Zebrafish Animal Model for the Study of Allergic Reactions in Response to Tick Saliva Biomolecules

Summary

Here, zebrafish (Danio rerio) is used as a model to study allergic reactions and immune responses related to alpha-Gal syndrome (AGS) by evaluating allergic reactions to tick saliva and mammalian meat consumption.

Abstract

Ticks are arthropod vectors that cause disease by pathogen transmission and whose bites could be related to allergic reactions impacting human health worldwide. In some individuals, high levels of immunoglobulin E antibodies against the glycan Galα1-3Galβ1-(3)4GlcNAc-R (α-Gal) have been induced by tick bites. Anaphylactic reactions mediated by glycoproteins and glycolipids containing the glycan α-Gal, present in tick saliva, are related to alpha-Gal syndrome (AGS) or mammalian meat allergy. Zebrafish (Danio rerio) has become a widely used vertebrate model for the study of different pathologies. In this study, zebrafish was used as a model for the study of allergic reactions in response to α-Gal and mammalian meat consumption because, like humans, they do not synthesize this glycan. For this purpose, behavioral patterns and hemorrhagic anaphylactic-type allergic reactions in response to Ixodes ricinus tick saliva and mammalian meat consumption was evaluated. This experimental approach allows the obtention of valid data that support the zebrafish animal model for the study of tick-borne allergies including AGS.

Introduction

Ticks are vectors of pathogens that cause diseases and are also the cause of allergic reactions, affecting the health of humans and animals worldwide^1,2. During tick feeding, biomolecules in tick saliva, especially proteins and lipids, facilitate the feeding of these ectoparasites, avoiding host defenses³. Some saliva biomolecules with glycan Galα1-3Galβ1-(3)4GlcNAc-R (α-Gal) modifications lead to the production of high anti-α-Gal IgE antibody levels after the tick bite, only in some individuals, which is known as α-Gal Syndrome (AGS)⁴. T....

Protocol

All methods described here have been approved by the Ethics Committee on Animal Experimentation of the University of Castilla La Mancha under the study "Evaluation of the immune response to inactivated M. bovis vaccine and challenge with M. marinum in the zebrafish model number PR-2017-05-12."

Ticks were obtained from the laboratory colony, where representative samples of ticks in the colony were tested by PCR for common tick pathogensto confirm the absence of pathogens, and maintained at the Institute of Parasitology, Biology Centre of the Czech Academy of Sciences (IP BC CAS), Czech Republic.All animal experiment....

Results

The protocol presented here is based on several aspects of previously published experiments^27,30 and results performed in our laboratory where the zebrafish model is established and validated for the study of AGS and the immune response to α-Gal because both humans and zebrafish do not synthesize this molecule¹³. This model allows the characterization and evaluation of a variety of allergic reactions as a result of the host response t.......

Discussion

Zebrafish is a cost-effective and easy-to-handle model that also has been a very feasible tool for the study of molecular mechanisms of the immune response, pathogen diseases, novel drug testing, and vaccination and protection against infections^33,34,35. The study on the behavior of zebrafish is useful since previous studies have found that some fish species remain motionless at the bottom of the tank when they are stressed, whi.......

Materials

Name	Company	Catalog Number	Comments
1.5 mL tube	VWR	525-0990
All Prep DNA/RNA	Qiagen	80284
Aquatics facilities
BCA Protein Assay Kit	Thermo Fisher Scientific	23225
Disection set	VWR	631-1279
Dog Food - Red Classic	Acana
ELISA plates-96 well	Thermo Fisher Scientific	10547781
Gala1-3Gal-BSA 3 (α-Gal)	Dextra	NGP0203
iScript Reverse Transcription Supermix	Supermix	1708840
Microliter syringes	Hamilton	7638-01
Plate reader	any
Phosphate buffered saline	Sigma	P4417-50TAB
pilocarpine hydrochloride	Sigma	P6503
Pipette tip P10	VWR	613-0364
Pipette tip P1000	VWR	613-0359
Premium food tropical fish	DAPC
Sponge Animal Holder			Made from scrap foam
Stereomicroscope	any
Thermal Cycler Real-Time PCR	any
Tricaine methanesulphonate (MS-222)	Sigma	E10521