The molecular characterization of poliovirus strains by the RT-PCR-RFLP assay and its use in the active surveillance for acute flaccid paralysis cases in Romania between 2001-2006

Abstract

Poliomyelitis, an acute disease of the the central nervous system can be controlled through the use of inactivated virus vaccine (IPV) or live attenuated vaccine (OPV). The goal of the Global Polio Eradication Initiative is to stop the global transmission of poliovirus. Our study is a retrospective and prospective study that was made because in 2002 was isolated from one acute flaccid paralysis AFP case and eight healthy contacts belonging to the same small socio-cultural group having a low vaccine coverage living in Babadag a town in Romania a serotype 1 vaccine-derived poliovirus (VDPV), recombinant S1/S2/S1. The 67 poliovirus strains isolated in Romania between 2001-2006 from acute flaccid paralysis cases (AFP) (n=20, age = 3 months - 10 years), facial paralysis cases (FP) (n=5, age = 3 months - 3.7 years) and healthy's AFP contacts (n=42, age = 2 months - 5.10 years) were molecular investigated to confirm the vaccine origin of these strains and for the detection of recombinant strains. The identification of these strains was achieved through reverse transcription (RT), polymerase chain reaction (PCR) and restriction fragment length polymorphisme assays (RFLP) applied to two sequences of the viral genome, which are located in VP1-2A (2870 nt - 3648 nt) and 3D (6086nt - 6516 nt) regions. For the strains investigated in VP1-2A region, the RFLP profils after digestion with 3 restriction enzymes (Rsal, Ddel, Hinf l) were Sabin-like only with one exception, VDPV strain. In 3D region after digestion with one, two or three enzymes the genomes of most poliovirus (PV) strains were found to be similar to the original vaccine Sabin strain. In 9 AFP cases the profils detected after RFLP in 3D region were S3/S1 (n=3); S3/52 (n=2); S2/S1 (n=2), S2/S1 + S2 (n=2). In 3 FP cases the profils were S3/S2 (n=2), 53/S1 (n=1). In 11 healthy contacts the RFLP profiles in 3D region were S3/S2 (n=6); S1/S1 +S3 (n=1); S3/S1 + S2 (n=1), S3/S2 + S3 (n=1), S2 + S3/S1 (n=1), S1/S2 (n=1).