Purely symbiotic and NO3-fed nodulated seedlings of Virgilia oroboides (Bergius) T.M. Salter, Cyclopia maculata (L.) Vent and Podalyria calyptrata Willd. were exposed to biologically effective ultraviolet-B radiation (UV-B) to assess the effects of above- and below-ambient UV-B on growth, symbiotic function and metabolite concentrations. Seedlings were grown outdoors either on tables under ambient or 34 or 66% above-ambient UV-B conditions (UV-B100 control, UV-B134 and UV-B166, respectively), or in chambers providing below-ambient (22% of ambient) UV-B (UV-B22) along with a UV-A control and a photosynthetically active radiation (PAR) control. Exposure of seedlings to UV-B166 radiation reduced (P < or = 0.05) leaf and stem dry mass by 34 and 39%, respectively, in C. maculata, and reduced leaf nitrogen concentration (%N) by 12% in V. oroboides. Nodule %N in C. maculata and stem %N in P. calyptrata also decreased (P < or = 0.05) in response to UV-B22 radiation compared with the UV-A control, but not compared with the PAR control. Concentrations of flavonoids, soluble sugars and starch were unaltered by the UV-B treatments. Application of 1 mM NO3 to UV-B166-treated seedlings increased whole-plant dry mass of V. oroboides and P. calyptrata by 47 and 52%, respectively. Dry mass of organs, nodule %N and total N concentration of these species also increased with NO3 application. However, NO3 supply decreased (P < or = 0.05) nodule dry mass, stem %N and leaf %N as well as root and leaf anthocyanin concentrations in C. maculata. In terms of UV-B x N interactions, dry mass of stems, roots, nodules and total biomass of NO3-fed C. maculata seedlings were reduced, and nodule %N, total N and leaf anthocyanins were depressed by the UV-B134 and UV-B166 treatments relative to UV-B100-treated seedlings. Although we found that above-ambient UV-B had no effects on growth and symbiotic function of V. oroboides and P. calyptrata seedlings, feeding NO3 to these species increased (P < or = 0.05) seedling growth. In contrast, purely symbiotic C. maculata seedlings were sensitive to the UV-B166 radiation treatment, and adding NO3 further increased their sensitivity to both the UV-B134 and UV-B166 treatments.