Seismic waterguns produce acoustic energy that has the potential to affect aquatic organisms. This study examined the effects of exposure to a 1,966.4 cm³ watergun array (two units) on the abundance, behavior, and spatial distribution of Asian carp (Hypophthalmichthys spp.) and native fish species (i.e., bigmouth buffalo, Ictiobus cyprinellus; black buffalo, Ictiobus niger; channel catfish, Ictalurus punctatus; flathead catfish, Pylodictis olivaris; freshwater drum, Aplodinotus grunniens; gizzard shad, Dorosoma cepedianum; smallmouth buffalo, Ictiobus bubalus). Waterguns were deployed, approximately centered and perpendicular to the shoreline, in a channel that connects the Illinois River to a set of backwater gravel pits. To evaluate the effect of waterguns, six mobile side-looking split-beam hydroacoustic surveys were conducted at fixed times over a period of 12 days, during and between replicated watergun firing periods. Acoustic data collected over a five day period before placement of the waterguns were used as a control. Proximity to the waterguns of the first detected Asian carp while the watergun array was operational (52.95 ± 7.41 m, mean ± SE) was not different relative to when turned off (36.53 ± 6.08). In contrast, the closest detected native fish were farther from the waterguns while firing (32.38 ± 3.32 m) compared to when they were off (15.04 ± 1.6 m). The waterguns did not affect the abundance of acoustically detected Asian carp within the survey area, but the abundance of native fish was reduced during the firing treatment (115.82 ± 10.19) compared to both the control (203.46 ± 12.12) and off treatment (175.84 ± 12.16). The cumulative abundance of Asian carp relative to distance from the waterguns, examined at 50 m intervals up to 480 m bi-directionally away from the waterguns, was sporadically different (i.e., reduced abundance while the waterguns were firing) between treatments (i.e., at the 250, 300, 350, and 480 m intervals). In contrast, there was a significantly lower cumulative abundance of native fish during the firing treatment at every distance interval tested. The spatial distributions, relative to distance away from the waterguns, of both fish groups were affected by the firing of the waterguns. During active operation of the waterguns, there were proportionally less targets of both fish groups detected within a 50 m radius. While firing, there were proportionally more native fish farther than 400 m away on either side of the waterguns, and a larger proportion of Asian carp farther than 400 m from the waterguns on the northern side of the channel (i.e., closer to the Illinois River main channel). Asian carp and native fish were both more likely to be detected closer to the HMS pits when the waterguns were turned off. The watergun array did not influence the directional movement of detected Asian carp. In contrast, native fish were less likely to swim towards the IL River while the waterguns were firing versus not firing. The size distributions of both Asian carp and native fish species that remained in the survey area did not change across watergun treatments. It is concluded that the seismic watergun array examined in this study had a greater impact on native fish species than on Asian carp. Due to the lowered abundance, but not complete suppression of fish, it is suggested that the watergun array (as tested) is not suitable as a barrier for areas where absolute exclusion of fish is desired. The results from the spatial distribution analyses suggest the tested watergun array could be more suitable as a herding tool to facilitate removals of large fish aggregations. Caution should be taken to the extrapolation to other fish species and to fish exposed to seismic waterguns in different environments (i.e., deeper water, reduced shoreline interaction) or exposure to a larger array of waterguns.