Toxoplasma gondii infection is relevant to animal production, food safety, and public health, but current epidemiological data for small ruminants in northeastern Portugal are limited. This cross-sectional study estimated seroprevalence and investigated factors associated with seropositivity in 2678 animals from 149 flocks sampled between May 2022 and December 2024. Serum samples were tested using an indirect enzyme-linked immunosorbent assay, and associated factors were evaluated using mixed-effects logistic regression with flock as a random effect. The overall true seroprevalence was estimated at 21.5% (apparent seroprevalence: 21.6%). At the flock level, 83.9% (125/149) of the sampled flocks were seropositive. True seroprevalence was higher in sheep than in goats (23.4% versus 14.4%), and seropositivity was detected in all surveyed municipalities. In the multivariable model, higher odds of seropositivity were observed in sheep (adjusted odds ratio 1.74), adult animals (2.78), non-native breeds (1.84), animals not kept in permanent housing (3.16), and animals from farms with cats (1.69). These findings indicate exposure throughout the surveyed area and associations with both host-related and farm-level characteristics. Strengthening farm biosecurity, limiting contamination of feed and water, and integrating animal health, environmental, and food safety measures may contribute to prevention within a One Health framework.

Q fever, caused by Coxiella burnetii, is a globally distributed zoonosis associated with small ruminants. In Mediterranean regions, where extensive farming systems prevail, understanding environmental and management drivers of infection is essential for public health. This study estimated the seroprevalence and identified multilevel factors associated with C. burnetii exposure among small ruminants in Trás-os-Montes, Portugal. A cross-sectional study was conducted across 105 farms (n = 1827 animals; 1596 sheep and 231 goats). Serum samples were analysed using a commercial ELISA. Animal-, herd-, and management-level variables were evaluated using multivariable logistic regression to identify factors associated with seropositivity. Overall apparent seroprevalence was 40.7% (95% CI: 38.5-43.0), with apparent spatial heterogeneity in seroprevalence estimates across municipalities. Older age (≥18 months; aOR = 2.02), larger herd size (≥100 animals; aOR = 1.35), mixed ovine-caprine rearing (aOR = 1.58), and permanent confinement (aOR = 3.88) were associated with higher odds of seropositivity. Furthermore, the introduction of animals without prior health screening (aOR = 5.64; 95% CI: 1.40-51.4) and sharing of drinking troughs (aOR = 1.31) were associated with higher odds of seropositivity, whereas the absence of a maternity pen (aOR = 0.57) and participation in livestock competitions (aOR = 0.71) were associated with lower odds of seropositivity. The high seroprevalence indicates substantial C. burnetii circulation in the study region. Management characteristics, particularly confinement and herd structure, were associated with seropositivity, although these associations should not be interpreted causally given the cross-sectional design. These findings support risk-based surveillance and targeted biosecurity measures within a One Health framework to reduce zoonotic risk.