This thesis focuses on the molecular characterization of tick-borne parasites of dogs in South Africa. Emphasis is placed on Babesia, Erhlichia, incidental and novel parasite infections that may cause morbidity or mortality in infected dogs. An outbreak of canine babesiosis in the Netherlands is also reported in this thesis. Molecular techniques were employed to isolate, amplify and characterize genomic DNA of these parasites to species level. During preliminary screening of blood samples collected from various sites in the country, that included the Onderstepoort Veterinary Academic Hospital, SPCAs and private clinics throughout seven provinces in South Africa, it was discovered that domestic dogs harboured a wide variety of tick-borne pathogens. The most frequently encountered parasites in South Africa were Babesia rossi, a novel Theileria species of dogs, B. vogeli and Ehrlichia canis respectively. The parasites occurred as single or mixed infections. Incidental infections that included B. gibsoni and Trypanosoma congolense were also detected using PCR. Although it was anticipated that zoonotic Ehrlichial infections of dogs would be detected, none were found. Babesia vogeli was reported for the first time in South Africa although, without any clinical significance. An outbreak of autochthonous canine babesiosis in the Netherlands was confirmed to have been caused by Babesia canis. Dermacentor reticulates was implicated in the transmission of the parasite to naïve dogs. Clinical significance of B. rossi and the novel Theileria sp. of dogs was evaluated. Babesia rossi was found to be of significant clinical importance. Genotyping of B. rossi isolates revealed that parasite genotypes could be correlated to disease phenotype. Additionally, specific genotypes could also be associated with fatalities. Although the characterization of the Theileria sp. in dogs was a first report in South Africa, the clinical significance of this infection in dogs appeared to be poorly resolved. The dangers of having non-endemic species becoming established in South Africa was highlighted with the incidental finding of a B. gibsoni infection in an imported dog. The results of this thesis have shown therefore that populations of dogs that live in tickendemic areas are exposed to single or multiple tick-borne pathogens. These pathogens continue to cause morbidity and mortality in susceptible dogs. Correct diagnosis (supported by molecular diagnostic tools) followed by appropriate treatment offers a better understanding and management of these tick-borne pathogens. Preventative measures should be fully evaluated and applied to prevent these tick-borne pathogens from adversely affecting the canine population in South Africa and elsewhere. / Thesis (PhD)--University of Pretoria, 2008. / Veterinary Tropical Diseases / unrestricted

No description available.

This study was conducted to understand the occurrence, diversity and epidemiology of tickborne diseases (TBDs) in a transhumant pastoral area of Karamoja Region, Uganda. We used participatory epidemiology (PE), involving focus group discussions (n = 24) with livestock keepers, 30 key informant interviews, review of previous surveillance data, clinical examinations, and laboratory confirmation of cases of TBDs, to define and prioritise cattle diseases, evaluate current control activities, and identify constraints to the control of TBDs. The livestock keepers regarded TBDs, particularly East Coast fever (ECF) and anaplasmosis, as the most important health problems in their cattle, based on morbidity and mortality rates, rates of transmission, treatment costs, difficulty in accessing the correct treatment, difficulty to control, and inadequate knowledge to manage the diseases. The main constraints to the control of TBDs were inadequate knowledge, inadequate veterinary services and limited availability of drugs. Hand picking of ticks was done by all pastoralists while hand spraying was done with under-strength acaricides, often at irregular intervals and with little acaricide wash. We determined the endemic status of TBDs in 20 randomly-selected cattle herds by estimating the proportion of annual ECF and anaplasmosis cases in different age groups of cattle using participatory approaches and clinical examinations, determining the diversity and abundance of ticks (161 cattle in 20 herds), and establishing the seroprevalence of antibodies to Theileria parva and Anaplasma marginale among cattle (n = 397 in 20 herds). Clinical examinations and informant interviews showed that TBDs affected all age groups of cattle. Two-thirds of the cattle were infested with moderate (37.3%, 11-50 ticks) to abundant (28.6%, > 50 ticks) numbers of ticks. Out of the 10,923 ticks collected, Rhipicephalus appendiculatus (54.4%) was the most abundant species followed by R. decoloratus (17.7%), Amblyomma variegatum (12%) and A. lepidum (11.6%). We collected tick species that are either rare in Uganda (A. lepidum, Hyalomma truncatum, A. gemma) or were not known to be present in the country (R. pulchellus). The true seroprevalence of antibodies to A. marginale was high (95.1%, 95% confidence interval [CI] 91.3% - 98.5%), while that of T. parva was low (16.5%, 95% CI 12.9% - 19.6%). We determined the presence of tick-borne haemoparasites among cattle and the prevalence of infections using reverse line blot (RLB) hybridisation and quantitative real-time polymerase chain reaction (qPCR), and characterised Theileria and Babesia species by sequence and phylogenetic analyses of the full-length 18S rRNA gene. The RLB hybridisation assay demonstrated the presence of tick-borne haemoparasites in most cattle (99.6%), mostly as mixed infections (97.5%). The most frequently-detected species were Theileria mutans (88.3%, 95% CI: 84.6-92.1%), A. marginale (73.8%: 68.3-79.2%), T. velifera (71.3%: 65.8-76.7%) and Anaplasma sp. Omatjenne (63.3%: 57.5-68.8%). Other virulent pathogens, namely Babesia bigemina (5.0%) and T. parva (2.9%), were also detected with RLB, but Ehrlichia ruminantium was not detected. The proportions of qPCR positive samples were 82.9% (A. marginale), 12.1% (A. centrale), 3.3% (T. parva), and 1.7% (E. ruminantium). Variations (5 to 9 nucleotides) in the 18S rRNA gene sequences of B. bigemina were identified as compared with previously published sequences. We assessed the 16S rRNA gene phylogeny of Anaplasma species from cattle and analysed the msp1α gene sequences of A. marginale to identify genotypes. There was genetic heterogeneity within A. marginale in cattle in the study area. Most A. marginale sequences (16/19) were closely related (99- 100% identities) and clustered with A. marginale strain Veld from cattle in South Africa, with strong bootstrap support, while three sequences clustered (100% identity) with A. marginale strain from Virginia, USA. The A. centrale sequences were closely related (100% identity) and clustered with the Israel vaccine strain. We found four different kinds of MSP1a tandem repeat sequences (UP39-F-M2-3) that correspond to one A. marginale strain unique to Uganda. One tandem repeat (UP39) was unique to Karamoja cattle. The findings from this study provide knowledge on the diversity and epidemiology of TBDs, which can be used to support diagnosis and strategic control of TBDs, and consequently improve cattle productivity and the livelihoods of pastoralists. More effective control and prevention measures against ticks and TBDs should urgently be implemented in the region. There is need to facilitate and promote immunisation of cattle against ECF. Given the scarcity of veterinarians in the region, the activities of community-based animal health workers [CAHWs] should be strengthened. The local governments in Karamoja Region should facilitate a more favourable business environment to attract private companies, who could scale-up veterinary drug supplies to the region. Pastoralists should be sensitised to improve their ecological knowledge of the seasonal activity of ticks for strategic tick control. / Thesis (PhD)--University of Pretoria, 2017. / Agricultural Technology and Agribusiness Advisory Services (ATAAS) / The National Agricultural Research Organisation (NARO) of Uganda / Veterinary Tropical Diseases / PhD / Unrestricted

Three studies relating to the vector competency of several species of ticks and <i>Simulium</i> spp. (blackflies) for <i>Ehrlichia risticii</i>, causative agent of Potomac horse fever (PHF) and <i>Amblyomma variegatum</i> for <i>Cowdria ruminantium</i>, causative agent of heartwater, are described. <i>Dermacentor variabilis, Rhipicephalus sanguineus, Amblyomma americanum and Ixodes scapularis</i> ticks were investigated for their ability to acquire and transmit PHF. Larval and nymphal ticks were exposed to <i>E. risticii</i> by feeding on mice inoculated with the organism. Molted exposed ticks were then allowed to feed on susceptible ponies or mice and were examined by light and electron microscopy. No evidence of transmission, either clinically or by seroconversion in mice or ponies was observed. Blackflies (<i>Simulium</i> spp.) were trapped in an area endemic for PHF and inoculated into mice in an attempt to demonstrate <i>I. risticii</i>. No evidence of seroconversion by mice to <i>E. risticii</i> was observed. Two laboratory colonies of <i>Amblyomma variegatum</i> ticks were investigated for their ability to acquire and transmit C. <i>ruminantium</i> and was febrile. Nymphs from both laboratory groups were simultaneously fed on a goat that had been infected with C. <i>ruminantium</i> and was febrile. Engorged nymphs from both groups were replete from feeding on three consecutive days. Nymphs from both groups were then incubated under identical conditions until molting. / Ph. D.

Tick-borne encephalitis is a vector-borne zoonotic disease with more than 12,000 annual clinical reported cases globally (WHO, 2021). Tick-borne encephalitis is caused by the flavivirus and transferred by Ixodes ricinus from roe deer to human and affects the central Nervous system. Climate change also increases tick-borne encephalitis incidences in Sweden and fluctuated considerably from year to year. A quantitative study design with secondary data was conducted to analyze the spatial and temporal pattern of Tick-borne encephalitis in Sweden from 1986-2020. The distribution of Tick-borne encephalitis within age and sex, along with other factors were also analyzed. The results showed that Tick-borne encephalitis with passing each year spread across the country. Male and age group, 50-59, have more incidence of Tick-borne encephalitis. Incidents were associated with climate conditions such as temperature and precipitation, which provided a favorable environment for Ixodes ricinus for its lifecycle activities, host searching, and disease transmission. Roe deer population, other vertebrates abundance, vaccination, population interest, economy, and land change are the critical factors that facilitate the disease incidence or control. People who visit forests for hunting, trekking, leisure, and professional activities without proper immunity and preventive measures are at risk to infect with Tick-borne encephalitis. The theory One health approach showed suitable performance for the control of this vulnerable climate zoonotic disease.

In Western Europe the incidence of tick-borne encephalitis (TBE) has increased over the last 30 years, coupled with changes in distribution. Modifications in the TBE enzootic cycle, through a combination of changes in temperature, vertebrate abundance and habitat suitability may have increased the risk of TBE in recent years. In Switzerland, analysis using satellite-derived climate data demonstrated that the environment of areas with TBE since the 1980s and areas that recently became endemic for TBE have become more similar between 2001 and 2009. This was coupled with an increase in April, May and June temperature, which could have affected the tick population and/or human exposure to ticks. Deer and boar abundance also changed in some cantons. In Germany, spatio-temporal modelling demonstrated the importance of temperature, vertebrate abundance and unemployment in the incidence and distribution of TBE between 2001 and 2009. Changes in TBE reporting, April, May and June temperature, vertebrate abundance and pesticide use may have contributed to increases in TBE in 1992 and 2001. Human exposure patterns, however, appear to be as important as the enzootic cycle in shaping the incidence of TBE, not only in determining the overall trend but also in interacting with the weekly, seasonal and yearly patterns of tick hazard to give the observed incidence. In Switzerland, in weeks with warm, sunny weather, human exposure to ticks is promoted and short-term increases in tick bites are seen. Human outdoor activity also shifts the seasonal pattern of tick bites, when compared with tick questing. There was no apparent increase in time spent in outdoor activities between the 1990s and 2000s in Italy, Germany and Austria, but survey data demonstrated that walking and hiking were already popular activities across Europe by the 1990s. The popularity of mushroom and berry foraging as a source of income in Latvia, Lithuania and Poland, coupled with the expense of vaccination, provide an inverse link between economic wellbeing and TBE risk. Correspondingly, in 2009, the economic recession was associated with an increase with TBE in these three countries.

This thesis is focused on the role of host cytoskeleton, primarily microtubules and microfilaments, during tick-borne encephalitis virus infection in human neuroblastoma cell line SK-N-SH and tick cell line IRE/CTVM19. The importance of cytoskeletal integrity and dynamics to the viral replication cycle were examined using specific chemical inhibitors showing the virus utilizes studied structures in both cell lines. Immunofluorescence microscopy revealed structural changes in the actin cytoskeleton during late infection in SK-N-SH cells. Moreover, differences in expression of cytoskeleton-associated genes in both cell lines were compared. Several genes with up-regulated expression in SK-N-SH cells were identified during late infection.

The proposed thesis contributes to the basic knowledge in tick (Ixodes ricinus) and tick-borne pathogens (Borrelia burgdorferi sensu lato, tick-borne encephalitis virus) ecology in particular studying the spatial distribution, host associations and its causes and consequences in Central European habitats.

Ticks are important vectors of a wide variety of pathogens including protozoa, bacteria and viruses. Many of the viruses transmitted by ticks are of medical or veterinary importance including tick-borne encephalitis virus (TBEV) and Crimean- Congo hemorrhagic fever virus causing disease in humans, and African swine fever virus and Nairobi sheep disease virus affecting livestock. Although several studies have elucidated tick antimicrobial mechanisms including cellular immune responses such as nodulation, encapsulation and phagocytosis and humoral immune responses such as the JAK/STAT pathway, complement-like proteins, antimicrobial peptides, lectin like pattern-recognition molecules and lysozymes, very little is known about the innate immune response of ticks towards viral infection. This study therefore aimed to identify molecules that might be involved in the response of ticks to viral infection. The hypothesis was that TBEV infection leads to changes in the expression of immunity-related transcripts and proteins in Ixodes spp. tick cells and that at least some of these might be antiviral. Ixodes scapularis-derived cell lines IDE8 and ISE6 were chosen since I. scapularis is currently the only tick species with a sequenced genome and an Ixodes ricinus-derived cell line, IRE/CTVM19, was used because I. ricinus is the natural vector of TBEV. Basic parameters required to study the responses of tick cells to infection were determined, including levels of virus infection, kinetics of virus replication and production, formation of replication complexes and uptake of dsRNA or siRNA. The cell lines IDE8, ISE6 and IRE/CTVM19 were infected with either of two tick-borne flaviviruses, TBEV and Langat virus (LGTV), or with the mosquito-borne alphavirus Semliki Forest virus (SFV). Infection was characterised using techniques including plaque assay, luciferase assay, immunostaining and conventional, confocal and electron microscopy. Two time points for transcriptomics and proteomics analysis of TBEVinfected IDE8 and IRE/CTVM19 cells were selected: day 2 post-infection (p.i.) when virus production was increasing and day 6 p.i. when virus production was decreasing. RNA and protein were isolated from TBEV-infected and mock-infected tick cells at days 2 and 6 p.i. and RNA-Seq and mass spectrometric technologies were used to identify changes in, respectively, transcript and protein abundance. Differential expression of transcripts was determined using the data analysis package DESeq resulting in a total of 43 statistically significantly differentially expressed transcripts in IDE8 cells and 83 in IRE/CTVM19 cells, while differential protein representation using Χ2 test statistics with Bonferroni correction in IDEG6 software resulted in 76 differentially represented proteins in IDE8 cells and 129 in IRE/CTVM19 cells. These included transcripts and proteins which could affect stages of the virus infection, including virus entry, replication, maturation and protein trafficking, and also innate immune responses such as phagocytosis, RNA interference (RNAi), the complement system, the ubiquitin-proteasome pathway, cell stress and the endoplasmic reticulum (ER) stress response. After verification of sequencing data by qRT-PCR, the ability of several of the identified transcripts or proteins to affect virus infection was determined by knockdown experiments in IDE8 and IRE/CTVM19 cells using wild type LGTV, LGTV replicons or TBEV replicons. Knockdown of genes encoding proteins including the ER chaperone gp96 and the heat-shock protein HSP90 resulted in increased virus production in both cell lines, hinting at an antiviral role. In contrast, knockdown of calreticulin, another ER chaperone, resulted in a decrease in virus production in IRE/CTVM19 cells but not in IDE8 cells, implying a requirement for virus production. This functional genomics approach has identified possible novel genes/proteins involved in the interaction between flaviviruses and tick cells and also revealed that there might be antiviral innate immune pathways present in ticks additional to the exogenous RNAi pathway.

This study examines, from a modelling point of view, the dynamics of infectious diseases in wildlife caused by macroparasites and by tick-borne infections. The overall aim was to investigate the important role played by parasite aggregation in the dynamics of both systems. For macroparasites we first developed some deterministic models that incorporate explicit mechanisms for generating aggregation in parasite distribution, specifically multiple infections and host heterogeneity. We explored the role of aggregation in host regulation and in determining a threshold value for parasite establishment. A large aggregation makes it more difficult for parasites both to regulate hosts, and to get established in a population at carrying capacity. Furthermore, the stabilization yielded by aggregation strongly depends on the mechanism that produces the aggregation. We then introduced some uncertainties into the host-macroparasite system, presenting an individual-based stochastic model that incorporated the same assumptions as the deterministic model. Stochastic simulations, using parameter values based on some real case studies, preserved many features of the deterministic model, like the average value of the variables and the approximate length of the cycles. An important difference is that, even when deterministic models yield damped oscillations, stochastic simulations yield apparently sustained oscillations. The amplitude of such oscillations may be so large as to threaten the parasites’ persistence. With respect to tick-borne diseases we presented a general model framework that incorporated both viraemic and non-viraemic routes of infections. We compute the threshold for disease persistence and study its dependence on the parameters and on host densities. The effects of tick aggregation and correlation between different tick stages on the host have both an important effect on infection persistence, if non-viraemic transmission occurred. In the case of Lyme Disease and Tick-borne Encephalitis (TBE) in Trentino (northern Italy) we showed some numerical results, using parameter estimates based on a detailed field study, and explored the effects of uncertainty on the endemic equilibrium of both diseases assuming only viraemic transmission for Lyme Disease while for TBE we permitted only non-viraemic transmission through co-feeding ticks. In conclusion we have examined the patterns and changes of aggregation in a number of contrasting systems and believe that these studies highlight both the importance of considering heterogeneities in modelling host-parasite interactions and, more specifically, modelling the biological mechanisms that produce aggregation in parasite distributions.