Fasciola's excretory-secretory material comprises chiefly cathepsin B and cathepsin L. These cysteine proteases are proposed as major mediators of parasitism, and are considered targets for vaccination. In order to assess the vaccine efficacy of these enzymes, single and multivalent recombinant protein vaccinations of adult-stage F. hepatica cathepsin L5, metacercarial-stage F. gigantica cathepsin L1 g and juvenile-stage F. hepatica cathepsin B were analysed in rats against F. hepatica challenge infection. The protective efficacy of anti-fluke vaccines was evaluated in terms of parasitological parameters (recovered fluke burden, fluke body size and wet weight) and pathological changes (liver damage score) in rats. The rats vaccinated with recombinant proteins were shown to have significantly fewer and smaller flukes than the control rats. A maximum protection of 83% was seen in the group vaccinated with a combination of cathepsin B and cathepsin L5. © 2010 Elsevier B.V. All rights reserved.