Design and Production of a Recombinant Hybrid Toxin to Raise Protective Antibodies Against Loxosceles Spider Venom.

Ana Moura-Da-Silva,Monica Colombini,Katia Barbaro,Lhiri Shimokawa-Falcão,Paula Calabria,Eliana Faquim-Mauro,Geraldo Magalhaes

doi:10.3390/toxins11020108

Abstract

Human accidents with spiders of the genus Loxosceles are an important health problem affecting thousands of people worldwide. Patients evolve to severe local injuries and, in many cases, to systemic disturbances as acute renal failure, in which cases antivenoms are considered to be the most effective treatment. However, for antivenom production, the extraction of the venom used in the immunization process is laborious and the yield is very low. Thus, many groups have been exploring the use of recombinant Loxosceles toxins, particularly phospholipases D (PLDs), to produce the antivenom. Nonetheless, some important venom activities are not neutralized by anti-PLD antibodies. Astacin-like metalloproteases (ALMPs) are the second most expressed toxin acting on the extracellular matrix, indicating the importance of its inclusion in the antigen’s formulation to provide a better antivenom. Here we show the construction of a hybrid recombinant immunogen, called LgRec1ALP1, composed of hydrophilic regions of the PLD and the ALMP toxins from Loxosceles gaucho. Although the LgRec1ALP1 was expressed as inclusion bodies, it resulted in good yields and it was effective to produce neutralizing antibodies in mice. The antiserum neutralized fibrinogenolytic, platelet aggregation and dermonecrotic activities elicited by L. gaucho, L. laeta, and L. intermedia venoms, indicating that the hybrid recombinant antigen may be a valuable source for the production of protective antibodies against Loxosceles ssp. venoms. In addition, the hybrid recombinant toxin approach may enrich and expand the alternative antigens for antisera production for other venoms.

Highlights

In view of the wide geographical distribution, the large number of individuals affected and the evolution of the clinical picture, the accidents with spiders of the genus Loxosceles, denominated loxoscelism, have received great attention from public health [1,2,3]
In order to know the main toxin transcripts present in the venom gland of Loxosceles gaucho, a transcriptomic approach was performed and the analysis showed that 22.36% of all sequences gave match to toxins already described in the database
Among the metalloprotease’s transcripts with identity greater than 95%, the largest group contained 45% of all metalloproteinase transcripts, and a sequence called LgALP1 was selected from this group to be part of the hybrid immunogen

Summary

Introduction

In view of the wide geographical distribution, the large number of individuals affected and the evolution of the clinical picture, the accidents with spiders of the genus Loxosceles, denominated loxoscelism, have received great attention from public health [1,2,3]. The loxoscelism is associated with a number of clinical symptoms including. Cutaneous loxoscelism may progress to systemic manifestations (cutaneous-visceral loxoscelism) and the symptoms of this clinical condition usually begin 24 h after the spider bites, which is characterized by anemia, jaundice, intravascular hemolysis, platelet aggregation, and, in more severe cases, renal failure [8]. Studies have shown that phospholipases D (PLDs) are the most abundant toxins able to elicit a cascade of adverse pharmacological events such as inflammation [13,17] dermonecrosis [11,13,18,19,20,21], platelet aggregation [21,22,23], hemolysis [13,23,24], and nephrotoxicity [25,26], among others

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Conclusion