Peters and Rothman Introduction The pea crab Pinnotheres pugettensis (Holmes, 1900) is known from Departure Bay, B.C. and Puget Sound, Washington, where it occurs in the ascidians Halocynthia aurantium, H. hilgendorfi igaboja and Ascidia paratropa, and from Monterey Bay, California where it was recently discovered in the ascidian Styela montereyensis (Dall, 872) by C. C. Lambert (Garth and Abbott, 1980). The range of P. pugettensis may extend even further south; a pea crab was found in one S. montereyensis collected at Pt. Loma, California (Fay and Johnson, 1971). Pinnotheres taylori Pinnaxa faba, and Fabia subquadrata, however have also been reported from Pacific ascidian hosts (Schmitt, et al, 1973). Pinnotheres pugettensis has not been extensively studied. Very little is known of its occurrence, it position in and relation to the host ascidian, its methods of feeding, and its developement. The present research was undertaken to try and answer some questions in these areas, using the P. pugettensis population in S. montereyensis from southern Monterey Bay. The ascidians studied often harbored the notodelphyid (Iiig) copepod Pygodelphys aquilonaris, known on this coast from studies in Washington, (e.g. Illg, 1958; Dudley, 1966). Some notes on its occurrence in S. montereyensis in Monterey are included. Peters and Rothman Materials and Methods Specimens of S. montereyensis were collected between April and June 1980. Subtidal collections were made from depths of 15 to 35 feet in kelp beds at Mussel Pt., and Pacific Grove;,Pt. Alones and Wharf no. 2, Monterey, sing SCUBA. Intertidal collections were made by hand at Pescadero Pt., Monterey Peninsula, as well as at Mussel Pt. and Wharf no. 2. Specimens were kept alive in tanks at Hopkins Marine Station, where they were suspended from styrofoam floats by plastic clips clasping the base of the stalk. Aquaria were supplied with running seawater at 10-15°0, a temperature generally within three degrees of ambient local ocean temperatures. Tunicate body volumes were measured by immersing the body (but not the stalk) of the contracted specimen in a 100 ml graduated cylinder of seawater and noting displacement. S. montereyensis shows no external evidence of presence or absence of a pea crab. After initial trials, all tunicates were dissected by making a longitudinal slit through the tunic from the base of the oral siphon at the anterior end down to the beginning of the stalk. The tunic was pulled aside to reveal the translucent mantle, within which a crab if present, could usually be observed. The mantle and the pharynx were then opened with a cut similar to the first incision, Peters and Rothman this one running along the length of the endostyle. By dissecting the tunicate in this manner the crab, in the atrial cavity, is minimally disturbed; when incisions were made down either side of the body laterally or on the dorsal side, the crab usually became active and broke through the pharyngeal wall into the pharyngeal cavity of the tunicate. Perhaps this type of action accounts for at least some of the reported instances of pea crabs being found in the pharyngeal basket of ascidians (e.g. Fay and Johnson, 1971). Crabs found in dissections were removed from the tunicate, measure with an ocular micrometer, and kept in running seawater in individual compartments of ice cube trays fitted with screened plexiglass tops. They were fed Artemia naupli intermittently, we are uncertain whether or not they were eaten. In tunicate and crab feeding experiments, phytoplankton suspensions (Chaetoceros gracilis and Isochrysis galbana), colored black with "Aquadag" colloidal graphite or red with powdered carmine, were used. Tunicates used in feeding experiments were placed in 10% formalin and seawater overnight and then dissected. Various methods of anesthesia were tried in the course of the dissections. The most successful procedure was to "tranquilize" the tunicates first in a solution of menthol in seawater for several minutes then place Peters and Rothman them in MS 222 (Ethyl m-aminobensoate, methanesulphonic acid salt 98%, Aldrich) diluted 1:2000 for five to ten minutes. This method, however, never completely relaxed the tunicates. Neither menthol nor MS 222 seemed to have any effect upon the crab. Distribution In the course of the study, 222 subtidal and 25 intertidal Styela montereyensis were collected. In the subtidal tunicates, 73 Pinnotheres pugettensis were found; 49 were females, and 24 were identified as males. However, sex identification of the smallest individuals was difficult. Eight male P. pugettensis were found outside of S. montereyensis in collecting bags and in which S vela were kept. aquaria The overall infection rate of the subtidal Styela population was 32.9%. No crabs were found in intertidal S. montereyensis. In order to obtain an accurate and useful measure of tunicate size the authors compared body volume and body length. Measurements of the stalk were omitted because its length was highly variable and did not correlate well with body size. When body length is plotted against body volume it becomes apparent that the variance in body volume is less then that for body length, (see Fig. 1). Consequently body volume was used as a measure of tunicate size. Peters and Rothman No statistically significant difference in body volume was found between intertidal and subtidal populations of Styela (Student's t-test, p-.02). An Rx C contingency test showed the difference in P. pugettensis infection rates between subtidal and intertidal Styela populations to be highly significant (p£.01). Some caution, however, must be exercisized when interpreting these results; 19 of the intertidal Styela were collected at Wharf no. 2, but none of the six subtidal tunicates found at the Wharf contained P. pugettensis. The difference in infection rates may be due to some other factor than differences in tidal height. Of the infected tunicates 62 contained one crab, four held two crabs, and one tunicate contained three much adult crabs. Male crabs were generally,smaller thangfemales. Males had an average carapace width of 4.Omm (SDt1.6imm). The smallest male had a carapace width of 1.Omm and carapace length of 0.9mm; the largest male was 6.8mmx 6.Omm. Female crabs had an average carapace width of 8.2mm (SDt1.5mm). The smallest female had a carapace 5.2mm x 5.2mm, while the largest was limmx iimm. There was a moderate positive correlation (r = 0.35) between crab carapace width and body volume of the tunicate (see Fig. 2). This positive relationship between host size and crab size has been found in other Pinnotheridae (Pearce, 1966a; Christensen and MeDermott, Peters and Rothman 1958 ; and Silas and Alagarswami, 1967). This correlation is better (r = 0.56) if the few cases of multiple infections are omitted. The rate of infection varied with tunicate body volume; additionally, as volume increased, fewer males and more females were found (see fig. 3). The infection rate of subtidal S. montereyensis by the copepod Pygodelphys aquilonaris was 30.2%. No copepods were found in Styela taken intertidally. An Rx C contingency test showed the difference between the intertidal and the subtidal infection rates to be highly significant (p«.01). However, subtidal Styela at Wharf no. 2 lacked copepods, just as they lacked pinnotherid crabs. The infection rate with P. aquilonaris varied with the size of the tunicate (see fig. 3). Multiple infections were fairly common; the largest number of copepods within a single host tunicate was 15 (see fig. 4). Copepods were found only within the ascidian pharynx. Position and feeding of crab in host Of the total of 73 Pinnotheres pugettensis found in Styela montereyensis, one crab was found within the branchial basket, while 72 were located in the anterior third of the atrial cavity. The ventral surface of the crab was always in contact with the pharynx, the dorsal surface towards the tunic. The Peters and Rothman crabs legs were held closely against the body; not grasping the tissues of the ascidian. Connective tissue ascidian strands and blood vessels joining the mantle to the pharynx were interrupted by the presence of the crab, but these same tissues in the area surrounding the crab remained intact, thus the crab was maintained in position in a sort of pocket. P. pugettensis was found in either the left or right side of the atrial cavity, but always with the mouthparts and chelae within 3mm or less of the dorsal lamina, and facing it. A crab on the right was positioned as in fig. 5a, a crab on the left was rotated 180 degrees. Crabs positioned dorsally in the atrium were oriented with the mouth in the closest proximity to the dorsal lamina of Styela (see fig. 5b). A consistent feature of S. montereyensis containing P. pugettensis in the atrium was a hole in that part of the pharyngeal wall located close to the mouth of the crab (see fig. 6 and 7). This hole was 1-3.5mm in diameter, its size positively correlated with the size of the crab. The hole appeared well healed, the edges were smooth, and the blood vessels of the pharynx were fused and diverted around the hole. Slight damage was occasionally observed to the dorsal lamina in the region next to the hole. (Additionally, when the crab infecting a tunicate was large the testes of the Peters and Rothman ascidian behind the pocket containing the crab were reduced in both size and number). Where more than one crab was present in the tunicate, each crab had its own hole; one crab occurred in the usual position while the additional crab was located either at the same level in the atrium on the opposite side of the pharynx, or lower down in the atrium on the same side as the first crab. The position of the crab in the host, and the neat hole in the pharyngeal wall of the tunicate, strongly suggested thatt the crab was feeding on the mucus rope formed at the ascidian dorsal lamina. S. montereyensis is a suspension feeder, like all ascidians, and collects food particles by passing surrounding water through mucus nets. These nets, which line the pharyngeal cavity, are secreted by the endostyle, then passed across the gill slits on both sides of the pharynx until they reach the dorsal lamina where the sheets from each side are rolled together into a rope and passed down to the ascidian esophagus (see Jørgensen, 1966; Fiala¬ Médioni, 1978; Flood and Fiala-Médioni, 1979). To test whether the crab might be feeding on this mucus rope, phytoplankton marked with "Aquadag" colloidal graphite or powdered carmine was pipetted into the region of the oral siphons of submerged, feeding tunicates. Each of 25 subtidally obtained Styelas was fed for 10-15 minutes. For an additional 15-45 minutes each ascidian was submerged in seawater containing a less concentrated 10 Peters and Rothman mixture of the marker and plankton, finally the animal was killed in a 10% formalin 90% seawater solution. Subsequently the tunicates were dissected and the pharynxes opened. The mucus rope was usually well stained, appearing bright red or black dependent upon the marker used. Nine P. pugettensis were found within these ascidians, and all were carefully removed. Six crabs had the mouthparts and chelipeds stained with the colored mucus, and one was found clutching the mucus rope. Of the above six crabs, three contained dyed food particles in their cardiac stomachs. P. pugettensis does not leave the host to forage, or even move about much within the host atrium, as indicated by the pocket created by the tissue strands and blood vessels encircling the crab. The hole in the ascidian pharynx was always small and almost always well healed, indicating the injury was not recent, and that the crab was not actively eating the pharyngeal tissue. Tunicates containing some of the larger crabs often showed additional scaring of the pharyngeal wall directly next to the body of the crab, but never another hole. S. montereyensis, in the lab, were able to completely heal holes cut in the pharyngeal wall within 1-2 weeks. Further, the crab was always 11 Peters and Rothman located posterior to the anal pore of the ascidian, and was in the wrong position to capture and feed on ascidian fecal pellets. Moreover, in the feeding experiments with stained food the crabs were eating colored material, yet upon dissection of the tunicate gut it was found that the dyed mucus had passed no further than the stomach. These observations all indicate that P. pugettensis is reaching into the pharynx with its chelipeds or mouth parts, through a single small and strategically placed hole in the pharyngeal wall, and feeding upon the mucus rope as it passes down the dorsal lamina. No evidence was found to support previous statements that P. pugettensis is a filter feeder (Wells, 1940, Peters and Rothman Christensen and MoDermott, 1958; Silas and Alararswami, 1967). An examination of crab mouthparts shows no obvious adaptions for filter feeding, and it seems unlikely that the crab could get sufficient quantities of particulate food through the network of the pharyngeal basket, even on occasions when the mucus sheet is not being secreted. It would seem far more advantagous for the crab to take advantage of the highly efficient filtering and collecting mechanisms of the tunicate. by simply stealing portions of the mucus rope as it is being passed back to the ascidian esophagus. The pea crab obtains both food and shelter from S. montereyensis. The food-robbing process probably produces only a small loss to the ascidian host. Further stress may be placed on the host by disrupted tissue strands and blood vessels, the hole in the pharynx, slight damage to the dorsal lamina, and some disruption of the gonads. The reduced testes would be expected to reduce the tunicates reproductive fitness. as is proposed by Berner (Silas and Alagarswami, 1967) Pinnotheres pisun for in the mussel Mytilus edulis, and Christensen and MeDermott (1968) for Pinnotheres ostreum in the oyster Crassostrea yirginica. However, this may be relatively insignificant in Styela, since testes are very numerous and those on the opposite side and below the crab appear unaffected. Additionally the ovaries appear undisturbed Peters and Rothman by the presence of a crab. No evidence was found in S. montereyensis of severe enlargement of the mantle or atrial cavity toaccommodate individual crabs as aurantium from shown for Halocynthia (-Tethyum) Puget Sound infested with P. pugettensis(see Wells, 1928). In our estimation, the damage sustained by S. montereyensis is relatively minor. No quantitative measure of food taken was made, but the S. montereyensis containing crabs were not significantly smaller than S. montereyensis without crabs. In general the destruction appeared much less severe then that caused by the behavior of some other commensal crabs, particularly those infecting bivalves, Fabia subquadrata, Pinnixa faba, P. littoralis (see Pearce, 1966, 1966a; Anderson, 1975) and Pinnotheres ostreum (see Sandoz and Hopkins, 1947; Christensen and McDermott, 1958) Larval Stages Twenty living gravid females were obtained from tunicate dissections in the course of this study. Eight of them subsequently hatched larvae. Immature eggs were maroon in color, changing to orange as they developed, and then to a light tan 4-5 days prior to hatching. One ovigerous female (9.0 x 9.0 mm) contained approximately 2700 eggs. Developing eggs overflowed the sides of the Peters and Rothman abdominal plate. A brooding female removed from the tunicate often brushed away the excess eggs with her legs and chelae, regardless of the developmental state of the eggs. The remaining eggs under the abdominal plate developed and hatched normally. Expectant females were placed in quart jars in a seawater bath, following the methods of Irvine and Coffin (1960) who were successful in raising the larva of Fabia subquadrata. When larva hatched they were divided into groups of 30-50 and placed in individual jars in the seawater bath. Depth of water in the jars and seawater bath was 15-20cm. Temperature of the bath was 10-15°0 during the course of experiment Larva were fed Artemia nauplii and had their water changed daily. All larval measurements were made using an ocular micrometer. Developement The eggs of P. pugettensis hatched directly to an advanced zoea stage. There was no free protozoea. Amolt occurred after 10-12 days to a second zoea stage. At 21-24 days this second zoea moltéd to a megalops stage. At 28-30 days this megalops molted to the first juvenile crab instar. 14 Peters and Rothman First Zoea (see fig. 8) Carapace 0.58 x 0.58 x 0.42mm; duration of stage 10-12 days. Characterized by a pronounced rostrum O.2imm long, and a curved dorsal spine 0.18mm long. Eyes sessile, 0.15 x 0.2 mm. First and second antennae 0.lomm long. Four setae on each of the second and third makillipeds, which are the longest appendages; rear thoracic appendages uniramous and undifferentiated. Abdomen six-segmented; telson 0.33 mm long with pronounced point on either side and six short spikes in center. Each abdominal segment with a pair of uniramous periopod buds 0.05-0.08mm in length. Zoea positively phototactic, found swimming at top of rearing containers. Second Zoea (see fig. 9) Carapace 0.6 x 0.6 x O.5mm; duration of stage 10-12 days. Differentiated from first stage by larger carapace, longer straight rostrum (0.Emm), and curved dorsal spine 0.3mm long. Eyes stalked and mobile, compound eye 0.19 x O.19mm, stalk O.5mm. Small ventral spike below the eyes O.15mm long. First antennae 0.15mm long with terminal setae O.05mm. Second and third maxillipeds still the longest appendages, but now each bears six terminal setae 0.35mm long. Rear thoracic appendages still undifferentiated. Abdomen six-segmented with uniramous periopods now O.imm long. Telson O.Amm long. still forking to two points each 0.15mm long; six 15 Peters and Rothman 16 central, thickened setae 0.07mm long present between the points. Zoea still positively phototactic; normally remains at bottom of jar but swims upward toward a strong light source held above the container. Megalops (see fig. 10) Carapace 0.58 x 0.58; duration 6-7 days. Eye stalks elongated and thickened, o.25mm long, compound eye 0.13 x 0.13mm. Antennule biramous, O.25mm long; antenna 0.25mm with two terminal setae O.25mm long. Chelipeds 0.34mm long with both digits of claws hooked and overlapping at tip; both chelipeds bent slightly inward at center. Last four thoracic appendages well developed. Abdomen six-segmented with telson small and rounded posteriorly. Pleopods, now used for swimming, 0.15mm long, each with 3 terminal setae O.35mm long. Megalops swam actively or settled and walked; when placed in bowls of sea water with S. montereyensis, they ignored the tunicates. Peters and Rothman Adults (see fig. 11) The only previously described P. pugettensis adult is the female (Holmes, 1900; Rathbun, 1918; Wells, 1928; and Schmitt et al, 1973). Males found during the present study differ from females in Adult numerous respects. females are characteristically adult translucent and soft shelled, males are brown and hard shelled. Females have rounded smooth chelae and legs with short trailing hairs, males have flattened legs with longer trailing hairs and their chelae are angular and sculptured to fit the contours of the front of the carapace. Female chelae are not sculptured to fit the front of the carapace. Males are characteristically smaller than females. There are at least five molts female and three in the male (see in the fig. 12). Specimens have been deposited in the Allan Hancock Foundation, University of Southern California, and the California Academy of Sciences, San Francisco. Eight males were found outside of tunicates, in our collecting bags and,aquaria where specimens of tyela were kept. No free-living males were found except these which were on or with specimens of Styela. Only one female was found outside a tunicate and that was in a stagnant bowl inadvertently left for several days -- all tunicates in the bowl were dead; four males and a female crab were found outside of the tunicates 17 Peters and Rothman 18 in the bowl. Several female crabs removed from tunicates were placed in screened boxes with S. montereyensis. They attempted to re-enter the tunicates. In one test, seven out of eight females chose the oral siphon and one chose the atrial siphon. In another experiment two females suceessfully entered a tunicate already containing another female; both intruders entered the oral siphon and were found one day later in the pharynx. When females were placed in a box with the ascidian Ciona intestinalis they ignored the tunicate, but climbed on a Styela montereyensis when offered. Only two males from nine trials successfully reentered a tunicate; they entered the atrial siphon. Of the cases of double and triple infestations, males were found with males, males with females, but no females with females were found. Discussion Pinnotheres pugettensis passes through two zoeal stages and a megalops stage. It is not known at which stage the crab invades the host. Phototactic response decreases at each successive larval stage, suggesting that, though possibly dispersed during the first zoeal stage, the larvae tend to settle towards the bottom and therefore toward their potential host. The relatively advanced first zoea and the short larval cycle of 28-30 days also suggest that P. pugettensis larvae 19 Peters and Rothman tend to remain in the micro-habitat of their host. No megalopae were ever seen inside a tunicate. This does not exclude the possibility that a megalops may enter and then molt into a juvenile crab, but megalopae and the one first stage juvenile instar collected did not appear attracted to Styela when placed in close proximity with the host. Only one crab, tentatively identified as a male, was found in the pharyngeal cavity of a Styela. On close examination there appeared to be no hole through the pharyngeal (carapace 0.9 x 1.0 wall into the atrial cavity. This crab was larger,than the first stage juvenile instar (carapace 0.58 x 0.69mm). but was the smallest crab found in a tunicate and the only one not found in the atrial cavity. It was also smaller than the smallest crab feeding hole found in the pharynx of any Styela (1.0 x 1.0 mm). Perhaps one of the juvenile instars is the first invasive stage, entering the tunicate first through the oral siphon and then breaking through the pharyngeal wall into the atrial cavity where it takes up residence. This might also explain the formation of a pocket by the invading crab, and the tendency of adult females removed from the host to reenter through the oral rather than the atrial siphon. Very small males (2.8 x 2.8 mm) were found in Styela with feeding holes, while most males found outside the tunicates were relatively large (4.0 x 4.Omm to 6.4x 6.4mm). There is one report of a male found free by F. A. Pitelka on the holdfasts of the sea grass Phyllospadix (cited in Schmitt, et al, 1973); Rathbun (1918) also reports two Peters and Rothman females that were collected free-living. No free-living females were collected or observed by the authors. The sizes of the adult females, their gross morphology, and their locomotory limitations make it hard to envision them free-living. Silas and Alagarswami, 1967, in a review of the literature find no evidence for free-living adult female Pinnotheres. The male being smaller and more motile might perhaps leave the host to breed, explaining the large number of males found outside of tunicates, though no breeding behavior was observed. Summary The commensal pea crab Pinnotheres pugettensis inhabits about 33% of the subtidal population of its ascidian host, Styela montereyensis, in southern Monterey Bay, California. No pea crabs were found in intertidal S. montereyensis. Of 247 tunicates dissected 62 contained one crab, four held two crabs and one tunicate contained three crabs. There was never more than one adult female P. pugettensis per tunicate. The crabs Iive in the in the anterior third of the atrial cavity of the tunicate, and feed by reaching into the pharyngeal cavity, through a small well-healed hole situated dorsally in the pharyngeal wall, and taking pieces of the rope of mucus and trapped food particles rolled up by the ascidian dorsal lamina, We saw no evidence of filter feeding by adult P. pugettensis. Eggs hatch as advanced zoeae, which molt to a second zoeal stage in 10-12 days, to a megalops at 20-24 days, and to the first juvenile instar at 28-30 20 Peters and Rothman 21 days. Adult males are dark brown and hard shelled, and generally smaller than the translucent and soft shelled adult females. Acknowledgement: We would like to thank Dr. Donald P. Abbott for his incredible energy and guidance, Dr. Isabella Abbott. Dr. Robin Burnett, Dr. Gary Wagenbach, Chuck Baxter and the spring students and staff of Hopkins Marine Station. Peters and Rothman 22 Anderson, G. L., 1975. The effects of intertidal height and the parasitic crustacean Fabia subquadrata (Dana) on the nutrition and reproductive capacity of the California sea mussel Mytilus californianus (Conrad). Veliger, 17 (3): 299-306. Christensen, A. M. and J.J. McDermott, 1968. Life-history and biology of the oyster crab, Pinnotheres ostreum (Say). Biol. Bull. Woods Hole, 114(2): 146-179. Dudley, P. L., 1966. Developement and systematics of some Pacific marine symbiotic copepods. 1-282. (University of Washington Press, Seattle). Fay, R. C. and J. V. Johnson, 1971. Observations on the distribution and ecology of the littoral ascidians of the mainland coast of Southern California. Bull. So. Calif. Acad. Sci., 70(3):114-124. Fiala-Medioni, A., 1978. A scanning electron microscope study of the branchial sac of benthic filter feeding invertebrates (Ascidians). Acta zool. (Stockh.) 59:1-9. Flood, P.R. and A. Fiala-Médioni, 1979. Filter characteristics of ascidian food trapping mucus films. Acta Zool. (Stochh.) 60:271-272. Garth, J. S. and D. P. Abbott, 1980. Brachyura: The true crabs: in: Morris, R. H., D. P. Abbott, and E. C. Haderlie, Intertidal invertebrates of California. 1-928. (Stanford University Press, Peters and Rothman Stanford Ca.). Holmes, S. J., 1900. Synopsis of the California stalk-eyed Crustacea. Occas. Pap. Calif. Acad. Sci., 7:1-262. Illg, P. J., 1958. North American copepods of the family Notodelphyidae. Proc. U.S. Nat. Mus., 107, 463-649 Irvine, J. A. and H. G. Coffin. 1960 Laboratory culture and early stages of Fabia subquadrata (Dana) (Crustacea, Decapoda). Publ. Dept. Biol. Sci., Walla Walla College, 28:1-24. Jorgensen, C. B., 1966. Biology of suspension feeding. 1-357. (Pergamon Press, Oxford). Pearce, J. B., 1966. The biology of the mussel crab, Fabia subquadrata from the waters of the San Juan Archipelago, Washington. Pacific Sci., 20:3-35 -. 1966a. On Pinnixa faba and Pinnixa littoralis (Decapoda: Pinnotheridae) symbiotic with the clam, Tresus capax (Pelecypoda: Mactridae): in: Barnes, H., ed., Some contemporary studies in marine science. 1-716. (Allen and Unwin, London). Rathbun, M. J., 1918. The grapsoid crabs of America. Bull. U.S. Nat. Mus., 97:82-83. Sandoz, M. and S. H. Hopkins, 1947. Early life- history of the oyster crab, Pinnotheres ostreum Peters and Rothman (Say). Biol. Bull., 93:250-258. Schmitt, W. L., J. C. McCain, and E. S. Davidson, 1973. Crustaceorum Catalogus editus a H. - E. Gruner et L. B. Holthuis. 3. Decapoda I, Brachyura I, Fam. Pinnotheridae. The Hague: Junk. 1-160. Silas, E. G. and K. Alagarswami, 1967. On an instance of parasitisation by the pea-crab (Pinnotheres sp. on the backwater clam (Meretrix casta (Chemnitz)) from India, with a review of the work on the systematics, ecology, biology and ethology of pea crabs of the genus Pinnotheres Latreille. Mar. Biol. Assoc. India Symp. Ser., 2(3):1161-1227. Wells, W. W., 1928. Pinnotheridae of Puget Sound, Puget Sound Biol. Stn. Publ., 6:283-314 —, 1940. Ecological studies on the pinnotherid crabs of Puget Sound. Univ. Washington Publ. Oceanogr., 2:19-50. 24 * . o. * s. . . . .. * 0 * .0 •* •0 9. 0 00. 0° — 0006: 10 uitu) HISNET AGOS HIVOINRI o 9 8 15 . 9. lkkkt — 0 o 9 0 o0e o0 0 o0 c 99 44 0 o (tu) HIdIM aSvavavs avas —18 2 o 00 10 100 e L ktakav- 20 9 2 o O 9 20 100 50 o 5 LE 0 G 1C olos Lelelselelee o 251 100 50 8 L 30 TUNICATE BODY VOLUME (cc) — k kk k —e o —e Jo e e ö e — - 0 o e a —o o o 291 No 9 SOOdado HIVN 20 aZaNON L 11 — — —.—— — —3 O — L 4 — T T 9 0%00 o 9089 a 8088 -0 08 o oo0 184 110 4 3. E 10 —5 (uiu) aovavavo 30 HIONaT Peters and Rothman Figure captions Fig. 1 Relation of body length to body volume in the ascidian Styela montereyensis. Fig. 2 Relation between carapace width of Pinnotheres pugettensis and body volume of Styela montereyensis Fig.3 Numbers of crabs (Pinnotheres pugettensis) and ascidicolous copepods (Pygodelphys aquilonaris) found, and percent infestation for each size class of Styela monter eyensis. N is total number of tunicates in each size class dissected. Numbers on bars indicate percent infestation. Fig 4 Numbers of male and female Pygodelphys aquilonaris found together in pharynges of individual Styela montereyensis. Number at each dot shows total number of ascidians containing each ratio. Female copepods outnumber males. 5 Diagrams of Styela montereyensis dissected by a Fig midventral cut through body wall and pharynx along endostyle and laid open, showing position and orientation of Pinnotheres pugettensis. Crab in the atrial cavity is seen through the pharyngeal wall. Hole through pharyngeal wall is within 3mm of dorsal lamina. 6 Ventral view of Styela montereyensis with pharynx Fig. laid open by a cut along the endostyle, showing hole through pharyngeal wall interrupting a pharyngeal fold next to the dorsal lamina. The crabs mouthparts are directly behind the hole. Peters and Rothman Fig. 7 a) Closeup of hole through pharyngeal wall of tyela montereyensis shown in figure 6 illustrating smooth edges of the hole. The tips of the crabs chelae are visible through the mesh of the pharyngeal wall. b) Another view of a hole in the pharyngeal wall of Styela montereyensis. Fig. 8 Pinnotheres pugettensis, first zoea. Pinnotheres pugettensis, second zoea. Fig. 9 Fig. 10 Pinnotheres pugettensis, megalops. Fig. 11 Pinnotheres pugettensis, male and female adults, dorsal and ventral views. Fig. 12 Relation of carapace length in Pinnotheres pugettensis to carapace width. Vertical lines signify points at which molting individuals were found, symbols at base of line indicate sex of molting individuals. Horizontal line indicates range of carapace width of ovigerous females. Peters and Rothman Hopkins natural history notes: 1) Approximately 350 Styela montereyensis were collected from the subtidal sites marked on the following map. 2) Styela were found to have excellent regenerative abilities, healing damage to the pharynx, mantle and tunic. Many radical operations were performed with a low mortality rate, one Styela had its tunic removed from siphons to stalk, and survived for two weeks, at which time it was sacrificed. Taking advantage of the Styelas ability to survive severe trauma, the authors sewed several windows into tunicates. A longitudinal incisison was cut through the tunic, then the mantle, or the mantle and pharynx were cut, again longitudinally. The window was then sewn into place using 000 stainless steel sutures. The window was made of 1/8 inch plexiglass with 1/32" holes drilled in its perimeter at 1/2" intervals. The window provided a clear view of the interior ofthe tunicate. The tunicates operated on survived for at least two weeks. This method is workable, but has several disadvantages, it is time consuming, care must be exercised to avoid the gonads on first incision or they will obsturct the window, and the amount of force necessary to sew through the tough tunic may disrupt the tunicate's internal order. u Su O 0 u m O 2 o. 2 0 Jo Aa M 30