Detection of koi herpesvirus DNA in tissues of infected fish

Abstract

A newly recognized herpesvirus, koi herpesvirus or KHV, causes a lethal disease in common carp, Cyprinus carpio, and its colourful strain known as koi or fancy carp. In this study, we report new outbreaks of the disease, present initial characterization of the KHV genome, and describe assays for detection of KHV DNA in infected cells and tissues of infected fish. Restriction endonuclease (RE) profiles of viral DNA derived from two epidemiologically distinct KHV isolates were identical to each other. Cloned KHV BamHI and SphI DNA probes specifically hybridized to KHV DNA, but not to DNAs derived from a variety of other fish herpesviruses. The KHV DNA probes detected KHV DNA in tissues of experimentally infected koi fish by DNA hybridization. The KHV specific polymerase chain assays (PCR) were developed for rapid detection and confirmation of KHV DNA in tissues of infected fish.

An Unusual Koi Herpesvirus Associated with a Mortality Event of Common Carp Cyprinus carpio in New York State, USA

“…comm. ), and three KHV primer sets (Gilad et al, 2002;Gray et al, 2002). The different reaction conditions for the different primer sets are outlined in Table 1.…”

Koi Herpes Virus: A Review and Risk Assessment of Indian Aquaculture

“…The incubation period of virus inside the host ranges between 7 and 10 days before the onset of clinical symptoms. The clinical signs are lethargy, loss of appetite, gill necrosis, haemorrhages on the body and uncoordinated swimming [29,60]. The infected fish begin to die within 1 or 2 days after the onset of the symptoms [34].…”

“…Viral DNA can be detected in the blood and kidney, 1 day after virus exposure [17,65]. Subsequently, DNA can be detected in the gill, intestinal tract, spleen and liver, but not in the brain [29]. Virus is most abundant in gill, kidney, and spleen during the course of infection [27].…”

“…Several diagnostic tests based on PCR [6,25,29,89], nested PCR [7,20], real-time PCR [27] and loop-mediated isothermal amplification (LAMP) [31,75,87,88] have been developed for detection of KHV. A highly sensitive PCR method for detection of TK gene of KHV is able to detect 10 fg of KHV DNA [6].…”

“…A highly sensitive PCR method for detection of TK gene of KHV is able to detect 10 fg of KHV DNA [6]. The sensitivity of this method is *10-1,000 times greater than other PCR methods [25,29]. PCR primers targeting DNA polymerase gene and the major envelope protein gene of KHV have a sensitivity of 100 femtogram (fg) and 1,000 fg of KHV DNA, respectively in infected gills [41].…”