A proteome-wide screen of Campylobacter jejuni using protein microarrays identifies novel and conformational antigens.

Jiayou Liu,Russell L Finley,Jodi R Parrish,Julie Hines,Linda Mansfield,Nicholas J Mantis

doi:10.1371/journal.pone.0210351

Jiayou Liu, Russell L Finley + Show 4 more

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https://doi.org/10.1371/journal.pone.0210351

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Abstract

Campylobacter jejuni (C. jejuni) is a foodborne intestinal pathogen and major cause of gastroenteritis worldwide. C. jejuni proteins that are immunogenic have been sought for their potential use in the development of biomarkers, diagnostic assays, or subunit vaccines for humans or livestock. To identify new immunogenic C. jejuni proteins, we used a native protein microarray approach. A protein chip, with over 1400 individually purified GST-tagged C. jejuni proteins, representing over 86% of the proteome, was constructed to screen for antibody titers present in test sera raised against whole C. jejuni cells. Dual detection of GST signals was incorporated as a way of normalizing the variation of protein concentrations contributing to the antibody staining intensities. We detected strong signals to 102 C. jejuni antigens. In addition to antigens recognized by antiserum raised against C. jejuni, parallel experiments were conducted to identify antigens cross-reactive to antiserum raised against various serotypes of E. coli or Salmonella or to healthy human sera. This led to the identification of 34 antigens specifically recognized by the C. jejuni antiserum, only four of which were previously known. The chip approach also allowed identification of conformational antigens. We demonstrate in the case of Cj1621 that antigen signals are lost to denaturing conditions commonly used in other approaches to identify immunogens. Antigens identified in this study include those possessing sequence features indicative of cell surface localization, as well as those that do not. Together, our results indicate that the unbiased chip-based screen can help reveal the full repertoire of host antibodies against microbial proteomes.

Highlights

Bacterial infections continue to claim lives and burden health care systems worldwide
E. coli BL21 strains that express GST-tagged C. jejuni fusion were described previously [43]. 5 μL of the stock strains were inoculated into 300 μL of LB medium containing 100 μg/mL carbenicillin and 1% glucose, and were grown overnight at 37 ̊C. 100 μL of each overnight culture was transferred from a well of a 96-well plate to a well of a 24-well 10-mL plates containing 6 mL of the same medium
We previously cloned over 90% of the C. jejuni open reading frames (ORFs) into a vector for expression of N-terminally-tagged GST fusion proteins in E. coli [43]

Summary

Introduction

Bacterial infections continue to claim lives and burden health care systems worldwide. With the rise in antibiotic resistance, the need for new vaccines and diagnostics has become urgent [1, 2]. New vaccines and diagnostic assays can be difficult to develop, in part because bacterial pathogens can encode thousands of proteins, possess complex antigen profiles, and elicit. A proteome-wide screen of Campylobacter jejuni using protein microarrays identifies novel antigens. N01-AI-30058 and Enteric Research Investigational Network Cooperative Research Center (ERIN CRC) grant U19AI090872. All funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. There was no additional external funding received for this study

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