Three counterparts from FARTC, R.K. Dey, D. Kumar and B.K. Mishra, who were previously assigned to the unit for Ichthyopathology and Fish Health Protection (UIFHP) worked all the time with the consultant. One previous member of the unit, Mr K. Suresh was transferred and a new member of the team will be assigned to the unit soon. Two counterparts from other CIFRI research station, A.K. Gosh and R.N. Pal were also working with the consultant during most of his assignment in Dhauli.
The work reported on incorporates reviewing the research and diagnostic activities of unit staff members and aid in formulating research programmes. In addition, group and individual training of assigned scientists in their respective fields consumed a significant part of the consultant's time.
During the previous assignment of the consultant, he aided the FARTC staff in setting up five research projects. The aim of these projects was two-fold: (i) to develop methodologies and capabilities for in depth studies of fish diseases, as well as of their impact on production in ponds, and (ii) to start a thorough inventory of fish health problems in composite fish culture, as practised in India. These basic steps were recommended in order to (i) enable the scientists involved to specialize in some narrower disciplines and thus form the basis for team-work both in research and in diagnostic services, and (ii) to form a proper basis for selection of research priorities within ichthyopathology.
The five investigations were designed for a three-year period (beginning with 1981), with extension after revision and necessary adjustments.
Preparations for implementation of this project are in progress. In the meantime, several treatment experiments were carried out. Further recommendations on this work are given in Section 2.3.1.
In accordance with the general proposals of the consultant in his previous report (see Section 4.2.7) several field and laboratory experiments were carried out to determine the efficacy of preventive drug application to fish in reduction of losses or for preliminary evaluation of the possibility to cure the sporozoan kidney infections. These experiments deserve special attention.
In a large-scale field experiment, the influence of prophylactic drug treatment of larvae and the value of improved food quality for increasing of survival was tested. There were four groups in the experiment and three ponds (repetitions) were used for each group. All 12 ponds were stocked with 5 million per ha of three days old rohu larvae. The experiment lasted for one month. In group A, the larvae were treated immediately prior to stocking by a short bath in an antibiotic and by a dip in KMnO4; during the experiment this group was fed a conventional supplemental food. In group B, larvae were not treated prior to stocking; the group was fed with a formulated complete diet. In group C, the treatments of groups A and B were combined, i.e., larvae were treated prophylactically prior to stocking and the complete diet was used. The control group D was fed the conventional supplemental food. At the end of the experiment, the survival was lowest (arithmetic mean 6.17 percent) in the control group D. In group A, survival was 8.41 percent, in group B 11. 3 percent and in group C 16.04 percent. Results indicate clearly that, under the conditions of the experiment, antibacterial treatment prior to stocking increased survival about 40 percent as compared to untreated groups, while feeding of the complete diet increased survival by over 80 percent. Combined treatment had an additive effect, which is understandable; mechanisms and time of action of these two treatments are quite distinct. Results will have to be analysed further by statistical methods but, even without a more detailed analysis, they clearly indicate the usefulness and practical significance of the two treatments applied.
The main finding in this experiment is actually the fact that very high losses occurred in all experimental ponds (mean for 12 ponds 90.5 percent, range 77.4 to 96.6 percent). These results exclude the possibility of interpreting losses as accidental. They were only partially caused by bacterial infection immediately after stocking or by inadequate quality and quantity of food distributed to the ponds. The main causes of losses will have to be determined in further experiments and this work deserves highest priority.
The stocking density applied in the experiment may be an important factor in occurrence of high losses. The density of 5 million/ha of larvae has been applied on the basis of some recommendations in India. The stocking rate of cyprinid larvae for rearing up to about one month of age is much lower in Europe; the maximum does not exceed 2 million/ha and 1 million/ha or lower densities are commonly applied. In commercial European operations, such stocking gives about 0.5 million/ha of fry at harvesting. The average yield (number/ha) of fry in this experiment is exactly in the same range. It seems, therefore, to the consultant that at least one experiment on a similar scale should be conducted to determine the optimum standard stocking density of larvae for the experimental ponds in Dhauli.
Methods for improved survival of larvae can be developed best by experiments in which monitoring of health status is carried out. Some proposals on these approaches are given in Chapter 2.3.3.
Based on the previous experimental findings that antibiotic treatment during handling of rohu prevents development of columnaris disease and of losses (mediated by handling stress and handling-induced lesions), experiments were conducted to determine whether antibiotic treatment of spawners can reduce post-spawning mortalities. Post-spawning antibiotic treatment by injection seems to reduce the post-spawning mortality: in the untreated control group, five out of 30 fish died (16.5 percent), while only one of 20 treated fish died (5 percent). These results warrant carrying out further experiments on post-spawning antibiotic treatment. Some suggestions are given in Section 2.3.3.
Mrigal fingerlings from a group with high incidence of a myxosporidian infection in kidneys were brought to the laboratory and divided into experimental groups. They were treated by several antiprotozoan drugs (Nitrofurazolidone, Ctenadazole, Metakelfin and Dihydroametine) in food. Results were inconclusive. The high incidence of myxosporidiosis may be having an impact on growth rate of fish populations in ponds and further drug treatment experiments are recommended. The chances of finding an efficient treatment may be improved by using an improved methodology of experimentation.
The toxicity of potassium permanganate and of formalin for rohu eggs incubated in hapas and the influence of these chemicals on fungus infection of eggs was studied. First the innoxious doses were determined. Fertilized eggs (95 percent fertilization) of one rohu spawner were divided into six lots of 50 000 eggs, for three experimental groups. Each lot was incubated in a separate hapa. In group A, eggs were treated with 2 000 ppm of formalin for 60 seconds, in group B with 250 ppm of KMnO4 for 60 seconds and group C served as control. Differences among groups were small but in favour of treatments. Efficacy of treatments for prevention of egg losses could not be ascertained because there were no fungus infections or abnormal losses in controls. These experiments should be continued, especially in situations where fungus or bacterial infections cause problems during incubation of eggs.
Preparatory steps for work with cell cultures have been completed; special procedures of washing and sterilization of glassware and other items, as well as the preparation of fish for taking of tissues, are routinely used. However, some difficulties were encountered during preparation of cell cultures, mainly due to inadequate results of attempts to tripsinize tissues and due to the contamination in vessels seeded with cells. During the consultant's assignment, conditions for aseptic work in the cell culture room were improved by installation of a hood, borrowed from the genetic laboratory of FARTC. The first primary monolayer cell cultures prepared from explants of catla ovaries and from trypsinized kidneys of catla were grown in test tubes and in MDB bottles. Cultures were of moderate quality. Results of trypsinization of tissues are still not satisfactory despite several experiments to improve the technique. Much better results will probably be obtained with the use of mechanical stirrers.
A sample of moribund rohu fry was examined virologically. This first attempt to isolate a virus from an Indian major carp on primary cell culture from rohu was conducted in a technically correct way but there was no indication of virus presence in inoculated cultures.
For further successful work and for completion of this investigation, additional equipment is urgently needed (see Appendix 1).
After completion of the first tasks defined in the expert's previous report, the investigations should be extended.
The person working on this investigation was transferred and only a summary of the work was available. Flexibacter columnaris was isolated from several sick fish and pure cultures were successfully kept in a maintenance medium. Pathogenicity tests showed the isolate to be pathogenic. Several other bacteria were also isolated. During the consultant's stay, one scientist (B.K. Mishra) received basic training on preparation of media, inoculation of agar plates with material from sick fish, obtaining and maintenance of pure cultures and on some basic biochemical tests for identification of bacteria. When the new member of the unit starts to work he will have to take over this work with the aid of Mr Mishra and learn additional methodologies.
The work on this investigation was concentrated on myxosporidian parasites. Observations on kidney samples often showed simultaneous presence of two morphologically distinct types of spores, indicating that two species of myxosporidia may be involved. Describing of these species is in progress. Securing of adequate literature is essential for this identification work.
Staining techniques for spores, such as Giensa, parasitological iodine, silver nitrate and malachite green have been mastered and used. Technique of spore isolation and concentration has been attempted and is in process of standardization. A method for long-term maintenance of spores was developed.
The technique of blood sampling and staining of smears has been mastered but it is not used. It might prove useful in some later studies.
Work on methods for detection of intestinal coccidia has not started yet.
Kidneys and livers of a considerable number of Indian major carps have been collected and fixed mainly in Bouin's fluid or in formalin. Paraffin embedding, sectioning into 5–7μ thick sections and staining by haematoxylin and eosin has become a routinely used technique. Freeze sectioning offers considerable advantages in diagnostic work and is an obligatory prerequisite for some special staining procedures. When carbon dioxide in containers will be obtained, the freezing microtome will be put in use.
Observations of stained slides of organs have slowed occurrence of various damages in the organs (see Section 2.2.4). So far, organs were collected mostly from sick fish. In the few normal fish, kidneys and livers were also slightly to moderately altered. Further collection of material, use of additional staining methods and improved record keeping has to be applied (see Section 2.3.5).
Samples for diagnostic examinations were taken or received from FARTC ponds, KVK/TTC ponds, State Government ponds at Kausalyagang, village ponds under the FARTC Lab-to-Land programme and a few from State Government farms outside Bhubaneswar.
The material for diagnostic work stems from a limited geographic area. This is quite understandable since the diagnostic activity at FARTC does not have any tradition yet. State and private fish culturists, as well as the extension workers, will have to get more acquainted with diagnostic capabilities at FARTC and become interested in obtaining quick diagnosis and recommendations for practically applicable and efficient treatments. To achieve this fully, the staff of the FARTC Unit for Ichthyopathology and Fish Health Protection (UIFHP) has to gain more experience through more diagnostic work, training, study of literature and research. The benefit from increased volume and quality of diagnostic work will come in the form of a constant supply of material for research as well as of informations on needed developments in upgrading prevention and treatments in praxis. Recommendations for increasing the inflow of samples are given in Section 2.3, in accordance with the initiatives of Dr V.R.P. Sinha, the national Project Director.
It is worth noting that some researchers in other units of FARTC (nutrition, genetics, etc.) increasingly use the diagnostic capabilities of the UIFHP and seek advice for treatments. Such cooperation is of mutual interest and benefit and should be vigorously pursued.
Dissection and macroscopic observations of organs as well as microscopic examination of scrapings and wet mounts for parasites is routinely used. Histopathological examinations are increasingly applied and proved to be very useful. More experience and knowledge is needed in this field. Bacteriological examinations are applied when necessary but procedures for identification of bacteria will have to be better developed for more accurate and fast work. Virological diagnostic methods are developed almost to the point of practical applicability.
Accumulation of specimens, slides and pathogens for documentation of reports and progress, as well as for training, extension and exchange with other laboratories is an important activity based on diagnostic work and on research. This activity has been started. Several specimens showing gross pathology have been preserved in formalin. Some pictures of fish were taken but their quality is not satisfactory due to the poor quality of the private camera used. Requisition of adequate equipment is urgently needed (see Appendix 1); histological slides are also being collected and a cabinet for systematized storage is needed. Microphotographs of various parasites and histopathological changes were taken with the aid of the consultant in order to help counterparts to develop knowledge on selection of material, magnifications and selection of typical lesions. Methods for maintenance of microorganisms will have to be adopted. The consultant advised on some simple methods but in the future freeze-drying and cryopreservation methods will have to be adopted too.
Up to now the largest and most frequently encountered group of diseases in diagnostic work were those caused by parasites. Diseases already known to occur in India, such as trichodinoses, ichthyophthiriasis, argulosis, dactylogyrosis and gyrodactilosis were infrequent. In most instances only a few of these parasites were found on healthy looking fish. Myxosporidioses manifested by formation of cysts on skin, fins or gills were found to affect some populations.
Several parasitic diseases which have so far not been reported in India seem to be quite important. The most frequent findings in all three Indian major carps were spores of two myxosporidia in kidneys. These spores were also found in other organs but they were less abundant. The infestation is a chronic process and defence of the fish is manifested by formation of cysts with parasites in kidneys. Heavily infected carps exhibited emaciation, discolouration, degeneration of kidney tubuli and slow growth. Mild and moderately infected fish did not show macroscopic symptoms of disease. These kidney myxosporidioses seem to be endemic, at least in the Dhauli area.
Severe damage of epithelium in kidney tubuli was found in one catla. The condition was caused by a parasite of the microsporidian type which infested most of the tubuli and replicated within them. The same fish was simultaneously infected by kidney myxosporidia. The fish was emaciated.
Among bacterial diseases, tail and fin rot, columnaris disease and bacterial gill disease were encountered. The sample of fish with tail and fin rot that was brought to the laboratory and kept in tap water recovered spontaneously. This indicates that environment (the water quality in the pond) triggered the bacterial infection and necrosis of skin. Occasional losses with manifestations of columnaris disease among catla and rohu fry kept in the laboratory usually occur after handling. Histological examination of such a case disclosed the presence of necrobiotic processes in liver, intestine, pancreas and spleen. Some of these processes are not typical for columnaris disease and another casual agent might have been simultaneously involved in this case.
Secondary bacterial gill diseases are frequent in fish culture in cold and temperate waters but were not studied in India. Histological examination of catla from a case of mass mortality of this species in a pond showed hypertrophy and hyperplasia of gill lamellae epithelium, typical of bacterial gill disease. This condition must have been triggered by an irritant in water that affected catla more than other species in the pond. Gills of catla were also affected by a mild myxosporidian gill infestation which could not much influence the health of fish. Several non-communicable diseases, such as hernia in catla, cholelithiasis in rohu, cataract in rohu fingerlings, bleedings in eyes, gas bubble disease in early rohu fry and tail deformities (lordosis) in rohu were noted.
Histopathological observations on moribund fishes from a laboratory case of gas bubble disease showed hyperplasia and hypertrophy of gill epithelium, sloughing off of the intestinal epithelium and dilated blood vessels in the liver. These findings fit only to some extent the gas bubble disease and may also be eventually caused by toxins of algae that were abundantly present and caused the oxygen hypersaturation in containers.
A particularly interesting case, demonstrating both the need of thorough checking in any mass case of abnormal health status (including abnormalities in reproduction) and the necessity of a multimethodological approach in diagnostic work, was encountered during the consultant's assignment. When fishing for adult catla in a pond with a history of inadequate gonadal development in previous spawning seasons, one obviously sick specimen was found among 15 fish. The disease condition was diagnosed on the spot as dropsy. Laboratory examinations revealed emaciation, anaemia, accumulation of fluid in the skin and the abdominal cavity, haemmorrhages in the skin, deformation and thickening of the posterior swim bladder chamber and a dark brownish liver. Myxosporidian spores were present in kidneys, ascitic fluid and other organs. Two species of bacteria were isolated from kidneys and the predominant one was preliminarily identified as Aeromonas hydrophila. The main histological finding was an enormously high presence of pigmented cells in the liver and spleen, especially around thickened walls of blood vessels. Capillary bleeding was also pronounced in these organs.
If only one of the above diagnostic techniques had been used, a wrong diagnosis might have been made: a parasitologist could have implied a primarily myxosporidian aetiology of the disease, while a bacteriologist alone could imply A. hydrophila as the primary agressor causing dropsy (or haemorrhagic septicemia). A myxosporidian infection can cause emaciation and anaemia but not the mass appearance of pigmented cells in the liver. It can, therefore, be regarded as concomitant, aggravating the health status. A. hydrophila can neither cause emaciation nor induce pigment accumulation but can be associated eventually with bleeding. Bacterial infections have most probably developed in the premortal stage. In this case, the histopathology gave the clue to the most important lesion - chronic blood vessel damage and chronic pigment accumulation in the liver. The nature of the pigment has been determined by a histochemical method - as hemosiderin. In other cases, some other diagnostic methods may prove to be essential for establishing a proper diagnosis.
The above thorough examination of a single fish has interesting and important implications. Examination of ovaries from two healthy looking catla caught in the same pond showed abundant presence of pathologically pigmented cells full of hemosiderin, as well as changes in blood vessels. Pathological hemosiderosis in liver and ovaries was found in several other dissected healthy looking catla which had been subsequently taken from the same pond. This indicates a probable link between infertility of catla in this pond and of the pigment formation as well as vascular pathology.
Chronic damage of blood vessels in certain organs, with bleeding and accumulation of hemosiderin is a rare manifestation in fish and has not been described in catla. The condition should be tentatively named “hemosiderosis in catla”. At the moment, only speculation can be made on its primary cause, which could be nutritional, toxic or infectious. Since reproductive unfitness of catla breeders seems to be one of the consequences of this disease, research on its aetiology deserves highest priority. It is recommended to start research on the following three aspects of catla hemosiderosis: (i) collection of epizootiological data by health monitoring (see Section 2.3.1); (ii) better definition of the disease by detailed examination of affected fishes (determination of haematocrit and haemoglobin values, blood smear examination and determination of red blood cell differential counts, distribution of bleeding and hemosiderin in various organs, histopathological examinations of all organs using various staining methods); (iii) experimentation on possible nutritional or bacterial aetiology (see Section 2.3.6). Depending on results of this research, further work will have to be directed to test the various hypotheses on aetiology and to develop methods for prevention of the hemosiderosis.
As can be seen from Section 2.1.2 of this report and from the recommendations on the further programmes, research efforts aimed at control of diseases are intensive. As the first practical result, prophylactic treatment of breeders after spawning with injection of antibiotics was recommended and is used at FARTC. More such practical applications could not be achieved yet, mainly for three reasons: (i) diseases for which treatments are known have not yet been encountered in a serious form in the relatively small pond area covered by the diagnostic work; (ii) applicable prevention and treatment methods are not yet available for several diagnosed diseases (myxosporidiosis, sanguinicolosis, etc.) and will have to be developed by research; (iii) members of the unit are young scientists that still need more training and experience. Implementation of control measures for diseases will certainly increase with time.
One of the difficulties in implementation of this project might be the specificity of pond culture practices in India which have not previously been taken sufficiently into account by the consultant. Contrary to the practice in European pond farms, the regular sampling (at 14-day intervals) of fish from each pond is not a common part of the technology in India. In Europe, this sampling provides the possibility of closely following up the growth rate, food conversion and the general health status of fish in all ponds. Fairly satisfactory sampling methods have been developed. More recently, a more thorough examination of samples for health status is performed at monthly or bimonthly intervals. The value of a few fish that have to be sacrificed for examination can be amply repaid by prevention of significant losses through timely application of proper treatments. But even in Europe some fish farmers are reluctant to adopt regular detailed examinations. At the present status of knowledge of fish diseases in India, such detailed examinations of fish at regular intervals are even more important than in Europe. The consultant is of the opinion that FARTC should develop more dependable, as well as less manpower and time consuming fish sampling methods than the ones presently used. Also, the sacrifice of fish for detailed health monitoring should be used in a few production ponds.
This research project can be implemented as recommended in Section 5.1 of the consultant's previous report (Field Document 2) with the following amendments:
(i) Development of more efficient sampling methods
A method of sampling by fishing in a corner of the pond is described below but other means of sampling can also be developed.
Net: The net should be about 25 m long and 2–3 m deep, with a mesh size adequate to the size of fish to be sampled. One net for fingerlings and one for adult fishes would be desirable. The strong bottom line should be weighted by 50 g pieces of lead at 15–20 cm intervals (totalling about 8 kg of lead). Leads on the line should have an elongated cylindrical form with tapered ends and a very smooth surface to prevent entirely any snagging or entangling. The top line must have plastic floats 20–25 cm apart. The size of the floats should keep the entire upper line floating. On one end of the net, the top and the bottom line should be connected to two pulling ropes, each about 25 m long. A strong 2.5–3 m long wooden pole with a strong metal ring or loophole (5 cm hole diameter) firmly fixed on one end is also needed.
Preparation of one corner in the pond: A deep corner of the pond and its banks of about 10–15 m in length at each side should be cleaned and kept free of any obstacles and weeds. The bottom and sides of the corner should be smoothed carefully to avoid completely any snagging of the net.
Attracting fish to the corner: Three days prior to sampling, as well as on the same day of sampling, the feed should be given in this corner only, every day at the same time (for instance at noon).
Preparation for netting: On the day of sampling, at least four hours before feeding time (for instance 19.00–20.00 h), the net should be set up in the corner. One end of the net (the one without pulling ropes) is fixed to the bank about 10 m from the tip of the corner. The net should be set up in the water neatly folded so that it does not obstruct more than 30 percent of the “entrance” to the corner and so that it can be pulled quickly to the other side of the corner. During placing of the net special care should be taken to prevent tangling the bottom leads with themselves and with the meshes of the net. The bottom pulling rope should be inserted through the loop (or ring) on the wooden pole and the pole put into the water on the spot where the net will be pulled to fence off the corner. Both ropes should be slightly tightened and placed on the bank.
Pulling of the net: After the dispensing of food in the corner, there should be no noise or walking around that part of the pond. The net should be closed (pulled) 1 ½–2 ½ h after dispensing the food; the exact timing should be improved by observations and experience. Four men are needed for this fishing: one pushes the pole with the bottom rope firmly to the bottom, one is pulling the bottom rope, one is pulling the surface rope and raising the surface line of the net somewhat when the fish start to jump, and the fourth man raises the surface line of the net on the end that is fixed to the bank. The hauling of the net should be done as quietly and inconspicuously as possible to the fish. For the whole operation, two times one half hour of four men will be needed.
Some training and experience are needed to perform this operation successfully.
(ii) This project will also serve as the source of materials for work on parasitological and histological investigations (see Sections 2.3.5 and 2.3.6).
(iii) The area monitored and the number of samples in this project can be increased by regular sampling of organs of Indian major carps in fish markets and in the future by examination of samples taken regularly from state ponds in several states.
Sampling of organs in markets is suitable for increasing the area of monitoring and the number of samples. Its applicability is limited to certain types of examinations where proper dissection and examination are not needed and where the grade of freshness of organs is not critical. This source of materials can therefore be used for parts of the investigations under Sections 2.3.4, 2.3.5, 2.3.6 and 2.3.7, i.e., for (a) study on virus presence in asymptomatic fish; (b) the study of incidence, intensity and seasonality of kidney myxosporidiosis in carps; (c) for study of histopathology in kidneys and livers of Indian major carps (if grade of freshness of organs will be satisfactory), and (d) for study on incidence of hemosiderosis in catla. An example of a sheet for collecting of data is given as Appendix 2.
Upon the initiative of Dr V.R.P. Sinha, FARTC will inform departments of fisheries of several neighbouring state governments about the interests of FARTC for fish health problems, ask for information about extent of losses in pond culture operations of these departments, about the current known role of diseases in these losses and on diseases that have been noted so far. At the same time, diagnostic services and fish health monitoring services will be offered. Proposed forms to accompany this initiative are attached as Appendixes 3, 4, and 5 of this report. This initiative should increase the inflow of material for health monitoring (and diagnostic work).
Work on projects defined in the consultant's previous report in Sections 5.2 to 5.5 should be continued, as already outlined, and completed during 1982 and 1983. Details on technical programmes were discussed, further outlined and demonstrated during the work of the consultant with the counterpart scientists. Objectives of this work, i.e., development and application of various specific methodologies for research and diagnostic work on fish diseases have already been partially reached. This allows the start of the research on individual diseases. Specializations of scientists engaged in foreign fish disease laboratories will also add to their theoretical knowledge and the technical level of methodologies used by them.
Work described in Section 2.1.2 of this report should be continued, except on treatments for myxosporidian infections. For the latter research, preparatory experiments and more data on the biology of parasites are needed, as outlined in the proposed investigations under Section 2.3.4 of this report.
Two scientists (D. Kumar and B.K. Mishra) with participation of other FARTC staff when needed.
Objectives are to develop and test practically applicable drug treatments for prevention of fish losses in various life stages and during various phases of fish culture operations.
Opening up of possibilities to reduce losses in various phases of fish production technology.
for reduction of losses in postspawning brood stock, intraperitoneal injections of antibiotics and vitamin-C alone and in combination will be tested, using adequate untreated controls; effective doses per 1 kg of body weight will be determined for all species of major carps.
during egg incubation, short bath treatments in non-toxic concentrations of formalin, potassium permanganate will be used daily in order to evaluate their effectiveness in preventing egg losses due to fungus and bacterial infections.
influence of pond pretreatment with an insecticide on survival in larval nursing period will be tested; 0.25 ppm of Malathion, five days prior to stocking, should be tried first; additional experiments may be needed to compare and evaluate other rapidly decomposing insecticides of relatively low toxicity to fish.
influence on survival of bath treatment of larvae prior to stocking in antibiotic solutions or potassium permanganate, alone or in combination, should also be evaluated.
efficiency of prophylactic pond treatment with 0.15 ppm malachite green after second and third week of fry rearing should be tested.
effects of drug application in food (two days prior to the netting) on survival after the handling stress during stocking operations or subsequent survival should be tested; terramycin (50 mg/kg of fish) and vitamin-C (150 mg/l kg of fish) alone and in combination can be tested.
Three years, with extension after revision.
One (B.K. Mishra).
Myxosporidian infections of major carps have not yet been described in India. Their occurrence in cultivated species of carps may have a damaging effect on growth and survival rates, species of Myxosporidia in kidneys have to be described and identified and their seasonal and age-dependent incidence in cultivated fish species determined, both under various pond and open water conditions. The influence of intensity of kidney invasion on growth and condition of fish will also be determined. In order to open up possibilities for prevention and treatment experiments, the biology of parasites has to be known.
The disease will be described and their impact on fish culture evaluated. The knowledge gained will also serve as a basis for prevention of damages by these diseases in fish culture.
on the basis of microscopic measurements of morphological characteristics and staining of spores from various fish species, the parasites will be described and their systematic position determined.
kidneys of major carps will be collected from ponds under the Fish Health Monitoring Research Programme and from the market, as well as from various other experiments and diagnostic cases; squash preparations will be examined microscopically and the intensities of invasions by different species of Myxosporidia determined arbitrarily.
data on length-weight relations in fishes will be compared with findings in kidneys in order to check whether a correlation between the intensity of invasion and the condition of fish exists.
the minimal age and size of fish which can be infected by each myxosporidian will be determined in laboratory experiments; pond mud will serve as a source of infection for larvae reared in plastic pools; the time elapsed between exposure to infection and development of spores will also be determined in such a laboratory experimental system.
Two years, with possibility of extension after revision.
One (R.K. Dey).
The kidney and liver are affected by pathological alterations in most of the systematic bacterial, parasitic and viral diseases and some localized parasitic diseases, as well as by some nutritional deficiencies in fishes. It is, therefore, essential to start a detailed study of their organopathology in species that are most important for the fish culture in India. These studies will contribute to a better characterization of the known diseases of Indian major carps, as well as to detect pathological changes which remained unnoticed so far.
Data obtained will be useful for improvement of routine diagnostic work, as well as for further research aimed at health protection of fishes in pond culture.
samples of kidney and liver tissues from diseased and healthy looking major carps will be collected from the following sources: (a) ponds where the fish health monitoring project will be carried out and samples taken at bimonthly intervals; (b) any outbreaks of mortality or disease in ponds covered by diagnostic work of the unit; (c) fish of satisfactory freshness in local markets; sampling will cover all ages (from fry on) during 18 months.
general data on source of samples and data about age, size, source of fish, stocking density, etc., and diagnostic findings obtained by other laboratory methods will be collected together with the material.
samples will be fixed in Bouin's fluid and buffered formalin, embedded in paraffin, sectioned and stained by haematoxylinlosin; other staining methods will be also used for demonstration of tissue sections to lesions and of metabolic disturbances; a collection of stained slides will be prepared; work will be also documented by photomicrographs.
findings will be grouped and described according to diseases and pathological conditions encountered and then analysed in each species, age and season for incidence, severity and joint occurrence.
Two years.
Five, three from the unit (D. Kumar B.K. Mishra and R.K. Dey) and two from the Nutrition Laboratory (Dr B.N. Singh and K. Kumar).
To define and describe better the recently encountered hemosiderosis in terms of incidence, seasonality, severity, consequences for the fish organism and pathology; to start the work on determining the cause of disease, testing first the hypothesis on nutritional and bacterial aetiology; to check the eventual influence of stocking density on occurrence and severity of disease.
Knowledge of this disease may help to develop methods to prevent or cure it and thus improve chances of better practical results in cultivation and reproduction of this important species.
catla of various origins and ages will be checked for presence of hemosiderin in livers and ovaries by microscopic examination of squash preparations under UV light and by histopathological methods.
brood fish stock in a FARTC pond will be examined thoroughly by various methods during seasons of the year, including examination for relation between development of gonads and the amount of hemosiderin.
experiments will be conducted in FARTC ponds to determine the influence of food composition and addition of antibiotics on intensity of hemosiderosis in a way to enable drawing of conclusions about the possible role of micronutrient defficiencies or bacteria in aetiology.
Two years, with possibility of extension after revision.
The laboratory and field occurrence of gas bubble disease needs to be explored because
of its metabolic impact and losses in fry. Counterparts were therefore advised on preliminary
experimentation aimed at induction of the disease under controlled conditions and
A disease condition of uncertain aetiology that causes heavy mortality among tiger shrimps (Penaeus monodon) cultivated in Bengal in paddies with low salinity water was discussed with Mr Pal. It was recommended that two research approaches should be pursued for better defining the disease and its aetiology, i.e., histopathological and bacteriological examinations of lesions in initial stages of the disease. For prevention, experiments on efficiency of bath treatment with antibiotics (selected on the basis of drug sensitivity testing of isolated bacteria) during transportation of seed was suggested.
Approaches to bacteriological monitoring of pond water and fresh water and fish in experimental ponds fed with treated sewage were discussed with Mr A.K. Gosh. It was suggested that total number of bacteria, Eschericnia coli and Salmonella-Shigella group be monitored in ponds without sewage, ponds with a mixture of fresh water and sewage, as well as in 100 percent treated sewage water. The monitoring should start at the time of pond filling and proceed at regular intervals. Fish should be examined at the same time intervals as water. To determine the influence of sewage on fish health, records on survival rates, disease problems and results of fish checking should be used.
