Morphometric variability of Fasciola hepatica eggs among definitive host species: A multivariate approach

Egg morphometry provides valuable information for investigating phenotypic variation and host-associated patterns in Fasciola hepatica. However, comparative studies evaluating egg morphometric variation across multiple definitive host species remain limited. Therefore, the aim of this study was to evaluate morphometric variation in F. hepatica eggs recovered from different definitive host species using univariate, multivariate, and host-level classification approaches. A total of 690 eggs obtained from 60 independent host individuals representing cattle, sheep, goats, horses, mules, donkeys, and guanacos were analyzed using four morphometric variables: length, width, shape ratio, and size. Significant differences among host species were detected for all morphometric variables (Kruskal-Wallis, p < 0.001). Multivariate analyses revealed significant host-associated morphometric differentiation (PERMANOVA, p = 0.001), with egg size and length representing the principal contributors to variation. To account for potential pseudoreplication, complementary host-level analyses were performed using host individuals as the statistical unit, confirming that morphometric differences among host species remained significant. In addition, Linear Discriminant Analysis correctly classified 63.3% of host individuals, demonstrating that egg morphometry retains moderate discriminatory power at the host level. Overall, the results support the existence of host-associated morphometric variation in F. hepatica eggs and are consistent with the marked phenotypic plasticity previously described for this parasite. The inclusion of guanaco-derived samples also provides novel information on egg morphology in a poorly documented South American wild host.