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Occurrence of Tick-Borne Diseases in Wild Rodent Species and Their Risks for Public Health

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Occurrence of Tick-Borne Diseases in Wild Rodent Species and Their Risks for Public Health OCCURRENCE OF TICK-BORNE HAEMOPARASITES IN SELECTED SOUTH AFRICAN WILD RODENT SPECIES IN THE MNISI COMMUNAL AREA Submitted in partial fulfilment of the requirements for the degree Master of Science (Animal/Human/Ecosystem Health) Faculty of Veterinary Science (Onderstepoort) University of Pretoria Liesl De Boni DECEMBER 2017 DECLARATION I, Liesl De Boni, declare that this mini dissertation is my own original work. Where other people’s work has been used (either from a printed source, internet or any other source), this has been acknowledged and referenced in accordance with University of Pretoria requirements. I have not used work previously produced by another student or any other person to hand in as my own. I have not allowed, and will not allow, anyone to copy my work with the intention of passing it off as his or her own work. 1 ACKNOWLEGEMENTS I would like to give a huge thanks to Prof. Marinda Oosthuizen. You were an enormous help to me during this degree and while writing this mini dissertation. Not only did you help me to choose the project, but you also provided all the support I needed in writing it up. I am very grateful for your guidance, assistance and support throughout the process. I am also very grateful to my co-supervisor, Prof. Kelly Brayton, whose valuable input helped to keep my work accurate and to the point. I would not have been able to complete the laboratory work so successfully and effectively without the help of Ms. Ilse Vorster, Mrs. Milana Troskie and Dr. Agatha Kolo. You all provided so much support for me when it came to the molecular techniques involved and were very generous with the information you shared when I came to write the dissertation. Agatha collected the rodent samples that I used all the way from Mnisi and she performed all the DNA extraction from the samples for me which saved a large amount of time and I am very grateful for that. Thank-you very much to the following people who were involved in the sample collection in the field in Bushbuckridge East, Mpumalanga Province: Prof. Sonja Matthee (Stellenbosch University), Prof. Luis Neves, Prof. Marinda Oosthuizen, Ms. Jeanette Wentzel and Dr. Agatha Kolo (all from Department of Veterinary Tropical Diseases, University of Pretoria). It was very kind of them to allow me to use their samples. The support and love that my husband, Angelo De Boni, always gives to me is what has made this possible. Studying while working full time was a major challenge for me and I believe it also became a challenge for you. Your understanding means the world to me. I am very lucky to have you. I must also thank you for helping me with the design of my RLB image. I would not have been able to do that alone. Thank-you to Mrs. Rina Serfontein for being a massive support when it came to all aspects of this MSc degree. Your prompt and friendly responses made everything a little easier. This work is based on the research supported by the Belgian Directorate General for Development Co-operation Framework (ITM/DGCD), the South African Agricultural Sector 2 Education and Training Authority (AgriSETA), the National Research Foundation (grant 92739 to Marinda Oosthuizen), and University of Pretoria Institutional Research Theme (Animal and Zoonotic Diseases) (to Marinda Oosthuizen). The funding provided made it all possible. Thank-you very much. Being involved with the Department of Veterinary Tropical Diseases while completing this MSc was a pleasure and a privilege for me and for that I thank all the staff members and students there. This dissertation emanates from project V119-15 approved by the Research Committee of the Faculty of Veterinary Science and the Animal Ethics Committee of the University of the University of Pretoria 3 OCCURRENCE OF TICK-BORNE HAEMOPARASITES IN SELECTED SOUTH AFRICAN WILD RODENT SPECIES IN THE MNISI COMMUNAL AREA Candidate: Dr Liesl De Boni Supervisor: Professor Marinda C. Oosthuizen Co-supervisor: Professor Kelly A. Brayton Co-worker: Dr Agatha O. Kolo Department: Veterinary Tropical Diseases Degree: MSc (Animal/Human/Ecosystem Health) SUMMARY In 2013, a disease survey of people in the Mnisi community, Bushbuckridge, Mpumalanga, South Africa revealed a surprisingly high level of exposure to tick bite fever, Bartonella, Q fever and leptospirosis. This indicated the need to educate the community about ecto- parasites, exposure to rodents and general hygiene. In addition, the potential contribution of rodent pathogen transmission to human febrile illnesses is considered a public health concern and warrants further investigation. Most human tick-borne diseases are considered emerging diseases because the number of reported cases annually is increasing. This is probably due to ecological changes, increased awareness and an increased number of susceptible individuals in the global population. Conservation efforts have increased numbers and ranges of host animals which maintain the infected tick population and humans are increasingly encroaching into tick dense areas. Thus, humans are more exposed to tick-borne infectious diseases. Increased awareness by doctors and the public has also contributed to the increased reporting of human tick-borne disease. With major medical advancements, there is an increased proportion of the population that is more susceptible to these infectious diseases. Vector-borne parasitic zoonoses are an important aspect of a constantly changing world because they are also constantly adapting to their new circumstances. Infectious organisms can change virulence, hosts and vectors. These diseases are, therefore, very much a part of 4 the future. With new advances in molecular biology, detection of infectious diseases is constantly improving. The economic impact and losses due to vector-borne diseases are wide and felt at both the human health and animal health levels. Losses occur due to the cost of human illness, deaths, reduced animal production and costs to control or prevent disease. In developing countries where animals are relied on for work, transportation and nutrition, the losses are more profound. The order Rodentia is the most abundant and diversified order. Rodents are distributed all over the world except for Antarctica. They often live in close contact with humans, their farm animals and their pets. Rodents pose a threat to humans in several ways. Not only do they act as reservoirs for infectious diseases and serve as hosts for vectors, they also compete globally with us for food, causing serious pre-harvest damage to cereals every year. Examples of rodent-borne human illnesses include rickettsiosis, babesiosis, human granulocytic anaplasmosis, leptospirosis, bartonellosis, tick-borne relapsing fever, Lyme disease and Q fever. More information about these diseases and their impact on public health in an African context is needed. This mini dissertation is divided into two parts. The first is a narrative literature review on what is known about the common rodent-borne bacterial and parasitic pathogens which have an important impact on public health and their relevance in Africa. The current knowledge is described and recommendations for further research are given. The second part is a laboratory project which focused on determining the incidence of selected tick- borne pathogens found in wild rodent species in the Mnisi community, a livestock-wildlife- human interface area in South Africa. Forty wild rodent blood samples underwent a broad- based screening approach using reverse line blot (RLB) hybridization and a more specific approach using quantitative real-time PCR (qPCR) techniques. The RLB results revealed that 7.5% (n=3) of the samples tested were positive for the presence of A. bovis DNA, while 2.5% (n=1) were positive for A. phagocytophilum DNA; none of the samples tested positive for B. microti (n=0). PCR products from 17.5% (n=7) of the samples hybridized with Theileria/Babesia group-specific probes, 17.5% (n=7) with the Anaplasma/Ehrlichia group-specific probes, 17.5% (n = 7) with both the Babesia 1 and 2 5 genus-specific probes and 7.5% (n = 3) with the Babesia 1 genus-specific probe. A total of 57.5% (n=23) of samples were negative (or below the detection limit of the test) for any species for which we tested. Results of the B. microti SYBR green real-time PCR (qPCR) proved difficult to interpret due to the wide melting temperature range observed; nine of the rodent samples (22.5%) produced melting temperatures within 4°C of that of the positive control (i.e. 79.16 - 86.17°C). Going forward we propose to sequence the parasite 18S rDNA gene of these nine rodent samples to determine whether they are indeed South African variants of B. microti and/or a novel species. The qPCR for A. phagocytophilum revealed 14 positive samples. This is an incidence of 35% (14/40) in a very small sample size which is relevant and warrants further surveillance for A. phagocytophilum in South Africa. Five of the 14 A. phagocytophilum PCR positive rodent samples were confirmed by gene sequence analysis. From the 12 partial msp2 gene sequences obtained, two Msp2 sequence types were identified (designated Ap1 and Ap2); Ap1 differed by one amino acid from Ap2. Ap1 was identical to A. phagocytophilum human strain Webster and A. phagocytophilum strain Dog2, while Ap2 was identical to A. phagocytophilum human strain HZ as well as A. phagocytophilum strains previously described from various hosts (including deer, ticks and mite). It should, however, be noted that we could only obtain partial msp2 gene sequences (334 bp) spanning the conserved area of the gene. Although valuable data, as we could confirm the presence of A. phagocytophilum DNA in our samples, full-length gene sequences will have to be obtained if we want to draw any clear conclusions on the genetic variability of the A.
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