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Noroviruses (NVs) cause human gastroenteritis through person-to-person transmission and via contaminated foods. In food poisoning, a major suspected cause is the consumption of raw oysters. We detected NVs from environmental water and oysters around a closed gulf where oysters are cultivated. We collected oyster and water samples once or twice a month for 30 months from October 2001 to March 2004. We then studied monthly changes in virus occurrence and in genetic relationships among 208 NVs isolated from water and oyster samples and from the feces of children suffering from acute gastroenteritis during the same period in the same region. In the analysis of untreated water flowing into farm sewage, NVs were detected year round. In other water samples -processed sewage, river water, and seawater-, oysters, and children's feces, NVs were detected mainly in winter. A comparison of NV nucleotide sequences showed genetic diversity, but some strains predominated in certain winter seasons. These predominant strains were detected across sample materials. In 2002/03, an identical strain was detected in sewage, river water, seawater, oysters, and feces. We also found that NV genetic types changed at the beginning of the season, in November or December, in both 2001/02 and 2002/03. This study showed a clear relationship between NVs detected in children's feces and those in environmental water and oysters. These results support the idea that NVs are transmitted from the feces of infected persons to oysters by the flow of water through farm sewage, rivers, and the sea, finally accumulating in the mid-gut gland of oysters.

Outbreaks of gastroenteritis caused by Norwalk-like viruses are often induced by the consumption of raw shellfish such as oysters. Incidences reach a peak during the cold season in Japan, when seawater temperatures fall below 10 degrees C. We investigated oysters' uptake and excretion of viruses, over varying lengths of exposure, monitoring the effects of changes in temperature and flow rate of seawater, and the presence of plankton. The study was performed using a poliovirus and an experimental circulatory system, which was framed on the same principle as a model practically used for the depuration of oysters. Polioviruses present in the seawater were taken rapidly into the midgut gland of oysters. However, virus levels detected in oysters at both 10 degrees C and 20 degrees C were decreased to approximately 1/1,000 to 1/10,000 within 6 hrs after the circulatory seawater was replaced by UV irradiated seawater. These results demonstrate the effectiveness of the circulatory depuration system for the elimination of poliovirus from oysters, and indicate that controlling the temperature and flow rate of the circulatory system could decrease the risk of NLV infection.

To clarify the environmental distribution of Vibrio vulnificus, sea water, sea mud, and oysters were examined at 13 sites, i.e. 4 sites in the Tokyo Bay (eastern Japan) and 9 sites (5 sites for oysters) in Tokushima Prefecture (western Japan). 1. V. vulnificus was isolated from 80 (54.8%) of the 146 samples of sea water examined. It was isolated from 19 (41.3%) of the 46 samples from western Japan and 61 (61.0%) of the 100 samples from eastern Japan. 2. It was isolated from 40 (40.8%) of the 98 samples of sea mud obtained in eastern Japan. 3. It was isolated from 655 (30.3%) of the 2,165 samples of oysters. They were 30 (9.7%) of 309 samples from western Japan and 625 (33.7%) of 1,856 samples from eastern Japan. 4. The density of V. vulnificus was 0.3-1.1 x 10(6) MPN/L in seawater, 0.3-1.1 x 10(5) MPN/100 g in sea mud, and 0.3-1.1 x 10(7) MPN/100 g in oysters. 5. Seasonally, V. vulnificus was isolated from 44 (6.2%) of the 713 samples in spring, 450 (72.6%) of the 620 samples in summer, 264 (51.8%) of the 510 samples in fall, and 17 (3.0%) of the 56 samples in winter. Thus, the isolation rates of V. vulnificus from sea water and oysters tended to be higher in eastern Japan than in western Japan and to be highest in summer, then, in fall.

Norwalk viruses(NVs) are well known pathogens that can cause acute nonbacterial epidemic gastroenteritis in adults as well as acute gastroenteritis in children. In food poisoning cases due to raw oysters, NVs is considered as an important pathogen and the molecular epidemiological studies of NVs are the most suitable method to trace these routes of infections in conjunction with oyster supply mechanism. Furthermore, the prevalence of genotypes of NVs which are detected from cases of acute gastroenteritis in children may provide us with useful information for the epidemiological study of acute nonbacterial epidemic gastroenteritis, because investigations of acute nonbacterial epidemic gastroenteritis might be revealed to be only a piece of infiltration of NVs in nature.

The application of recently developed molecular technique revealed the important role of the Norwalk virus(NV). The major role of the NV as agents of food and water-related outbreaks of gastroenteritis has now been recognized in the world. Moreover, a part of NV is also documented as important agents of gastroenteritis in childhood and infancy. A high rate of food-related viral gastroenteritis outbreaks was observed following ingestion of shellfish in Japan. Specific methods are not available for the prevention or control of NV gastroenteritis. Hand washing will be the most effective to decrease transmission within a family and a hospital. To control food-borne diseases a different strategy is necessary with virus and bacteria.

The Norwalk virus(NV) is widely known as a cause of nonbacterial food poisoning, infant diarrhea, and acute gastroenteritis in the winter months between November and March. While it is strongly suspected that NV that is excreted by humans flows into coastal seawaters via rivers and wastewater treatment facilities to contaminate oysters that are grown in farms in the area, light has yet to be shed on the behavior of this virus in the natural environment. We therefore conducted a polymerase chain reaction (PCR) survey of NV levels in the aquatic environment of the oyster bed area of the Shima region in Mie Prefecture, whereupon the NV was detected in marine sediment, oysters, and mule clams even during the summer months, when food poisoning is infrequent. In order to assess their similarity to human-derived strains, the detected viruses and their human-derived counterparts were subjected to genetic analysis, whereupon some of the detected viruses were found to be remarkably similar to those that were previously detected in humans infected with NV. In the interests of examining methods for decontaminating NV-contaminated oysters, we also conducted an assessment on a system of virus decontamination that focuses on seawater temperature and oyster metabolism, using Poliovirus Sabin strain. The decontamination system mentioned above was a closed loop, water circulating system, built on the same principles as those actually in use at oyster farms. Our experiment indicated that at seawater temperatures of both 10 degrees C and 20 degrees C, virus placed into the water tank was rapidly incorporated into the midgut glands of the oysters. Thereafter, when seawater irradiated with UV was circulated, the virus count in the oysters fell from 1/1,000 to 1/10,000 within 6 hours. These results indicated the utility of this system for virus decontamination, suggesting the possibility of significantly alleviating the risk of NV infection in humans by using this system to maintain the seawater temperature within the decontamination tank above a certain temperature, and to perform decontamination with an adequate water flow.

Through a year from April, 1999 to March, 2000, 20 samples, which consisted of raw sewage (2), chlorine-treated sewage (2), seawater (10) and naturally grown oysters (6), were collected monthly both from the sewage works at Mihama-ku, Chiba City and at a yacht harbor in Chiba City Bay, Japan. Astrovirus RNA were detected by reverse transcription-polymerase chain reaction (RT-PCR) and was typed by direct sequencing. Astrovirus positive products were detected from 9 samples (raw sewage; 1/2, chlorine-treated sewage; 2/2, seawater; 5/10 and oysters; 1/6) collected in April, 1999. In May, positive products were detected from 4 samples (raw sewage; 2/2 and seawater; 2/10). In June, only 1 positive product was detected from raw sewage. The number of positive samples showed a tendency to decrease and no positive products were detected from samples collected in July, 1999 to January, 2000. After that period, positive products were again detected from 3 samples (raw sewage; 1/2, chlorine-treated sewage; 2/2) collected in February, 2000. In March, the number of positive samples showed the peak and positive products were detected from 12 samples (raw sewage; 2/2, chlorine-treated sewage; 2/2, seawater; 7/10 and oysters: 1/6). Astrovirus positive products detected in April, May, June, July, 1999 and February, 2000 were classified into type 1 or 2 by sequencing, whereas in March, 2000 were type 1, 2, 3, 6 and 7.

SRSV, astrovirus in tvirus and HAV genomes were detected by RT-PCR in naturally grown oysters (total 112) collected in Chiba City bay, Japan, through a year from April, 1997 to March, 1998 and genogroup was typed by sequencing. SRSV positive products were detected by RT-PCR from 28 out of 112 oysters and all of them were grouped into G-II by sequencing. The highest incidence was observed in February, 1998. Furthermore, astrovirus positive products were detected sporadically from 15 out of 112 samples and the highest incidence was observed in January, 1998. On the other hand, HAV was detected from only 2 out of 112 and no adenovirus positive product was detected. The results indicated that both SRSV and astrovirus were predominantly distributed into naturally grown oysters in the winter season. SRSV and astrovirus seem to contribute mainly to the food-born outbreaks of gastroenteritis occurred by eating contaminated oysters as causative agents.