Abstract
Foot-and-mouth disease Virus (FMDV) is an economically important, highly contagious picornavirus that affects both wild and domesticated cloven hooved animals. In developing countries, the effective laboratory diagnosis of foot-and-mouth disease (FMD) is often hindered by inadequate sample preservation due to difficulties in the transportation and storage of clinical material. These factors can compromise the ability to detect and characterise FMD virus in countries where the disease is endemic. Furthermore, the high cost of sending infectious virus material and the biosecurity risk it presents emphasises the need for a thermo-stable, non-infectious mode of transporting diagnostic samples. This paper investigates the potential of using FMDV lateral-flow devices (LFDs) for dry transportation of clinical samples for subsequent nucleic acid amplification, sequencing and recovery of infectious virus by electroporation. FMDV positive samples (epithelial suspensions and cell culture isolates) representing four FMDV serotypes were applied to antigen LFDs: after which it was possible to recover viral RNA that could be detected using real-time RT-PCR. Using this nucleic acid, it was also possible to recover VP1 sequences and also successfully utilise protocols for amplification of complete FMD virus genomes. It was not possible to recover infectious FMDV directly from the LFDs, however following electroporation into BHK-21 cells and subsequent cell passage, infectious virus could be recovered. Therefore, these results support the use of the antigen LFD for the dry, non-hazardous transportation of samples from FMD endemic countries to international reference laboratories.
Highlights
Foot-and-mouth disease (FMD) virus (FMDV) [1] causes a highly infectious and contagious disease of wild and domesticated cloven-hoofed animals, with huge potential for rapid spread within susceptible animal populations
FMD viral genome was detected by 3D rRT-PCR on all sections of the lateral-flow devices (LFDs) washed with elution buffer one week and one month after visual development (Figure 3)
This study, demonstrates for the first time, that it is possible to recover full length Foot-and-mouth disease Virus (FMDV) RNA from positive LFDs which can be successfully used as a template for diagnostic rRT-PCR, full genome amplification, sequencing and recovery of infectious virus upon electroporation
Summary
Foot-and-mouth disease (FMD) virus (FMDV) (family Picornaviridae, genus Aphthovirus) [1] causes a highly infectious and contagious disease of wild and domesticated cloven-hoofed animals, with huge potential for rapid spread within susceptible animal populations. Many countries rely on the services of international laboratories such as those within the OIE/FAO FMD Laboratory Network to confirm and characterise the strain of virus collected in clinical material from suspected outbreaks. Maintenance of the cold chain is important since inadequately preserved samples are often degraded which generates difficulties for the reporting laboratory to fully characterise the field isolate, resulting in a loss of vital information required for control measures to be implemented. There is interest in developing and evaluating methods that can be used to preserve clinical samples at source in a form that is non-infectious and less subject to degradation during transit, but which can be fully recovered upon arrival at the reference laboratory
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