First-line Drug Susceptibility Testing Research Articles

Active tuberculosis case finding and detection of drug resistance among HIV-infected patients: A cross-sectional study in a TB endemic area, Gondar, Northwest Ethiopia

BackgroundTuberculosis (TB) patients co-infected with human immunodeficiency virus (HIV) often lack the classic symptoms of pulmonary tuberculosis, making the diagnosis difficult. Current practices in resource-limited settings often indicate that these co-infected patients are diagnosed when they clinically manifest disease symptoms, resulting in a delayed diagnosis and despite continued transmission. The aim of this study is to determine the prevalence of undiagnosed pulmonary tuberculosis cases through active case finding and including multidrug-resistant TB (MDR-TB) among HIV-infected patients. Materials and methodsA total of 250 HIV-infected patients, aged 18years and above were evaluated in a cross-sectional design between February 2012 and November 2012. Socio-demographic and clinical data were collected using a structured questionnaire. Sputum samples were collected from all participants for acid fast bacilli (AFB) direct smear microscopy and Mycobacteria culture. A PCR-based RD9 deletion and genus typing, as well as first-line anti-TB drug susceptibility testing, was performed for all culture-positive isolates. ResultsFollowing active TB case finding, a total of 15/250 (6%) cases were diagnosed as TB cases, of whom 9/250 (3.6%) were detected by both smear microscopy and culture and the remaining 6/250 (2.4%) only by culture. All the 15 isolates were typed through RD9 typing of which 10 were Mycobacterium tuberculosis species; 1 belonged to Mycobacterium genus and 4 isolates were non-tuberculous mycobacteria. The prevalence of undiagnosed pulmonary TB disease among the study participants was 4.4%, which implies the possibility of identifying even more undiagnosed cases through active case finding. A multivariate logistic regression showed a statistically significant association between the presence of pneumonia infection and the occurrence of TB (OR=4.81, 95% CI (1.08–21.43), p=0.04). In addition, all the isolates were sensitive to all first-line anti-TB drugs, except for streptomycin, seen in only one newly diagnosed TB patient, and MDR-TB was not detected. ConclusionThe prevalence of undiagnosed pulmonary TB infection among HIV-infected patients in Gondar was 4.4%. Additionally, the possibility of these undiagnosed TB cases in the community could also pose a risk for the transmission of the disease, particularly among family members. Active screening of known HIV-infected individuals, with at least one TB symptom is recommended, even in persons with opportunistic infections.

Genotyping and clinical characteristics of multidrug and extensively drug-resistant tuberculosis in a tertiary care tuberculosis hospital in China

BackgroundThere is a lack of information on the clinical characteristics of multidrug-resistant (MDR) tuberculosis (TB) and extensively drug-resistant (XDR) TB in the Jiangxi Province of China; furthermore, data have not been reported on the utility of mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) analyses in genotyping Mycobacterium tuberculosis strains isolated from this region. The aim of this study was to analyse the clinical features of patients with MDR and XDR TB from Jiangxi Province and to evaluate the discriminatory power of the 15-loci MIRU-VNTR method.MethodsA retrospective study was conducted on patients diagnosed with MDR and XDR TB at the Jiangxi Chest Hospital from July 2010 to June 2011. The RD105 deletion-targeted multiplex PCR (DTM-PCR) and the 15-loci MIRU-VNTR method were used to determine the genetic background of the identified MDR and XDR M. tuberculosis clinical isolates.ResultsOf 804 M. tuberculosis clinical isolates, 159 (159/804, 19.8%) of the isolates were identified as MDR with first-line drug susceptibility testing. Of the 123 available MDR isolates, 13 (13/123, 10.6%) were XDR. The RD105 deletion-targeted multiplex PCR method identified 85 (85/110, 77.3%) MDR and 12 (12/13, 92.3%) XDR isolates as the Beijing genotype. MIRU-VNTR cluster analysis demonstrated that 101 MDR and 13 XDR strains had unique genotype patterns; the remaining 9 MDR strains were in 4 clusters, namely 1 cluster with 3 strains and 3 clusters with 2 strains, resulting in a low clustering rate (4.06%). The Hunter-Gaston discriminatory index (HGDI) of the 15-loci MIRU-VNTR method was as high as 0.992. In addition, clinical surveys showed that 87 (87/110, 79.1%) MDR TB patients and 10 (10/13, 76.9%) XDR TB patients had been previously treated. Diabetes mellitus was the most common comorbidity in both MDR TB (16/110, 14.5%) and XDR TB (2/13, 15.4%) patients.ConclusionsBased on our preliminary data, the MDR and XDR M. tuberculosis clinical isolates identified at the Jiangxi Chest Hospital were genetically diverse and clustered at a low frequency. The 15-loci MIRU-VNTR method showed high discriminatory power and may be used as a first-line genotyping tool in investigating the molecular epidemiology of M. tuberculosis in Jiangxi, China. Decisive measures are urgently needed to effectively prevent and manage MDR and XDR tuberculosis in this province.

Prevalence of Multidrug-resistant Tuberculosis Among Newly Diagnosed Cases of Sputum-positive Pulmonary Tuberculosis from MGM Hospital Warangal

In spite of availability of effective chemotherapy and Bacille-Calmette-Guerian (BCG) vaccine, tuberculosis remains a leading infectious killer worldwide. Tuberculosis (TB) is presently recognized as one of the most common opportunistic infections seen in HIV seropositive patients, mostly presenting in the form of pulmonary and extrapulmonary infections. The prevalence of multidrug-resistant tuberculosis (MDRTB) is increasing throughout the world. Although previous treatment for TB is the most important risk factor for development of MDR-TB, A total of 100 clinically diagnosed and radiologically evident cases suggestive of pulmonary tuberculosis were selected for study. A total of 27 samples were found to be culture positive and 73 were culture negative, of these 26 stains were identified as Mycobacterium tuberculosis, one was identified as M.avium complex (MAC). All sputum-positive TB cases were subjected to mycobacterial culture and first-line drug-susceptibility testing (DST). MDR-TB was defined as TB caused by bacilli showing resistance to at least isoniazid and rifampicin. All the 27 isolates were concurrently subjected for minimal inhibitory concentration (MIC), Proportion method and BACTEC. Out of 27 cultures tested by the MIC methods, resistance was observed in 99.9 percent at MIC of 128 mg/l. In proportion method five isolate were shown resistance to Isoniazid (H); Rifampicin (R); but shown sensitive to other drugs Ethambutol (E); Pyrazinamide and Streptomycin.

Inconsistencies in drug susceptibility testing of Mycobacterium tuberculosis: Current riddles and recommendations.

Drug susceptibility testing (DST) of Mycobacterium tuberculosis is a crucial procedure to determine the effective drug regimen for patients' treatment. Reporting of erroneous DST results to the treating physician has adulterous effects on patients. As a first study of its type, the inconsistencies in reporting DST results of rifampicin and isoniazid from Saudi Arabia were assessed. An automated liquid culture-based DST and a molecular mutation detection technique were used. Performance of first-line drug susceptibility testing of 1904 clinical isolates showed 44 inconsistent results. The majority of the cases reported as MDR-TB from the referral laboratories could not reproduce the same results at a different site (Mycobacteriology Research Section). Of the 44 cases, 16 (36.3%) showed false resistance to isoniazid and rifampicin and on the other hand, 14 (31.8%) cases showed false susceptibility to the same drugs. The possible causes for the inconsistencies and recommendations to overcome the biases based on this experience are discussed.

Drug Resistance in Mycobacterial Isolates From Meningitis Cases

To the Editors: In countries with high burden of pulmonary tuberculosis (TB), the incidence of extrapulmonary TB (EPTB) is high. There has been a rising trend of tuberculous meningitis (TBM) in developing countries in the past 2 decades. There are few published reports on drug susceptibility profiling of EPTB, particularly TBM.1 Early and rapid detection of drug resistance in TBM is not always possible because bacterial load in cerebrospinal fluid (CSF) is too low to detect the resistance by both culture and molecular methods. Hence, it is important to have surveillance of drug resistance in EPTB to help in planning the treatment regimen. Here, we report the prevalence of drug resistance in Mycobacterium tuberculosis isolates from CSF of TBM cases. CSF samples from 370 clinically suspected pediatric TBM cases were referred to the TB laboratory, Department of Microbiology, Chhatrapati Shahuji Maharaj Medical University, Lucknow, India, for laboratory confirmation of TBM. Each CSF sample was processed for acid-fast bacilli smear examination, mycobacterium culture2 and demonstration of mycobacterial DNA by in-house nucleic acid amplification method.3 All M. tuberculosis isolates were subjected to drug susceptibility testing for first-line drugs: isoniazid, rifampicin, streptomycin and ethambutol by conventional 1% proportion method.2 The laboratory is under external quality control for first-line drug susceptibility testing (provided by the National JALMA Institute for Leprosy and Other Mycobacterial Diseases, Agra, India). Of the 370 cases, 106 (28.6%) patients had laboratory-proven TBM. CSF samples from all 106 laboratory-proven patients were polymerase chain reaction positive whereas 60 (16.2%) patients were positive by smear examination for acid-fast bacilli and 51 (13.7%) patients were culture positive for M. tuberculosis. Among 106 samples, 32 (30.2%) samples were positive by all the 3 tested methods. Of the 51 M. tuberculosis isolates, 34 (66.7%) isolates were pan-susceptible whereas resistance to isoniazid or streptomycin was detected in 7 (13.7%) and 5 (9.8%) isolates, respectively. Multidrug resistance (MDR) was detected in 1 isolate. Almost 50% of MDR-TB pulmonary cases worldwide are estimated to occur in China and India. Drug resistance in M. tuberculosis isolates from TBM cases is low despite the fact that, among pulmonary cases, prevalence of MDR in our setting was higher (19.8%)4 than other parts of the country.5 MDR among cases of pulmonary TB is well documented, but reports on MDR–EPTB and MDR–TBM are limited. Continuous surveillance of drug resistance in EPTB is needed. Amita Jain, MD Pratima Dixit, MSc Indu Jaiswal, MSc Department of Microbiology C.S.M. Medical University Ravindra Kumar Garg, MD, DM Department of Neurology C.S.M. Medical University Rashmi Kumar, MD Department of Pediatrics, C.S.M. Medical University Lucknow, India

Proficiency testing of first- and second-line anti-tuberculosis drugs in Italy

To the Editors: The emergence of drug-resistant tuberculosis (TB) is an increasing threat to public health in industrialised countries; thus, it is important to supervise mycobacteriology laboratories by performing periodic proficiency of anti-TB drug susceptibility testing (DST). In 1994, the World Health Organization (WHO) and the International Union against Tuberculosis and Lung Diseases developed a global project of anti-TB drug resistance surveillance to assist countries via a network of supranational reference laboratories (SRLs). Proficiency test (PT) results of first-line anti-TB drugs have been reported for the SRL network [1] and for some individual countries [2, 3]. The SRL in Rome, Italy, coordinated two PTs of first-line drugs in endemic countries in 2002–2006 [4] and two PTs of first-line drugs in Italy in 1998–2000 [5, 6]. The present study aims to verify whether the quality of DST in Italy changed after that time; to this end, a comprehensive survey of five PTs during a 13-yr period (1998–2010) is reported here, together with a pilot round of second-line drug PTs in 2010. Laboratories covering 18 out of 20 Italian regions participated in the PT exercise: 22 laboratories in 1998, 20 in 2000, 28 in 2003, 29 in 2007 and 30 in 2010. To maintain knowledge and skills, the laboratories were selected by the SRL on the basis of the number of patient samples analysed for DST. For instance, a mean of 88 first-line DSTs per laboratory (range 21–357) were performed in 2009. In 2010, 13 laboratories with a mean of 113 first-line and six second-line DSTs per laboratory in 2009 also performed the second-line drug PT. A mean of nine second-line DSTs per laboratory was performed in 2010. The Mycobacterium tuberculosis panels for first- and second-line drug PTs distributed by the Rome …

Evaluation of the VersaTREK System Compared to the Bactec MGIT 960 System for First-Line Drug Susceptibility Testing of Mycobacterium tuberculosis

The aim of this study was to evaluate the reliability of the VersaTREK system for Mycobacterium tuberculosis drug susceptibility testing compared with results obtained with the Bactec MGIT 960 system. A total of 67 strains were evaluated. Overall agreement was at 98.5%. Kappa indexes were 1.0 for isoniazid, rifampin, and ethambutol, 0.937 for pyrazinamide, and 0.907 for streptomycin. The VersaTREK system is validated for M. tuberculosis drug susceptibility testing.

Mycobacterium tuberculosis testing practices in hospital, commercial and state laboratories in the New England states

The mycobacterial laboratory is assuming an increasingly important role in tuberculosis (TB) control in the United States today. To assess mycobacterial laboratory capacity and practices in the New England states, USA. We surveyed 143 hospital and commercial laboratories and five of the six state public health laboratories in New England that offer testing services for Mycobacterium tuberculosis. The survey captured information on types of services offered and volume of testing, use of state laboratories for testing, and promptness of reporting results to TB control programs. State laboratories perform the majority of testing services, particularly for more specialized tests. All state laboratories surveyed perform species identification of acid-fast isolates, culture and first-line drug susceptibility testing. Less than 20% of hospital and commercial laboratories offer these services, and 78.6% of hospitals and commercial laboratories refer specimens to state laboratories for culture. Surveys of M. tuberculosis testing capacities in a region can help decision makers ensure maintenance of essential services. Hospital and commercial laboratories with lower testing volume might increase efficiency by referring more specimens to state laboratories. State health departments might consider organizing regional laboratory service networks to monitor the provision of services, improve efficiency and oversee quality improvement initiatives.

A survey on resistance to second-line drugs in patients with pulmonary tuberculosis in Shanghai, China

To determine the pattern of resistance to second-line drugs (SLDs) in patients with tuberculosis (TB) in Shanghai, China. All sputum culture positive patients detected at each district/county TB hospital/clinic in Shanghai during January to December 2009 were enrolled. First-line drug susceptibility testing was routinely performed for each clinical isolate. All clinical isolates with any resistance to first-line anti-TB drug were tested for second-line drug susceptibility. Of the 431 TB patients with any resistance to first-line anti-TB drugs included in this study, 27.8% (120/431) were resistant to any SLDs tested. The rank of drug resistance was as follows: ofloxacin (68/431, 15.8%) > kanamycin (37/431, 8.6%) > para-aminosalicylic acid (35/431, 8.1%) > capreomycin (28/431, 6.5%) > amikacin (23/431, 5.3%) > prothionamide (8/431, 1.9%). Mono-resistance to SLD was the most predominant pattern. Among patients with resistance to SLD, the percentages of mono-resistance were 2/2, 20/36, 22/24 and 43/58 for rifampin-resistant group, isoniazid-resistant group, other-resistant group, and MDR group, respectively. The high prevalence of resistance to SLDs in TB patients suggests the necessity to strengthen the TB control program for drug-resistant TB in Shanghai.

New and improved diagnostics for detection of drug-resistant pulmonary tuberculosis

Tuberculosis (TB) remains a global emergency and continues to kill 1.7 million people globally each year. Drug-resistant TB is now well established throughout the world and most TB patients are not being screened for drug resistance due to lack of laboratory resources and rapid accurate point-of-care tests. Accurate and rapid diagnosis of TB and drug-resistant TB is of paramount importance in establishing appropriate clinical management and infection control measures. During the past decade, there have been significant advances in diagnostic technologies for TB and drug-resistant TB. The purpose of this article is to review the current data, recommendations and evidence base for these tests. Second-line drug susceptibility testing (DST) is complex and expensive. Automated liquid culture systems and molecular line probe assays are recommended by the WHO as the current 'gold standard' for first-line DST. Liquid culture DST for aminoglycosides, polypeptides and fluoroquinolones has been shown to have relatively good reliability and reproducibility for diagnosis of extensively drug-resistant TB; however, DST for other second-line drugs (ethionamide, prothionamide, cycloserine, terizidone, para-aminosalicylic acid, clofazimine, amoxicillin-clavulanate, clarithromycin, linezolid) is not recommended. Automated liquid culture systems are currently recommended by the WHO as the 'gold standard' for second-line DST. In this review, we describe the phenotypic and genotypic methods currently available for the diagnosis of TB and drug-resistant forms of Mycobacterium tuberculosis and discuss future prospects for TB diagnostics. Current technologies for the detection of drug resistant M. tuberculosis vary greatly in terms of turnaround time, cost and complexity. Ultimately, the 'holy grail' diagnostic for TB must fulfil all technical specifications for a good point-of-care test, screen for drug resistance concurrently and be adaptable to the various health system levels and to countries with diverse economic status and TB burden.

High ethionamide resistance in Mycobacterium tuberculosis strains isolated in Kenya

SUMMARY Background: Increasing development of tuberculosis (TB) resistance to the currently available drugs including second-line anti-TB drugs that are being used for treatment of Multi-Drug Resistant TB (MDR-TB) patients has frustrated efforts to control TB worldwide. Ethionamide (Eth) is one of the drugs used in the regimen for treatment of these patients. Objective: To determine level of Ethionamide resistance among second-line anti-tuberculosis drugs in Mycobacterium tuberculosis (MTB) strains isolated in Kenya. Design: A retrospective lab-based study involving archived strains from previous studies carried out at the Centre for Respiratory Diseases Research (CRDR), Kenya Medical Research Institute (KEMRI) from 2002 to 2007. Setting: Centre for Respiratory Diseases Research (CRDR), Kenya Medical Research Institute (KEMRI). Methods: A total of 216 MTB strains with pre-determined first-line drug susceptibility testing (DST) results were used including 78 first-line resistant to individual and combined drugs, and 138 susceptible to streptomycin, rifampicin, isoniazid and ethambutol. The strains were subjected to DST to ethionamide among other second-line. Results: Thirty two [32/216 (14.8%)] strains showed resistance to second-line drugs. Resistance to Eth was the highest [18/32 (56.3%)] including co-resistance with isoniazid [8/18 (44.4%)]. Nine [9/18 (50%)] strains were fully resistant and 9 [9/18 (50%)] were intermediate resistant to Eth. Conclusion: Unexplainable high levels of Eth resistance is a cause for concern. This will impact negatively on the outcome of management of MDR-TB especially in Kenya where the use of this drug is almost mandatory. Close monitoring of Eth before initiating individual patient management may be necessary.

Extensively Drug-Resistant Tuberculosis, Lesotho

Extensively Drug-Resistant Tuberculosis, Lesotho

Scale-up of Multidrug-Resistant Tuberculosis Laboratory Services, Peru

Over the past 10 years, the Peruvian National Tuberculosis (TB) Program, the National Reference Laboratory (NRL), Socios en Salud, and US partners have worked to strengthen the national TB laboratory network to support treatment of multidrug-resistant TB. We review key lessons of this experience. The preparation phase involved establishing criteria for drug susceptibility testing (DST), selecting appropriate DST methods, projecting the quantity of DST and culture to ensure adequate supplies, creating biosafe laboratory facilities for DST, training laboratory personnel on methods, and validating DST methods at the NRL. Implementation involved training providers on DST indications, validating conventional and rapid first-line DST methods at district laboratories, and eliminating additional delays in specimen transport and result reporting. Monitoring included ongoing quality control and quality assurance procedures. Hurdles included logistics, coordinating with policy, competing interests, changing personnel, communications, and evaluation. Operational research guided laboratory scale-up and identified barriers to effective capacity building.

HIGH RATES OF MULTIDRUG-RESISTANT TUBERCULOSIS IN THE COUNTRY OF GEORGIA: A POPULATION-BASED STUDY.

Background Multidrug-resistant TB (MDR-TB, defined as resistance to at least isoniazid and rifampin) has emerged as a serious global public health problem, especially in former Soviet republics. The extent of the problem in Georgia has been incompletely defined because there have been no representative or population-based studies on the rates of drug-resistant TB in Georgia. Purpose The purpose of this study was to determine the prevalence and risk factors for MDR-TB in the country of Georgia. Methods/Study Design A prospective population-based study was carried out in the country of Georgia between July 2005 and May 2006. All patients diagnosed with AFB sputum smear positive (ss+) pulmonary TB in Georgia were enrolled and sputum specimens processed for culture, and subsequently, first-line drug susceptibility testing (DST) on one M. tuberculosis isolate recovered from each patient with a positive culture was performed. Specimens were collected from patients receiving care at 74 civilian TB facilities and 8 TB facilities within the penitentiary system located throughout Georgia. Demographic data, laboratory data, and data abstracted from medical records and patient interviews were entered into an Epi Info database and analyzed using SAS software (9.1). Results A total of 2,200 ss+ patients were enrolled, and 1,451 were culture positive for M. tuberculosis. DST was available for 1,426 patients (98%). Overall, 810 (57%) patients had resistance to at least one anti-TB drug, and 220 (15%) of 1,426 patients had MDR-TB; 794 (56%) were newly diagnosed patients with TB, 511 (36%) were previously treated for TB for > 1 month, and prior treatment status was not known for 121 (8%) who were excluded from further analyses. In univariate analysis, patients who had previously been treated for TB were significantly more likely to have MDR-TB (140/511 [27%]) than patients who were newly diagnosed and never previously treated (53/794 [7%]) (OR = 5.27, 95% CI 3.75-7.41, p Conclusions Drug-resistant TB, including MDR-TB, has emerged as a major public health problem in Georgia. Further TB control efforts being implemented to prevent the development of new cases of MDR-TB and to treat existing patients with MDR-TB are urgently needed.