Thermal Stability of Viruses in Coculture with Enterobacter cloacae

Introduction: Human noroviruses (hNoV) are the primary cause of foodborne disease in the United States. Thermal stability has been studied though there is limited data on the impact of bacteria-virus interactions on the stability of enteric viruses such as hNoV. It is presumed that viruses associated with environmental materials are physically more stable than nonassociated viruses.

Purpose: This study aims to investigate the thermal stability of viruses when associated with Enterobacter cloacae.

Methods: Tulane virus (TuV), murine norovirus (MNV), and Aichivirus (AiV) were selected for thermal stability evaluation along with E. cloacae. For each experiment, 400 µl 10⁵ PFU/ml of virus was added to 400 µl 10⁹ CFU E. cloacae/ml and exposed to 37, 56 and 63, and 72°C for up to 24 h, 30 min, and 10 min, respectively. To detect virus at 37°C not associated with E. cloacae, samples were filtered prior to analysis by plaque assay. At ≥56°C, E. cloacaewas inactivated by ≤ one minute; thus, both filtered and whole samples were analyzed at specific time intervals.

Results: Analysis of AiV has been completed. At 37°C, AiV remained stable at five logs for 24 h and was fully associated with E. cloacae by 6 h postinoculation. At 56°C, AiV with and without bacteria was completely inactivated at 10 min with D-values of 41 and 43 s, respectively. Five-log inactivation of AiV with and without E. cloacae was achieved by 30 s at 63 and 72°C. TuV and MNV experiments are ongoing.

Significance: The use of viral surrogates is critical in applied food safety research related to thermal inactivation. Often researchers study pathogens in monoculture, and this is not representative of the “real-world”. These data are the first step in understanding the role that microbe-microbe interactions may play in survivability.