E. coli Is a Poor End-Product Criterion for Assessing the General Microbial Risk Posed From Consuming Norovirus Contaminated Shellfish

The fecal indicator organism (FIO) Escherichia coli is frequently used as a general indicator of sewage contamination and for evaluating the success of shellfish cleaning (depuration) processes. To evaluate the robustness of this approach, the accumulation, retention, and depuration of non-pathogenic E. coli, pathogenic E. coli O157:H7 and norovirus GII (NoV GII) RNA were evaluated using a combination of culture-based (E. coli) and molecular methods (E. coli, NoV GII) after exposure of mussels (Mytilus edulis) to water contaminated with human feces. We simulated water contamination after a point-source release from a combined sewer overflow (CSO) where untreated wastewater is released directly into the coastal zone. All three microbiological indicators accumulated rapidly in the mussels, reaching close to maximum concentration within 3 h of exposure, demonstrating that short CSO discharges pose an immediate threat to shellfish harvesting areas. Depuration (72 h) in clean water proved partially successful at removing both pathogenic and non-pathogenic E. coli from shellfish tissue, but failed to eradicate NoV GII RNA. We conclude that current EU standards for evaluating microbiological risk in shellfish are inadequate for protecting consumers against exposure to human norovirus GII found in polluted marine waters.

Effect of Ionic Presence on Chlorine Disinfection of Animal Viruses

Work by Scarpino in 1972 indicated that disinfection rates are affected by ionic presence. When disinfecting coxackievirus A9 with H0C1 at pH 6.0 in the presence of 0.05M KC1 the effect is to increase the disinfection rate 50 times. Disinfection of coxackievirus A9 at pH 10.0 shows that 0.05M KC1 increases the disinfection rate 10 times. This rate difference due to 0.05M KC1 between the two pH's may be due to the charge on the hypochlorite ion, the change of charge on the surface of the virion, or to a change in the chemical equilibrium of chlorine due to the pH. Studies with poliovirus 1 and 0.05M KC1 showed the same ion effect. When the disinfection of poliovirus 1 with hypochlorous acid (H0C1) was compared with the disinfection of E.coli, the E.coli was disinfected quicker than poliovirus 1. This relationship that viruses are more resistant than the fecal indicator organism E.coli is generally accepted. However, poliovirus 1 was more sensitive to hypochlorite ion (OCl−) than E. coli, when 0.05M KC1 was present in the buffer. Whereas the ion effect was present for coxsackievirus A9 and poliovirus 1, it was not present for E.coli. We have been pursuing this ion effect phenomenon using both chlorine and chloramines. To determine whether this ion effect was present with chloramines, similar tests were run using monochloramine (NH2C1) as the disinfectant. Again the effect held. That is, the presence of KC1 in the buffers increased the efficiency of the disinfectant. This ion-effect was dose dependent between 0.05M KC1 and 0.1M KC1, where the effect was greater with 0.1M KC1. The OCl− and NH2C1 studies were both run at pH 10.0 and at 5°C. Tests were then run to determine whether the effect held for dichloramine (NHC12) at pH 4.5. The ion-effect was present using dichloramine. However, the level of dichloramine dropped from 11.5 ppm to 6.5 ppm during the test. The chloramine loss may be due to the shift of equilibrium as represented by the following equation: With the formation of nitrogen tri-chloride (NC13) the ion-effect could be dissipated if the addition of KC1 was made at a different time. When the chlorine species was formed in the presence of KC1 the effect was present. When the KC1 was added to the already formed specie the effect was not present in the case of monochloramine and much reduced in the case of hypochlorous acid. Therefore, the time of the KC1 addition to the system was critical to the ion-effect phenomenon.