The toxicity of surface waters and interstitial waters from sediments were determined at six study sites in Todos os Santos Bay, Bahia, Brazil, to evaluate the possibility of chronic environmental impact induced by 40 years of exposure to the local petroleum industry. Samples collected from four sites associated with the extraction, transportation and refinement of petroleum, and from two control sites, were tested at seven three-month intervals. Toxicological assays using acute mortality of brine shrimp (Artemia salina) nauplii and chronic abnormalities of sea urchin (Echinometra lucunter) and mangrove oyster (Crassostrea rhizophorae) larvae were employed. Friedman non-parametric analyses of variance integrated seasonal variations in species response patterns and revealed significant differences among the study sites. Ranging the among-site variations for each organism in each sampling period, on a scale from 0.00 (minimum response) to 1.00 (maximum response), permitted the calculation of a single mean value for each species and the ordination of the sites on a qualitative scale of relative impact. Although the ordinations varied with species, the reduction of three species response patterns to a common relative scale also permitted their integration into a single multispecies ordination of the study sites. A cluster analysis of the six sites and two aquatic substrates, based on their toxicity to all three species, illustrated the similarities and differences between locations. Interstitial waters were more toxic, revealing an integrated ordination of Station 6 ≤ Station 5 ≤ (Station 3 = Station2) ≤ (Station 4 = Station 1). The ordination based on surface waters was Station 6 ≤ Station 5 ≤ (Station 2 = Station 1 = Station 3) < Station 4. In combination, the three procedures served efficiently for the description and inferential testing of the multispecies responses and, complemented with additional data on species diversity and chemical contamination of the sediment, confirmed the existence of chronic impact within the study area.