Viruses of free-roaming and hunting dogs in Uganda show elevated prevalence, richness and abundance across a gradient of contact with wildlife.

Domestic dogs (Canis lupus familiaris) live with humans, frequently contact other animals and may serve as intermediary hosts for the transmission of viruses. Free-roaming dogs, which account for over 70% of the world's domestic dog population, may pose a particularly high risk in this regard. We conducted an epidemiological study of dog viromes in three locations in Uganda, representing low, medium and high rates of contact with wildlife, ranging from dogs owned specifically for traditional hunting in a biodiversity and disease 'hotspot' to pets in an affluent suburb. We quantified rates of contact between dogs and wildlife through owner interviews and conducted canine veterinary health assessments. We then applied broad-spectrum viral metagenomics to blood plasma samples, from which we identified 46 viruses, 44 of which were previously undescribed, in three viral families, Sedoreoviridae, Parvoviridae and Anelloviridae. All 46 viruses (100 %) occurred in the high-contact population of dogs compared to 63 % and 39 % in the medium- and low-contact populations, respectively. Viral prevalence ranged from 2.1 % to 92.0 % among viruses and was highest, on average, in the high-contact population (22.3 %), followed by the medium-contact (12.3 %) and low-contact (4.8 %) populations. Viral richness (number of viruses per dog) ranged from 0 to 27 and was markedly higher, on average, in the high-contact population (10.2) than in the medium-contact (5.7) or low-contact (2.3) populations. Viral richness was strongly positively correlated with the number of times per year that a dog was fed wildlife and negatively correlated with the body condition score, body temperature and packed cell volume. Viral abundance (cumulative normalized metagenomic read density) varied 124-fold among dogs and was, on average, 4.1-fold higher and 2.4-fold higher in the high-contact population of dogs than in the low-contact or medium-contact populations, respectively. Viral abundance was also strongly positively correlated with the number of times per year that a dog was fed wildlife, negatively correlated with packed cell volume and positively correlated with white blood cell count. These trends were driven by nine viruses in the family Anelloviridae, genus Thetatorquevirus, and by one novel virus in the family Sedoreoviridae, genus Orbivirus. The genus Orbivirus contains zoonotic viruses and viruses that dogs can acquire through ingestion of infected meat. Overall, our findings show that viral prevalence, richness and abundance increased across a gradient of contact between dogs and wildlife and that the health status of the dog modified viral infection. Other ecological, geographic and social factors may also have contributed to these trends. Our finding of a novel orbivirus in dogs with high wildlife contact supports the idea that free-roaming dogs may serve as intermediary hosts for viruses of medical importance to humans and other animals.

Comparison of the efficacy of crude methanolic extracts of Cassia occidentalis and Euphorbia hirta with levamisole-HCL against gastrointestinal nematodes of economic importance to goat production in Uganda.

Natural compounds from medicinal plants provide safe and sustainable alternatives to synthetic anthelmintics. In this study, we assessed in vitro and in vivo anthelmintic activity of Cassia occidentalis (NH-A) and Euphorbia hirta (NH-B) and compared it with levamisole-HCl. The shoots of NH-A and whole plant of NH-B were used to prepare extracts using 70% methanol which were used in the in vitro and in vivo assays. In vitro assays of crude methanolic extracts (CMEs) of NH-A and NH-B on larvae of mixed gastrointestinal nematodes (GINs) of goats revealed mortalities of 95.7% (at 100 mg ml-1) and 98.1% (at 50 mgml-1) 24 h postexposure. In vivo assays of NH-A administered orally at doses of 100, 300, 900, and 2700 mg kg-1 bwt revealed dose- and time-dependent anthelmintic effects in goats experimentally infected with mixed species of GINs. NH-B exhibited similar properties when administered at doses of 50, 100, 200, and 400 mg kg-1 bwt. Both NH-A (900 mg kg-1 bwt) and levamisole (7.5 mg kg-1 bwt) achieved a 100% reduction in fecal egg count per gram (EPG) on day 21 and day 14 respectively posttreatment. NH-B (400 mg kg-1 bwt) achieved 93.1% and 86.1% reduction in fecal EPG 7 and 14 days postexposure respectively compared with 88.2% and 82.3% reduction with levamisole-HCl 7 and 14 days postexposure. Our results show that extracts of both plants can disrupt helminth lifecycles by suppressing the egg-laying capacity in adult worms but also kill their infective larvae. Future studies should aim at establishing synergies or antagonisms between the two plant extracts and further development for control of helminths in goats.

Genetic mutations in sodium channel domain II and carboxylesterase genes associated with phenotypic resistance against synthetic pyrethroids by Rhipicephalus (Boophilus) decoloratus ticks in Uganda.

We previously reported emergence of super synthetic pyrethroid (SP) resistant Rhipicephalus (Boophilus) decoloratus ticks in Uganda. This study investigated the genetic basis of phenotypic resistance against SP in R. (B.) decoloratus and sought to identify novel diagnostic mutations for rapid detection of SP resistance in the above tick species. Genomic DNA was extracted from pooled larvae of 20 tick populations (19 of known SP susceptibility and 1 unknown susceptibility). The voltage sensitive sodium channel (VSSC) domain II S4-5 linker (SP target) and partial carboxylesterase (SP metabolizing enzyme) genes were amplified by PCR, cloned and sequenced. The resultant sequences were analyzed to determine single nucleotide polymorphisms (SNPs) associated with phenotypic resistance in the various tick populations investigated. Novel SNPs that introduced Eco RI and Eco RII restriction sites in carboxylesterase gene were identified in silco and validated with restriction fragment length polymorphism (RFLP) against 18 tick populations of known SP susceptibility. The study identified a super knock down resistance (kdr) mutation T58C in R. (B.) decoloratus VSSC associated with stable SP resistance. We further identified multiple nonsynonymous mutations in carboxylesterase of SP resistant ticks; one of which conferred novel EcoRII (G195C) restriction site for PCR-RFLP detection of SP resistance. In conclusion, this study is the first to report super kdr mutation in sodium channel domain II and multiple mutations in carboxylesterase genes that may concurrently mediate stable resistance against synthetic pyrethroids in R. (B.) decoloratus ticks from Uganda. The Eco RII based PCR-RFLP is a useful tool for rapid detection of stable SP resistant R. (B.) decoloratus ticks.

Descriptive analyses of knowledge, attitudes, and practices regarding rabies transmission and prevention in rural communities near wildlife reserves in Uganda: a One Health cross-sectional study.

BACKGROUND: Despite urban (domestic dog) rabies cycles being the main target for rabies elimination by 2030, sylvatic (wildlife) rabies cycles can act as rabies spillovers especially in settlements contiguous to wildlife reserves. Rural communities next to wildlife reserves are characterized by unique socio-demographic and cultural practices including bat consumption, hunting for bushmeat, and non-vaccination of hunting dogs against rabies among others. This study aimed to compare the knowledge, attitudes, and practices (KAPs) related to rabies transmission and prevention in the three districts of Uganda; (1) Nwoya, neighboring Murchison Falls National Park (MFNP) in the north, (2) Kamwenge neighboring Kibaale National Park (KNP), Queen Elizabeth National Park (QENP) and Katonga Game Reserve (KGR) in the west, and (3) Bukedea, neighboring Pian Upe Game Reserve (PUGR) in the east of Uganda. METHODS: A community-based cross-sectional survey was conducted in settlements contiguous to these wildlife reserves. Using a semi-structured questionnaire, data were collected from 843 households owning dogs and livestock. Data were collected between the months of January and April 2023. Stratified univariate analyses by district were carried out using the Chi-square test for independence and Fisher's exact test to compare KAPs in the three study districts. RESULTS: The median age of study participants was 42 years (Q1, Q3 = 30, 52) with males comprising the majority (67%, n = 562). The key findings revealed that participants from the Nwoya district in the north (MFNP) had little knowledge about rabies epidemiology (8.5%, n = 25), only 64% (n = 187) of them knew its signs and symptoms such as a rabid dog presenting with aggressiveness and showed negative attitudes towards prevention measures (15.3%, n = 45). Participants in the Kamwenge district-west (KNP, QENP, and KGR) had little knowledge and negative attitude towards wildlife-human interaction pertaining to rabies transmission and prevention especially those with no or primary level of education (20.9%, n = 27) while participants from Bukedea in the east (PUGR) had remarkedly poor practices towards rabies transmission, prevention, and control (37.8%, n = 114). CONCLUSIONS: Rabies from sylvatic cycles remains a neglected public health threat in rural communities surrounding national parks and game reserves in Uganda. Our study findings highlight key gaps in knowledge, attitudes, and practices related to rabies transmission and prevention among such communities. Communication and action between veterinary services, wildlife authority, public health teams, social science and community leaders through available community platforms is key in addressing rabies among the sympatric at-risk communities in Uganda.

Estimating the potential of beekeeping to alleviate household poverty in rural Uganda.

Robust evidence underpinning the role of beekeeping in poverty alleviation is currently lacking. This study estimated the production potential for beekeepers in Northern Uganda by quantifying current production assets (equipment and knowledge) and impact on rural income streams range of proposed interventions. Intervention scenarios evaluated the economic benefits to be derived from different hive types combined with year-round provision of a nectar source (Calliandra calothyrsus) planted at varying density. Findings show that the type and number of beehive combinations used influenced the amount of revenue streams generated by the beekeepers. Addition of 20 log hives increased incomes 10 times, 20 KTBs increased revenues 16 times and Langstroth 18 times. Adding Calliandra trees as a forage source to the baseline scenario yielded revenues up to 17.6 times higher than the baseline. Implying that good management plus the introduction of a reliable nectar source, to off-set dry season challenges (absconding), could improve beekeeping productivity in Northern Uganda. Further research is required to validate in situ the impact of modelled scenarios on both honey yield and other ecosystem service benefits.

Evidence-based tick acaricide resistance intervention strategy in Uganda: Concept and feedback of farmers and stakeholders.

The emergence of multi-acaricide resistant ticks has led to unprecedented level of acaricide failure in central and western Uganda. In the absence of a national acaricide resistance management strategy, the country's dairy sector is threatened by upsurge of ticks and tick-borne diseases. In this study, we developed a short-to-medium-term intervention approach called Evidence-Based Acaricide Tick Control (EBATIC): Identify, Test, Intervene and Eradicate (IT-IE). Furthermore, the perception of 199 farmers and extension workers, 12 key informants in four districts and 47 stakeholders in the animal industry in Uganda were assessed using semi-structured questionnaires. We report that the establishment of a specialized laboratory is pivotal in identifying and testing (IT) acaricide resistant ticks for prompt intervention and eradication (IE). The laboratory test results and the farm tick control gaps identified are very important in guiding acaricide resistance management strategies such as evidence-based acaricide rotation, development and dissemination of extension materials, training of farmers and extension workers, and stakeholders' engagement towards finding sustainable solutions. All the 47 stakeholders and 91.0% (181/199) of the farmers and extension workers reported that the EBATIC approach will help in solving the tick acaricide resistance crisis in Uganda. Similarly, all the 12 key informants and 92.5% (184/199) of the farmers and extension workers suggested that the EBATIC approach should be sustained and rolled out to other districts. The EBATIC stakeholders' dialogue generated both short-to-medium and long-term strategies for sustainable management of tick acaricide resistance in the country. Overall, the positive feedback from farmers, district veterinarians and stakeholders in the animal industry suggest that the EBATIC approach is a useful proof-of-concept on scalable intervention pathway against tick acaricide resistance in Uganda with possibility of adoption in other African countries.

C190A knockdown mutation in sodium channel domain II of pyrethroid-resistant Rhipicephalus appendiculatus.

This study investigated the target site mutations in the partial sequence of voltage-sensitive sodium channel (VSSC) domain II of synthetic pyrethroid (SP)-resistant Rhipicephalus appendiculatus. Genomic DNA was extracted from seven tick populations (two susceptible and five resistant) collected from central, eastern and southwestern Uganda. The PCR amplicons of the VSSC domain II were cloned and sequenced to determine novel single nucleotide polymorphisms (SNP). A non-synonymous mutation C78 A corresponding to C190 A was found in all the five SP-resistant ticks. The C78 A mutation led to amino acid substitution from leucine to isoleucine (L21I) which was previously reported to confer knockdown (kdr) mutation in R. (Boophilus) microplus. The genetic confirmation of SP-resistant R. appendiculatus in central and southwestern Uganda calls for an urgent strategy for controlling the ticks.

Chemical tick control practices in southwestern and northwestern Uganda.

Tick acaricide failure is one of the leading challenges to cattle production in Uganda. To gain an understanding into the possible drivers of acaricide failure, this study characterized the current chemical tick control practices in the southwestern (Mbarara, Mitooma and Rukungiri districts) and northwestern (Adjumani district) regions of Uganda. A total of 85 farms participated in a survey that utilized a semi-structured questionnaire. Moreover, ticks were collected to determine the most common species on the farms. Tick acaricide failure was mainly encountered in the districts where 95% (60/63) of the farms reared exotic cattle (dairy cross-breeds) under a paddocking (fenced) system. In the northwestern region, local cattle were reared in communal grazing areas. All farms used chemical acaricides for tick control, predominantly amidine (amitraz) (48%, 41/85) and co-formulated organophosphates and pyrethroids (38%, 32/85). The spraying method was the most common (91%, 77/85) acaricide application technique, with cattle crush (81%, 69/85) as a common means of physical restraint. Less than optimal tick control practices encountered included use of substandard equipment for spraying, inappropriate dilutions, frequent interaction between animals in neighboring farms despite lack of synchronized chemical tick control and malpractices in acaricide rotation. Only Rhipicephalus appendiculatus and R. (Boophilus) decoloratus ticks were found in the southwestern region, where 51% (32/63) of the farmers used high acaricide concentrations above the manufacturers' recommendation. Farmers in the northwestern region used 2.2 times less acaricide volume per cattle than those in the southwestern region, and more diverse tick species were encountered. Toxic effects of acaricide to cattle and workers were reported by 13% (11/85) and 32% (27/85) of the respondents, respectively. All 27 cases of human acaricide toxicity reported were from the southwestern region. Overall, our findings may inform strategies for more prudent chemical tick control and safe acaricide handling to benefit animal welfare, food safety and public health.

Molecular epidemiology of Babesia species, Theileria parva, and Anaplasma marginale infecting cattle and the tick control malpractices in Central and Eastern Uganda.

East Coast fever, babesiosis, and anaplasmosis are the major tick-borne diseases affecting cattle productivity in Uganda. The emergence of acaricide-resistant ticks is suspected to have caused a rise in hemoparasites. This study sought to detect and characterize hemoparasites among farms in acaricide-failure hotspots of central as compared to the acaricide-failure naïve areas in Eastern Uganda. Nested PCR assays were performed to determine the prevalences of Babesia bovis, Babesia bigemina, Theileria parva, and Anaplasma marginale in cattle blood samples sourced from randomly selected farms. Randomly selected isolates were sequenced to determine the genetic diversity of the parasites using the following marker genes: B. bovis spherical body protein 4, B. bigemina rhoptry-associated protein 1a, T. parva 104 kDa microneme-rhoptry antigen, and A. marginale major surface protein 5. Furthermore, partially and fully engorged adult ticks were collected for taxonomy, and tick-control practices were assessed using a semi-structured questionnaire. The prevalences of B. bigemina, T. parva, and A. marginale in cattle were 17.2, 65.1, and 22.0%, and 10.0, 26.5, and 3% in the central and eastern region, respectively. Whilst, B. bovis was not detected in the farms involved. The sequences for B. bigemina, T. parva, and A. marginale from the central region showed 99% identity with those from the eastern region. Of the 548 ticks collected, 319, 147, 76, and 6 were Rhipicephalus (Boophilus) decoloratus, Rhipicephalus appendiculatus, Amblyomma variegatum, and Rhipicephalus evertsi evertsi, respectively. The Rhipicephalus ticks were more abundant in the central region, whereas A. variegatum ticks were more abundant in the eastern region. Tick control malpractices were found in both Central and Eastern Uganda, and 42 of the 56 surveyed farms lacked appropriate restraining facilities and so they utilized either ropes or a 'boma' (enclosure). In summary, B. bigemina, T. parva, A. marginale and their co-infections were more prevalent in the central than eastern region; even though, tick control malpractices were observed in both regions. Therefore, an urgent tick and TBD control strategy is needed.

Demography and health of "village dogs" in rural Western Uganda.

"Village dogs" in developing economies are assumed to be heavily burdened by infectious disease. We followed a cohort of 61 village dogs in rural western Uganda prospectively for fifteen months to measure changes in health and demographic outcomes, and to examine risk factors for morbidity and mortality. The mean (±standard deviation) number of dogs per household was 2.4 (±2.0), of which 56.0% were male and 44.0% female. For females, average age at first estrus was 1.7 (±0.6)years with a mean litter size of 3.8 (±1.5). In the first, second and third parities, average puppy mortality per litter was 3.2 (±2.5), 2.4 (±2.1) and 3.4 (±2.9), respectively. The main causes of morbidity and mortality were infectious disease (46.1%), culling (euthanasia) by owners (30.8%), and attacks by baboons, Papio anubis (23.1%). Cox proportional hazard regression showed that a clinical diagnosis of anemia significantly predicted morbidity (HR=4.3 (95% CI: 1.1-17.8); p<0.05), and younger age significantly predicted mortality (HR=3.6 (95% CI: 1.2-10.6); p<0.05). Our results indicate that infectious disease is indeed important to the health and survival in village dogs in this setting, but that cultural practices related to ownership and interactions with wildlife also contribute substantially to morbidity and mortality.

Prevalence and Risk Factors Associated with Hemoparasites in Cattle and Goats at the Edge of Kibale National Park, Western Uganda.

Livestock production is a major sector of the Ugandan economy. Ugandan ruminant livestock (principally cattle and goats) are susceptible to hemoparasites that can cause serious clinical disease and production losses. Kibale National Park, in western Uganda, is a protected forest ecosystem surrounded by small-scale farms where cattle and goats are raised. We conducted a cross-sectional study of cattle and goats in this area and diagnosed hemoparasite infections by microscopy. We collected data on animal characteristics and management practices to assess risk factors associated with infection. We studied 186 cattle and 317 goats from 20 villages, including 16 villages directly adjacent to Kibale and 4 villages ≥3 km from the park boundary. Hemoparasites detected in cattle and goats were of the genera Theileria, Anaplasma, and Trypanosoma with a prevalence of 15.1%, 1.6%, and 4.3% respectively in cattle, and 10%, 6.0%, and 0.0%, respectively in goats. Trypanosomes infected approximately 8% of cattle in villages bordering Kibale but were never detected in cattle in "control" villages ≥3 km from the park. Trypanosomes were approximately 7 times more likely to infect animals in households that did not provide veterinary care to their animals than in households that provided routine veterinary care. Within cattle, Theileria infections were approximately 7 times more likely to occur in cross-bred cattle than in indigenous pure breeds. Anaplasma infections were approximately 3.5 times more likely to occur in cattle than in goats (no goats were diagnosed with Trypanosoma infection). These data suggest that proximity to the park, provision of veterinary care, and breed are significant risk factors for hemoparasites in this population of ruminants, and that, in general, cattle are more susceptible than goats.

Retrospective study on cattle and poultry diseases in Uganda.

Cattle and poultry enterprises are among the major contributors to food security and socioeconomic empowerment of households in Uganda. However, various diseases constrain their productivity. A two-year retrospective study between April 2012 and March 2014 was conducted using records for cattle and poultry diseases diagnosed at the Central Diagnostic Laboratory (CDL) to determine prevalent diseases in Uganda. The laboratory received 836 samples from poultry (36.3%) and cattle (63.7%). Of the 836 samples, 47.5% had a definitive diagnosis of disease causation. Most of the cattle and poultry diseases diagnosed were protozoan diseases (39.3%) followed by bacterial (21.4%), viral (17.1%), helminthiasis (11.1%), nutritional diseases (4%) and others (7.1%). For poultry, viral diseases (29.5%) and protozoan diseases (27.1%) especially newcastle disease (44.3%) and coccidiosis (100%) respectively, were the most diagnosed. While for cattle, hemo-protozoan parasites (52.1%) were the most prevalent, of which 92.9% were east coast fever infection. Bacterial infection (20.5%) in cattle were the second most diagnosed diseases and mastitis was the most diagnosed (46.2%). In summary, coccidioisis, collibacillosis, newcastle disease, gumboro disease, and avian helminthiasis were the most prevalent poultry diseases while in cattle, east coast fever, helminthiasis, mastitis, brucellosis and rabies were the most frequently diagnosed diseases. This study has identified the major diseases that hinder poultry and cattle production in Uganda. The data generated by CDL could be used for surveillance, monitoring and designing strategic interventions for control of poultry and cattle diseases in Uganda.

Emergence of multi-acaricide resistant Rhipicephalus ticks and its implication on chemical tick control in Uganda.

BACKGROUND: Acaricide failure has been on the rise in the western and central cattle corridor of Uganda. In this study, we identified the tick species associated with acaricide failure and determined their susceptibility to various acaricide molecules used for tick control in Uganda. METHODS: In this cross sectional study, tick samples were collected and identified to species level from 54 purposively selected farms (from 17 districts) that mostly had a history of acaricide failure. Larval packet test was used to screen 31 tick populations from 30 farms for susceptibility at discriminating dose (DD) and 2 × DD of five panels of commercial acaricide molecules belonging to the following classes; amidine, synthetic pyrethroid (SP), organophosphate (OP) and OP-SP co-formulations (COF). Resistance was assessed based on World Health Organization criteria for screening insecticide resistance. RESULTS: Of the 1357 ticks identified, Rhipicephalus (Rhipicephalus) appendiculatus and Rhipicephalus (Boophilus) decoloratus were the major (95.6%) tick species in farms sampled. Resistance against SP was detected in 90.0% (27/30) of the tick populations tested. Worryingly, 60.0% (18/30) and 63.0% (19/30) of the above ticks were super resistant (0% mortality) against 2 × DD cypermethrin and deltamethrin, respectively. Resistance was also detected against COF (43.3%), OP chlorfenvinphos (13.3%) and amitraz (12.9%). In two years, 74.1% (20/27) of the farms had used two to three acaricide molecules, and 55.6% (15/27) rotated the molecules wrongly. Multi-acaricide resistance (at least 2 molecules) was detected in 55.2% (16/29) of the resistant Rhipicephalus ticks and significantly associated with R. decoloratus (p = 0.0133), use of both SP and COF in the last 2 years (p < 0.001) and Kiruhura district (p = 0.0339). Despite emergence of amitraz resistance in the greater Bushenyi area, it was the most efficacious molecule against SP and COF resistant ticks. CONCLUSION: This study is the first to report emergence of super SP resistant and multi-acaricide resistant Rhipicephalus ticks in Uganda. Amitraz was the best acaricide against SP and COF resistant ticks. However, in the absence of technical interventions, farmer-led solutions aimed at troubleshooting for efficacy of multitude of acaricides at their disposal are expected to potentially cause negative collateral effects on future chemical tick control options, animal welfare and public health.