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Theileria sergenti Shintoku stock consists of 2 parasite populations bearing 2 allelic forms of p33/32, an immunodominant piroplasm surface protein. Parasite population changes during parasite passages among cattle and tick vectors, and during persistent infection in individual calves were analysed by using allele-specific polymerase chain reaction (PCR). The parasite DNAs were prepared from piroplasms from calves which had been infected with Shintoku stock by inoculation of sporozoite stabilates or parasitized erythrocytes, and from sporozoite stabilates which had been prepared from Shintoku stock-infected ticks. Changes in a dominant parasite population were demonstrated during transmission from calves to vector ticks and from infected ticks to calves. Parasite population changes were also apparent during persistent infection in cattle over several months, and this change is thought to occur under host immune pressure. The results of this study indicate that expression of diverse forms of p33/32 may play a role in parasite persistence within mammalian hosts and its transmission from tick vector.

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ABSTRACT. Benign Theileria species distributed in China and Korea were characterized by allele-specific polymerase chain reaction (PCR), based on the sequences of major immunodominant piroplasm surface protein genes. In China, all the isolates contained Chitose (C) type parasites. One out of 5 isolates tested was a mixed population of Ikeda (I), C and B-2 types, whereas, all the isolates from Korea consisted of I type parasites. Except for 4 isolates, 29 isolates from Korea consisted of more than two types of parasites. The present data showed that benign Theileria species distributed in these countries were mixed parasite populations. -KEY WORDS: allele, PCR, Theileria sergenti/ buffeli/orientalis.

ABSTRACT. Two pairs of PCR primers were designed to perform nested PCR targetting of a 540 bp fragment of the nucleocapsid (N) protein gene (N gene) of porcine epidemic diarrhea virus (PEDV). The N gene of PEDV was amplified with 4 PEDV strains and 11 small intestines of PEDV-infected piglets collected from 2 farms in Kagoshima prefecture, Japan. Nucleotide sequences of the PCR products from a Korean and two Japanese strains (KKN96-1 and S1) of PEDV isolated in 1993 and 1996, respectively, were almost identical. These results suggest that the PCR is an available tool for detection of PEDV from pigs in the field, and that the two Japanese strains (KKN96-1 and S1) were genetically similar to the Korean strain.-KEY WORDS: nested PCR, nucleocapsid protein gene, porcine epidemic diarrhea virus.

Bovine piroplasmosis caused by Theileria sergenti is a major cause of economic loss in grazing cattle in Japan. Infected calves show chronic anaemia with intraerythrocytic piroplasms and occasionally die in severe cases. We found that parasite stocks and isolates consist of genetically and antigenically mixed populations. To differentiate parasite populations bearing 3 allelic forms of p32/34, an immunodominant piroplasm surface protein, 3 sets of oligonucleotide primers were designed to amplify either of 3 alleles by polymerase chain reaction (PCR). By using this allele-specific PCR, we found that the majority of T. sergenti-infected calves in Japan harbored mixed parasite populations bearing C and I type parasites. To control Theileria infection, we produced 2 vaccine candidates: recombinant baculovirus p32 and synthetic peptide containing Lys-Glu-Lys (KEK) motif. Immunization with either recombinant p32 or synthetic peptide containing KEK sequences with Freund's complete adjuvant resulted in low parasitemia and reduced the clinical symptoms compared to control calves. Interestingly, the parasite with the p32 allelic form corresponding to the one used as the immunogen was suppressed.

ABSTRACT. An survey of Theileria parasite infection in cattle in Taiwan was carried out by polymerase chain reaction (PCR). A total of 491 blood samples, 105 from southern area and 386 from northern area, were collected from bovine in 16 different farms. From northern area, Theileria piroplasms could be seen in only 4 of 105 blood samples microscopically. However, when p32/34 genes (encoding immunodominant piroplasm surface proteins) were amplified by PCR, 15 blood samples were detected positive. They were analyzed by using allele-specific primers of 3 allelic forms of p32/34 and all contained C type of T. sergenti. Four blood samples were found infected with both C and B (T. buffeli) type parasites. Examination of 386 blood samples from southern area of Taiwan did not reveal any Theileria parasite microscopically, as well as by PCR amplification. -KEY WORDS: PCR, Taiwan cattle, Theileria sergenti.

Epitopes on a 32 kDa protein, which is an immunodominant major surface protein of Theileria sergenti, recognized by anti-merozoite monoclonal antibodies were characterized. The results of a competitive binding assay between monoclonal antibodies indicated that there were at least three epitopes in this protein. The presence of repeated epitopes was suggested by using two-site enzyme-linked immunosorbent assay. The protein was partitioned into the detergent phase of Triton X-114 extracts, indicating that the 32 kDa protein is an integral membrane protein. Periodate treatment of 32 kDa protein implies that one epitope of the epitopes recognized by monoclonal antibody has a carbohydrate moiety.