title: Production of monoclonal antibodies to Listeria monocytogenes and their application to determine the virulence of isolates from channel catfish.
abstract: We produced monoclonal antibodies (MAbs) to the extracellular proteins of Listeria monocytogenes EGD grown in Chelex-treated improved minimal medium. Ten of the positive hybridomas generated were chosen for further characterization. Seven of the MAbs reacted with a protein having a molecular mass of 60 kDa. These MAbs inhibited listeriolysin (LLO)-mediated hemolysis, and two of them were specific for LLO and none of the other thiol-activated toxins tested. In an enzyme-linked immunosorbent assay and Western blot analysis, five of the anti-LLO MAbs reacted with ivanolysin from Listeria ivanovii. Three of the 10 MAbs reacted with a 29-kDa protein on Western blots and neutralized the phosphatidylcholine-specific phospholipase C (PC-PLC) activity of L. monocytogenes. These three anti-PC-PLC MAbs did not react with phospholipases from five different gram-positive bacteria. However, the anti-PC-PLC MAbs recognized a 27-kDa extracellular protein from L. ivanovii and neutralized sphingomyelinase activity in a hemolysis test that demonstrates the antigenic relatedness of listerial phospholipases. These data indicate that listerial thiol-activated toxins possess species-specific epitopes and share group-specific epitopes. This is the first description of MAbs that neutralize listerial PC-PLC, and the data suggest that there is antigenic similarity between L. monocytogenes PC-PLC and L. ivanovii sphingomyelinase. The reactions of the MAbs with catfish isolates of L. monocytogenes suggested that some of the isolates examined lack the LLO and/or PC-PLC required for pathogenicity. The MAbs described here differentiated some catfish isolates from previously described type strain-pathogenic isolates and could be useful for detecting and determining the virulence of L. monocytogenes in food and clinical samples and for detecting L. ivanovii in veterinary clinical samples.
authors.affiliation: Veterinary Medical Research Program, College of Veterinary Medicine, Mississippi State University, Mississippi State, Mississippi 39762, USA.
authors.forename: S
authors.initials: S
authors.lastname: Erdenlig
authors.forename: A J
authors.initials: AJ
authors.lastname: Ainsworth
authors.forename: F W
authors.initials: FW
authors.lastname: Austin
mesh_terms: D000818:Animals; D000907:Antibodies, Bacterial / Q000096:biosynthesis; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000276:immunology; D001427:Bacterial Toxins; D015153:Blotting, Western; D004797:Enzyme-Linked Immunosorbent Assay; D006360:Heat-Shock Proteins / Q000276:immunology*; D006460:Hemolysin Proteins / Q000276:immunology*; D006461:Hemolysis; D007059:Ictaluridae / Q000382:microbiology*; D008089:Listeria monocytogenes / Q000276:immunology / Q000302:isolation & purification / Q000472:pathogenicity*; D010740:Phospholipases / Q000378:metabolism; D014118:Toxins, Biological; D010738:Type C Phospholipases / Q000276:immunology / Q000378:metabolism; D014774:Virulence
chemical_list: D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D001427:Bacterial Toxins; D006360:Heat-Shock Proteins; D006460:Hemolysin Proteins; D014118:Toxins, Biological; D010740:Phospholipases; D010738:Type C Phospholipases; C105598:phosphatidylcholine-specific phospholipase C; C038705:hlyA protein, Listeria monocytogenes