The physical habitat simulation sub-routine of the Instream Flow Incremental Methodology (IFIM) uses hydraulic modeling and suitability indices of target fish species to predict how differences in-stream flows affect the microhabitat occupation by fish species. This habitat modelling approach was adopted to assess the ecological effects of running flows on three neotropical fish species of different orders (Bryconamericus ornaticeps, Ancistrus multispinis and Geophagus brasiliensis).The study encompassed two reaches of an Atlantic Forest stream in Southeastern Brazil where topographic and hydraulic (depth, velocity and type of substrate) characteristics were measured to implement one-dimensional hydraulic simulation. Sub aquatic observation of fish was performed to collect data on microhabitat use and these data were used to develop habitat suitability curves that were used in the habitat simulation to obtain the habitat suitability index (HSI) and weighted usable area (WUA) versus flow curves. Upon these curves minimum and optimum environmental flows for the target fish species were proposed. Bryconamericus ornaticeps and A. multispinis selected microhabitats around 0.6 m depth, whereas G. brasiliensis showed a wider suitable range (0.35-0.9 m). All the three species were mainly observed in microhabitat with low flow velocity (0.1 m/s). Bryconamericus ornaticeps selected more frequently coarse substrate (e.g. boulders) but it appeared also over sandy substrate, whereas A. multispinis and G. brasiliensis selected preferably boulders. The range of 0.65-0.85 m 3 /s was found as the optimum to meet the needs of the three fish species. Our results agree with the necessary objective information to perform grounded management actions in the frame of a management program aiming at ecosystem conservation. Thereby it can be considered a successful pilot study in environmental flow assessment in an Atlantic Forest stream of Brazil.A sub-rotina simulação de habitat fĂsico da metodologia incremental -IFIM usa variĂĄveis hidrĂĄulicas e os Ăndices de aptidĂŁo das espĂ©cies alvo para predizer como diferentes vazĂ”es afetam a ocupação de microhabitat por espĂ©cies de peixes. Esta abordagem de simulação de hĂĄbitat foi adotada para avaliar os efeitos ecolĂłgicos sobre trĂȘs espĂ©cies de peixes tropicais de diferentes ordens (Bryconamericus ornaticeps, Ancistrus multispinis e Geophagus brasiliensis) em dois trechos de um riacho de Mata AtlĂąntica do sudeste do Brasil, durante a estação chuvosa de 2013 e a estação seca de 2014. CaracterĂsticas topogrĂĄficas e hidrĂĄulicas (profundidade, velocidade e tipo de substrato) foram mensuradas para implementação da simulação de habitat em uma dimensĂŁo; observaçÔes subaquĂĄticas dos peixes foram realizadas para avaliação do uso do microhabitat. As curvas de aptidĂŁo de habitat de cada variĂĄvel hidrĂĄulica foram utilizadas na simulação de habitat para obtenção das curvas de aptidĂŁo composta (HSI) e a ĂĄrea Ăștil ponderada (WUA) versus fluxo, e propor vazĂ”es ambientais mĂnimas e ideais para as espĂ©cies de peix...