BULLETIN OF MARINE SCIENCE, 61(2): 225–240, 1997 LARVAL AND POSTLARVAL DEVELOPMENT OF THE NEW ZEALAND PIPI, PAPHIES AUSTRALIS (BIVALVIA: MESODESMATIDAE) Simon H. Hooker ABSTRACT Pipi (Paphies australis) were conditioned, spawned and the resultant larvae reared to settlement in the laboratory. Adult pipi were successfully conditioned after 23 d at 22° C. Spawning was induced using a combination of increased temperature and a dilute sperm solution. Settlement occurred 18-22 d after spawning at a mean shell length of 264 µm. After settlement and metamorphosis pipi gradually began to take on the adult shape. Cultured juveniles were grown to 37 mm shell length both in the laboratory and later in the wild. Once in the wild, cultured pipi grew from 13 to 37 mm in 17 mo with a strong seasonal component to their growth. Microscopic examination revealed slight differ- ences in shell morphology between pipi larvae and the closely related tuatua (Paphies subtriangulata) and toheroa (Paphies ventricosa). Pipi were more rounded and smaller than both tuatua and toheroa at a similar stage of development. The results of scanning electron microscopic examination of larval shell hinge structure confirmed previous pre- liminary findings for pipi. The larval shell ligament is posterior to the center of the provinculum in pipi but central in both the tuatua and toheroa. These differences are sufficient to enable larval pipi to be distinguished from larval toheroa and tuatua. The pipi (Paphies australis) is a common burrowing bivalve of the Family Mesodesmatidae, one of the most familiar clams of New Zealand beaches. The family is less common in other parts of the world. Pipi are found throughout New Zealand, the Chatham Islands and in the Auckland Islands (Powell, 1979). There are four species in the family; Paphies ventricosa, (toheroa) Paphies subtriangulata (tuatua), Paphies donacina (deep water tuatua) and Paphies australis. Pipi are restricted to channels and sand banks at the mouths of harbors and occur in the top few centimeters of coarse sediments. The other Paphies species are most abundant on open coasts (Morton and Miller, 1968). Pipi have long been an important part of the diet of New Zealanders. They are an important part of the recreational harvest, and support a small commercial fishery with an average annual harvest of around 120 t (Haddon, 1989). The pipi is also a potential candidate for aquaculture (Hayden, 1988). Despite its popularity and the widespread distribution of the species there is no published scientific information available on pipi. Some information, usually resulting from short-term environmental studies, is contained in a number of unpublished reports (Creese, 1988; Richie, 1974; Richie and Mason, 1977; Mason and Richie, 1979; Dickie, 1986a; 1986b; 1986c; 1987a; 1987b; Venus, 1984), and an unpublished thesis (Grace, 1972). Bivalves show the greatest diversity of larval development in the mollusca (Chanley, 1969). Their larvae are an important part of marine plankton communities around the world (Thorson, 1946). Identification of bivalve larvae by shell characteristics, such as morphometric shell measurements, hinge structures and overall size, is well documented (Rees, 1950; Loosanoff et al., 1966; Chanley and Andrews, 1971; Lutz et al., 1982). It has been successfully applied to many species around the world. In New Zealand, provi- 225 226 BULLETIN OF MARINE SCIENCE, 61(2): 225–240, 1997 sional larval identification of many of the common bivalve larvae in the plankton was made by Booth (1977; 1979a; 1979b; 1983). However, positive larval identification usu- ally requires laboratory culture of the species (Loosanoff et al., 1966; Chanley and Andrews, 1971; Booth, 1983). Larval identification of New Zealand bivalves has been confirmed so far by laboratory culture for the rock oyster Saccostrea glomerata (Dinamani, 1973; 1976; Chanley and Dinamani, 1980), the Pacific oyster Crassostrea gigas (Chanley and Dinamani, 1980), the dredge oyster Tiostrea chilensis (Chanley and Dinamani, 1980), two commensal bivalve molluscs Arthritica crassiformis and Arthritica bifurca (Chanley and Chanley, 1980), the New Zealand cockle Chione stutchburyi (Stephenson and Chanley, 1979), four mussel species Mytilus edulis aoteanus, Perna canaliculus, Xenostrobus pulex, Modiolarca impacta (Redfearn et al., 1986), and two surf clams, the toheroa P. ventricosa (Redfearn, 1982), and tuatua P. subtriangulata (Redfearn, 1987). This paper describes the larval and early post-larval development of the pipi by larval culture techniques. This information is useful for identification of pipi larvae from plankton samples to help in the prediction of settlement patterns. Pipi larval culture and informa- tion on the growth of juvenile pipi may also help to establish the feasibility of pipi aquac- ulture. MATERIALS AND METHODS The following research was done in two batches (batch 1 and batch 2). All pipi used in this study were collected from the Whangateau Harbor, in northeastern New Zealand. Batch 1.—The following experiment was designed to investigate the feasibility of conditioning adult pipi and rearing their larvae, and to gather preliminary information on the possibility of sus- pended culture and growth rates of hatchery pipi. Conditioning.—In May 1991, 92 adult pipi (mean shell length = 61.0 mm, SE = 1.31), were collected from the Whangateau Harbor. Thirty were fixed in Bouin’s solution for histological ex- amination and the remaining 62 pipi were placed in 300-liter tanks of filtered (10 µm) seawater at ambient temperature. The water temperature was gradually increased over a period of 5 d to a constant temperature of 22°C ± 1°C. Each day the tank was emptied, cleaned and refilled with 10 µm filtered seawater at 22°C. Animals were fed daily with approximately 40 liters of live, near axenic microalgae, (Isochrysis galbana “Tahitian” CS-177 and Thalassiosira pseudonana, clone 3H CS-173; obtained from CSIRO, Australia). Histology.—All tissue sections were blocked in paraffin, cut at 5-7 µm thickness and stained with Mallory-Heidenhain. All histological sections were examined with a light microscope at 40x, 100x and 400x magnification. Each section (individual) was put into one of four categories (early active stage, late active stage, mature stage, partially spawned stage) as modified from Ropes and Stickney (1965) and Ropes (1968). A description of these categories is given in the account of pipi reproductive biology (Hooker and Creese, 1995). Spawning and Larval Development.—Twenty three days after collection of the adult pipi, they were removed from the conditioning tank, and 34 were preserved as above for histological exami- nation. The remaining individuals were spawned using a combination of temperature shock (27°C) and a dilute sperm solution. Spawning male and female pipi were separated so the eggs and sperm were collected separately and the eggs fertilized with a dilute sperm solution. The fertilized eggs were placed in 300-liter tanks of filtered (1 µm) seawater at a density of approximately 20 eggs ml- 1. Every 2 d the resultant larvae were sieved out, and the tank cleaned and refilled with fresh (10 µm) seawater at 20°C ± 1°C. Veligers were fed the same two microalgal species used to condition the adults at a concentration of approximately 105 cells per ml of culture tank. Pediveligers were settled in a downweller arrangement with a small amount of sand from the Whangateau Harbor. In HOOKER: CLAM LARVAL DEVELOPMENT 227 Figure 1. Morphometric measurements used in larval pipi (after Chanley and Chanley, 1980). early September 1991, 3 mos after the adults had spawned, the juveniles were moved to an upweller. In January 1992 they were individually tagged, using glue-on plastic mollusc tags (Hallprint, Aus- tralia). Tagged pipi were replaced in the upwellers and fed on raw seawater. In July 1992, 80 were put out in the Whangateau Harbor in hanging nets. The nets were suspended from a stationary raft, approximately 2 m from the bottom, in 8 m of water near the entrance of the Whangateau Harbor. Batch 2.—Once the techniques for larval rearing had been perfected, a second batch of pipi was reared to thoroughly describe the larvae as an aid to identification in plankton samples. Previous descriptions of mesodesmid larval development had ceased at time of settlement (Redfearn, 1982; 1987). It is useful however, to also describe the development through to the post metamorphic shape of the juveniles commonly found in field samples. Therefore, the second batch describes the larval and early post-larval development of pipi. In late October 1993, pipi were observed spawning at the mouth of the Whangateau Harbor. Adult specimens were collected from this site and taken to the laboratory. To obtain information on egg size, a haphazard selection of 10 females was “stripped” and their eggs measured. Thirty animals were placed in a 300-liter tank of 1-µm filtered seawater, at 28°C. These pipi took much longer to spawn than those in batch 1 and actual spawning was not observed, occurring during the night. Trochophore larvae were present in the tanks the following morning, and these were col- lected, a sample measured and the remainder re-suspended in the larval tanks. Veliger larvae were 228 BULLETIN OF MARINE SCIENCE, 61(2): 225–240, 1997 Figure 2. Histograms of the proportion of conditioned adult pipis in each gonad stage. A, wild pipis at the start of conditioning (7 May 1991). B, pipis conditioned in tanks for 23 d. C, wild pipis at the end of conditioning period (4 June 1991). cultured as previously described. Larvae were fed as in batch 1 but with the addition of two further microalgal species: Pavlova lutheri, CS-182 (at all developmental stages), and for late-stage ve- ligers, Chaetoceros gracilis was also fed. Measurements.—Every 2 d a sample of larvae was removed, photographed and video taped. A total of 599 larvae was measured covering the full range of sizes. The dimensions measured (Fig.