Title: British Society for Microbial Technology
1British Society for Microbial Technology
- The laboratory diagnosis of tuberculosis
- 25 years of progress
D A Mitchison St Georges, University of London
With assistance from FINDdiagnostics
2Diagnostic testing at different levels of health
system
3Sputum 25 years ago (1985)
- Poor countries Microscopy alone
- Richer countries. Microscopy, LJ culture ,
DST - Advanced countries. Microscopy, Liquid culture,
ID, DST
4Sputum bacteriology UK (1985)
- Direct smears
- Culture on LJ slopes (3-6 weeks)
- Identification as M. tuberculosis
- (Chemical PNB, niacin, catalase)
- Drug susceptibility tests (DSTs)
- (Rifampicin screen)
5- FIND and Carl Zeiss Micro Imaging GmbH have co-
developed a fluorescent LED microscope based on
the proven Primo Star platform. FIND/Zeiss
microscope offers superior optics, reflected
light illumination, easy switch from brightfield
to fluorescent light
6Direct smears
- Fluorescence v. Bright field microscopy
- Fluorescence
Introduced in 1940s. 5x more rapid than Bright
field BUT Mercury vapour bulb
Expensive. Limited life. Gradual
decline.
LED illumination introduced during past 5
years Find/Zeus collaboration
7Culture solid v. liquid
- Solid LJ slopes. 7H11 slopes or plates.
- Liquid Early attempts high contamination.
- 1971 Selective medium paper
- (Mitchison et al J Med Microbiol
1971 5 165) - Penta used in Bactec machine
- Automated liquid systems v. solid media
- Sensitive. Rapid.
- Contamination. NTMs v. TB.
8Genetic systems
Sensitivity
9Culture, identification DSTs
- HAIN MDTBDR plus PCR Line probe based 1.
Identifies as TB complex. - 2.DSTs for RIF INH (95)
- Can be used directly on sputum avoiding culture
10What to do about MDR TB?(MDR Resistance to INH
RIF)
- Genetic tests for reserve drugs not adequate yet.
Therefore cultures in liquid or on solid medium
necessary as well as genetic techniques.
11Reserve drugs
12MGIT 960 Reserve Critical Concentrations
1Rusch-Gerdes S et al. JCM 200644688-92. 2Rodri
gues C et al. IJTLD 2008121449-55. 3Kruuner A
et al. JCM 200644811-8.
13DSTs
- Phenotypic
- Classic on LJ slopes or 7H11 plates. Takes 7
weeks . - MGIT or other automated liquid tests.
- Microcolony methods
- Liquid medium Mods.
- Sensitive, time consuming, ?dangerous
- Solid medium Thin layer agar (TLA)
- Quicker. Less dangerous
14Phenotype DSTThin-layer agar plate (TLA) method
- 7H11 thin layer plates made selective
- Each plate with up to 6 strains in quadrants
- Control no drug
- PNB (p-nitrobenzoate) TB inhibited.
- INH 0.2 µg/ml
- RIF 2.0 µg/ml
- SM 2.0 µg/ml
- PZA 2,000 µg/ml nicotinamide
- etc
15What is drug resistance?
Defined from distribution of MICs on wild
strains
16Studies of early bactericidal activity define
the therapeutic margin
17Can high drug dosage still have an effect on
resistant strains?