New improved PCR method to detect AHPND bacteria

Vibrio parahaemolyticus is a specific type of bacteria that causes acute hepatopancreatic necrosis disease (AHPND) (also called early mortality syndrome or EMS) in shrimp, resulting in extensive damage to both domestic and international shrimp industries. Thus, a group of Thai and Taiwanese researchers previously developed an interim PCR method to detect isolates of AHPND bacteria until the lethal toxin gene could be identified. To be more precisely identify isolates of V. parahaemolyticus causing AHPND, Center of Excellence for Shrimp Molecular Biology and Biotechnology (Mahidol-BIOTEC cooperative center) collaborated with the Aquatic Animal Health Research Center, Charoen Pokphand Company to develop a better PCR detection method.

This new improved PCR method is based on the gene sequence of a protein discovered in a sub-fraction of cell-free culture broth from isolates of V. parhaemolyticus that cause AHPND. This protein is not present in V. parahaemolyticus or other bacteria that do not cause AHPND. This cell-free preparation caused the typical signs of acute AHPND (massive sloughing of hepatopancreatic tubule epithelial cells) when administered to shrimp by reverse gavage. Primer (AP3) was designed to amplify the gene sequence of one small protein from this toxic sub-fraction. The AP3 method gave 100% sensitivity, specificity, positive predictive value and negative predictive value for the detection of AHPND bacteria using 98 bacterial isolates characterized by bioassay as AHPND and non-AHPND bacteria.


The research has been supported by the National Research Council of Thailand, the Agriculture Research and Development Agency, the Thai Commission for Higher Education, Mahidol University, Faculty of Marine Technology at Burapha University, the National Science and Technology Development Agency, the Patani Shrimp Farmers Club, the Surathani Shrimp Farmers Club, the Thai Frozen Foods Association, Charoen Pokphand Company, SyAqua Co. Ltd. and Thai Union Co. Ltd.

Download: Announcement regarding free release of a new and improved PCR method for detection of AHPND bacteria

Posted on 25 June 2014