Laboratory Diagnosis

Acute Infections

The diagnosis of acute infections must be done during the window of opportunity for virus isolation, as early as 3 days postinfection to 8 or 10 days postinfection. Some animals may be virus isolation positive for only 2 to 3 days during the course of BVDV infection. A whole blood sample is the best sample to collect and submit for BVDV isolation to identify animals acutely infected. A viremia would be detectable in serum by virus isolation during a more narrow window of opportunity than from whole blood. In addition, swabs from mucosal or nasal surfaces can be collected and submitted for virus isolation. Since nucleic acid methods are not influenced by the development of neutralizing antibodies, these methods may remain positive for longer periods during the convalescent stage when neutralizing antibody would interfere with virus isolation. In acute outbreaks in dairy herds, the collection of samples from other herdmates in addition to the severely affected animals may aid in the ability to make positive diagnoses. Whole blood and clotted blood samples should be collected from any animals with mild diarrhea, slight increase in temperature, decrease in feed consumption, or decrease in milk production. Virus isolation should be done on whole blood samples and the serum stored for submission with a paired sample 30 days later for serology. To properly asses serology, paired acute and convalescent samples collected 30 days apart are required to identify four fold increases in serum antibody titers following convalescence. Specimens collected at postmortem examinations from lymphoid organs as mentioned previously should be submitted for virus isolation from any animals that die or fetuses from any abortions that may occur. Many BVDV associated abortions are virus isolation negative. Some aborted fetuses may already have produced antibodies, the presence of which will confirm intrauterine infection. Maternal serology is only seldom helpful because seroconversion has often taken place before the abortion. If the dam on the other hand is antibody negative BVDV can be ruled out as the cause of abortion. Most BVDV associated congenital defects occurring following infection after the onset of immunological competence and therefore calves with these defects have BVDV antibody. The diagnosis of BVDV induced congenital defects in calves should included both virus isolation and serology to detect BVDV-specific antibody prior to uptake of colostrum.

Persistent infections

The identification of persistently infected animals is most routinely done by virus isolation. The level of viremia in persistently infected animals is generally quite high (10⁶ CCID₅₀/ml of serum) but may vary from 10² to 10⁷ CCID₅₀/ml. In addition, the level of viremia may decline in individual animals over time. In most cases, for routine identification of persistently infected animals, serum is adequate for virus isolation. In young calves, maternal antibody will decrease the level of free virus in serum and virus isolation may be falsely negative. Due to colostral antibody, in young calves less than 3 months of age the best sample remains to be whole blood in which the mononuclear cells are separated for virus isolation. It has been suggested that serology can be used to identify seronegative animals as candidates for testing by virus isolation to identify persistent infections. Although persistently infected animals are immunotolerant, they may develop neutralizing antibody titers to BVDV. Therefore, serology should not be used as a screening method to identify animals to test for virus isolation. Since the number of persistently infected animals in any one herd will generally be low, any potentially infected animals should not be excluded from being tested. Persistent infections should only be determined by identification of BVDV by virus isolation in sequential samples collected 30 days apart. By testing the animal 30 days apart it is possible at the same time to test for a four-fold increase in antibody titer should the first virus isolation have been due to acute infection.