Purpose: With a focus on detection applications, this study sought to identify a broadly reactive bacterial ligand that might be used to facilitate norovirus capture and/or detection.
Methods: Seven previously characterized bacterial isolates (2 ATCC strains; 5 fecal isolates obtained from norovirus-positive stool samples) were used in this study. Isolates were grown in minimal media overnight and sonicated. These whole bacterial lysates were examined for three characteristics (1) histo-blood group antigen (HBGA) activity; (2) lectin binding; and (3) human norovirus binding. Western blots determined which bacterial glycoproteins bound the widest variety of viral strains and identified potential residues facilitating virus-bacteria interactions. Glycoproteins binding multiple human norovirus strains were isolated, purified and further analyzed using LC-MS.
Results: All of the bacterial species tested possessed various levels of HBGAs (relative densities: AB 1.0-15.8; B 0.5-5.1; H 0-0.8; Lea 0-2.1; Leb 0-1.3; Ley 0-0.7). Lectin binding narrowed the bacterial residues to specific sugars: N-acetyl galactosamine, relative density 0.2-5.7; α-D-mannose/glucose, relative density 0-10.7; α-L-fucose, relative density 0.8-4.9; and α-D-galactose, relative density 0.0-4.7. A virus overlay confirmed that select human norovirus strains bound these sugar residues with relative densities of: GI.6, 0-0.8; GII.4, 0-0.2; and GII.17, 0.0-0.8. Six bacterial strains bound the GII norovirus strains, via glycoproteins of 140 kDa, 35 kDa, and 17 kDa. A 35 kDa glycoprotein from a Bacillus fecal isolate bound the GI and GII human norovirus strains tested.
Significance: This is the first study to identify specific bacterial residues possessing human norovirus binding potential. With further development, these molecules may be exploited as potential ligands for human norovirus capture and/or detection.