2 Recommendations

RIF registant case detected by GeneXpert MTB/RIF found RIF sensitive by MGIT960

By Md. Shamim Hossain | 20 Jun, 2015

A very few cases are detected Rifampicin (RIF) resistant(R) by GeneXpert MTB/RIF method but RIF sensitive(S) by AST result in MGIT960 (liquid culture) method. Both the results are getting from the same specimen of the respective patient. After getting GeneXpert result the patient is started MDR-TB treatment according to treatment category. But some clinician raise the question about the laboratory result in these short of cases. They asking why a few cases are happening like this discordant type of result? Which result should be considered finally?



Fulgence Nzabintwali Replied at 3:08 PM, 20 Jun 2015

Use Gene gene x pert result

If necessary send sample to your supra national laboratory for extra molecular test

Bontle Mbeha Replied at 3:17 PM, 20 Jun 2015

The crucial thing to note is that genexpert picks DNA. It does not make a
distinction between viable and non viable organism. On the other hand,
culture only picks viable organism so if the organisms are responding to
treatment they will be sensitive with culture . I believe culture is still
the gold standard

Somsak Rienthong Replied at 10:30 PM, 20 Jun 2015

Xpert detected point mutation but sometime did not express so in Phenotypic DST can show RIF sensitive. For Xpert, if you know exactly the history of the patient, in new case should be confirm by other method, by other molecular or MGIT, and clinician can treat with according to the latter result. But in previously treated cases or suspected MDRTB if Xpert showed RIF resistant, MDR regimen should be treated. MGIT RIF sensitive, should be aware false result.

Prince James Replied at 2:49 AM, 21 Jun 2015

I agree with Somsak. Genexpert is a new test and discordance is being
noted and also being published all over the world. And also discordance
is being noted in both directions- xpert sensitive mgit resistant and
vice versa.
We should interpret results in clinical scenario. In a patient of
suspected MDR Tb, xpert showing resistance to rifampicin is more
reliable, than a mgit showing rifampicin sensitive in same sample. Pt
should be started on MDR Tb treatment.

Leen Rigouts Replied at 4:40 AM, 21 Jun 2015

It is known that liquid-medium-based phenotypic tests like MGIT960 can miss some rpoB mutants with demonstrated clinical significance (e.g. 511Pro, 533Pro). On the other hand, could Xpert also yield false-RIF resistant results in case of non-synonymous mutations for instance of in case of high Ct values (low bacillary load).

Bacteriologically, the following can be done/checked:
1. check the Ct values and Xpert intrepretation (check graphs)
2. perform rpoB sequencing (by the SRL)
3. perform DST on solid medium (less likely to miss resistance for these mutations)

Clinically, the patient should be closely followed and checked for MDR contacts etc.
It seems safer to wait for a confirmatory result (mol or phenotypic) before switching to the standard treatment.

Mathilde Jachym Replied at 5:24 AM, 21 Jun 2015

Gene xpert find mutations on the rpoB gene. The most frequent mutation occurs on the locus 531 and lead to rifampicine high level of resistance. Therefore Gene XPERT has a good specificity, but sometimes the resistance revealed by Gene Xpert is on another location e.i. 516. This mutation is not associated with a resistant DST but the sensitivity is not completely normal. There is probably a low level of resistance with a high risk of failure or relapse among these patients. The right thing to do is to perform sequencing but if it's not possible, the treatment should use high dose of rifampicin and perhaps for longer.

Urvashi Singh Replied at 11:32 AM, 21 Jun 2015

RIF resistance detection in the *Xpert assay* is based on the
hybridization (or the absence) of five molecular beacon probes
complementary to the wild type sequence of *rpoB* gene (responsible for 95%
of drug resistance mutations in the Rifampicin Resistance Determining
Region, RRDR, codons 507 to 533).

All SNPs in the RRDR are not known to lead to phenotypic RIF resistance,
the data on such mutations are contradictory. (Rigouts et al,2013, Van Deun
et al,2009,Jamieson et al 2014,Somoskovi et al,2006,Van Deun et al 2011,Van
Deun et al 2013)

While resistance detection by *phenotypic methods* is based on MIC.
MGIT-960 cutoffs for resistance have been shown to miss low level RIF
resistance mutations.Studies have suggested revision of critical
concentration from 1 mg/ml to 0.5mg/ml or lower for MGIT-960.
(Jamieson et al, 2014,Van Deun et al, 2013, Zhang et al, 2014).

I agree with Leen Rigouts and others.We could do:

1.DST on solid medium: to confirm phenotypic resistance
2.Sequencing RRDR: to define the mutation site (whether known to be
responsible for low level or high level resistance or neither)

Musarurwa Kwaramba Replied at 3:55 AM, 22 Jun 2015

I have a different view. Given that the patient is Gene Expert Rif resistant tells us of the absence of a wild type gene and presence of a mutant gene on rpo B gene. This does not tell us the amount of the mutant cell population present. The fact that phenotypically it grew susceptible strain from the same sample but not necessary same strain may mean a mixture of both high susceptible strain with a low resistant strain. My conclusion would be that all results are correct to a patient with mixed population of resistant (minority) and susceptibility (majority). And I would classify this patient as an MDR.

Anthony Byrne Replied at 4:25 AM, 22 Jun 2015

This is a very interesting and important point that demonstrates the difference between phenotypic (what is seen from cultured m.tb sensitivity result) and genotypic (the Gene Xpert Rif mutation result).

Both are true results (if we asume there is no chance of laboratory testing error, which is as we know, always possible) but mean different things.

The first (phenotypic) means that a proportion of the cultured organism will be killed (susceptible) to Rifampicin. The second (Gene Xpert) means that there are (at least) some detectable m.tb organisms in the sample that are carry the genétic mutation for Rifampicin resistance.

This needs to be kept in mind when put in The clinical context and a treatment regimen is designed. If first line treatment (instead of MDR-TB) treatment is given then it needs to be acknowledged that a proportion of patients (probably those with more extensive or cavitatory disease) will fail. The question that then needs to be asked (of The clinician and the TB program) is, "Is this risk (of treatment failure) acceptable ?"


Dr Anthony Byrne
Thoracic Physician
University of Sydney, Australia
Socios En Salud Lima, PerĂº

Md. Shamim Hossain Replied at 2:38 PM, 26 Jun 2015

Thanks to all the scientists to have the nice discussions. We got some nice suggestions and recommendations. From the replies of the nice recommendations we could conclude some important points in case of these types of patients:
1. To perform DST on solid medium- to confirm phenotypic resistance, as solid culture is still gold standard.
2. To perform rpoB/ RRDR sequencing to define the mutation site (by the SRL)
3. To check the Ct values and Xpert intrepretation (checking graphs)
4. Based on Xpert RR result, MDR treatment regimen should be started correlating with the clinical sign and symptoms like: Previous TB history of the patients, X-ray shadow, Diabetic status, HIV/AIDS status, Nutritional status, Alcohol or drug abuse etc.

Prof.Dr.Matiur Rahman Replied at 3:14 AM, 27 Jun 2015

Excellent discussion.We must remember no test is perfect and we must keep clinical correlation always in mind.I have also come across cases like this but a lot of possibilities pointed out by our worthy colleagues have to be confirmed before reaching to a conclusion and we must remember that Gene Xpert is a relatively new test and a lot more has to come about it's authenticity in the light of experience globally.

Masoud Dara, MD Moderator Replied at 6:44 AM, 27 Jun 2015

Dear colleagues,

Thank you Dr Hussain for your post and summary conclusion of the responses. In most settings identifying the index case is a daunting or even impossible task, but in order to shift the gears in TB prevention and eventual elimination, along other tools, we also need molecular epidemiology and detailed assessments to track the transmission chain. In addition to designing more effective interventions at population level, obtaining the history of successful or unsuccessful treatment of the index case (while keeping in mind potential mutation(s) and/or possibilities of acquiring further resistance), can help clinicians decide on the controversial cases as the one presented in this discussion.

All the best,

Ranjan Perera Replied at 5:18 AM, 28 Jun 2015

There are mutations in the rpoB gene which is the target Gene for GeneXpert which are not phenotipically expressed in MGIT. We do not know how these mutations are expressed in vivo, but for safe side, always consider such cases as resistant to Rif for clinical management.

Anita qaffari.MD Replied at 8:40 AM, 28 Jun 2015

A research ,,, reported that the BACTEC 460 and the BACTEC MGIT 960 systems missed certain strains associated with low-level rifampicin resistance.

Furthermore, using Xpert MTB/RIF and gene sequencing, identified patients whose TB isolates contained mutations to the rpoB gene but whose results from the BACTEC MGIT 960 system indicated that the isolates were rifampicin susceptible.
And also found that rpoB mutations conferring low-grade resistance were often missed by rapid phenotypic DST, particularly with the BACTEC MGIT 960 system.

Anita qaffari.MD Replied at 8:46 AM, 28 Jun 2015

(WHO) in 2011 issued a policy statement recommending the Xpert MTB/RIF assay (Cepheid, Sunnyvale, CA, USA) for the diagnosis of tuberculosis (TB) and rifampicin (RIF) resistance as a proxy for multidrug-resistant TB (MDR-TB).
Xpert MTB/RIF should be used as an initial diagnostic test in individuals suspected of MDR or HIV-associated TB. It should be used as an add-on test to smear microscopy in settings where MDR-TB or HIV are of lesser concern, especially in smear-negative specimens.

Mohammed suaudi Hassen Replied at 10:02 AM, 28 Jun 2015

Dear all Hi
I think such finding need further analysis, it should be excellent for
researcher to watch another method as
an option and competent to those . and i believe that culture is golden

*king Rigard*

Dr. Saswata Dutt Replied at 10:19 AM, 28 Jun 2015

Thank you so much Mr. Ranjan for your important comment. Please may I have
the reference (book and page number, article etc.) point for that, as it
has very important programmatic fall-out for PMDT in many developing
countries. And what about solid culture in relation to rpoB results?

VIDYANIDHI GUMMA Replied at 12:46 PM, 28 Jun 2015

Dear all,

Very good discussion and information were shared by scientist. I like to add , my views
1) If such discrepancies were found between phenotypic and genotypic( Xpert) testing very frequently encountered as per case studies, the very first priority task is to reconduct epidemiological study of mutations by sequencing as per geographic location in the country.
2) Reset the diagnostic algorithms based on mutations if necessary.
We all must aware , most of the countries already implementing, MDRTB treatment based on molecular testing (LPA or Xpert). after test results were obtained as a Rif Resistant or RIF+ INH resistant, they are going to initiated the treatment. So again the most priority is, country clinical and laboratory experts sit together to decide, what could be best strategy to set the treatment algorithm based on epidemiological results,case studies and it should be reviewed periodically, as so far history with Xpert testing, it contributed well for case finding and treatment initiation.

Jean de Dieu IRAGENA Replied at 4:03 AM, 29 Jun 2015

As reference on the use of Xpert MTB/RIF assay, please find attached the WHO Policy and implementation manual on the Xpert MTB/RIF assay in tree languages: English, French and Russian:
- Policy: http://www.who.int/tb/laboratory/xpert_policyupdate/en/
- Xpert MTB/RIF implementation manual: http://www.who.int/tb/publications/xpert_implem_manual/en/

Jean de Dieu

Lal Mani ADHIKARI Replied at 2:22 AM, 15 Jul 2015

Dear all scientists and Colleagues,

Its very much beneficial discussion I felt over the Xpert MTB/RIF implementation. As experience from Nepal, I would like to share how we are moving forward thinking lesson would be helpful for further resolution.

Since the MGIT 960 is still not in practice for public use, it may show discordant result phenotypically campared with GeneXpert. firstly, major thing to be kept in mind that the ultimate national policy on it help for immediate management of patients in such a case would be considered. here in Nepal, when there is RIF resistance, the prompt treatment with MDR regimen along with reffering of Sputa for further confirmation with Solid DST. secondly, the drug resistance survey as suggested by previous scientists if we could conduct would be also helpful to figure out the epidemiological girth and to determine how much it is probable for conferring resistance. thirdly, the clinical picture of the patient, existing coinfections, xray radiograph and the chronicity of the condition, previous TB treatment history will further assist to make a picture to trigger the treatment decision.

thank you,
Lal Mani

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