Long-term storage at tropical temperature of dried-blood filter papers for detection and genotyping of RNA and DNA viruses by direct PCR.
In tropical countries the diagnosis of viral infections of humans or animals is often hampered by the lack of suitable clinical material and the necessity to maintain a cold chain for sample preservation up to the laboratory. This study describes the use of filter papers for rapid sample collection, and the molecular detection and genotyping of viruses when stored over long periods at elevated temperatures. Infected blood was collected on filter papers, dried and stored at different temperatures (22, 32 and 37 degrees C) for various periods (up to 9 months). Two animal viruses, African swine fever, a large double-stranded DNA virus and Peste des Petits Ruminants, a negative single-stranded RNA virus, were used to validate the method. Filter papers with dried blood containing virus or control plasmid DNA were cut in small 5mm(2) pieces and added directly to the PCR tube for conventional PCR. Nucleic acid from both viruses could still be detected after 3 months at 32 degrees C. Moreover, the DNA virus could be detected at least 9 months after conservation at 37 degrees C. PCR products obtained from the filter papers were sequenced and phylogenetic analysis carried out. The results were consistent with published sequences, demonstrating that this method can be used for virus genotyping.
Michaud
V
V
CIRAD, UR Contrôle des Maladies, Montpellier F-34398, France.
Gil
P
P
Kwiatek
O
O
Prome
S
S
Dixon
L
L
Romero
L
L
Le Potier
M-F
MF
Arias
M
M
Couacy-Hymann
E
E
Roger
F
F
Libeau
G
G
Albina
E
E
D000349:Africa; D000357:African Swine Fever / Q000821:virology; D000358:African Swine Fever Virus / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D000818:Animals; D001800:Blood Specimen Collection / Q000379:methods*; D005838:Genotype; D006358:Hot Temperature; D029021:Peste-des-Petits-Ruminants / Q000821:virology; D018107:Peste-des-petits-ruminants virus / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D010802:Phylogeny; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013997:Time Factors; D014329:Tropical Climate