Detection of serpins involved in cellular immune response of Rhipicephalus microplus challenged with fungi

Abstract

The understanding of tick physiology and immune system is important to improve the effective control of this ectoparasite. Invertebrates' innate immune response is activated when the organism is challenged with pathogens. The present study describes the changes of serine proteinase inhibitors (serpins) and in the number of circulating haemocytes involved in cellular immune defence of Rhipicephalus microplus engorged females challenged with the entomopathogenic fungi Metarhizium anisopliae or Beauveria bassiana, or with the non-entomopathogenic fungus Fusarium oxysporum. The cell-free haemolymph was separated from haemocytes by centrifugation and cells were re-suspended in phosphate buffer pH 7.2. The proteins of haemocytes were analysed by SDS-PAGE and the segments of the 1D gel were submitted to protein digestion with trypsin. The peptides were analysed by liquid chromatography coupled with electrospray tandem mass spectrometry (LC-ESI-MS/MS). The analysis by mass spectrometry allowed the identification of several proteins through the search in the database built based on public banks of Ixodidae and Argasidae. In haemocytes, many proteins were identified highlighting serpins. The results showed that the entomopathogenic fungi M. anisopliae or B. bassiana reduced the amount of serpins, while F. oxysporum increased. The present study reports, for the first time, the variation of serpins in haemocytes of R. microplus engorged females infected by fungi.

Keywords:

• serpins
• Metarhizium anisopliae
• Beauveria bassiana
• Fusarium oxysporum
• proteomics
• cellular immune response

Acknowledgements

This research was supported by grants from the National Council for Scientific and Technological Development (CNPq) and the Rio de Janeiro State Research Foundation (FAPERJ). We also thank the Office to Improve University Research (CAPES) for providing the M.Sc. and Ph.D. scholarships. V.R.E.P.Bittencourt is CNPq researcher.