Background

In 1997 and 1998, a disease causing mass mortalities and associated with the presence of a herpes-like virus, occurred in koi carp from several countries including Germany, UK, the Netherlands, Israel and USA (Bretzinger et al. 1999).  The virus, koi herpesvirus (KHV, but subsequently renamed Cyprinid herpesvirus-3, CyHV-3), has since spread to many other countries around the world, devastating the carp and koi industries of SE Asia.   The virus affects both common and koi carp varieties specifically (Haenen et al. 2004)  The other finfish species present during these outbreaks were unaffected.  KHV  has not established in Australia to date.

In Australia, carp are widely regarded as an introduced environmental pest.  There are no native closely-related species.  This project is assessing the technical feasibility of using KHV as a biological control agent for carp in Australia based on information gathered from controlled experiments in a secure laboratory environment. 

Testing the relevance of these laboratory results in the field would be reserved for a future phase that would also, of necessity, include a discussion of the politico-socio-economic aspects of using KHV as a biological control agent, in a similar manner to how rabbit haemorrhagic disease virus has been used to successfully control rabbits.

Progress to date

UK and USA isolates of Koi herpesvirus (KHV) have been imported into the secure area of the Fish Diseases Laboratory at the Australian Animal Health Laboratory.

The cell-lines that are necessary for culturing these viruses have also been imported.  All of these reagents have been tested by a series of KHV-specific polymerase chain reaction (PCR) assays in order to:

1. confirm the presence of KHV in the virus samples, and

2. demonstrate the absence of virus in the uninfected cell lines.

In addition, these reagents have all been innocuity-tested, ie, to make sure that they are pure and contain no unexpected pathogens.  Recently, a further isolate of KHV and a new cell-line have also been imported from Indonesia, and this too has been tested in a similar manner. 

As part of establishing the tools to complete the project, procedures for the isolation of KHV in cell culture have been established, and small volumes of virus have been stored for future use.  We have now gained a lot of experience in, and developed an understanding of, how to handle this particular herpesvirus in cell culture.

Further tools essential for the completion of the project are the molecular methods for the detection and identification of KHV by PCR. These have also been established in the lab. We have 5 different conventional KHV-specific PCRs now available to us, and, following specificity and sensitivity testing, we generally use the official Japanese diagnostic PCR  (Yuasa et al, 2005) as our standard molecular test. 

Our expertise with these procedures has been tested in a KHV molecular diagnostics proficiency test that was co-ordinated by the Weymouth Laboratory in the UK.  Our results from that test matched perfectly with those of the co-ordinating lab.  We have also replicated a KHV real-time PCR in the lab. This PCR was developed by Gilad et al (2004), and we are currently setting it up as a quantitative PCR (qPCR).

Finally, we are in the process of developing another PCR for the detection of cyprinid herpesviruses in general.  We hope that this PCR will be able to detect any herpesvirus of cyprinids and any other currently unrecognized such viruses.  

The next step is to test the virus on live carp in laboratory conditions.  This part of the project has been delayed due to the unavailability of sufficient carp of the appropriate size (<10 cm) during the Spring and Summer (2006-07) because of the drought.   A catch in October 2006 found that the carp were not in a sufficiently healthy state (in the wild) to participate in the trial.

In mid-May 2007, a preliminary susceptibility trial was started using 16 large carp (30-35 cm) caught from the wild.  The team are conducting experiments to determine the persistence of KHV in the environment.