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Diagnosis of Genitourinary Tuberculosis Dr. Jayesh Dhabalia Dr. Ulhas Sathe Consultant Urologist Professor & Head Sahyog Speciality Hospital Department.

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Presentation on theme: "Diagnosis of Genitourinary Tuberculosis Dr. Jayesh Dhabalia Dr. Ulhas Sathe Consultant Urologist Professor & Head Sahyog Speciality Hospital Department."— Presentation transcript:

1 Diagnosis of Genitourinary Tuberculosis Dr. Jayesh Dhabalia Dr. Ulhas Sathe Consultant Urologist Professor & Head Sahyog Speciality Hospital Department of Urology, & Lithotripsy Centre LTMG Hospital, Mumbai Jhamnagar

2 INTRODUCTION General Incidence – India 1 One fifth of world TB population 1.8 million new cases / year 8 lac infectious cases One death every 90 sec, 45 / hour, 1080 / day, 3.88 lack /year

3 INTRODUCTION General Incidence – India – GUTB1 Most common extra pulmonary TB 30 % of all extrapulmonary TB 18% of infertile women 11% of hematospermia 5-25 year after primary pulmonary TB Primary GUTB - anecdotal cases in females

4 MICROBIOLOGICAL BACKGROUND Classification M tuberculosis complex M tuberculosis – GUTB M bovis – Very rare M microtii- X M africanum - X DIAGNOSTIC PROBLEM Atypical / NTM / MOTT NO GUTB DIAGNOSTIC PROBLEM FALSE POSITVE Mycobacteria leprae NO GUTB

5 MICROBIOLOGICAL BACKGROUND Bacterial characteristics – diagnostic difficulties Slow growing - divides every 15–20 hours - Delayed growth on culture Can survive in a dry state for weeks - Infectivity & contamination Obligate aerobic Acid-fast -waxy coating on the cell surface makes the cells impervious to Gram staining

6 GUTB - DIAGNOSTIC DIFFICULTIES Characteristics of M. tuberculosis bacteria Difficult and delayed lab diagnosis Index of suspicion not high enough

7 GUTB - DELAYED DIAGNOSIS After significant irreversible anatomical changes - major surgical procedure ( thimble bladder, multiple ureteric strictures, multiple infundibular stenosis) Irreversible loss of kidney function Infertility in both gender Early diagnosis in India – high index of suspicion

8 8 GUTB – DIAGNOSTIC MODALITIES CLINICALRADIOLOGICALLABORATORY Reliability increases with Progression of disease Multisystem involvement Early changes Needs confirmation Advanced disease Almost diagnostic Valuable for Early disease diagnosis Rapid diagnosis Drugs sensitivity INVASIVE – Endoscopy / Biopsy / Dye study

9 GUTB - CLINICAL EVALUATION SPECIFIC - Genitourinary tract Lower urinary tract – 50 to 80 % – Burning, frequency, urgency, urge incontinence – Dysuria, hematuria – Suprapubic pain / perineal discomfort – Decreased stream, straining, ineffective voiding – Slough in urine

10 GUTB - CLINICAL EVALUATION Upper urinary tract symptoms – Pain - kidney and ureter region – Gross hematuria- 10 % Genital – Male Hematospermia - 10 % Azoospermia S/S of chronic epididymorchitis Genital – Female Menstrual irregularities Pelvic pain syndrome Infertility – 18 %

11 Other systems Respiratory - 12 % patients Gastrointestinal - 10 % Lymphoreticular Constitutional - 10 to 15 % 1 – Evening rise of temperature – Weight loss – Anorexia GUTB - CLINICAL EVALUATION

12 GUTB - LABORATORY DIAGNOSIS Supportive Urine – R&M Tuberculin test Serodiagnosis Potentially diagnostic ZN staining Culture PCR – urine, tissue Histological diagnosis

13 GUTB - PROBLEMS IN LAB DIAGNOSIS Paucibacillary Intermittent bacterial shedding Fastidious / slow growth – difficult to culture Diagnostic difficulty due to atypical mycobacteria / MOTT / NTM Sensitivity of all tests - extra pulmonary TB

14 GUTB - LAB DIAGNOSIS SUPPORTIVE TESTS Urine – routine and microscopy Acidic urine, sterile pyuria, microscopic hematuria Guide for further investigation, especially in pauci- symptomatic patients Montaux Test (Robert Koch in 1890) If Positive – supports the diagnosis If Negative – can not exclude extrapulmonary TB Response – HIV, Immunocompromised, Post-transplant pts Problems in India Invariably positive - Exposure in childhood, BCG vaccination

15 GUTB – SERODIAGNOSIS Basis – by detecting I. Specific immunological host response 1.Humoral (serological) antibody immune response to M. tb Ig G – High levels in active TB Ig M – Immediate appearance, disappears later on Commercial ELISA test kit available 2.T cell–based cellular immune response – Different antigens detects different types of TB Test kit not available II. Direct detection of bacterial antigens & metabolites Test kit not available

16 GUTB – SERODIAGNOSIS PROBLEMS Low sensitivity and specificity At present supportive at best Potential role in future – Early diagnosis – Response measurement to treatment – Early detection of relapse LITTLE OR NO ROLE IN DIAGNOSIS

17 GUTB - POTENTIALLY DIAGNOSTIC LAB TESTS COLLECTION OF SAMPLES Urine Physiologically pooled ( overnight) early morning 1st sample Three consecutive days, ? 5 days Volume – 10 ml for AFB, culture, PCR Immediate processing – if not feasible – refrigerate at 4-8 o C Alkalinisation of urine - ? Increased yield 2 Tissue In neutral transport media – avoids dessication Swab specimen – not suitable

18 GUTB - POTENTIALLY DIAGNOSTIC LAB TESTS 1. Availability2. Sensitivity3. Specificity4. Cost5. Time of processing6. Antimicrobial sensitivity TEST UTILITY EVALUATION CRITERIAS

19 GUTB - DIAGNOSTIC LAB TESTS ZN / AFB SMEAR EXAMINATION First described by Franz Ziehl (1859 to 1926), a bacteriologist and Friedrich Neelsen (1854 to 1894) Requiring 10 4 bacilli ml 1 of sample to achieve a positive result 3 Can not differentiate between live versus dead bacteria

20 GUTB - DIAGNOSTIC LAB TESTS ZN / AFB SMEAR EXAMINATION 1. Take sediment and make a smear 2. Carbol-fuchsin solution - allow slides to stand in hot solution for 5 minutes. 3. Wash in running tap water. 4. 1% Acid alcohol until light pink and color stops running. 5. Wash in running tap water for 5 minutes Working methylene blue for 30 seconds. 8. Rinse in water. 9. Dehydrate, clear, and cover slip. RESULTS: Acid-fast bacilli bright red Background blue

21 ZN - AFB STAIN TEST UTILITY EVALUATION Availability- Universal Sensitivity - 30 – 40 % 4 Specificity- 95% 4 Processing time- 45 mins Cost- Cheap Antibiotic sensitivity- NA

22 GUTB - DIAGNOSTIC LAB TESTS FLUORESCENCE MICROSCOPY Microscopy with fluorochrome dyes such as auramine O or auramine-rhodamine To increase sensitivity over ZN – AFB staining

23 TEST UTILITY EVALUATION Availability - Poor Sensitivity - 10 % > ZN staining 5 Specificity - Similar Processing time - Less than ZN staining Cost - Significantly > ZN staining Antibiotic sensitivity - NA

24 MOLECULAR TECHNOLOGY POLYMERASE CHAIN REACTION (PCR) Principle- by detection of species specific DNA Technique – to amplify a single or few copies of a piece of DNA to generate millions of copies of a particular DNA sequence Targeting different gene sequences of M. tuberculosis, showed different positivity – IS % - Most commonly targeted 6 – 65kDa - 75%, 38 kDa - 72%,85B protein - 73%

25 Diagnostic Problems Few Indian strains lack copy of IS 6110 IS present in M.tb complex bacteria ( tuberculous / africanum / microti / bovis) False positive – old cases of TB, contamination False negative – extremely paucibacillary cases ( detection limit up to 10 copies / ml ) Cannot differentiate dead from live bacteria 25 MOLECULAR TECHNOLOGY POLYMERASE CHAIN REACTION (PCR)

26 MOLECULAR TECHNOLOGY - PCR Universal Sample Processing (USP) Technology AIM - To false negative rate by removing inhibitors of DNA amplification Potent inhibitor-heparin, hemoglobin, phenol & sodium dodecyl sulfate 4 Detected in 0 to 20% of the clinical specimen 4

27 Thermal cyclingConsisting of cycles of repeated heating and cooling of DNA for meltingEnzymatic replication of the DNA. Primers (short DNA fragments) containing sequences complementary to the target region As PCR progresses, the DNA generated is itself used as a template for replication DNA template is exponentially amplified MOLECULAR TECHNOLOGY - PCR TECHNICAL PROCESSING

28 MOLECULAR TECHNOLOGY - PCR Availability- Limited Sensitivity - 75% - 95% 6 Specificity % 6 Processing time - One day Cost - Rs 1600 Antibiotic sensitivity - NA TEST UTILITY EVALUATION

29 MOLECULAR TECHNOLOGY RNA PCR DNA PCR cannot differentiates dead from live organisms Principle – Reverse transcriptase copies the RNA target from DNA into a transcription complex, which is then amplified by RNA polymerase

30 MOLECULAR TECHNOLOGY TISSUE PCR Sampling limitations Availability- Limited Sensitivity - 95% Specificity - 95 to 97 % Processing time - One day Cost - Rs 1600 Antibiotic sensitivity- NA TEST UTILITY CRITERIA

31 M.TUBERCULOSIS CULTURE Gold standard ???? Highly specific and sensitive Detects presence of live bacteria Can differentiate between typical and atypical Antimicrobial sensitivity Biggest drawback – time required

32 M.TUBERCULOSIS CULTURE MEDIA Solid Egg-based - Petragnini medium and Dorset medium Middlebrook 7H10 Agar Middlebrook 7H11 Agar Blood based -Tarshis medium Serum based - Loeffler medium Potato based - Pawlowsky medium LJ media Liquid Dubos' medium Middlebrook 7H9 Broth Proskauer and Beck's medium Sula's medium Sauton's medium Radiometric – Bactec Nonradiomaetric – MGIT / MB Redox

33 M.TUBERCULOSIS CULTURE Solid Media LJ media Liquid Media Radiometric Non Radiometric Bactec 460 TB system MGIT MB Redox

34 M.TUBERCULOSIS CULTURE SOLID MEDIA Growth medium specially used for culture of Mycobacterium Processing time – 2-8 weeks Time increases in pauci bacillary specimen Further 2-8 weeks in typical v/s atypical -? Simultaneous culture with inhibition of M.tb Further 2-8 weeks for drug susceptibility LOWENSTEIN-JENSEN MEDIUM

35 M.TUBERCULOSIS CULTURE LOWENSTEIN-JENSEN MEDIUM Composition Malachite green,Glycerol, Asparagine, Potato starch, Coagulated eggs, Mineral salt solution (Potassium dihydrogen phosphate, Magnesium sulfate, Sodium citrate ) Penicillin and nalidixic acid -Inhibit growth of gram positive and gram negative bacteria,. Presence of malachite green in the medium inhibits most other bacteria.

36 M.TUBERCULOSIS CULTURE More sensitive and specific Shorter processing time Variable availability Simultaneous differentiation of atypical v/s typical LIQUID CULTURE MEDIA

37 M.TUBERCULOSIS CULTURE RADIOMETRIC BACTEC 460 TB system Principle – Detection of the metabolism of the bacteria – Detects 14 CO 2 liberation during the decarboxylation of 14 C labelled substrates palmitic acid Differentiates typical and atypical mycobacteria 4 – p-nitro benzoic acid (PNBA) test – The BACTEC NAP test

38 M.TUBERCULOSIS CULTURE NONRADIOMETRIC Principle – detection of metabolism of bacteria Types – Mycobacteria Growth Indicator Tube (MGIT) – MB Redox If bacteria present Oxygen consumed in metabolism Fluorescence Vial is positive

39 M.TUBERCULOSIS CULTURE LIQUID MEDIA (1) Mycobacteria Growth Indicator Tube (MGIT) Sensor Contains 4 ml of modified Middlebrook 7H9 broth with an oxygen quenching- based fluorescent sensor. Nonradiometric Liquid Culture Media

40 M.TUBERCULOSIS CULTURE LIQUID MEDIA (2) MB Redox Tube Four ml of modified, serum-supplemented Kirchner medium with a colourless tetrazolium salt as a growth indicator. During bacterial growth, the tetrazolium salt is reduced to a pink-, red-, or violet-colored formazan. Contains a special vitamin complex which provides for a considerable acceleration of the growth of mycobacteria Nonradiometric Liquid Culture Media

41 M.TUBERCULOSIS CULTURE SOLID V/S LIQUID MEDIA Solid mediaLiquid media Availability GoodVariable Sensitivity %80-95 % Specificity % Processing time 4-8 weeks12-15 days Cost 1500 Antibiotic sensitivity ++ Hence liquid media is preferred

42 TEST UTILITY EVALUATION RADIOMETRIC V/S NONRADIOMETRIC RadiometricNon radiometric BactecMGITMB redox Availability VariableMost commonUncommon Sensitivity % %80% Specificity 95-97% Processing time days17 days16 days Cost Antibiotic sensitivity +++ Technical difficulty Labor intensive Radiation hazards

43 GUTB DIAGNOSIS ENDOSCOPY & BIOPSY Indications – Suspected GU Koch's with equivocal radiology and lab test – Ureteric evaluation ( RGP) and stenting Risks – Bladder perforation, septicemia Problems - Morphology - If normal no biopsy - Changes - specific / nonspecific biopsy ? Need – Highly suggestive radiological findings


45 GUTB - ENDOSCOPY & BIOPSY Classical Histological features Granuloma formation Caseous necrosis Cavitation Chronic inrterstital inflammation


47 Differential diagnosis - Fungal infection: histoplasmosis & cryptococcus Infections: – Cat-scratch fever (caused by bartonella henselae) – Chronic pyelonephritis, sarcoidosis Wegener's granulomatosis: Drugs Very rare compared to M.tb infection 47 GUTB DIAGNOSIS ENDOSCOPY & BIOPSY

48 Availability- Universal Sensitivity - 18 to 56 % 6,12,13 Specificity - 95 to 97 % Processing time - four days Cost - Rs 500 Antibiotic sensitivity- NA

49 Bivalved resected specimen shows two foci of caseous necrosis in the upper pole (arrows). GUTB DIAGNOSIS ENDOSCOPY & BIOPSY

50 DIAGNOSTIC LAB TESTS COMPARISON AFB StainPCRLJ CultureBactecMGITMB redox AvailabilityUniversalLimitedCommonLimited Uncommon Sensitivity40 – 60 %95%40-70 %80-95 % %80% Specificity % Processing time 45 minsone day4-8 weeks12-14 days17 days16 days CostRs 500Rs 1600 Rs Antibiotic sensitivity NA ++++

51 GUTB LAB DIAGNOSIS PROBLEMS - No Gold Standards Current Literature - Not all test done in all patients - Different samples from different systems Sensitivity & Specificity - Maximum with combination of tests Recommendation – PCR with Bactac Radiometric Culture 51

52 Persistent Irritative voiding &/OR genital symptoms History of recurrent UTI, past H/O TB, Failed symptomatic t/t Urine – sterile pyuria, acidic, microscopic hematuria USG Specific NonSpecific IVP Urine-PCR/AFB stain & Culture Positive AKT Negative Endoscopy Morphology Specific / Non-Specific Biopsy –HPE Tissue PCR Positive Negative Clinical Decision AKT ? Normal No Biopsy DIAGNOSTIC ALGORITHM

53 Bibliography Das P, Ahuja A, Gupta SD. Incidence, etiopathogenesis and pathological aspects of genitourinary tuberculosis in India: A journey revisited. Indian J Urol 2008;24: Debra L. Piddington et al; Infection and Immunity, August 2000, p , Vol. 68, No. 8 10th Edn, Topley and Wilson. Bacteriology; Vol. 2, pg 1190 Negi SS, Khan SF, Gupta S, Pasha ST, Khare S, Lal S. Comparison of the conventional diagnostic modalities, bactec culture and polymerase chain reaction test for diagnosis of tuberculosis. Indian J Med Microbiol 2005;23:29-33 Nguyen Van Hung et al; Fluorescence microscopy for tuberculosis diagnosis ; The Lancet Infectious Diseases - Volume 7, Issue 4 (April 2007) A. K. Hemal, N. P. Gupta et al ; Polymerase Chain Reaction In Clinicall Suspected Genitourinary Tuberculosis: Comparison With Intravenous Urography, Bladder Biopsy, And Urine Acid Fast Bacilli Culture ; UROLOGY 56: 570–574, 2000 Venkataswamy MM, Rafi W, Nagarathna S, Ravi V, Chandramuki A. Comparative evaluation of bactec 460tb system and lowenstein-jensen medium for the isolation of M. tuberculosis from cerebrospinal fluid samples of tuberculous meningitis patients. Indian J Med Microbiol 2007;25: P Anargyros, D S Astill and I S Lim; Comparison of improved BACTEC and Lowenstein-Jensen media for culture of mycobacteria from clinical specimens ; J Clin Microbiol June; 28(6):

54 9.H.P. Chien; M.C. Yu; M.H. Wu; T.P. Lin; K.T. Luh; Comparison of the BACTEC MGIT 960 with Löwenstein-Jensen medium for recovery of mycobacteria from clinical specimens ; The International Journal of Tuberculosis and Lung Disease, Volume 4, Number 9, September 2000, pp (5) 10.L. Heifets, T. Linder, T. Sanchez, D. Spencer, and J. Brennan; Two Liquid Medium Systems, Mycobacteria Growth Indicator Tube and MB Redox Tube, for Mycobacterium tuberculosis Isolation from Sputum; Journal of Clinical Microbiology, March 2000, p , Vol. 38, No. 3 Specimens 11.W. K. Chew 1, R. M. Lasaitis, F. A. Schio and G. L. Gilbert; Clinical evaluation of the Mycobacteria Growth Indicator Tube (MGIT) compared with radiometric (Bactec) and solid media for isolation of Mycobacterium species; J Med Microbiol 47 (1998), Wong SH, Lau WY, Poon GP, et al: The treatment of urinary tuberculosis. J Urol 131: 297–301, Gow JG: Genito urinary tuberculosis, in Walsh PC, Retik AB, Stamey TA, et al (Eds): Campbells Urology, 6th ed. Philadelphia, WB Saunders, 1992, vol 1, pp 951–981

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