A specific interaction between the F and H proteins is required to enable fusion of the virus and host cell membranes and in some cases these proteins are not interchangeable between related viruses of the family Paramyxoviridae. For example, the F and H proteins of two ruminant morbilliviruses, rinderpest virus (RPV) and Peste-des-petits-ruminants virus (PPRV), are not interchangeable since viable virus could not be rescued from cDNA constructs where an individual glycoprotein gene of RPV was replaced with that from PPRV. To investigate which domain of the H protein, extracellular or cytoplasmic/transmembrane, was most important for preventing this interaction, two chimeric H gene constructs were made where the normal H gene of RPV was substituted with variant H genes where the transmembrane/cytoplasmic tail region (pRPV2C-PPRTm) or the whole ectodomain (pRPV2C-PPRExt) were derived from PPRV. Chimeric viruses were rescued from both the constructs and, while RPV2C-PPRTm virus grew to as high titres as the parent virus, RPV2C-PPRExt virus was extremely debilitated with respect to growth in tissue culture. Thus the ectodomain of H is the most important region required for effective interactions of the two glycoproteins for the recovery of viable virus. Nevertheless, the transmembrane/cytoplasmic domain of RPV alone can allow a chimeric virus to be rescued, which was not possible when the complete H gene was derived from PPRV. Both versions of the H protein and also the F protein were found to be incorporated into the envelope of the budded virions.

There has been much debate about the use of the so-called "vaccinate-to-live" policy for the control of foot-and-mouth disease (FMD) in Europe, according to which, spread of the FMD virus (FMDV) from future outbreaks could be controlled by a short period of "emergency" vaccination of surrounding herds, reducing the need for large-scale pre-emptive culling of at-risk animals. Since vaccinated animals may become subclinically infected with FMDV following challenge exposure, it is necessary to either remove all vaccinates (vaccinate-to-kill) or to detect and remove vaccinates in which virus is circulating or has established persistent infections (vaccinate-to-live), in order to rapidly regain the most favoured trading status of FMD-free without vaccination. The latter approach can be supported by testing vaccinated animals for the presence of antibodies to certain non-structural proteins (NSP) of FMDV, which are induced by infection with the virus, but not by vaccination with purified FMD vaccines. Using test sensitivity and specificity data established at a recent workshop on NSP assays [Brocchi E, Bergmann I, Dekker A, Paton DJ, Sammin DJ, Greiner M, et al. Comparative performance of six ELISAs for antibodies to the non-structural proteins of foot-and-mouth disease. Vaccine, in press], this paper examines the ways in which serological testing with NSP ELISAs can be used and interpreted and the effect that this will have on the confidence with which freedom from infection can be demonstrated within guidelines specified by the World Animal Health Organisation and the European Commission.

Foot-and-mouth disease virus (FMDV) can be excreted in milk and thereby spread infection to susceptible animals in other holdings. The feasibility of using real-time reverse transcription polymerase chain reaction (rRT-PCR) as a diagnostic tool for detection of FMDV in milk was assessed by studying the excretion of virus from experimentally-infected cattle. Fore- and machine milk samples were collected over a 4-week period from two dairy cows infected with FMDV and from two in-contact cows held in the same pen. The whole, skim, cream and cellular debris components of the milks were tested by automated rRT-PCR and results compared to virus isolation (VI) in cell culture. The onset of clinical signs of FMD in all four cows correlated with viraemia, and the presence of FMDV in other clinical samples. rRT-PCR results matched closely with VI in detecting FMDV in all milk components and generally coincided with, but did not consistently precede, the onset of clinical signs. rRT-PCR detected FMDV in milk up to 23 days post inoculation which was longer than VI. Furthermore, the detection limit of FMDV in milk was greater by rRT-PCR than VI and, in contrast to VI, rRT-PCR detected virus genome following heat treatment that simulated pasteurisation. rRT-PCR was also able to detect FMDV in preservative-treated milk. In conclusion, this study showed that automated rRT-PCR is quicker and more sensitive than VI and can be used to detect FMDV in whole milk as well as milk fractions from infected animals.

Pigs inoculated with the Alfort 187 isolate of classical swine fever (CSF) virus were used to study the immunological mechanisms associated with the humoral immune response in the disease. Quantitative and qualitative changes in the B-cell population (lambda light chain [C-lambda]-positive, immunoglobulins [Ig]-M-positive, and IgG-positive were demonstrated in the spleen, thymus and ileocaecal lymph node. Blood and serum samples were used to examine changes in leucocytes, albumin/globulin ratios and specific antibodies against CSF virus titration. Despite the lymphoid depletion shown by infected animals, an increase in B cells and potentially immunoglobulin-producing C-lambda(+) plasma cells was observed in the lymphoid organs from the onset of disease. The increase in C-lambda(+) B cells was matched by a parallel increase in IgM(+) cells, which attained peak values from 7 days post-inoculation (dpi), while IgG(+) cells increased from 11 dpi onwards. The enhanced biosynthetic capacity of these cells may have been linked to the initiation of a humoral response to CSF virus, and to the progressive decline in the albumin/globulin ratios of inoculated animals. Activation, proliferation and differentiation of B cells coincided with the presence of viral antigen, and with an intense phagocytic and biosynthetic activity of monocytes-macrophages and T lymphocytes. The previously reported increase of cytokine (TNF alpha IL-1 alpha and IL-6) production by monocytes-macrophages, and the release of IL-2, IL-4 and IFN gamma by T lymphocytes, may play a role in the initiation of the humoral immune response in CSE These changes may have influenced the late appearance of virus-specific antibodies in the study, as well as the progressive increase of immunoglobulins.