Document kmMGev2OgVQvV51jJ09QvovXE

CHRONIC TOXICITY TO FISH TEST SUB TANCE Identity:N-ethylperfluorooctasnuelfonamidoethanolm;ay also be referredto as N-ETFOSE Alcoholor FM-3422. (1-Octanesulfonamide,N-ethyl1,1,2,2,3,3,4,4,5,5,6,6,7,7,8,8,8-heptadecafluoro-N-(2-hydroxyethyl)-, CAS # 1691-99-2) Remarks: Materialisan off-whitew,axy solidof uncharacterizedpurity.The laboratoryconductingthe study refersto N-ETFOSE Alcoholas "78.01". METHOD: Method Followed: The methodology forthe egg and fryexposure closely followedthatpresented in"Proposed recommended bioassay procedureforegg and frystages offreshwaterfish" (U.S.EPA, 1972). Type (testtype):Flow-through GLP: No Year study performed: 1978 Species/Strain/SupplierF:athead minnow (Pimephalespromelas)eggs from the brood stockofthe U.S. EnvironmentalProtection Agency's EnvironmentalResearch Laboratoryin Duluth,Minnesota. Analyticalmonitoring: Temperature, dissolvedoxygen concentrationa,nd pH were monitoreddaily,alternatinbgetween aquaria such thateach aquarium was measured once each week. Remarks: One thousand mi water samples were taken from each aquarium at the initiatioofnthetest(day 0),when hatchingwas completed (day4),and weekly thereafte(rdays 11, 18,25, and 32) fordeterminationof N-ETFOSE Alcoholconcentration.Samples were taken witha beaker from a point approximatelymidway between the surface,bottom and sidesof each aquarium, and storedin1000 ml amber glassbottleswithfoilinedcaps. Allsamples taken duringthe testwere storedat room temperatureand shipped on May 31, 1978 to the3M Company. Exposureperiod: 4,11,18,25,and32days StatisticaMlethods: Means ofmeasured biologicaplarameters from duplicateaquariawere subjectedto analysisofvariance(Steeland Tonie, 1960, completelyrandomized blockdesign,P=0.05). Data forpercentagesurvivaland percentage hatch were transformedto arcsinsquare rootof percentage priorto analysis. Test Condition Remarks: Exposures were initiatewdithin48-hours afteregg fertilizatiaonnd continued through 30 days post-hatch. The diluentwater was wellwater pumped to a concretereservoirwhere itwas aeratedbeforeflowingtothe exposure system throughaged PVC pipe. The pH ofthiswater ranged from 7.0-7.2;totalhardness,31-38 mg/L as CaC03; alkalinit2y6,-29 mg/L as CaC03; and specificonductance,149-170 umhos/cm. Fivenominal concentrationsof N-ETFOSE Alcohol,rangingfrom 1.3-20gg/L, controlwater (wellwater),and controlwatercontainingsolvent (Dimethylsulfoxid(eDMSO) at24 mg/L) were deliveredtoduplicatetestaquaria. A 50 ml gas-tightsydnge was used to deliver46 tiolfa 0.84 mg/mi stocksolution of N-ETFOSE AlcoholinDMSO through a stainlessteelneedle and polyethylene tubingtothe mixingchamber ofthe diluter. Each glasstestaquarium measured 40 X 20 X 25 cm witha 19 cm high standpipedrainwhich maintaineda constanttestwater volume of 15.2 liters. The dilutedrelivered0.50 literosftestwater to each aquadum 185 times per day, yieldinga 90% testwater replacementtime of approximately9 hours. The aquariarestedina water bath containingcirculatinwgater heated by immersion coilheaterand regulatedby a mercury column thermoregulatordesigned to maintainthetestwater temperatureat250C. Upon arrivaalt E G & G ,Bionomics,the eggs were allowedto acclimatefrom 17.50C to thetesttem*peratureof250C over a pedod oftwo hours. Sixtyeggs were then randomly distributetdoeach of 14 egg cups which were then dipped in a 60 mg/lmalachitegreen solutionfor15 seconds topreventfungalgrowth. One egg cup was then suspended ineach ofthe 14 testaquaria.EFR incubation cups were glassjars(5cm O.D.,8 cm high)with40 mesh Nitex screen bottoms. An egg cup rockerarm apparatus,as describedby Mount (1968),was used to gentlyoscillattehe egg cups inthe testwater. Dead eggs were removed and counted dailyuntilhatchingwas completed. Percentage hatchwas calculatedbased on the number oflivefryper aquarium afterhatchingwas completed compared tothe number ofeggs per aquadum (60) atthe initiatioofnthe exposure. To initiatehe 30-day fryexposure,fortyfrywere randomly selectedfrom each egg cup and transferretdothe respectiveaquaria. Upon transferoffrytothe aquaria,the frywere fed newly hatched San Francisco Bay varietybrineshrimp naupliia,d libitumt,hreetimesdailythroughoutthe exposure period.The aquariawere brushed and siphonedto remove excess food and fecalmaterialthreetimesper week. At 30 days post-hatch,percentage survivalm,ean totallength,and mean wet weightwere determined forfryfrom each aquarium. The frywere measured individualtloycalculatea mean and standarddeviationtotallengthwhileeach frygroup (fryfrom one aquarium) was wet weighed tocalculatea mean wet weight. At theterminationofthe test,thefryfrom the controland the highconcentration (20 gg/L)were preservedin10% bufferedformalinwhilethefryfrom the other testaquariawere frozen.Ten formaln!-preservedfry(5from each replicatef)rom the controland the highconcentrationunderwent histopathologicaelxamination ofa transversesectionofthe nares and cephalicextensionofthe lateralline (See Table 3). The remainingpreservedfryand frozenfrywere analyzed at a laterdate (by3M Company) forN-ETFOSE Alcoholconcentrations. RESULTS The biologicadlatagenerated inthisstudyindicatethathatchabilitoyfeggs, and percentage survivalm,ean totallengthand mean wet weight offrywere not adverselyaffectedatany N-ETFOSE Alcoholconcentrationtested. Remarks: Water qualityparameters measured duringthe egg and fryexposure exhibited littvlaeriationbetween testdays and testchambers. Dissolvedoxygen ranged from 8.2-10.2mg/L. Temperature ranged from 23.5-26.50Cand pH ranged from 6.9-7.5. TABLE 1 PERCENTAGE HATCH OF EGGS, PERCENTAGE SURVIVAL, MEAN AND STANDARD DEVIATION TOTAL LENGTH, AND MEAN WET WEIGHT OF FATHEAD MINNOW (Pimephalespromelas)FRY EXPOSED TO N-ETFOSE ALCOHOL FOR 30 DAYS POST-HATCH., Nominal concentration (ggA) Control Solvent Control 1.3 2.5 5.0 10 20 Replicate A B A B A B A B A B A B A B Hatch % 93 92 92 93 90 92 97 95 88 92 93 92 92 87 Survival% 82 92 82 90 92 92 90 95 100 92 92 88 98 98 30 Days Mean lengthin mm L+SD) 21(3) 21(2) 20(3) 20(4) 21(3) 21(3) 21(3) 21(2) 20(4) 21(2) 21(3) 21(2) 21(2) Mean weight in mg 69 71 64 66 65 70 65 64 58 69 60 64 66 The 1000 mi watersamples takenduflngthestudy(describedin"Analytical Monito@ng")were analyzedat3M Company. Resultsindicateedxcellent recovery.See Table TABLE2 ANALYSES FOR N-ETFOSE ALCOHOL IN BIONOMICS SAMPLES TO VERIFY RECOVERY CAPABILITIES TheoretiCcoanlc. Sample I.D. (ppb) A5619 ControlA A5620 ControlB A5621 SolventControlA A5622 SolventControlB A5617 1.25 A5618 1.25 A5670 1.25 A5670 1.25 A5615 2.5 A5616 2.5 A5613 5 A5614 5 A5611 10 A5612 10 A5609 20 A5610 20 (a) receivebdrokeninshipment (b)replacesoriginaslamplesA5617;A5618 Determined (ppb) <0.1 <0.1 <0.1 <0.1 (a) (a) 1.1;1.0(b) 1.0;1.6(b) 2.5;2.6 1.2;1.3 5.2;5.2 4.8;5.1 10.2;9.9 9.7;10.1 20.8;19.8 19.8;20.6 TABLE 3 RESULTS OF HISTOPATHOLOGICAL EXAMINATION OF FRY OF FATHEAD MINNOW (PIMEPHALES PROMELAS) EXPOSED TO 20 @Lg/OlF N-ETFOSE ALCOHOL' TestMaterial Control Number of Observations 10 N-ETFOSE AJcohol 10 I a. Work performedundercontracttoEG Histopathological Findings 3/10Normal 6/10LiverFattyChange 3110GillHyperplasia 1(EpitheliuIm) 9/10 Normal 1 1/10GillHyperplasia & G BionomicsLaboratory NOTE: "Onlythosetissueswhichwere missingorcontaineddemonstrablechange are listedT.he onlytissuechanges observedwere hyperplasioafgilllamellaerpitheliuamnd fattcyhangeoftheliverT.hese changeswerejudgedtobe minimaland consistenwtith changesseen routineliynhealthyfish.Autolipiosfgiltlissuewas observedinseveral fish.Thischangewas probablydue tothepoorpenetratioonfthebufferedformalitno theposteriodrorsalportioonfthegilslpace." CONCLUSIONS The No EffectConcentration(NOEC) of N-ETFOSE alcoholforhatchability, growth,histopathologyand survivalofFathead minnow fryisgreaterthan 19.820.8 pg/L measured (20 pg/L nominal),the highestconcentrationtested. DATA QUALITY ReliabilityK:limischranking= 2. This studymeets allthe criterifaorquality testingb,uthas severaldeficienciesI.tlacksinformationon purityofthe test substance,the productionlotnumber from which the testsample was taken,and was notdone atthe reportedsolubililtiymiftorthe testsubstance. REFERENCES Thisstudy was conducted by E G & G, Bionomics,Wareham, Massachusetts, 1978 on the requestof3M Company. .ResearchReport'7HE EFFECTS OF CONTINUOUS AQUEOUS EXPOSURE TO 78.01 (N-ETFOSE ALCOHOL) ON HATCHABILITY OF EGGS AND GROWTH AND SURVIVAL OF FRY OF FATHEAD MINNOW (Pimephales promelas)."Report# BW-78-6-195, E G & G, Bionomics,AquaticToxicology Laboratory,790 Main Street,Wareham, Massachusetts,June 1978. Research Report "SUMMARY OF HISTOPATHOLOGICAL EXAMINATION OF FATHEAD MINNOW (Pimephales pr-omelas)EXPOSED TO 78.01 (N-ETFOSE ALCOHOL) FOR 30 DAYS." Report # BW-78-6-195, E G & G, Bionomics, AquaticToxicologyLaboratory,790 Main Street,Wareham, Massachusetts,June 1978. 3M TechnicalReport Summary, "AnalyticaMlethodology and Support",Fate of Fluorochemicals,ProjectNumber 9970612643, Report Number 008, Arthur Mendel, January 17,1979 OTHER Submifter: 3M Company, Environmental Laboratory,P.O. Box 33331, St.Paul, Minnesota,55133 Last changed: 5/18/00 THE EFFECTS OF CONTINUOUS AQUEOUS EXPOSURE TO 78.01 ON HATCHABILITY OF EGGS AND GROWTH AND SURVIVAL OF FRY OF FATHEAD MINNOW (Pimephales promelas). q2-?- RESEARCH REPORT SUBMITTED TO 3M COMPANY ST. PAUL, MINNESOTA REPORT #BW-78-6-195 E G & G, Bionomics Aquatic Toxicology Laboratory 790 Main Street Wareham, Massachusetts June, 1978 ABSTRACT Fathead minnow (Pimephales promelas) eggs and fry were continuously exposed to nominal 78. 01 concentrations ranging -from 20 to 1.3 Ug/l through 30 days post-hatch. Data were collected as percentage hatch of eggs, and survival, total length, and average wet weight of fry. Results of these data indicate that none of the above parameters were adversely affected by exposure to any of the 78.01 concentrations tested. TABLE OF CONTENTS SECTION PAGE I ii III IV v INTRODUCTION ........................................ MATERIALS AND METHODS ........................o...... 3 A. Exposure System ........o .............o .......... 3 B. Egg and Fry Exposure ...................o........ 4 C. Statistics ......................o ........ooo.... 7 RESULTS ......................... ........ 8 REFERENCES..oo.o ....ooo..o....o..*ooooooo.eoooooooo. 9 TA.BLE..ooo ......o ....o .... o o ...oo .............o... o. 11 SECTION I .INTRODUCTION The objective of this study was to determine the effects of 78-01 on fathead minnow (Pimephales promelas) eggs and fry during continuous aqueous exposure. Exposures were initiated within 48-hours after egg fertilization and continued through 30 days post-hatch. The effects on egg hatchability and on survival and growth of fry were measured and would be used to make an estimate of the@@MTC (minimum threshold concentration). The MTC is virtually synonomous with the term MATC (maximum acceptable toxicant concentration) developed by Mount and Stephan (1967). Mount and Stephan's term, however, was estimated after the performance of full life@:cycle chronic test where e--r-fectson reproduction and second generation fry were also measured. Macek and Sleight (1977) and McKim (1977) described egg and fry investigations as being reasonably accurate short-term estimations of potential long-term chemical hazards to fish, and as being similar to those estimations derived from definitive chronic toxicity studies. In the majority of the studies reported by the authors and of those'performed at this laboratory, the embryos and fry during early stages of development were generally the most sensitive stages to chemical exposure. Rarely was reproduction or survival and growth of second generation fry reduced at exposure levels lower than those that reduced survival or growth of the first generation fry. The authors demonstrated that for the great majority of toxicants, the quicker and more economical egg and fry tests yielded estimates of safe concentrations very similar to those derived from chronic toxicity studies. 2 SECTION II MATERIALS AND METHODS The methodology for the egg and fry exposure closely followed that presented in "Proposed recommended bioassay procedure for egg and fry stages of freshwater fish" (U.S. EPA, 1972). The test material used, labelled 78.01, a white powder, was obtained January 23, 1978 from the 3M Company, St. Paul, Minnesota. A. EXPOSURE SYSTEM A modified, proportional diluter similar to that described by Mount and Brungs (1967) with a 0.50 dilution factor was used . in this study. The diluent water was well water pump ed to concrete reservoir where it was aerated before flowing to the exposure system through aged PVC pipe.. The pH of this water ranged from 7.0-7.2; total hardness, 31-38 mg/t as CaCO3;, alkalinity, 26-29 mg/i as CaC03; and specific conductance, 149-170 limhos/cm. The diluter delivered five nominal 78-01 concentrations ranging from 20-1.3 Ug/t, control water (well water), and control water containing solvent (Dimethylsulfoxide, DMSO) to duplicate test aquaria. A 50 mi gas-tight syringe was used to deliver 46 pt of a 0.84 mg/ml stock solution of 78.01 in DMSO through a stainless steel needle and polyethylene tubing to the mixing chamber of the diluter. A'mariotte bottle-volumetric tube 3 metering device was used to deliver DMSO into the solvent control aquaria at a concentration equal to.the highest concentration of DMSO in the 78.01 exposure aquaria (24 mg/L DMSO). Each glass test aquarium measured 40 x 20 x 25 cm centimeters (cm) with a 19 cm high standpipe drain which maintained a constant test water volume of 15.2 L. The diluter delivered 0.50 1 of test water to each aquarium 185 times per day, yielding a 90% test water replacement time of approximately 9 hours (Sprague, 1969). The aquaria rested in a water bath containing circulating water heated by immersion coil heaters and-regulated by a mercury column thermorecjulator designed temperature at 250 C. to maintain the test water B. EGG AND FRY EXPOSURE On March 31, 1978, the exposure of fathead minnow eggs to 78.01 was-initiated. Eggs used were from the brood stock of the U.S. Environmental Protection Agency's Environmental Research Laboratory in Duluth, Minnesota. Upon arrival'at E G & G, Bionomics, the eggs were allowed to accl4-mate from 17.5 0 C to the test temperature of 250 C over a period of two hours. Sixty eggs were then randomly distributed to each of 14 egg cups which were then dipped in a 60 mg/i malachite green solution for 15 seconds to prevent fungus growth. One egg cup was then suspended in each of the 14 test aquaria. Egg incubation cups were glass jars (5 cm O.D., 4 8 cm high) with 40 mesh Nitex R screen bottoms. An egg cup rocker arm apparatus, as described by Mount (1968), was used to gently oscillate the egg cups*in the test water. Dead eggs were removed and counted daily until hatching was completed. Percentage hatch was calculated based on the number of live fry per aquarium after.hatching was completed compared to the number of eggs per aquarium (60) at the initiation of the exposure. To initiate the 30-day fry exposure, forty fry were randomly selected from each egg cup and transferred to the respective aquaria. Upon transfer of fry to the aquaria, the fry were fed newly hatched San Francisco Ba@@ variety brine shrimp nauplii, ad libitum, three times daily throughout the exposure period. The aquaria were brushed and siphoned to remove excess fqod and fecal material three times per week. At 30 days post-hatch, the fry from each aquarium were anesthetized with MS-222 (tricaine methane-sulfonate) and percentage survival, mean total length, and mean wet weight were determined. The fry were measured individually to calculate a mean and standard deviation total length while each fry group (fry from one aquarium) was wet weighed to calculate a mean wet weight. At the termination of the test, the fry from the control and the high concentration (20 pg/t) were preserved in 10% buffered formalin while the fry from the other test aquaria were frozen. 5 Ten formalin preserved fry (5 from each replicate) from the control and the high concentration were sent to the EnvirorAmental Pathology Laboratories, Inc., Carolina, Rhode Island for histopathological examination of a transverse section of the nares and cephalic extension of the lateral line. The remaining preserved fry and frozen fry were sent to the 3M Company, St. Paul, Minnesota, May 31, 1978. Temperature, dissolved oxygen concentrations, and pH were monitored daily, alternating between aquaria such that each aquarium was measured once each week. Temperature was measured with a mercury thermometer, dissolved oxygen with a YSI Model #54 dissolved oxygen meter and probe, while pH was measured with an Instrumentation Laboratory Model #175 portable pH meter and probe. One thousand mi water samples were taken from each aquarium at the initiation of the test (day 0), when hatching was.completed (day 4), and weekly thereafter (days 11, 18, 25 and 32) for determination of 78.01 concentration. Samples,were taken with a beaker from a point approximately midway between the surface, bottom and sides of each aquarium, and stored in 1000 mt amber glass bottles with foil lined caps. All samples taken during the test were stored at room temperature and shipped on May 31, 1978 to the 3M Company, St. Paul, Minnesota. 6 C. STATISTICS Means of measured biological parameters from duplicate aquaria were subjected to analysis of variance (Steel and Torrie, 1960, completely randomized block design, P=0.0,5). Data for percentage survival and percentage hatch were transformed to arc sin VpercentE-g-ep-rior to analysis. 7 SECTION III RESULTS Water quality parameters measured during the egg and fry exposure exhibited little variation between test days and test chambers. Mean measured dissolved oxygen was 9.0 + 0.5 mg/l and ranged from-8.2-10.2 mg/l. Mean temperature was 25 + 10c and ranged from 23.5-26.5 0C and pH ranged from 6.9-7.5. The biological data generated in this study (Table 1) indicate that hatchability of eggs, and percentage survival, mean total length and mean wet weight of fry were not adversely affected at any 78.01 concentration tested. Based on thi Fdatuumm,, thee MTCC for fathead minnows and 78.01 is estimated to be >20 pg/i based on noninal concentrations..,, 8 SECTION IV REFERENCES Macekl X.J. and B.H. Sleight, 111. 1977. Utility of toxicity tests with embryo and fry of fish in evaluating hazards associated with chronic toxicity of chemicals to fishes. Symposium Proceedings, ASTM, Memphis, Tennessee, Octoberf 1976: 137-146. McKim, J.M. 1977. Evaluation of tests with early life stages of fish for predicting long-term toxi&ity. J. Fish. Res. Bd. Can. 34: 1148-1154. Mount, D.I. 1968. Chronic toxicity of copper to fathead minnow (Pitnephales promelas, Rafinesque).. 215-223. Water Res. 2: Mount, D.f. and II.A. Brungs. 1967. A@ simplified dosing apparatus -20-29. for fish toxicology studies. Water Res. 1: 14ount,,D.I. and C.E. Stephen. 1967. A method for establishing acceptable toxicant limits for fish, malathion and the butoxyethanol ester of 2,4-D. Trans. Amer. Fish. Soc. 96: 185-193. Sprague, J.B. .1969. Measu.rerients of pollutant toxicity to fish. I. Bioassay methods for acute toxicity. Water 9 Res. 3: 793-831. Steel, R.G.D. and J.H. Torrie. 1960. Principles and pro- cedures of statistics. I-IcGraw-Hill,New York: 481 pp. U.S. EPA. 1972. Proposed recor=ended bioassay procedure for egg and fry stages of freshwater fish: 7 pp. 10 SECTION V TABLE Table 1 Percentage hatch of eggs, percentage survival, mean And standard total length, and mean wet weight of fathead minnow (Pimephales p exposed-to 78.01 for 30 days post-hatch. .... .......... Nominal concentration (ug/.E) Replicate Hatch 30 Days Survival. Mean length mc (mm) 20 10 5.0 2.5 1.3 control solvent -control A 92 98 21(2) B 87 98 21(2) A 93 92 21(3) B 92 88 23(2) A 88 100 20(4) B 92 92 21(2) A 97 90 21(3) B 95 95 21(2) A 90 92 21(3) B 92 92 21(3) A 93 82 21(3) B 92 92 21(2) A 92 82 20(3) B 93 90 20(4), SUBMITTED BY: PREPARED BY: APPROVED., @BY: E G & G, Bionomics Aquatic Toxicology Laboratory 790 Main Street Wareham, Massachusetts June, 1978 Jerry W. Dean tic 03.010giSt Stephen J. Elis artic Tovicologist George A. Cary Z,5ee Director, Aqua ic Biology SUMMARY OF HISTOPATHOLOGICAL EXAMINATIONS OF FATHEAD MINNOW (Pimephales promelas) EXPOSED TO 78.01 FOR 30 DAYS. 3 @zz. RESEARCH ]REPORT SUBMITTED To 3M COMPANY ST. PAUL, MINNESOTA REPORT #BW-78-9-300 SUBMITTED BY E G & G, Bionomics Aquatic Toxicology Laboratory 790 Main Street Wareham, Massachusetts September, 1978 INTRODUCTION An "egg and fry test" was conducted between March 31 and May 4, 1978 (tionomics Report #BI-7-78-6-195, June, 1978) to assess the sub-lethal effects of 78.01 on fathead minnows (Pimephales promelas). Effects on egg hatchability and on survival and growth of fry were measured. To further investigate possible effects due to exposure to 78.01, histopathological examinations of exposed fish were also performed. The results of these examinations are reported here. ,MATERIALS AND METHODS Upon termination of the 6gg and fry study, five fish from each replicate of the high 78.01 concentration (20 ug/y,, nominal) were preserved in 10% buffered formalin and sent., to the Environmental Pathology Laboratories, Inc. Carolina, 017.- Rhode Island. Control fish from a similar ongoing study were also examined. The fish were examined for gross lesions, and sagital sections were prepared by a pathologist. Fish were placed with identifying numbers into processing cassettes, dehydrated, cleared and infiltrated on an Auto-technicon tissue processor using the method of the Armed Forces Institute of Pathology. The fish were then placed in a vacuum oven and subsequently embedded in paraffin. All blocks were sectioned at 6 microns: one 2 slide was prepared fron, each block and stained with hematoxylin and eosin. Special staining procedures such as PAS, Trichrome and Acid Fast were used occasionally when requested by the pathologist. All blocks were sealed with paraffin and stored. Slides were labeled, boxed and delivered.w*ith all work sileets to the pathologist for examination. Two slides were prepared from each fish. Tissues processed and examined included but were not limited tc>;gill, thymus, liver, spleen, heart, gonad, kidney, foregut, hindgut, olfactory mucosa (nares), brain, ear, skin, muscle, pancreas and pharyngeal mucosa. RESULTS A summary of the examination of each fish is reported Table 1. Only those tissues which were missing or contained demonstrable change are listed. The only tissue changes observed were hyperplasia of gill lamellar epithilium and fatty change of the liver. These. changes were judged to be minimal and consistent with changes seen routinely in healthy fish. Autolipis of gill tissue was observed in several fish. This change was probably due .to the poor. penetration of the buffered formalin to the posterior dorsal portion of the gill space. 3 Based on these data', it is concluded that 30 days exposure to a nominal 78.03 concentration of 100 mg/l did not cause any significant, demonstrable, tissue changes in 30 day old fathead minnow fry. Table 1 Histopathological examination of fathead minnow (Pimephales promelas) exposed 30 days to 20 ug/i nominal 78.01 and control water. 20 ug/y._78.01 Fish 1 2 3 4 5 6 7 8 9 10 Control 2 3 4 Sex F.a Ukn b Ukn Ukn p p F p F F Ukn Ukn F F Organ Condition Missing Tissue Normal Normal Normal Normal Normal Normal Normal Normal Normal Gill Hyperplasia (epithelium) Ic Ear, thymus Spleen, gonad, nares Spleen, gonad, thymus Spleen, heart, gonad, nares Spleen, nares. Spleen, nares, thymus Spleen, heart- Spleen, heart, nares Spleen Spleen, heart,. thymus Liver-fatty change I Gill-hyperplasia (epithilium)I Liver-fatty change I Gill-hyperplasia (epithilium)I Normal Gon,-ld,nares, thymus Spleen, gonad, nares Thymus Spleen, nares Table 1 (cont.) Fish Sex control 5 md 6 F 7. Ukn 8 Ukn 9 F 10 Ukn a Female. b Unknown. c Lesions are minimal. d male Organ Condition Missing Tissue Normal Liver-fatty change I Spleeh, thymus, heart Spleen, heart Liver-fatty change I Spleen, heart, gonad, thymus, nares Liver-fatty change I Spleen, gonad, nares, thymus Liver-fatty change I Gill hyperplasia (epithelium) I Spleen, nares Normal Gonad, thymus SUBMITTED BY: PREPARED BY: APPROVED BY: E G & G, Bionomics Aquatic Toxicology Laboratory 790 Main Street Wareham, Massachusetts September, 1978 Stephen J. Ells Aqartic re(nwinologist George A. Cary Director, Aquatic.,,@iogy