Murine Norovirus Interaction with Pseudomonas aeruginosa Biofilm in a Dynamic Bioreactor.

Biofilms can constitute permanent threats to food safety and public health. Bacteria and viruses lodged in biofilm can escape cleaning and sanitizing agents. The aim of this study was to compare Pseudomonas aeruginosa developing and mature biofilms produced on agri-food surfaces in terms of interaction with murine norovirus. Whether they were mature or still developing the biofilms apparently accumulated murine norovirus in large numbers after 24 h of contact with medium which viral titer was 2.6 × 104 pfu ml-1 (≈ 8 log10 genome copies ml-1). This appeared unrelated to surfaces' nature and bacterial viable count but related to polysaccharide and protein contents. Virus releases may also occur mainly in connection with P. aeruginosa biofilm dispersal systems. These findings suggest that the effectiveness of surface cleaning agents and procedures for reducing the risks of biofilms-related viral contaminations need to be re-evaluated in relation with biofilm components. However, more repetitions and further in-depth specific studies are needed for confirmation of these findings and more clarifications on virus-biofilm interaction phenomenon.

Adhesion of Norovirus to Surfaces: Contribution of Thermodynamic and Molecular Properties Using Virus-Like Particles.

The aim of the study was to assess human norovirus and feline calicivirus (FCV) surface free energy, hydrophobicity, and ability to interact with fresh foods and food-contact surfaces. Virus-like particles (VLPs) of human norovirus (GI.1 and GII.4) and FCV were produced, purified, and analyzed for their surface free energy, hydrophobicity, and the total interfacial free energy of interaction [Formula: see text] with lettuce, strawberry, polyethylene, and stainless steel. GII.4 VLPs were further tested for adhesion at different pH, ionic strengths, and temperature. All the VLPs and the test materials showed low surface energies, as well as hydrophobic characters except for GI.1. Nearly all [Formula: see text] values were propitious for spontaneous adhesion. GII.4 VLPs adsorbed almost indifferently to stainless steel, polyethylene, and lettuce. Isoelectric point and high temperature generally promoted adhesion while ionic strength effect was surface-dependant. According to this study, all the materials assessed are of low-energy and hydrophobic nature except GI.1 VLPs. Interfacial free energies of interaction were favorable for spontaneous adhesion ([Formula: see text] < 0) of all VLPs to the test materials, except for GI.1 VLPs to both stainless steel and straweberry. It is also found that norovirus adhesion is more sensitive to physicochemical conditions than to surface character itself.

Stability of Secondary and Tertiary Structures of Virus-Like Particles Representing Noroviruses: Effects of pH, Ionic Strength, and Temperature and Implications for Adhesion to Surfaces.

Loss of ordered molecular structure in proteins is known to increase their adhesion to surfaces. The aim of this work was to study the stability of norovirus secondary and tertiary structures and its implications for viral adhesion to fresh foods and agrifood surfaces. The pH, ionic strength, and temperature conditions studied correspond to those prevalent in the principal vehicles of viral transmission (vomit and feces) and in the food processing and handling environment (pasteurization and refrigeration). The structures of virus-like particles representing GI.1, GII.4, and feline calicivirus (FCV) were studied using circular dichroism and intrinsic UV fluorescence. The particles were remarkably stable under most of the conditions. However, heating to 65°C caused losses of ß-strand structure, notably in GI.1 and FCV, while at 75°C the α-helix content of GII.4 and FCV decreased and tertiary structures unfolded in all three cases. Combining temperature with pH or ionic strength caused variable losses of structure depending on the particle type. Regardless of pH, heating to pasteurization temperatures or higher would be required to increase GII.4 and FCV adhesion, while either low or high temperatures would favor GI.1 adhesion. Regardless of temperature, increased ionic strength would increase GII.4 adhesion but would decrease GI.1 adhesion. FCV adsorption would be greater at refrigeration, pasteurization, or high temperature combined with a low salt concentration or at a higher NaCl concentration regardless of temperature. Norovirus adhesion mediated by hydrophobic interaction may depend on hydrophobic residues normally exposed on the capsid surface at pH 3, pH 8, physiological ionic strength, and low temperature, while at pasteurization temperatures it may rely more on buried hydrophobic residues exposed upon structural rearrangement.

Zeta Potential and Aggregation of Virus-Like Particle of Human Norovirus and Feline Calicivirus Under Different Physicochemical Conditions.

Although the spread of human norovirus reportedly depends on its ability to bind to food materials, the mechanism of the phenomenon remains unknown. Since protein size and electrical charge are reportedly important parameters in their adsorption, the current work is focused on determining human noroviruses isoelectric point (IEP), electrical charge and aggregate size at different pH, ionic strength (IS), and temperature. Using the baculovirus expression vector system, we produced and purified virus-like particles (VLPs) of GI.1 and GII.4 noroviruses and feline calicivirus, determined their IEP, and examined their size and electrical charge using a Zetasizer Nano ZS apparatus. Shape and size were also visualized using transmission electron microscopy. IEPs were found close to pH 4. Net charge increased as the pH deviated from the IEP. VLPs were negatively charged at all IS tested and showed a gradual decrease in charge with increasing IS. At low temperature, VLPs were 20-45 nm in diameter at pH far from their IEP and under almost all IS conditions, while aggregates appeared at or near the IEP. At increased temperatures, aggregates appeared at or near the IEP and at high IS. Aggregation at the IEP was also confirmed by microscopy. This suggests that electrostatic interactions would be the predominant factor in VLPs adhesion at pH far from 4 and at low ionic strength. In contrast, non-electrostatic interactions would prevail at around pH 4 and would be reinforced by aggregates, since size generally favors multiple bonding with sorbents.