Light-emitting Diode Fluorescence Microscopy Research Articles

Use of light-emitting diode fluorescence microscopy to detect acid-fast bacilli in sputum as proficient alternative tool in the diagnosis of pulmonary tuberculosis in countries with limited resource settings.

Despite recent advances in the development of more sensitive technologies for the diagnosis of tuberculosis (TB), in resource-limited settings, the diagnosis continues to rely on sputum smear microscopy. This is because smear microscopy is simple, cost-efficient and the most accessible tool for the diagnosis of TB. Our study evaluated the performance of light-emitting diode fluorescence microscopy (LED-FM) using auramine/rhodamine (auramine) and the fluorescein di-acetate (FDA) vital stain in the diagnostic of pulmonary TB in Bamako, Mali. Sputum smear microscopy was conducted using the FDA and auramine/rhodamine staining procedures on fresh samples using LED-FM to evaluate the Mycobacterium TB (MTB) metabolic activity and to predict contagiousness. Mycobacterial culture assay was utilized as a gold standard method. Out of 1401 TB suspected patients, 1354 (96.65%) were retrieved from database, which were MTB complex culture positive, and 47 (3.40%) were culture negative (no mycobacterial growth observed). Out of the 1354 included patients, 1343 (95.86%), were acid-fast bacillus (AFB) positive after direct FDA staining, 1352 (96.50%) AFB positive after direct Auramine, and 1354 (96.65%) AFB positive with indirect auramine after digestion and centrifugation. Overall, the FDA staining method has a sensitivity of 98.82%, while the sensitivity of Auramine with direct observation was 99.48%, and 99.56% with the indirect examination. This study showed that, using fresh sputum both auramine/rhodamine and FDA are highly sensitive methods in diagnosing pulmonary TB and could be easily used in countries with limited resource settings.

Comparison of Light Emitting Diode (LED) Fluorescent Microscopy with Ziehl-Neelsen Microscopy on Sputum Specimens for Diagnosis of Pulmonary Tuberculosis Keeping Culture as ‘Gold Standard’

Objective: To compare LED fluorescence microscopy and Ziehl-Neelsen staining in terms of their diagnostic performance in diagnosing pulmonary tuberculosis, taking sputum specimens from patients suspected of pulmonary tuberculosis.
 Study Design: Prospective longitudinal study.
 Place and Duration of Study: Microbiology Department, Armed Forces Institute of Pathology Rawalpindi, from Jan 2019 to Dec 2019.
 Methodology: Sputum samples from patients with clinical suspicion of pulmonary tuberculosis were stained using Ziehl-Neelsen (ZN) stain, fluorescent stain with Auramine O staining (AO) stain and Mycobacterial culture on Mycobacterial Growth Indicator Tube (MGIT 960), to detect Mycobacterium tuberculosis. WHO guidelines were followed to grade positive smears.
 Results: Among 206 patients with suspicion of tuberculosis, 143 (69%) were male, and 63 (30%) were female patients. The mean age of the patients was 53.67 ± 14.73 years. Out of 206 sputum samples, 64 were negative by all three techniques used. 142 (68%) specimens detected Mycobacterium tuber-culosison MGIT960. Within 142 culture-positive samples, only 40 samples were positive on Ziehl-Neelsen microscopy, whereas 97 samples were detected positive by LED fluorescent microscopy. In culture-negative samples, three were missed on Ziehl-Neelsen staining, which was positive with Fluorescent microscopy. Sensitivity and specificity for Ziehl-Neelsen smear microscopy were 26.7% and 96.8%, respectively. Sensitivity and specificity for Fluorescent smear microscopy were 64.8% and 92.2%, respectively.
 Conclusion: We concluded that the efficacy of LED fluorescence microscopy has proven to have many potential advantages over conventional Ziehl-Neelsen microscopy.

Laboratory Cost Analysis of Conventional and Newer Molecular Tests for Diagnosis of Presumptive Multidrug-Resistant Tuberculosis Patients.

Introduction:Tuberculosis (TB) remains a deadliest infectious disease. Lack of rapid test with low cost is one of the important challenges to eradicate the TB. The objective of the study was to analyze the laboratory costs of conventional and newer molecular tests, for diagnosis of presumptive multidrug-resistant TB (MDR-TB) patients.Methods:A detailed laboratory cost of various conventional tests (Ziehl − Neelsen [ZN] microscopy, light-emitting diode-fluorescent microscopy [LED-FM], culture and drug susceptibility testing [DST] using solid Lowenstein–Jensen media and liquid media [BACTEC MGIT 960]) was compared with rapid methods (GenoType MTBDRplus line probe assay [LPA] and GeneXpert MTB/RIF assay). Laboratory cost was also calculated in terms of cost per TB and MDR-TB case detected by using different diagnostic scenarios.Results:Cost per test for ZN microscopy, LED-FM, LPA, GeneXpert MTB/RIF assay, solid culture plus DST, liquid culture plus DST was found as $2.5 (INR 156.8), $2.0 (INR128.9), $18.6 (INR1210), $13.8 (INR 895.2), $21.5 (INR 1396.6), and $29.1 (INR 1888.2), respectively. The laboratory cost for detecting TB and MDR-TB by diagnostic scenarios involving molecular DST was found to be less as compared to involving only conventional liquid culture-based test.Conclusions:The implementation of rapid molecular tests with selective use of liquid culture-based DST may be less in cost as compared to the use of culture-based DST alone, at high burden reference TB laboratory.

Prevalence, Patterns, and Determinants of Malaria and Malnutrition in Douala, Cameroon: A Cross-Sectional Community-Based Study.

Background Malaria and malnutrition are major public health problems in developing countries. This studywas mainly focused on the prevalence, patterns, and predictors of these conditions and their associations. Methods A cross-sectional community study was conducted from February to March 2018 among 281 participants living in two districts in Douala. A questionnaire was used to collect sociodemographic information and parasitological and anthropometric data of participants. Nutritional status was determined using age, weight, and height. Body mass index for age (BMIAZ), height-for-age (HAZ), weight-for-age (WAZ), and weight-for-height (WHZ) Z scores were computed based on the World Health Organization growth reference curves. Malaria infection was diagnosed using light-emitting diode fluorescence microscopy. Results The overall prevalence of malaria was 18.9%, mostly asymptomatic cases. Malaria infection was associated with study site (p = 0.04), age (p = 0.01), WAZ (p = 0.0049), HAZ (p = 0.03), and BMI (p = 0.02). The overall prevalence of malnutrition was 43.1%, and stunting was the main form of malnutrition recorded in children under five years of age (23.6%). The risk of being stunted in this group was about quintupled in malaria-infected participants (ARR = 4.70; p = 0.02). In those aged 5-19 years, the prevalence of underweight was significantly higher in malaria-positive individuals as compared to their negative counterparts (p = 0.02). The overall prevalence of malaria and malnutrition cooccurrence was 8.5% and varied with age (p < 0.0001) and study site (p = 0.04). Conclusion and Recommendation. Malaria was associated with malnutrition among the study participants. Early detection and treatment of these ailments would reduce morbidity and mortality.

Disease of consumption to extensively drug resistance tuberculosis: Race of techniques to enhance diagnosis

Tuberculosis is one of the most ancient infectious diseases known to mankind. Detection of the causative organism, Mycobacterium tuberculosis, has experienced a series of advances from the late 19th century through early 21st century. The hierarchy of diagnosis ranges from smear microscopy with traditional Ziehl–Neelsen staining to advanced light-emitting diode–fluorescence microscopy staining to both solid and liquid culture, and more recently, to genotypic methods like nucleic acid amplification. Each procedure has its own merits and drawbacks in terms of specificity, sensitivity, minimum detectible number of bacilli, and most crucially, turnover time. The latest diagnostic tools, like geneXpert and line probe assays, have gone a step further by detecting drug resistance. Both early and accurate diagnosis based on the drug susceptibility test implies precise and prompt initiation of treatment, thereby curtailing not only the agony of patient but also the burden tuberculosis places on the health system.

Performance and operational feasibility of two diagnostic tests for cryptosporidiosis in children (CRYPTO-POC): a clinical, prospective, diagnostic accuracy study

SummaryBackgroundCryptosporidiosis is a common cause of diarrhoea in young children (aged younger than 24 months) in low-resource settings but is currently challenging to diagnose. Light-emitting diode fluorescence microscopy with auramine-phenol staining (LED-AP), recommended for tuberculosis testing, can also detect Cryptosporidium species. A lateral-flow test not requiring refrigerator storage (by contrast with most immunochromatographic lateral-flow assays) has also recently been developed for Cryptosporidium spp detection. We aimed to evaluate the diagnostic accuracy and operational feasibility of LED-AP and the lateral-flow test strip for cryptosporidiosis in children.MethodsWe did a prospective diagnostic accuracy study in two health-care facilities in Ethiopia, in a consecutive series of children younger than 5 years of age with diarrhoea (three or more loose stools within the previous 24 h) or dysentery (at least one loose stool with stains of blood within the previous 24 h). Stool samples were tested for Cryptosporidium spp by LED-AP and the lateral-flow test strip; accuracy of each test was estimated by independent and blind comparison with a composite reference standard comprising quantitative immunofluorescent antibody test (qIFAT), ELISA, and quantitative PCR (qPCR). Quantitative cutoff values for diarrhoea-associated infection were established in an embedded case-control substudy, with cases of cryptosporidiosis coming from the 15 districts in and around Jimma and the eight districts surrounding Serbo, and community controls without diarrhoea in the previous 48 h recruited by weekly frequency matching by geographical district of the household, age group, and enrolment week.FindingsStool samples from 912 children with diarrhoea or dysentery and 706 controls from the case-control substudy were tested between Dec 22, 2016, and July 6, 2018. Estimated reference-standard cutoff values for cryptosporidiosis positivity were 2·3 × 105 DNA copies per g of wet stool for qPCR, and 725 oocysts per g for qIFAT. LED-AP had a sensitivity for cryptosporidiosis of 88% (95% CI 79–94; 66 of 75 samples) and a specificity of 99% (98–99; 717 of 726 samples); the lateral-flow test strip had a sensitivity of 89% (79–94; 63 of 71 samples) and a specificity of 99% (97–99; 626 of 635 samples).InterpretationLED-AP has high sensitivity and specificity for cryptosporidiosis and should be considered as a dual-use technology that can be easily integrated with existing laboratory infrastructures in low-resource settings. The lateral-flow test strip has similar sensitivity and specificity and provides an alternative that does not require microscopy, although purchase cost of the test strip is unknown as it is not yet available on the market.FundingNorwegian Research Council GLOBVAC fund, The Bill & Melinda Gates Foundation, Norwegian Society for Medical Microbiology, University of Bergen, and Vestfold Hospital Trust.

Fluorescence microscopy for the diagnosis of smear-negative pulmonary tuberculosis in Ethiopia.

BackgroundDespite its low sensitivity, microscopy remains the main method for the diagnosis of pulmonary tuberculosis in most laboratories in Ethiopia. Few studies have evaluated the performance of light-emitting diode fluorescent microscopy (LED-FM) in bleach-concentrated smear-negative sputum specimens.ObjectiveThis study aimed to evaluate the diagnostic performance of LED-FM for smear-negative pulmonary tuberculosis in Ethiopia.MethodsA total of 194 adult patients with a cough lasting for more than two weeks, and who had three direct smear-negative sputum tests for Mycobacterium tuberculosis by Ziehl-Neelsen light microscopy, were included. All direct Ziehl-Neelsen-stained smear-negative sputum samples were cultured and were also visualised by LED-FM. Smears for LED-FM were performed from bleach-concentrated sputum sediment. The diagnostic performance of the LED-FM was compared to the culture method (the reference standard).ResultsOf the 194 smear-negative sputum specimens analysed, 28 (14.4%) were culture-positive and 21 (10.8%) were LED-FM-positive for M. tuberculosis. However, only 11 of the 21 (52.4%) LED-FM-positive patients had a confirmed tuberculosis diagnosis by culture. Light-emitting diode fluorescence microscopy (FM) had a sensitivity of 39.3% (95% confidence interval: 21.2–57.4) and specificity of 93.9% (95% confidence interval: 90.4–97.6). Ten LED-FM-positive specimens were culture-negative, and all of these specimens had scanty grading (1–19 bacilli per 40 fields on LED-FM).ConclusionThis study showed that implementation of LED-FM on bleach pre-treated and concentrated sputum can significantly improve the diagnosis of smear-negative pulmonary tuberculosis. However, all scanty grade, positive smears by LED-FM need to be confirmed by reference culture method.

Evaluation of ‘TBDetect’ sputum microscopy kit for improved detection of Mycobacterium tuberculosis: a multi-centric validation study

ObjectivesThe present study aimed to evaluate the performance of the ‘TBDetect’ kit-based bio-safe fluorescent microscopy filter (BioFM-Filter) microscopy in comparison with direct smear microscopy and culture for the detection of pulmonary tuberculosis (TB) in a multi-centric setting in India. MethodsThe TBDetect kit enables sputum concentration through filtration using the BioFM-Filter for improved and bio-safe smear microscopy. We evaluated the performance of the TBDetect kit in a six-site multi-centric validation study on sputum collected from 2086 presumptive TB patients. ResultsThe combined positivity of TBDetect microscopy performed on these sputum samples was 20% (n = 417/2086) vs 16.1% of light-emitting diode fluorescence microscopy (LED-FM, n = 337/2086) and 16% of Ziehl Neelsen (ZN) smear microscopy (n = 333/2086). The increment in positivity of TBDetect over both LED-FM and ZN smears was significant (p < 0.001). The overall sensitivity of TBDetect for six sites was ~55% (202/367, 95% confidence interval (CI): 50, 60%) vs 52% (191/367, 95% CI: 47, 57%) for LED-FM (p 0.14) and 50.9% (187/367, 95% CI: 46, 56%) for ZN smear (p < 0.05), using Mycobacterium Growth Indicator Tube culture (MGIT, n = 1949, culture positive, n = 367) as the reference standard. A bio-safety evaluation at six sites confirmed efficient sputum disinfection by TBDetect; 99.95% samples (1873/1874) were sterile after 42 days of incubation. Scientists and technicians at the study sites indicated the ease of use and convenience of TBDetect microscopy during feedback. ConclusionsTBDetect added value to the smear microscopy test due to its improved performance, convenience and user safety. These findings indicate that equipment-free TBDetect technology has the potential to improve TB diagnosis in basic laboratory settings by leveraging on the existing nationwide network of designated microscopy centres and primary healthcare centres.

A study to analyze the differences between GeneXpert MTB/RIF assay and LED fluorescent microscopy for the diagnosis of pulmonary tuberculosis in a tertiary care centre

cultures although being the gold standard for diagnosing tuberculosis has its limitations as it is time consuming. Thus newer methods using light-emitting diode fluorescent microscopy (LED-FM) and molecular methods like the GeneXpert MTB/RIF assay are being adopted as a mainstay for diagnosing tuberculosis around the globe. This observational descriptive study was carried out for a period of 21 months. Sputum samples from 110 adult subjects with a clinical suspicion of pulmonary tuberculosis (PTB) were stained for LED-FM microscopy and also inoculated into the GeneXpert assay. The positive smears were graded as per the Revised National Tuberculosis Control Programme (RNTCP) guidelines and positive GeneXpert assay results along with Rifampicin resistance were noted and analysed. Out of the 110 samples, (77.2%) were males and (22.7%) females. Of the 110 samples, 75 (68.1%) were positive for by the GeneXpert assay. In contrast with LED-FM, all 25 smear positive samples were also GeneXpert positive. Additionally, GeneXpert detected 10 smear negative cases. 10 (9%) positive GeneXpert isolates showed resistance to Rifampicin. The overall sensitivity of GeneXpert was greater but its specificity was comparable with LED-FM. The Xpert and LED-FM tests were comparable in specificity, but the GeneXpert assay out did the LED-FM as it was a more sensitive and rapid test for the diagnosis of tuberculosis and additionally detection of Rifampicin resistance. Our study fully agrees that the GeneXpert assay is a step ahead in the early diagnosis of tuberculosis even for smear negative cases of PTB. This assay is also a relatively simple and routine test that staff can perform with minimal training.

Performance of Xpert MTB/RIF in comparison with light-emitting diode-fluorescence microscopy and culture for detecting tuberculosis in pulmonary and extrapulmonary specimens in Bamako, Mali.

The diagnosis of tuberculosis (TB) has mostly been relied on a long-used method called sputum smear microscopy. In 2010, Xpert MTB/RIF assay was approved by the World Health Organization for simultaneous TB diagnosis and detection of resistance. Our current study was undertaken to compare the diagnostic performance of Xpert MTB/RIF assay to auramine staining-based light-emitting diode-Fluorescence Microscopy (LED-FM) considering culture as the gold standard method for pulmonary and extrapulmonary TB. Pulmonary and extrapulmonary specimens of suspected TB patients were examined in this study. From January 2016 to June 2019, sputum, urine, superficial swabs, gastric aspirates, and pleural infusion specimens were collected from new and previously treated TB individuals. Specimens were examined using Xpert MTB/RIF, LED-FM, and Mycobacterium culture techniques to evaluate their performance. A total of 697 suspected TB samples were included in this analysis, and of these, 469 (67.29%) were positive for all three used methods. The overall sensitivities, specificities, and positive and negative predictive values were 99.6%, 62.0%, 88.4%, and 98.2% for Xpert MTB/RIF and 88.0%, 95.6%, 99.0%, and 60.7% for LED-FM, respectively, compared to culture method. The sensitivity of Xpert MTB/RIF assay was observed to be higher than the LED-FM method, thus suggesting this molecular technique as a promising tool for the diagnosis of pulmonary and extrapulmonary TB, which will help in the management of TB infections in developing countries such as Mali.

Comparison of Ziehl–Neelsen light microscopy and fluorescent (light-emitting diode) microscopy with solid culture for the diagnosis of tuberculosis with special reference to extrapulmonary cases

Introduction Owing to the HIV pandemic, partially fuelled by breakdown of infrastructure and poverty, the increase in tuberculosis (TB) cases has reached unmanageable levels in the developing countries. Smear microscopy is still the main diagnostic modality. Efficient and prompt case detection followed by adequate treatment has drastically reduced TB transmission. Hence, an effort was made to compare Auramine-O (AO) and Ziehl–Neelsen (ZN) staining methods for demonstration of acid-fast bacilli with solid culture on Lowenstein–Jenson medium as a reference standard. Patients and methods A total of 100 sputum and 50 extrapulmonary samples from patients clinically suspected of TB were included. Smears were stained by ZN and AO method separately. All the samples were processed as per Revised National Tuberculosis Control Program (RNTCP) guidelines of India and inoculated on Lowenstein–Jenson medium. Mycobacterium tuberculosis was identified by their growth characteristics, nitrate reduction test, and susceptibility to para-nitrobenzoic acid. Results Of 150 samples, 27 (18%), 37 (24.67%), and 38 (25.33%) were positive by ZN stain, AO stain, and culture, respectively. Of 50 extrapulmonary samples, none were positive by ZN stain, four (8%) were positive by AO stain, and two (4%) by culture. AO method detected additional 8% of extrapulmonary TB cases which were missed by ZN and also detected more number of paucibacillary cases. Sensitivity and specificity of ZN were 71.05 and 100%, respectively. Sensitivity and specificity of AO were 86.84 and 96.43%, respectively. Conclusion The higher sensitivity and diagnostic accuracy of AO method indicate that the light-emitting diode fluorescent microscopy can be efficiently implemented in resource-limited settings with a high burden of TB.

Diagnostic Performance of Xpert MTB /RIF in Comparison with LED Fluorescence Microscopy and Culture in Suspected Cases of Pulmonary Tuberculosis

GeneXpert MTB/RIF assay (GeneXpert) is relatively new technology in India and it is important to generate data on its efficacy for diagnosis of Tuberculosis. Thus; present study was undertaken to compare the diagnostic performance of GeneXpert for diagnosis of pulmonary tuberculosis (PTB) in comparison with Auramine O staining based Light Emitting Diode- Fluorescence Microscopy (LEDFM) considering culture as gold standard. A total of 662 (451 BAL, 211 Sputum) respiratory samples obtained from patients suspected of PTB were tested for smear microscopy and GeneXpert assay and were compared with culture results. Rifampicin resistance was compared with Genotype MTBDR plus assay. Sensitivity, specificity, positive predictive value (PPV) and negative predictive values (NPV) of the assays were calculated using SPSS version 22. On comparing with culture the sensitivity, specificity, PPV and NPV of GeneXpert MTB/RIF was found to be 97.25%, 98.54%, 96.20% and 98.95% respectively whereas sensitivity, specificity, PPV and NPV of smear microscopy was found to be 73.08%, 100%, 100% and 90.74% respectively. Rifampicin resistance was detected in 11 (1.66%) samples by GeneXpert. Five (0.75%) samples among them were found to be MDR whereas one sample showed false positive result for rifampicin resistant when compared by Genotype MTBDR plus assay. GeneXpert MTB/RIF assay offers much higher sensitivity as compared to Auramine O staining based LED-FM and seems to be a promising tool for TB diagnosis. Further longitudinal studies are required to evaluate the potential of this assay at primary health care settings and diagnosis of Extrapulmonary TB.

Diagnosis of pulmonary tuberculosis with cartridge based nucleic acid amplification test and light emitting diode fluorescent microscopy: a comparative study

Background: Due to low sensitivity and inability to detect drug resistance, smear microscopy limits its impact on TB control. Culture methods and drug susceptibility testing is complex, time consuming, and takes around 6-8 weeks. A new diagnostic test, cartridge based nucleic acid amplification test (CBNAAT) was developed based on real-time polymerase chain reaction (RT PCR). Objective of this study to compare the results of CBNAAT for diagnosis of pulmonary tuberculosis with LED fluorescent microscopy and sputum culture.Methods: A cross-sectional study was conducted in the department of Chest and TB, CIMS, Bilaspur. Each Sputum sample of presumptive TB patients were tested with CBNAAT, sputum smear microscopy by light emitting diode (LED) fluorescent microscopy (FM) and solid and liquid culture for diagnosis of Tuberculosis. Results of CBNAAT, Fluorescent Microscopy and Culture for detection of Mycobacterium Tuberculosis were compared.Results: The sensitivity and specificity for CBNAAT were 97% and 100% respectively; while that for Fluorescent microscopy were 70% and 100% respectively. The positive and negative predictive value for CBNAAT was 100% and 96% respectively. The positive and negative predictive value for Fluorescent microscopy was 100% and 73% respectively.Conclusion: CBNAAT is having high sensitivity and specificity for diagnosis of pulmonary tuberculosis. It should be routinely used under national health programme to detect a tuberculosis case efficiently.

Newer Diagnostic Tests and their Application in Pediatric TB.

The diagnosis of childhood tuberculosis is a challenge due to the pauci-bacillary nature of infection and the difficulty in obtaining appropriate sample. In the past 2-3 decades, many new tests were introduced for the diagnosis of tuberculosis (TB) and some of them have been evaluated for their application in pediatric tuberculosis as well. There is an attempt to improve smear microscopy by introducing light-emitting diode (LED) fluorescence microscopy and there are also some automated digital microscopy platforms under evaluation. Introduction of automated liquid culture platform along with rapid molecular based identification methods have considerably reduced the time delay in mycobacterial culture. Recent addition of many nucleic acid amplification platforms like Amplicor PCR, Genprobe, Xpert MTB/Rif, line probe assays, loop mediated isothermal amplification etc are also been found to be useful. Latest techniques like microarray and gene sequencing are also being used in clinical laboratories with variable results. Indirect methods of TB diagnosis like T cell based assays including tuberculin skin test and interferon-gamma release assays have their role primarily in the diagnosis of latent TB. Biomarkers are the latest addition in the battery of TB diagnostic tests facilitating diagnosis using easily accessible samples like urine, blood and breath of patients. Many biomarkers are still under evaluation and some of them are found to have a potential role as promising diagnostic tests of future.

Light-emitting diode-fluorescent microscopy: Determining its sensitivity and specificity in diagnosis of pulmonary tuberculosis in a high-burden tuberculosis region and resource-limited country like India

Background: Tuberculosis (TB) remains a major global health problem. India accounts for the one-fourth of the global TB burden. For resource-limited countries like India having high burden of TB, smear microscopy is the most commonly used method for microbiological diagnosis. Although culture being highly sensitive and specific method, it is time-consuming. Under Revised National TB Control Program (RNTCP), Ziehl–Neelsen (ZN) stain-based microscopy and light-emitting diode-based fluorescent microscopy (LED-FM) are being recommended. Aim: The present study was done to determine the sensitivity and specificity of ZN stain-based microscopy and FM (LED-FM) keeping solid culture as gold standard. Materials and Methods: A total of 1503 sputum samples were collected from suspected cases of pulmonary TB (new or previously treated). They were processed and subjected to ZN (conventional light microscopy), Auramine-O (AO) staining (LED-FM) and solid culture on Lowenstein–Jensen medium for detection of acid-fast bacilli. Positive smears were graded according to RNTCP guidelines. Results: Sensitivity, specificity, positive predictive value, and negative predictive value of ZN staining technique were 84.93%, 95.74%, 96.50%, and 82.16%, respectively, and that of FM was 95.25%, 91.33%, 93.81%, and 93.31%, respectively. Conclusion: LED-FM being more sensitive, especially in detecting the paucibacillary cases and less time-consuming, has advantage over ZN method. Nation-wide implementation will lead to improved diagnosis, thus better control of TB.

Cost-effectiveness of GeneXpert and LED-FM for diagnosis of pulmonary tuberculosis: A systematic review.

BackgroundEarly and accurate diagnosis of tuberculosis is a priority for TB programs globally to initiate treatment early and improve treatment outcomes. Currently, Ziehl–Neelsen (ZN) stain-based microscopy, GeneXpert and Light Emitting Diode-Fluorescence Microscopy (LED-FM) are used for diagnosing pulmonary drug sensitive tuberculosis. Published evidence synthesising the cost-effectiveness of these diagnostic tools is scarce.MethodologyPubMed, EMBASE and Cost-effectiveness analysis registry were searched for studies that reported on the cost-effectiveness of GeneXpert and LED-FM, compared to ZN microscopy for diagnosing pulmonary TB. Risk of bias was assessed independently by four authors using the Consensus Health Economic Criteria (CHEC) extended checklist. The data variables included the study settings, population, type of intervention, type of comparator, year of study, duration of study, type of study design, costs for the test and the comparator and effectiveness indicators. Incremental cost-effectiveness ratio (ICER) was used for assessing the relative cost-effectiveness in this review.ResultsOf the 496 studies identified by the search, thirteen studies were included after removing duplicates and studies that did not fulfil inclusion criteria. Four studies compared LED-FM with ZN and nine studies compared GeneXpert with ZN. Three studies used patient cohorts and eight were modelling studies with hypothetical cohorts used to evaluate cost-effectiveness. All these studies were conducted from a health system perspective, with four studies utilising cost utility analysis. There were considerable variations in costing parameters and effectiveness indicators that precluded meta-analysis. The key findings from the included studies suggest that LED-FM and GeneXpert may be cost effective for pulmonary TB diagnosis from a health system perspective.ConclusionOur review identifies a consistent trend of the cost effectiveness of LED-FM and GeneXpert for pulmonary TB diagnosis in different countries with diverse context of socio-economic condition, HIV burden and geographical distribution. However, all the studies used different parameters to estimate the impact of these tools and this underscores the need for improving the methodological issues related to the conduct and reporting of cost-effectiveness studies.

Screening for Tuberculosis With Xpert MTB/RIF Assay Versus Fluorescent Microscopy Among Adults Newly Diagnosed With Human Immunodeficiency Virus in Rural Malawi: A Cluster Randomized Trial (Chepetsa).

Tuberculosis (TB) remains the leading cause of death among human immunodeficiency virus (HIV)-infected individuals globally. Screening for TB at the point of HIV diagnosis with a high-sensitivity assay presents an opportunity to reduce mortality. We performed a cluster randomized trial of TB screening among adults newly diagnosed with HIV in 12 primary health clinics in rural Thyolo, Malawi. Clinics were allocated in a 1:1 ratio to perform either point-of-care Xpert MTB/RIF assay (Xpert) or point-of-care light-emitting diode fluorescence microscopy (LED-FM) for individuals screening positive for TB symptoms. Asymptomatic participants were offered isoniazid preventive therapy in both arms. Investigators, but not clinic staff or participants, were masked to allocation. Our primary outcome was the incidence rate ratio (RR) of all-cause mortality within 12 months of HIV diagnosis. Prevalent TB was diagnosed in 24 of 1001 (2.4%) individuals enrolled in clinics randomized to Xpert, compared with 10 of 841 (1.2%) in clinics randomized to LED-FM. All-cause mortality was 22% lower in the Xpert arm than in the LED-FM arm (6.7 vs 8.6 per 100 person-years; RR, 0.78 [95% confidence interval {CI}, .58-1.06]). A planned subgroup analysis suggested that participants with more advanced HIV (World Health Organization clinical stage 3 or 4) disease had lower mortality in clinics randomized to Xpert than to LED-FM (RR, 0.43 [95% CI, .22-.87]). In rural Malawi, using point-of-care Xpert MTB/RIF to test symptomatic patients for TB at the time of HIV diagnosis reduced all-cause 12-month mortality among individuals with advanced HIV. NCT01450085.

Impact of introducing light-emitting diode fluorescence microscopy services for diagnosis of pulmonary tuberculosis under Revised National Tuberculosis Control Program India.

Context:Light-emitting diode fluorescence microscopy (LED-FM) has been recommended by the WHO and the Government of India over the conventional bright-field microscopy using Ziehl–Neelsen (ZN) staining for the diagnosis of sputum smear-positive tuberculosis (TB) suspects.Aim:The aim of this study was to assess the effectiveness of LED-FM in detecting TB cases.Setting and Design:This retrospective cohort study was carried out in December 2016 using secondary data of the years 2011–2012 from designated microscopy centers (DMCs) at Chandigarh, India.Methodology:Two peripheral DMCs where conventional ZN microscopy was used were taken as controls comparable to two peripheral DMCs which used LED-FM services in programmatic settings. The record of all suspected TB patients who underwent sputum smear examination pre- and post-LED-FM introduction was compared for measuring smear-positive cases, conformational grading, and time taken to read per slide examined. Chi-square was applied to access the statistical significance.Results:Out of total 8850 cases registered after the inception of LED-FM microscopy services, case detection rate was 13.3%, with significant decrease in case detection rate at tertiary level DMCs, there was significant increase in case detection rate at peripheral DMCs operating with LED-FM in contrast to ZN microscopy which was used previously. Scanty grade smear increased significantly by 9.0%. The time taken per slide examination decreased by 57.1% (from 7 min on an average to 3 min on an average) after LED-FM was used.Conclusion:LED-FM is easy to use, takes lesser time to examine slides, and has longer self-life. It also eases laboatory technician's task in reporting scanty grade positives which use to be challenging previously. The results of this study provide enough evidences to scale up the installation process and usage of LED-FM at DMCs for TB diagnosis in Indian settings.

Efficacy and role of Xpert® Mycobacterium tuberculosis/rifampicin assay in urinary tuberculosis.

Introduction:The aim was to study the accuracy of Xpert® (Cepheid Inc., Sunnyvale, CA, USA) Mycobacterium tuberculosis/rifampicin (MTB/RIF) assay as compared to a composite gold standard (urine culture, imaging, and biopsy) and to asses its utility as the initial test compared to smear microscopy to diagnose urinary tuberculosis.Methods:This prospective study included adult patients suspected to have urinary tuberculosis from March 2014 to December 2017. Three urine samples were collected from each patient and were subjected to Xpert MTB/RIF assay, acid-fast bacillus (AFB) smear microscopy, and liquid media (BACTEC Mycobacteria Growth Indicator Tube [MGIT] 960) culture. Imaging and tissue biopsies were performed as clinically indicated. Sensitivity, specificity, positive predictive value, and negative predictive value were calculated using the bootstrap method for 95% confidence intervals for the Xpert assay.Results:Xpert MTB/RIF assay was found to be superior to the currently best available light-emitting diode fluorescent smear microscopy as the initial test for urinary tuberculosis (sensitivity of 69.09% vs. 32.72%). The Xpert MTB/RIF polymerase chain reaction test was found to have a moderate sensitivity (69.09%) and high specificity (100%) as compared to the composite reference standard. The sensitivity of liquid AFB culture MGIT 960 as compared to the reference standard was 90.32%.Conclusions:Xpert MTB/RIF assay on an early morning first void urine specimen can replace smear microscopy as the initial diagnostic test for urinary tuberculosis.

Comparison of GeneXpert against Light-Emitting Diode Fluorescent Microscopy for the Diagnosis of Pulmonary Tuberculosis in Addis Ababa, Ethiopia

Background: Tuberculosis remains a global health problem despite availability of effective tools. Globally, there were 40% of the 10+ million patients with TB did not get diagnosed or notified. The study was aimed to compare the diagnostic performances of iLED-FM and GeneXpert test for the diagnosis of PTB in Addis Ababa, Ethiopia. Methods: Facility-based cross-sectional study was conducted on a total of 286 sputum samples collected from health centers and hospitals clients with presumptive TB from December 2016 to March 2017. Kappa value, Sensitivity, Specificity, Positive Predictive Value (PPV) and Negative Predictive Value (NPV) of iLED-FM and GeneXpert were calculated against the gold standard. Results: The sensitivity, specificity, PPV and NPV of iLED-FM was 80.15%, 95.48%, 93.75% and 85.06%, for GeneXpert 88.55%, 92.90%, 91.34% and 90.57%, respectively. Kappa value of iLED-FM was 0.765 and 0.817 for GeneXpert. Out of 55 HIV positive presumptive TB patients enrolled in the study, 19 (34.55%) were sputum smear positive by iLED-FM. However, 24 (43.64%) TB cases were detected by GeneXpert. Conclusion: The sensitivity of Xpert MTB/RIF assay was better than iLED-FM in the diagnosis of PTB. It should be implemented as primary diagnostic test in areas where overlapping synergy of TB and HIV/AIDS is high.