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Item Open Access Distribution and acaricide resistance status of the theileriosis vector, Rhipicephalus appendiculatus Neumann (Acarina:Ixodidae) in response to changing climate in Zimbabwe(University of the Free State, 2023) Nemaungwe, Tinotenda Mercy; van Dalen, Elsie Maria Susanna Petronella; Chidawanyika, FrankTick control is a key aspect of managing Tick-Borne Diseases (TBDs) with chemical control being the widely applied method. However, continued application of chemicals can lead to the development of resistance. A recent increase in theileriosis in Zimbabwe coupled with indications of shifts in outbreaks from seasonal to spontaneous, raised possibilities of acaricide resistance by the vector, Rhipicephalus appendiculatus. Factors behind changes in the spatio-temporal distribution of the tick and the disease it transmits are poorly understood. This study evaluated the distribution of the theileriosis vector, R. appendiculatus, its response to frequently applied acaricides and the implications of current management practices on acaricide resistance development in Zimbabwe. Initially, the thermal tolerance of larvae, nymphs and adult R. appendiculatus ticks were assessed followed by modeling the current and future distribution of suitable habitats using MaxEnt. Bioassays were then conducted to determine the tick’s resistance status to frequently applied acaricides namely amitraz and deltamethrin. Furthermore, a survey was undertaken to understand the relation between tick management strategies and acaricide resistance development in ticks of economic importance.Results indicated that suitable habitats for R. appendiculatus may expand into previously non-endemic areas between 2040 and 2060 if temperatures rise by 2 °C. This expansion may be aided by the ability of the species to tolerate higher temperatures during assays than those recorded in the environment. Results further showed that R. appendiculatus populations are still highly susceptible to amitraz and deltamethrin (Resistance ratios<4). In contrast, the indicator species R. decoloratus which was used for comparison had 50% of populations moderately resistant to amitraz and two dip tanks resistant to deltamethrin (4Item Open Access Enzootic geophagy by elephants (Loxodonta Africana) in relation to geochemical composition of mineral licks in Addo Elephant National Park, South Africa(University of the Free State, 2022) Darker, Kristen Nadine; Butler, H. J. B.Geophagy, the deliberate ingestion of soil, is a common occurrence amongst various animal species including mammalian herbivores such as elephants. Despite the documented instances of soil-eating, and several nonexclusive hypotheses, the real motivation behind the phenomenon remains controversial. In this study, six camera traps were set up throughout Addo Elephant National Park, South Africa at selected geophagy sites which captured visitation frequency as well as the demographic trend of elephant groups during site visits from April 2019 to May 2020. The geochemical and mineralogical composition of soils at these selected geophagy sites were analysed using X-ray diffractometry (XRD) and X-ray fluorescence spectrometry (XRF). Furthermore, the spatial distribution of five collared elephants (three matriarchs and two males) in relation to the six geophagy sites were investigated using kernel density estimations (KDE). Females had larger home ranges that incorporated more geophagy sites than males. Visitation frequency to geophagy sites were estimated using 500 m buffer zones from the centre of each site. Individuals visited at least three or more geophagy sites throughout the study period. Overall, essential elements Na, Ca and Mn were identified as main drivers for geophagic behaviour in the elephants of AENP. These essential elements (Na, Ca and Mn) are important for certain physiological demands such as bone and tusk growth in elephants and reproductive (pregnancy and lactation) demands in females. Geophagy is considered to be a contributing factor of movement patterns and area utilisation and may have important implications for conservation and management.Item Open Access Development of species-specific polymerase chain reaction (PCR), real-time PCR and loop mediated isothermal amplification (LAMP) assays for detection of anaplasma marginale strains in South Africa(University of the Free State, 2012-12) Khumalo, Zamantungwa Thobeka Happiness; Mtshali, M. S.; Thekisoe, O. M. M.Anaplasma marginale is a virulent intra-erythrocytic pathogen that causes bovine anaplasmosis, its closely related species, Anaplasma centrale causes mild sickness. The pathogen is transmitted biologically by tick vectors and mechanically through blood contaminated fomites. It has a worldwide distribution extending from tropical to subtropical regions in correlation with the vector distribution. Bovine anaplasmosis is often characterised by progressive anaemia, jaundice, decreased milk production, abortion and a sudden death. The commonly used method for the diagnosis of A. marginale of infected cattle in South Africa is microscopic examination of Giemsa stained blood smears anddetection of antibodies from serum using cELISA. However the diagnostic methods have limitations in cases of low parasitemia and in carrier cattle (microscopy) and they fail to differentiate closely related Anaplasma spp due to antigenic similarity (serology), the detection limitations of the diagnostic methods influenced the aim of this study which is to develop molecular species -specific assays for the detection of A. marginale strains in South Africa, specifically Including conventional polymerase chains reaction, real-time polymerase chain reaction and loop-mediated isothermal amplification assay. Chapter one of this study discusses bovine anaplasmosis and its causative agent A. marginale, the diversity of the strains transmission, distribution, clinical signs, treatment and economic importance of the disease. The first objective of this study was to develop a species specific conventional PCR for detection of A. marginale in cattle in South African regions based on msp1b gene. The conventional PCR primers were designed through visual inspection and were named F3 and 83 primers. In the specificity test, the primers were specific whereby the amplified only A. marginale DNA and did not amplify control DNA's: A. centrale, Babesia bovis, B. bigemina and Ehrlihia rumunantium. The sensitivity of the conventional PCR primers was examined using a 10 ng/ul DNA and the detection limit of the assay was 0.01 ng/ul, The assay was validated on field samples to confirm the infection of the cattle with A. marginale, out of 144 samples, (60%) infection rate was obtained with the newly developed onventional PCR, the homogeneity of the sequences were confirmed with the GenBank, the maximum similarity varied from 94 - 100%. The second objective of this study was to develop a species-specific real-time PCR for detection of A. marginale in cattle in South African regions based on msp1b gene. The real-time PCR primers and probes were designed using Genescript program, one set of primer (Prf 2, PrR2, and Pr82) was chosen to carry out the study as it showed high sensitivity with the detection limit of 0.001 ng/ul .The specific and sensitive TaqMan based real-time PCR was successfully developed for the of A. marginale infections in South Africa. Validation of the assay on field samples showed that the rate of infection was 74% in different sampled provinces of South Africa. The third objective of this study was to develop loop-mediated isothermal amplification for the detection of A. marginale in South African regions based on msp1b gene. The LAMP primers were designed using primer Explorer version 4, the LAMP primers were named LAF3, LA-83, LA-FIP, LA-BIP,LA-LF and LA-LB. The LAMP assay showed positive results with specific amplification, but as far as the validation of the assay false positive results were obtained, troubleshooting involved the addition of additives, changing of primer purification and manufacturers, however the results were not consistent, false positive results were obtained, speculations were that it could be possible contamination of the laboratory resulting in the amplification of control DNA and distilled water. The first three objectives of this study were achieved. The newly developed assays were further compared for specificity, sensitivity and detection performance on field derived samples. The developed assays are specific and sensitive; they form a good tool of diagnosis of bovine anaplasmosis, with each assay having its own unique characteristic over the other, they are sensitive giving a correct determination of the infection status, aiding in compiling of epidemiological information. These assays will aid in understanding the major constraint to develop control measures due to the genetic diversity of A. marginale, and will also help in constructing of phylogenetic tree between strains from South Africa and other countries.Item Open Access A survey of parasitic helminths in horses from communal farms in the northeastern Free State Province of South Africa(University of the Free State, 2004) Masangane, Flatha Euginia Sarah; Mbati, P. A.; Malatje, S. M.A survey of helminthes occurring in horses at Kestell in the northeastern region of the Free State Province was conducted for a period of ten months (June 2002-March 2003). The aim was to determine the factors influencing helminth parasites infections and to record the ticks of veterinary importance in horses in the northeastern Free State. Blood, fecal samples and ticks were collected from 24 horses on the same farms every month for a period of ten months. The age of horses was between five months and seven years. Collection was done from both males and females. Fecal samples were collected from horses for identification of helminthes, blood was collected to perform packed cell volume and ticks were collected for identification and recordings. McMaster, coprological and Visser sieve techniques were used for egg counts. A total of three helminth species and a protozoan were recovered. Dominating species were strongyle; with egg per gram (EPG) values ranging between 0 and 4400. An R test showed that there was significant difference between age (p = 0.0218), seasonality (p< 0.0001) and physical conditions (p < 0.0001) in the prevalence of strongyle. This means that the younger horses had higher infestation levels; colder months had lower infestation rates and the better the physical condition of the horse the lower the infestation. It also showed that there is a strong linear relationship between packed cell volume (PCV) and (r = - 0.23465; p- value= 0.0004). Larval identification was done through preparation of fecal cultures. Small strongyle larvae made up more than 80% of larval cultures in all samples cultured. Blood samples were collected to conduct a PCV test. The readings were found to be normal ranging between 24 and 44. Rhipicephalus evertsi evertsi was were the only two tick species collected from the horses. A questionnaire survey was concurrently carried out to determine the influence of socio-economic factors on the management of horses, which may favour helminth infestation in these species. From the owners interviewed 35.8% were pensioners and unemployed. A total of 67% of horse owners utilize their horses everyday for transport, but the frequency range from once or twice a week. Only 39% of owners use herbal or natural product or drugs to treat their animals, whereas 23% use commercial products. Horses belonging to 90% of the owners were given lucem hay all year round, a few owners fed their horses on mealies and other utilized the veld to feed their horses. The disease conditions reported were worms, ticks, eye lesion, hoof problems, skin problem and infectious diseases. 45% of owners asked their friends or neighbours who had knowledge of horses for advice when their animals became sick and 30% treat the animals themselves. 19% reported that they did nothing and 6% took their animals to the state veterinarian. Approximately 3% made use of animal health inspectors, private veterinarians and traditional healers. Many owners (87%) allowed their horses to roam free during the day and about 84% of owners kept their horses in kraals or small camps during the night. The information gathered from this study provides the first documentation study on helminthes of veterinary importance in horses in the northeastern Free State.Item Open Access A sero-epidemiological survey of parasites in cattle in the north eastern Free State, South Africa(University of the Free State, 2001) Mtshali, Moses Sibusiso; Mbati, Peter Amunga; De Waal, TheoGlobal economic losses due to parasitic infections in livestock are enonnous. There is little or no infonnation on parasites of veterinary importance afflicting livestock in the north eastern Free State, South Africa. A survey to determine the incidence of parasites in cattle (n=386) was conducted in the north-eastern Free State between August 1999 and July 2000. Giemsa-stained blood smears were negative for blood parasites. A total of 94% of the cattle were sero-positive for Babesia bigemina by Indirect Flourescent Antibody Test (IFAT) while 87% were sero-positive for Anaplasma by Enzymed-linked lmmunosorbent Assay (ELISA). All the animals were sero-negative for B. bovis and this is probably because the tick vector (Boophilus microplus) which transmits the disease is not present in the Free State Province. There was no significant difference in the incidence of either anaplasmosis or babesiosis between the seasons. Two tick species belonging to the family Ixodidae were found on cattle, namely, Boophilus decoloratus and Rhipicephalus evertsi evertsi. In the present study significant differences in seasonal burdens of B. decoloratus occurred, with the highest infestations recorded from February to June. The presence of R. evertsi evertsi throughout the year without any or with small fluctuations in winter months was observed, with a peak from February to May. The observation of negative blood smears but high incidence of positive serological results for Anaplasma and Babesia for the same group of cattle indicates that this area is endemic for these diseases but with a stable disease situation.Item Open Access Diagnosis of Babesia equi in horses belonging to resource-poor farmers in the northeastern Free State, South Africa(University of the Free State, 2005-10) Motloang, Makhosazana Yvonne; Mbati, P. A.Babesia (Theileria) equi, a tick transmitted protozoan parasite and one of the causative organisms of equine babesiosis is widespread in South Africa. This work reports on a study that evaluated the usefulness of the polymerase chain reaction (PCR) for detection of B. (Theileria) equi in horses belonging to resource-poor farmers in the northeastern Free State, South Africa. Blood samples from a total of 99 horses and seven from previously known B. (Theileria) equi infected horses on Kaai plaas farm (Onderstepoort Veterinary Institute) were examined for the presences of B. (Theileria) equi using Giemsa-stained blood smears, immunofluorescent antibody test (IF AT) and PCR technique. Giemsa-stained blood smears and serum samples were prepared from blood samples on arrival at the university laboratory. Aliquots of blood samples from EDT A coated vacutainers were transferred into marked cryogenic vials and stored at -35°C for further use in PCR. A PCR system that amplifies a 664 bp target of region of the l 6S rRNA gene and shown to specifically detect the genome of B. (Theileria) equi, was· used in this study. Examination of the 99 blood smears from the northeastern Free State revealed no parasites whilst serological analysis showed 98% of animals were seropositive for B. (Theileria) equi, 48% were seropositive for B. caballi and 58% had mixed infections for both Babesia species. However, PCR was negative for all the 99 horses from the northeastern Free State. Babesia equi parasites were demonstrated in blood smears from 4 out of 7 horses from the Kaai plaas farm. All these animals were also seropositive for B. (Theileria) equi while 3 out of 7 were seropositive for B. caballi. Three out of seven horses were seropositive for both B. (Thei/eria) equi and B. caballi. All seven Kaai plaas farm horses were PCR positive for B. (Theileria) equi infection. Questionnaire results indicated that 100% of resource-poor black farmers in the northeastern Free State did not know about equine babesiosis, its cause, symptoms, diagnosis or control. Based on the negative blood smears and PCR results, and the high prevalence of antibodies against B. (Theileria) equi as demonstrated by IF AT, the conclusion drawn from this study is that the status of equine babesiosis in the northeastern Free state region is endemically stable.Item Open Access A sero-epidemiological survey of parasites in small stock in the north eastern region of the Free State province, South Africa(University of the Free State, 2001) Mogaswane, Kagiso H. R.; Mbati, P. A.; De Waal, D. T.A sero-epidemiological survey was conducted in the north-eastern Free State region of South Africa to determine the parasites of veterinary importance infecting sheep and goats. Blood smears from sheep (n=37 I) and goats (n= l88) were negative for Anaplasma and Thei/eria or any other blood parasites. All the sheep and goats were seropositive for Theileria species by IF AT while 85% of sheep and 100% of goats tested seropositive for Anaplasma species by competition inhibition ELISA. The observation of the negative blood smears but high incidence of positive serological results for Anaplasma and Theileria species for the two animal groups indicates that this area is endemic but with a stable disease condition. Because the animals did not show any clinical. signs of infection the sheep and goats in the north eastern Free State are probably healthy carriers of Anaplasma and Theileria. Two tick species found to infect sheep and goats in the three study sites of Harrismith, Kestell and Qwa-Qwa were Rhipicephalus evertsi evertsi and Boophilus decoloratus. These two are known vectors of Anaplasma species and Thei/eria species, the two diseases found to infest sheep and goats in this region. Rhipicepha!us evertsi evertsi was the dominant tick species across the study sites. A seasonal pattern in tick infestation was observed, whereby there is a high peak during the wanner months and a very low peak during the colder months of the season. Results from this study will form an integral component in the development of disease control programs in the region.Item Open Access Development of molecular diagnostic methods (LAMP and PCR) for detection of Haemonchus contortus, fasciola spp and Trichostrongylus spp infections in livestock(University of the Free State, 2012-12) Mabe, Lerato Tshepiso; Tsotetsi, A. M.; Thekisoe, O. M. M.Helminths belonging to the genera Haemonchus and Trichostrongylus as well as those of the genus Fascia/a are the causative agents of various helminthiases in animals and sometimes in humans. In both host types infections are often acquired through the ingestion of infected food as well as drinking contaminated water. In animals they disrupt the efficient conversion of food material and absorption of nutrients resulting in weakness and death. Infected humans often suffer from intestinal obstruction, insomnia, vomiting, weakness and stomach pains and sometimes temporary asthma. Infections are associated with huge economic losses globally. Diagnosed is achieved through the observation of clinical manifestations. In low infections alternative diagnosis is based on the recovery and identification of faecal eggs and cultivation of L3 by microscopy. More accurate diagnosis has recently been achieved through the use of molecular diagnostic tools such as PCR and LAMP. The aim of this study was to develop rapid, sensitive, specific and accurate molecular diagnostic assays. In particular the study focused on LAMP and PCR for the detection of H. contortus, Fascia/a and Trichostrongylus spp. infections in four provinces of South Africa. The first study emphasized on the development of LAMP and PCR assays for detection of H. contortus infections in livestock. LAMP primers that specifically amplify the ITS2 gene of H. contortus were designed from this target gene. This set of primers was used to develop a PCR assay for species-specific gene amplification. Both assays were tested at various reaction conditions to optimize for primer annealing temperature. Sensitivity reactions were conducted using 10 fold serial dilutions of target DNA while primer specificity was determined using DNA extracted from closely related species. For the LAMP assay, the optimum annealing temperature was found to be 60°C and SS°C for the PCR assay. When tested for specificity both assays only amplified target DNA thereby proving to be specific. The sensitivity reactions for both the LAMP and PCR assay yielded a detection limit of 0.42 ng and 10-3 ng respectively as the lowest amount of the target DNA that can be detected by the assays. Screening of field samples by PCR yielded negative results on several occasions while the positive control amplified the target gene as expected. Failure to validate the assay using field samples was attributed to poor quality or lack of DNA. Validation of these assays is central to determining their efficacy and potential importance in diagnosing natural infections with high sensitivity and rapidity. Therefore we recommend close evaluation of DNA extraction from faecal material as well as ways of reducing or eliminating PCR inhibitors. The second study was aimed at developing a LAMP and PCR assay for the detection of Trichostrongylus spp. infections in livestock. In this study, the target gene used for LAMP primer design for genus-specific amplification was the ITS2 gene. Two primer sets were designed and from these primers, two PCR assays were developed. All these assays were subjected to various LAMP and PCR conditions respectively in order to determine suitable annealing temperatures for each assay. Various methods of DNA extraction were evaluated for troubleshooting together with the use of already published PCR primers. Both LAMP and PCR assays did not amplify the target gene at different ranges of annealing temperatures tested. Furthermore, no amplification was achieved from control DNA samples extracted using different methods. Negative results were obtained when the PCR was troubleshot using already published primers. The results achieved with the designed assays may be a direct consequence of improperly designed primers; however, the inability of all primer sets to function for LAMP and PCR as well as for already published primers suggests problems with the extraction of DNA. This can be attributed to ineffective disruption of worms or the presence of DNases that may degrade DNA and inhibit its amplification. The results of this study suggest the need to evaluate pre-treatment of worms prior to DNA extraction. Evaluation of the methods used to reduce the effects of PCR inhibitors during extraction and amplification of DNA may also be considered. The third study was aimed at developing a LAMP and PCR assay for detection of Fascia/a spp. infections in livestock. Two sets of primers for species-specific amplification by LAMP were designed by targeting the ITS2 gene of both F. hepatica and F. gigantica. Species-specific PCR assays were developed from the latter primers and gene of the parasites and the assays were then tested at various reaction conditions. A genus-specific PCR assay was subsequently developed and tested. No amplification of DNA was observed with the F.hepatica-specific LAMP assay whereas the assay developed for specific detection of F. gigantica produced false positive results. All PCR assays yielded negative results following many attempts to optimize for primer annealing temperature. Reagent contamination was eliminated as the source of non-specific amplification with LAMP suggesting that improper primer design was a possible cause of this problem. On the other hand failed attempts to optimize all the other assays suggest that there is need to evaluate pre-treatment of worms prior to DNA extraction as well as the methods reducing the effects of PCR inhibitors during amplification. Overall, this study successfully achieved the development and optimization of a LAMP and PCR assay for detection of H. contortus DNA. However, subsequent validation of the assays using field derived samples was not possible. The overall results of this study mostly point to faulty primer design and the presence of PCR inhibitors in extracted DNA samples, extracted from either tissue or faecal samples. According to previous studies the presence of inhibitory substances may interfere with the lysis step, inactivate the thermostable DNA polymerase and even interfere with nucleic acids. Therefore evaluation of more accurate methods of mechanical disruption of the worms, DNA extraction, primer design and the use of amplification facilitators may yield desired results. Therefore, these factors should be taken into account for successful development and validation of the molecular diagnostic tool for detection of helminth infections.Item Open Access Molecular phylogeny of South African Anopheles, Aedes and Culex (Diptera: Culicidae) based on COI, ITS2 and 28S DNA sequences(University of the Free State, 2021-11) Whitehead, Liezl; Swart, V. R.; Gryzenhout, M.; Koekemoer, L. L.Mosquitoes have a substantial impact on human and animal health as vectors of disease and consequently take a heavy toll on the economy. South Africa hosts a multitude of mosquito-borne pathogens, including numerous arboviruses and Plasmodium. Many pathogens are transmitted by species currently belonging to three genera, namely Aedes Meigen, 1818, Culex Linnaeus, 1758 and Anopheles Meigen, 1818. Mosquitoes have traditionally been classified by their morphology, which has produced several non-monophyletic taxa. However, the field of systematics has been revolutionised with the advent of molecular techniques, which allow researchers to gain novel insights into the hidden affiliations of taxa. Since few phylogenetic studies have focused on South African mosquitoes, the current study aimed to investigate the intrageneric relationships of South African Aedes, Culex and Anopheles species based on COI, ITS2 and 28S DNA sequences. Therefore, mosquitoes were sampled across various habitats within central South Africa, ranging from relatively pristine nature reserves to urban centres with substantial anthropological activity. Morphological variations of sampled specimens were noted and numerous morphotypes were sequenced for the three target DNA regions. The identities of sampled specimens were verified with BLAST queries and the generated sequences were incorporated into genus-specific multiple sequence alignments. The datasets included representatives of many major taxonomic subdivisions and all accessible South African species, which were obtained from international DNA repositories. Due to the limited availability of sequences, it was not possible to construct concatenated datasets consisting of all three target DNA regions. Therefore, the aim was rather to incorporate shared representative species within each of the independent COI, ITS2 and 28S datasets. These genus and gene-specific multiple sequence alignments were used to conduct maximum likelihood and Bayesian phylogenetic analyses. Several phylogenetic findings were corroborated by multiple gene regions and supported by the results of other authors. This included the polyphyly of several Aedes subgenera (Aedimorphus, Ochlerotatus and Stegomyia) and the non-monophyly of several Neomelaniconion species. The subgenus Culex was also often rendered non-monophyletic by two other subgenera (Barraudius and Oculeomyia) and similarly contained polyphyletic assemblages of Cx. pipiens, Cx. quinquefasciatus and the Pipiens Group. Finally, the current analyses also recovered non-monophyletic groupings of the subgenus Anopheles, the Laticorn Section and the Funestus Group within the genus Anopheles. These results emphasized consistent phylogenetic findings, revealing numerous challenges within the current systematic framework. Therefore, the study provided insight into the affiliations of numerous South African species, generated molecular and phylogenetic data for sampled individuals and provided distribution data for the associated specimens. These results expanded the foundation of phylogenetic and ecological data, which can be incorporated into other epidemiological, biogeographical and evolutionary investigations.Item Open Access Geographic distribution and habitat selection in the berg adder, Bitis atropos (serpentes, viperidae) on the Mpumalanga escarpment, and the consequences for conservation(University of the Free State, 2019-06-01) Jordaan, Adriaan; Heideman, N. J. L.; Buschke, F. T.Understanding how species use geographic space is foundational to ecology, biogeography and conservation biology. However, such essential geographic information remains unavailable for many species. This is especially true for reptiles, which, despite being the largest group of terrestrial vertebrates, remain poorly studied relative to other groups. Moreover, the number of reptile species keeps growing, as new species are discovered and described from existing species complexes. Therefore, in this dissertation, I set out to narrow this knowledge-gap by exploring geographic patterns in the Mpumalanga population of the berg adder (Bitis atropos). This population represents a distinct evolutionary lineage within a larger species complex and justifies reconsideration as a distinct species. I began by using species distribution modelling to estimate the total geographic range size of this snake lineage and showed how it has a restricted distribution that has experienced persistent habitat loss in recent decades. Combined, these results suggest that this lineage would be classified as "Vulnerable" according to IUCN criteria should it be recognised as a new species. I then carried out repeated field surveys to study Mpumalanga berg adder habitat-use within the Buffelskloof Private Nature Reserve. These field records were combined with topographic information to quantify habitat occupancy and account for imperfect detection. Analyses showed that the snakes were more likely to occur on north-west facing slopes with higher heat load index values and, therefore, higher ambient energy. This is important for behavioural thermoregulation in ectothermic species. By understanding distribution patterns and habitat use, this thesis is an important piece in a larger puzzle of reptile spatial ecology. Ultimately, the results presented here provide a deeper evidence base on the ecology and biogeography of a poorly-studied reptile to inform future conservation and management.Item Open Access Blood parasites of Free State and Lesotho reptiles(University of the Free State, 2003-07) Van As, Johann; Davies, Angela; Basson, LindaEnglish: The study of blood parasites of reptiles is a relatively new and unexplored field of research in South Africa. The Free State province and the Lesotho highlands provide a range of reptiles in which their intraerythrocytic parasite fauna were explored. Objectives of this study were to set a baseline of blood parasite diversity and to identify the enigmatic Sauroplasma Du Toit, 1937 and Serpentoplasma Pienaar, 1954 infections in lizards and snakes, respectively. Surveys were conducted in various localities in the Free State and Lesotho. Although low in diversity, 204 specimens representing 14 species of lizards, and 59 specimens representing 13 species of snakes were investigated for the presence of blood parasites. Three known infections were found: Sauroplasma thornasi Du Toit, 1937, Sauromella haemolysus Pienaar, 1954 and Hepatozoon (Haemogregarina) sebae (Laveran and Pettit, 1909) Smith, 1996. These were redescribed and S. thomasii and H. sebae were examined by aid of transmission electron microscopy. The investigation led to the discovery of six new records and possibly new species of haemogregarines named Hepatozoon sp. A-F, four new records and possibly new species of lizard malaria named Plasmodium sp. A-D, and a viral infection possibly of the Pirhemocyton type. Furthermore nine new host and distribution records for Sauroplasma in lizards and nine for Serpentoplasma in snakes are described. Ultrastructural investigations of S. thomasi in Cordylus giganteus A. Smith, 1844, Serpentoplasma in Bitis arietans arietans (Merrem, 1820) and H. sebae in Python sebae natalensis (Gmelin, 1789) were the first to examine the nature of infections in this manner. This is the first comprehensive survey of the biodiversity of blood parasites in reptiles in the Free State and Lesotho highlands.Item Open Access Effect of soil moisture and host plants on behaviour and survival of the common cutworm, agrotis segetum (Denis & Schiffermüller) (Lepidoptera: Noctuidae)(University of the Free State, 2001-11) Mabuda, Khathutshelo; Van der Westhuizen, M. C.; Du Plessis, M. J.English: The objectives of this study were to investigate the effect of light, soil moisture and host plants on behaviour of Agrotis segetum (Denis & Schiffermuller). A marking technique for larvae was developed to study behaviour at night. The marker consisted of al: 1 mixture of Humbrol paint and fluorescent powder. It had no adverse effect on larval behaviour and survival. Dusting of larvae with fluorescent powder also proved useful in tracking movement for short range studies. Powder trails could be detected for a distance of up to two meters. The effect of illumination with different light sources i.e incandescent, infrared and UV light on activity of larvae was evaluated. Larvae were starved for 24, 48, 72 and 96 hours. Significantly more larvae that had been starved for 72 and 96 hours remained on the soil surface in search of food after an hour of illumination with infrared and UV light. Light and dark adaptation periods of 24,48 and 72 hours had an effect on larval activity. Phototaxis of A. segetum was therefore influenced by their degree of starvation and dark adaptation. Larvae moved towards the light source when illuminated with UV light under field conditions possibly perceiving it as a sign of open space. To evaluate the feeding behaviour of A. segetum larvae under wet and dry conditions, a greenhouse trial with Chenopodium carinatum R. Br., Portulaca oleracea L. and Zea mays L. was conducted. Significantly more above ground plant sections of all the three plant species were damaged under both dry and wet soil conditions. Larvae survived for approximately one week at soil moisture levels 10, 20, 40, 60, 80 and 100 % in the absence of food. To evaluate survival in the presence of food at various moisture levels, Amaranthus hybridus L. and P. oleracea were buried in separate pots at soil moisture levels 0, 10 and 80 %. All larvae remained alive for up to 10 days at the 80 % moisture level for each weed species. Larval survival was lowest at ° % soil moisture in both the absence and presence of food. Larval development was supported until pupation on A. hybridus at 80 % soil moisture. Agrotis segetum moths were given a choice between six plant species for oviposition. Eggs were laid on all plant species. Oviposition was however significantly higher on A. hybridus and 1. purpurea. Significantly more eggs were laid on stems than on the leaves. When laid on leaves, the abaxial was preferred over the adaxial surface. There was no significant difference in the number of eggs laid when given a choice between Vetiveria zizanioides (L.) Nash and Pennisetum purpureum (K.) Schumach. Dry leaves of P. purpureum and V. zizanioides were more preferred for oviposition than fresh plant sections. Plants selected by moths for oviposition are not always appropriate hosts for newly emerged larvae. Larval survival was low on some of the plants preferred for oviposition. First instar larvae were notable to feed on the grasses, P. purpureum, V. zizanioides and Z. mays and died while in the first instar. Survival and mass gain of first instar larvae fed on A. hybridus and C. album was significantly higher than on the rest of the plant species. Presence of these plant species in uncultivated maize fields seems to contribute to high population levels due to their suitability as hosts. Leaf trichome density on the plant species evaluated could however not account for the differences in survival and development of A. segetum larvae. The type rather than density of trichornes could be inhibitory or stimulatory to oviposition and larval survival of A. segetum.Item Open Access Myxosporean parasites (Myxozoa: Myxosporea) infecting fishes in the Okavango River system, Botswana(University of the Free State, 2000-06) Reed, Cecile Catharine; Basson, Linda; Van As, Liesl L.English: The Class Myxosporea Butschli, 1882 comprises an enormous group of spore forming obligatory parasites that are able to infect any organ of the hosts in which they are found. Myxosporeans mostly parasitise fish and to date there are more than 1300 species known from fishes throughout the world. In Africa, approximately 84 species have been described from primarily freshwater fish. Research on myxosporeans in southern Africa is very limited, with only a few publications appearing from the coast of South Africa during the early 20th century. In Botswana no research has ever been conducted on myxosporean parasites infecting fishes. Botswana contains one of the worlds largest inland delta systems formed by the Okavango River, which flows in a southeasterly direction from Angola. Knowledge on the fishes as well as their parasites is essential for the well being of a unique body of water such as this. Since myxosporean parasites of fish hold the potential to be pathogenic to their hosts and have been known to cause serious economical losses in aquaculture industries throughout the world, it is thus essential to determine the distribution and biodiversity of these parasites in the Okavango River and Delta in Botswana. The main aims of this project were to investigate the available literature regarding African myxosporeans, compile a database of species infecting freshwater fishes in Africa and to investigate the taxonomic status, species biodiversity and prevalence of myxosporeans infecting fishes in the Okavango River and Delta in Botswana. Fieldwork for this project was conducted in June and July 1998 and 1999 in the Okavango and a total of 275 fishes belonging to 31 species from nine different families were examined for the presence of myxosporean parasites. Five myxosporeans from the genus Henneguya Thélohan, 1892 were collected from four different fish hosts. Only one of these was identified as a known species and was described in this dissertation. A comprehensive morphological description of the four unknown species was also provided. Eight myxosporeans from the genus Myxobolus Butschli, 1882 were also found infecting nine different fish hosts, of which four were identified as known species and described. Detailed descriptions of the four unknown species were provided. It was also found that some fish species showed very high gill infections of these parasites, which may cause some form of respiratory deficiency. This current investigation into myxosporean parasites infecting fishes in the Okavango River System in Botswana provided new insight into the distribution of myxosporeans in southern Africa and has also opened the door for future research in this unique group.Item Open Access Insect-fungal ecology on selected new crops in South Africa(University of the Free State, 2002-11) Swart, Vaughn Richmond; Louw, S. vdM.; Swart, W. J.English: This study investigates insect-fungal-plant interactions on three new crops (viz. cactus pear, pistachio and pigeonpea) in South Africa. Isolation of Drosophila. melanogaster and D. hydei flies from cactus pear (Opuntia ficus-indicay orchards showed that these two species harbour fungi. Inoculations of specific fungi obtained from the two Drosophila species onto fruit and cladodes of cactus pear showed that some of the fungi had the ability to cause rot and may even be pathogenic. Scanning electron microscopy (SEM) confirmed the harbouring of fungi on the bodies by of the two Drosophila species and it also showed that most of the fungal propagules were present on the legs of the flies. An insect succession study showed that both Drosophilidae flies and Nitidulidae beetles play a vital role in the degradation of the fruit by contributing to the rate of fruit decomposition and by spreading yeasts and fungi that cause rot. Further to this a dissemination potential experiment with Drosophilidae also showed that these flies are capable of transmitting fungi, which are capable of causing rot on 0. ficus-indica fruit. Isolation of Nysius natalensis bugs collected in Pistacia vera (pistachio) orchards showed that this species harbours fungi. Additionally, isolation of the heads and faeces of Sparrmannia flava beetles collected in pistachio orchards showed that this species also harbours fungi. More specifically, both these species harbour fungi known to be pathogenic to P. vera. SEM of N. natalensis confirmed that they harbour fungi on the exoskeleton and that most of the fungal propagules were present on the legs of the insects. It was also shown that N. natalensis cause feeding lesions on pistachio nuts, thereby providing access for fungal infection. Analysis of N. natalensis confirmed that their numbers are higher in late summer and that cover crops and surrounding vegetation have an influence on their population dynamics. Analysis of S. flava showed that they have a peak season during summer and that their flight activity is influenced by a combination of temperature, rainfall, windspeed, soil type and lunar phases. Isolation of Clavigralla tomentosicollis bugs collected in a Cajanus cajan (pigeonpea) cultivation showed that this speceis harbours fungi. Some of these fungi are known to be pathogenic to C. cajan. Scanning electron microscopy (SEM) confirmed the harbouring of fungal propagules on the exoskeleton of C. tomentosicollis. It was also shown that fungal propagules were present all over the body, most importantly the mouthparts. A dissemination potential experiment showed that this species is capable of transmitting fungal proapgules to pigeonpea plants. C. tomentosicollis prefers some pigeonpea cultivars above others.Item Open Access Aspects of geophagia amongst dairy cattle in a feedlot system(University of the Free State, 2017-03) Wiid, Emmarentia Elsabe; Butler, H. J. B.English: Modern confinements imposed on animals limit their access to natural resources required for optimal nutrition. This lack of resources may lead to nutrient deficiencies and one of the hypotheses for the deliberate consumption of soil is to supplement mineral deficiencies. Counteraction of acidosis, detoxification, buffering of unpalatable plant compounds as well as the use of soil as anti-diarrhoeal agent are other suggested motivations for geophagy. By means of direct observation as well as camera traps, the behaviour of geophagy amongst dairy cows in a confined feedlot system was documented. In addition, soil analysis was done to determine the geochemistry of ingested as well as excreted soil. Geophagy amongst cows at Amperplaas constituted less than 2% of daily activity. The non-lactating individuals invested less time in this behaviour than lactating individuals and the frequency as well as time spent on this behaviour decreased as the lactation phases progressed. Recorded geophagy occurrences were most during winter and thereafter spring and peak soil ingestion occurred during mid-day. At Amperplaas, all but one of the mineral licks were situated on the undisturbed elevated areas underneath the fence wire or around the wooden and iron poles. The average size of a typical mineral lick was about 40 cm by 15 cm with a depth of 10 cm. The soil collected from the mineral licks was alkaline and classified as sandy clay loam. Quartz, Plagioclase and K-Feldspar were identified in all samples while Ilmenite was quantifiable in all but one of the control sites. Silica concentrations were supportive of quartz dominance in the soil collected from mineral licks as well as soil from the stomachs and faecal matter. The most recent mineral lick contained the lowest silica, arsenic, copper, lead, scandium, and zirconium content. But this site had elevated calcium, iron, potassium, magnesium, sodium and phosphorus as well as bromine, nickel, strontium and zinc levels. When offered soil combined with different minerals, the preference of most cows was sodium chloride mixture and thereafter calcium-phosphorus. When comparing the estimated amount of soil ingested per second with the excreted amount of soil, it appears that the daily excretion rate of soil is almost equal to the ingestion rate, therefore not posing any threat for sand impaction. Milk collected at Amperplaas had low levels of calcium as well as potassium and therefore, soil is possibly ingested to supplement this deficiency. Results of this study confirm the ability of some animals to detect deficiencies in the body and to attempt to correct these deficiencies from available resources. Results also support the theories that this nutritional wisdom is partly from cultural transmission as well as innate behaviour.Item Open Access Diversity and seasonality of Chrysomelidae (Coleoptera) on kenaf, Hibiscus cannabinus Linnaeus (Malvaceae), in South Africa, with special reference to preference indices of Podagrica testacea (Chapuis) (Alticinae)(University of the Free State, 2005-05) Van Deventer, Francina Elizabeth; Louw, Schalk vdM.English: Kenaf (Hibiscus cannabinus L.) is being developed as a new fibre crop in South Africa and its potential in this regard has been under investigation since 2000. Kenaf is a short-day, annual, herbaceous plant. It belongs to the Malvaceae, a plant family notable for both its economic and horticultural importance. The kenaf stalk consists of two types of fibre, an outer ‘bast’ and an inner ‘core’, thereby making it a multi-purpose crop, with a variety of applications, for example, paper, building materials, adsorbents, textiles, livestock feed. Monitoring of all arthropods associated with kenaf was conducted at different trial-site localities throughout South Africa and leaf beetles (Chrysomelidae) were recorded as the most abundant phytophagous insect group. The aim of this study was, to first of all, determine the diversity of Chrysomelidae feeding on kenaf. In this regard, a total of 20 species was sampled during the study period. This data was analysed, and preference indices with regard to season, cultivar or cultivation method were determined for the most abundant species. The flea beetle, Podagrica testacea (Chapuis) (Alticinae), proved to be the most abundant and wide spread species, also causing the most damage by skeletonizing the leaves of the plants, characteristically resulting in a ’shot-hole’ appearance. This species occurred in high densities at four of the six locations (i.e. Winterton (western KwaZulu-Natal), Rustenburg (Northwest Province), Addo (Eastern Cape Province) and Makhathini (northern KwaZulu-Natal)) where kenaf trials were planted. The entomopathogen, Beauveria bassiana Balsamo (Vuillemin), was tested as a biopesticide on P. testacea flea beetles sampled at the Winterton (KwaZulu- Natal) site. Results obtained were inconclusive suggesting that the specific formulation used was not successful in controlling these specific flea beetles. Because of the close relationships that exist between plant diseases and insects, the possibility exist that leaf beetles, P. testacea and Monolepta cruciata Guérin- Méneville occurring on kenaf were acting as possible plant disease vectors, was investigated. Beetles from Winterton were incubated and a total of twelve different fungal species were isolated from these specimens, of which Alternaria spp., Fusarium spp. en Penicillium spp. were the most abundant. World-wide defoliation of kenaf plants by insect herbivores has serious consequences in terms of its successful cultivation. Selective artificial defoliation as a partial insect control mechanism has some potential. It implies removing leaves of the plant, thereby excluding the insects on the basis that their food source is no longer abundantly available. Artificial defoliation trials in this study did not demonstrate this, but rather contributed towards an understanding of the level of impact that simulated beetle herbivory exerts on kenaf yield. Overall the impact of Chrysomelidae in the agricultural environments covered by this study has demonstrated that they are prominent role-players with regard to phytophagy and disease transmission. Therefore, these beetles contribute to the wide variety of factors that should be considered and evaluated to justify the ultimate goal of successfully establishing a viable and sustainable fibre industry for South Africa. Future recommendations for the successful cultivation of kenaf in South Africa would be to invest in a resistant cultivar breeding program, for cultivars particularly suited to South African conditions. Accompanying this would have to be an understanding of the array of anthropocentric activities in the agroenvironment and their influence on the status of pests, crop cultivar and density, and environmental variables, and to what extent, all of these would contribute to the justification and scope of a pest management program.Item Open Access The bio-ecology of the grass leaf miner, Agromyza ocularis (Diptera: Agromyzidae), on wheat and barley in the Northern Cape Province, South Africa(University of the Free State, 2010-06) Adendorff, Joan; Prinsloo, G. J.; Louw, S. vdM.English: The grass leaf miner fly, Agromyza ocularis (Spencer) (Diptera: Agromyzidae) was first described from specimens collected at Ceres (1959), Giant's Castle (1961) and the Maseru (1963) district. These specimens were collected with a sweep net from grass and no information was recorded on the host range and biology of the species. During 2000, A. ocularis was recorded on irrigated wheat in the Prieska district, Northern Cape and since then the species has spread to many wheat and barley fields in the Prieska and Douglas area. Two distinct types of injury are caused by A. ocularis, namely feeding punctures and leaf mining. The adult femail cause's circular punctures on the upper leaf surface of the host plant with her ovipositor, the ovipositor puncture can provide a feeding or egg-laying site. Mines are caused by the larvae feeding on tissue inside the leaf. The mine commences at the egg laying site where the larvae hatch. The mines are usually linear in the direction of the stem attachment or apex of the leaf. The area mined is dead, necrotic tissue and could not be revived through plant growth compensation. Photosynthesis rate by the leaves is therefore diminished. A mature larva cuts a slit in the leaf epidermis, escapes from the mine, drops to the ground and burrows into the damp soil adjacent to the plant and pupates. The pupal stage can last 23 days at 25°C under laboratory conditions. A pupal diapause stage which lasts for ±10 months in the laboratory can occur. The function of this stage is not clear, but it could be a mechanism to survive through periods when food resources are not readily available and extreme high temperatures occur. The lifespan of the adult fly is 30 days at 25°C, also under laboratory conditions. This can lead to overlapping generations of flies in a field and increase of damage to the crop. The flies were found to be active in the field when mean daily temperatures ranged between 10°C and 30°C and high relative humidity prevailed due to irrigation. This could be the optimal conditions the flies need to survive and thrive. A number of alternative host plants for A. ocularis were identified, i.e. Phalaris minor (small canary grass), Bromus catharticus (rescue grass), Lolium perenne (perennial rye grass) and Avena fatua (common wild oats). These grasses were recorded on the edges of the wheat fields during the wheat growing season, but not during the peak of the summer or in the natural vegetation surrounding the fields. The appearance of the flies in the cultivated fields differed by 14 days between 2008 and 2009 growing seasons. In spite of this difference, the crops in both years were older than plant growth stage (GS) 5 (Joubert scale) and the plants had already entered the stem elongation phase when the leaf miner outbreak occurred. It thus seems as if the appearance of flies is not linked to the specific plant growth stage, but rather to climatic conditions. At plant GS 5 the number of tillers per plant is already set and the plants' energy is utilized to initiate the number of heads per plant and the number of florets per head. If the plant is heavily damaged by this time, the number of heads and florets per plant will be influenced. The leaf miners commence oviposition on the lower older leaves of the plant, which are then obviously mined first. The rest of the leaves are attacked as they develop. In 2008, 100% of both barley and wheat plants sampled in the Douglas area were damaged by leaf miners when evaluated at GS 5. This damage intensity continued in all subsequent investigations. The number of damaged tillers per wheat plant increased from 48% - 63% between GS 5 and GS 17, the latter being the stage immediately prior to flowering. The number of damaged wheat leaves per tiller increased from 33% - 58% between GS 5 and 17, whilst indices ranging from 10% to 100% of leaf area damage were recorded. This amply demonstrates the severity of damage that is caused by the leaf miner. The damage varies, however, between fields and between years and could be due to specific spatial and temporal parameters. Damage to the crop and its yield is thus expected. However, insecticide trials conducted under similar conditions during 2008, demonstrated no difference in yield, albeit that a number of larvae were still present per tiller. The plant therefore seems to be able to compensate for the damage inflicted by the leaf miner. If the optimal conditions under which the crop is produced, i.e. sufficient water and fertiliser, is considered, this could be possible. The testing of different insecticides in field trials between 2007 and 2008 provided variable results, with a double dosage of Unimectin® resulting in 80% reduction in larval numbers on barley, which is the only reduction figure which is according to the pesticide registration application (Act 36 of 1947). In 2008 only Abamectin® double dosage met these standards and was the most successful in larval reduction, resulting in figures of 53% - 85% on barley. In spite of all these variables no significant decrease or increase in yield could be measured on any of the treatments in any of the years, suggesting that the plants could absorb the damage levels through compensatory growth.Item Open Access Development of molecular and serological assays for diagnosis of bovine (taenia saginata) and porcine (taenia solium) cysticercosis in South Africa(University of the Free State (Qwaqwa Campus), 2016-09) Seipati, Eunice Sarah; Tsotetsi-Khambule, A. M.; Thekisoe, O. M. M.Cysticercosis is an infection of cattle and pigs caused by metacestodes of human tapeworms, Taenia saginata, and T. solium. Currently, meat inspection is the standard method used for diagnosis of cysticercosis and has proven to be a less sensitive and subjective. The current study was therefore aimed at improving and developing specific and sensitive molecular assays for detection of T. saginata and T. solium infections in cattle and pigs respectively. Furthermore, this study also sought to assess the potential of antigen detection ELISA for diagnosis of cystircercosis in South Africa. The currently available conventional PCR assays respectively targeting HDP2 and cox1 genes were optimised for use under South African conditions, whilst real-time PCRs (qPCR) targeting the cox1 gene of both T. saginata and T. solium were newly developed. The HDP2 gene PCR assay was successfully optimised, however no positive results were obtained in the field samples. However, the assay targeting the cox1 gene yielded positive results in both the control and field samples. All cyst samples collected from bovine and porcine carcasses tested positive, but only bovine blood samples tested positive with a prevalence of 94% (577/614) with no positive results obtained in the porcine blood samples. The T. saginata and T. solium qPCR assays were successfully developed with respective detection limits of 0.0013 ng/μl and 0.0034 ng/μl. Both assays only detected target species, thus showing good specificity. The assays respectively confirmed 63% (45/71) and 100% (2/2) T. saginata and T. solium cysticerci respectively, further detected 75% (458/614) of T. saginata and 33% (76/233) of T. solium infections in bovine and porcine blood samples. Furthermore, a bovine cysticercosis sero-prevalence of 5.6% (18/320) was obtained through HP10 AgELISA, although meat inspection recorded 0% prevalence. The study also showed that MoAb (HP10) antigen detecting ELISA is more sensitive than meat inspection in the diagnosis of taeniid infection in cattle. Both conventional and real-time PCR assays targeting the cox1 gene proved that they can be used as confirmatory tools for meat inspection results made at abattoirs and have the potential to be used as pre-mortem diagnostic tools for detection of T. solium and T. saginata infections in cattle and pigs. Further validation of the developed qPCR assays using known taeniid species positive blood samples is recommended.Item Open Access A review of the genus Caligus (Copepoda: Caligidae) from South Africa(University of the Free State, 2004-09) Grobler, Nicolaas Johannes; Van As, J. G.; Olivier, P. A. S.English: The ichthyoparasitic and fish louse copepod genus, Caligus Müller, 1785, is the most abundant copepod genus, found in almost any part of the world’s oceans, with the exception of one freshwater species. More than 315 nominal species have been described, of which only 26 species have been recorded off the coast of South Africa. Most of the information on this genus from South Africa is limited to very old and often incomplete taxonomic descriptions. Of the 26 known species of Caligus recorded from South Africa, only 15 species have reference material, either in the collection of the South African Museum, or in the collection of the Aquatic Parasitology Research Group, University of the Free State, Bloemfontein. These 15 species are redescribed, and the validity of each species is discussed. Additional morphological features for six species are given, based on studies with the aid of scanning electron microscopy. In addition to the species descriptions, a synopsis of host/parasite and parasite/host is given. A key to all 15 species reviewed in the present study is included. The reference material of 15 species has disappeared, mainly due to material on loan that got lost. These species will, however, remain as species recorded from South Africa, until such time that their status can be validated. However, for the purpose of this study, they are included as part of the South African fauna. Three of these 15 species, Caligus affinis Heller, 1866, C. africanus Oldewage & Van As, 1989 and C. labracis Scott, 1902, have been misidentified. Specimens of the misidentified C. affinis were subsequently collected from the same host, Pomatomus saltatrix (Linnaeus, 1766), in the present study, and a detailed description is included for this species (Caligus sp. B), but it is still uncertain whether this species is new. Caligus africanus is not a valid species and is therefore synonymised with Caligus tetrodontis Barnard, 1948. One new species (Caligus sp. A) has been collected from the Kob, Argyrosomus japonicus (Temminck & Schlegel, 1843), and a fully illustrated description is supplied. With the description of this new species, as well as the inclusion of Caligus sp. B, the total number of species recorded from the South African coast is 28. The genus Caligus can traditionally be divided into four major species groups based on the segmentation and armature of the exopod of the fourth leg. In this study, a fifth group is proposed to include eight species (two species from the present study) with the same characteristic features of the exopod of the fourth leg. In this group the exopod of leg 4 is either one- or two-segmented, the first segment bearing a terminal spine and the last segment bearing only two terminal spines. Future studies on the caligid copepods of South Africa should involve molecular work, as this trend may be useful when species are found which are close to each other, as was found in the present study with Caligus epinepheli Yamaguti, 1936, and C. rotundigenitalis Yu, 1933, or to explore phylogenetic relationships within the family to see whether a species belongs to another genus or not. During the course of the present study, which started as an Honours project in 1997, four papers have been published in international journals. Various conference proceedings have also been published during the course of this study. This present study forms a well-outlined reference for future studies on species of Caligus from South Africa.Item Open Access An examination of non-mammalian cynodont cranial endocasts(University of the Free State, 2010-09) Du Plessis, Dewald; Botha-Brink, J.; Butler, H. J. B.English: The palaeoneurology (study of the nervous system of extinct animals) of nonmammalian cynodonts, which were the ancestors of mammals, is poorly understood. It is thought that the relative brain size of cynodonts increased with time, and that this change significantly affected their physiology and behaviour. Forty-four cynodont skulls belonging to eleven genera from Permian, Triassic and Jurassic strata were scanned and analysed using neutron computerized tomography (NCT), at the SAFARI-1 Nuclear Research Reactor of the Nuclear Energy Corporation of South Africa (Necsa). Data concerning the endocranial casts was hereby obtained in a non-destructive manner. Four specimens, namely Galesaurus, Platycraniellus, Langbergia and Tritylodon produced successful scans and were used for further study. Measurements of the cranial material were used to calculate the size of the brain endocasts and the approximate body masses were determined from an equation using skull length. The encephalization quotient (EO), a measure of brain size relative to body size, was determined for each specimen using brain volume and body mass. Changes in the position of the brain inside the skull, and an overall increase in size of the brain in the more derived cynodonts indicate some development towards the mammalian condition. Taxonomic variation regarding the shape and size of the olfactory bulbs and other parts of the brain endocasts is discussed and related to differences in ecology or behaviour. The relatively deep and large olfactory bulbs in some cynodonts may be an adaptation to either nocturnal behaviour, dietary or habitat preferences. Relatively large cranial endocasts in Galesaurus, Platycraniellus, Massetognathus, Tritylodon, Chiniquodon and the tritheledonts may be related to habitat preference or social behaviour. The basal epicynodonts Galesaurus and Platycraniellus have the largest cranial endocasts of all the cynodonts studied. Their large cranial endocasts may be related to behavioural or ecological preferences, but the results require confirmation from a larger sample size. The present study provides an important foundation from which further cranial endocast studies on cynodonts can be developed.