Population Declines of Two Ecosystem Engineers in Pacific Northwest (USA) Estuaries Running head: Population declines of ecosystem engineers *Brett R Dumbaulda, Lee M. McCoya, Theodore H. DeWittb, and John W. Chapmanc a Agricultural Research Service, U.S. Dept. of Agriculture, Hatfield Marine Science Center, Newport, OR 97365, USA b United States Environmental Protection Agency, National Health and Environmental Effects Research Laboratory, Western Ecology Division, Newport, OR 97365 c Oregon State University, Dept. of Fish and Wildlife, Newport, OR 97365 *Corresponding author brett.dumbauld@ars.usda.gov 1 Abstract Population density and distribution of two species of burrowing thalassinid shrimp, Neotrypea californiensis and Upogebia pugettensis, were surveyed and compared over time in two estuaries along the West coast of the United States (USA) where these shrimp have been recognized as important ecosystem engineers. Since these shrimp construct deep burrows in the sediment, previous population studies have quantified abundance and temporal change at only a few discrete locations, potentially mischaracterizing true population trends. We used a rapid assessment of burrow openings and quantified the relationship between burrow openings and shrimp density (1.5 and 1.7 burrow openings per shrimp for Neotrypaea and Upogebia respectively) to estimate population abundance over broader scales. Burrow counts were collected using a gridded survey design in portions of these estuaries from 2006 to 2010 and compared with data collected from individual monitoring stations over a longer time frame. The Neotrypaea population on two large tide flats in Yaquina Bay, Oregon declined by 23% between 2008 and 2010 and by 48% on a large tide flat in Willapa Bay, Washington from 2006 to 2009. Upogebia had already disappeared from Willapa Bay by 2006 and declines were observed in Yaquina Bay, but the magnitude and long-term trajectory are not as clear for this species in this estuary. These shrimp population declines mirrored trends in density observed at discrete sampling locations over the same period, equate to large changes in secondary production and have likely resulted in dramatic changes to estuarine habitat and food webs. Keywords: Upogebia pugettensis, Neotrypaea californiensis, shrimp, population, burrows, map, GIS 2 Introduction The ghost shrimp Neotrypaea californiensis (hereafter Neotrypaea), and the mud shrimp, Upogebia pugettensis (hereafter Upogebia) are common inhabitants of intertidal soft sediments in NE Pacific estuaries from southeastern Alaska, USA to Baja Mexico (MacGinitie, 1930, 1934). Although occasionally found at low density, the majority of these shrimp occupy well defined dense beds (often exceeding 400 shrimp m-2) in estuaries from Northern California, USA to British Columbia, Canada (Bird, 1982; Swinbanks and Luternauer, 1987; Dumbauld et al., 1996, DeWitt et al., 2004). Shrimp beds occupy significant portions of the intertidal in many of these estuaries: >80% in Yaquina Bay and Salmon River estuaries in Oregon (DeWitt et al., 2004), 23% in Alsea Bay, Oregon (Chapman and Carter, in press), and 19% (4,300 ha) of the intertidal in Willapa Bay, WA (Dumbauld, unpublished data). Burrowing thalassinidean shrimp are important ecosystem engineers (sensu Jones et al., 1994; Berkenbusch and Rowden, 2003) because in the process of constructing and maintaining their burrows they greatly modify the substrate, enhance nutrient and carbon fluxes (Webb and Eyre, 2004; D'Andrea and DeWitt, 2009; Jordan et al., 2009; Pillay and Branch, 2011, Volkenborn et al., 2012), alter habitat structure by displacing seagrasses (Suchanek, 1983; Dumbauld and Wyllie-Echeverria, 2003; Berkenbusch et al., 2007), and change the composition of infauna, epifauna and demersal nekton in the system via burial and sediment disturbance (Ronan, 1975; Peterson, 1977; Posey, 1986a; Posey et al., 1991; Dumbauld et al., 2001; Berkenbusch and Rowden, 2007; Ferraro and Cole, 2007). Burrowing shrimps also directly alter local food webs by consuming phytoplankton and benthic microphytobenthos (Griffen et al., 2004; Abed-Navandi and Dworschak, 2005), modifying the abundance of benthic macrophytes, and serving as prey themselves for larger 3 consumers such as crabs, birds, fish and whales (Stevens et al., 1982; Posey, 1986b; Weitkamp et al., 1992; Harada and Tamaki, 2004; Dumbauld et al., 2008). These shrimp also directly affect shellfish aquaculture operations in the estuaries where they co-occur, since the shrimp’s burrowing activity suspends sediment and destabilizes the substrate which buries or smothers sessile bivalves. As a consequence a program to control shrimp on commercial oyster beds using a pesticide has been in place for over 50 years in coastal estuaries of Washington state USA (Feldman et al., 2000; Dumbauld et al., 2006). Thalassinidean shrimp abundance is difficult to measure directly because the shrimp excavate elaborate burrows with multiple openings to the sediment surface and the galleries of some species can extend to over a meter deep in the substrate (Thompson, 1972; Griffis and Suchanek, 1991; Felder, 2001; Kinoshita, 2002; Coelho, 2004). We have sampled with large cores to track density and population structure of Neotrypaea and Upogebia at several discrete locations for over 20 years in Willapa Bay, Washington and for six years in Yaquina Bay, Oregon and several other estuaries along the west coast of the USA. After several years of stable or increasing density, Neotrypaea abundance began declining in 1995 and Upogebia abundance declined dramatically after 2000 at our sampling locations in Willapa Bay. The decline in Upogebia was attributed, at least in part, to the coincident increase in prevalence of an introduced bopyrid isopod parasite Orthione griffennis which causes infected females of this shrimp to be unable to reproduce (Markham, 2004; Smith et al., 2008; Griffen, 2009; Dumbauld et al., 2011; Chapman et al., 2012; Chapman and Carter, in press). However this isopod does not infect Neotrypaea and there are a number of potential factors that could be responsible for these changes in both species including low recruitment, predation, habitat loss and sampling error. Given their wide 4 distribution, high density, commercial impact and especially their ability to dramatically alter ecology at the estuarine landscape scale, our primary focus here was to test whether trends observed at individual locations reflected larger scale changes in estuarine shrimp populations or instead were the result of contraction or movement of patches or beds within these systems. The coring technique used to sample these shrimp at discrete locations would be prohibitively time consuming and difficult to adopt for broader scale surveys of estuarine shrimp populations so we measure larger estuarine ecosystem level trends directly in this paper by first quantifying the relationship between the abundance of burrow openings made by Neotrypaea and Upogebia and the number of shrimp taken in cores at depth and then rapidly and comprehensively surveying burrow counts over broad areas. We re-surveyed populations of these species in large portions of both Willapa Bay, WA and Yaquina Bay. OR and quantified changes in population density and distribution over time to determine whether estuary-wide population changes were comparable to temporal trends in density observed at our discrete long term monitoring sites. We then discuss the relevance and potential magnitude of these changes to coastal estuarine ecosystem structure and function and to management efforts for shellfish production. Methods and Materials Study Sites This study was conducted in Yaquina Bay, Oregon (44o N, 124oW), and Willapa Bay, Washington (46 o N, 124 o W, Fig 1), drowned river valley estuaries located along the Pacific 5 Northwest coast of the continental United States. Forty-eight percent (8.2 km2) of the Yaquina estuary and 68% (227 km2) of Willapa Bay consists of intertidal sand and mudflats that are regularly exposed on semidiurnal low tides. Both species of shrimp occur at low density in subtidal areas (<50 m-2; T.H. DeWitt, unpublished data), but their distribution is thought to be limited mostly to intertidal areas by predation (Posey, 1986b). Upogebia densities are highest in the low to mid intertidal zone and Neotrypaea are most abundant in the mid to upper intertidal (up to 400 shrimp m-2 ; Bird, 1982; DeWitt et al., 2004; Dumbauld et al., 2011). Sampling at long-term monitoring sites Population structure and local density was quantified by collecting cores annually in dense shrimp colonies. Neotrypaea were collected from two sites in Willapa Bay; near the Palix River (1988 – 2009) and near Stony Point (2009-2010), and from one location on Idaho Flats in Yaquina Bay (2005 – 2010; Figure 1). Upogebia were collected from two sites in Willapa Bay; near the Cedar River (1988 to 2004 and 2007) and near Goose Pt. (2003 - 2009), and from one location on Idaho Flats in Yaquina Bay (2005 – 2010, Figure 1). The number of burrows at the surface was recorded within the area encompassed by each of 10 cores (60cm depth x 40 cm diameter) and the content then excavated and sieved (3-mm mesh) to collect all shrimps. Species, gender, size (carapace length, CL in mm), and presence of parasitic bopyrid isopods were recorded for all shrimp collected. Burrow Count to Shrimp Relationship 6 In order to efficiently determine and map the distribution and density of shrimp at the estuarine landscape scale we needed to quantify the relationship between burrow openings at the surface and the actual number of shrimp at depth. Two sources of data were used to calculate this relationship: 1) cores collected annually in dense colonies of both Upogebia and Neotrypaea at Idaho Flats from 2005 to 2010 (described above) and 2) cores collected at 42 stratified random sample locations on Sally’s Bend and Idaho Flat in Yaquina Bay in 2008. The 2008 survey was designed to capture a broader range of burrow and shrimp densities, since the annual single monitoring site surveys represented only dense shrimp colonies. These 2 sources of data were combined to create the most representative burrow to shrimp relationship. Broad Scale Shrimp Population Surveys We used a gridded survey design to visit 315 sites located at the vertices of a 100 X 100 m grid distributed across the two major tide flats, Idaho Flat and Sally’s Bend in Yaquina Bay in 2008 and 2010. A similar survey design was used to visit 230, 144, and 206 sites in 2006, 2009, and 2010 respectively in Willapa Bay where we used a larger 200 X 200 m grid across two major tide flats, Ellen Sands and Stony Point Sands. The areas surveyed for this study represent only a portion of the total shrimp population in each estuary because it was not feasible to map the entire distribution of shrimp every year. We used a hovercraft and/or visited each site on foot using a TrimbleGeoXT handheld GPS running Terrasync v. 4.0 (Trimble Inc., Sunnyvale CA) to navigate to each grid point. At each grid point, the number of burrow openings in a 0.25m2 quadrat was recorded; the contributing percentages of Neotrypaea, Upogebia, bivalves, and other large burrowing infauna estimated visually; and the dominant habitat characteristics and 7 qualitative sediment type (mud, sand, sandy mud, muddy sand) was recorded. Bivalve and polychaete worm burrows were counted only if their size was large enough to be confused with Neotrypaea or Upogebia burrows. Burrow inhabitants were qualitatively confirmed in the field by directly sampling with “yabby” pumps or by expert opinion based on burrow characteristics – lining, shape, elevation, topology, feces, and sediment. Survey data were downloaded, post processed, and exported as an ESRI shapefile using Trimble GPS Pathfinder Office v. 4.20 (Trimble Inc, Sunnyvale, CA). We combined the 2008 and 2010 survey data for Yaquina Bay using a Spatial Join (25m) in ArcMap 9.3 (ESRI, Redlands, CA) and added a field to designate the location (Sally’s Bend or Idaho Flat) for each of the sites. The 2006, 2009, and 2010 data for Willapa Bay were combined using R (R Core Development Team, 2012). All data were imported into R for analysis and maps were generated using the rgdal (Keitt et al., 2012) and raster (Hijmans and van Etten, 2012) packages. Data analyses Population change The average densities of shrimp found at long-term monitoring locations each year were compared using Welches t-test (no equal variance assumption), and the magnitude and direction of change in population densities were assessed using linear regression. The slopes and a 95% confidence interval (CI) of the slope estimates for relationships between burrow counts taken in 2006, 2009, and 2010 for Neotrypaea in Willapa Bay, and in 2008 and 2010 for both Neotrypaea 8 and Upogebia in Yaquina Bay were compared to a 1:1 relationship. If the confidence interval of the linear fit did not overlap the 1:1 line then the population was assumed to have undergone a significant change (population growth if above, and decline if below). The same locations were visited in 2008 and 2010 in Yaquina Bay. In Willapa Bay, the majority of sample sites overlapped in the 2006, 2009, and 2010 survey, but the number and extent of sites were different each year. Burrow count changes were therefore only compared at locations where data were taken in both of the years being compared. Total Burrow and Shrimp Estimates We applied the combined burrow to shrimp relationship across interpolated (inverse distance weighted) burrow distribution and density maps to estimate the abundance of shrimp occurring in the sample areas of Willapa Bay and Yaquina Bay each year. Although these estimates are based on a portion of the total estuary and therefore represent the total estimated burrows and shrimp in the sample area and not the total shrimp in each estuary, they were used to demonstrate the magnitude of population change at the estuary scale. Additional Analyses The density and distribution of both Neotrypaea and Upogebia in Yaquina Bay were also surveyed in 2002 (DeWitt et al 2004, unpublished manuscript). Different mapping techniques precluded direct comparison of categorical transect data collected in 2002 with the continuous grid data collected in 2008 and 2010. To allow comparison, we extracted the estimated species 9 and density designations from the 2002 map at the locations visited in 2008 and 2010 (see supplementary material S1 for method details and results). The combined burrow to shrimp relationships for each species were then applied across the predicted burrow distribution and density maps to estimate the abundance of both species occurring in the sample area of Yaquina Bay and extend temporal comparisons for these populations back to 2002. Results Population change at long-term monitoring sites Neotrypaea density at the Palix River site in Willapa Bay, Washington increased between 1988 and 1995 to a high of nearly 500 shrimp m-2 and then declined to fewer than 70 shrimp m-2 in 2009 (Figure 2). Shrimp disappeared from our standard monitoring area and became increasingly patchy at the scale of several hundred meters at this site making coring impractical. In response, we moved our monitoring location to Stony Point Sands in 2009, where mean Neotrypaea density was comparable, but where the distribution of shrimp was more homogenous and extensive. Focusing on the years when we performed broad scale mapping, a significant decline in the mean density of shrimp at the Palix River site was observed between 2006 (166 Neotrypaea m-2, Table 1) and 2009 (55 Neotrypaea m-2). The decline was not significant at Stony Point Sands between 2009 (67 Neotrypaea m-2) and 2010 (61 Neotrypaea m-2, Table 1). Upogebia density at the Cedar River site in Willapa Bay exhibited broad fluctuations between 1988 and 2001, but declined to zero in 2003 and remained at zero in 2004 and 2007. In 2003 we 10 moved our sampling location for Upogebia to a site near Goose Point where the density was measureable, but by 2006 this population declined to near zero as well (Figure 2). Neotrypaea density on Idaho Flat in Yaquina Bay, Oregon declined from 388 shrimp m -2 in 2005 to 286 shrimp m-2 in 2010 (Figure 3), but shrimp remained dense in comparison to the Willapa Bay sampling locations (Figure 2). Focusing on the years when we performed broad scale mapping, Neotrypaea density at our long-term sampling site declined significantly from 350 shrimp m-2 in 2008 to 286 shrimp m-2 in 2010 (Table 1). Upogebia density increased from 176 shrimp m-2 in 2005 to 375 shrimp m-2 in 2007 and then declined to 199 shrimp m-2 in 2010. The increase in Upogebia density observed in 2007 was apparently due to a significant recruitment event in 2006. The majority of small (0 - 4 mm CL) newly recruited shrimp likely passed through our sieves in 2006 and the shrimp sampled in 2007 (measuring 10-20 mm CL), represented one year old animals (Figure 4). Focusing on the years we performed broad scale mapping, Upogebia density declined significantly between 2008 and 2010 (291 shrimp m-2 to 199 shrimp m-2respectively). Prevalence of the parasitic bopyrid isopod Orthione griffenis which occurs only in reproductive sized Upogebia (Dumbauld et. al 2011), declined from a high level of 73% in 2006 to 20% in 2010 (Figure 4). There was a significant relationship with a positive slope between burrow counts and the number of shrimp extracted in the cores. We observed 1.52 burrow openings shrimp-1 (CI = ±0.14, R2 = 0.78) for Neotrypaea and 1.69 burrow openings shrimp-1 for Upogebia (CI = ±0.11, R2 = 0.88, Figure 5). Broad scale population change 11 Broad scale survey results showed the population of Neotrypaea on Ellen Sands and Stony Point Sands, in Willapa Bay declined significantly between 2006 – 2010 with model fits and confidence intervals for regression of year-to-year burrow density measurements having slopes significantly less than 1 (2006-2009 = 0.41 ± 0.07, 2009-2010 = 0.58 ± 0.10, Figure 6). The population of Neotrypaea on Idaho Flat and Sally’s Bend, in Yaquina Bay also declined significantly between 2008 and 2010 (slope = 0.69 ± 0.04, Figure 7). The population of Upogebia on Idaho Flat and Sally’s Bend, in Yaquina Bay showed a smaller decline between 2008 and 2010, but regression model fits and confidence intervals indicated statistical significance (slope = 0.76 ± 0.05, Figure 8). Extraction and analysis of 2002 data (supplementary material S1), indicates that the decline in density and abundance of both Neotrypaea and Upogebia in Yaquina Bay predates our 2008 survey with substantially more shrimp present in 2002 (supplementary material Figure S1, Table S1). Shrimp population size Populations of Neotrypaea and Upogebia on Idaho Flat and Sally’s Bend in Yaquina Bay and Ellen/StonyPoint Sands in Willapa Bay were estimated by interpolating burrow counts over the surveyed areas and using the burrow count to shrimp density relationships described above. In Yaquina Bay, an estimated 47.8 million (M) Neotrypaea burrows in 2008 translates to 31.5 M Neotrypaea (Table 2). The lower burrow counts in 2010 result in an estimate of 24.3 M Neotrypaea – a 23% decline. The relationship for Upogebia results in an estimate of 274 M burrows in 2008, which translates to 162 M Upogebia. The lower burrow counts in 2010 result in an estimate of 151 M Upogebia – a 7 % decline. For Willapa Bay, an estimated 539 M 12 Neotrypaea burrows in the overlapping mapped areas in 2006 translates to 356 M Neotrypaea. The lower burrow counts in the same areas in 2009 and 2010 result in estimates of 186 M and 149 M Neotrypaea – a 48% and 20% decline respectively. Discussion Quantifying the population abundance of large deep burrowing benthic marine organisms like polychaetes, bivalves, and thalassinidean shrimp at appropriate scales presents numerous logistical challenges to ecologists and managers. Consequently, these ecologically important organisms are either overlooked or under-sampled due to time and monetary constraints. Rare exceptions involve surveys of individual sites over long time periods or broader surveys repeated at very disparate points in time (see Reise et al., 2008 for examples). The primary goals of this research were to conduct broad scale surveys of thalassinidean shrimp populations in two eastern Pacific estuaries, compare the maps produced to obtain and evaluate temporal trends in shrimp distribution and abundance, and contrast these population trajectories with those from data collected at single long term monitoring locations. We then use these results to evaluate the potential magnitude and relevance of these changes in shrimp populations to coastal estuarine ecosystem structure and function and to management efforts for shellfish production. Neotrypaea and Upogebia population trajectories We observed broad scale population declines of both species of burrowing shrimp in both estuaries and these declines mirrored declines in shrimp density observed at single monitoring 13 locations within dense shrimp populations. Comparisons of the burrow density maps for the two largest tide flats in Yaquina Bay, Oregon revealed that the Neotrypaea population declined markedly (-71%) from 2002 to 2008 and by an additional 23% between 2008 and 2010. Neotrypaea density also declined from 350 shrimp m-2 in 2008 to 286 shrimp m-2 in 2010 (-18%) at our single Idaho Flat monitoring location in Yaquina Bay. In Willapa Bay, the density of Neotrypaea declined from 166 shrimp m-2 at the Palix river monitoring site in 2006 to 55 shrimp m-2 in 2009 and the magnitude of the drop (-67%) was similar to that observed in the broader population mapped over Ellen/Stony Point Sands (-48%) in the same years. Declines were also observed in both density and population size of Upogebia in Yaquina Bay, although the magnitude and long-term trajectory were not as clear for this species. Upogebia populations declined by 64% between 2002 and 2008 and by an additional 7% between 2008 and 2010. Density at our Idaho Flats monitoring location increased between 2005 and 2007, and then decreased between 2007 and 2010 to approximately the 2005 density (ca. 200 shrimp m-2). The increase in Upogebia density in 2007 was due to a large recruitment event of juvenile shrimps at this location in 2006 (Figure 4, and see Dumbauld et al. 2011). Precision of the broader population estimates for both shrimp species should be treated with some caution because they are based on interpolations of widely-spaced survey points, but we are confident in the direction and relative magnitude of the population trajectories. Relatively large fluctuations in density of both burrowing shrimp species have been previously reported at discrete locations in several NE Pacific estuaries. Burrowing shrimp have been a documented problem for shellfish growers since the late 1920’s (Stevens, 1929), but became a serious issue after an El Nino event in 1957 and a large increase in their populations. 14 Washington and Oregon shellfish growers subsequently sought relief and eventually implemented a treatment program on their beds utilizing the pesticide carbaryl (Fisheries and Ecology, 1985; Feldman et al., 2000). Bird (1982) reported 250 Neotrypaea m-2 in Yaquina Bay in 1978 and a subsequent decline to 130 shrimp m-2 in 1980. He also documented a recruitment event in 1978 that resulted in large increases in Neotrypaea densities in Siletz Bay and Alsea Bay, Oregon over the same time frame. Though commercial bait fishery landings could be influenced by other factors, Chapman and Carter (in press) report declines in Upogebia harvested since the 1990’s in several Oregon estuaries. In Willapa Bay, we document that Neotrypaea density increased during the early 1990’s and then declined from over 400 shrimp m-2 in 1995 to fewer than 50 shrimp m-2 in 2010, while Upogebia density fluctuated around 100 shrimp m-2 in the mid 1990’s until the population dropped to almost zero in 2002 (Figure 2). Together these records suggest that population sizes, densities, or areal coverage have increased and declined substantially and perhaps synchronously in the past, and that large-scale fluctuations in density may be a recurring feature of these shrimps’ population dynamics. There are a suite of potential factors that could be responsible for changes in shrimp populations including variable recruitment, predation, habitat loss and sampling error. Our results confirm that trends observed at individual locations over the last 2 decades reflected actual larger scale changes and were not the result of sampling error due to contraction or movement of patches or beds of shrimp within these systems. The declines and local extinctions in estuarine Upogebia populations from British Columbia, Canada, to Morro Bay, California have previously been associated with an introduced parasitic 15 bopyrid isopod, Orthione griffenis (Dumbauld et al. 2011; Chapman et al. 2012). This isopod severely reduces the fecundity of adult female Upogebia in proportion to its prevalence in these shrimp which varied from 20% to almost 100% in both Willapa Bay and Yaquina Bay populations we studied (from 2005 through 2010; also see Figure 4). These shrimp have pelagic larvae that leave the estuaries, develop in the nearshore coastal ocean, and then must return to estuaries and recruit as postlarvae. This lost reproductive output would be expected to reduce shrimp larval recruitment throughout the region and potentially be correlated with adult abundance in a given estuary, especially if larvae on average return to their natal estuary or nearby estuaries and/or if the estuary is isolated from others. It was not clear however from our annual density assessments at the single Idaho Flats location, why the Yaquina Bay Upogebia population had not declined as much as the Willapa Bay population. The concurrent broader population assessments that we present here suggest that this population was in fact declining dramatically at multiple sites within the estuary from 2002 – 2010, despite a significant recruitment event in 2006 and roughly maintaining its spatial footprint within the estuary. Genetic studies are underway to test whether recruiting larvae originate from natal estuaries, but surveys suggest that Yaquina Bay Upogebia remain one of the only significant source populations in the larger coastal Upogebia metapopulation (Chapman et al. 2012). Larval settlement and recruitment to surrounding and perhaps distant estuaries could currently depend on near-shore ocean circulation, the prevalence of Orthione griffenis, and the abundance of Upogebia in this single estuary in a given year. There can be no similar explanation, however, for even larger declines in Neotrypaea populations. The native parasitic isopod, Ione cornuta, infests and also effectively castrates 16 Neotrypaea, but is an order of magnitude less abundant than Orthione (< 1% prevalence, Dumbauld et al. 2011). Neotrypaea populations declined in both estuaries and in areas well outside oyster aquaculture beds in Willapa Bay, ruling out any influence of pesticide treatment for shellfish culture in that estuary. Further, though previous fluctuations in Neotrypaea density have been observed, Neotrypaea has not experienced widespread population crashes like Upogebia. While there have been recent Neotrypaea recruitment events in both estuaries (2010 in Yaquina Bay and 2012/13 in Willapa Bay), very low recruitment was observed in Willapa Bay from the mid 1990’s through 2011 and in Yaquina Bay from 2004 to 2010 (Dumbauld et al. 2004; Dumbauld et al. 2006; Dumbauld et al. 2011; Dumbauld unpublished data) and this coincides with decreased densities of larger and presumably older shrimps (Bosley and Dumbauld, 2011). Population declines of these two shrimp species may thus be due to coastal ocean conditions that influence either larval survival or larval transport into estuaries, and the effect merely exacerbated for Upogebia by the presence of Orthione. Inter-annual fluctuations and longer period regime shifts in atmospheric and coastal ocean conditions dramatically influence recruitment and year class strength of other invertebrates along this coast (Shanks and Roegner, 2007; Morgan et al., 2009a; Morgan et al., 2009b; Morgan and Fisher, 2010; Yamada and Kosro, 2010) and thalassinidean shrimp populations elsewhere (Wooldridge and Loubser, 1996; Tamaki et al., 2010). Climate cycles could alter current direction and duration in the coastal ocean as well as water chemistry in both the ocean and receiving estuaries during the approximately three and eight-week larval period that Upogebia and Neotrypaea respectively reside in the meroplankton (Hart, 1937; McCrow, 1972; Dumbauld unpubl. data ). However, we witnessed a 17 large Neotrypaea settlement event in Yaquina Bay in 2010 including an order of magnitude greater abundance of post-larvae entering the estuary than settled on the tideflats suggesting that post-larval competence, transport to the tideflats, or predation during the recruitment process could also be important. Ecosystem scale consequences of declining burrowing shrimp populations Since Neotrypaea and Upogebia act as ecosystem engineers (sensu Jones et al. 1994), declines in their populations must also change ecological processes and ecosystem services including nutrient cycling and organic matter processing, food web dynamics and fisheries production, infaunal and nekton biodiversity, and shellfish farming and harvest. Proportional loss estimates of 78% and 53% of the Neotrypaea and Upogebia populations respectively from 2002 – 2010 on the two largest mudflats in Yaquina Bay, correspond to losses of 86 M Neotrypaea and 167 M Upogebia on these tideflats over the 8 year period (Table 3). These changes equate to a loss of 998 tonnes (t) of Upogebia and 140 t of Neotrypaea using an average size distribution (from annual samples) and wet weight to carapace size relationship from the literature (Table 3). A 58% loss of Neotrypaea from 2006 – 2010 on a portion of the largest mudflat dominated by these shrimp in Willapa Bay translates to a loss of 207 M shrimp or 2,140 t. We surveyed the entire tideflat in this estuary in 2006 and estimate that there were 1.8 billion or about 18,356 t of Neotrypaea present. Though cultured oysters and clams also contribute substantially in Willapa Bay (a conservative estimate based on annual harvest is 330 t yr-1; Ruesink et al., 2006), these shrimp likely represented the largest single contributor to estuary secondary production in most estuaries along the US west coast before the declines we document here. For example the 18 biomass of adult native clams in all intertidal areas of Yaquina Bay is approximately 1,212 t (based on 2012 Yaquina Bay intertidal survey, A. D’Andrea, Oregon Dept. Fish and Wildlife, pers. comm.) versus a total of ~2,079 t of burrowing shrimp on just the two largest tideflats in 2002, but declining to just 942 t in 2010 (Table 3). Water filtration and phytoplankton-removal rates for Upogebia are dependent on shrimp-size, shrimp-abundance, and particle-concentration (Griffen et al., 2004), and these authors estimated that Upogebia populations present in Yaquina Bay in 1999 were capable of filtering the entire body of water overlying their burrows on a daily basis. A loss of 53% of this filtering capacity between 2002 and 2010 could result in lower water clarity, but a potential increase in the availability of phytoplankton to other fauna. D’Andrea and DeWitt (2009) demonstrated that dissolved inorganic nitrogen and carbon fluxes from the sediment to the water column were enhanced in a population density-dependent manner by Upogebia, with high-density patches of the shrimps producing disproportionately large effects on rates of organic matter re-mineralization (e.g., decomposition) in sediments and attendant efflux of DIN and DIC to the water column. They proposed that the surface area of Yaquina Bay mudflats was increased by 1.5 to 15 times by Upogebia burrows over which these enhanced fluxes occurred. Although the density-dependent functions have not been fully developed (DeWitt et al. 2004), Neotrypaea have also been shown to intermittently irrigate and oxygenate their burrows (Volkenborn et al., 2012) and populations would be expected to significantly influence sediment biogeochemistry and microbial diversity (DeWitt et al., 2004; Bertics et al., 2010). We estimate that the burrow lining surface area for both species of shrimp combined would have declined from 17.8 km2 to 8.0 km2 on the two largest tideflats in Yaquina Bay between 2002 and 2010, a 55% loss of exchange area for those biogeochemical processes (Table 3), yet the remaining area is still double that of the actual mudflat surface (3.5 km2). 19 Finally, loss of these shrimp would also be expected to substantially change estuarine habitat and food webs because both shrimps, but particularly Neotrypaea, are strong bioturbators. Tubebuilding polychaetes and crustaceans, bivalves and other sedentary species are excluded from areas where these shrimp are abundant due to the frequency of sediment disruption, resuspension of fine particles which can clog gills, and decreased sediment compaction (Ronan, 1975; Peterson, 1977; Posey, 1986a; Posey et al., 1991; Dumbauld et al., 2001). Thalassinidean shrimp bioturbation also directly influences the presence of seagrass habitat (Brenchley, 1981; Suchanek, 1983; Dumbauld and Wyllie-Echeverria, 2003; Berkenbusch et al., 2007), and Ferraro and Cole (2010, 2011) used their presence to define unique habitats that distinguish benthic macrofaunal and nekton communities in several Pacific Northwest estuaries. Decreases in the density and distribution of populations of both shrimp species has thus likely resulted in changes in the structure of infaunal invertebrate and epifaunal nekton communities, and potentially increased the spatial distribution of both native and introduced seagrasses (Zostera marina and Z. japonica). Both shrimp also serve as important prey for other consumers like Dungeness crab (Metacarcinus magister), sculpins, and sturgeon (Stevens et al., 1982; Armstrong et al., 1995; Dumbauld et al., 2008), and larval shrimps are prey for juvenile salmonids and other planktivorous fishes (Forsberg et al., 1975; J. Chapman, unpubl. data). Finally, burrowing shrimp cause substantial losses of cultured shellfish (Feldman et al., 2000; Dumbauld et al., 2004), so declining populations should result in reduced need for their control as prescribed in an integrated pest management plan for shellfish culture in Willapa Bay and Grays Harbor, Washington (Dumbauld et al. 2006). The shellfish growers in these two estuaries are currently evaluating alternative shrimp control measures including a less toxic pesticide, but did report lower shrimp abundance on their aquaculture tracts during this study period. 20 Population survey method considerations Sediment cores or suction devices like yabby pumps remain the only way to directly sample burrowing shrimp and obtain life history information because, unlike some burrowing crabs and lobsters that can be sampled at the surface (Mouton and Felder, 1996; Tuck et al., 1997; Macia et al., 2001; MacFarlane, 2002; Smith et al., 2003), these shrimps are believed to remain in their burrows for most of their lives (Dworschak, 1987; Astall et al., 1997; Candisani et al., 2001). Direct sampling techniques are laborious and expensive, however, and have prompted repeated attempts to relate shrimp density obtained from cores to counts of burrow openings measured at the sediment surface in order to quantitatively assess populations (Wynberg and Branch, 1994; McPhee and Skilleter, 2002; Rotherham and West, 2003). Rotherham and West (2007) used yabby pumps to sample Trypaea australiensis across a broad range of temporal and spatial scales in Australia because they found the burrow count shrimp relationship to be dependent on time and location. Butler and Bird (2007) found similar sources of variability, but suggested that shrimp per burrow relationships could be used when site-specific validation procedures were conducted and when the counts were used to detect large fluctuations in abundance. Our results suggest that a consistent and reasonable fit between burrow opening count and shrimp density held for Upogebia pugettenis and Neotrypaea californiensis in U.S. West Coast estuaries. Though we also found variability (e.g., largely due to errors in the identification and counting of burrow openings), we agree with these authors that trends can be reasonably assessed as long as sampling occurs during warmer months when shrimp are most active and quality assurance methods are used to minimize identification and counting errors. The ratios we obtained (1.5 21 burrow openings shrimp-1 for Neotrypaea and 1.7 burrow openings shrimp-1 for Upogebia) are similar to those derived in previous studies of these species (1-2 burrow openings per shrimp; Posey, 1986b; Posey et al., 1991; Dumbauld et al., 1996). While these ratios are lower than those based on the actual number of surface openings obtained from resin casts of the burrows (Thompson, 1972; Swinbanks and Luternauer, 1987; 2-3 burrow openings per shrimp; DeWitt et al unpubl. data; Griffis and Chavez, 1988), they can be reliably used in population estimations. This study was a first attempt to map and track populations of Upogebia and Neotrypaea populations over time at an estuary scale. Our primary goal was to compare these trends to those observed at single monitoring stations. Monitoring changes in shrimp populations in multiple estuaries within the region would be beneficial for establishing a comprehensive, quantitative understanding of the larger metapopulation status of these shrimps, how changes in their populations affect ecosystem processes, and for distinguishing probable mechanisms driving population change including the influence of coastal oceanographic processes on larval recruitment. While population trends estimated from stationary long-term monitoring stations located in high-density shrimp beds could mis-characterize the average population density and total population size within the system resulting in the false appearance of stability (Tamaki et al., 1996; Berkenbusch and Rowden, 1998; McPhee and Skilleter, 2002; Rotherham and West, 2007), we found that the direction of estuary wide change was adequately characterized by such a simple monitoring effort. Since wide area surveys are costly to conduct, we have explored a simpler and less expensive approach which tracks annual changes in the area and demographics of selected shrimp beds at our long-term monitoring locations, surveying their perimeters on foot using a GPS device and taking representative core samples within each bed (e.g. Chapman and Carter, in press). Used in conjunction with estuary-scale grid sampling conducted at 5 or 10-year 22 intervals, this smaller scale bed-mapping method would facilitate monitoring shrimp populations within and among estuaries and should provide ecosystem-scale information needed to understand and respond to changes in these populations. 23 Acknowledgements The authors are grateful to a host of field assistants including J. Connely, D. Scarborough, M. Schneider, D. Trobaugh and G. Wendling for the 2002 surveys and Katelyn Bosley, Cara Fritz, Roy Hildenbrand, and Roxanna Hintzman for 2008 and 2010 surveys and long-term density assessments. We especially thank the Lincoln County CSC-OYCC crew and their leader Jack Chapman for their dedicated 2008 field sampling effort. The manuscript was greatly improved by several reviewers including Bob Ozretich and we also thank Fiona Bird and Kristine Feldman for their reviews of a much earlier draft. This research was funded by the U.S. Department of Agriculture, Agricultural Research Service and the U.S. Environmental Protection Agency, Office of Research and Development, which have subjected it to agency review and approved it for publication. Mention of trade names or commercial products does not constitute endorsement or recommendation for use. 24 References Abed-Navandi, D., Dworschak, P.C., 2005. Food sources of tropical thalassinidean shrimps: a stable-isotope study. Marine Ecology Progress Series 291, 159-168. Armstrong, J.L., Armstrong, D.A., Mathews, S.B., 1995. Food habits of estuarine staghorn sculpin, Leptocottus armatus, with focus on consumption of juvenile Dungeness crab, Cancer magister. Fishery Bulletin 93, 456-470. Astall, C.M., Taylor, A.C., Atkinson, R.J.A., 1997. Behavioural and physiological implications of a burrow-dwelling lifestyle for two species of Upogebiid mud-shrimp (Crustacea: Thalassinidea). Estuarine, Coastal and Shelf Science 44, 155-168. 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Biological Invasions 12, 1791-1804. 37 Table 1. Mean density of shrimp and percent change in our long-term monitoring locations on Idaho Flat, Yaquina Bay, OR, for Neotrypaea and Upogebia, and at Stony Point Sands and Palix River, Willapa Bay, WA, for Neotrypaea. Also given are results (test statistic (t) , degrees of freedom, and p value for a Welches T- test on density. Willapa Bay, Washington Neotrypaea Palix River 2006 2009 Stony Point Sands 2009 2010 Yaquina Bay, Oregon Neotrypaea Idaho Flat 2008 2010 Upogebia Idaho Flat 2008 2010 -2 Mean Adults m ( ± Standard Error) 165.8 ± 36.56 55.2 ± 16.24 Percent Change -67 % Welch T-test t, df, p 2.76, 12.4, 0.017 66.6 ± 10.72 60.9 ± 7.08 -9 % 0.44, 7.6, 0.67 350.2 ± 15.05 286.0 ± 24.10 -18 % 2.26, 15.1, 0.039 290.9 ± 18.48 199.1 ± 8.50 -32 % 38 4.51, 12.7, 0.001 Table 2. Estimated number of total burrows, total shrimp, and percent change in our focal areas on Idaho Flat and Sally’s Bend, Yaquina Bay, OR, and Ellen/Stony Point Sands, Willapa Bay, WA (only in areas with overlapping data in each year). Note: these numbers do not represent the total number of shrimp in the estuary, are based on interpolations and should only be used as approximations . Burrows Shrimp (million) (million) Yaquina Bay 2008 Yaquina Bay 2010 48 37 32 24 -23 % Willapa Bay 2006 Willapa Bay 2009 Willapa Bay 2010 539 282 225 356 186 149 -48 % -20 % Burrows (millions) Shrimp (millions) 274 256 162 151 Neotrypaea Upogebia Yaquina Bay 2008 Yaquina Bay 2010 39 Percent Change Percent Change -7 % Table 3. Abundance, biomass, and total area of burrow surface linings of Upogebia and Neotrypaea on the Yaquina Bay tideflats in 2002, 2008 and 2010 and Neotrypaea on Willapa Bay tideflats in 2006, 2009 and 2010. Biomass and burrow lining areas per average shrimp were estimated by integrating wet weight and burrow area with carapace length (CL) over the average length frequencies of Upogebia and Neotrypaea in annual samples (2005 – 2010). Wet weight to carapace length (CL) estimates were: Uwt = 6.07E-4*CL2.99 and Nwt = 2.69E-4*CL3.66 for Upogebia and Neotrypaea respectively (from Dumbauld et al. 1996, Smith et al. 2008). Average burrow surface area per Upogebia: Us = (8.6E-5*CL2.09) was estimated from resin casts of Upogebia burrows collected in Yaquina Bay by the authors and from Bodega Bay, California by Thompson (1972) resulting in an average of Us = 0.05 m2 per burrow. For Neotrypaea we used an average Ns = 0.016 m2 per burrow from the literature (summarized in Griffis and Suchanek 1991). Note: these numbers do not represent the total number of shrimp in the estuaries, are based on interpolations, and should only be used for comparative purposes. Upogebia Yaquina Bay Willapa Bay Idaho Flats and Sallys Bend Ellen and Stony Pt. Sands (3.46 km2) (7.97 km2) 2002 2008 2010 2006 20009 2010 318 162 151 na na na 1,900 965 902 na na na 16.0 8.1 7.6 na na na Abundance (M) 110 32 24 356 186 149 Biomass (t) 180 52 40 3,670 1,920 1,530 Burrow surface area (km2) 1.8 0.5 0.4 5.7 3.0 2.4 Abundance (M) Biomass (t) Burrow surface area (km2) Neotrypaea 40 Figure Captions Figure 1. (A) Focal mapping area for Neotrypaea in Willapa Bay, Washington in 2006, 2009, and 2010, with our annual population monitoring sites at Palix River for Neotrypaea (,(19882009), Stony Point Sands for Neotrypaea (×, 2009 -2010), Cedar River for Upogebia (, 19882007), and at Goose Point for Upogebia (, 2003 – 2009) (B) Focal mapping area for Neotrypaea and Upogebia in Yaquina Bay, Oregon in 2008 and 2010, with our annual population monitoring sites at Idaho Flats (2005-2010) for Neotrypaea () and Upogebia (). Figure 2. Mean density of Neotrypaea (Palix River and Stony Point Sands, Willapa Bay) and Upogebia (Cedar River and Goose Point. , Willapa Bay) at site-specific monitoring locations in dense shrimp beds (n = 10 cores, error bars = ±SE). Broad scale survey years (2006, 2009, and 2010) for which changes in mean Neotrypaea density are compared to population changes are highlighted. Figure 3. Mean density of Upogebia and Neotrypaea collected at a long-term monitoring location in dense shrimp beds located on Idaho Flat in Yaquina Bay from 2005-2010 (error bars = ±SE). Broad scale survey years (2008 & 2010) are highlighted. Changes in mean density between 2008 and 2010 for Upogebia and Neotrypaea are compared to total population changes estimated from the broad scale survey in these years. Figure 4. Length frequency of Upogebia collected at a long-term monitoring site located on Idaho Flats in Yaquina Bay from 2005 – 2010. Shrimp parasitized by the bopyrid isopod 41 Orthione griffenis are shown as dark bars. Note large group of 1 year old shrimp that increased population density and abundance in 2007. Figure 5. Relationship between the number of burrows and the number of shrimp collected from the combination of 60 annual survey cores from 2005-2010 (filled circles) and 42 cores collected in a stratified random design across a range of shrimp density in 2008 (open circles) in Yaquina Bay, OR. Dashed lines = 95% CI. Figure 6. Neotrypaea burrow counts (m-2) at mapping locations at Ellen/Stony Point Sands, Willapa Bay, Washington in 2006, 2009 and 2010. The change in burrow counts at each location between each set of years is shown in the graphs on the right. The model fit line and confidence intervals do not overlap the 1:1 line, indicating a significant decline in burrow densities between 2006, 2009, and 2010. Figure 7. Neotrypaea burrow counts (m-2) at mapping locations in Yaquina Bay, Oregon in 2008 and 2010. The change in burrow counts at each location between years is shown in the graphs on the right. The model fit line and confidence intervals do not overlap the 1:1 line, indicating a significant decline in Neotrypaea burrow densities between 2008 and 2010. Figure 8. Upogebia burrow counts (m-2) at mapping locations in Yaquina Bay, Oregon in 2008 and 2010. The change in burrow counts at each location between years is shown in the graphs on the right. The model fit line and confidence intervals do not overlap the 1:1 line, indicating a significant decline in Upogebia burrow densities between 2008 and 2010. 42 WILLAPA BAY 7.5 Kilometers WASHINGTON OREGON - - YAQUINABAY 2.5 Kilometers Fig. 1. 500 Neotrypaea Palix River Stony Point Sands 400 Upogebia Cedar River 300 2006 200 2009 2010 Density m -2 Goose Point 100 0 1985 1990 1995 2000 Collection Year Fig. 2. 2005 2010 2008 500 Density m - 2 2010 400 300 200 Neotrypaea 100 Upogebia 0 2005 2006 2007 2008 Collection Year Fig. 3. 2009 2010 60 50 2005 40 Count Count 60 Parasitized 50 30 20 10 10 0 5 10 15 20 25 30 0 35 60 Count Count 30 10 15 20 25 30 35 30 20 10 10 0 5 10 15 20 25 30 0 35 60 2009 40 20 0 0 5 10 15 20 25 30 35 60 1+ shrimp 50 50 2007 40 Count Count 5 50 2006 40 30 30 20 10 10 0 5 10 15 20 25 Carapace Length (mm) 30 35 2010 40 20 0 0 60 50 Fig. 4. 30 20 0 2008 40 0 0 5 10 15 20 25 Carapace Length (mm) 30 35 Neotrypaea 2 600 Combined Fit: y=0.66x R =0.78 500 400 300 -2 Shrimp (numberm ) 200 100 0 Upogebia 600 2 Combined Fit: y=0.59xR =0.88 500 400 300 200 100 0 0 200 400 600 -2 Burrows (number m ) Fig. 5. 800 300 1:1 Sites Fit 95% Confidence Interval 200 Burrow Counts m 2 in 2009 250 150 100 50 0 0 50 100 150 Burrow Counts m 200 2 250 300 in 2006 300 1:1 Sites Fit 95% Confidence Interval 200 Burrow Counts m 2 in 2010 250 150 100 50 0 0 50 100 150 Burrow Counts m Fig. 6. 200 2 in 2009 250 300 500 1:1 Sites Fit 95% Confidence Interval 300 Burrow Counts m 2 in 2010 400 200 100 0 0 100 200 Burrow Counts m Fig. 7. 300 2 in 2008 400 500 600 1:1 Sites Fit 95% Confidence Interval 400 Burrow Counts m 2 in 2010 500 300 200 100 0 0 100 200 300 Burrow Counts m Fig. 8. 400 2 in 2008 500 600