Evaluation of Immunogenicity and Efficacy of Fasciola hepatica Tetraspanin 2 (TSP2) Fused to E. coli Heat-Labile Enterotoxin B Subunit LTB Adjuvant Following Intranasal Vaccination of Cattle.

Gemma Zerna,Hayley Toet,Travis Beddoe,Terry W Spithill,Vignesh A Rathinasamy,Glenn Anderson,Robert Dempster

doi:10.3390/vaccines9111213

Abstract

Fasciolosis, caused by the liver flukes Fasciola hepatica and F. gigantica, is an economically important and globally distributed zoonotic disease. Liver fluke infections in livestock cause significant losses in production and are of particular concern to regions where drug resistance is emerging. Antigens of the F. hepatica surface tegument represent promising vaccine candidates for controlling this disease. Tetraspanins are integral tegumental antigens that have shown partial protection as vaccine candidates against other trematode species. The Escherichia coli heat-labile enterotoxin’s B subunit (LTB) is a potent mucosal adjuvant capable of inducing an immune response to fused antigens. This study investigates the potential of F. hepatica tetraspanin 2 extracellular loop 2 (rFhTSP2) as a protective vaccine antigen and determines if fusion of FhTSP2 to LTB can enhance protection in cattle. Cattle were immunised subcutaneously with rFhTSP2 mixed in the Freund’s adjuvant and intranasally with rLTB-FhTSP2 in saline, accounting for equal molar ratios of tetraspanin in both groups. Vaccination with rFhTSP2 stimulated a strong specific serum IgG response, whereas there was no significant serum IgG response following rLTB-FhTSP2 intranasal vaccination. There was no substantial antigen specific serum IgA generated in all groups across the trial. Contrastingly, after the fluke challenge, a rise in antigen specific saliva IgA was observed in both vaccination groups on Day 42, with the rLTB-FhTSP2 vaccination group showing significant mucosal IgA production at Day 84. However, neither vaccine group showed a significant reduction of fluke burden nor faecal egg output. These results suggest that intranasal vaccination with rLTB-FhTSP2 does elicit a humoral mucosal response but further work is needed to evaluate if mucosal delivery of liver fluke antigens fused to LTB is a viable vaccine strategy.

Highlights

Fasciolosis is a globally distributed zoonotic disease caused by the liver flukes Fasciola hepatica and F. gigantica, which infect a wide range of mammals including domestic ruminants
The DNA sequences encoding labile enterotoxin’s B subunit (LTB)-FhTSP2 and FhTSP2 were cloned into the pPICZαA vector, and recombinant rLTB-FhTSP2 and rFhTSP2 were expressed as secreted proteins from P. pastoris to act as vaccine antigens
Both rLTB-FhTSP2 and rFhTSP2 were successfully expressed after methanol induction and purified from X33 P. pastoris culture supernatants as shown by SDS-PAGE (Figure 1)

Summary

Introduction

Fasciolosis is a globally distributed zoonotic disease caused by the liver flukes Fasciola hepatica and F. gigantica, which infect a wide range of mammals including domestic ruminants. Current vaccine candidates predominantly sit at the host–parasite interface with fluke excretory/secretory (E/S) molecules representing the lead F. hepatica vaccine candidates. These candidates, such as glutathione S-transferase (GST) and cathepsin L1 (CatL) protease, are yet to achieve substantial and reproducible protection with mean efficacies of 43% and 48% in cattle, respectively [10,11]. The tegument or surface tissue layer of flukes has direct contact with the host immune system and could contain antigens that stimulate the previously described potential immune-mediated killing mechanism, antibody dependent cellular cytotoxicity (ADCC), that is observed in rats, sheep, and cattle [9,12,13]

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