Evaluation of rotavirus dsRNA load in specimens and body fluids from experimentally infected juvenile macaques by real-time PCR

• Published in: Virology (New York, N.Y.) N.Y.), 2005-10, Vol.341 (2), p.248-256
• Main Authors: Zhao, Wei, Xia, Mingjing, Bridges-Malveo, Tamika, Cantú, Mayra, McNeal, Monica M., Choi, Anthony H., Ward, Richard L., Sestak, Karol
• Format: Article
• Language: English
• Subjects: Animals; Body Fluids - virology; Cell Line; Cerebrospinal Fluid - virology; Disease Models, Animal; dsRNA; Enzyme-Linked Immunosorbent Assay; Feces - virology; Macaca mulatta; Nested PCR; Plasma - virology; Polymerase Chain Reaction - methods; Real-time PCR; Rhesus; RNA, Double-Stranded - analysis; RNA, Viral - analysis; Rotavirus; Rotavirus - genetics; Rotavirus - isolation & purification; Rotavirus Infections - virology; RT-PCR; Sensitivity and Specificity; Viral Load; Viremia; Virus Shedding
• ISSN: 0042-6822; 1096-0341
• DOI: 10.1016/j.virol.2005.06.048

We recently established a non-human primate model of rotavirus infection that is characterized by consistent and high levels of virus antigen shedding in stools. Here, we report that starting from post challenge day (PCD) 2, 6 × 10 3 to 1.5 × 10 6 copies of rotavirus double-stranded RNA per nanogram of total RNA were detected by real-time PCR in MA104 cells that were 48 h pre-incubated with filtered stool suspensions of three experimentally infected juvenile macaques. The peak of virus load was detected at PCD 4–5, followed by decreased load at PCD 6–11, and very low levels at PCD 12. Such a pattern corresponded to virus shedding in stools as reported recently based on enzyme-linked immunosorbent assay (ELISA) results. In addition, plasma and cerebrospinal fluids (CSF) from six infected animals were tested for the presence of rotavirus. Rotavirus extraintestinal escape was revealed in three out of six animals by a combination of real-time and nested PCR. However, very low quantities of detected viral RNA (∼20 copies/ng of total RNA) were not suggestive of viremia. Thus, the rhesus model of rotavirus infection can be exploited further in studies with vaccine candidates designed to prevent or abrogate rotavirus infection.