Norimitsu Watabe

EXPERIMENTAL STUDIES ON CALCIFICATION IN MOLLUSCS AND THE ALGA COCCOLITHUS HUXLEYI

The calcium carbonate crystals of biological systems occur as calcite, as aragonite, and-less commonly-as vaterite. The reasons for the formation of a particular type of crystal in a given organism are not well understood, although studies of crystallization in solutions have indicated factors which may play a role. These include the presence of organic compounds, inorganic salts, and variations in temperature (Saylor, 1928 ; Kitano, 1962; Kitano et al. 1962; Curl, 1962; Wilbur and Yonge, in press). In molluscs calcium carbonate crystals are usually formed in association with an organic matrix, and it has been suggested that the chemical structure of the matrix may determine the type of crystal (Roche et al., 1951; Ranson, 1952; Watabe and Wilbur, 1960). Temperature also appears to have an influence on the type of crystal deposited by animals. Lowenstam (1954a, b ) found that in organisms which deposited both aragonite and calcite the proportion of aragonite was higher in those animals that lived a t higher environmental temperatures. The first, considered in SECTION I, concerns the influence of three conditions on crystal type: (1) the presence of organic matrices taken from aragonitic and calcitic shells; (2) alteration of temperature; and (3) the supply of nitrogen. In investigating the influence of the matrix we have determined the crystal types which form on decalcified matrices taken from various molluscan species and introduced into other species or recalcified in vitro. The effects of temperature on crystal type have been examined in two systems, the regenerating shell of the snail Viviparus intertextus and the alga Coccolithus huxbyi. The effect on crystal type of reducing the nitrogen content of the medium has also been observed in C. huxleyi. SECTION I1 concerns the relation of crystal formation to cell structure in C. huxleyi. This cell provides an especially interesting opportunity for study of this relationship in that one strain normally forms no CaC03 crystals but can be induced to calcify by reducing the nitrogen content of the medium.

Studies on shell formation: XI. Crystal—matrix relationships in the inner layers of mollusk shells

Details of crystal growth in the calcitostracum of Crassostrea virginica have been studied with the purpose of analyzing the formation of the overlapping rows of oriented tabular crystals characteristic of this part of the shell. Crystal elongation, orientation, and dendritic growth suggest the presence of strong concentration gradients in a thin layer of solution in which crystallization occurs. Formation of the overlapping rows can be explained by three processes observed in the shell: a two-dimensional tree-like dendritic growth in which one set of crystal branchings creeps over an adjacent set of branchings; three-dimensional dendritic growth; and growth by dislocation of crystal surfaces. Multilayers of crystals may thus be formed at one time. This is favored by infrequent secretion of a covering organic matrix which would inhibit crystal growth. The transitional zone covering the outer part of the calcitostracum and the inner part of the prismatic region is generally characterized by aggregates of small crystals with definite orientation. Growth in this zone appears to take place in a relatively homogeneous state of solution without strong concentration gradients. Thin membranes and bands of organic matrix were commonly observed in the transitional zone bordering the prismatic region. The membrane showed a very fine oriented network pattern.

TROPICAL MARINE ALGAE: GROWTH IN LABORATORY CULTURE

Mature tropical marine algae were transplanted from Jamaica, Bermuda, and Florida to aquaria at Duke University, North Carolina and The Queens University of Belfast, Northern Ireland. Genera cultured included: Caulerpa, Halimeda, Penicillus, Udo‐tea, Acetabularia, Batophora, Cymopolia, Dictyosphaeria, Neomeris, and Valonia. Methods of transport and culture are described. The use of adult plants was convenient, as well as essential for those genera and species in which spore or gamete stages are as yet unknown. Since only small amounts of seawater were needed for maintenance of the cultures, studies on these livinng marine algae can be made at institutes far from the sea.

Collagen in the Spicule Organic Matrix of the Gorgonian Leptogorgia virgulata

Decalcification of the calcareous spicules from the gorgonian Leptogorgia virgulata reveals an organic matrix that may be divided into water insoluble and soluble fractions. The insoluble fraction displays characteristics typical of collagen, which is an unusual component of an invertebrate calcium carbonate structure. This matrix fraction exhibits a collagenous amino acid profile and behavior upon SDS-PAGE. Furthermore, the reducible crosslink, dihydroxylysinonorleucine (DHLNL), is detected in this fraction. The composition of the matrix varies seasonally; i.e., the collagenous composition is most prevalent in the summer. These results indicate that the insoluble matrix is a dynamic structure. Potential roles of this matrix in spicule calcification are discussed.