Comparison of Biosafety and Diagnostic Utility of Biosample Collection Cards

Abstract

Six different biosample collection cards, often collectively referred to as FTA (Flinders Technology Associates) cards, were compared for their ability to inactivate viruses and stabilize viral nucleic acid for molecular testing. The cards were tested with bluetongue virus, foot-and-mouth disease virus (FMDV), small ruminant morbillivirus (peste des petits ruminants virus), and lumpy skin disease virus (LSDV), encompassing non-enveloped and enveloped representatives of viruses with double-stranded and single-stranded RNA genomes, as well as an enveloped DNA virus. The cards were loaded with virus-containing cell culture supernatant and tested after one day, one week, and one month. The inactivation of the RNA viruses was successful for the majority of the cards and filters. Most of them completely inactivated the viruses within one day or one week at the latest, but the inactivation of LSDV presented a greater challenge. Three of the six cards inactivated LSDV within one day, but the others did not achieve this even after an incubation period of 30 days. Differences between the cards were also evident in the stabilization of nucleic acid. The amount of detectable viral genome on the cards remained approximately constant for all viruses and cards over an incubation period of one month. With some cards, however, a bigger loss of detectable nucleic acid compared with a directly extracted sample was observed. Using FMDV, it was confirmed that the material applied to the cards was sufficiently conserved to allow detailed molecular characterization by sequencing. Furthermore, it was possible to successfully recover infectious FMDV by chemical transfection from some cards, confirming the preservation of full-length RNAs.

Keywords: Ahlstrom-Munksjö; Copan; FTA (Flinders Technology Associates) cards; Lipofectamine 3000; Macherey-Nagel; Whatman; biosample collection cards; real-time RT-qPCR; sequencing; transfection; virus isolation.

Figure 1

Virus inactivation on cards and filters during one day, one week, and one month of incubation. The percentage of all replicates still causing CPE in cell culture is shown for each virus, card, or filter and time. (a) BTV; (b) FMDV; (c) PPRV; (d) LSDV.

Figure 2

Stabilization and recovery of nucleic acid from cards and filters analyzed by real-time (RT)-PCR after one day, one week and one month of incubation, showing the mean ∆C_q values for each virus, sample time point, and card with their 95% confidence intervals indicated by the error bars. ∆C_q values were calculated between the eluted nucleic acid and the virus preparation used for spotting (positive control). (a) BTV; (b) FMDV; (c) PPRV; (d) LSDV.