Identification and antigenicity of the Babesia caballi spherical body protein 4 (SBP4)

Abstract

Background: The tick-borne intra-erythrocytic apicomplexan Babesia caballi is one of the etiological agents of equine babesiosis, an economically important disease of equids in most tropical and subtropical areas of the world. Discovering candidate antigens for improved diagnostic tools and vaccines remains needed for controlling equine babesiosis. This study describes the B. caballi sbp4 (Bcsbp4) gene and protein (BcSBP4) and analyzes its antigenicity in infected equids. Methods: BLAST searches of an uncurated B. caballi assembly genome using the B. bovis SBP4 as a query were carried out, followed by PCR amplification and sequencing of a newly identified BcSBP4. Characterization of this novel gene and protein was performed by bioinformatics analysis, western blots, immunofluorescence (IFA) and an in vitro neutralization test using anti SBP4 peptide antibodies. Antigenicity of recombinant BcSBP4 (rBcSBP4) was tested with sera from field animals (n = 18) using an indirect ELISA (iELISA). Results: Babesia caballi genome searches using B. bovis SBP4 as a query allowed identification of a novel gene termed Bcsbp4. The Bcsbp4 gene encodes for a protein of 30.58 kDa, which is fully conserved among B. caballi isolates from USA and Egypt. Bioinformatics analysis indicates that BcSBP4 contains a signal peptide and lacks additional transmembrane domains. Expression of BcSBP4 in blood stages of B. caballi was confirmed by western blot and IFA using antibodies against synthetic peptides representing putative B-cell epitopes of BcSBP4 predicted by in silico analysis. In vitro neutralization tests using anti-BcSBP4 peptide antibodies showed a marginal, but statistically significant inhibitory effect on the infectivity of B. caballi merozoites in horse red blood cells. Sera from eight B. caballi-infected equids, but none out of ten negative equid control sera, gave a positive signal in an rBcSBP4 based iELISA. Conclusions: The Bcsbp4 gene is expressed in B. caballi blood stages. The BcSBP4 protein is a potential candidate for developing a novel serological test that could detect B. caballi infection in equids in tropical and subtropical countries worldwide.

Babesia gibsoni Whole-Genome Sequencing, Assembling, Annotation, and Comparative Analysis

“…Spherical body proteins (SBPs) are specific proteins to Babesia spp. and Theileria spp., which are localized in the cytoplasm of erythrocytes and get involved in the secretion of apical organelles during the merozoite stage and lead to a unique mechanism of invasion ( 62 , 63 ). Multiple sequence alignments were performed between B. gibsoni genome and SBPs (SBP2, SBP3, and SBP4) from B. bovis.…”

Impact of Equine and Camel Piroplasmosis in Egypt: How Much Do We Know about the Current Situation?

“…Additionally, in many rural areas of Egypt, horses, donkeys, mules, and ponies are often used as working equids. These animals assist personnel in a variety of sectors, including agriculture and construction, help farmers in soil drilling and public transportation, and contribute to sustaining the livelihoods of millions of people [5,6].…”

“…This disease is caused by blood-borne piroplasmids, mainly Babesia and Theileria. It occurs frequently in rural areas of developing countries, including Egypt, where there is still a huge reliance on working equines, which include horses, donkeys, mules, and ponies [5][6][7]. Particularly, camels are an economically significant multipurpose animal that has been utilized traditionally as an important source of milk, meat, and wool, and they are widely distributed in Africa, the Middle East, and Northern India [8,9].…”

Rapid Detection of Equine Piroplasms Using Multiplex PCR and First Genetic Characterization of Theileria haneyi in Egypt

“…In rural areas of many developing countries, including Egypt, there is a huge reliance on working equids, which include horses, donkeys, mules, and ponies. These animals play important roles in sustaining the livelihoods of millions of people by providing support in industries that include agriculture, construction, tourism, mining, and public transport [1,2]. The health and welfare of domesticated equids are often overlooked in rural areas.…”

“…However, investigations on vector competence for T. haneyi has not been reported [8]. Infection with T. equi and B. caballi can cause severe economic losses in the equine industry due to the cost of treatment, especially for acutely infected horses, which, in the absence of appropriate treatment, can die [2]. It was found that T. haneyi rarely causes clinical signs in field horses [9], even in splenectomized horses experimentally infected with T. haneyi using the intravenous (IV) route [5,10].…”