Ticks and Tick-Borne Pathogens in Domestic Animals, Wild Pigs, and Off-Host Environmental Sampling in Guam, USA

Background: Guam, a United States of America (USA) island territory in the Pacific Ocean, is known to have large populations of ticks; however, it is unclear what the risk is to wildlife and humans living on the island. Dog (Canis familiaris), cat (Felis catus), and wild pig (Sus scrofa) sentinels were examined for ticks, and environmental sampling was conducted to determine the ticks present in Guam and the prevalence of tick-borne pathogens in hosts.Methods and Results: From March 2019-November 2020, ticks were collected from environmental sampling, dogs, cats, and wild pigs. Blood samples were also taken from a subset of animals. A total of 99 ticks were collected from 27 environmental samples and all were Rhipicephalus sanguineus, the brown dog tick. Most ticks were collected during the dry season with an overall sampling success rate of 63% (95% CI: 42.4–80.6). 6,614 dogs were examined, and 12.6% (95% CI: 11.8–13.4) were infested with at least one tick. One thousand one hundred twelve cats were examined, and six (0.54%; 95% CI: 0.20–1.1) were found with ticks. Sixty-four wild pigs were examined and 17.2% (95% CI: 9.5–27.8) had ticks. In total, 1,956 ticks were collected and 97.4% of ticks were R. sanguineus. A subset of R. sanguineus were determined to be the tropical lineage. The other tick species found were Rhipicephalus microplus (0.77%), Amblyomma breviscutatum (0.77 %), and a Haemaphysalis sp. (0.51%). Blood samples from 136 dogs, four cats, and 64 wild pigs were tested using polymerase chain reaction (PCR) and DNA sequencing methods. Five different tick-borne pathogens with the following prevalences were found in dogs: Anaplasma phagocytophilum 5.9% (95% CI: 2.6–11.3); Anaplasma platys 19.1% (95% CI: 12.9–26.7); Babesia canis vogeli 8.8% (95% CI: 4.6–14.9); Ehrlichia canis 12.5% (95% CI: 7.5–19.3); Hepatozoon canis 14.7% (95% CI: 9.2–28.8). E. canis was detected in one cat, and no tick-borne pathogens were detected in wild pigs. Overall, 43.4% (95% CI: 34.9–52.1) of dogs had at least one tick-borne pathogen. Serological testing for antibodies against Ehrlichia spp. and Anaplasma spp. showed prevalences of 14.7% (95% CI: 9.2–28.8) and 31.6% (95% CI: 23.9–40), respectively.Conclusion: Four different tick species were found in Guam to include a Haemaphysalis sp., which is a previously unreported genus for Guam. Dogs with ticks have a high prevalence of tick-borne pathogens which makes them useful sentinels.

Analysis of Exotic Parasitic Infection in a Large Group of Imported Dogs Following an Animal Welfare Investigation

Background: Increased dog relocation can cause dissemination of parasite and vector populations and this is being recognised in countries across Northern Europe, including the UK. Data regarding the prevalence of exotic infections entering the UK would be beneficial to vets to help assess pets entering the UK from abroad, and to help calculate risk of establishment of novel pathogens. One such group of dogs were seized as part of an RSPCA-led animal welfare investigation and blood tested for exotic pathogens. Methods: As part of the RSPCA investigation, 151 dogs were removed from the site. Blood tests were performed for Babesia. canis, Ehrlichia canis, Hepatozoon canis and Leishmania infantum by PCR, Br.canis by antibody serology and D.immitis by blood antigen. In addition to pathogen screening, a serology titre for Rabies was measured for each dog. A clinical examination was performed by a veterinary surgeon and clinical signs recorded. Clinical signs data were analysed by the Fisher-Freeman-Halton Exact TestResults: Overall, 24% of the dogs tested positive for an infection. Two dogs were positive for Br.canis antibodies and had no clinical signs indicative of infection. Leishmania infantum was identified in 10.5% of dogs with all but two cases being diagnosed in dogs whose microchip originated in Romania. Hepatozoon canis was identified in 9.6% of dogs, all of which had a Romanian microchip. Dirofilaria immitis was identified in 4.1% of dogs, B.canis in 2.3% of dogs and E.canis was only present in 1.5% of dogs tested. Only four dogs were found to have co-infections. No significant association was found between the pathogens detected and presenting clinical signs. Conclusions: This study demonstrates a range of exotic pathogens entering the UK including Br.canis and demonstrates the importance of screening imported dogs. The emphasis for early recognition of exotic pathogens in imported dogs has relied on screening based on relevant clinical signs and the country of origin. While these factors are useful, this study demonstrated no significant association between presenting clinical signs and the pathogens carried.

A Unique Case of Fatal Coinfection Caused by Leptospira spp. and Hepatozoon canis in a Red Fox Cub (Vulpes vulpes)

Red foxes are the most abundant wild carnivore species in Europe commonly exposed to pathogenic Leptospira and Hepatozoon canis. Despite high seroprevalence, the clinical disease caused by these pathogens in red foxes has never been reported. Herein, we report the first-ever case of a fatal Leptospira spp. and H. canis coinfection in a two-month-old red fox cub with acute haemolytic anaemia, mild bronchopneumonia, intraalveolar haemorrhage, and tubulonephrosis. The presence of pathogenic Leptospira spp. DNA was detected in the kidney and lung tissues of the infected animal. In contrast to our previous knowledge, we believe that such fatal cases due to concomitant infection by Leptospira spp. and H. canis, especially in young animals, may commonly occur in nature. However, further studies are required to identify other factors that possibly contribute to the severity and the pathogenic effect of Leptospira spp. and H. canis infections in red foxes.

A community approach of pathogens and their arthropod vectors (ticks and fleas) in dogs of African Sub-Sahara

Background Arthropod-borne pathogens and their vectors are present throughout Africa. They have been well-studied in livestock of sub-Saharan Africa, but poorly in companion animals. Given the socio-economic importance of companion animals, the African Small Companion Animal Network (AFSCAN), as part of the WSAVA Foundation, initiated a standardized multi-country surveillance study. Methods Macro-geographic variation in ectoparasite (ticks and fleas) and pathogen communities in dogs was assessed through molecular screening of approximately 100 infested dogs in each of six countries (Ghana, Kenya, Nigeria, Tanzania, Uganda and Namibia), both in rural and urban settings. The most important intrinsic and extrinsic risk factors within the subpopulation of infested dogs were evaluated. Results Despite the large macro-geographic variation in the dogs screened, there was no consistent difference between East and West Africa in terms of the diversity and numbers of ticks. The highest and lowest numbers of ticks were found in Nigeria and Namibia, respectively. Most often, there was a higher diversity of ticks in rural habitats than in urban habitats, although the highest diversity was observed in an urban Uganda setting. With the exception of Namibia, more fleas were collected in rural areas. We identified tick species (including Haemaphysalis spinulosa) as well as zoonotic pathogens (Coxiella burnetti, Trypanosoma spp.) that are not classically associated with companion animals. Rhipicephalus sanguineus was the most abundant tick, with a preference for urban areas. Exophilic ticks, such as Haemaphysalis spp., were more often found in rural areas. Several multi-host ticks occurred in urban areas. For R. sanguineus, housing conditions and additional pets were relevant factors in terms of infestation, while for a rural tick species (Haemaphysalis elliptica), free-roaming dogs were more often infested. Tick occurrence was associated to the use of endoparasiticide, but not to the use of ectoparasiticide. The most prevalent tick-borne pathogen was Hepatozoon canis followed by Ehrlichia canis. High levels of co-parasitism were observed in all countries and habitats. Conclusions As dogs share a common environment with people, they have the potential to extend the network of pathogen transmission to humans. Our study will help epidemiologists to provide recommendations for surveillance and prevention of pathogens in dogs and humans. Graphical abstract

Occurrence of Babesia Species and Co-Infection with Hepatozoon canis in Symptomatic Dogs and in Their Ticks in Eastern Romania

Although the distribution of Babesia spp. and Hepatozoon canis is well known in Romania, there is still a marked lack of information in many places of the country. This study aimed to investigate the occurrence of these haemoparasites in symptomatic dogs and in their ticks in Iasi, eastern Romania. Ninety owned dogs were subjected to clinical examination at the Faculty of Veterinary Medicine of Iasi and all detectable ticks (58 ticks from 15 dogs) were collected. Additionally, 124 ticks collected from the coat of other dogs (no. = 23) were included. Three Babesia species were found in dogs: Babesia canis (94.4%), Babesia vogeli (3.3%), and Babesia rossi (2.2%). All the dogs resulted negative for H. canis. The ticks were identified as follows: Ixodes ricinus (64%), Dermacentor reticulatus (33%), and Rhipicephalus sanguineus group (3%). B. canis (Minimum Infection Rate; MIR = 81%), B. vogeli (MIR = 3%), and Babesia microti-like piroplasm (MIR = 1%) were found in ticks. Moreover, 15 ticks were positive for H. canis, 6 were co-infected with B. canis, and 1 with B. microti-like piroplasm. This is the first molecular identification of B. rossi in two symptomatic dogs from Romania, although further studies are needed to investigate the vector competence of other ticks from Europe.

Detection of Anaplasma phagocytophilum, Babesia odocoilei, Babesia sp., Borrelia burgdorferi Sensu Lato, and Hepatozoon canis in Ixodes scapularis Ticks Collected in Eastern Canada

Tick-borne pathogens cause infectious diseases that inflict much societal and financial hardship worldwide. Blacklegged ticks, Ixodes scapularis, are primary vectors of several epizootic and zoonotic pathogens. The aim sets forth the pathogens and their prevalence. In Ontario and Quebec, 113 I. scapularis ticks were collected from songbirds, mammals, including humans, and by flagging. PCR and DNA sequencing detected five different microorganisms: Anaplasma phagocytophilum, 1 (0.9%); Babesia odocoilei, 17 (15.3%); Babesia microti-like sp., 1 (0.9%); Borrelia burgdorferi sensu lato (Bbsl), 29 (26.1%); and Hepatozoon canis, 1 (0.9%). Five coinfections of Bbsl and Babesia odocoilei occurred. Notably, H. canis was documented for the first time in Canada and, at the same time, demonstrates the first transstadial passage of H. canis in I. scapularis. Transstadial passage of Bbsl and B. odocoilei was also witnessed. A novel undescribed piroplasm (Babesia microti-like) was detected. An established population of I. scapularis ticks was detected at Ste-Anne-de-Bellevue, Quebec. Because songbirds widely disperse I. scapularis larvae and nymphs, exposure in an endemic area is not required to contract tick-borne zoonoses. Based on the diversity of zoonotic pathogens in I. scapularis ticks, clinicians need to be aware that people who are bitten by I. scapularis ticks may require select antimicrobial regimens.

Potential Role of Hepatozoon canis in a Fatal Systemic Disease in a Puppy

Canine hepatozoonosis caused by Hepatozoon canis is an emerging disease in Europe. Clinical pictures vary from subclinical to life-threatening and non-specific clinical signs are predominantly reported. A 2-month-old female puppy originating from Southern Italy was adopted and moved to Northern Italy. Then, the dog was brought to a local veterinary practice for gastrointestinal signs, migrating lameness and pruritic dermatitis, and then tested positive for Hepatozoon spp. gamonts at the blood smear. After treatment with imidocarb dipropionate and doxycycline, the dog showed an initial clinical improvement. However, gastrointestinal signs recurred, and diffuse superficial pyoderma appeared on the thoracolumbar region, along with fever, lethargy, and weight loss. Eight months from the first onset of clinical signs, the dog was referred to a veterinary clinic and subjected to complete blood count, urine and fecal analysis, along with abdominal ultrasonography, whole-body CT and gastroduodenal endoscopy. Skin biopsies and blood samples were subjected to a PCR-coupled sequencing protocol, which scored both positive for H. canis. Alterations were consistent with a pre-existing cholangiohepatitis and multiple acquired extrahepatic shunts secondary to portal hypertension. The dog was euthanatized due to a clinical worsening two months later. The potential role of H. canis in the systemic disease observed, clinic-pathological findings and epizootiological implications are discussed.

Phylogeography and population differentiation in Hepatozoon canis (Apicomplexa: Hepatozoidae) reveal expansion and gene flow in world populations

Background Hepatozoon canis is a protozoan transmitted to dogs and other wild carnivores by the ingestion of ticks containing mature oocysts and is considered the principal cause of canine hepatozoonosis in the world. Here, we examined ribosomal RNA 18S gene sequence variation to determine the genetic differences and phylogeographic diversity of H. canis from various geographical areas around the world. Methods We used 550 publicly available sequences of H. canis from 46 countries to assess haplotype relationships, geographical structure, genetic diversity indices, and relationships among populations. We performed neutrality tests and pairwise comparisons of fixation index (FST) values between groups and pairwise comparisons of FST values between populations. To determine whether populations are structured, analyses of molecular variance (AMOVAs) and spatial analysis of molecular variance (SAMOVA) were performed. Results The dataset of H. canis yielded 76 haplotypes. Differentiation among populations indicated that there is no phylogeographical structure (GST = 0.302 ± 0.0475). Moreover, when samples were grouped by continents a significant FST was obtained, meaning that populations were genetically differentiated. The AMOVA showed that 57.4% of the genetic variation was explained by differences within populations when all locations were treated as a single group and revealed that there is no population structure when populations are grouped into two, three, and four groups (FCT, p > 0.05), suggesting that dispersal between populations is high. SAMOVA revealed significant FCT values for groups K = 5. The Tajima’s D and Fu’s Fs show that populations have undergone recent expansion, and the mismatch distribution analysis showed population expansion (multimodal distribution). Conclusions The current molecular data confirmed that H. canis does not show phylogeographic or population structure. The haplotypes exhibit low genetic differentiation, suggesting a recent expansion due to gene flow among populations. These results provide pivotal information required for future detailed population genetic analysis or to establish control strategies of this parasite. Graphical abstract

Incidence of Canine Tick Vectors and Molecular Detection of Haemoparasites in Vectors and Hosts

Background: Ticks are of great importance in transmission of various canine tick borne diseases. Several characteristics of ticks make them outstanding vectors of pathogenic agents, the wide host range and slow feeding habit along with tendency to feed on several hosts during life cycle ensures ample opportunity to acquire and transmit pathogens. Methods: This study focuses on status of canine tick vectors and molecular detection of haemoparasites in these ticks and their host, in and around Kolkata. The blood and tick samples were collected from Dog Ward, Department of Teaching Veterinary and Clinical Complex; Faculty of Veterinary and Animal Sciences, WBUAFS at Belgachia; Veterinary Clinic of Kolkata Police Dog Squad at Alipore; Veterinary Clinic of Barrackpore Police dog Squad and samples from stray dogs were also collected from inside the University campus through a period of one year (August, 2016 to July, 2017). Result: The tick infestation was recorded at 41% with Rhipicephalus sanguineus, being the only tick. Nine primer sets were used for detection of Babesia spp, Ehrlichia canis, Ehrlichia chaffensis, Babesia gibsoni, Hepatozoon canis, Mycoplasma haemocanis, Anaplasma platys and Theileria annae from the respective tick samples and blood sample of hosts. Tick samples were found positive for Babesia spp, Ehrlichia spp. and H. canis where as the corresponding blood samples were found positive for Babesia spp, Ehrlichia spp., Mycoplasma spp. and H. canis. This study conclusively provides evidence of high rates of incidence of haemoparasitic infection or canine tick borne diseases infection and tick infestation, with at least four haemoparasites infecting the dog population and at least one tick species (Rhipicephalus sanguineus) infesting the dogs in and around Kolkata.