Major Piroplasm Surface Protein Research Articles

Haematological and Biochemical Profiles, Including Vitamin A and Zinc Status, in Clinical and Subclinical Oriental Theileriosis in Cattle

Aim: The present study investigated the clinical, haematological and biochemical profiles in cattle infected with Theileria orientalis, correlating disease severity with various parameters. Study Design: A total of 32 clinically ill, 10 subclinically infected, and 10 healthy control animals were included. Place and Duration of Study: Department of Veterinary Epidemiology and Preventive Medicine, College of Veterinary and Animal Sciences, Kerala Veterinary and Animal Sciences University, Pookode, Wayanad, Kerala, between November 2023 to September 2024. Methodology: Theileria piroplasms, observed via Giemsa staining, appeared as thick and thin rods with trailing cytoplasm and annular, pyriform, garland, and signet-ring shapes. PCR amplification of the T. orientalis major piroplasm surface protein (MPSP) yielded a specific 776 bp product in clinically and subclinically infected animals. Results: Haematological analysis revealed significant anaemia and thrombocytopenia in infected groups, without notable changes in leukocyte counts. Serum protein analysis indicated hypoproteinemia and hypoalbuminemia, while liver enzymes, particularly AST, were elevated in clinically infected cattle, suggesting hepatic involvement. Additionally, serum vitamin A and zinc concentrations were significantly lower in infected cattle, underscoring a possible role for oxidative stress in disease pathogenesis. Conclusion: These findings suggest that T. orientalis infection is associated with multi-systemic impacts, including haematological abnormalities, liver damage, and nutrient deficiencies, which are crucial for accurate diagnosis and effective management strategies.

Transient efficacy of buparvaquone against the US isolate of Theileria orientalis Ikeda genotype in sub-clinically infected cattle.

Theileria orientalis, an economically significant tick-borne hemoparasite, infects cattle globally. The T. orientalis Ikeda genotype, transmitted by Haemaphysalis longicornis ticks, is associated with clinical manifestations characterized by anemia, abortions, and mortality, although subclinical infections prevail. Despite the common occurrence of subclinical infections, therapeutic interventions targeting T. orientalis Ikeda in such cases are currently lacking, impeding effective parasite control measures. To address this critical knowledge gap, we assessed the efficacy of buparvaquone (BPQ) in eliminating the T. orientalis Ikeda, US isolate, in sub-clinically infected cattle. Twelve sub-clinically infected calves, identified by the presence of T. orientalis in peripheral blood alongside the absence of fever and anemia, were enrolled in the study. Six calves received two treatments of the BPQ label dose (2.5 mg/kg) at a 48-h interval, while additional three calves received the drug at a dosage of 6 mg/kg following the same regimen. Three untreated calves served as controls. Endpoint and quantitative PCR analyses revealed that BPQ exerted a transient effect on T. orientalis parasitemia. Parasites remained undetectable in peripheral blood until weeks 4 and 11 post-treatment in animals administered 2.5 mg/kg and 6 mg/kg of BPQ, respectively. Intriguingly, following recrudescence, administering 6 mg/kg to animals previously treated with 2.5 mg/kg did not result in a reduction in parasite load. Pharmacokinetic analysis data suggested that escalating the dosage led to a less than proportional increase in serum concentrations of BPQ. Moreover, a significant yet reversible decrease (p < 0.05) in blood urea nitrogen was observed in animals treated with the drug, irrespective of the dosage. Despite parasitemia relapse, animals treated with 6 mg/kg BPQ exhibited a noteworthy decrease (p < 0.05) in IgG levels specific to the T. orientalis major piroplasm surface protein compared to controls and animals treated with 2.5 mg/kg of the drug. BPQ did not demonstrate efficacy in clearing subclinical T. orientalis Ikeda infection. Future investigations are warranted to explore innovative therapeutic modalities that, in synergy with vaccines and diagnostic assays, can facilitate the development of comprehensive programs aimed at controlling and eradicating this parasite.

Latitude and seasons influence the prevalence of Theileria orientalis and affect the hematology of non-grazed dairy cows in Korea.

This pilot study aimed to investigate the effects of regional and seasonal variations on the prevalence of Theileria orientalis and the hematological profile of non-grazed dairy cows in Korea. A total of 365 clinically healthy lactating Holstein Friesian cows from 26 dairy farms in 7 provinces that were categorized into northern, central, and southern regions were sampled during the warm period from July to August and the cold period from October to December. The detection of T. orientalis major piroplasm surface protein gene and the hematology non-grazed dairy cows were analyzed using peripheral blood samples. The T. orientalis prevalence was 20.0% (73/365). The prevalence in the southern region was 35.9%, which was significantly higher than that in the central (21.6%) and northern (12.9%) regions (P &lt; 0.05). The prevalence during warm period was higher (43.0%) than that during the cold season (13.5%). The infected cows showed significantly lower erythrocyte counts in the southern region (5.8 ± 0.6 M/µl) and during the warm period (5.8 ± 0.7 M/µl) compared with those in the central and northern regions and during the cold season, which affected the extended RBC parameters, including hematocrit and hemoglobin concentrations. Our findings revealed the prevalence of T. orientalis in Korea, highlighting its high occurrence during warm periods and in certain geographical regions. Climatic factors could contribute to the health and productivity of cattle, as evidenced by the prevalence of T. orientalis and its negative impact on animals.

Dermacentor variabilis Does Not Transstadially Transmit the U.S. Isolate of Theileria orientalis Ikeda: A Controlled Acquisition and Transmission Study

Theileria orientalis Ikeda, an emerging U.S. bovine hemoparasite, causes anemia, abortion, ill-thrift, and occasionally death. While Haemaphysalis longicornis is the primary vector, it is possible that other U.S. ticks are capable of parasite transmission and may contribute to disease spread. Dermacentor variabilis is highly prevalent in the U.S., exhibits a similar geographical distribution to T. orientalis, and is a competent vector of the related parasite, Theileria equi. Herein, we conducted controlled acquisition and transmission studies using splenectomized calves to assess whether D. variabilis can transstadially transmit T. orientalis. D. variabilis nymphs were applied to an infected, splenectomized calf for parasite acquisition and subsequently incubated to molt into adults. Freshly molted adults were applied to two splenectomized T. orientalis-naïve calves to investigate parasite transmission. Calves were monitored for 59 days, and no evidence of parasite transmission was detected using PCR for the T. orientalis Ikeda major piroplasm surface protein gene, blood smear cytology, complete blood counts, or physical examination. Salivary glands from a subset of D. variabilis adults were assessed for T. orientalis using PCR, and the parasite was not detected. These findings support the conclusion that D. variabilis is not capable of transstadial transmission of the U.S. T. orientalis Ikeda isolate.

Theileria orientalis Buffeli pathotype in cows in a theileriosis-endemic region of India.

Oriental theileriosis caused by Theileria orientalis is a growing health concern of lactating cows in its endemic areas. Rapid and sensitive diagnostic tests are demand areas for appropriate and effective prophylactic and therapeutic measures. Quantitative polymerase chain reaction (qPCR) is the answer for both detection and quantification of parasites. Present study deals with qPCR for detection of parasitemia level of T. orientalis in apparently healthy and clinically affected cows. Major piroplasm surface protein (MPSP) gene present in T. orientalis was cloned in pUC57 vector and transformed into E. coli Top 10 cells. Single and mixed infections of hemoprotozoa other than T. orientalis, causing anemia were differentiated through blood smear examination and PCR tests. T. orientalis was detected in 108 (63.15%) ill and 48 (26.66%) healthy cows. Piroplasms detected per 1000 red blood cells (RBCs) was 0-1 in the healthy group as compared to 3-22 in those showing clinical signs. Parasitemia in ill cows ranged between 6.9 × 102 and 4.5 × 103 parasites / µl of blood which was significantly higher (p<0.05) than healthy group (2.6 × 102 - 5.7 × 102 parasites / µl of blood). Phylogenetic study of the isolates showed similarity with Buffeli type that unfolded its pathogenic form in apparently healthy and ill cows.

Multiplex PCR for rapid differential diagnosis of co-prevalent species of Theileria (Theileria annulata and Theileria orientalis) in cattle

Theileriosis is a tick-borne disease that causes enormous losses in the dairy industry. There are several species of Theileria that can infect bovines. Generally, more than one species are prevalent in any geographical area; thus, chances of co-infections are high. Differentiation of these species may not be possible by microscopic examination or serological tests. Therefore, in this study, a multiplex PCR assay was standardized and evaluated for rapid and simultaneous differential detection of two species of Theileria viz., Theileria annulata and Theileria orientalis. Species-specific primers were designed to target the merozoite piroplasm surface antigen gene (TAMS1) of T. annulata and the major piroplasm surface protein gene of T. orientalis, yielding specific amplicon of 229bp and 466bp, respectively. The sensitivity of multiplex PCR was 102 and 103 copies for T. annulata and T. orientalis, respectively. The simplex and multiplex PCRs were specific and showed no cross-reactivity with other hemoprotozoa for either primer. For comparative evaluation, blood samples from 216 cattle were tested by simplex and multiplex PCR for both species. Using multiplex PCR, 131 animals were found infected for theileriosis, of which 112 were infected with T. annulata, five were infected with T. orientalis, and 14 had mixed infections. This is the first report of T. orientalis from Haryana, India. Representative sequences of T. annulata (ON248941) and T. orientalis (ON248942) were submitted in GenBank. The standardized multiplex PCR assay used in this study was specific, sensitive, for the screening of field samples.

Oriental theileriosis associated with a new genotype of Theileria orientalis in buffalo (Bubalus bubalis) calves in Uttar Pradesh, India

Theileria orientalis is known to cause a benign infection in cattle and buffalo (Bubalus bubalis). However, the Ikeda and Chitose genotypes of the parasite cause lethal disease in beef and dairy cattle. Recently an outbreak of clinical oriental theileriosis occurred in buffalo calves in a Government Animal Husbandry and Agricultural Farm located in Uttar Pradesh, India. Examination of Giemsa stained thin blood smears revealed typical rod-shaped T. orientalis piroplasms in the erythrocytes. The clinical signs included pyrexia, nasal discharge, lacrimation, lethargy, inappetence and anaemia with varying degrees of paleness of the visible mucous membranes. Vascular congestion in internal organs, pulmonary emphysema and consolidation of lungs, focal areas of necrosis in the heart with mononuclear cell infiltration, focal mononuclear cell aggregation in the cortex and tubular degeneration of the kidney were significant necropsy findings. The T. orientalis major piroplasm surface protein (MPSP) gene was amplified by polymerase chain reaction (PCR) using specific primers. The nucleotide sequence analysis of the PCR product revealed 84.8% identity between the T. orientalis Uttar Pradesh isolate and other reference genotypes available in the public domain. Furthermore, the phylogenetic analysis of the MPSP gene sequence ratified that this is a new genotype of T. orientalis. This is the first report of a clinical outbreak of oriental theileriosis in Indian buffalo calves caused by a novel genotype of T. orientalis.

Theileria and Babesia infection in cattle – First molecular survey in Kazakhstan

Central Asia, including Kazakhstan, is an endemic area of Theileria and Babesia infections in cattle. Current data on the geographic distribution, prevalence, and genetic diversity of these pathogens in vertebrate hosts are lacking in Kazakhstan. The present study aimed to fill this gap, using molecular techniques for the first time.A cross-sectional survey was performed on adult cattle from 40 villages in nine administrative districts of the provinces of Turkistan and Zhambyl, southern Kazakhstan, in summer 2020. A total of 766 blood samples were screened for Theileria annulata (enolase gene), Theileria orientalis (major piroplasm surface protein gene, MPSP) and Babesia spp. (18 S ribosomal RNA gene) using polymerase chain reaction. The genetic variability of Theileria spp. was assessed by sequencing one amplicon from each village. All Babesia spp. positive amplicons were sequenced to identify the species involved.The overall prevalence of infections with T. annulata, T. orientalis and Babesia spp. was 83.0% (40 villages positive), 33.3% (31 villages) and 13.5% (36 villages), respectively. Co-infections with two or three species were present in 48.9% of all positive cattle. Theileria annulata showing a high polymorphism of the enolase gene occurred with similar frequency in both provinces. Theileria orientalis was detected for the first time in Kazakhstan being significantly (P = 0.014) more prevalent in Zhambyl than in Turkistan. Fourteen genotypes of T. orientalis were identified; two belonged to the moderately virulent MPSP-type 1 (‘Chitose’) and the others to MPSP-type 3 (‘Buffeli’) which is considered avirulent. The prevalence of Babesia infection was significantly (P < 0.000) higher in Turkistan than in Zhambyl. An unequivocal identification of the species involved was possible in 127 sequenced samples: Babesia occultans was the most common species, followed by Babesia bigemina and Babesia major, the latter being the first record in the country. The results show that Theileria and Babesia infections in cattle are widespread and occur with remarkably high prevalence in the southern Kazakhstan. They also provide first data on the genetic diversity of the species involved.

Detection of Theileria orientalis Genotypes from Cattle in Kyrgyzstan.

The ikeda and chitose genotypes of Theileria orientalis, which for many years were thought to be benign, cause a disease that results in significant economic losses in the cattle industry. This study was carried out in order to determine the genotypes of T. orientalis in cattle in Kyrgyzstan, and 149 archived DNA samples known to be T. orientalis were analyzed by the PCR amplification of the major piroplasm surface protein (MPSP) gene region. Single-Strand Conformation Polymorphism (SSCP) analysis was performed to uncover the nucleotide changes in the archived DNA samples, and 15 samples showing different band profiles were subjected to sequence analysis. As a result of the sequence analysis, it was seen that the samples belonged to the buffeli and chitose A genotypes. In order to identify mixed genotypes, PCR was performed using primers specific for these genotypes, and buffeli (type 3), chitose (type 1) and buffeli+chitose were found to be positive in 26.2%, 2% and 71.8% of samples, respectively. As a result of this study, we showed the presence of buffeli (type 3) and chitose (type 1) genotypes of T. orientalis in cattle in Kyrgyzstan. Comprehensive epidemiological studies are needed to understand the clinical infections caused by the pathogenic chitose A and to determine the geographical distribution and different genotypes of T. orientalis.

Molecular epidemiology and genetic characterization of Theileria orientalis in cattle.

Theileria orientalis (T. orientalis) is a benign species of Theileria that is found all throughout the world, including Egypt. The purpose of this work was to determine the current status of T. orientalis infection in cattle, as well as to define T. orientalis genetic diversity using the major piroplasm surface protein (MPSP) genes. In 2020, blood samples from 500 asymptomatic cattle were collected, which live in four different governorates at northern Egypt and examined using PCR assay based on MPSP gene. Overall, 44 blood samples (8.8%, 44/500) were positive for T. orientalis, and the highest prevalence rate was observed in Kafer ElSheikh while it was relatively low in Gharbia governorate. Moreover, based on multivariable analysis, the risk of infection with T. orientalis infection was higher in mixed breed cattle and in animals of age group more than 3years compared to other. Additionally, absence of acaricide application, infestation with ticks, and communal grazing system were identified as potential risk factors for T. orientalis infection. Phylogenetic analysis revealed that the isolates studied belonged to two MPSP genotypes, types 1 and 2. These findings reveal that the MPSP genotypes discovered in the present work displayed genetic variation that was linked to geographic area. Therefore, further research should be focused on developing an effective T. orientalis monitoring and preventive program.

Molecular Detection and Genetic Diversity of Tick-Borne Pathogens in Goats from the Southern Part of Thailand.

Tick-borne hemoprotozoan and rickettsial diseases affect the health and productivity of small ruminants in tropical and subtropical regions. Despite the large population of goats in the southern part of Thailand, there is limited information on the prevalence of tick-borne pathogens. In this study, polymerase chain reaction was used to detect the presence of Theileria spp., T. ovis, T. orientalis, Babesia ovis, Anaplasma ovis, and A. marginale in 262 goats from three provinces in the southern part of Thailand. In this investigation, Theileria spp. and A. ovis were detected while T. ovis, B. ovis, and A. marginale were not detected. Overall infection rates of Theileria spp. and A. ovis were 10.3% and 1.5%, respectively. The co-infections of two parasites was observed in 1.5% of goats. Sequence analysis showed the presence of T. luwenshuni and T. orientalis in the goat samples. This study is the first to use the molecular detection of T. orientalis in Thai goats, and presents genetic characterization using the major piroplasm surface protein (MPSP) gene. In the phylogenetic analysis, the T. orientalis MPSP sequence was classified as type 7. The A. ovis major surface protein 4 (MSP4) gene sequences shared high identities and similarity with each other and clustered with isolates from other regions. This study provides information about the prevalence and genetic diversity of tick-borne pathogens in goats in the study area, and is expected to be valuable for the development of effective control measures to prevent disease in animals in Thailand.

Genetic Diversity of Bovine Hemoprotozoa in South Korea

Tick-borne pathogens cause economically significant diseases in cattle. Theileria spp. are parasitic protozoa and the causative agent of bovine theileriosis. Here we report the distribution and risk factors of bovine Theileria using blood samples taken between 2018 and 2019. Of 737 tested cattle, nine animals (1.2%) were positive for Theileria orientalis infection by 18S rRNA gene amplification. Further analysis of the infected samples using the T. orientalis major piroplasm surface protein (MPSP) gene revealed five different genotypes circulating in the population: Types 1, 2, 3, 7, and N3. To the best of our knowledge, this is the first research to describe the existence of the T. orientalis MPSP genotype N3 in South Korea. Although the prevalence of bovine T. orientalis was low, our study offers data on the geographical distribution and prevalence of bovine Theileria spp. in South Korea. Further studies are warranted to determine the correlation of clinical symptoms with parasite MPSP genotypes. Our data provide epidemiological information to help control bovine theileriosis in South Korea.

A Targeted “Next-Generation” Sequencing-Informatic Approach to Define Genetic Diversity in Theileria orientalis Populations within Individual Cattle: Proof-of-Principle

Oriental theileriosis is an economically important tickborne disease of bovines, caused by some members of the Theileria orientalis complex. Currently, 11 distinct operational taxonomic units (OTUs), or genotypes, are recognized based on their major piroplasm surface protein (MPSP) gene sequences. Two of these genotypes (i.e., chitose and ikeda) are recognized as pathogenic in cattle, causing significant disease in countries of the Asia-Pacific region. However, the true extent of genetic variation and associated virulence/pathogenicity within this complex is unknown. Here, we undertook a proof-of-principle study of a small panel of genomic DNAs (n = 13) from blood samples originating from individual cattle known to harbor T. orientalis, in order to assess the performance of a targeted “next-generation” sequencing-informatic approach to identify genotypes. Five genotypes (chitose, ikeda, buffeli, type 4, and type 5) were defined; multiple genotypes were found within individual samples, with dominant and minor sequence types representing most genotypes. This study indicates that this sequencing-informatic workflow could be useful to assess the nature and extent of genetic variation within and among populations of T. orientalis on a large scale, and to potentially employ panels of distinct gene markers for expanded molecular epidemiological investigations of socioeconomically important protistan pathogens more generally.

Theileria orientalis Ikeda in host-seeking Haemaphysalis longicornis in Virginia, U.S.A.

The Asian longhorned tick, Haemaphysalis longicornis, has recently become established in the United States. In East Asia, Australia, and New Zealand, the native and previously introduced ranges, this tick is a vector of an important pathogen of cattle, Theileria orientalis. In 2017, the pathogenic Ikeda genotype of T. orientalis was associated with cattle mortalities in Virginia and in 2018 the exotic H. longicornis was detected at this same site. To investigate the possible role of this exotic tick in the epidemiology of theileriosis in Virginia, we tested host-seeking H. longicornis for piroplasm infections. We document the detection of exotic Theileria orientalis Ikeda genotype in 12.7 % (15/118) environmentally collected H. longicornis using both the 18S rRNA and major piroplasm surface protein (MPSP) gene targets. This is the first detection of a pathogen in H. longicornis in its introduced range in the United States and offers new insight into the animal health risks associated with the introduction of this exotic tick species to North America.

Molecular detection, prevalence and risk factors of Theileria orientalis infection among cattle in Peninsular Malaysia

There is need to confirm the presence of Theileria orientalis among the cattle population in Peninsular Malaysia and to evaluate the risk factors associated with the infection. To this effect, blood samples were collected from 1045 cattle from 43 farms throughout the entire States of Peninsular Malaysia. The collected blood samples were subjected to DNA extraction and subsequent PCR amplification of the major piroplasm surface protein (MPSP) gene of the haemoprotozoan. Representative positive amplicons were purified, sequenced and compared with other sequences of the MPSP gene of T. orientalis curated from the GenBank. A well-structured questionnaire was used to get information about each cattle, it’s demography, the bio-security, environmental and management factors. Univariate and multivariate analysis were used for the statistical evaluation, with significance set at p < 0.05. A total prevalence of 49.76% (520/1045; 95% CI: 46.73 – 52.79) was obtained. Types of breeds, age, production type, herd size, level of farm biosecurity, farm size, presence of other animal species in the farm, management systems and prophylaxis were significantly (p < 0.05) associated with the prevalence of T. orientalis. This study confirmed the presence of T. orientalis and establish that the haemoprotozoan is endemic among cattle in Peninsular Malaysia.

Genotypic Analysis of Piroplasms and Associated Pathogens from Ticks Infesting Cattle in Korea.

Tick-borne pathogens (TBPs) impose an important limitation to livestock production worldwide, especially in subtropical and tropical areas. Earlier studies in Korea have examined TBPs residing in ticks and animals; however, information on multiple TBPs in ticks infesting cattle is lacking. This study assessed the prevalence of TBPs in ticks parasitizing cattle. A total of 576 ticks, including 340 adults and 236 nymphs, were collected from cattle in Korea between 2014 and 2018. All ticks collected were identified as Haemaphysalis longicornis based on their morphological and molecular characteristics. Among piroplasms and other tick-associated pathogens, seven TBP genes, namely Theileria orientalis (5.0%), Anaplasma bovis (2.3%), Anaplasma capra (4.7%), Anaplasma phagocytophilum-like Anaplasma spp. (APL) clades A (1.9%) and B (0.5%), Ehrlichia canis (1.6%), and Candidatus Rickettsia longicornii (17.5%), were detected. Bartonella spp. and severe fever with thrombocytopenia syndrome virus were not found. To our knowledge, this is the first study to report the presence of the pathogens T. orientalis major piroplasm surface protein genotypes 3 and 7, A. capra, and APL in ticks from Korea. Cattle ticks may be maintenance hosts for many TBPs, and veterinary and medical clinicians should be aware of their high probability of infection and clinical complexity in humans.

Molecular, epidemiological and haematological evaluation in Theileria orientalis infected cattle from an endemic region in India

The objective of the present study was to investigate the epidemiological and haematological parameters with simultaneous molecular detection of Theileria orientalis infection of crossbred jersey (CBJ) cattle. Haematological values like mean hemoglobin (Hb) (7.31 ± 2.3 g/dl), packed cell volume (PCV) (21.69 ± 6.11%), red blood cells count (RBCs) (4.40 ± 1.6 M/μl), white blood cells count (WBCs) (6.93 ± 3.06 103/µl) and mean corpuscular hemoglobin concentration (MCHC) (33.56 ± 3.51 g/dl) were decreased significantly (p < 0.05), whereas mean corpuscular volume (MCV) (51.06 ± 6.14fl) and eosinophil count (0.39 ± 0.44 103/µl) were significantly (p < 0.05) increased in cattle infected with T. orientalis. Analysis of major piroplasm surface protein (MPSP) of 110 blood samples randomly collected from cattle from seven districts by PCR indicated that an average of 70% of cattle was positive for T. orientalis infection. In particular, Puri and Khorda districts were identified as relatively high-risk areas for T. orientalis infection, with infection rates of 76.66% and 72.4%, respectively. The phylogenetic analysis of isolated T. orientalis MPSP gene (MN334767) classified it into type 5. Earlier Indian isolates were classified into three types viz.type 1, type 3 and type 7 and this is the first time to detect type 5 in Odisha, India.

An appraisal of oriental theileriosis and the Theileria orientalis complex, with an emphasis on diagnosis and genetic characterisation.

Oriental theileriosis, a tick-borne disease of bovids caused by members of the Theileria orientalis complex, has a worldwide distribution. Globally, at least 11 distinct genotypes of T. orientalis complex, including type 1 (chitose), type 2 (ikeda), type 3 (buffeli), types 4 to 8, and N1–N3, have been described based on the sequence of the major piroplasm surface protein (MPSP) gene. Of these 11 genotypes, mainly ikeda and chitose are known to be pathogenic and cause considerable morbidity (including high fever, anaemia, jaundice and abortion), production losses and/or mortality in cattle. Mixed infections with two or more genotypes of T. orientalis is common, but do not always lead to a clinical disease, posing challenges in the diagnosis of asymptomatic or subclinical forms of oriental theileriosis. The diagnosis of oriental theileriosis is usually based on clinical signs, the detection of piroplasms of T. orientalis in blood smears, and/or the use of serological or molecular techniques. This paper reviews current methods used for the diagnosis of T. orientalis infections and the genetic characterisation of members of the T. orientalis complex, and proposes that advanced genomic tools should be established for investigations of these and related haemoparasites.

Ticks and accompanying pathogens of domestic and wild animals of Kerala, South India.

The objective of the present study was to detect the chosen nucleotide DNA or RNA sequences of the pathogens in ticks of domestic and wild animals of Kerala, South India based on molecular techniques. Among 602 ticks collected, 413 were from bovines (cattle and buffalo), 26 from goats, 101 from dogs and 62 from wild animals. Amblyomma integrum, Am. gervaisi, Dermacentor auratus, Haemaphysalis bispinosa, Ha. intermedia, Ha. shimoga, Ha. spinigera, Rhipicephalus annulatus, Rh. microplus, Rh. haemaphysaloides and Rh. sanguineus s.l. were identified from various domestic and wild animals of Kerala. The cDNA synthesized from the RNA isolated from fully or partially engorged adult female/nymphal ticks was used as template for the specific polymerase chain reactions (PCR). Out of 602 ticks examined, nucleotide sequences of pathogens were detected in 28 ticks (4.65%). The nucleotide sequences of tick-borne pathogens like Theileria orientalis, Babesia vogeli, Hepatozoon canis, Anaplasma marginale, An. bovis, Rickettsia sp. closely related to Ri. raoultii, Ri. massiliae, Ri. africae and Ri. slovaca were detected. The identification of the previously unreported nucleotide sequences of rickettsial pathogens from India is of particular interest due to their zoonotic significance. The phylogenetic analysis of the major piroplasm surface protein (MPSP) gene of T. orientalis amplified from Rh. annulatus ticks revealed that they were genetically close to type 7, which belong to the highly pathogenic Ikeda group.

Theileria orientalis Ikeda Genotype in Cattle, Virginia, USA.

Theileria orientalis Ikeda genotype is a parasite that causes a disease in cattle that results in major economic issues in Asia, New Zealand, and Australia. The parasite is transmitted by Haemaphysalis longicornis ticks, which have recently been reported in numerous states throughout the eastern United States. Concurrently, cattle in Virginia showed clinical signs consistent with a hemoprotozoan infection. We used amplicons specific for the major piroplasm surface protein and small subunit rDNA of piroplasms to test blood samples from the cattle by PCR. Bidirectional Sanger sequencing showed sequences with 100% identity with T. orientalis Ikeda genotype 2 sequences. We detected the parasite in 3 unrelated herds and from various animals sampled at 2 time points. Although other benign T. orientalis genotypes are endemic to the United States, detection of T. orientalis Ikeda genotype might represent a risk for the cattle industry in Virginia.