1. EXCERCISE 2: SUSPICION ON THE FARM BIG TROUT SITUATED ON BLACK RIVER DIAGNOSTIC METHOD USED BLUE GROUP

2. DIAGNOSTIC METHODS USED DISSECTION (increased mortality, exophtalmia, dark pigmentation, hemorrhagies on piloric caeca, visceral organs, enlarged spleen, serosa) PARASITOLOGICAL INVESTIGATION - NEGATIVE SUSPICION 1.BACTERIA-samples of organs plated onto BA and TSA – NEGATIVE AFTER 48h of incubation on 15 o C

3. VIRUS ISOLATION VIROLOGICAL EXAMINATION – Sampled organs:heart, kidney and spleen homogenized and inoculated on EPC and BF2 cell lines incubation on 15 o C for 7 days;

4. VIRUS IDENTIFICATION Identification – cell culture supernatant was tested by ELISA tests for the presence of viruses: VHSV, IHNV and IPNV Positive finding of VHSV, tests to other viruses were negative

5. COMMUNICATION WITH REFERENCE LABORATORY When virus was confirmed, cell culture supernatant was sent to NRL for confirmation of VHSV presence in the sample Diagnostic procedure in NRL: Cell culture (EPC, BF2) IFAT for presence of VHSV, IHNV & IPNV VHSV positive material was confirmed by means of RT-PCR

6. VIRUS GENOTYPING In the following two weeks PCR product was submitted to sequencing Results showed that it is VHSV, Genotype I

7. Investigation possible connection of virus genotype isolated in surrounding coutries Sequence of G gene showed 90 % similarity with VHSV isolates, genotype I isolated in Germany and Poland