Page 2 ABSTRACT Idotea resecata lives primarily on bay Zostera and Macro- cystis and is considered to be a typical bay species (Menzies, 1950). Idotea montereyensis is adapted to open coast Phyllospadix beds and is considered to be a typical open coast species. Both species have been labelled as lower intertidal forms. Tolerances to heat, salinity, and survival at low oxygen tensions were measured, and morphology of the pereaopods was also examined. It was found that I. montereyensis had longer LT-50's for the higher temperature and salinity stresses, enabling it to cope with both the lower intertidal environment of the Phyllospadix beds and also the higher intertidal tide pools into which it is sometimes washed. I. resecata has slightly greater abilities to deal with low oxygen situations, enabling it to cope with the low oxygen concentrations often found in Zostera beds. The pereaopods of I. montereyensis seem to be more adapted than those of I. resecata to gripping the substrate effectively in heavy wave action on the basis of morphology. Because I. montereyensis migrates vertically as it develops and has the physiological tolerances to cope with the range of environmental parameters associated with such a migration, it is felt that to consider it only a species of the lower intertidal is misleading. Page 3 INTRODUCTION In 1950 Menzies described the distribution of the California Idoteid isopods and while he found several species inhabiting open coastal situations, he only described one, Idotea resecata, as typically being a bay inhabitant. With eight species being described from subtidal to intertidal levels it seemed unusual that only one of these had the appropriateadaptations to inhabit bay localities. It was my intent to examine the bay inhabiting I. resecata, comparing some of its morphological and physiological adaptations to those of one of the isopods described by Menzies as a typical open coastal form. Idotea montereyensis was chosen as the open coast species to be studied because of its availability and the ease with which it could be collected. Menzies classifies both of these species as being lower intertidal, so variations due to differences in vertical distribution were expected to be minimal. In the spring, plentiful number of I. resecata live in the canopy of the Macrocystis pyrifera found off Hopkins Mar- ine Station in Monterey Bay, California. While the Macrocystis is not in the intertidal zone, I. resecata may also be found on Zostera marina which occurs intertidally at about the O.O tidal level. In these habitats, I. resecata is presumably exposed to less wave action than is present in a typical open coast rocky intertidal area. In contrast, I. montereyensis lives on the flowering plant Phyllospadix scouleri found along the open coast at approximately Page 4 the same tidal level as the bay Zostera. Here, individuals spend much of their time underwater but can be washed by wave action into tide pools at higher levels. Work by Lee (1966) seems to indicate that juveniles of this species occur in the inshore tide pool areas but move back out into the Phyllospadix beds as adults. Given these two species, one a typical bay inhabitant and the other a typical outer coastal species, it was decidedcto focus on those physical parameters that probably repre¬ sent the major differences between bay and open coastal habitats. These parameters were considered to be salinity, temperature, oxygen level, and wave action. Accordingly, data was gathered on salinity and temperature tolerances, respiration rates, and tolerance to low oxygen conditions. The peraeopods were also examined with an eye towards determing difference in holding ability under conditions of wave stress. RESULTS Tolerance to Physical Parameters--Salinity. Procedure: Concentrations of 25%, 50%, 75%, 100%, 125%, 150%, 175%, and 200% sea water were made from Instant Ocean (available from The Dolphin, San Jose). Tap water which had been kept in an open container overnight in a heat closet to drive out the chlorine was used for fresh water. 1. resecata and I. montereyensis were kept in holding aquaria in the laboratory for several days at ambient sea water temperature before tests were run. Subsequently samples of either 5 or 6 Page 5 animals were placed in finger bowls and covered by 200 ml of each of the solutions. The animals were chosen randomly and included males and females from approximately 6 mm to 25 mm in size. The bowls were placed in a 15° C constant tem¬ perature room and aerated continuously. At hourly intervals the bowls were examined and the number of live animals noted. Criteria for death were absence of pleopod movement, pleopod extension from the body, open uropods, lack of per- eaopod and antennal movement, and lack of response to mech- anical stimulation or removal from the water. Tests were then repeated four times, except in the case of 50% sea water which was repeated only once. Data from all replicates at each salinity were pooled. Results: The data from these experiments is presented in figure 1. At all salinities I. montereyensis survives at least as well, if not better than, I. resecata. In fresh water the stress is apparently too severe to allow much distinc- tion between the two species. At 25% and 50% sea water, however, the greater survival of I. montereyensis is marked. Both species seem to survive salinities equally well from 75% to 175% sea water for at least 23 hours. I. montereyens again appears to show greater survival in 200% sea water. Several of the survival curves are approximately sigmoid, suggesting that tolerance to salinity stress is normally dis- tributed. Isolated cases of mortality are due primarily to cannibalism. LT-50's at the various salinities were interpolated from the raw data and are summarized in figure 2. Both species are Page 6 euryhaline, but I. montereyensis clearly survives better at the more extreme salinities. The difference in LT-50 at both 25% and 50% sea water is quite marked. No obvious differential viability was noted relative to the size, sex, or reproductive state of the females in either species tested. Tolerance to Physical Parameters--Temperature. Procedure: To test temperature tolerance to 15° C, 22° c, 30° c, 35° C, and 40° C, finger bowls with 200 ml of 100% sea water were placed in heat closets in the dark at 30° C, 35° C, and 40° C, in a dark room at 15° C, and in a darkened box at room temperature 2° C). All bowls were aerated continuously and allowed to equilibrate to the experimental temperature for about one hour. In a manner similar to that utilized in the previous experiment, samples of 5 or 6 animals were introduced into the bowls and checked hourly to determine the number still alive. The criteria of death were the same as in the preceeding experiment. The experiments at 15°, 30°, and 35 were replicated four times while the experiment at 40' C was repeated only once. The data was pooled as in earlier experiments. Results: The pooled data is shown in figure 3. The severe heat stress of 40° C killed all animals in too short a period to show any distinction between the two species. At temperatures of 30° C and 35° C I. montereyensis again shows greater survival than I. resecata. Both species can survive temperatures of 15° C and 22° C for at least 23 hours with no significant Page mortality. Again, no differential mortality between sexes or size classes was noted. LT-50's at the various temperatures are summarized in figure 4. For the sake of comparison this figure also incorporates some of the data by Keever (1973) on another Idoteid isopod, I. stenops, a lower intertidal open coast isopod which does not migrate to higher tide levels as does I. montereyensis. Note that the I. stenops has markedly less tolerance to high temperature stress than either I. resecata or I. montereyensis. Tolerance to Physical Parameters--Ox gen. rocedure: Oxygen requirements of the isopods were investigated in two ways. First, respiration rates in oxygen saturated, filtered sea water were measured with a constant volume respirometer. This information was needed to interpret the experiments on low 0, tolerance. Eight male I. montereyensis and nine male I. resecata were used in these experiments, temperature being maintained constant at 15' C. Animals of similar size for each species were used in the experiment, ranging from about .Ol gm to about .23 gm. Second, tolerance to low oxygen was determined. Ten individuals of each species were used in each experiment. Nitrogen was bubbled through a stock solution of filtered sea water for varying lengths of time to lower the oxygen concentration. A sample was then taken and a Winkler oxygen determination performed to measure oxygen tension. Simultaniously twenty large test tubes Page 8 with animals in them were filled with the anoxic sea water and sealed. A second water sample for Winkler analysis was taken after the tubes were filled in the first few exper- iments to be sure that water of the same oxygen level was being placed into each tube. In all cases where this was done, the second sample was found to have essentially the same oxygen level as the first sample (+.02 ml 0,/ 1 sea water). The sealed tubes were checked every 10 minutes until only one or two animals were still alive, and an LT-50 for the ten animals of each species was subsequently interpolated for the particular experimental oxygen concentration. The experiment was then repeated nine times at different oxygen tensions. Results: Figure 5 is a graph of log respiration rate vs. log wet weight. While there is some indication tha a male I. resecata of a given weight may have a slightly higher respiration rate than a male I. montereyensis of the same weight, the difference is statistically insignificant. Therefore, the possible differences in oxygen levels between the bay and open coast habitats are not reflected in the respiration rates of the two species in fully oxygenated water. An experiment was performed on I. resecata to determine if body weight had any affect on time of death at a given low oxygen level. The experiment was performed in the same manner as the other low oxygen experiments except that the animals were weighed before the experiment. Figure 6 is a graph of log survival time vs. log wet weight at an oxygen concentration of .42 ml 0,/ 1 sea water. Two females and Page 9 eight males were used in this experiment and the females show no significant differences from the males. The least squares regression line has a slight positive slope, but does not differ significantly from 0. It seems, then, that weight has little affect on time of death, at least at this particular oxygen concentration, and that individual variation due to factors other than body size and sex may have greater affect on the time of death. The results of experiments on survival at low oxygen ten- sions are shown in figure 7. While the unexplained variation is not small for either species, the regression slope for I. resecata is significantly different from that of I. mon- tereyensis suggesting that if the oxygen stress is not extreme (tensions below about .60 ml 0/ 1 sea water), I. resecata may enjoy a better survival rate. When the oxygen stress is quite extreme the animals die off too soon to show a distinction between species, as was the case in the more extreme salinity and temperature stresses tested in previous experiments. Care must be taken in interpretting this data for several reasons. First, oxygen tensions above about 2.22 ml 0,/ 1 sea water were not investigated and an extrapolation of the regression lines to higher oxygen tensions is probably not valid. Further, the regression lines fitted to the data are linear. More data might reveal an exponential relationship between LT-50 and oxygen concentration although for both species the linear correlation is significant at the 1% level. Finally, it must be remembered that once the test tubes were Page 10 sealed the oxygen concentrations decreased with time as the animals respired. Consequently the data show LT-50's not at a constant oxygen concentration but rather in water which had an initially determined concentration. Differences in oxygen depletion rates both between the species and between differ- ent size classes probably accounts for some of the unex¬ plained variation, although the experiments shown in figures 5 and 6 suggest that this variation is small. The assumption is made that since the respiration rates of I. resecata and I. montereyensis are not greatly different in fully oxygenated water, as is shown in figure 5, the rates are not significantly different in low oxygen water. A further assumption is that time of death, shown not be affected much by animal size in I. resecata at an oxygen concentration of .42 ml 0/ 1 sea water, is not greatly affected by size at higher oxygen tensions. Observations made during the course of the experiment seem to bear out the assumption regarding the independence of size and time of death although the matter was not investigated directly at higher oxygen concentrations. Furthermore, death did not seem to be significantly correlated to sex or reproductive state of the females. Morphology and Behavior. Both isopod species studied are quite streamlined and are strong swimmers. They can both hold on to their usual substrate quite tightly, although some factors, such as removal of the substrate from water appears to influence their holding Page 11 ability. When a Phyllospadix blade is taken out of the water most I. montereyensis either drop off the blade or crawl back into the water. When Macrocystis is lifted out of the water, I. resecata tend to slide to the underside of the stipe or walk down it, although on occasion some may also drop off. In general, I. resecata maintains a stronger grip on its substrate when it is held out of water than I. montereyensis when it is held out of water. How the animals behave in surf is a different question, however. Holding ability in a natural situation is quite difficult to investigate and so I therefore directed my attention more towards examining the morph- ology of the pereaopods hoping to find differences in structure which would suggest differential holding ability and could be correlated with differences in wave action in the isopods' respective habitats. Figure 8 shows drawings of the 7th pereaopods of I. montereyensis and I. resecata drawn in poitions characteristic for the animals in grasping their substrate. In I. montereyensis the segments tend to be short and wide and there are more bristles on the propodus, possibly to increase frictional contact. In addition, the dactyl is more fully developed than in I. resecata. I. resecata in contrast has rather long and sparcely spaced bristles on the propodus and the dactyl is weakly produced. Characteristically, I. resecata keeps its pereaopods more extended than I. montereyensis because it is on a broader surface, either a Macrocystis blade or else the relatively large diameter stipe. I. montereyensis, on the other hand, Page 12 grips the thin, narrow Phyllospadix blades by holding them against its ventral surface with the dactyls. While I, esecata can pierce the surface of Macrocystis with its dactyl, 1. montereyensis seems to engage in a more secure stance becuase it actually encircles its substrate. The fact the 1. montereyensis is moved into higher intertidal zones, presumably by wave action (Lee, 1966), suggests that its holding ability, particularly in juveniles, which make up the predominant portion of those I. montereyensis found at higher levels, is not perfect. Nevertheless, the morphology of the pereaopods is consistant with the suggestion of a holding ability greater than that of I. resecata. This would have obvious advantages in light of its more turbulent habitat. DISCUSSION Experimental evidence has shown that while both species studied are euryhaline, I. montereyensis survives markedly better than I. resecata at salinities of 25% and 50% sea water, and somewhat better in both fresh water and 200% sea water. I. montereyensis also seems more tolerant of temperature stress, surviving better than I. resecata at both 30° and 35 C. While the evidence on oxygen stress is not clearcut, this is a suggestion that I. resecata performs slightly better than I. montereyensis at oxygen concentrations in the range of .60 to 2.22 ml 0,/1 sea water. Unfortunately a complete set of field data characterizing Page 13 the actual environmental conditions faced by these isopods is lacking. Some data in the literature, however, can provide guidelines relative to the ranges in temperature. salinity and O, tension experienced by both I. montereyensis and I. resecata. Johnson (1965) examined the temperatures in Tomales Bay, where I. resecata are commonly found living on Zostera, and reported that whereas ocean temperatures had an annual range of 12-15* C from winter to summer, the cove in Tomales Bay which he studied varied from 8-23° C during the year, a range of 15° C as compared to 3 C on the open coast. Conceivably the temperature range on the Zostera blades, where the I. resecata lives, might be even greater due to factors such as increased heat absorption due to its dark color. Some physical parameters of the kelp beds off Hop- kins Marine Station were monitored for 48 consecutive hours in the summer of 1971 (Pearse, 1971), and the daily surface temperature where populations of I. resecata are located was found to vary between about 12.5°-15° C. Three measurements on a spring day in 1973 showed the temperature during daylight hours to vary between 12-16° C. There is even less data on the habitat of I. montereyensis, but a bay Phyllospadix bed where I. montereyensis does not occur was found to have a late afternoon temperature of 15-16° C in the spring. Open coast Phyllospadix beds might be presumed to have temperatures approximately equivalent to those of the surface ocean temperatures because of the large amount of wåter movement and mixing between the intertidal Phyllospadix beds and the Page 14 offshore ocean surface. This would give the Phyllospadix beds an annual temperature variation of 12-15° C. Unfortunately there is little information on the temperature conditions of the tide pools into which I. montereyensis are periodically washed. Pearse's study showed the oxygen levels of the Macrocystis to vary considerably, ranging from 11 ml 0/ I sea water in mid afternoon to 4 ml/1 shortly before sunrise. Broekhuysen (1935) found oxygen levels in Zostera beds to ap- proach O at night and observed I. resecata swimming free of the beds under these conditions. A measurement of pre¬ sunrise oxygen levels in bay Phyllospadix showed them to be around 1.5 ml/1, whereas daytime oxygen levels in the same beds were in the meighborhood of 12 ml/1. To summarize what is known about the comparitive habitats of I. resedata and I. montereyensis, then, it seems that in addition to marked differences in wave action, bays tend to be more temperature variable than the open coast llospadix beds. The intertidal tide pools into which I. montereyensis is sometimes washed may in turn be more temperature variable than the bays, however. Oxygen levels in the kelp beds do not seem to get very low, but Broekhuysen's work indicates that I. resecata in Zostera may have to face quite low oxygen concentrations. I. montereyensis, on the other hand, is limitted to the open coast and probably does not see the same low levels of oxygen that I. resecata does. Indeed, O, concentration may be a factor limitting the horizontal distribution of I. monter- eyensis. Presumably oxygen levels in open coast Phyllo¬ Page 15 spadix beds never reach levels quite as low as those in the bay because of increased mixing due to heavy wave action. Little is known regarding salinity variation in these areas, but fresh water run-off could introduce salinity variation where there is not adequate mixing with a large salt water mass. Therefore bays might be expected to have greater salinity variation than well-mixed open coast Phyllo- padix beds. In intertidal tide pools, however, because of both increased temperature causing high evaporation rates and fresh water run-off, the tide-pool waters might be quite variable in relation to salinity. On the basis of this speculation, one might then characterize the habitat of I. montereyensis as being more variable in terms of temperature and salinity than that of I. resecata but more stable with respect to ambiant oxygen levels. Experimental results agree with this comparison of the habitats. In short, I. montereyensis does better at high salinity and temperature than I. resecata. However, I. resecata appears to withstand lower O tensions than I. montereyensis. This suggests that Menzies' original classification of the vertical distribution of the two species is misleading. He felt that both could be considered lower intertidal species, suggesting they both could be expected to have the tolerances of lower intertidal species. It has been shown, however, that I. montereyensis has the temperature and salinity tolerances to cope with the high variability of the intertidal tide pools into which it is periodically washed. Because Page 16 of this vertical migration, then, I. montereyensis should perhaps not be considered an animal typical of the lower intertidal zone. Keever's (1973) work on I. stenops lends support to this hypothesis. I. stenops is an Idoteid isopod which lives in the lowest intertidal zone among Laminaria and lower Egresia. In his study, Keever found the lowest oxygen concentration in areas frequented by I. stenops to be 4.2 ml 0,/1 sea water, and experiments showed that while this animal could sur- vive in water slowly brought to.70 ml/1, it became noticably stressed in levels below 2.8 ml/1. Both I. montereyensis and 1. resecata,however, swim normally in tubes with oxygen concentrations down to about.90 ml/1 for close to an hour. Evidently I. stenops is less tolerant of low oxygen concentra¬ tions than either I. resecata or I. montereyensis. Keever never found spring temperatures in areas with I. stenops to get above 12' C, and as might be expected, figure 2 shows that I. stenops has a temperature toler- ance less than that of either I. resecata or I. monter- éyensis. At 20' C the LT-50 for I. stenops is under 9 hours, while at a slightly higher temperature almost all I. res- ecata and I. montereyensis studied could survive for over 23 hours. In summary, the tolerance ranges of the animals studied correspond to the ranges of environmental parameters for each of their respective habitats. I. montereyensis, living in a more temperature and salinity variable environment Page 17 than either I. resecata or I. stenops, can tolerate higher levels of temperature and salinity. I. resecata, living in Zostera beds which approach low oxygen levels at night, performs better in low oxygen conditions. I. stenops, living in a more homogenious environment than that found in either the bay or in Phyllospadix beds and tide pools, has the least tolerance to extremes of all three species studied. Until more information is gathered, however. particularly field data, very little can be said conclusively about what affect the tolerance differences have on the actual distribution of I. montereyensis, I. resecata, or I. stenops. ACKNOWLEDGEMEN I would like to acknowledge Robert Keever for generously letting me use his data and Chris Harrold for being kind enough to wade into Phyllospadix beds for me at 4:30 am. I want to thank Kathy Dereimer for making sure I got out of bed for early morning collecting and Nat Howe for making so many tiresome trips to the storeroom to get materials. I am very grateful to Mrs. Connally for letting me use so many envelopes and just generally being such a sweetie. I very much want to thank Dr. Lee for providing encouragement, advice, ideas, and understanding throughout the quarter in addition to knowing an excellent Italian restaurant in Occidental, California. Finally, I would like to acknowledge everyone at Hopkins Marine Station in the spring of 1973 for making the place just what it was: a great spot to spend some time. LITERATURE CITED Broekhuysen, G. J. (1935) The extremes in percentages of dissolved oxygen to which the fauna of a Zostera field in the tidal zone at Nieuwdiep can be exposed. Arch. Neerl. Zool. 1(3), 339-346. Johnson, Ralph. (1965) Temperature variation in the infaunal environment of a sand flat. Limn. Ocean. 10, 114-120. Keever, Robert. (1973) unpublished manuscript on file at Hopkins Marine Station library. Lee, Welton L. (1966) Color change and the ecology of the marine isopod Idotea montereyensis (Maloney). Ecology. 47, 930-941. Menzies, Robert J. (1950) The taxonomy, ecology, and distribution ofnorthern California isopods of the genus Idot a with the description of a new species. Wasmann J. 3iol. 8, 155-195. Pearse, et. al. (1971) A Kelp Bed as a Classroom: Results of five week study of kelp beds in the Monterey Bay region. Unpublished manuscript on file at Hopkins Marine Station library. FIGURE CAPTIONS 1. Survival curves for I. resecata and I. montereyensis in solutions of different salinities. x- I. resecata, o- I. montereyensis. Salinities are expressed as % sea water. Experiments were terminated after 23 hours. 2. LT-50's for I. resecata and I. montereyensis in solu- tions of different salinities. Salinities are expressed as % sea water. Empty bars represent I. resecata, cross¬ hatched bars represent I. montereyensis. N=6 for 75% sea water; N=10 for 125%, 150%, and 175% sea water; N=12 for 50% sea water; N=22 for 100% and 200% sea water; and N-24 for 0% and 25% sea water. 3. Survival curves for I. resecata and I. montereyensis in different temperatures. x-I. resecata, o-I. montereyensis. Temperatures are expressed in C. Experiments were terminated after 23 hours. 4. LT-50's for I. resecata, I. montereyensis, and I. stenops in different temperatures. Empty bars represent I. resecata, cross-hatched bars represent I. montereyensis, and stippled bars represent I. stenops. Temperatures are expressed in C'. N=6 for 22°; N=10 for 40°; N=16 for 20°; and N=20 for 35 30, and 15° except for the I. stenops bar at 15% for which N=16. 5. Respiration rate as a function of wet weight in I. resecata and I. montereyensis at 15° C. x- I. resecata, o- yensis. Animals used ranged from.0186 gm to I. monter 2325 gm and respiration rates ranged from 3.60 to 20.27 microliters O,/hr. 6. Survival of I. resecata under low 0, tension as a function of wet weight. Log survival time in minutes is plotted against log wet weight in grams. Animals ranged in weight from .0195 gm to .3705 gm, and survival times ranged from 20 min to 65 min. The regression line is not significantly different from 0. Eight males and two females (denoted "F") were used in the experiment. 7. LT-50's for I.resecata and I. montereyensis at low oxygen tensions. LT-50 in minutes is plotted against initial O, concentration in ml 0/1 sea water. Each point represents an LT-50 for 10 animals. x- I. resecata, o- I. montereyensis. The regression coefficients are significantly different at the 1% level. 8. The left 7th pereaopod of I. montereyensis and I. resecata in characteristic grasping positions. 24 18 12. 6 O 24 18 12 10008088 6 L -24 V 18. 12 +e 288 O 6 O 24 12 alao00 24 eoere 18 12 6 0 ). 200 %0 75 90 50 90 2 90 100 %0 . 25 benoo Peg 3 6 9 IM 75 % 50% 250 0 O% 12 15 182124 rs. A 0 OH Og- 1 O 11 D 24 18. O t 24 18. 12. 6 O 24 18 Po 12 6. O. 24 18. 12. Og2ag2888 24 0088808 18 12. I 8 12 ( 40 ( 35 C 3090 22 16 2024 DUF . 1 OOOONODSONO + (Sanonog- C 0 10 u 2 t 0 0 1 DHIVAFNOIIValasa o0 0 11 1 -4.0 -3.0 2.0 WEIHT (gm) LOG WE 1.0 X X XX 00 0X 10 0 XX Z E Z Oul C Z 11 oX U og o e 0 .MONTEREYENSIS I. RESECATA S 1mm