Detection of Pseudorabies Virus from Oral Fluids of Experimental Infected Pigs

Pseudorabies is one of the important infectious diseases, causing enormous economic losses to swine industry. Pseudorabies virus (PRV) is the causative agent that is monitored mainly from nasal swabs and tissue samples collected from pigs. Oral swabs have been widely used for the detection of classical swine fever virus (CSFV), foot-and-mouth disease virus (FMDV), porcine circovirus type 2(PCV2) and porcine reproductive and respiratory virus (PRRSV) in swine herds due to its simplicity in sampling. However, there is no report about the application of oral fluids sampling for PRV surveillance till now. In this study, we attempted to collect the oral fluids of experimental infected pigs based on the oral cotton swabs for further PRV surveillance.

The new PRV-HNX isolate was propagated in Vero cells, and the virus titer was determined. 35-day-old PRV negative pigs were randomly divided into 2 groups of 6 and treated with PRV infection (2 mL, 1×107 TCID50/mL) and equal volume of DMEM, respectively. Sterilized cotton ropes were hung in the pens for about 30 min for pig biting to collect the oral fluids at 0d, 1-14d, 21d, and 28d post infection, respectively. Meanwhile, the nose swab was also collected. The oral fluids collected were freezing and thawing for three times, and then centrifuged at 8000 rpm/min for 5 min. The supernatant was used for DNA extraction. Then, the conventional gD-PCR and real-time PCR were performed to monitor the presence of PRV qualitatively and quantitatively.

Conventional PCR detection indicated that PRV was positive at 1-28d post infection, while it was negative prior to infection. And the PRV was negative in control group all the time. Data from real-time PCR showed that PRV was positive at 1-28d post infection. Additionally, the copies of PRV detected were consistently increasing at 1-4d post infection, and peaked at 4d post infection. Meanwhile, the method based on oral fluids sampling can detect more viruses than from nose swabs. Clinical validation test was also successful.

In the present study, we first demonstrated that oral fluids collected by oral cotton swabs can be used for PRV detection. Compared with nose swab, we concluded that the oral cotton swab has higher sensitivity and superiority to be applied for clinical PRV surveillance.

This work was supported by grants from the China Agricultural Research System (CARS-36).

Disclosure of Interest: None Declared.