex differences. In the PK age of the patients was 34.5 ± 0.9, in IR - 34.4 ± 1.5 years. The samples were obtained from 67.2 and 65.4 % of men, respectively. Result. The study of the M. tuberculosis indicates significant predominance of Beijing genotype strains in patients with TB-HIV of PK (92.2 %) compared to the IR (59.5 %) (х2 = 18.0; p < 0.01). The prevalence of MDR pathogens in TB-HIV patients exceeded 50 %. The mixed genotype detection in the PK and IR was high (14.1 and 12.7 % respectively). The level of virulent strains B0/W148 was 34.4 % in PK patients and 25.3 % in IR ones. Analysis of the results suggests the epidemic spread of MDR-TB in the immunocompromised individuals. Conclusions: The identified trends may indicate that Perm Kray have a process of active dissemination of transmissible strains of M. tuberculosis within HIV-infected population

ВЫЯВЛЕНИЕ МИКОБАКТЕРИЙ ТУБЕРКУЛЕЗА В КРОВИ КАК МЕТОД ДИАГНОСТИКИ ГЕНЕРАЛ ИЗОВАННОГО ТУБЕРКУЛЕЗА У ПАЦИЕНТОВ С ВИЧ-ИНФЕКЦИЕЙ

Objective: To study the informative value of the detection of mycobacteria in blood with the cultural method in patients with suspected tuberculous sepsis and to determine the most significant clinical and laboratory criteria for testing. Materials and methods: The investigation to detect M.tuberculosis was fulfilled in 159 HIV-positive patients with suspected tuberculosis sepsis. Blood culture was completed with culture medium Myco/F Lytic Culture Vials and analyzer BACTEC 9050. Results: Mycobacteria were detected in blood of 19 patients (11,9% of all patients): in 18 patients the growth of М. tuberculosis complex was detected (25,3% of all patients with diagnosed tuberculosis) and in 1 patient it was Mycobacterium avium complex (0,6% of all patients). It was shown, that the probability of M.tuberculosis detection was especially associated with the severity of the disease, immunosupression (less than 100 cells/mkl), hemoglobin quantity less than 90 g/l (levels were determined through the seeking for the most significant cutoffs). It was not proofed, that meningoencephalitis develops more often in patients with proven bacteremia. There were no evident differences in detection frequency of mycobacteria in sputum between patients with tuberculous sepsis and without it.Цель: изучить информативность культурального исследования крови на МБТ у больных ВИЧ-инфекцией с подозрением на туберкулезной сепсис и определить наиболее значимые предтестовые клинико-лабораторные критерии. Материалы и методы: обследование для выявления микобактерий в крови выполнено 159 больным ВИЧинфекцией с подозрением на туберкулезный сепсис. Посев крови производили на среду Myco/F Lytic Culture Vials и помещали в гемоанализатор BACTEC 9050. Результаты: микобактериальная инфекция кровотока выявлена у 19 пациентов (11,9% от числа тестируемых): у 18 человек обнаружен рост М. tuberculosis complex (25,3% от числа больных туберкулезом) и у одного больного – Mycobacterium avium complex (0,6% от числа тестируемых). Выявлено, что вероятность бактериемии наиболее связана с тяжестью состояния пациента; иммуносупрессией менее 100 клеток/мкл; снижением уровня гемоглобина менее 90 г/л (уровни были определены посредством поиска наиболее значимых порогов). Не удалось доказать, что менингоэнцефалит чаще развивается у лиц с подтвержденной бактериемией. Достоверных различий по частоте выявления МБТ в мокроте у больных с туберкулезным сепсисом и без него выявить не удалось

The spectrum of primary drug resistance of Mycobacterium tuberculosis in patients with tuberculosis in relation to human immunodeficiency virus status

Aim. To estimate the detection rate and spectrum of primary drug resistance of Mycobacterium tuberculosis (MBT) in patients with tuberculosis (TB) in relation to their human immunodeficiency virus (HIV) status in a region with high HIV infection rates (the Perm Territory) and to compare of drug-resistant MBT (DR-MBT) in patients with HIV/TB co-infection, by using phenotypic and molecular genetic testing (MGT) methods. Subjects and methods. The results of sputum bacteriological examination were analyzed in 178 HIV-infected patients and 354 non-HIV-infected individuals with a TB diagnosis made in the period July 1, 2014 to August 1, 2015. The diagnostic algorithm for all patients involved a duplicate sputum test for MBT by two techniques: fluorescence microscopy (FM) and inoculation into the Levenstein-Jensen dense culture medium. In patients with HIV/TB, the bacteriological examination was complemented with two more methods: detection of MBT DNA by a real-time polymerase chain reaction assay using the AmpliTube-RV system (Synthol, Russia); and inoculation into the Middlebrook liquid nutrient medium, by applying the automated BACTEC MGIT 960 system. Results. In patients with HIV/TB, the sensitivity of FM proved to be lower than in those with TB (24.2 and 32.8%, respectively; p0.05). The primary drug resistance of MBT in patients with HIV-TB was higher than that in HIV-negative individuals (60.2 and 41.6%, respectively; p<0.05). The phenotypic method (inoculation into the Levenstein-Jensen culture medium) and MGT revealed their agreement for the resistance of MBT to rifampicin (the most clinically significant drug in the choice of treatment policy) in 88.5% of the patients with HIV/TB. Conclusion. In patients with HIV/TB, the sensitivity of FM for detecting acid-resistant mycobacteria was lower than in those with TB and that of inoculations into the dense medium was comparable regardless of HIV status

Выявление микобактерий туберкулеза в мокроте у больных ВИЧ-инфекцией при использовании современного алгоритма этиологической диагностики заболевания

The objective. To establish the informativity of using phenotypic and molecular genetic methods of detection of mycobacterium tuberculosis (МВТ) from sputum to diagnose tuberculosis in patients with HIV infection. Patients and methods. 360 HIV-infected individuals with suspected tuberculosis were examined. A diagnostic algorithm included two tests of sputum for AFB/ МВТ by fluorescence microscopy, detection of МВТ DNA in real-time PCR and inoculation of liquid and solid media. Results. Tuberculosis was diagnosed in 49.4% of the examined patients. The greatest sensitivity and accuracy of МВТ detection was demonstrated by molecular genetic methods (71.3% and 0.85, respectively), which even exceeded tire results of culture diagnosis (sensitivity - 58.4 and 60.7%, accuracy - 0.79 and 0.81 for inoculation of liquid and dense media, respectively). An extremely low sensitivity was demonstrated by fluorescence microscopy (24.2%). No relation between the frequency of МВТ detection and the number of CD4+-lymphocytes was found in any method but for fluorescence microscopy showed a tendency of a better effectiveness in patients with profound immunodeficiency (in CD4 less than 100 cell/pl). Repeat analysis of inoculation of liquid media and detection of МВТ DNA (MGM) enhanced the diagnostic value of each method by 8.6 and 7.1%, respectively. In 73.5% of patients, tuberculosis was diagnosed within 2-3 days from the moment of turning for medical help. Conclusion. In general, an examination algorithm for detection of МВТ demonstrated its sensitivity in patients with co-infection (HIV/ТВ) in 87.1%. If molecular genetic methods can be used for rapid detection of a causative agent fluorescence microscopy looses its relevance for diagnosing tuberculosis.Цель. Установить информативность использования фенотипических и молекулярно-генетических методов детекции микобактерии туберкулезис (МВТ) из мокроты для диагностики туберкулеза у больных ВИЧ-инфекцией. Пациенты и методы. Обследование выполнено 360 ВИЧ-инфицированным с подозрением на туберкулез. Диагностический алгоритм включал двукратное исследование мокроты на КУМ/МБТ люминесцентной микроскопией, выявлением ДНК МВТ с помощью полимеразной цепной реакции реального времени и посевом на жидкие и плотные питательные среды. Результаты. Туберкулез установлен 49,4% пациентам от числа обследованных. Наибольшую чувствительность и точность в выявлении МБТ показали молекулярно-генетические методы (71,3% и 0,85 соответственно), которые даже превысили результаты культуральной диагностики (чувствительность - 58,4 и 60,7%, точность - 0,79 и 0,81 для посева на жидкие и плотные среды соответственно). Крайне низкую чувствительность показала люминесцентная микроскопия (24,2%). Не отмечено связи между частотой выявления МБТ и количеством СД4+-лимфоцитов ни по одному из методов, однако для люминесцентной микроскопии отмечена тенденция лучшей результативности у больных с глубоким иммунодефицитом (при CD4 менее 100 клеток/мкл). Повторный анализ посева на жидкие среды и выявления ДНК МБТ (МГМ) усилили диагностическую ценность каждого из методов на 8,6 и 7,1% соответственно. У 73,5% больных диагноз туберкулеза был установлен в течение 2-3 дней от момента обращения за помощью. Заключение. В целом алгоритм обследования для выявления МБТ продемонстрировал чувствительность у больных ко-инфекцией (ВИЧ/ТБ) 87,1%. При возможности использовать быстрые молекулярно-генетические методы детекции возбудителя люминесцентная микроскопия теряет свою значимость в диагностике туберкулеза

Detection of Mycobacterium tuberculosis in blood for diagnosis of generalised tuberculosis in HIV-positive patients

Objective: To study the informative value of the detection of mycobacteria in blood with the cultural method in patients with suspected tuberculous sepsis and to determine the most significant clinical and laboratory criteria for testing. Materials and methods: The investigation to detect M.tuberculosis was fulfilled in 159 HIV-positive patients with suspected tuberculosis sepsis. Blood culture was completed with culture medium Myco/F Lytic Culture Vials and analyzer BACTEC 9050. Results: Mycobacteria were detected in blood of 19 patients (11,9% of all patients): in 18 patients the growth of M. tuberculosis complex was detected (25,3% of all patients with diagnosed tuberculosis) and in 1 patient it was Mycobacterium avium complex (0,6% of all patients). It was shown, that the probability of M.tuberculosis detection was especially associated with the severity of the disease, immunosupression (less than 100 cells/mkl), hemoglobin quantity less than 90 g/l (levels were determined through the seeking for the most significant cutoffs). It was not proofed, that meningoencephalitis develops more often in patients with proven bacteremia. There were no evident differences in detection frequency of mycobacteria in sputum between patients with tuberculous sepsis and without it

The spectrum of primary drug resistance of Mycobacterium tuberculosis in patients with tuberculosis in relation to human immunodeficiency virus status

Aim. To estimate the detection rate and spectrum of primary drug resistance of Mycobacterium tuberculosis (MBT) in patients with tuberculosis (TB) in relation to their human immunodeficiency virus (HIV) status in a region with high HIV infection rates (the Perm Territory) and to compare of drug-resistant MBT (DR-MBT) in patients with HIV/TB co-infection, by using phenotypic and molecular genetic testing (MGT) methods. Subjects and methods. The results of sputum bacteriological examination were analyzed in 178 HIV-infected patients and 354 non-HIV-infected individuals with a TB diagnosis made in the period July 1, 2014 to August 1, 2015. The diagnostic algorithm for all patients involved a duplicate sputum test for MBT by two techniques: fluorescence microscopy (FM) and inoculation into the Levenstein-Jensen dense culture medium. In patients with HIV/TB, the bacteriological examination was complemented with two more methods: detection of MBT DNA by a real-time polymerase chain reaction assay using the AmpliTube-RV system (Synthol, Russia); and inoculation into the Middlebrook liquid nutrient medium, by applying the automated BACTEC MGIT 960 system. Results. In patients with HIV/TB, the sensitivity of FM proved to be lower than in those with TB (24.2 and 32.8%, respectively; p0.05). The primary drug resistance of MBT in patients with HIV-TB was higher than that in HIV-negative individuals (60.2 and 41.6%, respectively; p<0.05). The phenotypic method (inoculation into the Levenstein-Jensen culture medium) and MGT revealed their agreement for the resistance of MBT to rifampicin (the most clinically significant drug in the choice of treatment policy) in 88.5% of the patients with HIV/TB. Conclusion. In patients with HIV/TB, the sensitivity of FM for detecting acid-resistant mycobacteria was lower than in those with TB and that of inoculations into the dense medium was comparable regardless of HIV status

Выявление микобактерий туберкулеза в мокроте у больных ВИЧ-инфекцией при использовании современного алгоритма этиологической диагностики заболевания

The objective. To establish the informativity of using phenotypic and molecular genetic methods of detection of mycobacterium tuberculosis (МВТ) from sputum to diagnose tuberculosis in patients with HIV infection. Patients and methods. 360 HIV-infected individuals with suspected tuberculosis were examined. A diagnostic algorithm included two tests of sputum for AFB/ МВТ by fluorescence microscopy, detection of МВТ DNA in real-time PCR and inoculation of liquid and solid media. Results. Tuberculosis was diagnosed in 49.4% of the examined patients. The greatest sensitivity and accuracy of МВТ detection was demonstrated by molecular genetic methods (71.3% and 0.85, respectively), which even exceeded tire results of culture diagnosis (sensitivity - 58.4 and 60.7%, accuracy - 0.79 and 0.81 for inoculation of liquid and dense media, respectively). An extremely low sensitivity was demonstrated by fluorescence microscopy (24.2%). No relation between the frequency of МВТ detection and the number of CD4+-lymphocytes was found in any method but for fluorescence microscopy showed a tendency of a better effectiveness in patients with profound immunodeficiency (in CD4 less than 100 cell/pl). Repeat analysis of inoculation of liquid media and detection of МВТ DNA (MGM) enhanced the diagnostic value of each method by 8.6 and 7.1%, respectively. In 73.5% of patients, tuberculosis was diagnosed within 2-3 days from the moment of turning for medical help. Conclusion. In general, an examination algorithm for detection of МВТ demonstrated its sensitivity in patients with co-infection (HIV/ТВ) in 87.1%. If molecular genetic methods can be used for rapid detection of a causative agent fluorescence microscopy looses its relevance for diagnosing tuberculosis.Цель. Установить информативность использования фенотипических и молекулярно-генетических методов детекции микобактерии туберкулезис (МВТ) из мокроты для диагностики туберкулеза у больных ВИЧ-инфекцией. Пациенты и методы. Обследование выполнено 360 ВИЧ-инфицированным с подозрением на туберкулез. Диагностический алгоритм включал двукратное исследование мокроты на КУМ/МБТ люминесцентной микроскопией, выявлением ДНК МВТ с помощью полимеразной цепной реакции реального времени и посевом на жидкие и плотные питательные среды. Результаты. Туберкулез установлен 49,4% пациентам от числа обследованных. Наибольшую чувствительность и точность в выявлении МБТ показали молекулярно-генетические методы (71,3% и 0,85 соответственно), которые даже превысили результаты культуральной диагностики (чувствительность - 58,4 и 60,7%, точность - 0,79 и 0,81 для посева на жидкие и плотные среды соответственно). Крайне низкую чувствительность показала люминесцентная микроскопия (24,2%). Не отмечено связи между частотой выявления МБТ и количеством СД4+-лимфоцитов ни по одному из методов, однако для люминесцентной микроскопии отмечена тенденция лучшей результативности у больных с глубоким иммунодефицитом (при CD4 менее 100 клеток/мкл). Повторный анализ посева на жидкие среды и выявления ДНК МБТ (МГМ) усилили диагностическую ценность каждого из методов на 8,6 и 7,1% соответственно. У 73,5% больных диагноз туберкулеза был установлен в течение 2-3 дней от момента обращения за помощью. Заключение. В целом алгоритм обследования для выявления МБТ продемонстрировал чувствительность у больных ко-инфекцией (ВИЧ/ТБ) 87,1%. При возможности использовать быстрые молекулярно-генетические методы детекции возбудителя люминесцентная микроскопия теряет свою значимость в диагностике туберкулеза

Healthcare visits of HIV-infected persons before and after tuberculosis diagnosis and contamination of the environment of health-care facilities with mycobacterium tuberculosis

The objective. To study healthcare visits of HIV-infected persons before and after they were diagnosed with tuberculosis and contamination of healthcare facilities (HF) with mycobacterium tuberculosis. Materials and methods. We studied medical records of 169 HIV-infected patients who fell ill with tuberculosis. The frequency of visits of HIV-infected patients to various HF before and after tuberculosis diagnosis was determined. Molecular-genetic examination of 316 smears was performed for the presence of mycobacterium tuberculosis (MBT) DNA, taken from the working environment of four HF (AIDS Centre clinic, infectious inpatient hospital for HIV-infected persons, tuberculosis inpatient clinic, local general-duty polyclinic). Results. As has been found, before and after tuberculosis diagnosis HIV-infected persons most commonly apply for medical aid to the AIDS Centre polyclinic, more rarely to other HF. Tuberculosis in HIV-infected persons is most commonly diagnosed in the AIDS Centre polyclinic, more rarely in general-duty polyclinic, department of the infectious hospital for HIV-infected patients, somatic and surgical inpatient units. According to molecular-genetic examination of environmental smear sampling in various healthcare facilities, the maximal contamination with MBT DNA was found in the AIDS Centre polyclinic, in a less degree in the tuberculosis inpatient hospital and the infectious inpatient hospital for HIV-infected persons. In the general-duty polyclinic, no mycobacterium tuberculosis DNA was isolated. Conclusion. Predominant visits of HIV-infected persons before and after tuberculosis diagnosis to the AIDS Centre polyclinic and a higher level of contamination of healthcare environment in this facility with MBT DNA show that in the settings of a specialised polyclinic there are conditions for nosocomial infection with tuberculosis bacteria