Prediction, mapping and validation of tick glutathione S-transferase B-cell epitopes
Introduction
Tick control methods have long relied on the use of acaricides. Nevertheless, numerous methods of tick control have been suggested and explored, among which vaccinating cattle against ticks is considered one of the most sustainable alternatives (de la Fuente and Contreras, 2015; Ghosh et al., 2007; Manjunathachar et al., 2014). Indeed, remarkable progress has been made toward identifying tick vaccine antigens (de la Fuente and Kocan, 2006; Merino et al., 2013; Nuttall et al., 2006; Valle and Guerrero, 2018). Strikingly, however, only a few antigens are reported to induce partial protection against multiple tick species (de la Fuente et al., 2013; Kumar et al., 2017; Ndawula et al., 2019; Parizi et al., 2011; Sabadin et al., 2017; Trimnell et al., 2005). For example, recombinant Bm86 (a Rhipicephalus microplus protein) (Rand et al., 1989) was shown to induce a high protection against R. microplus strains from Australia (Willadsen et al., 1995) and Cuba (Valle et al., 2004), and a lower protection against R. microplus strains from South America (Andreotti, 2006; García-García et al., 2000). The variation in protection could be attributed to differences in Bm86 amino acid sequences among different R. microplus populations (Freeman et al., 2010; García-García et al., 1999). Furthermore, phylogenetic differences within the populations of R. microplus were determined (Burger et al., 2014). Nevertheless, Bm86 was shown to induce cross-protection against Rhipicephalus annulatus (Fragoso et al., 1998), Rhipicephalus decoloratus, Hyalomma anatolicum and Hyalomma dromedarii, but not against Rhipicephalus appendiculatus and Amblyomma variegatum (De Vos et al., 2001).
Considering that multiple tick species are present in most parts of the world, it is vital to develop vaccines that can induce cross-protection against different ticks. However, based on anti-tick vaccination reports, for instance using recombinant Bm86, it seems unlikely that single-antigen tick vaccines will induce a high cross-protection against multiple tick species. Therefore, constituting cocktail vaccines present a possibility to enhance the protection range and the efficacy of anti-tick vaccines. However, how to select antigens for constituting efficacious cocktail tick vaccines is still a challenging question to researchers in the field, due to antigenic competition (Shaffer et al., 2016; Taussig et al., 1973) and the amount of non-protective epitopes exposed to the immune system (Gershoni et al., 2007; Vyas et al., 2008). Epitope-based vaccines, by contrast, do not trigger undesirable immune responses, can induce a high specific immune response and, most importantly, they could induce longer immune protection (Childs et al., 2015; Gershoni et al., 2007; Vyas et al., 2008). The concept of constituting epitope-based vaccines has been demonstrated toward tick control (Aguirre et al., 2016; Patarroyo et al., 2002). Therefore, it is plausible that replacing conventional or whole antigen-based vaccines by an epitope-based approach could enhance the potency of cocktail anti-tick vaccines. The fundamental question, however, remains how to identify and locate epitopes among the sequences of the conventional or whole antigen-based anti-tick vaccine antigens.
There are numerous methods for identifying B-cell epitopes (Ahmad et al., 2016; Gershoni et al., 2007), of which location based on protein structure is regarded as the gold standard (Gershoni et al., 2007); however, x-ray crystallographic data on the structure of tick antigens remains scanty. Nevertheless, the potential of using computer-based tools as an alternative approach to locate epitopes has been exploited (Soria-Guerra et al., 2015). Compared to crystallography, epitope prediction using in silico algorithms is cheaper, quicker and readily applicable. Indeed, numerous in silico epitope prediction tools have been reported (Potocnakova et al., 2016; Sun et al., 2019), of which the linear prediction tools are more commonly used. It is suggested that most of the sequence-based predicted epitope peptides constitute the conformational-based predicted epitopes (Van Regenmortel, 1996), but incorporate structural information on epitope prediction may include epitopes that are discontinuous. Therefore, predictions based on protein structure and linear peptide sequence are likely to give more accurate and reliable results (Assis et al., 2014).
In this work, a combination of three sequence- and conformation-based epitope prediction tools were used, which were selected among the most accurate available (Potocnakova et al., 2016; Sun et al., 2019). Additionally, CBTOPE (Ansari and Raghava, 2010) and Scratch (Cheng et al., 2005) were specifically selected to complement the structural and conformation epitope prediction tools.
Previous work by our group has demonstrated that glutathione S-transferase (GST) is a suitable candidate for a cocktail tick vaccine, inducing non-homologous cross-reaction against different tick species (Ndawula et al., 2019). Therefore, in the present study we performed in silico prediction and mapping of conserved epitopes within the GST sequence of R. appendiculatus (GST-Ra), R. decoloratus (GST-Rd), Haemaphysalis longicornis (GST-Hl), A. variegatum (GST-Av) and R. microplus (GST-Rm). Moreover, the potential epitope peptides were synthesized and screened by immunological assay using anti-GST and anti-peptide sera. The predictions can partially explain the cross-reaction phenomenon observed. Finally, the study illustrates an approach to select epitopes toward developing epitope-based tick vaccine antigens.
Section snippets
Ethics statement
Rabbits used in the experiments were housed at Faculdade de Veterinária, Universidade Federal do Rio Grande do Sul facilities. This research was conducted in agreement with the Norms for Animal Experimentation Ethics Committee of Universidade Federal do Rio Grande do Sul (process number 38748).
GST protein sequences
The GST sequences used herein were deduced from the GSTs of R. appendiculatus (MK133338), R. decoloratus (MK133339), A. variegatum (MK133337), R. microplus (AF366931.1), H. longicornis (AY298731.1), R.
Results and discussion
Until now, vaccinating cattle against ticks is regarded as one of the most promising tick-control alternative approaches to replace the use of acaricides. Indeed, researchers have identified numerous antigens (de la Fuente and Kocan, 2006; Merino et al., 2013; Ndawula et al., 2019; Nuttall et al., 2006; Schetters et al., 2016; Valle and Guerrero, 2018), but none has so far matched the success exhibited with Bm86 under field conditions (de la Fuente et al., 2016, 2007, 1999, 1998). Combining
Conclusion
Ultimately, the data presented here support the hypothesis that GST-Ra, GST-Rd, GST-Hl, GST-Rm and GST-Av contain conserved epitopes, which may explain the cross-reaction among tick GSTs (Ndawula et al., 2019). Additionally, the study brings forward an approach to select B-cell epitopes. Most importantly, the epitopes predicted herein were found to be immunogenic. Based on these findings, our hypothesis is that the identified epitopes could be used to constitute epitope-based antigen tick
CRediT authorship contribution statement
Charles Ndawula: Conceptualization, Methodology, Formal analysis, Visualization, Writing - original draft, Writing - review & editing. Marina Amaral Xavier: Methodology, Formal analysis, Visualization, Writing - original draft, Writing - review & editing. Bianca Villavicencio: Conceptualization, Methodology, Formal analysis, Visualization, Writing - original draft, Writing - review & editing. Fernanda Cortez Lopes: Conceptualization, Methodology, Formal analysis, Visualization, Writing -
Declaration of Competing Interest
The authors certify that they have no affiliations with, or involvement in any organization or entity with any financial interest in the subject matter or materials discussed in this manuscript.
Acknowledgements
This study was financed in part by Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES): # Procad 88881.068421/2014-01; PGCI 23038.005296/2014-37; 88881.153222/2017-01; Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq): Grant # 465678/2014-9; 405763/2018-2, 302360/2018-2 and 441092/2016-0; CNPq-INCT and TWAS. Ndawula Charles Jr is a student under the TWAS-CNPq PhD Fellowship.
References (95)
- et al.
B-cell epitope mapping for the design of vaccines and effective diagnostics
Trials Vaccinol.
(2016) - et al.
Basic local alignment search tool
J. Mol. Biol.
(1990) - et al.
Phylogenetic analysis of mitochondrial genome sequences indicates that the cattle tick, Rhipicephalus (Boophilus) microplus, contains a cryptic species
Mol. Phylogenet. Evol.
(2014) - et al.
Design of the ATAQ peptide and its evaluation as an immunogen to develop a Rhipicephalus vaccine
Vet. Parasitol.
(2016) - et al.
Field studies and cost-effectiveness analysis of vaccination with Gavac(TM) against the cattle tick Boophilus microplus
Vaccine
(1998) - et al.
Vaccination against ticks (Boophilus spp.): the experience with the Bm86-based vaccine Gavac(TM)
Genet. Anal. - Biomol. Eng.
(1999) - et al.
Consensus motifs and peptide ligands of mhc class I molecules
Semin. Immunol.
(1993) - et al.
Protection against Boophilus annulatus infestations in cattle vaccinated with the B. Microplus Bm86-containing vaccine Gavac
Vaccine
(1998) - et al.
Control of ticks resistant to immunization with Bm86 in cattle vaccinated with the recombinant antigen Bm95 isolated from the cattle tick, Boophilus microplus
Vaccine
(2000) - et al.
Human Mu-class glutathione S-transferases present in liver, skeletal muscle and testicular tissue
Biochim. Biophys. Acta (BBA)/Protein Struct. Mol.
(1993)
Two serine protease inhibitors (serpins) that induce a bovine protective immune response against Rhipicephalus appendiculatus ticks
Vaccine
The development of veterinary vaccines: a review of traditional methods and modern biotechnology approaches
Biotechnol. Res. Innov.
Functional characterization of candidate antigens of Hyalomma anatolicum and evaluation of its cross-protective efficacy against Rhipicephalus microplus
Vaccine
The Aedes aegypti glutathione transferase family
Insect Biochem. Mol. Biol.
Economic importance of ticks and their effective control strategies
Asian Pacific J. Trop. Dis.
Control of multiple arthropod vector infestations with subolesin/akirin vaccines
Vaccine
Constituting a glutathione S-transferase-cocktail vaccine against tick infestation
Vaccine
In silico analysis of glutathione S-transferase supergene family revealed hitherto unreported insect specific δ- And ε-GSTs and mammalian specific μ-GSTs in Ixodes scapularis (Acari: Ixodidae)
Comput. Biol. Chem.
Cross immunity with Haemaphysalis longicornis glutathione S-transferase reduces an experimental Rhipicephalus (Boophilus) microplus infestation
Exp. Parasitol.
Multi-antigenic vaccine against the cattle tick Rhipicephalus (Boophilus) microplus: a field evaluation
Vaccine
Immunization of cattle with synthetic peptides derived from the Boophilus microplus gut protein (Bm86)
Vet. Immunol. Immunopathol.
Distinctive structure of the human GSTM3 gene - Inverted orientation relative to the mu class glutathione transferase gene cluster
Arch. Biochem. Biophys.
Mapping protective epitopes in the tick and mosquito subolesin ortholog proteins
Vaccine
Effect of recombinant glutathione S-transferase as vaccine antigen against Rhipicephalus appendiculatus and Rhipicephalus sanguineus infestation
Vaccine
An overview of bioinformatics tools for epitope prediction: implications on vaccine development
J. Biomed. Inform.
Antigenic competition and genetic control of the immune response. A hypothesis for intramolecular competition
Cell. Immunol.
A cross-reactive tick cement antigen is a candidate broad-spectrum tick vaccine
Vaccine
Mapping epitope structure and activity: from one-dimensional prediction to four-dimensional description of antigenic specificity
Methods A Companion to Methods Enzymol.
Performance of two Bm86 antigen vaccin formulation against tick using crossbreed bovines in stall test
Rev. Bras. Parasitol. Veterinária
Identification of conformational B-cell epitopes in an antigen from its primary sequence
Immunome Res.
In silico models for B-cell epitope recognition and signaling
Functional evolution of subolesin/akirin
Front. Physiol.
A novel combined scientific and artistic approach for the advanced characterization of interactomes: the akirin/subolesin model
Vaccines
B-cell epitopes of antigenic proteins in Leishmania infantum: an in silico analysis
Parasite Immunol.
Continuous and discontinuous protein antigenic determinants
Nature
The antigenic structure of proteins: a reappraisal
Annu. Rev. Immunol.
The protein data bank (www.rcsb.org)
Nucleic Acids Res.
Pcleavage: an SVM based method for prediction of constitutive proteasome and immunoproteasome cleavage sites in antigenic sequences
Nucleic Acids Res.
Analysis of Bm86 conserved epitopes: is a global vaccine against cattle tick Rhipicephalus microplus possible?
Rev. Bras. Parasitol. Vet.
Conjugation of peptides to carrier proteins via glutaraldehyde
Prediction of linear B-cell epitopes using amino acid pair antigenicity scale
Amino Acids
SCRATCH: a protein structure and structural feature prediction server
Nucleic Acids Res.
Trade-offs in antibody repertoires to complex antigens
Philos. Trans. R. Soc. B Biol. Sci.
From immunome to vaccine: Epitope mapping and vaccine design tools
Novartis Found. Symp.
Tick vaccines: current status and future directions
Expert Rev. Vaccines
Strategies for development of vaccines for control of ixodid tick species
Parasite Immunol.
A ten-year review of commercial vaccine performance for control of tick infestations on cattle
Anim. Health Res. Rev.
Cited by (11)
Insight Into the Dynamics of the Ixodes ricinus Nymphal Midgut Proteome
2023, Molecular and Cellular ProteomicsThe current strategies and underlying mechanisms in the control of the vector tick, Haemaphysalis longicornis: Implications for future integrated management
2022, Ticks and Tick-borne DiseasesCitation Excerpt :Therefore, greater emphasis should be placed on developing vaccines effective in different hosts, in addition to multiple protection against multiple tick species because H. longicornis may share hosts with other tick species (de la Fuente and Contreras, 2015). However, some vaccination studies have identified a recombinant GST of H. longicornis protein (rGST-Hl) capable of inducing immunoprotection against more than one tick species (Parizi et al., 2011; Sabadin et al., 2017; Ndawula et al., 2020). For instance, rGST-Hl induced protection level of about 67% overall efficacy ratio against the engorged adults of Rhipicephalus appendiculatus infestation in rabbit (Sabadin et al., 2017), and about 57% overall efficacy ratio against the engorged adults of R. microplus infestation in cattle (Parizi et al., 2011).
Prediction and validation of cross-protective candidate antigen of Hyalomma asiaticum cathepsin L between H. asiaticum and H. anatolicum
2022, Experimental and Applied Acarology
- 1
These authors contributed equally to the work.