Purification and Characterization of Pomphorhynchus kashmirensis Somatic Antigens Collected from Naturally Infected Local Fish Schizothorax

Abstract: The present investigation deals with the fish intestinal parasite Pomphorhynchus kashmirensis isolated from local fish species Schizothorax. The host fish were collected from two study sites viz the Dal Lake and the River Jhelum, Srinagar, India. The Pomphorhynchus kashmirensis were subjected to immunological studies, especially to reveal the nature of their somatic antigens. The nature of somatic antigens was studied by affinity chromatography and SDS-PAGE and their antigenic properties were also confirmed by Ouchterlony double diffusion test (ODD). A total of 363 fish specimens were collected and out of which 94 specimens were found to harbor the Pomphorhynchus kashmirensis constituting an overall prevalence of 25.89%. SDS-PAGE of partially purified somatic antigens of Pomphorhynchus kashmirensis through affinity chromatography resolved into five prominent polypeptides of molecular weight ranging from 29-66 kDa by using the known molecular weight marker. Furthermore, the antigenicity of the purified antigens was confirmed in ODD against hyper immune sera raised in rabbit and with homogenous immune sera of naturally infected fish. Only one precipitation arch was formed against hyper immune sera and many precipitation arches were formed against homogenous immune sera

Effect of Metal Toxicity in Cyprinus carpio Linnaeus and Schizothorax niger Heckel with emphasis on Biochemical and Histopathological Parameters.

Due to scanty information regarding the metal toxicity in water bodies of Kashmir valley and their effects on the aquatic fauna, the present study was designed with the following objectives: 1) To study the toxicity of some metals viz. copper, zinc, iron, and manganese in Dal lake and River Jhelum. 2) To study the concentration of these metals through Atomic absorption spectrophotometer in various organs/tissues viz. gills, liver, kidneys, and muscles of Schizothorax niger and Cyprinus carpio spp. 3) To study the subsequent effects of metals on biochemical parameters viz. total protein, albumin, globulin, blood glucose, urea, serum creatinine and cholesterol in both the fishes. 4) To study the subsequent effects of these metals, on the histomorphology of gills, liver, kidney and muscles of both the fishes.Digital copy of Thesis.University of Kashmir

Haematological and Biochemical Studies of Helminth Infected Goats in South Kashmir

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Purification and Characterization of Pomphorhynchus kashmirensis Somatic Antigens

Fish fauna is ecologically very important and plays a vital role in food chain. There is a great scope for the development of fishery resources in the valley of Kashmir so as to overcome the prevailing animal protein defiency in the diet of the local people. In country like India, intake of meat & milk is low, so fish has special importance as a supplement to ill- balanced cereal diet. Fish flesh is also a highly perishable commodity constituted by 60-80 %water & 13-20 % protein & being low in cholesterol & free from fats. Fish protein hydrosylates also have antioxidant properties. The flesh also contains phosphorus & vitamins. Freshwater fishes form one the important food sources in both the developed as well as underdeveloped countries. A lot of research work has been done on various aspects of these vertebrates but a meagre work is available on the molecular aspects particularly immunogenicity of parasites of the fish host. Therefore, study on the characteristics of the protein profile of the fish parasite Pomphorhynchus kashmirensis was undertaken. The fishes were collected from Dal Lake and River Jhelum and it forms the first study of its type in this part of the country. In order to have a glimpse of the related work done in the past, an attempt was made to review the available literature on the subject. Parkhouse, et al., (1987) characterized and studied the protective effect of nematode antigens. Bunyatova and Elchiev (1989) constructed electrophoretic spectrum of proteins of an acanthocephalan Leptorhynchoides plagicephalus. Coscia and Oreste (2000) investigated the presence of antibodies against protein antigens of the nematode parasite Pseudoterranova decipiens in the plasma and bile of the Antartic Teleost, Trematomus bernachii. Knopf, et al., (2000) studied the humoral response of the European eel Anguilla Anguilla elicited by an experimental infection with a swim bladder nematode Anguillicola crassus. Noga, et al., (2000) purified antimicrobial proteins from rainbow trouts and sunshine bass by RP-HPLC and SDS-PAGE. Saifullah, et al., (2000) worked on the excretory /secretory (ES) metabolic products released by Gastrothylax crumonifer (trematode:Digenea) during in vitro incubations and the somatic extract of the adult parasites were analyzed using PAGE. As obvious, it was not possible to review all the available literature on the said topic, the important ones were penned down in the dissertation. Studies on the fishes has showed that Pomphorhynchus kashmirensis is usually found in the intestine by boring its proboscis into it and thereby produces lesions and makes it prone to further secondary infections. Besides, it is also present in the body cavity, liver and spleen of its fish host. It causes tremendous damage to the intestinal walls at the site of its attachment. The lamina propia gets thickened and goblet cells in this region become more prominent and their number also gets increased (Yildiz, et al, 2004; Ahanger, et al., 2008).This parasite is considered to be one of the most dangerous parasite responsible for fish mortality and morbidity (Ahanger, et al., 2008). Hence the need was felt to study this parasite at molecular level so that our fish fauna are spared from them. A total of 363 fish specimens of Schizothorax species were collected and out of which, 203 fishes were collected and examined from Dal Lake and 160 fishes were collected and examined from River Jhelum during the present study. The host was collected with the help of local fishermen in live condition. Fishes were dissected & body cavity was thoroughly examined for any parasite. Intestines were placed in Petri dish containing normal saline (0.75%Nacl, Cable 1958) to allow adhering parasites to be released from the lumen. Pomphorhynchus was carefully removed from the intestines with the help of brush and needle. A regular record of this parasites was recorded and then subjected to various immunological and biochemical techniques in order to understand the nature of somatic antigens. Out of 203 specimens examined from the Dal Lake only 42 specimens were found infected with the Pomphorhynchus kashmirensis which constitutes the prevalence of 20.68%. Similarly out of 160 specimens examined from the River Jhelum only 52 specimens were infected with the Pomphorhynchus kashmirensis which constitutes 32.5% prevalence. Also, Pomphorhynchus kashmirensis showed a wide host range and was successfully establishing in various species of Schizothorax. The highest prevalence were found in S. niger (30 %) (26.19% in Dal and 34.85% in Jhelum) followed by S. curvifrons (27.11 %) (19.11% in Dal and 38% in Jhelum) and least prevalence were found in S. esocinus (17.89%) (13.72% in Dal and 22.73% in Jhelum). Pomphorhynchus infection also revealed definite seasonal prevalence of infection in all the three species of Schizothorax, with highest infection in summer and lowest in winter. There was a gradual increase in the prevalence rate from spring to summer and falls down with onset of autumn and least observed prevalence during winter season. In summer the prevalence was 34.54% (S. niger 46.34%, S. curvifrons 31.11% and S. esocinus 30%) and the least prevalence was found during the winter season 8.16% (S. niger 14.28 %, S. curvifrons 6.89 % and S. esocinus 9.52%). Gender wise observations were also made which revealed that the sex wise differences were not much prominent but in most cases males 30.30% (S. niger 32.50%, S. curvifrons 27.65% and S. esocinus 28.94%) were found to be more infected than females 27.27% (S. niger 27.14%, S. curvifrons 36.61% and S. esocinus 15.78%). To characterize the somatic antigens from Pomphorhynchus kashmirensis, the sera of the fish was used as a source of antibodies. For this purpose affinity chromatography is the most appropriate technique to be utilized. From this technique the eluted bound protein was dialyzed extensively against 20 mM Tris-saline buffer, pH 7.4, concentrated with PEG 20,000 and was designated as affinity purified P. kashmirensis somatic antigen (Aff-PSAg). The protein content of the antigen was determined spectrophotometrically. SDS-PAGE confirmation of purified antigen Aff-PSAg was done by SDS-PAGE followed by staining with Coomassie Brilliant blue. Electrophoretic separation of Aff-PSAg resolved into 5 prominent polypeptides of molecular weight ranging from 29 to 66 kDa which is inferred to the presence of 5 or more number of active somatic antigens of P. kashmirensis. To measure the antigenicity of the purified antigens the commonly used serotest viz., Ouchterlony double diffusion (ODD) was used. Crude somatic and partially purified pooled fractions of Pomphorhynchus kashmirensis were subjected to double immunodiffusion against rabbit hyper immune sera. Ouchterlony gel diffusion test of somatic antigens showed one precipitation arch against heterogeneous hyper immune sera and many precipitation arches against homogenous hyper immune sera. In nutshell, it is observed that the somatic antigens derived from the Pomphorhynchus kashmirensis can be used as good immunogens and hence can be exploited for mounting the protective immune response in fish. The results of the present study suggest that low molecular weight antigens of Pomphorhynchus kashmirensis deserve further investigation

Ichthyofauna of Lidder Stream with Emphasis on Haematobiochemical Study of Schizothorax SPP.

Jammu & Kashmir, lying between six mountain ranges and covering an area of 2,22,236 sq. kms. is located between 32° 17' and 36° 58' North latitude, and between 37° 26' and 80° 30' East longitude. The state commonly known as Kashmir is bounded in the north by Afghanistan and China, in the east by China, in the south by the state of Himachal Pradesh and the state of Punjab in India, and in the west by the North-West Frontier Province and the Punjab Province of Pakistan. Jammu and Kashmir geographically comprises of three regions: the foothill plains of Jammu; the lakes and blue valleys of Kashmir rising to alpine passes, the high altitude plains and starkly beautiful mountains of Ladakh which lies beyond narrow passes. Kashmir is rightly said to be the nature's grand finale of beauty. In this masterpiece of earth's creation seasons in strong individuality vary with one another in putting up exquisite patterns of charm and loveliness. This state holds the glorious history of the valiant kings, the placid lakes, the greenery of the forests and the amazing rivers. The sunny gardens, romantic house boats and Lakes add to the beauty of the region. The state of J&K measures about 425 kms from north to south and extends over 520 kms from east to west

Community structure of fish parasites and their intermediate hosts in relation to Trophic conditions of lakes.

The present study was undertaken to evaluate the helminth communities in populations of three fish species along the trophic gradient of lakes. Data were collected from three lakes of differing trophic status in the Kashmir Himalayas, namely Anchar (severely eutrophic), Dal (highly eutrophic) and Manasbal (moderately eutrophic) during June 2006 to May 2008. The main objectives of the study were: (1) to examine the physico-chemical characteristics of lakes and (2) to assess the helminth assemblage patterns in fish populations of lakes. In addition, a follow-up study was undertaken to study the density profiles of intermediate hosts during the second year (June 2007 – May 2008). Fish species studied included one native species Schizothorax niger Heckel and three non-native species – Cyprinus carpio communis Linnaeus, C. c. specularis Linnaeus and Carassius carassius (Linnaeus). A comparison of physico-chemical parameters and depth revealed that the study lakes presented a mosaic of different trophic gradients; Anchar was most affected by eutrophication, followed by Dal, whereas Manasbal was the least eutrophic among the three lakes. A total of seven helminth parasite species were recorded during the investigation period. These included three trematodes: Diplozoon kashmirensis Kaw, Clinostomum schizothoraxi Kaw, and Posthodiplostomum cuticola Dubios (Neascus cuticola Nordmann); two cestodes: Bothriocephalus acheilognathi Yamaguti and Adenoscolex oreini Fotedar and two acanthocephalans: Pomphorhynchus kashmirensis Kaw and Neoechinorhynchus manasbalensis Kaw from Acanthocephala. The populations of different intermediate host guilds such as cyclopoid copepods, gastropods, oligochaetes, ostracods, and amphipods showed interlake variations. The densities of intermediate hosts especially cyclopoid copepods, gastropods and oligochaetes were higher at Anchar, suggesting a positive response of the invertebrate host community to cultural eutrophication. However, the pattern exhibited by ostracods and amphipods were not so distinct which agreed well with the rare status of their parasites in lakes. The analysis of data showed a clear habitat effect on the abundance pattern of helminth species. Helminth infracommunity richness was highest in host populations of S. niger and C. carassius from the Anchar lake compared to two other lakes. On the otherhand, there was no observed pattern of community structure in the case of C. carpio in the three lakes.Digital copy of Ph.D thesis.University of Kashmir

Whirlpool: Improving Dynamic Cache Management with Static Data Classification

Cache hierarchies are increasingly non-uniform and difficult to manage. Several techniques, such as scratchpads or reuse hints, use static information about how programs access data to manage the memory hierarchy. Static techniques are effective on regular programs, but because they set fixed policies, they are vulnerable to changes in program behavior or available cache space. Instead, most systems rely on dynamic caching policies that adapt to observed program behavior. Unfortunately, dynamic policies spend significant resources trying to learn how programs use memory, and yet they often perform worse than a static policy. We present Whirlpool, a novel approach that combines static information with dynamic policies to reap the benefits of each. Whirlpool statically classifies data into pools based on how the program uses memory. Whirlpool then uses dynamic policies to tune the cache to each pool. Hence, rather than setting policies statically, Whirlpool uses static analysis to guide dynamic policies. We present both an API that lets programmers specify pools manually and a profiling tool that discovers pools automatically in unmodified binaries. We evaluate Whirlpool on a state-of-the-art NUCA cache. Whirlpool significantly outperforms prior approaches: on sequential programs, Whirlpool improves performance by up to 38% and reduces data movement energy by up to 53%; on parallel programs, Whirlpool improves performance by up to 67% and reduces data movement energy by up to 2.6x.National Science Foundation (U.S.) (grant CCF-1318384)National Science Foundation (U.S.) (CAREER-1452994)Samsung (Firm) (GRO award

DHA Supplemented in Peptamen Diet Offers No Advantage in Pathways to Amyloidosis: Is It Time to Evaluate Composite Lipid Diet?

Numerous reports have documented the beneficial effects of dietary docosahexaenoic acid (DHA) on beta-amyloid production and Alzheimer's disease (AD). However, none of these studies have examined and compared DHA, in combination with other dietary nutrients, for its effects on plaque pathogenesis. Potential interactions of DHA with other dietary nutrients and fatty acids are conventionally ignored. Here we investigated DHA with two dietary regimes; peptamen (pep+DHA) and low fat diet (low fat+DHA). Peptamen base liquid diet is a standard sole-source nutrition for patients with gastrointestinal dysfunction. Here we demonstrate that a robust AD transgenic mouse model shows an increased tendency to produce beta-amyloid peptides and amyloid plaques when fed a pep+DHA diet. The increase in beta-amyloid peptides was due to an elevated trend in the levels of beta-secretase amyloid precursor protein (APP) cleaving enzyme (BACE), the proteolytic C-terminal fragment beta of APP and reduced levels of insulin degrading enzyme that endoproteolyse beta-amyloid. On the contrary, TgCRND8 mice on low fat+DHA diet (based on an approximately 18% reduction of fat intake) ameliorate the production of abeta peptides and consequently amyloid plaques. Our work not only demonstrates that DHA when taken with peptamen may have a tendency to confer a detrimental affect on the amyloid plaque build up but also reinforces the importance of studying composite lipids or nutrients rather than single lipids or nutrients for their effects on pathways important to plaque development

The epidemiology of suicide and attempted suicide in Dutch general practice 1983–2003

BACKGROUND: Many patients attempting or committing suicide consult their general practitioner (GP) in the preceding period, indicating that GPs might play an important role in prevention. The aim of the present study was to analyse the epidemiology of suicidal behaviour in Dutch General Practice in order to find possible clues for prevention. METHOD: Description of trends in suicide and suicide attempts occurring from 1983–2003 in the Dutch General Practice Sentinel Network, representing 1% of the Dutch population. The data were analysed with regard to: 1) suicidal behaviour trends and their association with household situation; 2) presence of depression, treatment of depression and referral rate by GPs; 3) contact with GP before suicide or suicide attempt and discussion of suicidal ideation. RESULTS: Between 1983 and 2003 the annual number of suicide and suicide attempts decreased by 50%. Sixty percent of the patients who committed or attempted suicide were diagnosed as depressed, of whom 91% were treated by their GP with an antidepressant. Living alone was a risk factor for suicide (odds ratio 1.99; 95% CI 1.50 to 2.64), whereas living in a household of 3 or more persons was a relative risk for a suicide attempt (odds ratio 1.81; 95% CI 1.34 to 2.46). Referral to a psychiatrist or other mental health professionals occurred in 65% of the cases. GPs recalled having discussed suicidal ideation in only 7% of the cases, and in retrospect estimated that they had foreseen suicide or suicide attempts in 31% and 22% of the cases, respectively, if there had been contact in the preceding month. CONCLUSION: With regard to the prescription of antidepressants and referral of suicidal patients to a psychiatrist, Dutch GPs fulfil their role as gatekeeper satisfactorily. However, since few patients discuss their suicidal ideation with their GP, there is room for improvement. GPs should take the lead to make this subject debatable. It may improve early recognition of depressed patients at risk and accelerate their referral to mental health professionals