Perfluoroalkyl carboxylic acids (PFCAs) are persistent contaminants increasingly subjected to regulatory restrictions. To date, their effects on terrestrial plants remain poorly investigated. To address these knowledge gaps, a comparative assessment was conducted to identify the most sensitive plant species and the most responsive early-growth endpoints. Five PFCAs were selected according to their carbon-chain length (from 3 to 8 C-atoms). Seven plant species were exposed to a wide range of concentrations (from 0.01 up to 100 µg kg−1). Germination and root elongation were evaluated as developmental endpoints to assess both acute and sublethal effects. Across species, germination exhibited weak responses, whereas root elongation appeared to be the most sensitive screening parameter, displaying divergent species-specific patterns. Notably, Sinapis alba and Cucumis sativus emerged as the most responsive species, although they exhibited opposite responses. While mustard exhibited low-dose root stimulation, cucumber showed root inhibition. Interestingly, species within the same family (Brassicaceae and Cucurbitaceae) showed contrasting sensitivity, suggesting that PFCA phytotoxicity is species-specific rather than driven by taxonomic relatedness. This divergent pattern may be linked to distinct morpho-physiological traits of the root system architecture. The mechanisms underpinning the two opposite morphological effects remain to be investigated. Disruption of hormonal processes involved in development of RSA may represent a possible explanation underlying the contrasting root responses observed in this study.