Detection of Drug Resistant Mycobacterium Tuberculosis Strains Using Kit SIRE Nitratase®: a Multicenter Study

Abstract (1) Background: The Commercial Kit SIRE Nitratase® PlastLabor, is a drug susceptibility test kit used to detect Mycobacterium tuberculosis resistance to first-line TB treatment drugs. The present study aimed at evaluating its performance in a multicenter study. (2) Methods: To determine its accuracy, the proportion methods in Lowenstein Jensen medium or the BACTECTMMGITTM960 system was used as a gold standard. (3) Results: The study revealed that the respective accuracies of the kit with 190 M. tuberculosis clinical isolates, using the proportion methods in Lowenstein Jensen medium or BACTECTMMGITTM960 system as a gold standard, were 93.9% and 94.6%, 96.9% and 94.6%, 98.0% and 97.8%, and 98.0% and 98.9%, for streptomycin, isoniazid, rifampicin, and ethambutol, respectively. (4) Conclusion: Thus, the kit can rapidly screen resistance to streptomycin, isoniazid, rifampicin, and ethambutol. Additionally, it does not require sophisticated equipment; hence, it can be easily used in the laboratories of low and middle income countries.

Fast detection of Mycobacterium tuberculosis in culture-positive sputum samples by nitrate reductase activity

ABSTRACT Microscopy and bacterial culture are the main tools in the diagnosis of tuberculosis. Since the slow growth of Mycobacterium tuberculosis impairs rapid diagnosis strategies, especially in countries where the latter are the only available resources, the ongoing development of new and inexpensive tools based on mycobacterial metabolism optimizing growth detection with preliminary identification is greatly welcome. When compared to the other species from the M. tuberculosis complex, M. tuberculosis is a strong nitrate reducer. Current assay compares the nitrate reductase activity of M. tuberculosis from pulmonary specimens cultivated in nitrate-supplemented media. Fifty-five sputum samples were decontaminated and inoculated in conventional (Middlebrook 7H9, Ogawa Kudoh-OK) and in nitrate-supplemented media (Middlebrook 7H9-N, Ogawa Kudoh-N). An aliquot from the media directly reacted with Griess reagent (7H9-N and OK-N) every five days, or transferred to a nitrate substrate solution (7H9, OK). Nitrate to nitrite reduction was considered positive, revealed by the pink color, indicating bacterial growth. As reference method, the Mycobacteria Growth Indicator Tube (MGIT) was used for sensitivity calculations and statistical analysis. 7H9-N and OK-N assays proved to perform better in detecting M. tuberculosis than conventional assays (7H9 and OK). Indeed, broth nitrate-supplemented medium (7H9-N) was comparable to MGIT to detect M. tuberculosis, except in growth detection time. Results show that 7H9-N may be used as an alternative tool particularly in low-income countries since it is a simple and cheap technique, and does not restrict diagnosis to single-source products.

Evaluation of the commercial kit SIRE nitratase for detecting resistant mycobacterium tuberculosis in Brazil

Abstract INTRODUCTION: This study aimed to evaluate a new commercial kit, Kit SIRE Nitratase-PlastLabor, for testing the drug susceptibility of clinical Mycobacterium tuberculosis isolates. METHODS: The accuracy of the Kit SIRE Nitratase was evaluated by examining the susceptibility (streptomycin, isoniazid, rifampicin, and ethambutol) of 40 M. tuberculosis isolates, using the proportion method with Lowenstein-Jensen medium or the BACTEC MGIT 960 system. RESULTS: The detection accuracy for streptomycin, isoniazid, rifampicin, and ethambutol was 95%, 97.5%, 100%, and 80%, respectively. CONCLUSIONS: The exceptional accuracy demonstrated by Kit SIRE Nitratase for isoniazid and rifampicin makes the kit an attractive option for screening M. tuberculosis strain resistance.

Resposta de híbridos de milho ao uso de piraclostrobina na ausência de doenças / Corn hybrids responses to pyraclostrobin use in disease absences

Objetivando-se avaliar o efeito da aplicação de piraclostrobina em diferentes épocas e combinações de aplicação em dois híbridos simples de milho cultivados na safra de verão, realizou-se um experimento no município de Jataí, Goiás. Adotou-se o delineamento de blocos ao acaso no esquema fatorial 2 x 9 (híbridos x aplicações de piraclostrobina), com 4 repetições. As aplicações foram realizadas em diferentes combinações de fungicidas: presença ou ausência de piraclostrobina + tiofanato metílico + fipronil (100 g i.a.100 kg semente-1) no tratamento de sementes (V0) combinado com a aplicação de piraclostrobina (150 g i.a.ha-1) antes ou após a adubação de cobertura (V4 ou V6) combinado ou não com a aplicação de piraclostrobina + ciproconazole (137,25 g i.a.ha-1) em pré-pendoamento (VT) e uma testemunha (sem uso de piraclostrobina). Foram avaliados: atividade da enzima redutase do nitrato, índice de clorofila, altura de plantas, altura de inserção de espiga, diâmetro de colmo, porcentagem de folhas senescentes, severidade de Puccinia polysora Underw, massa de mil grãos, densidade e produtividade de grãos. Submeteram-se os dados ao teste F para verificação de significância; e para comparação de médias utilizou-se o teste de Tukey, ambos a 5% de probabilidade. Para a maioria dos caracteres avaliados observou-se diferença entre híbridos. Não foi observado efeito das aplicações de piraclostrobina e interação entre os fatores em questão. Conclui-se que não há efeito benéfico ou deletério da aplicação de piraclostrobina em diferentes épocas e combinações de aplicação sobre os dois híbridos simples de milho cultivados na safra de verão.(AU)

An experiment was conducted in Jataí, Goiás, Brazil, aiming to evaluate the effects of the application of pyraclostrobin at different times and with different application combinations in two simple hybrids of corn grown in the summer season. A randomized block design was adopted in a factorial 2 x 9 (hybrid x pyraclostrobinapplications) design, with 4 replications. The applications were performed in different combinations: presence or absence of pyraclostrobin + thiophanate methyl + fipronil (100 g a.i.100 kg seed-1) in seed treatment (V0) combined with pyraclostrobin (150 g a.i.ha-1) application before or after topdressing (V4 or V6) combined or not with pyraclostrobin + cyproconazole (137.25 g a.i.ha-1) application in pre-bolting (VT) and a control (without use of pyraclostrobin). The following evaluations were performed: activity of the nitrate reductase enzyme, chlorophyll index, plant height, corn cob insertion height, stem diameter, percentage of senescent leaves, severity of Puccinia polysora Underw, thousand grain weight, grains density, and grains yield. An F test was performed to analyze the significance, and for a means comparison, a Tukey test was used, both at 5% probability. Differences among hybrids were observed for most of the evaluated traits. There were no effects of the applications of pyraclostrobin and no interaction between the factors in question. It can, therefore, be concluded that there are not beneficial or deleterious effects of the application of pyraclostrobin at different times and with different application combinations in two simple hybrids of corn grown in the summer season.(AU)

Evaluation of colorimetric nitrate reductase assay for rapid detection of methicillin resistance in clinical isolates of Staphylococcus aureus.

Background & objectives: Methicillin resistant Staphylococcus aureus (MRSA) remains a major cause of health care-associated infections. Rapid detection of MRSA facilitates the early initiation of appropriate treatment and infection control. Hence, the present study was undertaken to standardize and evaluate the performance of rapid colorimetric nitrate reductase assay (NRA) for determining methicillin resistance in S.aureus. Methods: A total of 160 clinical isolates of S. aureus, (80 each of methicillin susceptible and methicillin resistant) were included in the study. Minimum inhibitory concentration (MIC) was determined by NRA and reference broth micro dilution (BMD) methods. Results of NRA were compared with BMD and analyzed. Results: For MRSA, the MIC values ranged from 4 to ≥ 16 μg/ml and for MSSA, ≤ 0.5 to 2 μg/ml. Category and essential agreement for NRA as compared with BMD were found to be 99.4 and 89.7 per cent, respectively. No minor or major discrepancy was observed. A single resistant isolate showed very major discrepancy. Interpretation & conclusions: Colorimetric NRA being an inexpensive test requiring no special equipment can be employed as an alternative method for rapid detection of MRSA in resource limited settings.

Simple Method for Measuring Endothelial Nitric Oxide Synthase Activity in Clinical Researches.

Objectives: This study aimed to evaluate a new simple colorimetric method for the assay of endothelial nitric oxide synthase activity in clinical researches and to compare between the role of vanadium chloride and nitrate reductase enzyme in the assay method. Methods and materials: The new method involved using RBCs lysate to measure eNOS of the endothelium, using NADPH recycling system to enable the colorimetric measurement of eNOS, and using vanadium chloride to converting nitrate to nitrite. The method repeated again using nitrate reductase then vanadium results were compared with nitrate reductase results. The clinical study involved sixty patients who were randomly divided into two groups; group one received atorvastatin 40 mg daily, group two (control group) received placebo capsules. Patients were examined both before and after six weeks for endothelial nitric oxide synthase (eNOS) and for international index of erectile function-5 (IIEF-5) scores. Results: In comparison to nitrate reductase, vanadium showed no significant difference in eNOS activity indicating similar role and activity. However, In comparison to control group, atorvastatin showed a significant increase in eNOS (48.16% for vanadium, and 44.3% for nitrate reductase, P<0.05). Also, atorvastatin showed a significant increase in IIEF-5 score (39.16%, P<0.05) and there was a significant correlation between eNOS and IIEF-5 score (P<0.05). Conclusion: The study recommended this method for assaying eNOS using RBCs lysate, NADPH recycling system and vanadium chloride. This method is simple, reliable, and suitable for routine use in clinical researches.

Evaluation of four colourimetric susceptibility tests for the rapid detection of multidrug-resistant Mycobacterium tuberculosisisolates

The purpose of this study is to evaluate four rapid colourimetric methods, including the resazurin microtitre assay (REMA), malachite green decolourisation assay (MGDA), microplate nitrate reductase assay (MNRA) and crystal violet decolourisation assay (CVDA), for the rapid detection of multidrug-resistant (MDR) tuberculosis. Fifty Mycobacterium tuberculosisisolates were used in this study. Eighteen isolates were MDR, two isolates were only resistant to isoniazid (INH) and the remaining isolates were susceptible to both INH and rifampicin (RIF). INH and RIF were tested in 0.25 µg/mL and 0.5 µg/mL, respectively. The agar proportion method was used as a reference method. MNRA and REMA were performed with some modifications. MGDA and CVDA were performed as defined in the literature. The agreements of the MNRA for INH and RIF were 96% and 94%, respectively, while the agreement of the other assays for INH and RIF were 98%. In this study, while the specificities of the REMA, MGDA and CVDA were 100%, the specificity of the MNRA was lower than the others (93.3% for INH and 90.9% for RIF). In addition, while the sensitivity of the MNRA was 100%, the sensitivities of the others were lower than that of the MNRA (from 94.1-95%). The results were reported on the seventh-10th day of the incubation. All methods are reliable, easy to perform, inexpensive and easy to evaluate and do not require special equipment.

Utilidad del método de la nitrato reductasa para la detección de resistencia a drogas antituberculosas de segunda línea / Usefulness of the nitrate reductase method to detect resistance to second-line anti-tuberculosis drugs

Introducción: el creciente hallazgo de cepas de Mycobacterium tuberculosis multidrogorresistentes extremadamente resistentes ratifica la importancia de ofrecer, de forma rápida, los resultados de susceptibilidad de M. tuberculosis a drogas de primera y segunda línea como única alternativa para evitar la transmisión. Objetivo: comparar el método de la nitrato reductasa y el de las proporciones para la detección de susceptibilidad a drogas antituberculosas de segunda línea en aislamientos clínicos de M. tuberculosis, recuperados de pacientes cubanos con tuberculosis multidrogorresistente. Métodos: se investigó, mediante el método de las proporciones en Löwenstein-Jensen y el de la nitrato reductasa, la susceptibilidad a la ofloxacina, la kanamicina y a la capreomicina en 34 aislamientos de M. tuberculosis multidrogorresistentes. Resultados: en tres aislamientos se evidenció un comportamiento extremadamente resistente por ambos métodos. Mediante el método de la nitrato reductasa los resultados estuvieron disponibles entre 7 y 14 días. La sensibilidad fue de 100 por ciento, 90,0 por ciento y 77,8 por ciento para la ofloxacina, la kanamicina y la capreomicina, respectivamente, mientras que la especificidad fue superior al 95,0 por ciento y el valor de kappa fue superior a 0,85 para las tres drogas. Conclusión: de acuerdo con los resultados alcanzados, consideramos que el método de la nitrato reductasa constituye una valiosa alternativa para la detección oportuna de tuberculosis extremadamente resistente en países con limitados recursos económicos(AU)

Introduction: the increase of multidrug resistant and extensively drug resistant tuberculosis underlines the urgent need to obtain early results of Mycobacterium tuberculosis susceptibility both to first and second line antituberculosis drugs in order to avoid dissemination of resistant isolates. Objective: the aim of this research was to compare the performance of the nitrate reductase assay and the proportion method for to detect the susceptibility to second line antituberculosis drugs in multidrug resistant clinical isolates of M. tuberculosis. Methods: the susceptibility to ofloxacin, kamamycin and capreomycin of 34 M. tuberculosis multidrug resistant isolates was investigated using the proportion method in Löwenstein-Jensen and the nitrate reductase assay. Results: three isolates were identified as extensively drug resistant by both methods. The results of the nitrate reductase assay were obtained between 7-14 days achieving 100 percent, 90.0 percent and 77.8 percent of sensitivity for ofloxacin, kamamycin and capreomycin, respectively while specificity was higher than 95.0 percent and kappa value was higher to 0,85 for all drugs. Conclusion: the nitrate reductase assay represents a useful tool for the rapid identification of extensively drug resistant tuberculosis in low resources setting(AU)

Resposta do cafeeiro à inoculação de fungos micorrízicos arbusculares, em latossolo vermelho, do cerrado / Response of coffee plants colonized inoculation with arbuscular mucorrhizal fungi, red latosol, in cerrado

O objetivo deste trabalho foi verificar a resposta de dois cultivares de café (sensível e tolerante ao alumínio - Al), à inoculação de Gigaspora margarita e Glomus etunicatum, em Latossolo Vermelho do cerrado, com diferentes saturações por bases (30, 45 e 53 %). O experimento foi realizado em casa de vegetação, com delineamento inteiramente casualizado e em esquema fatorial 2x3x3, consistindo de 2 cultivares de (tolerante e sensível a Al), 3 tratamentos com micorriza (com inoculação de duas espécies de FMA e sem inoculação) e 3 níveis de saturação por bases do solo (V%), com cinco repetições por tratamento. As variáveis foram: altura da planta, diâmetro do caule, área foliar, massa da matéria seca da parte aérea, massa da matéria fresca de raiz, atividade da redutase do nitrato, teor de clorofila, colonização micorrízica e número de esporos. Os isolados de micorrizas proporcionaram maior crescimento do cafeeiro em solo ácido com alta concentração de Al, porém esta resposta foi verificada para ambos os cultivares quando colonizados por G. margarita. Os cultivares avaliados não mostraram diferenças quanto à tolerância ao Al quando não micorrizados.

The aim of this study was evaluate the response of two coffee cultivars (tolerant and sensitive to aluminum - Al), inoculated or not by two arbuscular mycorriza fungi (AMF), Gigaspora margarita and Glomus etunicatum, in cerrado Oxisol, with different base saturation. This experiment was conducted under greenhouse conditions, with a complete randomized design, in a 2x3x2 factorial scheme, consisting of 2 cultivars (tolerante and sensitive to Al), 3 treatments with mycorrhizal (inoculated with two species of AMF and without inoculation) and 3 levels of soil base saturation (30, 45 and 53 V%), with five replicates per treatment. The variables were: plant height, stem diameter, leaf area, shoot dry weight, root fresh weight, nitrate reductase activity, chlorophyll concentration, root colonization and number of AMF spores. Mycorrhizae isolates promoted greater response of coffee plants, in acid soil with high concentration of Al, but this response was observed for both cultivars when plants were colonized by G. margarita. The cultivars evaluated showed no differences in Al tolerance when non inoculated.

Determinação de nitrato redutase em isolados de Fusarium graminearum / Determination of nitrate reductase in strains of Fusarium graminearum

Os fungos filamentosos como o Fusarium graminearum são fungos que produzem expressiva quantidade de metabólitos secundários. A pesquisa da atividade da enzima nitrato redutase, enzima diretamente associada ao processo de desnitrificação, em fungos do gênero Fusarium tem sido tema controverso. O objetivo deste estudo foi avaliar a atividade da enzima nitrato redutase em cepas de Fusarium graminearum. As cepas de F. graminearum foram cultivadas na presença de nitrato e nitrito, apresentando níveis de nitrato redutase diferentes. Os resultados obtidos apontam para a utilização da via assimilatória de nitrato pelos organismos de F. graminearum avaliados neste estudo.

Fusarium graminearum are filamentous fungi that produce significant amounts of pigments. Studying the activity of the enzyme nitrate reductase, which is closely associated with the process of denitrification in Fusarium, has been controversial. The aim of the present study was to evaluate two strains of F. graminearum that have grown in the presence of nitrate and nitrite, showing different levels of nitrate reductase. The results point the use of nitrate assimilation pathway for the fungus F. graminearum evaluated in this study.

Detection of drug resistant Mycobacterium tuberculosis among patients with and without HIV infection in a rural setting / Detección de Mycobacterium tubersulosis resistente a los antibióticos entre pacientes con y sin infección por VIH en un área rural

OBJECTIVE: To analyse the sensitivity of Mycobacterium tuberculosis by nitrate reductase assay (NRA) and the Hain molecular line probe assay (LPA) in sputa of tuberculosis (TB)/HIV co-infected patients in Guyana. DESIGN: Sputum samples were collected from known TB patients at Georgetown Chest Clinic and were analysed at the Reference Laboratory, Guyana, over the period April 2010 to April 2011. RESULTS: Both methods recorded greater sensitivity for rifampin (RIF) than of isoniazid (INH). Both methods detected four RIF resistant, two INH resistant and two multi-drug resistant (MDR) strains and they had greater negative agreement indices than positive agreement indices. CONCLUSION: It was established that the sensitivity of Mycobacterium tuberculosis by the NRA and Hain LPA in TB/HIV co-infected patients has acceptable correlation and that HIV infection does not affect drug susceptibility testing.

OBJETIVO: Analizar la sensibilidad de Mycobacterium tuberculosis por medio del ensayo de nitrato reductasa (NRA) y el ensayo de sonda lineal (LPA) molecular de Hain en esputos de pacientes co-infectados TB/VIH en Guyana. DISEÑO: Muestras de esputo de pacientes de la Clínica del Tórax en Georgetown diagnosticados con tuberculosis, fueron analizadas en el Laboratorio de Referencias, en Guyana, en el período de abril de 2010 a abril de 2011. RESULTADOS: Ambos métodos registraron una mayor sensibilidad a la rifampicina (RIF) que a la isoniacida (INH). Ambos métodos detectaron cuatro cepas resistentes a RIF, dos resistentes a INH, y dos resistentes a mútiples medicamentos (RMM). Asimismo, presentaban mayores índices de concordancia negativa que de concordancia positiva. CONCLUSIÓN: Se estableció que la sensibilidad de Mycobacterium tuberculosis por medio del ensayo de NRA y el LPA de Hain en pacientes co-infectados TB/VIH, guarda una correlación aceptable, y que la infección por VIH no afecta la prueba de susceptibilidad a los medicamentos.

Detección de la mutación S315T en el gen katg como estrategia para identificación de Mycobacterium tuberculosis resistente a isoniacida en un laboratorio de referencia / Detection of S315T mutation in the katg gene as a strategy for identification of isoniazid-resistant Mycobacterium tuberculosis in a reference laboratory

Introduction: The emergence of strains of Mycobacterium tuberculosis resistant to drugs is a public health problem. Aim: To characterize the resistance to isoniazid (INH) in M. tuberculosis. Methods: Phenotypic and genotypic methods were used to determine the contribution of mutations at 315 codon of katG gene to the phe-notypic expression of resistance. The analysis of susceptibility to antibiotics was performed by the proportional method of Canetti and nitrate reductase method.Genotypic analysis of INH resistance was performed by PCR-RFLP. Results: 193 strains of M. tuberculosis from patients with respiratory symptoms were analyzed. Nineteen (9.8%) strains resistant to INH were identified, of which 12 (63.2%) showed resistance to other drugs. Genotypic analysis allowed to detect the mutation S315T in the katG gene in 15 of 17 strains phenotypically resistant to INH, showing a sensitivity of 88.24%, 100% specificity, 100% positive predictive value, 92% negative predictive value and high concordance with phenotypic methods (kappa = 0.85 (p < 0.01). Conclusion: The S315T mutation in the katG gene is the predominant mechanism of INH resistance in our circulating strains. This feature could be used as a rapid diagnostic tool with potential to detect at least 88% of isoniazid resistant strains, with great impact on the therapeutic management of patients.

Introducción: La emergencia de cepas de Mycobacterium tuberculosis multi-resistentes a antimicrobianos constituye un problema de salud pública. Objetivo: Caracterizar la resistencia a isoniacida (HIN) en cepas de M. tuberculosis. Métodos: Se aplicaron métodos fenotí-picos y genotípicos para determinar la contribución de mutaciones en el codón 315 del gen katG a la expresión fenotípica de resistencia. El análisis fenotípico de susceptibilidad a antimicrobianos se realizó por métodos de las proporciones de Canetti y nitrato reductasa. El análisis genotípico se realizó por RPC-PLFR. Resultados: Se analizaron 193 cepas de M. tuberculosis aisladas de pacientes sintomáticos respiratorios. Se identificaron 19 (9,8%) cepas resistentes a HIN, de las cuales, 12 (63,2%) exhibieron resistencia a otros antimicrobianos. El análisis genotípico permitió detectar la mutación katG S315T en 15/17 cepas fenotípicamente resistentes a HIN, exhibiendo una sensibilidad de 88,24%, especificidad de 100%, valor predictor positivo de 100%, valor predictor negativo de 92% y alta concordancia al comparar con los métodos fenotípicos (kappa = 0,85 (p < 0,01). Conclusión: La mutación S315T en el gen katG representa un mecanismo de desarrollo de resistencia a HIN predominante en las cepas circulantes en nuestro medio, característica que podría ser utilizada como herramienta diagnóstica rápida para detectar al menos 88% de las cepas resistentes a isoniacida, con gran impacto en el manejo terapéutico de los pacientes.

Nitrate reductase assay using sodium nitrate for rapid detection of multidrug resistant tuberculosis

We validated the nitrate reductase assay (NRA) for the detection of multidrug-resistant Mycobacterium tuberculosis (MDR-TB) using sodium nitrate (NaNO3) in replacement of potassium nitrate (KNO3) as nitrate source. NaNO3 is cheaper than KNO3 and has no restriction on use which facilitates the implementation of NRA to detect MDR-TB.

Comparative evaluation of the microplate nitrate reductase assay and the rezasurin microtitre assay for the rapid detection of multidrug resistant Mycobacterium tuberculosis clinical isolates

The microplate nitrate reductase assay (MNRA) and the rezasurin microtitre assay (REMA) were used for the susceptibility testing of 73 clinical isolates and the results were compared with those that were obtained using the Bactec 460 TB and Bactec MGIT 960 systems. The REMA and the MNRA were performed in 96-well plates. For the REMA, the concentrations of isoniazid (INH) and rifampicin (RIF) ranged from 1.0-0.01 µg/mL and 2.0-0.03 µg/mL, respectively. For the MNRA, the INH concentration was between 1.0-0.03 µg/mL and the RIF concentration was between 2.0-0.06 µg/mL. For the MNRA, the sensitivity, specificity, positive predictive value, negative predictive value and INH/RIF agreement were 100/95.6, 97.6/100, 96.8/100, 100/98 and 98.6/98.6, respectively, and for the REMA, they were 100/91.3, 90.4/100, 88.5/100, 100/96.1 and 94.5/97.2, respectively. Our data suggest that these two rapid, low-cost methods may be inexpensive, alternative assays for the rapid detection of multidrug resistant tuberculosis in low-income countries.

Chromium-induced alterations in photosynthesis and associated attributes in Indian mustard.

Contamination of soil and water by chromium (Cr) is increasing enormously due to anthropogenic activities. The potential of plants to accumulate or stabilize Cr compounds for the purpose of remediation of Cr contamination has been recognized in recent years. We conducted pot experiments to study photosynthesis and associated attributes in cv Pusa Jai Kisan of Indian mustard under natural as well as Cr-loaded environmental conditions. High doses of Cr caused toxic effects in plants, as evident by a reduction in photosynthetic rate (24.3 to 8.7 Cmol CO2 m-2 s-1 at 80 DAS), nitrate reductase activity (3.76 to 1.30 Cmol nitrite g-1 f. wt. h-1 at 80 DAS) and the contents of chlorophyll (1.49 to 0.86 mg g-1 f. wt. at 80 DAS) and soluble protein (2.96 to 1.93 mg g-1 f. wt. at 80 DAS). Since plants lack a specific Cr-transport system, mineral nutrient contents also changed due to Cr toxicity. Cr accumulation in different plant parts was affected by both duration and dose of Cr treatments, with a maximal localization of Cr in roots (up to 0.77 mg g-1 d. wt) at initial stages (40 DAS) and in stem (up to 4.19 mg g-1 d. wt) at the later stage (80 DAS) of plant growth. Thus, Indian mustard was able to withstand Cr stress and protect itself from Cr toxicity by altering various metabolic processes. Owing to its ability to accumulate large amounts of Cr, it may be useful in the process of land reclamation.

Effects of cadmium on growth and metabolism of Phaseolus mungo.

An attempt has been made to assess the response of Phaseolus mungo L. under influence of cadmium chloride (Cd Cl2) with special reference to growth, morphology, yield and biochemical aspects. Surface sterilized seeds of Phaseolus mungo L. cv. T-9 were exposed to various concentrations of Cd Cl2 solution (10-2 M, 10-4 M, 10-5 M, 10-8 M and control) for 12 hr at room temperature and these seeds were transferred to petriplates and polythene bags in triplicate. 10-2 M conconcentration was found to have deleterious effects on seed germination, germination relative index, length and dry weight of root and shoot, shoot root ratio and seedling vigour index, plant height, phytomass, number of leaves and branches, leaf area and chlorophyll contents while 10-8 M revealed slightly promotory effects. Phytotoxicity percentage and chlorophyll stability index were maximum in (10-2 M) concentration, while minimum in 10-8 M conconcentration of Cd Cl2. Nitrate and nitrite reductase activity was markedly inhibited at higher conconcentration. Low dose of Cd (10-8 M) did not affect soluble sugar contents of seeds but it induced a significant increase at higher conconcentration (10-2 M). It however, did not affect protein contents of seeds accept at higher concentration.

Comparative evaluation under routine conditions of the nitrate reduction assay, the proportion assay and the MGIT 960 assay for drug susceptibility testing of clinical isolates of Mycobacterium tuberculosis

The performance of the nitrate reductase assay (NRA) was compared with the proportion method (PM) on Lowenstein-Jensen medium and the BACTEC MGIT960 assay under routine conditions using 160 clinical isolates of Mycobacterium tuberculosis with a high proportion of resistant strains. The mean time to obtain results was 8.8 days and the overall agreements between NRA and PM and NRA and M960 were 95 percent and 94 percent, respectively. NRA was easy to perform and represents a useful tool for the rapid screening of drug-resistant M. tuberculosis strains in low-resource countries.

Zinc and copper induced changes in physiological characteristics of Vigna mungo (L.).

The effect of deleterious concentration of zinc and copper provided either individually or in combination in the nutrient media was investigated in order to assess the effect of metal interaction in Vigna mungo (L.). Both metals showed negative effect and led to a marked decrease in seed germination (20%), seedling growth (91.7%) and nitrate reductase activity (85.7%) with the increase in metal concentrations. The present study also emphasizes on the response of catalase and peroxidase enzyme under zinc and copper stress. Both antioxidant enzymes exhibited an increasing trend under different treatment conditions but it was reverse at highly toxic metal concentration. The results showed active involvement of peroxidase enzyme in regulating oxidative stress rather than catalase enzyme, as the specific activity of peroxidase enzyme got increased by 8.94% under the combined metals stress whereas catalase activity got declined by 60.97% in comparison to control due to excessive stress. The combined effect of copper and zinc metal was more pronounced in comparison to their individual effects.

A rapid detection of multidrug-resistant Mycobacterium tuberculosis by a nitrate reductase assay on blood agar

The susceptibility of 49 Mycobacterium tuberculosis clinical isolates to isoniazid (INH) and rifampisin (RIF) (28 multi-drug resistant-tuberculosis samples) was determined by a nitrate reductase assay (NRA) on blood agar. Agreement between the NRA and other testing methods was found to be 93.8 percent for both INH and RIF. The sensitivity, specificity, positive predictive value and negative predictive value for INH were 92.8 percent, 94.2 percent, 86.6 percent and 97 percent, respectively. The sensitivity, specificity, positive predictive value and negative predictive value for RIF were 90.4 percent, 96.4 percent, 95 percent and 93.1 percent. In conclusion, we show here that blood agar can be used effectively for the NRA test.

Evaluation of the Performance of Nitrate Reductase Assay for Rapid Drug-susceptibility Testing of Mycobacterium tuberculosis in North India.

The objective of the study was to evaluate the performance of nitrate reductase assay (NRA) as a rapid, reliable and inexpensive method for drug-susceptibility testing (DST) of Mycobacterium tuberculosis against firstline antitubercular drugs, such as rifampicin (RIF), isoniazid (INH), streptomycin (STR), and ethambutol (EMB). In total, 286 isolates were subjected to test by proportion method (PM) and NRA. By comparing the results of NRA with those of the gold standard PM, sensitivities and specificities were 98.4%, 97%, 88.5%, and 94.2% and 100%, 100%, 94%, and 99% for RIF, INH, STR, and EMB respectively. The positive predictive values were 100%, 100%, 95%, and 98% for RIF, INH, STR, and EMB respectively. The negative values were 99%, 98%, 87%, and 96% for RIF, INH, STR, and EMB respectively. The median time of obtaining results was shorter using NRA (10 days) compared to PM (28 days). An excellent agreement was observed between the two phenotypic tests with the κ values of 0.98, 0.97, 0.81, and 0.93 for RIF, INH, STR, and EMB respectively. The results demonstrated that NRA is suitable for the early determination of INH and RIF resistance and has the potential to be a useful tool for rapid drug-sensitivity test of M. tuberculosis in resource-constrained settings.