1. Horseshoe crab (Limulus polyphemus) larvae abundance and distribution: patterns in a small estuary Jaymie Frederick 1, Ken Able 2, Rosemarie Petrecca 2 1 Maine Maritime Academy, Corning School of Ocean Studies, Pleasant St. Castine, ME 04420 2 Rutgers University Marine Field Station, Institute of Marine and Coastal Sciences, 800 c/o 132 Great Bay Blvd. Tuckerton, NJ 08087 The Atlantic Horseshoe crab, Limulus polyphemus, is an ancient species that is important to estuarine ecology, particularly for the food source its eggs are to shore birds (Karpanty et al., 2006) and fish (Nemerson and Able, 2004), as well as its economic importance; including bait and biomedical industries (Walls et al., 2002). Negative human impacts on the horseshoe crab population from bait fisheries and coastal development, have raised concern over the management of this species (Odell et al., 2005). In order to protect this species scientific studies need to address all aspects of its life cycle including settlement. A majority of the work has been done studying horseshoe crabs in large estuarine systems such as Delaware Bay (Botton et al., 2003; and Karpantry et al., 2006), however, an understanding of the role they play in small estuarine systems is relatively unstudied. The objective of this study is to obtain an understanding of the reproductive seasonality and distribution and abundance of larval horseshoe crabs in the small estuarine system of Little Egg Inlet, Tuckerton, NJ. L ARVAL S AMPLING On a weekly basis, a 1 meter (1mm mesh) circular plankton net was used to collect larval horseshoe crabs on the night flood tide. The net was deployed for three replicates in the mid-water column (2 meters below the surface) each for thirty minutes, off of the Little Sheepshead Creek Bridge, Tuckerton, NJ (Fig.2). Water flow was measured for each replicate tow. Salinity and water temperature were measured before the first tow and after the third tow. To look at potential spatial distribution and occurrence within the estuarine system, one sampling date in June and July, 2010additional samples were collected from two other sites; Jimmy’s Creek, and Thorofare Creek (Fig. 3). Data collected from these 2010 collections will be added to an existing data set of larval horseshoe crab abundance from Little Sheepshead Creek that originated in 2005. This data will be used to analyze seasonal and annual patterns, as well as to look for potential environmental patterns. Botton, M. L., R.E. Loveland, and A. Tiwari. Distribution, abundance, and survivorship of young-of-the-year in a commercially exploited population of horseshoe crabs Limulus polyphemus. Mar. Ecol. Prog. Ser. 2003; 265: 175-184. Karpanty, S. M., J. D. Fraser, J. Berkson, L. J. Niles, A. Dey, and E. P. Smith. Horseshoe crab eggs determine red knot distribution in Deleware Bay. Journal of Wildlife Management. 2006; 70(6): 1704-1710. Nemerson, D. M. and K. W. Able. 2004. Spatial patterns in diet and distribution of juveniles of four fish species in Delaware Bay marsh creeks: factors influencing fish abundance. Marine Ecology Progress Series 276:249-262. Odell, J., M. E. Mather, and R. M. Muth. A biosocial approach for analyzing environmental conflicts: A case study of horseshoe crab allocation. BioScience. 2005; 55(9): 735- 747. Rudloe, A. Aspects of the biology of juvenile horseshoe crabs, Limulus polyphemus. Bulletin of Mar. Sci. 1981; 31(1): 125- 133. Sekiguchi, K., H. Seshimo, and H. Sugita, 1988. Post-embryonic development of the horseshoe crab. Biol. Bull. 174: 337-345. Horseshoe crabs reproduce in Great Bay Little Egg Inlet Estuary. Larvae are available to sampling gear; this is supported by laboratory observations. In the six years of data, the peak of larval density has been in the middle of July. The spatial analysis started in 2010 suggests that larvae are distributed through out the system and as such it is likely that they play a significant ecological role. Overall, horseshoe crabs reproduce, survive hatching and early stages of development, and as such may play an important role in small estuaries such as the study site. Introduction & Objectives Results CONCLUSIONS ACKNOWLEDGEMENTS REFERENCES Special thanks to Daniel Gibson (Worchester Polytechnic Institute) for sharing his knowledge and enthusiasm of horseshoe crabs and to Jackie Toth (RUMFS) for calculating horseshoe crab densities for the plankton net data. I would also like to thank the NSF and RIOS for funding this project and supplying the materials and resourced needed to complete the project. L AB R EARING Eggs were collected from the beach next to Little Sheepshead Creek Bridge and brought back to the Rutgers University Marine Station (RUMFS) laboratory. In the laboratory eggs were kept in aerated containers (20cm diameter 7cm deep). After hatching out larvae (1 st instars) were put inflow through containers which were placed in the RUMFS flow through laboratory. Containers were marked with the egg collection date and the hatching date so that length of stage could be monitored. Individuals were measured and classified using the methods of Sekiguchi et al. (1988). The 2 nd instars, or individuals that have molted once since hatching, were fed brine shrimp. S WIMMING B EHAVIOR Swimming experiments were preformed with both larvae (1 st instars)and 2 nd instars. Fifteen trials were run for each stage during the day and another 15 for each stage at night. One individual was dropped into a 200ml graduated (Fig. 1)cylinder filled to 100ml, and watched their behavior for a period of 15 minutes. Experiments to see what influence flow rate had on behavior in the water column were performed in a 10 gallon aquarium. Three trials were run for each larvae and 2 nd instars during the day, and another three trials for each were done at night. Twenty individuals were used for each trial. Each trial consisted of two flow rates; “slow” between 5-10cm/sec and “fast” 5-20cm/sec. Materials and Methods Figure 3 Map of sampling sites; L-Little Sheepshead, Creek J- Jimmy’s Creek, and T- Thorofare Creek. RUMFS indicated by green dot. L AB R EARING Individuals in the lab were raised as far as the second instar stage (Fig. 4). Larvae ranged from 2.8 to 3.7mm; 2 nd instars ranged from 4.9 to 5.9mm. Aside from size, the 2 nd instars have longer telsons and overall shape is comparable to that of an adult (Fig. 5). On average it took 10-14 days for larvae to reach the 2 nd instar stage. S WIMMING B EHAVIOR Larvae were in the water column more than the 2 nd instars. On average the larvae were in the water column 19% of the time during the day, and 0.3% of the time at night. This finding is interesting as the literature states that the larvae are more active at night (Botton et al., 2003, and Rudloe 1981). The 2 nd instars landed on the bottom and there was no swimming behavior from any of the individuals in either the day or night trials. Larvae were more present in the water column during low flow than the 2 nd instars. The 2 nd instars were more influenced by the higher flow rate. P LANKTON S AMPLING S EASONAL AND A NNUAL V ARIATION Larval horseshoe abundance data has been collected for the last six years from Little Sheepshead Creek showing that horseshoe crabs have annually been spawning in this small estuarine system and that peak abundance in the water column is in mid-July (Fig. 6). There is large annual variation in density of larvae (Fig. 6). Some possible explanations include variation in number of spawning adults that enter the estuary system, environmental conditions, and larval availability to sampling gear. S PATIAL P LANKTON S AMPLING In the June spatial plankton samples, no larvae were caught at any of the sites. In July, larvae were found in samples from all three sites. While limited data was collected, it suggests that there could be a spatial relationship as todensity of larvae; Little Sheepshead Creek having the highest density and Jimmy’s Creek having the lowest (Fig. 7). Figure 1. Swimming behavior experiment. Figure 6. Average density of larvae for each weekly larval sampling date since 2005. Graphs show mean density + 1 s.d. to display the variation in density amongst the three tows taken each date. The y axis scale because of differences in density. Note that data for 2010 has only been collected up to July 20 th. 5.a 5.b 5.c 5.d 5.e5.f Figure 7. Mean density of larvae for all three tows at each sample site during the July spatial sampling project; + 1 s.d. See Figure 3 for location of sampling sites. Atlantic Ocean Little Egg Inlet Great Bay Figure 2. Plankton net sample off of Little Sheepshead bridge. Figure 5. 2 nd instars Figure 4. Trilobite larvae (right) and 2 nd instar (left)