Application of bacteriophage-displayed peptide libraries to study polysaccharide antigens of Aeromonas salmonicida

Animal diseases, Aquatic sciences

McCafferty, Calum Angus

University:University of Glasgow

关键词: Animal diseases, Aquatic sciences;   

Others  :  http://theses.gla.ac.uk/71566/1/10391194.pdf

来源: University of Glasgow

【 摘 要 】

This thesis describes the application of bacteriophage displayed peptide libraries to the salmonid fish pathogen Aeromonas salmonicida. Antisera to both the capsular polysaccharide and lipopolysaccharide of A. salmonicida were used to select phage from 6-mer and 15-mer bacteriophage peptide libraries displayed in the p III protein of bacteriophage fd tet. When rabbit polyclonal anti-capsular polysaccharide antiserum was used for selection of phage from the 6-mer library, 140 phage were recovered by elution with glycine /HCl buffer (first round elution) and capsular polysaccharide solution in the second and third round of elution. Phage were amplified to high concentration and analysed by enzyme linked immunosorbent assay (ELISA) and the insert sequences determined Biopanning with anti-capsular polysaccharide antiserum selected a range of phage with peptide insert sequences which contained similar 3 to 4 amino acid motifs. The most predominant motif was 'serine-glycine-serine'. ELISA demonstrated that several of these phage bound to anti-capsular polysaccharide antiserum suggesting that these phage peptide sequences might antigenically mimic some capsular polysaccharide epitopes to which antibodies were present in the antiserum. A monoclonal antibody to A. salmonicida lipopolysaccharide was also used for biopanning with the 6-mer library and this selected phage with sequences which could be classified into groups, several containing identical sequences and others with similar motifs. ELISA assays to detect binding of these phage to lipopolysaccharide antiserum were inconclusive. Several selected with anti-lipopolysaccharide monoclonal antibody displayed peptides similar to those selected by polyclonal anti-capsular polysaccharide antiserum, which may reflect the similar monosaccharide composition of these related polysaccharides.