Identification and antigenicity of the Babesia caballi spherical body protein 4 (SBP4).
Abstract: 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.
Publication Date: 2020-07-22 PubMed ID: 32698835PubMed Central: PMC7376649DOI: 10.1186/s13071-020-04241-9Google Scholar: Lookup
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- Journal Article
Summary
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The research focuses on the identification and analysis of a new gene in the tick-borne parasite Babesia caballi that causes equine babesiosis. This new gene, called Bcsbp4 and resulting protein (BcSBP4), could potentially be used to develop a serological test for detecting the disease.
Study methodology and findings
- To identify potential antigen candidates, a BLAST search was used on an uncurated B. caballi assembly genome with B. bovis SBP4 as a query. This step resulted in the discovery of Bcsbp4, a novel gene that was then amplified and sequenced.
- The Bcsbp4 gene was characterised through computational analysis, western blots, immunofluorescence, and in vitro neutralization tests using antibodies to the SBP4 peptide. This characterization revealed that the Bcsbp4 gene encodes for a 30.58 kDa protein conserved among B. caballi isolates from the USA and Egypt.
- Further analysis revealed the protein BcSBP4 has a signal peptide but lacks additional transmembrane domains, suggesting it may act as an antigen. The protein’s expression in blood stages of B. caballi was confirmed using antibodies against synthetic peptides predicted by in silico analysis.
- In vitro neutralization tests showed a marginal but statistically significant inhibitory effect on B. caballi merozoite infectivity in horse red blood cells.
- Antigenic testing of the BcSBP4 protein involved using an indirect ELISA (iELISA) with sera from 18 field animals. The test showed positive signals in eight B. caballi-infected equids, but not in ten negative controls.
Conclusions and implications
- The findings of this study suggest that the newly identified Bcsbp4 gene is expressed in B. caballi’s blood stages, and BcSBP4 protein is a potential antigen candidate for equine babesiosis.
- A test based on the BcSBP4 protein could potentially improve the detection of B. caballi infection in horses in tropical and subtropical regions worldwide, thereby helping to combat equine babesiosis, a serious economic threat to the equine industry.
- However, further validation with larger sample sizes and diverse geographical regions is likely needed to confirm the efficacy of BcSBP4 as a diagnostic tool or viable vaccine target.
Cite This Article
APA
Mahmoud MS, Kandil OM, Abu El-Ezz NT, Hendawy SHM, Elsawy BSM, Knowles DP, Bastos RG, Kappmeyer LS, Laughery JM, Alzan HF, Suarez CE.
(2020).
Identification and antigenicity of the Babesia caballi spherical body protein 4 (SBP4).
Parasit Vectors, 13(1), 369.
https://doi.org/10.1186/s13071-020-04241-9 Publication
Researcher Affiliations
- Parasitology and Animal Diseases Department, National Research Center, Dokki, Giza, Egypt.
- Parasitology and Animal Diseases Department, National Research Center, Dokki, Giza, Egypt.
- Parasitology and Animal Diseases Department, National Research Center, Dokki, Giza, Egypt.
- Parasitology and Animal Diseases Department, National Research Center, Dokki, Giza, Egypt.
- Parasitology and Animal Diseases Department, National Research Center, Dokki, Giza, Egypt.
- Department of Veterinary Microbiology and Pathology, College of Veterinary Medicine, Washington State University, Pullman, WA, USA.
- Department of Veterinary Microbiology and Pathology, College of Veterinary Medicine, Washington State University, Pullman, WA, USA.
- Animal Disease Research Unit, United States Department of Agricultural-Agricultural Research Service, Pullman, WA, USA.
- Department of Veterinary Microbiology and Pathology, College of Veterinary Medicine, Washington State University, Pullman, WA, USA.
- Parasitology and Animal Diseases Department, National Research Center, Dokki, Giza, Egypt. heba.alzan@wsu.edu.
- Department of Veterinary Microbiology and Pathology, College of Veterinary Medicine, Washington State University, Pullman, WA, USA. heba.alzan@wsu.edu.
- Department of Veterinary Microbiology and Pathology, College of Veterinary Medicine, Washington State University, Pullman, WA, USA.
- Animal Disease Research Unit, United States Department of Agricultural-Agricultural Research Service, Pullman, WA, USA.
MeSH Terms
- Animals
- Antibodies, Protozoan / blood
- Babesia / cytology
- Babesia / genetics
- Babesia / immunology
- Babesia / metabolism
- Babesiosis / blood
- Babesiosis / diagnosis
- Genes, Protozoan
- Horse Diseases / diagnosis
- Horses / parasitology
- Phylogeny
- Protozoan Proteins / immunology
- Protozoan Proteins / metabolism
- Serologic Tests / methods
Conflict of Interest Statement
The authors declare that they have no competing interests.
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Citations
This article has been cited 3 times.- Liu Q, Guan XA, Li DF, Zheng YX, Wang S, Xuan XN, Zhao JL, He L. Babesia gibsoni Whole-Genome Sequencing, Assembling, Annotation, and Comparative Analysis.. Microbiol Spectr 2023 Aug 17;11(4):e0072123.
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