Abstract
The present thesis concerns the study of myxosporea infections in Mediterranean maricultured species, with emphasis in the myxosporea infection caused by Enteromyxum leei Diamant, Lom & Dyková 1994. The objectives of the present thesis were: - The estimation of the myxosporea infections in greek aquaculture, in Sparidae family cultured fish, which are the largest part of Greek mariculture and the role of wild fish as myxosporea reservoirs in cultured fish, as well. - The study of E. leei transmission and immune response in receptor fish subjected to experimental infection of E. leei. - The estimation of the efficacy and the toxicity of orally administrated anti-coccidial drugs, commonly used for other cultured animals, for innovative treatments of E. leei in infection in sharpsnout seabream (Diplodus puntazzo Cetti 1777). The thesis is divided in four Chapters, Introduction, Material & Methods, Results and Discussion. In the Introduction, the relevant literature is reviewed. The Chapte ...
The present thesis concerns the study of myxosporea infections in Mediterranean maricultured species, with emphasis in the myxosporea infection caused by Enteromyxum leei Diamant, Lom & Dyková 1994. The objectives of the present thesis were: - The estimation of the myxosporea infections in greek aquaculture, in Sparidae family cultured fish, which are the largest part of Greek mariculture and the role of wild fish as myxosporea reservoirs in cultured fish, as well. - The study of E. leei transmission and immune response in receptor fish subjected to experimental infection of E. leei. - The estimation of the efficacy and the toxicity of orally administrated anti-coccidial drugs, commonly used for other cultured animals, for innovative treatments of E. leei in infection in sharpsnout seabream (Diplodus puntazzo Cetti 1777). The thesis is divided in four Chapters, Introduction, Material & Methods, Results and Discussion. In the Introduction, the relevant literature is reviewed. The Chapter is subdivided into four Parts. In Part A, the World, European and Greek aquaculture situation is described. In Part B, the pathological problems that are observed in aquaculture are described. In Part C, the literature of the most important myxosporea infections observed in Mediterranean aquaculture is reviewed. In Part D, the literature associated with the immune response of fish against infections and in particular protozoan parasites is reviewed. In Material and Methods, the protocol used in each experiment and the laboratory techniques used for the detection of parasites and for definition of immunological parameters are described. The statistical packages used for the statistical analysis of the results are also described. In the Results, in subchapter 3.1, an estimation of the seasonality and prevalence caused by myxosporea infections in wild and cultured fish species in Greece is described. E. leei has been observed in both cultured and wild fish neighboring cages of cultured fish. Sharpsnout seabream proved to be the main host of this parasite where the highest prevalence and mortality rate were observed, while white seabream (Diplodus sargus Cetti 1777) proved to be also a significant host. E. leei was also observed in wild gilthead seabream (Sparus aurata Linnaeus 1758) and mullets (Mugil cephalus Linnaeus 1758) neighboring cultured fish. The main target organ of all infected fish was the intestine. In cases where high prevalence was observed in cultured sharpsnout seabream and white seabream, the gallbladder and the gills were also target organs. This is the first time that renal tissue is found as a target organ of this parasite; E. leei mature spores being observed in the kidney of heavily infected cultured sharpsnout seabream. Polysporoplasma sparis Sitjà-Bobadilla & Álvarez-Pellitero 1995, has been observed in both cultured and wild fish neighboring cages of cultured fish. Gilthhead seabream proved to be the main host of this parasite, but it was also observed in cultured sharpsnout seabream. This is the first time that P. sparis is observed in wild fish such as: gilthead seabream and mullets and in one case in common Pandora (Pagellus erythrinus Linnaeus 1758), neighboring cages of cultured fish. The main target organ in all infected fish was the kidney. The parasite was found also in the renal interstitial tissue while in a few cases P. sparis spores were observed in the melanomacrophage centers of the kidney demarcated by two layers of connective tissue. Myxobolus sp. has been observed in both cultured and wild fish neighboring cages of cultured fish. This parasite was observed in cultured sharpsnout seabream and for the first time, internationally, in cultured white seabream. Also this is the first time that this parasite was observed in wild mullets in Greece, while it was also observed in wild white seabream. The main target organ in cultured infected fish (white seabream and sharpsnout seabream) and in wild white seabream, was the renal interstitial connective tissue and the melanomacrophage centres of the kidney where trophozoites and mature spores were observed. In wild mullet, mature spores of the parasite were observed only in the gut and the gallbladder; this is found for the first time internationally. All the developmental stages of Ceratomyxa diplodae Lubat, Radujkovic, Marques & Bouix 1989 and Ceratomyxa sparusaurati Sitjà-Bobadilla, Palenzuela & Álvarez-Pellitero 1995 were observed in sharpsnout seabream and gilthead seabream respectively, without causing any mortality. Also, unidentified species of the genus Ceratomyxa were mentioned for the first time in wild fish (gilthead seabream and bogue, Boops boops Linnaeus 1758) internationally. The main target organ of the parasite in both cultured and wild fish was the gallbladder. The parasite Henneguya sp. was observed in wild gilthead seabream and for the first time in cultured sharpsnout seabream. The main target organ of this parasite was the gallbladder. Finally, Kudoa sp. was observed for the first time internationally in the muscle of cultured white seabream. Myxosporean parasites outbreaks were observed in the seasons of highest temperature. Summer, in particular, was the season when the more statistically significant prevalence was observed in all studied fish species (wild and cultured). Also, an interaction in transmission of myxosporea parasites between cultured and wild fish was documented. In most cases, wild fish proved to be the reservoirs of myxosporea parasites transmitting the parasites in the cultured fish. This conclusion was based on the fact that simultaneous infection in cultured and wild fish occurred in most cases and on the fact that the prevalence was statistically higher in wild than in cultured fish. In the Results, in subchapter 3.2, the study of the life cycle of myxosporea Enteromyxum leei in sharpsnout seabream is described. Direct transmission of the infection was achieved by cohabitation of infected and healthy sharpsnout seabream for the first time. The progress of the infection was monitored by the detection of different developmental stages of the parasite in the intestine. E. leei infected fish were firstly observed on day 5 (prevalence of 20%). The total prevalence reached 100% on day 19 and was maintained at this high level until the end of the experimental period (day 55). In addition, the initial intensity of the different parasite stages was low, but on day 26 the intensity of trophozoites was suddenly increased, followed by the increased intensity of sporoblasts and mature spores. Also, two innate immune factors were examined in order to determine the influence of the infection on the defense immunity of the host. Particularly, ΝΟ secretion from blood monocytes was gradually increased after their stimulation from parasite antigens. In the present study, lysozyme levels were not detected in blood serum of infected fish despite the fact that in natural infections of the same fish with other myxosporea parasites a possible influence of myxosporea infection in lysozyme production had been found, a fact that was related to the infection’s intensity level. Also, the experimental transmission of E. leei in recovered sharpsnout seabream previously treated with Salinomycin and Amprolium (AS) proved to be unsuccessful, suggesting a possible development of protective immunity against this parasite. In the Results, in subchapter 3.3, different drugs, effective against other spore-forming parasites in other animals were tested in order to estimate their anti-myxosporean effect in naturally infected (prevalence 100%) sharpsnout sea bream (with E. leei). Two medicated diets were applied: a) Amprolium and Salinomycin (AS) and b) Fumagillin (F). Significant drug effect on reduction of prevalence, intensity of all developmental myxosporea stages and mortality were observed at the end of both treatments in comparison to untreated fish. The effect was most prominent in the case of Amprolium and Salinomycin combination, exhibiting a significant reduction in intensity, prevalence and mortality rate in treated fish without any histopathological evidence of toxic side effects or reduction on growth. Furthermore, sporoblasts and mature spores with distorted structure were observed in both tested treatments, but their prevalence was higher in the case of Amprolium and Salinomycin medication than in Fumagillin treated fish proving evidence of the effectiveness of the drug directly on the parasite. This data suggests that Amprolium and Salinomycin could be a promising treatment for E. leei infections in intensively cultured warm-water fish. From the findings of the present study the following conclusions can be drawn: - Myxosporea are widely distributed parasites in greek aquaculture causing serious economic effects. Cultured and wild fish are the hosts of these parasites and summer is the season that favours the myxosporea’s elevation. Also, there is an interaction in myxosporea infection between cultured and wild fish and wild fish seems to be the parasite reservoir in cultured fish. - The transmission of E. leei in sharpsnout seabream is direct and the parasite transmission is achieved through waterborne contamination. The intestine is the first ‘’target’’ organ of this parasite. The influence of the infection on the defense immunity of the host, was proved by ΝΟ secretion from blood monocytes in infected fish. During the 55day experiment, the parasitosis is still in a primary phase, which is proved by the presence of the parasite in the intestine and not in other ‘’target’’ organs and the deficiency of mortalities. - The Amprolium and Salinomycin combination proved to be a well promising, cheap and safe combination treatment for E. leei reducing the prevalence and intensity level of parasitosis as well as the mortality rate.
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