The structure and composition of the shell of living Lingula have been studied in detail to provide a measure for evaluating the evolution of the lingulid shell since early Palaeozoic times. Four constituents are identifiable at ultrastructural level. Apatite, specifically the fluorapatite francolite, occurs as coated granules, up to 10 nm in diameter, aggregated into spheroidal masses up to several microns in size, cylindroids up to 600 nm long and rare plates. Glycosaminoglycans (GAGs) are present throughout the shell as an elastic isotropic gel. Chitin is normally associated with a protein and is best seen under the scanning electron microscope after digestion of its associate by proteinases. As strands about 30 nm thick, chitin can form mats or be part of granular rods aligned as anastomosing ridges. Fibrillar collagens, with a periodicity of about 45 nm, occur mainly as sporadically developed mats throughout the body platform succession and as the core of the dorsal median septum. Elsewhere they appear sparingly as vertical and horizontal strands. At least ten proteins have been identified in the shell but it is not yet known which are covalently attached to GAGs, associated with the chitin or fabricated into membranes. The entire integument consists of: a periostracum, about 4 mu m thick, made up of closely packed fibrous tubes with axial strands presumably of chitin; a primary layer, about 40 mu m thick, composed mainly of GAGs; and a mineralized secondary layer of variable thickness. The secondary layer is a succession of isotopic and isochronic laminae with thicknesses in microns compared with areas frequently of several square millimetres. Compact laminae, composed of closely packed spheroidal aggregates of apatitic granules are succeeded: by botryoidal or walled laminae, in which apatitic aggregates form botryoidal masses or vertical walls in a GAGs matrix; or by rod and plate laminae with apatitic rods accreting into anastomosing ridges disposed transversely on the body platform and radially in peripheral regions. The preponderance of the botryoid and rod laminae in the body platform and of anastomosing ridges in the lateral areas of a valve respectively explains the complex patterns of c-axis orientation of apatite obtained by X-ray diffraction. Membranous laminae, consisting of chitinoproteinaceous membranes (and sporadic collagenous mats) in GAGs, occur throughout the succession; while stratified laminae, characterized by gentle inclined, alternating organic and apatitic units, each just over a micron thick, are especially well developed (with compact laminae) at the junction with the primary layer. Laminae group into well defined rhythmic sets beginning with an abrupt change from a membranous lamina to a biomineral succession founded on a compact lamina and passing gradually into a terminating membranous lamina which serves as the substrate for the next set. These rhythmic successions are secreted directly, without the intervention of extrapallial space, by outer epithelium which is anchored to the shell by variably persistent arrays of fine canals so that any cell is capable of secreting, in the right order, all the components of a rhythmic unit. When a compact lamina is being secreted apatitic granules are exocytosed through the digitate protrusions of the apical plasmalemmas of cells with clusters of large vesicles containing mottled electron-dense material and with mitochondria congregating apically. Termination of the compact lamina is marked by a sudden onset of GAGs secretion, probably from intercellular sources as well as from the cells, in which large vesicles of medium electron-dense material appear. Towards the end of a rhythmic cycle of secretion, large vesicles, which are mostly empty, and mitochondria are more dispersed throughout the cells. Two features of phylogenetic importance have been revealed. The outer layers of the integument, the periostracum, primary layer and the underlying stratified laminar set of the secondary layer, are found in living Discina and Lingula and must have been characteristic of the ancestor of all organophosphatic brachiopods. The association of the newly discovered collagen with GAGs, apatite and chitin is probably a unique feature of the metazoan exoskeleton. It does, however, compare most closely with the uncalcified cuticle of certain protostomes, especially the priapulids, and strengthens the claim of affinities between the Priapulida and lingulide brachiopods based on their respiratory protein being haemerythrin.
