Trace Elements Contents in <Emphasis Type="Italic">Asparagus curillus</Emphasis> (Buch.-Ham.) ex Roxb.

Concentration level of nine elements viz. Zn, Cu, Mn, Fe, Co, Na, K, Ca, and Li were determined in leaves and roots of Asparagus curillus (Buch.-Ham.) ex Roxb. collected from four different altitudes in three seasons by atomic absorption spectroscopy. The overall concentration of K was found to be highest, whereas the level of Cu was lowest. The maximum concentrations of Zn, Cu, Mn, Fe, Co, Na, K, Ca, and Li were found to be 97.0 ± 1.5, 28.0 ± 7.0, 44.0 ± 7.3, 1138.0 ± 18.5, 91.0 ± 6.2, 381.0 ± 7.8, 9508.0 ± 7.8, 3076.0 ± 6.4, and 78.0 ± 4.6 mg/kg, respectively.     

Quantification of Parasite Load in Clinical Samples of Leishmaniasis Patients: IL-10 Level Correlates with Parasite Load in Visceral Leishmaniasis

A rapid and accurate method to detect and quantify Leishmania parasite is urgently needed to facilitate early diagnosis of Leishmaniasis and monitoring of antileishmania therapy. In this study, real-time assay was applied to estimate parasite load in clinical samples of visceral leishmaniasis (VL) and post kala-azar dermal leishmaniasis (PKDL) patients. The mean parasite load in blood of VL patients (n = 31) was 8,372 parasites/ml, while the mean parasite load in bone marrow aspirate (BMA) was 194,962 parasites/million nucleated cells (n = 12). Parasite load was undetectable after treatment with amphotericin B (n = 9) in VL, while a residual parasite burden was detected in 2 of 6 patients following treatment with sodium antimony gluconate. Further, circulating levels of IFN-γ, TNF-α, IL-10, IL-6, IL-4 and IL-2 were analysed in VL patients (n = 29) by Cytometric Bead Array to evaluate correlation with parasitic load. Interestingly, IL-10 levels correlated significantly with parasite load (r = 0.82, P<0.0001). The mean parasite load in dermal lesions of PKDL patients was 9,502 parasites/µg tissue DNA at pre-treatment stage (n = 25), with no detectable parasites after therapy (n = 5). Parasite burden was distinctly higher (P<0.0001) in nodular lesions (n = 12) (19,586 parasites/µg tissue DNA) compared to papular/macular lesions (n = 13, 193 parasites/µg tissue DNA). Further, chronic PKDL lesions showed significantly (P = 0.0166) higher parasite load in comparison with acute lesions. Results indicate that chronic, nodular cases constitute the major parasite reservoir for anthroponotic transmission. Our results establish that the high parasite load in VL is strongly correlated with a high level of IL-10, implicating IL-10 as a marker of disease severity. The assay is applicable for diagnosis as well as prognosis of both VL and PKDL, providing a simple molecular tool to monitor the efficacy of antileishmanial drugs or vaccines.     

Genetic Diversity Among Cold-Tolerant Fluorescent Pseudomonas Isolates from Indian Himalayas and Their Characterization for Biocontrol and Plant Growth-Promoting Activities

In Uttarakhand, the Organic State of India, where soils in most farming situations are deficient in nutrients and loss of crops due to soil- and seed-borne pathogens is rampant, use of native plant growth-promoting rhizobacteria (PGPRs) possessing biocontrol (BC) activities holds promise. In view of this, 600 native cold-tolerant rhizospheric bacterial isolates were collected from Uttarakhand Himalayas, of which 336 were confirmed as fluorescent Pseudomonas spp. On the basis of specific biochemical tests, these were characterized into three major groups: P. fluorescens (308 isolates), P. aeruginosa (20 isolates), and P. putida (8 isolates). Most of the isolates could grow at 8°C after 12 h of incubation, confirming their cold tolerance. In vitro biocontrol assays revealed that of 336 isolates, 74 were antagonistic to Rhizoctonia solani and 91 to Fusarium solani, the two major pathogens associated with root-rot complex in vegetables widespread in the region. Simultaneously, good HCN producers (33 isolates), siderophore producers (80 isolates), and P solubilizers (49 isolates) were also identified, which could increase the biocontrol and plant growth-promoting efficacies of the putative PGPRs. Among the different species and biovars, P. fluorescens biovar-I had the maximum number of potential isolates with BC and plant growth-promoting (PGP) activities. In French bean, under polyhouse and field conditions, five isolates (Pf-173, Pf-193, Pf-547, Pf-551, and Pf-572) showed good BC and PGP activities as up to 93% reduction in root rot was achieved. A combination of all five isolates was found to be best with respect to BC and PGP activities. In a set of 59 fluorescent Pseudomonas isolates, RAPD-PCR analysis, using three random oligodecamer primers, revealed high diversity and formed ten distinct clusters, corresponding to the host of origin (annual or perennial) or habitat (farming situations) of the isolates. The amount of diversity revealed in the set of fluorescent Pseudomonas isolates could represent enormous diversity that exists in the wild that could be exploited for improved BC and PGP activities of the PGPRs. For the first time, this study led to a large-scale characterization and repositioning of fluorescent pseudomonads from the Indian Himalayas.     

Studies on substituted benzo[h]quinazolines,benzo[g]indazoles,pyrazoles, 2,6-diarylpyridines as anti-tubercular agents

Various substituted 5,6-dihydro-8-methoxybenzo[h]quinazolin-2-amine, 1-(3-(4-alkoxyphenyl)-7-methoxy-3,3a,4,5-tetrahydro-2H benzo[g]indazol-2-yl)ethanone, pyrazole and 2,6-diarylpyridine derivatives have been synthesized in good yields by an efficient methodology. The synthesized compounds (4–23) were evaluated for their in vitro anti-tubercular activity against Mycobacterium tuberculosis H37Rv strain. Compounds 6a, 6c, 8a, 19a and 19e exhibited significant anti-tubercular activity at MIC values 50, 100, 50, 25 and 100 μM concentration. In vitro cytotoxicity data using THP-1 cells indicated that most active compound 19a is safe as its MIC value is much lower than the cytotoxic value.     

midD-encoded ‘rhizomimosinase’ from Rhizobium sp. strain TAL1145 is a C–N lyase that catabolizes L-mimosine into 3-hydroxy-4-pyridone, pyruvate and ammonia

Rhizobium sp. strain TAL1145 catabolizes mimosine, which is a toxic non-protein amino acid present in Leucaena leucocephala (leucaena). The objective of this investigation was to study the biochemical and catalytic properties of the enzyme encoded by midD, one of the TAL1145 genes involved in mimosine degradation. The midD-encoded enzyme, MidD, was expressed in Escherichia coli, purified and used for biochemical and catalytic studies using mimosine as the substrate. The reaction products in the enzyme assay were analyzed by HPLC and mass spectrometry. MidD has a molecular mass of ~45 kDa and its catalytic activity was found to be optimal at 37 °C and pH 8.5. The major product formed in the reaction had the same retention time as that of synthetic 3-hydroxy-4-pyridone (3H4P). It was confirmed to be 3H4P by MS/MS analysis of the HPLC-purified product. The K _m, V _max and K _cat of MidD were 1.27 × 10^?4 mol, 4.96 × 10^?5 mol s^?1 mg^?1, and 2,256.05 s^?1, respectively. Although MidD has sequence similarities with aminotransferases, it is not an aminotransferase because it does not require a keto acid as the co-substrate in the degradation reaction. It is a pyridoxal-5′-phosphate (PLP)-dependent enzyme and the addition of 50 μM hydroxylamine completely inhibited the reaction. However, the supplementation of the reaction with 0.1 μM PLP restored the catalytic activity of MidD in the reaction containing 50 μM hydroxylamine. The catalytic activity of MidD was found to be specific to mimosine, and the presence of its structural analogs including l-tyrosine, l-tryptophan and l-phenylalanine did not show any competitive inhibition. In addition to 3H4P, we also identified pyruvate and ammonia as other degradation products in equimolar quantities of the substrate used. The degradation of mimosine into a ring compound, 3H4P with the release of ammonia indicates that MidD of Rhizobium sp. strain TAL1145 is a C–N lyase.