This paper not to be cited without prior reference to the author

•

International Council for theExploration of the Sea

C.M.1970/E: 15Fisheries Improvement Committee

REF.: K( Shellfish and Benthos Committee)

••

SOME OBSERVATIONS ON THE RESPONSE OF THE BENTHIC BIVALVE

TELLINA TENUIS TO POLLUTANTS

by

E.A. Stirling

:Marine Laboratory,Aberdeen, Scotland

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'SOME OBSr.:RVATI'ONS 'ON' THE RESPoNSE 'OF THE B:E1~HIG' BIVALvE~: .,. .'. .. • . . .. .. ••.. '. l ,.~" ".

TELLINA TENUIS TO POLLUTANTS,. : .:, ;;;~, ~f' ,,'.

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smnJARY' , ', ,A

E.A. StirlingMarine Laboratory,Aberdeen, Scotland

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. ) ,. , ..'.. .Experiments describcd hore sh~w how shcrt:"tenn toxieÜY,'töcits on phenol

and copper using subacute pollutant' concimtrations may'be used to':1.nvestigate:the action of pollutants on the bchaviour of the marine sand-dwelling bivalve,Tellina tenuis. Results of such tests are a useful preliminary to lon6-:termexperiments designed to investigate the effects of low levels of pollutants (in6rowth and metabolism.

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A preliminar,y step'in manY pollution investigations is·the,dcfinition,of,toxie concentrations of a pollutant to the species at risk. ,The st?-_ndardLDSOtest is used to indicate the ccncentration of' pollutant which will cause sO%'mortality under given conditions in astated time interval. In such testsanimOJ.s are orten subjocted to highly atypical conditions, and little-::.:' ~:, "significance can bo placed on behavioural responses observed during the test.In so.nd":'dwelling bivalves, ,for example~ it is ,us.uo.l'in LDsO:.tests: to ei:pose' ,aniniai'sto arange '~f pollutant concentrations in water"in,tho',ab6erice of'sand.Havi~g det,ermined ~h,e 'LD,O concentration," supp~ementa.ry,~härt·7~crmtests~.. , 'perf'ormed under more natural conditions,are of' usa in inves~igating,behavioural

respons es and' in def'ining modifications of', the action' ,of the pollutant which'result from properties of the', environment: Such acutebehavioural responses,;wl1en'def'ined,' may be Usedto' mcasure the 'short-term ef'fects of'.a range 'of "pollutant' concentrations' b elow the LDSO' level.. ' " :';; ~ ~ " , ,

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nrrRODUCTION

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" " :'Th1s'repO'rt 'descnbes' observations" of', short-term behavioural responses, of the 'manne sand-dwelling bivalve; Tellina tenuis, when ,eXposed"to s'ubacute'concentrations of pollutant' s olutions. ' The burrowL"lg response of animals when

, in contact with a sand surf'ace has, been used ,tomeasure the: ef'fect'"cifpollutants, under'different', ccnd?-tions~: Healthy-T. tenui1!, when. exposed :~n the surf'ace ':

:, of, sand, burrow into' the ' substrate; more' or 1038 immediately. , Any:interf'erence, :,W'ith".the'burro~ing behavi,ourcould cause the animals :to,;be ~ngerouSly.exposecl

... 'on the.'sand surface to predators or. to wave action~ . The, effect cf pollutantson ,this responsehas been' determined by recording the number.' cf aru.m~ls whichhad', disappeared from view in aseries of time intervals arter, exposure, of'animals at, thc sand"sürlace.' This rate of disappearance wilLbe referred to'"subsequontlyas the burrowing rate.' 'In 'all' expe:l-iments" 'airlmals were'exposedto 'the' pollutant f'olution for' a poriod of' 96 hours •. At the end' of tliis periodthe pollutant solution was replaced by clean sea water ,a~d,f'urtheI' data,for 'bgrrowing ~te ,~lere recprded•. The experimental temperature in a11 cases' was

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The effect of a pollutant solution on the burrowing rate of Tellina isshown by comparing the observed burrowing rate for animals in. clean sea ;vaterwith the rate in the pollutant solution~ Comparison' of data d.erived underdifferent experimental conditions' indicates ·howthis rate may be affected byphysical variables in the environment. " .

PHENOL• ." f ~.

96 Hour LDSO

Initial 96 hour LD50 tests perf'ormed by eXposing Tellina to a range. <?f ,ccncentrations ofphenol in the absence of sand indicated that LD50 concentrationat 9°C', is b'etween',700 and,800 ppm' of phen'~l.' ..', '",'.: , .-

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Subacute tests:

Methods. ":' .""'....

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, -"..• f •...• •Three types of experimental conditions were tested: 0. static system vdth

no water flow, a system with water flowing through 0. sand column, and 0. systemwith water floring over the sand surface, but not through the COlUIm itself. 'Conditions 'af water-IIiixing ,in .the,fieldprobably,lie somcwhere within thesethree'distinct situations;' :',:. ", :.. "":

'. -' '. . ., ' ,

•Static tests:

, .., . ,", ,.

(ÄY: ,Staticte~ts ,~e~ö ,cai-ried out using 2-litre pyrex'beakers ,contatlung 86mdepthof sand and 500~ml:of pollutimtsolution. 'Fifteen:ani.!nals of mixed sizes

'w'ere used in cach'test and' the burrowingrate, of all animals in clean s'ea' water, , \. ., , - " ' " ,

was recorded before adding the pollutantsolutions. ,At the start of thetest,. the.' sea water was run ,off' the, sand surf'ace and repla'ced by ,the pollutant solutionbut the latter was not \ mixed with· the sand'.' ,Phenol concentrations tested. were0, 5,10, 25,' 50,: 100 'and 500 ppm. The phenol solution vrasrenewed and theburrowing rate was determined every 24 houre. ' After 96 hours phenol was replacedby sea water, and further readingsofb~rrowingrate,were determined over'the' •subsequent 4-day periode ' ., " " "

(B) Continuous flow over sand surface

. 'EXperim~nts"to test' the effect 01"0., continu,?us flow of phenol 0v:er the sand •surf'ace on the burrowing rate in Tellina were also carried .out using 2-litrepyrex beakers'with' quantities :of sand and, pollutant, s'o~ution·.identicalto'thoseused in static'tests. Ten animals of mixed sizes,were used in each test;andtheburrowing rate" of all test animals .was determiIled in'·clco.n,sea water before

.' adding phenol. Animals 'which failedt"o burrow ,were removed ,after 75' minütesand were replaccd~ 'In these tests animals were, allowed', t (, become. establishod inthe sand befor'e: rerplacing the sea water_ by 500, ml, of phenoL The concenträtionstested were 0, 100,"300 arid 503 ppm.: Exchange:was ef'fccted by a continuousflcm of phenol into'and'out of tbe'test vessel at '0. ratcof,1.2'litres~per24 houremaintained by'a po'ris taltic pump., The phenol reservoir, supplying thc' inflow wasreplo.cod ~ver7i 24 haurs until 96 hours, had elapsed. ~t this. time thc head ofphenol in the'beakers was drainedoff' and replaced by clean sea'water, and cleansea water was supplied at the inflow. " ..,'.... . i." .".~.~.. _ . t .*" •.

In this experiment readings of bu~r~win'g rate weredeterirl~edoIily afto~'the phenol had been replaced by sea water. At no time ware animals in direct

2

,.

In static tests a ccncentration as low as 10 ppm of phenol gave .. a"noticeable reductioll in burrowing rate comparcd with control aniöals '(Graphs

.,1 and 2).' At concentrations of500j ,100 and 50.ppm ofphenol"burrowingstopped more' or 'less ;'completely. while ,anim?-ls. were 'exPösed~,t6,the' polltitant,t"-olution"~' . '.'. :, I' ' ~ , • .. , •• " .- •. '!:' 'lt.J ••••• ( ..••• '. ' •• t~. • ~ . • ~.1 _ •• '. • ' .. ~.. •••• '. ..~ -... • ..._.,."- .

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, Observations. fromboth static and, continuous ,flow tests indicate' that -:'Tollina.has quite, a 'high: capacitYlf0r:- rocoveY:j after. exPosuroto ';pherlOl: .:.:~pollution; .In the static tests ~ .in· C?9nc,e~t~tions of50 'ppDi 'phenol 'there~:;.:was considerable. recovery within:two,. days." and, even animals.'takon f'rOD' l.:~''''100 :ppm 'regained some, capacity ,to..burrow,after'foür" days, iri-clean s'eci.: water"(Graph :3) •. ·:IIn the oase of,ani~als take~ from 500 ppmof phenol, .-:sooe' did' ',~'oventually, die~but even at.this 9~ncenträtion 8omeanimo.ls.di:d rego.in 'tho',

. ability to dig.into,sand aftera.few,days in clean sea'water.' ',Rosul·ts fram'·continuous flow,~tests:eIs 0 show a' s'e"ciuence" of rocömmcnc'ement' of:burrowinB, on

. subs equent days .after, rcplacing .phenol by' .soa;,inter... Thc' ia.to- of,. recoyc~ '.~was in inverse relation tri tbe phenol concentration' experienced (s~~.' Grap'hs,5-7). ..,. ' ....' .,'.,~,~ , .. -'

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Subacute tests

Rosults

contact'with'the' free'phenol:solutiön;: they were exposed to pollutant' solutionin ·tho· inte~ti:tial water only." (~thoueh·the e:x:n.ct conccntration of phenol. "in. the interstitial waterwas not·measured;· the' co..'1.centrätion',compared with' ...that· in;'free": sea water i8 likely to bo' reduced"duo to adsorrtion arid:: ".: '."degradatioli'of ph'enol:,during passage throu'gh the,sand~) "After rcplacing ',: ':'phenol by sea water a 3-hour period wasnllowed .to· elapse ,to allnw·fl.ushing ",of the sand beforo.iletcrmining burrowing 'rote., ' Readings wcre taken 3 ~ 27and 75 h'ours 'o.ft·er·replacing:phenol'.::·': -.- ·~ .. ::;,.i:: ': ~ ,~:.:IY·,= L' ;l'i',~..)-:,.:·'l

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,'.~- . (eY: ContinF6us' flow'through·sand'''colUr.1ns'''· ,:;~~:.:':' .. :"<.: .'.' , :' "'c,:·.'L'.,; ',-:.:~ ... , •• ,,:.'1 .... ';•• ' ..... _.i ••; ...... ' •• '; 0*" o.. ;'~ -.' ",:•• "!;~ _" ':.~ i: .~ .. ':: ".; ': '\ ~':'.. ..! .... ;.; :

", ':" ':'·persp'ex sand' colUmns:·of'i6 tt.diametor wereused to t'est,the'effoct 'on ",::'(Telli'na of a- flow"of phenol"through:the"'sand 'column~r,As~f6r tosts described

\', . above, a>depth>of 8/cmof sandwasused,·'and this, wa's·· 'covcred·at ,the start of',Ithe' test by 500' ~l: 'of phcnol;'solution. ~:Asphenol'.was·~pur:cpcd'on.to· the··top :1'­of 'the '-column at o,'rato' (cif'1'.61itres· per 24 'hours)' which ,was' in .excess' ·of,

" . ,the ·outfl.'owrote:('6f 1 ~2 'lifres per,24 hours)·this hoad of polhitant increasedthrough the test periode (This inequality of inflow and outflow pumping rateswas ~ precaution to ensure that the head of water did not drop and leave th.esand column exposed.) As in the experiments testing continuous fl0".7 of':';";:,':"pollutant ovor tho sand surfaco, animals wero tested for burrowing rate andworo :allowed t obocome esto.blished in ;the' s'and' before adding the' pollutant

'.. 'solüti'ori.:--: Tho inflow'reS'erVoir of'pheriolwas:;replaced daily and the ..... ·..~':.,.. .!concentrations tested were 0 ~ 100, 300 and 500 ppm. Burrowing .rate ;was In :-- .:.recorded only after tbo phenol had been replaced by sea water and a lagporiod of two houm was allowed after this replacement and beforo. tald..ng thofirst reading to allow time for the sea water to flush through -the sand

..: col~.·. Readings were·,taken' 2;, 26 and:74 hoursafter" replacing the ,phenol by'scawater. ~ '. " ~.' .. , .' .', .' .. ' ." ,:::': r .. :,

••

••

Comparison of tho rate of recovery at each concentration undöi:thethree different experimental regimes shows :that most rapid r:-ecovery occurredwhon phenol had been floVling over tho sand surface onlY,:·and was, slowest inthe static experiments (Graphs. 1-7) •. ·This corrolates with the fact that in

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:' the'static expe:cl.ments, exposure of ani~als'to phenol:was more, complete.: .~,In' addition, after rcplacing phenol by clean sea water,: under.. static: conditionnthere was no water flow to nush residual phenol from tho sand•. In the' r,

static test~ unlike .the ;continuous flow, tests," sandrrcm .tlie vessel ~hich hadcontained the 500 ppm phenol solution,' had a distillCt.: phenolic smeil severaldays after ·tho phenol had been replaced by sea water.,;. .'; ," -;' .".: .": .•-::. ,

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Exposure of Tellina to higher concentrations of, phenol tested (Le. 100and 500 ppm) did not result simply in adduction of'the shell valves to 'excludethe pollutant solution. This appearedto be the case in ,the lowor concentrationstested. After being exposed to 100 or 500'ppm'of phenol'for'24 hOUrs,'however,the majority, of animals.were extended,from ~he:sh.ell,~nd·remainedin this stateduring the 'rest of the. period of exposure.' ,Durlng ,this. time ther'o wa:s. little,if any, response .to.tactile stimulation and .anilllals,appeared.morib~d•.·~(mever,when thephenol was replacedby.seawater even some animals in.this conditionregained the. ability to close tho shell valves .together and. to 'retract"the: .~ •

: .siphons and foot. 'After:several days; some animals. !V'ere ev.en'-capabl~~ .0f.:t)U~rowing •

~ . . .,' "••' .~ • .... ~ ••• ".. .. ~ •• ;'. ,. I ~.. J.. :; •• : •

..... The effect ·af copper, a persistent. no~-degradablepollutant, on .the .' . ,"burrowing'response of T.tenuis,was .tested aso. ,contr~st.w~t~~~e nön~~~~s~stentpollutant, phenol. .~.~ ,,' /:,', .. ' ..;.....~.":J~:.

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eOPPER·.., I"; ...

" .' 96 H~ur LDSO , .' > ":(. '.: : '

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: ," ,i., Initial 96 hour' LDSO tests were~perfor6ed'by,eXposing a sample of:Tollinaof mixed lengths to 0. range of concentrations of copper sulphate insea ,waterin thc absence of sand. These tests indicnted an LD50 concentration at 11

0 e ofabout 1.00 ppm of copper under these conditions. ,:'.:" ;';

.' ~Static. tests; ,only; were' cnrri'~ci' out using coppor.:: eoncentrati6n~'of" 0,0;25;'0~50,-0~75 and 1.00 ppm 'of'copper'in the form·'of copprir'sulphate ware,tested. Tests were carried out in polythene tanks of dimensions 42.x.29 cm:containing 5 cm depth of sand and 2 litres of pollutant solution, or soa water.Before Tellina wercadded'at the' start '01' the,experiment, ',the sand' was, soakedforrseveralhours' in acoppersi.ilphatc solution of t~e;test concentration; ...~Inthis 'way i t'was hoped that adsorption of copper .on to the sand' would ~be '.completed'and there would be no"subs'equent'reduction of"concentration in thewater' durine; the test. 'At the' start 01' the test ,the coppar solütions ~ere· ~exchanged'and'25 aiiimals'of ench 01' two sizc groups, Lc~ less than.and' ;.;'grenter than 10 rom.were ~dded toeach'tunk~ 'eopper solutions,were replaced t

ever,y,24 hours for 96 hours arid burrowing'rate was'measured on each occasion.After.96 hoUrsJ.copper solution.s;worein all cases'replaced'bY'sea water. I ·:

Exchange' of: soa water and~measu'i'ement of:burrowing' rate:was carried'out dailyfor a .furth~r thre'o' days .. · .. ;.', . ...... .:,: :~, ~ '... ',: ,." .' '.

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Subacute tests:

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. ., .Results ','. ..... ".•

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., c' ,; (~) .- 'Th'~ e~fect 01' animal' size - i ,~': . .... .~ ; . ". ~., • I ... I .' ':. .

;', :; Data ~howthat''the 'r~s'ponse'of 'animals' to'pollutant solutions differs;'markedly with size; in large animals. there was a much more marked change in

4

••

••

thc' burrowing rate wh~' animals were subjected to adverse experimental ..c6riditioris~(cioIDpure'G:ro.phs·8,.9 and.10:with 11,12 and er.' .Even in thc ...

. : 'hirge contiol 'animals' which were rnaintained in .sea,water' thröuChout 'the t'est;'there was a marked reduction during thc experiment in the ·ia.~e at which··· ..:....;;

. animals reburrowed into the sand. Although sane changes also occurredin .:'.'", :thä: burrowing 'rate for'small' control"animals, ·the effect was .much.less marked

(c'omparä ~r~phs·8 rind' .11) ;':. Comparison of ,the .'number .~f; ~~athS~'at ;eiich:,-öoppcr.concentratiori' for the :two:size groups :similarly:.indicates .that 'inortality'.· .. :--:. '-~""th ···al····· :.'" _', .. ,' _.I, ,..•:.J.J..ncreases WJ.. anJ..m ·sJ..ze.· .. :-c.•• ::r.. ::. c.,;'· i .1') .:\:l ..,;.;~·:..; -.. '.. , , .

... :~~~ '.:.::J ~.~. \ .. ,;.. "::: ~ "':: "'1.' .. :" '.J ! :.. _"';:}.::.= ' . . -" '.: "

(b) The effect· of"copper' co'nccntration : , ;~~;~..~;:. ;..~..: :~ .':, ", :"'; ~.'- :..:' .. : ' .',~ ~ .:;.~• I ~, ; , : "- , . ........

" . ", .All'concentratJ..ons· of.::eopper. tested,:,i.c •. 0.25,·0.50, ,0.75 and .1.00 ppmCu 'affected'· theburrowing~.ratefor all· sizes .of;·animals., rGönsider, .first thc

··:·data,'for·soall am.oals: at-:all concentrations ,tested thers"was',found to'be 'a"reduotiori in'both the:·rato,of.:burrowing,and i~.the number o:r.arumals which": had burrowed into' the sand' within .24 hours. r~ This :redueti~n was' more' ma.rked· ,at hi'gher"copper eoncentrations .and.the cffect, in some ·Casos; at.least, ...inereased;With time '( see' Graphs -11A;:.12A~andr13A). ',För aminals.. eXp''o'sed', to, :;:

·:·0~2.5;" 0;50 andO~75 ppm; cf' eopper there·;was. a6~rtain.degree: of:' rCc'overj-·or.·the' burrowing' rate when copper. solutionswere. repla.ced .by~.sca;wa.ter (see: ~".Graph 12B). Data given here, however, do not indicate iner-eased recoven;y'withtime. Animals whieh had been exposed to 1.00 ppm of copper showed no suchrecovery (Graph 13B).

Large animals similarly showed a reduction of tho burrowing responseafter exposure to copper pollution (Graphs BA, 9A and 10A). For this sizegroup, however, there was no evidonco of recovery when thc eopper solutionswere replacod by elean sea water (Graphs 9B and 10B). This result underlinesthe fact that tho effoct of eopper on Tellin~ increases with animal size •.

DISCUSSION

Comparison of data tor phenol with those for copper shows that forphenol, ardmals havo a much greater capaeity for roco7cry after they areremoved from the direct effect of pollutant in thc water. Thera are twopossible explanations of this result; the residual copper in tho saril. may actas a deterr'ent to burrcl1dng or there ma~.. be a direct irre'lcreiblo ph~tsiological

errect on the onire:l1 it:.~clf. Further tests are in progress, to 8ho'o": therelative irlportanoe or thece two effocts. For exo..~:ple, invest:i.gation of theburrowing rate of animals taken fr~ pollutant solutions and presentcd withan unconto.minated sand oe:d.ium, "lhen eompared \vith d:lta civen here shoulddemonstrate the importanee of avoidance of eontaminat'ed sand in depressingthe burrowing response.

Several observations made during the experiments described abovo dosuggest that avoiclanee may be an important factor. In tests where 0.25 and0 •.50 ppm of copper had been replaecd by clean sea watcr, several animals wereseen to ~robe the sand surface actively with the foot, for an extended periodof time tat least an hour in some eases). In SOmO cases where animals hadbeen probing the sand in this way, a groove across tho sand 5urface indicatedthat the animal had moved a considerable distance on the surface by means ofthis action of tho foot. The fact that some of these animals did eventuallydisappear from thc; surface suggests that tho animals though capabIo ofburrowing were deterred by some adverso property of the sand itself.

5

Further evidence of avoidance of sand contaminatedbycopper isthat ina few cases atthe highest copper concentration tested (i.e. 1.00 ppmcopper)animals which initially burr'Jwed into the sand, later reappeared at the surface.(Gra~hs 10Aand 13A.) ,

In many cases,particularly for lower'copper concentrationstested, ariimals~hi.ch :CaUed to burrow into copper-'contaminated 'sand .showed no signs of beingunhealthy or'moribund. Although' siphons ,were inmany cases extended, themajorlty were withdrawn on tactile stimulation. This ,further supports .thetheory that avoidance of sand contaminated by copper i5 a major factor con­tributing to the observed change in'the burrowing ,resp,onse.

During the burrowing tests described here, incidental observations whichweremade on the occurrence of siphon holes at the sand surface suggest thatthis may also be used to investigate subacute.,effects of pollutants ßespite thelarge variation found for control animals. ' Compared 'with controls, siphon' holes •occurred muchless f'requently for all concentrations of copper tested. As theproduction of' siphon holes will be directly related to the rate of waterfiltration forfeeding and respiration~ short-term experiments testing theeffects of pollutants on siphon hole development would be a useful preliminary •to long-term studies' on the effects of pollution on grawth and metabolism. '

•"

•

6

APPENDIX:

INDEX

GRAHIS 1-4 Tellina tenuis: Static subacute toxicity tests using phenol.The effect of a range of phenol concentrations on burrowingrate.

A

B

= Burrowing rate whilst exposed to phenol solution.

= Burrowing rate after previous exposure to phenol solution.Phenol replaced by sea water.

•

1

2

34

GRAPHS 5, 6and 7

= Control, no phenol

= 10 ppm Phenol

= 100 ppm Phenol

= 500 ppm Phenol

Tellina tenuis: Continuous flow subacute toxicity testsusing phenol. The effect of a range of phenol concentrationson burrowing rate.

A

B

= Continuous flow over sand surface.

= Continuous flow through sand column.

56

7

= Control, no phenol

= 100 ppm Phenol

= 500 ppm Phenol

Tests using small animalslength ~ 10 mm

Tests using large animalslength ~ 10 mm

Control, no copper )

0.50 ppm copper ~1.00 ppm copper )

Control, no copper ~

0.50 ppm copper )

)1.00 ppm copper

Burrowing rate whilst exposed to copper in watel'.

Burl'owing rate of' animals pl'eviously exposed to copper addedin solution. Copper replaced by sea watel'.

Tellina tenuis: static subacute toxicity tests using copperas copper sulphate. The effect of a range of copperconcentrations on burrowing rate.

GRAIHS 8-13

•A =

• B =

8 =9 =

10 =11 =

12 =

13 =

•

GRAFHS OF RESULTS

KEY OF SYMBOLS USED:

Burrowing rate of' animals bef'ore exposure to pollutant.

••

••

•

o

x

o

Burrowing rate immediately after exposing animals topollutant (1. e. Day 1).

Burrowing rate of' animals which had been in pollutant f'or2 days (i.e. on Day 3).

Burrowing rate of' animals which had been in pollutant f'or3 days (Le. on Day 4).

Burrowing rate immediately after replacing polluted waterby clean S.W. 96 hours after initial addition of' pollutant( 1. e. Day 5).

Burrowing rate 1 day after replacing polluted water byclean S.W. (1. e. Day 6).

Burrowing rate 2 days after replacing polluted water byclean S.VI. (Le. Day 7).

Burrowing rate 3 days after replacing polluted water byclean S.W. (Le. Day 8).

Burrowing rate 4 days after replacing polluted water byclean S.W. (Le. Day 9). .

.,

C/)HTAO I.s - /'dli l!.i{{!C.!

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3A JB

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