Fascioliasis is a zoonotic infection linked to significant economic losses in the livestock industry. Infection prevalence and estimated financial burden vary across locations owing to different diagnostic tests used. The accuracy of liver condemnation to estimate the prevalence and costs of fascioliasis has seldom been evaluated. We performed a pilot study to determine the prevalence and burden of Fasciola hepatica infection among cattle slaughtered at the municipal abattoir in the Anta province of the Cusco highlands in Peru. We compared liver condemnation with bile microscopy for the diagnosis of infection and prediction of carcass weight. Data were collected from 2009 slaughtered cattle for 1 year. The overall prevalence of Fasciola infection by bile microscopy was 62.5% (1247/2009). A higher prevalence was observed after the rainy season from March to August than from September to February (p < 0.01). Fascioliasis prevalence during the first 6 months was 77.4% (714/923), combining the results of condemnation and microscopy. Bile microscopy diagnosed more infections than liver condemnation (62.7% (579/923) versus 55.4% (511/923), McNemar test p < 0.01). The agreement of the bile microscopy testing with liver condemnation was fair (κ = 0.247). Animal age, gender, breed, and liver condemnation predicted carcass weight [F (df 4, 704) = 61.1, p < 0.001]. Liver condemnation and bile microscopy are complementary tools for evaluation of the prevalence and burden of fascioliasis in livestock. Large scale studies are warranted to confirm our results.