Abstract: According to the gene sequences in GenBank of Haplosporidium sp, three specific primers were designed for amplifying the specific fragments of Haplosporidium sp. The reaction parameters were optimized to develop the semi-nested PCR method for detection of Haplosporidium sp. The 333 bp specific DNA fragment of Haplosporidium sp was amplified, but not from other pathogenics such as Marteilia refringens,Perkinsus sp, Vibrio parahaemolyticu, Vibrio alginolyticu and Vibrio fluvialis by this semi-nested PCR. The sensitivity results showed that as little as 0.01 fg DNA plasmid of Haplosporidium sp was detected by this semi-nested PCR. Shellfish samples of Shandong coastal were detected by this semi-nested PCR. As the result, 6 Oyster and 8 Manila clam were Haplosporidium sp positive. It suggested that Haplosporidium sp existed in the cultivated shellfish in Shandong and this semi-nested PCR could be used as a sensitive tool to detect Haplosporidium sp in clinical samples.

Key words: shellfish; Haplosporidium sp; semi-nested PCR