›› 2009, Vol. 27 ›› Issue (2): 9-139.

• 实验研究 • Previous Articles     Next Articles

Identification of Calf-origin Cryptosporidium bovis Shanghai Isolate by Nested PCR

YUAN Zhong-ying1，SHEN Yu-juan1 *，CAO Jian-ping1，LIU Hui1，2，
CHEN Sheng-xia3   

  1. 1 National Institute of Parasitic Diseases，Chinese Center for Disease Control and Prevention，WHO Collaborating Centre for Malaria，Schistosomiasis and Filariasis，Shanghai 200025，China；2 Department of Parasitology，Zunyi Medical College，Zunyi 563003，China；3 School of Medical Science and Laboratory Medicine，Jiangsu University，Zhenjiang 212013，China
  • Received:1900-01-01 Revised:1900-01-01 Online:2009-04-30 Published:2009-04-30

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Abstract: Objective To identify a strain of Cryptosporidium in the feces of naturally infected calf in Shanghai. Methods Stool sample was examined by modified acid-fast staining. The size and morphology of the oocysts were micros-copically determined. Genomic DNA was extracted from the oocysts isolated from feces of a naturally Cryptosporidium-infected calf. According to the sequence of Cryptosporidium 18S rRNA gene, two pairs of primers were designed and syn-thesized. The PCR products was amplified by nested PCR and sequenced in double directions. Homology searches were done over the Web using the program Blast. Phylogenetic tree was constructed with NJ method by MEGA4.0 software. Results Oocysts of the Shanghai isolate were round or elliptical with a size of （5.6±0.49） μm ×（5.2±0.51） μm. Nested PCR resulted in fragments of approximately 810 bp, and the 18S rRNA nucleotide sequence had 100% identity with C. bovis from Brazil （GenBank Accession No: 151935628）. This isolate was clustered in the same clade with C. bovis from Brazil. It showed an identity of 99% with the sequences of C. bovis from Qinghai Ｐrovince of China, Mongolia, USA, and Tunisia. Conclusion The calf-origin Cryptosporidium derived from Shanghai has been identified as C. bovis.

Key words: Cryptosporidium bovis, 18S rRNA gene, Nested PCR, Phylogeny