In vitro time and temperature dependence effects of three molecular weight (53, 421 and approximately 1150 kilodaton) chitosans against human noroviral surrogates

Chitosan, a deacetylated chitin, is known to have a broad spectrum of antimicrobial activity. The objectives of this research were to determine the in vitro time and temperature dependent effects of chitosan with different molecular weights (53, 421 and ~1150 kDa) at two concentrations (35 percent and 0.7%) against the human noroviral surrogates, feline calicivirus (FCV-F9) and murine norovirus (MNV-1) at two temperatures (37oC and 18oC) over 2 hours. The virus titers were determined using standard plaque assays and compared to untreated virus controls. The tested chitosans were shown to decrease FCV-F9 titers, but had no observable effects on MNV- 1. Chitosan with molecular weight (M.W.) of 53 kDa at 0.35% decreased FCV-F9 titers by ~1.0 log PFU/ml, whereas at 0.7% decreased FCV-F9 titers by ~3.0 log PFU/ml after 2 h at 37oC and by ~1.0 log PFU/ml after both 1 and 2 h at 18oC. Chitosan with M.W. of 421 kDa at 0.35% decreased FCV-F9 titers by ~1.0 log PFU/ml whereas at 0.7% decreased FCV-F9 titers by ~2.0 at 37oC after 2 h. Chitosans with 421 and ~ 1150 kDa at 0.35% and 0.7% decreased FCV-F9 titers by ~1.0 log PFU/ml after 2 h at 18oC. Chitosan with ~ 1150 kDa at 0.35% decreased FCVF9 titers by ~1.0 log PFU/ml whereas at 0.7% decreased FCV-F9 titers by ~2.0 log PFU/ml at 37oC after 2 h. The results show that, in general, higher temperature and longer incubation cause greater inactivation of FCV-F9, with 53 kDa chitosan showing the greatest activity. Transmission electron microscopy did not show clear evidence of capsid damage of FCV-F9 treated chitosan and chitosan’s mode of action could not be clearly elucidated. Overall, the tested chitosans showed minimal effect against MNV-1. Thus, caution must be used in interpreting the data based solely on chitosan’s effects against FCV-F9 alone for the use of chitosan as a natural antiviral agent to control foodborne viral infections.