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1.
Several new methods to detect drug resistance in Mycobacterium tuberculosis have been proposed in recent years. Colourimetric methods that use redox indicators or the nitrate reduction assay have received increasing attention because of their simplicity and the absence of any requirement for sophisticated equipment or highly trained personnel. Several studies have evaluated their accuracy and performance in comparison with reference standard methods, particularly for the detection of resistance to rifampicin and isoniazid, which are the two most important drugs used for the treatment of tuberculosis. This review describes the development, evaluation and implementation of these methods as rapid alternative tests for the detection of multidrug resistance in M. tuberculosis. Based on published evidence and the high accuracy of colourimetric methods for detecting drug resistance in M. tuberculosis, these methods seem to be appropriate for implementation in high-burden low-resource countries. 相似文献
2.
C. Arnold L. Westland G. Mowat A. Underwood J. Magee S. Gharbia 《Clinical microbiology and infection》2005,11(2):122-130
The rapid technique of pyrosequencing was used to examine 123 samples (in the form of DNA extracts and inactivated sputum) of Mycobacterium spp. Of 99 Mycobacterium tuberculosis samples investigated for single-nucleotide polymorphisms (SNPs), 68% of isoniazid-resistant isolates analysed had an AGC --> ACC mutation in katG at codon 315, resulting in the Ser --> Thr substitution associated previously with isoniazid resistance. Of the rifampicin-resistant isolates, 92% showed SNPs in rpoB at codons 516, 531 or 526. Inactivated sputum samples and DNA extracts could both be analysed by pyrosequencing, and the method was able to differentiate rapidly between the closely related species of the M. tuberculosis complex (M. tuberculosis, Mycobacterium bovis, Mycobacterium africanum, Mycobacterium canetti and Mycobacterium microti), except between M. tuberculosis, M. canetti and one of two M. africanum strains. This low-cost, high-throughput technique could be used as a rapid screen for drug resistance and as a replacement for some of the time-consuming tests used currently for species identification. 相似文献
3.
Rapid detection of resistance in Mycobacterium tuberculosis: a review discussing molecular approaches 总被引:4,自引:3,他引:4
D. García de Viedma 《Clinical microbiology and infection》2003,9(5):349-359
The last few years have seen the development of several molecular designs to search for mutations encoding resistance to antituberculous drugs in Mycobacterium tuberculosis . Most of these are highly efficient for RIF-r detection and are well adapted to search for the most relevant INH-R mutations. In this review, these new molecular approaches are explained and are presented according to the molecular strategies on which they are based. In this sense, techniques based on DNA-sequencing, electrophoresis and hybridization are reviewed and the newer designs based on real-time PCR and microarrays are also included. Molecular methods are sure to transform standard approaches to the issue of resistance in the mycobacteriology laboratory. This will allow laboratories to speed up the performance of resistance assays and provide access to essential information for highly refined detection, follow-up and management of antibiotic resistance in M. tuberculosis . 相似文献
4.
L.D. Ghiraldi P.A.Z. Campanerut F.L.E. Spositto D.N. Sato C.Q.F. Leite M. Hirata R.D.C. Hirata R.F. Cardoso 《Clinical microbiology and infection》2011,17(12):1792-1797
The microscopic observation drug susceptibility assay (MODS) was evaluated to determine susceptibility to pyrazinamide in Mycobacterium tuberculosis, and compared with the broth microdilution method (BMM), absolute concentration method (ACM), and pyrazinamidase (PZase) determination. We tested 34 M. tuberculosis clinical isolates (24 sensitive and eight resistant to pyrazinamide) and the control strains M. tuberculosis H37Rv (ATCC 27294) and Mycobacterium bovis AN5. The MODS, BMM, ACM and PZase determination provided results in average times of 6, 18, 28 and 7 days, respectively. All methods showed excellent sensitivity and specificity (p <0.05). Of the methods studied, the MODS proved to be faster, efficient, inexpensive, and easy to perform. However, additional studies evaluating the MODS in differentiating pyrazinamide-resistant and pyrazinamide-susceptible M. tuberculosis must be conducted with a larger number of clinical isolates. 相似文献
5.
《Clinical microbiology and infection》2020,26(10):1332-1337
ObjectivesEffective use of antibiotics is critical to control the global tuberculosis pandemic. High-dose isoniazid (INH) can be effective in the presence of low-level resistance. We performed a systematic literature review to improve our understanding of the differential impact of genomic Mycobacterium tuberculosis (Mtb) variants on the level of INH resistance. The following online databases were searched: PubMed, Web of Science and Embase. Articles reporting on clinical Mtb isolates with linked genotypic and phenotypic data and reporting INH resistance levels were eligible for inclusion.MethodsAll genomic regions reported in the eligible studies were included in the analysis, including: katG, inhA, ahpC, oxyR-ahpC, furA, fabG1, kasA, rv1592c, iniA, iniB, iniC, rv0340, rv2242 and nat. The level of INH resistance was determined by MIC: low-level resistance was defined as 0.1–0.4 μg/mL on liquid and 0.2–1.0 μg/mL on solid media, high-level resistance as >0.4μg/mL on liquid and >1.0 μg/mL on solid media.ResultsA total of 1212 records were retrieved of which 46 were included. These 46 studies reported 1697 isolates of which 21% (n = 362) were INH susceptible, 17% (n = 287) had low-level, and 62% (n = 1048) high-level INH resistance. Overall, 24% (n = 402) of isolates were reported as wild type and 76% (n = 1295) had ≥1 relevant genetic variant. Among 1295 isolates with ≥1 variant, 78% (n = 1011) had a mutation in the katG gene. Of the 867 isolates with a katG mutation in codon 315, 93% (n = 810) had high-level INH resistance. In contrast, only 50% (n = 72) of the 144 isolates with a katG variant not in the 315-position had high-level resistance. Of the 284 isolates with ≥1 relevant genetic variant and wild type katG gene, 40% (n = 114) had high-level INH resistance.ConclusionsPresence of a variant in the katG gene is a good marker of high-level INH resistance only if located in codon 315. 相似文献
6.
《Indian journal of medical microbiology》2014,32(3):318-322
Detection of drug resistance plays a critical role in tuberculosis treatment. The aim of this study was to evaluate the performance of GenoType Mycobacteria Drug Resistance (MTBDR) assay (Hain Lifescience, Germany) and to compare it with radiometric BACTEC 460 TB system (Becton Dickinson, USA) for the detection of rifampicin (RIF) and isoniazid (INH) resistance in 84 Mycobacterium tuberculosis complex (MTBC) isolates. RIF resistance was identified in 6 of 7 (85.7%) isolates and INH resistance was identified in 8 of 14 (57.1%) isolates by the GenoType MTBDR assay. Compared with BACTEC system, the sensitivity, specificity, positive predictive value and negative predictive values were 85.7%, 98.7%, 85.7% and 98.7% for RIF resistance; and 57.1%, 100%, 100% and 92.1% for INH resistance, respectively. GenoType MTBDR assay is reliable when tested specimen is resistant to the tested drugs. Although test was more successful in the detection of RIF resistance, it exhibited low sensitivity for the detection of INH resistance. 相似文献
7.
V. V. Nikolayevskyy T. J. Brown Y. I. Bazhora A. A. Asmolov Y. M. Balabanova F. A. Drobniewski 《Clinical microbiology and infection》2007,13(2):129-138
Understanding the molecular epidemiology of tuberculosis (TB) and mutations in genes associated with drug resistance may contribute to the development of appropriate interventions to improve tuberculosis control. A structured questionnaire was used to collect basic epidemiological data from 589 patients with radiologically confirmed TB in the Odessa and Nikolaev regions of the Ukraine in 2003-2004. A non-commercial reverse hybridisation assay and DNA sequencing were used to detect mutations associated with rifampicin and isoniazid resistance. Genotyping was performed using multilocus variable number tandem repeat (VNTR) typing and spoligotyping. Mutations conferring rifampicin and isoniazid resistance were detected in 32.9% and 44.0%, respectively, of 225 Mycobacterium tuberculosis isolates from individual consecutive patients. Mutations in codon 531 and codon 315 of the rpoB and katG genes, respectively, were predominant among drug-resistant isolates. Multidrug (MDR) resistance rates were significantly higher among former prison inmates compared with non-prisoners (54.8% vs. 27.3%; RR 2.01; 95% CI 1.35-2.97) and the prevalence of mutations was higher in Beijing strains sharing the VNTR signature 223325173533424 than in other Beijing strains (71.4% vs. 45.7%; RR 1.74; 95% CI 1.17-2.57), suggesting that this group may be responsible for rapid transmission of MDR TB in the southern Ukraine. 相似文献
8.
《Indian journal of medical microbiology》2010,28(3):211-216
Objective: The objective of our study was to evaluate the use of a real-time polymerase chain reaction (PCR)-based technique for the prediction of phenotypic resistance of Mycobacterium tuberculosis. Materials and Methods: We tested 67 M tuberculosis strains (26 drug resistant and 41 drug susceptible) using a method recommended for the LightCycler platform. The susceptibility testing was performed by the absolute concentration method. For rifampin resistance, two regions of the rpoB gene were targeted, while for identification of isoniazid resistance, we searched for mutations in katG and inhA genes. Results: The sensitivity and specificity of this method for rapid detection of mutations for isoniazid resistance were 96% (95% CI: 88% to 100%) and 95% (95% CI: 89% to 100%), respectively. For detection of rifampin resistance, the sensitivity and specificity were 92% (95% CI: 81% to 100%) and 74% (95% CI: 61% to 87%), respectively. The main isoniazid resistance mechanism identified in our isolates is related to changes in the katG gene that encodes catalase. We found that for rifampin resistance the concordance between the predicted and observed phenotype was less than satisfactory. Conclusions: Using this method, the best accuracy for genotyping compared with phenotypic resistance testing was obtained for detecting isoniazid resistance mutations. Although real-time PCR assay may be a valuable diagnostic tool, it is not yet completely satisfactory for detection of drug resistance mutations in M tuberculosis. 相似文献
9.
Raut U Narang P Mendiratta DK Narang R Deotale V 《Indian journal of medical microbiology》2008,26(3):222-227
PURPOSE: To evaluate MTT method for detection of drug resistance to rifampicin and isoniazid in M.tuberculosis . This method utilises the ability of viable mycobacterial cells to reduce MTT( 3-4,5-dimethylthiazol-2-yl-2, 5-diphenyl tetrazolium bromide). METHODS: The method was standardised with known resistant and sensitive strains of M.tuberculosis and was then extended to 50 clinical isolates. An inoculum of 10 7 cfu/mL was prepared in Middlebrook 7H9 medium supplemented with oleic acid, albumin, dextrose and catalase. For each drug three tubes were used, one with INH(0.2microg/mL) or RIF(1microg/mL), another as inoculum control and third as blank control. These were incubated at 37 degrees C for four and seven days respectively for RIF and INH after which MTT assay was performed. Results were read visually and by colorimeter at 570 nm. Relative optical density unit (RODU) of 0.2 was taken as cut off. Results were compared with drug sensitivity obtained by proportion method using LJ medium. RESULTS: For rifampicin, concordance with proportion method was 90% by visual and 94% by RODU. Sensitivity and specificity was 86.8% and 100% respectively by visual method and 95.2% and 87.5% respectively by RODU. For Isoniazid, concordance was 94% and sensitivity and specificity was 94.7 and 91.7% respectively by both visual and RODU. CONCLUSIONS: MTT assay proved to be rapid and cheap method for performing drug sensitivity of M.tuberculosis. 相似文献
10.
《Indian journal of medical microbiology》2010,28(2):162-163
A low-cost method of detecting multi-drug resistant Mycobacterium tuberculosis (MDR-TB) with the possibility of quick adoption in a resource limited setting is urgently required. We conducted a study combining isoniazid and rifampicin in a single LJ medium, to detect MDR-TB strains. Combined and individual drug media showed 100% concordance for the detection of MDR-TB and susceptible strains by proportion method. Considering the results, combined isoniazid and rifampicin containing medium could be considered for use in settings where the sole detection of MDR-TB strains is justified. 相似文献
11.
结核分枝杆菌临床分离株异烟肼耐药相关基因突变的分子特征 总被引:1,自引:0,他引:1
目的 阐明结核分枝杆菌异烟肼(INH)耐药相关基因突变特征.方法 对137株结核分枝杆菌临床分离株(耐异烟肼菌株87株,异烟肼敏感菌株90株)的9个结构基因furA、katG、inhA、kasA、Rv0340、iniB、iniA、iniC和efpA以及两个调控区oxyR-ahpC基因间隔区和mabA-inhA启动子进行DNA片段扩增及序列分析.结果 82株(94.3%)INH耐药分离株的katG基因存在突变,其中katGSer315Thr突变占优势(55.2%).50株INH敏感的分离菌katG的463密码子没有突变.35株(40.2%)INH耐药的分离株katG的463有突变.87株INH耐药株中,20株(23.0%)的katG基因存在两重突变.13株(14.9%)分离菌inhA基因的启动子区存在突变,4.6%的分离菌有inhA结构基因突变,11.5%oxyR-ahpC基因间区存在突变.iniBAC区域和efpA中发现耐药性关联突变.结论 研究证实多个基因突变与异烟肼耐药之间的关系,并且为阐明结核分枝杆菌耐药机制提供线索. 相似文献
12.
Y.-J. Sun A. S. G. Lee S.-Y. Wong N. I. Paton 《Clinical microbiology and infection》2007,13(8):833-835
Previous studies have suggested that Mycobacterium tuberculosis kasA G312S and G269S gene mutations may represent sequence polymorphisms of the M. tuberculosis East-African-Indian (EAI) and T families, respectively, rather than relating to isoniazid resistance. The present study examined polymorphisms of these two codons in 98 drug-susceptible M. tuberculosis isolates (68 EAI and 30 T isolates). Twenty-eight isolates belonging to a sub-lineage of the EAI family had the kasA G312S mutation, but none of the 30 T isolates had the G269S mutation. The data suggest that the kasA G312S mutation is not related to isoniazid resistance, but represents a sequence polymorphism in a sub-lineage of the EAI family. 相似文献
13.
G. T. Noordhoek S. Mulder P. Wallace A. M. van Loon 《Clinical microbiology and infection》2004,10(4):295-301
The aim of this study was to evaluate the laboratory performance of nucleic acid amplification tests (NATs) for detection of the Mycobacterium tuberculosis complex. A proficiency panel consisting of eight sputum specimens and four specimens diluted in phosphate-buffered saline (PBS) was sent to 82 laboratories in 23 countries by the Quality Control for Molecular Diagnostics (QCMD) TB programme. The performance of different NATs was analysed in combination with a questionnaire on the applied methods. Seventy-eight participants (95.2%) contributed a total of 85 evaluable data sets. The percentage of correct results on the eight sputum samples was 86.3% (586/679). Of the sputum specimens considered as 'smear-negatives' (650 CFU/250 micro L), only 61.2% (104/170) were reported positive. The percentage of correct results for the three scored PBS samples was 75.7% (193/255). The total number of false-positive results was 11 (4.3%); these were reported for seven (8.2%) of the 85 data sets. In 32 (37.6%) data sets an 'in-house' NAT method was used, and in 53 (62.4%) sets a commercial assay was tested. The percentage of data sets achieving correct results on all sputum samples was 35.3% and 37.8%, respectively. For the PBS samples this was 45.8% and 41.5%. Overall, the results of this study demonstrated that the performance of NATs for the detection of M. tuberculosis has improved since previous studies. The percentage of false-positives has decreased considerably. However, a large number of procedures still lack sufficient sensitivity for application to smear-negative samples. 相似文献
14.
Objective: To evaluate the distribution of acquired resistance in isolates of Mycobacterium tuberculosis from treated patients in two periods, 1984–89 and 1990–95, in the Bursa (Southern Marmara) region.
Method: Susceptibility of 531 M. tuberculosis isolates to four commonly used drugs (isoniazid (INH), streptomycin (SM), ethambutol (EMB) and rifampin (RMP)) was determined by the absolute concentration method of Canetti et al.
Results: In 203 strains isolated in the years 1984–89, the total acquired resistance was 32.5%, and it was 37.5% in 328 strains isolated in 1990–95 ( p >0.05). Resistance to INH, SM, RMP and EMB was found in 23.6%, 16.7%, 6.4% and 3.9%, respectively, in the first period (1984–89), and in 26.2%, 20.4%, 25.3% and 8.2%, respectively, in the second period (1990–95). The increase in RMP resistance was statistically significant ( p <0.001). The incidence of multidrug-resistant strains was 12.3% in the first period, and 24.4% in the second period, a significant increase ( p <0.001).
Conclusions: We believe that progressive emergence of phenotypes resistant to INH+RMP in our region is caused by inadequate treatment for various reasons. In the present study, the fact that multidrug resistance occurred in nearly 25% of patients treated previously but still infective suggests that the approach to surveillance, patient therapy and follow-up programs should be fundamentally reconsidered in our region. 相似文献
Method: Susceptibility of 531 M. tuberculosis isolates to four commonly used drugs (isoniazid (INH), streptomycin (SM), ethambutol (EMB) and rifampin (RMP)) was determined by the absolute concentration method of Canetti et al.
Results: In 203 strains isolated in the years 1984–89, the total acquired resistance was 32.5%, and it was 37.5% in 328 strains isolated in 1990–95 ( p >0.05). Resistance to INH, SM, RMP and EMB was found in 23.6%, 16.7%, 6.4% and 3.9%, respectively, in the first period (1984–89), and in 26.2%, 20.4%, 25.3% and 8.2%, respectively, in the second period (1990–95). The increase in RMP resistance was statistically significant ( p <0.001). The incidence of multidrug-resistant strains was 12.3% in the first period, and 24.4% in the second period, a significant increase ( p <0.001).
Conclusions: We believe that progressive emergence of phenotypes resistant to INH+RMP in our region is caused by inadequate treatment for various reasons. In the present study, the fact that multidrug resistance occurred in nearly 25% of patients treated previously but still infective suggests that the approach to surveillance, patient therapy and follow-up programs should be fundamentally reconsidered in our region. 相似文献
15.
Use of a high-density DNA probe array for detecting mutations involved in rifampicin resistance in Mycobacterium tuberculosis 总被引:3,自引:0,他引:3
W. Sougakoff M. Rodrigue C. Truffot-Pernot M. Renard N. Durin M. Szpytma R. Vachon A. Troesch V. Jarlier 《Clinical microbiology and infection》2004,10(4):289-294
A Mycobacterium high-density DNA probe array designed to detect rpoB mutations conferring rifampicin resistance in Mycobacterium tuberculosis was evaluated. The rpoB hybridisation patterns produced by 41 susceptible (RifS) and 59 rifampicin-resistant (RifR) clinical isolates of M. tuberculosis were compared with the results of conventional dideoxynucleotide sequencing of the rpoB gene. For all the RifR isolates, the rpoB hybridisation patterns correlated with the rpoB sequencing results. Among the 59 isolates, 11 distinct amino-acid changes were detected by the DNA probe array. Of these, 36 (61%) corresponded to replacement of the serine residue found in position 531 (S531L in 34 isolates and S531W in two isolates), 16 (27%) affected histidine 526 (five H526D, five H526Y, four H526L, one H526N and one H526R), four (6.8%) replaced aspartate 516 with a valine, and one (1.7%) replaced glutamine 513 with a leucine. Deletion of the asparagine residue at position 519 was detected in one isolate susceptible to rifampicin, but yielding c. 0.1% resistant colonies on rifampicin-containing medium. No mutation was detected in the rpoB region from one isolate yielding c. 5% of resistant colonies on rifampicin-containing medium. Finally, a D516Y substitution was detected in association with an unexpected mutation, G523W, not tiled on the DNA probe array, but which could be detected by analysing the hybridisation pattern obtained with the wild-type probes covering codon 523. In conclusion, the Mycobacterium probe array is a promising approach to rapid detection of mutations involved in rifampicin resistance in M. tuberculosis. 相似文献
16.
K. A. K. Ängeby J. Werngren J. C. Toro G. Hedström B. Petrini S. E. Hoffner 《Clinical microbiology and infection》2003,9(11):1148-1152
We evaluated the BacT/ALERT 3D system for recovery and drug susceptibility testing (DST) of Mycobacterium tuberculosis (MTB). Of 2659 clinical specimens, MTB was detected in 92 using BacT/ALERT, compared to 94 using Löwenstein–Jensen culture. Detection time was 25% shorter with BacT/ALERT. Sensitivities were 92%, 96%, 78% and 100% for resistance to rifampicin, isoniazid, streptomycin and ethambutol, respectively, while specificity was 100% for all antibiotics, when BacT/ALERT was compared with the BACTEC 460 method on 50 MTB isolates. The BacT/ALERT system is fully automated and creates no radioactive waste. It seems to be a valid alternative for primary isolation, but further evaluation is needed regarding DST. 相似文献
17.
Two techniques have been approved by the United States FDA for diagnosis of tuberculosis in smear positive sputa: LCX M. tuberculosis, a ligase chain reaction procedure manufactured by Abbott Laboratories, and Amplicor, a polymerase chain reaction (PCR) procedure manufactured by Roche. However, these commercial methods are expensive and beyond the reach of laboratories in most developing countries. We compared the Roche Amplicor kit with an in-house PCR using a primer set for Mycobacterium tuberculosis/bovis directed at MPB 64 protein gene. It was able to distinguish between M. tuberculosis, M. avium, M. gordonii, M. intracellularae, and M. kansasii. Fifty-seven cytological samples were submitted to the laboratory for molecular diagnosis of M. tuberculosis. Both procedures were run on every sample submitted and the two methods agreed completely. The custom-made method is less expensive than the commercial technique. 相似文献
18.
《Clinical microbiology and infection》2021,27(9):1351.e1-1351.e4
ObjectivesTo evaluate the accuracy of two new molecular diagnostic tests for the detection of drug-resistant tuberculosis, the FluoroType MTB and MTBDR VER 2.0 assays, in combination with manual and automated DNA extraction methods.MethodsSputa from 360 Xpert Ultra Mycobacterium tuberculosis complex (MTBC)-positive patients and 250 Xpert Ultra MTBC-negative patients were tested. GenoType MTBDRplus served as reference for MTBC and drug resistance detection. Sanger sequencing was used to resolve discrepancies.ResultsFluoroType MTB VER 2.0 showed similar MTBC sensitivity compared with FluoroType MTBDR VER 2.0 (manual DNA extraction: 91.6% (294/321) versus 89.8% (291/324); p 0.4); automated DNA extraction: 92.1% (305/331) versus 87.7% (291/332); p 0.05)). FluoroType MTBDR VER2.0 showed comparable diagnostic accuracy to FluoroType MTBDR VER1.0 as previously reported for the detection of MTBC and rifampicin and isoniazid resistance.ConclusionsThe FluoroType MTB and MTBDR VER 2.0 assays together with an automated DNA extraction and PCR set-up platform may improve laboratory operational efficiency for the diagnosis of MTBC and resistance to rifampicin and isoniazid and show promise for the implementation in a centralized molecular drug susceptibility testing model. 相似文献
19.
显色法芯片检测结核分枝杆菌利福平和异烟肼耐药基因 总被引:6,自引:0,他引:6
目的建立显色法芯片检测结核分枝杆菌耐药基因的方法。方法设计4对地高辛标记引物,扩增结核分枝杆菌rpoB、katG、inhA和ahpC 4个基因部分片段,根据结核分枝杆菌利福平(RFP)和异烟肼(INH)4条耐药相关基因上8个位点的25种单核苷酸多态性设计探针制作芯片,扩增产物与芯片杂交,显色法判断结果;用该法检测46株结核分枝杆菌临床分离株。结果扩增产物琼脂糖电泳可见4条大小分别为165、181、245、315 bp DNA条带;结核分枝杆菌菌液浓度为1.6×103/ml时,该法仍可检测到各位点野生及突变信号;19种非结核分枝杆菌标准株和9种非分枝杆菌标准株扩增产物无DNA条带,与芯片杂交亦无信号,H37Rv结核分枝杆菌标准株芯片检测各位点均为野生型;5株结核分枝杆菌临床分离株重复检测5次,结果完全一致;6份PCR产物的测序结果与芯片检测结果完全一致;46株结核分枝杆菌临床分离株中,RFP耐药株28株,芯片检出突变株24株,突变率为85.7%,RFP敏感株18株,芯片检出突变株2株。INH耐药株31株,芯片检出突变株20株,突变率为64.5%,INH敏感株15株,芯片检出突变株4株。结论显色法芯片检测结核分枝杆菌耐药基因具有较高的敏感性和特异性,无需特殊仪器设备,有一定的推广应用价值。 相似文献
20.
目的 建立快速检测结核分枝杆菌异烟肼(INH)和利福平(RFP)耐药相关基因katG、inhA和rpoB突变的多重聚合酶链反应-单链构象多态性(multi PCR-single strand conformational polymorphism analysis,mPCR-SSCP)方法.方法 药敏试验检测134株结核分枝杆菌临床菌株对INH和RFP的耐药性.设计结核分枝杆菌INH和RFP耐药相关katG、inhA和rpoB基因PCR引物,建立mPCR-SSCP技术检测上述菌株katG、inhA和rpoB基因的突变,同时采用PCR直接测序技术(PCR-DS)检测上述基因片段突变情况,并对上述3种方法检测结果进行分析和比较.结果 134株临床菌株均含有katG、inhA和rpoB基因,其中42株(31.3%)对INH耐药、45株(33.6%)对RFP耐药.mPCR-SSCP和PCR-DS检测结果显示,92株INH敏感菌株katG和inhA基因均未发生突变,检测特异性均为100%;89株RFP敏感菌株中rpoB基因分别有2株和1株检测出突变,检测特异性分别为97.8%和98.9%;42株INH耐药菌株中分别有33株和36株katG和/或inhA基因突变,检测灵敏度分别为78.6%和85.7%;45株RFP耐药菌株中rpoB基因分别有41株和43株发生突变,检测灵敏度分别为91.1%和95.6%.结论 本研究建立的mPCR-SSCP能快速、简便、特异,并有一定的敏感性检测结核分枝杆菌异烟肼和利福平耐药相关基因katG、inhA和rpoB突变,具有临床应用前景. 相似文献