Foot-and-mouth disease virus (FMDV) remains an important pathogen of livestock more than 120 years after it was identified, with annual costs from production losses and vaccination estimated at €5.3–€17 billion (US$6.5–US$21 billion) in FMDV-endemic areas. Control and eradication are difficult because FMDV is highly contagious, genetically and antigenically diverse, infectious for a wide variety of species, able to establish subclinical carriers in ruminants, and widely geographically distributed. For early detection, sustained control, or eradication, sensitive and specific FMDV surveillance procedures compatible with high through-put testing platforms are required. At present, surveillance relies on the detection of FMDV-specific antibody or virus, most commonly in individual animal serum, vesicular fluid, or epithelial specimens. However, FMDV or antibody are also detectable in other body secretions and specimens, e.g., buccal and nasal secretions, respiratory exhalations (aerosols), mammary secretions, urine, feces, and environmental samples. These alternative specimens offer non-invasive diagnostic alternatives to individual animal sampling and the potential for more efficient, responsive, and cost-effective surveillance. Herein we review FMDV testing methods for contemporary and alternative diagnostic specimens and their application to FMDV surveillance in livestock (cattle, swine, sheep, and goats).

In this study, two sheep, eight dromedary camels and two Bactrian camels were inoculated with foot-and-mouth disease virus (FMDV) type A SAU 22/92. Five naive dromedary camels and four sheep were kept in direct or indirect contact with the inoculated camels. The inoculated sheep, which served as positive controls, displayed typical moderate clinical signs of FMD and developed viraemia and high antibody titres. The presence of the virus was also detected in probang and mouth-swab samples for several days after inoculation. In contrast, the inoculated dromedary camels were not susceptible to FMDV type A infection. None of them showed clinical signs of FMD or developed viraemia or specific anti-FMDV antibodies despite the high dose of virus inoculated. All the contact sheep and contact dromedaries that were kept together with the inoculated camels remained virus-negative and did not seroconvert when tested up to 28 days post-inoculation (p.i.). In comparison with the non-susceptible dromedaries, the two inoculated Bactrian camels showed moderate to severe clinical signs of FMD; however, the clinical signs of FMD appeared rather late, between 8 and 14 days p.i., compared to the inoculated sheep. Characteristic FMD lesions in the Bactrian camels, accompanied with severe lameness, were only observed on the hind feet. The presence of the virus in the serum samples of both Bactrian camels was detected by real-time RT–PCR in one of the animals on days 3 and 7 p.i. and in the second animal from days 1 to 3 p.i. and subsequently again on day 21 p.i. The Bactrian camels developed high titres of antibodies to the inoculated FMDV which appeared at 7–10 days p.i. and lasted up to 130 days p.i. Only low and transient amounts of FMDV were detected in the mouth-swab and probang samples collected from both Bactrian camels.

Internal initiation of translation is promoted by internal ribosome entry site (IRES) cis-acting elements. Using transcripts that correspond to the structural domains of the foot-and-mouth disease virus (FMDV) IRES, we have identified RNA–RNA interactions between separated domains (1–2, 3, 4–5, or HH) of the IRES structure. All the assayed domains were able to interact with the full-length IRES as well as with domain 3, although to a different extent, with the most efficient interactions being those occurring between domains 3 and 4–5, and domains 3 and 1–2. RNA–RNA complexes were stable over 1 h of incubation at 37 °C, and depended on Mg2+ and RNA concentration. Neither the antisense domain 1–2 nor tRNA interacted with domain 3, providing experimental evidence of the specificity for the sense strand of the IRES sequence. Additionally, domain 1–2 did not interact with 4–5, leading to the suggestion that domain 3 acts as a scaffold structure where the other domains bind. The thermal disassociation profile of these complexes indicated different strength in these interactions. Whereas 50% of the complexes between domains 3 and 4–5 were destabilized at 45 °C, those formed by domain 1–2 and 3 required temperatures higher than 51 °C. Efficient self-dimerization of domains 3 and 4–5 was found in the absence of other transcripts. Formation of domain 3 homodimer competed with formation of heterocomplexes with other domains, and conversely, domain 3 homodimers were competed out by the presence of the other domains. RNA interactions were also observed at physiological concentrations of Mg2+ and K1+. The identification of the RNA–RNA complexes reported here provide direct experimental evidence of tertiary interactions within IRES elements.