This document discusses diagnostic methods for genital tuberculosis. It covers direct visualization techniques like culture as well as non-culture methods like nucleic acid amplification tests. It notes that conventional culture methods using Lowenstein Jensen media are slow, taking 8 weeks for results. Newer liquid culture systems like MGIT and BacT/Alert allow for faster diagnosis within 1-4 weeks. Molecular tests like PCR and line probe assays can also rapidly detect Mycobacterium tuberculosis and determine drug resistance directly from clinical samples. The document emphasizes using a diagnostic algorithm that combines smear, culture and non-culture tests to improve definitive diagnosis of genital TB.

1. Diagnosis of Genital Tuberculosis
Direct Visualization Techniques
Culture : Conventional & Rapid
Dr Ashok Rattan,
Chairman : Laboratory Medicinee
1

2. Commonest method for diagnosis of TB
1882 2002
2

3. Slow diagnostic methods
Twentieth Century Mentality
AFB smear
Culture on LJ
Identification by Niacin &
Nitrate test
Sensitivity by RR
Culture report in 8wk
DST report in 8 wks
Treatment duration from 9 M to 18 M
3

4. Progress made in
laboratory diagnosis of tuberculosis
Fluorochrome stain for AFB smear Mid 1970s
BACTEC 460TB Early 1980s
DNA probes for AFB +ve respiratory Late 1980s
Automated broth systems for culture Early 1990s
HPLC on AFB +ve cultures Early 1990s
Drug susceptibility by Bactec Early 1990s
Nucleic acid amplification on all smear +ve 1996
Use of courier to transport specimen 1997
Lipa for identification and Rif resistance 1997
Use of NAA TB testing on smear –ve 2000
Reporting of results by fax 2000
Genotype MTBDR for subspecies 2003
Expert group recommends Hain’s test 2008
Expert group recommends Xpert system 2011 4

5. Genital Tuberculosis
Hospital Associated Infection
Tuberculosis :
1. Pulmonary
2. Extra pulmonary
• Genital tuberculosis:
represents 15 – 20% of all Extra pulmonary cases
12% of pulmonary tuberculosis cases
20% of infertility cases have tuberculosis
5

6. Microbiology
90 to 95% due to Mycobacterium tuberculosis
5 – 10% due to M bovis
Mostly secondary to TB elsewhere
Rarely primary : direct inoculation (? STD)
Routes of infection:
• Haematogenous : 90% from lungs, LN, Skeletal system
• Descending : via lymphatics
• Directly from GIT, mesenteric LN or peritoneum
6

7. Genital Organs Infected
Fallopian tube 95 – 100 %
Endometrium 50 – 60 %
Ovaries 20 – 30 %
Cervix 5 – 15 %
Vulva / Vagina 1%
Myometrum 2.5 %
Haematogenous spread: involvement of submucosa, endosalpingitis
7
Direct spread: exosalpingitis, tubercles on the surface

8. Diagnosis
• Combination of
• high index of suspecion
• thorough clinical examination
• Appropriate investigations
• High Risk Factors
• Previous pulmonary tuberculosis or contact with a case
• Socio economic background
• Drug use
• HIV status
• H/o chronic chest symptoms 8

9. Investigations
Full blood counts, ESR, HIV test
TST or Quantiferon Gold IT
Chest X Ray
Smear & Culture
Histopathology
NAAT : PCR, SDA, TMA, NASBA, LAMP, RT PCR
Imaging
Hysterosalpingogram 9

10. Culture : Conventional & Rapid
Samples:
• Biopsy of suspected lesion in accessible site
• Endometrial tissue (pre menstrual period)
• Menstrual fluid (on 1st day of cycle)
• Urine (early morning x 3)
• Blood for Quantiferon Gold IT
• Serological tests for IgG/IgM/IgA  NO diagnostic value
• WHO has expressed NEGATIVE opinion on their use
• Govt of India should BAN import of Serological test kits
• clinicians should STOP requesting it 10

11. AFB SMEAR
11

12. PNA
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13. Culture : Conventional
Slow : weeks
Lowenstein Jensen (LJ) Low sensitivity (102 AFB/ml)
13
Middlebrook 7 H 10 Agar

14. Dr Tanil Kocagoz
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15. Liquid Culture : Rapid
Bactec 460
C Palmatic Acid  14CO2
14
MGIT 960
15

16. Liquid Culture : Rapid
BacT/Alert 3D MB
16

17. Microscopically Observed Drug Susceptibility
(MODS)
Zip lock,
examined from 4 to 15 days daily, A/D 40
24 wells plate (BD),
12 wells used per sample, 4 drug free control, 8 containing drugs
Middlebrook 7H9 broth (BD) INH 0.1 & 0.4 ug/ml
OACD RMP 1 & 2
PENTA Etham 2.5 & 5
720 ul inoculum used SM 2 & 6
17

18. Thin Layer Agar
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19. MDR & XDR TB are Laboratory Diagnosis
Clinicians should not label “non responding” infections MDR
M. tuberculosis
Drug susceptible
INH + RIF Resistant
M. tuberculosis
= MDR
Drug susceptibile
FQ Injectable
XDR
Resistant Resistant
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20. Clinicians should not label “non responding” infections
MDR without Laboratory support
MDR TB XDR TB
2 most important INH INH
RIF RIF
1 line drugs
st
PZA PZA
Etham Etham
2 most important FQ FQ
Injectable Injectable
2nd
line drugs
Ethio Ethio
Cyclo Cyclo
PAS PAS
20

21. Sample & culture positive load for AFB Bactec at GGN
sample number = 1656
885
900
Culture Positive
800
Total Samples
700
600
500
379
400
245
300
200 157 158
69
100 31 55 21 43 44
10 2 4 3 3
0
P ulmo na ry L ymph N o de T is s ue E ndo me tria l B o dy F luids CSF Urine O the rs
& P us
21
Data courtesy Religare SRL Gurgaon Lab

22. Total samples processed = 1656 (smear +ve=302, --ve=1354)
Total culture positive =453 (M tb=393, MOTT=60)
Smear Mycobacteria MGIT LJ +ve Speed of Speed of
Result spp. +ve growth on growth on
MGIT in days LJ in days
Smear M.tb 258 191 9.64 26.12
Positive
(n=302) MOTT 21 10 6.6 28
Smear M.tb 133 66 17.15 34.28
Negative
MOTT 41 8 13.79 30.62
(n=1354)
Data courtesy Religare SRL Gurgaon Lab 22

23. Liquid Culture improves detection and speed of definitive
diagnosis
Speed Of Growth Of M.tb (MGIT 960 vs LJ Media)
166
132
63
55
46 48
23
23 24
13 2
0
1st wk 2nd wk 3rd wk 4th wk 5th wk 6th wk
Data courtesy Religare SRL Gurgaon Lab 23

24. Genital TB Algorithm
21st century approach
Tests Results
Smear + -- -- + --
&/or
culture
NAAT + + -- -- --
Quantiferon + + + -- --
Gold IT
Diagnosis Genital TB Genital TB Latent MOTT No
TB contamination evidence
of TB
infection
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25. Carry Home Messages : Diagnostic Methods
• Smear is rapid but insensitive (104 AFB/ml),
can be improved by using Fluorescent
Microscopy
• Liquid culture is sensitive & specific, faster
than solid culture but still slow
• Diagnostic tests : culture & non culture tests
should be used simultaneously in an
algorithm to improve definitive diagnosis
• Serological tests, available at present, have
no diagnostic value 25

26. Carry Home Messages : Diagnostic Methods
5. Laboratory must validate tests that they use
6. Laboratories must participate in EQAS
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Data courtesy Religare SRL Gurgaon Lab

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