We investigated the feasibility of using Flinders Technology Associates (FTA®) filter cards for the storage of allantoic fluid containing an infectious bronchitis virus (IBV), such as Arkansas-DPI, Connecticut, and Massachusetts, and for their identification by reverse transcriptase (RT)-polymerase chain reaction (PCR) and characterization by restriction fragment length polymorphism (RFLP) or nucleotide sequencing. FTA® paper is a cotton-based cellulose membrane containing lyophilized chemicals that lyses many types of bacteria and viruses. IBV was inactivated upon contact with the FTA®, as shown by the inability of the virus to be propagated in embryonating chicken eggs. RT-PCR of the S1 gene showed that viral RNA in allantoic fluid remained stable after storage on FTA® filter cards and that the stability was time and temperature sensitive for the large (1700 base pair [bp]) but not the small (383 bp) PCR products. Analysis of the amplified products showed that molecular characterization is feasible in allantoic fluid stored on FTA® under nonfavorable environmental conditions (41 C) for at least 15 days. The use of FTA® cards for the collection, transport, and storage of IBV–containing samples is safe, inexpensive, and adequate for molecular diagnosis. We propose that specimens coming from overseas on FTA® cards would be first analyzed by RT-PCR with primers yielding a 1700-bp product followed by RFLP of the positive cases. Negative cases would be analyzed with primers yielding a 383-bp product (to exclude detrimental effect of the storage conditions) followed by nucleotide sequencing of the positive cases.
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Vol. 49 • No. 1