My research career

I started my research activities following grant of Junior Research Award  Council of Scietific and Industrial Research, New Delhi in the year 1982 and joined the Unviversity Department of Zoology,L.N.Mithila University under the supervision of Late Dr.S.Pandey. Taking an start with the research problem was like a dream coming true.It aws difficult as well in the given state of the art of the Department. However, it couldd be started in an isolated corner of the  Department vieving the safety reasons. The problem I was working on related to induction of cancer thhrough highly contagious food contaLLOWminants of fugal origin.

In the following months I was selected for the post of Lecturer, in the services of L.N.Mithila University and joined at J  K College,Biroul on November 2,1982.The place is about 60 kms from Darbhanga. Following one year of the probation period I was awarded Teacher Fellowship of University Grants Commission  for three years. I rejoined The P G Department of Zoology at Darbhanga to complete of Doctoral Research under the supervision of  late Dr S pandey.The extensive work plan, however, could be completed only with an extension of one year of time because it was the tenure during which I was married to Rashmi and also blessed with the first son, Mitul.

Completion of Doctoral work was a real experience  due to poor ifrastructural facilities and poor knowhow of the experties at place of work. I highly acknowledge the cooperation extended  by Dr.B D Jones, Tropical Products Institute,London;  Dr.Dawe and Dr Harshberger of Smithsonian Institute, Director of Registry of Tumors in Lower Animals;Dr Nazar Ehshan , Professor and Headof Zoology, Patna Universityand Dr R A B Verma, Professor of Botany ,C.M.Science College,Darbhanga.

In the meantime ,I also applied for a Major Research Project to The Indian Council of Agricultural Research, New Delhi for financial support. Since I was then a research student ,Dr. S Pandey became the Principal Investigator and Dr. RA B Verma, Professor in Botany ,the Co Principal Investigator. I worked in this project as an Investigator.The Project was approved and the team started working.But, sudden demise of Dr. Pandey  left the work in midway

Award of Doctoral Degree could happen in 1989 .The delay was becasue of one of the examinner who suddenly left for abroad. A brief summary may be seen below:

Summary of my Doctoral work:

The thesis includes two sections.

SECTION I

1.  The fish feeds stored under poor sanitory conditions ( inadequate ventillation and improper protection against the relative humidity) are prone to fungal infestation.

2.  The fu :ngus ( Rhizopus sps.,Penicillium sps., and Aspergillus sps.) are found on all the three types of fish feeds ( mustard oil cake, mixed oil cake and ground pulses mixture) all the year around. Out of the seven species of the Aspergillus group, A.niger dominated in all the summer and winter samples but are overnumbered by A.flavus during the rains. A few soil fungi like A.terreus  and  A.nidulanse are also found in addition to A. rubur, which has not been reported from mustard or its derivatives.

3.  The range of temperature and relative humidity during summer and winter favour growth of  A.niger while the rainy months with temperature range between 20-30 °c and high relative humidity are congenial for the growth of A.flavus  and subsequent toxic metabolite production.

4.  The amount of moisture within the substrate is also a significant factor deciding the degree of mycobial spoilage. A direct product, like oil cakes , rapidly traps the moisture when stored in direct contact of moist soil without protective barrier. Such substrates are more prone to fungal spoilage with increased risk of subsequent toxic metabolite production.

5.  The amount of fat within the oil cakes increases the potentiality of the fungi (A.flavus) to produce their toxic metabolite (aflatoxin B1) in perticular during the rains.

SECTION II

6.  Morphology: The fish became lethargic with increased mucus secretion. they refuse to feed from day 60 onwards and the body color also darkens even at the lowest dose of the toxin (10 ppm). these symptoms are pronounced withincrese in the dose of the toxin , on day 4-5 at 40 ppm concentration.

7. Gross Anatomy : The fish show moderate to severe anemia with different ranges of hemorrhage primarily on the liver and parts of the gastrointestinal tract. The spleen and head kidney become swollen. the allterations include hyperamic swollen patches in descending order of severity from liver, excretory kidney, intestine to pyloric caeca.

                             The tumorous outgrowths ranging from 0.5- 3.0 mm are found on the liver, kidney and intestine. The first outgrowth on the liver appears at 30 ppm/60 day level and the number becomes 14 at 40 ppm/60 day level. The tumor on the kidney develops at 30 ppm/60 day and on the intestine at 40 ppm/ 80 day level.

8.  Histopathology:

Liver : Non  neoplastlar symptoms appearing in liver include periportal and centrilobular necrosis, swelling, edema, hyperamia, hemorrhage, sinusoidal dilation (moderate), vacoulization and toxic liver cirrhosis.

The neoplasic changes appear at 20 ppm/80 day level prominantly in cirrhotic liver. however , the classical form of trabecular hepatoma is observed at 40 ppm dose level. The malignant nature of the lesion becomes apparent due to envasive erosion of theb adjacent tissue structures and trace of malignant hepatocytes within the vascular system.

Excretory Kidney:  In addition to hyperamia, inflammation , edema, hemorrhage and necrosis sign of severe interstitial lyphoid tissue prolifaration is marked at 20 ppm/ 80 day level. Increased dose of toxin leads to adenocarcinoma arising from the tubular epithelium and outgrowths due to anaplastic proliferation of interstitial lymphoid tissue. The blood vessels also undergo malignant arteriosclerosis.

Head Kidney : The early symptoms simulate the condition as in the case of the excretory kidney. The sinusoidal dilation and reduction in the haematopoietic tissue  due to edema and focal necrosis are evident. The interstitial haematopoietictissue shows sign of prolifaration at 20 ppm/ 80 day, which becomes more pronounced wit increase in the dose and treatment period. A concomitant alteration characterising preneoplastic condition in the interrenal tissue is obserevd at 40 ppm dose level.

Pyloric caeca and Intestine:  Both of these organs develop hyperamia, hemorrhage, mucosal sloughening and atrophy in the similsr manner. At 20 ppm dose level the submucosa alongwith the columnar epithelial cells show signs of hyperplasia, ultimately appearing as anaplasia in advaced conditions.. No defined anaplastic lesion is marked in the pyloric caeca while in the intestine  adenocarcinomatous lesion ( poorly differentiated) are found raplacing the entire mucosal layer. these lesion remain organsed in atypical glandular masses.

Metastasis of malignant hepatocytes forming lesion around blood vessel between serosa and muscularis is disticnt. However, the evidences suggest a reverse phenomenon of metastasis of anaplastic cells from mucosa to the adjacent tissue system (local metastasis) and distant organs as well.

The muscularis becomes mostly hyperamic with swollen fiber cells conatining darkened nuclei. The interlacing arrangement of these cells suggest the manifestation simulating leimuoma but no apparent sign of proliferation is marked to designate the condition as neoplastic.

SpleenThe splen during advance stages show monotony of lymphocytes of different rangealongwith a few atypical cells. the symptoms simulate lymphatic leukemia. a marked elevation in the total lymphocytic count also support the present fidings of lymphocytic leukemia. However, a proper diagnosis requires prolonged treatment.

 

Area of Interest

  • Environmental Toxicology
  • Toxins of Biological Origin (algal and fungal toxins) with reference to freshwater aquaculture and human health
  • Wetlands: ecology,management and conservation
  • Fish consupmption and human health

      Research Experience

  • Junior Research Fellow, Council of Scietifhic and Industrial Research, New Delhi  - Feb.3, 1982- Nov. 1,1982
  • Investigator, Minor Research Project of UGC, New Delhi (No.F-25-6-1(639)-84-SR-1-V)
  • Teacher Fellow of UGC,New Delhi  -   Jan 10,1985 to Nov.1,1988
  • Investigator, ICAR sponsored Major Research Project on " Pathophysiological Changes induced in Fish due to mycotoxin contaminated ration"  1988-1991
  • Principal Investigator, ICAR sponsored Major Research Project on " Mycotoxins: Histopathological Manifestations in fish and possibilities of Human Uptake"  1999-2002
  • Principal Investigator, ICAR sponsored Major Research Project on "Biodiversity of blue-green algae and ecotoxicology of Microcystis aeruginosa in fish culture ponds of Darbhanga" 2003-2006
  • Principal Investigator, ICAR sponsored Major research Project on " Bioattainment of Peak Nutrition Value (PNV) in Fish  2004-2007

      Research Contributions

  •  Reported about the toxic fungal flora infesting agricultural products, their byproducts during storage condition and impact of climatic factors, recurrent flood on the incidence of the toxic fungal strains and subsequent toxic metabolite production.
  • Workedout the behavioural, clinical and histopathological traits of allatoxicosis, ochratoxin and sterigmatocystin poisonin in air-breathing teleost fish as well as some of the major carps.
  •  Workedout the chronology of events leading to a classical case of trabecular hepatoma.
  • Bisides liver cancer the aflatoxins also induce some alteration at hemic level simulating lymphatic leykemia in man.
  • Drawn the attention of  Fisheries Section of Indian Council of Agricultural Research, New Delhi towards fungal contamination of fish feeds in prevalent climatic conditions and potential hazards to our Inland Aquaculture practices through major research schemes.
  • Highlighted the threat to aquaculture practices due to increasing incidence of cyanobacterial bloomes awith special referce to Microcystis aeruginosa
  • Many of the aquaculture ponnds were thoroughly investigatod for physicochemical factors and compared with eutrophicated , unproductive ponds to ivvestigate the casative factors.
  • In most of the conditions of cyanobacterial blooms N:P ratio was found to have highly significant role, rather a causative factor.
  •  Possibilities of direct human exposure to aflatoxins throug dried vegetable substitutes and dry fruits specially makhana ( Euryale ferox slisb.)
  • Drawn the attention of WHO towards in kidney disorders in Bihar. The problem was investigated through Doctoral work and found that staple dietary items were often contaminated with a strain of fungi well recognised for creating such ailments.
  • Investigated the etiology of the nephropatheic disorders in man (in patients under treatment) through elaborate  survey, clinical and pathological examinations to trace the role of fungal contaminants.The data obtained through bioassay on Guineanpigs  were extraploated to come to an established conclusion. findins were further supported with trce of the fugal metabloites in human blood.
  •  Possibilities of human exposure to toxic fungal metabolites through fish flesh consumption has widely investigated
  • Effect of different cooking processes (prevalent in India) on the fate of fugal contaminants has been investigated.
  • In view of the understanding of fish flesh consumption for theraputic purposes, a major research project has been carriedout to investigate the peak nutrition value of fishes in their life cycle with special reference to omega 3 and omega 6 fatty acids. Role of these fatty acids in cardioprotection,many forms of cancer, arthritis are now fairly established. Fish as one of the most abundant natural resorce of these fatty acids have gained significance
  • Effect of differesnt culture conditions,culture practice,supplementary diets and feeding habit of the fish itself  on age dependent profile of these fatty acids have also been investigated
  • Studies on the effect of dietrary components (protein, cabohydrate and fat) on inhibiting carcingenic lesions of  reveale that most of dietary fat synnergistically affect the process of carcinoma development.

       Works Presently engaged in:

  • Impact of mycotoxin contamination on the ntritional value of the substrates (agricultural products and their byproducts)
  • validate the use of air-breathing teleost fish as model for experimental carcinogenesis
  • Histopathological,biochemical and histochemical correlation of the alterations induced due to mycotoxin intake for early diagnosis of the problem
  • Algal toxin: characterisation of the inducing factors and biological control maesures
  • Ochratoxin induced nephropathy 0n man and oter laboratory animals

     Special Assignments

External Editor:  A book  on "MAKHANA"( Euryale ferox salisb.) containing 36 artcles, punlished by Directorate of Information and Publication on Agriculture, Indian Council of Agricultural Research, New Delhi

Author: Fish Culture,Simple interpretations of the scientific aspects of freshwater aqua culture in question answer pattern, 125 pages, in hindi, published by Anulekha Publishers, Darbhanga

 

     Research Supervision

Ph.D Titles:

  1. Ochratoxin A in Agriculture products and its impact on human health( Awardee: Bandana Verma)

Summary:

An extensive survey of some agricultural commodities stored and marketed for human consumption viz. wheat, maize, pulses, flat rice, wheat flour and vegetable substitutes ( Tilauri, adauri, Kumhrauri, Biriya, dried vavetables etc.) was conducted from local market as well as domestic sources. The reperesentative samples of eache of them were subjected to culturebfor acreening of mycobial strains contaminating these substrates in natural conditions. the fungal strains appearing on these substrates belonged to Aspergillus, Penicillium, Curvularia, Syncephalastrum and Rhizopus genera. Among Aspergillus group the strain A.flavus was recorded as most dominant mycoflora followed by A.ochraceous. The incidence of :A.niger, A.parasiticus, A.versicollr, A.ruber, A.clavatus, A.nidulanse  was also recorded

During Winter 28 strains of A.ochraceous was isolated of which 50% were found to ochratoxigenic ellaborating various amount  of ochratoxin A. Incicence of ochratoxigenic strains during summer was relativel low(37.03%). Among the samples collected from domestic sources 41.2% of the strains were toxigenic.

Flat rice appear to be the most potent substrate for the growth and subsequent ellaboration of the toxic metabolites by A.ochraceous. However,the flat rice samples collected from domestic sources were comparatively less contaminated.Maize could be recorded as second substrate of choice for A.ochraceous followed by wheat and other commodities investigated.

The amount of ochratoxin A ellaborated by the present strains ranged between 5 to <80 ug/kg. Similarly, the strains isolated from domestic sources ellaborated in range of 2 to 100 ug/ kg of ohratoxin A.

Most of the contaminated samples were encountered at retail shops. Only 9.76% of the samples collected from whole-sale depots harboured toxic spores.

The agricultural products and their byproducts are always susceptible to fungal spoilage in pre- and post-harvest stages. However, from mold contamination point of view storage has been regarded as most critical. Durind storage , various factors like source of contamination, physical condition of the  substrate and type of storage decide the degree of mold spoilage. Most of the present substrates nbesides being a good source of glucose which is readily utilised as nitrogen source by many of the fungal strains, are already recognised as favoured  substrate for growt of A.ochraceous group of fungi.

In addition to the climatic factors, water content of the substrate is a critical factor playing significant role in mycobial spoilage. In general, water content of the present substrates varied between 6-9%, which normally do not favour fungal growth. But, their increase through various means like seepage, storage on damp places, exposure to high relative humidity, makes them prone to mycobial spoilage.

Natural contamination of the substrates with ochratoxin A  depends on the genotype  of the strain and the environment in which the mold is growing. It has been reported that all the species of the A.ochraceous group can produce one or two important mycotoxins like ochratoxin A and penicillic acid.

In present study majority of the strains were ochratoxigenic and maximum number of toxic strains were isolated freom vegetable substitutes (50 %) folloed by wheat (45.4%) and maize (40%). However, the strains capable of ella borating high amount of ochratoxin were frequent on flat rice.

Bioassay of crude ochratoxin A samples obtained from mass culture of the A.ochraceous strain ( capable of ellaborating 100ug/kg) was conducted on guinea pig (6 wk old). Test groups were fed with 25ug crude ochratoxin A for 20 and 40 days respectively. Histopathological observations were made on day 60 and 120 along with the control.

Clinical symptoms were recorded as faded body color, anemic appearance, raised hairs and loss in appetite. eye color whcih starts fading even earlier beocmes prominet in advanced conditions. The

 

2..Ochratoxin A in Fish feeds and its pathophysiological manifestations in fish ( Awardee: Shambhu Prasad)

                    Summary:

 

Ochratoxin A is a nartrally occorring mycotoxin produced by some strains os Aspergillus and Penicillium genera as their secondary toxic metabolites. a comprehensive survey of fish farmer storehouses and retail shops in different blocks of Darbhanga district was carried out to determine the incidence of A.ochraceous group of fungi. Altogether 868 fish feed samples collected during October 2000  to September 2001 were screened . A.flavus, A ochraceous, :niger, A.versicolor, A.nidulanse, A.tamari, Alöternaria spp. and Penicillium genera appeared as most frequent invaders with variable degree of occurrence. Degree of  A.ochraceous infestation on these fish feed samples varied between 13.13- 22.91 % being maximum during pre-winter (13.33%). During monsoon 18.47 % samples were found to be contaminated. However, A.flavus was recorded as the most dominant mycoflora on these substrates.

Among the agricultural commoditeis , oil cakes ( Mustard, Sunflower,Linseed), wheat bran, and corn flour appeared to be substrates of preference and the climatic conditions prevalent during prte-winter months was observed to be congenial for the growth of A.ochraceous group of fungi.

Maximum number of toxigenic strains of A.ochraceous were obtained from oil cake samples. In particular, mustard oil cake and summer and monsoon months were recorded as more vulnerable for these toxic strains. The quantity of ochratoxin elaborated by these strains reflected wide variation ( from trace to 100 ug/kg). however, in general, the present toxic strains were capable of elaborating ochratoxin between 10 -50 ug/kg.

Section II of the present thesis embodies bioassay conducted on air-breathing fish, Channa punctatus. Crude ochratoxin A extract were obtained through culture of the identified toxic strains of A.ochraceous. Four groups of healthy fish were subjected to oral administration of 25 and 50 ug ochratoxin A through diet respectively and observations were made on day 30 and 60 from each group along with the control.

Early clinical manifestations were marked by increase in mucous over body surface, darkening, swollen belly, bowel disorder, dilated pupils and increased opercular movement. Surgical inspection nrevealed moderate petechial hemorrhage of internal organs and marked inflamatory reaction of kidney, spleen and head kidney in advance condition of the treatment at high toxin dose level.

Histopathologically, excretory kidney appeared to be affected most. The characteristic response of tubular degeneration involving glomeulus in advanced was well pronounced. The interstitial lymphoid tissue indicated proliferative changes . However, in liver, the lesion characterised pre.neoplastic alterations. Stomach and pyloric caeca were grossly affected. The symptoms simulated chronic atrophic gastritis and acute gastritis respectively.Poorly defined adenocarcinoma arising from columnar epithelium cells of gastric mucosa was also marked .

A significant loss in interstitial lymphoid tissue of head kidney due to edema and focal autolytic degeneration constituted prominent findings but the chromaffin cells indicated moderate hyperplastic proliferation. In spleen, gross swelling of the organ was attributed to monotony of the lymphocytes/ lymphoblasts that intended to repalace all native cells. Distinctive appearance of spllenic follicles were completely lost.

the gonads respoded differently. While the ovaru refelected gross toxc response leading to atrecia and autolytic degeneration in different stages of maturity, the testis, in cotrast, was relatively less affected. Moderate autolytic degeneration of interstitial cells was marked in testis. No gross lesion in muscle and gills were marked.

 

 

3.Mycobial feed contaminants and their histopathological manifestationf in a major carp (Awardee: Meenakshi Das)

 

Summary:

 

Section I of the thesis deals with the mycobial contyaminants infesting fish feeds in general marketing and domestic storage conditions. Altogether, 1899 samples of fish feed ingredients like mustard oil cake (MOC),ground pulses mixture (GPM), rice bran (RB), rice flour (RF),wheat flour (WF), and wheat bran (WB) were collected during summer , winter and monsoon months of 2000-2001. The samples were visually screened for fungal spoilage. depending upon  the physical condition , the samples were categorised as fresh (F), partially infected (PI) and infected (I).

A homogenous mixture of the all the samples were subjected to culture for appearance of fungal flora infesting the substrates. the study was conducted with special reference to aspergillus group of fungi. The strains idetified from these substrates included A:flavus, A.parasiticus, A.ochraceous, A.niger, A.versicolor, A.ruber, A.fumigatus,A.candidus, Penicillium viradicatum and P.islandicum. The samples collected from retail shops were highly contaminated.

Investigations suggested that many of the samples appearing fresh visually harboured spores that appear on the substrates following culture. Among the strains invading the present substrates, A.flavus is well documented for its toxic and carcinogenic properties. Thus, A.flavus aws selected for further studies.

Incidence of A.flavus on the present substrates was variable in different season and on different substrates. Most of the present toxic strains were obtained from mustard oil cake and ground pulses mixture. Qualitative and quantitative analysis of the toxigenic potentials of the present strains revealed that most of the strains were capable of elaborating aflatoxin B1 and B2 only in amounts ranging between trace to 100 ug/kg. Section I, thus indicates about the gravity of the problemof mycobial spoilage of agricultural products and their byproducts incorporated as fish feed ingredients.

Section II embodies the result of bioassay conducted on a major carp, Cirrhinus mrigala by using crude aflatoxin extract through mass culture vof identified A.flavus strain in laboratory. Four groupf of ten healthy carps were fed with diet containing 25ug crude aflatoxin extract and were ethuanized for histopathological observations on day 10, 20, 30 and 40 alongwith the control.

Morphological features of crude aflatoxin poisoning was characterised by darking of the body color, increse in mucous over body surface, listlessness and dilated pupils with faded eye color. Anatomically, moderate hemorrhagic patch along the liver margins and localised inflamatory reaction associated with hyperamia constituted characteristic early response while in advance condition gross swelling of the liver and kidney and somtimes stomach as well, was marked.

Histopathological observations revealed that besides the routine toxicological response in all the present organs like liver, kidney, stomach and intestine, characterised by degenerative changes , the toxin induced carcinomatous alterations too. In liver, preneoplastic condition leading to trabecular hepatome was clearly marked. The kidney indicated onset of alterations that may ultimately be defined as adenocarcinome arising from tubular epithelial cells.However, the early pathological manifestations are triggered in lymphoid tissue.The autolytic changes in stomach and intestine were also marked but they involved mainly the mucosal layer. Submucosa, in contrast, aws hyperactive in bothe the cases (stomach and intestine) Differentiation of atypical glandular masses replacing all the native structures in submucosa was clearly marked. The condition was diagnosed as adenocarcinoma arising from columanar epithelial cells of mucosa.

Establishment of different forms of the described lesions , however, demand prolonged incubation of the present fish at the similar dose level. The carp, C.mrigala,thus appears to be a sensitive model for toxicological studies.

 

4.Dietary factors and aflatoxin B1 carcinogenecity in fish (Awardee: Rakesh Kumar Sinha)

 

Summary :

Reprts of hepatoma epidemics in parts of the world incriminaqting aflatoxins in its etiology and successful induction of comparable carcinogenic manifestations in rainbow trout, Indian major carps and air-breathing teleosts cofirmed that aflatoxins as the most devastating intruder in thefood chain of fish as well as human beings.

The fish culture for economic purposes need to be provided supplementary diets and the agricultural products and their byproducts form the bulk of the fsih feed formulations. Almost all of the items incorporated in fish feed are good sobstrate for mycobial spoilage. Array of literature explain mycotoxin contamination of these products and subsequent adverse  affects on the health of the cosumers.

Impact of dietary intakes on internal physiology of the living system either progressive or retrogressive hardly needs any explanation. Thus, the present work has been carried out with the assumption that a fish population reared on a diet with special biochemical profile is bound to refelct specific response. If the fish has been reared on diet contaminated with aflatoxins the toxigenic / carcinogenic manifestations may be induced od suppressed under the influence of the dietary intakes.

the present problem broadly covers three aspects : mycotoxin contamination of food and feeds, supplementaru dietary items in fish culture operation and their vulnerability to mycobial spoilage under prevalent climatic conditions and finally the organism-feed interaction. All these three aspects have been discussed in the Review of Literature.

A group of 240 healthy fish divided in three test groups of 60 each designated as "Diet group"depending on the three types of diet presently used. Each diet group was sub-divided in test group of 20 fish each to be administered with 15, 30 and 40 ppm of crude aflatoxin B1 intraperitonially throug propylene glycol. Observations were made on day 15, 30 and 45. at the end of each incubation period five fish from each group was selected at random along with the cotrol for anatomical and histopathological observations. Tissue were fixed in Bouin`s (alcoholic) and 5 u sections were  stained in H & E.

A tabular presentation of alterations in morphology and gross anatomy, frequency of gross pathological alterations , histopathology of liver. kidney, stomach, intestine and pyloric caeca, cytology of hepatocytes, measurement of cell and nuclear size and their nucleocytoplasmic ratio have been presented separately at different dose levels and diet. Histopathology have been adequately supported with photomicrographs at different magnifications.

test fish of all the test groups receiving variable dose of the toxin were invariably lethargic on day 30 onwards wit increase in mucous over  body surface and darkening of the body color. The clinical symptoms refect a positive correlation with increase in dose of the toxin and treatment period. However, the group reared on cottonseed oil cake supplement appeared to be the worst victims.

Similarly, gross anatomical alterations were prononced in the group of fish reared on cottonseed cake followed by mustard oil cake and soya flour spplement.In general, the anatomical alterations were observable at higher dose level in form of hemorrhage/hemorrhagic patches associated with localized swelling and hyperamic areas. Loss in consistency of the internal organs was distinct. The symptoms involved liver, viscera and visceral fats,excretory kidney. In parts of gastriintestinal tracts the symptoms were partially marked.

Histopathology of the liver revealed that in C.punctatus, an air-breathing teleost, symptoms like hyperamia, hemorrhage, cytoplasmic vacuolization, necrosis and bile duct proliferation characterrise the toxic response of aflatoxin B1 following 45 days of incubation. However, some concomitant findings include gross swelling of the hepatocytes, sinosoidal dilation, appearance of pale cell foci and affinity of the cytoplasm towards eoisinophilic stain. In fish subjected to 40 ppm of aflatoxin B1 symptoms characterizing onset of cirrhotic alterations were marked.

In the excretory kidney, the interstitium was edematous with signs of hemorrhage, inflamation and necrosis in early stages.In advance conditions the interstitium becomes hyperchromatic involving columnar epithelial cells with irregular nuclei and infrequent mitosis. Thickening of the basement membrane of the glomeruli and arteriosclerosis was also marked.

The histopathology of the stomach, intestine and pyloric caeca has been discussed under one head. The ealy lesion refelct hyperamia, hemorrhage, edema, necrosis, atrophy and mucosal sloughening characterising features simulating gastritis in man. the advancing lesion appear the columnar epithelium become hyperchromatic in nature and small glandular masses appear to differentiate in the submucosa.

There is paucity of literature on effect dietary components specially in fish subjected to aflatoxin intoxication. However,the present findings suggest that intake of dietary cotton seed cake that contains cyclopropenoid fatty acids (CPFA) increases the toxic effects of aflatoxin B1 apparently in comparison to diet based on mustard oil cake and soya flour.

Anatomically the chronology of the toxic effect in C.punctatus following aflatoxin B1 treatment and subjected to different dietary regimes may chronologically be expressed as :

                                             Cotton seed cake < mustard oil cake < Soya flour

 

5.Influence of dietary habits on lipid profile of some freshwater fish (Awardee: Deepika Srivastava)

 

Summary

 

Ph.D underway:

  1. Incidence of coronary heart diseases in relation to lipid profile ( Sharad Chandra)
  2. Fish Biodiversity in the Terai Plain of Eastern nepal ( Prakash Kumar)
  3. Eutrophication of ponds and its impact on freshwater aqua culture (Rupum Kumari)
  4. Studies of behavioural and hematological response to Mancozeb and Metalaxyl toxicity in the freshwater air-breathing fish, Channa punctatus( Farhat Zabeen)

M.Sc Desertations:

  1.  Studies on the Butterfies of Darbhanga
  2. Ecology and fisheries of a swapm of Darbhanga
  3. capture fisheries of madhubani. Trends in annual landing
  4. Isolation and characterization of bacterial pathogens in a freshwater fish
  5. Phytoplanktons in a fish culture pond of darbhanga

Special Training

  • CAS training Programme on  ADVANCED TECHNIQUES IN FISH IMMUNOLOGY  organised by central Institute of Fisheries Education Versova ,

 

 

        A brief Report on the Major Research Projects Completed

Project one

Mycotoxins: Histopathological Manifestatations in Fishes and Possibilitie of Human Uptake

F.No. 4(4)/97 ASR 1, A P Cess Fund ,ICAR, New Delhi

Summary

  1. During 1999-2000, starting from November, altogether 1774 samples were collected . This contained mustard oil cake (375), ground pulses mixture (380),rice bran (390),soya flour (25),rice flour (175),wheat flour (201), wheat bran (178) and formulated commercial rations (50)
  2. Out of the total (1774) only 894 samples appeared to be normal visually, but most of them like 45.8% of the winter, 50.7% of the summer and 72.2% of the monsoon samples developed fungal colonies following incubation increasing the the degree of mycobial spoilage upto 71.9%. The samples,thus , appearing normal in look too, were contaminated with varieties of spores that developed fungal colonies under favourable conditions.
  3. In general, water content of the substrate range betwwen 5-12 % during summer exhibiting a gain of 3.6% during winter and 5.3% during monsoon. Moisture gained by the substrates during monsoon gets hardly some chance  to loose during winter and is retained all through till rise in atmospheric temperature (following summer). Brsides, unplanned storage under poor sanitory conditions and low ventilation causes the internal temperature of the substrate to rise and provide a congenial range favouring sporulation, growth and subsequent toxic metabolite production.
  4. During sommer low RH and relatively low water content of the substrate apparently check the spoilage. However, during survey , many prevalent practices were observed that do help the substrate to gain moisture at an accelerated pace worsening the spoilage scenario of the substrates concernned.
  5. The climatic factors like atmospheric temperature and RH are beyoned control and infestation of the substrates also can not be fully checked , since the agricultural commodities from sowing to final storage are always at risk of contamination .B ut the water content of tye substrates before short term/ prolonged storage can be brought down to quit at a "low-risk range" by adopting simple "sun drying" in a proper way. The substrates handled with care in pre- and post-harvest conditions and stored at 5-6% water content level faces minimum risk of mycobial spoilage and subsequent human/ pet health hazards.
  6. Beside the present strains ( A.flavus,a.parasiticus,A.ochraceous,A.versicolor, A.rubur and Penicillium viridicatum) a number of other species, many of them well recognised for their toxigenic potentials, like A.niger,A.fumigatus, A.candidus,penicillium islandicum, Rhizopus sps., Mucor spshave been found to be common invaders on the substrates under investigation.
  7. Maximum number of aflatoxigenic strains of A.flavus and A.parasiticus were isolated from mustard oil cake (MOC). Wheat bran and wheat flour appear to be substrate of choice for ochatoxin A producing strains like A.ochraceous and P.viradicatum. In contrast, none of the A.versicolor strains obtained from MOC was positive to sterigmatocystin except one ( showing trace only). Incidence of A.versicolorwas high on ground pulses mixture.
  8. Altogether 51.1% A.flavus,38% of A.parasiticus and 50% of A.ruber appeared to be aflatoxigenic in quantities raqnging between 2-100 ug/kg. Similarly,42.8%of  A.ochraceous  and 37.5% of P.viradicatum were ochratoxigenic in variable quantities (15-50 ug/kg). Thirty five per cent of A.versicolor bore capabilities of producing sterigmatocystin (>8 to <10 ug/kg)
  9. Among air-breathing teleosts  H.fossilis appears to be most sensitive to all the three mycotoxins (aflatoxins, ochratoxin A and sterigmatocystin) followed by Clarius batrachus. Anabas testudineus tolerated the mycoxin insult fairly well in all the three cases. In major carps L.rohita  was more susceptible than Catla catla.
  10. Out of the three toxins tested , crude extracts of aflatoxins reflected highest toxic potentials followed by ochratoxin A. the toxic response of sterigmatocystin was delayed indicating low toxic potentials of the present A.versicolor strains.
  11. Morphologically ,the fish exhibit darkeningh of body color, increased mucus over body surface.The behavioural response of the fish become apparent following 30 days of incubation and onwards gradually becoming more and more pronounced . In early stages the treated fish are usually lethargic and exhibit loss of appetite. In advanced condition of the treatment they are listless ( easy to capture), refuse to feed, show poor taxis  and increased surfacing and opercular movement.
  12. The symptoms characterise chronic aflatoxicosis. In ochratoxicosis the fish frequently exhibit swollen belly with moderate to severe bowel disorder along with the symptoms listed above.
  13. Anatomically, all the fishes exhibit anemia and diffuse hemorrhage on liver, viscera and visceral fats. Locallised swelling with hyperamia was marked on the liver, kidney, intestine. In advanced condition, gross swelling also involved spleen and head kidney .
  14. At haematological level, significantly elevated leucocytic count dominated by immature lymphocytes , significant decrease in Hb content and total erythrocytic count constituted the major findings.
  15. The biochemical findings suggest that treated fish are faced with a serious metabolic crisis due to strong hyperglycemia and hypercholestererolemia with decrease in serum protein content. Manifestations   of hypermetabolic stage is obvious but it also indicate the ability of the test fishes to mobilize sufficient resources to cope with the stressfus  condition.
  16. At histopathological leve, characteristic trend of chronic hepatotoxicity of aflatoxins and nephrotoxicity of ochratoxin was prominantly marked in all the air-breathing and major carp models. Besides the toxin induced carcinomatous alterations in the gastro-intestinal tract. Gonads were also severly affected. Among air-breathing fish sensitivity was marked in descending order as H.fossilis<C.batrachus<A.testudineus. In major carps L.rohita was more sensitive than C.catla.
  17. The mycotoxin following entry into the fish systemthrough repeated dietary intakes intend to bio-accumulate (approximately upto 1% of the exposure level), their concentration gradually piles up, but their tissue clearance rate is also very high. They also find route via excretory pathways. If a fish has been exposed to a single acute dose, the trace of the parent compound goes beyoned detectable level within 20-25 days of the administration. But the property of ochratoxin A to bind with protein molecules provide avenue for further inveestigation.

 

Recomendations

 

             Toxins of mold origin are global problem but the climatic conditions of the Indian subcontinent makes it a congenial ground  for micro-toxic fungi. Fungal contamination of food and feeds are usually air-born and an array of toxic spores are  there in the environment, compteting with each other to infest and dominate over the substrate. Though, a plethora of information is available on different aspects of mycotoxins including detoxification but the most sensible point at which the  mycotoxins should be tackled lies in handling,harvesting,processing and storage of food and animal feeds. Here a thorough knowledge of ecology,physiology and taxonomy of molds become relevant. The following are some suggestions both from scientific and practical standpoint that may help to combat the mycotoxin problem in aquaculture sector but with the consideration that mycotoxin contamination is a recurrent and inevitable phenomenon. We can not eliminate them completely from our food and feeds.

SCIENTIFIC

  1. There is a need to facilitate wide survey programmes for identification of incident toxic microEfungi and their ecotoxicology in each agro-climatic zone of the country.
  2. Each commercially viable fish species produced through feed-based culture need to be evaluated for their toxic response against mycotoxins. Studies on immunisupressive potentials of mycotoxins should be given priority over other toxicological parameters.
  3. Metaboic transformation of parent mold metabolites inside the fish system,their trace in edible portions(bioaccumulation, tissue clearance rate, toxin-tissue ratio,clearance through excretory pathways etc.) demand further investigation through elaborate but precisely planned experiments. This will provide valuable information helpful in certifying "Safe Consumability" of aquaculture products.
  4. Specific  and sensitive analytical methods for detecting mycotoxins in fish flesh needs to be workedout.
  5. A national network of scientific facilities in existing research laboratories (with defined copverage area) to undertake "sample check programme" for different mycotoxins in the samples collected from the field by the farmers/middle man/concerned workers on no profit basis should be established.
  6. Above all is the establishment of a "National Aquaculture Feed Quarentine" that must be designed (by a committee of experts), imposed and monitored with utmost care and strict surveillance.
  7. Among a few chemical methods for decontamination of aflatoxins ammoniation of oil seed cakes, the chief fish feed ingredient, appears to be effective and innocuous.The method may be recommended for fish feed industries (not a part of the present investigation).

PRACTCAL

  1. It is important for the fish feed manufacturers to buy a raw product of known history ( time of production,processing details, storage specifications) after thorough visual inspection for mycobial spoilage. A grower may accordingly be advised to use resistant varieties of crops and adopt standard processing methods. A product with high water content (not properly dried) must be avoided.
  2. The raw feed ingredients or processed feed should be stored/ packed at minimum moisture level and in packing materials that are quite effective against moisture. Infected portions of the commodities, if any, must be eliminated by hand picking as far as possible.
  3. Prolonged storage, in any case, must be avoided. However, sun drying of the feed ingredients/ feeds, if possible, should be made a regular practice but not in conditions of high realtive humidity. Re-wetting of the subsstrate is detrimental.
  4. Feed before final application must be inspected thoroughly for mycobial spoilage
  5. Creating general awareness among people concerned with aquaculture practices through popular means hold the key value. This may be achived by utilising govermental agriculture extension services, public health services and persons related to food and nutrition networks. These extension workers should be sufficiently trianed on mycotoxin problems (incidence on food/feeds, probable hazards, practices facilitaing mold spoilage, inspection methods,sampling for mycotoxins,check programmes and applicable ways for prevention and control) and be instructed to create due awareness primarily among those related to farm and rural families in matters relating to feed, feed preparations,storage, preservation,sanitation and hygiene. Similarly, the farm organisations ,cooperatives, trading bodies may also be trained and be instructed/provided materials (in form of pictures,posters) to create awareness.
  6. Incorporation of mycotoxin problem as teaqching subject in graduate, post-graduate and professional courses that relate to aquaculture may also be an effective measure to combat the problem.
  7. Artificial feeding ,at least one month before crop harvesting, should be stopped.

 

Project Two

BIODIVERSITY OF BLUE-GREEN ALGAE AND ECOTOXICOLOGY OF Microcystic aeruginosa IN FISH CULTURE POND OF DARBHANGA

F.No. 4(25)/2002 ASR I, Project code 0614027, Dated 04.02.2004, A P Cess Fund, ICAR, New Delhi

Summary and Conclusion

  1. 1. The water bodies of Darbhanga (North Bihar) provides favourable condition for the growth of wide range of Cyanobacterial biodiversity. Altogether 187 species of Cyanobacteria belonging to 45 genera was recorded from 15 fish culture ponds.The number of species recorded from each pond was quite variable, probably, depending upon the ecological status of the specific water body. Maximum number of cyanobacteria were recorded during spring (109. sps) followed by monsoon (68. sps.), winter (66 sps.) and summer (45 sps.)
  2. Study of physico-chemical features of the present ponds studied in three groups ( ponds having history of recurrent algal blooms[A], those having trace of toxic bacteria but bloom never occured [B] and the ponds in which toxic bacteria were absent [C] ) revealed that the condition which favour growth of M.aeruginosa may be outlined as : Depth : above 2.5m  ; Temperature:22-35°c; Transparency:150-158 cm ; pH 9 and above ; Redox potential-501-582 mmhos/cm ; Co2-2.0 to7.0 mg/lit ; Iron-0.48 mg/lit; Sulphate-117-127 mg/lit ; Mangnesium-150-198 mg/lit; Phosphate-0.25 to 0.65 mg/lit; Total Phophorus-0.42 to 0.65 mg/lit; Total Nitrogen- 3.5 to 5.5 mg/lit; Ammonia- 0.01 to 0.09 mg/lit. It was thus, apparently marked that low concentration of  ammonia and nitrate and trace of nitrite favours blue-green algal blooms. The physicochemical factors are highly variable under multiple influences including climate. A congenial range , as soon as obtained in any water body, the dormant population of microcystis starts proliferating to attain bloom like condition.
  3. In natural conditions, population density of M.aeruginosa was maximum in five ponds of group ´A´having history of recurrent algal blooms. The maximum density was recorded as 38 x104 cells/ between May-June. In group B ponds representing trace of cyanobacteria but bloom like situation never encountered record of the population density varied between 0.5 x 104  to 2.0x 104cells/cm3. In the third group C, microcystis population showed a very low count. On the basis of color appearance of the water a color a chart has been developed which will enable even a layman to assess the population density of microcystis in his pond through comparison.( Refer link)
  4. In vitro culture of any micro organism requires a suitable medium which not only ensures proper growth of the organism but also provides opportunity to investigate into the the various physiological and metabolic activities of the organism. In case of M.aeruginosa, any dfined medium ensuring their growth is still not recommended. Besides, the two - ASM and Hughes media, 8 different formulations ( five of them modified and three new formulations) were tested. Of these, the BRL-III (Biotoxin Research Lab. III) supported a luxuriant growth repesenting a record  in vitro population of 10.86 x 104 to 12.3 x 104  cells/ m3 within 50 days of incubation. This media is modified form of Hughes by supplementing low nitrate (0.15 mg/l). It works exceptionally at pH 9.5.
  5. In the life cycle of microcystis the transitional stage between initial stationary phase and logirathmic phase is critical. The cells are usually very sensitive to unfavourable conditions. Thus , the period appears to be a potential phase during which control measures may effectively be taken up.
  6. Toxicity test on Daphnia magna subjected to static exposure of different concentration of  Microcystis aeruginosa toxic extract  indicate the present strain to be potentially toxic. The extract obtained from 35 day old culture was less toxic than those obtained from 150 days old culture. In the later case , 0.5 ml of the extract caused the death of 25% population of the test animal. The death record increased with increase in concentration of the extract reaching upto 76%  at 2.5 ml concentration within 1 hour of the exposure.
  7. Both air-breathing and major carps subjected to bioassay against microcystis endotoxin showed sign of lethargy, respiratory distress,low to excessive mucus secretion, gray spots on the lateral side of the body and muscle fasciculation. The air-breathing teleosts, however, appear to be  more tolerant to the toxic insult in comparison to major carps. In the major carp group, L.rohita was more susceptible than C.catla.
  8. At histopathological level micystis toxin grossly affects the liver. Vauolar degeneration and moderate necrosis being the chief symptoms. Edematous hypertrophy of the epithelial cells of the gill and cloudy swelling of muscle fibers with lymphocytic infiltration characterises microcystin poisoning. The gonads also become grossley affected. The stomach and intestine reveal necrosis of the columnar epithelial cells with sign of hemorrhage. The spleen, however, appeared normal.
  9. Ochromonas danica, a non-colonial, freshwater, flagellate protist was found to be a microcystis colony cosumer in field conditions.
  10. In vitro studies by introducing O.danica in microcystis culture (21.4 x 104 cells/cm3) the population was reduced to 0.5 x 104  cells/cm3 followin 35 days of incubation. The findings suggests that O.danica may be used as potential tool for biological control over microcystis bloom.
  11. It is beyoned doubt that the nutritional enrichment of the waterbodies and subsequent  eutrophication is accelerated due to anthropogenic activities leading to increased productivity of phytoplanktons. It causes a shift in the phytoplankton community toweards cyanobacteria and cosequent bloom during favourable climatic conditions.
  12. Experiments coducted on control measures suggest that the 25% simazine (90%) which is virtually non toxic to aquatic and pet animals may be used for chemical control of microcystis blooms.
  13. The N:P ratio is a key factor regulating development of microcystis bloom. Therefore, the management programmes to balance N:P ratio by decreasing P supply and addition of inorganic nitrogen above certain threshold  concentration is desirable.
  14. Ammonia , when used to control microcystis bloom favours slight increase in algal population initially, thereby showing a gradual decline.

 

Project Three

BIO-ATTAINMENT OF PEAK NUTRITION VALUE ( PNV) IN fISH

F.No.4(24)/2002 ASR I, Project code- 0664003, Dated 03.03.2004, A P Cess Fund, ICAR, New Delhi

 

Summary and Conclusion

  1. Making up of the biochemical profile of the fish is closely related to feed intake, behaviour, sex and changes in the climatic conditions. A substantial normal variation is observed in biochemical constituents of the fish muscle. In particular, the lipid component shows a greater variation.
  2. The prevalent climatic factors in the present area of investigation provides two congenial ranges for the growth of the fish: first, during January to June the second during October to middle of December. The first, on account of longer duration of four months has been reffered to as "period of maxima" and the following as the "period of minima" respectively.
  3. In this geographical area July to September  is a period of recurrent flood inundating all the waterbodies .This period characterise ingress of new water , low food level and severly altered physicochemical conditions.
  4. The major carp L.rohita subjected to traditional polyculture (TPC), semi intensive culture (SIC), and intensive culture (IC) gained weight almost double to their initial record within four months (Jan. to May). The pre-monsoon rains and the following monsoon months affect gain in weight significantly. During November onwards a wide gain in weight was recorded with a brief latency period during extreme of winter. The variation was more apparent in control group probably on account of food security in laboratory cultures and safety from climatic turbulences.
  5. The control fish, however, exhibit remarkable gowth in comparison to the experimental group during period of maxima due to abundance of natural food in form of phytoplanktons.
  6. Variation in the pattern of weight gain in fish subjected to three culture schedules was well marked as :  Intensive culture > semiintensive culture> Control> Polyculture.
  7. The impact of mustard oil cake ,coconut oil and sunflower oil used as dietary supplement in all the three culture conditions was also obvious and may be represented as: Sunflower oil > Mustard oil> Coconut oil.
  8. The fat content of the fish  appear to be dependent on food intake.The first peak of fat deposition is observed during June-July and the second, between September to December corresponding the periods of  maxima and minima.The level of added fats deplete reasonably with onset of maturation and practically attains a "no gain" stage or even less in periods of food scarcity.
  9. Average gain in iodine value of fat in fish reared on mustard oil supplement was recorded between -4.0 to +8.0 (TPC) ; -3.3 to +9.0 (SIC) ;and -1.7 to +10 (IC) in comparison to control . Similarly, in the second group group of fish reared on coconut oil supplement the iodine value ranges between -14 to +8.0 (TPC); +1.6 to +10.0 (SIC) and -2.0 to +9.0 .In the group of fish reared on sunflower oil supplement the iodine value ranged betwwen -14 to + 8.0 (TPC);  +2 to+32 (SIC) and -8 to +25 in IC group.
  10. Similarly, in the case of mustard oil the serum triglyceride ( mgm%) ranged between  -6 to +23 (TPC); +3 to +34 (SIC) and +1to +30 (IC). In case of coconut oil it ranged between  -19 to+23(TPC), +2 to +32 (SIC) and -8 to +25in IC group. In the third experiment with sunflower oil the serum triglyceride values varied between +2 to +28 (TPC), +4 to +30 (SIC) and +1 to +29 (IC).
  11. Thus, in terms of I value and serum triglyceride the SIC group again reflect age over  other culture systems and sunflower oil was marked as best dietary supplement in comparison to others repesented as :  Sunflower oil > Mustard oil > Coconut oil.
  12. In accordance to the effectof availabilty of food on growth and lipid/ serum trigyceride profile , as mentioned earlier , the essential fatty acid profile with special reference to omega 3 and 6 fatty acids are also affected. Rather , it may be inferred that  adverse climate that affect the food intake is finally reflected in EPA profile of the fish. The present fish, L.rohita has a rich profile of omega 6 fatty acids than omega 3. But it show a marked fluctuation in relation to availabilty of food vis a vis climatic conditions.
  13. In view of the facts it is quite imperative to determine the PNV ( Peak Nutrition Value) of fish with reference to the growth pattern and coinciding level of serum triglyceride because, it is this simple lipid class that consists all fats and oil. Besides, the method to determine serum triglyceride is simple and available widely ( in Clinics) . In contrast, determination of iodine value, though very simple, but authentic result demands analytical grade iodine monochloride which is very costly. 
  14. In light of the above findings it appears to be right choice to opt serum triglyceride as PNV/ Age Marker of the fish because: 
    • Rise in serum triglyceride also ensures an optimum protein content of the muscle
    • It is sensitive to seasonal/ climatic influences
    • It incorporates all "good fats" from human consumption point of view
    • Methods of analysis is widely available even in all diagnostic clinics    
  15. In the case of  Clarius batrachus, an air- breathing carnivorous teleost, gain in fat content of the fish is similarly comparable to that of weight gain. It becomes almost static for two-three months during monsoon and start rising again till maturity. The period of minima , due to abundance of zooplanktons in natural waterbodies, is significant for the growth and subsequent  enrichment of biochemical profile of this fish   .
  16. The iodine value of thefat shows a different pattern in comparison to major carp. It maintains the pace of gradual piling even during monsoon and the depletion is marked only on onset of physiological events leading tomaturation, whereas the serum triglyceride depletes during monsoon.
  17. The effect of feeding habit was well marked in EFA profile of this carnivorous fish characterised by relatively persistent level of omega 6 fatty acids.

 

Publications

              1.S.K.Verma and S.Pandey. 1982. Aflatoxin contaminated fish feeds. Proc.Indian Sc. Cong. p 87 (abstract)

2.S.Pandey, S.K.Verma and R.A.B. Verma, 1985. Effect of aflatoxin B1 on some organs of an air-breathing teleost,Channa puctatus.    Paper presented at the fourth Symposium on Fish Physiology. Goteborg, Sweden. In Proceedings p 61.

Abstract :

Among all the natural contaminants of fish feeds, aflatoxin  warrant special attention. Aflatoxin B1, the secondary metabolites of Aspergillus flavus and Aspetgillus parasiticus, has been established to be most potent carcinogen for a number of animal species including man. The presence of such carcinoogen in fish feeds is thus detrimental to the fish population. the probablity of this toxin being transferred to man through fish flesh could be a major health hazard.

Channa punctatus, an air-breathing teleost, was selected for this study. 0.5 ml of 10,20,30 40 ppm of aflatoxin B1 in propylene glycol was admonistered intraperitonially. Fish from each groupm were sacrificed at the interval of every 20 days along with the control.

Tumorous outgrowths were distinct more on the liver than on the kidney. However, no such change was observed on the intestine, stomach, spleen , gill and pyloric caeca. The main effects on the liver included necrotic changes in hepatocytes showin evidence of hepatitis mostly around the central vein. Stages simulating hepatoma were also observed. Increase in cell size and nuclear size as well as nucleocytoplasmic ratio in the kidney indicated some malignant changes which could lead to adenocarcinoma. Varying degree of degenerative changes was observed in the spleen, stomach, intestine , pyloric caeca and gill.

Key words : Aflatoxin B1, Fish pathology, Aflatoxicosis, Hepatoma

3.S.K.Verma, S.Pandey, R.A.B.Verma and S.K.Verma,1986. Mycoflora associated with contaminated fish rations and aflatoxin B1 contamination (Abstr.), In Proceedings of National Seminar on Fish and their Environment, Haridwar, P 27.

4.S.Pandey,S.K.Verma and R.A.B.Verma, 1987. Effect of aflatoxin B1 on inter-renal tissue of an air breathing teleost, Channa punctatus. Paper presented at the First Congress of Asia and Oceania Society for Comparative Endocrinology. Nagoya Japan. In Proceedings P 312-313.

Abstract:

Several carcinogenic agents induce neoplasia in fish. aflatoxin B1, a mycotoxin, led to to hepatic neoplasm in the rainbow trout. However, the effect of aflatoxin B1 inducing neoplasia in the interrenal tissue of fish has not been reported earlier. the present investigation provides information on this score.

four groups of 25 fish were injected with 40 ppm of crystalline aflatoxin B1 in 0.5ml propylene glycol by itraperitonial route. the control fish were injected with propylene glycol only. While the experimental groups were sacrificed on day 20, 40, 60 and 80, the control group was sacrifices on day 80 for histopaqthological observations. The interrenal tissue was fixed in Bouins ( alcoholic) for routine histology and the 6 um sections were stained in H & E.

In 23% experimental fish swelling and haemorrhage were apparent in the head kidney on day 40 onwards. The inerrenal tissue revealed hyperamia, reduction in haematopoietic tissue and edema. the foci of pale cells become distinct. these symptoms pronounced as the period of treatment was extended. a diffuse necrosis and hyperplastic interrenal cells were observed in 13.2% of fish (day 60) and 16% of fish (day 80). Four out of 25 fish had preneoplastic changes leading to carcinoma characterized by atypical compact cells with vesicular nuclei aroun dilated sinusoids and blood capillaries. A few cells with displaced nuclei like "signet-ring stage" suggests anaplastic nature of the lesion. The characteristic feature of well differentiated carcinoma (metastatic invasion to blood vessel and numerous mitotic divisons in affected tissue) were not clearly marked. The interstitial lymphoid tissue was dominated by blast and mature lymphocytes. A slightly elevated peripheral lymphocyte count in blood is indicative of some significant alteration.

The control fish, on the other hand , did not show any of the above changes.

Fourty ppm of aflatoxin B1 (I.P) is a significant dose to induce carcinoma iv Channa punctatus. However, induction of well differentiated interrenal cell carcinoma requires longer treatment which is being persued.

key Words : Aflatoxicosis, Interrenal cell carcinoma, Channa punctatus, Aflatoxin B1

 

5.S.K.Verma,S.Pandey and R.A.B. Verma . 1988. Effect of aflatoxin B1 on the liver of an air-breathing teleost,Channa punctatus. Paper presented at National Symposium on Animals ana Aquaculture, Kalyani, West Bengal, In Proceedins P 59.

Abstract :

The remarkable biological and morphological similarities between the neoplasm of fish and higher vertebrates render them useful model for copmparative study oc cancer. The effect of aflatoxin B1 on the Indian air-breathing teleost, Channa punctatus has been investigated in this regard.

Four groups of 25 fish were injected with 10,20,30 and 40 ppm of aflatoxin B1 in 0.5ml propylene glycol intraperotonially. The control group was injected with 0.5my of poropylene glycol only. Experimental groups were sacrificed on day 20,40,60 and 80 for histopathological observations along with the control.The tissues were fixed in Bouins (alcoholic) and 6 um sections were  stained with haematoxylene and eosin.

Non neoplastic symptoms appearing in liver included periportal and centrilobular necrosis, swelling, edema, hyperamia, hemorrhage, sinusoidal dilation, vacuolization and toxic liver cirrhosis. the neoplastic changes appeared at 20pp/80 day level prominantly in cirrhotic liver. However, ythe classical form of trabecular hepatoma was observed at 40ppm dose level. The malignant nature of the lesion became apparent due to envasive erosion of adjacent tissue structures and trace of malignant hepatocytes within the vascular system.

Key Words : Trabecular hepatome, Toxic live Cirrhosis , Fish patholgy, Aflatoxicosis.

6.S.Pandey and S.K.Verma, 1988. Toxigenic fungi associated with fish ration and their effect on the histopathology of Channa punctatus. Biojournal vol.1,No.2 P 113-118.

Summary

7.S.K.Verma and Sadhana Pandey, 1990.Studies on fungi contaminating fish feeds during storage condition. Biojournal , Vol.2,No.1 P163-170.

Abstract :

A general survey of 280 sample of  three types of fish feeds was carried out at block level in the disrict of Darbhanga to study the incidence of fungal mycoflora contaminating these substrates during storage condition. Studies revealed an alarming level of Aspergillus group of fungi dominated by Aspergillus flavus (during monsoon) and A.ochraceous ( during summer and late winter) on these substrates. A.nidulanse , A.rubur, A.parasiticus, A.tamarii and A.terreus showed a seasonal occurrence on different substrates. Fungi other than Aspergillus sps.dominating the substrate included Rhizopus sps. and Penicillium sps. Infrequent occurrence of Alternaria sps.,Syncephalastrum sps. and Curvularia sps. was also marked.

Key words:Mold Contamination , Fish Feeds, Storage Fungi

 

8.S.K.Verma and Sadhana Pandey, 1990.Effect of aflatoxin B1 on the haematopoetic tissue of an air-breathing teleost, Channa punctatus. J.Zool.Soc.India 40(122), P79-87

Summary

During recent years studies in fish neoplasia has met a sporadic growth provbablydue to the fact that aquatic animals are sensitive to carcinogenic stimuli. The present studies are designed to test and validate the use of Channa punctatus as a model for studies in oncology. A comprehensive report on hepatocarcinogenic potentials of aflatoxin B1, a toxic mold metabolite, in some species ofrainbow trout and an air-breathing teleost has been presented ( Halver and Mitchell,1967; Halver, 1969; Pandey et.al.,1985; Pandey et.al., 1988; Verma, 1988). However, the effect of aflatoxin B1 inducing neoplastic condition in hematopoietic tissue has not been earlier reported.

Four groups of 25 fish were injected with 40ppm pure crystalline aflatoxin B1 in 0.5ml propylene glycol. The control fish were injected with 0.5ml propylene glycol only. The experimental fish were sacrificed on day 20, 40, 60 and 80 alongwith the control. The tissue were fixed in Bouins (alcoholic) and 6 um sections were stained in H&E.

In teleostean fish haemopoiesis is associated with the interrenal, spleen and submucosa of the gut. the spleen and interrenal were swollen in most of the cas4s following 80 day treatment. The histpathological observations of the spleen revealed a monotony of the lymphocytes with some cells containing atypical nuclei. The interrenal showed dominance of atypical lymphocytes alongwith severe proliferation of interenal lobules. The symptoms simulate lymphocytic leukemia. A marked elevation in the total lymphocytic count also support the present finding of lymphocytic leukemia. However, a proper diagnosis requires a prolonged treatment.

Key Words: Haematopoietic tissue, Interenal Pathology,Lymphocytic Leukemia.

9.S.K.Verma, 1990, Aflatoxin B1 in fish feeds and experimental acute aflatoxicosis in an air-breathing teleost. Paper selected for Young Scietist Research Award by the Second Asian Fisheries Forum, Manglore, In Proceedings P-21

Abstract

A general survey of three types of fish feeds for the incidence of tozigenic mold infestation revealed an alarming level of aflatoxigenic Aspergillus flavus contamination during storage. About 89 % of the strains were toxigenic. The toxicity of crode toxxin sample was assed on Channa punctatus. Separate experiment was conducted using pure crystalline aflatoxin B1. Four different concentrations (10, 20, 30 and 40 mg/l) of pure toxin in 0.5ml of propylene glycol were administered intraperitonially to four groups fish alongwith  the control. Observations were made on day 20, 40, 60 and 80. The treate fish showed loss of appetite followed by listlessness and increased mucus over body surface. Hemmorrhagic patches and tumors were apparent on the liver. Histopathologically, liver was most prominently affected organ leading to classical trabecular hepatoma. The excretory kidney revealed some pre-neoplastic alteration arising from tubular epithelium. the head kidney was atrophic, but proliferation inter-renal lobules was prominent. The muscularis of the stomach showed severe proliferation. The intestine, pyloric caeca and gills were atrophic and necrosed. Formation of atypical galandular masses in mucosal region awx marked in the intestine. the spleenic pulps in the treated fish were inditinguishable and showe a monotony of lymphocytes. The total and differential counts of leucocytes revealed a rise of lymphocytes in the peripheral blood.

Key Words: Aflatoxicosis, Air-breathing teleost, trabecular hepatoma, Fish feeds

10.S.K.Verma and S.Pandey,1991. Effect of aflatoxin B1 on the histopathology of some endocrine glands of an air-greathing teleost, Channa puctatus. The Second Indian Fisheries forum. in Proceedings P-210

Summary

11.S.K.Verma,S.Pandey, B.S.Jha, B.K.Jha and A.K.Pandey, 1993. Incidence of aflatoxin B1 in some dietary items of North Bihar in pre-and post-cooked stages. Bulletin of Pure and Applied Science, Vol.1 No.12, P-9-14.

Abstract:

In recent years ohratoxin A, a secondary toxic mold metabolite ellaborated by some strains of  Aspergillus and Penicillium genera is being incresingly implicated in human and health problems. the present evaluates some of the staple dietary items stored and marketed for human consumption for the incidence of toxic mycoflora with special reference to A.ochraceous group of fungi, well known for producing ochratoxins. Besides, the role of prevalent climatic factors , impact of storage conditions and water content of the substrate on sporulation and growth of the fungi has also been discussed.

Key words ; A.ochraceous, Ochratoxin A, mycotoxins, .Foodstuff

12.S.K.Verma (1997). Aflatoxin B1 in fish feeds and experimental acute aflatosicosis in a teleost fish. Journal of Aquaculture, Vol.5, P 13-21.

Abstract:

A general survey of three types of fish feeds for the incidence of tozigenic mold infestation revealed an alarming level of aflatoxigenic Aspergillus flavus contamination during storage. About 89 % of the strains were toxigenic. The toxicity of crode toxxin sample was assed on Channa punctatus. Separate experiment was conducted using pure crystalline aflatoxin B1. Four different concentrations (10, 20, 30 and 40 mg/l) of pure toxin in 0.5ml of propylene glycol were administered intraperitonially to four groups fish alongwith  the control. Observations were made on day 20, 40, 60 and 80. The treate fish showed loss of appetite followed by listlessness and increased mucus over body surface. Hemmorrhagic patches and tumors were apparent on the liver. Histopathologically, liver was most prominently affected organ leading to classical trabecular hepatoma. The excretory kidney revealed some pre-neoplastic alteration arising from tubular epithelium. the head kidney was atrophic, but proliferation inter-renal lobules was prominent. The muscularis of the stomach showed severe proliferation. The intestine, pyloric caeca and gills were atrophic and necrosed. Formation of atypical galandular masses in mucosal region awx marked in the intestine. the spleenic pulps in the treated fish were inditinguishable and showe a monotony of lymphocytes. The total and differential counts of leucocytes revealed a rise of lymphocytes in the peripheral blood.

Key Words: Aflatoxicosis, Air-breathing teleost, trabecular hepatoma, Fish feeds

13.S.K.Verma and S.K.Varma .1997. Aflatoxins in MAKHANA, Euryale ferox  Salisb.,and possibilities of its carryover to man, In Book Published by DIPA, Indian Council of Agricultural Research, New Delhi , eds. Mishra,R.K;Jha, V,N and Dehadrai ,P.V.

(Full length paper supplemented - Invited Guest Lectures)

14.S.K.Verma, 1998. Bloom forming Cyanobacteria- A threat to aquaculture practices in India. Journal of Aquaculture (6), P49-55

Summary

 

15.Shishir K. verma , 1998.  Possibilities of cyanobacterial blooms in wetlands ( Chaurs and Mauns) of Darbhanga . In Proceedings of National Seminar on Perspectives in Wetlands, Department of Botany LNM University. P 23.

17.Shishir K. verma , 1998. Wetland of North Bihar-A profile. In Proceedings of National Seminar on Perspectives in Wetlands, Department of Botany LNM University. P 27.

18.S.K.Verma, 1999. Wetlands and sustainable living : A socio-economic perspective in North Bihar. Presented at Sixth World Wilderness Congress,Banglore (abstract)

19.Shishir K. verma ,A.K.Pandey, B.K.Jha and K.Pandey ,2000, Wetlands of Darbhanga:Abiotic and Biotic status, In Advaces in Zoology Environmental Degradation and Biodiversity. eds. Pandey B.N and Singh,B.K, P 181-192

Abstract:

The district of Darbhanga is a flood prone region of North Bihar, geogrtaphivally identified as area of tectonic depressions. The district comprises vast stretches of wetlands locally called as "chaurs" which are not only highly productive but hold significance from biodiversity point of view. Till date not less than 125 species of fish has been recorded.

The paper presents about abiotic and biotic status of these chaurs with special reference to seven important water bodies well recognised for high economic out put. the most important chaur having a national status belong to this district.

The physico-chemical status of these water bodies though indicate a moderate productive status but , a shift towards eutrophication was also apparent. At places, the condition emphasize a need of awareness as well as proper management.

Biotic status reflects a mixed pattern of phytoplankton dominance and a thick macrophytic vegetation. Among fishes the cat fishes dominate the cathes followed by air-breathing teleosts.

Key words : Wetlands, Chaurs, Physico-chemical status, Biodiversity.

20. Rashmi Verma, Shambhoo Prasad and Shishir K Verma. 2000.  Effect of dietary intake of crude aflatoxins on the blood biochemistry of Channa punctatus. In Proceedings of The Fifth Indian Fisheries Forum, , Central Institute of Freshwater Aquaculture, Bhubaneshwar, P 119.

Abstract:

The fish, Channa punctatus, obtained from local marketwere acclimatized and divided in three groups of 10 fish each. A single dose of 20 ppm crude aflatoxins was  administered daily through oral route. Test fish were sampled o nday 80 and 160 along with control fed Gordons Diet without any toxin supplement.The observed effects indicate elevation in blood glucose level (33.33%) which has been attributed to an imbalance of blood glucose homeostasis eithrr due to decreased utilization of blood glucose or increased gluconeogenesis. Aflatoxin B1 has been found to cause depletion of serum proteins by 14.76% which is suggestive of their inhibited synthesis by aflatoxin B1. the serum cholesterol under aflatoxin stress reveals a case of hypercholesterolemia as evident from the increased serum cholesterol  content (24.07%). It has been opined that aflatoxin B1 either stimulated de novo syntheis of the precursor material rto meet the requirements of an enhanced corticosteroidogenesis or depresses breakdown of cholesterol probably under the influence of impaired thyroid function. it has been concluded that the present fish in response to aflatoxin feeding are faced with a serious metabolic crisis lrading to the mobilisation of sufficient energy reserve.

Key Words: Blood Biochemistry, Channa punctatus, Crude aflatoxins, Bioassay.

 

21. Rashmi Verma,Shashi Bhusan Shashi, Prasant Kr. Saha and Shishir K Verma, 2001. Physico-Chemical characterisation of fish culture ponds favouring survival and growth of toxic cyanobacteria. In proceedings of Seminat on Fish Health and Management, L.N.Mithila  University, P28.

Abstract:

Toxic cynobacterial bloom in India is perceived as a rare event. In present communication 20 fish culture ponds located in Darbhanga town were evaluated for physico-chemical profile and population density of toxic cyanobacteria during summer and monsoon months of the year 2000.

On the basis of the physico-chemical profile culture ponds were categorised as productive  (P),normal (N) and nonproductive (NP). Studies indicatde that pH, redox potential, total alkalinity and dissolved oxugen arte dependable parameters for characterising a water body capable of upholding and subsequent growth of cyanobacterial population. Among chemical factors alteration in N:P ratio appeared to be the most critical affecting the event. Besides, iron, sulphur,calcium and mangnesium in concentration above threshold level also seems significant.

A lis of toxic algae recorded from these ponds have been presented but the discussion has been made with special reference to Microcystis aeruginosa, which was found to be most privileged to alteration in physico-chemical profile. Attemts have been made to establish correlation between physico-vchemical profile and subsequewnt toxic bloom formation.

Key Words : Cyanobacterial bloom, Toxic algae, Freshwater ponds,Microcystis

22. V.Jha, Prashant Kr. Saha, Shishr K Verma and Shambhu Prasad. 2001. Management of fisheries through integrated aquaculture with Makhana, Singhara and other water crops in North Bihar. In proceedings of Seminat on Fish Health and Management, L.N.Mithila  University, P 52.

Abstract:

North Bihar is rich in water resources, Chaurs ( land depressions caused by shifting river cources over millenia), mauns (ox-bow lakes), dead river courses and homogenic pond constitute the lentic water bodies of the area. Rivers like Ghaghra, Gandak, Burhi Gandak, Kosi, Kamala, Baghmati-kareh,Adhwara group Mahananda etc. and fine network of their tributaries constitute the lotix systems of the region. there has been a rapid increase in the areas under water inundation in the last few years. Faulty management of embankments on alost all major rivers in the state has brought a colossal 69 lakh hectare  areas under water logging during monsoon months. Of this, 9 lakh hectares remain under prolonged inundation of 4 to 6 months. It is beyoned the capacity of the state coffers to address itself to meet this adverse situation of such a high magnitude. Attempts, however should be made to derive optimum productivity in thes water bodies under the rotational anc concurrent methods of integratyed aquaculture with crops like Makhana ( Euryale ferox), Singhara (Trapa bispinosa, var natans) deep water rice (DWR) varieties etc. In fact fish farmers in this region have already been practicing integrated qauaculture under indigenous processes. There is a need to augment the process through proper scietific management. The pare reviews the state-of-art of the process as prevalent in lentic water bodies of North Bihar.

Key Words: Fisheries Management, Integrated aquaculture, North Bihar, Lentic water bodies.

23. Meenaxi Das, Shambhu Prasad and Shishir K. Verma. 2002. Toxigenic Mold on Fish Feeds- I: Impact of Climatic Factors, Journal of the Indian Fisheries Association, vol. 2, P 77-92.

Absract:

The present communication is a survey report carried out to asses the inciedence of toxic mycoflora on seven types of agricultural products/ sbyproducts incorporated during fish culture as supplementary dietary items. samples were obtained from various sources at Darbhanga, madhubani and Samastipur disricts during summer, monsoon and winter months. Out of the total 1774 samples , only 894 appeared to be fresh visually refelecting average incidence of contamination around 46.4 %. However, the apparently fresh samples , when subjected to culture, 2.9% of them were also found to be contaminated. Thus degree of fungal spoilage in fish feed ingredients in parts of N.Bihar appears gto be significantly high (73.5%). thw present study ellaborates the facts with special reference to aspergillus flavus and Aspergillus parasiticus  (elaborating aflatoxins);A.ochraceous and Penicillium viradicatum (elaborating ochratoxins) and A.versicolor (elaborating sterigmatocystin). The other srains already known for toxigenic potentials that appeared on the present substrates included A.niger, A.fumigatus, A.candidus, P.islandicum, Rhizopus spp.and Mucor spp. Studies indicate that the prevalent climatic factors like temperature and relative humidity facilitate a congenial condition almost all through the year and in particular during summer and monsoon months. But water content of the substrate is a vital factor that further accelerates the mycobial spoilage.A thorough sun-drying of the agricultural commodities before prolonged storage to bring water content  below the " low risk limit" may significantly reduce the incidence of molds.

Key words :  Toxigenic molds, Fish feeds, Fungal spoilage, Climatic factors.

24. BANDANA verma and Shishir K.Verma, 2002.  Ochratoxin A in staple dietary items stored and marketed for human consumption. Indian Journal of Environmental and Ecoplanning, 6 (2),P 229-240.

Abstract :

In recent years Ocratoxin a , a secondary toxic mold metabolite elaborayted by some strains of Aspergillus and Penicillium genera is being increasingly implicated in the human and animal health problems. The present paper evaluates the staple dietary items stored and marketed for  human consumption for incidence of A.ochraceous group of fungi, well known for producing ochratoxins. Beside the role of prevalent climatic factors , impact of storage conditions and water cdontent of the substrates on sporulation and growth of the fungi has also been discussed .

Key Words . A.ochraceous, OchratoxinA, Mycotoxin, Foodstuff

 

25. Shishir K. verma,  2003. Freshwater toxic Blue-green algal Blooms:A response to Extra Nutritional Enrichment In : Ecology of Polluted Waters eds. Arvind Kumar. Daya Publishing House, New Delhi, P 1161-1175.

ABSTRACT:

Toxic algae have been reported from all aquatic environment throughout the world. In freshwater, vcertain species of this group fix atmospheric nitrogen but many of them of the cynophycae group  have gained recognition as water contaminants. Thyey form dense unialgal growth often calle3d as "bloom" and are capable of ellaborating potentially lethal toxins. Inspite of numerous reports of livestock, pets and human beings being victim of algal toxins in India it is still perceived as a rare event. During the last few decades a major shift in phytoplankton community of freshwater  bodies mainly towards toxic blue-green algae is being noticed. It is due to extra nutritional enrichment of the waterbodies specially phosphorus component.

Study of twenty fish culture ponds of DARBHANGA (N.Bihar) where mass mortality of fish is a recurrent phenomenon revealed that  it is difficult to predict blooming of toxic algae but monitoring of some of the physico-chemical parameters may trigger the alarm of their onset.

Blooming of toxic algae is practyically rare in productive ponds having ideal physico-chemical conditions but the risk increases with gradual enrichment of the water bodies being maximum in defined eutrophic ponds  having altered N:P ratio. While phosphorus is essential for the growth of the organisms, nitrogen is frequently considered as growth  limiting factor. Imbalance in the ratio of these two factors , by any means, accelerates pace of toxic algae dominance.

Key words  :  Cyanobacteria, Toxic Algae, Blue-Green Algae, Europhication, Freshwater.

26. Baidyanath Kumar, Shishir K Verma,Meenaxi Das and Nagina Kumar Dubey, 2006. In vitro growth response of Micrcystis aeruginosa, a bloom producing tox ic cyanobacterium. J.Haematol. & Ecotoxicol. 1(2) P 51-60

Abstract:

In vitro growth response of  Microcystis aeruginosa, a toxic bloom producing cyanobacterium, was evaluated on ten nutrient media (I to X) in which  three were self formulated and abbreviated as BRL-I,II and III (Biotoxin Res. Lab.) medium. In all the nutrient media growth of M .aeruginosa startyed at slow rate upto seven days of incubation. Mineral media I,II,III,IV,V and VI supported very slow growth. Following 35 days of incubation in media I,II,IV,V,VII aand IX a decline in the population density was recorded. In media I (ASM), II and VII the population density declined to almost nil after 56 days of incubation. Medium IX (BLR-III) supported a luxuriant growth representing a record population of 10.86 x 104 to 12.3x 104 cells/cm3 between 35 to 49 days of incubation.

Key words: In vitro growth,Toxic bloom, Micrcystis aeruginosa, BRL-media, ASM-media.

27. Baidyanath Kumar, Meenaxi Das, Nagina Kumar Dubey and Shishir K Verma . 2006. Toxic Assessment of Micrcystis aeruginosa.J.Haematol. & Ecotoxicol. 1(2) P 20-32.

Abstract: 

The effect of cellular extract (endotoxin) from M.aeruginosa wa studied on Daphnia magna employing static toxicity test on air-breathing teleosts and major carps. The endotoxin prooved to be toxic to all the experimental animals. 1.0ml of endotoxin extracted from 42 days old culture caused 50% mortality of the D.magna population. Similarly, 1.0ml of the endotoxin extract when administered intraperitonially caused the death of Heteropneustus fossilis, Clarius batrachus,labeo rohita and Catla catla. However, the major carps L.rohita and C.catlawere found more susceptible to microcystis endotoxin than H.fossilis and C.batrachus.

Key words : Toxic Assessment, Microcystis aeruginosa , Daphnia magna, Bioassay,Toxic bloom

28. Shishir K. Verma , 2007. Mycotoxins in human food chain via fish flesh consumption. (Invited lecture) In proceedings: 18th All India Congress of Zoology and National Seminar on Applied Zoology and Environmental Sience with Special Refernce to Eco-restoration and Management of Bioresources, University of Lucknow . P 155

Abstract

Entry of mycotoxins in human food chain through contaminated dietary items is an age-old problem and has been investigated widely. the present problem, however, could be envisaged with the possibilities of bioaccumulation of these mycotoxins and their carryover to man through dietary intakes of animal origin. Establishment of the fact that some of the mycotoxins that find entry into the fish through contaminated supplementary diets into human food chain is a long and torturous pathway. Attempta have been made to present the chronology of events illustrating the problem.A brief record of micro fungi infesting 1774 fish feed samples collected during summer, monsoon and winter months have been discussed with influence of the climatic factors. Isolation, culture and extraction of the identified fungal strains for aflatoxins, ochratoxin A and sterigmatocystin alond with bioassay on major carps highlighting chief pathological targets have been dealt with. Studies on biaccumulation, biomagnification ant tissue clearance rate have led to the establishment of the fact that if a fish has been exposed to a single acute dose of aflatoxin B1, the trace of parent compound goes beyoned detectable level after 20 to 25 days of the administration. But, the property of Ochratoxin A to bind with the protein molecules provide avenue for further investigations.

28.Shishir K. verma , 2009. Aflatoxin B1 induced changes in the ovary of an air-breathing fish. In Recent Advances in       Hormonal Physiology of Fishand Shellfish Reproduction eds. B.N.Singh and A.K.Pandey, Narendra Publishing House, P 187-292.

Abstract

In order to record the effect of aflatoxin on the ovary of Channa punctatus,  fish were exposed to 10, 20, 30 and 40 ppm of pure crystalline AFB1 for 80 days. Acute toxic response of AFB1 was apparent at high dose levels (30 and 4o ppm) within 20 days. The ovary became atrophic due to severe cellular necrosis dominated by atretic oocytes. In fish receiving 10 and 20 ppm AFB1, the ovary appeared to be relatively normal containing ova in various stages of maturation. Studies indicated that the toxin affects the movement of intranuclear elements during cell divison acausing acentric localization of the nucleolus and chromatid bivalents. In addition, the toxin induces some proliferative cxhanges in the thecal layer which apparently became multilayered conaining cells with increased nulearity.

Key Words : Aflatoxin B1, Cellular Necrosis, Proliferative Changes,Ovary, Channa punctatus  

 

                        ( To be added)

 

 

Invited Guest Lectures:

 

1.  Mycotoxins and Freshwater Aquaculture in the 21st. Century

International Symposium on Fish for Nutritional Security in the 21st. Century, December,2001.

Central Institute of Fisheries Education (Deemed University-ICAR), Versova, Mumbai 400 061.

 

2. Aflatoxins in Makhana and their carryover to man

In Book Published by DIPA, Indian Council of Agricultural Research, New Delhi , eds. Mishra,R.K; Jha, V,N and Dehadrai ,P.V.

 

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