Louisiana's streams provide habitat for a diversity of ecologically and economically important fish species. However, most stream systems in the state have been subject to substantial anthropogenic alterations related to municipal, agricultural, and forest resource development. Extensive reservoir construction has also resulted in altered dendritic connectivity in many drainages, which has been shown to significantly alter the function and biodiversity of streams throughout the U.S. This study explored interactions among stream connectivity, watershed characteristics, stream physicochemistry, and fish assemblage composition and abundance in 21 headwater streams within the Red River basin of Louisiana, with half flowing directly into a reservoir. In 2014 and 2015, fish were sampled by electrofishing and seining, and habitats and physicochemistry were characterized. Six fish species were exclusively found in reservoir streams, three fish species were only found in river tributaries, seven fish species were more abundant in river tributaries, and all richness metrics (total, stream specialist, and generalist) calculated for seined fishes were higher in river tributaries. Notably, Grass Pickerel (Esox americanus) was more abundant in river tributaries. Because of its potential role as an obligate glochidial host for the Louisiana Pearlshell Mussel (Margaritifera hembeli), the influence of stream fragmentation on Grass Pickerel may have significant implications regarding conservation of this endangered mussel. Our results parallel trends observed in similar studies indicating decreased species richness and abundance in streams truncated by reservoirs.