BULLETIN OF MARINE SCIENCE. 33(2): 363-372,1983 CORAL REEF PAPER BREEDING PATTERNS OF THREE SPECIES OF CARIBBEAN BRITTLE STARS (ECHINODERMATA: OPHIUROIDEA) Philip V Mladenov ABSTRACT Reproductive periodicity of three species of brittle stars at Barbados, W.I. was studied for one year using a gonad index technique. Ophiothrix oerstedi has a synchronous annual re- productive cycle with a breeding season that extended from mid-July until early December. Ophiothrix suensoni reproduces continuously, but individual reproductive cycles are asyn- chronous and there was higher spawning activity in the late summer and early fall. Ophiocoma echinata has an asynchronous annual reproductive cycle with a prolonged breeding season. Males of O. oersledi and O. suensoni have significantly larger gonads than females at certain times during the breeding season; a large volume of spermatozoa may be required to com- pensate for sperm wastage during spawnings. Detailed information is presently available for reproductive periodicity of 19 species of tropical brittle stars. Contrary to earlier suggestions, prolonged breeding seasons and continuous reproduction are common in tropical brittle stars. Brittle stars are an abundant and diverse component of invertebrate faunas associated with coral reefs, coral rubble and soft sediments of the tropics (Glynn, 1973; Bray, 1975; Clark, 1976; Singletary, 1980; Sloan, 1982). Until recently, inferences concerning reproductive periodicity of tropical brittle star species were based mainly on sporadic, qualitative observations made by Mortensen (1921; 1931; 1937; 1938) of gonad ripeness and laboratory spawnings. Stephenson (1934) was the first investigator to study the reproductive cycle of a tropical brittle star throughout a I-year period. Summarizing Mortensen's data, Boolootian (1966) concluded that tropical ophiuroids have a breeding season of only I to 2 months duration and are generally spring and summer spawners. In recent years, the reproductive cycles of 16 species of tropical brittle stars have been studied for a period of at least I year using various quantitative gonad index techniques (Hen- dler, 1973; 1979; Stancyk, 1974; Singletary, 1980). These studies show that some tropical brittle star species have lengthy breeding seasons of at least 4 months duration, while some species reproduce continuously throughout the year. In this paper, in an effort to supplement our present understanding of breeding patterns of tropical brittle stars, I present information on reproductive periodicity of three species of Caribbean brittle stars, Ophiothrix (Ophiothrix) oerstedi Ltitken, Ophiothrix (Acanthophiothrix) suensoni Ltitken, and Ophiocoma echinala (La- marck) studied at Barbados, West Indies. The reproductive cycle of the first two species has not previously been studied, whereas the reproductive cycles of several populations of O. echinata from the Atlantic coast of Panama have been studied by Hendler (1979). All three species treated in this study are widely distributed in Caribbean waters (Clark, 1933; Parslow and Clark, 1963). O. oerstedi, a member of the family Ophiothricidae, ranges from southern Florida southwards to Trinidad and the north coast of South America at depths from 0.5 m to 400 m. Both O. suensoni, also of the family Ophiothricidae, and O. echinala, which belongs to the family Ophiocomidae, range from Bermuda southwards to Brazil but have not been reported on the United States coast north of Florida; the bathymetric range of the former is 2 to 525 m, while the latter is found in water less than 5 m deep. O. oerstedi may attain a disc diameter of more than 14 mm, but adults are generally 363 364 BULLETIN OF MAR[NE SCIENCE, VOL. 33, NO.2, [983 less than about 10 mm in disc diameter; very large specimens of 0, suensoni can have a disc diameter of nearly 20 mm, although the disc diameter of adults is usually less than 12 mm; 0, echinata is a large brittle star with a disc diameter that may exceed 30 mm, but typical adults are less than 25 mm in disc diameter (Clark, 1933; Mladenov, 1976), MATERIALS AND METHODS Approximately 20 adult individuals of each of the three species were collected on a regular basis from June 1975 to May 1976. The mean size of specimens sampled was 8.0 mm ± 1.1 mm (n = 305), 7,6 mm ± 1.7 mm (n = 314) and 18,7 mm ± 2.6 mm (n = 215) for Ophiothrix oerstedi, Ophiothrix suensoni and Ophiocoma echinata respectively. Animals with broken or regenerating arms were never collected. Sampling frequency for each species depended upon the observed state of reproductive activity at a particular time of the year; ifgonad size was changing rapidly, then collections were made at least twice monthly; when reproductive activity was low, collections were made only monthly until evidence of increased reproductive activity was noted. Ophiothrix oerstedi was collected by snorkeling from depths of 3 to 10m from a portion of the fringing reef just offshore from the Bellairs Research Institute of McGill University, which is located midway along the west coast of Barbados, West Indies (13°11'N, 59°38'W). O. oerstedi is particularly common within the interstices of the finger-coral Porites porites. Small chunks of P. porites were separated from a larger bed of this coral and returned to the laboratory in plastic buckets. Here the coral was broken into fragments and the brittle stars gently removed. Ophiothrix suensoni was collected by S.C.U.B.A. from depths of 15 to 25 m at a site on the offshore banking reef (which parallels the west coast of Barbados) approximately 1 km south of the Bellairs Research Institute. O. suensoni is epizoic on a variety of sponges, most notably Cal/yspongia spp., Xestospongia spp. and Verongia spp. During each collection, several sponges were placed in buckets and returned to the laboratory where the brittle stars were carefully removed from the sponge surface. Ophiocoma echinata was collected at low tide at depths of less than 1 m at a site on the southwest coast of Barbados known as Grave's End (13°4'N, 59°36'W). Specimens obtained by overturning coral rubble were transported to. the laboratory in insulated containers. All specimens were processed on the day of collection. The animals were first killed by a 2-min immersion in freshwater, and then transferred to a 10% formalin in seawater solution for 24 h. Killing in freshwater prevented arm autotomy, while the period in formalin hardened the gonadal tissue slightly, making its removal easier. The aboral covering of the disc of each specimen was removed to expose the gonads. The sex was noted, and all of the gonads were carefully removed from the disc with fine forceps and placed in a pre-weighed aluminum pan. The arms and disc remains were placed in another pre-weighed pan. The tissues were dried to constant weight at 75° to 80°C (2-3 days), cooled for 2 h in a desiccator (to minimize the amount of atmospheric moisture absorbed by the tissues during cooling), and then weighed to the nearest 10-5 g. Empty pans were pre-weighed following identical drying and cooling procedures. The gonad index (GI) was defined as: GI = dry wt of gonads x 100 total dry wt of organism (including gonad weight) Gonad indices for each species at each sampling date were plotted as frequency distributions and a mean index value calculated. All three species are dioecious and male and female data were plotted separately and the mean index values of the two sexes at each sampling date tested for statistical conformity using Wilcoxon's signed-ranks non-parametric test (P = 0.025, two-tailed) (Sokal and Rohlf, 1969). Microscopic examination of smears made from the preserved gonads of selected specimens of all three species was carried out on most sampling dates, prior to gonad index calculations, in order to compare gonadal development with gonad index values. The gonadal tissue of the smears was returned to the appropriate pan before drying and weighing took place. In addition, testis smears were made from fresh tissue obtained from approximately five adult males of each species collected at roughly monthly intervals throughout the sampling period and examined microscopically for the presence or absence of active spermatozoa. Information obtained from gonad indices and gonad smears was supplemented, whenever possible, with records of spawnings occurring in the laboratory. In order to induce laboratory spawnings, individuals of all three species were held in finger bowls following the procedure described in Mladenov (1979). This method was routinely successful in inducing spawning only for Ophiothrix oerstedi. MLADENOV: BREEDING PATTERNS OF CARIBBEAN BRITTLE STARS 365 RESULTS Ophiothrix oerstedi The mean gonad indices attained their highest values of the sampling period in July, reaching 5.8 for males and 4.6 for females (Fig. 1). The mean indices for both sexes then declined gradually, with some fluctuations, throughout the interval from August until mid-December, indicating a gradual reduction in gonad size over this time, and thus a prolonged period of spawning. From mid-December until February, the mean gonad indices remained at a very low value and the population was reproductively inactive. In March the gonad index of some mem- bers of the population was increasing; by early May the gonad index of most individuals was increasing, suggesting that gametogenesis and gamete accumu- lation were well underway by this time. The preserved gonads of all individuals examined microscopically during the mid-December to February resting phase of the reproductive cycle were very small and lacked spermatozoa or contained only small (less than 30 /.tm in di- ameter) oocytes. From March to June gonads were larger but still lacked sper- matozoa or oocytes larger than 150 /.tm in diameter. Microscopic observation of preserved gonads from specimens sampled during the period from mid-July to early December revealed that the gonads always contained large (greater than 200 /.tm in diameter) oocytes or a mass of tailed sperm, although the relative size of the gonads often varied distinctly from one individual to the next.