INSTRUMENT INTRODUCTION ITS USE SPECIMEN USED SAMPLE PROCESSING RESULT INTERPRETATION REPORT GENEXPERT MTB/RIF IN CONTEXT OF NEPAL

1. GENEXPERT MTB/RIF INSTRUMENT
Presented By:-Raghwendra Sah
GMC 2016 Batch
BSc.MLT Program12/29/2017
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2. Objectives
• Instrument introduction
• Use
• Specimen
• Sample processing
• Result
• GeneXpert MTB/RIF Nepal
• Conclusion
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3. Instrument introduction
• GeneXpert MTB/RIF is cartridge based
nucleic acid amplification test.
• GeneXpert MTB/RIF is automated diagnostic
test.
• It can identify Mycobacterium tuberculosis
(MTB) DNA and resistance to rifampicin (RIF)
by Nucleic Acid Amplification Test (NAAT).
• It was co-developed by the laboratory of
professor David Alland at the university of
medicine and dentistry of New Jersey
(UMDNJ)
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4. GENEXPERT MTB/RIF INSTRUMENT
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5. The GeneXpert Dx System automates and integrates sample
preparation nucleic acid amplification, and detection of the target
sequence in simple or complex samples using real-time Polymerase
Chain Reaction(PCR).
The system is suited for in vitro diagnostic applications that require
hands-off processing of patient samples (specimens) and provides
both summarized and detailed test results data in tabular and graphic
formats.
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6. GeneXpert Models
• There are different GeneXpert Dx instruments on the basis of
module present:
• The GeneXpert I instrument consists of one module (or one site)
to process one sample.
• Similarly, GeneXpert II, IV, XVI, consists of 2, 4,16 module (or 2, 4,
16 site) to process one sample.
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7. • The samples are prepared and processed in the
single-use, assay specific GeneXpert cartridges.
• The sample and applicable reagents are fed into a
cartridge, and then load the cartridge into one of
the available instrument modules.
• Each cartridge consists of following components:-
a) Processing chamber
b) Optical window
c) Valve
d) Reaction tube
GeneXpert cartridge
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Fig. Xpert MTB/RIF cartridge (top view).

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USES
1. To detect MTB.
2. To demonstrate mycobacterium is rifampicin resistance or not.
3. To process the sample of various site for various disease such
as CSF sample for Tuberculous meningitis(TBM), sputum
sample for MTB and pleural effusion for Tuberculous pleural
effusion.
4. To detect the extrapulmonary tuberculosis.
5. To detect MDR tuberculosis(multi drug resistant tuberculosis)
in HIV patient.

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Specimen
• Generally three types of specimens are used.
• They are:-
1. Sputum
2. CSF
3. Pleural effusion

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Sample processing

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Results interpretation
The results are interpreted by the GeneXpert DX System from measured fluorescent
signals and embedded calculation algorithms and will be displayed in the “View Results”
window. Lower Ct values represent a higher starting concentration of DNA template;
higher Ct values represent a lower concentration of DNA template.
MTB Detected
MTB target DNA is detected.
• MTB Detected—The MTB result will be displayed as High, Medium, Low or Very Low
depending on the Ct value of the MTB target present in the sample. Table 1 lists the Ct
value ranges for the displayed MTB results.
Table 1. MTB result name and Ct value range

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Fig. GeneXpert Dx System—Privileged User View Results window, MTB Detected Low, Rif
Resistance DETECTED

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• Rif Resistance DETECTED, Rif Resistance NOT DETECTED, or Rif
Resistance INDETERMINATE will be displayed only
in MTB DETECTED results and will be on a separate line from the MTB
DETECTED result.
• Rif Resistance DETECTED; a mutation in the rpoB gene has been
detected that falls within the valid delta Ct setting.
• Rif Resistance INDETERMINATE; the MTB concentration was very low
and resistance could not be determined.
• Rif Resistance NOT DETECTED; no mutation in the rpoB gene has been
detected.
• SPC— NA (not applicable); SPC signal is not required since MTB
amplification may complete with this control.
• Probe Check—PASS; all probe check results pass.

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MTB detected but RIF not detected
 MTB target DNA is detected.
 RIF target DNA is not detected, SPC meets acceptance criteria.
• RIF NOT DETECTED—RIF target DNA is not detected
• SPC— Pass; SPC has a Ct valid range and endpoint above the
endpoint minimum setting.
• Probe Check—PASS; all probe check results pass.

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Fig GeneXpert DX System—Privileged User View Results window, MTB Detected Medium, Rif
Resistance NOT DETECTED

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MTB not detected
MTB target DNA is not detected, SPC meets acceptance
criteria.
• MTB NOT DETECTED—MTB target DNA is not detected
• SPC— Pass; SPC has a Ct valid range and endpoint
above the endpoint minimum setting.
• Probe Check—PASS; all probe check results pass.

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Fig GeneXpert Dx System—Privileged User View Results window, MTB NOT DETECTED

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Reasons to Repeat the Assay
Repeat the test using a new cartridge or initiate alternate procedures if
one of the following test results occurs:
• An INVALID result indicates that the SPC failed. The sample was not
properly processed or PCR was inhibited.
• An ERROR result indicates that the Probe Check control failed and the
assay was aborted possibly due to the reaction tube being filled
improperly, a reagent probe integrity problem was detected, or because
the maximum pressure limits were exceeded or there was a GeneXpert
module failure.
• A NO RESULT indicates that insufficient data were collected. For
example, the operator stopped a test that was in progress.

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Report

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Pathology
Volume 49, Issue 1, January 2017, Pages 70-74
Microbiology
Evaluation of the GeneXpert MTB/RIF assay on extrapulmonary and respiratory samples other
than sputum: a low burden country experience
Author links open overlay panelSushilPandeyJacobCongdonBradleyMcInnesAlinaPopChristopherCoulter
https://doi.org/10.1016/j.pathol.2016.10.004Get rights and content
Summary
The aim of this study was to assess the performance of the GeneXpert MTB/RIF assay on
extrapulmonary (EP) and respiratory (non-sputum) clinical samples of patients suspected
of having tuberculosis (TB) from Queensland, Australia.
A total of 269 EP and respiratory (non-sputum) clinical samples collected from Qld
patients who were suspected of having TB were subjected to the GeneXpert MTB/RIF
analysis, Ziehl–Neelsen (ZN) staining, Mycobacterium tuberculosis (MTB) culture and
drug susceptibility testing. Phenotypic and genotypic data were compared.
The overall performance analysis of the GeneXpert MTB/RIF assay for detection of MTB
complex demonstrated sensitivity of 89%, specificity of 95%, PPV of 89% and NPV of
95% using culture as a reference standard. The GeneXpert MTB/RIF analysis of acid-fast
bacilli (AFB) smear positive samples and AFB smear negative samples showed
sensitivities of 100% and 77%, respectively.

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Looking at individual EP and respiratory (non-sputum) sample types, the sensitivity
ranged from 60% to 100% although the specificity ranged from 33% to 100% with the
specificity of lymph node tissue biopsy being the lowest. The GeneXpert MTB/RIF assay
detected 11% more TB cases than culture and 27% more cases than ZN microscopy.
Due to insufficient numbers of presenting rifampicin resistance cases, performance
analysis of the GeneXpert MTB/RIF assay on rifampicin resistance could not be carried
out.
The GeneXpert MTB/RIF assay is potentially valuable for TB diagnosis in the majority of
the EP and respiratory (other than sputum) samples in our setting. Although the
GeneXpert MTB/RIF assay provides rapid diagnostic results, the overall sensitivity to rule
out the disease is suboptimal for some specimen types. Performance varied according to
specimen type and AFB smear status. The sensitivity and specificity of lymph node tissue
was 63% and 33%. Care must be taken when using the GeneXpert MTB/RIF assay for
detection of MTB in lymph node tissue samples. All samples should be cultured
regardless of the GeneXpert MTB/RIF assay result

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Limitation
•The detection of MTB is dependent on the number of organisms present in the
sample, reliable results are dependent on proper specimen collection, handling,
and storage.
•Erroneous test results might occur from improper specimen collection, failure to
follow the recommended sample collection procedure, handling or storage,
technical error, sample mix-up, or an insufficient concentration of starting material.
•A positive test result does not necessarily indicate the presence of viable
organisms. It is however, presumptive for the presence of MTB and Rifampicin
resistance.
•Test results might be affected by antecedent or concurrent antibiotic therapy.
Therefore, therapeutic success or failure cannot be assessed using this test because
DNA might persist following antimicrobial therapy.

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GENEXPERT MTB/RIF NEPAL

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GENEXPERT MTB/RIF NEPAL
IOM, in close collaboration with NTP, has successfully
implemented TB REACH WAVE 2 Projects in Nepal
“Early and improved case detection of TB through the use of
GeneXpert technology in Nepal”
Started implementation in Nepal under W2Y1 (Oct. 2011- Feb.
2013)
Awarded for rollover to year 2 (W2Y2: Mar. 2013 - Apr. 2014)
Recently awarded for Wave4, with several new interventions in
the field (Jun 2014 – May 2015 with possibility of no cost
extension)

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TB case finding through use of point-of-care GX instruments
Increasing TB awareness and referrals among general population.
Increasing TB awareness and referrals among PLHA
To establish a referral system of smear-negative specimens for
GeneXpert testing and specimen of Rifampicin resistant cases for
Culture and DST

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FIG. DISTRIBUTION OF TB BURDEN AMONG DISTRICT BASED ON CNR
(ALL FORMS OF TB)

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INDIA
INDIA

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Populations
FIg: REGION WISE DISTRIBUTION OF
ALL TB CASE
FIG: ECO-TERRAIN WISE DISTRIBUTION OF
ALL TB CASE

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Populations
Table:Case notification (all forms of TB), 2015/16

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Populations
Table: Annual Gene Xpert test

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Populations

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Smear-negative individuals with high suspicion of TB
Individuals with high risk of MDR TB
Retreatment (Failure, Relapse and Return after default)
Close contacts of MDR TB patients
HIV+ with high suspicion of TB
Target group

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FIG: PERCENT DISTRIBUTION OF TB HIV TESTING AND
ENROLLMENT IN ART

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Microscopy
SS+SS-
Xpert/ Rif
Treat with
FLD
Confirm result with LPA or
Conventional DST
Treat with SLD
Further Clinical
Management
No MTB MTB+ / Rif SenMTB+ / Rif Res
CXR
Abnormal CXRNormal CXR
Smear not done
Test algorithm

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Conclusion
• In the various district of Nepal, mainly, of the terai region
,people are suffering from the tuberculosis.
• Among them most of the people have developed rifampicin
resistant.
• Only few of them have developed MDR MTB.
• And also the accuracy and specificity is almost 100%.
• The reliability and efficiency is also too good.
• The report is given within 2 hour.
• So, GeneXpert MTB/RIF instrument can used in NEPAL.

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https://www.ghdonline.org/uploads/all_in_one_-HPM40003.pdf
https://aidsfree.usaid.gov/sites/default/files/find_template.pdf
http://tbevidence.org/documents/rescentre/sop/XpertMTB_Broch_R9_EU.pdf
https://www.cdc.gov/tb/publications/factsheets/pdf/xpertmtb-rifassayfactsheet_final.pdf
http://nepalntp.gov.np/seminar/files/Nepal%20IOM%20TB%20Reach.ppt
https://www.ncbi.nlm.nih.gov/pubmed/27405129
http://onlinelibrary.wiley.com/doi/10.1111/tmi.12655/full
https://www.ncbi.nlm.nih.gov/books/NBK254320/
https://www.researchgate.net/publication/304527235_ADVANTAGE_OF_GENEXPERT_MTB
RIF_FOR_THE_DIAGNOSIS_OF_PULMONARY_TUBERCULOSIS_SUSPECTS_IN_NEPAL
http://nepalntp.gov.np/seminar/files/Nepal%20IOM%20TB%20Reach.ppt
https://www.google.com.np/search?q=GENEXPERT+MTB%2FRIF+NEPAL&oq=GENEXPERT+
MTB%2FRIF+NEPAL&aqs=chrome..69i57.20032j0j7&sourceid=chrome&ie=UTF-8
https://www.sciencedirect.com/science/article/pii/S1201971213002142
http://nepalntp.gov.np/theme/images/uploads/1500804055nnual_Report_2015.pdf
Xpert MTB/RIF Implementation Manual book
Refrence

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For more information, please contact:
NEXT GENERATION LABTECHNOLOGIST OF NEPAL
THANKS!!! THANKS!!! THANKS!!!

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The rifamycins are a group of antibiotics that are synthesized either naturally by the
bacterium Amycolatopsis rifamycinica or artificially.
The rpoB gene encodes the β subunit of bacterial RNA polymerase. It codes for 1342 amino
acids, making it the second-largest polypeptide in the bacterial cell.[1] It is the site of mutations
that confer resistance to the rifamycin antibacterial agents, such as rifampin.[2] Mutations in
rpoB that confer resistance to rifamycins do so by altering residues of the rifamycin binding site
on RNA polymerase, thereby reducing rifamycin binding affinity for rifamycins