lab methods: tuberculosis diagnosis · 2019-12-18 · grace lin, ms research scientist, microbial...

Curry International Tuberculosis Center, UCSF 2001 Center Street, Suite 700

Berkeley, CA; Office (510) 238-5100

LAB METHODS: TUBERCULOSIS DIAGNOSIS TB CASE MANAGEMENT AND CONTACT INVESTIGATION INTENSIVE

OCTOBER 8-11, 2019

LEARNING OBJECTIVE

Upon completion of this session, participants will be able to:

1. Utilize rapid identification methods for drug susceptibility for tuberculosis to improve patient

outcomes

INDEX OF MATERIALS

Lab methods: tuberculosis diagnosis slide outline Presented by: Grace Lin, MS

page 1-30

ADDITIONAL REFERENCES

CDC. Availability of an Assay for Detecting Mycobacterium tuberculosis, Including Rifampin-Resistant Strains, and Considerations for Its Use – United States, 2013. MMWR 2013;62 (No.41) October 18, 2013.

CDC. Guide to the application of genotyping to tuberculosis prevention and control. Page last updated September 1, 2012. www.cdc.gov/tb/programs/genotyping/manual.htm.

CDC. Report of an expert consultation on the uses of nucleic acid amplification tests for the diagnosis of tuberculosis. Page last updated September 1, 2012. www.cdc.gov/tb/publications/guidelines/amplification_tests/background.htm.

WHO. Drug-resistant tuberculosis. Frequently asked questions. January 2012. http://www.who.int/tb/challenges/mdr/tdrfaqs/en/# (Accessed November 10, 2014).

CDC. Updated Guidelines for Using Interferon Gamma Release Assays to Detect Mycobacterium tuberculosis Infection, United States. MMWR 2010; 59 (No.RR-5) See CDC factsheet Interferon-Gamma Release Assays (IGRAs). Page last updated September 1, 2012. http://www.cdc.gov/tb/publications/factsheets/testing/IGRA.htm.

Chang-Hong, S., Xiao-Wu, W., Hai, Z., et al. Immune responses and protective efficacy of the gene vaccine expressing Ag85B and ESAT6 fusion protein from Mycobacterium tuberculosis. DNA Cell Biol. 2008 Apr;27(4):199-207.

Honscha, G., Von Groll, A., Valenca, M., et al. The laboratory as a tool to qualify tuberculosis diagnosis. Int. J. Tuberc Lung Dis. 2008;12(2):218-20.

Perez-Martinez, I., Ponce-De-Leon, A., Bobadilla, M., et al. A novel identification scheme for genus Mycobacterium, M.tuberculosis complex, and seven mycobacteria species of human clinical impact. Eur. J. Clin. Microbiol. Infect. Dis. 2008;27(6):451-459.

Barman, P., Gadre, D., A study of phage based diagnostic technique for tuberculosis. Indian J. Tuberc. Jan. 2007; 54(1):36-40.

Haldar, S., Chakravorty, S., Bhalla, M., De Majumdar, S., Tyagi, JS. Simplified detection of Mycobacterium tuberculosis in sputum using smear microscopy and PCR with molecular beacons. J. Med. Microbiol. 2007;56(Pt.10):1356-62.

Nahid, P., Pai, M., Hopewell, P. Advances in the diagnosis and treatment of tuberculosis. Proc. Am. Thorac. Soc. 2006; 3:103-110.

http://www.cdc.gov/tb/programs/genotyping/manual.htm

http://www.cdc.gov/tb/publications/guidelines/amplification_tests/background.htm

http://www.who.int/tb/challenges/mdr/tdrfaqs/en/

http://www.cdc.gov/tb/publications/factsheets/testing/IGRA.htm

Curry International Tuberculosis Center, UCSF 2001 Center Street, Suite 700

Berkeley, CA; Office (510) 238-5100

Somoskovi, A., Dormandy, J., Mitsani, D., Rivenburg, J., Salfinger, M. Use of smearpositive samples to assess the PCR-based genotype MTBDR assay for rapid, direct detection of the Mycobacterium tubercuslosis complex as well as its resistance to isoniazid and rifampin. J. Clin. Microbiol. 2006;44(12):4459-4463.

Lin, G., Probert, W., Lo, M., Desmond, E. Rapid detection of isoniazid and rifampin resistance mutations in Mycobacterium tuberculosis complex from cultures or smearpositive sputa by use of molecular beacons. J. Cli. Microbiol. 2004;42(5):4204-4208.

Barnes, P., Cave, D. Molecular epidemiology of tuberculosis. NEJM. 2003;349(12): 1149-1155. Review article.

Dowdy, D.W., Maters, A., Parrish, N., Beyer, C., Dorman, S.E. Cost-effectiveness analysis of the gen-probe amplified mycobacterium tuberculosis direct test as used routinely on smear-positive respiratory specimens. J. Clin. Microbiol. 2003;41(3):948-53.

Drobniewski, F.A., Caws, M., Gibson, A., Young, D. Modern laboratory diagnosis of tuberculosis. Lancet Infect. Dis. 2003;3(3):141-47.

Van Der Zanden, A.G., Te Kopppele-Vije, E.M., Vijaya Bhanu, N., et al. Use of DNA extracts from Ziehl-Neelsen-stained slides for molecular detection of rifampin resistance and spoligotyping of Mycobacterium tuberculosis. J. Clin. Microbiol. 2003;41(3):1101-08.

Gillespie, S. Minireview. Evolution of drug resistance in Mycobacterium tuberculosis:clinical and molecular perspective. Antimicrob. Agents Chemother. 2002;46(2):267-274.

Mostowy, S., Behr, M.A., Comparative genomics in the flight against tuberculosis: diagnostics, epidemiology and BCG vaccination. Am. J. Pharmacogenomics. 2002;2(3):189-196.

Somoskovi, A., Mester, J., Hale, Y.M., Parsons, L.M., Salfinger, M. Laboratory diagnosis of nontuberculous mycobacteria. Clin. Chest Med. 2002;23(3):585-597.

Breese, P.E., Burman, W.J., Hildred, M., et al. The effect of changes in laboratory practices on the rate of false-positive cultures for Mycobacterium tuberculosis. Arch Pathol. Lab Med. 2001;125(9):1213-1216.

Somoskovi, A., Parsons, L.M., Salfinger, M. The molecular basis of resistance to isoniazid, rifampin, and pyrazinamide in Mycobacterium tuberculosis. Respir. Res. 2001;2(3):164-168.

Hale, Y.M., Desmond, E.P., Jost, K.C., Jr., Salfinger, M. Access to newer laboratory procedures: a call for action. Int. J. Tuberc Lung Dis. 2000;4(12 suppl 2):S171-175.

Grace Lin, MS

Research Scientist, Microbial Diseases Laboratory

California Department of Public Health

TB Case Management and Contact Investigation Intensive

Oakland, CA

October 8-11, 2019 1

Laboratory Methods:Tuberculosis Diagnosis

Grace LinResearch Scientist

MDL, CA Dept of Public HealthAt Curry Center

10 /8/19

Mycobacterium tuberculosis Complex(MTBC)

• M. tuberculosis

• M. bovis• M. africanum

• M. caprae

• M. microti

• M. pinnipedii

• M. mungi, M. orygis, M. canettii, M. suricattae(proposed)

Curry- Case management & investigation 10-8-19 2

All cause TB

Grace Lin, MS




Oakland, CA

October 8-11, 2019 2


MTBC

• Acid-fast bacilli (AFB)– NTM (non-TB-mycobacteria) are also AFB

– When AFB smear is positive, it can be MTBC or NTM

• Slow growing– Easily be overgrown by other bacteria

– Require special procedures to process specimen• Decontamination and concentration sediment

– Delayed turnaround time (TAT) • Molecular testing significantly shortens TAT

SFDPH retreat 9-18-2019 4

Specimen

Sputum: the most common specimen type

• Non-sterile source

– Rinse month with water before collecting

– First morning, deep cough sample—best yield

– 3-5 ml, collect in sterile, leak-proof containers

– Refrigerate if cannot send to lab immediately

• For diagnosis: 3 samples (CDC), 2 (WHO)

– Collect on different days or at least 8 hr apart.

Other specimen types:

BAL, pleural fl, lymph node, other tissues, CSF, etc


Grace Lin, MS




Oakland, CA

October 8-11, 2019 3


Sediment

CultureMolecular TestingXpert

PSQ, MDDR, WGS

MTBC NTM

cDST Genotyping

Smear

1 day 6 weeks

Specimenprocessing

Smear and Culture workup

Case 1

• 60 yr Chinese male came to U.S. to attend his son’s phD graduation

• Cough x 3+ weeks

• Lose weight? (pt said, no, no…)

• A sputum sample was collected for AFB smear & culture at Alameda PH lab

Curry- Case management & investigation 10-8-196

Grace Lin, MS




Oakland, CA

October 8-11, 2019 4


Specimen Processing Liquification by NALC

Decontamination by NaOH

Concentration by centrifugation


AFB Smear• Fluorochome stains

– AR (Auramine-rhodamine)

• Fluorescent microscope.

• Golden orange-yellow rods

• More sensitive than ZN or Kinyoun.

• Carbo fuchsin-based stains

– ZN (Ziehl-Neelsen) or Kinyoun stain

• light microscope.

• AFB--red. Non-AFB--blue

ZN stain

AR stain

AFB

Grace Lin, MS




Oakland, CA

October 8-11, 2019 5

Case 1

•Smear: AFB negative

•Let’s ask Dr. Chen if a molecular test should be ordered? •Yes! Please run Xpert.•FDA approved for testing sputa, smear-pos or

neg •But, test performance for smear-neg samples

is not as good as for smear-positive samples.



Xpert MTB/RIF Assay

Grace Lin, MS




Oakland, CA

October 8-11, 2019 6

Xpert MTB/RIF assay

• A great molecular test for: – MTBC detection and RIF-R detection– Cannot differentiate M. tb from M. bovis

• Realtime PCR, 5 molecular beacon probes, A to E– rpoB gene— 81 bp core region

• > 95% of RIF-resistant strains have mutations in rpoB• Assuming if mutations detected → RIF-R

– Yes, most times, but not always.

11Curry- Case management & investigation 10-8-19

12

Molecular Beacon probe(Hairpin structure—head & two arms)

Head (MTBC sequence)

Two arms (5-7 nt)

Fluorophore Quencher

At “rest” stage, two arms bind together forming a stem.Fluorophore is quenched. No signals are produced.

Grace Lin, MS




Oakland, CA

October 8-11, 2019 7


MB’s head does not bind to mutant SQ.Arms remain closed. No signals produced.

Mutation Detection with Molecular Beacons (Head containing wildtype SQ)

Mutant Sequence Wildtype Sequence

MB in actionMB at rest

MB’s head binds to wildtype SQ.Arms open. Fluorophore is away from quencher. Signals produced.

Critical Rules for Interpretation(Set by the Xpert software)

• MTBC detected – If 2 or more probes have signals = MTBC

• RIF-R detected– If a probe does not generate signals (Ct = 0), or

– If the highest and lowest Ct differs by more than 4 (Δ Ct max > 4)

• See more rules in package insert


Grace Lin, MS




Oakland, CA

October 8-11, 2019 8

Xpert Clinical Report

Procedure: MTB complex Nucleic Acid Amplification Test

Source: sputum

Final report (4 possibilities): 1. MTB complex not detected by PCR

2. MTB complex detected by PCR. RIF resistance not detected

3. MTB complex detected by PCR. RIF resistance detected

4. Detection of MTB complex was indeterminate


Case 1Xpert Clinical Report

Procedure: MTB complex Nucleic Acid Amplification Test

Source: sputum

Final report: MTB complex not detected by PCR


Grace Lin, MS




Oakland, CA

October 8-11, 2019 9

Xpert Raw Report

• Hidden information in raw reports

– Does SPC (specimen control) have signals?

– How many probes are up (having signals)?

– MTBC detected because how many probes are up?

– RIF-R detected by which probe(s)?

– Ct values?


18

Ct End-pt • SPC has signals: • No inhibitory substances.

• No probe has signals• MTBC: Not detected

• Does NOT rule out MTBC• MTBC DNA too low• No MTBC DNA (NTM?)• Wait for culture to grow

or test another specimen

10

SPC


Grace Lin, MS




Oakland, CA

October 8-11, 2019 10

Ask Dr. Chen for advice

•Test another sputum.

•MMWR 2/27/15•For smear-neg specimens, if test 2 sputa,

sensitivity increases from 55% to 69%.


Collect another sputum

• Jonny sets off to collect another sputum•He thinks how to help lab to produce better results….•A better specimen would help!!!

•Quality•First morning, deep deep cough

•Quantity•5 ml

•Send to lab ASAP or refrigerate. (Why?)


Grace Lin, MS




Oakland, CA

October 8-11, 2019 11

21

Ct values are high• DNA: low• Smear: negative5 probes up• MTBC detected• No mutation• Δ Ct max < 4

(33.9-31.8=2.1)• RIF-S

Ct End-ptCt End-pt

threshold


SPC

No RIF resistance detected.Great!

The 2nd sample worked!It was a better specimen.

What if the 2nd sample did not work?* MTBC may be present but too few cells

* Detection limit ~ 100 colonies/mL* Could be NTM/other microbes…..


Grace Lin, MS




Oakland, CA

October 8-11, 2019 12

Worst false results by Xpert

(when testing smear-negatives)

• Two probes positive = MTBC– a problematic rule

• NTM may be mis-identified as MTBC with RIF-R

–2 probes have low signals barely cross the threshold

–Means 3 probes are negative (no signals) = at least 3 mutations

–RARE to have 3 mutations within 81 bp of rpoB core

–Of 3754 tested, 2 samples had 3 mutations (CA data)• None detected by 3 probes

• One detected by 2 probes (A & B)

• One detected by 1 probe (D)

Curry- Case management & investigation 10-8-1923SFDPH retreat 9-18-2019

Next day, another TB suspect walks in…….


Grace Lin, MS




Oakland, CA

October 8-11, 2019 13

Case 2

• 82 yr male, US-born, no travel hx

• Cough x 4 weeks, weight loss

• No prior TB treatment

• Smear negative, culture pending

• Shall Xpert be ordered?

–Yes

–RIF-R suspected? No…..


25


Procedure: MTBC nucleic acid amplification test

Source: sputum

Final report:

MTB complex detected by PCR

Rifampin resistance detected

What?!

MUST Ask lab for Xpert raw report.


Grace Lin, MS




Oakland, CA

October 8-11, 2019 14

27

4 probes up• MTBC detectedProbe B: no signals• Mutations detected. • RIF-R? not always!Actions?

Ct Ct End-pt


Threshold

When Xpert detects RIF resistance

•Submit for sequencing• CA PSQ or CDC MDDR services• Verify if rpoB mutations are present• Identify the mutation SQ if present

• Silent or missence? • Disputed mutation? (Mutations conferring phenotypic R)• Predict levels of resistance to RIF & RFB*

• Additional molecular results• INH and 2nd-line drugs• Helpful for formulating treatment regimens

• *Ref: Berrada Z. Elsevere, Diag Micro & ID, 85(2016)


28

Grace Lin, MS




Oakland, CA

October 8-11, 2019 15

PSQ Detected a Silent Mutation

• 514TTT (433TTT using MTB codon)

– Great! It does not confer RIF-R.

– The most common silent mutation

• Found in ~70% of the mutations detectable by probe B

– It was interpreted as RIF-R by Xpert

• Xpert detects presence/absence of mutations, does not know if it is silent

– Other mutations detected by Probe B may confer RIF-R

• Before SQ results are available, evaluate RIF-R risk factors

Curry- Case management &

investigation 10-8-1929

Another TB suspect……


Grace Lin, MS




Oakland, CA

October 8-11, 2019 16

Case 3

•40 yr female from Philippines

•A nurse assistant works at SFGH

•Cough x 4 weeks, weight loss

•10 yr ago treated for TB

•A sputum sample was collected

•Smear 1+, culture pending

•Xpert test was ordered




Procedure: MTBC nucleic acid amplification test

Source: sputum

Final report:

MTB complex detected by PCR

Rifampin resistance detected

Not Surprised, unfortunately!

Ask lab for Xpert raw report.


Grace Lin, MS




Oakland, CA

October 8-11, 2019 17

33

4 probes up• MTBC detected

Probe E: no signal• Mutation detected• RIF-R detectedProbe E detects most common mutation in MDR

Actions• Send for sequencing

Ct End-ptCt End-pt




Pyrosequencing (PSQ)

• Realtime sequencing

• TAT: 1 day

• Can detect

– MDR:

• R to INH & RIF

– XDR:

• R to INH, RIF, fQs, injectable

drugs (AK/CM/KM)

Grace Lin, MS




Oakland, CA

October 8-11, 2019 18


Criteria for Requesting PSQ• Confirmation of RIF-resistance detected by Xpert.

• Drug-resistant TB is suspected– Immigrants from countries of high DR prevalence

– Contact of DR patients

– Previously treated cases / Not responding to treatment

• Patients have wide exposure or to vulnerable population

• Patients with vulnerable conditions (HIV+, transplant, etc.)

• Patients have adverse reactions to INH or RIF– Need to treat with 2nd-line drugs

• Laboratory issues– Smear-pos but culture-neg for undiagnosed TB

– Mixed cultures, cDST cannot be performed

– Confirmation of DR from culture-based DST

36

RIF (rpoB)

INH (katG)

PSQ

Grace Lin, MS




Oakland, CA

October 8-11, 2019 19

37

PSQ of pncAfor Identification of M. bovis

• Differential identification of:

– M. bovis

– MTBC-not-M. bovis

• M. bovis is PZA-R

• ID of M. bovis before culture grows

– Avoid PZA treatment


Grace Lin, MS




Oakland, CA

October 8-11, 2019 20

MDDR at CDC

• Require pre-approval

• Able to detect XDR. Molecular targets are:

– INH: katG & inhA.

– RIF: rpoB

– EMB: embB

– PZA: pncA

– fQ: gyrA

– Injectable drugs: rrs, eis, tlyA


mDSTGeneXpert PSQ

(MDL)

Sanger

(CDC)

Targeted NGS

(tNGS)

WGS

Specimen Sputum,

Raw sample or

sediments

All sources

sediments or cultures

All sources

sediments or cultures

All sources

Sediments or cultures

All sources

Pure cultures

TAT 2-3 hours 1-2 days 1-2 days 4-5 days

(~1 week)

4-5 days

(~1 week)

Drug RIF only INH, RIF, fQs,

AMK, CAP

INH, RIF, EMB, PZA

fQs, AMK, CAP,

KAN

Any drug if genetic

targets are known

Any drug if genetic

targets are known

Results Mutations

present or not.

(Mutation

identity not

provided)

Wildtype or mutant

sequences provided

(Hetero-R

can be detected but

not as sensitive as

tNGS)

Wildtype or mutant

sequences provided

(Hetero-R

can be detected but

not as sensitive as

tNGS)

Wildtype or mutant

sequences provided

Hetero-R

(Show % of R, best)

Wildtype or mutant

sequences provided

Hetero-R

(Not as reliable

/sensitive as tNGS)

Curry- Case management

& investigation 10-8-1940

3

Grace Lin, MS




Oakland, CA

October 8-11, 2019 21


Sediment

CultureMolecular TestingXpert

PSQ, MDDR

MTBC NTM

cDST Genotyping

Smear

1 day 6 weeks

Specimenprocessing

Workflow

Culture


Grace Lin, MS




Oakland, CA

October 8-11, 2019 22


Media

• To assure better recovery, liquid & solid media should be inoculated for primary isolation.– Different media provide different nutrients

• Liquid media– MGIT (BD)

– BacT/Alert (bioMerieux)

– Myco (VersaTrek)

• Solid media– Egg-based: LJ

– Agar-based: Middlebrook 7H10, 7H11• Plates: allow to view colonial morphologies to discern MTBC, NTM

or non-AFB, also possible to pick colonies if mixed

Liquid medium instruments Continuously monitor growth.

Detect growth faster than solid media


MGIT 960

MGIT tubes fluoresce when O2 is reduced/depleted.

Positive

Negative

Grace Lin, MS




Oakland, CA

October 8-11, 2019 23


Rough colonies on 7H10

MTBC


Cellular Morphology(seen on smears made from positive cultures)

MAC MTBC

Grace Lin, MS




Oakland, CA

October 8-11, 2019 24



MTBC Cording Clumps(seen on smears made from positive cultures)

47

Early cording clumps Older cording clumps


MTBC MAC

Colonial Morphology

(Microscopic)

Grace Lin, MS




Oakland, CA

October 8-11, 2019 25


Culture Identification • Cultures take 2-3 weeks to grow (smear 4+, may grow in 1 wk)

• DNA probes (AccuProbe)– MTBC

– M. kansasii

– MAC

– M. gordonae

• MALDI-TOF (mass spectrometry)

– Matrix assisted laser desorption ionization-time of flight

– Protein profiles

• Conventional methods

– pigmentation, biochemical tests, growth rate, etc.

• DNA sequencing

These 4 probes identify ~90% of AFB isolated in clinical labs.


cDSTMust set up from pure cultures

• Agar proportion (“conventional”)—21 days

• Modified proportion methods using liquid media

– MGIT (4-14 days, average 7-8 days)

– VersaTrek (4-13 days)

• Sensititre microdilution MIC method, (7 to 21 days)

– Not FDA approved yet

– Use growth from solid media to test, delayed TAT.

– Not available for testing PZA

– Drug panels can be customized for testing BDQ, LZ CF, etc.

Grace Lin, MS




Oakland, CA

October 8-11, 2019 26


Interpretation for ResistanceAgar Proportion method

(used at CDC)

• Resistant:

– growth in drug quadrant ≥ 1 % of growth in the control (with no drug).

• SIRE, ETA, KAN, OFX, RFB, PAS. (PZA by MGIT).

MGIT DST at MDL

• 1st-line: RIEP

• 2nd-line: MACE (MFX, AMK, CAP, ETA)

• Plan to validate for testing LZN, BDQ


Grace Lin, MS




Oakland, CA

October 8-11, 2019 27


Verification of Resistancedetected by MGIT

• Visual check MGIT tubes

– MGIT may appear turbid due to contamination.

• Quick confirmation of R by sequencing

– Mutations detected R results are confirmed

– Mutations not detected, retest from a pure culture.

• Review purity plate

– Even if culture is pure, contamination may be introduced during DST set-up. Need to check smear morphology on MGIT/VT showing R.

• Check smear morphology

– To differentiate MTBC vs NTM

– If smear morphology is not clear-cut, make sub onto 7H10 or 7H11 plates. Check colonial morph in a few days.

When you have doubtsin DR results

• It is critical to communicate with lab

• Ask lab how they verify the DR results

– DST must be set up from pure culture

– Insidious presence of NTM may not be detected initially

– Contamination may be introduced during set-up


Grace Lin, MS




Oakland, CA

October 8-11, 2019 28

When lab testing is not successful

• When need to retest, find out why, get a better sample

• No sequences yielded / MTBC not detected by Xpert

– MTBC DNA too low or not present

– If MTBC is strongly suspected, get a better specimen: deep cough sputum, higher volume.

– If MTBC is not strongly suspected, wait for culture to grow. ptmay be infected with NTM/other microbes.

• If culture is contaminated

– When collecting anther sputum, make sure pt rinses mouth thoroughly with clean water (bottled water)

– Deliver sample to lab ASAP or refrigerate. Curry- Case management & investigation 10-8-19 55

Time Frame for Results

Test Testing TimeTurnaround Time

(from date rec'd at lab performing tests)

AFB Smear 1 hr 1 day

Xpert 3 hr 1-2 days

PSQ 6 hr 1-2 days

MDDR (CDC) 1 day 1-2 days

Culture Positive 6 wk incubationAvg. 2-3 wk for positives

6 wk for negatives

ID by Accuprobe 1.5 hr 3-4 wk (tested once/wk)

Susceptibility 1-3 wk 4-8 wk (tested once/wk)


Grace Lin, MS




Oakland, CA

October 8-11, 2019 29


Genotyping

• Epidemiology surveillance

• Investigation of outbreaks

• Investigation of cross-contamination

• Differential diagnosis of relapse, reinfection, specimen collection error, or lab errors

Case 4*5 years ago, a 45 yr old male from Mexico was treated for pan-S

TB. At 6 wks into treatment, culture was converted and remained negative at the completion of treatment.

*Now, MTBC was isolated and tested R to RIPE by MGIT.

** Quite unusual, what happened?

• False DR? (mixed or contaminated?)

– No, culture was pure

• Wrong isolate? No, another current isolate yielded same results

• Acquired resistance or infected with a different strain?

• Let’s see Genotyping results………[to be continued…..]


Grace Lin, MS




Oakland, CA

October 8-11, 2019 30

Genotyping results (cont.)

• If the current and old isolates have different genotypes

– Reinfected with a different strain

• If the current and old isolates have the same genotype

– If the genotype is rare, likely it is acquired-R• A possibility is that the uncle acquired-R and he reinfected with the

uncle’s

– If the genotype is very common, • Possibly reinfected with a different strain which has the same

genotype. [WGS may have strong power to sort this out.]


Case 5

• A 64 yr old female came from Taiwan 40 years ago to attend a graduate school.

• She was seen by a family doctor at a private hospital because of coughing for more than 4 weeks.

• 3 sputa were collected and sent to a commercial lab for AFB & culture work-up. All 3 smear were negative.

• 6 weeks later the family doctor received a lab report stating M. tuberculosis complex isolated, Drug susceptibility testing pending. Other 2 sputa yielded no growth.

• Title 17 requires reporting of TB. The case was forwarded to a county TB control program.

• Something wrong?Curry- Case management & investigation 10-8-19 60

Grace Lin, MS




Oakland, CA

October 8-11, 2019 31

Thank you!


lab methods: tuberculosis diagnosis · 2019-12-18 · grace lin, ms research scientist, microbial...

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