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→Virus Identification
=====Laboratory Tests=====
=====Laboratory Tests=====
======Virus Identification======
======Virus Identification======
The virus is identified by complement fixation (CF), immunofluorescence, PCR, ELISA, virus isolation or plaque reduction neutralisation (PRN) tests.
The virus is identified by complement fixation, immunofluorescence, PCR, ELISA, virus isolation or plaque reduction neutralisation tests.
*The Complement Fixation Test can be used to identify Eastern or Western EEV in infected mouse or chicken brains, cell culture fluid or amniotic-allantoic fluid
*Immunofluroescence: virus may be identified in brain tissue or cell culture using direct immunofluorescent staining.
*PCR: EEE and WEE viral RNA may also be detected by reverse-transcription PCR. EEE virus nucleic acid in mosquitoes and tissues has been identified by the polymerase chain reaction (PCR) using primers selected from the capsid gene (15). An alternate identification procedure is by hybridisation with an oligonucleotide probe. A reverse-transcription PCR method for detection of WEE RNA and alternative methods for EEE RNA detection have also been described (5, 7).
*ELISA can be used to detect virus in brain tissue. An antigen-capture ELISA, developed for EEE surveillance in mosquitoes, can be used where virus isolation and PCR facilities are unavailable (1).
Virus isolation
Virus isolation
Virus isolation is the most definitive method for diagnosis of EEE or WEE. EEE virus can usually be isolated from the brains of horses, unless more than 5 days have elapsed between the appearance of clinical signs and the death of the horse. EEE virus can frequently be isolated from brain tissue even in the presence of a high serum antibody titre. WEE virus is rarely isolated from tissues of infected horses. Brain is the tissue of choice for virus isolation, but the virus has been isolated from other tissues, such as the liver and spleen. It is recommended that a complete set of these tissues be collected in duplicate, one set for virus isolation and the other set in formalin for histopathological examination. Specimens for virus isolation should be sent refrigerated if they can be received in the laboratory within 48 hours of collection; otherwise, they should be frozen and sent with dry ice. Newborn mice, chicken embryos and a number of cell culture systems can be used for virus isolation. Virus may be isolated from the CSF of acutely infected horses
Virus isolation is the most definitive method for diagnosis of EEE or WEE. EEE virus can usually be isolated from the brains of horses, unless more than 5 days have elapsed between the appearance of clinical signs and the death of the horse. EEE virus can frequently be isolated from brain tissue even in the presence of a high serum antibody titre. WEE virus is rarely isolated from tissues of infected horses. Brain is the tissue of choice for virus isolation, but the virus has been isolated from other tissues, such as the liver and spleen. It is recommended that a complete set of these tissues be collected in duplicate, one set for virus isolation and the other set in formalin for histopathological examination. Specimens for virus isolation should be sent refrigerated if they can be received in the laboratory within 48 hours of collection; otherwise, they should be frozen and sent with dry ice. Newborn mice, chicken embryos and a number of cell culture systems can be used for virus isolation. Virus may be isolated from the CSF of acutely infected horses
======Serology======
======Serology======