Journal of Experimental Marine Biology and Ecology 420–421 (2012) 26–32

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Journal of Experimental Marine Biology and Ecology

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Ontogenetic trajectories of otolith shape during shift in habitat use: Interactionbetween otolith growth and environment

Matthias Vignon ⁎Univ Pau & Pays Adour, UMR ECOBIOP, INRA/UPPA, UFR Côte Basque, Allée du parc Montaury, 64 600 Anglet, FranceINRA, UMR ECOBIOP, INRA/UPPA, Pôle d'Hydrobiologie de l'INRA, Quartier Ibarron, 64 310 St Pée sur Nivelle, France

⁎ UPPA, UFR Sciences & Techniques de la Côte Basq64600 Anglet, France. Tel.: +33 5 59 57 44 48; fax: +3

E-mail address: matthias.vignon@univ-pau.fr.

0022-0981/$ – see front matter © 2012 Elsevier B.V. Aldoi:10.1016/j.jembe.2012.03.021

a b s t r a c t

a r t i c l e i n f o

Article history:Received 12 September 2011Received in revised form 19 March 2012Accepted 23 March 2012Available online 3 May 2012

Keywords:Coral reefGeometric morphometricsGrowthOntogenyOtolith morphology

Otolith morphometrics has been shown to provide a practical basis for stock discrimination and ageing. Thisapproach has been extensively used by fisheries scientists over the last decades. However, the determinantsof otolith shape are not fully understood and shape apparently results from the synergistic action of variousconfounding factors. In this study, I used a geometric morphometric approach to quantitatively investigatethe concomitant effect of local environmental conditions and ontogeny on otolith shape. I specifically focusedon the ontogenetic trajectories of otolith shape in a coral reef fish (Lutjanus kasmira) during an ontogeneticshift in habitat use, from juveniles settled in the estuary to adults inhabiting either the channel or the outer-reef off French Polynesia. Data emphasize that both ontogeny and environmental conditions influence otolithshape in an interactive way, potentially mediated by growth rate. More specifically, during the early life stagesliving in the estuary, otolith shape is mainly linked to fish size, indicating an ontogenetically determineddevelopment that induces an overall reshaping of the otoliths. In contrast, the transition from the estuarine tothe reef life is considered as a crucial phase in fish life history, as revealed by an important change in theontogenetic rate and direction of the otolith development for both habitats. After the shift in habitat use, otolithdemonstrated divergent ontogenetic growth patterns, not in terms of heterochrony but in the magnitude anddirection ofmorphological changes. This indicates that growth axis can be completely reshapedby environmentalconditions, with respect to allometric component. This information is fundamental if otolith shape is to be used infisheriesmanagement as an effective tool for modeling age-structured populations and stocks as a function of theuse made of the habitat during life span.

© 2012 Elsevier B.V. All rights reserved.

1. Introduction

Discrimination of stocks, populations and cohorts is one of the mostimportant issues in fisheries management (Begg et al., 1999; Carvalhoand Hauser, 1994; Hammer and Zimmermann, 2005), with researcheffort concentrated on measures of vital life history characteristicssuch as growth, survival and reproductive success. In this context,fisheries scientists have paid attention to the proper techniques forstock identification (Begg and Waldman, 1999; Cadrin et al., 2005;Coyle, 1998). The discovery of otolith growth increments (Pannella,1971) has allowed otoliths to be used for life-history studies of fishes atboth the daily and the yearly scale (Begg et al., 2005; Campana, 2005;Campana and Neilson, 1985; Campana and Thorrold, 2001). Theirpeculiar chronological and chemical properties has allowed otoliths tobe considered as permanent recorders of environment/habitat charac-teristics experienced by a fish during its life history (Campana andCasselman, 1993; Campana and Neilson, 1985), hence inferring stock

ue, 1 Allée du parc Montaury,3 5 59 54 51 52.

l rights reserved.

structure, population connectivity and dynamics. As a consequence,otolith's growth patterns have been extensively studied over the last20 years for fisheries purposes.

Similarly, the saccular otolith (sagitta) is characterized by highmorphological variability that has proven to be a useful tool fordiscriminating between local fish stocks and populations (e.g. Bollesand Begg, 2000; Castonguay et al., 1991; Mérigot et al., 2007;Stransky, 2005). Although stock discriminations made using otolithmorphometrics are common, the determinants of otolith shape arenot fully understood. It is currently recognized that otolith growthand shape occur under a dual regulation: overall shape is regulatedgenetically (Cardinale et al., 2004; Lombarte and Lleonart, 1993;Reichenbacher et al., 2009; Vignon and Morat, 2010), but there is astrong variability related to sex, age, year class and diet (Begg andBrown, 2000; Cardinale et al., 2004; Castonguay et al., 1991; Gaglianoand McCormick, 2004) aswell as by local environmental conditions orfactors such as depth, water temperature and substrate type (Gaglianoand McCormick, 2004; Hüssy, 2008; Lombarte and Lleonart, 1993;Mérigot et al., 2007). In addition, ontogenetic allometry has also beenconsidered as a significant factor in otolith shape determination(Cardinale et al., 2004; Lombarte and Lleonart, 1993). Thus, the

100

50

1020

1 km

Papetoai

OpunohuBay

5 km

N

EstuaryChannelOuter slope

Moorea Island

Channel

Outer slope

Estuary

Fig. 1. Map of Opunohu Bay (north coast of Moorea Island, French Polynesia) showinglocation of collection sites and bathymetry: estuary (■), channel (▲) and outer slope(●). Arrows indicates ontogenetic shift in habitat use.

27M. Vignon / Journal of Experimental Marine Biology and Ecology 420–421 (2012) 26–32

understanding and characterization of the otolith shape variationduring ontogeny is of interest since it may act as an importantconfounding effect, hampering discrimination among individuals. Inthe literature, most of the studies were based on indirect evidence,implying that otolith shape vary to some extent according to age (Birdet al., 1986), size (Campana and Casselman, 1993;Mérigot et al., 2007)and year-class (Bolles and Begg, 2000; Campana and Casselman, 1993;Castonguay et al., 1991). In contrast, there are few comprehensivereports that have directly focused on ontogenetic trajectories ofotolith shape (Capoccioni et al., 2010; Hüssy, 2008; Monteiro et al.,2005) and none have specifically investigated the synergistic influenceof environmental factors and ontogeny on otolith shape. Althoughotolith shape varies during ontogeny and may be affected by a widerange of environmental factors, in an age-dependent manner, it is stillunclear to what extent and how they affect otolith shape. Becausestudying the otolith of wild-collected specimens integrates all pastontogenetic and environmental variations, such understanding wouldstrengthen otolith-based fisheries recommendations andmanagement.

The specific investigation of environmental factors affecting otolithshape is difficult to analyze forwildfish in natural systems and generallyrequires habitat manipulations, most of them being conducted inexperimental design under controlled parameters (e.g. Cardinale et al.,2004; Gagliano and McCormick, 2004). Alternatively, the study offish that undergo an ontogenetic shift in habitat use could provide aninteresting approach to evaluate the relative contribution and potentialinteraction of ontogenetically constrained processes and environmen-tally induced plasticity.

In coral reef ecosystems, mostmarine fishes have a stage-structuredlife history consisting of a pelagic larval stage that can disperse followedby a relatively sedentary juvenile and adult stage. At the end of theplanktonic stage, postlarvae colonize shallow waters and occur innurseries.Mortality is high during and immediately after the settlementevent (Sale, 2002), withmangroves, seagrass beds and estuary typicallyoffering attractive habitats that promote survival and growth forsettling postlarvae and developing juveniles. However, as an individualgrows, its morphology and behavior change and ultimately fish at theadult stage can no longer obtain the resources they require for lifeprocesses and they move to new location that supports their lifeprocesses. Adults ultimately settle on various substrates/habitats fromthe shoreline and fringing reef to channel, barrier reef and outer slope.Many coral reef fish exhibit such bipartite life-history characterized byan ontogenetic shift in habitat use (Gillanders et al., 2003; Moura et al.,2011; Nagelkerken et al., 2002). Around Moorea Island (SocietyArchipelago, French Polynesia), juveniles of the bluestripe snapperLutjanus kasmira (Forsskål, 1775) typically occur in estuarine/mangrovenurseries, and adults are found frequently in large aggregations aroundcoral formations in either the lagoon, the vertical slopes of channels orthe surrounding outer-reef. Migration patterns as adult between thesethree habitats and residence of specimens in a specific habitat haverecently been investigated using both otolith shape and parasites(Vignon et al., 2008). While fishwere all sampledwithin a few hundredmeters, they exhibited significantly different parasite fauna and otolithshapes according to their locality. Data from Vignon et al., 2008 haveemphasized that adults rarely move between their three adjoininghabitats, and more interestingly that large aggregations in the channelare formed predominantly by resident individuals with limited localmovement. In this system, larvae start growing in the same environ-mental conditions and subsequently shift to contrasted habitats,allowing investigating the ontogenetic trajectories of otolith shape,with specific consideration for its interaction with environmentalconditions. It is also of interest in the perspective of understandingshape flexibility and reshaping capability of otoliths.

The objective of the present study is to examine the synergisticinfluence of environmental factors and ontogeny on otolith shapevariation. More specifically, I investigated the ontogenetic trajectoriesof otolith shape in a coral reef fish that exhibits a shift in habitat use

during growth (the juveniles of L. kasmira settle in the estuary,whereas adults subsequently live in the channel or the outer-reef). Ialso evaluated the degree to which growth variability occurs amongindividuals and to which extent environmental conditions affect theontogenetic trajectories of otolith shape.

2. Material and methods

2.1. Sites and fish collection

A total of 222 L. kasmira (38 larvae, 44, 71 and 69 individuals,respectively in the estuary, the channel and the outer slope) wascaught during 2006 and 2007 in the north coast of Moorea Island (17°30′ S, 149° 50′W; Society Archipelago, French Polynesia, Fig. 1). Allfish except larvae were speared at a depth ranging between 3 and20 m. Three larvae were directly sampled on the reef using handnets and soft anesthetic (mix of ethanol/clove oil 4:1). Thirty-fiveadditional recruiting larvae (passing through the reef crest a fewhours/days before they settle onto the reef) were sampled on theouter reef crest using a net fixed directly on the substrate. The net waslocated in the surf zone at the highest point of the crest. Standardlength (SL) of individuals was measured to the nearest mm. The ageof individuals was estimated by transverse sectioning of each otolithaccording to standard procedure (e. g. Morales-Nin, 1989).

2.2. Otolith shape analysis

Otoliths were extracted, cleaned gently with water and air-dried.Each unbroken otolith was placed with the sulcus acusticus orientedtowards the observer and then examined under a stereomicroscopefitted with a digital camera (XC-77CE) linked to a computer. The highcontrast images were thresholded and binarized for contour extrac-tion by ImageJ software (freely available at http://rsb.info.nih.gov/ij/).There are two main alternative methods for quantitative descriptionof a curve, such as an outline: Fourier Series which approximates thecontour of the object by passing through a series of progressivelymore complex trigonometric functions and Semi-landmark-basedmethods that incorporate information about curves into the classicallandmark-based formalism (Bookstein, 1997). Although the two

28 M. Vignon / Journal of Experimental Marine Biology and Ecology 420–421 (2012) 26–32

approaches allow very good approximations of the shape of an object,produce roughly equal efficiency of discrimination/classification andgenerally lead to similar conclusions about shape (e.g. Loy et al., 2000;Ponton, 2006; Sheets et al., 2006; Van Bocxlaer and Schultheiß, 2010),the results of Fourier Series do not encourage intuitive understandingof shape differences between objects (Bookstein et al., 1982). Themathematics of Fourier analysis precludes the decomposition ofshape into single characters that describe localized change sincelocalized features are necessarily spread across multiple harmonics.In contrast, the aligned landmark coordinates using geometricmorphometrics are often fitted to an interpolation function such asa thin-plate spline, which can be decomposed into global (affine) andlocal (nonaffine) components. Otoliths have recently been success-fully studied using geometric morphometrics (Monteiro et al., 2005;Ponton, 2006; Ramírez-Pérez et al., 2010; Vignon and Morat, 2010),providing the proper framework in which to visualize and quantifygrowth and ontogenetic trajectories. In this context, while shapeanalyses of otolith 2D contour are generally performed through Fourieranalysis, a semi-landmark-based method was preferred for analyzingallometric patterns.

2.3. Morphometric analysis

The ontogenetic shape changes of otoliths were studied usinglandmark-based geometric morphometric methods (Bookstein, 1991).The outline of each specimen was stored as pixel coordinates that werereduced to 100 points equally spaced (using contour length as aparameter) along the contour, starting at the intersection of the majoraxis with the anterior margin of the otolith (Rostrum). This referencepoint (biologically corresponding among individuals) is referred to as alandmark and the remaining 99 points as semi-landmarks. Thecoordinates of the landmark and semi-landmarks were digitized usingthe software tpsDig version 2.10 (from the TPS package, Rohlf, 2006).Generalized orthogonal least square Procrustes superimpositions oflandmark and semi-landmark coordinates were then computed withtpsRelw version 1.45, minimizing bending energy with respect to amean reference form (Bookstein, 1997) to determine the criteria forsliding semi-landmarks along outlines. Partial warp scores (PW)including both uniform and nonuniform components were calculatedand used as descriptors of variation in shape (Bookstein, 1991). Theallometric patterns of shape variation were calculated by multivariateregressions of PW scores on log-transformed SL (Monteiro, 1999;Zelditch et al., 2003) using TpsRegr, version 1.28 [the matrix B of least-squares estimates of the regression coefficients was calculated by usingthe standard formula B=(XtX)−1XtW where the matrix W is thedependent array of variables (PW scores), and the matrix X containsthe array of independent variables in its columns. The t operatorindicates transposition]. The SL was log-transformed (log-SL) becausethe allometric relationships are better described by a linear model thattakes into account the progressive decrease of the rate of shape changeduring growth. The fit of the regression models was evaluated by theexplained variance of the model and by Goodall's F-test.

The amount of the overall shape differences between otoliths wasestimated using the Procrustes distances (PD), which is a propermetric for shape dissimilarity in the Kendall shape space. This distancewas used as a univariate measure of shape difference, but needs to beconsidered as an overall measure of multivariate shape components(i.e. partial warps). Dynamics of shape changes are visualized by plotsof PD between each otolith and the average shape of the smallestlarvae found in the estuary and the reef crest, on their SL. The averageshape of larvae was calculated based on 38 larvae with SLb40 mm,corresponding to the length of fish at colonization (Lecchini andGalzin, 2005).

More than one principal component generally shows associationsto size and non-allometric shape variation may carry along allometricfractions of variance. Such overlap may confound the identification

of underlying shape variations. In this context, I used a multivariateanalysis of covariance (MANCOVA) to specifically disentangle theunique shape changes that are associated with pure growth allometryand that portion of shape variation that is independent from thoseallometric factors.MANCOVAwas performed using thewhole ontogenydataset, testing the null hypothesis of homogeneity of linear allometricmodels (the expectation that the ontogenetic trajectories between twohabitats do not differ in their slope. Multivariate regression lines arethen either parallel or coincident). In this test, uniform components andnon-uniform PW scores are considered as variates, log-SL as covariateand habitats as grouping factor. This statistical analysis was performedusing the program TpsRegr. When the allometric models differed forthe two habitat, the differences were analyzed by comparing the anglebetween the habitat-specific multivariate regression vectors usingVecCompare6 and SpaceAngles6b [from the integrated morphometricspackage [IMP], (Sheets, 2003)] (for detailed explanation, see Zelditchet al., 2003). Briefly, to examine how similar the directions of shapechange are between the two ontogenetic trajectories, I computed twowithin-habitat growth vectors. The range of angles between such vectorswithin each habitat is calculated using a bootstrapping procedurefrom the IMP software (each within-habitat dataset is resampled withreplacement and the calculation of the angles between the trajectories isrepeated. N=3600). This range was compared with the angle betweenthe vectors of both habitats. The two trajectories are statistically differentif the angle between two growth vectors is significantly different fromzero. This null hypothesis was tested by comparing the angle betweentwo growth vectors from different habitats to the 95% confidenceinterval of angles produced by comparisons of growth vectors withineach habitat.

To visualize localized otolith shape differences among habitats andontogenetic stages, I generated thin-plate spline deformation grids(Bookstein, 1991)with scaling optionα=0along the first canonical axisusing tpsRegr. Thin-plates were not exaggerated (×1 magnification).

3. Results

3.1. Allometry and allometric models

Otoliths from the larvae are highly uniform in shape (Fig. 2), asrevealed by a mean PD among pairs of otoliths of 0.008±0.003 (as acomparison, the PDs of the most similar otoliths found in the channeland the outer reef are respectively 0.10 and 0.11). The PD betweenaverage shape of larvae from the estuary and from the reef crest isonly 0.005, indicating insignificant differences between the two larvalsources. This consequently allows to reliably analyze the dynamics ofshape changes during ontogeny that is expressed as the PD betweeneach otolith and the average shape of larvae. The null hypothesis ofisometric growth is rejected. Otolith shape is highly significantlyallometric when partial warp scores and uniform components areregressed onto log-SL, as supported by the Goodall test (Table 1,Goodall's F-test Pb0.0001). The significant regression model in shapespace accounts for 29.9% of the total variance when all individuals areincluded in the analysis. The degree of allometry in otolith fromadults is smaller but significant relative to that in juveniles (64.5% and57.3% of explained variance, respectively for the channel and theouter slope, compared to 76.8% in the estuary, Table 1). The lowerpercentage of variance explained in adults indicates that somevariability in shape is possibly due to factors other than size or thatthe ontogeny could be nonlinear (based on log-values of standardlength). As revealed by the sharp decrease of F-values in the channeland the outer reef compared to the estuarine (Table 1), results indeedtend to indicate that otolith shape is highly uniform at smaller than atlarger sizes.

MANCOVA is highly significant (Pb0.0001), confirming the allome-tric trend in otolith shape in adults, and showing that the same linearmodel cannot be employed for the description of the ontogenetic

Fig. 2. Overall superimposition of otolith outlines from the 38 fish larvae (same scale).The mean Procruste distance among pairs of otoliths is 0.008±0.003. The distancebetween average shape of larvae from the estuary and from the reef crest is 0.005.

0.00

0.05

0.10

0.15

0.20

SL (cm)

0 5 10 15 20 25 30

Proc

rust

es D

ista

nce

EstuarineChannelOuter slope

Fig. 3. Dynamics of shape changes during ontogeny expressed as the relation betweenProcruste distance and standard length of fishes. Deformation grids (thin-plate, ×1magnification) depict shape differences among four individuals from the differenthabitats. Procruste distances and deformation grids were computed using 38 larvae(SLb40 mm) as reference. Error bar corresponds to the mean Procruste distance amongpairs of otoliths from these 38 larvae.

29M. Vignon / Journal of Experimental Marine Biology and Ecology 420–421 (2012) 26–32

allometries in the habitats (Table 2). The differences in the modelscould be attributed to different rate in shape changes or to differentontogenetic trajectories of shape. Dynamics of shape changes duringontogeny for each habitat, expressed as the relation between PD andSL, are given in Fig. 3. The trajectories are biphasic and express anasymptotic trend, with a decrease in the rate of ontogenetic shapechanges, up to an apparently stable stage. The main changes in growthrate of these biphasic trajectories clearly appear to correspond to theshift in habitat use. The PDs between the average shapes at settlementin the estuary and at the adult stage show that the ontogenetictrajectory is longer in the outer reef, emphasizing that otoliths undergomore shape modifications during ontogeny (Figs. 3 and 4). In addition,analysis of the angles between vectors of ontogenetic allometrieswithin and between-habitats showed that themean angle between thechannel and the outer-reef is 127.9° and thus higher than the ranges ofthe within-habitat angles (31.5° for the channel and 53.4° for the outerreef, angles are in decimal degree, Table 3). As a conclusion otolithsfrom the two habitats therefore differ significantly in both their rateand trajectory of shape changes.

4. Discussion

4.1. Otolith shape variation

As already emphasized, the intraspecific quantitative significanceof allometric shape changes in otoliths were poorly investigated(Capoccioni et al., 2010; Hüssy, 2008; Monteiro et al., 2005),particularly in relation to habitat shift. This study provides newinsight into otolith shape formation mechanisms. It allows examiningwhether shape development is an ontogenetically determined processand determining the effect of environmental factors in snappers frompostlarvae/juveniles to adults. Otolith growth continues throughoutlifetime and is based on a genetically guided mechanism (Gauldieand Nelson, 1990). However, while otolith shape is geneticallyconstrained, such growth patterns in the calcified structures may be

Table 1Fit of multivariate regressions of otolith-shape versus log-values of standard length(Log-SL) for coral reef fishes.

% Explainedvariance

Goodall's F-test

F-value P

All individuals 29.918 79.2392 b0.0001Estuarine 76.818 259.8848 b0.0001Adults (channel) 64.511 210.1518 b0.0001Adults (outer reef) 57.278 201.8864 b0.0001

affected/influenced by a wide range of exogenous factors, producingvariation among conspecific individuals that have experiencedcontrasted life histories.

During the early life stages, otoliths are still small with a relativelylarge accretion rate and can therefore be reshaped by environmentalfactors in a significant manner (Hüssy, 2008). The ontogeneticdevelopment from the circular, larval otolith to the oblongate adultotolith is indeed known for a wide range of species (see Hüssy, 2008).Interestingly, although otoliths of post-larvae and juveniles are proneto distort/deform, ontogenetic trajectories in the early life-stages arecongruent, as revealed by a singular restricted shape disparity amongindividuals from the estuary. Although fish were collected in the wild,inter-individual differences are unlikely to exhibit large variationsin this particular system. Results from the present study appearconsistent with the observations of Capoccioni et al. (2010) on eelsthat otoliths are highly uniform in shape at the smaller sizes comparedwith the larger ones. This contrasts with one study focusing on 11species of the genus Merluccius that emphasizes an importantenvironmental influence on otolith growth increment in immaturefish, compared to adults (Lombarte et al., 2003). Data generallysuggest that the development of otolith shape is indeed an ontoge-netic process reaching beyond the earliest life stages, that can reshapethe overall otolith outline, regardless of individuals and small-scaleenvironmental conditions. The reason for such early stabilitywould bea biological constraint posed by otolith function as a sound transducer(Gauldie and Nelson, 1990). Practically, proteins within the endolym-phatic epithelium act as a significant component of the mechanismcontrolling the shape of the otolith and endolymph physiologicalconditions allowing bio-mineralization are under genetic control andregulation (Borelli et al., 2003; Morales-Nin, 2000; Söllner et al.,2003). This may explain why the early shape development appears asan ontogenetically determined process, with otolith growth followingthe accretion of either calcium carbonate or protein. In any case, therate of this shape change is high, suggesting the need/constraint fora rapid morpho-functional shift.

Table 2Full factorial MANCOVA of partial warps and uniform components scores, pooledsample (N=222). The level of significance is assessed using Wilk's test.

λWILKS F P

Intercepts >0.00010000 b5.000 b0.0001Log-SL 0.00046401 2059.353 b0.0001Habitat 0.00028220 54.185 b0.0001Log-SL∗habitat 0.00000000 6467076.364 b0.0001

Affine Non-affine

Specific componentsOverall shape deformation

Lar

vae

Est

uary

Out

er s

lope

Cha

nnel

Est

uary

Fig. 4. Morphometric differences of otoliths between the three main habitats occupied by fishes during ontogeny. Complete deformation (left) of major growth warps and specificcontribution (right) of localized (non-affine) and uniform (affine) components. All at ×1 magnification.

30 M. Vignon / Journal of Experimental Marine Biology and Ecology 420–421 (2012) 26–32

In contrast, the transition from the estuary to the reef is consideredas a crucial phase in fish life history, as revealed by an importantchange in the ontogenetic trajectory of the otolith development forboth habitats (Fig. 3). Growth rates of fish and otoliths depend notonly on the recent but also on the growing inertia due to past growth(Gutiérrez and Morales-Nin, 1986). The strong autoregressive natureof otolith growth causes inertia in the growth processes that buffersthe otolith from exogenous influences and induces a lag betweenotolith response and environmental perturbations (Maillet andCheckley, 1991; Morales-Nin, 2000). This makes unlikely that shiftin habitat use directly affects the ontogenetic direction of otolithdevelopment. However, the present data surprisingly emphasize thatshift in habitat use not only changes the magnitude and rate of otolithdevelopment but also changes the overall direction of the otolithdevelopment (Fig. 4 and Table 3). After their initial growth innurseries (i.e. estuary), the larvae colonize surrounding reefs andsettle. While such ontogenetic shift in habitat use does not coincidewith important changes inmorphology and physiology (Dahlgren andEggleston, 2000; McCormick and Makey, 1997), evidence suggeststhat the magnitude of environmental change and/or feeding behaviorexperienced by snappers migrating to the reef can significantly act onthe otolith shape. This indicates that the growth axis (ontogeneticgrowth direction) of otolith can be completely reshaped by environ-mental conditions during ontogeny. Similar findings were recently

Table 3Pairwise comparisons for hyperplane angle differences between the three habitat-specific growth vectors. Mean and 95% confidence interval result of bootstrappingprocedure (N=3600) comparing the multivariate regression vectors. Angles are indecimal degree.

Pairs ofgrowth axes

Within habitatvariation

Between habitatvariation

Vectorcorrelation

Lagoon 16.5 (12.8–19.5) 72.6 (64.7–93.3) 0.30Channel 31.5 (25.7–26.8)Lagoon 16.5 (12.8–19.5) 83.9 (83.5–84.4) 0.11Outer reef 53.4 (41.2–62.4)Channel 31.5 (25.7–26.8) 127.9 (114.5–150.3) −0.61Outer reef 53.4 (41.2–62.4)

obtained in two tropical damselfishes during settlement in Australia'sGreat Barrier Reef (Baumann and Gagliano, 2011). Data from theseauthors illustrate an apparent rapid (i.e. within 12–15 days) change inotolith morphology between pre- and post-settlement stages. How-ever, their data still remain inconclusive regarding the non-exclusivemechanisms responsible for this apparent morphological change(selective mortality or flexible growth pattern).

On a theoretical basis habitat heterogeneity may mediate therelationship between organisms and their environment. However,most of the studies were carried out over large geographic scales,such as latitudinal gradient and marine ecological regions (e.g. Bollesand Begg, 2000; Lombarte and Lleonart, 1993). In contrast, studiesdealing with small-scale variations (i.e. over a few hundred meters)are scarce (Vignon et al., 2008) and mainly considered local pollutionor anthropogenic disturbances. The paucity of studies at a small scalemay reflect the lack of expectation that local habitat heterogeneitywould affect otolith shape over short distances in the absence ofgenetic differences. However, I found significant differences in otolithshape between the adults inhabiting the channel and the outer reef.In addition, shape divergence increase with age/size, emphasizingthat habitat can mediate the developmental ontogeny of otolith in asignificant manner, putatively through growth rate. Previous analyseshave suggested that adults rarely move between adjoining habitatsand large aggregations are formed predominantly by residentindividuals with limited local movement. In this context, environ-mental conditions experienced by adults may locally vary amongindividuals on a permanent basis. Various environmental factors suchas water temperature (Cardinale et al., 2004), depth (Gauldie andCrampton, 2002; Lombarte and Fortuno, 1992), type of substrate(Mérigot et al., 2007) and feeding conditions (Gagliano andMcCormick,2004) are thought to influence fish growth that in turn can affectotolith growth, hence producing variations in otolith shape in theabsence of genetic differences (Simoneau et al., 2000). For example,Gauldie and Nelson (1990) found that faster growth produced longthin crystals compared with shorter more compacted ones in slowergrowing fish, which could influence the overall shape of the developingotolith. While uncoupling of somatic and otolith growth rates maytemporarily occur (Baumann et al., 2005; Fukuda et al., 2009;Mosegaard

31M. Vignon / Journal of Experimental Marine Biology and Ecology 420–421 (2012) 26–32

et al., 1988; Wright et al., 1990), numerous studies have documenteda link between otolith shape differences and somatic growth rates(Campana and Casselman, 1993; Casselman, 1990; Reznick et al., 1989;Simoneau et al., 2000). In Moorea Island, hydrodynamism generated bytide and draining of the lagoon create original habitats in the channel.This zonation may results in different prey availability or physico-chemical composition of the water between habitats distant from eachother by only a few hundred meters. Considering differential consump-tion and growth rates between habitats (Vignon et al., 2008), this mayexplains to some degree the differences among individuals as adultsinhabiting the channel and the outer reef. Nevertheless, I recognize thatfurther investigations are required to specifically disentangle the effectof underlying factors and investigate whether differences are related tochanges in otolith microincremental growth.

As a conclusion, differences among individuals in the early life-stages seem to be governed by the genetically determined nature ofotolith shape development, while differences among adults may bethe result of a combination of genetic and environmental effects, henceincreasing the interindividual disparity. In this context, different shapeswould result in different mechanical efficiencies for hearing at differentfrequencies (Gauldie and Nelson, 1990; Popper et al., 2003), suggestingthat shape-based functionality may impose a strongest selectionpressure on post-larvae and juveniles than on adults. Alternatively,increasing disparity may reflect the individual phenotypic response toenvironmental conditions, suggesting developmental constraints inlarval and juvenile stages. As a consequence, the different ontogenetictrajectories of snappers from the adjacent localities are unlikely to beinfluenced by environmental conditions experienced by the larvaewithin the estuary. Instead they are more likely to result from differentenvironmental conditions experienced by adults.

4.2. Implications in fisheries management

Based on the incremental properties of calcified structures, thechronological characteristics of otoliths allow accurate estimates ofgrowth and age at both the daily and the yearly scale. Traditionally,daily otolith increments are counted along a linear transect from thecore to the edge of thin transversal sections of otolith (commonlyalong the longest axis of the cross-section). As a consequence, thesevere change in otolith morphology during ontogeny renders otolithincrement widths along the transect inconsistent proxies for dailysomatic growth through habitats (Baumann and Gagliano, 2011). At awider scale, knowledge of the age profile of a fish population is ofprimary importance for creating age structured population models,hence allowing stock assessment and management (Cadrin et al.,2005; Campana and Thorrold, 2001). However, traditional large-scaleroutine reading of age (i.e. based on pattern of growth rings) isdifficult, excessively time consuming and still not very reproducible,hence limiting sample size and producing ageing errors that propagateup to stock assessment (Brander, 1974; Kimura and Lyons, 1991). In thiscontext, some pattern recognition systems have been proposed foraccomplishing automatically this task based on extracting differentkind of features fromfish otoliths (Bermejo, 2007; Bermejo et al., 2007).The use of two dimensional images may provide a useful ageingtechnique avoiding any extensive handling or manipulation of theotoliths. Several attempts such as the European project FABOSA (FishAgeing By Otolith Shape Analysis) intended to develop an automaticfish age prediction technique based on themeasurement of the externalshape of otoliths (Doering and Ludwig, 1990; Doering-Arjes et al., 2008;Steward et al., 2009). Using this approach, descriptions of age-class andhabitat achieved high discriminant accuracy. Alternatively, includingotolith morphometrics into ageing processes has the potential to makeageing unbiased, accurate and fast (Bermejo, 2007; Doering-Arjes et al.,2008). This emphasizes that the synergistic combination of direct agereading and shape information achieves a greatest accuracy. In eithercase, the incorporation of age and its interaction with environmental

conditions appears as an important component in an otolith shape-based classifier that remains to be included in traditional large-scaleroutine reading of age.

In the context of stock discrimination, future studies should alsoaddress the interaction between ontogeny and environmental factorssince fish of various sizes are generally involved (Burke et al.,2008; Campana and Casselman, 1993). The lack of age structuredpopulation models hinders researchers from refining strategies formanaging fishery that should integrate data on both juveniles andreproductively mature adults. This study highlights the importancefor considering ontogenetic trajectory to distinguish between groupsof fish that are separated at a particular point in their life history. Theontogenetic trajectories showed that larger individuals were moreefficiently classified than the smaller sized ones in the different habitats.The present data suggest that a differential habitat conditioningmediates otolith development, hence producing otolith shape differ-ences during the later growth stages, not in term of heterochrony (i.e.alteration in the timing of ontogeny such that the relative time ofappearance or rate of development of a feature is either accelerated ordelayed) but in the magnitude and direction of morphological changes.

In both contexts (stock discrimination and age reading), a numberof confounding effects must be considered to ensure that classifica-tion is unbiased and accurate. Because otolith shape apparentlyresponds to some degree to various factors, their relative importanceis not fully understood and they can act as confounding effects. Thesomatic and otolith growth rate may vary among habitats, andconsequently, habitat and otolith development may strongly interactduring ontogeny. This study highlights that habitat and ontogenyinduces an important change in otolith shape that must be taken intoconsideration in a synergistic manner. Although ontogenetic changesin otolith morphology have been reported, many studies to date havechosen otolith morphometrics without consideration of potentialspecies-specific otolith ontogenetic growth patterns. This importanteffect can compromise stock discrimination and age reading studies ifvariables are not standardized. Thus, the development of an otolithgrowth model is needed for standardization/calibration if otolithshape is to be used as an effective tool for management of fisheriesresources in the future. Unfortunately, few papers have thoroughlyinvestigated the presence of allometric changes in otolith shape andthe present data suggest that both habitat and ontogeny inducecomplex and non-uniform (i.e. non-affine) changes in otolith shapethat affect the overall otolith shape. Further specific investigations arestill required to comprehend the complex nature of otolith shape andthe use of morphometric geometrics appear as a promising tool forichthyologists that can be used in a wide range of contexts. This mayalso be helpful for the recent interest in automated pattern recognitionssystems developed to accomplish tasks based on extraction of differentkind of features from fish otoliths.

Acknowledgments

Financial support for this research was partly supported by USR3278 EPHE-CNRS CRIOBE. The author thanks Fabien Morat for his helpin photographing otoliths and Edward Beall for revising this manu-script. The author is also grateful to the anonymous reviewers whomade highly valuable suggestions for improving the manuscript. [RH]

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