Title: Increasing transmission and antimicrobic resistance of Mtbc
1Mycobacterial Services
- Increasing transmission and antimicrobic
resistance of Mtbc - introduction of new technologies require
adjustments in laboratory services
2Safety in the Mycobacteria Lab
- Active TB in Mycobacteriologists
- 100 x General Population
- 3 x Other Laboratory Workers
- Skin Test Conversions in TB Labs
- 8-30
3Safety in the Mycobacteria Lab (continued)
- Non-behavioral risk factors for infection
- Poor directional airflow
- Equipment failure
- Biological safety cabinets
- Centrifuges
- Autoclaves
- Specimens
- Number of positives processed
- Organism concentration
4Safety in the Mycobacteria Lab (continued)
- Behavioral risk factors for infection
- Lack of adequate safety guidelines
- Noncompliance with guidelines
- Aerosol production
- Pouring, pipetting mixing liquids
- Homogenizing cutting tissues
- Dropping or spilling culture flasks
- Preparing smears
- Needle sticks
5Laboratory Biosafety Levels
- Biosafety Level - 2
- Personnel trained in handling pathogens
- Lab directed by competent scientist
- Access to lab is limited during work periods
- Extreme precautions with sharps
- Aerosol-producing procedures conducted in
biologic safety cabinet
6Laboratory Biosafety Levels (continued)
- Biosafety Level - 3
- Training in handling agents acquired by
inhalation able to cause lethal disease - Supervised by scientists experienced in working
with such agents - P3 laboratory facilities and procedures
7Levels of Laboratory ServiceATS Recommendations
- Level I
- Collect, smear transport specimens
- Level II
- Smear culture ID sensi testing of Mtb
- Level III
- Smear culture ID sensi testing of all
mycobacterial isolates
8Level I Laboratories
- Tasks
- Collect good specimens
- Prepare examine smears
- Ship to Level II/III for culture sensitivities
- Smear proficiency test scores of gt 80
9Level I Laboratories (continued)
- Characteristics
- Receive minimum of 10 AFB smears/week
- Use fluorochrome stain
- Quantitate smear AFB report within 24 hr.
- Use certified biologic safety cabinet
- Send specimen for culture within 24 hr.
- Meet Dept. of HHS Biosafety Level 2 (BL-2)
requirements
10Sputum Specimen Collection
- Collection of good specimen is critical
- First morning expectoration
- Rinse mouth with water
- Produce specimen by deep coughing
- Expectorate directly into lab-provided tube
- If specimen inadequate, induce sputum
- Dispatch immediately to lab
- Containers to be closed and sealed
- Refrigerate if not processed immediately
11AFB Smears
- Level I Laboratories
- Recommended specimen sterilization
- Open in BSC sterilize with Clorox x 15 min
- Stain read on open bench
- Specimen cannot be used for culture
- Probably safe - not currently recommended
- Open, prepare, dry heat fix x 2h in BSC
- Stain read on open bench
- Dispatch sealed container to Level II/III lab
12Level II Laboratories
- Tasks
- Perform all procedures done by Level I
- Isolate organisms in pure culture
- Identify M. tuberculosis complex
- Perform susceptibility testing of Mtb
- Refer MOTT isolates for ID sensitivities
- Retain recovered isolates for 6 months
- Proficiency testing of isolation, ID and
sensitivity procedures
13Level II Laboratories (continued)
- Characteristics
- Receive minimum of 20 specimens/week
- Use Class II or III biologic safety cabinet
- Detect growth in liquid media in 10 days
- ID with DNA probes, BACTEC NAP or mycolic acid
profile within 2-3 weeks - Determine sensis of Mtb isolates in 3-4 w
- Meet Dept of HHS Biosafety Level 3 (BL-3)
requirements
14AFB Stains
- Concentrated Smear for Stain Culture
- Done in Level III laboratory BSC
- Liquify decontaminate
- Centrifuge gt 3000G x 15min in sealed cups
- Prepare smear, dry, heat fix x2 h
- Stain and read within BL-3 facility
- Flourochrome (auramine) stains preferred
15Level III Laboratories
- Tasks
- Identify all mycobacterium
- Perform susceptibility tests on MOTTs
- Proficiency testing on MOTT identifications
- Work concentrated in very few labs
16Level III Laboratories (continued)
- Characteristics
- Perform 20 susceptibilities per month
- USE DNA probes, HPLC or other rapid ID methods
- Do not rely only on NA amplification for ID
- Meet Dept HHS Biosafety Level 3 (BL-3)
requirements
17New Technologies
- Fluorescent microscopy
- Radiometric culture systems
- NAP test, DNA probes, HPLC
- Radiometric susceptibility testing
- Direct amplification of mycobacterial nucleic
acid from specimens
18New Technologies (continued)Direct Amplification
Techniques
- AMTD approved for smear respiratory specimens
- Requires 4-5 hours for completion
- Detects only M. tuberculosis complex
- Specificity approaches 100
- Sensitivity 90-100 less in smear negative
specimens
19AFB Stains
- Sensitivity on expectorated sputum
- M. tuberculosis 65
- MOTT 20
- Cum Sensitivity in series of 3 sputa
- M. tuberculosis 80
20AFB Stains
- Cum sensi of smears in culture series that yield
both a () culture for M. tuberculosis and one or
more positive smears - First specimen 86
- Second specimen 95
- Third specimen 95
- Fourth specimen 100
21AFB Stains (continued)
- Specificity
- Abs. False () 0.05
- Rel. False () 1.8
- Specificity 99.9
- PPV 98.1
22Culture
23Culture (continued)
24CDC Isolation Guidelines
- Initiation of Isolation
- Member of high risk population
- Contact with infectious TB patient
- Born in area of high TB prevalence
- Medically under-served populations
- HIV, elderly, homeless, imprisoned
- Alcoholics, IV drug users
- Appropriate clinical findings
25CDC Isolation Guidelines (continued)
- Initiation of Isolation
- Member of high risk population
- Appropriate clinical findings
- Cough gt 3 weeks
- Bloody sputum,
- Anorexia, weight loss
- Fever, night sweats
26CDC Isolation Guidelines
- Discontinuation of Isolation
- Patients with proven infectious TB
- Appropriate Rx (optimally 2-3w)
- Clinical Improvement
- 3 consecutive negative smears (q2w)
- Pt discharged to suitable locale
- Patients suspected of having infectious TB
- Alternate diagnosis confirmed
- Pt determined to be noninfectious
27Availability of AFB Smears
- CDC recommendations
- Results available within 24 hours
- No longer refer to STAT smears
28The End
29Timeline Comparison of TB MethodologyNew verses
Old
- Specimen
- Received
- Week 0 1
2 3 4 5
6 7 8 9 10 11 12 13 14 15
16 17 18 - ______________________
__________________________________________________
____________ -
-
- Mycobacteria growth
- Bactec 460 System
- TB DNA Probe ID
- (after growth)
- Bactec Sensitivities
MTD