The ects of Temperature and Desiccation on Pagu samuelis an! Pagurus granosimanus at Pacific urove, alifornia Richard Forward Hopkins Marine Station Stanford University Pacific Grove, California Introduction A crude survey of the hermit crab population indicated a marked lifference in the intertilal areas occupied by Pagurus samuelis (Stimpson, 1857) and Pagurus granosimanus (Stimpson, 1859). It has since been determine by concurrent work of R. Belknap and John Markham that P.samuelis ranges from O.3 to 1.3 feet intertidally and P. granosimanus from -.6 to .55 feet. Melody Bollay (1964) demonstrated that P. samuelis coul withstand gross exposure longer than P. granosimanus. This suggests that desiccation and temperature tolerance may be limiting species distribution, and it is these physical factors which I investigated. Metod The hermit crabs were obtained intertidally at Hopkins Marine Station and kept alive in aquariums supplied with running sea water. Only adult crabs as jescribed by Schmitt (1921) were used. The two species were first tested to determine water temperature tolerance and their lethal water temperature (taken as that temperature at which 508 of the crabs can live indefinitely). Twenty five crabs of each species were placed in plastic dishes, to prevent random mixing. in an aquarium. Using a Precision Thermo-Regulator the 14°0. sea water was heated to the jesired temperature in order to simulated the slow heating occuring in exposed tidepools. At intervals of 1. 4, and 8 hours after reaching this temperature the number of crabs alive was recorled. Air was pumped into the aquarium causing water circulation, thereby preventing stratification of heated water. Shell temperatures were investigated by exposing crabs out of water to bright sun light between 11 A.M. and 3 P.M. Since most P. samuelis and P,granosimanus occupy Tegula funebralis (A. Adams 1854) shells (Bollay, 1964), only these were used. A 3/16 inch hole was drilled in the upper most portion of the shell,and a small (1.5mm. temperature sensitive probe inserted. Temperatures were measured on a YSl Model 43 Single Channel Tele-thermometer. The crabs were placed in a plastic dish, whose bottom was covered by sand. Readings were taken every minute until the animal either vacated the shell or became completely inactive. To study normal body temperatures of each species I inserted the probe, used in the shell temperature experiment, into the thorax of animals(ut in half)and read temperatures on the Tele-Thermometer. Using essentially the same procedure, I determined body temperatures at death from exposure. Equal numbers of each species both in and out of the shell were again exposed in plastic dishes whose bottom was covered by sand. The time till death for each animal was recorded, and species behavior observed. At death the animals were cut in half. and the probe inserted into their thorax. Desiccation experiments were begun by comparing rates of water loss when exposed to bright sunlight in open plastic dishes. I used five crabs of each species both in the shell and out; their total weights were taken every 15 minutes. Weight loss was attributed to water loss and computed as weight loss per unit of original body weight. Since different rates of water loss were observed, I next tested the amount of desiccation tolerated by each species before death. Each crab was removed from its shell, weighed and placed in 72 individual 50 ml. glass beakers within a desiccator, whose atmosphere was kept dry by caleium chloride. The time till death and weight at death were recorded. The weight loss was attributed to water loss and calculated as percent of original body weight. Tabulations of results within species were divided into three groups, females, gravid females and males. Since similar hermit crabs are isotonic to normal and slightly concentrated sea water (Prosser, 1950, p.24) possibly they might die in sea water from which was removed that peroent of water they lost at death. Therefore I weighed and placed four orabs of each species in 1.5X sea water which I prepared by removing 38% of its original water. The number alive and total crab weights were recorded after 24 and 41 hours. Their failure to die within the concentrated sea water indicates their bodies can withstand higher salt concentrations and suggests that possibly drying of the gills could cause jeath in the desiccator. Therefore five orabs of each speices were placed in the desiccater, and at 2 hour intervala 0.02 cc. sea water was injected onto their gills by a 1/4 co. Tuberculin syringe. The time till death was recorded and compared with that for normal death by desiccation. Another possible cause of death during desiccation may be waste product accumulation, which cannot be removed in the absence of excess water. This was tested by placing three crabs of each species (in their shells) within individual air tight jars containing wet toweling (thereby keeping the atmosphere saturated and preventing desiccation). The toweling was remoistened with sea water every 10 hours, the weight of the crabs taken originally, after 24 and 41 hours, and number dead recorded at these times. Résults The death rates plotted in figure 1 and 2 show P. samuelis can tolerate high water temperatures better than P. granosimanus. More exadt temperature levels indicated the lethal temperature for P.granosimanus to be 29.7°0. and P.samuelis, 31.5°0.. Although during an inconclusive survey of exposed tidepools, the highest temperature I observed was 29.0°0., I feel higher values could be reached for short periods during a low tide on a very hot day. Thus if P.granosimanus were exposed one hour to 31°0. they would begin dying, and one hour exposure to 32.9°0. would begin killing P.samuelis. Since the two species live in an intertidal region, some ability to withstand exposure out of water is necessary. The shell temperature experiments (Table I and figure 3) were conducted under the most extreme field conditions, for crabs are rarely found exposed on dry sand, but rather are under rocks or nestled in the algae; however, the results jemonstrate ability to live under these conditions. Although not indicated by my data, shell temperatures appear to depend on the degree of sun exposure, for a passing cloud rapidly lowered the temperature two to three degrees, while its disappearance increased the temperature. The representative graph (Figure 3) shows P, samuelis more active and better able to control shell temperature for a longer time, almost double that of P.granosimanus. A successful attempt at control was considered any act by the crab which lowered shell temperature. The surprising observation was the apparent behavioral response of P samuelis to control its increased shell temperature. The crab would come partially out of the shell, begin walking,as if airing the shell's interior, and then withdraw deep into the shell, thereby 130 lowering the interior temperature. As indicated by the data (Table I) the average number of successful attempts was 6,and the average temperature drop found to be 0.5 0. P.samuelis vacated the shell either due to high temperature, or because the probe was inhibiting complete withdrawal into the shell. P.granosimanus appears to lack a behavioral control of shell temperature, for only 5 out of 10 crabs made one successful attempt at lowering the temperature. Osually its "walking-withdrawal" response increased the temperature. The shell vacating tendency also seemed absent, as 9/10 of the crabs hung half out of the shell in an apparent narcotized state at about 29.0°0., which interestingly is olose to their lethal water temperature. Body temperature experiments revealed some interesting trends. As expected of poikilotherms both species' normal body temperatures (Table II)were only slightly higher than the surrounding water temperature. When exposed without their shells, the time till death was the same for both species, 45-65 minutes (Table III). The body temperatures at death seemed dependent on exposure conditions, as the second day's values were higher than the first. But generally the mean body temperaturés of P.samuelis are higher than those of P.granosimanus. Body temperatures at death within the shell (Table IV) again depend on exposure conditions with the second days' results being higher than the first. Contrary to results without the shell, P.granosimanus generally has a mean temperature higher than P.samuelis. The times till death for P.granosimanus ranged from 65 to 90 minutes, while those for P.samuelis were from 65 to 220 minutes. This indicated o the shell acts as a protective agent against exposure. O The behavioral responses of P. granosimanus seam to correlate with those observed during the shell temperature experiment, as 2 of the 16 crabs vacated their shells, and the rest seemed to die hanging to of their body out of the shell. However only one third of the P.samuelis individuals vacated the shell while the rest withdrew tightly into its interior. This suggests the 80% exit in the shell temperature experiment was higher than normal,and probably due to the probe obstructing the animals complete withdrawal into the shell. It is surprising that P.samuelis which occupies a higher inter- tidal zone than P, granosimanus should lose more water and faster, (Figure 4amp;5) especially since other crabs living higher in the inter- tidal have acquired the ability to tolerate exposure. (Pearse 1929) This problem appears resolved when the walues for percent of desiccation (Table VæpVI) before death were determined. Clearly P.samuelis can withstand a greater water loss than P.granosimanus. Within each species the trend in amount lost goes from male, withstanding the least, to female, and then gravid females, withsta ding the greatest loss. The experiment of removing toe females' eggs demonstrated they still lost the greatest amount of water before death, which suggests a physiological change within the gravid hermit crab. The relation between weight an' water lost before death within each species generally suggests that the larger animals can tolerate a lower percentage of weight lost as water than the smaller. This correlates with the casual observation that the larger crabs are lower in the intertidal zone. If desiccation leath results from loss of body water theg might dan die when placed in 1.5X sea water, Presumably their internal body water should decrease osmotically, and likewise their weight. Table VII disproves these assumptions, for the animals did not die and did gain weight. This suggests that the maintenance of a constant amount of internal water balances the increased salt concentration. (Also the denser sea water makes the animals weigh more.) The results in Table VIII show that neither specles remains alive if only its gills are moistened. Normally P.granosimanus died from 6-11 hours after being placed in the desiccator and P.samuelis 7-134 hours. The hours till death when I moistened their gills show that both species die within the normal death time. Thus the cause of death during desiccation is questionable. Gill drying appears not to be the cause, but since the crabs seem to adjust to more concentrated sea water, body water loss may be the primary mortality factor. The data in Table IX rule out the possibility of waste product accumulation causing death, for only one P.granosimanus died after A mata, 41 hours, Tais time is much longer than the normal time interval observed for mortality within the desiccator. P.samuelis il and /3 and P.granosimanus 1 and 2 seem to lose weight after 24 hours probably because of initial weighing inaccuracies due to water held within the shell. Discussion I feel the foregoing data can be correlated by a theoretical anemals explanation for the causes of death when exposed out of the shell and within it. When exposed out of the shell both species died between 45 and 65 minutes with P.samuelis having a higher body temperature. As shown in Figure 5 the amount of body water lost at the time of death of P.granosimanus would be 14-19.5% and that of P.samuelis 16-21.54. As Table VI shows,the percent water lost before death for P.granosimanus is about the same as the amount lost in this time interval. This suggests P.granosimanus is dying from water loss, and its low temperature could be due to water evaporation. P.samuelis, although losing water faster (according to Table 7) has not lost enough water to cause death. This sugrests the crabs are dying from limited water loss combined with high temperature (as indicated by their higher body temperatures). However hermit crabs are almost never observed exposed outside their shells within the intertidal zone. Table IV shows that P.samuelis within the shell lives longer and has a lower body temperature at death than P.granosimanus. Interpolating from Figure 4 in the time interval till death P.ganosimanus should hawe lost 21.5-22.0% of its water. These high values woul' suggest lesiccation as the killer, but initially each crab stores within the shell free water which it extrudes when exposed. Therefore much of this weight loss results from extruded water, and I seriously doubt the animal has lost enough water to cause death. Thus I would attribute le th mostly to higher temperatures, (as indicated by its body temperature) combined with limite? water loss. 131 Figure 4 demonstrates P.samuelis over the time interval for death when exposed (Table III) should have lost between 31.5 and 45.0 of its body weight as water. Although they did extrude free water when exposed,these losses are still much higher than the observed desiccation death values. Since P.samuelis can maintain lower shell temperatures, probably by water evaporation, and its body temperature at death is far below the lethal water temperature, I feel P.samuelis died from desiccation while maintaining lower body'and shell temperatures by means of the behavioral response previously described. Therefore by testing the effects of physical factors, I can suggest how P.samuelis is better adapted to withstand more extreme conditions encountered higher within the intertiial zone, but not why it inhabits this area. P.samuelis as compared to P.granosimanus can tolerate higher water temperatures possibly encountered in high exposed tidepools. When exposed to the sun within the shell, P.granosimanus appears to die mainly from high tempevatures,and it is this harmful factor against which P.samuelis seems adapted. Temperature control supersedes water conservation, necessitates the ability to withstand greater desiccation and elicits the evolution of a behavioral temperature-controlling response. Summary 1. fests were performed on P.samuelis and P.granosimanus to deterwine the effects of temperature and desiccation. 2. F.sanwelis can tolerate higher water temperature than P.Kranosinanus with P. sammelis lethal temperature at 31.500. ani E.granosimanus 29.700 3. Shell temperature measurements on exposure to sunlight showed t at P.Samuelis maiatained a lower shell temperature longer an' possessed an a parent behavioral response for controlling temperature. (In response to increase temperature the animal woul' ap arently air the shell by walking and then withiraw deep into its interior, thereby lowering the shell temperature.) 4. Both species ile at 45-65 minutes out of the shell when exposed to the sun with P.samuelis body temperature at death higher than P. granosiranus. 5. When exposed within the shell P.samuelis livei longer and ha a lower body temperature at jeath than P.granosimanus. 6. Pisamuelis loses more water and faster than P.granosimanus both in and out of the shell, but t e former can lose about 27.06 (nean value) of its body weiwht as water before death, the latter oily 18.36 (mean value). In both species rales can tolerate the least loss, then females; females with egges tolerate the greatest loss. 7. Generally P.samuelis is a'apted to a higher position in the intertidal zone due to its ability to withstan' an' control temperature. Table I Shell Temperature Experiment of Attemps to control Temp. at which First attempted Temp. to control temp. P. granosimanus P.samuelis P.Granosimanus P.samuelis none 26.3°. 24.90 none 25.20 24.9 27. nne 24.1 50 2.80 23. 25.8°0. noe 26.8 ne 22. 3 mean 25.00c. mean.5 mean mean 24.3 Temp. vacate Temp. hang out of shell shell P.samuelis P.granosinanus 28.0 00c. 26.00 27.00 30.4 c 380 28. 26.90 no exit 29.( 28 26. 33 30.0 30.80 left shel! no eit mean 29.270. mean 20. Temp. "ise time time P.granosimanus P.samuelis 00 8 min. 19.3°0.-28. min. .-2 19. 22. 2°0.-27.7 21.8°0.-26 10 min. 9 min. 23.5°0.-28.80 20.2°0.-30.00 min. min. nin 23.00c.-2 min. 20.580.-28.200 80 C.-24 o0.-27.080 in. 13 min. 20.9°0.-29.0 19.0°c.-29. min. 25 mir. min. 24.5 20.0,C.-32.0 nin. 'c.-29. 23.0°0.-27.2 3.-35.590 min. min 22.0 21.5°C.-32.5 19.29 C.-29.00 11 min. min. 23.0 C.-27. 48 min. 6 min. 23.5°0.-24. mean mean values 21.2°0.-29.1°0. 11 min. 21.4°c.-28.2°0. 19 min. C Body Temperature of fagurus Under Different Conditions Table II Normal Body Temp.°. P. samuelis P. granosimanus 14.5 14.2 14.5 14.5 14.6 14.8 14.4 14.6 14.5 14.5 mean 14.5°0.. mean 14.500. Table III Boiy Temp. "0. at Death out of the Shell When Exposed to Sun Trial 41 Trial #2 (Next Day) Time till death 50-65 min. Time till death 45-65 min. P. samuelis samuelis P.granosimanus Paganosimanus 7.5 29.5 22. 25.8 28.8 3.2 25.5 24.3 26.8 25.5 27.0 26.5 26.5 27.0 25.0 24.. mean 28.2%0. Rean 27.10. 25.0 23.8 s -1 S - 24.7 25.0 25. mean 25.6°0. mean 24.200. s = 1 s 2.8 Table IV Body Temp. 90. at Death When Exposed to Sun in the Shell Trial 1 P. samuelis P.granosimanus Time till death time till death 65-220 min. 65 min. temp. temp. 24.0 26.5 24.5 25.8 26.0 vacated shell vacated shell 27.0 26.0 25.9 26.5 mean 25.0°. mean 26.3°. s =.4 Trial #2 (Next Day) P.samuelis P.granosimanus time till death time till death 65-160 min. 65-90 min. temp. temp. 28.2 28.2 18.0 vacated shell 26.5 29.0 27.9 30.0 vacated shell 29.1 29. 27.9 29.0 28.1 31.0 mean 28.4°0. nean 28.800. s =1 s =.9 Desiccation Experiment Table VI P. granosimanus Female Male lost before lost before Weight weight death death gms. gms. 1.344 1.068 16.8 13.2 1.394 .462 17.1 13.3 18.4 .960 1.490 13.8 19.3 .63. 14.0 1.140 19.6 .462 14.7 .680 335 23.0 1.140 14.9 Mean percent 19.02 .910 15.8 1.349 16.1 -1.1 1.244 16.7 16.8 1.130 .470 17.0 18.4 .998 18.6 1.007 .336 19.1 957 19.2 19.8 .972 19.9 1.829 20.7 .630 643 20.8 2. 608 Mean percent 16.83 s=2.8 Femaleswith eggs removed Weight Zlost before death 18.6 .910 .889 20.9 24.6 420 4ean percent 21.47 s=1.8 Gravid Female lost before Weight death gms. .709 14.7 .850 15.3 1.1? 15.5 1.024 15.7 584 16.9 380 17.1 17.3 .896 .863 17.4 .986 17. 1.150 18.2 18.4 .760 541 18.8 .670 19.4 1.170 19.6 20.3 1.130 .780 20.5 0.7 .990 20.8 .847 21.2 .848 399 21.3 23.0 .907 .93 24.0 26. Mean percent 19.23 s= 2.9 Table V Desiccation Experiment P. Samuelis male Female weight weight Blost before lost before gms. death death gms. 1.134 321 21.3 23.4 .726 21.4 440 25.0 22.4 .850 385 25.1 1.014 25.4 22.5 .562 .480 25.5 349 22.9 560 22.9 26.8 1.103 .763 301 27.2 395 27.4 23.4 1.162 28.6 .490 23.6 1.176 .524 .670 24.1 28.6 236 24.2 29.0 1.312 24.4 29.7 1.310 .478 25.2 413 .588 30.2 25.4 .606 Mean percent 27.38 25.5 1.312 28.2 s = 2.1 537 28.4 660 .965 28.6 28.6 29.0 .972 236 29.0 29.4 .510 29.8 .525 30.0 1.167 359 31.4 31.9 Mean percent 25.87 s =3.2 Females with eggs removed weight 3 lost before death gms. .824 28.5 .632 30.1 521 Mean percent 30.52 s =1.3 Gravid Female weight % lost before death gms. .814 24.3 .692 24.0 .958 25.3 25.4 .860 .806 1.440 31.2 629 .586 32. 440 32.5 .946 32.9 36 4 660 Mean percent 29.53 s =4.1 Placement in 1.5X Sea Water Table VII P. granosimanus P.samuelis total weight alive total weight alive time 4.075 gms. originally 3.028 gms. 4.113 gms 3.161 gms 24 hr. 4.259 gms 41 hr. 3.197 gms. Table VIII Time till death of first ten crabs under normal conditions in the desato P. samuelis P.granosimanus Hr. Hr. 30 10 121 10 134" 11 desiccator when gillare moistened Time till death P.Samuelis P.granosimanus 10t hr. hr. 104" 10 102 Table IX Moist Air Experiment P. granosimanus P. samuelis weights in the shell weights in the shell originally after 24 hr. after 41 hr after 24hr. after 41 hr. originally 5.582 gms. 5.415 gms. 5.387 gms. 5.209 gms. 5.053 gms. 5.081gms. 6.957 6.7 6.796 5.304 5.260 5.217 4.767 " 4.757 dea 4.787 7.333 7.316 7.362 C O No. Alive L Fig. N Ali O O Temp° 10 Fig. 3 C (D A Fig. 4 B u6 O O 1 8 Fig. 5 Figure 1. Water Temperature tolerance by P.samuelis. 4) Begin 14°C., Final Temperature 30.100., time to reach temperature 2 hr. 5min. (Same result for all temperatures below 30.100. B) Begin 14°C., Final Temperature 31.0°0., time to reach temperature 2 1/3 hr. C) Begin 14°0., Final Temperature 31.5 (Lethal Temperature), time to reach temperature 2 1/3 hr. D) Begin 13°0., Final Temperature 31.9°0. time to reach temperature 2 hr 34 min.. E) Begin 14'C., Final Temperature 32.8°0., time to reach temperature 2 hr. 20 min.. Figure 2. Water Temperature Tolerance by P.granosimanus 4) Begin 14°0., Final Temperature 28.9°0., time to reach temperature 14 hr. (Same result for all temperatures below 28.700.) B) Begin 14°0., Final Temperature 29.700. (Lethal Temperature), time to reach temperature 1 3/4 hr.. G) Begin 14°0., Final Temperature 30.1°0., time to reach Temperature 2 hr. 5 min. D) Begin 14°0., Final Temperature 31.000., time to reach temperature 2 1/3 hr. E) Begin 13°0., Final Temperature 31.900., time to reach temperature 2 hr. 34 min., (Same result for 32.9°0.) Figure 3. Shell Temperatures on Exposure to Sun A. Temperature rise for P.granosimanus. Plot ends as animal becomes narcotized at 29.200. B. Temperature Rise for P.samuelis. Plot ends as animal vacates its shell at 28.0°0. O Figure 4. Water Lose of Animals In Shells on Exposure to Sun 4) P. samuelis B) P.granosimanus Arrows indicate interval of time during which animals died. Figure 5. Water Lose of Animals Out of Shell on Exposure to Sun A) P.samuelis B) P.granosimanus Arrows indicate interval of time during which animals died. Bibliography Bollay, Mellody, (1964), Distribution and Utilization of Grastropod Shells by the Hermit Grabs Pagurus samuelis, Pagurus granosimanus, and Pagurus hirsutiusculus at Pacifle Grove, Callfornia, The veliger 6; Supplement: 71-76, o text figures, 1 table. Pearse, A.S., (1926), Observation on Certain Littoral and Terrestrial Animals at Tortugas Florida, with special Reference to Migrations from Marine to Terrestrial Habitats, Papers from the Tortugas Lab. of Carnegie Institute of Wash., Volume XXVI: 205-225. Prosser, O. Ladd, ed., (1950), Comparative Animal Physiology, 888pp., ed.1, W.B. Sanders Company, Philadelphia, London, p 24. Schmitt, Waldo L., (1921), Marine Decapods of California. Univ. Calif. Publ. 200l., 23: 1-470; plts; 1-50; 165 text figs.