Role of clinical microbiology laboratories in diagnosis of tuberculosis


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Role of clinical microbiology laboratories in diagnosis of tuberculosis

  1. 1. Role of Medical Microbiology Laboratories in Diagnosis of TuberculosisDr.T.V.Rao MDImportance of Microscopy:Smear examination for the detection of Acid fast bacilli continues to be the gold standard in Diagnosis ofTuberculosis. In spite of several inadequacies, Microscopy for AFB detection is economical, specific andthe man power can be trained easily. The detection of AFB in sputum smears helps for higher casedetection, and contains the spread of Tuberculosis in the Society.The smear will become Positive when one has bacilli more than 5,000 - 10, 000 / 1 ml of sputum.Multiple smear examinations, at least three morning specimens are advised and appropriate collection ofspecimens will increase yield to > 43 %. If efforts were taken in educating patients for 1- 2 minutes inmethods to collect the sputum, will yield higher results. Sputum induction procedures are helpful.Todays emphasis to identify AFB, in smears is more demanding with associated HIV/AIDS, as fewbacilli are excreted. Concentration of specimens and digestion of thick and mucous associate specimenswith Sodium hypochlorite, Sodium hydroxide, N-acetyl -cystine - Sodium hydroxide will increase rate ofdetection to > 18 % in sensitivity and incremental yield of 9 %( positive after treatment with abovechemicals - positives with direct Ziehl Nelsons straining ) Sodium hypochlorite is beneficial in HIVpositive patients as it is Mycobactericidal and also kills human Immunodeficiency virus, but not suitablefor culturing specimens.Need for Florescent MicroscopyThe developing world should explore the Fluorescence microscopy, which will improve the sensitivity ofMicroscopy in patient who excrete few bacilli as in association HIV infection, The role of Ziehl Neelsensmethod of staining and conventional Microcopy is losing the sensitivity with ever increasing work load,technicians opting to see few fields, monotonous nature of work, the lack of accountability, and interInstitutional quality control protocols. Many systematic reviews indicated use of Florescent Microscopywill increase 10% higher sensitivity and 9 % in incremental yield when compared with Z.N method ofstaining. About 15 times as many fields of view can be scanned by Fluorescent Microscopy as byconventional Microscopy in the same period. The developing countries face crunch to buy Fluorescentmicroscopes and to maintain the regular availability of florescent dyes. It is utmost important to developcentralized and dedicated centers for Microscopy to have control on peripheral laboratories. Negativesmears by conventional Microscopy needs further attention with optimal microscopy, concentrationmethods to detect AFB to reduce early mortality among the infected and to contain the spread in society.
  2. 2. Culturing for Mycobacterial IsolationSputum culturing remains a gold standard for diagnosing Mycobacterial infections. A Positive growth canbe demonstrated with few bacilli to as low as 10 - 100 of viable bacilli per I ml of sputum. Cultures showgrowth of AFB even when patients where on treatment and negative by smear examination. A simplemeasure with decontamination of specimens and inoculation of at least 150 - 200 µl of concentrate onculture medium will increase the success in culturing. In spite of best decontamination procedures, 1 - 4% of the isolates are false Positive. The greatest limitation of culturing on Lowenstein - Jensen mediumand other equivalent medium is long periods (2 - 12 weeks) for isolation of bacteria.Advances in Diagnostic Methodologies.1. Mycobacterial growth in Incubator tube MGIT (Mycobacterium Growth Indicator Tube) is one newculturing method, costlier to install and automated system. Economic limitations and timely availability ofreagents (closed system committed to the manufactures.) continue to hamper the growth of technologyin developing world2.. Recent success with MODS (the Microscopic Observation of drug susceptibility Assay ) developed inPeru gained the success as affordable, and primary drug resistance can be performed with simpleefforts, But inverted microscope is essential to read the results at frequent intervals. Contamination orhazard to technical personnel is minimal. Even the district laboratories can report resistance to Isoniazidand Rifampicin In spite of several controlled studies on MODS assay is poor to discriminate between,M.tuberculosis from Non Tuberculosis Mycobacterium. The success of MODS is a great breakthrough indetection of MDR strains provided the prevalence of NTM prevalence is low MODS assay can identifypatients with TB in approximately on third of time required for culturing on L J medium.Emerging and Rapid Diagnostic methods.-1 Fast plaque with phage amplification technology, tested in areas with high rates of HIV infection, hadcontradicting results, needs more understanding.2.Quanti - Feron TB test - Done on Blood specimens, based on the principle of ELISA and enzymelinked immunospot. With higher production of Interferon γ (Inf-γ) by cells in response to Mycobacteriumtuberculosis, than to the other environmental Mycobacterium in particular to Mycobacterium aviumcomplex. The testing results correlated with Tuberculin skin test reactivity, but still hampered in BCG
  3. 3. vaccinated.3Elispot - Tested by Elisa methodology detects Interferon γ produced by T lymphocytes in response tolatent TuberculosisInfection. Elispot gained more clinical acceptability and advantageous, being negative in majority of BCGvaccinated individualsBoth the above testing methods were limited to high end laboratories and cost of testing remained themajor limitation in many developing countries. More helpful to diagnose the latent TuberculosisInfections.Molecular Technology:The fast gains of Polymerase technologies by amplification of DNA (PCR) are limited to controlledstudies interpreted in relation to clinical context and performance of the laboratory .Rapidly changingmolecular technologies, out dating earlier hardware, other equipment and patented primers, added tolimitations in the Developing world. Mainly used as restricted research tool, and unaffordable to theneedy poor.Many extra pulmonary tuberculosis cases were benefited with Molecular technology.Future Goals in Control of Tuberculosis ;Stop TB partnership, Global Plan for 2006 to 2015 call for strengthening of network to facilitate detect allTB cases including smear negative tuberculosis. The Emphasis should focus on Sputum concentrationmethods, promoting the use of Fluorescent Microscopy. Helping the smaller laboratories to initiateculturing, and antibiotic sensitivity testing. The present affordable option may remain with utilizing themethodology of MODS .The Developing world wishes to utilize this upcoming technology for practical,and simple way to detect the MDR tuberculosis even at district Laboratories. Yet there is no fool proof, sensitive and specific test, which is inexpensive and rapid method forDiagnosing the Tuberculosis.Great challenges include detection and controlling of MDR TB. Strengthening the Smear Microscopy,and more aggressive provisions for enforcing the Fluorescent microscopy, may reduce the incidence ofspread of tuberculosis. We have to watch the Impact of X-MDR in the Indian continent. Theundergraduate and postgraduate Medical students should be taught with more emphasis on control of
  4. 4. drug resistant tuberculosis The best options with implementation of International standards fortuberculosis care with initiation of Major global health participation may bring hope to reduce theincidence of Tuberculosis by ; Share