Marek Matras, Jerzy Antychowicz, Jeannette Castric and Sven M. Bergmann
The KHV free carp were experimentally infected with CyHV-3 suspension by immersion at temperatures of 18ºC±1 and 25ºC±1. Skin, brain, gills, liver, spleen, kidneys, and leukocytes were investigated for the presence of KHV DNA, and sera for the presence of antibodies against the virus. Similar investigations were performed with non-infected carp, designed as control groups. The results of KHV DNA detection in the infected carp kept at 25ºC ±1 were compared with the results of the virus isolation onto common carp brain cells using leukocyte co-cultivation method and results of specific antibody detection. The appearance of specific antibodies at various time intervals after detecting the clinical symptoms of KHVD in naturally infected carp was additionally studied. At 18°C water temperature, KHV DNA was not detected up to the 6th d post infection (dpi). Then it was found in samples from skin, gill, liver, spleen, and kidneys and much later from the brain. At 25°C water temperature, KHV DNA was already detected on the 3rd dpi in samples from skin and gills, and from the 5th dpi it was found in all examined internal organs, except the brain where it was found at 6th dpi. At 25°C, isolations of CyHV-3 have succeeded between the 7th and 11th dpi. The first specific antibodies were found no sooner than on the 21st dpi. The serological examination performed in naturally infected carp in 15 carp pond farms showed that first specific antiviral antibodies can be detected 2 weeks after the appearance of clinical symptoms of KHVD.
Marek Matras, Magdalena Stachnik, Ewa Borzym, Joanna Maj-Paluch and Michał Reichert
Koi herpesvirus (KHV) has infected farmed common carp in Poland clinically and asymptomatically since 2004. The role of non-carp species as vectors of virus transmission is well known except for in the case of KHV. The aim was to better understand this virus’ infection and transmission pathways in common carp, looking at the potential vector role of fishes kept with them.
Material and Methods
Eight species were experimentally infected with KHV by immersion in a suspension at 20°C ±1 and transferred to a tank after 45 minutes. Specimens were euthanised at intervals up to 56 days post infection (dpi) and tissue was examined for KHV DNA. Surviving infected fishes were introduced at intervals, each time into a separate tank, to naïve common carp for experimental infection. These were observed daily for symptoms, sacrificed along with controls after three months, and dissected to provide tissue samples. Also fish from 14 species collected from a farm with a history of KHV were sampled from 3 to 22 months after disease was confirmed. Organ sections from single fish were collected in a single tube.
Viral DNA was detected in tench and roach samples up to 49 dpi, but in three-spined stickleback and stone maroko samples only up to 14 dpi. Transmission of KHV to naïve carp occurred after cohabitation. KHV DNA was detected in three fish species three months after the farm outbreak.
We confirmed that grass and Prussian carp, tench, roach, and brown bullhead can transfer the virus to naïve common carp.
Ewa Borzym, Joanna Maj-Paluch, Magdalena Stachnik, Marek Matras and Michał Reichert
The aim of the study was to identify the genotype of Polish isolates of salmonid alphaviruses (SAV) and to find the origin of the virus. Samples for virus isolation included the kidneys, spleen, and liver pooled from 10 fish. A typical cytopathic effect was observed after inoculation of samples on cell lines. Total RNA was extracted from cell culture supernatant and submitted to RT-PCR with primers amplifying two informative regions of the genome: a conserved region in the E2 gene and a variable region in the nsP3 gene. The sequences revealed that the strain from Poland belonged to subtype SAV 2, indicating a very strong genetic identity with isolates from Italy and France.