Manganese (Mn) is an essential micronutrient and has a broad range of functions for all plant growth and reproduction. It plays a crucial role in NAD-ME C4 photosynthesis, including as a constituent of the water splitting protein of photosystem II (PSII) and as an activator of NAD-ME which catalyzes the release of CO2 from malate in bundle sheath cells (BSC). Switchgrass (Panicum virgatum L.), a perennial NAD-ME C4 grass, is native to much of the United States. Switchgrass is also considered as a promising biofuel species for sustainable production of bioenergy feedstock. As a NADME C4 species, switchgrass may have a high requirement for Mn for optimum growth. However, little is known about switchgrass responses to Mn availability at present. To study the influence of Mn on biomass production and photosynthetic characteristics, one lowland ('Alamo') and one upland ('Cave-in-Rock') switchgrass ecotype were grown in 19-L pots filled with either washed sand, vermiculite, or perlite, and fertilized with nutrient solutions with Mn concentrations ranging from 0 to 200 [micro]M under field conditions in three consecutive years. In the last year (perlite), pearl millet (Pennisetum glaucum L. R. Br.) ('KGraze') was also grown. Shoot Mn concentration was highly responsive to increasing Mn in the nutrient solution in all experiments and for all entries. When grown in washed sand and vermiculite, no Mn treatment effects on biomass production were found for either switchgrass ecotype. In perlite, a significant decrease in biomass production grown in the 0 [micro]M Mn treatment compared to 10-25 [micro]M Mn treatments was only observed for Alamo and KGraze, and not for Cave-in-Rock. Late in the season, relative chlorophyll contents of both switchgrass ecotypes were significantly lower in the 0 [micro]M Mn treatment than other treatments, but, in KGraze, relative chlorophyll content was low early in the season, increased throughout the season, resulting in a less pronounced, but still significant Mn treatment effect, at late stages. Leaf Mn concentration of all entries increased with increasing Mn concentration in the nutrient solution. In switchgrass, leaf Mn concentration was significantly greater early compared to late in the season in the absence of Mn in the nutrient solution; however, this was not the case for pearl millet. In switchgrass, the absence of Mn in the nutrient solution significantly decreased photosynthetic rates and maximum PSII efficiency (Fv/Fm) late in the season. In contrast, in pearl millet the effect of 0 ë_M Mn in the nutrient solution on net photosynthesis and Fv/Fm was more pronounced early in the season. Chloroplast ultrastructure in mesophyll and bundle sheath cells were only affected by Mn availability in the lowland switchgrass ecotype. Manganese availability did not influence NAD-ME, NADP-ME and PEPCK activities in switchgrass, but NADME and PEPCK activities were reduced in pearl millet early in the season in the absence of Mn in the nutrient solution. Based on these results, Mn limitation for the oxygen evolving compex of PSII rather than for NAD-ME was the primary limitation of low Mn availability on net photosynthesis. Overall, switchgrass and pearl millet exhibited distinct temporal responses to limited Mn availabillity.