Appendix 18
Quantification and duration of Foot-and-Mouth disease virus RNA in bovine esopharyngeal fluid as determined by real-time RT-PCR Zhidong Zhang , Ciara Murphy, Melvyn Quan, Jeanette Knight and Soren Alexandersen Institute for Animal Health, Pirbright, Woking, Surrey, GU24 ONF, U.K.
Abstract: Foot-and-mouth disease virus (FMDV) load was quantitated in experimentally infected cattle in order to improve knowledge about critical features of the initial virus-host interactions which determine disease outcome. A quantitative real-time RT-PCR was used to measure FMDV RNA in oesophageal-pharyngeal fluid (OP-fluid) samples from cattle experimentally infected with type O FMDV by inoculation and contact. The dynamics of FMDV load in OP-fluids exhibited remarkable similarities with viral replication patterns observed in nasal and mouth swab samples for the first week of infection. Viral RNA was recovered at 24 hours post infection (hpi), and rapid viral replication led to peak levels of RNA viral load by 30 hpi to 53 hpi. The extent of virus declined after the peak. Complete clearance of viral RNA occurred in some animals between 7 and 18 dpi. However, viral RNA persisted in OP-fluids at detectable levels beyond 28 dpi in persistently infected cattle (socalled carriers) and was still detectable in some animals at 57 dpi when the experiments were terminated. Viral RNA could not be detected in nasal and mouth swabs from FMDV carriers and non-carriers at any time after 7-18 dpi. An association between the extent of virus replication (growth rate) during acute infection and the outcome of infection (persistence or non-persistence) could not be established. The most significant predictor of the outcome of FMDV infection (persistence or non-persistence) was the extent of viral “clearance” (decay rate) following peak levels. Animals with a slow decay rate became carriers while more rapid clearance of viral RNA was characteristic of non-carriers. Introduction Foot-and-mouth disease virus (FMDV) causes a highly contagious viral disease of domesticated and wild ruminants and pigs. Of considerable importance in the control of FMD is the persistent infection that can occur following clinical or sub-clinical infection in both vaccinated and non-vaccinated ruminants (so-called carriers). There is field evidence to indicate that these carrier animals can precipitate new outbreaks of disease 2. A carrier is defined as an animal from which live virus can be recovered for longer than 28 days after exposure 9,11. Persistent FMDV is eventually eliminated from the carriers, but during the period of persistence, there is considerable variation in the levels of virus recovery from oesophago-pharyngeal fluid (OP-fluid) samples. On the other hand, persistent infection does not occur in all infected ruminants, i.e. only a proportion of infected ruminants become FMDV carriers 2. The mechanism for these phenomena remains to be fully understood although the immune status of the animal prior to contact has been shown not to influence the development of a carrier state. A number of studies on experimentally infected cattle have unequivocally shown the importance of pharyngeal area tissues in FMDV infection and replication during acute disease or persistence 4,5,7,13. During persistence, infectious virus can be isolated from the OP-
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