De novo assembly, transcriptome characterization and marker discovery in Indian major carp, Labeo rohita through pyrosequencing.

Labeo rohita, one of the Indian major carps, is the most popular culture species in Indian subcontinent due to its consumer preference and delicacy. A selective breeding program for harvest body weight has resulted in an average genetic gain of 17% per generation. Transcriptome resource for this species is scanty. Here, we have characterized the liver and muscle transcriptomes of rohu using Roche 454 GS-FLX next generation sequencing platform. In total, 1.2 million reads were generated, de novo assembly and clustering resulted in 4171 transcripts. Out of these, 4171 had significant blast hit against NCBI nr database, and 2130 transcripts were successfully annotated. In total, 289 SSRs were identified with an identification rate of 5.8%, and dinucleotide repeat motifs were observed to be the most abundant SSRs. Further, 2231 putative SNPs were identified with high confidence. Validation of eight putative SNPs using Sanger sequencing resulted in 100% true SNPs. Significant allelic imbalance of M1, M4 and M5 loci between growth selected and control individual were observed. Furthermore, 13 transcription factors were identified in the present study belonging to six different transcription factor families. The present study demonstrated the utility of RNAseq to develop genomics resources in non-model fish species, and the marker resources developed would support the genetic improvement program of this species.

Molecular characterization, computational analysis and expression profiling of Dmrt1 gene in Indian major carp, Labeo rohita (Hamilton 1822).

Sexual dimorphism of fish morphology, physiology and behavior is diverse and complex in nature. Doublesex and mab-3 related transcription factor (Dmrt) is a large protein family whose function is sexual development and differentiation in vertebrates. Here, we report a full-length cDNA sequence of Labeo rohita (rohu) Dmrt1 of 907 bp length having 798 bp of open reading frame encoding 265 amino acids. The molecular weight of rohu DMRT1 protein was found to be 28.74 KDa and isoelectric point was 7.53. DMRT1 protein contains 23 positively and 24 negatively charged amino acids with a GRAVY score of -0.618. A characteristic DM domain was found in DMRT1 protein, which is a novel DNA-binding domain. Phylogenetic analysis showed maximum similarity with Cyprinus carpio when compared with DMRT1 of other vertebrates. Molecular docking study identified active sites to be targeted for drug designing. Rohu DMRT1 was observed to interact with other proteins such as FOXL2, CYP19a1a, AMH and SOX9a. Differential expression study revealed higher expression in testis tissue implying its role in male sex differentiation and testicular development. The information generated in the present work could facilitate further research to resolve the issues related to gonadal maturation and reproduction of commercially important aquaculture species.

Capturing transformation of flood hazard over a large River Basin under changing climate using a top-down approach.

Existing flood modeling studies over coastal catchments involving different combinations of model chain setup imparting complex information fails to entail the needs of policy or decision-makers. Thus, a comprehensive framework that pertains to the requirements of practitioners and provides more perspicuous flood hazard information is required. In this paper, a novel approach translating complex flood hazard information in the form of decision priority maps derived using a rational combination of models (physical and statistical) is elucidated at the finest administrative scale. The proposed methodology is illustrated over a highly flood-prone deltaic region in Mahanadi River Basin, India, to characterize impacts of climate change for a 1:100 years return period flood event under future conditions (2026-2055). The modeled flood events are further analyzed to capture the transformation dynamics of flood hazard classes (FHCs) in near-future, for prioritizing areas with greater hazard potential. Interestingly, the results capture a high transformation characteristic from low to high FHCs in agriculture-dominated areas, which are significantly greater than the areas experiencing flood hazard reduction. The results show a significant increase of 12.5% and 27.35% in areas with high FHCs under RCP4.5 and RCP8.5 scenarios, respectively. Moreover, a notable climate change response is indicated under both climate change scenarios, with approximately 22% (RCP4.5) and 25% (RCP8.5) in villages showing a drastic increment in flood hazard magnitude. The results thus highlight the importance of identifying and prioritizing the areas for flood adaptation where a relative change in flood hazard potential is higher due to climate change. Therefore, we conclude that this study can provide an insight into the implication of new approaches for effective communication of flood information by bridging the gaps between scientific communities and decision-makers in appraisal for better flood adaptation measures.

Population structure and genetic diversity of hatchery stocks as revealed by combined mtDNA fragment sequences in Indian major carp, Catla catla.

Catla catla is the second most important Indian major carp due to high growth rate and acceptance to consumers for food value. It is widely cultured in the Indian subcontinent as monoculture or polyculture. In the present study, genetic diversity among hatchery stocks (total 218 samples of catla) collected from different geographical regions of India was examined using mtDNA fragment sequence of Cyt b (306 bp) and D loop (710 bp). High numbers (57) of population specific haplotypes were observed in the present study. The results revealed significant genetic heterogeneity for the sequence data (FST = 0.27546, p < .05). Analysis of molecular variance revealed significant genetic differentiation among different catla populations. The information generated in present study could be useful to develop broad genetic base populations of catla.

Role of inflammatory and hemostatic biomarkers in Alzheimer's and vascular dementia - A pilot study from a tertiary center in Northern India.

INTRODUCTION: A reliable plasma biomarker in differentiating between Alzheimer's disease (AD) and Vascular dementia (VaD) is the need of the hour, in most memory clinics. Even though there is no disease modifying treatment, it is important to know the type of dementia for both symptomatic treatment and prognostication. METHODS: Neuropsychological assessment, MRI brain, FDG-PET brain and CSF biomarkers of AD (Aß42 and total tau) were used for establishing the diagnosis of Mild Cognitive Impairment (MCI), AD or VaD. RESULTS: 68 diagnosed patients of AD/MCI/VaD were included. FDG PET brain, plasma fibrinogen, d dimer, IL6 and CRP were done in all 68 patients while 48 patients underwent CSF biomarker analysis. Sixteen patients had MCI, of which 11 were MCI-AD and 5 were MCI-VaSC. There were 41 patients with AD (Mild AD-9, Mod AD-23, Severe AD-9) and 11 patients with VaD. Alzheimer group (MCI-AD and AD) and Vascular group (MCI VaSC & VaD) consisted of 52 and 16 patients respectively. Alzheimer and Vascular groups did not exhibit significant difference in IL6 and CRP levels. Plasma fibrinogen levels were significantly higher in VaD and vascular group as compared to Alzheimer group. But MCI-VaSC was not significantly different from MCI-AD. Plasma d dimer levels were significantly higher in all vascular subgroups compared to Alzheimer subgroups except between MCI-VaSC and MCI-AD. CONCLUSION: Hemostatic biomarkers were higher in Vascular group compared to Alzheimer group whereas there was no difference in inflammatory biomarkers. But the sensitivity and specificity of fibrinogen and d-dimer were not high enough for routine clinical use. Further studies in a larger sample are required to confirm these results.

Phytostabilisation potential of lemon grass (Cymbopogon flexuosus (Nees ex Stend) Wats) on iron ore tailings.

The present pot culture study was carried out for the potential phytostabilisation of iron ore tailings using lemon grass (Cymbopogon flexuosus) a drought tolerant, perennial, aromatic grass. Experiments have been conducted by varying the composition of garden soil (control) with iron ore tailings. The various parameters, viz. growth of plants, number of tillers, biomass and oil content of lemon grass are evaluated. The studies have indicated that growth parameters of lemon grass in 1:1 composition of garden soil and iron ore tailings are significantly more (-5% increase) compared to plants grown in control soil. However, the oil content of lemon grass in both the cases more or less remained same. The results also infer that at higher proportion of tailings the yield of biomass decreases. The studies indicate that lemongrass with its fibrous root system is proved to be an efficient soil binder by preventing soil erosion.

Surgical outcome in relation to duct size at the porta hepatis and the use of cholagogues in patients with biliary atresia.

BACKGROUND: Small ductules communicating with the bile ducts have been described at the porta hepatis in extrahepatic biliary atresia (EHBA) and these form the basis for hepatic portoenterostomy. The use of cholagogues like dehydrocholic acid (DHC) and ursodeoxycholic acid (UDCA) to enhance bile flow postoperatively has been reported. AIMS: This communication describes our experience with the use of cholagogues following surgery in EHBA and attempts to correlate the outcomes with the diameter of the ductules. MATERIAL AND METHODS: Fifty five EHBA patients treated by the Kasai procedure form the basis of this study; 35 patients treated during 1979-1986 and administered DHC (3-5 mg/kg) postoperatively and 20 patients treated during 1999-2002 and administered UDCA (15 mg/kg) postoperatively. The diameter of ductules was measured using an optical micrometer on 5 microm serial sections; the ducts were classified as type I (no demonstrable ducts, n = 14), type II (< 50 microm, n=22) and type III (> 50 microm, n = 19). The clinical outcome was categorized as 1 (jaundice free survival at 5 years follow-up, n = 7), 2 (initial good response but deteriorated after one year, n = 27) and 3 (expired within one year following surgery, n = 21). The response to surgery was monitored using biochemical liver function tests (LFT), hepatobiliary scintigraphy (HIDA scan) and occurrence of cholangitis. RESULTS: Age did not affect the size of ducts in both DHC and UDCA groups but patients in the DHC group were older than those treated with UDCA (mean age DHC: 105.22 +/- 33.53 days, UDCA: 74.68 +/- 23.73 days; p = 0.009). There was no statistically significant difference between duct size and postoperative LFT in both groups (DHC p = 0.1, UDCA p = 0.5). Bile excretion on HIDA scan was significantly better with larger ducts (DHC p = 0.003, UDCA p = 0.025); overall UDCA showed significantly better bile excretion (p = 0.003) but this was not reflected in the surgical outcome. There was no significant difference in the surgical outcome of those treated with DHC or UDCA but a significantly higher incidence of cholangitis was seen with smaller ducts in the UDCA group (p = 0.02). CONCLUSIONS: There was no correlation between duct diameter and postoperative LFT but type III ducts were associated with better bile flow on HIDA scan. Cholangitis was seen more often with type I and II ducts in both DHC and UDCA groups. UDCA administration seemed to be beneficial in patients with type III ducts in increasing bile flow and reducing cholangitis.

Profile of trauma related mortality at Manipal.

BACKGROUND: The main objective of the present study is to analyse the pattern of death related to trauma/injuries at Manipal in Udupi district, Karnataka state, India. MATERIAL AND METHODS: Retrospective data (January 2001 to December 2003) were collected from autopsy reports maintained in the department of forensic medicine, and the inquest reports from the police. Out of the total (n=470) autopsies, 344 victims had traumatic fatality. The demographic data, type of trauma, duration of survival, body areas involved was recorded. RESULTS: 470 autopsies were performed during the period of study. Of these 344 (73%) were traumatic fatalities. Males were preponderant (71%). More than half of the victims were between 1-40 years. The youth age of 21-30 years were affected maximum due to all type of trauma except for falls and assault in this area of the country (India). The leading type of trauma was Road Traffic Accident (RTA) (52%) followed by burn (24%). Females were predominant with 74% in trauma due to burns. In majority of cases where the death was reported in the hospital, among them more than 50% of victims were survived in hospital for more than 24 hours. The victims those expired within 3 hours were in majority from RTA and those who survived for more than 24 hours were in majority from burns. Majority (258) of the victims had head injuries followed by thoracic injuries (169), and abdomen (125). Twenty four percent of the victims died on the spot or before reaching hospital. CONCLUSION: In this area of the country the trauma due to RTA is a leading cause of death among the traumatic death followed by burns.

Evaluation of an in situ forming hydrogel wound dressing based on oxidized alginate and gelatin.

Wound dressings that can be formed in situ offer several advantages over the use of preformed dressings such as conformability without wrinkling or fluting in the wound bed, ease of application and improved patient compliance and comfort. Here we describe such an in situ forming hydrogel wound dressing from gelatin, oxidized alginate and borax. Periodate oxidized alginate rapidly cross-links proteins such as gelatin in the presence of borax to give in situ forming hydrogels that are both non-toxic and biodegradable. The composite matrix has the haemostatic effect of gelatin, the wound healing-promoting feature of alginate and the antiseptic property of borax to make it a potential wound dressing material. The hydrogel was found to have a fluid uptake of 90% of its weight which would prevent the wound bed from accumulation of exudates. The water vapour transmission rate (WVTR) of the hydrogel was found to be 2686+/-124 g/m2/day indicating that the hydrogel can maintain a moist environment over wound bed in moderate to heavily exuding wound which would enhance epithelial cell migration during the healing process. The wound healing efficacy of hydrogel was evaluated in experimental full thickness wounds using a rat model which demonstrated that within 2 weeks, the wound covered with gel was completely filled with new epithelium without any significant adverse reactions. These in situ forming hydrogels fulfil many critical elements desirable in a wound dressing material.

Long term tissue response to titanium coated with diamond like carbon.

Diamond like carbon (DLC) coatings were deposited on to Ti substrates by plasma enhanced chemical vapor deposition technique. Ti and DLC/Ti samples were implanted in skeletal muscle of rabbits. The samples were explanted after 1, 3, 6 and 12 months and the tissue-cell interaction was studied. Our data indicate both DLC/Ti and bare Ti to be compatible with skeletal muscle.

Bancroftian filariasis: a 13-year follow-up study of asymptomatic microfilariae carriers and endemic normals in Orissa, India.

The natural history of human filarial infections leading to development of disease has been a subject of intense debate. The models proposed so far have largely been based on cross-sectional data on microfilariae (Mf) and disease prevalence in filariasis endemic areas. In an attempt to study the parasitological and clinical consequences of filarial infection in Beldal (Orissa, India), an area endemic for Bancroftian filariasis, cohorts of 59 asymptomatic Mf carriers (AS) and 187 asymptomatic and amicrofilaraemic subjects or 'endemic normals' ('EN'), were followed-up and a fraction (73% and 46% respectively) re-examined after 13 years to monitor (a) Mf prevalence, (b) Mf density, (c) circulating filarial antigen (CFA) and (d) chronic disease manifestations. The Mf prevalence and density were also monitored in Mf carriers after 1 and 4 years. Both Mf prevalence and density decreased progressively in the cohort of Mf carriers over a period of 13 years in Beldal. Only 37% of them continued to be microfilaraemic and the Mf density in these subjects was only 10% of the original level. However, loss of circulating Mf in this cohort did not result in loss of CFA and 95% remained CFA positive regardless of Mf status. About 23% of males in the 'EN' cohort developed hydrocoele while only 5.7% of male Mf carriers, who were not treated with DEC, had developed hydrocoele after 13 years. A cohort of Mf carriers in another area, Jatni, was also examined after 10 years to study the parasitological and clinical outcome. In this area, about 59% of the Mf carriers continued to be microfilaraemic after 10 years. These results reveal that in Mf carriers adult filarial worms persist for several years and that loss of circulating Mf with or without chemotherapy with DEC (single 12-day course) does not influence adult worm survival. The findings have been discussed in the context of 'static' and 'dynamic' models describing the relationship between infection and disease in human filariasis.

Hypotonic swelling-induced Ca(2+) release by an IP(3)-insensitive Ca(2+) store.

Hypotonic swelling increases the intracellular Ca(2+) concentration ([Ca(2+)](i)) in vascular smooth muscle cells (VSMC). The source of this Ca(2+) is not clear. To study the source of increase in [Ca(2+)](i) in response to hypotonic swelling, we measured [Ca(2+)](i) in fura 2-loaded cultured VSMC (A7r5 cells). Hypotonic swelling produced a 40.7-nM increase in [Ca(2+)](i) that was not inhibited by EGTA but was inhibited by 1 microM thapsigargin. Prior depletion of inositol 1,4,5-trisphosphate (IP(3))-sensitive Ca(2+) stores with vasopressin did not inhibit the increase in [Ca(2+)](i) in response to hypotonic swelling. Exposure of (45)Ca(2+)-loaded intracellular stores to hypotonic swelling in permeabilized VSMC produced an increase in (45)Ca(2+) efflux, which was inhibited by 1 microM thapsigargin but not by 50 microg/ml heparin, 50 microM ruthenium red, or 25 microM thio-NADP. Thus hypotonic swelling of VSMC causes a release of Ca(2+) from the intracellular stores from a novel site distinct from the IP(3)-, ryanodine-, and nicotinic acid adenine dinucleotide phosphate-sensitive stores.

Human bancroftian filariasis - a role for antibodies to parasite carbohydrates.

Studies on immune responses to parasites have been undertaken in filariasis with a view to understand protective immunity, pathogenesis of the disease process and mechanisms of immune deviation. However none of the investigations conducted so far on antibody responses have addressed the issue of immunogenicity of filarial carbohydrate antigens in human lymphatic filariasis. In this communication we report details on relative protein and carbohydrate contents of various developmental stages of filarial parasites and antibody responses to filarial proteins (Fil.Pro) and carbohydrates (Fil.Cho) in different clinical spectrum of human bancroftian filariasis. As expected, antibodies of IgM and IgG2 subclass recognized primarily Fil.Cho while IgG4 filarial antibodies recognized exclusively Fil.Pro. Reactivity of IgG3 to Fil.Cho was similar to that of IgG2 while IgG1 more readily recognized Fil.Pro than Fil.Cho. The IgG2 and IgG3 antibodies to Fil.Cho were found to be significantly more in patients with chronic filarial disease and in endemic normals when compared with microfilariae (mf) carriers while IgG4 antibodies to Fil.Pro were significantly more in mf carriers. The dichotomy in reactivity of filarial IgG2, IgG3 and IgG4 was dependent on active filarial infection as indicated by presence of circulating filarial antigen (CFA). Individuals with CFA were found to possess significantly more IgG4 to Fil.Pro than those without CFA while IgG2 and IgG3 levels to Fil.Cho was significantly more in CFA negative subjects when compared to those with CFA. Although IgG1 reacted more readily with Fil.Pro, unlike IgG4, their levels were significantly more in CFA negative subjects when compared to those with active filarial infection. Absorption of sera with phosphorylcholine (PC) resulted in no significant loss of reactivity to Fil.Cho indicating that most of the anticarbohydrate antibodies were recognizing non-PC determinants in human filariasis. Elevated levels of IgG2 and IgG3 antibodies to Fil.Cho in individuals free of filarial infection indicate a possible role for carbohydrate antigens in induction of protective immunity in human filariasis.

The influence of photodynamic therapy on the wound healing process in rats.

In photodynamic therapy (PDT), photosensitisers (PS) are used along with lasers for the treatment of tumors. The combined effect of photosensitisers and lasers on the wound healing process is studied using delta-aminolevulinic acid (ALA) (5 mg/kg) and hematoporphyrin derivative (HPD) (5 mg/kg) as photosensitisers in the open excision wounds of rats. The lasers used were He-Ne laser (3 J/cm2) and Nd:YAG laser (30 J/cm2). This study is important for understanding the healing process involved after PDT. Open excision wounds treated with He-Ne lasers in animals that received ALA as photosensitiser showed complete wound closure at the earliest by 13 +/- 1 days, and with results obtained for HPD and the combination of lasers with complete closing by 14 +/- 1 days. However, the control group of animals that received ALS or HPD with no laser treatment showed wound healing on the twentieth and eighteenth days with a deviation of one day and two days, respectively. ALA with the combination of Nd:YAG and He-Ne lasers and HPD with He-Ne laser alone does not show quicker wound healing effects. Histopathological results also gave similar results. Tensile strength measurements do not vary significantly from control group to the test group. ALA along with He-Ne laser of HPD along with the combination of He-Ne and low power Nd-YAG lasers are found to be ideal methods for quickening the wound healing process in rat.

Human Bancroftian filariasis: loss of patent microfilaraemia is not associated with production of antibodies to microfilarial sheath.

Antisheath antibodies have been incriminated in elimination of circulating microfilariae in human filariasis since a very significant inverse association has been consistently demonstrated between the two parameters. An attempt was made in the present study to seek empirical proof for the above proposal. Two cohorts of 43 and 73 microfilariae (mf) carriers were examined after 13 and 10 years, respectively, for mf as well as antisheath antibodies. The first cohort was also examined for the presence of circulating filarial antigen (CFA). Of the 43 mf carriers examined after 13 years, 62.8% were free of circulating mf although only 3.7% of them had demonstrable antisheath antibodies. Approximately 93% of this cohort (with or without current microfilaraemia) tested positive for CFA after 13 years indicating continued presence of adult filarial worms in the host after loss of mf in circulation. When the second cohort of 73 mf carriers were examined after 10 years, 30 were found to be amicrofilaraemic and only 6.66% of them had demonstrable antisheath antibodies. We conclude that, in human Bancroftian filariasis, elimination of circulating microfilariae may not be mediated by antibodies to microfilarial sheath.

Synergistic effect of released aspirin/heparin for preventing bovine pericardial calcification.

Calcification is a frequent cause of the clinical failure of bioprosthetic heart valves fabricated from glutaraldehyde pretreated bovine pericardium (GATBP). Aspirin, a potent antiplatelet drug, and heparin, an anticoagulant, are commonly used for postimplant complications such as thrombosis and thromboembolism. Aspirin and heparin were embedded in chitosan/polyethylene vinylacetate co-matrix to develop a prolonged release form. The effect of these drugs towards the bioprosthetic calcification was investigated by in vitro and in vivo models. In vitro and in vivo evaluation suggest that the released aspirin/heparin from the co-matrix had a synergistic effect in inhibiting GATBP calcification. In vivo subcutaneous co-implantation was performed with PEG-20,000 grafted bovine pericardium (PEG-GABP), aspirin, and heparin. Biochemical, histological, and scanning electron microscopic evaluation of retrieved samples demonstrated a significant reduction in calcium deposition and alkaline phosphatase activity on PEG-GABP compared to GATBP. It seems that the aspirin/heparin combination synergistically inhibits the pericardial calcification in addition to their antithrombotic function.

Effects of double cross-linking technique on the enzymatic degradation and calcification of bovine pericardia.

The strength, resorption rates, and biocompatibility of collagenous biomaterials are profoundly influenced by the method of cross-linking. The in vitro and in vivo calcification and enzymatic degradation of bovine pericardia (BP) after a series of surface modifications were studied as a function of exposure time. Collagenase degradations of modified BP were monitored by scanning electron microscopy and tensile strength measurements. Bovine pericardium was modified by a combination of different tissue fixatives such as glutaraldehyde (GA), carbodiimide (EDC), diisocyanate (HMDIC), and polyethylene glycol (PEG). GA-PEG-EDC-PEG and GA-PEG-HMDIC-PEG combination treated BP retained maximum stability in collagenase digestion compared to GATBP. In vitro calcification studies and in vivo rat subcutaneous implantations of modified pericardium have shown substantial reduction in the calcification of double cross-linked BP with PEG modification. Further, the biocompatibility aspects of pericardial tissues were established by platelet adhesion and octane contact angle. It seems that cross-links involving amino and carboxyl residues may provide new ways of controlling biodegradation and calcification.

Setaria digitata infections in cattle: parasite load, microfilaraemia status and relationship to immune response.

A total of 110 cattle were examined in an area endemic for Bancroftian filariasis for the prevalence of infection of the bovine filarial parasite Setaria digitata. About 12.5% of cattle were found to harbour both adult worms in the peritoneum and microfilariae (mf) in circulation; 70% of the cattle were amicrofilaraemic but with an adult worm infection. A third group of cattle (16.5%) was free of detectable mf and adult worms. The presence of adult worms and/or mf did not influence the antibody levels to any of the four antigen preparations of S. digitata. However, there was a significant inverse relationship between the presence of antibodies to microfilarial sheaths and the absence of circulating mf as shown by the immunoperoxidase assay. Cattle immunoglobulin containing high titres of anti-sheath antibodies cleared circulating microfilariae very effectively in Mastomys coucha thus demonstrating the protective nature of anti-sheath antibodies in eliminating circulating microfilariae in vivo.