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Foundations of TB Nurse Case Management

El Paso, TexasMarch 27 – 28, 2012

An Overview of the Laboratory’s Role in TB Diagnosis and TreatmentRole in TB Diagnosis and Treatment

Thor Elliott, MT (ASCP), AASMarch 27, 2012

Thor Elliott, MT (ASCP), AAS has the following disclosures to make:

•No conflict of interests•No relevant financial relationships

with any commercial companies pertaining to this educational pertaining to this educational activity

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Practical Guide to the Laboratory

Thor Elliott MicrobiologistTB Lab/Program Consultantthorelliott@rocketmail.com@

Objective

To describe the current laboratory methods for the detection, identification, drug susceptibility testing, and genotyping of Mycobacterium tuberculosis.Mycobacterium tuberculosis.

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Requirements to get a high quality specimen to the lab

Collection

Labeling

Requisitions

ShippingShipping

Collection

Garbage in Garbage outout.

Sputum not Spit

5 – 10 mls

Other fluids

Why do they always fill it when they submit urine?

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Labeling

UF Sticker

N Name

DOB

Date

Seal with parafilm or tape.

Requisitions

One Universal Form per specimen, not patient.

Fill out completely.

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Universal Forms

Shipping

Universal Form outside of plasticoutside of plastic container

Seal box with tape

Always follow local/state/federal shipping guidelinesshipping guidelines.

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Standard diagnostic techniques for the detection and identification of Mycobacteria

ReceivingP i Processing

Smears Cultures Probes HPLC HPLC Colonies Reports

Receiving

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Processing

BSL – 2 with BSL – 3 practicespractices

Very controlled and safe

Unpacked/Sorted Digested Concentrated Concentrated Set up Storage 3- 4 hour process

Smears

Read within 24 hours

P iti Positives are contacted by phone and faxed report

Negatives mailed out

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Nucleic Acid Amplification Test-NAAT

New PatientM t b iti ( lMust be a positive smear (unless extenuating circumstances)

Respiratory (sputum or bronch wash) No Tb meds Performed within 24 hours of smearPerformed within 24 hours of smear 4 hour process Very delicate

Cultures

Liquid culture (MGIT) on all specimens

M it d 24/7 d h Monitored 24/7 read every hour

Grown for six weeks or until positive

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Positive MGIT

St k d 7H11S Streaked on 7H11S media

Kinyoun stained

Positive stains are Identified

Negative stains are monitored

Kinyoun Stains

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The Monster

Cording

HPLC

Selection and SaponificationSaponification

Extraction

Sample Preparation

Chromatographs

4+ hours

2 days

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Chromatograph

Colonies

Slow grower

Rough Rough

Non-pigmented

35◦ C - 37◦C

Morphology is the final answer

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Report Timeline

PreliminarySmear Negative

PreliminarySmear Positive ( Smear Negative (same

day)

Final Final at 6 weeks

Smear Positive (same day)

Probe (if applicable 24hrs)

Identification (approx. 2 weeks)

Sensitivity if TB (10 days to 2 weeks))

Final If NTM final with ID

TB biochem report and final ID (batched)

Importance of acid-fast bacilli smear microscopy as a primary diagnostic tool

Primary tool for determining risk and treatment The hardest call we make is a 1+

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Smear Negative

Smear Positive

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Quantifying

Observed AFB Report

4 – 40 per 100 fields*1+

4 – 40 per 10 fields* 2+

4 – 40 per field*3+3+

> 40 per field* 4+

(using 40X objective)

Drug susceptibility testing of M. tb

Set up from positive MGITMGIT

Growth tube Done as a set Streptomycin

1.00ug/ml Isoniazid 0.10ug/ml Isoniazid 0.10ug/ml Rifampin 1.00ug/ml Ethambutol 5.00ug/ml Pyrazinamide

100.0ug/ml

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Drug susceptibility testing of M. tuberculosis

10 days to 2 weeks to complete (if growth is pure)pure)

Resistant results are rerun and sent to CDC for confirmation and second line drug testing.

MDR-TB Resistant to at least isoniazid and rifampin

XDR-TB Resistant to any floroquinolone, and resistant to at

least 1 of 3 injectible 2nd line drugs (ex: amikacin, kanamycin, or capreomycin)

Rapid methods for the identification and drug susceptibility testing of M. tuberculosis as they compare to the traditional methods

Genetic testingM l l B C lif i ( t b t Molecular Beacon – California (must be at least 3+)

MDDR – CDC

Special circumstance

Result will show predisposition not Result will show predisposition not definitive

Cost and labor prohibitive to do all patients.

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CDC program for genotyping M. tb isolates

1 specimen from every patientevery patient

Spoliogotype

Miru 1

Miru2

CDC program for genotyping M. tb isolates TBGIMS – TB

Genotype InformationGenotype Information management System

ubmitter Numb RVCT Number spoligotype Miru Cluster_nameed_false_Date Shipped Date Received patient_dob4113439 20040001 000000000003771 223325173533 KS 002 24 Mar 04 30 Mar 04 16 Feb 584113439 20040001 000000000003771 223325173533 KS_002 24-Mar-04 30-Mar-04 16-Feb-584237023 20060029 000000000003771 223325173533 KS_002 N0 25-May-06 26-May-06 09-Sep-804243582 20060024 000000000003771 223325173533 KS_002 N0 25-May-06 26-May-06 01-Jan-5610009689 20060028 000000000003771 223325173533 KS_002 NO 02-May-06 04-May-06 08-Sep-824223101 20050038 000000000003771 223325173533 KS_002 NO 19-Oct-05 20-Oct-05 05-Nov-534159793 20040064 000000000003771 223325173533 KS_002 NO 01-Sep-04 02-Sep-04 12-Nov-774108219 20040006 000000000003771 223325173533 KS_002 24-Mar-04 30-Mar-04 08-Aug-624132391 20040009 000000004020771 225313153223 KS_003 NO 03-Jun-04 04-Jun-04 18-Sep-674132405 20040035 000000004020771 225313153223 KS_003 NO 03-Jun-04 04-Jun-04 29-Sep-634113415 NAC 676773777777600 2z2324253322 KS_004 22-Jan-04 26-Jan-04 18-Jan-304113415 NAC 676773777777600 2z2324253322 KS_004 24-Mar-04 30-Mar-04 18-Jan-304231948 20050039 777776777760771 123325153323 KS_005 NO 19-Oct-05 20-Oct-05 20-Apr-554143550 20040013 777776777760771 123325153323 KS_005 NO 03-Jun-04 04-Jun-04 12-Nov-674072030 20030085 777776777760771 123325153323 KS_005 22-Jan-04 26-Jan-04 12-Dec-474092066 20030087 777776777760771 123325153323 KS_005 22-Jan-04 26-Jan-04 20-Feb-594092063 20040004 777776777760771 123325153323 KS_005 24-Mar-04 30-Mar-04 26-Oct-644235856 20050043 777776777760771 123325153323 KS_005 No 01-Dec-05 02-Dec-05 12-Aug-674152967 20040033 777776777760771 123325153323 KS_005 19-Aug-04 20-Aug-04 21-Aug-644167148 20050006 777776777760771 123325153323 KS_005 NO 09-Feb-05 10-Feb-05 29-Mar-62

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Questions?

La Senorita Margarita Guadalupe Hernandez

Contact InformationContact Information

Thor Elliott, MicorbiologistTB Laboratory/Program Consultanty g

thorelliott@rocketmail.comthorelliott.com

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