Reniform nematode (Rotylenchulus reniformis) resistance is being transferred to Gossypium hirsutum from its distant relatives. Reports of fluometuron damage to LONREN lines with nematode resistance from G. longicalyx raised concerns about introducing herbicide sensitivity from other nematode resistance sources. The research objective was to evaluate 15 sources of reniform nematode resistance for their reaction to fluometuron three weeks after planting in a replicated greenhouse trial: two G. herbaceum accessions, four G. arboreum accessions, three G. barbadense accessions, three G. hirsutum accessions, and three G. hirsutum lines with resistance introgressed from G. barbadense (FR-05) or G. longicalyx (LONREN-1 and LONREN-2). The control genotype was G. hirsutum cultivar Deltapine 161 B2RF. Across all herbicide rates tested, mean herbicide injury ratings for G. arboreum accessions were greater than the control, whereas G. barbadense GB 713 and TX 110 were less. Regression analysis of herbicide rates indicated that injury increased linearly with increasing herbicide rate for all accessions, although G. arboreum A2-083 had more injury than the control. Regression analysis of herbicide rates indicated that biomass decreased linearly with increasing herbicide rate for all accessions, although G. barbadense GB 713 and Pima PHY 800 exhibited greater biomass reduction than the control. Across all herbicide rates tested, mean electron transport rates of all G. herbaceum and G. arboreum accessions and G. barbadense Pima PHY 800 were lower than the control. The relationship between herbicide rate and electron transport rate was curvilinear, with similar decreases in electron transport rate in response to increasing herbicide concentration for all lines. Increased sensitivity to fluometuron could be introduced into G. hirsutum through crosses with distantly related species, but with the exception of G. arboreum A2-083, the lines did not respond to the herbicide differently from the control.