Prevalence of gastro-intestinal and haemoparasitic infections among domestic cats of Kerala.

The population of domesticated cats has drastically increased during the last decades. With the recently identified rise in cat population an upsurge in the parasitic infections associated with domestic cats is evident. A total of 122 domestic cats were screened for gastro-intestinal and haemoparasites. Screening for gastro-intestinal parasites revealed an overall prevalence of 19 per cent (23/122). Ancylostoma spp. was the major gastro-intestinal parasite noticed (61 per cent) followed by Toxocara cati (13.04 per cent), Isospora spp. (8.7 per cent), Diphyllobothrium latum (4.35 per cent) and mixed infection of these (13 per cent). Blood smear examination revealed Cytauxzoon spp. in three cats (2.46 per cent) and Babesia spp. in two cats (1.6 per cent). None of the cats were positive for gamonts of Hepatozoon spp. Molecular analysis revealed Hepatozoon spp. infection in seven cats (5.7 per cent), Cytauxzoon spp. in 29 cats (23.8 per cent) and Babesia spp. in two cats (1.6 per cent). Present study highlights the inevitability of molecular techniques in the identification of haemoparasites. Establishment of proper preventive measures are required to control parasitic infection among domestic cats.

Molecular detection and characterization of vector-borne agents in common opossums (Didelphis marsupialis) from northeastern Brazil.

Opossums are synanthropic marsupials able to interchange among wild, peri­urban and urban environments, playing an epidemiologically important role as hosts for emerging pathogens and ectoparasites of relevance in public health. The present study aimed to detect and molecularly characterize vector-borne agents in a population of common opossums (Didelphis marsupialis) from the Island of São Luís do Maranhão, northeastern Brazil. Of the 45 animals analyzed, one (2.22%) was positive in the nested PCR assay based on the 18S rRNA gene of piroplasmids. The obtained sequence was phylogenetically positioned in a clade containing sequences of Babesia sp. previously detected in Didelphis aurita, Didelphis albiventris and associated ticks from Brazil. Eight (17.77%) samples were positive in PCR for Ehrlichia spp. based on the dsb gene; four samples were sequenced and positioned into a new clade, sister to E. minasensis and Ehrlichia sp. clade detected in Superorder Xenarthra mammals. No samples tested positive in the screening PCR assays based on the 16S rRNA gene of Anaplasma spp. Two samples were positive in the qPCR for Bartonella spp. based on the nuoG gene. Seven animals (15.56%) were positive in the nPCR based on the 16S rRNA gene of hemoplasmas. Of these, three were positive in a PCR based on the 23S rRNA gene. The phylogenies based on both 16S rRNA and 23S rRNA genes corroborated to each other and positioned the sequences in the same clade of hemoplasmas previously detected in D. aurita and D. albiventris sampled in Brazil. Finally, three (6.66%) animals were positive in the PCR for Hepatozoon spp.; the obtained 18S rRNA sequence was positioned into the H. felis clade.The present study showed, for the first time, the circulation of piroplasmids, Hepatozoon spp., Ehrlichia spp., hemoplasmas and Bartonella spp. in D. marsupialis sampled in northeastern Brazil, with description of putative novel genotypes of Ehrlichia and Hepatozoon and copositivity by different vector-borne agents. The present work consolidates the "South American Marsupialia" piroplasmid clade, adding one more genotype of Babesia sp. to this clade.

Arthropod-Borne Pathogens in Wild Canids.

Wild canids, as well as other wild animal species, are largely exposed to bites by ticks and other hematophagous vectors where the features favoring their presence and spread are found in wooded and semi-wooded areas. Much of the information about arthropod-borne infections concerns domestic and companion animals, whereas data about these infections in wild canids are not exhaustive. The present study is a narrative review of the literature concerning vector-borne infections in wild canids, highlighting their role in the epidemiology of arthropod-borne bacteria and protozoa.

Novel Ehrlichia and Hepatozoon genotypes in white-eared opossums (Didelphis albiventris) and associated ticks from Brazil.

White-eared opossums (Didelphis albiventris) are well adapted to anthropized areas. The increased contact with domestic animals and humans mediates the transmission of arthropod-borne pathogens. Despite the worldwide occurrence of tick-borne Anaplasmataceae and Hepatozoidae species in a variety of vertebrates, few studies reported serological evidence or molecular detection of theses agentes in marsupials. Up to now, while Ehrlichia/Anaplasma spp. have only been detected in marsupials from Brazil, Hepatozoon spp. have been reported in marsupials from Chile, Australia and Brazil. The present work aimed to investigate, using molecular techniques and blood smear analysis, the presence of Ehrlichia spp., Anaplasma spp., and Hepatozoon sp. in the blood and ticks collected from D. albiventris in urban forest fragments from midwestern Brazil. Between May and December 2017, 43 D. albiventris (27 males and 16 females) were captured for blood and tick collection in the city of Campo Grande, state of Mato Grosso do Sul, midwestern Brazil. Ticks (46 Amblyomma dubitatum nymphs and 24 Amblyomma spp. larvae) were collected from 14 out 43 (32.5%) of the white-eared opossums. Panoptic-stained blood smears were performed using peripheral blood (tail tip) of the captured opossums. DNA extracted from blood and tick samples were subjected to PCR/qPCR assays for Anaplasmataceae agents (rrs, gltA, groEL, sodB, and dsb genes, and 23S-5S intergenic region) and Hepatozoon spp. (18S rRNA gene), followed by Sanger sequencing, BLASTn and phylogenetic analyses. An inclusion resembling Ehrlichia morulae was found in a white-eared opossum's monocyte from a blood smear stained with Panoptic. Five (11.63% [5/43]) white-eared opossums' blood samples and 7 (25% [7/28]) tick samples (2 pools of Amblyomma spp. larvae and 5 pools of A. dubitatum nymphs) were positive for Anaplasmataceae via a PCR assay targeting the conserved rrs gene. Phylogenetic analysis based on the rrs gene positioned three sequences obtained from opossums and ticks together as a subclade within the Ehrlichia canis clade. However, all samples were negative in a qPCR assay specific for E. canis based on the dsb gene. Phylogenetic analyses positioned the gltA and 23S-5S ITS sequences obtained from opossums' blood samples in a separate clade from the other validated Ehrlichia species. One (2.3% [1/43]) opossum blood sample was positive for the 18S rRNA gene of Hepatozoon sp. The phylogenetic analysis positioned the Hepatozoon sp. sequence obtained from a D. albiventris specimen in a clade with a sequence previously detected in a black storm petrel (Oceanodroma melania) from Mexico. All the other sequences of Hepatozoon sp. previously detected in marsupials from Brazil were positioned in a separated clade. The present work showed the occurrence of putative novel genotypes of Ehrlichia sp. and Hepatozoon sp. in white-eared opossums and associated A. dubitatum ticks from midwestern Brazil.

Molecular detection of tick-borne pathogens in Rhipicephalus sanguineus sensu stricto collected from dogs in the steppe and high plateau regions of Algeria.

Epidemiology and distributions of canine tick-borne diseases as well as their veterinary and zoonotic significance are poorly understood in Algeria. The present study describes a molecular investigation of important tick-borne pathogens in Rhipicephalus sanguineus sensu stricto collected from domestic dogs in steppe and high plateau areas of central and eastern Algeria. In total, 1,043 ticks were collected from 147 dogs, including 756 ticks from 124 dogs in the steppe region of Djelfa and 287 ticks from 23 dogs in the high plateau area of Bordj Bou Arreridj. Ticks were divided into 384 pools (309 pools from Djelfa and 75 pools from Bordj Bou Arreridj) and tested for genomic materials of Crimean-Congo hemorrhagic fever virus (CCHFV) as well as DNA for Anaplasma phagocytophilum, Anaplasma platys, Ehrlichia canis, Rickettsia spp., Babesia spp., and Hepatozoon spp. using PCR, sequencing and phylogenetic analysis. Hepatozoon spp. was most prevalent, with 160 positive pools (41.7%), and 12 of these were sequenced and identified as Hepatozoon canis. Babesia spp. was detected in 50 samples (13.0%), of which 11 were sequenced and identified as Babesia vogeli. A. platys and E. canis were detected in 92 (24.0%) and 15 (3.9%) of tested samples, respectively. Rickettsia spp. were detected in 24 (6.3%) samples, including 11 samples identified as R. massiliae, 6 samples identified as R. conorii conorii, and 7 samples could not be identified to species level. All 384 pools tested negative for CCHFV and A. phagocytophilum. In addition to detection of R. conorii conorii, R. massiliae, and E. canis, the present study provides the first molecular data for occurrence of A. platys, B. vogeli, and H. canis in Rh. sanguineus s.s. infesting dogs in Algeria. Further large scale studies should be conducted to better understand the epidemiology, distributions, and importance of canine tick-borne pathogens in Algeria.

First Record of Hepatozoon spp. in Alpine Wild Rodents: Implications and Perspectives for Transmission Dynamics across the Food Web.

Among the Apicomplexa parasites, Hepatozoon spp. have been mainly studied in domestic animals and peri-urban areas. The epidemiology of Hepatozoon spp. is poorly investigated in natural systems and wild hosts because of their scarce veterinary and economic relevance. For most habitats, the occurrence of these parasites is unknown, despite their high ecosystemic role. To fill this gap for alpine small mammals, we applied molecular PCR-based methods and sequencing to determine the Hepatozoon spp. in 830 ear samples from 11 small mammal species (i.e., Apodemus, Myodes, Chionomys, Microtus, Crocidura and Sorex genera) live-trapped during a cross-sectional study along an altitudinal gradient in the North-Eastern Italian Alps. We detected Hepatozoon spp. with an overall prevalence of 35.9%. Two species ranging from 500 m a.s.l. to 2500 m a.s.l. were the most infected: My. glareolus, followed by Apodemus spp. Additionally, we detected the parasite for the first time in another alpine species: C. nivalis at 2000-2500 m a.s.l. Our findings suggest that several rodent species maintain Hepatozoon spp. along the alpine altitudinal gradient of habitats. The transmission pathway of this group of parasites and their role within the alpine mammal community need further investigation, especially in consideration of the rapidly occurring environmental and climatic changes.

Potential Role of Birds in the Epidemiology of Coxiella burnetii, Coxiella-like Agents and Hepatozoon spp.

Birds may be involved in the epidemiology of infectious and/or parasitic diseases which affect mammals, including humans. Q fever, caused by Coxiella burnetii, is an important zoonosis causing economic losses mainly due to pathologies induced in ruminants. Even though birds are known to be potential reservoirs of C. burnetii, their role in the epidemiological cycle of the pathogen is not completely verified. In recent years, new bacteria identified as Coxiella-like agents, have been detected in birds affected by different pathologies; the potential role of these bacteria as pathogens for mammals is not currently known. Hepatozoon spp. are haemoprotozoa, causing arthropod borne affections within several vertebrate classes. The infection of vertebrate host develops after ingestion of the arthropod final hosts containing oocysts; different tissues and blood cells are then colonized by other parasite stages, such as merozoites and gamonts. In avian hosts, there are several recognized Hepatozoon species; however, their life cycle and pathogenicity have not been fully elucidated. Referring to a carrier role by avian species and their ticks in the epidemiology of canine hepatozoonosis, the only clinically relevant affection caused by this parasite genus, they would act as carriers of infected ticks and, when Hepatozoon americanum is involved, as paratenic hosts, as well.

Occurrence and Molecular Identification of Hemoparasites in Wild Mammals Kept in Rehabilitation Centers in Brazil.

PURPOSE: Hepatozoonosis and piroplasmosis are diseases caused by apicomplexan protozoa that affect different types of animals, including mammals. The present study aimed to evaluate the occurrence of Hepatozoon spp. and piroplasms in wild mammals kept in captivity in rehabilitation centers in the states of Minas Gerais and Goiás, Brazil. METHODS: For this, blood samples from 152 animals were collected and analyzed by conventional optical microscopy and polymerase chain reaction (PCR). In addition, positive PCR samples were submitted to sequencing for molecular characterization of the specimens found. RESULTS: Microscopic analysis revealed 53 of the 152 animals (28.3%) parasitized by piroplasms. No Hepatozoon sp. was observed. On the other hand, using the primers HepF300/HepR900 and Piro1F/Piro5R, both amplifying fragments of the 18S rDNA gene, eight animals (5.2%) were positive for Hepatozoon spp. and 40 (26.3%) for piroplasms. From the sequencing of the positive samples Hepatozoon canis, Hepatozoon felis, Theileria cervi, Theileria equi and Cytauxzoon felis were identified. In addition to the aforementioned hemoparasites, some animals were found parasitized by microfilaria. Such data ratify the presence of hemoparasites in captive wild animals, and are unprecedented in the two geographical regions covered by the present study. 19.7% of mammals harbored ectoparasites of the genera Amblyomma and Rhipicephalus. CONCLUSION: Wild mammals are infected by several pathogens that can also infect domestic animals, some of them potentially zoonotic which can directly contribute to mortality and species reduction. Therefore, a deep understanding of the parasites, the hosts and the diseases is extremely necessary so that prevention, control and treatment measures are effectively applied.

Molecular detection of vector-borne agents in ectoparasites and reptiles from Brazil.

Trombidiformes and Mesostigmata mites, as well as Ixodida ticks, infest ectothermic tetrapods worldwide, potentially acting as vectors of bacteria, viruses and protozoa. The relationship among ectoparasites, transmitted pathogenic agents (e.g., Borrelia spp., Coxiella spp., Hepatozoon spp., and Rickettsia spp.) and ectothermic hosts has been scarcely investigated. This research focuses on a large collection of Brazilian herpetofauna screened for the presence of arthropod ectoparasites and vector-borne microbial agents. Reptiles (n = 121) and amphibians (n = 49) from various locations were infested by ectoparasites. Following genomic extraction, microbial agents were detected in 81 % of the Acari (i.e. n = 113 mites and n = 26 ticks). None of the mites, ticks and tissues from amphibians yielded positive results for any of the screened agents. Blood was collected from reptiles and processed through blood cytology and molecular analyses (n = 48). Of those, six snakes (12.5 %) showed intraerythrocytic alterations compatible with Hepatozoon spp. gamonts and Iridovirus inclusions. Hepatozoon spp. similar to Hepatozoon ayorgbor and Hepatozoon musa were molecularly identified from seven hosts, two mite and two tick species. Rickettsia spp. (e.g., Rickettsia amblyommatis, Rickettsia bellii-like, Rickettsia sp.) were detected molecularly from four mite species and Amblyomma rotundatum ticks. Phylogenetic analyses confirmed the molecular identification of the above-mentioned microbial agents of mites and ticks related to snakes and lizards. Overall, our findings highlighted that the Brazilian herpetofauna and its ectoparasites harbour potentially pathogenic agents, particularly from the northern and south-eastern regions. The detection of several species of spotted fever group Rickettsia pointed out the potential role of ectothermic hosts and related arthropod ectoparasites in the epidemiological cycle of these bacteria in Brazil.

Ticks and Associated Pathogens From Rescued Wild Animals in Rainforest Fragments of Northeastern Brazil.

The Ixodidae family comprises ticks that are hematophagous ectoparasites and are considered vectors of several hemoparasites from the Anaplasmataceae family and the genus Hepatozoon, Babesia, and Rickettsia. These ectoparasites parasitize domestic and wild animals belonging to several vertebrate groups. Ticks are highly adapted to different biomes and thus possess a wide geographical distribution. In the Brazilian state of Bahia, localized in the Northeast region, there are large rainforest fragments. Studies have rarely been carried out on ticks, and their hemoparasites, that parasitize wild animals in this region. Thus, this study aimed to identify the tick species parasitizing wild animals rescued in rainforest fragments of Bahia and investigate the presence of hemoparasites in tick tissues. During a 2-year period, 238 ticks were collected from 41 wild mammalians, reptiles, and amphibians. These ectoparasites were taxonomically classified according to their morphological characteristics. The ticks identified belonged to five different species from the Ixodidae family: Amblyomma varium, Amblyomma rotundatum, Amblyomma nodosum, Ixodes loricatus, and Rhipicephalus sanguineus. For the first time, an A. rotundatum parasitizing the Mesoclemmys tuberculata turtle was described. PCR assays using DNA extracted from salivary glands or midgut of the ticks were performed to detect specific DNA fragments of hemoparasites from the genus Rickettsia, Ehrlichia, Babesia, Hepatozoon, and from the Anaplasmataceae family. The results showed positive detection of the Rickettsia genus (7.9%), Anaplasmataceae family (15.8%), and Hepatozoon genus (15.8%). Specific DNA from the Ehrlichia and Babesia genera were not detected in these samples. Specific DNA from members of the Anaplasmataceae family was detected in A. varium for the first time. The present work showed that amphibians, reptiles, and mammals from Bahia's Atlantic Forest areparasitized by different tick species, and that these ectoparasites present pathogens in their tissues that impact both humans and animals due to their zoonotic potential.

Prevalence of protozoan parasites in small and medium mammals in Texas, USA.

Wildlife interaction with humans increases the risk of potentially infected ticks seeking an opportunistic blood meal and consequently leading to zoonotic transmission. In the United States, human babesiosis is a tick-borne zoonosis most commonly caused by the intraerythrocytic protozoan parasite, Babesia microti. The presence of Babesia microti and other species of Babesia within Texas has not been well characterized, and the molecular prevalence of these pathogens within wildlife species is largely unknown. Small (e.g. rodents) and medium sized mammalian species (e.g. racoons) represent potential reservoirs for specific species of Babesia, though this relationship has not been thoroughly evaluated within Texas. This study aimed to characterize the molecular prevalence of Babesia species within small and medium sized mammals at two sites in East Texas with an emphasis on detection of pathogen presence in these two contrasting wild mammal groups at these sites. To that end, a total of 480 wild mammals representing eight genera were trapped, sampled, and screened for Babesia species using the TickPath layerplex qPCR assay. Two sites were selected for animal collection, including The Big Thicket National Preserve and Gus Engeling Wildlife Management Area. Molecular analysis revealed the prevalence of various Babesia and Hepathozoon species at 0.09% each, and Sarcocystis at 0.06% . Continued molecular prevalence surveys of tick-borne pathogens in Texas wild mammals will be needed to provide novel information as to which species of Babesia are most prevalent and identify specific wildlife species as pathogen reservoirs.

Canine vector-borne protozoa: Molecular and serological investigation for Leishmania spp., Trypanosoma spp., Babesia spp., and Hepatozoon spp. in dogs from Northern Algeria.

Dogs are competent reservoirs/hosts of several protozoan pathogens transmitted by blood-feeding arthropods. Throughout their long history of domestication, they have served as a link for the exchange of parasites among livestock, wildlife, and humans and therefore remain an important source of emerging and re-emerging diseases. In Algeria, while canine leishmaniosis (CanL) is well known to be endemic, no data are available on other vector-borne protozoans. Here, we investigate the occurrence and diversity of trypanosomes, piroplasms and Hepatozoon spp. and update the epidemiological status of CanL in dogs from Kabylia, northern Algeria. A total of 227 dogs from three regions of Kabylia were enrolled, including 77 dogs with clinical signs. Dogs were clinically examined and were tested for L. infantum antibodies using a Rapid Immuno-Migration (RIM™) and a quantitative indirect Immunofluorescence Antibody Test (IFAT). PCR screening and sequencing were performed for vector-borne protozoa. Sixty two percent (141/227) of dogs presented at least one infection, whereas 26% (59/227) were co-infected. L. infantum antibodies were detected in 35.7% (81/227) of dogs including 88.7% (68/77) of sick dogs. Molecular investigation revealed prevalence of: 6.6% (15/227), 13.2% (30/227), 41% (93/227) for Trypanosoma spp., B. vogeli and H. canis, respectively. T. evansi (3.1%) and potential new subspecies of T. congolense had been identified. Dog''s clinical status correlates positively with L. infantum antibody titers and the presence of co-infections. Susceptibility to CanL varied according to the dog's aptitude and guard dogs were more infected (51%) (P-value = .001). B. vogeli infection was more frequent in juveniles than adults (32% vs 9%, P-value < .001) and in females than males (21% vs 10%, P-value = .02). To the authors' knowledge, this is the first report on vector-borne protozoa infected dogs in Algeria. Current results are important not only for animal health, but also to avoid serious public health and livestock problems.

Molecular Survey of Babesia microti (Aconoidasida: Piroplasmida) in Wild Rodents in Turkey.

Babesia microti (Aconoidasida: Piroplasmida) (Franca, 1910) is an important tick-borne zoonotic parasite with rodents serving as reservoir hosts. In the present study, 536 rodents were captured from Burdur, Bartin, Giresun, and Yozgat provinces of Turkey between the years 2010 and 2012, and blood samples were examined for the presence of Babesia spp. using conventional PCR which targeted the 18S rRNA gene. The sequence analysis of PCR amplicons was tested for B. microti as well as for Hepatozoon spp., and Sarcocystis spp. Overall, 5.8% of the rodents were positive for B. microti: 41% in Myodes glareolus, 7.7% in Chionomys roberti, and 2% in Apodemus spp., whereas no Babesia DNA was detected in Mus macedonicus and Microtus spp. Six rodents were positive for Hepatozoon spp. and one rodent was positive for Sarcocystis spp. Overall, 14.9 and 4.5% of rodents captured from Bartin and Giresun provinces, respectively, were PCR positive for B. microti, whereas none of rodents captured in Burdur and Yozgat were positive for Babesia spp. The sequence data of B. microti from rodents revealed that all sequences belonged to the zoonotic genotype. Sequences of B. microti obtained from rodents of the Bartin province were genotypically closer to European isolates, whereas those obtained from rodents of the Giresun province were closer to Russian and Mongolian isolates.

Molecular characterization and identification of Hepatozoon species Miller, 1908 (Apicomplexa: Adeleina: Hepatozoidae) in captive snakes from Brazil.

Species of Hepatozoon are parasites frequently recorded in snakes. The species identification of this genus was based mostly on the gametocyte morphology and morphometric calculations. For more reliable results, molecular characterization, an initial step for the correct identification of Hepatozoon species, has been used. This study aimed to determine the prevalence and identification of Hepatozoon species in captive snakes from Brazil. To that end, morphological, morphometric, and molecular data were obtained. A total of 157 snakes; 128 venomous (Crotalus durissus) and 29 non-venomous (Epicrates crassus and Boa constrictor) were screened for Hepatozoon blood parasites. Using light microscopy, 20 (12.78%) snakes were found positive for Hepatozoon spp., of which 6/29 (20.7%) were non-venomous and 14/128 (10.9%) were venomous; all with low parasitemia. Polymerase chain reaction (PCR), performed with the primers HepF300/Hep900, confirmed all 20 (100%) samples positive for hemogregarines. Species of Hepatozoon were identified from eight sequenced samples. Two previously described species, Hepatozoon cuestensis and Hepatozoon musa, were identified. The present study is the first to report H. musa within the snake hosts E. crassus and C. durrisus. In addition, a potentially new Hepatozoon species from B. constrictor was identified.

Prevalence of Hepatozoon and Sarcocystis spp. in rodents and their ectoparasites in Nigeria.

Using polymerase chain reaction targeting the 18S rRNA gene and DNA sequencing the prevalence and diversity of Apicomplexa and Piroplasmida infections in rodents from Nigeria was studied. Overall, 13 of 194 (7.7%) rodent blood samples tested were positive for Hepatozoon spp. while 2 (1.0%) were positive for Sarcocystis dispersa. Hepatozoon spp. DNA was detected in all the rodentspecies tested except Neotoma spp., and was most prevalent (50%) in the African giant rat (Cricetomys gambianus), followed by Mus musculus (18.2%), Rattus rattus (6.3%) and Rattus norvegicus (4.1%). The Hepatozoon spp. DNA sequences from the rodents were 98-100% identical to each other and to Hepatozoon spp. DNA sequence from small mammals deposited in GenBank. Five of the sequences from R. rattus (n = 2) and R. norvegicus (n = 3) were 98-99% identical to Hepatozoon felis (KY649442.1). Sarcocystis dispersa DNA was detected in one R. rattus (2.1%) and one R. norvegicus (0.8%). These findings suggest that rodents are involved in endemic cycles of Hepatozoon spp. and Sarcocystis spp. agents of veterinary importance.

Association between canine leishmaniosis and Ehrlichia canis co-infection: a prospective case-control study.

BACKGROUND: In the Mediterranean basin, Leishmania infantum is a major cause of disease in dogs, which are frequently co-infected with other vector-borne pathogens (VBP). However, the associations between dogs with clinical leishmaniosis (ClinL) and VBP co-infections have not been studied. We assessed the risk of VBP infections in dogs with ClinL and healthy controls. METHODS: We conducted a prospective case-control study of dogs with ClinL (positive qPCR and ELISA antibody for L. infantum on peripheral blood) and clinically healthy, ideally breed-, sex- and age-matched, control dogs (negative qPCR and ELISA antibody for L. infantum on peripheral blood) from Paphos, Cyprus. We obtained demographic data and all dogs underwent PCR on EDTA-blood extracted DNA for haemoplasma species, Ehrlichia/Anaplasma spp., Babesia spp., and Hepatozoon spp., with DNA sequencing to identify infecting species. We used logistic regression analysis and structural equation modelling (SEM) to evaluate the risk of VBP infections between ClinL cases and controls. RESULTS: From the 50 enrolled dogs with ClinL, DNA was detected in 24 (48%) for Hepatozoon spp., 14 (28%) for Mycoplasma haemocanis, 6 (12%) for Ehrlichia canis and 2 (4%) for Anaplasma platys. In the 92 enrolled control dogs, DNA was detected in 41 (45%) for Hepatozoon spp., 18 (20%) for M. haemocanis, 1 (1%) for E. canis and 3 (3%) for A. platys. No Babesia spp. or "Candidatus Mycoplasma haematoparvum" DNA was detected in any dog. No statistical differences were found between the ClinL and controls regarding age, sex, breed, lifestyle and use of ectoparasitic prevention. A significant association between ClinL and E. canis infection (OR = 12.4, 95% CI: 1.5-106.0, P = 0.022) was found compared to controls by multivariate logistic regression. This association was confirmed using SEM, which further identified that younger dogs were more likely to be infected with each of Hepatozoon spp. and M. haemocanis, and dogs with Hepatozoon spp. were more likely to be co-infected with M. haemocanis. CONCLUSIONS: Dogs with ClinL are at a higher risk of co-infection with E. canis than clinically healthy dogs. We recommend that dogs diagnosed with ClinL should be tested for E. canis co-infection using PCR.

A PCR survey of vector-borne pathogens in different dog populations from Turkey.

In the present study, a total of 192 blood samples were collected from pet dogs, kennel dogs and shepherd dogs in Konya district, Turkey, and tested by specific PCR for the presence of vector-borne pathogens. Several pathogens were identified, most of which can cause substantial morbidity in dogs. PCR results revealed that 54 (28.1%) dogs were infected with one or more pathogens. Positive results were obtained for Babesia spp. in 4 dogs (2.1%), Hepatozoon spp. in 8 dogs (4.2%) and Mycoplasma spp. in 46 dogs (24%). Three dogs (1.6%) were infected with two or three pathogens. The sequence analysis of the positive DNA samples revealed the presence of Babesia canis vogeli, Hepatozoon canis, Hepatozoon sp. MF, Mycoplasma haemocanis and Candidatus Mycoplasma haematoparvum. Ehrlichia canis and Anaplasma platys were not detected. Regardless of ownership status, vector-borne diseases were common in these dog populations. There was significant difference of pathogen prevalence among the different dog populations. Mycoplasma spp. was more frequent in the kennel dogs (31.9%) than in the pet (21.4%) and shepherd dogs (13.8%). Additionally, the frequency of Babesia spp. and Hepatozoon spp. was higher in the shepherd dogs which account for three quarters and half of the total number of Babesia spp. and Hepatozoon spp., respectively. To our knowledge, this is the first report of Mycoplasma infection in dogs in Turkey. The results of the present study provide a foundation for understanding the epidemiology of canine vector-borne diseases (CVBDs), and for strategies to control these diseases in Turkey.

Molecular detection of Hepatozoon spp. in domestic dogs and wild mammals in southern Pantanal, Brazil with implications in the transmission route.

Hepatozoon parasites comprise intracellular apicomplexan parasites transmitted to vertebrate animals by ingestion of arthropods definitive hosts. The present work aimed to investigate the occurrence of Hepatozoon spp. in wild animals, domestic dogs and their respective ectoparasites, in southern Pantanal region, central-western Brazil, by molecular techniques. Between August 2013 and March 2015, 31 coatis (Nasua nasua), 78 crab-eating foxes (Cerdocyon thous), seven ocelots (Leopardus pardalis), 42 dogs (Canis lupus familiaris), 110 wild rodents (77 Thichomys fosteri, 25 Oecomys mamorae, and 8 Clyomys laticeps), 30 marsupials (14 Thylamys macrurus, 11 Gracilinanus agilis, 4 Monodelphis domestica and 1 Didelphis albiventris), and 1582 ticks and 80 fleas collected from the sampled animals were investigated. DNA samples were submitted to PCR assays for Hepatozoon spp. targeting 18S rRNA gene. Purified amplicons were directly sequenced and submitted to phylogenetic analysis. A high prevalence of Hepatozoon among carnivores (C. thous [91.02%], dogs [45.23%], N. nasua [41.9%] and L. pardalis [71.4%]) was found. However, ticks and fleas were negative to Hepatozoon PCR assays. By phylogenetic analysis based on 18S rRNA sequences, Hepatozoon sequences amplified from crab-eating foxes, dogs, coatis and ocelots clustered with sequences of H. canis, H. americanum and H. felis. The closely related positioning of Hepatozoon sequences amplified from wild rodents and T. macrurus marsupial to Hepatozoon from reptiles and amphibians suggest a possible transmission of those Hepatozoon species between hosts by ectoparasites or by predation. Hepatozoon haplotypes found circulating in wild rodents seem to present a higher degree of polymorphism when compared to those found in other groups of animals. Although rodents seem not to participate as source of Hepatozoon infection to wild carnivores and domestic dogs, they may play an important role in the transmission of Hepatozoon to reptiles and amphibians in Pantanal biome.

Molecular detection of tick-borne protozoan parasites in a population of domestic cats in midwestern Brazil.

Some tick-borne pathogens that infect domestic cats have been considered emergent in veterinary medicine. Occurrences of Hepatozoon spp., Babesia spp. and Cytauxzoon spp. have been described in several regions of Brazil. This paper offers a comprehensive analysis of the 18S rRNA gene of a Hepatozoon sp. strain detected in domestic cats in the metropolitan area of Cuiabá, in Midwestern Brazil. Based on a molecular analysis, we detected the presence of Hepatozoon species circulating among cats in this region. The aforementioned strain is closely related to other isolates of H. felis detected in wild felids. Moreover, a phylogenetic analysis indicates that this genotype is grouped into a clade of 18S rRNA sequences previously described for the genus Hepatozoon in wild felids around the world. Hepatozoon felis strains detected in cats from Spain and Israel showed, respectively, 98% and 97% identity to our sequence and are clustered on a separate branch of the phylogenetic tree. This finding suggests a high diversity of Hepatozoon genotypes occurring in cats in Europe and South America. None of the analyzed cats were positive for Babesia spp. or Cytauxzoon spp. by PCR analysis.

Tick-borne agents in domesticated and stray cats from the city of Campo Grande, state of Mato Grosso do Sul, midwestern Brazil.

Anaplasmataceae agents, piroplasmids and Hepatozoon spp. have emerged as important pathogens among domestic and wild felines. The present work aimed to detect the presence of species belonging to the Anaplasmataceae family, piroplasmas and Hepatozoon spp. DNA in blood samples of domesticated and stray cats in the city of Campo Grande, state of Mato Grosso do Sul, midwestern Brazil. Between January and April 2013, whole blood samples were collected from 151 cats (54 males, 95 females and two without gender registration) in the city of Campo Grande, state of Mato Grosso do Sul, Brazil. DNA extracted from cat blood samples was submitted to conventional PCR assays for Theileria/Babesia/Cytauxzoon spp. (18S rRNA, ITS-1), Ehrlichia spp. (16S rRNA, dsb, groESL), Anaplasma spp. (16S rRNA, groESL) and Hepatozoon spp. (18S rRNA) followed by phylogenetic reconstructions. Out of 151 sampled cats, 13 (8.5%) were positive for Ehrlichia spp. closely related to Ehrlichia canis, 1 (0.66%) for Hepatozoon spp. closely related to Hepatozoon americanum and Hepatozoon spp. isolate from a wild felid, 1 (0.66%) for Cytauxzoon sp. closely related do Cytauxzoon felis, and 18 (11.9%) for Babesia/Theileria (one sequence was closely related to Babesia bigemina, eight for Babesia vogeli, five to Theileria spp. from ruminants [Theileria ovis, Theileria lestoquardi] and four to Theileria sp. recently detected in a cat). The present study showed that Ehrlichia spp., piroplasmids (B. vogeli, Theileria spp. and Cytauxzoon spp.) and, more rarely, Hepatozoon spp. circulate among stray and domesticated cats in the city of Campo Grande, state of Mato Grosso do Sul, midwestern Brazil.