News & Events
PCR Testing for KHV and FLB in Tilapia Now Available for Farmers in Hawaii
Jul 20, 2012
One of the major outcomes of the Center for Tropical and Subtropical Aquaculture (CTSA) supported project entitled, “Operational Biosecurity and Diagnostic Surveillance” was a local diagnostic laboratory capable of employing polymerase chain reaction (PCR) techniques for the detection of selected aquatic pathogens. Through a private-public partnership between Moana Technologies Inc., and the College of Tropical Agriculture and Human Resources (CTAHR) Aquaculture/Aquaponic Extension Program, the capacity to conduct PCR assays have been developed and validated, and the service is currently being used for the detection of two fish pathogens: 1) Koi Herpes Virus (KHV) and 2) Francisella-like bacteria (FLB)1. As the project winds down, it becomes important that this capacity remain available to Hawaii’s aquaculture/aquaponic stakeholders. Moana Technologies LLC is offering PCR testing services for both KHV and FLB.
The tests and associated costs are:
1) PCR Test for KHV + Quality Control
2) PCR Test for FLB + Quality Control
Costs: $75.00/sample for the target pathogen (e.g. FLB) + quality control test. $45.00/sample for target pathogen only.
An example of what a positive test result would look like is provided in Figure 1, and this figure represents part of the validation process for detection of FLB in tilapia spleen (positive control: ADP case # 10-87). Using the primers specific for FLB, its DNA is replicated from a tissue sample that results in a distinct band clearly visible on a gel (left gel Lanes 2 and 3). As an added measure of assurance to the test, primers specific for fish DNA are also replicated on the same samples and they too result in bands (right gel Lanes 2 and 3). These bands, however, migrate differently on the gel indicating that they are a different sequence and size of DNA. This “positive control” for fish DNA becomes particularly important with samples that test negative for FLB, as no band(s) will be present in a negative sample being tested. The only assurance that the sample does contain fish DNA, and that the processing of the sample has not compromised the test result is the positive result for fish DNA.
CONDITIONS FOR TESTING AND REPORTING OF RESULTS:
The nature of the DNA molecule and sensitivity of the PCR assay require that samples be collected and processed in a particular manner. As important KHV is an Office International des Epizooties (OIE)2 listed pathogen and a positive sample will trigger a mandatory reporting process that is already in place and under the oversight of the HDOA State Veterinarian. Lastly, FLB is an emerging pathogen and data on its prevalence in the islands are being monitored by CTAHR’s Aquaculture/Aquaponic extension program and the Hawaii Department of Agriculture (HDOA). To minimize costs, maintain the integrity of the analyses and remain compliant with reporting procedures the following conditions for testing will need to be strictly adhered to:
1. Sample collection, preservation and submission for PCR testing will be conducted by CTAHR’s Aquaculture/Aquaponic Extension Personnel.
2. PCR analyses will be conducted by MOANA Technologies Inc., following already established protocols.
3. Results of the PCR testing will be reported back simultaneously to CTAHR’s Aquaculture/Aquaponic extension program and the HDOA aquatic veterinarian.
4. CTAHR’s project work group will report the results back to the client. Any discussions about the results are to be done with the CTAHR project work group and/or HDOA aquatic veterinarian. Data collected will be used as part of a growing database on the prevalence of these two pathogens in Hawaii.
5. Arrangements for collection of samples on islands other than Oahu will need to be made with CTAHR’s Aquaculture/Aquaponic extension program. Shipping costs will need to be included in addition to the PCR testing.
CTAHR Aquaculture/Aquaponic Extension Program
Department of Molecular Biosciences and Bioengineering
1995 East West Road AgSci 218
Honolulu, HI 96822