Polyelectrolyte Flocculation as an Aid to Recovery of Enteroviruses from Oysters

An improved recovery method and testing strategy were devised for recovery of low numbers of enteric viruses from each of three commercially important shellfish species. Effective recovery of virus depended as much upon details of the test strategy adopted for use of the improved method with each species as on the method itself. The most important test details involved sample composition, pool size, and method of use of cell cultures. Recovery sensitivity measured permitted detection of 25 to 3 plaque-forming units of enteroviruses and 100 to 27 plaque-forming units of reovirus through their recovery in cell culture, with effectivenesses averaging 64 and 46%, respectively. Test samples prepared by the improved recovery method were virtually cytotoxicity free. Optimal recovery of virus on 45-cm2 cell culture monolayers was obtained with 1-ml inocula adsorbed for 2 h. The most effective recovery of virus from shellfish samples was made by a sequential adsorption procedure which allowed equal exposure of an entire sample to each of two or more cell cultures. Removal of nonviral contaminants from test samples by antibiotic treatment was preferable to the use of ether or membrane filtration procedures. Surveillance ofthe sanitary quality of shellfish and shellfish-growing waters has been carried out since 1925 in the United States, when a certification program administered by the U.S. Public Health Service was instituted (1). This monitoring program, which developed into the National Shellfish Sanitation Program (NSSP), was instituted after a typhoid epidemic resulting in 1,500 cases with 150 deaths was attributed to sewage-polluted oysters (13). This epidemic, coming after pmany previously reported outbreaks of enteric disease attributed to shellfish, helped to focus critical attention upon the health hazards associated with the consumption of polluted shellfish. Since then a number of shellfishassociated outbreaks of infectious hepatitis and viral gastroenteritis have been reported. These reports have been reviewed and summarized recently by Gerba and Goyal (5). All of the commercially important shellfish, oysters, hardand soft-shell clams, and mussels, have been incriminated as vehicles for the transmission of enteric virus diseases to humans.

Section: Discussionmentioning

confidence: 99%

Improved method and test strategy for recovery of enteric viruses from shellfish

Metcalf¹,

Moulton²,

Eckerson³

1980

“…Microbiol. 1981, Q51, p. 209), cationic flocculant treatment (11), and filtration through positively charged filters (10). We found that freon treatment and positively charged filters were inadequate for removing cytotoxicity from crab concentrates.…”

Section: Discussionmentioning

confidence: 85%

Concentration of rotavirus and enteroviruses from blue crabs (Callinectes sapidus)

Seidel¹,

Goyal²,

Rao³

et al. 1983

A simple method for concentration and detection of rotavirus and enteroviruses in the blue crab is described. Virus was separated from tissue homogenates at pH 9.5, concentrated by adsorption to protein precipitates at pH 3.5, and recovered by elution of precipitates at pH 9.2. Test samples of 12 to 15 ml were produced from an initial 100 g of crab tissues. Cat-floc precipitation was used to remove sample toxicity for cell cultures. Recovery effectiveness averaged 52% with poliovirus 1, echovirus 7, and coxsackievirus B5 and 23% with rotavirus SAil.

“…ND, Not done. No 77 (69-100) 80 (69-100) 14 (7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25) a Oyster samples (40 to 100 g) were inoculated with 20 to 2,000 PFU of virus, diluted 1:1 with water, and adjusted to pH 8.9. Catfloc was added if indicated, and the samples were centrifuged.…”

Section: Resultsmentioning

confidence: 99%

“…Prior to 1978, ultrafiltration or ultracentrifugation was used for concentration of virus from shellfish samples (11,12). To determine the suitability for recovery of rotavirus and poliovirus from Canadian Atlantic oysters, several of these methods were compared by using oysters inoculated in the laboratory.…”

mentioning

confidence: 99%

Methods for recovering poliovirus and rotavirus from oysters

Speirs

Pontefract

Harwig

1987

Polioviruses and rotaviruses are potential indicators of sewage pollution of water and shellfish. Several methods for detecting these viruses in oysters were assessed. Elution-precipitation involving Catfloc for clarification and skim milk for subsequent flocculation resulted in the recovery of an average of 79% of poliovirus type 1 and 37% of rotavirus SA-li from oyster homogenates inoculated with low numbers of these viruses. Adsorption-elution-precipitation did not improve the recovery of poliovirus and was detrimental to the recovery of rotavirus. Ultrafiltration or ultracentrifugation resulted in improved recovery of rotavirus but also in higher toxicity of oyster extracts to cell cultures. We recommend the use of the described elutionprecipitation method for detecting viral pollutants in samples of oysters.