Liquid Culture Testing Research Articles

Evaluation of GeneNAT Real-Time Polymerase Chain Reaction Analyzer and Pre-loaded Chip-Based Mycobacterium tuberculosis and Multidrug-Resistant Tuberculosis Detection in the Diagnosis of Pulmonary Tuberculosis.

Tuberculosis (TB) is still the second causative agent of death worldwide after COVID-19. It is caused by Mycobacterium tuberculosis (MTB) infection. The aim of the current study was to compare the performance of GeneNAT real-time polymerase chain reaction analyzer and pre-loaded chip-based MTB screening and multidrug-resistant tuberculosis (MDR-TB) detection kit (Smart SureTM MTB & MDR-TB, Genetix Biotech Asia Pvt. Ltd., New Delhi, India) against the established WHO-approved GeneXpert Ultra (MTB/rifampicin (RIF)), line probe assay (LPA), and mycobacteriagrowthindicatortube(MGIT) culture at point of care (POC) level. A total of 450 pulmonary TB (PTB) suspect patients were enrolled from October 2023 to March 2024 at the Intermediate Reference Laboratory, Department of Medicine, All India Instituteof Medical Sciences, New Delhi, India. GeneXpert and GeneNAT tests were done directly from sputum specimens. However, processed sputum specimens were used for both LPA (GenoType MTBDRplus) and liquid culture and drug susceptibility testing (MGIT culture and drug susceptibility testing (DST)). On comparing with GeneXpert, for the detection of MTB and rifampicin (RIF), Smart SureTM showed a sensitivity of 98.18% and 97.5% with a specificity of 100% and 98.92%, respectively. While comparing mutations in therpoB gene with LPA, the Smart SureTM MDR-TB kit exhibited sensitivity and specificity of 96.77% and 99.12%, respectively. For katG and inhA genes, sensitivity and specificity were 97.6% & 85.71% and 98.66% & 98.01%, respectively, for both genes. Smart SureTM MDR-TB showed comparable results with MGIT-DST with sensitivity and specificity of 96.88% & 96.15% and 98.99%& 99.02%, respectively, for both RIF and isoniazid (INH) drugs. The GeneNAT system test may provide the status of RIF and INH resistance in PTB cases in a short time with the use of minimal specimens. It required very little infrastructurewith less skilled laboratory staffin comparison with other WHO-approved diagnostics used in resource-limited countries with TB and drug-resistant TB burdens.

Effectiveness of Histopathological Examination of Ultrasound-guided Puncture Biopsy Samples for Diagnosis of Extrapulmonary Tuberculosis

ObjectiveTo evaluate the diagnostic value of histopathological examination of ultrasound-guided puncture biopsy samples in extrapulmonary tuberculosis (EPTB). MethodsThis study was conducted at the Shanghai Public Health Clinical Center. A total of 115 patients underwent ultrasound-guided puncture biopsy, followed by MGIT 960 culture (culture), smear, GeneXpert MTB/RIF (Xpert), and histopathological examination. These assays were performed to evaluate their effectiveness in diagnosing EPTB in comparison to two different diagnostic criteria: liquid culture and composite reference standard (CRS). ResultsWhen CRS was used as the reference standard, the sensitivity and specificity of culture, smear, Xpert, and histopathological examination were (44.83%, 89.29%), (51.72%, 89.29%), (70.11%, 96.43%), and (85.06%, 82.14%), respectively. Based on liquid culture tests, the sensitivity and specificity of smear, Xpert, and pathological examination were (66.67%, 72.60%), (83.33%, 63.01%), and (92.86%, 45.21%), respectively. Histopathological examination showed the highest sensitivity but lowest specificity. Further, we found that the combination of Xpert and histopathological examination showed a sensitivity of 90.80% and a specificity of 89.29%. ConclusionUltrasound-guided puncture sampling is safe and effective for the diagnosis of EPTB. Compared with culture, smear, and Xpert, histopathological examination showed higher sensitivity but lower specificity. The combination of histopathology with Xpert showed the best performance characteristics.

Laboratory Diagnosis of Genital Tuberculosis: A Narrative Review

Genital tuberculosis is a leading cause of infertility in high tuberculosis burden countries like Bangladesh. Diagnosing this condition can be challenging due to the diverse ways it can present, which requires a physician to have a high level of suspicion. The fallopian tubes are mainly affected, and the endometrium in 50% of cases1. Diagnostic modalities usually include histopathology, microscopy, solid and liquid culture tests, Polymerase Chain Reaction (PCR) tests, immunohistochemistry tests, radiology imaging and laparoscopic and hysteroscopic procedures.
 Bangladesh Journal of Infectious Diseases, December 2023;10(2):92-100

Comparison of Xpert MTB/RIF Ultra with Xpert MTB/RIF for the detection of Mycobacterium tuberculosis and rifampicin resistance in a primary-level clinic in rural China

The Xpert MTB/RIF Ultra (Ultra) is not yet used for the diagnosis of tuberculosis (TB) in China. We compared the performance of the Xpert and Ultra for detecting Mycobacterium tuberculosis and rifampicin resistance in a primary-level clinic in rural China. Sputum samples from suspected pulmonary TB patients were collected and subjected to smear microscopy, liquid culture, Xpert and Ultra tests. We then compared the sensitivity and specificity of Xpert and Ultra for diagnosing TB against liquid culture. Whole-genome sequencing was performed to predict rifampicin resistance and the results were compared with the Xpert and Ultra tests. The sensitivities of Xpert and Ultra were 88.1% and 95.1%, and the specificities were 91.9% and 84.4%, respectively. Among the 61 smear-negative culture-positive patients, the sensitivities of Xpert and Ultra were 80.3% and 91.8%. All Xpert-positive patients were Ultra-positive. Among culture-negative Xpert or Ultra-positive patients, 69.6% were taking anti-TB drugs or had a previous history of TB. Of the samples that Ultra classified as trace, nearly 25% were probably false-positives. Both Xpert and Ultra accurately detected all rifampicin-resistant patients. In conclusion, Ultra was more sensitive than Xpert, especially for smear-negative patients but had decreased specificity with more false-positives, especially with Ultra trace results.

Tuberculosis screening in respiratory department inpatients in comprehensive medical institutions

Objective: To explore the effect of screening tuberculosis patients in the respiratory department of general hospitals, and to provide a basis for the development of patient screening strategy. Methods: Clinical information and sputum samples of inpatients in the respiratory department of a general hospital in Longhua District, Shenzhen from December 2018 to December 2020 were collected. Sputum samples were sent to the tuberculosis laboratory of the Shenzhen Longhua Center for Chronic Disease Control (designated tuberculosis diagnosis and treatment institution) for sputum smear, liquid culture and Gene-Xpert test. Results: A total of 407 sputum samples (23 cases of suspected tuberculosis by chest imaging and 384 by clinical manifestations) were collected from 3 724 hospitalized patients. A total of 88 patients with positive etiology were detected by the three methods, and the positive rate was 21.6% (88/407), among which 15 patients with suspected tuberculosis were detected by imaging reports, and the positive rate of etiology was 19.0% (73/384) in the reported patients without imaging reports. At least 1.96% (73/3 724) of the hospitalized patients were estimated to be tuberculosis positive during the study. Pneumonia (30.1%,22/73), cough (15.1%,11/73) and pulmonary infection (15.1%,11/73) were the main characteristics in the patients with positive pathogens. Conclusions: Screening for tuberculosis among inpatients in the respiratory department of general hospitals is an effective way to detect patients who were radiographically reported to have probable tuberculosis. It is of great significance to carry out active screening in key departments of general hospitals for tuberculosis detection and control.

Optimising Xpert-Ultra and culture testing to reliably measure tuberculosis prevalence in the community: findings from surveys in Zambia and South Africa

ObjectivesPrevalence surveys remain the best way to assess the national tuberculosis (TB) burden in many countries. Challenges with using culture (the reference standard) for TB diagnosis in prevalence surveys have...

Enhanced hexazinone degradation by a Bacillus species and Staphylococcus species isolated from pineapple and sugarcane cultivated soils in Kenya

In this work, hexazinone-degrading bacterial strains from Kenyan tropical soils with long term application history were isolated and identified. Non-sterile soils from sugarcane and pineapple cultivated fields with over 15 years' hexazinone application history degraded 82.2% and 93.4% initially applied hexazinone under laboratory conditions after 146 days of incubation, respectively. In contrast, non-sterile soils without history of application degraded 48.8% and 36.8% of hexazinone after 146 days, respectively. From liquid culture tests, using soils with prior application history from sugarcane and pineapple cultivated soils, two hexazinone-degrading bacterial strains were isolated and identified as Staphylococcus gallinarum and Bacillus toyonensis and Bacillus thuringiensis, respectively. The pure isolates rapidly degraded hexazinone up to 38.4% and 53.2% of 50 mg/L in 46 days, respectively, with microbial hexazinone metabolite B previously reported detected. These bacterial strains from Kenyan soils have been identified for the first time as prospective hexazinone-degraders from pineapple and sugarcane-cultivated soils.

Digital Chest Radiography Enhances Screening Efficiency for Pulmonary Tuberculosis in Primary Health Clinics in South Africa.

Optimized tuberculosis (TB) screening in high burden settings is essential for case finding. We evaluated digital chest X-ray with computer-aided detection (CAD) software (d-CXR) for identifying undiagnosed TB in three primary health clinics in South Africa. The cross-sectional study consented adults who were sequentially screened for TB using the World Health Organization (WHO) 4 symptom questionnaire and d-CXR. Participants reporting ≥1 TB symptom and/or CAD score ≥60 (suggestive of TB) provided 2 spot sputum for Xpert MTB/RIF Ultra (Xpert Ultra) and liquid culture testing, respectively. TB yield (proportion of screened tested positive) and number needed to test (NNT; no of tests to identify one TB patient) were calculated. Risk factors for microbiologically confirmed or presumed (on radiological grounds) were determined. Among 3041 participants, 45% (1356 of 3041) screened positive on either d-CXR or symptoms. TB yield was 2.3% (71 of 3041) using Xpert Ultra and 2.7% (82 of 3041) using Xpert Ultra plus culture. Modelled TB yield (identified by Xpert Ultra) by screening approach was: 1.9% (59 of 3041) for d-CXR alone, 2.0% (62 of 3041) for symptoms alone and 2.3% (71 of 3041) for both. The NNT was 9.7 for d-CXR, 17.8 for symptoms and 19.1 for d-CXR and/or symptom. Males, those with previous TB, untreated HIV or unknown HIV status, and acute illness were at higher risk of developing TB. d-CXR screening identified a similar yield of undiagnosed TB compared to symptom-based screening, however required fewer diagnostic tests. Due to its objective nature, d-CXR screening may improve case detection in clinics.

GeneXpert on patients with human immunodeficiency virus and smear-negative pulmonary tuberculosis.

ObjectivesVietnam is a high-prevalence country for tuberculosis (TB). Xpert MTB/RIF is a novel PCR-based diagnostic test that is substantially more sensitive for detecting M. tuberculosis than traditional smear-based techniques. However, locally-derived evidence of Xpert MTB/RIF in HIV-infected people is limited. This study evaluates the performance of the Xpert MTB/RIF in HIV-infected patients with smear-negative pulmonary TB (SNTB).MethodsThis was a cross-sectional study in 3 hospitals. The performance of Xpert MTB/RIF was compared with the reference standard of liquid culture and phenotypic drug-susceptibility testing for rifampicin (RIF) resistance.ResultsOut of 123 patients, the median age was 37.0 (IQR: 32.0–41.0) and 81.3% were male. The area under the receiver operating characteristic curve, sensitivity (Se) and specificity (Sp) of Xpert MTB/RIF for pulmonary TB diagnosis were 0.72 (95% confidence interval [CI]: 0.63–0.81), 66.7% (95%CI: 54.8–77.1) and 77.1% (95%CI: 62.7–88.0), respectively, while Se and Sp of Xpert MTB/RIF in detecting RIF resistance were 50.0 (11.8–88.2) and 86.4% (95%CI: 72.7–94.8).ConclusionThe performance of Xpert MTB/RIF in HIV-infected patients with SNTB for the diagnosis of TB and RIF-resistance was low. Further studies are required to evaluate the results of Xpert MTB/RIF assay in HIV-infected patients with SNTB and the role of Xpert repetition on the same specimens.

Phosphoproteomic Characterization of Primary AML Samples and Relevance for Response Toward FLT3-inhibitors.

Phosphoproteomic Characterization of Primary AML Samples and Relevance for Response Toward FLT3-inhibitors.

Bioaugmentation of animal feces as a mean to mitigate environmental contamination with anthelmintic benzimidazoles

Anthelmintics are used to control infestations of ruminants by gastrointestinal nematodes. The limited metabolism of anthelmintics in animals result in their excretion in feces. These could be piled up in the floor of livestock farms, constituting a point source of environmental contamination, or used as manures in agricultural soils where they persist or move to water bodies. Hence the removal of anthelmintics from feces could mitigate environmental contamination. We hypothesized that a thiabendazole-degrading bacterial consortium would also degrade other benzimidazole anthelmintics like albendazole, fenbendazole, ricobendazole, mebendazole and flubendazole. In liquid culture tests the consortium was more effective in degrading compounds with smaller benzimidazole substituents (thiabendazole, albendazole, ricobendazole), rather than benzimidazoles with bulky substituents (fenbendazole, flubendazole, mebendazole). We then explored the bioaugmentation capacity of the consortium in sheep feces fortified with 5 and 50 mg kg−1 of thiabendazole, albendazole and fenbendazole. Bioaugmentation enhanced the degradation of all compounds and its efficiency was accelerated upon fumigation of feces, in the absence of the indigenous fecal microbial community. The latter contributes to anthelmintics degradation as suggested by the significantly lower DT50 values in fumigated vs non-fumigated, non-bioaugmented feces. Overall, bioaugmentation could be an efficient means to reduce environmental exposure to recalcitrant anthelmintic benzimidazoles.

Spectrum of Drug Resistance in Musculoskeletal Tuberculosis.

Very few studies report resistance pattern exclusively in musculoskeletal tuberculosis (MSK-TB). This study of 100 pus samples from patients of MSK-TB with active disease in whom Mycobacterium tuberculosis (MTB) was detected by cartridge-based nucleic acid amplification test (CBNAAT), revealed the pattern of resistance among newly diagnosed and previously treated cases. Liquid culture and drug susceptibility testing (DST) using MGIT 960 was done for 11 anti-tubercular drugs. Among these 100 cases; 22% were AFB positive; MGIT 960 detected MTB in 58.33% (35/60) new cases and 30.0% (12/40) previously treated cases. Five new and 10 previously treated cases had drug resistance and 12 were detected rifampicin resistance (Rif-R) by CBNAAT. Among new cases MGIT-DST detected mono-INH resistant in 2.86% (1/35), mono-STR resistant in 2.86% (1/35), MDR-TB in 5.7% (2/35) and pre-XDR in 2.9%(1/35).Among previously treated cases Rif-R was found in 10% (4/40) where MTB was not detected by MGIT and MGIT-DST detected mono-INH resistant in 8.33% (1/12); MDR-TB in 8.33% (1/12) and pre-XDR in 33.3%. There were no cases of XDR-TB. High disease burden of various type drug resistance were seen more commonly in previously treated cases and was not uncommon in new cases of MSK-TB. Both CBNAAT and DST are essential for detecting resistance pattern in MSK-TB.

The transition to Xpert MTB/RIF ultra: diagnostic accuracy for pulmonary tuberculosis in Kampala, Uganda

BackgroundThe World Health Organization (WHO) has endorsed the next-generation Xpert MTB/RIF Ultra (Ultra) cartridge, and Uganda is currently transitioning from the older generation Xpert MTB/RIF (Xpert) cartridge to Ultra as the initial diagnostic test for pulmonary tuberculosis (TB). We assessed the diagnostic accuracy of Ultra for pulmonary TB among adults in Kampala, Uganda.MethodsWe sampled adults referred for Xpert testing at two hospitals and a health center over a 12-month period. We enrolled adults with positive Xpert and a random 1:1 sample with negative Xpert results. Expectorated sputum was collected for Ultra, and for solid and liquid culture testing for Xpert-negative patients. We measured sensitivity and specificity of Ultra overall and by HIV status, prior history of TB, and hospitalization, in reference to Xpert and culture results. We also assessed how classification of results in the new “trace” category affects Ultra accuracy.ResultsAmong 698 participants included, 211 (30%) were HIV-positive and 336 (48%) had TB. The sensitivity of Ultra was 90.5% (95% CI 86.8–93.4) and specificity was 98.1% (95% CI 96.1–99.2). There were no significant differences in sensitivity and specificity by HIV status, prior history of TB or hospitalization. Xpert and Ultra results were concordant in 670 (96%) participants, with Ultra having a small reduction in specificity (difference 1.9, 95% CI 0.2 to 3.6, p=0.01). When “trace” results were considered positive for all patients, sensitivity increased by 2.1% (95% CI 0.3 to 3.9, p=0.01) without a significant reduction in specificity (− 0.8, 95% CI − 0.3 to 2.0, p=0.08).ConclusionsAfter 1 year of implementation, Ultra had similar performance to Xpert. Considering “trace” results to be positive in all patients increased case detection without significant loss of specificity. Longitudinal studies are needed to compare the benefit of greater diagnoses to the cost of overtreatment.

Blood transcriptional biomarkers for active pulmonary tuberculosis in a high-burden setting: a prospective, observational, diagnostic accuracy study

SummaryBackgroundBlood transcriptional signatures are candidates for non-sputum triage or confirmatory tests of tuberculosis. Prospective head-to-head comparisons of their diagnostic accuracy in real-world settings are necessary to assess their clinical use. We aimed to compare the diagnostic accuracy of candidate transcriptional signatures identified by systematic review, in a setting with a high burden of tuberculosis and HIV.MethodsWe did a prospective observational study nested within a diagnostic accuracy study of sputum Xpert MTB/RIF (Xpert) and Xpert MTB/RIF Ultra (Ultra) tests for pulmonary tuberculosis. We recruited consecutive symptomatic adults aged 18 years or older self-presenting to a tuberculosis clinic in Cape Town, South Africa. Participants provided blood for RNA sequencing, and sputum samples for liquid culture and molecular testing using Xpert and Ultra. We assessed the diagnostic accuracy of candidate blood transcriptional signatures for active tuberculosis (including those intended to distinguish active tuberculosis from other diseases) identified by systematic review, compared with culture or Xpert MTB/RIF positivity as the standard reference. In our primary analysis, patients with tuberculosis were defined as those with either a positive liquid culture or Xpert result. Patients with missing blood RNA or sputum results were excluded. Our primary objective was to benchmark the diagnostic accuracy of candidate transcriptional signatures against the WHO target product profile (TPP) for a tuberculosis triage test.FindingsBetween Feb 12, 2016, and July 18, 2017, we obtained paired sputum and RNA sequencing data from 181 participants, 54 (30%) of whom had confirmed pulmonary tuberculosis. Of 27 eligible signatures identified by systematic review, four achieved the highest diagnostic accuracy with similar area under the receiver operating characteristic curves (Sweeney3: 90·6% [95% CI 85·6–95·6]; Kaforou25: 86·9% [80·9–92·9]; Roe3: 86·9% [80·3–93·5]; and BATF2: 86·8% [80·6–93·1]), independent of age, sex, HIV status, previous tuberculosis, or sputum smear result. At test thresholds that gave 70% specificity (the minimum WHO TPP specificity for a triage test), these four signatures achieved sensitivities between 83·3% (95% CI 71·3–91·0) and 90·7% (80·1–96·0). No signature met the optimum criteria, of 95% sensitivity and 80% specificity proposed by WHO for a triage test, or the minimum criteria (of 65% sensitivity and 98% specificity) for a confirmatory test, but all four correctly identified Ultra-positive, culture-negative patients.InterpretationSelected blood transcriptional signatures met the minimum WHO benchmarks for a tuberculosis triage test but not for a confirmatory test. Further development of the signatures is warranted to investigate their possible effects on clinical and health economic outcomes as part of a triage strategy, or when used as add-on confirmatory test in conjunction with the highly sensitive Ultra test for Mycobacterium tuberculosis DNA.FundingRoyal Society Newton Advanced Fellowship, Wellcome Trust, National Institute of Health Research, and UK Medical Research Council.

Evaluation of modified petroff’s method and manual MGIT culture system to facilitate detection of acid-fast bacilli in smear-negative presumptive cases of pulmonary tuberculosis

Background: Tuberculosis is a major public health problem in India. According to RNTCP guidelines, all efforts are to be made to diagnose tuberculosis on the basis of detection of AFB/ MTB in the clinical specimen; hence all those cases where AFB/ MTB have not been detected are presumptive cases of tuberculosis. In this background, the present study aims to detect AFB/ MTB in the sputum of clinico-radiologically presumptive cases of pulmonary tuberculosis using direct ZN microscopy, Modified Petroff’s method followed by ZN microscopy & MGIT liquid culture method. Materials and Methods: A total of 94 sputum samples which were negative for AFB by direct ZN microscopy at the DMC of the institute, were subjected to Modified Petroff’s method followed by microscopy & MGIT liquid culture test to find out any additional yield of bacteriologically confirmed disease. Result: Out of 94 specimens, 20 (21.28%) were positive for AFB on ZN microscopy post Modified Petroff’s method. Among all the specimens, 30 (31.91%) were found to be positive by MGIT liquid culture method with average time to detection growth (TTD) around 16.73 days (2-38 days). Conclusion: Modified Petroff’s method followed by ZN microscopy & MGIT test for mycobacterial culture can improve case detection.

Screening of pyrene degradation bacteria strain from the Kochia scoparia roots and its potential for promoting growth

The endophytic bacteria were isolated from the roots of polycyclic aromatic hydrocarbon (PAHs)-tolerant plant. We investigated their ability of PAHs degradation and plant growth promo-ting, with the aim to provide theoretical support for bacterial-plant cooperative soil remediation. Kochia scoparia living in coking plant area were selected for strains isolation. Eight endophytic bacteria strains, which used pyrene and 1-aminocyclopropane-1-carboxylic acid (ACC) as sole carbon and nitrogen source, were isolated from the roots of K. scoparia. Three endophytic bacteria, KSE4, KSE7 and KSE8, displayed high degradation efficiency in pyrene degradation experiment. They were identified as Bacillus sp., Pseudomonas sp., and Sphingobacterium sp., respectively. The abili-ties of those three strains to produce ACC deaminase and their effects on seed germination of K. scoparia were examined under pyrene stress through liquid culture tests. The results showed that the activity of ACC deaminase decreased with increasing pyrene concentration (0-15 mg·L-1). KSE7 had the strongest promotion effect. When pyrene concentration reached to 15 mg·L-1, the germination rate and bud length of K. scoparia increased by 44.8% and 61.1%, respectively. Our results indicated that KSE7 is a promising bacterial strain for soil remediation in coking plant area.

Pleural tuberculosis: A concise clinical review.

Tuberculosis (TB) is the leading infectious cause of death worldwide, and the commonest cause of death in people living with HIV. Globally, pleural TB remains one of the most frequent causes of pleural exudates, particularly in TB-endemic areas and in the HIV positive population. Most TB pleural effusions are exudates with high adenosine deaminase (ADA), lymphocyte-rich, straw-coloured and free flowing, with a low yield on mycobacterial culture. TB pleurisy can also present as loculated neutrophil-predominant effusions which mimic parapneumonic effusions. Rarely, they can present as frank TB empyema, containing an abundance of mycobacteria. Up to 80% of patients have parenchymal involvement on chest imaging. The diagnosis is simple if M. tuberculosis is detected in sputum, pleural fluid or biopsy specimens, and the recent advent of liquid medium culture techniques has increased the microbiological yield dramatically. Where the prevalence of TB is high the presence of a lymphocyte-predominant exudate with a high ADA has a positive predictive value of 98%. In low prevalence areas, the absence of an elevated ADA and lymphocyte predominance makes TB very unlikely, and pleural biopsy should be performed to confirm the diagnosis. Pleural biopsy for liquid culture and susceptibility testing must also be considered where the prevalence of drug resistant TB is high. Treatment regimens are identical to those administered for pulmonary TB. Initial pleural drainage may have a role in symptom relief and in hastening the resolution of the effusion. Surgical intervention may be required in loculated effusions and empyemas.

Drug resistance patterns among extra-pulmonary tuberculosis cases in a tertiary care centre in North India.

xtra-pulmonary tuberculosis (EPTB) is a growing public health concern, and data on drug resistance are limited. Specimens from 2468 clinically diagnosed EPTB patients received at the Intermediate Reference Laboratory (IRL) of a tertiary centre in India were subjected to Ziehl-Neelsen staining, Xpert® MTB/RIF testing, liquid culture and drug susceptibility testing (DST) using automated BACTEC MGIT™ 960™. Line-probe assay (LPA) was performed on all culture-positive isolates. Gene sequencing was performed on rifampicin-resistant/multidrug-resistant TB (RR/MDR-TB) and phenotypic/genotypic discrepant isolates. The culture positivity rate was 18.9% (483/2553). The sensitivity and specificity of Xpert in diagnosing EPTB were respectively 70.8% (95%CI 66.5-74.8) and 97.7% (95%CI 96.9-98.3), with liquid culture as the reference standard. Prevalence of RR/MDR-TB was 10.1% (49/483). Prevalence of pre-extensively drug-resistant TB (pre-XDR-TB) was 18.4% (09/49), whereas the prevalence of XDR-TB among MDR-TB patients was 2% (01/49). The sensitivity of genotypic DST for the detection of rifampicin resistance was 92.7% (95%CI 81.1-98.5) and specificity was 99.3% (95%CI 97.5-99.9), with 100% concordance between Xpert and LPA. The burden of drug resistance, including M/XDR-TB, among EPTB patients is high. Novel molecular tests can help in early diagnosis and treatment to prevent disease progression and amplification of resistance.

1-(2-aminophenyl)-1H-1,2,3-triazole-4-carboxylic acid: activity against Gram-positive and Gram-negative pathogens including Vibrio cholerae

We report a new synthetic aromatic ε-amino acid containing a triazole moiety with antimicrobial potential against Gram-positive, Gram-negative and pathogenic bacteria including Vibrio cholerae. Structure–property relationship studies revealed that all the functional groups are essential to enhance the antimicrobial activity. The 1-(2-aminophenyl)-1H-1,2,3-triazole-4-carboxylic acid was synthesized by click chemistry. From X-ray crystallography, the amino acid adopts a kink-like structure where the phenyl and triazole rings are perpendicular to each other and the amine and acid groups maintain an angle of 60°. The agar diffusion test shows that the amino acid has significant antibacterial activity. The liquid culture test exhibits that the minimum inhibitory concentration (MIC) value for Bacillus subtilis and Vibrio cholerae is 59.5 µg ml−1. FE-SEM experiments were performed to study the morphological changes of bacterial shape after treatment with compound 1. The antimicrobial activity of the amino acid was further studied by DNA binding and degradation study, protein binding, dye-binding assay and morphological analysis. Moreover, the amino acid does not have any harmful effect on eukaryotes.

Regulation of root development in Arabidopsis thaliana by phytohormone-secreting epiphytic methylobacteria.

In numerous experimental studies, seedlings of the model dicot Arabidopsis thaliana have been raised on sterile mineral salt agar. However, under natural conditions, no plant has ever grown in an environment without bacteria. Here, we document that germ-free (gnotobiotic) seedlings, raised on mineral salt agar without sucrose, develop very short root hairs. In the presence of a soil extract that contains naturally occurring microbes, root hair elongation is promoted; this effect can be mimicked by the addition of methylobacteria to germ-free seedlings. Using five different bacterial species (Methylobacterium mesophilicum, Methylobacterium extorquens, Methylobacterium oryzae, Methylobacterium podarium, and Methylobacterium radiotolerans), we show that, over 9days of seedling development in a light-dark cycle, root development (hair elongation, length of the primary root, branching patterns) is regulated by these epiphytic microbes that occur in the rhizosphere of field-grown plants. In a sterile liquid culture test system, auxin (IAA) inhibited root growth with little effect on hair elongation and significantly stimulated hypocotyl enlargement. Cytokinins (trans-zeatin, kinetin) and ethylene (application of the precursor ACC) likewise exerted an inhibitory effect on root growth but, in contrast to IAA, drastically stimulated root hair elongation. Methylobacteria are phytosymbionts that produce/secrete cytokinins. We conclude that, under real-world conditions (soil), the provision of these phytohormones by methylobacteria (and other epiphytic microbes) regulates root development during seedling establishment.