DAO 66:163-170 (2005)  -  doi:10.3354/dao066163

In vivo titration of white spot syndrome virus (WSSV) in specific pathogen-free Litopenaeus vannamei by intramuscular and oral routes

C. M. Escobedo-Bonilla1,2, M. Wille1, V. Alday Sanz3, P. Sorgeloos1, M. B. Pensaert2, H. J. Nauwynck2,*

1Laboratory of Aquaculture & Artemia Reference Center, Faculty of Bioscience Engineering, Ghent University, Rozier 44, 9000 Ghent, Belgium
2Laboratory of Virology, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, 9820 Merelbeke, Belgium
3INVE technologies, Hoogveld 93, 9200 Dendermonde, Belgium
*Corresponding author. Email:

ABSTRACT: White spot syndrome virus (WSSV) is a devastating pathogen in shrimp aquaculture. Standardized challenge procedures using a known amount of infectious virus would assist in evaluating strategies to reduce its impact. In this study, the shrimp infectious dose 50% endpoint (SID50 ml–1) of a Thai isolate of WSSV was determined by intramuscular inoculation (i.m.) in 60 d old and 135 d old specific pathogen-free (SPF) Litopenaeus vannamei using indirect immunofluorescence (IIF) and one-step polymerase chain reaction (PCR). Also, the lethal dose 50% endpoint (LD50 ml–1) was determined from the proportion of dead shrimp. The median virus infection titers in 60 d old and 135 d old juveniles were 106.8 and 106.5 SID50 ml–1, respectively. These titers were not significantly different (p ≥ 0.05). The titration of the WSSV stock by oral intubation in 80 d old juveniles resulted in approximately tenfold reduction in virus titer compared to i.m. inoculation. This lower titer is probably the result of physical and chemical barriers in the digestive tract of shrimp that hinder WSSV infectivity. The titers determined by infection were identical to the titers determined by mortality in all experiments using both i.m. and oral routes at 120 h post inoculation (hpi), indicating that every infected shrimp died. The determination of WSSV titers for dilutions administered by i.m. and oral routes constitutes the first step towards the standardization of challenge procedures to evaluate strategies to reduce WSSV infection.


KEY WORDS: Litopenaeus vannamei · WSSV · In vivo titration · Intramuscular inoculation · Oral inoculation


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