UNESCO-IHE INSTITUTE FOR WATER EDUCATION
Prevalence and pathology of protozoan and monogenean parasites from fry and fingerlings of cultured Clarias gariepinus (Burchell, 1882) in Uganda
Akoll Peter
MSc Thesis (ES 05.17) October 2005
Prevalence and pathology of protozoan and monogenean parasites from fry and fingerlings of cultured Clarias gariepinus (Burchell, 1882) in Uganda
Master of Science Thesis by Akoll Peter
Supervisors
Dr. Robert. Konecny (Institute of Ecology, Department of Limnology, University of Vienna, Althanstraße 14, 1090 Vienna, Austria)
Dr. Justus. Rutaisire (Faculty of Veterinary Medicine, Department of WARM, Makerere University Kampala, P.O. Box 7062, Kampala, Uganda)
Examination committee
Prof. J. O’keeffe, PhD (UNESCO-IHE, Delft), Chairman Dr. R. Konecny, PhD, MSc (University of Vienna, Austria) Dr. J.J.A. van Bruggen, PhD, MSc (UNESCO-IHE, Delft)
This research is done for the partial fulfilment of requirements for the Master of Science degree at the UNESCO-IHE Institute for Water Education, Delft, the Netherlands
Delft October 2005
The findings, interpretations and conclusions expressed in this study do neither necessarily reflect the views of the UNESCO-IHE Institute for Water Education, nor of the individual members of the MSc committee, nor of their respective employers.
This work is dedicated to my beloved parents Richard and Rita Masai and above all God for the strength He gave me till now.
Abstract
Aquaculture is considered the best option for the dwindling capture fisheries in Uganda. Among the economically important aquaculture fish species with very successful breeding technologies is African catfish, Clarias gariepinus. However, farmers are constraint with massive fry and fingerling mortalities, especially in intensive culture system. Despite the fact that parasites and diseases are reckoned to be the causes of these mortalities, little has been done to authenticate this claim in Uganda. Therefore this study investigated the cause of mortalities with the aim of establishing information on the prevalence and pathology of Protozoa and Monogenea parasites occurring on C. gariepinus. Using routine parasitological and histological techniques, a total of 334 fry and fingerlings of cultured C. gariepinus from three hatcheries were examined for parasite infestations. Five ectoparasites and one endoparasite species were recorded. Of these, the protozoan Trichodina sp. and the monogenean Gyrodactylus sp. were the dominant parasites. The major routes of entry of parasites into the hatcheries were through surface water supply and possibly via the semi aquatic organisms like amphibians. During initial stages of infection, the prevalence, mean intensity and mean abundance of these parasites increased with fish age. At the later stage, Trichodina sp. declined probably due to host resistance and competition for space with Gyrodactylus sp. Trichodina sp appeared less susceptible to the 40ml/l formalin treatment than Gyrodactylus sp. Major pathological changes observed were oedema and hyperplasia in the gills, infiltration of melanomacrophage centers into the skin epidermis and mild epidermal cell hypertrophy of parasitized fish. Hyperplasia and oedema of gill and subsequent gill fusion seem to have interrupted respiration and hence caused death of the host. Concomitant occurrence of Trichodina sp. and Gyrodactylus sp. aggravated pathological effects on fish thus increasing mortality. Epistylis sp., Apiosoma sp. and Trichophrya sp. were considered less pathogenic. The results were discussed in relation to parasite occurrence and parasite induced fish mortalities.
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Keywords: Uganda, Clarias gariepinus, Trichodina sp., Gyrodactylus sp., histopathology, aquaculture.
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Acknowledgements
The successful completion of this study greatly gained contribution from the good will of many people. First and foremost, my sincere gratitude go to my supervisors, Dr. Robert Konecny and Dr. Justus Rutaisire for their tireless efforts in guiding during field study, providing of literature and editing my work.
Special thanks go to Dr. Helmut Sattmann, Director of the Department of invertebrates, Natural History Museum, Vienna, for the support, good hospitality and above all provision of enormous amounts of literature from the commencement of this research project to the end. Thanks go to Dr. Oskar Schachner of the Department of Fisheries, Reptiles and Amphibians, University of Veterinary Medicine Vienna for providing practical support in preparations for sampling and helping in histopathological analysis. Michael Schabuss of University of Vienna for sparing his time to receive me at the airport and editing of my work is greatly acknowledged
I am very grateful to Dr. G. Mbahinzireki, Officer in Charge, Kajjansi Research Station, Mr. Digo Tugumisirize, Proprietor of Sunfish farm Ltd and Mr. Paul Ssebinyansi, Proprietor of Ssebinyansi fish farm for permitting me collect fish samples from their farms. Support from their employees is also highly appreciated.
I am deeply indebted to Assoc. Prof. JD. Kabasa, the Dean, Assoc. Prof. RT. Muwazi, deputy Dean and Dr. T. Amongi, all of the Faculty of Veterinary medicine, Makerere University Kampala and Prof. HA. Joachim, head of Department, Prof. Heinrich Prosl and Prof. R. Edelhofer of the Institute of Parasitology, University of Veterinary Medicine, Vienna for permitting and providing me with facilities to work. Not forgetting Miss Baerbel Ruttkowski, Mr. Hans Homola and the entire staff of the Institute of Parasitology for their support and kind hospitality during my work. Technical assistance to produce excellent histological slides during the study has been provided by William G. Muyombya, MF. Nakamya, Monica Nambi, Jane Atima, of Department of Anatomy, Faculty of Veterinary Medicine Makerere University and Prof. Peter Böck, head of the institute of Histology and Embryology, University of Veterinary Medicine, Vienna.
I am thankful to the people of the Institute of Environment and Natural resources especially Dr. Frank Kansiime, the Director, for allowing me utilise their facilities. Also Dr. FB. Bugenyi, Assoc. Prof. YK. Kizito, Ms. M. Masette and Head of Department, Dr. IG. Basuta, all of Zoology, Makerere University are acknowledged for the assistance they accorded during data collection. Thanks go to Mr. Markus and Denis Fruler of environment Agency Austria for their valuable contribution and not forgetting Franz Jirsa for his valuable discussions on the subject.
I am also deeply indebted to my uncles Malika M.Balayo, Lukoye Alex and their wives: Hajarah Malika and Kibone Lukoye respectively for their continuously supported and encouraged during my study. Also thanks to my parents, brothers, sisters, course mates and all other people whose positive and negative criticisms helped me to produce this quality work. Sincere thanks go the Austrian Ministry of Foreign Affairs and the Royal Government of The Netherlands for funding this.
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Table of Contents Abstract...... i Acknowledgements ...... iii List of figures ...... vi List of tables ...... vii List of symbols ...... viii 1 INTRODUCTION...... 1 1.1 Background...... 1 1.2 Problem statement ...... 3 1.3 Justification...... 3 1.4 Aim of the study ...... 4 1.5 Study objectives...... 4 1.6 Hypotheses ...... 5 1.7 Limitations...... 5 1.8 Organisation of the thesis ...... 5 2 LITERATURE REVIEW...... 7 2.1 Aquaculture in Africa ...... 7 2.2 Aquaculture in Uganda...... 7 2.2.1 Potential cultivable fish species in Uganda ...... 8 2.3 Fish parasites...... 10 2.3.1 Protozoa...... 10 2.3.2 Phylum: Platyhelminthes...... 15 3 METHODS AND MATERIALS ...... 19 3.1 Study area...... 19 3.1.1 Wakiso district...... 19 3.1.2 Mpigi district ...... 23 3.2 Sampling design ...... 24 3.3 Sample size...... 25 3.4 Water and fish sampling methods ...... 26 Biological terms used ...... 30 4 RESULTS...... 31 4.1 Water quality ...... 31 4.2 Parasites...... 33
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4.2.1 Protozoa...... 34 4.2.2 Monogenea ...... 40 4.2.3 Epitheliocystis ...... 42 4.3 Occurrence of parasites ...... 43 4.4 Clinical signs and pathology ...... 56 4.5 Histopathology ...... 56 5 DISCUSSION...... 64 Water quality ...... 64 Parasites: Occurrence and pathology...... 67 Trichophrya sp...... 68 Apiosoma sp...... 69 Epistylis sp...... 70 Epitheliocystis...... 71 Trichodina sp. and Gyrodactylus sp...... 72 6 CONCLUSIONS AND RECOMMENDATIONS...... 82 Conclusions ...... 82 Recommendations to farmers ...... 84 Recommendations to researchers ...... 85 REFERENCES ...... 86 APPENDICES...... 98
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List of figures
Figure 2.2.1: Photograph of African catfish, Clarias gariepinus fingerling from hatcheries in Uganda ...... 9 Figure 3.1: Location of sampling sites in Uganda ...... 19 Figure 3.1.1a: Kajjansi Research Station (KRS) nursing tanks...... 21 Figure 3.1.1b: Sunfish Farm nursing tanks ...... 22 Figure 3.1.2: Fish farmer and Water supplier (Ssebinyansi farm) nursing ponds ...... 24 Figure 3.4: The fish sampling equipments ...... 26 Figure 4.1a: Correlation of water physical parameters from C. gariepinus hatcheries in Uganda ...... 32 Figure 4.2.1a: Photomicrographs of Trichodina sp. from cultured C. gariepinus Uganda in fresh smears ...... 36 Figure 4.2.1b: Photomicrograph of fresh Trichodina sp. indicating measured points in fresh smear...... 37 Figure 4.2.1c: Apiosoma sp. photomicrograph from C. gariepinus fingerlings from Uganda in fresh smears ...... 38 Figure 4.2.1d: Epistylis sp. photomicrographs from C. gariepinus fingerlings from Uganda in fresh smear ...... 39 Figure 4.2.1e: Trichophrya sp. photomicrograph from C. gariepinus fingerlings from Uganda in fresh smears ...... 40 Figure 4.2.2 – i: fresh specimen of Gyrodactylus sp. found on C. gariepinus fry and fingerlings from Uganda...... 41 Figure 4.2.2 – ii: Histological specimen of Gyrodactylus sp. (G) found on gills magnification of C. gariepinus from Uganda...... 41 Figure 4.2.3: Epitheliocystis on the gills of Clarias gariepinus from hatcheries in Uganda...... 43 Figure 4.3.1: Occurrence of Trichodina sp. and Gyrodactylus sp. on C. gariepinus from fish farms in Uganda...... 45 Figure 4.3.2: Occurrence of Trichodina sp. and Gyrodactylus sp. on C. gariepinus from Sunfish farm...... 49 Figure 4.3.3a: Variance – mean ratio of Gyrodactylus sp. on fish by age from Sunfish farm...... 50
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Figure 4.3.3b: Frequency distribution of Gyrodactylus sp. on fish by age (4 – 10 weeks) from Sunfish farm...... 51 Figure 4.3.4: Occurrence of Trichodina sp. and Gyrodactylus sp. on C. gariepinus from KRS...... 53 Figure 4.3.5: Frequency distribution of Gyrodactylus sp. on fish by age from KRS .. 54 Figure 4.3.6: Occurrence of Trichodina sp. and Gyrodactylus sp. on C. gariepinus from Ssebinyansi farm...... 55 Figure 4.3.7: Frequency distribution of Gyrodactylus sp. on fish by age from Ssebinyansi farm ...... 56 Figure 4.5a: Normal gill lamellae of five-week-old fish ...... 57 Figure 4.5b: Gill lamellae oedema...... 58 Figure 4.5c: Hyperplastic gill lamellae...... 59 Figure 4.5d: Rodlet cells present in gills...... 59 Figure 4.5e: Normal structure of C. gariepinus skin ...... 61 Figure 4.5f: Reappearance of goblet cells after formalin treatment...... 62 Figure 4.5g: Caudal fin with a heavy infiltration of the Melanomacrophage centers 62 Figure 4.5h: Attachment of Gyrodactylus sp. on the fish skin ...... 63 Figure 4.5i: Attachment of Trichodina sp. on the fish skin ...... 63
List of tables
Table 3.2: Sampling schedule...... 25 Table 3.3: Sample size...... 25
Table 3.4: Categorisation of parasite intensities on fish...... 27
Table 4.1a: Water quality measurements from three hatcheries in Uganda...... 31 Table 4.1b: Average water inflow, outflow rates and retention time measured at Sunfish farm ...... 33 Table 4.2.1a: Occurrence of Trichodina sp. on cultured C. gariepinus fry and fingerlings in Uganda ...... 35 Table 4.2.1b: Dimensions of Trichodina sp. on fry and fingerlings of C. gariepinus from Uganda...... 37 Table 4.2.2: Occurrence of Gyrodactylus sp. in cultured C. gariepinus fry and fingerlings in Uganda ...... 42
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List of symbols cm Centimetres cm2 Square centimetres DPX Distyrene tricresyl phosphate Xylene e.g. For example FAO Food and Agricultural organisation fish/m Fish per metre h Hours
LC50 Lethal concentration at fifty percent l/sec Litres per second Ltd Limited m metres m2 Square metres MAAIF Ministry of Agriculture Animal industry and fisheries mg/l Milligrams per litre ml/l millilitres per litre mm millimetres NEMA National Environmental Management Authority sp. Species ppt Parts per thousand UK United Kingdom µm Micrometers
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Fish parasites in Uganda Akoll P.
1 INTRODUCTION
1.1 Background
Information on occurrence, prevalence and pathogenicity of fish parasites and diseases is essential in aquaculture. Such information enables aquaculturist to apply correct control measures for any disease outbreak, reduce cost of production, time and manpower while increasing production and profits.
Uganda's fisheries industry in the financial year 2002/2003 contributed US$ 90million to the countries foreign exchange. The majority of the fish came from capture fishery while aquaculture contributed less than 0.1%. In addition, increment of 160,000 tonnes of fish above the current level of 220,000 tonnes is required by 2015 to maintain the consumption rate per caput. This implies that the increase in production of food fish is considered feasible if aquaculture is dramatically increased (MAAIF, 2004). Also, due to over-exploitation of Uganda's fishery, the government has encouraged both private investors and central governments to devote to aquaculture as a way of supplementing the capture fishery (NEMA, 1998; 2000; FAO, 2002). Therefore, to achieve a highly productive and profitable venture, all the constraints faced in aquaculture must be addressed. The major constraints reported to holdback productivity in fish farming include parasites and diseases (Hecht and Endemann, 1998).
Due to the difficulties experienced in Tilapia culture, particularly, those resulting from overproduction and consequent stunted growth and narrow range of environmental conditions (water temperature range: 16 – 35°C, maximum tolerated ammonia concentration: 2.4 mg/l and minimum tolerated oxygen concentration: 1 mg/l) attention is focused on other fish groups. Amongst them included the African catfish, Clarias gariepinus (Burchell, 1882).
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C. gariepinus is characterised by fast growth rate, feeding on a large variety of agriculture by-products, tolerating adverse water quality conditions – water temperature range 8°C to 35°C, for egg hatching: 17°C to 32oC, salinity 0 to 12 ppt, with optimal ranging from 0 to 2,5 ppt, Oxygen: 0 to 100% saturation, wide tolerances range for pH and turbidity and for Unionised ammonia 96 h LC50 (adult fish)= 6.5 mg/l, 96h LC50 (larvae and early juveniles) = 2.3 mg/l. Due to a wide range of conditions under which catfish can live, it can be cultured in high densities resulting in high net yields. It also fetches a higher price as Tilapia's. Upon the successful trials on artificial propagation, which commenced at Sunfish Farm Ltd in 1999 and an increased private sector interest, several farmers adopted the species (Rutaisire, 2005). The species is currently focused as a complement for Tilapia to supplying proteins and generating income, therefore, a target aquaculture candidate in Uganda. In addition, catfish has a huge market as a bait of the lucrative Nile Perch fishery. Mkumbo and Mlaponi (2002) estimated that about 4 million hooks were deployed on Tanzania shores of Lake Victoria, 41% of which used C. gariepinus as bait. As a result, many hatcheries are sprouting up to cover the demand.
However, production of this fish (C. gariepinus) is limited by heavy mortalities of fry and fingerlings as well as high nutritional costs in hatcheries. The causes of heavy mortalities have been attributed to poor nutrient composition; poor environmental conditions as well as parasites, bacterial, viral and fungal infections. Preliminary investigations indicated a possibility of combined effect of parasitic infection (Rutaisire 2005). Parasites such as Trichodina sp., Myxobolus sp., Henneguya sp., Gyrodactylus sp. and Dactylogyrus sp. have been reported to cause most mortality in catfish fry (Basson et al. 1983); Lewis, 1991; Woo, 1995; Tonguthai et al., 1995; Klinger and Floyd, 2002; Reed et al., 2003). Monogeneans especially of genera Gyrodactylus and Dactylogyrus cause extreme loss in aquaculture. However, effects of these parasites, their occurrence and prevalence are not very well known in Africa (Hecht and Endemann, 1998). Such information is vital in formulation of adequate disease control measures. This research is focused on identifying both protozoan and monogenean parasites occurring in C. gariepinus fry and fingerlings as well as investigating their pathological effects on the host and recommending possible control measures.
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1.2 Problem statement
Despite the fact that a lot of research has been done on the biology and ecology of C. gariepinus (Adeyemo et al., 1994; De Graaf and Janssen, 1996; Brzuska, 2004), little is known about the parasites and their effects on the fish host especially in Uganda. Therefore, information is very scanty and unspecific on the occurrence of parasites and effects on the C. gariepinus host. This project is thus aimed at establishing information on the parasites occurring in cultured C. gariepinus fry and fingerlings in hatcheries, prevalence and their histopathological effects. In addition, some private farmers are intensifying aquaculture in order to supplement the over-exploited capture fishery; there is therefore a need to increase fish fry production in a commercially viable manner. It is well known that fry production is the most crucial stage in fish culture, due to their vulnerability to parasites, diseases and any stressful change in their environment. Consequently, heavy fry and fingerling mortalities attributed to parasitic infections must be investigated to lower production already constrained with high nutritional costs, (Axis StorPoint CD, 2001; Hecht and Endemann, 1998). Furthermore, it’s reported that even at low intensities of infection, both protozoans and monogeneans can cause mortality in fry (Woo, 1995).
1.3 Justification
Aquaculture is being focused as the most important supplement of the highly exploited capture fishery in Uganda (NEMA, 1998; 2000). Therefore, the government has set programmes such as the National Agriculture Advisory Services (NAADS) to extend services as well as encouraging people to get involved in aquaculture. The Plan for Modernisation of Agriculture (PMA), which is part of the Poverty Eradication Action Plan (PEAP), aimed at transforming subsistence farmers in agricultural sector to commercial ones. As a result, some farmers have invested in intensive production of C. gariepinus fry. Intensive culture is characterised by high stocking densities (> 1000 fish/m2), high feeding intensities and use of manufactured feeds. However, under such culture conditions, fish tend to be crowded, much confined and release a lot of waste all of which predisposes fish to diseases and parasitic infections. Hence, any slight outbreak of disease or parasite infections can easily be transmitted resulting in high
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Fish parasites in Uganda Akoll P. mortalities (Woo, 1995). Hecht and Endemann (1998) also observed that as aquaculture expands, parasites and the related diseases become eminent. Unfortunately, little has been done on fish parasites and their effects on C. gariepinus in Uganda. Work done in Uganda by Baker (1960, 1963), Khalil (1971) and Paperna (1972, 1996) on feral fishes indicated Ciliophora, Myxozoa and Sarcomastigophora as well as several helminths to occur even in C. gariepinus. However, no specific research on the effect of Protozoa and Monogenea on larvae and juveniles has been carried out (Axis StorPoint CD, 2001).
Losses due to parasitic infestation and associated diseases are great. Bykhovskaya- Pavlovskaya et al. (1964) for instance estimated hundreds of thousands of roubles in Russian feral fishes in the early 1960 and Klontz (1985) estimated the losses to be in the range of 10% - 30% of the total production cost in cultured fish. Due the fact that they can impose heavy economic impact in aquaculture, the situation needs to be addressed. On this basis, documented information on parasites and their effects on the host is essential in reducing such economic losses in fish production and hence the focus of this project.
1.4 Aim of the study
The primary goal for the study is to record the protozoan and monogenean parasites found in Clarias gariepinus fry and fingerlings in hatcheries and their histopathological effects.
1.5 Study objectives
• To characterize the important physical and chemical parameters of the ponds (Temperature, Dissolved oxygen, pH, Nitrite and Ammonia). • To investigate the prevalence of protozoan and helminth parasites on C. gariepinus fry and fingerlings. • To assess the histopathological effects of the parasites on the fish fry and fingerlings.
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1.6 Hypotheses
• Protozoan parasites do not occur on/in C. gariepinus fish fry and fingerlings in hatcheries. • Protozoan parasites occurring on/in C. gariepinus fish fry and fingerlings in hatcheries have a commensalistic rather than parasitic relationship with fish. • Helminths parasites do not occur in C. gariepinus fish fry and fingerlings in hatcheries and are not pathogenic to the host.
1.7 Limitations
The study was to establish the cause of fish mortality in ponds by investigating the prevalence of protozoan and monogenean parasites and assess their pathology. However, there were some limitations to this study • There is lack of information on the normal histology of C. gariepinus. • Due to shortage of time, diurnal changes of water quality were not considered.
1.8 Organisation of the thesis
Already presented in section 1 was the introduction containing the background of the study, the problem that stirred the research, the study aim and the specific objectives. In the proceeding sections, first, in section 2, the literature review providing highlights of the previous studies. Subsections 2.1 and 2.2 give information on the development of aquaculture in Africa and the extent of the practise in Uganda as well as data on cultivable species, potential as aquaculture candidates and their loopholes. Subsection 2.3 deals with parasites encountered in fish, extent in work done in Africa and Uganda with emphasis on Protozoa and Monogenea. Under which, Protozoa and Monogenea parasitizing C. gariepinus both in feral and culture conditions as well as their pathology are stated. Secondly, section 3, materials and methods present information on the choice and use of particular tools and strategies for data gathering and analysis as well as study areas. Thirdly, section 4, results under which both field and laboratory data collected are presented. Fourthly, section 5, discussion, the data presented under results are explained, relating results obtained to other work as well as fitting it into the context of the field.
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Fifth, section 6, conclusion and recommendation provide the significance of the finds and fulfilment of the aim while under recommendations, presentation of mitigation measures and further research gaps. Finally, references, where authors of various articles, books, magazines etc are acknowledged.
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2 LITERATURE REVIEW
In this chapter, related work on general aquaculture in Africa and Uganda as well as parasites of fish with emphasis on Protozoa and Monogenea, their occurrence and pathogenicity is presented.
2.1 Aquaculture in Africa
In comparison to the rest of the world’s aquaculture production, Africa’s contribution is insignificant. The continent as whole contributes a mere of 0.9 % to the total world aquaculture production (FAO, 2002; 2003). The production however increased from 37, 000 tonnes in 1984 to 189, 000 tonnes in 1998. In 2002 the total aquaculture production of Africa amounted to 451,537 metric tons and the major fish species cultured include the freshwater carp and tilapia. Leading producers are Egypt (376,296), Nigeria (30,663), Zambia (4,200), South Africa (4,177), Madagascar (7,966), Ghana (6,000) and Uganda (4,915) all in tonnes. These figures reveal the low level intensity of aquaculture in sub-Saharan Africa as compared to 553,933 tonnes produced by Norway.
Among the constraints of economic importance in farmed fish are diseases and parasites (Bykhovskaya-Pavlovskaya et al., 1964; Ellis, 1985; Klontz, 1985; Pike and Lewis, 1994; Woo, 1995; Axis StorPoint CD, 2001). However, in Africa, Hecht and Endemann (1998) revealed that very little research has been undertaken on diseases and parasites of fish in aquaculture. This was attributed to the low intensity of aquaculture in the region. However, with the current rapid development of African aquaculture, the prevalence of diseases and parasitic infections will also increase.
2.2 Aquaculture in Uganda
Fish has become one of the highest income earner to Uganda complementing coffee, contributing 4.9 % of the total export and fetching US$ 90 million (Ministry of Finance, Planning and Economic Development, 2003).
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Previously, fishing industry depended ultimately upon capture fisheries, however, with increased demand both for local and export; the sector is threatened with over exploitation. This has called for supplementation with the only option being through aquaculture (Ministry of Finance, Planning and Economic Development, 2003; NEMA, 2000; FAO, 2003).
2.2.1 Potential cultivable fish species in Uganda
Fish farming in Uganda started in 1950’s on subsistence level with Oreochromis niloticus (Nile tilapia), and Tilapia zilli as the culture fish species. In 1994, on realization of unsustainability of capture fishery and drastic decline in species diversity, aquaculture was strengthened. As a result, culture fish species for Uganda increased and currently, the most important include: Oreochromis niloticus (Nile tilapia), Tilapia zilli, Clarias gariepinus (African Catfish), Cyprinus carpio (Common carp; introduced species), Labeo victorianus whose reproductive and artificial breeding techniques are described by Rutaisire (2003), Lates niloticus (Nile perch) and Bagrus docmac (both of which are still under trials at Kajjansi Research Station (KRS)).
Oreochromis niloticus (English name: Nile tilapia; Ugandan local name: Ngege)
This is a widely spread cultured species in Uganda and it’s culture started as early as 1953. However, due to high reproduction rates even in pond (usually at 3 – 4 months after stocking), stuntedness is a consequence usually due to competition. Efforts to reduce this include the following: First, introduction of predator like C. gariepinus, which, according to De Graaf (1994), the African catfish (C. gariepinus) is considered a “lazy” predator, as it prefers to feed on the artificial food supplied. However, they can eliminate the offspring of O. niloticus if they are stocked in high densities (10,000 fingerlings/ha). Secondly, monoculture (culture of males only) e.g. Popma and Masser (1999) states that in monosex grow out ponds under good conditions, males generally reach a weight of ≥ 200 grams in 3 to 4 months, ≥ 400 grams in 5 to 6 months and 700 grams in 8 to 9 months. However, sorting efficiency is only up to 95% hence possibilities of further reproduction. Finally, male sex reversal by chemical treatment is under trial at KRS.
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Clarias gariepinus (English name: African catfish; Ugandan Local name: Male)
This study will focus on C. gariepinus (Fig. 2.2.1), which is among the major commercial fish species cultured in Uganda. It culture started in early 1990s. Due high tolerance to adverse environmental conditions – tolerating high stocking densities in ponds, less selective feeding behaviour and rapid growth rate – C. gariepinus is considered a very good aquaculture candidate. In addition to air-breathing ability it’s a potential fish for intensive culture systems without prerequisite pond aeration or high water exchange rates. The growth rates are high especially during the early stages of development. During the first year it is reported to increase between 200 and 300mm total length (TL) and in subsequent years length increments vary between 80 and 150mm (Axis StorPoint CD, 2001). Together with successful attempts to spawn it artificially in ponds has increased its favour as an aquaculture species worldwide.
Figure 2.2.1: Photograph of African catfish, Clarias gariepinus fingerling from hatcheries in Uganda However, still some deficits have remained with artificial production of fry in hatcheries. First, there is high nutritional cost during the changing from endogenous to exogenous feeding – ontogenic shift. During this period, fry must be fed on either rotifer of <50µm or Artemia nauplii. However, use of locally available cheap rotifers is a problem because of lack of synchronization of feeding shift with rotifer production and the importation of Artemia nauplii is rather expensive. Secondly, there are high mortalities of both fry and fingerlings, which have been attributed to fungal, bacterial and parasitic infections.
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Although it is reported that low infections of protozoa have low or no pathology on adult catfishes from the natural conditions (Axis StorPoint CD, 2001), severe impacts may be observed under culture conditions. In larvae and fry on the other hand, protozoan parasites were found on the skin as well as the gills and reported that even under low infections result in high mortalities. This further increases the cost of production of fish on both small and large-scale basis.
Therefore, in the next section, discuss parasites occurring in fish, then focus on Protozoa and Monogenea then later highlight those using C. gariepinus as a host. Further, general information on the pathology of these parasitic groups is provided.
2.3 Fish parasites
Parasites can be encountered with any fish species and within any type of aquatic system. They range from viruses, bacteria, parasitic protozoans, acanthocephalans, nematodes, trematodes, cestodes, and crustaceans (Lewis, 1991). In Africa however, very little work has been done on fish parasites. Thus not many parasites have been reported in C. gariepinus. Those reported to be problematic in African aquaculture include: ectoparasites; protozoans like Chilodinella sp., Trichodina sp.; monogeneans; leeches; crustaceans and larval bivalve molluscs. Endoparasites recorded include: several protozoans, for example Ichthyophthirius multifilis, haemoflagellates, apicomplexans, myxosporeans, microsporeans, as well as digenean trematodes, cestodes, acanthocephalans and nematodes (Hecht and Endemann, 1998).
2.3.1 Protozoa
Protozoa are single celled eukaryotic organisms belonging to the Kingdom Protista, which includes members of the Phyla of Ciliophora (ciliates), Sarcomastigophora (flagellates), Myxozoa (myxosporidians), Microsporea and Apicomplexa. They range from free-living through various forms of commensalisms to parasitism in most animals, plants, and even other protozoans (Lucy and Ernest, 1994; Woo, 1995). More than 65,000 species have been described. About 10,000 species are parasitic with over 2400 parasitizing fishes (Lucy and Ernest, 1994).
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The most abundant protozoan groups parasitizing fish are: the myxosporidians followed by the microsporidians and the ciliates – also are the most complex group (Bykhovskaya-Pavlovskaya et al., 1964; Klinger and Floyd, 2002).
Several authors have described structural and morphological characteristics of protozoan parasites in fish for instance: Bykhovskaya-Pavlovskaya et al. (1964); Lom and Halder (1977); Lom and Noble (1984); Mehlhorn (1988); Lucy and Ernest (1994) as well as Woo (1995). Reproduction and transmission of protozoan parasites is very diverse. Despite the fact that most of them have direct life cycle, many have indirect life cycles. During direct life cycle, Protozoa are usually released either through faeces, especially for intestinal parasites or when the host dies and the spores or cysts can also be shed directly into water via lesions. These can gain entrance into a new host through ingestion or active attachment of the trophont. On the other hand, indirect cycle like haemoflagellates usually require an intermediate host (usually blood sucking leeches). Detailed individual life cycles can be obtained in Mehlhorn (1988); Kreier (1994); Woo (1995); Overath et al. (1999).
Occurrence and pathology of Protozoa in African fishes
Pathogenic effects of protozoan parasites vary greatly from species to species. Among the common pathological observations are atrophy, hypertrophy, hyperplasia, general anaemia, necrosis and host death. Additional and different histopathological changes can be observed by different species within a phylum. Most of these pathological studies were done on European, American and Asian fishes (Rintamaki et al., 1994; Woo, 1995; Kent et al., 1997) but in comparison with some work from South Africa (Basson and Van As, (1987), Egypt (El-mansy and Bashtar, 2002) and West Africa (Fomera, et al., 1992; Faye, et al., 1994) same effects were observed.
Phylum: Sarcomastigophora
Ectoparasites like Piscinoodinium sp. (Oodinid) are associated with excess mucus secretion, skin ulcers, sloughing of the epithelium necrosis, gill filament degeneration and hyperplasia. The conditions are more pathogenic in young fish; they can kill the host after 1 – 2 weeks of infestation (Noga and Levy, 1995).
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Ichthyobodo sp. and Cryptobia sp. (Family Bodonidae) causes hyperplasia of the epithelium cells of the gill lamellae in catfish; hyperplasia of malpighian cells, oedema followed by degeneration and sloughing of the epithelium in salmonids (Woo, 1995). On the other hand, haemozoic – blood endoparasites like trypanosomes (haemoflagellates) cause anaemia (Woo and Poynton, 1995). Trypanosomes so far recorded in Africa are Trypanosoma mukasai, T. toddi and T. tobeyi (Baker, 1960). It is also reported by Baker (1960) that T. toddi and T. tobeyi do occur in C. gariepinus.
Phylum: Myxosporea
Myxozoans are the largest group of the protozoan parasites and known to occur in almost all organs and tissues of fish. They are reported to cause heavy infections with prevalence up to 100%; extensive lesions as well as mortalities in both captured and cultured fishes (Lom and Dykova, 1995). Several species of this group have been reported or described - Bykhovskaya-Pavlovskaya et al. (1964); Landsberg, (1986); Lewis (1991); Lom and Dykova (1992); Lucy and Ernest (1994); Ghaffar et al. (1995); Lom and Dykova (1995); Kent et al. (1997); Bartholomew et al. (1997); Palenzuela et al. (2002). Myxozoans are recorded to have the longest infection in fish for instance Myxidium lieberkueluni was recorded to take one year in the urinary bladder of the pike (Lom and Dykova, 1995).
Histopathological changes caused to the host are granulomatous inflammation due to tissue response; hyperplasia, hypertrophy as well as atrophy are observed. Coelozoic infections result in inflammations, which turn into jaundice and/or necrosis, hyperaemia, oedema and haemorrhage. Therefore, myxosporean infections result in both regressive (atrophy and necrosis) and progressive (hypertrophy and hyperplasia) pathological changes (Lom and Dykova, 1995).
Nine species of Myxozoa out of 137 described in Africa were reported to parasitize Clarias sp. (Reed et al., 2003). They include: Myxobolus gariepinus; M. comoei; M. clarii; Henneguya branchialis; H. clariae; H. fusiformis; H. samochimensis; H. laterocapsulata and H. suprabranchiae.
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Fish parasites in Uganda Akoll P.
Phylum: Microsporea
These are obligatory intracellular parasites with a very wide range of hosts: from protozoa to man. In fish, microsporidians are recorded to causes severe diseases in particularly cultured fish. They can form xenomas (extensive hypertrophy or tumour- like out growth with fragmentations of the nucleus). This is also known as hyperbiotic. For non-xenoma forming species, hypobiosis of cells do occur – regressive changes (Dykova, 1995; Lom and Nilsen, 2003). However, the end result is the same for both manifestations i.e. infected cell under go dystrophy, atrophy and necrosis.
From a catalogue of described genera and species of microsporidians parasitic in fish by Lom (2002), 34 species from seven genera: Loma, Pleistophora, Glugea, Microfilum, Neonosemoides, Nosemoides and Microsporidium were reported from African waters. Of these, three species belonging to two genera of Loma and Pleistophora also occurred on freshwater fish species while thirty species from the six genera of Loma, Pleistophora, Glugea, Microfilum, Nosemoides and Microsporidium parasitized marine fishes and one species – N. tilapiae (Faye, Toguebaye & Bouix, 1996) of the genus Neonosemoides was found on Tilapia zillii, T. guineensis and Sarotherodon melanotheron of brackish water off the coast of Benin. However, of all the genera mentioned above from Africa, none was found on Clariidae, thus, microsporidians parasitizing C. gariepinus are not well documented.
Phylum: Ciliophora
Ciliates are the most complex (Bykhovskaya-Pavlovskaya et al., 1964; Mehlhorn, 1988; Lom and Dykova, 1992), most common and widely distributed (found in any water system) group of protozoan parasites and/or commensals of fishes (Wellborn, 1967; Basson et al. 1983; Dickerson and Dawe, 1995). They are ectoparasites: found on gills and skin of fish and other organisms except for Trichodina oviducti, T. urinaria, and genera of Vauchomia and Paratrichodina, which are endoparasites (Lom, 1995). Heavy losses due to ciliates are usually associated with unfavourable stressful environmental conditions for the fish host. Therefore, they are known to cause problems in farmed fish’s especially young fish, which are kept in high densities in hatcheries (Basson and Van As, 1991; Dickerson and Dawe, 1995; Lom, 1995; Rintamaki-kinnunen and
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Fish parasites in Uganda Akoll P.
Valtonen, 1997).
Ciliate infections are characterised by: hyperplasia of gill epithelial cells (if the occur on gills), skin lessons, haemorrhages, excessive mucus secretion, sloughing of the skin and general oedema. Necrosis together with proliferation of the epithelial cells of the secondary gill lamellae which may later fuse reduce the respiratory surface or even render the gills non-functional (Lom, 1995; Dickerson and Dawe, 1995; Matthews, 1994). However, the pathological manifestation varies and depends upon the intensity of infection (Lom, 1973a; Lom, 1995; Hecht and Endemann, 1998). Ciliates so far recorded to occur specifically in C. gariepinus are Trichodina marietinkae (Basson and Van As, 1991) and Ichthyophthirius multifilis (Hecht and Endemann, 1998)
Phylum: Apicomplexa
This group of protozoans occurs in a wide variety of organisms of kingdom animalia (helminths to man) but little is known about those parasitizing fish (Molnar, 1995). However, Molnar (1995) mentioned that about 60 species of haemogregarinids, 2 species of cyrilics, 1 species of hepatzoon, 5 species of babesiosomas and 2 species of dactylosomas have been described in fish. Molnar (1995) added that though majority are relatively of low pathogenicity, lethal infections have been reported in farmed fish and few cases in feral. Apicomplexans are endoparasites occurring in the intestine, gall bladder, swim bladder, liver, connective tissues surrounding the oocytes and kidney. The damage to tissues depends on the intensity of infection (Bykhovskaya-Pavlovskaya et al., 1964; Mehlhorn, 1988; Molnar, 1995; Hecht and Endemann, 1998; Azevedo, 2001). Among the most common apicomplexans occurring in Africa include Eimeria vanasi and Goussia cichlidarum (Hecht and Endemann, 1998). Hecht and Endemann (1998) added that some species of Eimeria are reported to parasitise C. gariepinus.
Therefore, information on the groups of protozoans occurring in African fishes still remains scanty. In addition, parasites are widely spread for instance; Basson and Van As (1993) identified Trichodinid protozoans in South Africa previously recorded in Europe. This indicates that once temperate parasites are introduced in tropics may survive within closely related species of fish or vice versa. Therefore, compilation and documentation of such information will be of great importance especially in formulation
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Fish parasites in Uganda Akoll P. of regulations as well as control measures.
2.3.2 Phylum: Platyhelminthes
Members of the phylum Platyhelminthes also called flatworms are dorsoventrally compressed, un-segmented bilateratelly symmetrical bodies that lack coelom (aceolomates). Unlike unicellular Protozoa, Platyhelminthes are multicellular, also called metazoans. Based on classification presented in (Williams and Jones, 1994; Cone, 1995; Cribb et al., 2002; Myers et al., 2005) flatworms belong to the Kingdom Animalia and has of four classes – Turbellaria, Monogenea, Trematoda and Cestoda. The phylum has over 20000 species described thus, the largest group without coelom of which some are free living while others are parasitic.
Turbellaria consists of about 3000 species dominating marine environments though some do occur in freshwater as well like Planarians. Although most members of this class are free living, few are reported to be parasitic e.g. order Temnocephalida (Campbell, 2001). Class Trematoda consists of order Digenea and their allies of the order Aspidogastrea both constituting to about 9000 species are all parasites usually on both the cold-blooded and the warm-blooded vertebrates. The order Aspidogastrea is also reported to parasitize marine mollusks (Myer, 2001; Cribb et al., 2002). Most are endoparasites and only few are ectoparasitic. The class also has indirect and complex life cycles requiring in most cases an intermediate host. Cestoda are all endoparasites of vertebrates with over 5000 species so far described. Most require at least intermediate host to complete their life cycle as adults in final host. Monogenea have about 3000 – 4000 species, most of which are ectoparasites of aquatic organisms (Cribb et al., 2002). This class is of great economic importance in regard to fish health and fish farming around the globe. Schäperclaus (1979) placed genus Gyrodactylus as number one pathogenic parasite of fish above all other helminths. Cribb et al. (2002) noted that two monogenean families of Gyrodactylidae (Van Beneden & Hesse, 1863) and Dactylogyridae (Bychowsky, 1933) dominate literature and mostly either during description of new species belonging to the two genera occurring in fish or as a cause of massive fish mortality. Therefore, based on this information and economic implications due monogeneans, they are will be dealt with in detail.
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Monogenea (Bychowsky, 1937)
Members of Monogenea are mainly ectoparasites of poikilothermic organisms especially fish (Cribb et al. 2002), except few like Oculotrema hippopotami that occur in the eyes of hippopotamus, some genera of Dactylogyridae like Acolpenteron and Kritskyia found attached on the urinary duct and bladder (Cone, 1995; Guidelli et al., 2003, Viozzi and Brugni, 2003) and Enterogyrus found attached to the wall of the foregut of Tilapia Zillii, Oreochromis niloticus and O. mossambicus (Williams and Jones, 1994; Cone, 1995; Paperna, 1996). Concerning fish parasites, the greatest diversity has been reported for bony fish but only eight families are reported to occur often on cartilaginous fish (Cribb et al 2002).
The class has two orders. First, Monopisthocitylea, which includes members of the dominant and economically important families like of Gyrodactylidae, Dactylogyridae, Monocotylidae, Diplectanidae, Capsalidea and others. Secondly Polyopisthocotylea having families like Chimaericolidae, Diplozoidae and many others are reported to cause problems in Aquaculture.
Morphologically, monogeneans are dorsal – ventrally flattened with opisthaptor or haptor armed with hooks and/or suckers for attachment on to the host. Detailed description of the morphology of monogeneans is provided in Bykhovskaya- Pavlovskaya et al. (1964), Beoger and Kritsky (1993), Milan and Radim (1994), Cone (1995), Paperna (1996). Monogeneans are hermaphrodites and are either oviparous (egg laying) or viviparous (live bearing) modes of reproduction. They have a direct mode of transmission requiring no intermediate host. With exception to Gyrodactylus, which are viviparous, all monogeneans lay eggs; develop in free-swimming ciliated larvae called oncomiracidium, which on contact with the suitable host develop into adult. Detailed information on life cycles of Monogenea can be obtained in Williams and Jones (1994), Cribb et al. (2002), Milan and Radim (1994), Soleng et al. (1999), Paperna (1996), Reed at al. (2002). In the next subsection, additional information on the occurrence as well as pathogenicity
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Fish parasites in Uganda Akoll P. of monogeneans on C. gariepinus and freshwater fish generally in Africa and Uganda in particular is provided.
Occurrence and pathology of Monogenea in African fishes
Monogenea are the most abundant and ranked the most important pathogenic ectoparasites of fish among other Platyhelminthes (Schaperclaus, 1979; Williams and Jones 1994; Cone, 1995). Despite numerous reports of problems in fish associated with monogeneans and description of new species worldwide, relatively few reports on the subject come from Africa and Uganda in particular. Fish helminths occurring in Africa were summarised by Khalil (1971). With the current new discoveries and description, the list must be longer than provided by Khalil (1971). Example of some of the discoveries and descriptions are Paraquadriacanthus nasalis by Ergens (1988) and five new species of genus Cichlidogyrus by Douellou (1993). In addition, information regarding the pathology and distribution of some of these monogeneans was by Paperna (1996). This clearly shows the lack of research capacity or poor documentation of information, hence a likelihood of existence of gaps in parasitological information. Regarding occurrence, ten monogeneans genera that include, Dactylogyrus, Gyrodactylus, Diplectanus, Macrogyrodactylus, Pseudogyrodactylus, Enterogyrus, Cichlidogyrus, Cleidodiscus, Acolpenteron and Pseudoacolpenteron have been reported from African fish (Hecht and Endemann, 1998). However, Khalil (1971) listed up to 26 genera: Afrogyrodactylus, Gyrodactylus, Macrogyrodactylus, Afrocleidodiscus, Ancyrocephalus, Annulotrema, Bagrobdella, Characidotrema, Cichlidogyrus, Cleidodiscus, Dactylogyrus, Dogielius, Eutrianchoratus, Gussevstrema, Heteronchocleidus, Heterotesia, Nanotrema, Neodactylogyrus, Onchobdella, Protoancylodiscoides, Protogyrodactylus, Quadriacanthus, Shilbetrema, Archidiplectanum, Diplectanum and Diplozoon. Those genera and/or species parasitizing Clarias sp. include: Gyrodactylus (e.g. G. groschafti, Macrogyrodactylus – M. clarri, and M. congolensis (Khalil and Mashego, 1998; El- Naggar et al. 2001), Quadriacanthus. Paperna (1996), however, added that monogeneans parasitizing Clarias spp. and Cichlids in Africa belong to similar families occurring in the Mid East.
The pathogenic effects of Monogenea vary from species depending upon the nature of
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Fish parasites in Uganda Akoll P. haptor, site of attachment and mode of feeding. Dactylogyrus for instance having a haptor that provide tender attachment of the parasite on to the host while Gyrodactylus having a powerful haptor that is usually inserted deep into the hosts’ tissues. Moreover, unlike the thick epithelium of the skin, parasites preferring gill cause more damage to the host due to the thin gill epithelium. Even so, common pathological changes due to parasites attached on to gills are excessive production of mucus, decolouration of gills, hyperplasia of epithelium leading to fusion of gill lamellae, which eventually cause clubbing and fusion of gill filaments, necrosis of tissues. Monogeneans that occur more often on skin cause sloughing of the skin hence decolouration, excessive mucus production. Like in the gills, hyperplasia of the skin epithelium and lesions due to attachment discs act as point of secondary infection. General outcome of parasite infestation is death of the host. Detailed information on pathological effects of different monogenean species has been described in Williams and Jones (1994), Cone (1995), Paperna (1996), Reed et al. (2002), Buchmann et al. (2004), Mansell et al. (2005).
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Fish parasites in Uganda Akoll P.
3 METHODS AND MATERIALS
3.1 Study area
Three C. gariepinus hatcheries located in the Wakiso and Mpigi districts of Uganda (Fig. 3.1) where aquaculture practices are somehow well developed were sampled.
Ssebinyansi farm Wakiso
Kajjansi Research Station Sunfish farm
Figure 3.1: Location of sampling sites in Uganda.
Source: Mapquest (2005)
3.1.1 Wakiso district
The district is located in the central region of Uganda neighbouring districts of Luwero and Kampala in the North, Lake Victoria (Mukono) in the East, Kalangala on Lake Victoria in the south and Mpigi in the west. The district lies at an altitude ranging from 1,000m to 1,552m above sea level about eight kilometres from Kampala on Entebbe road. Temperature ranges from a minimum of 16°C and a maximum of 27°C; with an annual average temperature of 21.9°C. Sampled farms located in this district were Kajjansi Research Station (KRS) and Sunfish farm Ltd.
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Fish parasites in Uganda Akoll P.
In the next paragraphs, detailed information of each fish farm on water supply, feeding practise and facilities available for fish hatching and nursing are presented.
Kajjansi Research Station (KRS)
This is a government funded research centre engaged in practicing general research in all fish and fish related activities especially, development of fish farming techniques. Among the fish species hatched or produced under artificial hatching experiments are Oreochromis niloticus, C. gariepinus, Bagrus sp. Labeo victorinus mirror carp, Goldfish etc. Development of C. gariepinus commenced in early 1990s and is currently practised at extensive level of farming with stocking rates of 2 to 100 fish/m2 based on the age. Hatching is done artificially after injection of pituitary extract from the same species for induction of ovulation especially in females. Ovulated females are stripped; spread on hatching trays and eggs placed in plastic or concrete and tiled tanks supplied with groundwater for incubation. Water is however aerated by free fall into a holding tank and warmed to optimum temperature of 25 – 28°C for hatching prior to entering the incubation tanks. Three days post-hatching, fish are fed on zooplankton obtained from ponds fertilised two weeks pre-hatching. After three weeks of hatching, the stocking density of fry was reduced (thinned) and transferred to concrete untiled tanks (Fig 3.1.1a) supplied with surface water from Kajjansi stream, filtered through a fine mesh (undisclosed size but able to hold back clay particles). During this time, fish are feed on dry protein rich feeds and some zooplankton. In these tanks, fish stay for two weeks before being transferred to earthen ponds fed intermittently with surface water filtered to remove large debris. Apart from zooplankton obtained from pond water, fish are constantly provided with dry feed ratios and stay in these ponds till they are sold off.
KRS therefore, uses underground water for hatching and surface water for nursing and raising fish. However, groundwater is used till fish are three weeks old and at the start of supply of surface water, the water is filtered using a fine mesh net. Fish are fed on both artificial and live feeds in a more or less same proportion. The hatchery has concrete tiled and plastic tanks for hatching and raising fish up to five weeks old before transferred to earthen ponds supplied intermittently with more or less unfiltered water.
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Fish parasites in Uganda Akoll P.
Figure 3.1.1a: Kajjansi Research Station (KRS) nursing tanks
Sunfish farm
This privately owned farm is located not more than 50m away from KRS practicing intensive production of C. gariepinus. The farm is also involved to the least extent in raising Oreochromis niloticus, Lates niloticus, C. gariepinus and Cyprinus carpio. Like at KRS, hatching is artificial involving injecting females with pituitary extracts in physiological solution and stripping. Fertilised eggs are spread onto hatching tray in plastic or concrete tiled tanks receiving groundwater. The water is also aerated by free flow into holding tanks and heated to appropriate temperatures for hatching prior to supplying incubation tanks. Two weeks post-hatching, thinning and sorting is done, transferred to tanks supplied with a mixture of ground and surface water. Surface water is drawn from a reservoir located less than 100m from the ponds and tanks, which receives water from Kajjansi stream (same stream that supplies KRS but form up stream). At this farm filtering nets function as a barriers to wild fish from entering the nursing tanks. Thus the farm is faced with accumulation of clay and other debris – sieved out manually after every two or three days. During sieving process, fish would also enter a siphon and recollected at the end of the process hence fish would get stressed frequently.
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Fish parasites in Uganda Akoll P.
At four weeks post-hatching, fish are transferred to tanks (Fig 3.1.1b) supplied with flow through surface water only from the reservoir, at stocking densities of above 100 fish/m2 under a constant temperature provided by a greenhouse. It should be noted that fortnightly, fish were manually graded (fish are put on a flat surface then with fingers, larger fish are separated from the rest into another container). Coupled with bruises from manual grading and the tender age (low immune system and softer skin), fish are stressed and hence may be predisposed to parasite and disease infections. Feeding of fish at this farm is mostly on artificial (locally made) although zooplankton is supplemented during the ontogenic shift up to four weeks. However, from four weeks on, fish are fed on dry protein rich ratio. In this farm, 30 and 40mg/l formalin bath treatment was applied at seven and nine weeks respectively after experiencing massive mortalities.
Sunfish farm uses groundwater for hatching and surface water for nursing and raising C. gariepinus. Moreover, supply of surface water starts as early as two weeks post- hatching. Feeding is mostly on artificial feeds supplemented with live feeds for three days old to ca. four weeks old fry. Due to intensive nature of the farm, hatching is done in plastic and concrete tiled tanks while fish nursing is done in thoroughly polyethylene- paper-lined ponds and occasionally in happas except for brood stock, which are kept always in earthen ponds.
Figure 3.1.1b: Sunfish Farm nursing tanks.
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Fish parasites in Uganda Akoll P.
3.1.2 Mpigi district
Mpigi district is also located in the central region of Uganda, bordered by districts of Luwero and Mubende in the north, Wakiso in the East, Sembabule in the southwest, Masaka and Kalangala across Lake Victoria in the south. It lies at an approximate altitudinal range of between 1,182m - 1,341m above sea level and has a minimum temperature of 11.0°C and maximum of 33.3°C with an average of 22°C. One sampling site – Fish Farmer and Water Supplier hatchery (Ssebinyansi farm) was located in this district and practiced extensive system (Fig 3.1.2).
Ssebinyansi farm
Fish Farmer and Water Supplier hatchery commonly known as Ssebinyansi farm after the owner, is an extensive privately owned farm. The farm is also engaged in hatching of C. gariepinus. Like the Sunfish farm and KRS, Ssebinyansi farm also grows Oreochromis niloticus for subsistence consumption. Hatching of catfish is semi natural: involving induction of ovulation in females by injection of hypophysis solution, then kept in tanks supplied with groundwater equipped with artificial substrate like sisal and covered with black polyethylene papers. Following spawning, usually after 14 hours, parents are removed and eggs thinned by splitting the substrate. The temperature is then gradually raised to about 24°C to accelerate hatching. Three- day-old larvae are fed on zooplankton harvested from ponds fertilised two weeks prior hatching and this continues till fish are four weeks. After four weeks of hatching, fry are transferred to net cages made of fine mesh (happas) in earthen ponds (Fig 3.1.2) supplied exclusively with groundwater. The happas are however changed every fortnight with larger mesh sized nets following grading or sorting. In here, fish depend mostly on nature production of ponds, which are fertilised two weeks prior to fish transfers. However, dry feed supplements are also provided occasionally. Ponds were only drained into drainage channel by seepage. Therefore, there was no direct connection between the pond water and drainage channel water, which flows into a wetland. It’s therefore worth noting that Ssebinyansi farm use exclusively groundwater for hatching, nursing and raising of C. gariepinus. The farm depend to a large extent on natural productivity of ponds for feeding, which are fertilised two weeks prior to hatching and transfers.
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Fish parasites in Uganda Akoll P.
The farm also has concrete tiled tanks for hatching and earthen ponds with and/or without happas for nursing of fish. Based on this information, the farms had more or less similar hatching methods, hatching water and first week feeding practice. They differed in nursing and raising equipments, water supply especially when surface water first introduced and proportion of provision of dry feeds.
Figure 3.1.2: Fish farmer and Water supplier (Ssebinyansi farm) nursing ponds.
3.2 Sampling design
One cohort (those individuals of a stock hatched in the same time of breeding season) was chosen and followed from all hatcheries for this study, to avoid possible inherited variations in susceptibility to infections. Due to the delays in hatching at KRS and At Ssebinyansi farm sampling reached seven and five weeks respectively. At Ssebinyansi farm, 10 week old fish from the previous cohort was sampled to provide a clue of the health status at the farm. Sampling started one week post-hatching when fish were expected to have almost fully developed organs. Three sampling times per week i.e. once per farm for all three farms, with seven days interval between each sampling day was followed (Table 3.2) till ten weeks e.g. at Sunfish farm or when the sample duration expired e.g. at KRS and at Ssebinyansi farm. During each sampling time, 20 fish were collected from each hatchery but on most occasions, farmers never allowed 20 fish to be sacrificed, thus the number collected each sampling time varied between 12 to 30 fish.
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Fish parasites in Uganda Akoll P.
Water quality parameters were measured insitu concurrently with fish samples for parasite investigations. This was aimed at obtaining static information on the relationship of water quality and parasite as well as effects on fish.
Table 3.2: Sampling schedule
Sampling day Hatchery sampled Monday Sunfish farm Wednesday KRS Friday Ssebinyansi farm
During analysis, at least one hour was spent on each fish especially those being parasitized.
3.3 Sample size
A total of 334 fry and fingerlings of cultured C. gariepinus were investigated for the presence of parasites from three farms. Sampling was done weekly at mid morning and followed up of a cohort commencing at one week until ten weeks. A summary of the total number of fish per age and farm is shown in table 3.3.
Table 3.3: Sample size; by age, farm and total (- Indicate no data collected)
Farms Age Sunfish KRS Ssebinyansi farm Total
1 9 11 10 30
2 28 27 15 70 3 17 14 - 31 4 18 26 16 60 5 21 10 12 46 6 10 10 - 20 7 15 15 - 30 8 12 - - 12
9 14 - - 14
10 15 - 6 21
Total 162 113 59 334
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Fish parasites in Uganda Akoll P.
3.4 Water and fish sampling methods
All water parameters – dissolved oxygen, ammonium, nitrite temperature and pH were measured insitu using water kits and probes. Temperature and oxygen were measured using YS1 Environmental DO 200 probe while for pH, ammonium and nitrite using the Visocolor Test kit, model 300 (Germany). The measurements where repeated using the Hach test kit, Model OX-2P for dissolved oxygen, Lemotte Freshwater Aquaculture for ammonium and nitrites and Corning 313 pH/T°C meter (UK) probe for pH and temperature.
C. gariepinus fish specimens were captured using sieves from hatching tanks and scoop nets from nursing tanks or ponds as well as happas (Fig 3.4a and 3.4b respectively) from all the selected hatcheries (Sunfish farm, KRS and Ssebinyansi farm) for parasitological examination between the period of 21st April and 30th June 2005. Fish samples from farms were transported alive in locally improvise containers to the laboratory where parasite investigation was done immediately or within twelve hours in case of fish samples from distant farm of Ssebinyansi farm.
a b
Figure 3.4: Shows the fish sampling equipments; a – Sieve, b – Scoop net
In the laboratory, small fish (equal or less than 2cm) were put on a slide, total length measured using a mathematical pair of divider and ruler, and examined directly under a compound microscope for ectoparasites on the skin and fins. Then the fish were dissected carefully using a needle to expose internal organs for endoparasite investigations.
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Fish parasites in Uganda Akoll P.
However, any fish sample showing high ectoparasite infestations was fixed for histopathological investigations. Fish (larger than 2cm) were put in a Petri dish (containing pond water in which the fish was reared and carried to the laboratory) and killed by piercing into the brain before being examined under a dissecting microscope for ectoparasites. Similarly, prior the examination, total length is measured using the same technique as for smaller fish.
Skin scrapings from the fish were taken with a cover slip (22 x 22mm) from near the operculum to the caudal peduncle on the lateral side of the body length, dorsal part of the head, ventral region of the fish from head to just after the anal point and from the dorsal, the ventral and the caudal fins. Smears of the mucus taken from these various sites were mounted on glass slides, observed immediately unstained under light microscope. Parasites observed in these fresh smears were recorded, identified at least to genus level and counted. Intensities of infections were categorized into classes (Table 3.4) after enumeration of protozoa per field of view (area of 26cm2) or monogeneans from the whole fish.
Table 3.4: Categorisation of parasite intensities on fish Infection category Parasite intensity Low X ≤ 5 Moderate 5 < X ≥ 10 Heavy X ≥ 10
Slides with different intensities of parasites were fixed with either 70% alcohol (for helminths) or 4% formalin (for protozoa), air dried, fixed onto the slide with methanol, stained with eosin just for contrasting before mounting a coverslip using DPX and kept as a permanent slide for demonstration. These slides are deposited in the Department of Invertebrates, Natural History Museum, Vienna. Whole gill rakers were carefully cut out, put on a microscope slide and examined for ectoparasites, nodule forming protozoan parasites or cysts at X100 and/or X400 magnifications. In addition, mucus smears from the gills as well as blood smears were examined in wet mounts unstained under a compound microscope for presence of protozoa. Following the same intensity categorisation system above, gills with different infestation intensities were fixed in 70% alcohol or formalin fixatives for histological sections.
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The gastrointestinal tract was opened longitudinally and smears of faeces, and scrapings from the intestinal wall examined in unstained wet mounts under a microscope. Additionally, small pieces of well-developed, recognised fish organs (e.g. kidney and liver) were crushed under cover slip and examined thoroughly under compound microscope (magnification X100 and X400) in a wet unstained mount while some were fixed for histology. Photographs and/or video clips (especially for fast moving organisms) for positive slides were taken either at X100 or X400 or X1000 magnification for documentation.
To describe the distribution of parasites throughout hatcheries and to correlate the parasite infections to age classes, the parameters prevalence, mean intensity and mean abundance were used after Bush et al. (1997). These are parameters used in assessing the parasite infections within the host population. Prevalence describes presence or absence of parasites within a host population or sample. The parameter is convenient for the Protozoa, bacteria and viruses (microparasites) due to difficulty in enumeration but it is also used for helminths (macroparasites) (Anderson, 1993; Bush et al. 1997). Mean intensity and mean abundance provides a clue on the severity of the infections. Mean intensity concerns only infected part of the population (or sample) and mean abundance considers the whole population (or sample) – infected and noninfected. To measure mean intensity and mean abundance, either direct counting e.g. for ectoparasites especially for helminths or indirect techniques for microparasites e.g. protozoa are used. Therefore, to determine these parameters, specimens of Gyrodactylus sp. on at least the entire body and fins were enumerated. While mean numbers of protozoans counted from five fields of view (area = 26cm2) at a total magnification of X100 were used for estimation of the parameters. The mean number of protozoa was then expressed per cm2. However, in quantification of Protozoa, several methods were according to McGuigan and Sommerville (1985), Barker at al. (2002) and Khan (2004). Furthermore, plotting frequency distribution graphs also assesses dispersion patterns of macroparasites and hence parasites induced mortality. The distributions can be under–dispersion (regular, variance < mean), random (variance = mean) and over–dispersion (aggregated, variance > mean) (Anderson, 1993; Esch and Fernändez, 1993).
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The description of Trichodinids is base on silver nitrate impregnated air-dried smears, which aid in studying the adhesive disc and haematoxylin stained specimen for studying nuclear apparatus (Van As and Basson, 1989; Lom and Dykova, 1992; Lom, 1995). In addition, Lom (1995) has clearly stated the species described from fresh smears are worthless and misleading. However, during this study, due to resource limitation, measurement of the adhesive disc apparatus (Fig. 4.3.1b), fresh samples were used for purposes of testing for the difference in species occurrence between farms. All measurements were obtained using Leitz Wetzlar microscope equipped with X10 micrometer scale ocular. Following standard presentation of variables proposed by Lom (1958), Basson and Van As (1989) all measurements distinguishing characteristics are in micrometers. In each case minimum and maximum values are given, followed in parentheses by arithmetic mean and standard error. During measurement smears would dry out as a result of heat from the microscope lamp. This caused Trichodina sp. to lose body morphology, thus no body diameter measurements were made once the smear dried but denticle measurements continued. Similarly, no measurements were done on Gyrodactylus sp. from dried smear as it burst out completely. Consequently, measurements from fresh specimen were made as fast as possible. However, the identification of the parasites was done with the aid of determination keys provided by Bykhovskaya-Pavlovskaya et al. (1964); Khalil (1971); Lom and Dykova (1992); Woo (1995); Reed et al. (2002).
For histopathology, fixed samples and organs were trimmed, labelled carefully and put in tissue processing cassettes before being put in an automatic tissue processor. In the tissue processor samples were dehydrated in a series of ethanol: 70%, 85%, 95% and 100% (at two hours interval), cleared off alcohol with Xylene in two changes (at an interval of four hours) and impregnated with molten wax in two changes at an interval of six hours. The tissues were removed from the processor and embedded in 60°C paraffin wax. The embedded tissues were blocked on wooden blocks, trimmed and frozen (to harden the tissue) prior to sectioning. Using a Chadwell health (England) sectioning machine equipped with a R. Jung (Heidelberg, Germany) non disposable wedged microtome, sections of 5µm were cut, flushed with 70% alcohol on a slide, floated in lukewarm water bath to straighten the section before put on a slide again.
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Slides with samples were put in an oven overnight for adhesion before staining with haematoxylin and eosin, Masson´s Trichrome and Giemsa stains.
All numerical results were analysed by EXCEL (Microsoft) and SPSS 12.0 (Systat Software Inc) computer software packages applying Mann Whitney test (U test) and Kruskal Wallis test (H-test) for non-parametric data and T-test and One-way Analysis of Variances (ANOVA) for parametric data. Graphs were created using SigmaPlot 9.0 (Systat Software Inc).
Biological terms used
Fry – were the young fish with total length of 0.5cm to below 3.0cm. Fingerlings – were the young fish with total length of 3.0cm and above but less than 20.0cm The definitions of fry and fingerlings depend very much on the fish species. Pathology – study of diseases, causes, and progress of the resultant changes in the diseased organism. Histopathology – the study of changes due to a disease on cellular level. Mean intensity – is the number of parasites of a given taxonomic group per infected host in a given population or sample population. Mean abundance – is the number of parasites of a given taxonomic group divided by total number of individuals examined. Thus it is the number of parasites per the total individuals sampled. Prevalence – it is the proportion or percentage of fish infected with parasites or microorganisms to the total number of individuals examined. It is the number of infected individuals divided by total number of individuals examined multiplied by 100. Infection – refers to invasion of the host by foreign organisms e.g. Bacteria, Fungi and Viruses Infestation – is the presence of large number of parasites. However, the term infection is used to include infestation and its derivatives
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4 RESULTS
4.1 Water quality
Pooled data of water quality measurements from all farms indicated that water had a mean temperature of 24 ± 0.38°C (mode: 23.8, range: 21.8 – 28.9). Dissolved oxygen on the other hand was 6.6 ± 0.16mg/l (mode: 6.7, range: 4.2 – 8.5) and pH was 6.8 ± 0.11 (mode: 7, range: 5.5 – 7.8). Chemical parameters: nitrite and ammonium never exceeded 0.05 and 0.2mg/l respectively. Measurements from individual farms showed a different pattern particularly for physical parameters (Table 4.1a). In this respect, water temperature was significantly higher at Sunfish farm than other farms (ANOVA,
3.192,48 = 18.6, P < 0.05). Sunfish farm had a mean water temperature of 25°C with a median of 25.3°C compared to a mean water temperature of 23°C measured from both KRS and Ssebinyansi farm with medians of 23°C and 23.5°C respectively (Table 4.1a). However, dissolved oxygen and pH between farms where not significantly different at
the set significant level of 0.05 (ANOVA, 3.362,26 = 0.17, P = 0.84 and 3.192,26 = 1.16, P = 0.32 respectively).
Table 4.1a: Water quality measurements from three hatcheries in Uganda (Mean ± SE).
Hatcheries Sunfish farm KRS Ssebinyansi farm Parameter Range Range Range Mean Mean Mean Min Max Min Max Min Max Temp (°C) 25.5±0.51 21.8 28.9 23.0±0.28 22.0 24.1 23.2±0.31 22.0 23.7 Oxygen 6.7±0.26 4.2 8.5 6.7±0.14 6.2 7.2 6.4±0.24 5.7 7.0 (mg/l) pH 6.6±0.14 5.5 7.5 7±0.25 5.9 7.7 7±0.24 6.5 7.8
The pooled data of the measured dissolved oxygen concentration from all farms were ranging from 4.2mg/l to 8.5mg/l. Sunfish farm had a wider range of 4.3mg/l compared to KRS and Ssebinyansi farm, which had a range of 1mg/l and 1.5mg/l respectively.
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Similarly, pH followed the same trend with Sunfish having a range of 2 while KRS and Ssebinyansi farm had 1.8 and 1.3 respectively. Pooled water temperature data from all hatcheries had no significant correlation with dissolved oxygen values, n = 29, rs = -0.17, P = 0.389 (Fig 4.1a). Similar trends were observed from individual hatcheries.
9.00
8.00
7.00
Oxygen (mg/l) Dissolved 6.00
5.00
22.00 24.00 26.00 28.00 Temperature (°C)
Figure 4.1a: Correlation of Water physical parameters from Clarias gariepinus hatcheries in Uganda.
Water flow through the raising and/or nursing facilities at the farms varied tremendously. Sunfish farm had water flowing through the system daily with flow rates presented in table 4.1b. The rates however, varied widely both between tanks and days, that’s from 0.32 l/sec to 1.03 l/sec, and therefore, the retention time of water in tanks ranged from 1.6 hours to five hours. For calculation of retention time, volumes of water in tanks were divided by outflow rates. Calculations were based on the assumption that inflow rates were equal to the outflow rates, as shown in table 4.2b; therefore water level was more or less constant. Also the tanks holding fish were of the same size: 6m long, 2m wide and 0.5m deep. Measurements of oxygen concentrations showed that inflow concentration ranged from 6mg/l to 10mg/l and outflow ranged from 4mg/l to 9mg/l. The difference between oxygen concentration in the inflow and outflow ranged from 1mg/l to 3mg/l.
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In contrast KRS had intermittent water flow through the system depending upon the rate at which algal blooms are observed (water turned green). This could occur at most once or at least twice a week. Therefore, water inflows and outflows were not measured. Unlike Sunfish and KRS, Ssebinyansi farm had no water flowing through the system. The farm depended only on groundwater, which rises very slowly from the ground, hence had no large amount of over flow from the ponds.
Table 4.1b: Average Water inflow, outflow rates and retention time measured in different tanks at Sunfish farm, Uganda.
Number of ponds Inflow rate Outflow rate Retention time measured (l/sec) (l/sec) (hours) 1 0.37 0.37 4.5 2 0.72 0.72 2.3 3 0.80 0.80 2.1 4 0.80 0.80 2.1 5 0.32 0.32 5.2 6 1.03 1.03 1.6 7 0.85 0.84 2.0
4.2 Parasites
From a total of 334 individual fry and fingerings of C. gariepinus during the study, four Protozoa genera, one genus of Monogenea and Epitheliocystis were encountered from the hatcheries. The protozoans included Trichodina (Ehrenberg, 1838), Apiosoma (Blanchard, 1885), Epistylis (Ehrenberg, 1830), and Trichophrya (Claparede and Lachmann, 1858) while the monogenean was Gyrodactylus (Von Nordmann, 1832). Genera of Trichodina and Gyrodactylus were found on fish from all hatcheries. Apiosoma sp. was found 5 fish from KRS with a mean intensity of 1.8 individuals/fish and on three fish from Sunfish farm with a mean intensity of 2.7 individuals/fish. Epistylis sp. was found on one fish from KRS harbouring one colony of the parasite and one fish from Ssebinyansi harbouring 3 colonies. One fish specimen from Ssebinyansi farm was found harbouring three individuals of Trichophrya sp.
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Thus fish from KRS and Ssebinyansi harboured five genera of parasites while fish from Sunfish had four genera of parasites. Apart from Trichodina and Gyrodactylus all the other genera – Apiosoma, Epistylis, and Trichophrya – occurred in a very low intensity; a maximum mean intensity recorded was 3 colonies of Epistylis. All the Protozoa belong to phylum Ciliophora, while Gyrodactylus sp. on the other hand is a helminth belonging to the class Monogenea sub-class of Monopisthocotylea. These parasites preferred the skin and fins but Trichodina sp. and Gyrodactylus sp. could occur on gills under moderate and heavy infection. Epitheliocystis consisting of Rickettsia or Chlamydia–like inclusions belonging bacterial groups was found on gills of fish specimen from Sunfish farm. In the proceeding sub-sections, description of these parasites and their occurrence are presented.
4.2.1 Protozoa
Four genera of phylum Ciliophora in the Kingdom Protozoa were found on C. gariepinus fish during the parasite investigation.
Trichodina (Ehrenberg, 1838)
Among the ectoparasites, the protozoan Trichodina sp. (Figs 4.2.1a) did occur on C. gariepinus from all farms and it was the most dominant protozoan parasite. Pooled data showed that the number of parasitized fish was not significantly different from the non- parasitised (U-test P>0.05). Considering each age group independently, it was observed that one week old fish fry had no Trichodina sp. occurring on them. Whereas two weeks old fish had two individuals out of seventy individuals examined parasitized, which was significantly lower than the non-parasitized fish (U-test, P<0.05, n = 70). In contrast to older fish: three to ten weeks from all hatcheries, the number of parasitized fish was significantly higher than non-parasitized fish (U-test, P<0.05, n = 334, Table 4.2.1a). Scattered individuals of Trichodina sp. were observed moving all over the entire fish body but preferred the caudal fin, pectoral fin, pelvis fin, near the anal opening and caudal peduncle and occasionally occurred on the gills.
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Table 4.2.1a: Occurrence of Trichodina sp. on cultured C. gariepinus fry and fingerlings in Uganda (Mann Whitney (U) test, Level of significance: nd P>0.05, * P<0.05, ** P<0.01, *** P<0.001)
Age Parasitised Non- Total (n) Degrees of Significance parasitised freedom level 1 0 30 30 29 *** 2 2 68 70 69 *** 3 14 17 31 30 *** 4 42 18 60 59 *** 5 44 2 46 45 *** 6 20 0 20 19 *** 7 30 0 30 29 *** 8 11 1 12 11 *** 9 14 0 14 13 *** 10 15 6 21 20 ** Total 192 142 334 324 nd
The parasite moved with a sliding, circulating motion over the substrates with the use of cilia (C) that were clearly visible under a light microscope (Fig 4.2.1a). Morphologically, the parasite is characterized with proteinaceous skeleton of adhesive disc projections called denticles which consists of a semicircular outer projection: the Blade (B) and inner sharp pointed projection: the Ray (R). These denticles are interconnected to form a characteristic denticle ring visible in the ciliate from both fresh (Fig 4.2.1a & b) and stained smear, surrounded by striated border membrane. The parasite has distinct cilia observable both in fresh and stained specimen. When viewed laterally and occasionally when moving, Trichodina sp. was dome- shaped or bell- shaped (Fig 4.2.1a –ii) and it appeared as a flat disc when at rest especially on a flat surface (Fig 4.2.1a –i).
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B C
R
C
ii i
Figure 4.2.1a: Photomicrographs of Trichodina sp. from cultured C. gariepinus, Uganda in fresh smears. (i) X400, (ii) X200. Legend: B – Blades, R – Rays, C – Cilia
Various size measurements on the adhesive disc for denticle length, denticle ring diameter, ray lengths, blade lengths and number of denticles together with body diameter for species comparison, are shown in table 4.2.1b. It must be noted that all measurements are in micrometers (µm), unless stated. Based on all the measurements there was no significant difference (P > 0.05) between individuals of Trichodina sp. observed on fish from the three hatcheries sampled in Uganda. This clearly suggests that it was one species of genus Trichodina parasitizing C. gariepinus fish.
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Table 4.2.1b: Dimensions of Trichodina sp. on fry and fingerlings of C. gariepinus from Uganda Regions measured Mean ± SE Mode Min – max Number of specimen (n) Body diameter 64 ± 0.80 62.5 50 – 85 87 Denticle ring diameter 44 ± 0.43 42.5 30 – 55 116 Central circle 15 ± 0.26 15 7.5 – 27.5 116 Denticle length 14 ± 0.15 15 10 – 20 116 Blade length 5 ± 0.06 5 2.5 – 7.5 104 Ray length 7 ± 0.11 7.5 3.75 – 10 104 Blade-Ray division 2.5 ± 0.04 2.5 2.5 – 3.75 28 Number of denticles 24 ± 0.14 24 22 – 28 77
ST
B
R
CP
DD
BD
Figure 4.2.1b: Photomicrograph of fresh Trichodina sp. indicating measured points in fresh smears (magnification X400).
Legend: BL – Blade length, RL – Ray length, DL – Denticle length, DD – Denticle ring diameter, CP – Central circle, ST – Striated membrane, BD – Body diameter.
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Apiosoma (Blanchard, 1885)
Apiosoma synonymous to Glosatella (Buetschii, 1889) (Fig 4.2.1c) was found occurring of eight fish individuals of five weeks old from KRS and Sunfish farm. At KRS, Apiosoma sp. appeared on five of the fish kept in earthen pond at a mean intensity of 1.8 individuals/fish and a mean intensity of 2.7 individuals/fish on three fish from nursing tanks at Sunfish farm. However, no measurements were made because of their fast retractor movements. This protozoan was found occurring single, on outer surface of the operculum and laterally near the anal point of the fish. The parasite was sessile, bell-shaped but more or less cylindrical body towards the stalk and possessed a distinct adoral row of cilia (easily visible in fresh specimen). The stalk is relatively long and originates from a wide base (scopula).
Figure 4.2.1c: Apiosoma sp. photomicrograph from C. gariepinus fingerlings from Uganda in fresh smears (magnification X400)
Epistylis (Ehrenberg, 1830)
Epistylis sp. (Fig 4.2.1d) was found on the skin of ten week old fingerling kept in earthen pond at Ssebinyansi farm and of seven week old fish from KRS. This species measured (13.5 ± 1.27 x 8.8 ± 0.79µm, n = 5). The organism was observed swimming with the aid of cilia with scopula being directed forward (Fig 4.2.1d - i).
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This ciliate had cilia at the adoral end, clearly visible vacuoles and the body was bell- shaped but with relatively short conical stalk originating from a thin scopula. This genus usually occurs in colonies (Fig 4.2.1d – ii) but may occur solitary (Fig 4.2.1d – i)
i ii
Figure 4.2.1d: Epistylis sp. photomicrographs from C. gariepinus fingerlings from Uganda in fresh smear (i) Solitary swimming (magnification X400) (ii) colonial (magnification X100)
Trichophrya (Claparede and Lachmann, 1858)
Trichophrya sp. synonymous to Capriniana (Mazzarelli, 1906) (Fig 4.2.1e) was found on one fish during the whole sampling period. Three individuals were found the four week old fish from earthen pond at Ssebinyansi farm. This parasite was sedentary and occurred solitary near the opercula opening. It possessed tentacles at the adoral end and appeared bell-shaped with a very long stalk. The scopula was however not clearly visible during the study.
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Figure 4.2.1e: Trichophrya sp. photomicrograph from C. gariepinus fingerlings from Uganda in fresh smears (magnification X400)
4.2.2 Monogenea
Gyrodactylus (Von Nordmann, 1832)
The only metazoan parasite found parasitizing C. gariepinus fish during the study was Gyrodactylus sp. Incase of this parasite, pooled data showed that the number of infected fry and fingerlings were significantly smaller than the non-parasitized fish investigated (U-test, P<0.001). It was observed during the study that Gyrodactylus sp. did not occur on fish less than three weeks old (Table 4.2.2). However, from four weeks old fry to ten weeks fingerlings, the occurrence of Gyrodactylus sp. became significantly abundant on fish. Under low and moderate infestation levels, Gyrodactylus sp. preferred pectoral, anal, ventral and caudal fins but under moderate and heavy infestations, the parasite spread to gills, barbels and the head. The helminth had opisthaptor armed with marginal hooks (numbering 12 – 16) surrounding two strong median hooks (Fig 4.2.2). The parasites measured 0.7 ± 0.02 mm in size, which ranged from 0.60 mm to 0.72 mm (n = 6). Most individuals where observed to posses embryos easily discernable by the presence of median hooks in the abdomen. The anterior end was bifurcated with no eyespots.
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i
Figure 4.2.2 – i: Fresh specimen of Gyrodactylus sp. found on C. gariepinus fry and fingerlings from Uganda (magnification 100X)
G
G
G G
ii
Figure 4.2.2 – ii: Histological specimen of Gyrodactylus sp. (G) found on gills magnification of C. gariepinus from Uganda (magnification X400)
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Table 4.2.2: Occurrence of Gyrodactylus sp. in cultured C. gariepinus fry and fingerlings in Uganda (Mann Whitney (U) test, Level of significance: nd = P>0.05, * P<0.05, ** P<0.01, *** P<0.001)
Age Parasitised Non-parasitised Total (n) Degree of freedom Significance level 1 0 30 30 29 *** 2 0 70 70 69 *** 3 0 31 31 30 *** 4 10 50 60 59 *** 5 25 21 46 45 *** 6 16 4 20 19 *** 7 30 0 30 29 *** 8 10 2 12 11 *** 9 11 3 14 13 *** 10 14 7 21 20 ** Total 116 218 334 333 ***
4.2.3 Epitheliocystis
Epitheliocystis – cyst-like proliferated cells were observed on the gills of C. gariepinus from hatcheries in Uganda. This was the sole representative of internal parasite encountered during the study. Out of 56 fish specimen used for histopathology, 8 fish (12.5 % prevalence) had Epitheliocystis. On average, 2 cysts with a size range of 20µm – 70µm in diameter were observed at the apex of gill filament, at the apex of secondary gill lamellae and at the base of secondary gill lamellae (Fig 4.2.3). Histopathological examinations under a light microscope, Epitheliocystis swellings appeared as well defined spherical basophilic globules filled entirely with more or less homogenous mass of inclusions. Host tissues showed insignificant pathological changes except for the infected cells, which were hypertrophic.
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i ii
iv iii
Figure 4.2.3: Epitheliocystis on the gills of Clarias gariepinus from hatcheries in Uganda, Site of occurrence (i) at the apex of the gill filament (ii) at the apex of the secondary lamellae, (iii) at the base of the secondary gill lamellae (H&E; magnification X400)
4.3 Occurrence of parasites
In this section, prevalence, mean intensities and mean abundances of dominant parasites – Trichodina sp. and Gyrodactylus sp. – of C. gariepinus, in correlation to age and at different hatcheries are presented. First, overall distribution of the pooled data these parasites from hatcheries in Uganda and then individual hatcheries will be analysed.
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Overall occurrence of parasites from hatcheries in Uganda
The results reveal vividly that Trichodina sp. and Gyrodactylus sp. increased in prevalence drastically with increase in size and age of fish (Fig 4.3.1). Trichodina sp. however, appeared as early as two weeks after hatching and increased drastically to 100% after six weeks post-hatching or four weeks post infection. Gyrodactylus sp. in contrast, appeared at the fourth week post-hatching whose prevalence also increased tremendously to 100% after four weeks post infection. It is also observed that seven weeks old fry had both Trichodina sp. and Gyrodactylus sp. was occurring on the entire specimen collected. The distinct fluctuations at the age of eight and ten weeks were due to preventative measures taken after massive mortalities were experienced. However, after treatment, Trichodina sp. quickly reinfected the entire host population again while Gyrodactylus sp. never reinfected to all the hosts during the study.
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100 (a) 90 80 70 60 50 40
Prevalence (%) 30 20 10 0 12345678910 15 (b)
10
Mean intensity 5
0 12345678910 15 (c)
10
5 Mean abundance Mean 0 12345678910 A ge (w eeks) Trichodina sp. Gyrodactylus sp.
Figure 4.3.1: Occurrence of Trichodina sp. and Gyrodactylus sp. on C. gariepinus from fish farms in Uganda (n = 334, bar = SE)
Mean intensities and mean abundances of Trichodina sp. and Gyrodactylus sp. followed the same pattern as the prevalence (Fig 4.3.1). Both mean intensity and mean abundance of parasites increased with age of the fish. Trichodina sp. mean intensities and mean abundances increased more or less exponentially reaching the highest mean values of 13 individuals/cm2/fish on six weeks old fish.
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Similarly, the mean intensity of Gyrodactylus sp. increased drastically from mean of 2 individuals/fish on four and five weeks old fish to mean intensity and mean abundance of 9 individuals/fish on six weeks old fish and reached a maximum mean value of 15 individuals/fish on nine weeks old fish. Mean abundance of Gyrodactylus sp. showed the same trend as mean intensity but six weeks fish for example had low mean abundance than seven despite having the same mean intensity. It should be noted that mean intensities values are equal to mean abundances of parasites when the prevalence equals 100 % but mean intensity is higher than mean abundance when prevalence is less than 100 %.
It was observed in week seven, nine and ten after establishment that at high mean intensities of Gyrodactylus sp., the mean intensities and mean abundances of Trichodina sp. decreased (Fig 4.3.1). On application of formalin as parasite control measure, at seven and nine weeks, the prevalence, mean intensity and mean abundance of both Trichodina sp. and Gyrodactylus sp. reduced. After the treatment, the mean intensity and mean abundance of Gyrodactylus sp. quickly increased but its prevalence never reached 100 %. In contrast, mean intensity and mean abundance of Trichodina sp. remained more or less constant but redistributed to the whole host population hence attained 100 % prevalence.
Comparison of occurrence of parasites from individual hatcheries
There was no significant difference in the prevalence of Trichodina sp. and
Gyrodactylus sp. (ANOVA, 3.522,19 = 0.36, P = 0.70 and 3.522,19 = 0.72, P = 0.50 respectively) on the fish from Sunfish, KRS and Ssebinyansi farms. Prevalence of Trichodina sp. and Gyrodactylus sp. parasites increased drastically from 0% to 100%, a trend exhibited in all farms. However, bath of fish in formalin as parasite control measure at week seven and nine caused fluctuations in prevalence of parasites at Sunfish farm (Fig 4.3.2). On the other hand, parasites prevalence remained at 100 % throughout the study at both KRS and Ssebinyansi farm. After infestation, Trichodina sp. and Gyrodactylus sp. took three and four weeks respectively to spread to all the fish. Due to the delays in commencement hatching at Ssebinyansi farm, sampling ended at five weeks old fish, but there is a likelihood of proceeding age groups having similar trends in mean intensity, mean abundance and prevalence like other farms.
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Combined mean intensities of both Trichodina sp. and Gyrodactylus sp. occurring on fish from Sunfish and KRS were not significantly different from each other but were significantly higher on fish samples from Ssebinyansi farm (H-test, n =3, P < 0.001, X2 = 29.7, df =2). On the other hand, the combined mean abundances of Trichodina sp. and Gyrodactylus sp. on fish from KRS and Ssebinyansi farm were not significantly different from each other but were significantly lower than from Sunfish farm (H-test, n =3, P < 0.001, X2 = 30.2, df =2). Likewise, mean intensities of Trichodina sp. on fish from Ssebinyansi farm were significantly lower than from Sunfish farm and KRS (H- test n = 3, P<0.029, X2 = 7,df = 2). There was no significant difference of Trichodina sp. mean intensities on fish from Sunfish and KRS (H-test, P>0.05). Mean abundance of Trichodina sp. on fish from KRS was significantly higher compared to Sunfish and Ssebinyansi farm (H-test n = 3, P < 0.001, X2 = 27.5, df = 2). Also, the mean intensities of Gyrodactylus sp. occurring on fish from Sunfish farm were significantly higher than from KRS and Ssebinyansi farm (H-test n = 3 P < 0.001, X2 = 28, df = 2), but no difference between KRS and Ssebinyansi farm were observed. The mean abundances of Gyrodactylus sp. were significantly higher compared to KRS and Ssebinyansi farm (H- test n = 3, P < 0.001, X2 = 27, df = 2) and no significant difference was observed between KRS and Ssebinyansi farm.
Occurrence of parasites from individual hatcheries
In this section, a closer look at the occurrence of the dominant parasites in each hatchery is presented. Furthermore the dispersion of Gyrodactylus sp. within the host population is shown by frequency distributions.
Sunfish Farm
Prevalence, mean intensities and mean abundances of parasites on fish from Sunfish followed the same trend as the combined trend of all hatcheries studied. Therefore, emphasis will be on comparing the occurrence of parasites from different age classes (Fig. 4.3.2). The mean intensity of Trichodina sp. on fish of week one, two and three old were significantly lower than on fish of four – ten weeks (H-test P < 0.05) but were not significantly different from each other.
MSc. Thesis 47 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Four week old fish harboured Trichodina sp. with intensities higher than of one, two, three, and five weeks old but lower than of six weeks old and not different from seven, eight, and nine weeks old fish (H-test, P < 0.05). The mean intensity of Trichodina sp. on fish of five weeks was not significantly different from that of four and ten weeks but higher than of one, two and three weeks old fish and lower than six, seven, eight and nine weeks old fish. Six weeks old fish harboured the highest intensity of Trichodina sp., which was significantly different other age groups (H-test, P < 0.001). From seven weeks old fish till ten weeks, no significant difference in Trichodina sp. intensity was observed. The results therefore, indicate that the intensity of Trichodina sp. on fish from Sunfish farm increased up to a maximum mean of 15 ± 2 individauls/cm2/fish at 6 weeks, four weeks after appearance and exposure to surface water and then declined to more or less constant intensity.
Likewise, mean intensity of Gyrodactylus sp. on fish of one, two, three, four and five weeks old were significantly lower that from fish of six, seven, eight, nine and ten weeks old (H-test, P < 0.001). The age classes six, seven and nine weeks harboured a very high intensity of Gyrodactylus sp. compared to other age classes (H-test, P < 0.001). Like Trichodina sp., Gyrodactylus sp. attained the highest mean intensity of 29 ± 7 individuals per fish at six weeks post hatching. This however, gradually decreased to 23 ± 2 individuals/fish at seven weeks and drastically on addition of 40 ppm of formalin at seven and nine weeks. It is worth noting that the same trend of intensity and abundance of Trichodina sp. and Gyrodactylus sp. were observed from Sunfish farm (Fig. 4.3.2).
MSc. Thesis 48 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
100 (a)
80
60
40
Prevalence (%) 20
0 12345678910
30 (b)
20 Mean intensity intensity Mean 10
0 12345678910
30 (c)
20
10 Mean abundance abundance Mean
0 12345678910 Trichodina sp. Age (weeks) Gyrodactylus sp.
Figure 4.3.2: Occurrence of Trichodina sp. and Gyrodactylus sp. on C. gariepinus from Sunfish farm (n = 162, bar = SE)
Frequency distribution of Gyrodactylus sp. from Sunfish
Variance – mean ratios plotted (Fig. 4.3.3a) were above unity indicating that the distribution of the monogenean Gyrodactylus sp. was over-dispersed, viz; few fish had many parasites while many fish had no or very few parasites. Frequency distributions
MSc. Thesis 49 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P. for each age class (Fig. 4.3.3b) support the observation. Again, it’s evident that administration of formalin reduced the distribution towards unity.The effect was more pronounced at week eight where Gyrodactylus sp. was tending to a uniform distributed on its hosts (Fig. 4.3.3b).
120
100
80
60
40 Variance / Mean Ratio
20
++ ++ ++ 0 12345678910 Age (weeks)
Figure 4.3.3a: Variance – mean ratio of Gyrodactylus sp. on fish by age from Sunfish farm (n = 162, bar = SE, ++ = no parasite recorded).
MSc. Thesis 50 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
12 12 Week 4 10 Week 5 10 8 8 6 6 4 4 2 Number of hosts of Number 2 0 Number of hosts 0 0 1020304050 0102030 Number of Gyrodactylus sp. Number of Gyrodactylus sp. 12 12 10 Week 6 10 Week 7 8 8 6 6 4 4 Number of hosts Number 2 Number of hosts 2 0 0 0 50 100 150 200 250 300 350 0 100 200 300 400 500 Number of Gyrodactylus sp. Number of Gyrodactylus sp.
12 12 10 10 Week 9 Week 8 8 8 6 6 4 4
2 Number of hosts 2 Number ofNumber hosts 0 0 0 10203040506070 0 50 100 150 200 250 300 350 Number of Gyrodactylus sp. Number of Gyrodactylus sp.
12 10 Week 10 8 6 4
Number of hosts of Number 2 0 0 5 10 15 20 25 30 35 Number of Gyrodactylus sp.
Figure 4.3.3b: Frequency distribution of Gyrodactylus sp. on fish by age (4 – 10 weeks) from Sunfish farm (n = 162, bar = SE)
MSc. Thesis 51 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Kajjansi Research Station (KRS)
Results from KRS (Fig. 4.3.4) show that mean intensities and abundances of Trichodina sp. on one, two, three and four weeks old fish were not significantly different from each other but were significantly lower than intensities from five, six and seven weeks old fish (H-test, P < 0.05). Similarly, five, six and seven age groups carried Trichodina sp. of mean intensities and abundances not significantly different from each other. Like Sunfish, mean intensities and abundances of Trichodina sp. from KRS increased with increase in fish age and reached a maximum of 27± 1 individuals/cm2/fish at six weeks –exactly four weeks post appearance and exposure to surface water. The mean intensity however, declined in the proceeding age class seven. The decrease coincided well with a significant number of Gyrodactylus sp. Thus the presence of Gyrodactylus sp. is suspected to be the cause of decline in Trichodina sp. probably due to competition for space.
MSc. Thesis 52 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
100 (a) 80
60
40
Prevalence (%) 20
0 1234567 30 (b)
20 Mean intensityMean 10
0 1234567 30 (c) 20
10 Mean abundance abundance Mean 0 1234567 Trichodina sp. A ge (w eeks) G yrodactylus sp.
Figure 4.3.4: Occurrence of Trichodina sp. and Gyrodactylus sp. on C. gariepinus from KRS (n = 113, bar = SE) Gyrodactylus sp. in this farm occurred on fish of six and seven weeks only with mean intensity from seven week old fish being significantly higher than from six weeks (P < 0.001.
Frequency distribution of Gyrodactylus sp. from KRS
As indicated earlier, Gyrodactylus sp. was observed only on fish from six and seven weeks old fish. The parasites on both age groups six and seven showed over-dispersed distribution (Fig. 4.3.5). The classes with had variance to mean ratios of 9.1 and 19.7 respectively.
MSc. Thesis 53 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
When compared with fish of the same age from Sunfish farm, clearly shows that few fish in the later farm harboured many parasites than the former. The observation prompts a suggestion that these few fish at sunfish farm died of high burden of parasites.
7 12 6 12 10 10 8 8 6 6 4 4
Number of hosts Number 2 Number of hostsNumber 2 0 0 0 5 10 15 20 0 50 100 150 200 250 300 350 Number of Gyrodactylus sp. Number of Gyrodactylus sp.
Figure 4.3.5: Frequency distribution of Gyrodactylus sp. on fish by age from KRS (n = 113, bar = SE)
Ssebinyansi Farm
The mean intensities of Trichodina sp. on five week old fish were significantly higher than on one, two and four weeks old fish. Like other Sunfish farm and KRS, intensities and abundances increased with increase in age. Although, no fish samples were collected between six and nine weeks, probably the same pattern of mean intensities and abundances occurs. It was also observed for ten weeks old fish (belonging to the already existing cohort) that at higher intensities of Gyrodactylus sp., Trichodina sp. shows low intensity (Fig. 4.3.6)
MSc. Thesis 54 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
100
80 (a)
60
Prevalence (%) 40
20 0 ** ** ** 12345678910 20
15 (b)
10 Mean intensity Mean
5
0 ** ** ** 12345678910 20
15 (c)
10
5 Mean abundance abundance Mean 0 **** ** 12345678910 Trichodina sp. Age (weeks) Gyrodactylus sp.
Figure 4.3.6: Occurrence of Trichodina sp. and Gyrodactylus sp. on C. gariepinus from Ssebinyansi farm (n = 59, bar – SE and ** – no sample taken)
Frequency distribution of Gyrodactylus sp. from Ssebinyansi Farm
Gyrodactylus sp. was observed on fish of five and ten weeks. The distribution of this parasite on the groups showed over-dispersion (Fig. 4.3.7). The fish from this farm however had relatively lower variance/mean ration compared to Sunfish. Six weeks old fish had a ratio of 8.7 against 113 from Sunfish and ten week old fish had 6.5 compared to 30 from Sunfish farm. This may explain why there was not or insignificant mortality at Ssebinyansi farm to be attributed to parasites.
MSc. Thesis 55 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
12 5 12 10 10 10 8 8 6 6 4 4
Number of hosts 2 Number ofNumber hosts 2 0 0 0 5 10 15 20 25 60 80 100 120 140 160 Number of Gyrodactylus sp. Number of Gyrodactylus sp.
Figure 4.3.7: Frequency distribution of Gyrodactylus sp. on fish by age from Ssebinyansi farm (n = 59, bar – SE)
4.4 Clinical signs and pathology
Parasite infested fish were lethargic i.e. swimming sluggishly to water surface, concentrated at the sides of the pond and laid vertically in the water column. However, they would move rapidly for a very short distance when disturbed. There was decolouration of the skin where some fish became more yellowish while others became paler. Gills of high moribund fish were paler with localised haemorrhage at sites of attachment of Gyrodactylus sp. and Trichodina sp. Skin lesions were also observed more often particularly around preferred sites of parasites like caudal, ventral and dorsal fins. In many cases, skin lesions were overgrown with fungus, which confounded the problem and could have probably caused loss of the caudal fin observed in five fish specimen. Amputated barbels were also observed during heavy Gyrodactylus sp. infestations. On rare occasions, yellowing of the intestinal fluids (jaundice) was observed.
4.5 Histopathology
Following standard routine methods of histology, slides were made from different sites of low, moderate and heavily infested fish from all age groups. The sites include gills, caudal fins, ventral region near the anal opening, lateral sites, dorsal region, longitudinal sections of the internal organs and both dorsal and ventral regions of the head.
MSc. Thesis 56 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
To understand and appreciate abnormal changes, a brief description of basic structure and composition of the normal tissues, organs or systems where pathological changes were observed is provided.
Gills
These are the main respiratory organs in fish and other aquatic organism and most delicate part of the fish. It is however, a preferred site for living by several organisms, both parasitic and non-parasitic because of good supply of nutrients and relative protection from the external environment (Roberts 1978). Typically, gills have four gill arches each supporting gill filaments, which consists of secondary (S) and primary (P) lamellae both collectively called gill lamellae. The basic functional unit is the secondary lamellae where gaseous exchange and osmoregulation take place. Histological examinations of the normal gill through cross -sectioning (Fig. 4.6a) showed one single layer of cells (simple squamous epithelium) surrounding blood capillaries.
S
P
Figure 4.5a: Normal gill lamellae of five-week-old fish (H&E, magnification 400X)
During the study, histological examinations of parasitized fish revealed various pathological changes in gill morphology. First, there was accumulation of fluids in the intercellular spaces of the epithelial and pillar cells a condition known as oedema. The condition is as a result of host response to injury causing an increase the capillary permeability subsequently increasing infiltration of body fluids from the blood capillaries into the intercellular spaces but poor reabsorption (Robert, 1978).
MSc. Thesis 57 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
The accumulation of these fluids in the intercellular spaces consequently caused the enlargement of the secondary gill lamellae, a condition known as gill oedema (Fig. 4.5b). The condition was observed on all gill samples of fish harbouring moderate to heavy intensities of parasites. The severity of the condition increased with parasite intensity. The solid arrow (Fig. 4.5b) shows empty spaces, which were filled with body fluids and open arrow showing associated Trichodina sp.
Figure 4.5b: Gill lamellae oedema from five-week-old fish (Masson’s Trichrome, magnification 400X). Legend: Solid arrow shows fluid accumulation points and open arrow shows Trichodina sp. parasite
Secondly there was also proliferation of the gill epithelial cells leading to increase in size of the gill lamellae, known as hyperplasia. The sections (Fig. 4.5c) revealed extensive gill hyperplasia resulting in the fusion of the gill lamellae indicated with open-ended arrows. This condition was observed frequently in fish with heavy infestation of Gyrodactylus sp. and Trichodina sp.
MSc. Thesis 58 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Figure 4.5c: Hyperplastic gill lamellae from five-week-old fish (H&E, magnification 400X); open- ended arrow shows fused gill lamellae and closed-ended arrow shows Trichodina sp.
In addition, capsule-like cells with epically located nucleus called rodlet cells where observed in the epithelium of the gills. These cells where observed more frequently and in high intensities within heavily infested fish. The cells where however associated more with oedematous gills and early stages of gill hyperplasia (Fig. 4.5d).
Figure 4.5d: Rodlet cells (arrow) present in gills (magnification 200X)
MSc. Thesis 59 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Skin
Typical skin of clariids consists of epidermis and dermis layers (Grizzle and Rogers, 1976). The epidermis is composed of epithelial cells, mucus secreting cells – mucus cells (goblet cells) and alarm substance secreting cells – club cells (Fig. 4.5e). These cells are present in all regions of the fish though with different intensities. Furthermore, the relative intensities of occurrence of goblet cells and club cells in an individual fish and age are different. However, goblet cells in normal skin of Clarias sp. are present in lower intensities compared to club cells (Grizzle and Rogers, 1976). Goblet cells are also more superficial but the club cells never extend to the surface, thus their substances are only released when injured. Under pathological conditions, either the goblet cells increase in number or reduce depending on the type of infection, however, no reports on the changes in the intensities of club cells. Melanomacrophage centers (MMCs) – the main phagocytes in fish – on the other hand always occur in the least numbers (Agius and Roberts, 2003). They are reported to increase in intensity and size when the fish is exposed to stressful conditions and/or presence of infectious organisms. The dermis on the other hand located below epidermis is composed of connective tissues and adipose tissue filled with fat granules (Fig. 4.5e). Under pathological conditions, dermis usually undergoes necrosis and tissue hyperplasia.
MSc. Thesis 60 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
CT C
A
C
M
Figure 4.5e: Normal structure of C. gariepinus skin (Masson’s Trichrome stain, 400X magnification and M = mytomes, CT = connective tissue, C = club cells, A = adipose tissue, Arrow point to goblet cells, circles surround = melanocytes)
Sagittal and longitudinal sections through the skin and whole fish show minor tissue changes of both epidermis and dermis layers. There was no significant difference in the intensity of club cells in skin sections from the abdominal, dorsal and ventral regions in the low, moderate and heavy infested fish (ANOVA, 0.192,23 = 0.09, P = 0.99).
However, there was significant increase of club cells in the tail (ANOVA, 3.522,19 = 26.6, P < 0.05). Heavily infested fish had an average of 90 ± 3.8 (67 – 109) club cells compared to 52 ± 4.0 (48 – 56) in low and 54 ± 2.1 (46 – 61) in moderately infested fish. After application of formalin, there was outstanding reappearance of goblet cells which were very few or even absent in age classes before the treatment (Fig. 4.5f). In addition, there was enormous development and infiltration of melanomacrophage cells into the epithelium (Fig. 4.5g).
MSc. Thesis 61 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Figure 4.5f: Reappearance of goblet cells (circled) after formalin treatment (H & E 200X magnification)
Figure 4.5g: Caudal fin with a heavy infiltration of the Melanomacrophage cells (H&E, 200X magnification).
MSc. Thesis 62 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Figure 4.5h: Attachment of Gyrodactylus sp. on the fish skin (H&E, 400X magnification).
It was observed (Fig. 4.5h) that the attachment of Gyrodactylus sp. was more superficial. However, there is massive sloughing of the epithelium and general lack of adipose tissue except at the abdominal cavity. Similarly, attachment of Trichodina sp. reveals no significant effect on the skin (Fig. 4.5i). There was relatively small increase in the sizes (mild hypertrophy) of the epidermal cells observed at the points on attachment on Trichodina sp.
Figure 4.5i: Attachment of Trichodina sp. on the fish skin indicated by arrows (H&E, 400X magnification).
MSc. Thesis 63 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
5 DISCUSSION
Water quality
The physical and chemical parameters of water measured during the study were seemingly not contributing significantly to the fish mortalities observed except for occasional low dissolved oxygen. The relatively high water inflow and out flow rates for the intensive system at Sunfish farm – a farm which experienced the highest fish mortalities in nursing and raising tanks during the study – clearly shows that tanks had a very short water retention time. Thus the high flow rates could not allow the accumulation of ammonium, nitrite and changes in pH, which are attributed to fish kills (Schäperclaus, 1979; Durborow et al., 1997;) in water. Further, pooled data also show that water temperature had no correlation with dissolved oxygen values. This indicated that despite the maintenance of more or less constant temperature of ca. 24 °C in nursing and raising tanks, water temperature had no significant influence on dissolution of oxygen. The difference between the inflow and out flow oxygen concentrations, which ranged from 1 mg/l to 3 mg/l was attributed to microbial activity within the tanks rather than temperature. Although catfish can tolerate low dissolved oxygen concentrations – up to 1 mg/l, fry and fingerlings have lower tolerance abilities (Axis StorPoint CD, 2001). The low oxygen concentration measures on few occasions and a big difference between in and out flow oxygen concentrations may have had detrimental effect on the fish. These may have stressed and predisposed fish to succumb to parasite infestations. Notably, gill deformities shown by histopathological sections clearly indicated that fish had had their respiration compromised, thus, low oxygen concentrations could have been extremely detrimental. Although previous studies found that poor water quality parameters cause fish mortalities under culture conditions especially due to ammonia accumulation (Schäperclaus, 1979; Noble and Summerfelt, 1996; Durborow et al., 1997; Hargreaves and Tucker, 2004), ammonia never accumulated to devastating levels in this study.
MSc. Thesis 64 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
It is worth noting that other farms – KRS and Ssebinyansi farm, which had intermittent water flow through the pond, also had no accumulations of toxic nitrogen components as well as reduction in oxygen.
It is useful to recall that all water parameters according to the sampling design were measured at the same time as parasitic samples were taken. This thus never allowed for the follow up of the diurnal fluctuations of water parameters especially dissolved oxygen, ammonia accumulation and pH changes. Hargreaves and Tucker (2004) observed that several parasitological researches achieve constant concentrations of ammonia, which does not reflect diurnal variations and attributed this to deficiencies in methodology. Like, Sunfish farm receives water from a stream but is stored in a reservoir prior to supplying the ponds. In the reservoir, diurnal variations in water parameters are expected which were however not established based on the applied sampling design during the study. Therefore, a detailed study should be conducted with emphasis on water quality measurements and the relationship with the fish mortality and parasite infections.
Although water quality parameters measured appeared good and hence assuming good general maintain and hygiene of the farms, it was observed that parasites gained entry to the farm through the incoming surface water – for farms that relied on surface water for fish farming – and probably attachment on semi aquatic organisms such as amphibians, reptiles, which move from one water pool overland to another – for farms that used groundwater for fish farm. Incase of Sunfish farm and KRS (Fig. 4.3.2 & 4.3.4) the immediate appearance of parasites after change of water source from underground water to stream water supports the theory that source of parasites is the incoming water. At Sunfish farm, Trichodina sp. appeared in two weeks when the fish nursing ponds were supplied with a mixture of surface and groundwater. Similarly at KRS, Trichodina sp. appeared in three weeks immediately the fish were transferred to nursing tanks supplied with surface water. In fact at Sunfish farm for example, the distance between the water reservoir to fish tanks and ponds was less than 100m. Therefore, after dislodge from wild fish in the reservoir Trichodina sp. and Gyrodactylus sp. were to travel less than 100m to reach the cultured fish in ponds.
MSc. Thesis 65 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Previous studies have shown that Trichodina sp. can survive for about 16 hours off their suitable host (Madsen et al., 2000), which was long enough time to travel 100m, reach the pond alive and find new hosts. Further, the speed of water measured at the inlet (which may be more or less equal to the speed of water in the pipes from the reservoir) of up to 1l/sec could be high enough to transport these parasites quickly into the ponds.
However, no information was found regarding the time Gyrodactylus sp. would remain viable between dislodging from the original host and infecting a new one. It is most likely that Gyrodactylus sp. may survive long without a suitable host than Trichodina sp. due to the higher position in the evolutionary tree of the former. Several studies have also shown that most parasites into hatcheries and pond have their origin in wild fish and gain entry through water supply. Valtonen and Koskivaara (1994) observed that indoor kept and artificially fed Salmon fingerlings became infested with parasites from water source supplying the farm. Khan (2004) also suspected T. murmanica present on cultured cod originated from seawater supplying the farm. Khan (2004) added that although water was filtered to remove debris, the system was incapable of removing ciliates.
Low mean intensity and abundance of especially Gyrodactylus sp. at KRS compared to Sunfish farm supplied with water from the same stream could due to the low stocking density at KRS. Probably the reservoir at Sunfish farm could have acted as a breeding and population build up grounds for parasites. After establishment in the reservoir – whose ecology may not be significantly different from that of the ponds compared to stream ecology – the parasites may have had their population exploded in ponds. This may have been favoured also with high stocking density.
Ssebinyansi farm that ultimately depended on groundwater for fish farming had parasites appear much later. In this case, parasites may have gained entry into ponds through attachment on semi aquatic organisms like toads that migrated from the drainage channel. It is noteworthy that nursing ponds were not directly connected a drainage channel flowing into a wetland harbouring different types of aquatic organisms including wild fish. However, semi aquatic organism can migrate to the ponds from drainage channel.
MSc. Thesis 66 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Given the low host specificity particularly with Trichodina sp. (Van As and Basson, 1992; Lom and Dykova, 1992), the parasites may attach themselves on the semi aquatic organisms – toads. Due to free migration from one water pool to the other, semi aquatic organisms may carry these parasites and transmit them to fish. Supportive work from previous studies on the transmission of Gyrodactylus sp. via attachment on aquatic organisms was not obtained. Nevertheless, there is a likelihood of the parasite being introduced into the ponds by semi aquatic organisms. In this respect, more research is needed to establish the importance of semi aquatic organism in Gyrodactylus sp. transmission into fishponds.
The later appearance and the low intensities of Trichodina sp. and Gyrodactylus sp. observed at the farm could be attributed to low probability for those parasites to attach, successfully be transferred to another water body and find a susceptible host for establishment. All these may highlight the need for agent research to establish the source of parasites in farms depending ultimately on groundwater. Upon their findings, appropriate control and preventative measures can be set.
Thus, water quality parameters appeared to be in tolerable range for catfish but water supply was a major route for parasites into the hatcheries. Also due to already impaired gills, low dissolved oxygen measured occasionally probably contributed to stress and hence increased mortality of fish.
Parasites: Occurrence and pathology
As indicated in the results, six parasites species from six genera [Trichodina, Apiosoma, Epistylis, Trichophrya (Protozoa), Gyrodactylus (Monogenea) and Epitheliocystis] where observed on skin and gills of C. gariepinus from sampled three hatcheries in Uganda. Trichodina sp. and Gyrodactylus sp. being the dominant parasites were considered for assessment of pathology, however, Trichophrya sp., Apiosoma sp., Epistylis sp. and Epitheliocystis are briefly discussed.
MSc. Thesis 67 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Trichophrya sp.
Features observed from fresh smear categorically enabled us to follow taxonomic description of the parasite provided by Lom (1995) and Lom and Dykova (1992). The parasite belong to class Kinetophragminophora due to the fact that cilia may be lost in adults, subclass Suctoria because adults become sedentary with or without cilia and tentacles, order Suctorida having same the description as subclass and family Trichophryidae based on the fact that larvae have cilia which are lost once established on a suitable substrate like fish and develop tentacles. The presence of tentacles rather than cilia in the specimen observed is in agreement with the stated description of the genus. Several species of Trichophrya sp. are reported to parasitize gill lamellae (Lom, 1995; Rintamaki-Kinnunen and Valtonen, 1997), skin and fins (Post, 1992) of various fish. The parasites are associated with gill necrosis especially under heavy infections, which occur in fish predisposed to various stressors. Thus the parasites are more prevalent in cultured and/or aquaria fish, which are overcrowded or malnourished and supplied with high organically polluted water.
Despite the fact that studies have shown Trichophrya sp. cause gill, skin and fins hyperplasia and necrosis (Lom and Dykova, 1992; Post, 1992; Lom, 1995) especially under heavy infection, pathological effects of these species could not be assessed during the study. This was attributed to low prevalence and mean intensity of occurrence of the parasite. Furthermore, its pathological effects (if any) may have been masked by the dominating Trichodina sp. and Gyrodactylus sp. However, various reports from different authors show that minimal or no pathological changes are as a result of the presence of Trichophrya sp. For instance, Valtonen and Koskivaara (1994) found the parasite in very high intensities and even resisted salt and formalin treatment, no significant effect was observed on fish. Rintamaki-Kinnunen and Valtonen (1997) observed no mortalities associated with both Apiosoma sp. Epistylis sp. and Trichophrya sp. Post (1992) also stated that several species of Trichophrya sp. occur as commensals on fish gill lamellae. Therefore, with all this evidence from other authors, Trichophrya sp. may have not been involved in causing mortality at the low levels of mean intensities and abundances recorded during the study. And therefore, it can be considered less pathogenic to C. gariepinus fry and fingerlings
MSc. Thesis 68 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Apiosoma sp.
The parasite was characterized with solitary mode of living and possession of epical ring of cilia and stalks produced from adhesive point of sessile ciliates – scopula. According to Lom and Dykova (1992), the parasite belongs to the class Oligohymenophorea due to their distinct oral ciliary apparatus, subclass Peritrichia because of the bell-shaped and cylindrical body, order Peritrichida based on the presence of two or three rows of cilia (depending on species) at the anterior part (adoral) and family Epistylididae because they are sessile organisms especially when adults and possession of stalks produced from scopula.
As a consequence of low intensity of infestation, preferred site of occupation of this parasite was not ascertained, however, according to Lom (1995); Schmahl et al. (1989), the preferable sites of occurrence are the skin and fins. Furthermore, effects on fish due to Apiosoma sp. could not be established and assessed during the study because of very low prevalence as well as mean intensities and abundances. The results show that the parasite occurred on four fish (1.2%) and intensity range of 1 – 3 individuals per fish. The parasite also occurred amongst other dominant parasites (Trichodina sp. and Gyrodactylus sp), making it difficult if not impossible to assess tissue changes on/in fish. However, the parasite may cause suffocation of fish once present on gill in high intensities (Lom and Dykova, 1992).
Although the presence of Apiosoma sp. is reckoned to cause fish mortalities, the parasite is associated more with secondary infections like bacteria e.g. Aeromonas hydrophila (Lom and Dykova, 1992) and virus. Conversely, other authors have observed no mortalities or economically devastating effects of this parasite on fish (Lom, 1973b; Rintamaki-Kinnunen and Valtonen, 1997; Isaksen, 2003). As a result of low mean intensity and abundance of Apiosoma sp. on fish investigated and evidence from the previous researchers, this parasite can be considered less pathogenic as well to C. gariepinus and thus of low economic importance to aquaculturists.
MSc. Thesis 69 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Epistylis sp.
The low intensity and prevalence of this parasite made it difficult to assess pathogenicity as well as site of predilection. Systematically, this parasite is at the same taxonomic position as Apiosoma sp. (Lom and Dykova 1992) but is distinguished from Apiosoma sp. by the formation of colonies (Fig. 4.2.1d – ii). Furthermore Apiosoma sp. has a slightly longer circular stalk originating from a wide scopula (Fig. 4.2.1c) while Epistylis sp. had shorter more or less conical stalk originating from a thin scopula (Fig. 4.2.1d). Epistylis sp. is reported to induce mortality in fish (Lom and Dykova, 1992; Lom 1995; Phimmachak and Chanthavong, 2003) only in heavy infections but due a low intensity in occurrence during this study, the effects on fish could not be assessed. Lom, (1995) stated the Epistylis sp. has been alleged of causing mortalities in fingerlings of Micropterus salmoides in North America. However, like Apiosoma sp., Epistylis sp. has been encountered on several occasions on fish with no or minimal pathological effects (Rintamaki-Kinnunen and Valtonen, 1997; Isaksen, 2003). The parasite is considered as an ectocommensal i.e. using fish as living or moving substrate than as an ectoparasite. Nevertheless, it has been associated more with promotion of secondary infections than causing direct mortality (Lom and Dykova, 1992; Durborow, 2003).
It has been noted from literature Lom (1966, 1973b, 1995); Lom and Dykova (1992); Durborow (2003) that sessile ciliates are encountered in fish farms with water having high organic material from which they seem to derive nutrients viz organic debris and waterborne bacteria. The parasite therefore occurs in very high intensities and prevalence in highly organically polluted water and debris-laden sediments. Also, same authors point out that heavy infestations on the surfaces of fish clearly indicate that the host has been predisposed to debilitating environments and have reduced immune systems. However, the relatively good water quality as shown per the field measurements during the study, seems to explain the low prevalence and intensity of these parasites. Moreover, the flow through system in fry tanks is likely to wash out most of the debris from the system. The low intensity could also be due to inhibition by massive occurrence of Trichodina sp. observed from all hatcheries studied. Lom (1966) observed that sessile ciliates in an experiment disappeared whenever Trichodinids occurred in high intensities.
MSc. Thesis 70 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Further, Lom (1966) also observed that sessile ciliates – Apiosoma and Epistylis occurred in high intensities in winter but reduced or even disappeared in summer. The author hypothesised that fish become active during warmer periods rendering their surfaces unsuitable for settlement of these ciliates and the reverse true for cold winter season. Therefore, if Lom’s hypothesis holds, Clarias gariepinus fry and fingerlings cannot support massive populations of these sessile ciliates due to their relatively high activity.
Low pathological effects of these sessile ciliates may be attributed to their mode of attachment, where the scopula is just cemented on the surface of the hosts rather than penetration into the host tissues. However, Lom (1966, 1973b) stated that due to strong attachment on the surface could interfere with normal functioning of the occupied tissues e.g. reduced respiratory function when they occur on gills, hence documented mortality of fish. However, due to low intensity, such effects of reduced respiratory function may have not build up in this study.
Epitheliocystis
Epitheliocystis infections of up to 12 % prevalence were recorded in fry and fingerlings of cultured C. gariepinus from Uganda. Epitheliocystis is a common condition affecting gills or other tissues like kidney and spleen (Work et al., 2003) of both wild and cultured fish and have been associated with fish mortalities. The condition affects several cells including epithelial cells, chloride cells, or even kidney cells (Crespo et al., 1999; 2001). In this study, the infected cells appeared to be the gill epithelial cells due the fact that no other cell layer surrounded them. According to Lom and Dykova (1992), Epitheliocystis agents are grouped together with fish parasites rather than fungi, bacteria and viruses despite their inclusions. However, from ultrastructure studies, Epitheliocystis contained prokaryotic bacteria Chlamydia or Chlamydia-like organism, which elicits host reaction resulting into hypertrophy (Szakolczai et al., 1999; Crespo et al., 1999, 2001; Frasca Jr. et al., 2003). Similarly Lom (1995) and Paperna (1996) reported that Epitheliocystis are hypertrophic cells within the gill epithelium filled fine granules Chlamydia-like organisms having some features of rickettsiae. Therefore, Chlamydia bacteria or Chlamydia-like organisms or rickettsiae-like organisms are most likely causatives of the condition.
MSc. Thesis 71 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
In present study, relatively low mean intensity (ca. 2 cysts per fish) and relatively low prevalence (12.5 %) recorded, this condition may not have induced mortality. Previous studies observed that Epitheliocystis induced mortality when occurring at high infection percentages. Szakolczai et al (1999) suspected Epitheliocystis agents to have induced heavy mortality of pacu fish (Piaractus mesopotamicus) after recording an infection proportion of 30 to 40%. Furthermore, 6 –10 consecutive gill lamellae of sea bream had fused due Epitheliocystis occurring at a prevalence of 11% in juveniles and 50% in fingerlings (Crespo et al., 2001). Despite the low prevalence of Epitheliocystis observed in this study, infections should not be ignored as it may compromise with respiration after occlusion of the interlamellae spaces. In addition to the tender ages of the fish studied, which were already laden with parasite infestation, the relative low intensity of Epitheliocystis may have contributed to fish stress. Thus, information presented inhere should provides a baseline for further research into Epitheliocystis infections.
Trichodina sp. and Gyrodactylus sp.
These two parasites will be discussed concurrently due to the fact that they more or less occurred together and had same site of predilection hence presence of one seemed to affect the other. In addition, concomitant occurrence seemed to have their pathogenicity aggravated and hence more devastating.
Taxonomy
The measurements from fresh smears revealed that Trichodinids observed from all farms belong to a same species. Based on Lom and Dykova (1992) and the morphological features observed in the field, the Trichodina sp. belongs to the class Oligohymenophora due to their distinct oral ciliary apparatus. Because of the dome- shape or bell-shape and conspicuous cilia round the adoral (apical) pole, the parasite is placed in subclass Peritrichia. Moreover, due to flat disc-shape of the body assumed when the parasite is at rest especially on a flat surface, they are put under the order Mobilina. The parasite is classified under family Trichodinindae because of the presence of proteinous skeleton of denticles of the adhesive disc. Gyrodactylus sp. on the other hand belongs to the class of Monogenea, Platyhelminths (Boeger and Kritsky, 1993; Cribb et al. 2002).
MSc. Thesis 72 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
The genus is identified by possession of hooks of the next generation protruding in the central region of its body, opisthaptor armed with marginal hooks 12 – 16 surrounding two strong median hooks.
Sites of predilection
Caudal, anal, pelvic, and pectoral fins were the sites of preference at low infections. Later during investigations, the parasites spread to gills, head and barbels and the previous sites became increasingly more heavily infested in moderate and heavy infections. These sites of preference are the same as those reported in Buchmann and Bresciani (1998) for G. derjavini on rainbow trout. Buchmann (1999) suggested that parasites are attracted towards the lectin-rich goblet cells in the skin through lectin– carbohydrate interaction between fish and the monogeneans. Therefore the author suggested that Gyrodactylus sp. should prefer fins and ventral side of the body, due to high intensity of goblet cells. However, no or very few goblet cells were visible in these sites of predilection, maybe due to differences in fish species. Buchmann and Bresciani (1998) also observed that G. derjavini preferred the tail and corneal with low goblet cells in later stages of infestation. This was attributed to avoidance of host resistance mechanisms released through mucus in response to parasite invasion.
Occurrence
It was noted that Trichodina sp. appeared as earlier as two weeks post hatching, compared to Gyrodactylus sp., which appeared four weeks post hatching viz two weeks after change of water source from use of groundwater to surface water. The observation was attributed to the mode of attachment and movement behaviour of the parasites. Under light microscope, Trichodina sp. was observed to be frequently migrating all over the fish body. Barker et al. (2002) also observed that T. murmanica frequently moved randomly on winter founder fish and debris in attempt to find a suitable substrate. Probably during these movements, the parasite were dislodged and washed off the host by water currents. Lom (1973a) observed the Trichodinella epizootica was loosely attached to the host. Since Trichodinella are in the same taxonomic group with Trichodina, the early appearance of Trichodina sp. could have been due to its loose attachment on the host, which were dislodged and wash off its wild host by water currents into the pond.
MSc. Thesis 73 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
In contrast to Trichodina sp., Gyrodactylus sp. was seen firmly held on the host with occasional movements. Actually, Gyrodactylus sp. only moved when exposed to dry conditions during laboratory investigations. As a consequence of relatively strong attachment onto the host provided by haptors (Lester, 1972) and more of sedentary behaviour, water currents could not easily wash off Gyrodactylus sp. Therefore, difference in behavioural movement of Trichodina sp. and Gyrodactylus sp. coupled differences in ability to attach firmly to the hosts, may be the reason for differences in appearance on the hosts during this study.
The mean intensities and mean abundances of Trichodina sp. increased with fish age during the initial stages of infection. However, after attaining a peak at six weeks, both the mean intensity and abundance decreased to a more or less stable state. The reduction in mean intensities and mean abundances of Trichodina sp. could be due to preference of young fish probably due to low host resistance and direct competition with Gyrodactylus sp. for space. The combination of the two factors may have resulted into suppression of Trichodina sp. Preference of fry and fingerlings by Trichodina sp. and protozoa in general has been reported in previous studies for different fish species. Barker et al. (2002) in an experiment observed increase in the intensity of Trichodina sp. on winter founder after infection, which markedly declined after two weeks post infection and completely disappeared after five weeks of infection. The author attributed the decrease and disappearance of Trichodina sp. to host induced resistance. These results are similar to those of Pojmanska (1994) who observed that Protozoa decreased with age while Dactylogyrus sp. (Monogenea) increased. The same author attributed decrease in protozoan intensities to the development of host resistance to new infections. Rintamäki et al. (1994) also found a protozoan Chilodonella spp. decreasing with age. Similarly, McGuigan and Sommerville (1985) found higher prevalence and mean intensity of Trichodina sp. among smaller fish of less than 20 cm than larger fish of greater than 20 cm but no explanation was provided for these differences.
In contrast, Rintamäki-kinnunen and Valtonen (1996, 1997) observed that the prevalence and mean intensity of Protozoa infections increased with fish age. In their study, salmon yearlings were highly preferred by the parasites with mean intensity of 23.1 than the fingerlings.
MSc. Thesis 74 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
It is worth noting that in their study no monogenean was involved suggesting that Gyrodacytylus sp. found in the current study may have out competed Trichodina sp. Valtonen and Koskivaara (1994) also found Trichodina nigra infecting salmon skin increasing with age. During their study, Trichodinids seemed to have had no competitor because the trematode Diplostomum spathaceum and copepods present on the same host occurred at different sites – in the eyes and gills respectively. Nevertheless, there are reports showing that endoparasites can as well suppress ectoparasites. For instance, Larsen et al. (2002) observed that endoparasite (Anisakis sp larvae) reduced susceptibility of salmon trout to infections of ectoparasite (G. derjavini). These authors suggested that endoparasite provoked host immune mechanism which inhibited attachment and survival of G. derjavini. In this current study, however, it is likely that Gyrodactylus sp. suppressed the occurrence and intensity of Trichodina sp. as well. This maybe due to direct competition for space because both parasite species preferred same sites on the host: fins and body, but this is worthy more studies. Gyrodactylus sp. therefore, had its mean intensity and mean abundance increase with age except when interrupted with formalin treatment. The parasite also seemed to prefer relatively large and old fish but had its mean intensity and abundance declined slightly after the peak at six weeks. Buchmann and Bresciani (1998) obtained same results in an experimental study in which an increase in mean intensity of G. derjavini on rainbow trout with age was observed. Decrease of the mean intensity of the parasite after the peak is the same with experimental observations by Mansell et al. (2005) and implicated the decline to an increase in host immune response.
Frequency distribution plots revealed that infestation of Gyrodactylus sp. on fish host was generally over–dispersed both with pooled data from all farm, with data from individual farms and with age groups. Due to over-dispersion, a few hosts harboured many parasites while many hosts harboured very few parasites. The overdispersion observed could be explained by the spatial distribution of hosts due to artificial manipulation at the farms and direct reproduction due to viviparity of Gyrodactylus sp. and likely occurrence of offspring on the same host as parents. The reasons are supported with Esch and Fernändez (1993) and Anderson (1993) who indicated that overdispersion of parasites arises due to differences in host immune resistance, spatial distribution, direct reproduction on/in the host and behaviour.
MSc. Thesis 75 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
It has been stated that over–dispersion causes mortalities in heavily parasite infested hosts (Esch and Fernändez, 1993; Anderson, 1993). Thus mortalities of C. gariepinus observed could be due to heavy infections in hosts harbouring the parasites. Esch and Fernändez (1993) and Anderson (1993) also stated that lethal qualities of parasites would be manifested through killing the heavily infested host. They added that under such circumstances, parasite load would reduce, which was not observed during the current study. This was probably due to constant input of parasites through the constant supply surface water into the farm
Treatment of parasite infection
Both Trichodina sp. and Gyrodactylus sp. appeared susceptible to 40ml/l formalin treatments. Results from Sunfish farm (Fig. 4.4.2), though reflected in the pooled data (Fig. 4.4.1) showed fluctuations in the prevalence, mean intensity and abundance at eight and ten weeks after application of formalin at seven and nine weeks. However there was an increase in prevalence of Trichodina sp. with more or less mean intensity and abundance and increase mean intensity and mean abundance of Gyrodactylus sp. in during the next sampling date at the ninth week, exactly one week after first treatment. This suggests that the parasites are either partially affected or there is new input from the water supply. There is higher possibility that these parasites recolonised from the water supply, however, further experimental studies should be conducted. Although some studies have shown that formalin treatments are not effective on parasite infections, others revealed that formalin is very effective. Isaken (2003) observed that formalin treatments did not affect the patterns of ciliate infections in Atlantic salmon in a hatchery from Norway. Also T. murmanica were not eradicated from Atlantic cod after treatment of with 1:4000 formalin bath (Khan, 2004). Conversely, Rintamäki- kinnunen and Valtonen (1996) reported that formalin reduced the intensities of Trichodina sp. but was partially effective against G. salaris whose intensity decreased in the proceeding samplings. During the current study, second formalin treatment applied at nine weeks showed a drastic decline in the prevalence and mean intensity of both Trichodina sp. and Gyrodactylus sp. Therefore, we established that continuous application of formalin can keep these parasites at a very low intensity and prevalence. For effective control and/or elimination of these parasites, 40ml/l formalin (37 %) bath treatment for 1 hour must be repeated every three days (Stoskopf 1993).
MSc. Thesis 76 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Other highly recommended control and/or preventative drugs for ectoparasites are Toltrazuril (Baycox, Bayer) and Praziquantel (Droncit, Bayer) (Schmahl et al., 1989; Mehlhorn et al., 1988, Stoskopf, 1993).
Results also indicated that the duration of spread of Trichodina sp. and Gyrodactylus sp. to new hosts at initial infections was more or less same. Both parasites took about four weeks to peak in prevalence, mean intensity and mean abundance. The same duration to reach the maximum mean intensity was observed by Wells and Cone (1990) in their experiment. The intensity of Gyrodactylus colemanesis and G. salmonis peaked on 27th day – ca. four weeks post infection. Thus when Gyrodactylus sp. invades a system, it is likely to take about four weeks to reach the stationary phase. This rate of colonisation and multiplication clearly indicates that this parasite can quickly spread and become devastating once in the system.
Despite the fact the both parasites took more or less same duration to reach the maximum population, they had differences in reinfection post formalin administration. Trichodina sp. reinfected all its hosts – the prevalence increased to 100 % at nine weeks after the formalin application but mean intensity and abundance remained more or less the same. On the other hand, Gyrodactylus sp. never reinfected to all the hosts – the prevalence never reached 100% after treatment but the mean intensity and abundance increased. The differences were probably due to the reproduction mode, strength of attachment to the host, movement behaviour, competition, mode of transmission and susceptibility to formalin bath. The viviparity of Gyrodactylus sp., allows accumulation of the offspring and parents on the same host resulting in high intensity and abundance. Further the confinement of the offspring to the same host as parents, may explain the low rate of invasion of Gyrodactylus sp. to new hosts. As indicated early, Gyrodactylus sp. was usually found firmly attached on their hosts and had occasional movements unlike Trichodina sp., which was frequently found moving around. Besides this, Gyrodactylus sp. has strong hook development for attachment as well as minimal movement (Lester 1972); water currents cannot dislodge the parasite easily. The mode of transmission of Gyrodactylus sp. offspring to new hosts is not clear but overcrowding probably accelerates it.
MSc. Thesis 77 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Buchmann et al. (2004a) in an experimental study noted that water current played less significant role in attraction of Gyrodactylus sp. towards the host. It is therefore suggested that Gyrodactylus sp. are attracted by chemical products like lectins from the skin through parasite–host interaction (Buchmann and Bresciani, 1998; Buchmann, 1999). Therefore, offspring could easily be attracted to hosts they are “born” possibly due to high chemical intensity than the far away host whose chemical intensity may be diluted in water before reaching the parasites. This may explain the high intensity and abundance build up of Gyrodactylus sp. on a few infested fish rather than spreading to the entire host population. Conversely, Trichodina sp. have a binary fission mode of reproduction where daughter separate longitudinally (Lom and Dykova, 1992; Lom, 1995). However, during this period the attachment apparatus of the parasites are also under division, hence, presuppose loose grips onto the host. Thus the parasite can be dislodged by water current to another host hence reducing the mean intensity and abundance on previous host but increase the prevalence of parasites fish. In addition to their weak attachment, the offspring may easily dislodged during their constant movement is search of new host. Incase Trichodina sp. was out competed, presumably for space by Gyrodactylus sp., may cause dislodging in search for new sites for attachment. Thus with the aid of water current, Trichodina sp. may be transported to another host resulting in high prevalence but low mean intensity. It is also possible that Trichodina sp. is less susceptible to formalin than Gyrodactylus sp. The former parasite could have just dropped off some hosts resulting in decline in prevalence but formalin did not affect their mean intensities and abundances. This supports observations of (Isaken, 2003; Khan, 2004) who reported less susceptibility of Trichodina sp. Conversely, Gyrodactylus sp. could have been killed during the treatment but increase in mean intensity in subsequent sampling was probably due to reinfection from the water supply.
Histopathology
Infected fish were lethargic, accumulated on the water surface and rested in a vertical position in addition to skin discoloration. Although most fish diseases can manifest these macroscopic and behavioral changes, the observed features coincided with the presence and the intensity increased with the intensity of parasite infections.
MSc. Thesis 78 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Still several authors observed similar changes in fish infested with parasites (Roberts, 1978; Stoskopf, 1993; Cone, 1995). Further, fungus usually associated with parasites as a secondary infection (Roberts, 1978; Stoskopf, 1993; Cone, 1995) became more pronounced in heavily parasitized fish. This was suggestive of mechanical injury due to haptors of Gyrodactylus sp. and may be Trichodina sp. at the points of attachment on the skin and gills that provided avenues for fungus entry and proliferation. Several researchers have reported association of fungus with ectoparasites (Roberts, 1978; Stoskopf, 1993; Williams and Jones, 1994; Cone, 1995). It is also well known that fungal diseases are very pathogenic (Roberts, 1978; Stoskopf, 1993; Cone, 1995). Thus mortalities of C. gariepinus observed could also be attributed to fungus, which was promoted by the presence of parasites. However, further studies are required to describe and to assess the pathogenicity of fungi occurring in catfish fishery in Uganda.
Microscopically, significant proliferative pathological changes were observed in the gills. These included oedema within intercellular spaces of epithelial and pillar cells and hyperplasia of gill lamellae. Subsequently there was fusion of the secondary gill lamellae. These changes occurred concomitantly with increase in the intensity of parasite infections. Mansell et al. (2005) also observed that the occurrence of hyperplastic lamellae increased as the G. derjavini infection progressed. The poor re- absorption of body fluids back to the capillaries (Roberts, 1978), characteristic of oedematous tissues clearly indicates poor of ion balance within the fish. In addition to poor ionic balance, accumulation of body fluids in the intercellular spaces reduces the rate of gaseous exchange. Thus, these abnormal changes in the gills may have caused dysfunction of the general fish system and subsequently, mortality may have followed.
Furthermore, preference of water surface by infected fish presumably to gain more oxygen was indicative of compromised respiration. Therefore, presence of parasites coupled with occasional low oxygen in tanks was purported to be contributing significantly to the massive mortalities of catfish fry and fingerling. In addition, the presence of rodlet cells suggested the host reaction to parasite infections and poor environmental conditions. Reite (2005) observed that occurrence of rodlet cells were common in all salmonids caught in their natural environment than those kept under controlled conditions with respect to water quality.
MSc. Thesis 79 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
In the same study, Reite (2005) observed that cyprinid species, which were infested with helminths, had their epithelial tissues consisting of fairly high number of rodlet cells than normal fish. Therefore the occurrence of the rodlet cells, accumulation of infected fish on the water surface appeared to support that the pathological changes in gills were likely due to parasites.
Occurrence of Trichodina sp. alone in fish, however, seemed not to be highly pathogenic because no mortalities were observed in such instances. However histological section showed Trichodina sp. in association with the oedematous gills suggesting that the parasite may cause osmoregulation obstruction within gills. Similarly, Longshaw et al. (2004) found numerous Trichodina sp. on three pipefish but no observable host response except some increased mucus production and mild epithelial hyperplasia. During this study, Gyrodactylus sp. was always in association with Trichodina sp., thus the damage due Gyrodactylus sp. alone could not be evaluated. However, previous reports reveal that Gyrodactylus sp. is very pathogenic and responsible for massive mortalities in fish (Williams and Jones, 1994; Cone, 1995; Paperna, 1996). Therefore, sole occurrence of Trichodina sp. in fish can be considered less pathogenic. But the pooled effect of the two parasites – Gyrodactylus sp. and Trichodina sp. – seems additive and aggravated the pathogenic effects of the parasites, similar to the observation by Barker et al. (2002).
Minor pathological changes were observed in the skin samples of the fish specimen examined. There was infiltration of melanomacrophage cells (MMCs) especially at sites of parasite infestation. These are the main phagocytotic cells in fish – responsible for removing dead cells and engulfed antibodies at sites of infection (Agius and Roberts, 2003). Therefore, their presence may indicate death of some cells probably due to parasitic injury. However, little information exists in regard to reaction of these cells during ectoparasitic infestation. At points of parasite attachment, was mild hypertrophy of the skin epidermal cells. This change also indicates poor ion balance within the host cells (Roberts, 1978; Stoskopf, 1993). Thus the infected fish appeared to have had compromised ionic balance which further decreased general physiological function and hence death.
MSc. Thesis 80 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
In this study goblet cells showed no correlation with either age or parasite intensity. The cells occurred in very low numbers when present or were absent in low, moderate and heavy parasitized fish. However, various studies revealed a reduction in mucus cells (Pottinger et al. 1984, Wells and Cone, 1990) while others revealed an increase (Buchmann and Uldal 1997) in the presence of Gyrodactylus sp. No apparent changes in epidermal tissues and cell intensities were observed in the skin of stickleback fish infested with G. alexanderi (Lester, 1972). The author observed that G. alexanderi did not penetrate the epithelial cells but only exerted pressure them. Furthermore, (Appleby et al., 1997) reported that G. salaris infesting Atlantic salmon elicited no significant changes in mucus cells as well as epithelium and there was no correlation between the intensity of parasites and mucus cell intensity, epithelium thickness and epithelial cells. Therefore, the reports and results clearly show a wide diversity in manifestation of histopathological changes of ectoparasites on the fish skin. This may range from no or minor changes in skin epidermis to increase in goblet cells as well as epidermal wall thickness or to a decrease in goblet cells and reduction in epidermis thickness. Nevertheless, after formalin treatment, there was a relative increase in goblet cells compared to their counterparts before treatment. The increase in goblet cells after formalin application could have been a host response to the toxic effect of formalin through cell proliferation. In low concentrations (≤ 50 ppm) – a common practice at the sunfish farm – formalin treatment is observed to elicit proliferation of goblet cells (Buchmann et al., 2004b; Speare et al., 1997).
During the study, it was observed that there was a general increase in the intensity club cells in the caudal fin. The increase was concomitant with age and intensity of parasites. Since there is scanty information about club cells especially in regard to parasite infestation, probably the increase in club cell concentration was age related though parasites ought to have accelerated the production. However, fish specimen from eight weeks (after formalin treatment) showed a decrease in club cells intensity. The decrease was attributed to suppression by goblet cells during host response. Still extensive work is required to establish the importance of club cells in host defence
MSc. Thesis 81 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
6 CONCLUSIONS AND RECOMMENDATIONS
Conclusions
In addition to provision of cheap source of food (proteins), acting as a gene bank and production of fish for restocking, aquaculture – fish farming – is an important source of income for the alleviation of poverty. For these purpose, the Ugandan government extensively promotes fish farming. However, most of the studies in this field are dealing with development of breeding technologies, little is invested in fish health. Among the cultivable species, African catfish, Clarias gariepinus has become popular in Uganda. The fish being produced extensively for purposes of income generation, food, restocking and as bait for Nile perch. However, several farmers are facing a problem of massive mortalities of fry and fingerlings especially in intensive culture of the systems. The mortalities are attributed to many factors including poor nutrition, poor environmental conditions, parasites and their associated diseases. However, there is little or lack of information on fish diseases and parasites. Therefore, the study was conducted in three farms over three months to establish information on the occurrence, the prevalence and the pathology of protozoan and monogenean parasites in aquaculture conditions. Methods involved an application of routine parasitological and histological techniques. In addition, important physical and chemical parameters in aquaculture were concurrently measured. Based on the results and discussion, the following conclusion can be drawn for the study: • Except for occasional low of dissolved oxygen, water quality parameters were within the tolerable range for African catfish. Temperature was 21°C – 28°C (tolerable range is 8°C – 35°C), pH was 5 – 8 (tolerable range is 3 – 10), ammonia was approximately 0.11mg/l (maximum tolerable concentration for larvae is 2.3mg/l) • Surface water inflow was the major route for parasites to invade fish farms receiving water supply from the stream while semi-aquatic organisms (amphibians, reptiles etc) are probably involved in transmission of parasites to fish farms depending ultimately on groundwater.
MSc. Thesis 82 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
• Epistylis sp., Apiosoma sp. and Trichophrya sp. were present in very low prevalence (> 2%) and intensity (> 5 individuals/fish) while a protozoan Trichodina sp. and monogenean Gyrodactylus sp. were the dominant parasites found in all fish hatcheries. • Together with information from literature, Epistylis sp., Apiosoma sp. and Trichophrya sp. were considered less pathogenic parasites and may be of less economic importance to aquaculturists. • Epitheliocystis was also found on gills of young African catfish but with relatively low intensity and prevalence. Therefore, further studies are required to assign a correct status of their pathogenicity in fish. • Although the prevalence, the mean intensity and the mean abundance of both Trichodina sp. and Gyrodactylus sp. increased with age at the initial stages of infection, the mean intensity and the mean abundance of Trichodina sp. decreased after attaining a peak at six weeks probably due to host immunity and competition with Gyrodactylus sp. for space. • Probably the higher likelihood of parasites getting new host and higher parasite abundance in surface water may have contributed to higher mean intensities and mean abundance at the surface water-supplied farms (Sunfish farm and KRS) than at the groundwater-supplied farm (Ssebinyansi farm). • Trichodina sp. seemed to be less susceptible to 40mg/l formalin treatment than Gyrodactylus sp. Reappearance of Gyrodactylus sp. on fish could be through recolonisation from the water supply. Differences in the appear at the initial infection and reappearance after formalin treatment of Trichodina sp. and Gyrodactylus sp. were reckoned to the mode of attachment, frequency of movements and mode of reproduction. • Frequency distribution of Gyrodactylus sp. showed over-dispersion a clear indication of parasite-induced mortality for heavily infested hosts. The causes of over–dispersion where artificial manipulation of the farms creating spatial distribution and viviparous mode of reproduction of the parasite.
MSc. Thesis 83 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
• No mortalities were observed in fish infected with Trichodina sp. but the parasite caused hypertrophy of the skin epidermal cells and was associated with gill oedema. However, concomitant occurrence of Trichodina sp. and Gyrodactylus sp. aggravated the pathogenic effect on fish, hence increasing mortality. • When occurring together, the parasites were associated with oedema and hyperplasia in the gills and resulted in gill lamellae fusion, eliciting of the release of rodlet cells into gill lamellae as well as infiltration of melanomacrophage cells into the skin epidermis and mild hypertrophy of the epidermal cells. These pathological changes appeared to interfere with fish respiration and ion exchange, which may have reduced general fish physiology and subsequently causing fish death. • Reappearance of mucus cells after formalin treatment was probably a host reaction to toxicity of the chemical rather than to parasites. Increase of club cell intensity was related with age but parasites may have accelerated the process.
Recommendations to farmers
• Farms supplied with surface water should use filter systems capable of reducing the parasite loads into the ponds, such systems may include sand filters, installation of a series of different filter nets in water distribution systems. • Intensive systems should adapt a routine aeration program irrespective of conditions to avoid unpredicted oxygen depletion in ponds. At least water should be flow into ponds from 10 or more cm above the water level and have a constant monitoring scheme. • Formalin for control must be applied at a rate of 40 ml/l for 1 hour every 3 days after parasite infestation. However, other drugs are Toltrazuril (Baycox) applied at rates of 50mg/l for 20 minutes once a week against Trichodina sp. and 10 – 20 mg/l for 4 hours on the first treatment, 1 hour duration in the subsequent treatments at 2 day intervals for 6 times against monogeneans but with intensive inspection on fish behaviour; Praziquantel (Droncit) at a rate of 10mg/l for 3 hours for monogeneans. • Investigations of parasite invasions should be considered as part of the farm management objectives and conducted at least every two weeks.
MSc. Thesis 84 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Recommendations to researchers
• Information on the histology of Clarias gariepinus for different age groups is urgently required. Such information is very vital for fish pathologists working without control experiments or on monitoring scheme. • Long-term studies of parasites in aquaculture are important to establish the parasite population dynamics and hence recommendation of best control and preventative measures. • Studies on bacterial, viral and fungal diseases of fish either associated with or without parasites are needed urgently. • Studies to ascertain an optimum stocking rate with minimum stress on fish for intensive system are needed. • Effects of disease and parasite outbreak in fish farms on wild fish population in the drainage channel should be assessed. This will reduce on the reintroduction into water source supplying the farms. • Studies on the diurnal changes of water parameters at fish hatcheries must be conducted to elucidate the effect of water quality on induction of mortality or predisposing fish to diseases. • Research on the assessment of economic losses due to parasites and diseases is required to provide the best control and preventative measures.
MSc. Thesis 85 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
REFERENCES
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APPENDICES
Appendix 1: Physical – chemical parameters of water from study hatcheries
Dissolved Temperature Oxygen Nitrite Ammonium Date Fish Age Farm (°C) (mg/l) pH (mg/l) (mg/l) 21st 04 1 Sunfish 25.2 7.9 5.5 0.05 0.2 22nd 04 2 Sunfish 23.8 8.5 5.8 0.05 0.2 23rd 04 4 Sunfish 28.4 8.0 7.5 0.05 0.2 24th 04 5 Sunfish 28.6 5.6 7.0 0.05 0.2 25th 04 8 Sunfish 28.9 7.7 7.5 0.05 0.2 22nd 04 8 Sunfish 26.3 5.8 6.5 0.05 0.2 25th 04 9 Sunfish 27.3 5.9 7.0 0.05 0.2 26th 04 9 Sunfish 27.5 5.2 7.0 0.05 0.2 2nd 05 10 Sunfish 25.5 5.7 6.5 0.05 0.2 3rd 05 10 Sunfish 26.2 6.9 6.8 0.05 0.2 4th 05 1 Kajjansi 23.0 6.7 5.9 0.05 0.2 9th 05 3 Sunfish 21.8 7.3 6.0 0.05 0.2 10th 05 4 Sunfish 22.5 7.9 6.5 0.05 0.2 11th 05 2 Kajjansi 23.0 6.2 6.7 0.05 0.2 13th 05 10 Fish farmer 22.0 6.9 7.8 0.05 0.2 16th 05 4 Sunfish 23.8 6.7 6.0 0.05 0.2 16th 05 5 Sunfish 25.3 5.6 7.0 0.05 0.2 18th 05 3 Kajjansi 24.1 7.0 7.7 0.05 0.2 20th 05 1 Fish farmer 23.5 6.2 6.5 0.05 0.2 23rd 05 5 Sunfish 24.9 5.4 7.0 0.05 0.2 25th 05 4 Kajjansi 23.0 7.0 7.5 0.05 0.2 25th 05 2 Sunfish 24.0 6.8 6.0 0.05 0.2 27th 05 2 Fish farmer 23.4 6.4 6.5 0.05 0.2 30th 05 7 Sunfish 24.3 6.4 7.0 0.05 0.2 01st 06 5 Kajjansi 23.8 7.2 7.5 0.05 0.2 08th 06 6 Kajjansi 22.0 6.8 6.5 0.05 0.2 11th 06 4 Fish farmer 23.6 5.7 7.0 0.05 0.2 15th 06 7 Kajjansi 22.3 6.2 7.0 0.05 0.2 17th 06 5 Fish farmer 23.7 7.0 7.0 0.05 0.2
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Appendix 2: Parasites counted from fish samples and calculated mean intensity and abundance Intensity of Trichodina sp. is a mean of individuals in five fields of view expressed as individuals per cm2 per fish.
Gyrodactylus sp. expressed as individuals per fish
TL – Total length, Tr. sp. – Trichodina sp., Gy. Sp. – Gyrodactylus sp. Intensity Mean intensity Mean abundance Farm Age TL Tr. sp Gy.sp Tr.sp Gy. sp Tr. sp Gy. sp Sunfish 1 1.0 0 0 0.0 0.0 0.0 0.0 1 1.0 0 0 0.0 0.0 0.0 0.0 1 0.9 0 0 0.0 0.0 0.0 0.0 1 1.0 0 0 0.0 0.0 0.0 0.0 1 1.0 0 0 0.0 0.0 0.0 0.0 1 0.9 0 0 0.0 0.0 0.0 0.0 1 1.0 0 0 0.0 0.0 0.0 0.0 1 0.8 0 0 0.0 0.0 0.0 0.0 1 1.0 0 0 0.0 0.0 0.0 0.0 KRS 1 0.9 0 0 0.0 0.0 0.0 0.0 1 0.8 0 0 0.0 0.0 0.0 0.0 1 0.8 0 0 0.0 0.0 0.0 0.0 1 1.0 0 0 0.0 0.0 0.0 0.0 1 0.9 0 0 0.0 0.0 0.0 0.0 1 1.0 0 0 0.0 0.0 0.0 0.0 1 1.0 0 0 0.0 0.0 0.0 0.0 1 0.8 0 0 0.0 0.0 0.0 0.0 1 1.0 0 0 0.0 0.0 0.0 0.0 1 0.9 0 0 0.0 0.0 0.0 0.0 1 0.7 0 0 0.0 0.0 0.0 0.0 Ssebinyansi 1 0.9 0 0 0.0 0.0 0.0 0.0 1 1.0 0 0 0.0 0.0 0.0 0.0 1 0.9 0 0 0.0 0.0 0.0 0.0 1 0.9 0 0 0.0 0.0 0.0 0.0 1 0.9 0 0 0.0 0.0 0.0 0.0 1 0.8 0 0 0.0 0.0 0.0 0.0 1 0.9 0 0 0.0 0.0 0.0 0.0 1 1.0 0 0 0.0 0.0 0.0 0.0 1 1.0 0 0 0.0 0.0 0.0 0.0 1 1.0 0 0 0.0 0.0 0.0 0.0 Sunfish 2 0.9 0 0 0.0 0.0 0.0 0.0 2 1.0 2 0 1.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 0.8 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0
MSc. Thesis 99 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Intensity Mean intensity Mean abundance Farm Age TL Tr. sp Gy.sp Tr.sp Gy. sp Tr. sp Gy. sp 2 0.9 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.1 4 0 2.0 0.0 0.1 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 KRS 2 1.0 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.2 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0
MSc. Thesis 100 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Intensity Mean intensity Mean abundance Farm Age TL Tr. sp Gy.sp Tr.sp Gy. sp Tr. sp Gy. sp Ssebinyansi 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 0.9 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.1 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.1 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 2 1.0 0 0 0.0 0.0 0.0 0.0 Sunfish 3 1.6 0 0 0.0 0.0 0.0 0.0 3 1.3 13 0 0.9 0.0 0.4 0.0 3 1.4 25 0 1.8 0.0 0.8 0.0 3 1.2 7 0 0.5 0.0 0.2 0.0 3 0.9 1 0 0.1 0.0 0.0 0.0 3 1.0 0 0 0.0 0.0 0.0 0.0 3 1.3 0 0 0.0 0.0 0.0 0.0 3 1.0 0 0 0.0 0.0 0.0 0.0 3 0.8 9 0 0.6 0.0 0.3 0.0 3 0.9 17 0 1.2 0.0 0.5 0.0 3 1.2 0 0 0.0 0.0 0.0 0.0 3 0.9 2 0 0.1 0.0 0.1 0.0 3 1.1 0 0 0.0 0.0 0.0 0.0 3 1.2 0 0 0.0 0.0 0.0 0.0 3 1.2 0 0 0.0 0.0 0.0 0.0 3 1.3 0 0 0.0 0.0 0.0 0.0 3 0.7 0 0 0.0 0.0 0.0 0.0 KRS 3 0.8 2 0 0.1 0.0 0.1 0.0 3 1.1 5 0 0.4 0.0 0.2 0.0 3 1.0 11 0 0.8 0.0 0.4 0.0 3 1.0 0 0 0.0 0.0 0.0 0.0 3 0.9 4 0 0.3 0.0 0.1 0.0 3 1.0 0 0 0.0 0.0 0.0 0.0 3 1.0 0 0 0.0 0.0 0.0 0.0 3 0.9 8 0 0.6 0.0 0.3 0.0 3 0.9 0 0 0.0 0.0 0.0 0.0 3 1.0 13 0 0.9 0.0 0.4 0.0 3 1.0 9 0 0.6 0.0 0.3 0.0 3 0.9 0 0 0.0 0.0 0.0 0.0 3 1.0 0 0 0.0 0.0 0.0 0.0 3 1.1 0 0 0.0 0.0 0.0 0.0
MSc. Thesis 101 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Intensity Mean intensity Mean abundance Farm Age TL Tr. sp Gy.sp Tr.sp Gy. sp Tr. sp Gy. sp Ssebinyansi ------Sunfish 4 1.0 28 0 0.7 0.0 0.5 0.0 4 1.2 25 19 0.6 1.9 0.4 0.3 4 1.7 71 0 1.7 0.0 1.2 0.0 4 1.2 115 5 2.7 0.5 1.9 0.1 4 1.2 89 0 2.1 0.0 1.5 0.0 4 1.0 30 11 0.7 1.1 0.5 0.2 4 1.1 165 76 3.8 7.6 2.8 1.3 4 1.3 25 0 0.6 0.0 0.4 0.0 4 1.2 22 16 0.5 1.6 0.4 0.3 4 1.2 16 3 0.4 0.3 0.3 0.1 4 1.3 15 0 0.3 0.0 0.3 0.0 4 1.1 14 2 0.3 0.2 0.2 0.0 4 1.1 17 0 0.4 0.0 0.3 0.0 4 1.5 49 9 1.1 0.9 0.8 0.2 4 1.2 52 41 1.2 4.1 0.9 0.7 4 1.4 55 0 1.3 0.0 0.9 0.0 4 1.2 21 23 0.5 2.3 0.4 0.4 4 1.8 105 0 2.4 0.0 1.8 0.0 KRS 4 1.0 9 0 0.2 0.0 0.2 0.0 4 1.3 11 0 0.3 0.0 0.2 0.0 4 1.0 23 0 0.5 0.0 0.4 0.0 4 1.0 0 0 0.0 0.0 0.0 0.0 4 0.9 0 0 0.0 0.0 0.0 0.0 4 0.9 25 0 0.6 0.0 0.4 0.0 4 1.2 16 0 0.4 0.0 0.3 0.0 4 1.1 12 0 0.3 0.0 0.2 0.0 4 1.0 18 0 0.4 0.0 0.3 0.0 4 0.9 23 0 0.5 0.0 0.4 0.0 4 1.0 9 0 0.2 0.0 0.2 0.0 4 0.9 7 0 0.2 0.0 0.1 0.0 4 1.0 10 0 0.2 0.0 0.2 0.0 4 1.0 4 0 0.1 0.0 0.1 0.0 4 1.0 3 0 0.1 0.0 0.1 0.0 4 0.9 13 0 0.3 0.0 0.2 0.0 4 1.2 19 0 0.4 0.0 0.3 0.0 4 1.0 0 0 0.0 0.0 0.0 0.0 4 1.0 0 0 0.0 0.0 0.0 0.0 4 1.1 6 0 0.1 0.0 0.1 0.0 4 1.1 0 0 0.0 0.0 0.0 0.0 4 1.0 12 0 0.3 0.0 0.2 0.0 4 1.1 2 0 0.0 0.0 0.0 0.0 4 1.0 11 0 0.3 0.0 0.2 0.0 4 1.1 27 0 0.6 0.0 0.5 0.0 4 1.0 0 0 0.0 0.0 0.0 0.0
MSc. Thesis 102 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Intensity Mean intensity Mean abundance Farm Age TL Tr. sp Gy.sp Tr.sp Gy. sp Tr. sp Gy. sp Ssebinyansi 4 1.6 15 0 0.3 0.0 0.3 0.0 4 1.4 0 0 0.0 0.0 0.0 0.0 4 1.5 0 0 0.0 0.0 0.0 0.0 4 1.4 0 0 0.0 0.0 0.0 0.0 4 1.1 0 0 0.0 0.0 0.0 0.0 4 1.6 0 0 0.0 0.0 0.0 0.0 4 1.7 91 0 2.1 0.0 1.5 0.0 4 1.8 0 0 0.0 0.0 0.0 0.0 4 1.5 23 0 0.5 0.0 0.4 0.0 4 1.4 4 0 0.1 0.0 0.1 0.0 4 1.4 0 0 0.0 0.0 0.0 0.0 4 1.2 0 0 0.0 0.0 0.0 0.0 4 1.8 11 0 0.3 0.0 0.2 0.0 4 1.5 0 0 0.0 0.0 0.0 0.0 4 1.5 0 0 0.0 0.0 0.0 0.0 4 1.4 0 0 0.0 0.0 0.0 0.0 Sunfish 5 1.7 43 21 1.0 0.8 0.9 0.5 5 3.6 36 7 0.8 0.3 0.8 0.2 5 2.4 49 0 1.1 0.0 1.1 0.0 5 1.8 10 56 0.2 2.2 0.2 1.2 5 2.4 97 0 2.2 0.0 2.1 0.0 5 1.9 103 3 2.3 0.1 2.2 0.1 5 2.8 12 17 0.3 0.7 0.3 0.4 5 2.0 79 21 1.8 0.8 1.7 0.5 5 2.8 31 46 0.7 1.8 0.7 1.0 5 2.2 28 42 0.6 1.7 0.6 0.9 5 1.9 33 8 0.8 0.3 0.7 0.2 5 2.1 87 0 2.0 0.0 1.9 0.0 5 2.0 71 7 1.6 0.3 1.5 0.2 5 1.8 19 12 0.4 0.5 0.4 0.3 5 1.6 0 0 0.0 0.0 0.0 0.0 5 1.7 11 0 0.3 0.0 0.2 0.0 5 2.4 16 11 0.4 0.4 0.3 0.2 5 1.8 21 0 0.5 0.0 0.5 0.0 5 1.7 19 0 0.4 0.0 0.4 0.0 5 2.2 32 5 0.7 0.2 0.7 0.1 5 1.6 27 3 0.6 0.1 0.6 0.1 5 2.1 22 17 0.5 0.7 0.5 0.4 5 1.5 0 0 0.0 0.0 0.0 0.0 5 1.8 26 10 0.6 0.4 0.6 0.2 KRS 5 1.2 78 0 1.8 0.0 1.7 0.0 5 1.7 62 0 1.4 0.0 1.3 0.0 5 1.2 105 0 2.4 0.0 2.3 0.0 5 1.4 170 0 3.9 0.0 3.7 0.0 5 1.3 112 0 2.5 0.0 2.4 0.0 5 1.3 96 0 2.2 0.0 2.1 0.0 5 1.3 107 0 2.4 0.0 2.3 0.0 5 1.5 194 0 4.4 0.0 4.2 0.0 5 1.4 200 0 4.5 0.0 4.3 0.0 5 1.5 187 0 4.3 0.0 4.1 0.0
MSc. Thesis 103 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Intensity Mean intensity Mean abundance Farm Age TL Tr. sp Gy.sp Tr.sp Gy. sp Tr. sp Gy. sp Ssebinyansi 5 1.8 124 7 2.8 0.3 2.7 0.2 5 2.2 100 22 2.3 0.9 2.2 0.5 5 1.4 78 6 1.8 0.2 1.7 0.1 5 1.6 65 9 1.5 0.4 1.4 0.2 5 2.3 39 4 0.9 0.2 0.8 0.1 5 1.9 36 16 0.8 0.6 0.8 0.3 5 1.7 130 0 3.0 0.0 2.8 0.0 5 1.7 78 19 1.8 0.8 1.7 0.4 5 1.6 39 2 0.9 0.1 0.8 0.0 5 1.6 13 0 0.3 0.0 0.3 0.0 5 2.0 132 24 3.0 1.0 2.9 0.5 5 2.2 115 0 2.6 0.0 2.5 0.0 Sunfish 6 2.3 210 294 10.5 18.4 10.5 14.7 6 2.0 117 301 5.9 18.8 5.9 15.1 6 1.6 91 623 4.6 38.9 4.6 31.2 6 1.8 76 189 3.8 11.8 3.8 9.5 6 2.1 250 119 12.5 7.4 12.5 6.0 6 2.9 120 203 6.0 12.7 6.0 10.2 6 2.1 167 483 8.4 30.2 8.4 24.2 6 1.7 112 287 5.6 17.9 5.6 14.4 6 2.2 74 42 3.7 2.6 3.7 2.1 6 1.9 23 140 1.2 8.8 1.2 7.0 KRS 6 2.0 286 14 14.3 0.9 14.3 0.7 6 2.0 246 21 12.3 1.3 12.3 1.1 6 1.6 263 5 13.2 0.3 13.2 0.3 6 1.8 241 9 12.1 0.6 12.1 0.5 6 1.9 289 0 14.5 0.0 14.5 0.0 6 2.2 285 1 14.3 0.1 14.3 0.1 6 2.1 211 0 10.6 0.0 10.6 0.0 6 1.7 250 0 12.5 0.0 12.5 0.0 6 2.0 198 7 9.9 0.4 9.9 0.4 6 1.9 224 0 11.2 0.0 11.2 0.0 Sunfish 7 2.9 65 119 2.2 4.0 2.2 4.0 7 3.6 53 133 1.8 4.4 1.8 4.4 7 2.8 97 343 3.2 11.4 3.2 11.4 7 4.1 62 217 2.1 7.2 2.1 7.2 7 4.0 45 161 1.5 5.4 1.5 5.4 7 2.8 56 252 1.9 8.4 1.9 8.4 7 3.2 90 511 3.0 17.0 3.0 17.0 7 3.1 38 231 1.3 7.7 1.3 7.7 7 3.9 26 644 0.9 21.5 0.9 21.5 7 2.9 52 364 1.7 12.1 1.7 12.1 7 3.4 66 322 2.2 10.7 2.2 10.7 7 2.1 34 189 1.1 6.3 1.1 6.3 7 2.2 55 182 1.8 6.1 1.8 6.1 7 3.2 92 455 3.1 15.2 3.1 15.2 7 3.6 29 483 1.0 16.1 1.0 16.1
MSc. Thesis 104 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Intensity Mean intensity Mean abundance Farm Age TL Tr. sp Gy.sp Tr.sp Gy. sp Tr. sp Gy. sp KRS 7 2.2 27 217 0.9 7.2 0.9 7.2 7 2.7 34 189 1.1 6.3 1.1 6.3 7 4.2 49 203 1.6 6.8 1.6 6.8 7 3.6 61 147 2.0 4.9 2.0 4.9 7 3.4 47 133 1.6 4.4 1.6 4.4 7 2.9 65 329 2.2 11.0 2.2 11.0 7 2.6 52 126 1.7 4.2 1.7 4.2 7 2.3 23 175 0.8 5.8 0.8 5.8 7 4.1 37 182 1.2 6.1 1.2 6.1 7 2.9 25 238 0.8 7.9 0.8 7.9 7 3.1 22 315 0.7 10.5 0.7 10.5 7 3.6 42 252 1.4 8.4 1.4 8.4 7 3.2 71 161 2.4 5.4 2.4 5.4 7 2.7 83 140 2.8 4.7 2.8 4.7 7 4.0 78 259 2.6 8.6 2.6 8.6 Sunfish 8 5.9 77 0 7.0 0.0 6.4 0.0 8 4.9 136 83 12.4 8.3 11.3 6.9 8 6.5 0 28 0.0 2.8 0.0 2.3 8 5.9 29 35 2.6 3.5 2.4 2.9 8 4.6 16 19 1.5 1.9 1.3 1.6 8 4.4 29 43 2.6 4.3 2.4 3.6 8 3.5 9 67 0.8 6.7 0.8 5.6 8 3.6 58 17 5.3 1.7 4.8 1.4 8 5.0 14 0 1.3 0.0 1.2 0.0 8 5.5 39 31 3.5 3.1 3.3 2.6 8 4.3 28 41 2.5 4.1 2.3 3.4 8 3.9 105 53 9.5 5.3 8.8 4.4 Sunfish 9 7.0 19 94 1.4 8.5 1.4 6.7 9 6.2 13 0 0.9 0.0 0.9 0.0 9 5.2 17 159 1.2 14.5 1.2 11.4 9 4.5 21 87 1.5 7.9 1.5 6.2 9 5.4 120 0 8.6 0.0 8.6 0.0 9 3.4 9 350 0.6 31.8 0.6 25.0 9 5.1 72 126 5.1 11.5 5.1 9.0 9 4.7 91 210 6.5 19.1 6.5 15.0 9 4.2 65 0 4.6 0.0 4.6 0.0 9 5.6 31 378 2.2 34.4 2.2 27.0 9 4.1 55 189 3.9 17.2 3.9 13.5 9 4.6 76 287 5.4 26.1 5.4 20.5 9 5.0 36 168 2.6 15.3 2.6 12.0 9 4.9 95 119 6.8 10.8 6.8 8.5
MSc. Thesis 105 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Intensity Mean intensity Mean abundance Farm Age TL Tr. sp Gy.sp Tr.sp Gy. sp Tr. sp Gy. sp Sunfish 10 5.6 90.0 0.0 6.0 0.0 4.3 0.0 10 7.1 25.0 14.0 1.7 1.0 1.2 0.7 10 6.9 15.0 7.0 1.0 0.5 0.7 0.3 10 5.2 36.0 9.0 2.4 0.6 1.7 0.4 10 5.6 65.0 77.0 4.3 5.5 3.1 3.7 10 5.3 85.0 28.0 5.7 2.0 4.0 1.3 10 6.7 13.0 0.0 0.9 0.0 0.6 0.0 10 6.5 0.0 0.0 0.0 0.0 0.0 0.0 10 5.8 14.0 35.0 0.9 2.5 0.7 1.7 10 5.0 0.0 21.0 0.0 1.5 0.0 1.0 10 6.1 0.0 0.0 0.0 0.0 0.0 0.0 10 5.3 0.0 12.0 0.0 0.9 0.0 0.6 10 7.0 0.0 0.0 0.0 0.0 0.0 0.0 10 5.4 10.0 0.0 0.7 0.0 0.5 0.0 10 6.4 5.0 0.0 0.3 0.0 0.2 0.0 Ssebinyansi 10 8.4 0.0 119.0 0.0 8.5 0.0 5.7 10 8.7 25.0 91.0 1.7 6.5 1.2 4.3 10 9.7 25.0 147.0 1.7 10.5 1.2 7.0 10 10.5 70.0 77.0 4.7 5.5 3.3 3.7 10 10.8 35.0 126.0 2.3 9.0 1.7 6.0 10 9.5 18.0 91.0 1.2 6.5 0.9 4.3
MSc. Thesis 106 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Appendix 3: Trichodina measurements from Sunfish farm (– No measurement taken)
Sample Body Denticle ring Central Denticle Blade Ray Blade - ray Number of number diameter diameter portion length length length division Denticles 1 57.5 47.5 15.0 15.0 5.0 7.5 2.5 - 2 57.5 40.0 11.3 13.8 3.8 6.3 2.5 - 3 50.0 40.0 10.0 11.3 3.8 6.3 2.5 - 4 60.0 42.5 15.0 12.5 3.8 5.0 2.5 - 5 55.0 35.0 10.0 12.5 5.0 7.5 2.5 - 6 57.5 50.0 20.0 15.0 3.8 7.5 2.5 - 7 62.5 47.5 20.0 12.5 5.0 7.5 2.5 - 8 50.0 37.5 15.0 12.5 3.8 5.0 2.5 - 9 60.0 45.0 17.5 12.5 5.0 5.0 2.5 - 10 52.5 42.5 12.5 15.0 5.0 7.5 2.5 - 11 50.0 40.0 15.0 10.0 3.8 5.0 2.5 23 12 75.0 42.5 15.0 12.5 5.0 5.0 2.5 25 13 82.5 42.5 15.0 15.0 5.0 7.5 2.5 23 14 72.5 45.0 12.5 17.5 5.0 7.5 - 24 15 70.0 45.0 15.0 17.5 5.0 7.5 - 23 16 85.0 50.0 27.5 12.5 5.0 7.5 - 25 17 62.5 30.0 12.5 15.0 5.0 7.5 - 23 18 62.5 42.5 15.0 15.0 5.0 7.5 - 23 19 77.5 45.0 15.0 15.0 5.0 7.5 - 25 20 65.0 40.0 17.5 12.5 5.0 5.0 - 23 21 60.0 42.5 15.0 15.0 5.0 7.5 - 23 22 0.0 40.0 12.5 15.0 5.0 7.5 - 23 23 75.0 45.0 15.0 15.0 5.0 7.5 - 26 24 67.5 40.0 15.0 13.8 5.0 6.3 - 25 25 0.0 47.5 17.5 15.0 5.0 7.5 - 25 26 62.5 42.5 12.5 15.0 5.0 7.5 - 24 27 65.0 45.0 15.0 15.0 5.0 7.5 - 24 28 - 50.0 17.5 15.0 5.0 7.5 - 24 29 - 42.5 15.0 15.0 5.0 7.5 - 23 30 - 47.5 17.5 15.0 5.0 7.5 - 26 31 - 50.0 17.5 15.0 5.0 7.5 - 23 32 - 42.5 15.0 15.0 5.0 7.5 - 24 33 65.0 42.5 17.5 15.0 5.0 7.5 - 25 34 57.5 42.5 12.5 15.0 5.0 7.5 - 23 35 75.0 ------25 36 67.5 40.0 17.5 15.0 5.0 7.5 - 22 37 72.5 42.5 15.0 15.0 5.0 7.5 - 24 38 62.5 47.5 15.0 15.0 5.0 7.5 - 23 39 65.0 47.5 15.0 15.0 5.0 7.5 - 23 40 65.0 45.0 15.0 15.0 5.0 7.5 - 25 41 65.0 47.5 15.0 15.0 5.0 7.5 - 25 42 62.5 47.5 15.0 15.0 5.0 7.5 - 24 43 57.5 37.5 15.0 12.5 3.8 6.3 - 23 44 67.5 45.0 17.5 15.0 5.0 7.5 - 24 Mean 64.3 43.5 15.4 14.3 4.8 7.0 2.5 24
MSc. Thesis 107 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Appendix 4: Trichodina measurements from KRS farm (– No measurement taken)
Sample Body Denticle ring Central Denticle Blade Ray Blade - ray Number of Number diameter diameter portion length length length division Denticles 1 82.5 45.0 12.5 15.0 5.0 7.5 2.5 - 2 62.5 37.5 12.5 15.0 5.0 7.5 2.5 - 3 57.5 40.0 12.5 15.0 5.0 5.0 2.5 - 4 55.0 47.5 15.0 15.0 5.0 7.5 2.5 - 5 - 50.0 17.5 15.0 5.0 7.5 2.5 - 6 - 42.5 15.0 12.5 5.0 7.5 2.5 - 7 - 50.0 20.0 15.0 5.0 7.5 2.5 - 8 - 47.5 17.5 15.0 5.0 7.5 2.5 - 9 - 52.5 20.0 15.0 5.0 7.5 2.5 - 10 - 45.0 20.0 12.5 5.0 5.0 2.5 - 11 - 45.0 15.0 15.0 5.0 7.5 2.5 - 12 - 50.0 17.5 15.0 5.0 7.5 2.5 - 13 - 42.5 12.5 15.0 5.0 7.5 2.5 - 14 - 40.0 15.0 12.5 3.8 5.0 3.8 - 15 - 50.0 17.5 15.0 5.0 7.5 2.5 - 16 - 45.0 12.5 12.5 - - - - 17 - 42.5 12.5 15.0 - - - - 18 - 42.5 15.0 15.0 - - - - 19 57.5 45.0 12.5 15.0 - - - - 20 52.5 42.5 12.5 12.5 - - - - 21 65.0 45.0 15.0 15.0 - - - - 22 57.5 45.0 15.0 12.5 - - - - 23 57.5 45.0 15.0 12.5 - - - - 24 67.5 45.0 15.0 15.0 - - - - 25 67.5 50.0 15.0 15.0 - - - - 26 62.5 50.0 20.0 15.0 - - - - 27 65.0 47.5 15.0 15.0 - - - 24 28 62.5 37.5 10.0 15.0 5.0 7.5 - 23 29 55.0 32.5 12.5 15.0 3.8 3.8 - 23 30 - 37.5 12.5 12.5 5.0 5.0 - 24 31 75.0 50.0 17.5 15.0 5.0 7.5 - 24 32 - 42.5 15.0 11.3 5.0 6.3 - 24 33 57.5 40.0 10.0 15.0 5.0 7.5 - 22 34 62.5 45.0 15.0 15.0 5.0 7.5 - 23 35 65.0 42.5 12.5 12.5 5.0 6.3 - 24 36 57.5 37.5 12.5 11.3 5.0 6.3 - 23 37 60.0 42.5 12.5 15.0 5.0 7.5 - 24 Mean 62.1 44.3 14.7 14.2 4.9 7.0 2.6 24
MSc. Thesis 108 UNESCO-IHE, DELFT
Fish parasites in Uganda Akoll P.
Appendix 5: Trichodina measurements from Ssebinyansi farm (– No measurement taken)
Sumple Body Denticle ring Central Denticle Blade Ray Number of Number diameter diameter portion length length length Denticles 1 57.5 47.5 17.5 10.0 2.5 5.0 - 2 60.0 37.5 12.5 12.5 3.8 6.3 - 3 60.0 47.5 17.5 10.0 2.5 5.0 - 4 75.0 45.0 17.5 15.0 5.0 7.5 - 5 67.5 45.0 12.5 15.0 5.0 7.5 25 6 57.5 35.0 10.0 12.5 5.0 5.0 24 7 67.5 55.0 17.5 17.5 7.5 7.5 23 8 65.0 37.5 15.0 15.0 5.0 5.0 26 9 65.0 47.5 17.5 15.0 5.0 5.0 22 10 70.0 47.5 20.0 15.0 5.0 7.5 28 11 62.5 35.0 7.5 12.5 3.8 6.3 23 12 75.0 50.0 20.0 15.0 5.0 7.5 23 13 62.5 45.0 15.0 15.0 5.0 7.5 24 14 62.5 47.5 12.5 17.5 5.0 10.0 23 15 72.5 42.5 12.5 15.0 5.0 7.5 25 16 62.5 42.5 12.5 15.0 5.0 7.5 24 17 60.0 42.5 15.0 15.0 5.0 7.5 28 18 62.5 40.0 15.0 12.5 5.0 5.0 24 19 62.5 40.0 12.5 12.5 5.0 5.0 26 20 70.0 45.0 15.0 15.0 5.0 7.5 24 21 65.0 47.5 17.5 15.0 5.0 5.0 23 22 67.5 37.5 12.5 12.5 5.0 6.3 26 23 72.5 50.0 17.5 15.0 5.0 7.5 23 24 75.0 47.5 15.0 15.0 5.0 7.5 25 25 65.0 37.5 15.0 12.5 3.8 6.3 24 26 60.0 35.0 11.3 13.8 3.8 5.0 24 27 65.0 52.5 17.5 20.0 5.0 7.5 24 28 72.5 45.0 15.0 15.0 5.0 7.5 26 29 80.0 42.5 12.5 15.0 5.0 7.5 26 30 - 35.0 10.0 12.5 5.0 6.3 25 31 - 47.5 15.0 17.5 5.0 7.5 24 32 - 47.5 17.5 15.0 5.0 7.5 25 33 - 40.0 12.5 12.5 5.0 5.0 24 34 - 42.5 17.5 15.0 5.0 7.5 24 35 - 50.0 15.0 17.5 5.0 7.5 24 36 - 37.5 17.5 10.0 3.8 5.0 24 Mean 66.2 44 15 14 5 7 24
MSc. Thesis 109 UNESCO-IHE, DELFT