Age Determination and Age Determination and Validation in Validation in

Chondrichthyan FishesChondrichthyan Fishes

Presented by Kara Baca Presented by Kara Baca

and Sharon Homer-and Sharon Homer-DrummondDrummond

MethodologyMethodology

Age determination process:Age determination process: Collection of hard part samplesCollection of hard part samples Preparation of the hard partsPreparation of the hard parts Examination (age reading)Examination (age reading) Assessment of validity and Assessment of validity and

reliabilityreliability Interpretation (modeling growth)Interpretation (modeling growth)

Hard parts usedHard parts used

Vertebral centra Vertebral centra Dorsal fin spinesDorsal fin spines Neural archesNeural arches Caudal thornsCaudal thorns

Vertebral CentraVertebral Centra Larger, more Larger, more

anterior centra anterior centra should be should be usedused

Whole or Whole or sectionedsectioned Vertebrae Vertebrae

should be should be sectioned sectioned sagittally sagittally (longitudinally)(longitudinally)

Dorsal Fin SpinesDorsal Fin Spines Used Used

particularly in particularly in dogfish sharksdogfish sharks

Spines from Spines from second dorsal second dorsal fin are fin are preferredpreferred

Used whole or Used whole or longitudinally longitudinally cutcut

Neural ArchesNeural Arches

May be useful for species that have May be useful for species that have poorly calcified vertebral centrapoorly calcified vertebral centra

Preliminary studies with sixgill Preliminary studies with sixgill sharkssharks

Caudal Thorns and Other Caudal Thorns and Other StructuresStructures

Caudal thorns used Caudal thorns used with vertebral with vertebral centra to determine centra to determine age in bathyrajid age in bathyrajid speciesspecies

Evidence for growth Evidence for growth bands found in bands found in upper jaw of the upper jaw of the wobbegong wobbegong Orectolobus Orectolobus japonicusjaponicusSerra-Pereria et al., 2005

Preparation of Samples Preparation of Samples (Centra)(Centra) Thaw (if frozen) or wash (if Thaw (if frozen) or wash (if

preserved in alcohol)preserved in alcohol) Clean excess tissue and Clean excess tissue and

separate into individual separate into individual centracentra

Sectioning typically done Sectioning typically done with low-speed diamond-with low-speed diamond-bladed sawbladed saw

After mounting sections to After mounting sections to slides, sand with wet fine-slides, sand with wet fine-grit sandpaper to grit sandpaper to approximately 0.3 to 0.5 approximately 0.3 to 0.5 mm and air-drymm and air-dry

Binocular dissecting Binocular dissecting microscope is generally microscope is generally used for analysisused for analysis

Age DeterminationAge Determination View wide bands View wide bands

(opaque) separated by (opaque) separated by distinct narrow bands distinct narrow bands (translucent)(translucent)

Tend to occur in Tend to occur in summer (wide) and summer (wide) and winter (narrow)winter (narrow)

Each pair of Each pair of wide/narrow bands wide/narrow bands considered to considered to represent an annual represent an annual growth cyclegrowth cycle

Validity must be testedValidity must be tested Age determination for Age determination for

spines are nearly spines are nearly identicalidentical

Dorsal fin spinesDorsal fin spines

http://journal.nafo.int/35/21-calis.html

Banding in Neural ArchesBanding in Neural Arches

Distinct bands in Distinct bands in sixgill sharkssixgill sharks

Potential use in Potential use in age age determinationdetermination

Staining & imageryStaining & imagery Imagery & enhancementImagery & enhancement

scanning EM (atypical)scanning EM (atypical) steromicroscopysteromicroscopy binocular light microscopybinocular light microscopy X-radiograophy (atypical)X-radiograophy (atypical) X-ray spectrometryX-ray spectrometry Graphite microtopographyGraphite microtopography

StainingStaining silver nitrate (McFarlane et silver nitrate (McFarlane et

al., 2002)al., 2002) cobalt nitrate (Hoenig and cobalt nitrate (Hoenig and

Brown, 1998)Brown, 1998) ammonium sulfide (Hoenig ammonium sulfide (Hoenig

and Brown, 1988)and Brown, 1988) haemotoxylinhaemotoxylin cedar wood oil, alizarin red, cedar wood oil, alizarin red,

crystal violet, salts (Cu, Fe, crystal violet, salts (Cu, Fe, & lead)& lead)

HistologyHistologyGoldman, 2004

Evaluating Precision Evaluating Precision Average percent error (APE)Average percent error (APE)

May serve as good relative indicator of precision May serve as good relative indicator of precision within and between readerswithin and between readers

However, tells which reader was less variable, not However, tells which reader was less variable, not which was better or if either was biasedwhich was better or if either was biased

Goldman’s approach to estimating precision:Goldman’s approach to estimating precision: Calculate percent reader agreement (=[No. Calculate percent reader agreement (=[No.

agreed/No. read] x 100) within and between agreed/No. read] x 100) within and between readers for all samplesreaders for all samples

Calculate percent agreement plus or minus one Calculate percent agreement plus or minus one yearyear

Calculate percent agreement with individuals Calculate percent agreement with individuals divided into appropriate length or disk width divided into appropriate length or disk width groupsgroups

Test for biasTest for bias Criticism: varies widely among species and ages Criticism: varies widely among species and ages

within a specieswithin a species

Tests for BiasTests for Bias Age bias plot- Age bias plot-

graphing one graphing one reader vs. the reader vs. the other and other and referencing referencing equivalence line of equivalence line of the 2 readers (45° the 2 readers (45° line through the line through the origin)origin)

Chi-square tests of Chi-square tests of symmetry, such as symmetry, such as Bowker’sBowker’s

Evans-Hoenig testEvans-Hoenig test

Back-CalculationBack-Calculation

Method for describing the growth Method for describing the growth history of each individual sampledhistory of each individual sampled

Numerous methods – formulas that Numerous methods – formulas that follow hard part or body proportion follow hard part or body proportion hypothesis are recommendedhypothesis are recommended Proportional relationship between animal Proportional relationship between animal

length or disk width and radius of length or disk width and radius of vertebral centrum or distance from focus vertebral centrum or distance from focus to each annulus within a given centrumto each annulus within a given centrum

Verification & Verification & ValidationValidation

Confirmation of age Confirmation of age determination by determination by

indeterminate and indeterminate and determinate methodsdeterminate methods

Why validation & Why validation & verification?verification?Caillet, 1990Caillet, 1990

Apex Predators website: Apex Predators website: “Sharks are long-lived “Sharks are long-lived animals that grow very slowly and do not produce animals that grow very slowly and do not produce many young. In many parts of the world, sharks many young. In many parts of the world, sharks are fished commercially, thus, in order to ensure are fished commercially, thus, in order to ensure proper management of the stocks, age and growth proper management of the stocks, age and growth data must be obtained. With this data, we can data must be obtained. With this data, we can determine the longevity of the species as well as determine the longevity of the species as well as maximum age, age at maturity, growth rate, and maximum age, age at maturity, growth rate, and differences in growth between males and females.”differences in growth between males and females.”

The process of evaluating growth zone The process of evaluating growth zone deposition deposition in fishes can be divided into in fishes can be divided into verification and validation.verification and validation.

Verification: Verification: “… conforming an age estimate by “… conforming an age estimate by comparison with other indeterminate methods”comparison with other indeterminate methods”

Validation: Validation: “… proving the accuracy of age “… proving the accuracy of age estimates by comparison with a determinate age”estimates by comparison with a determinate age”

Some methods Some methods of validationof validation

Chemical tagging of Chemical tagging of wild fishwild fish

Mark-recapture of Mark-recapture of known-age fish*known-age fish*

Bomb carbon Bomb carbon dating*dating*

Growth ring Growth ring frequencyfrequency distinction between distinction between

absolute age (*) & absolute age (*) & periodicityperiodicity

Centrum age Centrum age analysisanalysis

Relative marginal Relative marginal increment analysisincrement analysis

Some methods Some methods of verificationof verification

Size mode analysisSize mode analysis Primarily a Primarily a verificationverification tool tool Progression of discrete Progression of discrete

length modes over timelength modes over time Size modes taken from a Size modes taken from a

random sample of all size random sample of all size classes from a populationclasses from a population

Mean or median sizes in age Mean or median sizes in age classes assessed by other classes assessed by other means (e.g.: age classes means (e.g.: age classes predicted by von Bertalanffy predicted by von Bertalanffy growth function applied to growth function applied to physical assessment)physical assessment)

Random sample size modes Random sample size modes and mean or median sizes at and mean or median sizes at age comparedage compared

Coincidence supports Coincidence supports assumption of age classesassumption of age classes

Monitor discrete length Monitor discrete length modes across a given time modes across a given time period at discrete intervalsperiod at discrete intervals

Tag-recaptureTag-recapture Initial in field:Initial in field:

CaptureCapture MeasureMeasure WeighWeigh TagTag ReleaseRelease

Recapture and assess for Recapture and assess for growth in length and/or growth in length and/or weightweight

Compared to von Compared to von Bertalanffy growth Bertalanffy growth functions, TET, functions, TET, oxytetracycline (OTC) or oxytetracycline (OTC) or other methods, based on other methods, based on size changes between size changes between tagging and recapturetagging and recapture

file:///D:/age%20validation/Tetracycline%20or%20OTC_files/age1.jpg

Simpfendorder, 2000: dusky sharks

Chemical marking, tag-recapture Chemical marking, tag-recapture & lab& lab

Validation of absolute age focuses on validating “temporal Validation of absolute age focuses on validating “temporal periodicity of … growth increment formation”periodicity of … growth increment formation”

Tetracycline (TET): standard for marking free-swimming animalsTetracycline (TET): standard for marking free-swimming animals OTC: marks by binding to CaOTC: marks by binding to Ca2+2+ & depositing to active & depositing to active

calcification sitescalcification sites• 25mg/kg bw IM25mg/kg bw IM• + external tag+ external tag• highly visible marks in vetebral centra & dorsal fin spines under UVhighly visible marks in vetebral centra & dorsal fin spines under UV• use body growth & calcification changes to compare ‘time at liberty’ use body growth & calcification changes to compare ‘time at liberty’

[release] w/growth band deposition (# rings in vetra centra or spine [release] w/growth band deposition (# rings in vetra centra or spine post-injection = time hacks)post-injection = time hacks)

• lab or captivitylab or captivity Tag-recapture + markingTag-recapture + marking

• tag + OTC followed by recapturetag + OTC followed by recapture• tag + TET followed by recapturetag + TET followed by recapture

Other tagsOther tags• Fluorescein (not well-evaluated in elasmo.; 5-10mg/kg bw, Fluorescein (not well-evaluated in elasmo.; 5-10mg/kg bw,

higher=mortality)higher=mortality)• Calcein (not well-evaluated in elasmo. .; 5-10mg/kg bw, Calcein (not well-evaluated in elasmo. .; 5-10mg/kg bw,

higher=mortality)higher=mortality) Define age Define age && indiv. growth characteristics indiv. growth characteristics Combining information from mark w/observed growth Combining information from mark w/observed growth

changeschanges

Tetracycline groupTetracycline group

Broad spectrum Ab’s that block bacterial Broad spectrum Ab’s that block bacterial protein synthesis by inhibition of aminoacyl-protein synthesis by inhibition of aminoacyl-t-RNA binding to the rRNA A site.t-RNA binding to the rRNA A site.

TET = tetracyclineTET = tetracycline higher water solubility at pH = 7, than other Ab’s higher water solubility at pH = 7, than other Ab’s

in same groupin same group OTC = oxytetracyclineOTC = oxytetracycline Inject TET or OTC at capture while taggingInject TET or OTC at capture while tagging

25 mg/kg25 mg/kg absorbed & deposited at growth sitesabsorbed & deposited at growth sites only mark glows under UVonly mark glows under UV pairs > mark = growth over timepairs > mark = growth over time

Centrum edge analysisCentrum edge analysis Centrum edge analysis:Centrum edge analysis:

Compares opacity (width) &/or translucency Compares opacity (width) &/or translucency (density) of centrum edge over time(density) of centrum edge over time

• measure or grade edgemeasure or grade edge• compare to time of year or seasoncompare to time of year or season• detail: analyze Cadetail: analyze Ca2+2+ & PO & PO4 4

–– at edge by x-ray or at edge by x-ray or electron microprobe spectrometryelectron microprobe spectrometry

• not used much yet:not used much yet: recaptured nurse sharks marked with TET (Carrier recaptured nurse sharks marked with TET (Carrier

and Radtke, 1988)and Radtke, 1988) 4 ray species in the Irish Sea marked with TET 4 ray species in the Irish Sea marked with TET

(Gallagher et al., 2004)(Gallagher et al., 2004) tiger sharks (Winter and Dudley, 2000)tiger sharks (Winter and Dudley, 2000)

http://journal.nafo.int/35/10-gallagher.html

Winter and Dudley, 2000

Relative marginal increment Relative marginal increment analysisanalysis

RMI = MIR (ratio): (VR-RRMI = MIR (ratio): (VR-Rnn)/(R)/(Rnn-R-Rn-1n-1)) ParametersParameters

• MIR – marginal increment ratioMIR – marginal increment ratio• VR = vetebral radiusVR = vetebral radius• RRnn = distance from center to outer edge of last complete = distance from center to outer edge of last complete

bandband• RRn-1n-1 = distance from center to outer edge of next-to-last = distance from center to outer edge of next-to-last

complete bandcomplete band Direct: plot against month to find band formation trend Direct: plot against month to find band formation trend

lineline Assess seasonal band &/or ring depositionAssess seasonal band &/or ring deposition

• measure margin (growth) area of centrum from last growth measure margin (growth) area of centrum from last growth ring to centrum edgering to centrum edge

• divide by width of last fully formed annulus (none for age 0)divide by width of last fully formed annulus (none for age 0)• plot values against month of captureplot values against month of capture• determine periodicity of band formationdetermine periodicity of band formation

Poss. combine w/stable isotope analysisPoss. combine w/stable isotope analysis• fish workfish work• no elasmo. work yetno elasmo. work yet• proved viable in other chondrichthyansproved viable in other chondrichthyans

Carlson and Baremore, 2004

Neer et al., 2005: Bull shark growth

Captive rearingCaptive rearing Laboratory or Laboratory or

aquaria growthaquaria growth Particularly useful if Particularly useful if

poor ring formation, poor ring formation, calcification, etc.calcification, etc.

Morphometric data Morphometric data reveals age and reveals age and growthgrowth

Periodicity of growth Periodicity of growth zone formation in zone formation in vetebral centravetebral centra

Difficult to determine Difficult to determine true baseline & true baseline & captivity effectscaptivity effects

www.pac.dfo-mpo.gc.ca

Bomb carbon (radiocarbon) Bomb carbon (radiocarbon) agingaging

Tied to global oceanic rise in Tied to global oceanic rise in 1414C following C following cold war atomic testingcold war atomic testing

Synchronous uptake in marine Synchronous uptake in marine carbonatescarbonates

Dated marker of calcified structures Dated marker of calcified structures allowing growth band datingallowing growth band dating Born < 1956: lowBorn < 1956: low Born 1956-1965: higherBorn 1956-1965: higher Born > 1965: decliningBorn > 1965: declining

Regional identificationRegional identification Validation of annulus formation & Validation of annulus formation &

absolute ageabsolute age

Campana et al., 2002

Campana et al., 2002

Current status of studiesCurrent status of studies Caillet, 1995 was last major reviewCaillet, 1995 was last major review

115 publications listed in chapter since then (several not included in 115 publications listed in chapter since then (several not included in chapter that were published late 2004 – mid-2005)chapter that were published late 2004 – mid-2005)

68 newly studied spp. among those68 newly studied spp. among those Methods: most studies combine (mult. validation & mult. Methods: most studies combine (mult. validation & mult.

verification)verification) most use structure calcificationmost use structure calcification

• 1°1°lyly vetebral centra (70% of studies reviewed in chap.) vetebral centra (70% of studies reviewed in chap.)• dorsal spines (7%)dorsal spines (7%)• neural arches (1 study) & jaws (1 study)neural arches (1 study) & jaws (1 study)

captive growth (9%)captive growth (9%) tag-recapture (7%)tag-recapture (7%) embryonic growth (4%)embryonic growth (4%) precision analyses: several growth parameter valuesprecision analyses: several growth parameter values

1° reviewed methods = MIR or ratio (50%) + centrum edge1° reviewed methods = MIR or ratio (50%) + centrum edge modal analysis (25 studies, 22 spp.)modal analysis (25 studies, 22 spp.) tag-recapture (20 studies, 19 spp.; many combined w/OTC marking)tag-recapture (20 studies, 19 spp.; many combined w/OTC marking) lab growth (21 studies; 18 of which used OTC marking)lab growth (21 studies; 18 of which used OTC marking) bomb carbon study (Natanson et al., 2002, validated pobeagle growth bomb carbon study (Natanson et al., 2002, validated pobeagle growth

& indicated shortfin mako growth of 1 ring/yr.)& indicated shortfin mako growth of 1 ring/yr.)• combined w/tag-recapture & OTCcombined w/tag-recapture & OTC

Combining validation & verification seems to be the most robust Combining validation & verification seems to be the most robust meansmeans

Growth modelsGrowth models von Bertalanffy (VBGF): fish growthvon Bertalanffy (VBGF): fish growth

size at a given point in time depends on anabolism & catabolismsize at a given point in time depends on anabolism & catabolism identification of the growth coefficients are dependent on the given identification of the growth coefficients are dependent on the given

studystudy easy population comparisonseasy population comparisons fitting allows for fine-tuningfitting allows for fine-tuning problems:problems:

• small sample size: poor parameter estimationsmall sample size: poor parameter estimation• tt00 (theoretical size) should be replaced by L (theoretical size) should be replaced by L00 (length at birth) as 3 (length at birth) as 3rdrd

parameter (more to come)parameter (more to come) Gompertz: larval & early growthGompertz: larval & early growth

S-shaped model function (like Ricker logistic function)S-shaped model function (like Ricker logistic function) also can fit to fine-tune for a given studyalso can fit to fine-tune for a given study growth rate = log of asymptotic disk width or length - actual growth rate = log of asymptotic disk width or length - actual

asymptotic disk width or lengthasymptotic disk width or length most commonly used for skates & raysmost commonly used for skates & rays possibly most appropriate for ovoparous spp.possibly most appropriate for ovoparous spp. possibly most appropriate for spp. that grow in V more than weight or possibly most appropriate for spp. that grow in V more than weight or

LL possibly better estimate captive growth (slowed or atypical growth possibly better estimate captive growth (slowed or atypical growth

due to stress)due to stress) Best fit functions: choice of model based on length-at-age or Best fit functions: choice of model based on length-at-age or

weight-at-ageweight-at-age nonlinear least-squares regression analysisnonlinear least-squares regression analysis maximum likelihood functionmaximum likelihood function

VBGFVBGF Traditional: LTraditional: Ltt = L = L∞∞(1-e(1-e-k(t-t-k(t-t

00))))

substitutesubstitute LLo o and L for t and tand L for t and t00

ModificationsModifications Neer et al., 2005 (following Fabens, 1965) to Neer et al., 2005 (following Fabens, 1965) to

estimate bull shark growth (modified to fit observed estimate bull shark growth (modified to fit observed size-at-age data)size-at-age data)• LLtt = L = L∞∞(1-be(1-be-k(t-t-k(t-t

00))) = predicted length at time t) = predicted length at time t

• b = b =

• LL∞∞ = theoretical asymptotic length = theoretical asymptotic length

• k = growth coefficientk = growth coefficient

• LL00 = length at birth = length at birth

compared to fitted models to sex-specific size-at-age compared to fitted models to sex-specific size-at-age data w/traditional VBGF modeldata w/traditional VBGF model• theoretical longevity estimated at 95% Ltheoretical longevity estimated at 95% L∞∞ (5x ln2 k (5x ln2 k-1-1))

• growth model parameters derived from least-squares growth model parameters derived from least-squares regressionregression

Neer et al., 2005

VBGF growth coefficients VBGF growth coefficients (k)(k)

k = average rate to reach maximal length k = average rate to reach maximal length or size from length at birthor size from length at birth

Studies fitted to VBGF allow estimates of LStudies fitted to VBGF allow estimates of L∞∞ (asymptotic size), k, & L(asymptotic size), k, & Loo or t or too high k = fast growth rate (0 – 1)high k = fast growth rate (0 – 1)

Large variance among spp. & studiesLarge variance among spp. & studies k in chimaera: 0.05 – 0.47k in chimaera: 0.05 – 0.47 k in sawfish: 0.07 – 0.08k in sawfish: 0.07 – 0.08 k in guitarfish, torpedo rays & stingrays: 0.2 – k in guitarfish, torpedo rays & stingrays: 0.2 –

0.50.5 k in skates: 0.05 – 0.5k in skates: 0.05 – 0.5 k in whale shark: 0.03 – 0.05k in whale shark: 0.03 – 0.05 affected by: sample size, aging methods, affected by: sample size, aging methods,

verification, validation, & growth and model verification, validation, & growth and model fittingfitting

Estimating maturity & Estimating maturity & lifespanlifespan

TTmatmat estimated by various means & shows great variance estimated by various means & shows great variance among spp. & studies (sex, spp., method, study)among spp. & studies (sex, spp., method, study)

chimaeras: 2.9 – 6.0 yearschimaeras: 2.9 – 6.0 years• CC.. monstrosa monstrosa: 11.2 – 13.4 years: 11.2 – 13.4 years

sawfish: 10 yearssawfish: 10 years guitarfish, torpedo rays, stingrays: 1 – 9 yearsguitarfish, torpedo rays, stingrays: 1 – 9 years skates: 4 – 13 yearsskates: 4 – 13 years angel shark: 11 yearsangel shark: 11 years sixgill shark: 5 – 21 yearssixgill shark: 5 – 21 years carpet sharks: 16 – 25 yearscarpet sharks: 16 – 25 years dog sharks: 4 – 45 yearsdog sharks: 4 – 45 years

Related to longevity estimates (5x ln2 kRelated to longevity estimates (5x ln2 k-1-1) ( well duh!)) ( well duh!) chimaeras: 5 - 10 yearschimaeras: 5 - 10 years

• CC.. monstrosa monstrosa: 29 years: 29 years sawfish: 30 - 44 yearssawfish: 30 - 44 years guitarfish, torpedo rays, stingrays: 1 – 9 yearsguitarfish, torpedo rays, stingrays: 1 – 9 years skates: 9 - 50 yearsskates: 9 - 50 years angel shark: 35 yearsangel shark: 35 years carpet sharks: 9-35 yearscarpet sharks: 9-35 years dog sharks: 12 - 70 yearsdog sharks: 12 - 70 years

Back to whyBack to why

Better knowledge = better Better knowledge = better estimation of growth potentialestimation of growth potential

Better estimation of growth Better estimation of growth potential = better management potential = better management strategiesstrategies

Real consequences mean that Real consequences mean that precise, careful, redundant work precise, careful, redundant work is neededis needed

Key references and citationsKey references and citations

Goldman, K.J. 2004. Chapter 6: Age and growth of elasmobranch fishes. unidentified book.Goldman, K.J. 2004. Chapter 6: Age and growth of elasmobranch fishes. unidentified book. Serra-Pereira, B., Figueiredo, I., Bordalo-Machado, P., Farias, I., Moura, T. and Gordo, L.S. 2005. Age Serra-Pereira, B., Figueiredo, I., Bordalo-Machado, P., Farias, I., Moura, T. and Gordo, L.S. 2005. Age

and growth of and growth of Raja clavataRaja clavata Linnaeus, 1758 – evaluation of ageing precision using different types of Linnaeus, 1758 – evaluation of ageing precision using different types of caudal denticles. Elasmobranch Fish. Sci. ICES CM. N-17:1-10.caudal denticles. Elasmobranch Fish. Sci. ICES CM. N-17:1-10.

Gallagher, M.J., Nolan, C.P. and Jeal, F. 2004. Age, growth and maturity of the commercially important Gallagher, M.J., Nolan, C.P. and Jeal, F. 2004. Age, growth and maturity of the commercially important ray species from the Irish Sea. e-journal NW Atl. Fish. Sci. V35(art.10):ray species from the Irish Sea. e-journal NW Atl. Fish. Sci. V35(art.10):http://journal.nafo.int/35/10-gallagher.html.

Carlson, J.K. and Baremore, I.E. 2005. Growth dynamics of the spinner shark (Carlson, J.K. and Baremore, I.E. 2005. Growth dynamics of the spinner shark (Carcharhinus Carcharhinus brevipinnabrevipinna) off the United States southeast and Gulf of Mexico coasts: a comparison of methods. ) off the United States southeast and Gulf of Mexico coasts: a comparison of methods. Fish. Bull. 103:280-291.Fish. Bull. 103:280-291.

Natason, L.J., Casey, J.G. and Kohler, N.E. 1999. Growth of the tiger shark, Natason, L.J., Casey, J.G. and Kohler, N.E. 1999. Growth of the tiger shark, Galeocerdo cuvierGaleocerdo cuvier, in the , in the western North Atlantic based on tag returns and length frequencies; and a note on the effects of western North Atlantic based on tag returns and length frequencies; and a note on the effects of tagging. Fish. Bull. 97:944-953.tagging. Fish. Bull. 97:944-953.

McFarlane, G.A., King, J.R. and Sauders, M.W. 2002. Preliminary study on the use of neural arches in McFarlane, G.A., King, J.R. and Sauders, M.W. 2002. Preliminary study on the use of neural arches in the age determination of bluntnose sixgill sharks (the age determination of bluntnose sixgill sharks (Hexanchus griseusHexanchus griseus) Fish. Bull. 100:861-864.) Fish. Bull. 100:861-864.

Gburski, C.M. 2005. Ageing procedures for the big skate (Gburski, C.M. 2005. Ageing procedures for the big skate (Raja binoculataRaja binoculata), longnose skate (), longnose skate (Raja Raja rhinarhina), Alaska skate (), Alaska skate (Bathyraja parmiferaBathyraja parmifera), Aleutian skate (), Aleutian skate (Bathyraja aleuticaBathyraja aleutica) and Berring skate ) and Berring skate ((Bathyraja interruptaBathyraja interrupta) at the Alaska Fisheries Science Center. In submission.) at the Alaska Fisheries Science Center. In submission.

Calis, E., Jackson, E.H., Nolan, C.P. and Jeal, F. 2005. Preliminary age and growth estimates of the Calis, E., Jackson, E.H., Nolan, C.P. and Jeal, F. 2005. Preliminary age and growth estimates of the rabbitfish, rabbitfish, Chimaera monstrosaChimaera monstrosa, with implications for future resource management. e-journal NW Atl. , with implications for future resource management. e-journal NW Atl. Fish. Sci. V35(art.21):Fish. Sci. V35(art.21):http://journal.nafo.int/35/21-calis.html

Simpfendorfer, C.A. 2000. Growth rates of juvenile dusky sharks, Simpfendorfer, C.A. 2000. Growth rates of juvenile dusky sharks, Carcharhinus obscurusCarcharhinus obscurus (Lesueur, (Lesueur, 1818), from southwestern Australia estimated from tag-recapture data. Fish. Bull. 98:811-822.1818), from southwestern Australia estimated from tag-recapture data. Fish. Bull. 98:811-822.

Wintner, S.P. and Dudley, S.F.J. 2000. Age and growth estimates for the tiger shark, Wintner, S.P. and Dudley, S.F.J. 2000. Age and growth estimates for the tiger shark, Galeocerdo Galeocerdo cuviercuvier, from the east coast of South Africa. Mar. Freshwater Res. 51:43-53., from the east coast of South Africa. Mar. Freshwater Res. 51:43-53.

Neer, J.A., Thompson, B.A. and Carlson, J.K. 2005. Age and growth of the Neer, J.A., Thompson, B.A. and Carlson, J.K. 2005. Age and growth of the Carcharhinus leucasCarcharhinus leucas in the in the northern Gulf of Mexico: incorporating variability in size at birth. J. Fish. Biol. 67:370-383., northern Gulf of Mexico: incorporating variability in size at birth. J. Fish. Biol. 67:370-383.,

Campana, S.E., Natanson, L.J. and Myklevoll, S. 2002. Bomb dating and age determination of large Campana, S.E., Natanson, L.J. and Myklevoll, S. 2002. Bomb dating and age determination of large pelagic sharks. Can. J. Fish. Aquat. Sci. 59:450-455.pelagic sharks. Can. J. Fish. Aquat. Sci. 59:450-455.

Conrath, C.L., Gelsleichter, J. and Musick, J.A. 2002. Age and growth of the smooth dogfish (Conrath, C.L., Gelsleichter, J. and Musick, J.A. 2002. Age and growth of the smooth dogfish (Mustelus Mustelus caniscanis) in the northwest Atlantic Ocean. Fish. Bull. 100:674-682.) in the northwest Atlantic Ocean. Fish. Bull. 100:674-682.

www.pac.dfo.-mpo.gc.cawww.pac.dfo.-mpo.gc.ca. Age determination of elasmobranchs.. Age determination of elasmobranchs. www.psrc.mlml.calstate.eduwww.psrc.mlml.calstate.edu. Age growth and demographic studies.. Age growth and demographic studies. www.flmnh.ufl.eduwww.flmnh.ufl.edu. Pacific shark research center. Moss Landing Marine Laboratories.. Pacific shark research center. Moss Landing Marine Laboratories. www.na.nefsc.noaa.govwww.na.nefsc.noaa.gov. Apex predators: age and growth in sharks.. Apex predators: age and growth in sharks.