To Explore The Evaluation Of Xpert MTB/RIF For The Detection Of Tuberculosis In CSF And Rifampicin Resiatnce

Objective: Tuberculous meningitis (TBM) is a central nervous systeminfection disease caused by Mycobacterium tuberculosis(MTB) with highfatality rate and disability rate. The key factors to improve prognosis areaccurate diagnosis and reasonable treatment. An automated nucleic acidamplification test, the MTB/RIF has recently been developed, which is a fullyautomated real time hemi-nested PCR system that can detect the amplifiedM. tuberculosis DNA sequences and its mutations responsible forRifampin-resistance within two hours. The aim of this study was to determinethe sensitivity and specificity of the MTB/RIF assay for the diagnosis oftuberculosis. To explore the sensitivity and specificity of the system,60patients with suspected tuberculous meningitis and30patients with other CNSdiseases are involved in the test. We also analyze the resistant condition ofrifampin.Methods: The case group were60cases（60shares of CSF）clinicaldiagnosis of tuberculous meningitis patients, and the control group were30cases（30shares of CSF）non-tuberculous meningitis patients. All of thepatients were examined CSF routine, biochemical, cytology and all of the casegroup were examined modified Ziehl-Neelsen(Z-N) stain. And we also scorethem according to the latest TBM clinical diagnostic criteria, screening out of60cases: the definite TBM38cases, probable TBM11cases, possible TBM11cases. All patients are detected special target DNA of MTB complex byusing Xpert MTB/RIF system. Collecting1mL CSF and then reserving in-80℃refrigerator. We melted the CSF in room temperature and mixed the CSFwith1:2ratio of processing solution. To make sure the CSF and the processingsolution are fully mixed, we add a vortex step to sample processing beforeputting the mixture into the system. Results are reported as positive(at least two of the five probes give positive signals) or negative for M.tuberculosis.Positive results are graded in four categories: very low(CT>28),low(22<CT<28), medium(16<CT<22) or high(CT<16). Rifampicin resistanceis reported as susceptible or resistant(not all the five probes gives positivesignal). By comparing PCR positive rates between the two groups, we analyzethe sensitivity and specificity of the Xpert MTB/RIF. By comparing positiverates between PCR and modified Ziehl-Neelsen(Z-N) stain, we analyze thedifferences between the two methods. By analyze the detection rate ofanti-RIF and the mutation, we analyze the condition of anti-RIF and theresistance mechanism.Results:1The detection result of MTB PCR from CSF: Positive results by thePCR were found in11adults of60patients with tuberculous meningitis, and0positive in30adults from the control group. The sensitivity and specificity ofthe Xpert MRB/RIF for MTB respectively are18.83%and100%. Bystatistical analysis P <0.05, the difference between the two groups wasstatistically significant.2The detection result of MTB PCR in each group in case group: Positiveresults by the PCR were found in9adults of38patients in definite group,2positive in11adults in probably group, and0positive in11adults in possiblegroup. By statistical analysis P＞0.05, there was no statistically significantdifference among the three groups.3Qualitative estimation of bacterial load: The majority of Xpert resultswere categorized by Xpert as ‘very low’(n=9/11)or ‘low’(n=2/11).No CSFsamples reported ‘medium’ and ‘high’ bacterial load by Xpert.4Detection of Rifampicin resistance: Rifampicin resistance was detectedin four cases during the study by Xpert MTB/RIF. Xpert testing for rifampicinresistance showed an‘indeterminate result’in two cases. The drug resistancerate in the study is36.36%. The failing probes that leading to resistance are Eprobe(n=3/4) and D probe(n=1/4).The failing probe that leading toindeterminate result is E probe(n=2/2). 5The comparison between the Xpert MTB/RIF of MTB and modifiedN-Z stain in the case group: There were38of60shares CSF from the casegroup stained by modified N-Z stain. We found that9shares CSF in38sharesStain positive were positive by PCR testing. And meantime, We found that2shares CSF in22shares Stain negative were positive by PCR testing. Bystatistical analysis P＜0.05, the difference between the two methods wasstatistically significant.Conclusion: Xpert MTB/RIF can provide a specific method for the earlydiagnosis of TBM. Because the load of bacteria in CSF is very low(thebacterial load are very low and low with Xpert), the detection of MTB withXpert is hard with a sensibility of18.83%. The sensibility of PCR is lowerthan N-Z stain. We can use the two methods together in detection of MTB toimprove the sensitivity and specificity. Due to the gene mutation, MTB cangenerate the ability of resistance to anti-tuberculosis drugs, which can lead tothe poor therapeutic effect. Rifampicin resistance was detected in four casesduring the study by Xpert MTB/RIF. The failing probes that leading toresistance are E probe and D probe, who cover the areas that531site and526site are located.