COMI A Os) DAMOS - Mussel Watch Oct 1786 Western Long Island Sound Disposal Site Monitoring Project June 1,1984 - June 1, 1985 Disposal Area — Monitoring System DAMOS Contribution 51 October 1988 ay fo BO FUMENT OS. US Army Corps \ Woods Hole Oceanograp! of Engineers 1 nes Sei New England Division ~ ener — A ec Ke (Oe Ww SEA VC Ci | DEMCO DAMOS - MUSSEL WATCH WESTERN LONG ISLAND SOUND DISPOSAL SITE MONITORING PROJECT JUNE 1, 1984 - JUNE 1, 1985 CONTRIBUTION #51 OCTOBER 1988 Written by: S.Y. Feng Marine Sciences Institute The University of Connecticut Groton, Connecticut 06340 Submitted to: Regulatory Branch New England Division U.S. Army Corps of Engineers 424 Trapelo Road Waltham, MA 02254-9149 NM 0 Submitted by: Tt Science Applications International Corporation Admiral's Gate 221 Third Street Newport, RI 02840 (401) 847-4210 iil M MOQ 0301 0 0 US Army Corps of Engineers New England Division yeti? nievovial } ait] éh HPolilagsd. etait Pit fade. week an : WA TABLE OF CONTENTS AcknowledgementS ...---e-eesccecscrcccccsccccce 500000000000000 i Executive Summary ....--cccccccccserecses creer scecsrcsescccees Li Introduction Western Long Island Sound Disposal Site ................. iL Eastern Long Island Sound Reference Station ............. 1 Project .ObjJectives ..... ccc eee cece cree creer ese cc cc cccccccccce 1 Experimental Field Operations ....... ..c.cecsnscsceccescscceccccccscne 2 Laboratory ProcedureS ..-eceeesceceererercee D000000000000 3 Trace Metal AnalySeS .....-cecccccscrecesrcsssecrrces 3 Analyses of Polychlorinated Biphenyls ............-. 3 Gas Chromatograph/Mass Spectrometry Identification of Phthalate Esters ......-2-c-s2cee- Sb000000000000000 3 Histopathological Studies ............eseseeeeeeeees 4 Statistical Analyses of the Data ..... 50 DG SOOO Sets eee Results Trace Metal Concentrations ......-.cccccccccccrccreceesees 5 Polychlorinated Biphenyl (PCBs) Concentrations 5000000000 6 Bis (2-ethyl hexyl) Phthalate ............---------- eee 7 MopAeeUbeleS Gaoococc0dGuKdaDDbODGbD0O0OD DO0000D0DD0DODDDOUDNS 8 Histopathological Studies S600d0D0D DO ODDO ODDO O DOD DNDOODDNDR 9 Discussion Trace Metal and PCB Concentrations .........cecccccceces 12 I@aleeplenyl Issa coecootodcuncc acco dD Og DK OCDOD CO S00 ODNbE 14 GOMGMUSHOTWS soos sosooodaabooo boo ao D OKO Dod Ob ODD ODO ODO ODOC OOOO NS I7/ YSNEETOEMEGS Gaeeel cooconnccan0 DDG FCO OD OOKKDDODODDNDOOS 5600000050 20 WEDS sacacccooc000000 SdOD OUD DOD ODU OOOO OOO OOUO OOO OOOO OOCODODDDS 23 ITIMCAURIS ad doagocshoococsocouc006 Soetlst ar ot enous st aieren cnepemetoienens VOBEAO OOOO 43 Appendix Tables la,b - 4a,b summarize tissue trace metal ......... 54 concentrations (expressed in ug/g wet weight and ug/g dry weight) observed in mussels deployed at RIr, WLISrN, WLISc and 500MW Tables 5-8 list PCB concentrations (expressed in ug/g ... 64 wet weight and ug/g dry weight) found in mussels deployed at RIr, WLISrN, WLISc and 500MW Table 9 summarizes PCB concentrations (expressed in ..... 70 ug/g wet weight and ug/g dry weight) found in mussels maintained at LATr eee abgaeeh eee Ae anoissrsHeono2 (ets? ra Shialge << ioe bien ate ladsat a ee ee Lt o's aan ara ee aes ee navi Tae? <2 te Ze peeve asgae: eutet+ wit isawue dune BO ih eae Sitphew sow p\oy af Seseexqney anolvasda a i ‘Bovedeeh, ala sae fi hewrelds isdesew web) BN ove bate a Maetey a) See ae ONE peceaa aries tetaaarog ao4 : _eMn08: betes prea Rares ne habibaenue aaa ero ban: fe DaROT (tdpiow Yap ons f is95..0 ay stag FS 7 4: Acknowledgements I wish to express my sincere thanks to my graduate students: Messrs. E. Miller, S. Tettelbach, L. Crockett, R. Niesenbaum, Ms. S. Kelly, Ms. H. Crawford, and Ms. R. Bishop, as well as my technician Mrs. E. Haddad for their support in the field and laboratory. Special thanks are due to Mr. Robert DeGoursey, Institute Diving master, for his management of the demanding field diving program. Sampling of mussels would have been impossible without his logistic support and vigilance in maintaining the mussel platforms. I am grateful to Mr. John Volk, Director, Division of Aquaculture, State of Connecticut Department of Agricultural and Natural Resources, and Captain Speer for providing the shellfish for sampling mussel platforms maintained at the Western Long Island Sound Disposal site. I am also indebted to Mr. Steve Congdon, Project Manager, New England Division, U.S. Army Corps of Engineers, for supplying records of the volume of dredged material disposed in Long Island Sound waters, as well as providing the necessary funds through Dr. Robert W. Morton of Science Applications International Corp. to carry out the tasks. Finally I should like to thank Mrs. Joyce Lorensen for her devoted and competent clerical assistance. Executive Summary Biological monitoring of Western Long Island Sound Disposal Site (WLIS) was conducted from June 1984 to June 1985 using mussels (Mytilus edulis) suspended from underwater platforms. The mussel monitoring platforms were deployed at four stations: WLISc and 500MW, on and near the disposal site, WLISrN, the reference station, and RIr or LATr, the master reference station at Eastern Long Island Sound. Monitoring criteria included the tissue concentrations of nine trace metals and three PCB Aroclors (1242, 1254 and 1260), the tissue wet and dry weight ratios, and the mortalities and histopathology of the mussel populations. In addition, extrinsic factors such as water temperatures, quantities of dredged material disposed and period of disposal activities were also recorded. The objectives of the mussel watch project were to determine (1) whether or not increases in tissue concentrations of trace metals and polychlorinated biphenyls were associated with on-going open-water disposal of dredged materials and (2) whether or not such increases were correlated with changes in mortalities, in tissue wet/dry weight ratios, in gonadal development, as well as in histology. Trace Metal Data. Trace metal results indicated similar significant spatial and temporal variations as reported in our previous 1983 and 1984 studies at the Central Long Island Sound (CLIS) and WLIS disposal sites (Feng 1984, 1985). Spatial variations in Cd, Cu and Zn tissue concentrations were noted among the mussel populations held at the four stations; levels of these three metals were significantly lower in the population at the master reference station, RIr, than in the other three populations. The reference station, WLISrN, displayed the same variations in tissue concentrations of Cu, Cd, and Zn, as the two disposal site stations, WLISc and 500MW, indicating that the concentrations were not associated with the disposal of dredged materials at the WLIS disposal site. Temporal or before-during disposal differences in the tissue concentrations of Co, Cu, and Fe were observed in the three mussel populations deployed in western Long Island Sound. In all cases the three tissue trace metal concentrations before disposal were significantly lower than those during disposal. Although no post-disposal data were available, based on our previous experience, we could predict that the elevated levels of the trace metals encountered in mussels during the disposal period were transient and of short duration. PCB data. Similar conclusions as presented for the trace metals could also be drawn from the PCB results. There was a discernible increase of the tissue PCB concentrations in all four of the mussel populations. Hence, no spatial difference could be attributed to the disposal activity at WLIS. Heightened levels of Aroclor 1242 and total PCBS associated with the disposal operation were observed at WLISc and WLISrN. Because stat the resources provided for the present investigation did not allow for examining all sources of PCB in the environment, i.e., dry fallout, riverine contribution, PCB associated with seston, etc. (dredged material was not the only source), even in cases where correlations were established, e.g. before-during disposal difference in PCBs, causation could not be assumed. The general conclusion that disposal of dredged material played a minor role in elevating the tissue trace metal and PCB concentrations, was further augmented by the results obtained from employing the procedures of stepwise multiple regression analysis. It was shown that intrinsic variables, i.e., wet/dry ratios and shell length, not disposal volume, could account for a major proportion of variance observed in the tissue trace metals. The disposal volume as an independent variable entered only ca. 28% of the cases, and it ranked third or fourth when it entered the regression model. For the tissue PCB concentrations obtained at WLISc and 500MW, 50-80% of the variance could be explained by the intrinsic variables. The only exception was the Aroclor 1242 concentration at WLISc where the volume of dredged material disposed accounted for 74% of its variance. Mortalities. The patterns of cumulative mortalities of the four mussel monitoring populations were similar to those of the trace metals levels. Generally there was no spatial difference that could be attributed to the disposal operation between the reference population, WLISrN and the two populations closest to the disposal site, WLISc and S500MW. The sharp increase of cumulative mortalities from January to March 1985 at WLISc appeared to be associated with the accelerated disposal activity at the site. Because this was the only population to exhibit such a trend, it is surmised that the effect of disposal on mortality, if any, was limited to the immediate disposal area. Histopathology. Among the seven criteria examined, there was evidence to suggest that retardation of gonadal development, Leydig tissue staining characteristics, and the absence of crystalline style and changes in kidney tubules could be associated with the placement of mussels at WLISc and 500MW and by inference with disposal activities. However, these results should be considered only as a presumptive and the interpretation tempered with caution for the following reasons: 1. Field experimental results are by nature correlational, therefore, no causation can be assumed; and 2. The histopathology results presented herein were derived from mussels which survived mortalities of undetermined causes and hence, represented the "fittest" particularly at the three transplanted stations in the western Sound (WLISc, 500MW and WLISrN), where the average cumulative mortality was more than 70%. abalat The results could be biased in favor of lesser spatial association with the observed abnormalities. If all the dead and dying mussels had been recovered by more frequent sampling and their tissues examined, more definitive conclusions might be reached. lv Introduction Western Long Island Sound Disposal Site The Western Long Island Sound disposal site (WLIS) located 8.33 kilometers southeast of Stamford Harbor is near a historic site (Eatons Neck) and one of the deepest disposal sites (40 meters) in Long Island Sound (Fig. 1). It was opened in March 1982 and began to receive dredged materials in April 1982; a total of 43,063 cubic meters (56,325 cubic yards) of dredged materials were deposited that year. During its second (1983) and third (1984) years of operation, 114,405 and 126,923 cubic meters (149,635 and 166,008 cubic yards) of dredged materials were disposed respectively. In the fourth year (1985), 298,518 cubic meters (390,445 cubic yards) were disposed (Table 1, Fig. 2). During the time of this study, June 1984 to June 1985, a cumulative volume of 304,622 cubic meters (398,445 cubic yards) of dredged material were disposed. The largest volumes of dredged material were deposited between January and June of 1985, with April claiming the highest monthly volume of 119,518 cubic meters (156,330 cubic yards). Most of the dredged materials came from various Connecticut and New York marinas as well as boat yards operating in Western Long Island Sound, and differed presumably in their chemical contents from those disposed at the Central Long Island Sound disposal site, which were of industrial harbor origin. Eastern Long Island Sound Reference Station Ram Island reference station (RIr) (Fig. 3) in Fishers Island Sound is located approximately 260 meters south of Ram Island and served as the master monitoring reference as well as the source of mussels for the other three mussel monitoring platforms. The depth of this location varies from 10 to 12 meters. The sea bottom is generally paved with mixed gravel, shells, and mud; its boulder-dotted surrounding is interspersed with outcrops of relic clay banks. The Latimers Light reference station (LATr) has similar sea bottom features as the Ram Island station and was the source of mussels for restocking the three experimental platforms in March 1985 when Ram Island mussels were not available due to natural mortality. Project Objectives The principal objective of this study was to monitor possible deleterious effects on the environment during on-going open water disposal of dredged materials in Long Island Sound; al the DAMOS Mussel Watch Project was concerned with the following questions: 1. Is there any evidence suggesting that increases in trace metals and polychlorinated biphenyls (PCBs) in Mytilus edulis were associated with on-going open water disposal of dredged materials? 2. Is there any physiological change, e.g., tissue wet/dry weight ratios, gonadal development in M. edulis that could be attributable to the increase in tissue trace metal and PCBs concentrations? SNe Is there any discernible histopathological change that could be correlated with the increase in tissue trace metal and PCBs concentrations? Experimental Field Operations Monitoring at the WLIS disposal site was initiated in June 1984 with the deployment of three experimental platforms (WLISIN, WLISc and 500MW) and a master reference platform (RIr) (Fig. 4). Our original plan of using Latimers Light mussels as a source population stock had to be modified due to the lack of mussels of the required size class (>2.5 cm) sufficient to stock the four platforms. An older mussel population from Ram Island Reef, which generally exhibited similar trace metal concentrations as those found in the Latimers Light population, was found in sufficient numbers for our purpose. More than 6,000 mussels were collected from this location, sorted, counted, bagged and held briefly at the Marine Research Laboratory dock until they were deployed at the three experimental stations. The rationale of using one population of mussels as sentinel organisms for all disposal site stations has been discussed in our previous reports (Feng, 1984, 1985). The deployment of the three mussel monitoring platforms at western Long Island Sound was accomplished on June 27, 1984 at (1) the center of the disposal mound (WLISc), (2) 500 meters west of the mound center (500MW) and (3) a reference station located 2.22 kilometers south of the mound (WLISrN). On the following day, June 28, 1984, a platform was also deployed at Ram Island Reef (RIr) serving as a master reference station for the other three stations. Each platform was stocked with 1350 mussels in 27 mesh bags (50 mussels per bag). Details of the field operation are summarized in Table 2. Our standard field sampling procedure requires that ten replicate baseline samples be collected for trace metal, PCBs and wet/dry weight ratio determinations when the stations were first established. Whenever restocking of the platforms was deemed necessary due to predation, mortality and irretrievable losses of the platform, replicate baseline samples were always obtained. During each monthly sampling period, weather permitting, triplicate samples of eight mussels each were collected. Laboratory Procedures In the laboratory the mussels were cleaned, measured, examined for infection by the pea crab, Pinnotheres maculatus, then shucked and homogenized. An aliquot of the homogenized sample was weighed (wet weight) and lyophilized using a Virtis Model 10-010 freeze drier. After being dried overnight in the apparatus, the freeze dried tissue was weighed again, and designated as the "dry weight" used in calculating the wet/dry rato. In addition, ten mussels each from RIr, WLISrN, WLISc and 500MW were collected monthly and fixed with one valve cracked in neutral buffered formalin for histological studies. Trace Metal Analyses. The protocol for trace metal analyses used in this study was the same as that previously reported (Feng, 1984). Analyses of Polychlorinated Biphenyls. Aroclors 1242, 1254 and 1260 in Mytilus edulis tissues were extracted, concentrated, cleaned and gas chromatographed according to the procedures of Arimoto and Feng (1983a). In the past, quantification of each Aroclor was accomplished manually. During the current study, quantifications of Aroclors were achieved by employing an Apple II+ microcomputer equipped with two disk drives, a ADALAB data acquisition/control card, 128K RAM card and CHROMATOCHART software (Interactive Microware, Inc., State College, PA). After the injection of the standards, the software computes retention time and actual concentrations for each peak of Aroclors 1242, 1254 and 1260 as external standards and the results are stored in the memory. The concentration of PCBs of the injected unknown samples is computed using the stored results of the external standards. A comparison of the results obtained from the manual method with that of the present procedure has shown a deviation of no more than 5%. Gas _ Chromatograph/Mass Spectrophotometer Identification of Phthalate Esters. An anomalous peak present in the PCB chromatograms was identified by injecting aliquots of the sample into a HP 5890 gas chromatograph equipped with a HP 5870 MSD (Mass Selective Detector). The detected ion peak was matched 3 first with the 16 Phthalate ester standards. The identified sample Phthalate ester was further confirmed by matching with its known standard mass spectrum. Because the extraction procedure was designed for analyzing PCBs, the presence of Phthalate esters in the mussel extracts could only be considered as a qualitative study. Any quantitative determination of this trace organic compound would require the use of a specific extraction procedure. Histopathological Studies. To aid fast penetration of the neutral- buffered formalin into the soft tissues of the mussels, one valve of each animal was cracked with a sharp blow using the handle of a knife, and the shell liquor drained. The mussel was immersed in the fixative so that air was not trapped inside the mantle cavity. To standardize the section, cross sections of the mussels were cut just anterior to the foot. The sections were further processed and stained with hematoxylin and eosin using the standard histological procedure by the Histology Laboratory of the Department of Pathobiology, University of Connecticut, Storrs, Connecticut. Finished histological preparations were examined with an Olympus VANOX microscope at magnifications of 4X, 10X and 40x. Each specimen was critically scrutinized for stages of gonadal development, staining characteristics of the Leydig tissue, tissue integrity of the gill, kidney tubules, and intestinal epithelium, as well as the degree of leucocytic infiltrations. In addition, the prevalence of parasitic infections by trematodes, pea crabs, and sporozoans were also recorded. Statistical Analyses of the Data. Prior to conducting any statistical procedure, the data set of trace metal and PCB concentrations was tested for normality using the procedure of Shapiro and Wilk (1965). If necessary, data sets were normalized bys transtLormacvonsin( Logi: (ec) 0 elena (eS)] Or msec pyie Statistical analyses were performed using an IBM 3081 computer and the software for analysis of variance (ANOVA) outlined in SAS User’s Guide: Statisticsia(SAS. Enst.3 Ench,. 782). For the one-way ANOVA, the data were classified by station, while for the two-way ANOVA, the data were categorized by station (spatial) and sampling period (temporal), i.e. before and during disposal. In the present study, samples were collected before and during the disposal period; post disposal samples were not collected because sampling was not continued beyond June 1985. If the null hypothesis was rejected, Tukey’s multiple range test (Sokal and Rohlf, 1969) was applied to discern which set(s) was different. The frequency of the occurrence of a given parameter, e.g., gonadal development obtained from histopathological studies, was tested using the G-statistics (Sokal and Rohlf, 1969). This procedure tests the null hypothesis that the stages of gonad. development or any other parameter, are independent of stations 4 where the mussels have been maintained. The ratios of immature and mature individuals derived from histological examinations of the four populations were further analyzed by a replicated goodness of fit test (Sokal and Rohlf, 1969) in which the total G is partitioned to yield additional information. Results Trace Metal Concentrations. The mean tissue concentrations of nine trace metals found in the mussel populations maintained at Ram Island reference station (RIr), Western Long Island Sound reference station (WLISrN), Western Long Island Sound disposal site center (WLISc) and 500 meters west of the disposal site (SOOMW) from June 1984 to June 1985 are presented with the results of two-way ANOVA and Tukey’s test in Table 3. In addition, tissue trace metal data organized on a temporal basis and expressed in terms of both lg per g of wet and freeze-dried weight by station are included in the Appendix (Table la,b to 4a,b). Trace metal concentrations exhibiting statistically significant differences were revealed by the two-way ANOVA ( p < 0.05) (Table 3). The analyses indicate that the concentration of Cd, Cu and Zn show highly significant between-station differences. However, as revealed by the Tukey’s Test, such differences occur only between the mussel populations maintained at RIr and the other three stations which manifested a general uniformity in the three trace metal concentrations (Table 3A). Therefore, the observed significant between-station differences were attributable solely to the RIr population which exhibited the lowest concentration of most trace metals examined (Figs. 5, 6 and 7). Because the original mussel populations deployed at the WLISc, 500MW, and WLISrN were supplemented with LATr mussels in March 1985 and sampled concurrently with the remaining original mussels, trace metal data for the newly imported mussels obtained during April, May and June 1985 were grouped separately and subjected to one-way ANOVA. The results shown in Table 4 are remarkably similar to those presented in Table 3; the same three trace metals, Cd, Cu and Zn, showed significant between-station differences. The results, therefore, indicate that the disposal of dredged materials did not induce a significant localized increase in the trace metal concentrations at any of the experimental stations. Temporal differences (before-during disposal differences) in tissue cobalt, copper and iron concentrations were observed in mussel populations deployed at WLISc, 500MW, and WLISrN (Table Spe isles SB) yo In all cases before disposal, the three tissue trace metal concentrations were significantly lower than the during disposal trace metal concentrations. Unfortunately, the project was terminated in June 1985 before disposal was 5 completed. Therefore, no post-disposal samples were available. However, based on our previous experience, we could predict that the elevated levels of trace metals observed during the disposal period were generally of short duration. The elevated level of copper during disposal at WLISrN was the only inconsistency encountered in this analysis. Such an occurrence could be interpreted as being coincidental with the disposal period or attributable to other unknown environmental factors at the site; further analyses by stepwise multiple regression favor the latter interpretation. Stepwise multiple regression analyses (Table 5) show that the volume of dredged materials disposed was correlated with the concentrations of chromium and nickel at WLISc, of copper, iron and nickel at 500MW. In contrast, for the reference populations maintained at WLISrN and RIr, dredged volume as an independent variable was not entered into the regression models. The results are consistent with our previous studies that show the intrinsic variables, W/D and L, generally could account for the major proportion of variance observed in the tissue trace metals. There are two lines of evidence suggesting that dredged material disposal played a minor role in the uptake of trace metals by the mussels; these are (1) the dredged volume entered only ca. 28% of the cases and (2) it was the third or fourth variable entered into the model. The figure of 28% was derived from the assumption that if the elevated levels of the nine trace metals were associated with the dredged volume at WLISc and 500MW, one would expect that the dredged volume entered all 18 cases (2 stations x 9 trace metals) of the stepwise multiple regression analyses. In the present study, the dredged volume entered the regression model as an independent variable only on 5 occasions (5/18, or ca. 28%). Polychlorinated Biphenyl (PCBs) Concentrations. The mean concentrations of the Aroclors and the total PCBs from the four mussel monitoring populations are summarized in Table 6. Temporal variations in tissue Aroclors and PCBs are presented in the Appendix (Tables 5-9). The mean concentrations of Aroclor 1242, 1254 and 1260 fall in two distinct groups: the RIr and the three populations in western Long Island Sound. The latter has nearly twice the Aroclor concentrations of the RIr population. When the data sets were subjected to two-way ANOVA, the null hypothesis that there were neither spatial (station) nor temporal (before-during disposal) differences in Aroclor concentrations among the four mussel populations, was rejected. PCBs showed significant between-station differences (Table 6, For Aroclors 125441260: F=6.92, d.f.=3,35, P=0.0009; For Total PCBs: F=4.43, d.f.=3,35, P=0.0097), while the Aroclor 1242 and total PCBs exhibited temporal or before-during disposal differences in mussel populations held at WLISrN and WLISc (level of Significance see F values listed in Table 6B). Further analyses of the concentrations of Aroclors 1254+1260 and total PCBs using 6 Tukey’s test revealed that no between-station differences occurred among the three mussel populations (WLISc, 500MW and WLISrN) and that the Aroclors and PCB concentrations were Significantly less in the RIr population than in the other three. The concentrations of Aroclor 1242 and total PCBs were different before and during disposal in the mussel populations placed at WLISrN and WLISc; both compounds were significantly higher during disposal operations (p < 0.046 to 0.001). The findings of Significant temporal and spatial variations in Aroclors and total PCB concentrations are similar to that of the trace metal data (Cd, Cu, Zn, Fe) discussed in the previous section. Stepwise multiple regression analyses (Table 7) show that the volume of dredged materials was entered only into the disposal site stations (WLISc and 500MW). At WLISc, the volume of dredged materials disposed was the first variable entered into the regression model and accounted for 73.6% of the variance in Aroclor 1242. Sixty to 80% of the variance of other Aroclors and total PCBs could be explained by such intrinsic factors as W/D ratio and shell length. Similar observations were made at 500MW, i.e., 50-75% of the variance in Aroclors 1242, 1254, 1254+1260 and total PCBs were attributable to the intrinsic variables. However, the dredged volume did account for 9, 13, and 21% of the variance in total PCBs, Aroclor 1260 and Aroclors 1254+1260 respectively at this station. Again the analyses suggest that the disposal operation was not a major factor for the uptake of PCBs in the monitoring populations. This observation is consistent with changing levels of PCBs associated with dredging operations reported in San Francisco Bay (Anderlini et al., 1975), Puget Sound (Engler, 1979), Eastern Long Island Sound (Arimoto and Feng, 1983a), and Central and Western Long Island Sound (Feng, 1984, 1985). A significant compositional change in tissue Aroclors in favor of higher chlorinated Aroclors (1254+1260) were first reported in 1984 (Feng, 1984) when the mussels were transplanted from LATr to CLIS and WLIS disposal and reference sites. Such changes have again been evident in the present study (Table 8). The percent relative concentrations of Aroclors 1242 and 1254+1260 showed significant station-differences as revealed by one-way ANOVA (F=6.15, d.f.=3,39, p<0.0016). Further analyses of the data by Tukey’s test indicate that the RIr mussel population was Significantly different from the other three mussel populations in their relative tissue concentration of Aroclor 1242 and Aroclors 1254+1260 (Table 8). It is apparent that the 45:55 Aroclor 1242/Aroclor 1254+1260 ratio for the RIr population differed markedly from 33:67 for the other three populations. Bis (2-ethyl hexyl) Phthalate. During the analysis of tissue PCB concentrations, it was found that ca. 40% of the GC chromatograms (or 67 samples) showed an anomalous peak near the end of the run (Fig. 8). Contamination of the samples has been 7 ruled out by incorporating procedural blanks which showed no such anomalous peak. Gas Chromatography/Mass Spectrometry analysis of the samples was then carried out using a Hewlett-Packard 5890 GC and 5870 SMD system equipped with a 12 m cross linked methyl silicone (0.33 1) fused silica capillary column. The total ion concentration pattern showed a single peak with the retention time of 25.92 minutes, which matches with the thirteenth peak of Bis (2-ethyl hexyl) Phthalate (BEHP) standard (Fig. 9A, B). The confirmation of this trace organic compound is shown in the mass spectrum (Fig. 9C, D). Bis (2-ethyl hexyl) Phthalate is the most common plasticizer in use by industry. According to Giam (1976), the national annual production of BEHP was 200 million kilograms in the 1970’s or six times the amount of the known pollutants, the PCBs. The solubility of BEHP in water is 0.04-0.40 ppm (Giam et al., 1984). Di-n butyl Phthalate (DNBP), a related Phthalate ester which has a solubility of 10-13 ppm in water (Giam et al., 1984) was found nearly 100% associated with particulates in Thames River water samples according to Dr. A. Libbey, Associate Professor of Chemistry, Department of Chemistry, University of Connecticut. BEHP, which is much less soluble in water than DNBP, is also expected to show a close affinity with particulates in environmental samples. Possession of such a property could be useful in serving as a particulate-borne or particulate- transported pollutant tracer. Frequency of occurrences of BEHP in Mytilus edulis maintained in both eastern (ELIS) and western Long Island Sound throughout the year is presented in Table 9. The highest frequency at ELIS was during July, August and September, while at WLIS it was during October, November and December (p < 0.05). The reason for the discrepancy is not known and would have required further studies outside the scope of this investigation. Mortalities. Prior to examining the prepared tissue sections for evidence of histopathologic changes, an attempt was made to obtain a first approximation of the possible adverse effects on the mussels by analyzing the cumulative mortalities. Figure 10 shows the differences in cumulative mortalities between the populations at RIr and in western Long Island Sound. The populations differed in the times required to reach 50% cumulative mortalities, i.e., 2, 2.5 and 3 months for the WLISc and 500MW, WLISrN, and RIr populations, respectively. The maximum cumulative mortalities: 50% for RIr and 70-90% for WLISrN, WLISc and 500MW were attained in September, three months after the deployment of mussels. WLISc population appears to be the only exception, and shows a steady increase of the cumulative mortalities from 65 to 90% during the period of January through March when disposal of dredged materials was taking place at this site. This observation suggests that the apparent disposal- associated mortality is limited to the immediate environment of the disposal site. The RXC contingency table of dead and live mussels observed in the four populations throughout the year is 8 used to test the hypothesis of independence of mortalities from the stations (Table 10); the G-test statistics obtained: G=234.2, v2 0.005(4) = 14.860, p << 0.001 reject the hypothesis. Therefore, the mortalities were associated with the stations where the mussel populations were deployed. At the reference station RIr, the 45% mortality was significantly lower than that of the LATr (52%) and the three populations, WLISc, 500MW and WLISrN (71-73%). In investigating the effects of stock and location on growth and mortality in Mytilus edulis in Nova Scotia waters, Dickie et al. (1984) reported that location was the major factor -in determining growth, while stock influenced mortality. Furthermore, the biomass and potential yield were determined approximately equally by location and stock. In the present study, because only one stock was used, the genetic effect was, therefore, removed. Thus, the observed differences in mortality (Table 10 and Fig. 10) and W/D ratio (potential yield) (Fig. 5) were site-specific. Histopathological Studies. Histological sections from the reference and disposal site populations were examined by using seven parameters: (1) the stage of gonadal development, (2) staining and morphological characteristics of Leydig tissue, (3) integrity of the intestinal epithelium, and intestinal content, (4) integrity of the style sac epithelium and the presence of crystalline style, (5) changes in plycate organ, (6) degree of leucocytic infiltrations, as well as (7) prevalence of parasitism. The first two parameters are indices of the mussel’s reproductive status, while parameters 3, 4, and 5 are indices of feeding and excretion status. The degree of tleucocytic infiltrations, except during the post spawning resorption of remanent ova, is an indicator of inflammation which could identify overt histopathology. The last parameter is to assess any tissue damage inflicted upon the host (mussels) by parasites; this information is important in separating environmentally induced histopathological manifestations from those caused by parasitisn. Whenever possible, the parameters were scored for quantitative presentation and subject to proper statistical treatment. For scoring the degree of reproductive conditions, the gonads were categorized in five classes: castration, early development, immature stage and spent. These stages are defined as follows: a. Castration (C): destruction or replacement of the gonadal tissue by sporocysts of Proctoeces maculatus; sex usually undeterminable. b. Early development stage (E): only germinal tissue present; sex undifferentiated. Cc. Immature stage (I): clearly recognizable germinal follicles are present; male follicles with a thick layer of spermatids around the periphery and a few spermatozoa in the center; female follicles relatively small with peripheral oocytes and occupying rather limited areas of the mantle tissue. ad. Mature stage (M): Male follicles with or without a thin layer of spermatids but filled with mature spermatozoa; female follicles packed with mature ova; both male and female follicles having displaced most of the Leydig tissue in the mantle; gametes in gonoducts. e. Spent (S): empty or near empty follicles with extensive leucocytic infiltration and phagocytosis. To facilitate statistical analyses of the data, the four developmental stages: E, I, M and S were pooled into two categories: the immature stage (I) and the mature stage (M). The immature stage consisted of individuals classified as in the early development and in the immature stage; the mature stage combined individuals exhibited mature ova and/or contained remnant ova and spent germinal follicles. The frequencies of the two condensed developmental stages: iy vandi ey Me ling ithe sour populations are presented in Table 11. The data were subjected to a replicated goodness of fit test (G-test) (Sokal and Rolf, 1969). In the analysis, the ratios of immature and mature individuals at WLISrN, WLISc and 500MW were treated as "replicate" samples which were tested against the ratio of immature and mature (8:73) mussels found at RIr. The analyses show that Gp, Gp and Gy are all significant. The three populations of WLISC, 500MW and WLISrN have an excess of immature individuals; the proportions of immature or mature individuals appeared to have been sampled from different populations, in spite of the fact that all three experimental populations were originated from RIr. The 500MW population has a nonsignificant excess (G=2.958, p>0.05) and the WLISrN and WLISc populations deviate significantly from the expected 8:73 ratio (G=5.968, p<0.025 and G=19.386, p<0.005, respectively). Hence, Gp is highly significant. The Gp is also highly significant, because the consistent trend favors immature mussels in the three experimental populations located in the western Sound. The significant heterogeneity G, indicates that the magnitude of favoring immature animals is not uniform in all cases. Based on analyses of the partitioned G values, the ratios of immature to mature mussels in the four populations are not homogeneous; it can also be seen that the heterogeneity of the ratios is contributed by two populations: WLISrN and WLISc, which is verified by the simultaneous test procedure (STP). These analyses suggest that the proportions of immature mussels at WLISrN (19%) and WLISc (27%) are higher than the expected proportion at RIr (10%) and are associated with the locations. 10 In the course of conducting histopathological studies, we noticed that the staining characteristic of the Leydig tissue, the site of glycogen storage, ranged from light to intense red. On close inspection, it was revealed that the intense red staining was due to the presence of many large amoeboid cells with eosinophilic cytoplasm in the region. These cells are known as adipogranular cells which play an important role in the glycogen metabolism and gametogenesis (Lowe et al., 1982). The results summarized in Table 12 suggest that the four populations are fairly homogenous (Gy=4.922, d.f.=3, P=ns). The trend is in favor of the lightly stained Leydig tissues indicating reduced adipogranular cell activities and in general agreement with high percentages of mussels being in the mature stage (Table 11). The only significant difference is found in WLISrN which shows the lowest proportion of lightly stained Leydig tissues (59%) as contrasted with that of the other three populations (69-77%) (Table 12, Gp=4.206, d.f.=1, p<0.05). Based on these analyses, it would appear that glycogen synthesis in these mussels was probably not adversely affected. As far as feeding was concerned, no discernible or at best marginal significant differences were found in the intestinal content of the four populations (Table 13, G=2.966, d.f.=3, P>0.05). However, an examination of the presence or absence of the crystalline style within the style sac as indicators of feeding activities revealed that significantly fewer mussels had styles at 500MW and WLISc than those maintained at RIr and WLISrN (Table 14, 500MW: G=7.456, d.f.=1, p<0O.01; WLISc: G=14.073, ad.f.=1, p<0O.001). Apparently, the presence of crystalline style is a more sensitive indicator of feeding activities than the presence of food in the lumen of intestine. In the RIr and WLISrN mussels, significantly higher percentages of the plycate organ showed enlargement of the lumen as contrasted with those deployed at WLISc and 500MW (Table 15, WLISc: G=9.475, d.f.=1, p<0.005; 5S0OO0OMW: G=7.741, d.f.=1, p<0.010). If one accepts the assumption that the enlarged lumen of plycate organ represents the normal functioning of the organ, then the non-enlarged lumen of plycate organ suggests functional atrophy of the organ. Occasionally, Proctoeces-induced enlargement of the lumens of the plycate organ was encountered. However, there were no significant differences in the prevalence of P. maculatus infection in the four mussel populations. The apparent dysfunction of the plycate organ could be associated with disposal. A survey of the histological slides for parasitic infections yielded the following information: 11 1) the prevalence of Proctoeces maculatus in the four mussel monitoring populations was not significantly different (Table 16, G=1.967, d.f.=6, p = 0.975), 2)) the prevalence of Chytridiopsis mytilovum in the WLISc population was significantly higher than the other three populations (Table 17, Gy=3.837, d.f.=3, p<0.05; WLISc: G=5.217, d.f.=1, p<0.025), and 3) the prevalence of Pinnotheres maculatus differed significantly between the RIr population and the ones deployed at WLISrN, WLISC and S500MW (Table i8, Gy=11.170, d.f.=3, p<0.025). There was an overall 12% reduction of P. maculatus infection in the transplanted populations as compared to the RIr population. Based upon the study of 310 slides, no _ significant differences in leukocytic infiltration were noted in all four mussel populations (Table 19, G=8.389, d.f.=9, p=0.50). Discussion Trace Metal and PCB Concentrations. In the present investigation, it was found that a suite of three trace metals, Cd, Cu, and Zn, showed no discernible difference among the three mussel populations, WLISc, 500MW and WLISrN. These observations support the interpretation that any changes in the trace metal concentrations of these elements in the monitoring populations could not be attibuted to dredged material disposal. 1S ss significant to note that similar inferences were made in the experimental populations during 1983 and 1984 disposal operations in central and western Long Island Sound, respectively (Feng, 1984, 1985). This indicates that much more pervasive factors than the episodic disposal of dredged materials are operating in Long Island Sound. Factors contributing to the general uniformity of trace metal concentration in the mussel populations held at reference and experimental stations in both central and western Long Island Sound could be the result of physical, chemical and biological processes and/or of disposal management processes. The capping procedure implemented during the 1983 operation at CLIS could effectively reduce the release of toxic metals into the water column, thus reducing the available trace metals to the mussels and consequently mitigating the environmental impact of disposing large quantities of relatively contaminated dredged material, e.g., the Black Rock Harbor material. A second factor is the amount of contaminant input by natural process which could be orders of magnitude larger than the amount of dredged materials 12 disposed in the Sound. Moreover, it is assumed that releasing of trace metals and other constituents from the sediment due to bioturbation (sediment reworking by infauna) over a vast area of the seafloor in the Sound is a continuous process. The nepheloid layer which prevails at the sediment-water interface throughout much of the area of the Sound with silt-clay bottom (including the disposal sites) is presumably the result of bioturbational activities of the resident infauna. It is believed that bioturbation of shallow sea sediment is possibly the most important factor in vertical transport of contaminants to the water column, while physical factors such as storm events, circulation patterns, residence time of water masses, etc., play important roles in large scale mixing of contaminants in the Sound. It is reasonable to assume that vertical and horizontal mixing of water masses in western Long Island Sound (the MNarrowest part of the estuary) are maintained most of the time except during the warm months when large areas of hypoxia exist due to temperature stratification of the water column (Rhoads, 1987; Welsh, personal communication). Bohlen (1980) reports that dredging-induced sediment resuspension is generally small in comparison to the transport resulting from natural storm events. Additionally, sediment resuspension produced by commercial FIShingeeactivicles sacouldisalsoumbew. a prLacctonmicontr bucingme to particulate-bound contaminant transport; studies in the Gulf of Mexico have shown that resuspension due to shrimp trawling activities could be 10-100 times greater than that generated by maintenance dredging of shipping channels in a year (Schubel et auleenes 957/9)) . ; In recent years, lobster and scup fisheries have become major commercial activities in Long Island Sound; these activities may play a role in sediment resuspension with the subsequent release and transport of contaminants including trace metals and particulate-associated PCBs. Thus, sediment resuspension generated from bioturbation, mixing due to tidal currents, storms and anthropogenic activity could serve as a driving force that renders the environment more uniform, which in turn is reflected in the homogeneous concentration of trace metals and PCBs found in the three experimental mussel populations located at WLISc, 500MW and WLISrN. In addition, the lack of distinction between WLISrN and the two stations nearest to the disposal site suggests that environmental conditions in the general western Sound are unfavorable as compared to the reference station in the eastern Sound. The results of this portion of the study support the hypothesis that the intrinsic and extrinsic factors played more of a role in changes in tissue trace metal levels than did disposal activities. Temporal and spatial variations in PCB levels behave in the same manner as the tissue trace metal concentrations. The lack 13 of demonstrable differences in the tissue trace metal and PCB concentrations among the three populations located in the western Sound both on (WLISc and 500MW) and off (WLISrN) the disposal site lends further support to the argument presented above. The finding of compositional change of Aroclors in favor of Aroclors 1254+1260 in the disposal site populations reflects the high concentration of suspended matter in this region; it is known that more highly-chlorinated biphenyls (biphenyls with more chlorine radicals) such as Aroclors 1254 and 1260 are closely associated with particulates and that lesser chlorinated isomers such as- Aroclor 1242 are water soluble (Duinker et al., 1982a,b). One could, therefore, predict that invertebrates which are deposit and suspension feeders would contain more highly chlorinated biphenyls. Such a prediction was verified in Mytilus edulis (Feng, 1984; the present study) and in the benthic invertebrates Macoma balthica, Arenicola marina and Crangon crangon from the Dutch Wadden Sea (Duinker et al., 1983). Thus, in order to resolve the question whether the Aroclor compositional change in the experimental mussel populations was due to environmental availability of the Aroclors or to biological processes (e.g., selective uptake or depuration of Aroclors), data on the composition of Aroclors in both water (dissolved and particulate-associated Aroclors) and sediment from both the reference and disposal site would be required. Biological Effects. Mortality is a common phenomenon in all living organisms which can be ascribed to aging, diseases, and the effects of environmental changes. The key issue is how natural mortalities can be separated from those induced by the effect of pollutants. In the present study, the heightened cumulative mortalities found in the three mussel populations located in the western Sound are probably the result of abnormal reactions superimposed on the natural seasonal mortality (the latter exemplified by the reference population at RIr). When the cumulative mortality curves depicted in Figure 10 are scrutinized carefully, one notices that the onset of heightened mortalities at WLISc and 500MW occurred during July and August; for the populations held at WLISrN and RIr, the onset was delayed and took place during August and September. Even though the magnitude of the mortalities of the two populations was different, the initial elevated mortalities found at the stations in the western Sound could not have been associated with disposal activities, because there was little or no disposal of dredged material during the summer months at the WLIS disposal site. However, there is a noted increase in cumulative mortalities at the WLISc station from January to March of 1985. This correlates with the time of renewed disposal activity at the WLIS disposal site; because this is the only station to show a marked increase in mortality, it can be surmised that the effect of disposal on mortality, if any, is limited to the immediate disposal area. The high cumulative mortalities found at WLISc, 500MW, and WLISrN in part, are probably related to other environmental conditions, 14 e.g., the hypoxia of bottom waters prevalent in the region during the summer. Gonadal development of Mytilus edulis under normal conditions is the result of interactions of environmental factors, especially temperature, salinity, light and food, as well as endogenous factors such as the influence of parasitism. In the present study, the reproductive activity may also have been affected in mussels living on and near dredged material disposal sites by the changing physical and chemical conditions of the environments. The relatively high proportion of immature gonads observed at WLISrN (19%) and WLISc (27%) (Table 11) suggest that the reproductive development of the mussels is location-specific and reflect differences in the depth at which the mussels were held, availability of food, disposal, and other associated local environmental parameters. Bayne et al. (1978) demonstrated that when mussels were experimentally exposed to temperature and food ration stresses, they produced smaller and fewer eggs than unstressed controls. Also the ripe gametes of the stressed animals occupied a smaller proportion of the mantle tissue than that of the controls. In this study, temperature probably did not play an important role in determining the gonadal development. According to Reid et al. (1979), the bottom temperatures of WLIS and Fishers Island Sound in September 1972 were 18° - 20° C and 16° = 18° C, respectively. During the winter, the bottom temperatures were rather uniform throughout Long Island Sound and varied within a narrow limit of 4° - 6° C. The reduced proportion of mature mussels at WLISrN and WLISc implies a concomitant reduction of areas occupied by female germinal follicles in the mantle. This view is generally supported by qualitative observations of the histological slides which show diminution of female germinal follicles. Quantitative studies conducted by Arimoto and Feng (1983b) revealed that the New Haven disposal site mussel population had significantly smaller ova (1880 + 140 2) than the New Haven reference mussel population (1520 + 140 u2) (p<0.01). The reduction of areas occupied by the gametic follicles or of ovum size could also be induced by the food ration stress and the infection caused by Proctoeces maculatus. While it is difficult to conduct food rationing experiments in the field, it is possible to determine whether the mussels were feeding and carrying out normal extracellular digestion by examining the presence of crystalline styles and intestinal content in the histological slides. There were no discernible or at best Marginal significant differences among the four mussel populations as far as feeding was concerned (Table 13, G=2.966, p>o.5). Moreover, at WLISc and 500MW where the depth is greater than 30 meters, diatom tests were readily identifiable in the 15 intestinal contents, indicating that feeding was not inhibited, and food was not limited. However, even though feeding was taking place, this did not necessarily mean that the ingested food was assimilated. The generally poor condition and significantly lower frequency of crystalline styles in the 500Mw and WLISc mussel populations appeared to support this assertion (Table 14). Examinations of the effect of parasitism on gonadal development revealed that in light infections, the development of gametic- follicles was inhibited, while in severe cases, the mussels were totally castrated. Such parasite-induced castration was observed in only 34 of the 409 slides examined, constituting approximately 8%; moreover, 35% of the observed castrations were seen in the RIr population. Also, the infection among the four populations was not significantly different (Table 16). Thus, parasitism probably exerted only a limited stress on the mussels at the three experimental stations, WLISc, 500MW, and WLISrN. In addition to the infection caused by Proctoeces maculatus, Chytridiopsis mytilovum infection in the transplanted populations was significantly higher (65-80%) than that of the RIr population (52%) (Gy=3.837, d.£.=3, p<0.05). Because C. mytilovum invades ova of M. edulis, this observation provides additional support for the assertion that gonadal development in the WLISrN and WLISc populations was retarded; this was reflected in the higher proportion of immature individuals at these locations. The prevalence of Pinnotheres maculatus infection in the transplanted populations was 15% less than that of the RIr population. Once a female crab enters a mussel host, it is generally "imprisoned" for life due to its large size, which prevents its escape. The reduction in occurrence of this infection could only be explained by the death of the female crabs in situ. This finding suggests that P. maculatus is more susceptible or sensitive to the changing environment at WLIS than the other two parasites. However, the reduction of percent infections could also be interpreted as a lack of new parasite invasions into the mussel host. Leucocytic infiltration, a well known indicator of tissue inflammation which has been used by mammalian pathologists in identifying foci of abnormal tissues was of limited use in the present study. Unlike that in mammalian hosts, leucocytic infiltration in marine bivalve mollusks is associated with the normal resorption of unspawned gametes. It is, therefore, a part of the normal function of the molluscan leucocytes. Furthermore, they also play an important role in nutrition and defense (Feng ee. Fadel Oi7u7))ns In the present study, some of the heightened leucocytic infiltration could be attributed to the presence of Proctoeces maculatus and to the post spawning resorption of 16 remanent gametes, but none appeared to be associated with disposal operations. The histopathological studies indicated that the retardation of gonadal development, the absence of crystalline style and the dysfunction of plycate organ could be associated with mussels at all three populations transplanted in the western Sound. However, these results should be considered only as presumptive and interpreted with caution for the following reasons: 1). Field experimental results are by nature correlational, therefore, no causation can be assumed. 2) The results could also be correlated with other unknown or uninvestigated factors, and 3) The results of histopathological studies presented herein were derived from mussels which survived mortalities of undetermined causes; hence, they represented the "fittest", particularly at the three transplanted stations (WLISc, 500MW and WLISrN) where the average cumulative mortality was more than 70%. The results could be biased in favor of lesser spatial association with the observed abnormalities. ies2 etal the dead and dying mussels could have been recovered by more frequent sampling during July, August and September with subsequent examination of their tissues, more definitive conclusions might be reached. The chances of recovering dead and dying mussels would be improved in future projects by augmenting the initial deployment with a monthly introduction of mussels at each station, followed by sampling these introduced mussels at monthly intervals concurrent with the sampling schedule of the mussels deployed initially. This approach would achieve two objectives: the monthly deployment should reveal more clearly the relationship between uptake rates of trace metals and PCBs and associated biological effects (mortalities, histopathology, W/D ratios, etc.) and allow recovery of more morbid mussels, while the initial deployment should represent cumulative effects. Moreover, the indications suggested in this study should be followed by vigorously controlled laboratory experiments in an attempt to determine causations of the observed abnormalities. Conclusions This investigation suggested that the disposal of dredged material has only a limited influence on the trace metal and PCB concentrations of mussel populations deployed on or near the disposal site in western Long Island Sound. Similar tissue concentrations of Cd, Cu, and Zn were found in all three 17 transplanted populations in the western Sound (both on and off the disposal site), suggesting that disposal activities had no effect on tissue concentrations of these elements. Significant temporal differences were found in tissue Co, Cu, and Fe before and during disposal; unfortunately, no post-disposal samples were available. However, similar results were found during earlier transplanted mussel investigations for disposal site monitoring at central and western Long Island Sound; very limited and transient changes could be associated with disposal activities (i.e., metal levels elevated during disposal quickly returned to background levels in post-disposal samples). Therefore, one could reasonably predict that the elevated levels of Co, Cu, and Fe were of short duration and would return to background following cessation of disposal activities. Variations in concentrations of tissue PCBs (both Aroclors and total PCBs) exhibited a pattern similar to the trace metal concentrations: no difference was found among the three populations in the western Sound, while a significant difference existed between the populations in the western Sound compared to the eastern Sound. This again supports the conclusion that the disposal operation was not a major factor for the uptake of PCBs in the experimental populations. Significant temporal differences in PCB uptake were also demonstrated, but because of the lack of post-disposal samples, it could not shown that this was a long-term change; however based on prior experience, one could predict that these elevated levels were of a transient nature. Concurrent with the previous disposal site mussel investigations carried out in Long Island Sound, some of the measured variables of physiological change (tissue wet/dry weight ratios and overall length) generally could account for the major proportion of variance observed in the tissue trace metal and PCB concentration. Histopathological studies revealed that the only effects which could be associated with disposal activities were a fewer number of mussels with crystalline styles, a higher incidence of parasitic infection by Chytridiopsis mytilovum, and a higher incidence of mussels with possible functional atrophy of the plycate organ. Otherwise, there was no effect of dredged material disposal on mussel gametogenesis, glycogen synthesis, gut content (a measure of feeding), parasitic infections by Pinnotheres maculatus, or destruction of the gonadal tissue by sporocysts of Proctoeces maculatus. The general lack of distinctions among the three transplanted mussel populations has been attributed to the uniform environmental condition in the western Sound mediated by complex physical and biological processes (e.g., water temperature, bioturbation, tidal currents, storms, commercial fishing activities). These considerations, coupled with the correlational nature of field data, suggest that causative effects cannot be assumed. Therefore, the observed biological effects from the histopathological studies can only be 18 considered as presumptive, because organisms respond to a variety of environmental factors in the field. The underlying objective of the DAMOS mussel watch program is to detect any potential for far-field impacts of dredged material disposal; this objective is accomplished by monitoring tissue uptake of selected trace metal and hydrocarbon compounds as an indicator of off-site transport of particulate-associated contaminants. To date, all mussel monitoring studies associated with dredged material disposal at the containment sites in Long Island Sound have shown the effects of disposal on trace metal and hydrocarbon tissue uptake are very limited spatially and of short duration (i.e., associated with the immediate disposal event and then a return to background levels). These results support the interpretation that sites in Long Island Sound are being effectively managed as containment sites, and any far-field impacts of dredged material disposal are non-existent or undetectable. 19 References Cited Anderlini, V.C., J.W. Chapman, A.S. Newton, R.W. Risebrough, B.E. Cole, B.W. deLappe, D.C. Girvin, A.T. Hodgson, S.J. McCormick, L. Nelbach, R.K. Okazaki, M.H. Panietz and T.T. Schmidt. 1975. Pollutant availability study, dredge disposal study, San Francisco Bay and Estuary, Appendix I: Pollutant Availability. U.S. Army Engineer District, San Francisco, 88 pp. + appendix. Arimoto, R. and S.Y. Feng. 1983a. Chapter 10. Changes in the levels of PCBs in Mytilus edulis associated with dredged- material disposal. pp. 199-212. In: Wastes in the Ocean. Vol. II, Ed. D.R. Kester, B.H. Ketchum, I.W. Duedall and P. Kilho Park. John Wiley & Sons, Inc., New York. Arimoto, R. and S.¥. Feng. 1983b. Histological studies on mussels from dredge spoil dumpsites. Estuarine, Coastal & Shelf Sci. 17, 535-546. Bayne, B.L., D.H. Holland, M.N. Moore, D.M. Lowe and J. Widdows. 1978. Further studies on the effects of stress in the adult on the eggs of Mytilus edulis. J. Mar. Biol. Assoc. U.K. 58, 825-841. Bohlen, W.F. 1980. Chapter 103. A comparison between dredge induced sediment resuspension and that produced by natural storm events. pp. 1700-1706. In: Sediment Resuspension Comparison Coastal Engineering 80. Dickie, L.M., P.R. Boudreau and K.R. Freeman. 1984. Influences of stock and site on growth and mortality in the blue mussel (Mytilus edulis). Can. J. Fish. Aquat. Sci., 41, 134-140. Duinker,...J.C., MoDods. Hllebrand),) Rak. GeNOLting: andiaes- Wellershaus. 1982a. The river Weser: processes affecting the behaviour of metals and organochlorines during estuarine mixing. Neth. J. Sea Res. 15, 141-169. Duinker wed GC.) seMal.di ne Hilshebrand, sekiihe sNwoltcancim aancumsr Wellershaus. 1982b. The river Elbe: processes affecting the behaviour of metals and organochlorines during estuarine mixing. Neth. J. Sea Res. 15, 170-195. Duinker 4 )idisC. 4 MaTed. wHilebrand” iand@mueP. arBoon 1983. Organochlorines in benthic invertebrates and sediments from the Dutch Wadden Sea; identification of individual PCB components. Neth. J. Sea Res. 17(1), 19-38. 20 Engler, R.M. 1979. Bioaccumulation of toxic substances from contaminated sediment by fish and benthic organisms. pp. B25 3/54 In: Management of Bottom Sediments Containing Toxic Substances, Proceedings of the fourth U.S.-Japan Experts! Meeting. October 1978, Tokyo, Japan. EiNCip Solo, SoSo weiNe; Evelel Go Webel 1977. Roles of Mytilus coruscus and Crassostrea gigas blood cells in defense and nutrition. pp. 31-67. In: Comparative Pathobiology, Vol. Big Safle (No LENA, hiss Eval Wao OnSnejo jlapmrn wewlsil, Co, , New York. Feng, S.Y. 1984. DAMOS-Mussel Watch. Central Long Island Sound Disposal Site and Western Long Island Sound Disposal Site Monitoring Project 1983 (February 1 - September 30, 1983). Submitted to Science Applications Inc., Newport, RI. 78 pp. Feng, S.-Y. 1985. DAMOS-Mussel Watch. Central Long Island Sound Disposal Site and Western Long Island Sound Disposal Site Monitoring Project 1983 (October 1, 1983 - September 30, 1984). Submitted to Science Applications International Corp., Newport, RI. 87 pp. Giam, C.S. 1976. Chapter 4. Trace analyses of phthalate (and chlorinated hydrocarbons) in marine samples. 0 Gilo77S. In: Strategies for Marine Pollution Monitoring. Ed. E.D. Goldberg. John Wiley and Sons, New York. Giam, C.S., E. Atlas, M.A. Powers, Jr. and J.E. Leonard. 1984. Phthalic acid esters. Pp. 67 —1l4)27° ers Handbook of Environmental Chemistry Vol. 3C Anthropogenetic Compounds. Ed. Otto Hutzinger. Springer-Verlag, New York. 220 pp. Lowe, D.M., M.N. Moore and B.L. Bayne. 1982. Aspects of gametogenesis in the marine mussel Mytilus edulis L. Sie Mar. Biol. Ass. U.K. 62, 133-145. Reid, R.N., A.B. Frame and A.F. Draxler. 1979. Environmental baselines in Long Island Sound, 1972-1973. NOAA Technical Rept. NMFS SSRF-738. U.S. Dept. of Commerce. 31 pp. Rhoads, D.C. 1987. REMOTS reconnaissance mapping of near-bottom dissolved oxygen: Central to Western Long Island Sound, August 1986. SAIC Technical Report #SAIC-87/7502&132; submitted to U.S. EPA Region I. 56 pp. SAS User's Guide: Statistics 1982 Edition. SAS Institute Inc., Box 8000, Cary, NC 27511. 584 pp. Schubel, J.R., H.H. Carter and W.W. Wise. 1979. Shrimping as a source of suspended sediment in Corpus Christi Bay (Texas). Estuaries 2, 201-203. 21 Shapiro, S.S. and M.B. Wilk. 1965. An analysis of variance test for normality (complete samples). Biometrika 52, 541-611. Sokal, R.R. and F.J. Rohlf. 1969. Biometry. W.H. Freeman and Co., San Francisco. 776 pp. 22 0° 001 p° 66 0°08 8°0p eet el 0°2 0°2 0°2 0°2 0°2 0°2 0°2 “[OA aAtyeinun) % Spo 86 560° 96 OSB'BIE O25‘°291 S08‘ 25 s99°82 000'8 000°8 000'R on0'R N00'R 00°8 QAN0'A “[OA dat yeynwny 0st *2 Spell OLE*9ST SIZ°60I Ovl‘b2 99°02 ae 20 se 25 =e a2 000°8 aun OA A, yWUOW une Kew jdy Jey qa4 ue 330 AON 120 das Bny a¢ une 37eQ S8 68 S86 2UNL 03 HREI auNr wosj 2714S {eSOdstp punos pues; buoy Usaq{Say ayy ye paz!sodap (Sp4ek I1qQNd UL) S{elsazew pabpaap Ayyyuow “tT atqey 23 “d{dues INC ayy 2q 0) Prsapysuod pue SB “[{ INC UO paydues tayep Syyy ye pauyeygo Sa{dwes Ou - UO4}IUNYL PY )-NYBO) cvs "YUO})IUNj| ee JuswdiNbs Buyayp yO aiNedIq pauye1Qo sa[ Gees OW oo “Sdjdues Gp ‘uel ayy aq 0) PasapysuCr pue py “BZ °220 vO patduws » “$194 (jU0D BuytNpayrs 0) anp arqeijere you S{assar Spauyeyqo yOu san sajdues pAb] 4290130 :210N SS ee p86 vet P2yd0,say dwes pai dues Praydwes 2867 320 22u48 0°¢L6Cb a | @ues 2Rr0} 89s buy yd038a0 pa| dues pajdwes pai dwes pai dues 02| i oa ee put Joy SL assne Ai yiuow pajdues €°C9092 pd | Gues 9334 (09 SN ONU | juo) aw) ee —— OO 9d | Oues payroysau pai dwes ory pajdwes Pajdwes pa | dwes Pay chues Dd) dues pakoydap 8'°S/6C9 pue W40jIPId 0°Sf B92 pa| Gwe s : a ee fesodsip oy 02) Gurs 0dNI0}SI5 40424 AHOOS 91 oPaj dwes pajdwes pojdwes prj dwes Pal dues pai dwes pakoydap 2°S(6Cy pue Paacw ws0j3040 WsO Field 6° 1C892 pd|Guas Pa) Gwe s 3514 SC Ooo SSSSSSSSSSsee 000Pd| Games Pdyd018d4 Pai dees opay dues pri dues pajdwes 9d dwes prj dues paduas pako\ dap L960 Og ws0j0ld C*v2ngee 03| Gus W4S 1A —_- ——_————— eee SSSSSSSSSSSSSSSSSSsSSSSsSsssS pd | Ones Pai Gues Pay dwes pa dwes pai dues Pai dues pai dwes pajdues paujeygo 8° 1660p Sai dues aus b° v8092 -aseg 9 prakoyd ayy ap wioyjeyd (2-92 sew St 984 Ct 4 Ut 929 C1 9 Ut 320 6 ¥ 2 AON 61-81 was 22-2 Say 92-52 Aine u2-¢2 aune $ayeujps00) 5861 3-wv¥0) vB6t voyrers “GB6l Punt Oy yB6E aunf wory Suoyjesadg Pyayy jo Abo; ouGI4) swe s6O1g YJOM GIssny °2 ayqey 24 Table 3. Summary of mean wet/dry weight ratios (W/D), shell length (1) and tissue trace metal concentrations (ppm + 1 S.D.) in Mytilus edulis deployed at Ram Island Reef (RIr), Western Long Island Sound reference (WLISrN), Mound Center (WLISc) and 500 meter west of the mound (500MW) from June 1984 to June 1985. All WLIS stations were restocked with Latimers Light mussels in March 1985 and sampled concurrently with the original RIr mussels. Stations RIr WLISrN WLISc 500MW W/D 7.48(1.54) 8.52(2.45) 8.61(2.47) 8.67(2.84) L 7.06(0. 22) 6.48(0.75) 6.57(0.75) 6.49(0.83) Cd* 0.94(0. 48) 1.47(0.74) 1.43(0.68) 1.37(0.72) Cr 2.90(1.61 4.34(5.76) 3.48 (2. 32) Bo AMCs) Co* 0.83(0.84) 0.91(0.91) 0.87(0.94) 0.80(0.%) Guz 7.68(2.06) 10.96 (3. 32) DW GIs Lk) NEO CB G22 970) ex 232(81) 263(118) 311(165) 277(130) Hg 0.198(0.053) 0.180(0.075) 0.200(0.065) 0.178(0.059) Ni ESI Is SS) OSS ZS )) 3.42(1.53) 3.82(1.86) Zn* 124(37) 154(43) 159(41) 137(35) V ae SG aesis))) 1.62(0.89) ‘1.51(0.68) 1.62(0.88) n 11 1l ll 10 “Two-Way ANOVA Significant using trace metal concentrations weighted with W/D ratios. 25 Table 3, Continued Tukey Groupings ( a = 0.05): A. Between-Station Differences RIr 500MW WLISC WLISrN Cd 0.94 es Bz 1.44 1.47 RIr WLISrN WLISc 500MW Cu 7.68 10.96 WIS 27/ 11.28 RIr 500MW WLISc WLISrN Zn 124 137 151 154 B. Before-During Disposal Differences Trace Metal. Jo | SStatvonl SOUriIngiimalBet One nim ntl Gs ii ilatl Alina Co WLISc 1.29 0.56 555 0.050 cu WLISPN 12.79 9.44 en25 0.034 WLISc 12.73 10.04 9.48 0.013 SOOMM 134th 0 Moles 25.20 0.001 ee WLISc 381 253 12.00 0.007 500MM 363 191 aie 0.003 26 Table 4. Summary of mean wet/dry weight ratios {W/D), shell lenath (L) and tissue trace metal concentrations (ppm+ 1 S.0.) in Mytilus edulis deployed at Latimers Light (LATr) and restocked at WLISrN, WLITSc and 500 MW during April, May and June 1985. Stations LATr WLISrN WLISc 5O0MW W/D 6.10(0.04) 5.69(0. 30) 6.10(0.50) 5.90(0. 37) L 5/5 01((0535)) 55 35 ((05 20) 5.41(0.11) 55101059) Cd* 0.48(0.04) 0.69(0.18) 0.78(0.11) 0.79(0.18) Gr 2.89(0.14) Aso U7) 2.44(0.24) 2.18(0. 26) Co 0.44(0.07) 0.40(0.12) 0.42(0.11) 0.40(0.09) Cu* 7.68(0.86) 9.09(0.49) 10.53(0. 39) 10.70(0.52) Fe 193(50) 193(105) CSVA((\74)) 237(26) Hg 0.134(0.010) 0.094(0. 068 ) 0.125(0.016) 0.128(0.027) Ni 3.45(0.77) 3.43(0. 39) 3.62(0.68) SoU (Ws25)) zn* 85(6) 102(17) 100(9) 105(26) V 1.14(0.40) 2.04(1. 34) 1.74(0.50) 2.22(0.93) n 3 3 3 One-way ANOVA significant uSing trace metal concentration weighted with — W/D ratios. Tukey Groupings (a = 0.05) LATr WLISrN WLISc 500MW We 0.48 O60 Ooms OSI LATr WLISrN WLISC 500MW Cu 7.68 9.09 LORS Si eek LATr WLISc WLISrN 500MW zn 85 00 02 0s 27 Table 5. Stepwise multiple regression analyses for tissue trace metal concentrations involving the variable “disposed volume." % = tne amount of variance in the trace metal concentration ex- plained by the model. Variable code: L = shell length, W/D = wet/dry tissue weight ratio, T = temperature, D = disposal volume. iia 1 ciakereT TS AIONS. a) Sateen WLISc 500MW Metal Variables % > 4 Gr i W/D 65.1 W/D 52.6 2 L 73.6 L 58.1 3 D 88.8 Cu 1 W/D 47.7 wW/D 24.8 2 L 66.0 L 40.0 3 T 79.2 YT Vsd 4 D 91.9 Fe 1 W/D ARO W/D 29.8 2 L 84.1 L 43.8 3 T 95.6 ilk 87.3 4 D 97.4 D 95.3 Ni 3 W/D 6.5 W/D 43.7 2 L INO = iL 60.7 3 v 78.3 4 D 88.3 Disposal volume (D) as a variable was not ent in i : F ered in th i regression models dealing with Cd, Co, Hg, Zn, and V. @ other multiple 28 ESAN WO Wl c38c bc ISM MA SP9d [&10L O1N0°O 82°22 SO] QS IST IN p9v0°O f£E°S S6 €S$ N4St 1M evel 40[ I04y d 4 Butsung asojag 1013845 S401 04 saduatassiqg pesodsiq Bbultung-asojog “g 62 12 G2 BET SAId [e10L oT bol Evl GL 092I+S2T 40, I04y MWOOS NJSTIM 2STIM JTY S9IUINIJ JIG UOLZLIS-udaMyag “Yy (G0°0 = vo ) sbutdnouyg Kayny "e4ep pawsoysuesy Bol uo pausojsad auam SYAQNY AeM-OM] "suolyeys Buowe aduasayjyip-2 !saduasay sip [esodsip Butunp-asojsag-[ Syuedtytubls WAONY AeM- OM] O\ a ea ae ea ee IRS ae eee ir eat OSE Mad Ped Se ee eS (€ER°0)6b°9 (b8°2)29°B = (9ST) NZ2 (pI) Za (22) 6 (29)€6 (9p) €8 OT MOOS (S2°0)25°9 = (2b°2)19°8 (S6)£E2 (99)SST (2b) 09 (99) S6 (RE) BZ Il 3S 11M (SZ°0)8b°9 §=(Gb°2)25°8 = (OTT) vEZ (9/2) 2ST (68) £9 (85) b6é (2E) 02 Ul N4S 11M (22°0)90°2 = (bS°T)8b°Z (6b)SbT (9€)28 (Tb) 9€ (6€)9b (12) b9 IT JTY ee a er eee Se ee ee es ee ee 71 Q/M $99d O92T+bS2l 0921 bS2l cbel u uOLyeYS L + [90] 4JO| 204y _ 40 904y 40| 20uy 40| 20uy : Gal é “a°S | + Ucay if SSS EE ee ee eee er ee ee ee eee “GR6I aUNe 07 bRET aun( WO4s MW OOS PUR ISTIM “N4STIM “4Ty 3e pakoidap sttnpa snpiqAW ut suolr04y pue (SAdq) spAuaydiq PpayeuLso,YydA[Od [e302 JO vOLze4qUsdUOD (4) YyBua, {yaus ‘(G/M) O1jes yybtana Asp/yam ueaw °9 alqey 29 Table 7. Stepwise multiple regression analyses for tissue Aroclor concentrations involving the variable “dredge volume.” &% = the amount of variance in the Aroclor concentration explained by the model. Variable code: wW/D = wet/dry weight ratio, L = shell length, T = water temperature, M = month, D = dredged volume. Station WLISc 500MW Aroclor Variable % % 1242 1 D 73.6 L 74.8 2 L 88.5 1254 ] W/D 60.4 W/D 61.7 2 T 82.5 1260 1 M 38.7 M 53.9 2 D 67.3 J L 68.3 L 49.0 1254+1260 2 T HAS D 70.3 3 M 87.9 1 L 80.1 L 55.9 Total PCBs 2 T 72.2 3 D 81.5 Dredgedvolume as a variable was not entered in the other two mussel populations maintained at RIr and wLISrN. 30 Table 8. Changes in the mean relative tissue concentration of Aroclors when Ram Island Reef mussels were deployed at WLISrN, WLISc and 500MW (June 84-June 32) )) 6 Aroclor Relative Concentration % Station 1242 1254 1260 1254+1260 RIr x 44.6 32.4 2380 55.4 S.0. 10.4 Da 22.8) 10.4 n 11 1l 1] 11 WLISTN x Boe 43.6 23,51) 67.3 S,0. 5.9 21.9 Dil 2 5.9 n ll i 11 ih WLISc x 33.4 39.1 27.5 66.6 $0). 8.8 238 20.7 8.8 n 11 11 SL ll SUOMW x a3 36.6 Bon 68.7 $0. 6.4 24.6 Dak 6.4 n 10 10 10 10 One-way ANOVA NewSloie W2ae8 PF Boll, Coico S dod, PS O0ONS Ngoehole UA5N8 (p SeO stn Clos 2 S565 iP Ss Osun Aroclor 1Z260k 1h =" 0539), dnt i=) 34395, P= 07595 Nixes Ue NANe «PS 6S, Getic S SSI, P S OOo Tukey Groups ( a@ = 0.05): Station Aroclor 500MW wLISrN wLISc RIr 1242 ORS, 0.32736 0.33378 0. 44629 1254+1260 0.638683 0.67264 0.66622 OSS 3 Syl “quasajysip Aj yuedtytubts you aue xKoqg ayy ULYyyIM |dUa44NII0 yo Sabeyuadvad pue satduanbas4 G0°0 > d ‘Ggrz =!£)90 0x ‘E="3'P “9EZ*6=9 8919517095 1881-9 Oot Wats cH (Ali 62 6 S SAILS ddUd4a Jay $113 00T 6°L L’bp Be C LT Sayts [esodstq S11 $(eOL ony Kew qa4 AON se 30] Bny Kew qa4 AON wotyzeys $aduUasNIIQ yo ADuanbauy ¥X SadUaIINIDQ yo ADuanbaw4 *SUOLJOYS JIUAIDJIA S{[7} pue sazis [esodsip STM 3@ paurequrew sitmpa snyiqzAW ut aqereuyyg ([Axay {AYyJa-2) Stg yo SadUauuNd90 jo Aduanbasajy °6 ayqel 32 Table 10, The number of dead and alive mussels found in the five populations maintained at RIr, LATr, WLISrN, 500 MW and WLISc from June 1984 through June 1985. Summary of replicated goodness of fit tests. Goodness of Fit Test Station Dead Alive r pi df G P RIr 329 404 733 0.449 ] 0.000 NS LATr 340 314 654 0.520 1 13.256 <0.001 WLISrN 641 262 903 0.710 1 251.439 <<0.001 500MW 25) ZRz 862 O75 1 - ASS oENO) <<0.001 IBIS CoO) 28S e039) 0.728 U 3315953 <<0.001 r 2691 1500 4191 =n Total G7 5 866.058 <<0.001 Pooled Gp Ul 631.856 <<0.001 Heterogeneity Gy 4 234.202 <<0.001 33 Table ll. Gonadal development of Mytilus edulis deployed at RIr, 500 MW, WLISrN and WLISc from June 1984 through March 1985. Summary of replicated goodness of fit tests. I = immature, M = mature. SS ae aa eee ee Gonadal Development Goodness of Fit Test Station I M rt pi df G fe) RIr 8 73 81 0.099 ] 0.000 NS 500MW 10 48 58 0.172 1 2.959 NS WLISrN 15 64 79 0.190 1 5.967 <0.025 WLISc Ge 38) _81 0.272 U 19.386 <<0.001 B , 55 244 299 =n Total Gy 4 28). 312 <<0.001 Pooled G, ] 90957 <<0.001 Heterogeneity Gy 3 8.355 <0.05 34 Table 12. The staining characteristics of Leydig tissues of Mytilus edulis maintained at RIr, 500 MW, and WLISc and WLISrN from June 1984 eRrouE March 1985. Summary of replicated goodness of fit tests. L = lightly stained, M + H = moderately and heavily stained. Staining Characteristics Goodness of Fit Test Station Ba M+H 5 pi df G P RIr 65 27 92 0.706 ] 0.000 NS 500MW 49 15 64 0.766 ] 1.128 NS WLISc 54 24 78 0.692 ] 0.076 NS WLISrN 4 30 me 0.594 U 4.206 <0.05 r 212 96 308 = n_ Total Gr 4 5.410 NS Pooled Gp ] 0.486 NS Heterogeneity Gy 3 4.922 NS 35 Table 13, The presence and absence of food contents in the intestine of Mytilus edulis maintained at RIr, WLISrN, WLISc and SOOMW. Food Content Station Present Absent r pi RIr 62 27 89 0.697 WLISrN 49 20 69 0.710 5OOMW 43 17 60 OPa7 il WLISc 43 29 _W2 0.597 z 197 93 290 =n G-test Statistics: G = 2.966, df = 3, X20.05(3) = 7.815, P>0.05. 36 Table 14. The presence and absence of crystalline style within the style sac of Mytilus edulis maintained at RIr, WLISrN, WLISc and 500MW. Summary of replicated goodness of fit tests. WS = with crystalline style, WOS = without crystalline style. Style Sac Goodness of Fit Test Station enw WOS r Pi df G P RIr 81 3 84 0.964 ] 0.000 NS WLISrN 65 6 7] 0.915 ] 3.585 NS 500MW 52 7 59 0.881 ] 7.456 <0.01 WLISc mos 10 _ 68 0.853 U 14.073 <0.001 z 256 26 282 =n Total Gr 4 25.114 <0.001 Pooled G, 1 18.411 <0.001 Heterogeneity Gy 3 6.703 >0.05, 37 <0.10 Table 15. Frequencies of enlarged plycate organ encountered in Mytilus edulis deployed at RIr, WLISrN, WLISc and 500 MW from June 1984 through March 1985. Summary of replicated goodness of fit tests. E = enlarged tubules, N = not enlarged tubules. Plycate Organ Goodness of Fit Test Station SE N 5 pi df G P RIr 52 39 9] 0.571 ! 0.000 NS WLISrN 42 3] 73 0.575 ] 0.005 NS WLISc 82 48 80 0. 400 1 9.474 <0.005 500MW 5 _38 _60 0.397 U 7.741 <0.010 iF 15] 156 307 = n_ Total Gr 4 17.220 <0.005 Pooled Gp 1 7.850 <0.010 Heterogeneity Gy 3 9.370 <0.025 38 Table 16. Frequencies of Proctoeces maculatus infection in Mytilus edulis maintained at RIr, WLISrN, WLISc and 5S00MW from June 1984 through March 1985. N = 0 sporocyst, L = 1-9 sporocysts, M = 10-99 sporocysts, H>100 sporocysts per section. ; Proctoeces Infection % Proctoeces Infection Station N L M+H Totals N L M+H Totals RIr 70 16 18 104 67s) lise4 Ws 3 100 WLISrN 65 19 20 104 6265 NES 19.2 100 WLISc 73 22 13 108 67.6 20.4 12.0 100 500 Mw 66 13} 14 93 71.0 14.0 550 100 Pepa mes ie es 2 = G-test Statistics: G=1.9670, d.f.=6, x 0.975(6) 71-237» P0.05 500MW 17 9 26 0.654 1 2.013 >0.05 WLISc eZee eS 15: 0.800 at Sr 2A5/ <0.025 >} 62 36 98 =n Total Gr 4 9.243 >0.05 Pooled Gp it 5.405 <0.025 Heterogeneity Gy 3 3.837 <<0.05 40 Table 18. Frequencies of Pinnotheres maculatus infection in Mytilus edulis maintained at RIr, WLISrN, 500MW and WLISc from June 1984 through March 1985. Summary of replicated goodness of fit tests. I = infected, NI = noninfected. Pinnotheres maculatus Goodness of Fit Test Station I NI DD pi df G P Rir 156 62 218 0.716 1 0.000 NS WLISrN LOS 6H 164 0.628 1 5.858 <0.025 500MW US — Dak 124 0.589 1 2) oz <0.005 WLISc 635 5257 SS) 0548 1 14.560 <0.001 D> 395 226 621 =n Total Gr 4 Be) 5 D0) <0.001 Pooled Gp 1 18.380 <0.001 Heterogeneity Gy 3 SeeL710 <0.025 There is no difference in the frequency of Pinnotheres infection among the three WLIS populations (G = 1.815, x2 0.05(2) = 5.991, p >> 0.05). 4l Table 19. Frequencies of leukocytic infiltration in Mytilus edulis maintained at RIr, WLISrN, WLISc and 500M/. N, L, M and H denote respectively no, light, moderate and heavy leukocytic infiltrations in the tissue sections. Leukocytic Infiltration % Leukocytic Infiltration Station N L M H Total N L M H Total | i RIr 19 40 23 12 94 20 43 26 He 100 © WLISrN ll 33 16 14 74 15 45 22 18 100 © WLISc 19 33 20 7 79 24 42 25 tS) 100 500MW 7 28 16 WAS) lil 44 25 20 100 one é 2) . , G-test Statistics: G=8.389, d.f.=9, x 0.50(9) 78-343; P=0.50 42 STAMFORD v. WLIS tl WLIS-II1 STATIONS LORAN-C CO-ORDINATES 26831.9 43975.2 26835.0 43975.8 WLISc SOOM w 26824. 3 43963.1 WLISeN WLISRN ~ 5 ST %e FLLOYD NECK 3 The Western Long Island Sound (WLIS) disposal site depicting the locations of three mussel monitoring platforms at WLISc, 500 MW and WLISrN, as well as the disposal buoy. Figure l. 43 = ~aeuvr ee wou} 3115 Lesodsip STAM ayy 1° pasodsip (suaqau d1qgnd) 5] e149} ell pabpesq °z aunbis SIVIHVSLVW GS5q34q i (S86L - P86L) HLNOW nr AW dv VW 34d vr dd ON 30 ds nv aly nr STIM LV GaSOdSId SIVIYSALVW Ga5qsya oot Oct ost ams SS (spuesnoyl) Sd3LlawW olagnd 44 FISHERS ISLAND SOUND STATIONS LORAN-C CO-ORDINATES RIr 26086 .7 43982. LATr 26063.3 43977.0 si a GROTON ee ere yo ae ee. or) NLON DISPOSAL SIE | SC/NES | | NAUT. MILES of RIr and LATr. 45 Figure 3. The Eastern Long Island Sound reference sites showing the locations | TO SURFACE BUOY Figure 4. A monitoring platform with attached bags of mussels. Each platform was equipped with sonic beacon (B), a remotely operated acoustic release (not shown), a surface float, and a subsurface float (P). The subsurface float was released if the surface float was pulled free. 46 0} Asenuee wor} paqyonpuod sem IS]qM 2e UOLyeuado jesodsig “GR6T aune “(VE alge, aas) SsaqLs STIM 249 pue JTY 7e paurezULew S_assnw uaaMzeq pazI97JaP 949M SUOLZeUQUaIUOD [eal aDeAZ BYI UL SadUBIA}JIP questytubis “MLIOOS PUR IST IM “NAST IM “4ty ye pafo[dap suotyetndod jassnw ut outz pue *1addo9 ‘wn Luped JO UOLJeAZUBDUOD ansstz ayy pue O1zeU YYHLaM anssly Aup/aN ayy JO UOL JEL AeA jevodway $86i | Gsm! Gamat bapa» aml He Jae 9 8f9° bil” LL 8/8" aauvnds -¥ GOél GN 0 8 UYU f f¢ be6l 0S Oot OST 002 MWO0S =F ISL IN=€ wagpanez | O52 JTYeT NOLLVIS becl tw 8 Nea NYO ee ee p BLL” 6LL° SLL” bel: aauvnds -Y 90°0 0S‘0 00°T 0S °T 00°2 G-b : Sah 0S°¢ vst oo HOLLVLS ~ NZ qo c06l beet Re mH YM) 19 2 f anos uw ¢ 2sl° MHOOS =b WL IST IM=€ ava NAST IM=2 Tbb° JTMEl aauwnbs -y NOLLWIS $86l bO61 Sb6" MHIO0S =b t 122° IST IM=€ 096° N!STIM=2 19L° ATU=l aauvnbs-uy —- NOTLVIS 00°€T 00°ST Nd . 41q/39M 47 Sr aaa Ee ee Se Re ee eee ag ee ee ee ee t t ae ae ir ai treo EE a Le aD NA acca aa 1 jesodsitp Bbulunp pue au0jaq pajde7,ap 949M UOLZeUZUBIUOD [|e Za! ade} au} UL Sa0uatas Jip UBD LSU -B1S “MNOOS PUe DSTIM SN4YSTIM S4TY 2e Plau SuoLye{ndod |assnw ut uouL pue uaddod ‘4, eqod JO UOL}e4QUaDUOD aNssiz ayy pue sqYyHtam ansstzy Aup/zaM JO UOLZeY |YY JO UOLZeLUeA [euodwal, ‘9g aunbiy Ge6l vast f ¢ W YU WH 4 fF GN O 8 YU ff fF 9 8 OT cl 2 | m4 vT tase aot MINONS = b au b99° N4SITH=-2 = | «(009 te ISM? 695. anu=t | aes yee 81 aauvnds-Y NOLLWIS 00L aauvnds -¥ NOTLVLS ro £861 beet ff WU NU-se Tt OhN=O 8 UW ft ft f f W UW it Qn oO 6 Mr f 00°TT 059° MWO0S =b 1 MHIO0S =b 00°ET 168 NAST IM=2 5 : : ° 096° NIST IM=2 £68 JTY=T 00°€ iol" squel aauvnbds-¥ NOTLWIS: 00°ST gauuvnbs-¥ NOTLWIS no Aud/LIM 48 “quedstytubis ALLeotystqeis you a4am suotzeuado jesodsip 6 F 17S | t [Ip DULANP-940Jfaq pue SUOLzeIS eee Sauda ILP JI SWSHHO MHONG Pue OSTIM SN4STIM S4TY 2e pauLezuLew SuoLzei{ndod jassnw t Lpeuea pue Aundsaw Swntwosyd *[ax¥ILU JO UOLZeAQUaDUOD ANSSly ay. JO UOLQeLUeA [P4Odma, *, a4nBL4 00°0 00°T 00°C N0°€ 00°47 ell’ MNOOS = ‘ lbl: IST TM=€ 00°S b£9° NAST IN=2 ZRE° ATu=l 0°9 a3uvnds -¥ NOTLWIS 00°L Ceci O61 fh) WE = iS he ce NG OS cu ORS EF 080° 021° 09T° 002° 072 ° 659° MWO0S = bla" IST IM=€ 188° NAST IM=2 O8Z° 1b6" A1Y=T ajuvnbs-y = ss NOTAVLS 89° aauvnbds -Y¥ OH 0Sb- O3NVNDS -Y 00°O 00°C 00°% 00°9 00° MOOS =b 10° OT 3S) IM=€ NAST Wee Jyuel 00°2T NOLIVLS 00° #T OO°T 00°C 00° 00° 00°S 00°9 mi00S-b 400°Z 2S} IM=E NAS] WN=2 : suet 700°8 NOLLVIS 00°6 WO IN 49 e ee ee Figure 8. Two GC chromatograms of polychlorinated biphenyls from mussel tissue extracts: A. normal chromatogram, B. exhibiting a large anomalous peak. 50 “ayereuyud ({Axay (AWQa-Zz)sig se (ys aunbt 4) paauasqo yead Sno[ewoue ayy JO UOLJeULLLJUOD puke UOLzedLytqQUap!L Auqauiouzdads ssey/ydesboyewouy) seg “6 aunbr 4 “puepueys ay} JO 2eY? YILM SH} ay yo Buryozew yoexa Butmoys punoy [e4quay punos puetsy buoy usazsamM wouy aldwes tassnw ayy Jo wnaqoads ssey “gq “(paepueys) azepeuqud ({Axay [Ayza-2) stg yead YET ay JO WN4ydads sseW °9 “spuepueysS WazSa 92eLeYIWd OT ByQ JO SUOLZeUZJUBDUOD UOL [eJO] “gq “ajdwes jassnw (2SI1M) Punoy saquay punos pues] Bbuo7 usazsaM yO SuOLzesQUaDUOD UOL [RIO] “YY bpIS E Q° '8@Se6H60:-cte 399 JTL 630 2 Q° 1G8S8a1r6ea-ct# 39 JIL Q0azZ Baar aaag AGHA a0n0\ T# 39 (UIW G26° G2) GOrI Yeas CUIW GEG Se) EDI YEAS 51 “MWNOOS PUE ISTIM *NASTIM ‘4IY 3e pakotdap suopyeindod tassnw uy AzyLeysow aAyyeyNUIND “OL aunbyy suo 2 WAG suse IS Tle Gl EEE elf ilies 2 peecare eee el: ee 5361 - 86> HLNOW oud? (WY asd Ge al Opbsiesse i kil ALTWLAOM SATLY INNS LE ALT TIBPLYOM SnT Le Inns 52 APPENDIX 33 (11°) (S9°T) (9€°) (100°) CC Sieatg) (2€°) (00°) (20°) (20°) cl” 66°et ie 820° bO°E€? 82° T 90° 61° 80° -SB8/82/€ (bI°) (22°b) (01°) (100°) (82°b) (Glia) (20°) (20°) (00°) 69° L0°02 98° 9£0° 82° ES 8b I Sb° 02° 1 WSs GR/ET/2e (10°) (62°) (€1°) (S00°) (T€°S) (91°) (cm) (92°) (10°) Gls Ol'ST As 620° €L°S2 c0°T €2° Ob vI° S8/€/1 (20°) (Sb°e) (21°) (100°) (Tb °6) (21°) (10°) (12°) (20°) L\° 91 °ee OL° SC0° SS* bb 80°T el” RL* b2° b8/E1/21 (10°) (2S°2) (90°) (100°) (EB=2)) (S0°) (00°) (b0°) (10°) £1’ €6°91 ce" c20° 29°61 Sg)” S0° LT" et” bB8/e/TT (10°) (p2°2) (81°) (100°) (68° 1) ( ) (T0*) Gules) (10°) - 60°9T bp” c20° fc Le L8° 80° cS° GO| hen b8/61/6 (0°) (2€°2) (GS) (100°) (06°) (91>) (10°) (0°) (10°) b0° Cb et Si CO €S ot Silat SOs Gs 0) vR/22/8 (02°) (€9°T) (80°) (200°) (1S°b) (82°) (€0°) (91°) (20°) €0°1 SS*I2 Ob* TEOR 02° ¢E 62° 90° bS° Tale ba/92/L (S0°) (£2£°2) Cl) (€00°) (09° b) (81°) (20°) (60°) (10°) Siles BI°St Cie 1€0° 1Z°9e 82° T £05 62° L0° b8//2/9 (b0") (21°9) (92°) (»00°) GAO) (HE) (20°) (S2°) (€0°) 6t° 10°61 SG £20° S2m6S 0 *T 60° 09° bi” b8/92/b A uZ tN H a4 ny 09 Ad Pd 97eQ Hur jdues (anssty jo 4M 4am 6/64) "Q°S + UOLJPujUadUOD [eJaW UeAaW "(ATY) A2LS adUauVasau pue{s] wey 3e pakojdap (stinpa snjiqAW) stassnw ut (7y6tam aM) suolzesqzuaoUOD ;eZaW Jo AreuMNS *eT a[qey 54 a ES a ek a (p0°) (85°T) (pT*) (S00°) (06°S) (AS?) (10°) (elites) (10°) St’ 2l° 2 gf ° 810° GI°92 Glee 60° 81° 80° G8/b2/S0 (O1*) (92°T) (20°) (100°) (86°T) (i) (10°) (20°) (00°) 12° 68° IT lites €20° £S°l2 68° 90° 9° 60° G8/92/b A uZ UN 6H a4 ny 0) 9) oe) 37eQ But | diue ¢ : (9nssty jO° 4M yam 6/6") “Qqrs 4 UOLJe4yUaDU0) [PAW Ue|y a EE a aa a Panutjyuo) “eT atqey 55 (€b°O) (6b°9) (20°2) (000°) (pl bt) (22°11) (20°0) (/1°0) (91°0) 21 °b Gl°bd 61°2 091 °0 v2 ££ RE °2 l£°0 alee lb°O G8/82/£0 (€2°0) (98° /2) (Sb°0) (€00°) (19°81) (0S5°0) (pT °0) (/1°0) (S0°0) Geb 91°22 bys (22° 16° 6££ bl 6 L8°2 19°2 0€ *T GB/ET/2 (S2°0) (0S°8) (26°T) (220°) (28°22) (bb°O) (82°0) (08°2) (12°0) 2e°1 Ip’ 8bl aes 8R2° 89° 262 80° OI b2°2 bl°b cial GB/E/T (S2°0) (21° 2) (92°) (210°) (Tb°€6) (28°0) (60°0) (00°2) (/2°0) 9f° | Lp°18T 2L°S €82° 00° 29€ 18°8 86°0 £p°9 R6°1 pB/ET/Z1 (T1°O) (0°SE) (22°0) (b20°) (86°E€1) (0€ °0) (90°0) (0$°0) (/1°0) 18° 1 p2°C8l Ip‘2 2b2° bL° O12 16°9 65°0 LB°1 2b°1 p8/2/11 (61°0) (€1°62) (pE°T) ($20°) (9S°2) (62°0) (S0°0) (E21) (81°0) 16°0 GE OET 9p°€ GLI° 20°812 16°9 G9°0 92°p 66°0 b8/61/6 (p£°0) (p9° pT) (00°2) (v10°) (6S°b) (26°0) (80°0) (Ob°0) (F0°0) 9£°0 62° SOI bob c9ol° 19°€S1 12°6 cpa NG°2 laa) bR/22/8 (90°1) (6£°9) (6€ °0) (600°) (02° 1€) (18°T) (91°0) (90°) (Nv°0) 0S°9 G9°9ET 6b°2 C6I° G2°8£2 02°8 9£°0 Gb'e 120 p8/92/L (82°0) (S2°91) (82°0) (€10°) (0° b2) (pZ°) (O10) (75°0) (10°0) 00° G9°86 p1°2 102° {2°edl 2e°8 S1‘0 06° Gb°O 08/ 12/9 (€2°0) (92°02) (68°T) (p10°) (62°65) (20°T) (O1°O) (22°10) (€2°0) LE °1 9/° CET 09°€ p8t° 95° 6bE Gl°8 85°0 b0°b R9°O b8/92/b A uz N by a4 nj 0) 49 oe) a7eg bul dues (anssty jo ym Aup 6/61) *Qg°s + uoyyeuyuadu0) [eJaW UeAaW aR REE me::J£Z-__,_:,:_:-,-,_, ee ee ee "(ATY) 9718S aduasvasau puelsy| wey ye paxoidap (sitnpa snjiqfi) stassnw ut (qyb1am Asp) Suotyesqzuaouo0d [eyaw jo Aueuuns ‘qt arqel 56 a Eee Sha ec 2d eS ee oe (O10) (S8°b) (€5°0) (S/0°) (08°ST) (18°O) (€0°0O) (99°0) (€0°0) 66°0 2S °€8 92°2 Ells 26° £9 G2°L 9£°0 Ge *T 25°0 68/2/50 (02°0) (1S°6) (19°0) (p10°) (Sb°Z) (29°0) (S0°0) (22°0) (20) 61° p2°8/ ras 251° 89° O8T £8°S £v°O 10°T RS°0 68/92/60 A uz UN 6H a4 ny) 0) 4) Pd a7eg Bur jdues RR a ee UT + _u0lje4jUadU0) |e aW UeAW “panutyuoy “qt aqey 57 (10°) (19°e) (Aft) (00°) (S2°T) (02°) (00°) (11°) (00°) Tate b6°61 Sie €00° Ol et 09°T 90° 6T” Sil SB/TI/L (20°) (8S °0) (€0°) (€00°*) (25°S) (60°) (00°) (0° (10°) 6£° 06° LT co” 120° 62° tb sis) I Ol Tb° el” $8/22/5 (60°) (p9°T) (91°) (200°) (16°2) (02°) (00°) (0€ *) (10°) 65° €S°ST 89° £20° Sb°lb SS*T 90° 18° 60° G8/S2/b (€0°) (/6°0) (v0°) (000°) (p8*b) (ST") (10°) (90°) (00°) Co Sv 8st Con b2e0° €0°SE 18° 60° UG Sie G8/92/€ (90°) (12°€) (20°) (900°) (86° IT) (0) i) (S0°) (10°) (20°) UG™ St °9T Soe 620° 80° Lb Bs) \l GOs cc° Ge° G8/11/2 (10°) (ST°b) (ST°) (€00°) (15°S) (90°) (20°) (€2°) (20°) Gls cl’ lt SSa 1c0° 6b ° 82 etl L0° Bb° Cou b8/82/21 (10°) (p1°2) (0°) (S00°) (19°9) (92) (10°) (90°) (10°) LOm SOMO 6e° Tc0° Ov *S2 bil L0° Gc" ie b8/1t/2l (€0°) (9S*b) (pI°) (100°) (€2°T) (20°) (10°) (b2°) (20°) La 12°02 BE ° 020° 68°82 08° L0° Ne ° A bR/6/1T (10°) (/6°S) (82°) (00°) (bI°€T) (92°) (0°) (€0°C) (10°) 60° St° dt 60°T 610° 06 °OE (fe) Gls Sb°2 St” b8/81/6 (20° (€S°T) (0°) (»00°) (OTT) Cie) (€0°) (0°) (20°) 90° 66°8T Of * 220° 8G SN Olt 60° Cle oa G8/52/8 (€0°) (20°T) (€0°) (200°) (22°€) (02°) (€0°) (40°) (20°) 02° Ge) IG cb” S20" 6b °S2 bet b0° (HO le: bB8/S2/L (ANSSty JO YM Yam B/HN) *Q°S + UOLzR4QUaDUO) jeJaW UAW a er “(NUSTIM) 8934S adUaaJa4 punos puelsy Buo] usazsay ye paXojdap (SLINPS SN[ LIAN) Stassnw ut (Yy6Lam 79M) SUOLJesqUaoUOD |eyYaW Jo ArewWNS ez atqey} 58 CU) (89°€1) (09°0) (100°) (62°9) (88°0) (20°) (19°0) (b0") s9°0 80°02I LO°€ 610° 06°2/ £9°6 ££°0 Glee gn °O GB/II/L (81°) (Glee) (62°0) (010°) (€8°6€) (Sb°0) (20°) (9b°0) (10°) tiles Of * 26 [oa6 €Il- p0°6€2 66°8 £5°0 (82°? 89°0 G8/22/5 (19°) (81°) (60°T) (OO) == (2282) (65°0) (€0°) (98°T) (10°) come G/°/8 GB°e OSI° Ge °992 99°8 f£°0 6S‘b 25°0 G8/S2/b (€1°) (0) (81°0) (600°) (29°62) (1S°0) (40°) (9b°0) (10°) be °2 99°9£T BE °2 G/I° b2°lS2 82a 99° cst 91° G8/92/E (69°) (85°SE) (02°0) (910°) (€b°S2T) (Glcp) (28°) (bT°0) (12°) po °2 66° 081 99°¢ GIe° 62°02S b6°LI IGS°€ bbz 19°2 GB/1T/2 (i) (26°82) (1S°2) (020°) (68°19) (91°T) (E22) (Ov °€) (bT°) 2s 1 p2°202 ££°9 822° £6°62E 2£°ST 98° 18°S 6b°2 bB/R2/21 (0°) (22°21) (p0°T) (220°) (2S°Tb) (ST°2) G@|E?)) (S8°) (Tf£*) 28° €2°E81 Ov’b 822° 29°9/2 bS° 21 €L° G8°2 G6 °T bB/11/2T (/b°) (pi°19) (p6°T) (120°) (25°9€) (1£°0) (ST°) (€0°C) (be°) 26°1 0f° 222 9f°b NEB? 18°22€ 06°8 g/° Abas v6" p8/6/1T (S0° (89° bf) (€1°6) (Gao?) (ESA) (09°1) (1S°) (92°62) (bI°) 98° 2€° 091 96° OI BLT” €2° LOE 12°8 Of °T Nelaultc Iv'T p8/21/6 (01°) (pS°S) (6£°0) (@20°) UAE 2@) (SE °0) (92°) (0£°) (OI) bb’ 20° bbl 92°2 891° Ip 2 Ov°8 0L° 00°T Gl°t p8/12/8 (22°) (6£°9) (91°0) (GIO?) — CAE Ee) (6€°T) (12° (9p°) Gli) tbl Ob IST G6°2 281° bb °S8ZT 02°8 O72 bS°I Gl° p8/S2/L A uZ UN 6H a4 ny 0) 49 8) a3eq z Bury dwes (anssty jo 3M Aup 6/6) *G°S + uolzesqUazUO) |e aW UeaW “(N4ASTIM) 89LS adUa4ajfau puNoS pueR[s] Bu0q U4a}SaM ye pako[dap (stinpa sn tzAW) Spassnw ut (YHLam A4p) suotzesquaouod jexaw jo Auvuuins -*qz alqe} 59 (Oy) (S2°S) (22°) (»00°) (SZ°b) (1€*) (10°) (40°) (20°) ce SO°LT CS)” c20° bb° be 09°T L0° 6£° bt” G8/92/9 (10°) (8S°T) (/T°) (€00°) (20°2) (01°) (10°) (01°) (10°) 02° cS °9T Bhs)e L10° SL” UKs 02° 80° St° Sle G8/22/S (0°) (99°T) (€0°) (b00°) (98°) (0°) (10°) (60°) (10°) ice Ge 91 Com €20° b6°cbv b6"T S0° 8b° a S8/S2/b (€0°) (0S°2) (90°) (100°) (€1°Z) (90°) (10° (00°) (00°) 62° 12°02 WG £20° cL 6b 9b°T 80° 61° 8t° G8/92/€ (10°) (62°€) (2°) (200°) (92°S) (60°) (»0°) (0€*) (10°) 81° 90° bT 1b° 120° 02° Lb Semel 82° ge G* G8/11/2 (0°) (€8°T) (20°) (€00°) (E9-2) (01°) (10°) UA) (10°) 8° 65°02 BE ° €20° Te ep fb’ 60° SGe 12° b8/82/21 (10°) (00°T) (20°) (100°) (62°S) (S0°) (00°) (St) (10°) OS €b°st 6b" 820° G2°eb LO*T 60° bS° 8T° b8/TI/et (€0°) (p1°S) (61°) (100°) (b€°2) (01°) (10°) UGE (10°) Sl CW) Ni 09° c20° 09° TE €6° 90° 09° ils b8/6/TT (20°) (96°b) (40°) (900°) (Ib'b) (/2°) (10°) (61°) (£0°) 80° 61°02 (S15 920} Se°9T £0°T 0) 6C° OC b8/81/6 (20°) (69°2) (€0°) (€00°) (p12) (lb") (b0°) (90°) (S0°) Wt 99°02 0c * €20° Tb st Sb°T S0° 82° (lh v8/12/8 (10°) (6€°¢) (€0°) (€00°) (8/°2) (€0°) (20°) (S0°) (10° S105 1O°8t O€ ° b20° 9€ * ec San v0“ NE N= bB/S2/L A uz : tN H | nd 0) A) Pd 9yed Se Ss 2 Ss See 25 SS Sees ee ae Bury dures (9NSStY JO JM JOM 6/6") ZFS 5 UOLJOUJUIIUO) [PJoW ueayW a ee A a “(9S11M) Punow g 4a}uUaj) punos pue[sy Buo7} usazsay ye paXo(dap (stinpa sn tqAW) SLassnw ut (34y6LamM yam) suolyesqUadUOD jeqow jo Axewuins "ee atqey 60 (91°0) (16°21) (S/°0) (800°) (€p°€T) (8€ °O) (20°) (20°) (Tiles) 99°] 28° SOI €B°€ (eye £6°L12 80° OI Ly° vb°2 98°0 68/92/90 (p0°0) (22°11) (90°) (120°) (8211) (65°0) (S0°) (29°) (0°) 82°T SZ° bOI Ll'b LOT" £0°6£2 62° OI 6b" 61°2 28°0 G8/22/5 (81°0) (00°8) (8170) (220°) (09°8) (22°0) (0°) (9°) (90°) R2°2 88° 68 98°2 OfT" 08° /E2 €L°0T 62° R9°2 s9°0 G8/S2/b (€2°0) (65° TE) (€9°0) (€00°) (79°16) (bb °0) (»0°) (/0°) (0°) 9l°2 bO° 161 661 LS2° 2L°OLb 6L°E1 61° 2R°T €L*1 G8/92/¢ (82°0) (00° 2b) (15°2) (220°) (b° bd) (49°0) (»9°) (10°b) (€0°) Ob°2 L0° 881 61°2 G82" v0°629 LL°91 29° €£°R le G8/11/2 (Zb°0) (S2°62) (26°0) (6£0°) (16° 6b) (2R°T) (OT") (9€°T) (61°) 981 St°2t2 G6°E Op2° 0S°S€b EL oT LR° Ob'2 Clane GR/82/21 (21°0) (S0°€) (62°0) (€00°) (98° Ob) ( 2£°0) (20°) (AIT) (91°) 90°1 95°8SI 90°S BB2° 92°E£b 00° U1 06° 60'S 28° 1 b8/TT/21 (8€°0) (Tb ° 2b) (Lp °2) (S20°) (06°€S) Gita) (1t") (Al 2) (22°) Sb b2° E81 08°9 Eb2° bl°bSE 9£ “OT OL’ G1°9 bet b8/6/11 (0) (60°92) (010) (000°) (bS°2T) (€5°0) (80°) (02°) (v1°) 19°0 80° 291 19°2 061° 22° 621 61°R 62° 02°€ 2° T p8/81/6 (2b°0) (25°11) (60°0) (S10°) (56°) (81°2) (81°) (vb°0) (22) 92°0 S9°9ET 20°2 eSie 06° 121 08°R Le° 68 I Gaal b8/12/8 (60°0) (9€°S2) (22°0) (€20°) (25°91) (SE°0) (ST°) (1€°0) (€0°) 68°0 2b d21 Hela L9T° bb’ SST 0L°R 62° OUT 22°0 08/G2/L A uZ UN 6H a4 ny 07 4) Pd a7eQ) Hur | dwe SSS aaa ee ety (anssty jo ym Aup 6/6") “q's + UOLYL4ZUBIUO) [eJaW UeayW a ee a "(2ST1M) punow g 493ua9 punos puetsy Buo7] usazsaMm ye pakoidap (stinpa SN{tqAW) sjassnu ut (1yB1am Aup) suotqesquaouos jeyaw Jo Aseuwns “gE apqeyi 61 (20°) (S0°) (01°) (€00°) (16°S) (92°) (10°) (10°) (10°) 9f° 92°12 89° 220° 12° 6£ YLT 10° RC* 91° G8/12/9 (20°) (ST°T) (12°) (100°) (»S°2) (91°) (10°) (60°) (10°) cen 2°91 19° L10° 8b °9£ /8°1 80° fe" €T° G8/22/S (90°) (22°1) (pT°) (200°) (€S°T) (10°) (00°) (90°) (10°) GS° fb°ST b9° 920° b2°Sb 08 *T 60° BE‘ It’ G8/92/b (p0°) (90°C) (20°) (b00°) (02°6) (ST°) (10°) (b0°) (20°) Of ° €2°6 00° 920° €S°1S L9°1 80° 81° (Be G8/92/£ (10°) (Sp°t) (S0°) (200°) (Sb°S) (OT°) (10°) (20°) (20°) (Ee £9°9] Of ° 620° 6£ bb IS‘ 62° 9I° G2° b8/11/2 (10°) (09°€) (61°) (€00°) (89°2) (€1°) (1?) (vb°) (20°) 10° 9f° IT 6S° vIo° 01°22 S8° 60° Tilesat GI° b8/11/2T (20°) (68°2) (S0°) (200°) (0S°2) (11°) (10°) (pt°) (£0°) LV 91°ST Of ° £20° 02° 0€ 86° 90° Se Ae 8/6/11 (20° (09°€) (90°) (200°) (€5°) (ST°) (10°) (20°) (20°) 90° 8p vl pb’ L10° 28°81 16° 80° Ob* GI° b8/81/6 (€0°) (€£°C) (b0°) (€00°) (€£°€) (60°) (20°) (€0°) (20°) 60° LE°LT be° £20° /8°81 Zleal v0" 9T° bi’ b8/12/8 (10°) (81°1) (pT°) (100°) (99°T) (91°) (20°) (b0°) (10°) are 96° oI fb° 610° 69° 8I (PO. £0° 21° (\) pB/S2/L A uz UN 6H a4 n) 07 wis) Pp) a3eq Hut} dwe — See {| S$ (anssty jo ym yam 6/6N) “Q's + uoijesjuadUuO) [eJaW UeAaW ee “punos puelsy Buoz usaqsam ut (mMWOOS) Punow g Jo ySam 62 S49} 9W 00G 32 pakojdap (si[npa sniiqyAW) Stassnw ut (qYybLam yam) suoLzeszUaDU0D [eJaW yo AuewwNS “ep alqe| (O01 °0) (22°¢) (SS°0) (810°) (€8 TE) (Ob°T) (0°) (20°) (20°) [Gare 19° bET 66 °€ Obl” EL °Lb2 el Sb 0 bb'2 66°0 GR/ 12/90 (1170) (98°) (02°T) (900°) (SZ°OT) (22°) (S0°) (8b°) (0°) G2°T S0°€6 6b°£ 160° 66° 202 99° OT Sb°0 16°T bZ°0 G8/22/60 (€b°0) (Sb°TT) (b0°T) (010°) (86° TT) (bb*) (0°) (Tb*) (90°) Teleae 91°18 99°€ Lvl’ 2b°9S2 12°OT 62°0 812 £9°0 G8/92/b0 (€2°0) (bS°92) (9€ °0) (220°) (91°96) (97°) (b0°) (SS°) (pT°) b9°2 G9° 691 S6°1 Sec SO0°LSb 99° bT bl°0 b9°T 6b°T G8/92/E (b2°0) (8£°SE) (98°0) (210°) (9b° bz) (1b°T) (/L°) (/€°) (2°) 20°2 15° 961 ese 182° 98° 21S G9" LI Ibe BRT 16°2 GR/TT/2 (p2°0) (86°60) (1S°€) (20°) (€8°S9) (S¢°T) (02°) (/b°Z) (€T") 90°1 80°29 95°8 002° Ob SIE €S*21 9/°0 98° C1 60°2 v8/TT/2T (60°0) (/8°ST) (p€°2) (210°) (29°2S) (SS°) (90°) (S9°T) (A?) 82° 1 80° 291 BE *b G22° bb °€2€ 9€ ° OT 69°0 SEE 8L°T p8/6/T1 (61 °0) (p8° £2) (62°0) (600°) (S2°TT) (28°) (60°) (Fates) (p1°) 9S°0 90°S2T cB°e Spl’ 20° b9T L£°8 69°0 6b°€ f£°T »8/81/6 (S2°0) (9€°S2) (0€°0) (120°) (11°S2) (19°) (ST°) (92°) (ST*) £9°0 OD? LZ 6/°1 89T° 95° BEI 02°8 92°0 Aten 66°0 b8/12/8 (S1°0) (Sb°8) (9/°0) (800°) (96° b2) (SE°) (ST°) (SE) (800°) 88°0 /6°801 SO°E cbt: GO°ZET 06°R G2°0 06°0 €2°0 v8/S2/L A uZ UN 5H a4 ny 0) AJ os) a7eq Huy due nm pe ee ee DU tires (anssty 40 ym ap 6/61) ‘a°s + UOLJP4ZUIIUO) [PAW UeAaYW a ee ES nk ee Pr “punos pue{sy] 6uo7 usaqsamy ut (MWOOS) punow g ay jo aSaM S49}OW OOG 3 paXkojdap (stinpa Sn| LAW) spassnw UL (qyuBLam AUp) suotzeuzuadu0d Leyow jo Auewuns -qp alqey 63 (9) (61) (€) (€1) (2) vs (5) (€) (9) (2) (S) S2 cbt ot LS é Ol 6 Lv ST G8 S8/82/€ (b) (22) (2) (21) (0) (0) (2) (21) (€) (81) LI Sol l fb I S 9 BC Ol c9 GR/ET/2 (>) (92) (2) (91) (b) (9€) (2) (02) (2) (ST) cl UG 9 RS S 9b cl l €9 G8/E/1 (2) (0S) (9) (€b) (9) (€b) (0) (0) (1) (2) c¢ bét Sl ect ST eel 0 0 9 es bB/E 1/21 (>) (2S) (2) (02) (2). (S2) (€) (pb) (€) (c€) cl SEL S 8S é be t ce l Gl b8/2/T1 (S) (2€) (€) (02) (T) (>) (€) (RT) (2) (21) 6 el 9 Sb i S S Ob c Le b8/61/6 (1) (1) (0) (1) (0) (€) : (0) (b) (0) (0) SI vil 6 bl \ 6 R G9 S Ov bR/22/8 (92) (621) (€1) (88) (0) (2) (€T) (68) (£1) (58) 6£ €S2 €2 OST I 8 ce ebl 91 col vR/92/L (py) (12) (2) (8) (0) (1) (2) (6) (€) (81) 14 lvl eT L8 I 8 él 6 a bS ¥v¥hB/ 12/9 (€) (21) (€) (21) (0) (2) (€) (ST) (0) (4) 82 vol 81 ecl I 6 LI ell ¥¥OT ¥89 b8/92/b {eyo} O9ZT+bS2l 09cT bSel chet 372d 40{ 904 40 [204 but; dure : °0°S + SUOLJe4QUaDUO) gy UeaW "(ATY) azts GIUdaJ9A PULLS] wey ye pako|dap (Si[npa snj LAW) s_passnw ut suotyeuquasuoy Ad JO Auewwnsg °G arqey 64 in DULL ASPG xxx w "0°S +_7M yam B/bu ut passaidxa xy "0°S + 3M Aup 6/6u ut passaudxa y (6) (Tf) (9) (61) (€1) (ST) (9) (>) (€) (21) Ov [02 Le tcl 9L p6 0S 62 bt 81 S8/b2/S (2) (9) (2) (02) (2) (6b) (S) (TE) (S) (be) ae lvl cl 08 6 cg c BI Ot 19 G8/92/b {e 70} O9cI+bS2l 09cT bSel cbel o7eQ 40} 904 AO; 304 "0°S + SUOLJRIJUaDUO) gIg UeAaW a ee ee ees *panutyuod ‘*G arqel (28) (8) (6S) (6b£) (»9) (6/€) (8) (9b) (£2) (9€T) 88 v0S 19 9bE 9S 6I€ S le 82 BST G8/S2/b (9) (pb) (S) (€€) (2) (01) (€) (€2) (2) (ST) 8I cel Ul cB é LT 6 S9 l eS G8/92/€ (8) (88) (9) (89) (€) (8€) . (€) (9€) (2) (22) 61 L0e cl cel e Le 6 G6 l Si G8/ 11/2 (2) (12) (9) (29) (8) (28) (2) (£2) (1) (6) bt 191 6 901 8 26 I a S 9S bR/R2/2T (pb) (bb) (€) (6€) (S) (2b) (9) (69) (1) (9) rat 981 cl 621 8 68 c Ob S LS b8/T1/21 (>) (SS) (2) (0€) (»2°0) (b) (2) ((2) (2) ((2) 9T LLi Ol Out é ce 8 06 9 v9 68/6/11 (2) (€2) (2) (21) (1) (8) (2) (21) (0) (8) cl OTT Ol 26 I UT ow 08 é be v8/81/6 (21) (26) (2) (19) (€) (S2) (2) (0S) (S) (6€) be 09¢ Se 16 p Te Te 09T 6 69 b8/12/8 (C) (61) (€) (S2) (1) (b) (€) (12) (2) (v1) GE 0£2 4 col c BI 14 bbl ¥¥ QT ¥89 bR/S2/L (eyo) O9el+bScl 092T bSel cbcl 97ed 40} 904 40{ 904 40| 30.y But yduwes "Q°S + SUOL}e4qUaDUD) Gg UeaW Seta “(NAST IM) A248 adUauasau punos pue;sy 6uo7 usaqsay ye pakojdap (SL{Npa snitqAyy) stassnu ut suoLzeszuadu0d Gd Jo Axewwns *9 alqey 66 9SSIudXd » (8) (2S) (S) (8€) (1) (2) (p) (££) (€) (61) 8b 5 3 ce aa 9 le le 96T Ll Ill G8/TI/L (pT) (29) (6) (6€) (€) (pT) (9) (2) (S) (2) 0S 692 62 9SI 9 I€ €2 bel Ie a G8/22/S {e301 O9cI+bSel 092T bSel cbel a3eg * JO] 904 "O°S + SU0Lze4zUaDU0} gjq UraW a eee ee ee *panutyuos “*g alqey 67 (€1) (99) (11) (8S) (€2) (S21) (21) (29) (2) (OT) 9S Cute l€ a LT 96 vb 8/ Se BET G8/S2/b (6) (96) (>) (bb) (1) (€1) (€) (1£) (S) (€S) Le €Se ST Sbl c O€ cl SII Il BOT G8/92/€ (1) (81) (1) (ZT) (€) (2b) (€) (SE) (1) (8) a Bol 9 08 € cb c RE S 89 SR/ 11/2 (9) (b9) (S) (pS) (S) (bS) (0) (0) (2) (12) at SLI eT Tel eT Tel 0 0 p bb 08/82/21 (2) (69) (9) (0S) (6) (26) (pv) (6€) (2) (ST) LI va cl BIT l tl p bb S 9S vB/T1/21 (>) (€€) (2) (ZT) (€) (2€) (€) (SE) (2) (02) eT Sb 8 SR 9 $9 ; é 02 9 09 68/6/11 (21) (19) (5) (Sb) (1) (L) (8) (6€) (€) (9T) 4 bbl ST Ul é a eT 86 ) RE b8/81/6 (S) (/£) (S) (22) (0) (0) (>) (22) (2) (21) cet SU be O9T c 02 I2 Ov R SS b8/1c/8 (01) (89) (8) (>S) (1) (OT) (9) (bb) (2) (v1) 62 002 Td 6b é Sif 6 cel val ¥1S bR/G2/L {e101 092T+bS2l 09¢l bSet chal 97eq 40} 904 Buty dwe °Q°S + UOLJeuyZU|aDUO) gq UeaW SSS ee Se ES ee ee ee eee "(9ST1M) punow g 4aquag Punos pue|sy buoy usaqzsay ye pakoydap (St[npa SN[ LAW) SLassnw ut suotzeazuacuod gg 4o Axeuuns "f apqey 68 nes +_7M 7am 6/bu ut passaudxa yy “0°S + IM Aup 6/6u ut passaudxa y (€2) (ObT) (62) (b9) (Sb) (ve) (€2) (0£) (S) (12) OvT Ovs SIT bOb C82 SG bol 681 Le 9ET §8/12/9 (TE) (881) (TE) (€81) (bE) (961) (02) (TIT) (2) (01) OOT 12S 69 Che 9b b92 EG 621 le 8/1 GA/ 22/5 (6) (Zb) (OT) (SS) (02) (E11) (11) (6S) (2) (8) 8b 1Z2 Le €St ST S8 al 89 I2 RIT §8/92/b (2) (2b) (>) (02) (1) (€) (€) (81) (€) (22) O02 89T el ell c te Ot 06 9 SS GR/92/€ (1) (92) (2) (82) (b) (1S) (2) (92) (1) (81) Ul Let l S8 9 OZ l ST D eb S8/11/2 (pb) (pb) (1) (01) (1) (01) (0) (0) (€) (9€) ST Ol2 or Ovl OT Obl 0 0 S OL bB8/11/el (S) (b2) (1) (82) (S) (2S) (9) (69) (b) (6b) 61 lod Il tcl 8 £8 e Ov l RL bB/5/ 11 (8) (pS) (2) (pS) (1) (8) (9) (9b) (1) (01) 02 Cll SI 621 4 91 eT etl S bb bB/R1/6 (p) (22) (€) (02) (0) (0) (€) (02) (1) (2) le 002 I2 est e 02 8T cet 9 Lb b8/12/8 (8) (Tb) (9) (bE) (1) (b) (S) (0€) (2) (18) eC 9€¢ be bét é 81 ce 9S ¥¥6 ¥¢09 p8/S2/L [P30] O92 +bSel 09cI bSel chet a7e0 40) 04y 40] D0uy Bur; due "0°S + SU0L}e4QUaDU0) Gq UeaW ° punos pues] buoy usaysay ut (MWo0S) Punow g JO 359M s4azauM OOG 3& pakoidap (Stinpa SN[LQAW) SLassnw ut suotze1quaou0s 9d JO Auewwns “g ayqey, 69 "O°S +_JyBLam yam 6/6u ut passaudxa yyy "0°S + WUuBtam Aap 6/6u wt passaudxa yy auL[aseg » tt Tae (82) (8p) (91) (82) (€) (pb) (ST) (S2) (21) 091 262 16 69T Of 8T €8 IST 69 GR/L2/9 (S) (SE) (€) (22) (€) (22) (0) (0) (2) be a 4 LT Set LT Get 0 0 g G8/be/S (6S) (08) (/€) (SS) (8b) (9b) (ST) (6) (71) bee c8b Tee Ice L8¢ 082 88 1S bor GB/TT/b (21) (SE) (2) (81) (21) (ST) (S) (€) (S) Cl 602 cb LO él 68 {€ 8T vue lf ¥58/92/£ eyo) O92I+bS2l bS2l 33eg 40] 304 Hut dure "0°S + SU0L}eu{UaDUO) QJq UeAW a ee ENS Sh “(4LW1) 8741S aduasayay ssauiye7 ye pakoidap (stinpa SNL LAW) S_assnw ut suotzeuquazuod Id JO Aaewuns 6 ayqet 70