Detection and identification of Theileria infection in sika deer ( Cervus nippon ) in China.

The sika deer ( Cervus nippon ) is a first-grade state-protected animal in China and designated a threatened species by the World Conservation Union. To detect hemoparasite infection of sika deer, blood samples were collected from 24 animals in the Hubei Province Deer Center. Genomic DNA was extracted, and the V4 hypervariable region encoding 18S rRNA was analyzed by reverse line blot hybridization assay. PCR products hybridized with Babesia / Theileria genus-specific probes but failed to hybridize with any of the Babesia or Theileria species-specific probes, suggesting the presence of a novel, or variant, species. Here 18S rRNA and internal transcribed spacer (ITS) genes were amplified, cloned, and sequenced from 7 isolates. Alignment and BlastN of the cloned sequences revealed high similarities to the homologous 18S rRNA genes and ITS genes of Theileria cervi (AY735122), Theileria sp. CNY1A (AB012194), and Theileria sp. ex Yamaguchi (AF529272). Phylogenetic analysis based on the 18S rRNA gene and ITS sequences showed that all cloned sequences were grouped within the Theileria clade. Phylogeny based on the 18S rRNA gene divided the organisms into 2 groups. Group 1 was closest to Theileria sp. ex Yamaguchi (AF529272), and group 2 was distinct from all other identified Theileria and Babesia species. These results suggest the existence of Theileria sp. infection in sika deer in China. To our knowledge, this is the first report of cervine Theileria sp. in China.

Prevalence of coccidian infection in suckling piglets in China.

To determine the prevalence of coccidian infection in suckling piglets in China, fecal samples from 779 litters of suckling piglets were collected on 80 different farms in 17 provinces from September 2009 to December 2010. These samples were examined through saturated saline flotation technique. The prevalences of coccidian infection ranged from 0 to 32.5% among different provinces and the average was 16.7% (130/779). The highest prevalence of 19.9% (69/346) was found in 8-14 day-old litters of suckling piglets. Seven coccidian species were detected in the positive litters of suckling piglets, including Isospora suis (63.9%), Eimeria debliecki (46.9%), Eimeria polita (19.2%), Eimeria suis (20.8%), Eimeria perminuta (13.9%), Eimeria scabra (4.6%), and Eimeria yanglingensis (1.5%). 55.4% of the positive litters of suckling piglet infected more than one coccidian species. The results of this investigation will provide the relevant basic data for control strategies against porcine coccidiosis on pig farms in China.

Development and evaluation of real-time PCR assay for the detection of Babesia orientalis in water buffalo (Bubalus bubalis, Linnaeus, 1758).

Babesia orientalis is the causative agent of babesiosis in water buffalo (Bubalus babalis, Linnaeus, 1758). In this study, a TaqMan real-time PCR assay was developed for quantitative detection of B. orientalis in water buffalo. Hybridization probe and oligonucleotide primers were designed based on the v4 region of 18S rRNA gene. Detection limit was determined at 2 parasites. Blood samples were collected from experimentally infected water buffalo, as well as from 180 field samples, which were collected from 4 different geographical locations to the north and south of the Yangtse River. The parasite was detected by real-time PCR on day 2 until day 39 post-infection, while reverse line blot (RLB) was on day 6 until day 36 in experimentally infected water buffalo. For the results of 180 field samples, statistical analysis showed no significant difference in relative effectiveness of real-time PCR and RLB. The analysis also indicated that there was no difference in the prevalence of B. orientalis between the regions of south and north of the Yangtse River by both the real-time PCR assay and RLB detection. These results indicated that the parasite infection has spread to the north of the Yangtse River.