Usefulness of Xpert MTB/RIF Ultra for rapid diagnosis of extrapulmonary tuberculosis in Tunisia.

Extrapulmonary tuberculosis (EPTB) remains a challenging diagnosis. The purpose of this study was to assess the accuracy of Xpert MTB/RIF Ultra (Cepheid, USA) for rapid diagnosis of EPTB in Tunisia. Eight hundred and forty-seven extrapulmonary samples collected from 2017 to 2021, were subjected to Xpert MTB/RIF Ultra. Microscopy and culture were performed for all the specimens. The accuracy of Xpert Ultra was evaluated in comparison to the culture. Xpert Ultra diagnosed EPTB with a global sensitivity of 80.66% (74.3-85.75) and specificity of 70.87% (67.31-74.20). The molecular test was most accurate when performed in cerebrospinal fluids, bones and joints and cutaneous specimens showing a sensitivity of 100% and a specificity ranging from 70.60 to 91.11%. In lymph node samples comprising aspirates and biopsies, the sensitivity of Xpert Ultra was high 87.50% (77.23-93.53), however, the specificity was 51.08% (44.67-57.46). For pleural samples, the Xpert Ultra sensitivity was 77.50% (68.34-84.68) ranging from 71.43 to 80% in pleural biopsies and fluids respectively. The specificity in all pleural specimens was 79.56% (74.40-83.91). Xpert Ultra showed promise in the diagnosis of EPTB. The performances varied according to the site of the disease. The test may be more valuable if used in combination with other diagnostic modalities.

Accuracy of the QIAxcel Automated System for MIRU-VNTR Genotyping of Mycobacterium tuberculosis in Two Limited Resource Settings.

Mycobacterial interspersed repetitive units variable number tandem repeat (MIRU-VNTR) typing of Mycobacterium tuberculosis complex (MTBC) isolates, based on 24 loci, is still widely used as the standard for routine molecular surveillance of tuberculosis (TB). QIAxcel system is proposed as an affordable tool that could replace conventional gel electrophoresis and provide high concordance with the reference methods regarding MIRU-VNTR typing of MTBC. We aimed to evaluate the QIAxcel accuracy for allele calling of MIRU-VNTR loci in two regional reference laboratories. A total of 173 DNA were used for the study. Results obtained with QIAxcel were compared to the reference results obtained with an ABI 3730 DNA analyzer. In Albania, the overall agreement with the reference method was 97.92%. A complete agreement result was obtained for 17 loci. In Tunisia, the overall agreement with the reference method was 98.95%. A complete agreement result was obtained for 17 loci. Overall agreement in both centers was 98.43%. In our opinion, use of QIAxcel technology has the potential to be reliable, given an optimized algorithm. Inaccuracies in sizing of long fragments should be solved, especially regarding locus 4052.

Whole-Genome Sequencing of Drug-Resistant Mycobacterium tuberculosis Strains, Tunisia, 2012-2016.

To investigate transmission of drug-resistant strains of Mycobacterium tuberculosis in Tunisia, we performed whole-genome sequencing on 46 multidrug-resistant strains isolated during 2012-2016. Core-genome multilocus sequence typing grouped 30 strains (65.2%) into 3 clusters, indicating extensive recent transmission and Haarlem clone predominance. Whole-genome sequencing might help public health services undertake appropriate control actions.

Diagnosis of lymph node tuberculosis using the GeneXpert MTB/RIF in Tunisia.

INTRODUCTION: GeneXpert MTB/RIF is a fully-automated diagnostic molecular test which simultaneously detects tuberculosis (TB) and rifampicin (RIF) drug resistance. The purpose of this study is to evaluate the performance of the GeneXpert MTB/RIF test for the detection of Mycobacterium tuberculosis complex (MTBC) in lymph node specimens and to show the place of Mycobacterium bovis as a major cause of TB lymphadenitis. MATERIAL AND METHODS: This study was conducted simultaneously in the National Reference Laboratory for Mycobacteria of Ariana and the Central Laboratory of Sfax, from January to December 2013. In total, 174 lymph node specimens were processed simultaneously for Ziehl-Neelsen, auramine and immuno-histochemical staining. Conventional culture on both Lowenstein-Jensen and liquid medium (Bactec MGIT 960 BD system) and the new molecular-based GeneXpert MTB/RIF assay system were performed. Positive cultures were confirmed using molecular identification (Genotype MTBC Hain Lifescience). RESULTS: Among the 174 samples tested, the GeneXpert detected the DNA of MTBC in 134 samples (77%). Standard bacteriological assays, including AFB microscopy and culture, were positive, respectively, in 41 (23.6%) and 79 (45.4%) specimens. M. bovis was isolated in 76% of positive cultures. GeneXpert sensitivity and specificity results were assessed according to smear and culture results, clinical and histological findings. The sensitivity and specificity of the Xpert assay were 87.5% (126/144) and 73.3%, respectively. CONCLUSION: The implementation of the GeneXpert MTB/RIF assay may dramatically improve the rapid diagnosis of lymph node TB.

Detection of isoniazid and rifampin resistance of Mycobacteriumtuberculosis by a multiplex allele-specific polymerase chain reaction (PCR) assay.

BACKGROUND: The use of molecular techniques is a major improvement for the rapid routine detection and control of multidrug-resistant tuberculosis (MDR-TB). MATERIALS: In this study, the multiplex allele-specific polymerase chain reaction (MAS-PCR) was developed to simultaneously detect the most frequent mutations associated with isoniazid (INH) and rifampin (RIF) resistance in a single assay. RESULTS: The assay was tested with 53 clinical isolates. Among them, 27 were MDR strains, 17 were mono-resistant to INH, one was mono-resistant to RIF, and eight were susceptible. The MAS-PCR assay showed a specificity of 100% in detecting drug resistance. An equivalent sensitivity of 92.6% in detecting MDR and RIF-resistance was found. The sensitivity for the detection of INH-resistance was 88.6%. CONCLUSIONS: The MAS-PCR assay was a simple and rapid method for detecting the INH and RIF-resistance in Mycobacterium tuberculosis (MT) clinical strains. It is also easy to perform and to interpret. The assay is inexpensive and a less-demanding technique.