Environ. Sci. Technol. 1997, 31, 3035-3036
Response to Comment on “Elevated Accumulation of Tributyltin and its Breakdown Products in Bottlenose Dolphins (Tursiops truncatus) Found Stranded along the U.S. Atlantic and Gulf Coasts” SIR: The intent of our study, as stated in ref 1, was to determine contamination levels and tissue distribution of butyltin compounds in dolphins stranded along the U.S. Atlantic and Gulf Coasts. Tissues of bottlenose dolphins were taken from those animals stranded dead; a few dolphins were from mass mortalities that occurred in 1990. These individuals afforded an opportunity to examine total butyltin (BTs ) MBT + DBT + TBT) concentrations in diseased animals, which were compared with the data available for marine mammals from different regions. Studies linking chemical contaminants and disease in dolphins were correlative, and in our opinion, no conclusive evidence exists as yet. Our statement in conclusions (1) imply that butyltin compounds, in addition to several other stressors, should also be considered as a class contaminants that may have contributed to immune suppression in dolphins. This has been stated as “greater accumulation of butyltin compounds may also have contributed to the immunedysfunction in dolphins” and “further studies are needed to establish the relationship ...”. We have neither stated that this study was designed to establish an empirical relationship between butyltin compounds and dolphin mortalities nor have we concluded that TBT is the cause of such mortalities. Green and his co-workers have ramified and misinterpreted our results. It is worth emphasizing that our studies, including the present one, have provided pioneering evidence that dolphins do accumulate considerable levels of butyltin compounds, particularly in their liver and kidney (1-12). Considering this evidence and toxicological studies, which unequivocally documented on the immune suppressing potential of BTs (13), we believe that BTs may also have contributed to immune suppression in dolphins. We have exercised adequate caution in interpreting our data, and our conclusions are adequately substantiated by suitable references. The authors of the preceding polemic stated that “a closer look at their data interpretations suggest ...”. The following clarifications will reveal that they have ignored or failed to understand several important concepts and basic factors that influence BT accumulation in marine mammals. The dolphin that contained a total BT concentration of 11 340 ng/g has been claimed as an outlier by Green and his co-workers without examining nature of the sample. This animal was collected in 1989, when TBT use was recently restricted (restriction on TBT use was implemented in 19881989), while the others were collected in later years. Examination of liver, kidney, and blubber of this animal, by three independent analyzers, invariably revealed the presence of BT concentrations as mentioned, which could be explained by increased exposure when TBT use was in effect. The BT concentration measured in this animal provided evidence that the dolphins stranded in the 1980s (period of maximum use of TBT) may have experienced greater exposure to BTs. We did not have more dolphins collected in the 1980s when this manuscript was accepted for publication. However, later investigations with a few more dolphins collected in 19881989 showed that total BT concentrations (8000-11 000 ng/g wet wt) in liver were similar to those measured in the 1989 individual. It is an outlier when compared with animals
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1997 American Chemical Society
collected in later years, but it belongs to a population exposed to elevated levels of BTs during the periods of maximum use. Due to the difficulties in obtaining a large number of dolphins with similar biological parameters, we did not design our study for statistical analysis. Green et al. assert that our paper (1) does not make clear “whether the sampled animals died from the same causes as the animals involved with the mass mortality event”. We have mentioned in the Introduction and Materials and Methods that the dolphins were stranded along the U.S. Atlantic coast during 1989-1994. Several references cited in our Introduction describe mass mortalities or stranding events, including causes, symptoms, and the possible link between contaminants/stressors and dolphin deaths. Regardless of whether the samples originated from mass mortalities in 1987-1988 or from strandings in later years, the dolphins were diseased (infected by microorganisms), immunologically suppressed, and contained elevated concentrations of PCBs in tissues. Dolphin deaths were linked to immune suppression because of exposure to various stressors including chemical contaminants. These details were the subject of several earlier investigations, which are referred to in our Introduction. The scope of our investigation is to examine the accumulation of butyltin compounds in dolphins, not to review all those facts about dolphin mortalities. If Green et al. had followed the literature on dolphin strandings, they would have been clear about the basic concepts and thoughts of our research. Green et al. have ignored two important variables, sampling period and age of dolphins, in their comparison of concentrations of BTs in U.S. dolphins with those of our earlier studies on dolphins from the Mediterranean Sea and Japanese coastal waters (2-4). When the dolphins of similar age collected in a similar year were compared, BT concentrations were similar or higher for the U.S. dolphins. The concentration measured for the U.S. dolphin collected in 1989 is the greatest value reported, even when compared with those of the cetaceans collected in 1980s from Japanese coastal waters. In any case, in order to avoid such misinterpretations, we have made corrections to our sentence as follows: “In general, the range of BT concentrations in bottlenose dolphins from the U.S. Atlantic coast was comparable or higher than those of the diseased Mediterranean dolphins (17) and finless porpoise (Neophocaena phocaenoides) and Risso’s dolphins (Grampus griseus) from Japanese coastal waters (18)”. As stated in Materials and Methods (1), the age groups of dolphins analyzed in this study were subjective, which were based on length estimates. The teeth of the dolphins were not available for actual age determination. Based on our classification, calves included those animals that ranged in length from 120 to 190 cm (a few months to 2 yr). Green et al. have compared BT concentrations in two male bottlenose dolphin calves with those of two Atlantic spotted dolphins and ignored that the mean length of bottlenose dolphin was 148 cm, whereas that of the Atlantic spotted dolphin was 169 cm. Thus, the bottlenose dolphins were younger than the spotted dolphins if the same growth curve was assumed for both species (as Green et al. did). Despite this, the bottlenose calves had greater BT concentrations than the spotted dolphins. Studies on BTs in dolphins from different parts of the world, including this study, showed that there is no significant difference in concentrations between sexes (512). Based on this, the mean concentration of butyltins in bottlenose dolphin calves (n ) 5; 650 ng/g) was about 2-fold greater than those of spotted dolphins (n ) 2; 350 ng/g), although the former were younger than the latter. The overall mean concentration, as presented in our Table 1, was 3-4fold less for off-shore species, which is consistent with those
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observed in other studies (4, 5, 8-12). Green et al. have neglected the basic factors that influence the magnitude of BT concentrations in marine mammals while making their comparisons. Green et al. claim that BT concentrations in captive bottlenose dolphin were 1.4-145-fold less those in freeranging dolphins and when the outlier is removed the difference is 1.4-32-fold. Again, they have failed to select a similar age group of dolphins while making their comparison. The captive animal is an adult female (21 yr), and it is inappropriate to compare with several “free-ranging” neonatal (i.e., newborn) dolphins to evaluate the magnitude of difference. Our comparison of mean concentration of BTs in livers of adult free-ranging bottlenose dolphins showed 38-fold greater concentration than those of captive animal, while the highest concentration was 145-fold greater (adult dolphin collected in 1989). In order to avoid misinterpretations, we have made corrections to the sentence as follows: “Total butyltin concentrations in the liver, kidney, and muscle of a captive adult female bottlenose dolphin were lower than the concentrations found in free-ranging dolphins”. Furthermore, Green et al. have assumed that the captive dolphin has spent 8 yr in the Mississippi Sound. It is not mentioned in the manuscript except the fact that the dolphin was collected in Mississippi Sound in 1978. It was neonatal when collected. Even if their baseless assumption that the animal has spent 8 yr in Mississippi Sound is considered, BTs could be eliminated during 13 yr of captivity in later years. The y-axes of Figures 4 and 5 have been multiplied by 5 prior to plotting. Since it was not mentioned in captions, we have added this statement later. This would answer Green et al.’s concern about the range of data plotted in these figures. We did not make any quantitative interpretations based on these figures, and all discussions remain correct. Green et al.’s concern about the data for Japanese cetaceans showing the values from 30 to 550 ng/g are for Dall’s porpoise, dwarf sperm whale, and ginko-toothed whale. These animals were specifically analyzed for our comparison because refs 16 and 18 of ref 1 did not include whales or off-shore animals, whereas the study on U.S. marine mammals included pygmy sperm whale and spotted dolphins. Inclusion of data for whales from Japan has made our comparison between U.S. and Japanese cetaceans more accurate. We have indicated in the text about inclusion of Dall’s porpoise, dwarf sperm whale, and ginko-toothed whale, and a manuscript on BTs in offshore dolphins has been submitted for publication (12). Green et al. claim that the attention accorded to total butyltins is questionable because mono-(MBT) and dibutyltin (DBT) are less toxic. In fact, they eliminated the concentrations of MBT and DBT from data while comparing the concentrations with Japanese cetaceans. First, they ignore the fact that these dolphins have been exposed primarily to TBT, and MBT and DBT are the breakdown products of TBT. Breakdown of TBT to MBT and DBT should not be interpreted as the toxic effect of BTs being insignificant or questionable. Secondly, they fail to know that DBT is also a potential immunotoxicant (ref 13 and references cited therein). Thirdly, they disregard various factors that influence the proportion of individual butyltin compounds in total BT concentrations (e.g., metabolic potential, chronology of exposure, decomposition of carcass, storage of samples, etc.). We object to Green et al.’s highly improbable and invalid assumption of neglecting MBT and DBT concentrations while assessing the risks of butyltin compounds. They show that, by neglecting MBT and DBT values, Japanese dolphins had greater TBT concentrations. They have overlooked the fact that the Japanese animals were collected during the periods of maximum usage of TBT. Thus, fresh inputs resulted in greater TBT proportions in the total BT concentrations. Furthermore, their estimate of 100 ng/g TBT for bottlenose dolphins is inaccurate, as they had included captive as well as off-shore
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animals. The value 960 in our Table 3 should be 690 (typographical error). This has not affected any of our interpretations or discussions. Green et al. claim that our study does not contain any histopathological, toxicological, or epidemiological analysis. Our objective was to examine the concentrations and accumulation feature of BTs in dolphins. During the course of this study, circumstantially, we found that the concentrations BTs were elevated when compared with those of the dolphins from other areas. This led us to speculate on the possible toxic effects of BTs in dolphins. As we have stated in our concluding remark, it is the next step to conduct histopathological and toxicological studies to establish etiological relationship, if any, and to evaluate threshold concentrations of BTs in dolphins. It is informative to note that our laboratory at the Department of Environment Conservation, Ehime University, has made extensive studies on butyltin contamination in marine mammals, which has provided us an opportunity to understand various factors that influence the accumulation of BTs in marine mammals. Our interpretations and discussions are based on our experience and knowledge gained from these investigations on butyltin compounds in marine mammals from different regions in the world (Pacific Ocean including the Bering Sea and the Gulf of Alaska, Mediterranean Sea, Baltic Sea, Asian coasts and Indian Ocean, Black Sea, Caspian Sea, Baikal Lake and the Atlantic and Gulf coasts of the U.S., Antarctic Ocean). While some of these results have been published (1-10), a few others are in preparation for publication. Green et al.’s argument against the role of BTs in immune suppression in dolphins is plagued by the lack of any evidence.
Literature Cited (1) Kannan, K.; Senthilkumar, K.; Loganathan, B. G.; Takahashi, S.; Odell, D. K.; Tanabe, S. Environ. Sci. Technol. 1997, 31, 296301. (2) Kannan, K.; Corsolini, S.; Focardi, S.; Tanabe, S.; Tatsukawa, R. Arch. Environ. Contam. Toxicol. 1996, 31, 19-23. (3) Iwata, H.; Tanabe, S.; Mizuno, T.; Tatsukawa, R. Environ. Sci. Technol. 1995, 29, 2959-2962. (4) Kim, G. B.; Tanabe, S.; Iwakiri, R.; Tatsukawa, R.; Amano, M.; Miyazaki, N.; Tanaka, H. Environ. Sci. Technol. 1996, 30, 26202625. (5) Kim, G. B.; Tanabe, S.; Tatsukawa, R.; Loughlin, T. R.; Shimazaki, K. Environ. Toxicol. Chem. 1996, 15, 2043-2048. (6) Kannan, K.; Senthilkumar, K.; Sinha, R. K. Appl. Organomet. Chem. 1997, 11, 223-230. (7) Kannan, K.; Falandysz, J. Mar. Pollut. Bull. 1997, 34, 203-207. (8) Iwata, H.; Tanabe, S.; Miyazaki, N.; Tatsukawa, R. Mar. Pollut. Bull. 1994, 28, 607-612. (9) Kim, G. B.; Lee, J. S.; Tanabe, S.; Iwata, H.; Tatsukawa, R.; Shimazaki, K. Mar. Pollut. Bull. 1996, 32, 558-563. (10) Iwata, H.; Tanabe, S.; Mizuno, T.; Tatsukawa, R. Appl. Organomet. Chem. 1997, 11, 257-264. (11) Madhusree, B.; Tanabe, S.; Amaha, A. O.; Tatsukawa, R.; Miyazaki, N.; Ozdamar, E.; Aral, O.; Samsun, O.; Ozturk, B. Fresenius Anal. Chem. (in press). (12) Tanabe, S.; Prudente, M.; Mizuno, T.; Hasegawa, J.; Iwata, H.; Miyazaki, N. Environ. Sci. Technol. (submitted for publication). (13) Fent, K. Crit. Rev. Toxicol. 1996, 26, 1-117.
Kurunthachalam Kannan* 201 Pesticide Research Center Michigan State University East Lansing, Michigan 48824
Shinsuke Tanabe Department of Environment Conservation Ehime University Tarumi 3-5-7 Matsuyama 790, Japan ES972007V
