Prevalence and Genetic Diversity of Human Sapoviruses in Shellfish from Commercial Production Areas in Galicia, Spain

Abstract

The prevalence of human forms of Sapovirus, an emerging pathogen of human gastroenteritis, was investigated in an 18-month survey from class B mollusc-harvesting areas in two Galician rias (northwest Spain). The detection and quantification of Sapovirus was performed by reverse transcription-real-time PCR, according to the recently developed standard method ISO/TS 15216-1:2013, and genotyping by reverse transcription-nested PCR. The bivalve species studied were wild and cultured mussels (Mytilus galloprovincialis), clams (Venerupis philippinarum and Venerupis decussata), and cockles (Cerastoderma edule). Sapovirus was detected in 30 out of 168 samples (17.9%), with cockles being the species with the highest prevalence of positives (28.1%), followed by clams (22.6%), wild mussels (14.3%), and cultured mussels (12.9%). The estuary in the south of the region demonstrated a higher percentage of positive samples (21.8%) than the one in the north (14.4%). Viral contamination levels for the positive samples ranged between 1.9 ؋ 10 3 and 1.4 ؋ 10 5 RNA copies/g of digestive tissue. Thirteen Sapovirus sequences could be obtained based on partial capsid gene sequence and were classified into four genotypes: GI.1 (2 samples), GI.2 (8 samples), GIV.1 (2 samples), and GV.1 (1 sample). Sapovirus (SaV), within the family Caliciviridae, is an etiological agent of human gastroenteritis, primarily in children (1, 2). In 1977 its type strain, Sapporo virus (Hu/SV/GI/Sapporo/77/JP), was first detected during an outbreak in a Japanese orphanage (3). Since then, five genogroups have been identified based on complete capsid gene sequences (4, 5). Genogroups I, II, IV, and V comprise human strains, whereas genogroup III was detected only in swine strains. The SaV genome is a single-stranded positivesense RNA molecule of approximately 7.3 to 7.5 kb with two or three open reading frames (ORF), depending on the genogroup (4, 5). ORF1 encodes nonstructural proteins such as viral protein genome-linked (VPg), protease, RNA-dependent RNA polymerase (RdRp), and a capsid protein (VP1), while ORF2 and ORF3 encode proteins of unknown function (5, 6). The development of molecular techniques became crucial for the study of SaV, as it cannot be cultivated in the laboratory. Reverse transcription-PCR (RT-PCR) is the method used worldwide to detect SaV. It has a high sensitivity and can be used to analyze the virus genetically when it is coupled with the sequencing of amplicons (7-10). Reverse transcription-quantitative real-time PCR (RT-qPCR) also is widely used to detect the virus. It is highly sensitive and useful for determining quantitative results (11).SaV is transmitted via the fecal-oral route, either through person-to-person contact or from contaminated food and water (5, 12). SaV outbreaks are less common than those caused by human norovirus (NoV), the major cause of human gastroenteritis, being detected principally in Southeast Asia. However, SaV recently was included in the U.S. Environmental Protection Agency's Candidate List (CCL3) as an...

Low prevalence of Aichi virus in molluscan shellfish samples from Galicia (NW Spain)

“…; Varela et al . , ). Thus, 8 of 10 AiV positive samples from Ría de Area‐Betanzos showed in addition contamination with NoV GI (three samples) or GII (three samples).…”

“…Bivalve molluscan shellfish are often associated with viral food‐borne disease (Vossen ) as they obtain their food filtering small particles suspended in water, being able to concentrate and retain pathogens including enteric viruses (Varela et al . ).…”

Temporal variations in genotype distribution of human sapoviruses and Aichi virus 1 in wastewater in Southern Arizona, United States

“…Thereafter, data on the occurrence of these viruses in aquatic environments have been accumulated rapidly within the past decade owing to development and/or improvement of PCR methods for the detection of these viruses in environmental samples (Kitajima and Gerba 2015;Oka et al 2015). Recent environmental studies carried out in multiple geographical regions have collectively demonstrated a high prevalence of both sapoviruses and AiV-1 worldwide (Kitajima et al 2010a(Kitajima et al , 2011aSano et al 2011;Fioretti et al 2016;Varela et al 2016;Haramoto and Kitajima 2017), indicating that these viruses might be more prevalent in the environment than previously appreciated.…”

An overview of 20 years of studies on the prevalence of human enteric viruses in shellfish from Galicia, Spain

“…; Varela et al . ). A total of 78 samples including wild‐ and raft‐cultured Mediterranean mussels ( M. galloprovincialis ), Manila and carpet shell clams ( Ruditapes philippinarum and R. decussatus respectively) and cockles ( C. edule ) were collected from the same five harvesting areas from Moaña Bay in Ría de Vigo.…”

“…; Varela et al . , ). Further studies are needed to determine their real importance for the public health in our region as well as to elucidate the role of bivalve molluscs as transmission vehicles of these pathogens, as suggested for SaV in Japan by Iizuka et al .…”