Open Access Research

Application of highly sensitive saturation labeling to the analysis of differential protein expression in infected ticks from limited samples

Margarita Villar1*, Alessandra Torina2, Yolanda Nuñez3, Zorica Zivkovic4, Anabel Marina3, Angela Alongi2, Salvatore Scimeca2, Giuseppa La Barbera2, Santo Caracappa2, Jesús Vázquez3 and José de la Fuente15*

Author Affiliations

1 Instituto de Investigación en Recursos Cinegéticos IREC (CSIC-UCLM-JCCM), Ronda de Toledo s/n, 13005 Ciudad Real, Spain

2 Intituto Zooprofilattico Sperimentale della Sicilia, Via G. Marinuzzi n°3, 90129 Palermo, Sicily, Italy

3 Centro de Biología Molecular "Severo Ochoa" (CSIC-UAM), 28049 Cantoblanco, Madrid, Spain

4 Utrecht Centre for Tick-borne Diseases (UCTD), Department of Infectious Diseases and Immunology, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 1, 3584CL, Utrecht, The Netherlands

5 Department of Veterinary Pathobiology, Center for Veterinary Health Sciences, Oklahoma State University, Stillwater, OK 74078, USA

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Proteome Science 2010, 8:43  doi:10.1186/1477-5956-8-43

Published: 12 August 2010

Abstract

Background

Ticks are vectors of pathogens that affect human and animal health worldwide. Proteomics and genomics studies of infected ticks are required to understand tick-pathogen interactions and identify potential vaccine antigens to control pathogen transmission. One of the limitations for proteomics research in ticks is the amount of protein that can be obtained from these organisms. In the work reported here, individual naturally-infected and uninfected Rhipicephalus spp. ticks were processed using a method that permits simultaneous extraction of DNA, RNA and proteins. This approach allowed using DNA to determine pathogen infection, protein for proteomics studies and RNA to characterize mRNA levels for some of the differentially expressed proteins. Differential protein expression in response to natural infection with different pathogens was characterized by two-dimensional (2-D) differential in gel electrophoresis (DIGE) saturation labeling in combination with mass spectrometry analysis. To our knowledge, this is the first report of the application of DIGE saturation labeling to study tick proteins.

Results

Questing and feeding Rhipicephalus spp. adult ticks were collected in 27 farms located in different Sicilian regions. From 300 collected ticks, only 16 were found to be infected: R. sanguineus with Rickettsia conorii and Ehrlichia canis; R. bursa with Theileria annulata; and R. turanicus with Anaplasma ovis. The proteomic analysis conducted from a limited amount of proteins allowed the identification of host, pathogen and tick proteins differentially expressed as a consequence of infection.

Conclusion

These results showed that DIGE saturation labeling is a powerful technology for proteomics studies in small number of ticks and provided new information about the effect of pathogen infection in ticks.