Limpets on Pollicipes page 2 Introduction The limpets Collisella digitalis (Eschscholtz, 1833) Collisella pelta (Eschscholtz, 1833) and Acmaea strigatella (Fritchman, 1960) are common grazers of micro algae in the mid-tide region of the exposed rocky intertidal of central California. Examination of plates of the gooseneck barnacle Pollicipes polymerus (Sowerby, 1833) living in this range reveals an environmentally different habitat for morphologically altered forms of these and possibly other species of limpets. Though the relationship of limpets on Pollicipes has been reported and described, (Giesel 1969) little is known about the taxonomy, morphology and behavior of these limpets. The following paper discusses the problems of taxonomy, and the difference in morphology and behavior of the limpets on Pollicipes, and those found on adjacent rocks. All collection and field work was done during the Spring of 1980 at Point Pinos and at the Hopkins Marine Station. (Collections excluded Collisella scabra (Eschscholtz, 1833) and limpets smaller than 4 mm in length.) Limpets on Pollicipes page 3 Distribution in field Methods The ratio of Pollicipes to limpets was determined by counting limpets on 61 randomly selected barnacle clusters from the midportion of their tidal range at Point Pinos. Cluster sizes ranged from three individuals to 400 per cluster with 60% of the clusters having at least 50 Pollicipes. The density of limpets on rocks within four feet of Pollicipes clusters (R-limpets) was determined by counting limpets inside a surveying square (.04 m2). Fifty-six determinations were made, taking note of the fraction of each square covered by clusters or algae. The density of limpets on Pollicipes (P-limpets) was determined by measuring the plate surface area available to limpets on Pollicipes, during dissection of Pollicipes. Pollicipes were estimated to double the surface area available to limpets. Results The lowest ratio of Pollicipes to limpets was 2.67. The largest cluster without limpets was 400 Pollicipes. The mean ratio was 29.73 and the standard deviation was 34.085. No limpets were on 188 of the clusters. For density values see table 1. page 4 Limpets on Pollicipes Areas with macroalgae are more densely populated by both the P-limpets and the R-limpets. The density of P-limpets per Pollicipes is less than that of R-limpets on rocks. A qualitative analysis of the mass ratio shows it is smaller than that of the ratio of individuals. Morphology of P-limpets vs. R-limpets Methods Morphological traits of P-limpets and R-limpets were scored: A. Color: "white", "black", and "patterned" if two or more colors were present B. Curvature of bottom of shell: "flat" if no more light passed between the middle of the shell and a flat surface than any other part of the shell bottom, "slight curve" if more light passed through the middle, "large curve" if a .25 mm dissecting pin fit between the shell and the surface. C. Ribbing: "suggested" if seen only on outer surface and "ribbed" if seen from underneath D. Length, height, width, distance posterior end to apex: measured using a micrometer and expressed in ratios. E. Radular ribbons: Microscope observations using the methods of Giesel (1969) Limpets on Pollicipes page 5 F. Shell color patterns: Direct observations based on descriptions by Giesel (1969) Results The bottom edges of a greater portion of the R-limpet shells are curved, than those of P-limpets. The results are summarized in table 2. A majority of black limpets with a white apex and an owl-shaped spot in the apex have a curve. Interestingly, 80% of C. digitalis collected from above the intertidal on rock surfaces are curved, and all have an owl-shaped spot in the apex. The mean length to heigth and length to width ratios of P-limpets are significantly larger than those of the R-limpets. (Table 3) The ratio of length posterior to apex to total length of P-limpets showed the white, black and patterned P-limpets are three different groups. (Table 4) Black shells with white apex are not statistically different from the rest of patterned shells. 30 percent of all limpets had a ratio greater than .84 (forward apex), indicating they may be C. digitalis or A. strigatella, rather than C. pelta. A low mean was observed for the black shells with a white apex and lacking ribs. Twenty four percent of P-limpets showed signs of ribbing, but only 128 had ribs visible on the inside of the shell. Five percent showed signs of ribbing in combination with a curved dorsal surface and a far-forward apex, ratio greater than.84. Limpets on Pollicipes page 6 The typical color patterns described by Giesel (1969) for C. digitalis were poorly represented in the P-limpets, while C. pelta and A. strigatella features were prevalent. (Table 5) A preliminary qualitative analysis of the apex position of 79 P-limpets indicated 68 possible C. digitalis and 11 possible C. pelta. Of these, a high percentage of radulae had a high radular length to width ratio and a low percentage of radulae without a shoulder. A high percentage of radular central plates of limpets whose apex was subcentral had central plate overlap. The central plate divergence accounted for 538 of all P-limpets. (Figure 1) Orientation and distance traveled Methods The positions of P-limpets were marked in the middle intertidal zone on clusters larger than 70 Pollicipes with fingernail polish and noted again the 2nd, 6th, 7th, 8th, and 13th day. Not all clusters were checked each day, due to adverse weather conditions. The resulting limpet sample sizes were 67, 18, 9, 18, and 8 respectively. A total of 60 P-limpets were painted with fingernail polish on seven clusters no smaller than 70 Pollicipes, in exposed and protected areas. Eighty P-limpets were painted on a vertical wall with western Limpets on Pollicipes page 7 exposure and no protection from waves, on clusters of varying sizes in the middle Pollicipes range. The 140 limpets were checked every six hours for 72 hours for their presence on rocks. Results P-limpets tend to move on their clusters without leaving them. A low percentage of limpets retain their orientation for a 24 hour period. (Table 6) On the average, P-limpets are not found more than two neighboring Pollicipes away from the original after 24 hours. In two cases, two limpets moved to the same Pollicipes, and remained there for more than three days. (Figure 2 ) Feeding patterns Methods The gut contents of 60 P-limpets, 20 R-limpets and the surface of 20 Pollicipes was analysed for: Hildenbrandia sp., Ectocarpus sp. blue green algae, red algae, fungi, Enteromorpha sp., and diatoms. The rock surface was checked for Ectocarpus sp. within three feet of the clusters. Limpets on Pollicipes page 8 Results Two of the 140 P-limpets observed left the Pollicipes clusters within 72 hours. One did not return to the clusters within the period, and the other dissapeared. No other marked limpets were seen on the rock surface. The results of gut contents analysed are summarized in table /. Red algae was not observed on Pollicipes plates, and Ectocarpus was not seen in the scrapings of the rock surface. Red algae and Ectocarpus were found in both the guts of the P-limpets and the R-limpets. P-limpets and R-limpets both eat all the food types surveyed. Transplants Methods Painted R-limpets and P-limpets were transplanted to Pollicipes clusters of size 50 Pollicipes or more. The limpets remaining on each cluster were counted one day later, and an area of ! m radius was checked for marked limpets. Where controls were used, equal numbers ofdtested and control limpets were transplanted. Limpets on Pollicipes page 9 Results P-limpets can adjust significantly better to new Pollicipes under natural conditions than R-limpets. See table 8. Between 68 and 302 of R-limpets transplanted to each cluster were observed climbing onto the surrounding rock, and moving upward. Only one R-limpet moved downward on the rock. Substrate Choice Methods A Pollicipes cluster attached to its granite rock and a separate granite rock (both 30 cm diameter) were placed in an outdoor salt water tank with 108 P-limpets and 108 R-limpets around their base. Salt water spray kept the system moist and limpets were counted on each surface the 2nd, 4th, 5th, and 7th day. Results Ihe results are summarized in Figure 3. P-limpets showed a greater preference for the Pollicipes cluster than R-limpets. Fourteen percent of R-limpets and 82 of P-limpets did not choose either the cluster or the rock, and remained at the bottom of the tank. Limpets on Pollicipes page 10 Dessication Resistance Methods P-limpets and R-limpets were allowed to acclimatize for two days on a fresh surface of granite and on Pollicipes plates cleaned with NaOH and attached with Epoxy Glue to a 35 x 15 cm board. The limpets were dried under room temperature conditions for two days and subsequently sprayed with equally strong streams of water to dislodge dead and weakened limpets. The remaining limpets were counted. Results A greater percentage of R-limpets remained on both surfaces. See table 19. Low salinity resistance (seal efficiency) Methods After two days of acclimatization under moist, laboratory conditions, 33 R-limpets and 34 P-limpets on a granite rock and 89 R-limpets and 122 P-limpets on a live Pollicipes cluster were submerged in fresh water. The limpets were counted after three hours and sprayed as above after 1I hours. The remaining limpets on each Limpets on Pollicipes page 11 surface were analysed for signs of life such as a soft, contracting foot, a retracting head, and moving tentacles. Results The survival rates were greatest for P-limpets on Pollicipes and for R-limpets on the rock. See figure 4. The tight seal of limpets to surface is not released immediately upon death. The remaining P-limpets were qualitatively judged more difficult to remove from the cluster than the R-limpets. Early during the experiment up to 44% of P-limpets were observed climbing upon each other's backs, forming stacks up to 6 limpets high, while on rocks. The corresponding behavior was observed for only 3% of R-limpets. Discussion The taxonomy of P-limpets presents an interesting challenge. Though it seems that approximately 9% are C. digitalis, 40% C. pelta, and the remaining 51% a combination of A. strigatella and others, enough discrepancies exist between various morphological features to preyent clearcut identifications of individuals. The features of many white shells, particularly the position of the apex, indicate c. digitalis is the prevalent species on Pollicipes, but the same limpets lack the inner shell patterns and radular ribbon structure of C. digitalis found on rocks. The 100% lack of curvature of white shell bottoms suggests a separate group of limpets, with similar genetic Limpets on Pollicipes page 12 background. A close examination of the radular ribbons and shell color patterns of P-limpets and those described by Fritchman, (1960) clearly shows that standard morphological features are not enough for the identification of P-limpets. P-limpets have shell adaptations for living on the Pollicipes plates. The identical color patterns of shells and Pollicipes, especially noted in the upper part of the mid-tidal range serve well as protection against preditors such as Oyestercatchers, (Giesel, 1969). The narrower and higher profile of P-limpets than that of R-limpets, indicates an adaptation for living on Pollicipes plates. A narrower base may be of advantage during stress periods. A wide base would overlap the edges of Pollicipes, and cause dessication. A narrow base may further facilitate feeding from the surface of Pollicipes since less surface is covered by the limpet's foot. While individual P-limpets have adjusted to living on small surfaces, they enjoy two to fourteen as much space as the corresponding R-limpets. Pollicipes clusters act as protective living quarters, absorbing the force of waves through flexibility, retaining moisture, and providing shade. Without traveling more than 1 cm, a P-limpet finds another Pollicipes, possibly covered with microalgae, in exactly the same environmental conditions as the previous Pollicipes. The crossing between two individual Pollicipes is dangerous, because the limpet needs to extend its foot through space, and may account for the recorded loss of limpets. page 1 3 Limpets on Pollicipes The greater density of P-limpets on clusters near macroalgae suggests a relationship of P-limpets to rock cover. Giesel (1969) describes the transition of limpets 4.5 mm or larger to Pollicipes clusters. A higher initial density of R-limpets on the rock, acting as a source for P-limpets may explain the higher P-limpet density on Pollicipes near algae. Direct observations that P-limpets remain on clusters and do not leave to feed on rocks agree with most of those of Giesel (1969). Over several months, he observed P-limpets leaving the clusters only twice during cool and moist conditions. The major food types for limpets grow on the surface of Pollicipes, but are abundant only in the grooves between plates. Since limpets were observed scraping the plates both in the field, and in the laboratory, they possibly act as surface cleaners of Pollicipes plates. The preference of P-limpets for Pollicipes and their greater survival on Pollicipes during transplants and stress tests than that of R-limpets suggests P-limpets are adapted for the life on Pollicipes and benefit from it. Their low rate of survival off Pollicipes suggests a strong dependence on the clusters. A study and comparison (Conrad) of limpets on mussels, Mytilus californicus may answer more clearly the questions about predation, density, feeding Mytilus lives within the habbits and surface - limpet relationship. range of Pollicipes, has a similar surface, yet offers a larger area per individual Mytilus, and drastically differs in colors. Limpets on Pollicipes page 14 Summary 1. The density of individual limpets living on rocks within four feet of Pollicipes clusters (R-limpets ) per surface of rock is greater than that of the limpets living on Pollicipes (P-limpets) per surface area of Pollicipes plates. 2. The morphology of P-limpets differs from that of R-limpets. 3. Many standard taxonomical descriptions of limpets do not apply to P-limpets. 4. P-limpets move from an individual Pollicipes to the next, but rarely leave the cluster. 5. P-limpets scrape food from the surface of Pollicipes. 6. P-limpets adjust better to new Pollicipes clusters and R-limpets adjust better to new rocks. 7. P-limpets are attracted to clusters at a greater rate than that of R-limpets. 8. P-limpets are better adopted to survive stresses on Pollicipes than R-limpets, and R-limpets survive stresses better on rocks and unnaturally arranged Pollicipes plates. page 15 Limpets on Pollicipes Acknowledgements I am gratefull for the help received from Izzie Abbott, Don Abbott and Chuck Baxter. Many thanks are due to all staff of the Hopkins Marine Station. Above all, however, I thank Robin Burnett for his advice and especially his patience during the last four days of the spring quarter. Limpets on Pollicipes page 16 Literature Cited Fritchman 1I, Dr. Harry K, 1960. Acmaea paradigitalis SP. NOV. (Acmaeidae, Gastropoda ), The Veliger 2 (3): 53-57 Giesel, James T, 1969. On the Maintenance of a Shell Pattern and Behavioral Polymorphism in Acmaea digitalis, a Limpet, Evolution 24: 98-119 March, 1970 Limpets on Pollicipes page 17 Table Legend Table 1: Density values for limpets and the ratios of Pollicipes to limpets on various surfaces at Point Pinos, in the intertidal. Table 2: Curvature of limpet shell bottoms measured on flat surface using light and a dissecting pin (.25 mm diameter). No curve: no more light passes between the shell and the flat surface in the center of shell, than towards the anterior and posterior ends Slight curve: more light passes through center Large curve: dissecting pin fits between center of shell bottom and flat surface. Table 3: Mean and standard deviation values for limpet shell length to heigth and length to width ratios. Values are significant for p «.001 (Student's T-test). Table 4: Mean and standard deviation values for ratios of distance posterior end to apex over total P-limpet shell length. Pairwise combinations of groups I, II, and III determined statistically significant for p «.001 (Student's T-test) page 18 Limpets on Pollicipes Table 5: Percentages of 232 P-limpets showing particular color patterns, and proposed species based on the color patterns described by Giesel (1969). Table 6: A summary of limpet movement and orientation changes in terms of percent of each group observed a given number of days after limpets were marked. Table 7: Chart of type and relative amount of limpet food found in the digestive track of limpets and on the surface of Pollicipes. Table 8: Results of transplant experiments showing percent of originally transplanted limpets remaining on each surface 24 hours after the transplant occured. Table 9: Results of a dessication experiment, given in terms of percent of limpets adhering to original surface after stress period. Limpets on Pollicipes I 2 0 9 page 19 2 00 otatakaa- 5 - — * Limpets on Pollicipes page 20 Table 2 Curvature of shell bottom as seen on flat surface P-limpets R-limpets AII A B C 578 1002 88 02 No Curve 102 138 08 298 08 Slight Curve 328 298 Large Curve 08 638 1008 579 127 56 51 10 Sample Size 105 A: All-white shells B: Black shell, white apex, brown post-apical spot C: Owl-shaped spot in the apex Table 3 Ratios of shell dimentions P-limpets R-limpets 2.49 2.96 Length/heigth: mean .38 .46 s.d. 1.38 1.34 Length/width: mean .09 s.d. 208 149 Sample size Limpets on Pollicipes page 21 Table 4 Posterior end to apex over total P-limpet shell length ratio Shell type Sample size Mean S.D. .06 1 White .80 .08 11 Black .76 .06 III Patterned .72 IV Black with .72 .06 white apex 230 .76 07 Total * Shells with two or more colors in various patterns "* Shells with distinct black-white boundary Table 5 Shell marking of 232 P-limpet shells Possible Internal External color 8 Species Apex Color Apex 9 C. digitalis Variable Variable Owl-shaped spot 40 C. pelta Variable Post-apical spot Black White 4 A. strigatella Variable Blue-Wht. No Spot Blue-Wht. Variable 34 A. strigatella Variable Variable ---------- Transitional ----- 13 Any Limpets on Pollicipes page 22 Table 6 P-limpet movement and orientation change Sample size + days % on other Orientation and type (after Ist) Pollicipes Zchanged %same glost 60 disturbed 10 77 60 not " 26 60 not " 10 18 60 not " 0 11 60 not " 12 12 60 not " 10 18 Table 7 Food type of limpets Location (Gutaof limpets and surface of Pollicipes) Food type 8 of 26 % of 60 % of 20 P-limpets R-limpets Pollicipes plates 45 23 Hildenbrandia Ectocarpus Blue Green A. Red Algae 38 Fungi Enteromorpha 25 Diatoms c Limpets on Pollicipes page 22 Table 8 The reaction of limpets to new Pollicipes clusters and rock surface after transplants f and origin of limpets % of transplanted limpets present next day on Pollicipes cluster on rock (control) 20 Flat cement wall 50 5 ft. Above cluster 50 6 ft. Above cluster 18 3 ft. Above cluster 36 17 50 Cluster level (Immediate transplant) 16 50 Cluster level (kept in lab ! day) 86 88 Other clusters " No transplant performed Limpets on Pollicipes page 23 Table 19 Dessication of P-limpets and R-limpets on Pollicipes and on Rocks + limpets before Surface limpets still P-limpets R-limpets adhering after dessication P-limpets R-limpets Pollicipes plates Rock Limpets on Pollicipes page 24 Figure legend Figure 1: Results of analysis of the radular ribbons of P-limpets using techniques described by Fritchman (1960) la: Plot showing divergence patterns of central plate lb: Plot showing percentage of radulae with central plate overlap Ic: Plot showing percentage of radulae with spatulum overlap Id: Plot of radular length/width ratio le: Plot showing percentage of radulae with shoulder * Preliminary determination of possible species by qualitative analysis of apex position and other features of shell Figure 2: Results of limpet movement observation, showing percentage of limpets that moved within given number of days over a given number of individual Pollicipes. Figure 3: Plot showing result of surface preference experiment byrgiving the rate of increase and decrese of percentage of P-limpets and R-limpets on: a: Pollicipes clusters b: rock Figure 4: Results of the fresh water (seal efficiency) stress test, given in percentage of limpets remaining and surviving the stress over the given period of time and on the given surface. Limpets on Pollicipes total C. digitalis C. pelta Percentages given by Fritchman (1960) 100 80 60 8 radulae 20 EN : Parallel P: Posterior A: Anterior Figure la Page 25 Limpets on Pollicipes 100 80 8 radulae 40 20 100 80 60- % radulae 40 20 —9— Figure 1b Figure 1c page 26 Limpets on Pollicipes 100—e— 80 60- % radulae 20 100 80 % radulae -e — Figure 1d — : Figure le page 27 Limpets on Pollicipes % limpets page 28 Day1 50 S Day 5 50 25 N Day 6 50 25 Day 8 50 25 Day 13 50 25 S 2 3 4 + neighboring Pollicipes from original (in straight line) Figure 2 off Limpets on Pollicipes katatatatatata- page 29 — 1 Limpets on Pollicipes ktatatataatatataa- page 30 page 3 Limpets on Pollicipes Limpets 90 rema ining Limpets 80 remaining 70 60 8 limpets 50 40 Limpets living 20 10 f Hours after start of experiment R-limpets on rock P-limpets on rock R-limpets on Pollicipés P-limpets on Pollicipes Figure 4