<I>Gyrodactylus salaris</I> Malmberg, 1957, a monogenean parasite of salmonid fish, has caused the death of up to 95% of salmon parr in 37 Norwegian rivers. In order to prevent further spread of this parasite, a reliable method of identifying <I>G. salaris</I> and distinguishing it from other closely related species is required. This study, the first investigation of <I>Gyrodactylus</I> genetics, has demonstrated that DNA technology can provide methods of gyrodactylid species identification suitable for routine use. DNA was extracted from <I>G. salaris</I> and two other species common on salmonid fish; <I>G. derjavini</I> and <I>G. truttae</I>. The small subunit ribosomal RNA (srRNA) gene was amplified from this DNA by polymerase chain reaction (PCR). The complete nucleotide sequence of the srRNA gene from <I>G. salaris</I> was determined. This was used to predict a secondary structure for gyrodactylid srRNA and to construct molecular phylogenies of platyhelminths including <I>Gyrodactylus. </I>Fragments of the srRNA gene from each species were compared by Denaturing Gradient Gel Electrophoresis. Mobility differences in <I>G. truttae</I> fragments were found and one fragment showed variation between and within species. The V4 region of srRNA was amplified from single specimens of gyrodactylids using a combined lysis and PCR reaction and sequenced. Examination of these sequences enabled prediction of Restriction Fragment Length Polymorphisms (RFLPs) between species and the design of oligonucleotide probes specific for each species. Digestion of the srRNA gene V4 region with two restriction enzymes produced restriction fragment polymorphisms that can be used to discriminate between <I>G. salaris</I>, <I>G. derjavini</I> and <I>G. truttae</I>. The digoxigenin labelled oligonucleotides GsV4, GdV4 and GdV4 are specific for <I>G. salaris, G. derjavini</I> and <I>G. truttae</I> respectively and can detect PCR amplified DNA from single specimens in dot blots. Both RFLP and probe methods of identifying <I>Gyrodactylus </I>species are suitable for use in diagnostic laboratories.

No description available.

No description available.

No description available.

The life-cycle of Rhipidocotyle campanula (Dujardin, 1845) has been experimentally demonstrated and the species identity confirmed. Sporocysts were recovered from digestive glands of the freshwater-mussel Anodonta anatina, and in heavily infected 'hosts from the reproductive, system. Both cercariae and glochidia are released simultaneously in mussels where the reproductive system is partly invaded by sporocyst tubules. The development of the cercariae in the sporocyst tubules has been studied briefly fusing histological and histochemical methods. The liberation of cercariae varied- between >1000/mussel/day to none, and is intermittent. Behaviour of the cercariae including swimming, response to light and gravity, survival and entry, into the secondary host, is described. The morphology of the cercaria has been studied in detail using electron microscopy, histochemistry and histological methods and its significance analysed in relation to free-living existence. This is the first attempt to study the cercaria of R. campanula in any detail. Cercariae enter the secondary host passively and encyst in the subcutaneous fatty tissue beneath the lining of the pharynx, and in some cases in the gill arches. This is the only species of bucephalids where cercariae enter the secondary host passively. Encysted metacercariae attain maximum development after 5-6 weeks and survive nearly 200 days, but spontaneous excystation takes place only in cysts 80-90 days old. Adult flukes were recovered from the posterior intestine and rectum of the perch (Perca fluviatilis) six weeks after feeding fully developed metacercariae. Spermatogenesis and oogenesis were studied in the adults until egg formation. Miracidial development could not be observed. External morphology using SEM of cercaria, metacercaria and adult has been studied and compared. This is the first report of SEM study of these stages of R. campanula. A brief review of the literature is given and the problems of systematics and taxonomy of the family Bucephalidae the taxonomic position of R. campanula are discussed.Tabular summaries are given for the previous life-cycle studies of bucephalids, synopses of bucephalid trematodes and all previous reports of bucephalid cercariae.

Ph.D. / The Branchiura is a group of ectoparasitic crustaceans which infect mostly fishes. The subclass contains four genera including Argulus, Chonopeltis, Dolops and Dipteropeltis. Despite their reputation as pests our understanding of their biology is meager. In the first part of the study the anatomy and physiology is described. The species studied include Argulus japonicus, Dolops ranarum and Chonopeltis australis. Live specimens of A. japonicus, D. ranarum and Chonopeltis australis were collected, fixed in Todd‟s fixative and processed for transmission electron microscopy. The results gleaned from A. japonicus and D. ranarum indicated that the ultrastructure and physiology of the cells were similar to the digestive epithelia of free-living copepods. The anterior midgut consists of Resorptive cells (R cells) and the enteral diverticula consist of two cell types including R and Fibril cells (F). The R cells contain lipid droplets and F cells contain rough endoplasmic reticulum indicating that the absorption of nutrients and digestion occurs in the anterior midgut and enteral diverticula. The lipid droplets in both species are large in size and number. Blister cells/B cells and R‟ (apostrophe) cells occur in the posterior midgut and are involved in the processing of digestive waste. The results from C. australis were different from A. japonicus and D. ranarum and reveal the possible presence of F/R cells with multiple functions of both absorption and enzyme secretion. The cells contain few lipid droplets and instead contain many hexagonal-shaped crystalline structures that are interpreted to be proteinaceous. The differences in diet are thought to be influenced by morphological differences in the feeding appendages. The second part of the study involved the supraoesophageal ganglion or brain of C. australis. This genus is unique among the Crustacea since the antennules are absent. This trait poses implications regarding the brain morphology since particular parts of the brain impart nerves that innervate certain appendages in arthropods. Specifically the deutocerebrum or midbrain innervates the antennules. A study was conducted to discover whether the antennular nerves were absent. Specimens of C. australis were fixed in AFA, sectioned and stained with Heidenhain‟s AZAN. Results showed that the antennular nerves were absent and the deutocerebrum was smaller in comparison to the other brain segments indicating degeneration. Moreover, C. australis possesses a reduced number of sensory sensilla on its external surface compared to related genera. The loss of antennules does not affect its sensory capabilities in host searching. However, the reduced numbers of sensilla and its feeble swimming capabilities are more likely to reduce its efficiency in host searching.

A two-year study of the actinosporean fauna of oligochaetes was conducted at an Atlantic salmon fish farm located at the extreme north of Scotland. The actinosporean fauna and their morphological characteristics, the ultrastructural development of four different actinosporean collective groups, the epidemiology of all actinosporean types identified,the complete life cycle of Sphaerospora truttae, the circadian and seasonal spore release patterns of actinosporean types and the myxospores of S. truttae, the viability of actinosporeans and their responses to fish mucus were determined. Twenty one actinosporean types belonging to seven collective groups: Synactinomyxon (3 types), Aurantiactinomyxon (4 types), Echinactinomyxon (5 types), Raabeia (6 types), Neoactinomyxum (l type), Triactinomyxon (1 type) and Siedleckiella (1 type) are described. Six types were identified to previously described forms; Synactinomyxon "A" of McGeorge et al. (1997); Synactinomyxon tubificis Stole, 1899, S. longicauda Marques, 1984, Aurantiactinomyxon-type of McGeorge et al. (1997), Echinactinomyxon radiatum Janiszewska, 1957, Raabeia-type of McGeorge et ai. (1997). The remainder appeared to be new types of the collective groups. Temperature was found to have a significant effect on the spore morphology and caused statistically important differences in the spore dimensions, especially on the caudal processes. Synactinomyxon-type 1, Aurantiactinomyxon-type3, Echinactinomyxon-type5 and Raabeia-type4 were studied at the TEM level to determine the developmental stages of each type. All actinosporean types studied had uninucleate cells as the earliest stage of development. Formation of a subsequent binucleate cell stage was either due to the division of the nucleus in a uninucleate cell or the plasmogamy of two uninucleate cells. The earliest pansporocyst formation seen was two outer somatic cells surrounding two inner generative alpha and beta cells in all actinosporean types studied. However, the formation of an early pansporocyst followed a four-nuclei stage only in Raabeia. Subsequently, the number of somatic and generative cells increased as a result of mitotic divisions and reached 8 alpha and 8 beta cells at the end of the division stages. Echinactinomyxon had only four somatic cells in pansporocyst, whilst Synactinomyxon, Aurantiactinomyxon and Raabeia had eight. Following the copulation of each pair of alpha and beta cells, 8 zygotes were formed. Then, two mitotic divisions of each zygote resulted in a four-cell stage of each sporoblast. Valvogenesis and capsulogenesis was followed by the formation of 8 mature spores inside each pansporocyst. Over the two year sampling programme the overall infection prevalence of oligochaetes with actinosporeans was 2.9%. The infection prevalence was higher in the first year (3.3%) than the second year (2.3%). The infection prevalences of individual types were between 0.001% and 0.9%. Summer was the preferred season of spore release (4.1%), followed by autumn (2.9%) , spring (2.8%) and winter (1.6%), Some parasites such as Echinactinomyxon-typel released spores throughout the study period, whilst Synactinomyxon-type2 was recorded only in summer. There was also a positive relationship between the number of actinosporean types released and water temperature. A one year sampling programme also indicated that Sphaerospora truttae had two distinct life cycle phases, extrasporogonic and sporogonic, in the fish. Extrasporogonic stages were first detected at the beginning of July 1996 with a prevalence of 50% and were seen over an 8-10 week period. Sporogonic stages first became detectable in the kidney tubules at the beginning of September 1996. As well as sporogonic stages, many developing pseudoplasmodia were also observed at this time. Pseudoplasmodia were always present along with mature spores. The infection prevalence stayed above 80% throughout the period of infection. Experimental infections showed that Echinactinomyxon-type5, was the alternate life cycle stage of S. truttae in the oligochaete Lumbriculus variegatus. The time taken from the exposure of Atlantic salmon to Echinactinomyxon-type5 spores to formation of mature Sphaerospora truttae spores was 4.5 months (138 days). However, infections of Atlantic salmon with presporogonic and immature spores of S. truttae were first seen at 3.5 months post-exposure (110 days). In addition to S. truttae, the life cycle of Chloromyxum truttae was also completed at 4.5 months (138 days) post - exposure at 12-16°C using Aurantiactinomyxon-type4 spores released from Tubifex tubifex. Worms infected with Synactinomyxon-type 1, Aurantiactinomyxon-type I, Echinactinomyxon-type1 and type5, Raabeia-type4 and Neoactinomyxum-type showed inconsistent spore release patterns over five subsequent days at ambient temperatures. Up to 5000 spores an each day were released from infected worms with the exception of Echinactinomyxon-type5 which released up to 80,000 spores per day. Experimentally there was a positive relationship between the numbers of spores shed and water temperature. The spore release of worms infected with Synactinomyxon-type I, Aurantiactinomyxon-type 1, Echinactinomyxon-type I, Raabeia-type4 and Neoactinomyxum-type spores were also studied at 3 h intervals and showed that peak release occurred between 22.00 and 01.00 h. Studies on the spore release patterns of Sphaerospora truttae myxospores from Atlantic salmon showed that mature spores were first released at the end of November, peaked around April and then decreased sharply. Number of mature spores present in the kidney of the fish showed a similar pattern of abundance. Polar filaments of Echinactinomyxon-type I, Raabeia-type4 and Aurantiactinomyxon-type I spores discharged in response to mucus from Atlantic salmon, brown trout, 3-spined stickleback and common carp. However, the response to the mucus from each fish species was different. In each case majority of discharges occurred within the first 5 min of exposure to mucus although there were further discharges up to lh. The viability of Synactinomyxon-type I, Echinactinomyxon-type I, Raabeia-type4, Aurantiactinomyxon-typel and Neoactinomyxum-type spores had a negative correlation with increasing temperature. In general, the spores remained viable for 6-7 days at 4°C, 4-5 days at 13°C and 4 days 22°C.

M.Sc. / Please refer to full text to view abstract / Prof. A. Avenant-Oldewage

M.Sc. / Recent years have seen a remarkable increase in scientific research and studies conducted regarding the unique group of parasitic isopods known as gnathiids. The taxonomy of gnathiids are based solely on the morphological characteristics of the free-living adult males, which means that the female and larval life stages are often not collected and described with the males, and when collected it is seldom possible to even identify these stages when not in the company of there adult males. Numerous studies have been conducted in Australia regarding cleaning behaviour of coral reef fishes, where these gnathiid larvae are extremely abundant and are often collected. These types of studies as well as others are increasing all around the world, and because of the fact that the larvae cannot be easily identified, the results and data collected from these various studies are often misinterpreted or inconclusive and makes no sense. The fact that gnathiid larvae may serve as possible vectors for certain fish blood parasites and have various physiological and ecological effects on their fish hosts, which may lead to mortality, can have detrimental economical effects on the fisheries industry. This study was thus conducted to improve the identification methods that are currently applied to the larval life stages of gnathiid isopods through various means including increasing the number of detailed descriptions available, constructing a taxonomic key and lastly conducting a basic phylogenetic analysis of a group of known gnathiid larvae in order to aid researchers with future identifications. It was hypothesised that firstly, although there are some gnathiid descriptions from coral reefs, mostly of adult males, it is likely that there are still a great number of unknown and not yet described gnathiid species, because coral reefs are one of the most highly biodiverse marine ecosystems. Secondly, that the distribution of coral reef gnathiids is much wider than currently known or documented and lastly, that it might be possible to identify gnathiid larvae species in the absence of adult males, through the use of detailed taxonomic descriptions of these larval stages and there live colouration patterns, and to then construct a taxonomic key to aid researchers with future identifications.

M.Sc. / The Bushveld smallscale yellowfish, Labeobarbus polylepis (Boulenger, 1907), is an ecologically, socially and economically important species. These indigenous freshwater fish are found above an altitude of 600m, and occur in the Inkomati and Phongolo River Systems and the southern tributaries of the Limpopo River System. In the past, it was suspected that morphological differences exist between the different L. polylepis populations, due to the occurrence of the rubberlip formation of individuals from the Elands River. Specimens of five L. polylepis populations were collected from the Phongolo, Assegaai, Elands and Komati rivers and Ngodwana Dam, Mpumalanga, South Africa. A L. natalensis population was collected from the Umvoti River and used as an out-group. Nine meristic counts and 46 morphometric measurements were taken. The measurements were changed into percentage ratios based on the fork length of each individual. The data was statically analysed and includes Multidimensional scaling techniques (MSD’s) and Principle Component Analysis (PCA’s). Statistical analysis split the five populations into three groups. The one group consists of fish from the Phongolo and Assegaai rivers, the second group consists of fish from the Elands River and Ngodwana Dam and the third group is mainly Komati River fish. The third group is the only group that does not overlap with any other group. There were morphological differences between the groups, but they were not significant. The L. natalensis population is morphologically similar to L. polylepis populations obtained from the Phongolo and Assegaai rivers. Twenty enzyme coding loci in two L. polylepis populations from the Phongolo and Elands rivers were analysed by horizontal starch gel-electrophoresis. Electrophoretic analysis of heart, muscle and liver tissue samples revealed genetic variation at 15% (Elands River) and 35% (Phongolo River) of the protein coding loci studied. Average heterozygosity values based on Hardy-Weinberg expectation were 0.019 (Elands River) and 0.059 (Phongolo River), with a genetic distance value of 0.004 between these populations.