172
D. Polo et al.
Discussion
The main objective of this study was to obtain a broad picture of the MNV-1
removal dynamics, as a model of human norovirus behavior, in clams and mussels
subjected to depuration processes. The inadequacy of current European regulations
to assess the sanitary quality of shellfish and classification of harvesting areas to
prevent viral contamination is well known (Anonymous 2004; Le Guyader et al.
2003; Romalde et al. 2002).In addition, enteric viruses concentrated by shellfish
can persist under depuration conditions that are sufficient for bacteria removal. How
infectious viruses can persist within the shellfish and their removal kinetics under
depuration conditions are important questions to be addressed.
The results of this study showed differences in the viral uptake and the removal
rate for viruses between clams and mussels subjected to an artificial bioaccumulation process with MNV-1. Fluctuations of the viral loads in consecutive depuration
stages were detected, which can be attributed to the heterogeneity distribution
of viral load. After 24 h of bioaccumulation the average viral uptake in clams
was 73.8 % higher than in mussels. However, only three out of five depuration
experiments with clams showed some reduction in viral quantification. The average
reduction in these three experiments was 0.5 log units (41.4 % reduction in RNA
copies/g DT). Mussels showed viral reduction in all the depuration trials with an
average reduction of 0.8 log units (74 % reduction in RNA copies/g DT).
Bioaccumulation rates and removal dynamics of MNV-1 seem to be different in
manila clams and mussels. Higher bioaccumulation levels were reached in clams,
but higher depuration rates have been observed in mussels.
These results indicate a different behavior ofMNV-1 in these two bivalve species.
Nappier et al. (2008), also reported a statistically higher bioaccumulation and
retention rate of MNV, NoV and HAV in Crassostrea ariakensis than in C. virginica.
Other studies carried out in our laboratory have shown a different bioaccumulation
and removal pattern in Manila clams artificially contaminated with HAV and MNV1 (unpublished data).
Although there are other studies in which viral bioaccumulation and subsequent
depuration of enteric viruses in bivalve molluscs is determined, to our knowledge,
this is the first to compare the rate of bioaccumulation and subsequent removal
dynamics of murine norovirus by qRT-PCR in these two molluscs.
The observed differences in MNV-1 uptake and removal dynamics could be
related to viral properties and/or to the existence of specific ligands. The specific
binding of NoV strains to digestive tract of shellfish through an A-like human histoblood group antigens (HBGAs) has been demonstrated (Le Guyader et al. 2006b;
Tian et al. 2006, 2007). Bivalve shellfish could, therefore, specifically concentrate
different viruses or specific strains on the basis of these receptors/ligands, while
other virus could be accumulated to a lesser extent by nonspecific mechanisms of
attachment, like mechanical entrapment and ionic bonding (Burkhardt and Calci
2000; Di Girolamo et al. 1977; Schwabm et al. 1998), and consequently, be more
rapidly depurated.
D. Polo et al.
Discussion
The main objective of this study was to obtain a broad picture of the MNV-1
removal dynamics, as a model of human norovirus behavior, in clams and mussels
subjected to depuration processes. The inadequacy of current European regulations
to assess the sanitary quality of shellfish and classification of harvesting areas to
prevent viral contamination is well known (Anonymous 2004; Le Guyader et al.
2003; Romalde et al. 2002).In addition, enteric viruses concentrated by shellfish
can persist under depuration conditions that are sufficient for bacteria removal. How
infectious viruses can persist within the shellfish and their removal kinetics under
depuration conditions are important questions to be addressed.
The results of this study showed differences in the viral uptake and the removal
rate for viruses between clams and mussels subjected to an artificial bioaccumulation process with MNV-1. Fluctuations of the viral loads in consecutive depuration
stages were detected, which can be attributed to the heterogeneity distribution
of viral load. After 24 h of bioaccumulation the average viral uptake in clams
was 73.8 % higher than in mussels. However, only three out of five depuration
experiments with clams showed some reduction in viral quantification. The average
reduction in these three experiments was 0.5 log units (41.4 % reduction in RNA
copies/g DT). Mussels showed viral reduction in all the depuration trials with an
average reduction of 0.8 log units (74 % reduction in RNA copies/g DT).
Bioaccumulation rates and removal dynamics of MNV-1 seem to be different in
manila clams and mussels. Higher bioaccumulation levels were reached in clams,
but higher depuration rates have been observed in mussels.
These results indicate a different behavior ofMNV-1 in these two bivalve species.
Nappier et al. (2008), also reported a statistically higher bioaccumulation and
retention rate of MNV, NoV and HAV in Crassostrea ariakensis than in C. virginica.
Other studies carried out in our laboratory have shown a different bioaccumulation
and removal pattern in Manila clams artificially contaminated with HAV and MNV1 (unpublished data).
Although there are other studies in which viral bioaccumulation and subsequent
depuration of enteric viruses in bivalve molluscs is determined, to our knowledge,
this is the first to compare the rate of bioaccumulation and subsequent removal
dynamics of murine norovirus by qRT-PCR in these two molluscs.
The observed differences in MNV-1 uptake and removal dynamics could be
related to viral properties and/or to the existence of specific ligands. The specific
binding of NoV strains to digestive tract of shellfish through an A-like human histoblood group antigens (HBGAs) has been demonstrated (Le Guyader et al. 2006b;
Tian et al. 2006, 2007). Bivalve shellfish could, therefore, specifically concentrate
different viruses or specific strains on the basis of these receptors/ligands, while
other virus could be accumulated to a lesser extent by nonspecific mechanisms of
attachment, like mechanical entrapment and ionic bonding (Burkhardt and Calci
2000; Di Girolamo et al. 1977; Schwabm et al. 1998), and consequently, be more
rapidly depurated.