Rift Valley fever virus (RVFV) is a mosquito-borne virus and a member of the family Phenuiviridae and genus Phlebovirus. The single stranded RNA genome consists of three segments, Large (L), Medium (M) and Small (S). Rift Valley fever (RVF) is a mosquito-borne zoonotic disease that may cause large epidemics in ruminants and humans. Infection in humans causes influenza-like symptoms but the disease can also be more severe and fatal. Outbreaks in livestock are classified by abortion storms and young and newborn animals are severely affected with a high mortality rate. Rift Valley fever causes severe health and economic consequences in the areas where it occurs. Since the first recorded incidence of RVF in Kenya in 1930, South Africa has had three major countrywide outbreaks: in 1950-1951, 1973-1975 and 2008-2011. The disease is characterized in southern Africa by large epidemics at long, irregular intervals. The epidemics are usually associated with conditions favourable for proliferation of mosquito populations, such as high rainfall and flooding. Rift Valley fever has previously been isolated from 12 different mosquito species in South Africa including 5 Aedes spp., 3 Culex spp., 3 Anopheles spp. and 1 Eretmapodites sp. The presence of the virus and patterns of occurrence of the disease in the eastern parts of South Africa are poorly understood. Multiple studies were conducted; the aim of the first study was to detect the presence of RVFV in far northern KwaZulu-Natal Province, South Africa and to estimate the incidence rate of seroconversion. Cross-sectional studies were performed in communally farmed cattle (n=423) and goats (n=104), followed by longitudinal follow-up of seronegative livestock (n=253) 14 times over 24 months, representing 160.3 animal-years at risk. Exposure to RVFV was assessed using an IgG sandwich ELISA and a serum neutralization test (SNT) and seroconversion was assessed using SNT. Initial overall seroprevalence was 34.0% (95%CI: 29.5-38.8%) in cattle and 31.7% (95%CI: 22.9-41.6%) in goats, varying by locality from 18-54%. Overall seroconversion rate in cattle was 0.59 per animal-year (95% CI: 0.46-0.75) and in goats 0.41 per animal-year (95% CI: 0.25-0.64), varying significantly over short distances. The high seroprevalence in all age groups and evidence of year-round viral circulation provide evidence for a hyperendemic situation in the study area. The second study investigated the seroprevalence and associated risk factors of RVFV in antelope in the Tembe Elephant Park (TEP) and the Ndumo Game Reserve (NGR), using 326 sera from nyala (Tragelaphus angasii) and impala (Aepyceros melampus) routinely culled over a two-year period. The overall seroprevalence of RVFV was 35.0% (114/326; 95% CI 29.8-40.4%); the presence of antibodies in juveniles (6/21; 28.6%; 95% CI 11.3-52.2%) and sub-adults (13/65; 20.0%; 95% CI 11.1-37.8%) confirmed that infections had occurred subsequent to the 2008-2011 RVF outbreaks in South Africa. Seroprevalence was highest in adults and inversely associated with distance from a swamp or floodplain. The third study aimed to investigate the diversity, abundance, and seasonal dynamics of mosquitoes in the study area, and to screen mosquitoes for RVFV. Monthly collections of adult mosquitoes were carried out from January 2017 to June 2018 at three sites using CO2-baited tent traps. Mosquitoes were identified, pooled and screened for RVFV by quantitative reverse transcriptase (RT)-polymerase chain reaction (PCR) directed toward amplification of a 217-bp fragment of the L segment. A total of 34,848 mosquitoes of 7 genera and 48 species, were captured; Culex (Cux.) tritaeniorhynchus (31%), Cx. (Cux.) antennatus (29%), Aedes (Adm.) durbanensis (12%) and Cx. (Cux.) neavei (10%) were the most abundant species collected. Genera differences were noted between the collection sites. Cumulative rainfall and average minimum temperatures 30 days prior to collection were positively associated with the number of mosquitoes collected while maximum temperatures were only associated with the number of Culex mosquitoes caught. A single pool of Ae. durbanensis was found to be positive for RVFV genomic RNA. The same pool was also positive for Chikungunya virus (Family Togaviridae, genus Alphavirus) (CHIKV) and Sindbis virus (Family Togaviridae, genus Alphavirus) (SINV). The RVFV isolate was closely related to one obtained from Ae. (Neo.) circumluteolus at Simbu pan in 1955, ±20 km from the collection sites for this study. Further investigation should be done on the human health implications of the presence of these three zoonotic arboviruses. It is possible that these viruses are causing disease among the communities in the area and that the diseases are under-reported. The results of this study show that RVFV is circulating in the area in domestic ruminants and wildlife, in the absence of apparent clinical disease, at a rate that varies by location, season and year. It appears that, under similar ecological conditions, domestic and wild ruminants may play a similar role in maintenance of viral circulation, and either or both may serve as the mammalian host in a vector-host maintenance system. The study also demonstrates the presence of a wide variety of mosquito species, several of which are known to be competent RVFV vectors. / Thesis (PhD)--University of Pretoria, 2019. / Veterinary Tropical Diseases / PhD / Unrestricted

In Mnisi, a rural community in South Africa, Rhipicephalus sanguineus is one of the most prevalent ticks found on dogs. The community lies at the wildlife/livestock/human interface where humans are at risk of tick-borne diseases. The aim of this study was to investigate the diversity of the bacterial microbiome in R. sanguineus that may impact human health. Over a 12-month period, R. sanguineus (n=1,788), Rhipicephalus simus (n=61), Rhipicephalus turanicus (n=73), Amblyomma hebraeum (n=68), Haemaphysalis leachi (n=219) and Hyalomma truncatum (n=1) ticks were collected from 64 dogs. Genomic DNA was extracted from salivary gland and midgut tissues of 62 R. sanguineus tick pools (1 pool = 10 ticks); identifications were confirmed using Cytochrome c oxidase I barcoding. The 16S rRNA gene was amplified using barcoded primers and sent for Pacific Bioscience’s circular consensus sequencing. Characterisation of the bacterial microbiome of midgut and salivary gland pools revealed a total of 260,312 sequences with Proteobacteria (85.44%) being the most prevalent phylum found; with Anaplasma (21.69%), Coxiella (12.12%) and Ehrlichia (19.94%) species dominating the microbiome. Further classification of Ehrlichia revealed 95.46% Ehrlichia canis and 4.54% Ehrlichia species Anaplasma consisted of 15.36% Anaplasma centrale, 75.82% Anaplasma platys and 8.82% Anaplasma species Phylogenetic analysis indicated that the A. centrale and A. platys clustered with various other published A. centrale and A. platys sequences, respectively. It also confirmed that all Ehrlichia species sequences detected in this study are E. canis sequences (94.46%). Furthermore, we determined that the Coxiella sequences detected in the study belong to the R. sanguineus Coxiella-like endosymbionts group. Assessment of risk factors for R. sanguineus infestation indicates that higher average monthly temperatures have a significant association with an increased risk of R. sanguineus tick infestations on dogs. Additionally, rearing chickens at the household was significantly associated with a decreased risk of R. sanguineus tick infestations on dogs. Our study indicated that R. sanguineus could be a potential reservoir for important bacterial pathogens of zoonotic importance. / Dissertation (MSc (Veterinary Science))--University of Pretoria, 2019. / Veterinary Tropical Diseases / MSc (Veterinary Science) / Unrestricted

Ticks and tick-borne diseases (TBDs) cause significant losses, through their effects on health, production of animals and humans worldwide. Notably, the countries located within the tropics and subtropics such as Uganda are the most affected due to abundance and distribution of the tick vector. Unfortunately, there is little data in Karamoja Region regarding tick species and the pathogens they transmit. Therefore, a cross-sectional study was undertaken to determine the various tick species, and to detect the tick-borne pathogens within the ticks collected from cattle in Karamoja Region, northeastern Uganda. Between June 2017 and early September 2017 (wet season), a total of 4,897 ixodid ticks were collected from 100 cattle in 20 purposively-selected herds. Three genera of ticks, namely Amblyomma (96.8%), Hyalomma (0.6%) and Rhipicephalus (2.6%) were identified. From the ticks collected, the most dominant species was A. lepidum (93.85%), followed by A. variegatum (2.0%), R. evertsi evertsi (1.0%) and A. gemma (0.98%). Tick species that have not been reported in recent studies in Uganda were found amongst cattle in Karamoja, and these were R. pravus, R. praetextatus and R. turanicus. A representative number of ticks, from each tick species identified in the present study were placed in pools of 1 to 10. Subsequently, a reverse line blot (RLB) hybridization assay was performed to screen for the presence of tick-borne pathogens. Out of the 40 tick pools, 30 (75%) were positive for tick-borne pathogens of the genera Ehrlichia, Anaplasma, Babesia and Theileria. The RLB assay results showed that 57% (n=17) of the tick pools were positive for single infections, while 43% (n=13) had mixed infections. The most frequently detected tick-borne pathogens were T. parva (10 pools), T. velifera (10 pools), T. mutans (9 pools) and Theileria sp. (sable) (5 pools). Other pathogens detected were E. ruminantium, B. microti, B. rossi, T. separata and B. bigemina. The tick-borne species B. microti, B. rossi, Theileria sp. (sable) and T. separata are not common in cattle, or not known to infect cattle, but were detected from the ticks collected. The detection of B. microti in this study may point to incidental infections with implications for human health. There could have also been a possibility of cross reactions during the RLB analysis leading to the detection of B. microti in this study. These findings provide knowledge of the distribution of ticks and epidemiology of tick-borne pathogens in cattle and may provide support for control of TBDs and improve cattle productivity. / Mini Dissertation (MSc (Tropical Animal Health))--University of Pretoria, 2019. / Veterinary Tropical Diseases / MSc (Tropical Animal Health) / Unrestricted

Rift Valley fever (RVF), is a mosquito-borne viral disease affecting humans and some species of ruminants including sheep, cattle, goats, buffalos and to a lesser extent wild animals. It is a re-emerging disease responsible for major losses in livestock production, with negative impacts on livelihoods of both commercial and resource- poor farmers in sub-Saharan African and some countries in the Middle East. It remains a threat to both endemic and non-endemic countries where competent mosquito vectors exist. The RVF virus (RVFV) causes the disease and though only a single serotype exists, differences in virulence and pathogenicity of the virus have been observed in a wide range of affected mammalian host species. This necessitates the need for a detailed genetic characterization of various isolates of the virus and whether the causal factors for host tissue tropism can be explained. Therefore, the aims of this study were to obtain comprehensive information on the genetic composition of the RVFVs circulating in South Africa between 2008 and 2010 and to differentiate these isolates based on cell infectivity and genomic parameters. In the first chapter the status of some published literature on the disease as well as the virus are reviewed. Viral characteristics, replication, assembly and release of the viral particle from the cell as well as virus-host receptors documented are also mentioned in this chapter. Chapter two focused on the genetic composition of RVFVs that caused outbreaks during 2008- 2010 in South Africa. Complete genome sequence analysis of isolates from different hosts and tissues collected at discrete foci of outbreaks were analysed and compared with virus sequences from earlier outbreaks in South Africa and from other countries. Phylogenetic analysis indicated that viruses that caused outbreaks during 2008-2010 were most probably reassortants, resulting from exchange of portions of the genome of different isolates, particularly of Segment M. In addition, the analysis indicated that the viruses were not introduced from outside the country but mutated in time and caused the outbreaks when the environmental conditions became favourable. Although no clear association between the virus genotype and phenotype has been established, various amino acid substitutions have been implicated for changes in the phenotype. The third chapter describes the characterization of isolates derived from different hosts (bovine and ovine), but from the same tissue (liver). The isolates from bovine liver presented a different growth phenotype in a cell culture-based system as well as some amino acid substitutions when compared with isolates from ovine livers. Although the codon usage patterns of the six isolates were the same, they differed with those of their hosts. Further investigation of the coding regions of the genome, molecular modelling of glycoproteins and codon usage bias failed to explain the phenotypic changes. The fourth chapter focused on an attempt to identify RVFV glycoprotein receptors using the yeast two-hybrid (Y2H) system. Baby hamster kidney cells were chosen as host cells in the laboratory because hamsters are known to be highly susceptible to RVFV. The complexity of the cDNA library constructed from BHK cells were assessed by random sequencing of 100 clones and revealed that 51 clones were genes from mRNA from the Syrian/Golden hamster using BLAST. The constructed library can also be used to study other animal pathogens such as bluetongue virus and African horse sickness virus. The constructed bait plasmids did not show any autoactivation or toxicity in yeast, thus making them suitable to be used in the Y2H system. Twelve unique clones (4 clones using transformants of the glycoprotein Gn and 8 clones using transformants of glycoprotein Gc) were screened from the cDNA library. Identification and further characterization of the clones is necessary. Sampling of the isolates that caused the 2008-2010 outbreaks in South Africa and full genome sequencing indicated that the isolates were genetically distinct, grouping in different clades, namely C and H. Reassortment have been identified in some of these isolates, particularly in their M segments. The majority of isolates that emerged in the outbreaks accumulated mutations over time while circulating in South Africa. The impact of these mutations on the pathogenicity of RVFV should be further investigated. Sequencing should be done on clinical samples directly to have a better idea of the phenotype and the effect of amino acid substitutions. Different phenotypes observed between cattle and sheep in tissue culture systems should be further investigated including investigation of different phenotypes in vivo using small experimental animals. The study has laid a foundation in understanding the pathogenicity of RVFV and necessitates the importance of understanding molecular mechanisms of the virus. / Thesis (PhD)--University of Pretoria, 2019. / Veterinary Tropical Diseases / PhD / Unrestricted

Foetal loss can be devastating to a cattle farmer. In the Free State province many commercial cattle farms contend with foetal loss due to abortion. The causes of most of these abortions are never diagnosed because of inappropriate samples submitted; diagnostics being too expensive or non-submission of samples due to ignorance. The aim of this study was to investigate the apparent seroprevalence of Brucella species and Coxiella burnetii in commercial cattle of five epidemiological units. The overall apparent seroprevalence in this study was 22% and 11% respectively for Brucella species and Coxiella burnetii antibodies. Although the presence of antibodies does not lead to a definitive diagnosis, it is now known that there are bacterial challenges in these commercial cattle, and this could form the basis of future studies. Improved education of the public as well as communication with the human health sector is necessary to effectively control brucellosis and Q-fever. / Dissertation (MSc)--University of Pretoria, 2019. / Veterinary Tropical Diseases / MSc / Unrestricted

In rural areas of developing countries like South Africa, people typically depend on goat farming for both meat and milk production due to the shortage of grazing and the higher maintenance cost of cattle. An understanding of the functionality of the mammary gland and subsequent drug transport into milk are important factors in determining milk withdrawal periods and drug residues in milk intended for human consumption. Tight cellular monolayers, cultured to resemble the in vivo blood-milk-barrier, are used to evaluate the transepithelial transport of drugs into milk in vitro. The aim of this study was to culture and maintain tight monolayers of primary Boer goat mammary epithelial cells that would be a barrier to paracellular drug movement. Cells were cultured and maintained similarly to the method described by Pantschenko and colleagues (2000), with some adaptations and with MCF10a as growth medium. The formation of tight barriers was evaluated by measuring transepithelial electrical resistance (TEER) and the paracellular movement of dextran-FITC. An aggregated monolayer was established which had the characteristic cobblestone appearance, typical of epithelial cells, with no fibroblasts seen microscopically. On day 11 the monolayers appeared to be confluent under microscopic examination, they presented a significant barrier to the movement of FD70 dextran (Papp = 0.001), and the transepithelial electrical resistance (TEER) was greater than 200 ?.cm2. At day 18 of culture, macroscopically the cells started to stack and cell debris formed, presumably due to overgrowing and cell differentiation, and the monolayers were no longer appropriate for use. Furthermore, cryopreservation techniques were performed on the cells and these cells were frozen, stored, and regrown as viable epithelial cells. Primary Boer goat mammary epithelial cells, cultured and maintained using the methods described in this dissertation, form tight monolayers that are a significant barrier to the paracellular movement of relatively large molecules like dextran70, with TEER values appropriate for xenobiotic transcellular flux studies between day 11 and 18 of culture. This timeframe corresponds with the time in which drug transfer studies are typically done in cell cultures from other species. Viable cryopreservation of Boer goat mammary epithelial cells is a useful tool that can be used to enhance these studies. / Dissertation (MSc)--University of Pretoria, 2015. / tm2016 / Paraclinical Sciences / MSc

The main aim of this study was to isolate and characterise antifungal and antibacterial compounds from leaf extracts of Combretum molle which belonging to the Combretaceae family. C. molle is one of the commonly used medicinal plants in southern Africa for numerous ailments. Three animal fungal pathogens, namely, Candida albicans, Cryptococcus neoformans, Aspergillus fumigatus and five plant fungal pathogens, namely, Aspergillus niger, Aspergillus parasiticus, Fusarium oxysporum, Penicillium janthinellum, Rhizoctonia solani and four nosocomial bacteria Staphylococcus aureus, Enterococcus faecalis, Escherichia coli and Pseudomonas aeruginosa were used as test microorganisms for bioactive compounds in leaf extracts of C.molle. Experiments for phytochemical analysis were done using different C. molle leaf extracts which were made using acetone, methanol, ethanol, ethyl acetate, chloroform, butanol and hexane as extractants. Thin Layer Chromatography (TLC) fingerprints of different leaf extracts were developed in three mobile phase systems, EMW, CEF and BEA and detected with vanillin-sulphuric acid spraying agent. The different extracts of C. molle showed the presence of many different compounds with distinct retardation factors (Rf), separated according to their polarities. Bioautography was carried out to determine the number of active compounds and their Rf values. The TLC plates were developed in three mobile systems, each sprayed with either fungal or bacterial strains. In BEA bioautograms of A. fumigatus, clear zones of inhibition were observed at Rf values of 0.12, 0.23, and 0.40. In EMW bioautogram of C. albicans, clear zones of inhibition were observed at Rf value of 0.73, 0.81, 0.87. C. neoformans had weak growth inhibition. Most of the fungal and bacterial strains tested in the bioautography displayed susceptibility to the active compounds, with P. janthinellum and P. aeruginosa showing exceptional sensitivity. The minimum inhibitory concentrations (MIC) values ranged from 0.02 to 2.5 mg/ml against the tested pathogens. The acetone and ethyl acetate extracts had the best inhibitory activity against P. janthinellum with an MIC value of 0.02 mg/ml. The acetone extract of C. molle gave the highest total activity (775 ml/g) against P. janthinellum. C. albicans was the most resistant pathogen with an average MIC value of 0.56 mg/ml compared with the other tested strains. Extracts were active against both Gram-positive and Gram-negative strains. P. aeruginosa extracts had the highest average MIC value (0.24 mg/ml) among the tested bacterial strains. In general, there was good overall inhibitory activity by different extracts of C. molle. Bioassay-guided fractionation of DCM extract of the leaves of C. molle yielded 32 fractions. Further fractionation led to the isolation of five compounds (C1, C2, C3, C4 and C5). Compound C1 was selected for structure elucidation due a larger quantity isolated and higher antimicrobial activity compared with the other isolated compounds. Nuclear magnetic resonance (NMR) spectroscopy and mass spectroscopy (MS) was used to show that compound C1 was taraxerol, belonging to the taraxerane group. Antimicrobial activity of the isolated compound against P. janthinellum had an MIC value of 0.08 ug/ml. Although the compound taraxerol have been discovered in other plant species, it is reported for the first time from C. molle in the study. The results illustrate that crude extracts and compound taraxerol from C. molle can be used as either an antibacterial or antifungal, and warrants further investigation. / Dissertation (MSc)--University of Pretoria, 2017. / Paraclinical Sciences / MSc / Unrestricted

Despite the occurrence of S. Typhimurium infections, little is known on the genetic diversity, virulence characteristics and antimicrobial resistance profiles of poultry S. Typhimurium in South Africa. Therefore, S. Typhimurium (n=141) isolated from organs (n=115) and environments (n=26) of diseased poultry between 1995 and 2002 were screened by PCR for bacteriophages, plasmids and Salmonella pathogenicity islands (SPIs) - encoded virulence genes (virulotyping) which are essential for invasion (invA, sopB, gtgB, sspH1, sopE, spvC, and pefA), survival (sifA, gipA, sodC1, gtgE, mig5, and sspH2) and serum killing (rck, and srgA) of the pathogen in the host. Isolates were also characterized by: pulsed field gel electrophoresis (PFGE) for genetic relatedness, and plasmid profiling (n=43). Furthermore, isolates (n=141) were tested for susceptibility to 16 antimicrobials by disk diffusion and further screened by PCR for the carriage of 27 resistance genes, and integrons. Multi-resistant S. Typhimurium definitive phage type (DT) 104 were determined by disk diffusion and confirmed by PCR. All isolates carried SPIs-encoded genes: invA, sopB, and sifA. Bacteriophages-encoded genes (sspH2, sspH1, sodC1, gtgB, and gtgE) occurred in more than 74.5% of the isolates expect for gipA (57.6%), and sopE (19%). The occurrence of plasmid-encoded genes (pefA, mig5, rck, spvC, and srgA) ranged from 48.2% to 74.5%. Two sample t - test showed that virulence genes: gtgB, spvC, gipA, gtgE, mig5, rck and srgA were more frequent (p ? 0.05) in S. Typhimurium isolates from environments. Virulotyping clustered 141 isolates into 59 virulotypes with 97 isolates clustering in 5 predominant virulotypes while 44 were single isolate virulotypes. PFGE grouped 140 isolates into 55 pulsotypes with 66 isolates clustering in 5 major pulsotypes, 51 isolates clustering in small pulsotypes (containing less than 5 isolates) while 33 were single isolate pulsotypes. Ten plasmid profiles ranging from 2kb to 90kb were observed. The most common plasmid profile contained the 90kb plasmid and was observed in 12/43 isolates. Major virulotypes and plasmid profiles corresponded approximately to pulsotypes and clustered isolates recovered from the same farms or during the same period. Virulotyping and PFGE showed identical discriminatory index (D=0.93). Multidrug resistance (resistance to ? 2 antimicrobials) was observed in 97.2% of isolates. High levels of resistance phenotypes and their respective resistance genes were observed for: streptomycin (94.3%) conferred by ant3'Ia (60.3%) and str (50.4%), sulphonamides (87.2%) conferred by sul1 (66%) and sul3 (31.9%), ciprofloxacin (79.4%) conferred by qnrA (79.4%), tetracycline (61%) conferred by tetB (35.5%) and tetG (28.4%), and cefotaxime (55.3%) conferred by blaSHV (57.4%). Two sample t - test revealed that isolates from poultry organs were more resistant (p?0.05) to ampicillin, amoxicillin clavulanic acid, chloramphenicol, tetracycline and sulfamethoxazole - trimethoprim while isolates collected from poultry environments were more resistant to cephalothin, cefotaxime, ceftazidime, colistin sulphate and nalidixic acid. Using the Kappa statistics, there were agreements ranging from good to perfect between phenotype and genotype. In addition, for every phenotypic resistance recorded, at least one corresponding resistance gene was detected. DT104 strains and class1 integrons were observed in 34.7% and 83% of the isolates respectively. Multi-resistant S. Typhimurium (97.2%) also carried SPIs - encoded virulence genes involved in invasion and survival in the host. In addition, more than 50% of resistant isolates to each of the antimicrobials also carried at least 12 virulence genes: invA, sopB, sifA, sspH2, sspH1, sodC1, gtgB, gtgE, pefA, mig5, spvC, and srgA. A significant number (44.9%) of the DT104 strains that were clustered in the same pulsotype X25 also belonged to virulotype V3a which contained 13 virulence genes: invA, sopB, sifA, sspH2, sspH1, sodC1, gtgB, gtgE, pefA, rck, mig5, spvC, and srgA. Most of isolates that belonged to the same antimicrobial resistance profile (phenotype and genotype) carried at least 8 common virulence genes. In conclusion, these data indicate that S. Typhimurium isolated from diseased poultry carry virulence genes that are usually incriminated in Salmonella human outbreaks. Virulotyping and PFGE showed the same discriminatory index (D=0.93) indicating that virulotyping can be an alternative subtyping method in laboratories where PFGE is not available. Salmonella Typhimurium are also genetically diverse since they were recovered from multiple farms and during a period spanning 8 years. Furthermore, isolates were resistant to multiple antimicrobials used in poultry operations (streptomycin, sulphonamides, and tetracycline) and those used to treat human salmonellosis: ciprofloxacin, and cefotaxime. Multidrug resistant isolates carried most of virulence genes. This relationship between virulence and antimicrobial resistance suggests that the adaptation of isolates against antimicrobial effects may induce expression of virulence factors. The increasing incidence of DT104 threatens the public health since DT104 strains are associated with hospitalizations and deaths in humans. Salmonella Typhimurium carried mobile genetic elements (bacteriophages, integrons and plasmids) which pose a public hazard as they propagate virulence and resistance genes with emerging new pathogenic bacteria as a result. Therefore, monitoring and surveillance of salmonellosis and prudent antimicrobials use need more efforts to ensure animal health and food safety for consumers in South Africa. / Dissertation (MSc)--University of Pretoria, 2016. / Paraclinical Sciences / MSc / Unrestricted

Shiga toxin-producing E. coli (STEC) is a food pathogen causing infections characterised by mild watery to severe bloody diarrhea and complications such as the hemolytic uremic syndrome (HUS). Humans acquire STEC through consumption of contaminated foods of animal origin, vegetables and water. Cattle are the main reservoir of STEC. The severity of STEC infections in humans depends on a number of virulence factors encoded in the bacterium’s genome. The seven major STEC serogroups most frequently incriminated in severe human disease outbreaks and HUS worldwide include O157, O45, O103, O111, O121, O145 and, O26, commonly referred to as the "top/big seven". Although STEC has been incriminated in human disease in South Africa, data on the role of played by cattle in human disease and virulence characteristics of cattle STEC are lacking. Therefore, the objectives of this study were to (i) investigate the presence of the seven major STEC serotypes in healthy beef cattle (cow-calf operations) and (ii) characterise isolates by serotype, virulence genes and markers, and antimicrobial resistance profiles. Polymerase chain reaction (PCR) was carried out to identify STEC serotypes (O and H antigens) and characterize the isolates by virulence factors and markers. The disk diffusion technique (Kirby Bauer test) was used to determine the antimicrobial resistance profiles of STEC isolates against a panel of 15 antimicrobials. Five hundred and seventy-eight STEC isolates (N=578), which had been previously recovered from 559 cattle from five beef farms were screened for STEC O26, O45, O103, O111, O121, O145 and O157. Confirmed STEC belonging to serogroups O26, O45, O103, O111, O121, O145 and O157 to isolates were characterised for major virulence genes including stx1, stx2, eaeA and ehxA. Furthermore, 140 isolates were characterised for xiii Shiga toxins (stx) subtypes, plasmid and pathogenicity island-encoded genes, and antimicrobials resistance profiles. / Dissertation (MSc)--University of Pretoria, 2017. / Paraclinical Sciences / MSc / Unrestricted

Introduction: Campylobacter spp. is the most frequent cause of bacterial gastroenteritis in humans globally. Campylobacter spp. infections are characterized by acute watery or bloody diarrhoea, fever, weight loss and abdominal cramps. Campylobacteriosis complications include extra-intestinal diseases such as Guillain-Barre Syndrome (GBS) or its variant the Miller Fisher Syndrome (MFS). Consumption of contaminated foods of animal origin including undercooked meat, contaminated dairy products has been associated with foodborne campylobacteriosis in humans. Cattle are considered an important reservoir of Campylobacter spp. and a source of foodborne Campylobacteriosis. Antimicrobial treatment failure in most bacterial infections including campylobacteriosis has emerged and led to the increase of animal and human health care costs. The use of antimicrobials in cattle for therapy in both cattle and humans and for growth promotion in exerts selective pressure on bacterial pathogens, which may result in the emergence of antimicrobial resistant Campylobacter spp. strains which can be transferred from animals to humans along the food chain or through contact between animals and humans. In South Africa, studies on the occurrence and antimicrobial resistance profiles of Campylobacter spp. of public health importance are lacking. The main objectives of this study were to: 1) investigate the occurrence of Campylobacter spp. in beef cattle on cow-calf operations in Gauteng and North West Provinces and 2) determine the antimicrobial resistance profiles of Campylobacter spp. isolates. The overall aim of the study was to contribute to monitoring and surveillance of Campylobacter spp. of public health importance in South Africa. Methodology: A total of 537 fresh faecal samples from beef cattle consisting of 453 from adult cows and 102 from calves were collected on 5 cow-calf operations in Gauteng and North West provinces. The samples were screened for Campylobacter spp., including C. jejuni subsp. jejuni, C. coli and C. upsaliensis by culture and the polymerase chain reaction (PCR). Furthermore, 86 Campylobacter spp. isolates consisting of 46 C. jejuni subs. jejuni, 24 C. coli and 16 C. upsaliensis were tested for antimicrobial resistance against a panel of nine antimicrobial agents including azithromycin, ciprofloxacin, erythromycin, gentamicin, tetracycline, florfenicol, nalidixic acid, telithromycin and clindamycin by the broth microdilution method. Results: Out the 537 cattle faecal samples tested in this study, PCR revealed that 29.4% (158/537) [16.23%-42.57%] 95%CI of cattle carried Campylobacter spp. Among the 158 Campylobacter spp. positive cattle, 62.6% (99/158) carried C. jejuni subsp. jejuni, 25.3% (40/158) C. coli, 10.1% (16/158) C. upsaliensis and 3.1% (5/158) cows that had mixed infections. Three cows harbored both C. jejuni and C. coli, one cow carried C. jejuni and C. upsaliensis and one cow carried both C. coli and C. upsaliensis. Further antimicrobial resistance profiling of 86 Campylobacter spp. isolates (46 C. jejuni isolates, 24 C. coli and 16 C. upsaliensis) by the broth microdilution method revealed that the highest resistance rates for clindamycin (36%), nalidixic acid (19.7%), tetracycline (18.6%) and erythromycin (17.4%). However, lower resistance rates against florfenicol (3.4%), gentamicin (4.6%), telithromycin and ciprofloxacin (5.8%) were observed. The isolates were multidrug resistant against tetracycline/clindamycin, erythromycin/tetracycline/clindamycin, and nalidixic acid/clindamycin. Conclusion: Little is known about the occurrence rates of Campylobacter spp. in beef cattle in South Africa. The prevalence of Campylobacter recorded in this study was consistent with various studies that have reported Campylobacter spp. prevalence rates within the same range in cattle in a number of countries with C. jejuni subsp. jejuni as the most predominant species. Campylobacter spp. isolates were mainly resistant to clindamycin, nalidixic acid and tetracycline. Findings from this study highlight the importance of beef cattle as a reservoir and a potential source of clinically relevant and antimicrobial resistant Campylobacter spp. isolates in South Africa. / Dissertation (MSc)--University of Pretoria, 2018. / Paraclinical Sciences / MSc / Unrestricted