Bioactive phytocompound mulberroside C and endophytes of Morus alba as potential inhibitors of HIV-1 replication: a mechanistic evaluation.

OBJECTIVES: Despite considerable advancement in antiretroviral therapy, development of safe, effective, and multi-targeted drugs for HIV still remains a big challenge. Endophytes are untouched and, hence, an important and novel sources in drug discovery endeavours. The present study was conducted to identify the anti-HIV compounds from Morus alba and endophytes isolated from it. METHODS: The extracts of isolated endophytes were screened using high-performance liquid chromatography (HPLC). Further, all samples were analysed for their cytotoxicity using a thiazolyl blue tetrazolium bromide assay. Subsequently, anti-HIV activity was performed using cell-based and cell-free assay. At the end, potential endophytes were identified using gene sequencing. RESULTS: A total of 27 endophytes were isolated from the eight stem bark samples of M. alba. Of the 27 endophytes, extracts of total of four endophytes showed a profile similar to the M. alba plant when analysed by HPLC. Further experimentation with extracts of these four endophytes, along with an extract of M. alba stem bark and its bioactive molecule, mulberroside C, revealed that all these six samples have good inhibitory potential for HIV. Among them, mulberroside C and two endophytic fungal extracts showed very potent anti-HIV activity. Subsequently, mechanistic studies at the molecular level showed that out of six test samples, three acted as protease inhibitors. Further, all four potential endophytes were identified using gene sequencing. CONCLUSIONS: The overall findings of these studies can help in the development of a novel anti-HIV candidate from mulberroside C, an extract of stem bark of M. alba and extracts of these endophytes. However, further validation and clinical studies are required to develop an anti-HIV drug.

Ecological health and water quality of village ponds in the subtropics limiting their use for water supply and groundwater recharge.

Ponds are a typical feature of many villages in the subtropics, and have been widely used as important sources of water for agriculture, aquaculture and groundwater recharge, as well as enhancing village resilience to floods and drought. Currently many village ponds are in a very poor state and in dire need of rejuvenation. This paper assesses the current water quality status and ecological health of twelve sub-tropical village ponds, situated in western Uttar Pradesh, India. This assessment is used to evaluate their wastewater treatment needs in relation to potential village uses of the water. Physico-chemical (Secchi depth, Total phosphorus and Total nitrogen) and biological (Phytoplankton chlorophyll-a) indicators highlight hypertrophic conditions in all the ponds. The study indicates that the status of village ponds requires significant investments in wastewater treatment to restore their use for many purposes, including aquaculture, although some may still be acceptable for irrigation purposes, as long as pathogenic bacteria are not abundant. We propose increased implementation of decentralised systems for wastewater treatment, such as septic tanks and constructed wetlands, to reduce the organic and nutrient loads entering village ponds and allow their use for a wider range of purposes.

Clinical evaluation of an in-house-developed real-time RT-PCR assay for serotyping of dengue virus.

In the present study, an in-house-developed real-time RT-PCR (rRT-PCR) for serotyping of dengue virus (DENV) was evaluated for its performance, using 612 clinical samples. Compared to the composite reference standard, the in-house-developed rRT-PCR had an overall sensitivity of 97.5% and a specificity of 100%. The assay had a sensitivity of 100%, 95.6%. 96.9% and 100% for detecting DENV-1, DENV-2, DENV-3 and DENV-4, respectively. We recommend periodic evaluation of real-time RT-PCR assays for detecting DENV serotypes with a large number of samples and the use of at least two assays that target different regions of DENV genomes.

Effects of gamma irradiation on chickpea seeds vis-a-vis total seed storage proteins, antioxidant activity and protein profiling.

The present work describes radiation—induced effects on seed composition vis—à—vis total seed proteins, antioxidant levels and protein profiling employing two dimensional gel electrophoresis (2D—GE) in kabuli and desi chickpea varities. Seeds were exposed to the radiation doses of 1,2,3,4 and 5 kGy. The total protein concentrations decreased and antioxidant levels were increased with increasing dose compared to control seed samples. Radiation induced effects were dose dependent to these seed parameters while it showed tolerance to 1 kGy dose. Increase in the dose was complimented with increase in antioxidant levels, like 5 kGy enhanced % scavenging activities in all the seed extracts. Precisely, the investigations reflected that the dose range from 2 to 5 kGy was effective for total seed storage proteins, as depicted quantitatively and qualitative 2D—GE means enhance antioxidant activities in vitro.

DNA polymorphisms in chickpea accessions as revealed by PCR-based markers.

Chickpea is a food legume which is alleged to be a preferred source of protein next only to milk. Germplasm of cultivated chickpea available is deficient in desired genetic variation. Genetic manipulations therefore, necessitate the genetic exploitation of its related annual and wild species. 42 RAPD and 41 ISSR markers were employed to ascertain polymorphism across 20 genotypes which were collected from 10 different geographical areas of the world. RAPD marker detected 51% genetic polymorphisms while ISSR marker detected 54 %. With an average of 6.5 each RAPD primer amplified 5—8 bands. Similarly with an average of 7.9 each ISSR primer amplified 4—12 bands. The cluster dendrogram demonstrated a similarity coefficient range from 0.80 to 0.92 due to RAPD markers, whereas with ISSR primers the cluster dendrogram showed similarity coefficient of 0.60 to 1.00. Accessions from same geographical area seem to be genetically similar than those from geographically distant and isolated ones. When however compared, interestingly the ISSR dendrogram showed more correlation with pedigree data than the RAPD dendrogram. The variability index worked out in the present study ranges from 0.79 to 0.96. Since the ultimate reason for such studies is selection of diverse genetic accessions for their recommendation to breeding programmers, the accessions like ICC6263, ICC6306 and ICC17160 can be recommended as parents. Further breeding programmes can therefore be planned to procure additional variation complexes in chickpea genetic stocks.

Purification and dose-dependent toxicity study of abrin in swiss albino male mice.

Abrin, a phytotoxin obtained from the seeds of the Abrus precatorius plant, is highly toxic with an estimated human fatal dose of 0.1­1 µg/kg. In this study, abrin was purified and characterized through SDS PAGE and mass spectrometry analysis; further study on toxicity was carried out to investigate the alteration in biochemical, and hematological variables through histopathological observations in mice. The intraperitoneal LD50 value of purified abrin for mice was found to be 0.91µg/kg of body weight. Mice were exposed to 0.4 and 1.0 LD50 abrin doses intraperitoneally and observed on days 1, 3, and 7. Plasma GOT and GPT levels increased significantly at both doses. At 1.0 LD50 dose, alkaline phosphatase, bilirubin, urea, uric acid, and creatinine levels increased, whereas albumin, total protein, glucose and cholesterol levels decreased significantly. Abrin intoxication also altered the hemoglobin, WBC, and RBC counts significantly at 1.0 LD50 dose. Liver GSH levels decreased while lipid peroxidation increased significantly in a dose­dependent manner. Biochemical changes were supported by the histological investigation, which also showed the degenerative changes in organs. In conclusion, abrin intoxication caused toxic effects and severe damages on studied organs mediated through alteration in biochemical and hematological variables, lipid peroxidation, and degeneration.

Inhibition of P. falciparum PFATP6 by curcumin and its derivatives: a bioinformatic study.

Curcumin, a yellow spice has been shown to have many pathological uses including cancer and malaria. Recent experimental data have shown the inhibitory effect of curcumin and its two derivatives on the growth of Plasmodium falciparum in cell culture at low micromolar concentrations. Previous studies have suggested that Ca(2+)­ATPase (PfATP6) of P. falciparum is the target of many antimalarial drugs. However, the mechanism of inhibition of Ca(2+)­ATPase (PfATP6) is not known. In addition, it is not clear which specific isomeric form of curcumin is the most potent inhibitor of P. falciparum. Here we address this issue using bioinformatics tools. We generated a molecular model of Ca(2+)­ATPase (PfATP6) of P. falciparum and carried out molecular docking of all curcumin analogues of Zinc database of compounds (zinc.docking.org). Two molecular docking programs Glide and FlexX were used to determine binding feasibility of 351 analogues of curcumin. The comparison of docking parameters showed, more than 20 analogues are better ligands of PfATP6 than curcumin itself. . The binding of curcumin and its analogues to PFATP6 is mediated by both hydrophobic and polar interactions. Our results suggest that curcumin analogues are promising lead compounds for the development of antimalarial drugs.

Analysis of the PB2 gene reveals that Indian H5N1 influenza virus belongs to a mixed-migratory bird sub-lineage possessing the amino acid lysine at position 627 of the PB2 protein.

Outbreaks of highly pathogenic avian influenza (HPAI) H5N1 virus were reported for the first time in India during February 2006. Herein, we have sequenced and analyzed the PB2 genes of five influenza virus isolates obtained from three affected states (Gujarat, Madhya Pradesh and Maharashtra) in India during the outbreaks. In the phylogenetic analysis, the Indian isolates were grouped in the mixed-migratory bird sub-lineage of the Eurasian lineage. From the phylogenetic tree, it is evident that viruses were probably introduced to India from China via Europe because they share a direct ancestral relationship with the Indian isolates. The virus might have spread through migratory waterfowls that survived the HPAI H5N1 infection. These viruses were able to replicate in cultured cells of avian and mammalian hosts and posses lysine at position 627 of the PB2 protein, indicating that they might be able to cross the host barrier to infect mammals.

Anti-ulcer activity of Adhatoda vasica Nees.

Adhatoda vasica Nees (Acantheceae), commonly known as Vasaka, is a well-known plant in indigenous systems of medicine and is used for its beneficial effects, particularly in bronchitis. The present investigation was carried out to study the anti-ulcer activity of Adhatoda vasica leaves using two ulcer models (1) Ethanol-induced, and (2) Pylorus ligation plus aspirin-induced models. Adhatoda vasica leaf powder showeda considerable degree of anti-ulcer activity in experimental rats when compared with a control. The highest degree of activity (80%) was observed in the ethanol-induced ulceration model. Results of the study suggest that in addition to its classically established pharmacological activities, the plant also has immense potential as an anti-ulcer agent of great therapeutic relevance.

HPTLC method for the estimation of alkaloids of Cinchona officinalis stem bark and its marketed formulations.

We report a sensitive method for the estimation of quinine (Qn), cinchonine (Cn), and cinchonidine (Cnd) and a new method based on fluorescence enhancement and detection and quantification of quinidine (Qnd) from Cinchona stem bark and its formulations, using HPTLC. Standard solutions of Qn, Qnd, Cn, and Cnd were applied on precoated HPTLC plates and developed with chloroform/diethylamine (9.6:1.4 v/v). The plates were scanned and quantified at 226 nm for Qn, Cn, Cnd and for Qnd at 366 nm in fluorescence and reflectance mode ([symbol: see text] K400 filter). The method was validated for precision, accuracy and repeatability. Further, the stem bark of Cinchona officinalis and some herbal and homeopathic formulations were evaluated for their individual alkaloid content applying the developed method.

A rapid method for isolation of andrographolide from andrographis paniculata nees (kalmegh).

A simple and rapid method for isolation of andrographolide from the leaves of Andrographis paniculata is reported. This involves extraction of the leaf powder by cold maceration in a 1:1 mixture of dichloromethane and methanol and isolation of andrographolide directly from the resulting extract by recrystallisation. The identity of the compound was confirmed through IR, UV, mass and melting point, and co-chromatography with a reference standard on TLC. The purity of the compound was confirmed by TLC, UV absorption spectrum, HPLC and differential scanning calorimetry, the latter of which gave the melting point of andrographolide as 235.3°C.

Sensitive high-performance thin-layer chromatographic method for the estimation of diospyrin, a tumour inhibitory agent from the stem bark of Diospyros montana Roxb.

Diospyrin, a tumour inhibitory agent from the stem bark of Diospyros montana was isolated and characterised. A sensitive high-performance thin-layer chromatographic (HPTLC) method was developed for the estimation of diospyrin. The method was validated for precision (intra- and inter-day), repeatability and accuracy. The method was found to be precise, with the RSDs for intra-day in the range of 0.72-1.85% and RSDs for inter-day in the range of 1.06-2.95%, for different concentrations. Instrumental precision and repeatability of the method were found to be 0.086 and 0.937 (% CV), respectively. Accuracy of the method was checked by performing the recovery study at two levels and average percentage recovery was found to be 97.87%. The developed HPTLC method was adopted for the estimation of diospyrin content of the stem bark of D. montana from different regions, which varied from 0.35 to 0.47% (w/w) in the samples.