Molecular evidence for trans-stadial transmission of Anaplasma platys by Rhipicephalus sanguineus sensu lato under field conditions.

This study investigated the transmission of Anaplasma platys by Rhipicephalus sanguineus sensu lato (Ixodida: Ixodidae). Engorged nymphs (n = 404) removed from A. platys-infected dogs were incubated at 28 °C until moulting. Unfed adults were obtained and divided into 48 pools. Unfed ticks collected from the grounds of the dog shelter, comprising 1800 larvae (n = 18 pools), 3100 nymphs (n = 62 pools) and 85 adults (n = 10 pools, including three male and seven female pools) were sorted into 90 pools. All pools were screened by polymerase chain reaction for the 16S rRNA gene of A. platys. Of 48 pools of unfed adults obtained from engorged nymphs, 12 were positive for A. platys; the infection rate maximum likelihood estimate (MLE) was 3.36 [95% confidence interval (CI) 1.84-5.68]. Anaplasma platys was detected in five of 23 male pools (MLE 2.82, 95% CI 1.06-6.20) and seven of 25 female pools (MLE 3.83, 95% CI 1.72-7.57). Of seven pools of unfed adult females collected from the shelter grounds, one was positive for A. platys (MLE 1.74, 95% CI 0.11-8.22). Among 62 unfed nymph pools, eight were infected with A. platys (MLE 0.27, 95% CI 0.13-0.52). No A. platys DNA was detected in the larva pools. The present results reveal molecular evidence for the trans-stadial transmission of A. platys by R. sanguineus s.l.

Molecular survey of haemoplasmas in shelter dogs and associations with Rhipicephalus sanguineus sensu lato.

This study aimed to assess the occurrence of canine haemoplasma infection in domestic dogs and its possible trans-stadial transmission by Rhipicephalus sanguineus sensu lato (Ixodida: Ixodidae) in shelter dogs in Diyarbakir Province in southeast Turkey. Blood samples (n = 282) collected from domestic dogs were analysed by polymerase chain reaction (PCR) for the presence of canine haemoplasma. Fully engorged nymphs (n = 204) were removed from dogs that were positive for canine haemoplasma by PCR and maintained in an incubator at 28 °C for moulting. Unfed ticks (n = 2185) comprising 2100 nymphs and 85 adults collected from the grounds of the same shelter were also screened. Of 282 dogs, 108 [38.3%, 95% confidence interval (CI) 32.6-44.2] were PCR-positive for canine haemoplasmas. Mycoplasma haemocanis (Mhc) infection (26.2%, 95% CI 21.2-31.8) was observed in a significantly higher number of dogs than was Candidatus Mycoplasma haematoparvum (CMhp) infection (6.7%, 95% CI 4.1-10.3). Co-infections were seen in 15 (5.3%, 95% CI 3.0-8.6) dogs. None of the tick specimens examined were found to be positive for haemoplasma. Partial sequences of the 16S ribosomal RNA (rRNA) gene shared 99-100% identity with the corresponding published sequences for Mhc and CMhp. The present results revealed no trans-stadial transmission of canine haemoplasma species by R. sanguineus s.l. in field conditions.

Transstadial Transmission of Hepatozoon canis by Rhipicephalus sanguineus (Acari: Ixodidae) in Field Conditions.

This study investigated possible transovarial and transstadial transmission of Hepatozoon canis by Rhipicephalus sanguineus (Latreille) ticks collected from naturally infected dogs in a municipal dog shelter and the grounds of the shelter. Four hundred sixty-five engorged nymphs were collected from 16 stray dogs that were found to be infected with H. canis by blood smear and PCR analyses and maintained in an incubator at 28 °C for moulting. Four hundred eighteen nymphs moulted to adults 14-16 d post collection. Unfed ticks from the shelter grounds comprised 1,500 larvae, 2,100 nymphs, and 85 adults; were sorted according to origin, developmental stage, and sex into 117 pools; and screened by 18S rRNA PCR for Hepatozoon infection. Of 60 adult tick pools examined, 51 were infected with H. canis. The overall maximum likelihood estimate (MLE) of infection rate was calculated as 21.0% (CI 15.80-28.21). Hepatozoon canis was detected in 31 out of 33 female pools (MLE 26.96%, CI 17.64-44.33) and 20 out of 27 male pools (MLE 14.82%, CI 20.15-46.41). Among 42 unfed nymph pools collected from the shelter, 26 were infected with H. canis, and MLE of infection was calculated as 1.9% (CI 1.25-2.77). No H. canis DNA was detected in any of the gDNA pools consisting of larva specimens. Partial sequences of the 18S rRNA gene shared 99-100% similarity with the corresponding H. canis isolates. Our results revealed the transstadial transmission of H. canis by R. sanguineus, both from larva to nymph and from nymph to adult, in field conditions. However, there were no evidence of transovarial transmission.

Molecular and Parasitological Survey of Ovine Piroplasmosis, Including the First Report of Theileria annulata (Apicomplexa: Theileridae) in Sheep and Goats from Turkey.

Blood and tick samples were collected from 333 apparently healthy sheep and 257 goats as well as 10 sheep exhibiting clinical signs of babesiosis in Adana, Gaziantep, and Adiyaman Provinces in southern Turkey. Fully engorged female ticks were selected and maintained in an incubator until they oviposited. The tick carcasses and their egg masses were examined. Piroplasms compatible with Babesia spp. and Theileria spp. were observed in both symptomatic and asymptomatic small ruminants. Genomic DNA isolates from blood of ovine, tick samples, and egg masses were screened for piroplasms by utilizing 18S rRNA polymerase chain reaction and reverse line blotting (RLB) assays. Parasitemia ranged from 0.01% to 5.6% of erythrocytes in clinical cases. RLB showed positivity in 239 (40.5%) of the sampled apparently healthy sheep and goats and revealed the presence of three Theileria and one Babesia species. Theileria ovis was the most prevalent (35.4%), followed by Babesia ovis (5.4%), Theileria annulata (3.9%), and Theileria sp. MK (0.3%). Thirty-two small ruminants infected with T. ovis were also infected with B. ovis One animal infected with T. ovis was also infected with Theileria sp. MK. Ticks were identified as Rhipicephalus bursa, Rhipicephalus turanicus, Hyalomma excavatum, Haemaphysalis parva, and Hyalomma anatolicum Egg masses of two female R. bursa carcasses were infected with B. ovis This is the first report of theileriosis caused by T. annulata in sheep and goats in Turkey.

Molecular and Parasitological Survey of Bovine Piroplasms in the Black Sea Region, Including the First Report of Babesiosis Associated with Babesia divergens in Turkey.

Clinical cases of babesiosis were evaluated, and the frequency of bovine Babesia and Theileria parasites was determined in cattle. Blood samples and thin blood smears were collected from 23 cattle exhibiting clinical signs of babesiosis. In addition, tick and blood samples were collected from 100 apparently healthy cattle cograzing from the same area. Egg masses obtained from fully engorged female ticks were included. DNA isolated from blood and tick samples was screened for Babesia and Theileria by reverse line blot assay. Piroplasms compatible with Babesia spp. were observed microscopically for symptomatic cattle as circular, oval, elongated, or pear-shaped bodies. Parasitemia ranged from 0.08 to 0.9% for Babesia bovis, 2.5 to 15.4% for Babesia bigemina, and 7.4% for Babesia divergens. Reverse line blot showed positivity in 13 (13%) of the sampled clinically normal cattle and revealed the presence of three Babesia species. Babesia bovis was the most prevalent (9/100, 9%), followed by Babesia occultans (3/100, 3%) and B. bigemina (1/100, 1%). One animal infected with B. bigemina was also infected with B. bovis. The single animal infected with B. divergens showed symptoms of babesiosis. Ticks were identified as Rhipicephalus annulatus, Rhipicephalus turanicus, and Ixodes ricinus. One female R. annulatus and its egg mass were infected with B. bigemina. Neither Theileria annulata nor Theileria buffeli/orientalis infections were observed in cattle or ticks. This is the first report of clinical babesiosis caused by B. divergens in cattle from Turkey.

Application of the Reverse Line Blot Assay for the Molecular Detection of Theileria and Babesia sp. in Sheep and Goat Blood Samples from Pakistan.

BACKGROUND: The present study was designed to detect the presence of tick-borne parasites (Theileria and Babesia spp.) in 196 blood samples collected from apparently healthy sheep and goats from two provinces, Punjab and Khyber Pukhtoon Khwa, in Pakistan. METHODS: Reverse line blot (RLB) assay was applied for the parasitic detection by the amplification of hypervariable V4 region of the 18S ribosomal RNA (rRNA) gene. A membrane with covalently linked generic and species specific oligonucleotide probes was used for the hybridization of amplified PCR products. RESULTS: Parasites were detected in 16% of the ruminant blood samples under study. Two Theileria species, T. lestoquardi and T. ovis, were identified in samples. 25, of the total 32, infected animals were from Khyber Pukhtoon Khwa. CONCLUSION: Sheep were more prone to tick borne haemoprotozans as 81% infected samples were sheep as compared to 19% goats (P > 0.001). Risk factor analysis revealed that male (P = 0.03), animals infested by ticks (P = 0.03) and herd composed of sheep only (P = 0.001) were more infected by blood parasites.