Interpretation of Otolith Microstructure in Juvenile Winter Flounder ( Pseudopleuronectes americanus ): Ontogenetic Development, Daily Increment Validation, and Somatic Growth Relationships

nterpretation of Oto ith Microstructure in e Winter F Pseudopieuron

ncrement

Susan M. Sogard2 Marine FieEd Station, Institute sf Marine and Coastal Sciences, Rutgers University, Great Bay Bouleva~d, kckeston, Nj 08087, USA

Sogard, S. M. 1991. Interpretation of otolith microstructure in juvenile winter flounder (Pseudopleusonectes arnericaners): ontogenetic development, daily increment validation, and somatic growth relationships. Can. j. Fish. Aquat. Sci. 48: 1862-1 871.

In winter flounder (Pseudopleuronectes amesicanus), sagittae developed secondary origins of ealci urn carbonate deposition during metamorphosis just prior to completion of eye migration. Sagittae and lapilli of larvae were bilaterally symmetrical, but those of postmetamorphic individuals showed increasing morphological asymmetry between the left and right side. in juveniles marked with oxytetracycline and maintained in field enclosures for 18 d, increment deposition on sagittae was daily if somatic growth following marking was good (90.25 mmd-I), but less than daily in individuals with poor or negative somatic growth (43.25 mmd-'1. Narrowly spaced increments or divergence of otolith growth from the main rostral-postrostral growth axis, where counts were made, may have limited detection of daily deposition. Lack of detectable daily increments occurred primarily in larger juveniles (950 mm totat length), which had lower absolute growth rates than newly settled juveniles. In oxytetracycline-marked fish there was a significant correspondence between otolith growth and somatic growth in both length and weight. The strength of this relationship, which varied with the specific radius used, was highest ( r = 0.854) for the rostra1 radius of the left sagiata; increment widths along this radius are reliable estimators of prior somatic growth rates.

Chen la plie rouge (Pseudopleuronectes americanus), la sagitta est le si&ge de depbts de carbonate de calcium d'origine secondaire durant la metamorphose, juste avant la fin de la migration de l'oeil. La sagitta et le lapillus des larves prksentent une symetsie bilat6rale, mais chez les sujets ayant passe la m6tamsrphose, elles pr6sentent entre le cat$ gauche et le c6t6 droit une asymetrie morpholsgique croissante- Chez des juv$niles marques 5 Iroxyt6tracyc1ine et gardes 110 jours en enceintes sur le terrain, le dep8t d'accroissement dans la sagitta etait quotidien si la croissance somatique aprgs le marquage etait suffisante (>0,25 mm par jour) et moins que quotidien chez les sujets dont la croissance somatique etait mediocre ou negative (<0,25 mm par jour). Il se peut que la detection du dep8t quotidien ait 6t$ Iimitee par le fait que les accroissements etaient peu espaces ou que le developpement des otolithes s'est fait dans un plan divergent de l'axe principal de croissance rostral-postrostral, 82 oir les comptages ont $t6 faits. Les cas o12 il n'y a pas eu d'accroissement quotidien detectable ont surtout kt4 vus chez les juveniles de grande taille (950 mm longueur totale), le taux de croissance absoiu etant plus faible chez ces sujets que chez ceux qui cornmenpient le stade juvenile. Chez les poissons marques 2 Ifoxyt6tracycline, on a constate une correspndance significative entre le developpement des otolithes et la croissance somatique en longueur et en poids. Le degr6 de certitude de cette relation, qui variait selon le rayon utilise, etait maximal (r = 8,854) dans le cas du rayon rostra1 de la sagitta de gauche; les largeurs d'accroissement sur ce rayon sont donc des facteurs d'estirnation fiables du taux de croissance somatique anterieure.

Received july 20, 19918 Accepted April 4, 7 99 7 ('JA657)

atterns in otolith microstructure of larval and juvenile fishes are widely and profitably used to characterize life history traits. Increment deposition has been demon-

strated to be daily in a broad variety of species, allowing precise aging of individuals (see Campana and Neilssn 1985 for a recent review). The ability to estimate individual growth histories is supported by several experimental studies demonstrating a significant comespondence between increment widths and somatic growth in length or weight (IVilson and Larkin 1982; Volk et al. 1984; Neilson and Geen 1985; Bradford and Geen 1987; Mhossaini and Pitcher 1988; Radtke 1989). Such age and

growth information at the individual level provides the funda- mental tools for demographic analysis of fish populations.

The left md right otolith of most teleosts are largely sym- metric to each other in shape, and either may be used in aging studies (Campma and Neilssn 1985). Adult flatfishes, however, are characterized by varying degrees of morphological contrast between the Ieft and right sagitta (Smith and Daikr 1977; Nolf 1985). Such bilateral asymmetry is perhaps more likely in flatfishes, with their unique anatomical transforma- tions associated with eye migration during metiamoqhssis. Some skeletal and neurological stmctms shift in position to

'Institute of Maine and Coastal Sciences Publication No. 91-35. accommodate the migrating eye (Graf and B~~~~ 1988; 'Present address: Hatfield Marine Science Center, Oregon State A h ~ s m m et 1984), but the otolith$ do not. Thus9 when the

University, Newport, OR 97365, USA. flounder assumes its 90' rotation in body orientation, the flat,

1862 Can. J . Fish. Aquar. Sci., Vol. 48, 6996

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TYPICAL TELEOST WINTER FLOUNDER

LEFT BIDE L, L = LAPILLUS 8, S = SAGlTTA

FIG. 1 . Top view sf the position sf the otdiths in a typical telesst and in a winter flounder. The fish are shown in their usual ~OTjntation relative to the substrate.

sagittal faces of the otoliths are parallel to the benthos and spa- tially oriented with one on top of the other (Fig. 1).

In addition to information on age and growth of individual fish, the timing of specific events has also been i n f e d from otolith microstructure. Life history transitions, such as hatching or metmorphosis, often result in changes in otolith deposition, md thus a visible record of the transition event. A particularly striking change in otolith deposition occurs in the s t q flounder (Platichthys stebbatus) at metmorphosis f Campana 1984a). Calcium carbonate deposition shifts to new foci on the outer surface of the larval otolith, resulting in secondary growth centres. Deposition emanating from the primary growth centre (central primordiaam) continues until this region is enclosed by growth from these secondary centres (also known as accessory primordia or peripheral nuclei). A similar pattern of development occurs in sagittae of European plaice (Pbusonectes pkaessa) (Alhossaini et al. 1989).

The use of otolith growth to infer past fish growth relies on establishment of a functional relationship between the width of dziily increments and the daily somatic growth. Detailed information on this relationship is critical to determining the pre- cision of back-calculated lengths-at-age, but direct comparison of otolith and somatic growth in individuals is rarely accom- plished (Wilson and Larkin 1982; Bradford and C e e n 198'7; Alhossaini and Pitcher 1988). Several studies have shown the relationship between otolith growth and somatic growth to be allometric in response to varying environmental conditions. 'This uncoupling results in relatively large otoliths in fish with poor somatic growth (Mosegaard et al. 1988; Reznick et d.

1989; Seeor and Dean 1989; Secor et al. 1989; Wright et al. 1998). Species-specific differences in the scaling of otolith to somatic growth will affect the reliability of increment widths as estimators of prior growth rates.

Hn the present study, H describe the development of secondary growth centres and subsequent bilateral asymmetry in trans- forming winter flounder (Pseudupleu~snectes arnericanus) and evaluate the question of daily deposition of increments e m - nating from these secondary centres. In addition, 1 directly test the correspondence of otolith growth to somatic growth over short (10 d) time periods, using individually marked juveniles enclosed in natural habitats. These results are used to evaluate the utility of otoliths in estimating ages and growth rates of winter flounder from natural populations.

Materials and Methods

Sagittae from a series of premetamorphic larvae through late young-sf-the-yea winter flounder were examined to determine the time of development of secondary growth centres. Larvae were collected with a 1 -m-diameter, 1 -mm-mesh plankton net fished weekly on night flood tides in Little Sheepshead Creek, New Jersey. L m a e used for otolith analysis occurred in plankton collections from April 18 through May 3 1 , 1989. Juveniles were collected by push net, throw trap, and bait trawl from a variety of shallow habitats (<2 m) throughout the summers of 1988 and 1989 in the Little Egg Harbor and Great Bay estuaries of New Jersey.

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Larvae were classified into five arbitrary stages based on the degree of migration of the left eye: stage 1 = eyes bilaterally

etric; stage 2 = left eye just visible from right side; stage 3 = left eye directly on top of dorsum; stage 4 = left eye just past centre of dorsum; stage 5 = eye migation com- plete. Stage 5 individuals were fully metamorphosed and dis- tinguished from epibenthic juveniles only on the basis of cap- ture by plankton nets. Both sagittae of randomly selected fish in each stage were excised, placed in immersion oil, md examined under a microscope at 100 )< for the presence of secondary growth centres. Sagittae from epibenthic juveniles were embedded in Spum resin and polished in the sagittal plme to the cen- trd primordium on both sides, using a series of 400-1500 grit sandpaper md alumina powder (6.3 pm) .

Two radii on each juvenile sagitta were measured to provide quantitative andy sis of left-right asymmetry. These were from the central primordium to the most anterior part of the rostmm a d from the central primordium to the opposite, post~ostmm edge. Measurements were made with an image analysis system attached to m Olympus BH-2 microscope, using a magnification on the monitor of 160 or 410 x depending on the size of the otolith. The relationship between otolith radial measurements and length (SL) of the fish was determined by regression mdysis. ' F L I ~ slopes of these regressions were then compared between left and right otoliths to test for asymmetry. Two additional measures of potential asymmeq, the total length of the sagittae md the total number of postmetamovhic increments (those emanating from the innernost secondary growth centre), were csmpared between left and right with paired comphsons $-tests.

A test of the rate of increment deposition involved matching the number of increments following a tetracycline-induced fluorescent mark on the otolith with the actual number of days elapsed. Juvenile flounders (22-84 mm total length (TL), n = 216) were immersed for 24 h in a 500 mg- l - ' solution of oxytetracycline dihydrate in natural seawater (263-25 ppt) diluted with distilled water to about 17 ppt. Immediately following immersion, the flounders were measured, weighed, and individually marked with a small injection of acrylic paint. They were then transferred to 1-m%ages in the field (depth at low tide = 30-40 cm) for experiments comparing growth mong different estuarine habitats (Sogad 1990). The cage design consisted of a wood frame with stiff plastic mesh (3 mm in diameter) on the sides (46 cm high) and top and galvanized steel edges (5 cm deep) around the bottom for embedding in the sediment; they were open on the bottom to allow foraging on natural substrates. The fish were recovered from the cages after 10 d, measured, weighed, md sagittae removed and cleaned.

The number s f increments following the tetracycline mark was counted under light microscopy at 400-1000 X magnification. To provide an unbiased count of rings past the mark, 18 rings in from the otolith edge were counted under incan- descent light. The UV light was then turned on, the position of the tetracycline-marked increment detected by epifluorescence, and the discrepancy in increment counts by the two methods noted. Counts were made along the longitudinal axis from the outer edge sf both the rostrum md postrostrum. It was assumed that increment detection and differences in increment widths would be greatest on the longest axis of the otolith. Because fluorescence from the tetracycline mark was usually very dis- tinct in whole otoliths but was faint or unclear in otoliths that had been embedded and polished, d l counts were initially made

TABLE 1. Percentage of winter flounder k ~ ~ - ~ a e with secondary growth centres in the left or right sagitta. Developmental stage refers t6 position of migrating eye (see text).

Percent with Size s m o n d q centres

Developmen~l range stage (warn SL> Left Right n

on whole otoliths in immersion oil. Inc~ments on each otolith were counted on three different dates by the same reader and the results were averaged; the reader did not know the source of the otolith or results of the previous counts.

Confirmation sf these increment counts was subsequently attempted with scanning electron microscope (§EM) examination. For SEM analysis, sagittae were ground and polished in the sagittal plane, etched with 2% EDTA (pH 7-41 for 1-5 wain, and sputter coated with gol&palladium. The position of the tetracycline-mwked increment was determined by matching with photomicrographs taken under UV illumination.

Btoliths from oxytetracycline-marked fishes held in field cages were also used to determine the relationship between otolith and somatic growth. The distance from the fluorescent mark to the edge of the sagitta was measured for the four radii sf right rostrum, right postrostrum, left rostmm, and left postrostmm, at a monitor magnification of 410 or 1640 x . Individual measurements of otolith and somatic growth during the 16-d trials were compared using least squares regressions.

Results

Development of Secondary Growth Centres

Secondary growth centres in larval winter flounder sagittae initially appeared midway through eye migration (Table 1; Fig. 2B). All of the sagittae from stage 1 and 2 larvae were spherical. Almost one third of stage 3 larvae and half of stage 4 I m a e had developed at least one secondary growth centre, first evident as a small bud-like protrusion on the sagitta surface. All postlarvae with completed eye migration (stage 5) had well- developed secondary growth centres, as did all epibenthic juveniles examined (n ) 200, close-up view in Fig. 3). There was a slight tendency for secondary growth centres to initially develop on the right sagitta (Table 1). Secondq growth centres were present on the right but not the left sagitta for four larvae; only one larva had the opposite pattern.

ANOVAs c o m p ~ n g total length between individuals with and without secondary growth centres were conducted to examine whether development of these features was related to larval size as well as stage of eye migration. No significant difference was found for stage 3 larvae (F = 3.40, p = 0.080), but stage 4 larvae with secondary growth centres were significantly larger than stage 4 larvae without them (F = 8 -33, p = 8.01 3). Thus, va.riation in the timing of formation was in part due to variation in fish size.

Bilateral Asymmetry

The left and right sagittae were virtually identical in appear- mce prior to eye migration (Fig. 2A). The two otoliths diverged

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LEFT RIGHT

FIG. 2. Left a d right sagithe of winter flounder at various stages of development, showing appearance of secondary growth centres a d bilateral asymmetry. (A) Stage 2 Bma, 6.6 m SL; (B) stage 4 lama, 7.0 m SL, with developing s e c o n ~ powth centres om both the left and right sagitta; (C) recently metamorphosed juvenile, 10.6 m SL; (B) juvenile, 19.0 rnm Sk; (E) la& juvenile, 63.0 mm SL. Scde bar = 200 prn for Fig. %A-2D and 525 prn for Fig. 2E. L w d stages are defined in the text. Lmd otoliths were whole and photographed in immersion oil; juvenile otoliths were polished on both sides. BtolitRs in Fig. 2A-2C show no particular orientation; those in Fig, 2D and 2E are oriented with the anterior edge (rostrum) toward the right and the dorsal edge toward the top. The rostrum (R) and postrostmm (PR) are denoted on the left sagitta in Fig. 2E.

in morphology as growth increased from the secondary growth Regressions of radii (left rostrd, left postrostraI, right rostral, centres, with greater deposition on the right sagittib toward the md right pstrostHd) on standud length indicated highly sig- anterior, rostra1 section and greater deposition on the left sagitta nificant relationships p ig . 4). Left and right sagittae diverged t o w 4 the posterior, posfrostral region (Fig. 2D and 2E). in their deposition pattern with increasing fish size, as evi-

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FIG. 3. Photomicrograph of the central region of the left sagitta of a winter flounder (4 1.5 mrn SL). Arrows point to secondary growth centres. Scale bar = 50 km.

denced by significant differences in the slopes sf the regression lines for both the rostrum (t = 9.57, p < 0.001) and postrostmm ( d = 9-82, p < 0.001).

Although the totall length of the sagitta often differed between left and right (Fig. 5), there was no significant tendency for one side to be larger than the other (paired t-test, t = 0.03, p > 0.05). Paired counts of juvenile increments (those from the innermost secondary growth centre to the otolith edge) on the sagittae also showed no tendency for one side to have more rings than the other (Fig. 5; paired t-test, 8 = 1 -24, p > 0.85).

Dimensions of lapilli were not quantified, but visual obser- vations suggested that they also show an increasing degree sf bilateral asymmetry with increasing fish size. Lagilli were cir- cular in outline and similar on both sides in l w a e (n = 151, but left and right lapilli of juveniles diverged in morphology, primarily in the longitudinal axis (Fig. 6; n = 46). Secondary growth centres did not occur in lapilli.

Daily Increment Validation

Winter flounder juveniles held in cages in the field varied markedly in growth over the 10-d-Bong experiments. Growth in length during an experiment ranged from - 1.5 to + 13.3 m, a d growth in weight ranged from - 1.3 to + 1.6 g. Growth varied depending on the caging location in the estuary and the initial size of the fish when placed in enclosures (Sogad 1990). Flounders starting an experiment at a larger size had slower absolute growth rates than smaller flounders; negative growth occumed only in individuals >50 m TL at the start sf experiments. The wide variability in growth allowed testing sf the frequency of increment formation and the relationship between otolith mQ somatic growth under a variety of environmental conditions that generated growth rates ranging from negative to very fast-

The extent of somatic growth occurring in the 10 d after fish were marked influenced the frequency of detectable incre-

ments. The number sf increments between the tetracycline mark and the edge of the sagitta matched the number of days elapsed for faster growing fish (Table 2; Fig. 7). Counts were not significantly different from 10 except on the right postrostrum (d-tests). In contrast, sagittae from fish that grew <2.5 mm during the 10-d experiments averaged significantly < 18 increments along all four sagittal radii @-tests, all p < 0 .Q 1). Within this latter group, counts were variable and unrelated to the amount of somatic growth (all csnelation coefficients of increments counts with somatic growth were <8.38 and not significant). Lapilli from the two groups had similar results, with counts underestimated in poorly growing fish (2 = 3.9, sra = 3.1, pa = 7) but not different from 10 for fish with good growth (3 - 9.6, SD = 0.4, n = 15).

Although some material was added to the otoliths of all fishes examined, including those with negative somatic growth, the pattern of otolith deposition seemed to shift in some slowly growing individuals. In these, deposition was not concentrated along the longitudinal (rostral-postrostral) axis (Fig. 71, but was displayed-by growth in width rather than lengthy~he lack of material following the mark was not due to grinding and polishing because counts were made on whole otoliths. Little or no deposition occumd in the rostra% and postrostral regions, where increment counts were made.

Examination of 23 sagittae with SEEM produced equivocal results. The tetracycline-marked increment, determined by matching with UV photomicrographs, was typically deeply etched. Increments on either side of the mark, however, varied greatly in degree of etching. Otsliths of the fast-growing flounders had some well-etched and discernible increments deposited during the caging period, but adjacent increments, visible under light microscopy, could not be identified with SEM. Hn a few slow-growing QuveniIes, the tetracycline mark under UV microscopy could be traced in dorsal or ventral sections, but merged with the otolith edge prior to entering the rostrum or

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Standard length (mm)

Standard length (mm)

FIG. 4. Relationship of radial lengths of polished sagittae tto standard length in juvenile winter flounder. Top - rosh-al radius; bottom - pstrostral radius. Least squares regression lines are plotted and parmeters given for left and right r d i d measurements.

p skosmm. Tracing the tetracycline mark of these otoliths under SEM examination confirmed the lack of deposition on the longitudinal axis during the caging period. SEM did improve resolution of closely spaced increments not apparent under light microscopy in other slow-growing individuals with at least some rostra1 or postrostrd deposition, but still indicated <I0 increments. Resolution problems may have been responsible for some of the failure to consistently detect daily increments.

Otolith and Somatic Growth Relationships

Distances measured from the deeply etched tetracycline mark to the sagitta edge on %EM photographs were consistent with those measured on whole otoliths under light microscopy. Oto- lith deposition in the outermost portions of the rostrum and postrostrum ranged from 0 to 202 pm over the 10-Q experiments.

Otolith growth during caging experiments was significantly related to somatic growth in both length and wet weight (Table 3; Fig. 8). The strength of this correspondence varied according to the radius used in regression analysis, with the left rostrum providing the best correlations (? = 0.73 for growth

Lef t sogitta length (mm)

Le f t seg i t ta i nc rements

FIG. 5. Lengths (top) of the left and right sagitta from 61 juvenile winter flounder md co~nts (bottom) of juvenile increments (fmm the innermost secondary growth centre to the edge) on the left and right sagitta of 29 winter flounder. Increments were counted toward the rostrum of the right sagitta and toward the postrostrum of the left sagitta. Counts represent duration of the postmetamorphic stage, ignoring age of larvae prior to metamorphosis. For both length measurements and increment counts the data do not deviate significantly from the displayed line of me-to-one correspondence.

in length and 63.68 for growth in weight). Initial and final weights of each fish were Iog-transformed prior to calculating the growth in weight. For somatic growth in length, the lack of evidence of nonlinearity and the homoscedastic distribution of residuals around the linear fit suggested an isometric relationship over the size range of winter flounder examined.

Growth s f winter flounder otoliths occurs incrementally md at the periphery. Sagittal growth has only one primordium as a focus for growth during the larval period. Initiation of see- ondary growth foci at the periphery is associated with metamorphosis. Deposition from these rapidly overgrows the nuclear region faid down during the larval period, The physical appearance of the centre region of juvenile winter flounder sagittae (Fig. 3) is remarkably similar to ha t of s t q flounder (Campma 1984a) and European plaice (Kuakiri et al. 1989; Mhossaini et a1 . 1989). Other pleuronectifoms displaying sex-

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LEFT

FIG. 6. Left and right lapilli of a juvenile winter flounder (48 1 rn SE). Scale bars = 100 pm. Both Iapilli are oriented with the anterior edge toward the rigfat and the dorsal edge toward the top.

TABLE 2. Mean number of otolith increments 10 d after an oxytetracycline mark applied to winter flounder held in field enclosures. Rounden were divided a posteriori into two groups: poor growers grew <2.5 m during the 10-d experiments and g o d growers grew B2.5 mm. Counts were made along the ms$fal md postrostra1 axis of both the left and right sagitta, but increments along a11 four radii were not always legible, resulting in the different n va1nes.

Left Left Right Right rostrum postrostrum Postgum pstros4nam

- - - -- -- -

Good growers Mean 9.63 9.8 1 10.33 8.91 SD 1.04 1.85 2.54 2.17 n 19 3 1 18 21

ondary growth centres in juvenile sagittae include summer flounder (Pawbichthys dentatus) (S. Szedlrnibyer and It(. Able, unpubl. data), srndlmsuth flounder (Etropus micrsstom~) (pers . obs .), and windowpane flounder (Scophthlmus q u o - sus) (pers. obs.). Although the timing sf secondary growth centre development in these species has not yet been determined, a common rnorphslsgical pattern is evident. Secondary growth centres are not, however, unique to flatfish, occurring in many species that maintain normal body orientation as adults (Nishimura and Yamada B 884; Brothers B 984; Carnpana 1984b). The cause of the abrupt change in the pattern of deposition is presently unknown.

This study provides the first documented evidence of diverg- ing ontogenetic patterns in otolith growth between l e t and right otoliths, although Smith and Daiber (1977) noted asymmetry in sagittae of adult summer flounder. This bilateral asymmetry implies that the paired otoliths are not functioning as mimr images of each other. The cause sf this bilateral variation in

deposition is also unknown, but could be due to a mechanical consequence sf the 90" shift in body orientation or a fpmctional modification to facilitate hearing a d balance control. Graf and Baker (1988) conducted hemilabyrinthectorny operations on adult winter flounder and found that their responses differ from those observed in species that maintain bilated symmetry. They concluded that the bilateral asymmetry s f the labryinth system was necessary for the continued 98" shift in body orientation of flatfish. Thus, the asymmetry sf the otoliths is d s s likely to be related to function. Smith and Daiber (1977) noted greater deposition toward the rostrum of the left mgitta md towad the postrostrum of the right sagitta sf summer flounder, a bothid (left-eyed). 'This pattern is opposite to that found in winter flounder, a pleuronectid (right-eyed). Thus, relative to the substrate the otolith growth pattern is the same for the two species.

The appearance of secondary growth centres provides a vd- uable time maker of r ne tmo~hss i s in winter flounder. Although there is some variabilit; in the timing of their formation (Table I), due in part to l m d size, secondary growth centres are present in all winter flounder with completed eye migration. Because eye migration is a rapid process completed in about 1 wk (Chambers and Leggett 1987), estimation of the date sf metmofphosis from the date of secondiigy growth centre appearance is likely to v q within only a few days.

Daily increments have been documented in stsliths of larval winter flounder, but enumeration of the Imd increments was imprecise under light microscopy even at m early age, and SEM techniques were required (Radtke and Scherer 1982). The long larval stage duration (approximately 2 mo) (Pearcy 1962; Chambers and Leggett 1983) and small size at metamorphosis (7-9 mm) (Lippson and Morm 1974; Chambers and Leggett 1987) in winter flounder result in slow otolith growth and problems in resolving the nmowly spaced increments in the larval stage. RobBematic interpretation of larval stage increments can be bypassed and valuable infomation obtained on age and

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FIG. 7. Sagithe of two juvenile winter flounder with fluorescent marks (mows) produced by immersion in an oxytetracycline bath; photographed under UV light at 1106 x . Otolith growth outside the marks mcmmd over 18 d. The individual in Fig. 7A grew 10.0 warn (TL) during this period whereas the individual in Fig. 71% grew 0.2 mm. The latter otolith &mmstrates no growth at the outermost tip of the pstrostmman, although growth did occur dorsal to this area (at mow).

vation (Mashall and Parker 1982; Radtke and Dean 1982; C m p m a 1983; Neilsom and Geen 1985; Rice et d. 1985; k k - m a n and Wey 1987; Secor et al. 1989). Secsr a d Dean (1989) outlined a daily increment packing model, in which a minimurn amount of material is added daily to the otolith, even if the corresponding somatic growth is negative. Under their model, increments deposited less frequently than daily (Geffen 1982; h u g h et al. 1982; McGurk 1984; Siegfried a d Weinstein 1989) are aEributed to difficulty in resolving narrowly spaced increments. Jones and Brothers (1 987) improved the cone- spondence between increment counts and days elapsed by using SEM, and Campma et d. (1987) concluded that the initial increments deposited in l m a l herring otoliths were too mmow to be resolved with light microscopy.

ABhossaini and Pitcher (1 988), however, obtained results for juvenile European plaice similar to my results for juvenile winter flounder. They also found increments to be less than daily in poorly growing juveniles, md SEM techniques did not elu- cidate additional increments. Etching problems occurred for both species, preventing discrimination between resolution dif- ficulties and actual less than daiHy deposition. For a few winter %%sunder, however, tetracycline marks indicated no growth in the rostrum or postrostrarm (Fig. 71, although growth occuned in other parts 0% the otolith. All of these individuals had extremely low or negative growth during caging experiments, and calcium cabonate deposition did not appear to completely cover the otolith saBPface, resulting in wondaily increments in the sections where counts were made. For juvenile s tmy flounder, C m p m a (1983) also faad that increment deposition appeared to cease in some individuals.

In this study, increment formation rates s f < 1 e24 h- ' were found primariiy in otoliths of slowly growing fish, which were &all larger individuals. Based on caging results and length- frequency c o m p ~ s o n s of wild-caught fishes, growth rates of winter flounder were very high after settlement from the p l d - ton, but declined rapidly in older juveniles (Sogxd 1990). Dur- ing caging experiments, negative somatic growth and zero deposition dong the Ionpitudinal axes of the sagittae occurred only i r flounders >50 m TL md only in certain habitats. This lqw growth rate may not be encountered in free-ranging flounders, but misinterpretation of increment patterns appears

growth patterns from juvenile stage increments deposited out- more likely in older juveniles. A means of recognizing declin- side secondary growth centres. ing growth rates in the field, by detection of decreased incre-

Several studies have found deposition of daily increments to ment widths or some other measure, would be valuable in sug- continue during periods of poor somatic growth or even star- gesting potential interpretation problems.

TABLE 3. Results of otolith growth (pm) regressed on somatic growth in total length (mm) and wet weight (g) for juvenile winter flounder during '10-d caging experiments. Sagittal growth was measured dong four radii (left rostmE, left passtrostrd, right ro-ostaaI, md right postrostml). Initial and final wet weights were log-trmsfomd pkor to mdy sis .

Radius pa S l o p Intercept .+' F P

Sagitbcal growth vs. growth in length

Left rostrum 104 8.71 24.02 0.73 273.4 <0.001 Left p~strostmm 115 4.86 16 0.46 96.4 <$.bd01 Wight rosmm 103 8-42 46.03 0.55 125.1 <0.001 Right pstrostmm 106 5.34 23.05 0.58 143.3 <O.W1

Sagittal growth VS. growth in weight

Left rostrum 104 198.04 25.29 0.68 221.8 <0.001 Left postrostmm 115 104.25 11.38 0.43 83.9 CO.Wl Right rostrum 103 287.59 46.89 0.53 114.9 C0.001 Wight pstr~stmrn 106 124.48 22.83 0.60 158.9 <0.001

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FIG. 8. Relationship sf otolith growth to somatic growth (TL) over a 10-d period for juvenile winter flounder held in cages in different habitats in the field (8 = 0.73: n = 104). Otolith growth was measured along the rostra1 radius of the left sagitta. Results of the regression analyses for measurements along d l four radii are presented in Table 3.

Plankton sampling was conducted by Rutgers Marine Field Station personnel. I thank Steve Weiss, Susan Engels, and David Witting for their efforts in weekly sampling and Dan Rmlke for his dedicated assistance with caging experiments. John Dean, EIisabeth Laban, David Secor, and Mary Bougherty provided valuable instmetion in otolith preparation and SEM techniques. 1 thank Ken Able for editorial comments md logistical support. Steve Carnpana, Chis Chambers, and David Secor provided constructive reviews sf the maaausc~pt. H am aspeci J l y grateful to David Slecsr for his advice on otolith analysis md interpretation. Funding was provided by grants from the Electric Power Research Institute, the National Audubon Society, the Mamasquan Marlin and Tuna Club, the New Jersey Marine Sciences CsnsoK%ium, the Leathern Fund, and the Center for Coastal and Envi- r~nn~ental Studies, Rutgers University.

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Documents

Interpretation of Otolith Microstructure in Juvenile Winter Flounder ( Pseudopleuronectes americanus ): Ontogenetic Development, Daily Increment Validation, and Somatic Growth Relationships