Abstract:
One of the principle mechanisms utilised by ticks to obtain a blood meal is the subversion of the host’s
haemostatic response. This is achieved through the secretion of saliva containing anti-haemostatic proteins
into the feeding lesion. Lineage-specific expansion of predicted secretory protein families have been
observed in all previously studied ticks and occurred in response to adaptation to a blood-feeding environment.
Of these, the predominant families are common between both hard and soft ticks. One of these
families, namely the Kunitz domain-containing protein family, includes proven tissue factor pathway
inhibitor-like (TFPI-like) anti-haemostatics such as ixolaris and penthalaris that play a crucial role during
tick feeding. Although Kunitz-type proteins have been found in Rhipicephalus microplus, the TFPI-like
Kunitz protein family has not yet been studied. We report a comprehensive search for TFPI-like Kunitz
domain-containing proteins in R. microplus expressed sequence tag libraries, resulting in the identification
of 42 homologues. The homologues were bioinformatically and phylogenetically studied, including
the application of an intensive Bayesian Markov Chain Monte Carlo (MCMC) analysis of the individual
Kunitz domain nucleotide sequences. We show that the R. microplus TFPI-like Kunitz protein family
groups into two main clades that presumably underwent ancient duplication, which indicates that a
whole genome duplication event occurred at least 150 million years ago. Evidence for recent and ancient
gene and domain duplication events was also found. Furthermore, the divergence times of the various
tick lineages estimated in this paper correspond with those presented in previous studies. The elucidation
of this large protein family’s evolution within R. microplus adds to current knowledge of this economically
important tick.
