Diagnosis of oyster herpes-like virus: development and validation of molecular, immunological and cellular tools "VINO" - Final Report 4th January 1999 to 3rd January 2002

Little information is available on viral infections that affect bivalve molluscs. Such a lack of data is due ta a certain inadequacy of the diagnosis methods that are employed when massive mortality events occur. Most laboratories involved in mollusc pathology still analyse samples through light microscopy. World-wide, there is thus currently a lack of information concerning the occurrence of bivalve herpesvituses. This is probably due to the lack of suitable diagnostic tools. The basic method for identification and examination of suspect samples is predominantly histopathology. This enables the identification of any cellular changes, but is not conclusive identification of bivalve herpesviruses. This technique doesn't allow, by itself, to detect viruses unless it is completed by other methods, such as transmission electron microscopy, the study of cytopathogenic effects in cell cultures or the detection through specific reactives. At present, no bivalve cell-line is available: the detection of cytopathogenic effects in a homologous system is thus impossible. Since invertebrates lack antibody-producing cells, the direct detection of viral agents remains the only possible tool. In these condition, the use of transmission electron microscopy is a necessity for visual confirmation. However, histology and transmision electron microscopy are time consuming and inadequate for epidemiological studies. Viral detection in bivalves may be performed on two kinds of biological material. When mortality events occur, moribund animais may be collected in the affected farms. This fresh material may be immediately used for nucleic acid extraction and further analysis. On the other hand, collected infected organisms can be frozen or fixed and remain archived for long periods of time, constituting a bank of reference material. The aim of the VINO project was therefore the development and validation of molecular, immunological and cellular tools for the diagnosis of, and studies on, the bivalve herpesviruses. The main objective was to develop these 'state of the art' diagnostic techniques. They should be applicable for identification of viruses during disease outbreaks. In addition, these techniques must also be suitable for the detection of subclinical infections and latent virus. The specific objectives of the programme were: 1 - Obtaining the complete oyster herpesvirus (OsHV-1) DNA sequence with determination of the genome structure. 2 - Comparing OsHV-1 with viruses belonging to the Herpesviridae family on the basis of sequence data and genome structure. 3 - Developing molecular tools for OsHV -1 detection. 4 - Developing immunological tools for OsHY-1 detection. 5 - Developing cellular tools for OsHV -1 detection using oyster primary cell cultures and vertebrate cell lines. 6 - Application of developed diagnostic tools for OsHV -1 detection in oyster samples from different geographical locations.