INTERNATIONAL COUNCIL FOR
THE EXPLORATION OF THE SEA
C.M.199O/G:29
Demersal Fish Committeel
Ref. Pelagic Fish Committee/
Baltic Fish Committee
•
ANNUAL CHECK FORMATION IN OTOLITHS AS A STARTING
POINT FOR AGEING BY MEANS OF COMPUTER ANALYSIS
by
H.C. Welleman
Netherlands Institute Car Fishery Investigations
P.O. Box 68.1970 AB Umuiden
The Netherlands
INTERNATIONAL COUNCIL FOR
TUE EXPLORATION OF TUE SEA
C.M. 1990/G :29
Dcmersal Fish Commiuee
Ref: Pelagic Fish Committee I Baltic Fish Committee
Annual check formation in otoliths as a starting point for ageing by means of
.
computer analysis•
. by
Henny C. Welleman
Netherlands Institute for Fishery Investigations
P.O. Box 68,1970 AB Umuidcn
The Netherlands .
•
Abstract
This paper is an aUempt to summanze the basic principles behind commonly used ageing
rmitines of otoliths. Age dctermination is based on the intcrpretation of optically distinct
zones. However, there is still no theoretical basis for a1locating a time seale to th6 visible ring
pauem. The reliability of the decision-making process in ageing ean be improved when
subjective interpretations are eliminated and when the chemistry beyond contrast enhancing
techniques is fuHy understood. An analytic approach of age determination in terms of calcium
mineralization and physiology is therefore in this study the first step in the development of an
automatie ageing system.
1. Introduction
Growth and mortality represent two population processes that are essential to the assessrrient
and management of fisheries. Estimation procedures for rates of change in weights and
numbers depends heavily on kriowledge of the age structure of the populations and ageing of
fish can be considered the back bOne of fish stock assessment. Interests in ageing methods
go back many years and calcareous structures were first used for ageing pike in 1759
(HEDERSTRÖM, 1759).' Scales, otoliths, opercular bones, fin rays, spines, cIeithra and
vertebrae are commonly used in modem times. The basic principle is that the age of fish can
be deduced from the ring features identified on these structures, when a time scale can be
allocated to the visible pattern: This latter assumption can often still be questioned and even
these days there appears to be no general theoretical basis for the age reading of hard tissues.
Sometimes, age reading is considered more an artthen a scientific activity. Even though, the
annual nature of the ring features is widely accepted and it is believed that, whenever proper
techniques can be developed, a large amount information on the life history of the individual
fish is accessible.
Ageing of fish continues to be a labour intensive routine activity infisheries research. There
is a cIear need for less time consuming and ~ore objective ways to produce age compositions
and it has been suggested that the use of image analysis systems could improve the efficiency
and reliability of age detenIiimition (e.g. FAWEu., 1974). As a first step, an understanding of
the structural characteristics, chemical cOluposition and formation of otoliths would make it
feasible to deterriline suitable enhancement-technique.
a
Not all studies relating to age reading can be summarized in a short paper. Thercforc a
seIection of topics in otolith reading has been made. .arie of thc major advance iri ageing of
fish, thc discovery of daily growth zoncs (PANNELLA, 1971), is biiefly discussed. The
currerit trend in this topic has reccntly been weIl rcviewed, but thc implications for the
determination of yearly, cvents in otoliths and thc process of ageing, are not yet fully
unders~Ood (BEAMISH AND ~CF~A,NE, .1?87). ~liercas daily growth increinents appear
to form as a consequence of dIUrnal rhythm In the mobilization of calcium (MUGlYA, 1987)
and neuropeptidc prodtiction (GAULDIE AND NELSON, 1988), anrimil check rings ofotoliths
are supposed to have a more complicated origin. Soine basic chemical and mineralogical
aspects of otolith mineralization mechanisms will 00 discussed arid related to currently used
ageing techniques.
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2. Otolith microstructure
In teleost fishes, the sagiuae are the largest ofthe three otoliths und appareritly thc most easy
to usc in ageing. Theyare composed of calCium carbonate crystals deposited on an organic
matrix (CARLSTROM, 1963). The mineral part is aragonite andthe organic part is called
otoHn (DEGENS er al., 1969). In comparison with skeletal bones there are some basic
differences. The organic compound of bonc are mucoproteins with a different amino aCid
composition than otolin. Most importantly, osteogencsis in bone is either cellular or acellular
rind osteoblasts or scleroblasts are involved, while no such cells have bien repoi1ed iri otolith
mineralization.
OtoÜths are sUITounded by a fluid medium in the labyrinth. The oniy conriectivc iissue is the .
sensory epithelium of the macula actistica. SensOlY hairs of this nervous-end material project
in a membrane that cover the 'sunace of the otolith. This niembrane is probably permeable to
material diffusing from the endolymph fluid towards the otolith. An ultrastructural study of
the.otolithic membrane (DUNKELBERGER er al.~ 1980) revealed two distinct iories, a
strtictured gelatinous layer arid a non-strueturoo subcupular meshwork. The gelatinous layer
extends from thc otolith sunaee to the tips of the sensory hairs only over the sensory region.
The subcupular meshwork, not limited to the sensory region, consists of a loose network of
fibrous material. This fibrous materiat'was relatively more abundant over areas of the nonserisory region. At the periphery of the otolith, a continuation of fibrous material into the
otolith was [ound (DUNKELBERGER er al., '1980). The fuH significance of this otolithic
membrane in regulating the calcium depositiori arid/or organic iriatriees within the otolith hus
not yet been established.
2.1. The organic matrices
In aimost all eases ofbiomIneralizatiori, proteinous macromolecules are fouod in solutionand
many of them have one p~lrtieularcharaeteristic:they are highly acidic (LOWENSTAM AND
WEINER~ 1989). These protdns (otoHn, in case of otoliths) assemble to a so called organic
matrix, a three-diinerisiorial framework arid they are rieh in highly oxygenated aspartic and
glutamic acid and low in aromatic arid basic amino acids. This fibrous protein is uniform in
composition and unlikely to be affectoo by eriviforimeriÜ1I or phylogenetic events at the
inolecular level (DEGENS er aI., 1969).
Orgariic material is fourid between individual calcified laycrs, between individual crystals in
each layer, as weIl as within subunits of the crystuls (respectively termed as intedamellar.
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intercrystaiIine and intraclysiailine; DUNKELBERGER et al., 1980). In the latter type it has not
!>een resol,,:ed: ~hethe.r eachindividual su~unit is an independent crystal or, very unlikely,
Just a portlOn of a smgle crystal. The ughtly" packed fibers of the intercrystalline and
~Iiterhim~llarma~~es were s~milar to the observed fibers in the subcupular component of the
surroundmg otohthlC membrane. ,
, :
Over a wide yariety of fish species the amino acid composilion ofotolin is rather uniform.
The total organic coritent in different species varies within a range between 0.2 and 10 % of
the total otolith weight (DEGENS et al, 1969).
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2.2. The mineral form cif calcium carbonate
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StructUres of calcium carbOnate depositoo in biomineralizatio~ are mostly calcit6 or aragonite.
Calcite is the most stable fomi at low teinperature and low pressure, and aragonite (and
vaterite) transform to calcite in aquatic environments under normal conditions. Electron
diffraction studies of teleost otoliths revealed that they were crystalline in the form of
aragonite (DEGENS et al.~1969; GAULDIE AND NELSON, 1988). For instance, otoliths of
plaice have been described as polycrystalline aragonite (CARLSTROM, 1963). The great
stability of biological aragonite is assumed to becausedby the highly stable lattice of
aragonite formed in the biological system, although the exact reasons remain to be studied
(KITANO etaI., 1980).
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Aragonitic crystals in otoliths Ure orientated with their crysiallographic c-axis perpendicular to
their projected surfaces. Electron micrographs showed twinning of the individual crystals.
This twinning often occurs in biological mineralization and in the presence of foreign ions. It
is supposed to be an efficient mechanism for rapid crystal grciwth (GAULDIE AND NELSON, .
1988).
.
Nevertheless some otoliths do contain more than one mineral fOrIn of calcium carbOnate
(GAULDIE AND NELSON, 1988). In a few studies, the sagittae were v~lteritic, or partially ,
aragonitic rind pai-tially vateriiic (CAMPANA, 1983b). Also calcite is knowri to occur as a thin
hiyer on the distal surface of the otolith (GAULDIE, 1990).
Differences between geologically and biologically fonned aragonltic minerals.
Theelectron diffraction patterns of otoiithic fragments and geological aragonite have been
used to recognize the crystal structures. Growth in the three-dimensiorial plane of mineral
aragonite is always observed to ejqjose the (010) face only whereas the aragonitic
biorriinerals also exposed the (001) face (MANN et a1., 1983). In biologically formed
aragonite impurity with several inorganic elements in trace quantities occurred while the
geological sampIe of aragonite was pure CaC03.
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The ratio of strontium to calcium concentrations in otolith aragonite has beeil found to be
correlatoo negatively with temperature, but in a different manner corripare to geologically
fonried aragonite. Thererore, assumirig a constant level of Sr/Cri in the environment. the
amount of stroillium iricorPorated irito otolith aragonite could be goveined by physiological
processes (RADTKE, 1984).
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The outstanding difference is that rilineral crystals have sharp edges and are different in size
while the single crystals froin biological sources have rounded edges and a friirly uniform
diameter of about 1000 A. The observed differences in the faces together With the assümption
of the basic building blocks of,these, 1000 A single crystals indicate that biological and
geological minerals differ iri the way in which theY. are formed. MANN al. (1983)
suggested that the matrix-mediated precipitation of otoliths is sufficiently different from the
mode of mineral formation to explain these obseived differences in inorphology.
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i Otolith growtti
There is Ütth~ knowledge aboui the eximceiiular proeess how calcium is depositcd. In the
arialysis ofthe inineralization process attention should bei paid fOf mechanisms of the
nucleation of the mineral, the transformation of unstable mineral to a more stable foriri, the
polymorph determination of the calcium carbonate deposition, and the control of crystal
growth, and crystal orientation. Although the. knowledge aboüt these mechanisms is
incomplete. the organic matrix is supposed to playa principal role. Thematrix hypothesis
assumes that the organic material is deposited aS a crystal nucleating suostrate (WATABE er al,
1982). The organie components supply locations for the initial fixation of ions, in .which
aspanie acid itself appears to be highly charged. This implies an active role in regulating the
mineral formation.
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· DEGENS et al. (1969) suppose that oxygen ions of the bicarbonate areöound in the polyhedci
strUcture of otolin. This will result in a more stable confirmation and crystal nucleation is
initiated by the arrangement of calcium to form metal ion coordination polYhedra's. Then .
carbOnate groups will oe attached to therruitrix by hydrogen linkages arid the oxygen from
the carbonate will exchange with oxygen from the metal ion coordination polyhedra's into a
stable structure. "
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Acidie glycoproteins from molluscs, echinooemis. tooth enamel and bone all adopt the ß~
sheet conforrriation in presence of calcium. The result of this orientatiori is that the .
carboxylate groups of the aspartic and glutrimie acid residues ure perPendicular to the plane of .
the sheet. This possible mode of calcium binding to the surface of the ß-shect could be an
explanation for the manner in which the proteins functiori. Funhermore. in vitro experiments
showed that the ß-sheet can interaet with cenain specifie crystal faces only (LOWENSTAM
AND 'VEINER, 1989). Sincc otolin is rich in aspanic and glutamiC acids~ in theory a similar
sort of mechanism may occur.
Ceased growth of aragonite could oe explained if nucleation sites on the matrix are blocked,
· for instance when calcite at those sites interferes with the aragonite gfowth. However X-ray
diagrams did not indicate any traces of c~l1cite (DEGENS er al.• 1969). But thereare more
possible exphinations for growth interruptions: the same authors refer to MORRIS AND
KITTLEMAN (1967) who suggested the presence of other onhorhombie minerals because
theyfound a significarit concentration of soditiiri in otoliths (calciurri substitution by sodium
in aragonitic minerals is cither unlikely).
To some extent mirieralization must be adynamie process because otoliths are sUIToimded by
a membrane in a fluid medium. Initiation, inhibition and contral of the proposed extracellular
mechanisins of otolith mineralizatiori are likely to be regulated by,orie or several of the
following possible principal factors: the organic matrix. hormonalor neurosecretory faetors.
arid the environmerit (temperaiüre, salinity. demerit concentrations). However. little is
known of the factors governing the crystal mOdificatioris.
.
Organic matrix
The formation of crystal riucleätiori is possible when concentraiioris of ioris exceed their
solubility product. CRENSHAW(l982) proposed the ionotropy theor)r as a likely mechanisrri .
of crystal micleation. The hydrophilie sites of thesoluble pari of the matrix (the acidic
groups) birid calciurri arid arotirid this sites a second carbonate enriched layer is forined.
Crystal riuclei (seeds) are formed whenever a local increase in pH or a decrease in caroon
dioxide occurs. Thc survival of the cfystal seed is deterinined by ion-tränsport processes or
the lcicäl concentratiori of the calcitim-binding matrix.
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· Orgänic material probably provides binding sites for ions or removes substances which
inhibit mineralizätiori ('VILBUR. 1980; MANN er al.• 1983). The macromolecules are secreted
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into the extracellular space where they assemble into a cOlttinuous sheetlike strUcture. Once
calcificatiori has begun, the .organic matrix is likely a mold for the mineralization (DEGENS el
ti/.~ 1969). LoWENSTAM ANDWEINER (1989) proposed that"thestructure of thenucleation
site is resporisible for the formation of aragonite or calcite, but the composition of the
surrou.nding ,endolymphe or the otolithic membrane could also be responsible. In this process
an interfering physiologicaI control cari not be excluded since the stable isotopic composition
and additives of mineralized parts are not in equilibrium with the environment.
One of the vanous explanations for reduce(l growth of cfystals is tanning of the organic iay~r
at its surface (WILBUR, 1976). A newly secreted layer of the organic material then. again
initiates a new layer of crystals and this alterriation coritinues. In this supposed mechanism,
crystal thickness depends upon the secretion frequency of inhibiting organic materiaL The
width of the crystallayer changes when the secretion of organic material does not prOceed at .
a constarit rate. The growth of thecrystallayer is iriterrupted by organic material and as a
result abipartite structure can be seen (WILBUR, 1980). The study of DUNKELBERGER el al.
(1980) showed that aragonitic crystals in otoliths ofthe mlllnmichog (Fundulus heteroclitus)
appeared to be relatively short and Iimited in length by the intercrysialline matrix.
Furthermore, a relatively thick interIameIlar matrix limits each individual growth laYer. Dut
agairi there is very little knowledge about the structures in orgariic matrices that are
responsible for crystal growth inhibition. In moIlusc sheIIs it is obvious that two distinct
organic structures are present in connection with the initiation rind gfowth of calcium
carbonate crystals. The 'sheetIike'. interlameIlar matrix is composed of glycine and alanine
and limits or ceases crystal growth. The'envelope' structure which intimately surrounds the
surface of growing crystals, is thotight to be involved in the acceleration of crystal growth
(BEVELANDER AND NAKAHARA; 1980).
A defidency of ions to the mineral site, for instance through trace quüntities of interacting
inolecules, is probably a more common and main reason why crystals stop growirig. Besides
lack of ions, the mechanical properties of the crystal can be changed by these iritemctioris. An
interaction during crystal formation retards growth on these faces causing them to grow more
slowly. Although there is no documeritation avmlable, LOWENSTAM AND WEINER (1989)
suspect that these type of effects are widely utilized by organisms for coritrolling
biomineralization.
Internal regulation
The otolith grows in the eridolymphatic sac, a fluid medium, arid the only connedive tissue is
the inacula acustica. Thus calcium carbOnate deposition sites on otoliths are isolated from the
environment und the calcium concentrntion in this pIasma is low in comprinson with seawater (SIMKISS, 1974). This is in agreement with the observation thatJhe regulation
mechanism of cation plasma levels in fish is quite effective (NORDLIE AND WHITTIER,
1983). Dy studying the composition of the eridolymph fluid during mineralizatiori, one might
be able to uriravel the chemical drivirig force for the calcium carbonate mineralization rate and
type.
MUGlYA (1974) found ihat injected hibeied calcium is tr~msportedthrough the blood
cripillaries andsecreted into the otolith fluid by some macular ceIls iri at least 3 hours. These
experiments have revealed that a large quantity of calcium deposition in rainbOw trout takes
place on the medial surfacenear the.macuhi cells except for the sulcus acusticus (Figure 1).
The author suggests tliat both organic arid inorganic constituerits of teIeost ötoliths are
secreted, at least iri part, by a ceIlular activity in the mricular region.
MuäiYA al. (1981) detefinined that there is a strong coi"relation bet~een the plasma
calcium level of fish bloo(l"and the calcium deposition rate on the otolith. They assurne that
el
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this is caused by the diei rhyihm in branchial uptake of calcium from theenvironment.'
Experiments with coho salmon, exposed to experimental stress conditioris, support the
hypothesis. Stress resulted iri microstructural check formation on otoliths and intenered with
a lowered calcium tiptake from th6 water (CAMPANA, 1983a).
Thecalcium physiology of otolith gr~wth appears to Oe much different from bones and
scales. Comparative aspects of thecalcium dynamics in differerit calcified structures have
beeil described by leHn AND MUGlYA (1983). Experiments with coho sahnon; in water with
radioactive labeled calcium, showed that the retention of radioactivity iri otoliths was
exceptionally large compared to bone arid scales. Even when the fish were put in nonnidioactive water after the exposure, the radioactivity of otoliths increased by 312% during
the first 14 days. This result supports the belief that once depositedcalcium is hardly
withdrawn from otoliths after initial mineralization and contradicts the suggestion of
PANNELLA (1980) that resorption produces checks refleeting growth discontinuities.
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The daily, deposition of calcium carbonate upori the organic matrix ceases neur dawn
(PANNELLA, 1980). An eridogenous circadian rhythm eritrained to the photoperiod may be
directly responsible for the reduced branchial uptake of calcium (MUGlYA er al., 1981;
TANAKA er al., 1981). Moreover, GAULDIE AND NELSON (1988) concluded that there are
strong physiological arguments for daily deposition of inicro-increments, in the anti-sulcul
section; tied to diel rhythms of the brain of the fishes .. The authors hypothesize that
neuroproteiris are transported down the palIial axon from the central ganglia arid secreted at
the nervous end-organ (the macula region).
As a resuit of the seaso~al change iri width of daily micfo-increments the optical pattern of
translucent and opaque zones in otoliths could be formed(RADTKE er al., 1985). This
deduction provides a physiological mechanism for the argument that opaque and translucent
zones are optical markers of seasonal changes in the growth rate of otoliths. GAULDIE (1990)
stated: ''Thus the iriforination in the anti-sulcul section that is used to estimate age of fish is
, on a firm scientific footing which would allow one to confidentlyassay the validity of the
patterns of opaque and hyaline (translucent) wnation in the anti-sulcul section of the otolith".
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4. Aga determination
. arie of the visible periodicities in otoliths i8 the alternation of opaque and transiucent zones,
commonly interpreted to detennirie the age of fish (\VILLIAMS AND BEDFORD, 1974). In
temperate ,!egions, intens,ive growth is reflected,by t.he ~orni~~ion of the opaque spring
summer zone. Probably due to a much reduced growth rate In wInter, the translucent zone is .
being formed (IMMERMAN, 1908). Although originally riarrow translucerit zones were
interpreted as ffalse' or 'check' rings (JENSEN, 1965), in ihis paper the term fcheck' is
refeiTirig to translucent zones regardless of theirwidth, corresporiding to the annually, or
otherWise, oCcurring decreases in somatic growth. The reasori for this charige is given by
GAULDIE (1990). Since JENSEN (1965) introducoo a standard terminology to describe otolith
marks, the interpretation of which features could be equivalerit to .mnual marks or not has
been changed. GAULDIE (1988) staied that on the crystal level there are no criteria io
differentiate between anriual check rings and additiorial check rings. If this, statement is
correct, it would follow that contrast-enhancing techriiqües which are specific for annual
checks are bOund to fail.
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Beside the descnption of false and annual rings, there is a more confusirig disagreemeni on
the terms hyalirie and opaque, especially in relation with the zone with richest in organic
matter. According to DANNEVIG (1956), MOLANDER (1947) and IRIE(1960), the organic
material is present only in the opaque zones, while the hyaline zones consist entirely of
aragonite. However, the burning techriique ofM0LLER CHRISTENSEN (1964) applied by
BLACKER(l974) showed that the optical dark parts (after burning) correspond to the parts of
. the hyaline zones, suggesting that this zone has the higher concentration of organic
compoimds. This phenomenon made the. term hyaline to become ambiguousbecause it is
being used to describe both an optical feature as weIl as a structural feature. Therefore, in this
paper a terminologieal distinction is niade in the otolith features deperiding on the aspect that
is being consideroo according to Table 1.
Table 1.
general
inte retation
check zone
rowth zone
Used descriprlons arid aIiocatCd components of otoliths.
optical feature
after buming
translucent
o a ue
dark
li ht
organic material
inorganic
material
short crystals
Ion c· stab;
The ring pattern is orten hard to deflrie ohjeCtively arid a variety of recipes is used 10 enhance
the contrast in ring features. It is generally believed that a large amount of information on the
individual life-history could become accessible if proper techniques wen~ employed..
However, in the literature. only theoretiCal mechariisms which could explain the bipartite
nature of otoliths have been proposed so far. At a microstructural level, the overall
conclusion can be drawn that daily ring features are a result of the rhythm iri neuropeptide
regulation and the mobilisation of calcium. These two processes are governed within the
organism but uridoubtedly are also irifluenced by erivironmerital factors (e.g. photoperiOd,
temPerature). Since calcium meiabolism of fish is poorly understood and protein syrithesis is
not easyto measure, it is hard to believe that any attempt to pOint out one principal factor thai
determines otolith growth is realistic.
The macrostrUctuml ring pattern is supposed to be a resuit of the change in width of daÜy
incremerits and the length of crystals would then be the major difference between tmnslucent
and opaque iones. On the other hand, the success of the burning techniques is explairi~ by
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the diiTerences in protdn conterit between the two zones. It is ~f courSe possible that both '
suggestioris are true. Beeause the protein"covers the crystal faces and/or smaUer crystals
reflect less light, the dark zone is optically trarislucent (fable 1)~ ,
Structural difTerences between the zonesare orten enharicro by treatments such as acid
etching, staining or bumirig of otoliths. The most common arid widespread methOd is still
M0LLER CHRISTENSEN's buming technique (1964). A main disadvantage of this methOd is
that the chemisuj beyond the technique is not fully understoOd. Wheriever, as stated before,
the protein content and thecrystallength in the two zones are bOth distinet, it is probable that
bumirig not only affects the organic layers but the crystal configurn.tion too. Aragonite is not
the most stable forin ofcalcium carbonate (crystallized at low temperature and low pressure)
arid heatirig will probably charige the mineral form to calche (MANN er aI., 1983). The
degradation rate of minerals is not only temperriture dependent but also influenced by the size
of the crystak Smaller crystals will degradate at a lower temperature. In practice the buming
technique has another disadvantrige. Great care has to be taken riot to crumble bumed otoliths
arid it is difficult to achieve standardization of the burning process.
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Receritly, auempts have beeri made todye fish otoliths in a simple way which is suitable for
mass proeessing. RICHTER AND DERMOIT (1990) succeeds in staining otoliths of several
economic species. The appearance of the ring features in treated otoliths resembled the
bumirig technique. However, strikirig was the result that collagen (related to otolin) specific
stains were not the most suitable ones. The äuthors found that, iri general, the darker the
stain; the beuer the contrast enhancerrient. This does not exclude the possibility that the
rougher zones are more easily siained, because after sectioning the zones consisting of
smaller crystills appear to have a relatively rough sUrface.
Considenngthese two contrast enh~uicing techniques, oniy one conclusiori can be dfaw~. It
is not certain ihat they act only as a chemical reagent and ure physically neutral. Only if it is
cleur what kind of rriechanism is involved, the interPretation of the contrast-enhanced
stiuctures can be deduced in a straightforward manner.
5~ Prospects
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The original ~im of this study was t6 achieve some irisights in the formation of transluceni
and opaque zones before starting to develop ari automatie ageirig method by meäns of iinage
analysis. In principle, a video camera fiued to ä stereomicroscope and combined with a
computer can be used in ageing (FAWELL, 1974, McGOWAN et al., 1987).1t is expected thiu
image analysis can reduce the amount of subjective interPretation in the ageing process. Tbe
setting of a threshold level for automatic identification of annual checks would serve as a
fixed kirid of error in jUdgement that could be rdatively easily detectable in due course
compared to subjective thresholds by expert r e a d e r s . ,
One of the possible advarice that cari be exjJected from an image analysis system is that,
besides number of growth zones arid width of zones, non-conventional data can be retrieved
such as shape factors and trarisformation coefficients (e.g. fourier, DE PONTUAL AND
PROUZET, 1988). Sophisticated image processing computer products are riow within reach
and can be applied for rriany biological topics (e.g. egg and larväe recognition and coimtirig,
zooplankton determination, sorting of fish, as weIl as pattern recognition and counting in
otoliths). Digitized inforination from the camera ori a video monitorcan be displayed at
resolution of, at least, 512 times 512 pixels each with the fuIl 8 bits of information. This
mearis that 256 intensities (grey levels) are available for image pröcessirig, which is far
beyond the detail that can be derived by direct visual reading. It is important to distinguish
fuH automatic or semi-automanc devices. In the latter ari operator moves a cursor ori the video
image and thus selects the coordinates of the cursor arid the corresporiding grey level values
of the original image that are stored to the computer for further processing. Tbe infonnation
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on groWih zones is thereby usercontrolledand in this case validation of automatie couriting is .
just as critical as it is in conventional methods. The potentiorial for automatie readirig can be .
developed bY. incorporating numerical techniques. One of thein, the graphie display of grey
levels along a specified counting path in a histogram, has aIre:idy been applied in different
ageing studies (MCGOWAN et al., 1987; GANDELIN AND LAVAL, 1987).
To. start with, video images should be of ihe highest possible quaÜty. Although image
enhancement by mathematic transformations eari lead to spectacular results, the desired
information needs tobe present in the, original image. Even though a new techriology may
give the impression of greater objectivity, there is always the riskthat arbitrary decisions,
such as contrast enhancing preparation of otoliths, have resultect in bias. Therefore pllysical .
changes in the ring structures should be excluded wheriever only cheinical reactions are
interpreted and vice versa. The inost suitable enhancemerit techriique for mass processing is
not (yet) available. In case of staining, some deeisive experiments with ehemieally neutral
dyes, for instance indian ink,need to be done. When working witll automatie ageing
systems, this will be more important than trying to understand the possible clues behind the
formation of yearly checks and a-periodic checks since the structural argument faiIs uritiI
now. The latter kind of checks should become detectable in a consequent manner by using
automatie ageing routines.
5~
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a
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Mar.
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vertiJ:ol transverne Yiev
•
+
y
med1 el S1 de
Iantenor I
3ulcU3 acU3ticU3
(m.a.cv.lar regio Il)
1ateral S1 de
anti-3ulcul3ectiDn
Figure 1. The sagittal view of the medial (concave) and lateral surface of an otolith. The
two axes in the upper picture represents the often used transverse and horizontal
sectioning through the nucleus.
\
- 11 -
•