Accumulation of sediment-associated viruses in shellfish

The present study focused on the importance of contaminated sediments in shellfish accumulation of human viruses. Epifaunal (Crassostrea virginica) and infaunal (Mercenaria mercenaria) shellfish, placed on or in cores, were exposed to either resuspended or undisturbed sediments containing bound poliovirus type 1 (LSc 2ab). Consistent bioaccumulation by oysters (four of five trials) was only noted when sediment-bound viruses occurred in the water column. Virus accumulation was observed in a single instance where sediments remained in an undisturbed state. While the incidence of bioaccumulation was higher with resuspended rather than undisturbed contaminated sediment, the actual concentration of accumulated viruses was not significantly different. The accumulation of viruses from oysters residing on uninoculated sediments. When clams were exposed to undisturbed, virus-contaminated sediments, two of five shellfish pools yielded viral isolates. Bioaccumulation of undisturbed sediments by these bivalves was considered marginal when related to the concentration of virus in contaminated sediments; they would only represent a significant threat when suspended in the water column. Arguments were advanced for water-column sampling in the region of the water-sediment interface to provide an accurate determination of the virological quality of shellfish harvesting waters.     

Inefficient accumulation of low levels of monodispersed and feces-associated poliovirus in oysters.

The accumulation of low levels (0.002 to 0.18 PFU/ml) of both feces-associated and monodispersed poliovirus by oysters (Crassostrea virginica or C. gigas) and clams (Mercenaria mercenaria) was investigated. These levels were chosen to duplicate the conditions present in light to moderately polluted waters. Experiments were performed in both small- and large-scale flowing seawater systems, developed to mimic the natural marine habitats of shellfish. Under these experimental conditions, viral accumulation by physiologically active shellfish was only noted when water column concentrations exceeded approximately 0.01 PFU/ml. Bioaccumulation increased with increasing concentrations of both monodispersed and feces-associated viruses. At virus concentrations below this level, viruses were seldom detected in either clams or oysters. Evidence indicated that the lack of accumulation was not the result of inefficient extraction or detection methods. The modified Cat-Floc-beef extract procedure used in the experiment was found to be capable of detecting as few as 1.5 to 2.0 PFU per shellfish. Evidence is presented to indicate that an uptake-depuration equilibrium was present at virus exposure levels of 0.10 PFU/ml, but not at 0.01 PFU/ml. The results suggested that viral accumulation by shellfish may not be efficient at water column concentrations below congruent to 0.01 PFU/ml.     

Bioaccumulation Efficiency,Tissue Distribution,and Environmental Occurrence of Hepatitis E Virus in Bivalve Shellfish from France

Hepatitis E virus (HEV), an enteric pathogen of both humans and animals, is excreted by infected individuals and is therefore present in wastewaters and coastal waters. As bivalve molluscan shellfish are known to concentrate viral particles during the process of filter feeding, they may accumulate this virus. The bioaccumulation efficiencies of oysters (Crassostrea gigas), flat oysters (Ostrea edulis), mussels (Mytilus edulis), and clams (Ruditapes philippinarum) were compared at different time points during the year. Tissue distribution analysis showed that most of the viruses were concentrated in the digestive tissues of the four species. Mussels and clams were found to be more sensitive to sporadic contamination events, as demonstrated by rapid bioaccumulation in less than 1 h compared to species of oysters. For oysters, concentrations increased during the 24-h bioaccumulation period. Additionally, to evaluate environmental occurrence of HEV in shellfish, an environmental investigation was undertaken at sites potentially impacted by pigs, wild boars, and human waste. Of the 286 samples collected, none were contaminated with hepatitis E virus, despite evidence that this virus is circulating in some French areas. It is possible that the number of hepatitis E viral particles discharged into the environment is too low to detect or that the virus may have a very short period of persistence in pig manure and human waste.     

Human enteroviruses in oysters and their overlying waters.

The presence of enteroviruses in oysters and oyster-harvesting waters of the Texas Gulf coast was monitored over a period of 10 months. Viruses were detected in water and oyster samples obtained from areas both open and closed to shellfish harvesting. Viruses were detected periodically in waters that met current bacteriological standards for shellfish harvesting. No significant statistical relationship was demonstrated between virus concentration in oysters and the bacteriological and physiochemical quality of water and shellfish. Viruses in water were, however, moderately correlated with total coliforms in water and oysters and with fecal coliforms in oysters. Total coliforms in water were realted to total coliforms in sediment were related only to total coliforms in sediment. Among the physiochemical characteristics of water, turbidity was related statistically to the organic matter content of water and to fecal coliforms in water. There was a marked effect of rainfall on the bacteriological quality of water. Of a total of 44 water samples, 26 yielded virus in concentrations from 4 to 167 plaque-forming units per 100-gallon (ca. 378.5-liter) sample. Of a total of 40 pools of 10 to 12 oysters each, virus was found in 14 pools at a concentration of 6 to 224 plaque-forming units per 100 g of oyster meat. On five occasions, virus was found in water samples when no virus could be detected in oysters harvested from the same sites. This study indicates that current bacteriological standards for determining the safety of shellfish and shellfish-growing waters do no reflect the occurrence of enteroviruses.     

Human enteroviruses in oysters and their overlying waters.

The presence of enteroviruses in oysters and oyster-harvesting waters of the Texas Gulf coast was monitored over a period of 10 months. Viruses were detected in water and oyster samples obtained from areas both open and closed to shellfish harvesting. Viruses were detected periodically in waters that met current bacteriological standards for shellfish harvesting. No significant statistical relationship was demonstrated between virus concentration in oysters and the bacteriological and physiochemical quality of water and shellfish. Viruses in water were, however, moderately correlated with total coliforms in water and oysters and with fecal coliforms in oysters. Total coliforms in water were realted to total coliforms in sediment were related only to total coliforms in sediment. Among the physiochemical characteristics of water, turbidity was related statistically to the organic matter content of water and to fecal coliforms in water. There was a marked effect of rainfall on the bacteriological quality of water. Of a total of 44 water samples, 26 yielded virus in concentrations from 4 to 167 plaque-forming units per 100-gallon (ca. 378.5-liter) sample. Of a total of 40 pools of 10 to 12 oysters each, virus was found in 14 pools at a concentration of 6 to 224 plaque-forming units per 100 g of oyster meat. On five occasions, virus was found in water samples when no virus could be detected in oysters harvested from the same sites. This study indicates that current bacteriological standards for determining the safety of shellfish and shellfish-growing waters do no reflect the occurrence of enteroviruses.     

A persistent, productive, and seasonally dynamic vibriophage population within Pacific oysters (Crassostrea gigas)

In an effort to understand the relationship between Vibrio and vibriophage populations, abundances of Vibrio spp. and viruses infecting Vibrio parahaemolyticus (VpVs) were monitored for a year in Pacific oysters and water collected from Ladysmith Harbor, British Columbia, Canada. Bacterial abundances were highly seasonal, whereas high titers of VpVs (0.5 x 10(4) to 11 x 10(4) viruses cm(-3)) occurred year round in oysters, even when V. parahaemolyticus was undetectable (< 3 cells cm(-3)). Viruses were not detected (<10 ml(-1)) in the water column. Host-range studies demonstrated that 13 VpV strains could infect 62% of the V. parahaemolyticus strains from oysters (91 pairings) and 74% of the strains from sediments (65 pairings) but only 30% of the water-column strains (91 pairings). Ten viruses also infected more than one species among V. alginolyticus, V. natriegens, and V. vulnificus. As winter approached and potential hosts disappeared, the proportion of host strains that the viruses could infect decreased by approximately 50% and, in the middle of winter, only 14% of the VpV community could be plated on summer host strains. Estimates of virus-induced mortality on V. parahaemolyticus indicated that other host species were required to sustain viral production during winter when the putative host species was undetectable. The present study shows that oysters are likely one of the major sources of viruses infecting V. parahaemolyticus in oysters and in the water column. Furthermore, seasonal shifts in patterns of host range provide strong evidence that the composition of the virus community changes during winter.     

Concentration of Escherichia coli in sediments as an indicator of the sanitary status of oyster (Crassostrea virginica) aquaculture sites

The objective of the study was to determine whether there are differences in Escherichia coli counts in relation to seasonal and/or spatial distribution patterns in a conditional shellfish‐growing zone located in the Richibucto estuary, New Brunswick, Canada. E. coli concentrations in surface water, sediments, suspended and bottom cultured American oysters (Crassostrea virginica) were determined. Contamination levels in water were significantly lower than in sediments, bottom and suspended cultured oysters (P < 0.01; P < 0.05, P < 0.01, n = 303, respectively). No significant difference in fecal contamination was observed between suspended and bottom cultured oysters (P > 0.05, n = 303). Seasonal variations (temporal) had a significant influence on fecal coliform contamination of all components (P < 0.01, n = 303, linear model). E. coli concentration levels were as much as ten times higher in sediments than in water. Furthermore, results suggest that E. coli concentration levels in sediments are a more reliable indicator of the sanitary status of a grow‐out operation than E. coli levels in water. Considering only sampling dates when contamination levels in oysters exceeded the established threshold of 230 MPN, less than 50% of the cases were predicted by water contamination while as many as 89.9% of the samples were predicted by sediment contamination using the proposed threshold of 75 MPN per 100 g. This paper emphasizes the importance of acknowledging the significant role sediments play in the dynamics of contamination by E. coli in areas that are currently exploited or are being considered for shellfish aquaculture. The presence of E. coli in sediments should not be overlooked as an integral factor in assessing the environmental sanitary status.     

A Persistent, Productive, and Seasonally Dynamic Vibriophage Population within Pacific Oysters (Crassostrea gigas)

In an effort to understand the relationship between Vibrio and vibriophage populations, abundances of Vibrio spp. and viruses infecting Vibrio parahaemolyticus (VpVs) were monitored for a year in Pacific oysters and water collected from Ladysmith Harbor, British Columbia, Canada. Bacterial abundances were highly seasonal, whereas high titers of VpVs (0.5 × 10⁴ to 11 × 10⁴ viruses cm⁻³) occurred year round in oysters, even when V. parahaemolyticus was undetectable (<3 cells cm⁻³). Viruses were not detected (<10 ml⁻¹) in the water column. Host-range studies demonstrated that 13 VpV strains could infect 62% of the V. parahaemolyticus strains from oysters (91 pairings) and 74% of the strains from sediments (65 pairings) but only 30% of the water-column strains (91 pairings). Ten viruses also infected more than one species among V. alginolyticus, V. natriegens, and V. vulnificus. As winter approached and potential hosts disappeared, the proportion of host strains that the viruses could infect decreased by ~50% and, in the middle of winter, only 14% of the VpV community could be plated on summer host strains. Estimates of virus-induced mortality on V. parahaemolyticus indicated that other host species were required to sustain viral production during winter when the putative host species was undetectable. The present study shows that oysters are likely one of the major sources of viruses infecting V. parahaemolyticus in oysters and in the water column. Furthermore, seasonal shifts in patterns of host range provide strong evidence that the composition of the virus community changes during winter.     

Effects of sediment resuspension on phytoplankton production: teasing apart the influences of light, nutrients and algal entrainment

1. Wind‐induced sediment resuspension can affect planktonic primary productivity by influencing light penetration and nutrient availability, and by contributing meroplankton (algae resuspended from the lake bed) to the water column. We established relationships between sediment resuspension, light and nutrient availability to phytoplankton in a shallow lake on four occasions. 2. The effects of additions of surficial sediments and nutrients on the productivity of phytoplankton communities were measured in 300 mL gas‐tight bottles attached to rotating plankton wheels and exposed to a light gradient, in 24 h incubations at in situ temperatures. 3. While sediment resuspension always increased primary productivity, resuspension released phytoplankton from nutrient limitation in only two of the four experiments because the amount of available nitrogen and phosphorus entrained from the sediments was small compared with typical baseline levels in the water column. In contrast, chlorophyll a entrainment was substantial compared with baseline water column concentrations and the contribution of meroplankton to primary production was important at times, especially when seasonal irradiance in the lake was high. 4. Comparison of the in situ light climate with the threshold of light‐limitation of the phytoplankton indicated that phytoplankton in the lake were only likely to be light‐limited at times of extreme turbidity (e.g. >200 nephelometric turbidity units), particularly when these occur in winter. Therefore, resuspension influenced phytoplankton production mainly via effects on available nutrients and by entraining algae. The importance of each of these varied in time. 5. The partitioning of primary productivity between the water column and sediments in shallow lakes greatly influences the outcome of resuspension events for water column primary productivity.     

Virus-contaminated oysters: a three-month monitoring of oysters imported to Switzerland

Molluscan shellfish are known to be carriers of viral and bacterial pathogens. The consumption of raw oysters has been repeatedly linked to outbreaks of viral gastroenteritis and hepatitis A. Switzerland imports over 300 tons of oysters per year, 95% of which originate in France. To assess the level of viral contamination, a 3-month monitoring study was conducted. Therefore, the sensitivities of several previously described methods for virus concentration were compared, and one protocol was finally chosen by using dissected digestive tissues. Eighty-seven samples consisting of five oysters each were analyzed for Norwalk-like viruses (NLVs), enteroviruses, and hepatitis A viruses from November 2001 to February 2002. The oysters were exported by 31 French, three Dutch, and two Irish suppliers. Eight oyster samples from six French suppliers were positive for NLVs, and four samples from four French suppliers were positive for enteroviruses; two of the latter samples were positive for both viral agents. No hepatitis A viruses were detected. The sequences of NLV and enterovirus amplicons showed a great variety of strains, especially for the NLVs (strains similar to Bristol, Hawaii, Mexico, and Melksham agent). The data obtained indicated that imported oysters might be a source of NLV infection in Switzerland. However, further studies are needed to determine the quantitative significance of the risk factor within the overall epidemiology of NLVs.     

贝类中人源诺如病毒污染净化技术研究进展

人源诺如病毒是全球引起人急性胃肠炎的食源性病原体之一。牡蛎、贻贝等贝类消化腺组织中含有诺如病毒受体类似物,可从污染水体中富集高浓度病毒,因此,生食或食用加工不当的受污染贝类极易造成诺如病毒感染。污染贝类的净化处理技术已成为诺如病毒防控领域中的研究热点,例如消杀试剂、臭氧处理工艺、新型非热消杀技术以及最近报道的具有抗病毒作用的益生菌等。诺如病毒活性检测对确定贝类中的病毒污染水平和评价消杀技术效果有重要作用,只有完整和具有感染力的病毒才会对人体健康造成威胁。因此,本文在前期工作基础上,进一步对诺如病毒活性鉴定方法、贝类产品消杀技术以及贝类养殖净化工艺等研究进展进行综述,以期为完善食源性病毒防控技术提供参考。     

Virus-Contaminated Oysters: a Three-Month Monitoring of Oysters Imported to Switzerland

Molluscan shellfish are known to be carriers of viral and bacterial pathogens. The consumption of raw oysters has been repeatedly linked to outbreaks of viral gastroenteritis and hepatitis A. Switzerland imports over 300 tons of oysters per year, 95% of which originate in France. To assess the level of viral contamination, a 3-month monitoring study was conducted. Therefore, the sensitivities of several previously described methods for virus concentration were compared, and one protocol was finally chosen by using dissected digestive tissues. Eighty-seven samples consisting of five oysters each were analyzed for Norwalk-like viruses (NLVs), enteroviruses, and hepatitis A viruses from November 2001 to February 2002. The oysters were exported by 31 French, three Dutch, and two Irish suppliers. Eight oyster samples from six French suppliers were positive for NLVs, and four samples from four French suppliers were positive for enteroviruses; two of the latter samples were positive for both viral agents. No hepatitis A viruses were detected. The sequences of NLV and enterovirus amplicons showed a great variety of strains, especially for the NLVs (strains similar to Bristol, Hawaii, Mexico, and Melksham agent). The data obtained indicated that imported oysters might be a source of NLV infection in Switzerland. However, further studies are needed to determine the quantitative significance of the risk factor within the overall epidemiology of NLVs.     

The depuration dynamics of oysters (Crassostrea gigas) artificially contaminated with hepatitis A virus and human adenovirus

Within the country of Brazil, Santa Catarina is a major shellfish producer. Detection of viral contamination is an important step to ensure production quality and consumer safety during this process. In this study, we used a depuration system and ultraviolet (UV) disinfection to eliminate viral pathogens from artificially infected oysters and analysed the results. Specifically, the oysters were contaminated with hepatitis A virus (HAV) or human adenovirus type 5 (HAdV5). After viral infection, the oysters were placed into a depuration tank and harvested after 48, 72 and 96 h. After sampling, various oyster tissues were dissected and homogenised and the viruses were eluted with alkaline conditions and precipitated with polyethylene glycol. The oyster samples were evaluated by cell culture methods, as well as polymerase chain reaction (PCR) and quantitative-PCR. Moreover, at the end of the depuration period, the disinfected seawater was collected and analysed by PCR. The molecular assays showed that the HAdV5 genome was present in all of the depuration time samples, while the HAV genome was undetectable after 72 h of depuration. However, viral viability tests (integrated cell culture-PCR and immunofluorescence assay) indicated that both viruses were inactivated with 96 h of seawater recirculation. In conclusion, after 96 h of UV treatment, the depuration system studied in this work purified oysters that were artificially contaminated with HAdV5 and HAV.     

Assessing the bioaccumulation of contaminants from sediments by fish and other aquatic organisms

Contaminated sediments that are not acutely toxic to aquatic organisms but contain bioaccumulable toxic substances present a common, yet poorly understood problem for regulatory decision makers. In order to recommend options to minimize bioaccumulation of these toxic substances, decision-makers need estimates of 1. which substances are available for accumulation by aquatic organisms; and 2. the potential impacts of such accumulation. The most direct and meaningful approach to estimating bioavailability is measurement of contaminant uptake by aquatic organisms exposed to the sediments of concern. Reasonably reliable methodologies exist for performing such exposures in the laboratory and in situ using marine or freshwater organisms. Such methods can demonstrate short-term potential for bioaccumulation of toxics from the sediments, but not necessarily the biological significance or long-term impact of any accumulated residues in the organisms and transfer of those residues through the food chain. Since most contaminated sediments contain a mixture of toxic substances, determination of the biological significance of their accumulation is not likely in the near future. Thus, the direct measurement of significant bioaccumulation of toxic substances from the sediments remains the most immediately useful index in a decision-making process.Contribution Number 631 of the National Fisheries Center-Great Lakes     

多氯联苯在模拟水生态系统中的分布、积累与迁移动态研究

利用微宇宙模拟水生态系统对多氯联苯在水体环境中的行为 ,包括多氯联苯在水体、沉积物、生物体 (皇冠草、河蚬、鲢 )不同分配相及生物体不同组织、器官中含量、分布以及迁移、富集等时间变化动态过程进行了研究。结果表明 ,水草对PCB有一定程度的吸收 ;河蚬对PCBs有明显的吸收积累 ,并且吸收达到平衡的时间较长。鱼体对PCB的吸收积累在不同组织中有明显的差别 ,在内脏和肌肉组织中积累较高 ,鳃组织中积累较低。     

Sediment Composition Influences Spatial Variation in the Abundance of Human Pathogen Indicator Bacteria within an Estuarine Environment

Faecal contamination of estuarine and coastal waters can pose a risk to human health, particularly in areas used for shellfish production or recreation. Routine microbiological water quality testing highlights areas of faecal indicator bacteria (FIB) contamination within the water column, but fails to consider the abundance of FIB in sediments, which under certain hydrodynamic conditions can become resuspended. Sediments can enhance the survival of FIB in estuarine environments, but the influence of sediment composition on the ecology and abundance of FIB is poorly understood. To determine the relationship between sediment composition (grain size and organic matter) and the abundance of pathogen indicator bacteria (PIB), sediments were collected from four transverse transects of the Conwy estuary, UK. The abundance of culturable Escherichia coli, total coliforms, enterococci, Campylobacter, Salmonella and Vibrio spp. in sediments was determined in relation to sediment grain size, organic matter content, salinity, depth and temperature. Sediments that contained higher proportions of silt and/or clay and associated organic matter content showed significant positive correlations with the abundance of PIB. Furthermore, the abundance of each bacterial group was positively correlated with the presence of all other groups enumerated. Campylobacter spp. were not isolated from estuarine sediments. Comparisons of the number of culturable E. coli, total coliforms and Vibrio spp. in sediments and the water column revealed that their abundance was 281, 433 and 58-fold greater in sediments (colony forming units (CFU)/100g) when compared with the water column (CFU/100ml), respectively. These data provide important insights into sediment compositions that promote the abundance of PIB in estuarine environments, with important implications for the modelling and prediction of public health risk based on sediment resuspension and transport.