Currently, little is known about the seasonal variation of corticosterone (CORT) levels, either baseline or stress response, within freshwater turtles. We conducted a seasonal CORT study with a species of freshwater turtle, Graptemys flavimaculata (Yellow-blotched Sawback; family Emydidae), that is endemic to the Pascagoula River system of southeastern Mississippi. Graptemys flavimaculata is commonly observed while basking on deadwood snags, with us using basking traps and dip nets as active capture methods. We caught both male (n  =  60) and female (n  =  49) turtles during the months of April–October during 2007 and 2008. Immediately after capture, we collected an initial blood sample, confined the turtle for 35 minutes, and then took a second blood sample. Competitive binding radioimmunoassays were done to determine CORT levels. Time zero CORT levels for both sexes were generally lower than previously reported levels for other turtle species, likely due to the trapping methods used. By time 35 min, CORT levels were significantly elevated relative to initial levels. We did not detect seasonal differences in time zero CORT levels for males or females. However, seasonal differences in time 35 CORT levels for males were detected, with higher levels observed during summer months (July, August) relative to spring (April) and fall (October) months. Seasonal differences were also detected in time 35 CORT levels of females, with lower levels observed during July (nesting season) relative to September. This study provides novel information and insight into the differences in seasonal physiological demands for this species, and aquatic turtle species in general, while also providing probable connections of physiology to seasonal life history events.