The results of statistical analyses conducted on monthly trawl and seine data collected from a multiyear fisheries-independent monitoring program, between July 2006 and March 2009 on the lower Apalachicola River, FL were used to determine which regions of the river, and what time of year should be further studied to determine if and how freshwater flow alterations affect the nekton. Differences in nekton community structure were clear between each of the 3 predefined habitat regions for shoreline seine catches. Deepwater trawled habitats showed a distinct difference between the upper river channel and both distributary regions, and less pronounced differences between the two distributary regions. The strongest pattern in seasonality of nekton communities coincides with seasonal fluctuations in recruitment of juveniles into the estuary and the periods of greatest salinity differences in the marshgrass-dominated lower distributary region for shoreline and deepwater habitats. Seasonal variations in community structure were evident and mostly likely dominated by recruitment, whereas the response of organisms to fluctuations in salinity may be dictated by their relative position within the lower reaches of the Apalachicola River system. Our results suggest that future studies of the effects of changes in flow on nekton assemblages in the lower Apalachicola River would best be performed during the dry season in the upper and lower portions of the distributary region.