ABSTRACT
Sapovirus particles are typically 41-46 nm in diameter and have a cup-shaped depression and/or 10 spikes on the outline. The sapovirus genome is a single-stranded, positive sense RNA molecule of approximately 7.5 kb that is polyadenylated at the 3′ end. Sapovirus can be divided into ve genogroups (GI-GV; Figure 11.1), among which GI, GII, GIV, and GV are known to infect humans, whereas sapovirus GIII infects porcine species. The sapovirus GI, GIV, and GV genomes are each predicted to contain three main open reading frames (ORFs), whereas sapovirus GII and GIII have two ORFs. Sapovirus ORF1 encodes for nonstructural proteins, including the VPg, protease and RNA dependent RNA polymerase (RdRp), and a major capsid protein (VP1). Human sapovirus ORF2 and ORF3 encode proteins of yet unknown functions. Previously, sapoviruses have been classied based on either their partial RdRp and/or capsid sequences, but since the discovery of recombinant sapovirus strains [16,17] classication numbering schemes have conicted between the different research groups. Ideally,
11.1 Introduction .....................................................................................................................................................................119 11.1.1 The Virus ...........................................................................................................................................................119 11.1.2 Morphology and Biology ...................................................................................................................................119 11.1.3 Diagnosis ........................................................................................................................................................... 120
11.2 Methods .......................................................................................................................................................................... 121 11.2.1 Sample Preparation ........................................................................................................................................... 121
11.2.1.1 Sample Handling .............................................................................................................................. 121 11.2.1.2 Phenotypic Characterization ............................................................................................................ 122 11.2.1.3 RNA Extraction ................................................................................................................................ 123
11.2.2 Detection Procedures ........................................................................................................................................ 123 11.2.2.1 Reverse Transcription ....................................................................................................................... 123 11.2.2.2 Nested PCR ...................................................................................................................................... 124 11.2.2.3 Real-Time RT-PCR ........................................................................................................................... 124
11.3 Conclusion ...................................................................................................................................................................... 125 References ................................................................................................................................................................................. 126
both RdRp and capsid genes should be analyzed. However, this is not always practical, due to amplication difculties, time, and cost of reagents. These days many phylogenetic studies are using the 5′ terminus of the capsid gene for classication.
