CHARACTERIZATION OF VACUOLAR BODIES IN SPARTINA ALTERNIFLORA: I. FORMATION, DEVELOPMENT, MORPHOLOGY, AND ULTRASTRUCTURE

Spherical, golden bodies, 0.5 to 25 um in diameter, were noted in outer bundle sheath and mesophyll of cells of fresh sections of Spartina alterniflora leaves. Attempts to further characterize these structures with light and electron microscope (EM) techniques after dehydration failed initially because these bodies were soluble in organic solvents such as alcohol. Subsequently, it was found that certain heavy metals would stabilize the structure so that dehydration techniques could be used. The resultant stabilized bodies as viewed with EM were found to reside in the cell vacuole. Two major structural components were recognized: (1) a granular matrix and (2) a non-electron dense internal vesicle. Internal vesicles were not always present. No membranes were visible with this technique. These vacuolar bodies also were found to occur in parenchyma cells of roots, peduncles, glumes, and rhizomes of S. alterniflora. The material which forms the vacuolar body matrix is probably produced in the cytoplasm and gradually accumulates in the vacuole, forming larger and more numerous vacuolar bodies during the growing season. The matrix material remains in the leaf cells following their senescence. PREVIOUS STUDIES on vacuolar body formation within plant cells suggest three mechanisms by which vacuolar bodies are formed: (1) the formation of vacuolar bodies within the central vacuole as substances are secreted into and accumulate within that vacuole (Reihles, 1936; Shumway, Rancour, and Ryan, 1970); (2) the formation of vacuolar bodies within cytoplasmic vacuoles, with the cytoplasmic vacuoles subsequently fusing with the main vacuole (Chafe and Durzan, 1973); and (3) the formation of vacuolar bodies by a pinocytotic-like process of the tonoplast (Matile and Moor, 1968; Fineran, 1971). The development of vacuolar bodies has been studied in meristematic cells of plants by Matile and Moor (1968) and Fineran (1971). These authors observed that although some vacuolar bodies increased in size after formation, all vacuolar bodies disappeared as the meristematic cells differentiated. Vacuolar bodies within various plants have been described as: (1) smooth surfaced, solid or hollow spheres (Buvat, 1937); (2) irregularly shaped masses with concentric lamination (Reihles, 1936; Graham, 1972); (3) irregularly shaped, amorphous masses of material (Shumway et al., 1970; Maitra and Deepesh, 1972; Chafe and Durzan, 'Received for publication 29 June 1976; revision accepted 1 March 1977. Paper No. 5010 of the Journal Series of North Carolina Agr. Exp. Sta., Raleigh, N.C. June 1976. A portion of this study was supported by the Institute of Environmental Sciences of the University of North Carolina, Chapel Hill. 1973); or (4) rough surfaced, solid spheroids (Greyson and Mitchell, 1969). Vacuolar bodies (originally designated as cytoplasmic vacuoles) were observed within cells of Spartina alterniflora Loisel and described by Akers and Anderson (1973). These bodies were vesiculate, smooth surfaced spheroids and, therefore, resembled the vacuolar bodies studied by Buvat (1937). They differed in that: (1) vacuolar bodies of S. alterniflora were not found in meristematic cells; (2) these vacuolar bodies increased in size and number in certain mature cells; and (3) that these vacuolar bodies were unstable in preparation procedures in which alcohol was used. The latter factor in part explains why they have not been described previously. This paper reports on the formation, development, morphology, and ultrastructure of vacuolar bodies in S. alterniflora. MATERIALS AND METHODS-Plant materialSpartina alterniflora plants from a regularly flooded salt marsh located near Sunset Beach, North Carolina, were collected for three consecutive years. Some plants were transplanted to pots in the greenhouse where they were flooded daily with tap water and fertilized at two-week intervals. Greenhouse transplants were used for observation on vacuolar body morphology. Plants collected directly from the marsh were used to study vacuolar body distribution, development, and ultrastructure. Light microscope methods-Standard paraffin methods such as those described in Johansen