Antigen Fingerprinting of Polyclonal Antibodies Raised in Immunized Chickens with Tick Total Proteins: A Reservoir for the Discovery of Novel Antigens

Abstract

Identification of tick-protective antigens remains the limiting step in vaccine development. The authors have generated several B cell epitope candidates by fingerprinting Rhipicephalus (Boophilus) microplus proteins that were characterized through bioselection of random peptide phage display libraries against polyclonal antibodies antitick proteins. From 280 clones selected and sequenced, 107 distinct reactive clones were validated by dot-blot assays. Eight consensus motifs were generated, and the most frequent ones were PxxKxH, NxxKxxL, and HTS (68.2%, 65%, and 42%, respectively). The consensus sequences identified potential vaccine targets by alignment with the protein database of R. microplus, which may have putative roles in the host response. Sequences that did not align with known proteins but shared extensive homology among each other were classified as conformational epitopes. Sequence alignments also recognized multiple targets, and the most predominant proteins were identified. Finally, immunized mice sera recognized tick proteins, demonstrating that functional epitope profiles can be identified through selection of phage-displayed peptide libraries with hyperimmune sera and revealing that the epitope-displaying phages can be used as potential vaccine immunogens.

Transmembrane proteins – Mining the cattle tick transcriptome

“…These proteins are involved in processes such as absorption and metabolism of cholesterol, as well as lipoprotein assembly (Chang et al, 2006(Chang et al, , 2009Alpy and Tomasetto, 2014). All three proteins have high VaxiJen scores and contain antigenic motifs previously identified by Prudencio et al (2011) (Table 2, Fig. 3).…”

“…Homologous sequences for all of these 30 proteins were identified in I. scapularis. Six protein sequences contained consensus sequences representing potential R. microplus vaccine targets as identified by Prudencio et al (2011). To the best of our knowledge 19 proteins have not been identified using proteomics approaches in ticks, and only 11 have been previously reported (Table 2).…”

“…For all the transcripts identified, orthologous sequences could be identified in I. scapularis highlighting the possibility of further development of candidate sequences for possible cross-species tick vaccines. Six proteins contained consensus sequences identified by Prudencio et al (2011), representing potential targets for further evaluation in R. microplus. To the best of our knowledge 19 proteins have not been identified using proteomics approaches before in ticks while 11 have been previously reported (Table 2).…”

“…The final set of transmembrane proteins confirmed with LC-MS/MS analysis (Table S1) were also screened for their presence in the midgut transcriptome of R. decoloratus and were considered present if it showed a log 2 fold change of ≤1.7 -with P value <0.0 5- (van Zyl et al, 2015). Matching refseq and GenBank numbers from I. scapularis and consensus sequences as described by Prudencio et al (2011) were also identified.…”

“…Published proteins identified in any tick species by mass spectrometry are also shown. Consensus sequences identified according to Prudencio et al (2011) are indicated in the motif column. Contiguous sequence numbers are indicated with additional short numbers referred to in Fig.…”