Mycobacterium Tuberculosis Complex Research Articles

Advanced integrative molecular platform for high-throughput screening of drug-resistant tuberculosis

A real time-polymerase chain reaction-based test in lyophilized form, was developed to simultaneously identify Mycobacterium tuberculosis complex (MTC) by targeting IS6110, rrs as dual markers, as well as mutations causing rifampicin and isoniazid resistance. The test was evaluated for pulmonary and non-pulmonary specimens from sample isolation to PCR analysis. The test demonstrated limit of detection of 25 CFU/mL for MTB, 200 CFU/mL for rpoB and inhA/katG targets with >95 % CI. The specificity for MTC was supported by a comprehensive clinical validation (n = 100). This load-and-go molecular platform, with features of high throughput, long shelf-life, room temperature storage provides simultaneous detection of MTC and its drug-resistant mutations in minimal time. The test named “PathoDetect TM MTB-RIF and INH resistance detection kit” has been approved by Central Drugs Standard Control Organisation, Indian Council of Medical Research and would have implications for tuberculosis elimination programs.

Tuberculosis in wild animals in India.

India is renowned for its complex megadiverse ecosystems and abundant biodiversity. Bovine tuberculosis (bTB) often remains synonymous with Mycobacterium bovis infection in cattle. The domain of tuberculosis (TB) among wild animals, induced by members of the Mycobacterium tuberculosis complex organisms (MTBC), is often underexplored and underreported in India. Within this context, instances of wild animal tuberculosis (wTB) have manifested across both captive and free-roaming animals. The sources contributing to wTB in animals can be human, animal, or environmental factors, thus illuminating the complex transmission pathways. The diagnosis of wTB continues to pose a formidable challenge, a consequence of the expansive taxonomic diversity in both the host and the pathogen. Complications inherent in acquiring samples from wildlife, the absence of standardized diagnostic protocols, limited insights into infection prevalence, and resource constraints compound diagnosis. Amidst these, adopting the comprehensive One Health paradigm surfaces as an imperative, accentuating the interconnectedness bridging human, animal, and environmental health. Recognizing key stakeholders and fostering intersectoral collaboration to provide enhanced diagnostic techniques driven by skilled personnel and advanced infrastructure play pivotal roles in a comprehensive strategy. Additionally, leveraging vaccination efforts contributes to effective control. A national wTB surveillance program is a cornerstone, ensuring an integrated and holistic approach to disease management. Through this review, we delve into the current landscape of wTB in India, unveiling its multifaceted challenges, and further explore the multifarious strategies that the One Health approach proffers in this dynamic endeavor.

Clinical applications of metagenomic next-generation sequencing in the identification of pathogens in periprosthetic joint infections: a retrospective study

BackgroundThis study aimed to evaluate the application of metagenomic next-generation sequencing (mNGS) technology to identify pathogens in periprosthetic joint infection (PJI).MethodsA retrospective analysis was conducted on 65 patients suspected of having PJI between April 2020 and July 2023. The patients were categorized into PJI (46 patients) and non-PJI (19 patients) groups based on the 2018 International Consensus Meeting criteria. Clinical data were collected, and both conventional bacterial culture and mNGS were performed. The diagnostic performance of the two methods was compared and analyzed.ResultsmNGS exhibited a sensitivity of 89.13%, a specificity of 94.74%, a positive predictive value of 97.62%, a negative predictive value of 78.26%, and an overall diagnostic accuracy of 90.77%. Compared to microbial culture, mNGS demonstrated superior diagnostic sensitivity while maintaining similar specificity. A total of 48 pathogens were successfully identified using mNGS, with Coagulase-negative staphylococci, Streptococci, Staphylococcus aureus, and Cutibacterium acnes being the most common infectious agents. Notably, mNGS was used to identify 17 potential pathogens in 14 culture-negative PJI samples, highlighting its ability to detect rare infectious agents, including Cutibacterium acnes (n = 5), Granulicatella adiacens (n = 1), Mycobacterium tuberculosis complex (n = 1), and Coxiella burnetii (n = 1), among others, which are not detectable by routine culture methods. However, mNGS failed to detect the pathogen in 4 culture-positive PJI patients, indicating its limitations. Among the 46 PJI patients, 27 had positive culture and mNGS results. The results of mNGS were concordant with those of culture at the genus level in 6 patients with PJI and at the species level in 18 patients. Furthermore, the present study revealed a significantly greater proportion of Staphylococcus aureus in the sinus tract group (45.45%) than in the non-sinus tract group (14.29%), indicating the association of this pathogen with sinus formation in PJI (P = 0.03). Additionally, there was no significant difference in the occurrence of polymicrobial infections between the sinus tract group (27.27%) and the non-sinus tract group (33.33%) (P = 0.37).ConclusionsMetagenomic next-generation sequencing can serve as a valuable screening tool in addition to traditional culture methods to improve diagnostic accuracy through optimized culture strategies.

Opinion review of drug resistant tuberculosis in West Africa: tackling the challenges for effective control.

Drug-resistant (DR) tuberculosis (TB) is a major public health concern globally, complicating TB control and management efforts. West Africa has historically faced difficulty in combating DR-TB due to limited diagnostic skills, insufficient access to excellent healthcare, and ineffective healthcare systems. This has aided in the emergence and dissemination of DR Mycobacterium tuberculosis complex (MTBC) strains in the region. In the past, DR-TB patients faced insufficient resources, fragmented efforts, and suboptimal treatment outcomes. However, current efforts to combat DR-TB in the region are promising. These efforts include strengthening diagnostic capacities, improving access to quality healthcare services, and implementing evidence-based treatment regimens for DR-TB. Additionally, many West African National TB control programs are collaborating with international partners to scale up laboratory infrastructure, enhance surveillance systems, and promote infection control measures. Moreso, novel TB drugs and regimens, such as bedaquiline and delamanid, are being introduced to improve treatment outcomes for DR-TB cases. Despite these obstacles, there is optimism for the future of DR-TB control in West Africa. Investments are being made to improve healthcare systems, expand laboratory capacity, and support TB research and innovation. West African institutions are now supporting knowledge sharing, capacity building, and resource mobilization through collaborative initiatives such as the West African Network for TB, AIDS, and Malaria (WANETAM), the West African Health Organization (WAHO), and other regional or global partners. These efforts hold promise for improved diagnostics, optimized treatment regimens, and provide better patient outcomes in the future where drug-resistant TB in WA can be effectively controlled, reducing the burden of the disease, and improving the health outcomes of affected individuals.

Assessment of the knowledge, attitudes, and practices toward human tuberculosis amongst rural communities in Chad.

Tuberculosis (TB) is an infectious zoonotic disease caused by bacteria belonging to the Mycobacterium tuberculosis complex. In sub-Saharan African countries such as Chad, TB is endemic and causes a high burden on humans and animals through morbidity, mortality, and reduced productivity in livestock. To effectively prevent and control the disease, strong coordination between policymakers, health and veterinary services, civil society organizations, and communities is needed. It also requires an understanding of the knowledge the communities have regarding TB. However, such knowledge is under-investigated, especially in rural areas. How knowledge affects people's attitudes and practices is also unclear. The main objective of this study was to investigate the knowledge, attitudes, and practices (KAP) of Chadian rural communities to better involve them in TB surveillance programs. A survey was conducted in 2021 in five rural health centers. Face-to-face interviews were conducted with persons suspected of having TB, and data on KAP were recorded and analyzed. In total, 139 participants were enrolled. Overall, the knowledge and attitude of the participants were found to be good to moderate, with 126 (90.6%) and 97 (69.7%) having good knowledge and attitude, respectively. However, their practices were found to be rather weak, with only 40 (28.7%) participants having good practices. Men were found to have good knowledge about the disease significantly more often than women. Poor attitude was significantly associated with a mobile lifestyle compared to settled lifestyles and with farmers (mainly engaged in agriculture) compared to breeders (livestock keepers). Poor health practices were associated more with men than women and with settled lifestyles compared to a mobile lifestyle. Good practices were found to be in line with good knowledge and good attitudes; however, in the analyses, the association was not significant [OR knowledge = 5.83 (95% C.I. 0.6842.83), p = 0.112; OR attitude = 2.09 (95% C.I. 0.875.04), p = 0.100]. Furthermore, attitude was not associated with knowledge [OR = 1.03 (95% C.I. 0.303.55), p = 0.964]. Our study highlights the need for targeted sensitization and awareness campaigns for communities with poor knowledge and attitudes regarding TB. These campaigns should also include practical training to increase the level of good practice rather than simply providing knowledge.

TUBERCULOSIS OF THE WRIST

INTRODUCTIONTuberculosis (TB) is a public health challenge. However, musculoskeletal involvement represents 10–15% of all extrapulmonary cases. Upper extremity involvement is extremely rare. The slow progressive course of clinical symptoms and lack of radiological signs lead to difficulties in establishing early diagnosis. Hence, the patients who have tuberculosis of the wrist are usually misdiagnosed. We report 5 cases of tuberculosis of the wrist seen in our unit from the year 2012 to 2021.METHODSCases were retrospectively evaluated on demographics, nature of history, clinical presentation, culture finding, and histopathological findings from 2012 to 2021 at our unit.RESULTSA total of 5 cases were evaluated retrospectively. Three patients were more than 60 years old and two were less than 30 years old. Four of the patients presented with wrist swelling 2 of them had wrist pain and only 1 patient had discharge from the wound. The duration of the symptoms ranges from 2 months to 3 years. Only one of the patients had a history of pulmonary TB contact whereas the others didn't. All patients underwent surgery. All of the patients had positive cultures for Mycobacterium tuberculosis complex and histopathological examination showed necrotizing granulomatous inflammation from specimens taken intraoperatively, which confirmed the diagnosis of tuberculosis of the wrist.DISCUSSIONSOur cases show that the common presentation of tuberculosis of the wrist was comparable to other literature. Most of the patients presented with chronic wrist swelling with or without wrist pain. The diagnosis of the disease was delayed an average of 10 - 12 months from symptoms onset to diagnosis. S. Bayram et al reported a case where the diagnosis was made 45 months later. Due to its rarity, it often is misdiagnosed, resulting in delays in the proper treatment.CONCLUSIONThe diagnosis of the TB wrist remains difficult because of insidious and non-specific. presentation. However, early diagnosis is essential to avoid delays in treatment and complications. Hence, chronic wrist pain, and swelling with or without unexplained bone erosion around the wrist area must be highly suspected of being osteoarticular tuberculosis.

Tuberculous Paraplegia: Outcome of Operative Intervention.

Tuberculosis of the spine, a common manifestation of extra-pulmonary tuberculosis is characterized by vertebral destruction, paradiscal involvement, abscess collection and varying degrees of neurologic affectation. The primary disease caused by mycobacterium tuberculosis complex infects the lungs, lymph nodes of the mediastinum and gastrointestinal tract/ viscera with spinal involvement being secondary and caused by haematogenous spread. Tuberculous paraplegia arises as a complication of spinal involvement. To determine the outcome of operative intervention in tuberculous paraplegia. Methodology: This was a retrospective study involving 10 patients with tuberculous spinal involvement with varying degrees of neurological deficit as defined by both Tuli and ASIA grading. The VAS score, ESR, ASIA grade (both pre-op and post op), Tuli's grade (pre-op and post op) were used to analyze the therapeutic effects of the surgery. The mean pre-operative VAS score was 5.9 ±1.8, which significantly decreased to 2.2 ±1.3 six weeks post operatively. The mean pre-operative ESR and CRP was 78.9 ± 11.3mm/hr and 83 ± 13.5 respectively; which both showed a statistically significant decrease post-operatively, p<0.05. All cases achieved an increase of more than one ASIA grade post-operatively. Early surgical intervention is beneficial in patients with tuberculous spinal disease with neurologic involvement.

Diagnostic Accuracy of the Abbott SD Bioline MPT64 antigen test for identification of MTB Complex in a U.S. Clinical Mycobacteriology Laboratory

Identification of the Mycobacterium tuberculosis complex (MTBC) from culture and differentiation from other non-tuberculous mycobacterial species is required for rapid diagnosis and accurate treatment. However, gaps exist in culture-based identification of acid-fast bacilli (AFB) positive cultures for rapid rule-out of MTBC in the United States. The SD Bioline™ MPT64 (Abbott Inc, South Korea) lateral flow assay (LFA) has high sensitivity and specificity for the detection of MTBC in liquid culture but has not been evaluated in a clinical mycobacteriology laboratory in the United States. We conducted a diagnostic accuracy study of this LFA for detection of MTBC versus NTMs on AFB positive cultures. A total of 362 tests were performed, with a sensitivity and specificity of 100 % (362/362) across all tests. The SD Bioline MPT64 assay provides accurate test results with AFB-positive liquid cultures and could fill the current gap for rapid rule-out of MTBC in U.S.-based clinical laboratories.

Resistance Rates of Mycobacterium tuberculosis Complex Isolates to Primary Anti-tuberculosis Drugs: A 5-Year Retrospective Study

Background/Aim: Tuberculosis remains a major global health problem with a high morbidity and mortality rate, approximately a quarter of the population is infected with tuberculosis. Drug susceptibility testing is an essential tool for identifying and managing drug-resistant tuberculosis. This study was conducted to evaluate the drug susceptibility pattern of Mycobacterium tuberculosis complex strains isolated from a university hospital. Methods: A total of 10900 samples sent to the microbiology laboratory with the suspicion of tuberculosis clinically between January 2018 and January 2022 were analyzed retrospectively. The automated BACTEC MGIT 960 (Becton Dickinson, USA) was used for sample culture and susceptibility testing. The obtained data were statistically analyzed with the Statistical Package for Social Sciences (SPSS version 20). Results: Out of the 154 isolated positive samples, males and females constituted equal parts of the study population (50%). The majority of tuberculosis cases were in the age group 56–75 years (42.2%), Pulmonary TB was detected in (90.3%) of the patients, while extrapulmonary TB cases were observed in (9.7%). As a result of susceptibility studies on positive samples, isoniazid resistance was 5.2%; streptomycin resistance 1.3%; ethambutol resistance was detected at a rate of 0.6%, while no rifampicin resistant sample was found. Both streptomycin and isoniazid resistance were seen together in 1.3% of the samples. Conclusion: A similar resistance pattern of the first-line antituberculosis drugs was observed in other studies conducted in different provinces of Turkey. The absence of multi-drug resistant and extensively drug-resistant tuberculosis in our study indicates that the tuberculosis surveillance program implemented in our region was successful.

Prosthetic Joint Infections Caused by Mycobacterium tuberculosis Complex-An ESGIAI-ESGMYC Multicenter, Retrospective Study and Literature Review.

While tuberculosis remains a significant global health concern, prosthetic joint infections (PJIs) caused by members of the Mycobacterium tuberculosis complex are exceptionally rare. Our objective is to perform a retrospective search of new cases of this disease and analyze all cases available in the literature of tuberculous PJIs, aiming to detect factors that may influence patient outcomes. The ESGIAI and ESGMYC study groups were used to collect information on non-published cases of tuberculous prosthetic joint infections (PJIs). Additionally, a literature review of all published cases of tuberculous PJIs was conducted. All identified cases in the retrospective study and in the literature review were merged and included in the statistical analysis, involving both univariate and multivariate analyses. Fifteen previously unreported cases of tuberculous prosthetic joint infections (PJIs) from four countries were detailed. Among them, ten patients were female, with a median age of 76 years. The hip was affected in 13 cases. Seven patients experienced co-infection with another microorganism. Treatment approaches varied, with 13 patients undergoing implant removal, one treated with DAIR (debridement, antibiotics, and implant retention), and one case was treated with an unknown treatment method. All patients received antibiotic therapy and achieved a cure. The literature review that was conducted detected 155 published cases. Univariate analysis revealed a statistical significance for previous tuberculosis, joint, and no importance of surgery for cure. Tuberculous prosthetic joint infection (PJI) is a rare condition, typically presenting as a localized chronic infection. Antibiotic treatment is essential for the management of these patients, but neither surgical treatment nor duration of treatment seems to have importance in the outcome.

Bacterial diversity dominates variable macrophage responses of tuberculosis patients in Tanzania

The Mycobacterium tuberculosis complex (MTBC) comprises nine human-adapted lineages that differ in their geographical distribution. Local adaptation of specific MTBC genotypes to the respective human host population has been invoked in this context. We aimed to assess if bacterial genetics governs MTBC pathogenesis or if local co-adaptation translates into differential susceptibility of human macrophages to infection by different MTBC genotypes. We generated macrophages from cryopreserved blood mononuclear cells of Tanzanian tuberculosis patients, from which the infecting MTBC strains had previously been phylogenetically characterized. We infected these macrophages ex vivo with a phylogenetically similar MTBC strain (“matched infection”) or with strains representative of other MTBC lineages (“mismatched infection”). We found that L1 infections resulted in a significantly lower bacterial burden and that the intra-cellular replication rate of L2 strains was significantly higher compared the other MTBC lineages, irrespective of the MTBC lineage originally infecting the patients. Moreover, L4-infected macrophages released significantly greater amounts of TNF-α, IL-6, IL-10, MIP-1β, and IL-1β compared to macrophages infected by all other strains. While our results revealed no measurable effect of local adaptation, they further highlight the strong impact of MTBC phylogenetic diversity on the variable outcome of the host–pathogen interaction in human tuberculosis.

Comparative Study of Vitamin D Levels in Newly Diagnosed Tuberculosis and a Normal Population.

Background and Objectives: Tuberculosis (TB) is an ancient disease caused by Mycobacterium tuberculosis, a member of the Mycobacterium tuberculosis complex. It contributes to significant morbidity and mortality. Treatment of TB poses a considerable challenge because of emerging drug resistance and the longer duration of therapy. Various past studies, both in vitro and in vivo, have established the role of vitamin D in the pathogenesis and treatment of TB. Results of in vivo studies are inconsistent, and this study aims to determine vitamin D levels and their association with newly diagnosed TB (pulmonary and extrapulmonary) cases and normal populations. Material and Methods: A Prospective Case-Control study with 116 subjects (58 cases and 58 controls) was conducted over two years. 29 cases of pulmonary TB and 29 cases of extrapulmonary TB constituted 58 cases of TB. Vitamin D levels were measured and compared in both the cases and controls. Data analysis was carried out using SPSS software 22.0. Results: The prevalence of vitamin D deficiency was 68.96% in the cases, while it was 51.72% in the controls. The reported median and quartile of serum vitamin D levels were 14.35 ng/mL (8.65, 25.48) in the TB group and 19.08 ng/mL (13.92, 26.17) in the control group. There was a significant statistical difference between the TB and non-TB populations with a p-value of 0.029 on the Mann-Whitney test. Conclusion: Vitamin D deficiency was more prevalent in individuals with TB than those without TB.

GenoMycAnalyzer: a web-based tool for species and drug resistance prediction for Mycobacterium genomes

BackgroundDrug-resistant tuberculosis (TB) is a major threat to global public health. Whole-genome sequencing (WGS) is a useful tool for species identification and drug resistance prediction, and many clinical laboratories are transitioning to WGS as a routine diagnostic tool. However, user-friendly and high-confidence automated bioinformatics tools are needed to rapidly identify M. tuberculosis complex (MTBC) and non-tuberculous mycobacteria (NTM), detect drug resistance, and further guide treatment options.ResultsWe developed GenoMycAnalyzer, a web-based software that integrates functions for identifying MTBC and NTM species, lineage and spoligotype prediction, variant calling, annotation, drug-resistance determination, and data visualization. The accuracy of GenoMycAnalyzer for genotypic drug susceptibility testing (gDST) was evaluated using 5,473 MTBC isolates that underwent phenotypic DST (pDST). The GenoMycAnalyzer database was built to predict the gDST for 15 antituberculosis drugs using the World Health Organization mutational catalogue. Compared to pDST, the sensitivity of drug susceptibilities by the GenoMycAnalyzer for first-line drugs ranged from 95.9% for rifampicin (95% CI 94.8–96.7%) to 79.6% for pyrazinamide (95% CI 76.9–82.2%), whereas those for second-line drugs ranged from 98.2% for levofloxacin (95% CI 90.1–100.0%) to 74.9% for capreomycin (95% CI 69.3–80.0%). Notably, the integration of large deletions of the four resistance-conferring genes increased gDST sensitivity. The specificity of drug susceptibilities by the GenoMycAnalyzer ranged from 98.7% for amikacin (95% CI 97.8–99.3%) to 79.5% for ethionamide (95% CI 76.4–82.3%). The incorporated Kraken2 software identified 1,284 mycobacterial species with an accuracy of 98.8%. GenoMycAnalyzer also perfectly predicted lineages for 1,935 MTBC and spoligotypes for 54 MTBC.ConclusionsGenoMycAnalyzer offers both web-based and graphical user interfaces, which can help biologists with limited access to high-performance computing systems or limited bioinformatics skills. By streamlining the interpretation of WGS data, the GenoMycAnalyzer has the potential to significantly impact TB management and contribute to global efforts to combat this infectious disease. GenoMycAnalyzer is available at http://www.mycochase.org.

Detection value of third-generation sequencing to identify the pathogenic organisms in prosthetic joint infection

To compare the detection value of third-generation sequencing (TGS) with pathogenic microbial culture in prosthetic joint infection (PJI). Arthrocentesis was performed on 29 patients who underwent hip and knee revision surgeries. In the PJI group, TGS detected 85.71 % of positive cases, while pathogenic microbial culture detected only 42.85 %. TGS identified 17 different pathogenic microorganisms, including Staphylococcus epidermidis, Staphylococcus aureus, Streptococcus lactis, and Mycobacterium tuberculosis complex. In the loosening group, TGS was positive in one patient, while microbial culture was negative in all cases. TGS showed higher sensitivity (85.71 % vs. 42.85 %), comparable specificity (93.33 % vs. 100 %), and similar positive predictive value (92.31 % vs. 100 %) compared to culture.However, TGS had a higher negative predictive value (87.5 % vs. 65.22 %).Additionally, TGS provided faster results (mean time 23.8±3.6 h) compared to microbial culture (mean time 108.0±9.4 h).These findings suggest that TGS holds promise for detecting pathogenic microorganisms in PJI and has potential for clinical application.

Impact of the COVID-19 pandemic on the real-world diagnostic infrastructure for tuberculosis-An ESGMYC collaborative study.

We determined the impact of the COVID-19 pandemic on mycobacterial diagnostic services. 40 laboratories from 22 countries completed an online questionnaire covering the redeployment of the laboratory infrastructure and/or staff for SARS-CoV-2 testing, staff shortages and supply chain disruptions. 28 laboratories reported monthly numbers of samples processed for mycobacterial investigations and monthly numbers of M. tuberculosis complex (MTBC) PCRs performed between October 1st 2018 and October 31st 2020. More than half (23/40) of the participating TB laboratories reported having performed COVID-19 diagnostics in the early phase of the pandemic, in part with negative impact on the mycobacterial service activities. All participating laboratories reported shortages of consumables and laboratory equipment due to supply chain issues. Average monthly sample numbers decreased by 24% between January 2020 and October 2020 compared to pre-pandemic averages. At the end of the study period, most participating laboratories had not returned to pre-pandemic average MTBC PCR throughput.

Clinical and morphological features of lung injury long-term after SARS-CoV-2 recovery

To study the clinical and histological profile of lung tissue in patients with persistent pulmonary disease, respiratory symptoms and CT findings after SARS-CoV-2 infection. The study included 15 patients (7 females and 8 males) with a mean age of 57.7 years. All patients underwent laboratory tests, chest computed tomography, echocardiography, and pulmonary function tests. Pulmonary tissue and bronchoalveolar lavage samples were obtained by fibrobronchoscopy, transbronchial forceps (2 patients), and lung cryobiopsy (11 patients); open biopsy was performed in 2 patients. Cellular composition, herpesvirus DNA, SARS-CoV-2, Mycobacterium tuberculosis complex, galactomannan optical density index, and bacterial and fungal microflora growth were determined in bronchoalveolar lavage. SARS-CoV-2 was also identified in samples from the nasal mucosa, throat and feces using a polymerase chain reaction. The results showed no true pulmonary fibrosis in patients recovered from SARS-CoV-2 infection with persistent respiratory symptoms, functional impairment, and CT findings after SARS-CoV-2 infection. The observed changes comply with the current and/or resolving infection and inflammatory process. Thus, no true pulmonary fibrosis was found in patients after SARS-CoV-2 infection with persistent respiratory symptoms, functional impairment, and CT findings. The observed changes comply with the current and/or resolving infection and inflammatory process.

Dual-centre evaluation of the FluoroType MTBDR version 2 assay for detection of Mycobacterium tuberculosis complex and resistance-conferring mutations in pulmonary and extrapulmonary samples from Denmark, Germany and Sierra Leone

ObjectivesWe evaluated the ability of FluoroType MTBDR version 2 (FTv2; Hain Lifescience), a second-step real-time PCR assay, to simultaneously detect Mycobacterium tuberculosis complex (MTBC) DNA and mutations conferring resistance to rifampicin (RIF) and isoniazid (INH), in pulmonary and extrapulmonary samples from patients and compared them with corresponding cultures. MethodsFTv2 MTBC was evaluated on 1815 and 432 samples from Denmark (DK) and Germany (DE), respectively. RIF and INH resistance mutations were assessed in the German samples and 110 samples from Sierra Leone and subsequently compared to phenotypic antimicrobial susceptibility testing and a composite reference DNA (CRD) based on the GenoType MTBDR line-probe assay and Sanger sequencing or whole-genome sequencing. ResultsOf the 584 (557 smear-negative) Danish and 277 (85 smear-negative) German sputum samples, 42 (16) and 246 (54) were culture positive, and 44 (18) and 222 (35) were FTv2 positive, providing an FTv2 sensitivity and specificity of 0.86 (0.63) and 0.98 (DK), 0.90 (0.65) and 1.00 (DE), respectively. The count, sensitivities, and specificities for all pulmonary samples were 1434, 0.79, and 0.99 (DK) and 347, 0.86, and 1.00 (DE), respectively; for extrapulmonary samples, 381, 0.33, 0.99 (DK) and 83, 0.50, and 1.00 (DE). The valid count, sensitivity, and specificity compared with CRD for detecting resistance mutations were RIF 355, 0.99, 0.96, and INH 340, 1.00, and 0.98, respectively. DiscussionFTv2 reliably detects MTBC DNA in pulmonary and extrapulmonary samples and detects resistance mutations for INH and RIF resistance in inhA promoter, katG, and rpoB genes.

Retrospective and prospective evaluation of the FluoroType®-Mycobacteria VER 1.0 assay for the identification of mycobacteria from cultures in a French center.

Rapid, reliable identification of mycobacteria from positive cultures is essential for patient management, particularly for the differential diagnosis of Mycobacterium tuberculosis complex (MTBC) and nontuberculous mycobacteria (NTM) species. The aim of the present study was to evaluate a new "In-Vitro-Diagnostic"-certified PCR kit, FluoroType®-Mycobacteria VER1.0 (Hain Lifescience GmbH) for NTM and MTBC identification from cultures. Mycobacteria identification isolated from positive cultures during routine practice at the Lyon university hospital mycobacteria laboratory obtained by hsp65 amplification/sequencing were compared retrospectively and prospectively to those obtained by and the FluoroType®-Mycobacteria VER1.0kit. The overall agreement between hsp65 amplification/sequencing and the FluoroType®-Mycobacteria VER1.0kit was 88.4% (84/95); 91.2% (52/57) for the retrospective period and 84.2% (32/38) for the prospective period. There were 9 (9.5%) minor discrepancies (species in the FluoroType®-Mycobacteria VER1.0database and identified at genus level): 4 during the retrospective period, 5 during the prospective period; and 2 (2.1%) major discrepancies (species in the FluoroType®-Mycobacteria VER1.0database and identified incorrectly to species level): 1 during the retrospective period (M. kumamotonense identified as M. abscessus subsp massiliense by the kit) and 1 during the prospective period (M. chimaera identified as M. smegmatis by the kit). Including concordant results at genus level and minor discrepancies, 17.9% (17/95) of strains were identified as Mycobacterium sp. by the FluoroType®-Mycobacteria-VER1.0kit. The good performance of the FluoroType®-Mycobacteria-VER1.0kit with few major discrepancies could enable its use for first-line identification of positive mycobacteria cultures. However, an alternative identification method at least for reference laboratories is needed owing to the non-negligible proportion of NTM strains were identified at genus level.

Mycobacterium tuberculosis complex and detection of rifampicin and isoniazid resistance, with phenotypic study and massive genome sequencing

Objective: The objective of this study is to compare the results obtained from the detection of MTBC (Mycobacterium tuberculosis Complex) and drug resistances between two commercial molecular methods, phenotypic Drug Susceptibility Testing (DST) and whole-genome sequencing. Materials and methods: Clinical samples positive for MTBC were selected. Identification and study of mutations associated with rifampicin and isoniazid resistance were carried out using 2 commercial molecular techniques and phenotypic DST, using critical concentration in liquid medium. In parallel, the analysis of Mycobacterium tuberculosis complex genome was performed by sequencing complete genomes.

Mycobacterium tuberculosis complex in domestic goats in Southern Spain

Tuberculosis (TB) is a zoonotic infectious disease caused by bacteria belonging to the Mycobacterium tuberculosis complex (MTC), which can affect a wide variety of domestic and wild animal species. Although the role of goats as a reservoir of MTC bacteria has been evidenced, information about the circulation of MTC strains in this species is still very scarce. The aim of the present study was to determine the seroprevalence, spatial distribution, risk factors and MTC spoligotypes circulating in goats from Andalusia (Southern Spain), the Spanish region with the largest goat census and a hotspot area of TB in both cattle and wild ungulates. A total of 2155 serum samples from 80 goat flocks were analyzed by an in-house ELISA using the P22 protein complex as a coating antigen. Antibodies against MTC were detected in 473 goats (21.9%, 95% CI: 20.2–23.7) and the true seroprevalence was 22.3% (95% CI: 20.6–24.1). Seropositivity was found in 72 (90.0%) of the 80 flocks analyzed. The generalized estimating equation model showed that the management system (higher seroprevalence on intensive and semi-intensive farms), and the presence of hospital pens inside the regular stables, were risk factors potentially associated with MTC exposure in goats in Southern Spain. The spatial analysis identified a significant spatial cluster (p < 0.001) in Eastern Andalusia. A total of 16 different MTC spoligotypes, including five of M. caprae and eleven of M. bovis, were identified in goats between 2015 and 2022 in the study area, with SB0157 as the most frequently isolated. The results obtained indicate widespread and non-homogeneous spatial distribution of MTC in goat herds from Southern Spain. The high individual and herd-level seroprevalence values found suggest that goats could play a significant role in the maintenance and transmission of MTC in the study area. Our results highlight the importance of implementing control measures in this species.