AUSTRALIA–INDONESIA JOINT SYMPOSIUM IN SCIENCE AND TECHNOLOGY

Jakarta, 13-17 September 2006

A novel PCR-based detection system specific for pathogenic Vibrios in shrimp hatchery
Professor Antonius Suwanto, Bogor Agricultural University, Bogor and
Research and Development Center, CPI, Jakarta

Vibrio harveyi, and V. campbelli have long been identified as an important pathogen in shrimp aquaculture. Surveillance and quantification of Vibrio has become a routine procedure in shrimp quality control systems. Current detection of Vibrios most frequently utilizes specific and differential media such as Thiosulphate Citrate Bile Sucrose (TCBS) agar. While it can specifically detect and enumerate Vibrio, it has limited capacity for discriminating Vibrio harveyi – the species of interest – from other Vibrio species.

Here we report the development of a PCR-based detection system specific to Vibrio harveyi. We have designed 2 novel primer sets, one specific to the Vibrio group and the other specific to Vibrio harveyi. A total of 29 Vibrio isolates were tested with these primers. Amongst these isolates, V. harveyi ATCC 14126 and V. charcariae ATCC 35084 type strains were used as positive controls. Other Vibrio isolates were gathered from shrimp hatcheries and environmental samples. All Vibrio isolates generated positive results with our Vibrio-specific primer set. From these 29 isolates, 13 were positive when tested with the V. harveyi specific primer set. In parallel with PCR detection, a microbiological survey using Vibrio harveyi Agar (VHA) (Harris et al., 1996) was conducted on 25 Vibrio isolates. Using VHA, 13 isolates gave positive morphological characteristic of V. harveyi and these results were in complete agreement with our PCR-based detection system.

PCR analysis can be conducted directly from colonies on agar plates, and can be completed within 2 hours. By contrast, the microbiological method (VHA test) required at least 48 hr to generate typical V. harveyi morphology. Thus, the PCR-based method described here has potential application in the aquaculture industry where rapid and accurate V. harveyi-campbelli detection is needed.