Utilization of primary and secondary biochemical compounds in cotton as diagnostic markers for measuring resistance to cotton leaf curl virus.

Introduction: Cotton (Gossypium hirsutum L.) is one of the most important staple fibrous crops cultivated in India and globally. However, its production and quality are greatly hampered by cotton leaf curl disease (CLCuD) caused by cotton leaf curl virus (CLCuV). Therefore, the aim of the present study was to investigate the biochemical mechanisms associated with CLCuD resistance in contrasting cotton genotypes. Methods: Four commercial cotton varieties with susceptible (HS 6 and RCH-134 BG-II) and resistant (HS 1236 and Bunty) responses were used to analyze the role of primary (sugar, protein, and chlorophyll) and secondary (gossypol, phenol, and tannin) biochemical compounds produced by the plants against infection by CLCuV. The resistant cultivars with increased activity of protein, phenol, and tannin exhibited biochemical barriers against CLCuV infection, imparting resistance in cotton cultivars. Results: Reducing sugar in the healthy plants of the susceptible Bt cultivar RCH 134 BG-II exhibited the highest value of 1.67 mg/g at 90 days. In contrast, the lowest value of 0.07 mg g-1 was observed at 60 DAS in the highly diseased plants of the susceptible hybrid HS 6. Higher phenol content (0.70 mg g-1) was observed at 90 DAS in resistant cultivars, whereas highly susceptible plants exhibited the least phenol (0.25 mg g-1) at 90 DAS. The lowest protein activity was observed at 120 DAS in susceptible cultivars HS 6 (9.4 mg g-1) followed by RCH 134 BG-II (10.5 mg g-1). However, other biochemical compounds, including chlorophyll, sugar, and gossypol, did not show a significant role in resistance against CLCuV. The disease progression analysis in susceptible cultivars revealed non-significant differences between the two susceptible varieties. Discussion: Nevertheless, these compounds are virtually associated with the basic physiological and metabolic mechanisms of cotton plants. Among the primary biochemical compounds, only protein activity was proposed as the first line of defense in cotton against CLCuV. The secondary level of defense line in resistance showed the activity of secondary biochemical compounds phenol and tannins, which displayed a significant increase in their levels while imparting resistance against CLCuV in cotton.

Synthesis of sulpha drug based hydroxytriazene derivatives: Anti-diabetic, antioxidant, anti-inflammatory activity and their molecular docking studies.

Herein, we report synthesis, characterization, anti-diabetic, anti-inflammatory and anti-oxidant activities of hydroxytriazenes derived from sulpha drugs, namely sulphanilamide, sulphadiazine, sulphapyridine and sulphamethazine. Before biological screening of the compounds, theoretical prediction using PASS was done which indicates probable activities ranging from Pa (probable activity) values 65-98% for anti-inflammatory activity. As per the predication, experimental validation of some of the predicted activities particularly anti-diabetic, anti-inflammatory and anti-oxidant was done. Anti-diabetic activities have been screened using two methods namely α-amylase and α-glucosidase inhibition method and IC50 values were ranging from 66 to 260 and 148 to 401 µg/mL, while for standard drug acarbose the values were 12 µg/mL and 70 µg/mL, respectively. Docking studies have also been done for antidiabetic target pancreatic alpha amylase. The molecular docking studies in α-amylase enzyme reveal that the middle phenyl ring of all the compounds mainly occupies in the small hydrophobic pocket formed by the Ala198, Trp58, Leu162, Leu165 and Ile235 residues and sulphonamide moiety establish H-bond interaction by two water molecules. Further, anti-inflammatory activity has been evaluated using carrageenan induced paw-edema method and results indicate excellent anti-inflammatory activity by hydroxytriazenes (71 to 97%) and standard drug diclofenac 94% after 4 h of treatment. Moreover, antioxidant effect of the compounds was tested using DPPH and ABTS methods. All the compounds displayed good results (24-488 µg/mL) against ABTS radical and many compounds are more active than ascorbic acid (69 µg/mL) while all other compounds showed moderate activity against DPPH radical (292-774 µg/mL) and ascorbic acid (29 µg/mL). Thus, the studies reveal potential of sulfa drug based hydroxytriazenes as candidates for antidiabetic, anti-inflammatory and antioxidant activities which have been experimentally validated.

How safe is the use of chlorpyrifos: Revelations through its effect on layer birds.

AIM: The present study was aimed to investigate the immunological competence of chlorpyrifos (CPF) insecticide after oral administration in layer chickens. MATERIALS AND METHODS: A total of 20 White Leghorn birds were given CPF in drinking water at 0.3 ppm/bird/day (no observable effect level dose) for a period of 3-month. Immune competence status of layer birds and chicks hatched from CPF-treated birds were estimated at 15 days interval in layer birds and monthly interval in chicks using immunological and biochemical parameters. RESULTS: There was a significant decrease in values of total leukocytes count, absolute lymphocyte count, absolute heterophil count, total serum protein, serum albumin, serum globulin, and serum gamma globulin in the birds treated with CPF as compared to control. Similarly, immune competence tests such as lymphocyte stimulation test, oxidative burst assay, and enzyme-linked immunosorbent assay tests indicated lower immunity in birds treated with CPF as compared to control. Subsequently, chicks produced from CPF-treated birds were also examined for immune competence, but no significant difference was observed between chicks of both the groups. CONCLUSION: The exposure to CPF produced hemo-biochemical and other changes that could be correlated with changes in the immunological profile of layer chickens suggesting total stoppage of using CPF in poultry sheds.

Effect of endosulfan on immunological competence of layer birds.

AIM: The present study was aimed to investigate the immunological competence of endosulfan insecticide after limited oral administration in White Leghorn layer chickens. MATERIALS AND METHODS: A total of 20 White Leghorn birds were given endosulfan in drinking water at 30 ppm/bird/day (no observable effect level dose) for a period of 3-months. Immune competence status of layer birds and chicks hatched from endosulfan offered birds were estimated at 15-day interval in layer birds and at monthly interval in chicks using immunological, biochemical parameters, and teratological estimates. RESULTS: There was a significant decrease in levels of total leukocytes count, absolute lymphocyte count, absolute heterophil count, total serum protein, serum albumin, serum globulin, and serum gamma globulin in the birds fed with endosulfan as compared to control. Similarly, immune competence tests such as lymphocyte stimulation test, oxidative burst assay, and enzyme-linked immunosorbent assay tests indicated lower immunity in birds treated with endosulfan as compared to control. Subsequently, chicks produced from endosulfan-treated birds were also examined for immune competence, but no significant difference was observed between chicks of both the groups. CONCLUSION: The exposure to endosulfan in limited oral dosage was able to exhibit hemo-biochemical and other changes that could be correlated with changes in the immunological profile of layer chickens suggesting cautious usage of endosulfan insecticide in poultry sheds.

Immunostimulatory and growth promoting potential of Tinospora cordifolia (Thunb.) Miers on fingerlings of Amur Carp.

The enhancement of immunity of fish through application of natural immunostimulants would help in minimizing the risk of occurrence of diseases in aquaculture. The present study was designed to evaluate the growth promoting and immunostimulatory effect of Guduchi, Tinospora cordifolia (Thunb.) Miers ex Hook F. leaf powder in fingerlings of Amur carp (Cyprinus carpio haeYnatopterus Martens). The fingerlings were divided into four treatment groups, Ti, T2, T3 and T4. Experimental diets were prepared by mixing rice bran, deoiled mustard cake, soybean meal and vitamin mineral mixture. The leaf powder of T. cordifolia was incorporated into diets D2, D3, and D4 @ 0.25%, 0.5% and 0.75%, respectively. In control diet D1, leaf powder of T. cordifolia was not incorporated. T1 group fishes were fed with D, diet, T2 with D2, T3 with D3 and T4 with D4 @ 5% body weight per day for 90 days. Fingerlings fed with diet D4 achieved significantly improved Specific Growth Rate (1), Feed Conversion Ratio (2.76), and Gross Conversion Efficiency (0.361) as compared .to the control and all other treatments (P <0.05). Hematological and biochemical parameters, total leuckocyte count (53.849x103/[L), total erythrocyte count (3.50x106/iL), hemoglobin concentration (17.17%), total serum protein, albumin and globulin (12.35, 1.56 and 10.78 g/dL) in D4 diet fed fishes showed increase as compared to the control and all other treatments (P <0.05). The above research revealed that Tinospora cordifolia leaf powder has significant growth promoting and immunostimulatory potential in Amur carp raising.

Can mycobacterial genomics generate novel targets as speed-breakers against the race for drug resistance.

The rise of multi-drug resistant and extensively drug resistant M. tuberculosis around the world poses a great threat to human health, and necessitates development of new, effective and inexpensive anti-tubercular agents. The availability of knowledge on molecular biology of M. tuberculosis infection coupled with whole genome sequences, transcriptomic, proteomic and metabolomic data sets have provided insights on the genes/proteins indispensable for initiation and maintenance of persistence, cross-talk with and/or sensing the host immune response, and finally the reactivation of persistent M. tuberculosis to a growing state. The review will focus on analysis of current state of M. tuberculosis genomic resources, host-pathogen interaction studies in the context of pathogen persistence, and the efforts made or required in the development and utilization of computational tools, models and metabolic network analyses to speed up the process of drug target discovery, particularly eradicating the dormant infections.

Investigation of an outbreak of infectious pustular balanoposthitis in cattle breeding bulls at a frozen semen bank.

Infectious pustular balanoposthitis (IPB) is one of the reproductive disorders caused by bovine herpesvirus 1 (BoHV1) that can be transmitted through artificial insemination. A herd of 63 breeding bulls at a frozen semen bank in Odisha state in India experienced a suspected outbreak of IPB, with 11 bulls showing clinical signs of the infection. Clinical signs were noticed in two bulls initially and a few days after in the other nine animals. Serum samples from 53 bulls were examined for anti-BoHV1 antibodies using a virus neutralisation test (VNT) and a competitive enzyme-linked immunosorbent assay (cELISA); the remaining ten bulls were not included in the study because it was difficult to restrain them at that time. Paired serum samples were collected 21 days apart from ten clinically affected bulls (the eleventh clinically affected bull was not included in the study for the reason stated above). In the neutralisation test, the paired serum samples showed a two- to fourfold increase in anti-BoHV1 antibody titre; in the cELISA, the paired samples were also found positive for anti-BoHV1 antibodies. Serum samples from 43 in-contact bulls were collected about day 22 after the first observation of clinical infection in the herd. Among these serum samples, a total of 30 were found positive for anti-BoHV1 antibodies in the VNT and a total of 30 were found positive in cELISA. Ten samples were positive in one test but not the other and 25 tested positive in both tests. In all, 35 serum samples from in-contact bulls tested positive in either one or both of the two types of test. An overall agreement of 76.74% was found in detection of anti-BoHV1 antibodies in the two tests. Sensitivity was higher than specificity in detection of anti-BoHV1 antibodies in the serum samples. The glycoprotein C region of the genomic DNA of BoHV1 was amplified from semen samples by polymerase chain reaction. The findings from the outbreak indicate that continuous monitoring of breeding bulls at frozen semen banks is warranted to avoid the risks associated with artificial insemination.

Antioxidant capacities, phenolic contents, and GC/MS analysis of Rhodiola imbricata Edgew. root extracts from Trans-Himalaya.

Our aim was to assess the antioxidant capacities and phenolic constituents of methanol and aqueous extracts of Rhodiola imbricata Edgew. root from Trans-Himalayan cold desert of Ladakh. The 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) radical scavenging capacity of the root extracts increased in a dose-dependent manner (up to 0.1 mg/mL) and root extract concentrations required for 50% inhibition of radical scavenging effect (IC50 ) were recorded as 0.013 and 0.014 mg/mL (for DPPH) and 0.016 and 0.017 mg/mL (for ABTS) for methanol and aqueous extracts, respectively. The total antioxidant power of the extract was determined by ferric reducing antioxidant power (FRAP) assay. Total polyphenol and phenolic acid content of methanol and aqueous extracts were 112.24, 59.06, 39.02, and 16.95 mg gallic acid equivalent/g of extract, respectively. Total flavonoid and flavonol contents were estimated to be 30.2, 17.67, 20.68, and 7.38 mg quercetin equivalent/g of extract, respectively. In all antioxidant capacity assays, the methanol extract exhibited significantly higher antioxidant capacity than that of aqueous extract due to the presence of significantly higher amount of vital phytoconstitiuents, viz. polyphenol, phenolic acid, and flavonol. GC/MS analysis showed that phytosterols, alkyl halide, phenols, and fatty acid esters were major phytochemical clusters. On the other hand, monoterpenes, fatty acids, tocopherols, aliphatic hydrocarbons, and ethers were found to be present in comparatively less amount in the methanol extract. Hence, our study signifies that this high-altitude medicinal herb could be used as the natural source of antioxidants and supports its use in traditional system of medicine to ameliorate oxidative stress and high-altitude maladies.

Development and validation of simultaneous spectrophotometric methods for drotaverine hydrochloride and aceclofenac from tablet dosage form.

Two simple spectrophotometric methods have been developed for simultaneous estimation of drotaverine hydrochloride and aceclofenac from tablet dosage form. Method I is a simultaneous equation method (Vierodt's method), wavelengths selected are 306.5 and 276 nm. Method II is the absorbance ratio method (Q-Analysis), which employs 298.5 nm as λ(1) and 276 nm as λ(2) (λmax of AF) for formation of equations. Both the methods were found to be linear between the range of 8-32 µg/ml for drotaverine and 10-40 µg/ml for aceclofenac. The accuracy and precision were determined and found to comply with ICH guidelines. Both the methods showed good reproducibility and recovery with % RSD in the desired range. The methods were found to be rapid, specific, precise and accurate and can be successfully applied for the routine analysis of drotaverine and aceclofenac in their combined tablet dosage form.

Pox outbreaks in sheep and goats at Makhdoom (Uttar Pradesh), India: evidence of sheeppox virus infection in goats.

Sheeppox and goatpox outbreaks occur often in India incurring huge economic loss to the small ruminant industry. This paper describes two sheeppox outbreaks, of which one occurred in an organized sheep breeding farm at Makhdoom (Uttar Pradesh), India, during 2007 and another in goats at the Central Institute of Research on Goats, Makhdoom (Uttar Pradesh), India during 2008. In the first outbreak, a local Muzaffarnagari sheep breed was affected (n=477) with morbidity and mortality rates, respectively, of 100% and 53.9% accompanied by significant productivity losses. In the 2008 outbreaks, a small number of goats were affected without any mortality. The tissue and swabs collected from both the outbreaks were processed and inoculated onto Vero cells, and the causative agent of the outbreaks, capripox virus (CaPV), was isolated. The identity of the virus was confirmed as CaPV based on electron microscopy, experimental pathogenesis in sheep, capripox-specific conventional and real-time PCRs. Sequence analysis of the P32 envelope protein gene revealed that the causative agent of both outbreaks was confirmed as sheeppox virus (SPPV) implying SPPV infection not only in sheep but also goats in India.

Isolation and characterization of Indian isolates of camel pox virus.

In this study, we isolated and identified three camel pox viruses (CMLV) from two outbreaks of camel pox infection in camels associated with eruptions on cheeks, nostrils, limbs, scrotum, and sheath that occurred at different places of Bikaner district, Rajasthan (India). The scab specimens collected were subjected for virus isolation in Vero cell culture, and the isolated viruses were characterized by employing polymerase chain reaction (PCR) and sequencing. The causative agent was identified as CMLV, based on A-type inclusion, B5R and C18L genes-specific PCRs and partial sequencing of these genes, which clearly confirmed that the outbreaks were caused by CMLV and identity of CMLV isolates. Further, phylogenetic analysis of partial C18L gene sequences have showed that Indian CMLV are clustered together with other reported isolates/strains.

A method for content uniformity determination of atenolol and losartan potassium in combined tablet dosage form.

A simple, accurate, rapid, specific and reproducible UV spectrophotometric method was developed for estimation of content uniformity of atenolol and losartan potassium in its combined tablet dosage form. The method involves formation and solving the simultaneous equation using 226.4 and 254 nm as two wavelengths for atenolol and losartan, respectively. Developed method was employed to determine the atenolol and losartan content in ten individual tablet units of five market formulations. Methanol was used as solvent. The method was validated. From the results, it was concluded that all brands are within the content uniformity limit, 85-115%.

Occurrence of canine parvovirus type 2c in the dogs with haemorrhagic enteritis in India.

Canine parvovirus 2 (CPV-2) causes a highly contagious and often fatal disease in dogs. Since its sudden emergence in the early 1970s, CPV-2 has been evolving through the generation of novel genetic and antigenic variants (CPV-2a/b/c) that are unevenly distributed throughout the world. In the present study we have examined 36 clinical cases of dogs suspected of CPV collected during year 2006. A fragment of the VP2 gene of the virus was analyzed using polymerase chain reaction (PCR), restriction endonuclease (RE) and DNA sequence analysis. Out of the 36 samples analyzed, 16 were found positive for CPV-2a/2b by conventional PCR. DNA sequencing was done for 6 PCR positive samples, out of which three were characterized as CPV-2c, indicating that this CPV type 2c is currently circulating in India.

Venous air embolism during microelectrode recording in deep brain stimulation surgery in an awake supine patient.

Venous air embolism (VAE) is a potentially serious complication in neurosurgery. It occurs most commonly during craniotomy in the sitting position. Diagnosis is difficult in awake patients on spontaneous ventilation due to different clinical manifestations. Prompt diagnosis and treatment are essential. We report venous air embolism in an awake patient undergoing deep brain stimulation for Parkinson's disease and discuss the pathogenesis, prevention and management of VAE.

Time course of Newcastle disease virus-induced apoptotic pathways.

Newcastle disease virus (NDV) causes economically significant Newcastle disease (ND) in almost all birds worldwide. Previous studies have shown that NDV induces caspase dependent apoptotic pathways in infected cells. In the present study, time course induction of apoptotic pathways in Vero cells is described. In NDV-infected cells, caspase-8 activity, percentage of cells showing TRAIL expression was higher at 24h p.i. (post-infection) compared to 48 h p.i. In contrast, caspase-9 activity, efflux of cytochrome c, loss of mitochondrial membrane potential was higher at 48 h compared to 24h p.i. The caspase-3 activity was high both times. Based on these results, it was concluded that at 24h p.i., NDV induces apoptosis through extrinsic apoptotic pathway while at 48 h p.i. predominantly through intrinsic apoptotic pathway.

Bovine herpes virus infections in cattle.

Bovine herpes virus 1 (BHV-1) is primarily associated with clinical syndromes such as rhinotracheitis, pustular vulvovaginitis and balanoposthitis, abortion, infertility, conjunctivitis and encephalitis in bovine species. The main sources of infection are the nasal exudates and the respiratory droplets, genital secretions, semen, fetal fluids and tissues. The BHV-1 virus can become latent following a primary infection with a field isolate or vaccination with an attenuated strain. The viral genomic DNA has been demonstrated in the sensory ganglia of the trigeminal nerve in infectious bovine rhinotracheitis (IBR) and in sacral spinal ganglia in pustular vulvovaginitis and balanoposthitis cases. BHV-1 infections can be diagnosed by detection of virus or virus components and antibody by serological tests or by detection of genomic DNA by polymerase chain reaction (PCR), nucleic acid hybridization and sequencing. Inactivated vaccines and modified live virus vaccines are used for prevention of BHV-1 infections in cattle; subunit vaccines and marker vaccines are under investigation.

Newcastle disease virus-induced cytopathic effect in infected cells is caused by apoptosis.

The velogenic Newcastle disease virus (NDV) causes highly infectious and economically significant Newcastle disease (ND) in birds of various species. In cell culture NDV induces cytopathic effect (CPE) characterized by rounding, vacuolation, syncytia formation and cell death. Aside from cell to cell fusion caused by the F and HN glycoprotein of the virus molecular events leading to cell death are not known. In the current study, NDV-infected Vero cells, at 48 h p.i., showed nuclear condensation, cytoplasm blebbing, DNA fragmentation, and phosphatidylserine translocation to the cell surface. In addition, virus-infected cells demonstrated decreased DNA content and an increased Bax to Bcl-2 ratio, p53 level and caspase 3, 8, 9 expression compared to mock-infected cells. Based on these results, it was concluded that CPE in NDV-infected cells was caused by to the induction of apoptosis with the involvement of p53 and the Bax, dependent apoptotic pathways.

Newcastle disease virus as an oncolytic agent.

Cancer is a major cause of deaths in humans. Though there has been significant progress in cancer therapy, the limited efficacy and toxicities of current chemo- and radiotherapies have provided an impetus for the search of new therapeutics. A therapeutic approach, which uses viruses for the treatment of cancer termed, oncolytic virotherapy has recently emerged. Newcastle disease virus (NDV) is one such virus with an inherent oncolytic property. NDV causes a highly infectious disease in poultry worldwide. In humans it is reported to have oncolytic and immuno-stimulatory effects. It specifically replicates in tumour cells while sparing normal cells and cause oncolysis. For many years different strains of the NDV have been investigated for treatment of various human cancers. Recent advances in reverse genetics provided investigators the tools to produce recombinant NDV with improved oncolytic property.

Rotavirus diarrhea in bovines and other domestic animals.

Rotavirus diarrhea is the major cause of death of millions of children in developing countries besides causing economically significant malady in neonates of many domestic animals. In neonates, the infection is non-viremic, have very short incubation period, and manifests profuse diarrhea and severe dehydration. Concurrent infection with secondary pathogens may augment the disease severity. Diarrhea occurs due to virus-mediated destruction of absorption efficient enterocytes, activation of enteric nervous system, or due to a rotavirus enterotoxin. Diagnosis of the infection relies on conventional techniques like isolation in MA 104 cell lines, electron microscopy, electro-pherotyping, and various serological tests. Presently, diagnosis and molecular typing is performed using serotype specific RT-PCR, sequencing or genomic hybridization techniques. As the rotaviruses are known to exhibit extreme genetic diversity and outplay disinfection procedures, eradication of the pathogen is often difficult. Hence, for prevention, good management practices coupled with vaccination of dam for protecting young ones, has to be practiced. Recently, new generation prophylactic strategies including DNA vaccines, subunit vaccines, virus-like particles (VLPs) and edible vaccines have been found to induce sufficient levels of passive immunity. Aside to the infection in animals, zoonotic significance of the animal rotaviruses has to be further unearthed. In this review, efforts have been made to highlight the importance and prevalence of the disease in bovines, its pathogenesis along with preventive measures, salient features of rotaviruses and their inter-species transmission abilities, zoonotic implications, and a concise account of the infection in various domestic animals and poultry.