African Swine Fever Diagnosis Adapted to Tropical Conditions by the Use of Dried-blood Filter Papers

Abstract

The performance of Whatman 3-MM filter papers for the collection, drying, shipment and long-term storage of blood at ambient temperature, and for the detection of African swine fever virus and antibodies was assessed. Conventional and real-time PCR, viral isolation and antibody detection by ELISA were performed on paired samples (blood/tissue versus dried-blood 3-MM filter papers) collected from experimentally infected pigs and from farm pigs in Madagascar and Côte d'Ivoire. 3-MM filter papers were used directly in the conventional and real-time PCR without previous extraction of nucleic acids. Tests that performed better with 3-MM filter papers were in descending order: virus isolation, real-time UPL PCR and conventional PCR. The analytical sensitivity of real-time UPL PCR on filter papers was similar to conventional testing (virus isolation or conventional PCR) on organs or blood. In addition, blood-dried filter papers were tested in ELISA for antibody detection and the observed sensitivity was very close to conventional detection on serum samples and gave comparable results. Filter papers were stored up to 9 months at 20-25°C and for 2 months at 37°C without significant loss of sensitivity for virus genome detection. All tests on 3-MM filter papers had 100% specificity compared to the gold standards. Whatman 3-MM filter papers have the advantage of being cheap and of preserving virus viability for future virus isolation and characterization. In this study, Whatman 3-MM filter papers proved to be a suitable support for the collection, storage and use of blood in remote areas of tropical countries without the need for a cold chain and thus provide new possibilities for antibody testing and virus isolation.

Clinical Validation of Two Recombinase-Based Isothermal Amplification Assays (RPA/RAA) for the Rapid Detection of African Swine Fever Virus

“…African swine fever belongs to a transboundary animal disease. Since it was first described in Kenya in 1921 (Montgomery, 1921), ASFV has been found in other 25 African countries (Randriamparany et al, 2016). In the 1950s, ASF incursions were subsequently reported in European countries, including Portugal, Spain, France, Italy, Belgium, the Netherlands, Malta and the virus further spread to the Caribbean and South America (Brazil, Cuba, Haiti, Dominican Republic) (Ekue et al, 1989).…”

Simplifying sampling for African swine fever surveillance: Assessment of antibody and pathogen detection from blood swabs

“…It is important to note that swabs can be directly dipped in AVL or ELISA buffer without cutting to run one assay, if cutting is inconvenient (see Petrov et al., ). FTA cards have also shown promising results (Braae et al., ; Michaud et al., ; Randriamparany et al., ; Uttenthal et al., ); however, a trained person must handle the samples as it is difficult to avoid contamination during close contact with bloody carcasses and additional transport devices are needed. In studies by Petrov et al.…”

“…Randriamparany et al. () also demonstrated the suitability of dried blood for antibody detection. No problems with specificity were encountered.…”

“…In later work by Randriamparany et al. () and Michaud et al. (), ASFV diagnosis and characterization were successfully performed with dried blood on filter paper, for both experimental and field samples and over extended periods of time.…”

How could an African swine fever outbreak evolve in an enzootic context? The case of Imerintsiatosika, Madagascar in 2015

“…When a breeder called for a butcher for a very sick or recently deceased pig, the field team came also and ask the consent of breeders, to take spleen. Conventional PCR analysis was done at the “Institut Pasteur de Madagascar” (IPM) following the protocol described by Randriamparany [38].…”