Distribution and transport of bay anchovy (Anchoa mitchilli) eggs and larvae in Chesapeake Bay

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1 Estuarine, Coastal and Shelf Science 6 (4) 49e429 Distribution and transport of bay anchovy (Anchoa mitchilli) eggs and larvae in Chesapeake Bay E.W. North ), E.D. Houde 1 University of Maryland Center for Environmental Science, Chesapeake Biological Laboratory, USA Received 28 February 3; accepted 8 January 4 Abstract Mechanisms and processes that influence small-scale depth distribution and dispersal of bay anchovy (Anchoa mitchilli) early-life stages are linked to physical and biological conditions and to larval developmental stage. A combination of fixed-station sampling, an axial abundance survey, and environmental monitoring data was used to determine how wind, currents, time of day, physics, developmental stage, and prey and predator abundances interacted to affect the distribution and potential transport of eggs and larvae. Wind-forced circulation patterns altered the depth-specific physical conditions at a fixed station and significantly influenced organism distributions and potential transport. The pycnocline was an important physical feature that structured the depth distribution of the planktonic community: most bay anchovy early-life stages (77%), ctenophores (72%), copepod nauplii (O76%), and Acartia tonsa copepodites (69%) occurred above it. In contrast, 9% of sciaenid eggs, tentatively weakfish (Cynoscion regalis), were found below the pycnocline in waters where dissolved oxygen concentrations were!2. mg l ÿ1. The dayenight cycle also influenced organism abundances and distributions. Observed diel periodicity in concentrations of bay anchovy and sciaenid eggs, and of bay anchovy larvae O6 mm, probably were consequences of nighttime spawning (eggs) and net evasion during the day (larvae). Diel periodicity in bay anchovy swimbladder inflation also was observed, indicating that larvae apparently migrate to surface waters at dusk to fill their swimbladders. Overall results suggest that wind-forced circulation patterns, below-pycnocline dissolved oxygen conditions, and diel changes in vertical distribution of larvae and their copepod prey have important implications for potential transport of bay anchovy early-life stages. Ó 4 Elsevier Ltd. All rights reserved. Keywords: biologicalephysical interactions; larval transport; predatoreprey interactions; bay anchovy; zooplankton; Chesapeake Bay 1. Introduction The mechanisms by which organisms respond or react to the biological and physical environment are critical in estuaries, the site of important spawning grounds and nursery areas for many fish species. Many factors, including physics, larval development, food abundance, and predation act and interact to affect the small-scale distributions and dispersal of fish early-life ) Corresponding author. University of Maryland Center for Environmental Science, Horn Point Laboratory, P.O. Box 775, Cambridge, MD 21613, USA. 1 Present address: University of Maryland Center for Environmental Science, Chesapeake Biological Laboratory, P.O. Box 38, Solomons, MD 688, USA. addresses: enorth@hpl.umces.edu (E.W. North), ehoude@ cbl.umces.edu (E.D. Houde). stages, including those of bay anchovy (Anchoa mitchilli) in Chesapeake Bay. Bay anchovy is the most abundant fish in Chesapeake Bay and in many coastal areas of the western North Atlantic. It is a pelagic, small (!1 mm), short-lived (!3 years) fish that plays an important ecological role in Chesapeake Bay as a major prey for piscivores such as striped bass (Morone saxatilis), bluefish (Pomatomus saltatrix), and weakfish (Cynoscion regalis) (Baird and Ulanowicz, 1989; Hartman and Brandt, 1995). It is a pelagic, serial spawner with a reproductive season in Chesapeake Bay that extends from May to September peaking in July (Luo and Musick, 1991; Zastrow et al., 1991). Spawning occurs at salinities from to 32 psu (Dovel, 1971; Olney, 1983; Houde and Zastrow, 1991) and peaks at temperatures from 26 to 28 (C(Houde and Zastrow, 1991). Bay anchovy spawns between 18 and /$ - see front matter Ó 4 Elsevier Ltd. All rights reserved. doi:.16/j.ecss

2 4 E.W. North, E.D. Houde / Estuarine, Coastal and Shelf Science 6 (4) 49e429 hours (Luo and Musick, 1991; Zastrow et al., 1991), producing daily cohorts of eggs that hatch into yolk-sac larvae at 18e27 h after fertilization, depending upon temperature (Houde and Zastrow, 1991). Larvae begin feeding about 2 days after hatching (Houde and Zastrow, 1991). Larval anchovy first feed on microzooplankton such as copepod nauplii, rotifers, and tintinnids, and shift to larger copepodites and adult copepods as they grow (Houde and Zastrow, 1991; Auth, 3). Their growth rate is temperature-dependent (Houde, 1974) and is about.5e.8 mm d ÿ1 in Chesapeake Bay (Rilling and Houde, 1999b; Auth, 3). Physical conditions may influence the spatial and vertical distribution of bay anchovy early-life stages. The occurrence and survival of early-life stages below the pycnocline may depend on dissolved oxygen (DO) concentrations because larvae avoid waters with low DO (Breitburg, 1994; Keister et al., ) which can limit the viability of bay anchovy early-life stages (Chesney and Houde, 1989; Houde and Zastrow, 1991). Temperature also potentially could influence the distribution of bay anchovy early-life stages, especially in highly stratified conditions. The reported range of occurrence for temperature is 13.e3. (C for eggs and 15.e 3. (C for larvae in Chesapeake Bay (Houde and Zastrow, 1991). In addition, circulation patterns such as residual eddies (Hood et al., 1999) and plume fronts (Peebles, 2) can form retention areas that affect early-stage distributions. Prey and predator concentrations may influence the spatial occurrence and vertical distribution of bay anchovy eggs and larvae. Adult bay anchovy may spawn where food abundance is high, leading to an association between high concentrations of early-life stages and the copepods that serve as prey for adult anchovy (Peebles et al., 1996; Peebles, 2). The location of larval prey may also influence larval distributions, and larvae may follow the vertical migrations of copepod prey. Predation may affect the distribution of bay anchovy eggs and larvae by: (1) causing direct mortality; (2) stimulating predator-avoidance movements by larvae; and/or (3) influencing adult spawning-site selection because adults may avoid spawning in areas of high gelatinous zooplankton abundance (Dorsey et al., 1996). Major predators include the gelatinous zooplankton scyphomedusan (Chrysaora quinquecirrha) and the lobate ctenophore (Mnemiopsis leidyi) (Purcell et al., 1994). Ontogenetic changes in swimming ability, buoyancy regulation, and larval behavior also may influence larval distributions (Boehlert and Mundy, 1988). Potential for buoyancy regulation increases when the swimbladder first inflates, although precise control may not be possible until later in the development process (O Connell, 1981). Bay anchovy larvae could affect their transport within or out of estuaries by regulating their vertical position in the water column in relation to currents rather than by swimming horizontally (Norcross and Shaw, 1984; Miller, 1988). Several transport mechanisms have been documented for larval fish including tidally-timed vertical migration, also referred to as selective tidal stream transport (Rowe and Epifanio, 1994), migration around the depth of null velocity (Fortier and Leggett, 1983), and migration in relation to both time of day and tides (Weinstein et al., 198). In estuaries with two-layer circulation, such as Chesapeake Bay, diel vertical migration could result in up-estuary transport if larvae move into landward-flowing waters during the day in summer when days are longer than nights. It is also possible that random movements of larvae, coupled with frequent spawning by adults, could lead to retention of a sizable fraction of larvae in the estuary. The objective of this research was to identify the mechanisms and processes influencing small-scale vertical distributions and potential transport of bay anchovy eggs and larvae. The study was designed to determine how currents, time of day, physics (temperature, salinity, DO), ontogeny (egg and larval stages), food abundance, predation, and weather act or interact to affect egg and larval distributions. The temporal scale of sampling was designed to detect diel and tidally-timed vertical migrations of fish larvae. The research consisted of: (1) an initial survey in Chesapeake Bay to determine areas of maximum egg and larvae abundance; (2) depthstratified sampling at a fixed location to describe vertical distributions of early-life stages in relation to physical and biological factors; (3) length measurements of larvae to classify larvae by ontogenetic stage; and (4) an analysis of environmental data to evaluate factors that influenced physical conditions and organism distributions. 2. Methods Data were collected in Chesapeake Bay from 18e27 June 1996 on the 1 ft RV Cape Henlopen. The first two days of the cruise consisted of an ichthyoplankton and CTD survey along the axis of Chesapeake Bay (Fig. 1). After the initial survey, sampling effort was concentrated at a fixed station located in mid-bay (37( 45# N) from to 23 June and from 26 to 27 June (Fig. 1) Axial survey The initial survey along the axis of Chesapeake Bay was conducted to determine the spatial distributions and abundances of fish early-life stages and gelatinous zooplankton and to locate suitable areas for intensive depth-stratified collections. Twelve stations at 15 nautical-mile (w26 km) intervals were occupied from

3 E.W. North, E.D. Houde / Estuarine, Coastal and Shelf Science 6 (4) 49e Baltimore zooplankton was preserved in 1% formalin-plus-ethanol solution and transferred to % ethanol within six days of collection Fixed station sampling Latitude Potomac River Sampling Station CBOS CBBT Longitude Fig. 1. Chesapeake Bay, USA. Location of fixed-station sampling ( ) on e23 and 26e27 June 1996, current velocity measurements (C) at Chesapeake Bay Observing System (CBOS) buoy, and water level records (:) at Baltimore, MD, and Chesapeake Bay Bridge Tunnel (CBBT), VA. Open circles (B) represent axial sampling stations on 18e19 June the Bay mouth (37( # N) to the head of the Bay (39( 26# N). At each station, a CTD cast (water-column temperature, salinity, DO, and fluorescence) preceded ichthyoplankton collections. Ichthyoplankton and gelatinous zooplankton were collected in an openingeclosing 1 m 2 Tucker trawl with 28-mm mesh nets, a flowmeter, and a depthetemperature recorder. A pair of 2-min collections was made in a single deployment at each station, one from near bottom (within 1.5 m) to the pycnocline, and the second from the pycnocline to the surface. On average, each collection filtered w1 m 3 of water. Gelatinous zooplankton was separated from the sample with large strainers (w5 mm pores), identified, and biovolumes of each species were determined. Remaining Samples were collected at a fixed station to determine vertical distribution and abundance of fish early-life stages, microzooplankton, and gelatinous zooplankton in relation to each other and water-column hydrography. The site of the fixed station was determined from information on egg and larval abundances in the axial survey (visual inspection of samples) and with consideration of water-column depth, shipping traffic, and gelatinous zooplankton concentrations. The first occupation of the station (e23 June) was for 81 h and the second (26e27 June) for 28.5 h. At the fixed station, a suite of biological and physical measurements was obtained on each tide (every w6 h). First, current velocity was measured with an Acoustic Doppler Current Profiler (ADCP). Then, a CTD was deployed to measure water-column temperature, salinity, DO, and fluorescence, and to collect microzooplankton. Following the CTD cast, depth-stratified sampling for ichthyoplankton and gelatinous zooplankton was conducted at 5-m intervals from the surface to 25-m (near bottom). After two sets of depth-stratified samples were complete, another CTD cast was made and additional ADCP measurements were obtained. The entire suite of measurements (details below) was completed in w2.7 h Current velocity measurements Instantaneous measures of current velocity were determined from 6-min averages of raw data from the RV Cape Henlopen s hull-mounted khz ADCP. Raw data were collected in 1-m depth intervals (from 4 to 25 m) for 6e min with the vessel at constant speed and heading (parallel to the channel) every w2.7 h during the first fixed-station occupation, and every w1.7 h during the second occupation. Current velocities were separated into longitudinal (along-channel) and lateral (cross-channel) components. The longitudinal direction was set by determining the principal axis, the direction in which velocity variance was at a maximum Microzooplankton Depth-stratified microzooplankton samples were collected during the CTD casts in -l Niskin bottles attached to the CTD rosette. Samples were collected at 2.5, 7.5, 12.5, 17.5, and 22.5 m depths. They were filtered onto a 35-mm mesh and preserved in 5% formalineseawater. In the laboratory, zooplankton were identified and enumerated under a dissecting microscope. All organisms were enumerated in samples from 12.5, 17.5, and 22.5 m depths (79% of the samples). When the

4 412 E.W. North, E.D. Houde / Estuarine, Coastal and Shelf Science 6 (4) 49e429 number of organisms was high (21% of the samples, usually from 2.5 and 7.5 m depths), samples were diluted to 25 ml and three 1-ml aliquots were enumerated. The mean number of organisms from the three aliquots was multiplied by 25 ml to estimate the total number of organisms in the sample Ichthyoplankton and gelatinous zooplankton Depth-stratified collections of ichthyoplankton and gelatinous zooplankton at the fixed station were made in the 1-m 2 Tucker trawl. This gear was deployed as in the initial axial survey except that the trawl was fished for 2 min in the 5e m, e5 m, 15e m, e15 m, and 25e m depth intervals. To minimize potential bias due to time and tidal stage effects during a station occupation, the order of depth intervals sampled was randomly assigned in each set of stratified samples, except that the e5 m and 5e m depth intervals were always sampled during one deployment, and in that order. Catches were processed as at the axial survey stations, except that (1) gelatinous zooplankton processing was limited to total biovolume measurements (species were identified in three sets of depth-stratified collections) and (2) samples were preserved in a 2% formalin-plus-ethanol solution when gelatinous zooplankton biovolumes were especially large. In the laboratory, fish eggs and larvae were identified and counted under a dissecting microscope. Although entire samples were sorted for larvae, eggs were counted in a subsample (1/2 to 1/8 of the whole sample, divided with a plankton splitter) when numbers were O. Larvae from the fixed-station collections were separated into two groups based on swimbladder inflation (inflated or not inflated). All larvae were measured with a computer-based digitizing system Analysis The time series of depth distributions of physical factors, ichthyoplankton (no. m ÿ3 ) and zooplankton (no. l ÿ1 ) concentrations, and gelatinous zooplankton biovolumes (ml m ÿ3 ) at the fixed station was contoured (Golden Software Surfer v.6.1). In these plots, time was depicted in hours since midnight of the first day of fixedstation occupation. The gridding method was kriging with an isotropic linear variogram model. Grid-line geometry was no finer than half the average distance between measurements in the Y-direction (depth), and no finer than the average distance between measurements in the X-direction (time). Abundances (per m 2 ) in a depth interval of ichthyoplankton, microzooplankton, and biovolumes of gelatinous zooplankton in each tow were estimated by multiplying organism concentrations by the sampled depth interval. For fixed-station samples, the abundance estimates from collections during a single net deployment (e5 and 5e m depth intervals only) were adjusted for a 1-m overlap between the sampled depth intervals (A i Z 4c i Cðc i Cc j Þ=2 where A i Z abundance in ith depth interval, c i Z concentration in ith depth interval, and c j Z concentration in the jth depth interval). Abundances in each depth interval were summed to determine total water-column abundance (no. m ÿ2 or ml m ÿ2 ) at the station. Mean depths of organism occurrence at the fixed station were calculated for each set of depth-stratified samples by averaging net tow depths (the midpoint of each depth interval) weighted by the concentration of fish eggs and larvae (North, 1). To investigate potential relationships in the distribution of organisms, correlation coefficients (SAS 6.12, PROC CORR) were calculated for organism abundances as well as for mean depths of occurrence. Data were log e -transformed and Pearson correlation coefficients were calculated for normally distributed data. Spearman rank-order correlation coefficients were calculated for data that were not normally distributed (abundances of bay anchovy yolksac and O13 mm larvae). Correlation analyses of mean depths of occurrences also included pycnocline depth and the depth of the 3. mg l ÿ1 DO contour. Pycnocline depth was estimated as the depth at which the BrunteVa isa lla frequency was maximum during each CTD cast (Mann and Lazier, 1996). The depth of the 3. mg l ÿ1 DO contour was calculated by linear interpolation of CTD DO measurements adjacent to 3. mg l ÿ1. The CTD DO measurements were calibrated with pre- and post-cruise Winkler titrations conducted by E.M. Smith during spring (28 Aprile5 May) and summer (17e26 July) research cruises onboard the RV Cape Henlopen in Potential advection of water (and planktonic organisms within it) during fixed-station sampling was estimated by calculating displacement (km) using interpolated ADCP current velocities (Golden Software Surfer v.6.1, applying kriging with an isotropic linear variogram model). Initial interpolated ADCP current velocities (grid-line geometry was 1 m in the depth direction and 1.67 h in the time direction) were reinterpolated with half the distance between grid points in the timeedirection. This process was repeated until the distance between grid points in the timeedirection was w7 s. Potential displacement during each time step (i) was calculated by multiplying interpolated velocity (v) at each 1-m depth by the time-step duration (t ¼ 7 s). Cumulative potential displacement (D c ) at time c was calculated by summing displacement at t ¼ c with displacement during all previous time steps (D c ¼ Sv i t for t ¼ to t ¼ c). Estimates of lower layer water mass movement based on our displacement calculations (18.5 cm s ÿ1 ) were similar to those reported at the mouth of the Choptank River, a Bay tributary ( cm s ÿ1 ) (Sanford and Boicourt, 199).

5 E.W. North, E.D. Houde / Estuarine, Coastal and Shelf Science 6 (4) 49e Simulated transport analysis Displacement calculations were used to estimate potential length-specific transport of bay anchovy larvae by size class to determine if diurnal vertical migrations that increase in range as larvae develop could affect dispersal up- or down-estuary. All size classes of larvae were hypothesized to remain at 5 m during the night to isolate the effects of daytime migrations. During the day, larvae 4e5 mm in length were hypothesized to be at 5 m, while the mean depth of successive 1-mm size classes was 1 m deeper than the next smallest size class. Potential transport (km) during the first fixed-station occupation was estimated by calculating cumulative displacement for the 5-m depth interval during night (sunset to sunrise) and adding it to cumulative displacement at depths specific for each length class during the day (sunrise to sunset) Statistical analysis A statistical analysis was conducted to determine if advection (displacement), photoperiod (dayenight), or time periods of differing physical conditions described a significant amount of variability in the abundances of organisms. Time periods of differing physical conditions were chosen based upon water-column hydrography. A repeated measures, multiple-regression analysis was conducted on log e -transformed abundances (no. m ÿ2 or ml m ÿ2 ) with displacement, dayenight, and time period as fixed effects, and station occupation (first or second) as a random effect (SAS 6.12, PROC MIXED). Covariance in time was modeled with a first-order autoregressive covariance structure. The dayenight variable was coded day Z and night Z 1 where night represented time from sunset to sunrise. Explanatory variables passed tolerance and condition index multicollinearity tests (SAS 6.12, PROC REG and PRINCOMP). All regression models passed ShapiroeWilks normality tests (SAS 6.12, PROC UNIVARIATE) as well as tests for homogeneity of variance (plots and correlation analyses of KresidualsK versus predicted values). Coefficients of determination (R 2 ) from repeatedmeasures, regression models with fixed and random effects are unreliable so are not reported. Instead, F values are included in tables of results to indicate the relative amount of variance accounted for by fixed effects. The random effect station occupation was not significant in any of the models, so was excluded from tables of results Environmental monitoring data Physical conditions during the cruise were evaluated with environmental monitoring data of the Chesapeake Bay Observing System (CBOS), NOAA National Ocean Service (NOS), and the Chesapeake Bay Program (CBP) Monitoring Program. Wind (RMP) and current velocities (at 2.4 and 18.9 m depths) from the mid-bay CBOS buoy in June 1996 were used in conjunction with NOS water level measurements at Baltimore and at the Chesapeake Bay Bridge Tunnel (Fig. 1) to identify forces that influenced physical conditions during fixedstation sampling. Water level measurements and CBOS current velocities were filtered with a Lanczos low pass filter with a 34-h half-power point to reveal lowfrequency water level and circulation patterns (courtesy W.C. Boicourt and C. Derry). June salinity (psu) and DO (mg l ÿ1 ) measurements from channel stations of the CBP Monitoring Program were used to provide historical context to the field sampling results in June Mean salinity and DO concentrations were calculated at the depths of CTD measurements from 1985e1995 June surveys and compared to June 1996 monitoring results. 3. Results 3.1. Historical context Freshwater flow into the Chesapeake Bay during the spring of 1996 was far above the long-term mean (Boynton et al., 1997). The high input of freshwater strongly affected June salinity structure in mid-chesapeake Bay. Upper layer (above-pycnocline) salinities were lower in June 1996 (Fig. 2a) than the mean of the prior years, as were lower layer (below-pycnocline) DO concentrations. The intersection of the psu isohaline with the surface was O8 km further downestuary in June 1996 compared to its mean location in 1985e1995 and, at the fixed station, the June 1996 surface salinity was w5 psu lower than the June long-term average. Dissolved oxygen concentrations!1 mgl ÿ1 extended 5 km further down-estuary in June 1996 compared to their mean locations in June 1985e1995. At the fixed station, lower layer DO concentrations in June 1996 were w1 mgl ÿ1 less than the June long-term average Predominant organisms The most abundant eggs and larvae collected were bay anchovy. Mean abundances (pooled axial and fixedstation surveys) of anchovy eggs and larvae were 183: m ÿ2 G 33:2 s.e. and 4:4 m ÿ2 G :69 s.e., respectively. Sciaenid eggs were the second most abundant fish egg (mean abundance Z 53:4m ÿ2 G 8:4 s.e.). Although the sciaenid eggs potentially were spawned by 4e7 different species (Olney, 1983; Daniel and Graves, 1994), most probably were weakfish (Cynoscion regalis) because most positively-identified sciaenid larvae were weakfish (5 out of 72 post-yolk-sac sciaenid larvae).

6 414 E.W. North, E.D. Houde / Estuarine, Coastal and Shelf Science 6 (4) 49e429 Depth (m) 3 a) June 11-12, 1996 salinity (psu, lines) and dissolved oxygen (shaded contours) (mg l ) Biovolume (ml m -2 ) Abundance (no. m -2 ) Abundance (no. m -2 ) Abundance (no. m -2 ) b) Bay anchovy eggs upper-layer lower-layer c) Bay anchovy larvae d) Sciaenid eggs e) Gelatinous zooplankton Distance from Head of Bay (km) Fig. 2. (a) Salinity (psu) contour lines and dissolved oxygen (mg l ÿ1 ) shaded contours along the axis of Chesapeake Bay for June 11e12, Data from Chesapeake Bay Monitoring Program. Black dots indicate depths of CTD measurements. Dissolved oxygen data up-estuary of 5 km were not available. Triangles mark the location of axial survey stations and an arrow marks the location of the fixed sampling station. Bar graphs depict axial survey abundances (no. m ÿ2 ) of (b) bay anchovy eggs, (c) bay anchovy larvae, (d) sciaenid eggs, and (e) gelatinous zooplankton biovolume (ml m ÿ2 ) above (upper layer) and below (lower layer) the pycnocline. Stations from 174e257 km were sampled at night. Biovolumes of gelatinous zooplankton were mostly composed of the ctenophore, Mnemiopsis leidyi, (99% in axial survey collections, 98% in fixed-station collections) with the hydromedusa Nemopsis bachei constituting the remainder. Mean biovolume of gelatinous zooplankton was 77:6 mlm ÿ2 G 4:2 s.e. During fixed-station sampling, most copepods (96%) were Acartia tonsa copepodites (mean abundance Z 1:8! 5 m ÿ2 G :14! 5 s.e.)

7 E.W. North, E.D. Houde / Estuarine, Coastal and Shelf Science 6 (4) 49e and adults (mean abundance Z 2:23! 4 m ÿ2 G :31! 4 s.e.) Axial survey In axial survey collections, bay anchovy eggs (Fig. 2b) were two orders of magnitude more abundant than anchovy larvae (Fig. 2c), and one order of magnitude more abundant than sciaenid eggs (Fig. 2d). Peak abundances of bay anchovy eggs and larvae, sciaenid eggs, and ctenophores (Fig. 2bee) occurred downestuary of the Potomac River (174 km), although ctenophores occurred in significant biovolumes as far up-estuary as 58 km. In the upper Bay (e89 km), anchovy eggs (max: 34.6 m ÿ2 ) were relatively uncommon and anchovy larvae were absent (Fig. 2b,c). Within lower Chesapeake Bay (O174 km), bay anchovy eggs were most abundant where ctenophore biovolume was minimal (23 and 285 km), and were virtually absent where ctenophore biovolume peaked (257 km). Anchovy eggs, larvae, and ctenophores were most abundant above the pycnocline at all stations (Fig. 2bee). In contrast, sciaenid eggs were abundant in both layers at the most down-bay stations but were predominantly in below-pycnocline waters up-estuary of 257 km, and apparently associated with salinities O18 psu Fixed-station sampling Fixed-station sampling was conducted near 1 km where depths were O25 m, anchovy larval abundances peaked, and gelatinous zooplankton biovolumes were moderate (Fig. 2c,e). Maximum abundance of bay anchovy larvae (17.9 no. m ÿ2 ) during the fixed-station occupation was comparable to peak abundance during the axial survey (11.5 no. m ÿ2 ) Physics Contour plots of physical factors at the fixed station show a well-defined pycnocline, below which temperatures (Fig. 3a) were lower than those above the pycnocline. Dissolved oxygen concentrations!3. mg l ÿ1 generally occurred just below the pycnocline (Fig. 3b)and decreased with depth, frequently measuring!2 mgl ÿ1 near bottom. Highest fluorescence values (RFU, raw fluorescence units) occurred within and above the pycnocline and peaked in surface waters during afternoons a) Salinity (psu) contour lines and temperature (ºC) shaded contours ºC Depth (m) b) Dissolved oxygen (mg l 1 ) contour lines and fluorescence (RFU) shaded contours RFU Depth (m) c) Along-channel current velocity (cm s 1 ) Velocity (cm s 1 ) Depth (m) Time (hrs) Fig. 3. Time series of fixed station (a,b) CTD and (c) ADCP measurements, e23 and 26e27 June Two variables are plotted in panel (a), salinity (psu) contour lines and temperature ((C) shaded contours, and in panel (b), dissolved oxygen (mg l ÿ1 ) contour lines and fluorescence (RFU) shaded contours. Panel (c) depicts along-channel current velocity (cm s ÿ1 ) for first (2e83 h) and second (15e179 h) station occupations. Measurement locations indicated by black dots. White and gray bars at the top of (b) indicate day and night. Negative current velocity is up-estuary.

8 416 E.W. North, E.D. Houde / Estuarine, Coastal and Shelf Science 6 (4) 49e429 (Fig. 3b). Daytime peaks in DO concentrations corresponded to those of fluorescence, a result of photosynthesis in the upper layer. During the initial 3 h of the fixed-station occupation, the pycnocline was shallow (w6 m), temperatures in the upper layer were the highest of the time series, and those below the pycnocline were the lowest (Fig. 3a). Tidal currents were reversing in the upper and lower layers (Fig. 3c). After 3 h, conditions changed. The pycnocline deepened (w9 m), water temperatures and salinity increased in the lower layer and decreased in the upper layer. These changes corresponded to strong upestuary currents below the pycnocline where tidal currents were no longer reversing. This change in physical conditions likely was related to a wind event that occurred from 3e45 h during the fixed-station occupation (Fig. 4a, shaded area, June e23). Strong winds blowing from the north and north-west reduced water levels in the upper Bay (Fig. 4b, Baltimore record), and enhanced two-layer estuarine circulation. This response can be seen in the CBOS filtered current record where residual surface velocities increased in the down-estuary direction while lower layer velocities increased in the upestuary direction (Fig. 4c). Just prior to the second station occupation (Fig. 4, shaded area, June 26e27), another strong wind event from the north (Fig. 4a) reduced water levels in the upper Bay (Baltimore record, Fig. 4b), increased water level in the lower Bay (CBBT record, Fig. 4b), and resulted in enhanced two-layer circulation (Fig. 4c). At the start of the second occupation of the fixed station, the pycnocline had deepened (w15 m) and intensified, probably a result of mixing during the wind event (Fig. 3a). The 3. mg l ÿ1 DO contour also was driven deeper during the wind event as was fluorescence (Fig. 3b). The strong up-estuary current velocity throughout the water column (Fig. 3c, 16e17 h) likely was due to a flooding tide combined with a barotropic response to the removal of wind stress, producing an up-estuary pulse of water (a seiche effect). The subsequent gradual elevation in pycnocline depth during the second fixed-station occupation apparently resulted from the decrease in wind mixing as well as the barotropic response. Potential displacement of water during the two fixedstation occupations demonstrates the dynamic change in physical conditions and potential consequences for organism transport (Fig. 5). For the first h of the time series, displacement in both the surface (!9 m) and Fig. 4. (a) Wind velocity ( m s ÿ1 ) at mid-bay CBOS buoy, (b) filtered water level (cm) at Baltimore, MD, and Chesapeake Bay Bridge Tunnel (CBBT), VA, and (c) CBOS filtered along-channel current speed (cm s ÿ1 ) in the upper and lower layers in June, Vectors in (a) point in the direction to which the wind was blowing. Negative current speed in (c) is up-estuary. Shaded areas indicate timing of fixed-station sampling.

9 E.W. North, E.D. Houde / Estuarine, Coastal and Shelf Science 6 (4) 49e Displacement (km) down-estuary 4 m m 16 m up-estuary 23 m Time (hr) 4 m 25 m 17 m Fig. 5. Potential displacement (km) over time for the first and second fixed-station occupations. Lines represent potential displacement of water at each 1-m depth interval. Displacement at each time point was calculated by multiplying the along-channel current velocity estimate at that time by the duration between velocity estimates (velocity (km s ÿ1 )! time (s) = displacement (km)) and then summing with displacement estimates from all previous time points. Specific depths are marked by open circles and labeled. Negative displacement is up-estuary. lower layers (O9 m) was within that expected by tidal excursion. Between 25 and 45 h, enhanced gravitational circulation related to the northern wind event resulted in potential displacement of the surface layer w km down-estuary and the lower layer w km up-estuary. The upper layer was nearly stationary between 45 and 75 h, suggesting that collections in the upper layer during this time period were sampling the same water mass and its associated organisms. In contrast, lower layer displacement was non-stationary after 3 h, indicating that up-estuary residual currents continually transported water and organisms within it past the fixed station. The fixed-station time series was divided into four periods, each with different physical characteristics, to examine the influence of advecting water masses on biological collections. During the first period, 2e h, the pycnocline depth was shallow and displacement was near zero in both the upper and lower layers. The second period, e4 h, was characterized by changing physical conditions and rapid displacement in both the upper and lower layer. From 4e83 h, the third period, upper layer displacement was near zero, pycnocline depth was stable, and lower layer salinity continually increased, as did lower layer displacement up-estuary. Finally, the second fixed-station occupation can be considered a fourth period characterized by rapidly changing physical conditions: decreasing pycnocline depth, a strong barotropic up-estuary response throughout the water column, and a transition in the upper layer from wind-mixed towards restratification. Fig. 6 summarizes water mass movement during the first three periods. Waters sampled in the lower and upper layers during the first period (Fig. 6a) were separated by as much as km by the end of the second period (Fig. 6b). Organisms that were located at the fixed station during the first period could have been separated by as much as 6 km by the end of the third period (Fig. 6c) Biology All stages of bay anchovy (Figs. 7a, 8), ctenophores (Fig. 7c), copepod nauplii (Fig. 9a), and Acartia tonsa copepodites (Fig. 9b) were located in highest concentrations above the pycnocline during both the first and second fixed-station occupations (Table 1). In contrast, only % of sciaenid eggs were found above the pycnocline (Table 1, Fig. 7c). Although only about half of A. tonsa adults (49%) were found above the pycnocline during the first occupation, most (85%) were above the pycnocline during the second occupation (Table 1, Fig. 9c). Periods of different physical conditions explained a significant amount of the variability in abundances of anchovy eggs, yolk-sac larvae, 6e9 mm larvae, ctenophore, and copepod nauplii in multiple-regression models (Table 2). This suggests that the different upper layer water masses sampled during periods one, three and four contained different biological communities. In multiple-regression models, displacement included a measure of spatial association within water masses as well as an indication of the direction of water mass movement. Significant parameter estimates for bay anchovy yolksac, 3e6 mm, 6e9 mm, and 9e13 mm larvae indicated that abundances decreased as upper layer waters moved down-estuary past the fixed station, and that sciaenid egg abundance increased as lower layer waters moved upestuary past the fixed station Anchovy and sciaenid eggs Mean abundances of anchovy eggs were highest during the first period (2e h) while abundances of

10 418 E.W. North, E.D. Houde / Estuarine, Coastal and Shelf Science 6 (4) 49e429 a) Period Depth (m) corresponded to peaks in egg concentrations near the pycnocline (Fig. 7b), suggesting that most of these eggs were newly spawned. The significant dayenight effect for sciaenid eggs in the multiple-regression model (Table 2) almost certainly represented effects of evening spawning activity. b) c) Period 1 Period 2 Period 1 Period 2 Period 3 Displacement (km) Displacement (km) Displacement (km) sciaenid eggs were highest during the second period (e4 h) (Table 3). Anchovy egg abundance declined by the fourth period. Bay anchovy egg concentrations peaked near surface at sunset and at night (Fig. 7a), presumably because adults spawn above the pycnocline at night. The deeper mean depths of anchovy eggs (Fig. a) just before nightfall probably represented unhatched eggs, possibly dead (Iseki and Kiyomoto, 1997), from the previous night s spawn that were diffusing and sinking. In contrast, the relatively shallow mean depths of occurrence of sciaenid eggs (Fig. a) just before sunset Depth (m) Depth (m) Fig. 6. Cumulative displacement (km) of water at the fixed station at the end of (a) period one (2e h), (b) period two (e4 h), and (c) period 3 (4e83 h). Lower and upper layer water masses during period 1(2e h) potentially were separated by as much as 6 km by the end of period 3 (within 3.5 days) due to wind-induced, enhanced two-layer circulation Prey and predators Copepod nauplii and Acartia tonsa copepodites, the most common prey items in bay anchovy larval guts in Chesapeake Bay (Auth, 3), peaked in concentration and abundance in the upper layer during the first two periods (Fig. 9a,b, Tables 1 and 3). Nauplii were concentrated in the upper m of the water column, while copepodites were found throughout the upper layer, especially at night. It is likely that most nauplii were A. tonsa because most of the copepods (96%) collected during fixed-station sampling were this species. Peak concentrations of adult A. tonsa were generally shallower during night and deeper during day, often occurring within or below the pycnocline during day (Fig. 9c). Like nauplii and copepodites, abundance of adult A. tonsa declined between the second and fourth periods (Table 3). The significant dayenight effect for A. tonsa copepodite and adult abundances in the multipleregression model (Table 2) could indicate that these stages aggregated at depths not sampled by the Niskin bottles or that they were able to evade Niskin-bottles capture during the day. Ctenophores were present throughout the upper layer (Table 1) and often peaked in concentration during afternoon and night (Fig. 7c). Ctenophore biovolumes increased from the first to the fourth period (Table 3). Mean depths of ctenophores, copepod nauplii, and Acartia tonsa copepodites and adults overlapped (Fig. b). However, mean depths of nauplii tended to be shallower than ctenophore and copepodite mean depths during the first occupation (2e83 h), while A. tonsa adults tended to occur deeper, especially during the day. Mean depths of nauplii, copepodites and ctenophores appeared to track pycnocline depth during the first occupation and were significantly correlated with pycnocline depth for the entire fixed-station sampling period (Table 4a). At the beginning of the second occupation (15e17 h), mean depths of copepods and ctenophores no longer overlapped pycnocline depth, possibly because the wind event, that had deepened the mixed layer, also dispersed organisms through it. The negative correlations between ctenophore biovolumes and both copepod nauplii and Acartia tonsa copepodite abundances (Table 4b) suggest that high concentrations of nauplii and copepodites did not cooccur with high concentrations of ctenophores, possibly the result of differing levels of copepod production

11 E.W. North, E.D. Houde / Estuarine, Coastal and Shelf Science 6 (4) 49e Depth (m) Depth (m) Depth (m) a) Bay anchovy eggs (no. m -3) shaded contours and salinity (psu) contour lines b) Sciaenid eggs (no. m -3) shaded contours and salinity (psu) contour lines c) Ctenophore biovolume (ml m -3) shaded contours and salinity (psu) contour lines Time (hrs) (no. m -3) (no. m -3) (ml m -3) Fig. 7. Time series of (a) bay anchovy and (b) sciaenid egg concentrations (no. m ÿ3, shaded contours) and (c) ctenophore biovolume (ml m ÿ3, shaded contours) with salinity (psu) contour lines for the first (2e83 h) and second (15e179 h) fixed-station occupations. Black dots indicate mid-point of trawl depth intervals. White and gray bars indicate day and night. and/or mortality from ctenophore predation within these water masses Bay anchovy larvae Abundance (Table 3) and upper layer concentrations (Fig. 8) of bay anchovy yolk-sac and 3e6 mm larvae peaked during the first period (2e h) and were more than an order of magnitude lower in the third and fourth periods. Larvae O13 mm increased in abundance from the first to the fourth period (Table 3). Eighty-five percent of larvae collected during the fourth period were O9 mm. These large larvae probably were not the survivors from eggs and newly-hatched larvae during the first period, but represented a new group advected into the region. Larvae hatched from eggs spawned during the first period could not have been Ow7.5 mm by the fourth period because bay anchovy larvae hatch at w1.9 mm standard length (Houde and Zastrow, 1991) and grow at a rate of w.5e.8 mm d ÿ1 (Rilling, 1996; Auth, 3). Larvae O6 mm showed clear peaks in concentration at night (Figs. 8c, 11), suggesting that evasion of the Tucker trawl during the day can confound determination of diel vertical distribution of large anchovy larvae. In fact, 67% of larvae O5 mm, 75% of larvae O9 mm, and 95% of larvae O14 mm were collected at night. The dayenight effect accounted for most of the variability in abundances of larvae O9 mm in multiple-regression models (Table 2) and was significant for larvae O6 mm. Abundances of bay anchovy early-life stages were positively and most highly correlated with abundances in adjacent length classes (Table 4b). Abundances of bay anchovy larvae!13 mm were positively correlated with prey abundance (copepod nauplii and copepodites). Abundances of yolk-sac and 3e9 mm larvae were negatively correlated with biovolume of ctenophores, a potential predator. In contrast to mean depths of their predators and prey, mean depths of bay anchovy larvae were often located near, but did not always track, pycnocline depth (Fig. c). The difference in mean depths of occurrence for large and small larvae indicated that ontogenetic factors influenced the vertical distribution of larvae. During the first 3 h of fixed-station sampling, the average mean depths of larger larvae (6e13 mm) were nearer to surface than smaller larvae (!6 mm) during the night but were deeper during day.

12 4 E.W. North, E.D. Houde / Estuarine, Coastal and Shelf Science 6 (4) 49e429 a) Bay anchovy yolk-sac larvae (no. m -3) shaded contours and salinity (psu) contour lines (no. m -3) Depth (m) Depth (m) Depth (m) b) Bay anchovy larvae 3-6 mm (no. m ) shaded contours and salinity (psu) contour lines c) Bay anchovy 9-13 mm (no. m -3) shaded contours and salinity (psu) contour lines (no. m -3) (no. m -3) Time (hrs) Fig. 8. Time series of bay anchovy larvae concentrations (no. m ÿ3, shaded contours) with salinity (psu) contour lines. (a) Yolk-sac larvae, (b) larvae 3e6 mm, and (c) larvae 9e13 mm for the first (2e83 h) and second (15e179 h) station occupations. Black dots indicate mid-point of trawl depth intervals. Trawls in which no larvae were captured are marked by an x. White and gray bars indicate day and night. The occurrence of larval mean depths nearer to surface during the early evening (Fig. c) may be related to diel periodicity in swimbladder inflation and the physiological requirement of clupeoid larvae to gulp air at the surface to inflate their swimbladders (Blaxter and Hunter, 1982). When collections were made O45 min after sunset, the mean depths of O6 mm larvae were nearest to surface in early evening (, 45, 7 h) and deepened through the night (Fig. c). During the entire fixed-station occupation, most larvae with inflated swimbladders consistently occurred during night (Fig. 11). The diel periodicity in swimbladder inflation increased with larval development: 5% of 6e7 mm larvae and O8% of O11 mm larvae had inflated swimbladders at night. Inflated swimbladders were observed in some larvae as small as 4e5 mm. Results of the simulated transport analysis demonstrate the strong effect of shifts in vertical distributions on potential transport of anchovy larvae (Table 5). A 1-m difference in daytime mean depth resulted in R2 km difference in potential transport after 3.5 days. If bay anchovy larvae make diurnal vertical migrations that became deeper during the day as they develop, then potential transport of larger larvae is up-estuary while transport of younger larvae is down-estuary. Alternatively, Table 5 demonstrates the error in potential transport estimates that could be induced by net evasion. For example, if the majority of 9e mm larvae actually were located at m during the day but most larvae were collected at 12 m because of net evasion, then the estimate of potential transport (ÿ1.9 km) would be opposite in direction from the actual transport (6.1 km). 4. Discussion Our results demonstrate that complex and interacting biological and physical factors determine the characteristics of larval fish nursery areas in estuaries. A suite of factors influenced the distribution and potential transport of bay anchovy early-life stages, including windforced circulation patterns, pycnocline depth, DO concentrations, salinity, and time of day. In addition, prey and predator distributions and larval developmental stage may be important factors that influence transport of bay anchovy larvae. Wind events as short as 12e24 h had both direct and indirect effects on distribution of anchovy early-life stages. Direct wind mixing deepened the upper layer of

13 E.W. North, E.D. Houde / Estuarine, Coastal and Shelf Science 6 (4) 49e a) Copepod nauplii (no. l ) shaded contours and salinity (psu) contour lines -1 (no. l ) Depth (m) Depth (m) Depth (m) b) Acartia tonsa copepodites (no. l ) shaded contours and salinity (psu) contour lines c) Acartia tonsa adults (no. l -1) shaded contours and salinity (psu) contour lines Time (hrs) Fig. 9. Time series of copepod concentrations (shaded contours) with salinity (psu) contour lines. (a) Copepod nauplii (no. l ÿ1 ), (b) Acartia tonsa copepodites (no. l ÿ1 ), and (c) A. tonsa adults (no. l ÿ1 ) for the first (2e83 h) and second (15e179 h) station occupations. Black dots indicate depth of Niskin bottle collection. White and gray bars at top of panels indicate day and night (no. l ) (no. l ) the water column, broadening the depth zone where first-feeding anchovy larvae usually were located. Wind events may diminish first-feeding larval survival by dispersing prey organisms (Lasker, 1975) or reduce anchovy egg abundance by inhibiting adult spawning (Moser and Pommeranz, 1999). Wind events also may indirectly, but significantly, impact anchovy early-life stages by forcing a restructuring of the biological community. Wind-forced enhanced gravitational circulation that began after the first period quickly provided the plankton community above the pycnocline with potential access to a different water mass below it. This two-layer movement can expand or reduce suitable habitat for larval fish and may affect prey and predator populations that could vertically migrate into the surface layer. In addition, small-scale differences in residual current velocities (i.e., shear) may have restructured plankton communities within the upper and lower layers. For example, organisms in waters separated in depth by only 2 m at the beginning of the wind event could have been separated horizontally by 3 km within h (Fig. 5). Results clearly demonstrated that the pycnocline is an important frontal boundary (Largier, 1993) that structured the plankton community and controlled the overlap of bay anchovy larvae, their prey and predators in the water column. Previous research in Chesapeake Bay also indicated that the pycnocline influenced distribution of organisms. For example, Purcell et al. (1994) found ctenophores to be most abundant above the pycnocline in mid-chesapeake Bay. Also in the mid-bay, bay anchovy larvae were in greatest abundance above the pycnocline (MacGregor and Houde, 1996). In contrast, in baywide surveys, Rilling and Houde (1999a) and Auth (3) did not find significant differences between early-stage abundances above and below the pycnocline over a broad range of physical conditions. In the lower Bay, Govoni and Olney (1991) reported that bay anchovy eggs and ctenophores were separated by the pycnocline at a wellstratified station (eggs above, ctenophore below) but cooccurred at a well-mixed station. This suggests that degree of stratification can influence the importance of the pycnocline as a frontal boundary as well as control predatoreprey interactions. The importance of the pycnocline as a frontal boundary also may depend upon the DO conditions in the lower layer. The observed low concentrations of organisms (except sciaenid eggs) below the pycnocline suggested