Bacterial leaf scorch caused by Xylella fastidiosa is a serious problem in the landscape, but its importance in the nursery trade is undetermined. Therefore, a survey was conducted in 2003 and 2004 in a commercial wholesale and production nursery in Maryland to determine the natural occurrence of X. fastidiosa in nursery plants and surrounding vegetation using enzyme-linked immunosorbent and nested polymerase chain reaction assays specific for the bacterium. Xylella fastidiosa was associated with crape myrtle (Lagerstroemia indica), Japanese stewartia (Stewartia pseudocamellia), Schipka laurel (Prunus laurocerasus 'Schipkaensis'), and Japanese flowering cherry (Prunus serrulata 'Kwanzan') in the nursery, as well as with mimosa (Albizia julibrissin) and sassafras (Sassqfrass albidum) growing in the perimeter of the nursery. All were previously unknown hosts of the bacterium. Xylella fastidiosa was also detected in dogwood (Cornus florida) and willow oak (Quercus phellos) in the nursery, as well as in red oak (Quercus spp.), box-eider (Acer negundo), and wild grape (Vitis spp.) surrounding the nursery, previously reported hosts of the bacterium. The red oak and box-elder plants on the edge of the nursery showed leaf scorch symptoms and harboured high populations of the bacterium. However, the Xylella-positive nursery plants and other plants from the local environment were symptomless, and the bacterial populations were low. These data suggest that red oak and box-elder are sources of the X. fastidiosa in the nursery plants.