Identification of immunogenic proteins and generation of antibodies against Salmonella Typhimurium using phage display.

Torsten Meyer,André Frenzel,Michael Hust,Stefan Dübel,Thomas Schirrmann,Gerald F Gerlach,Janin Stratmann-Selke,Katrin Strutzberg-Minder,Sebastian Miethe

doi:10.1186/1472-6750-12-29

Torsten Meyer, André Frenzel + Show 7 more

Open Access

https://doi.org/10.1186/1472-6750-12-29

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Abstract

BackgroundSolely in Europoe, Salmonella Typhimurium causes more than 100,000 infections per year. Improved detection of livestock colonised with S. Typhimurium is necessary to prevent foodborne diseases. Currently, commercially available ELISA assays are based on a mixture of O-antigens (LPS) or total cell lysate of Salmonella and are hampered by cross-reaction. The identification of novel immunogenic proteins would be useful to develop ELISA based diagnostic assays with a higher specificity.ResultsA phage display library of the entire Salmonella Typhimurium genome was constructed and 47 immunogenic oligopeptides were identified using a pool of convalescent sera from pigs infected with Salmonella Typhimurium. The corresponding complete genes of seven of the identified oligopeptids were cloned. Five of them were produced in E. coli. The immunogenic character of these antigens was validated with sera from pigs infeced with S. Tyhimurium and control sera from non-infected animals. Finally, human antibody fragments (scFv) against these five antigens were selected using antibody phage display and characterised.ConclusionIn this work, we identified novel immunogenic proteins of Salmonella Typhimurium and generated antibody fragments against these antigens completely based on phage display. Five immunogenic proteins were validated using a panel of positive and negative sera for prospective applications in diagnostics of Salmonela Typhimurium.

Highlights

IntroductionTwo species are in the genus Salmonella: S. bongeri and S. enterica [1]
Generation of the Salmonella Typhimurium genomic phage display library Sonication of Salmonella DNA did not lead to clonable DNA fragments, whereas the sonication of E. coli DNA
Genomic DNA was digested with a mixture of the 4 base pair cutters DpnI and AluI and the 6 base pair cutter AfeI to construct the Salmonella Typhimurium genome library (Figure 2A)

Summary

Introduction

Two species are in the genus Salmonella: S. bongeri and S. enterica [1]. Salmonella enterica is classified in serogroups and serovars on the basis of their O- and H-antigens (somatic and flagellar antigens) [2,3]. 2800 Salmonella enterica gene families and more than 2500 serovars are known. More than 1500 serovars belong to the subspecies Salmonella enterica subspecies enterica [4]. These pathogens cause foodborne gastrointestinal infections, usually through raw poultry and pork, but it can be found in non-alcoholic beer or seafood. The subspecies enterica is the cause of 99% of human Salmonella

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