Endocrine disruption and differential gene expression in sentinel fish on St. Lawrence Island, Alaska: Health implications for indigenous residents

Abstract

People living a subsistence lifestyle in the Arctic are highly exposed to persistent organic pollutants, including polychlorinated biphenyls (PCBs). Formerly Used Defense (FUD) sites are point sources of PCB pollution; the Arctic contains thousands of FUD sites, many co-located with indigenous villages. We investigated PCB profiles and biological effects in freshwater fish (Alaska blackfish [Dallia pectoralis] and ninespine stickleback [Pungitius pungitius]) living upstream and downstream of the Northeast Cape FUD site on St. Lawrence Island in the Bering Sea. Despite extensive site remediation, fish remained contaminated with PCBs. Vitellogenin concentrations in males indicated exposure to estrogenic contaminants, and some fish were hypothyroid. Downstream fish showed altered DNA methylation in gonads and altered gene expression related to DNA replication, response to DNA damage, and cell signaling. This study demonstrates that, even after site remediation, contaminants from Cold War FUD sites in remote regions of the Arctic remain a potential health threat to local residents - in this case, Yupik people who had no influence over site selection and use by the United States military.

Keywords: Bering Sea; FUD site; Formerly used defense site; Global distillation; Polychlorinated biphenyls; Yupik.

Figure 1

Total PCB concentrations (per wet weight) of ninespine stickleback and Alaska blackfish at study sites along the Suqitughneq River at Northeast Cape, St. Lawrence Island (see Fig. S1 for a remote image of the sites). Sites 13 and 14 are located upstream of the FUD site, while all other sites are downstream of it. Site 12 is at the confluence of the Suqitughneq River and the streamlet draining the former barrel-storage area and main operations center. Blackfish were not caught at Sites 2–5 and stickleback were not caught at Sites 10, 12, and 14 during the sampling period represented by these data (2012–2013). Site was a significant predictor of total [PCB] for blackfish (ANOVA F_8,30 = 2.468, p = 0.0347) but not for stickleback. Because data were not normally distributed, and individual sites had small sample sizes, we also analyzed the data with the non-parametric Kruskal-Wallis test. Again, site was a significant predictor of total [PCB] for blackfish (KW = 26.62, p = 0.0008). Blackfish collected at site 12 had significantly higher total [PCB] than blackfish collected at sites 13 and 14 (Dunn’s multiple comparisons test, p = 0.0023 and 0.0112, respectively). Additionally, blackfish collected at site 9 had significantly higher total [PCB] than blackfish collected at site 13 (p = 0.0383).

Figure 2

Difference in [PCB] for each PCB class (based on the number of chlorines in the molecule, and arranged along the horizontal axis from one to ten chlorines) between fish collected downstream vs. upstream of the FUD site at Northeast Cape, St. Lawrence Island for both ninespine stickleback (Panel A; n = 5 upstream fish and 42 downstream fish) and Alaska blackfish (Panel B; n = 10 upstream fish and 29 downstream fish). The concentration of lighter PCBs (those containing three or fewer chlorines) were not significantly higher or lower in upstream vs. downstream sites, a finding consistent with atmospheric deposition of these congeners. For blackfish, concentrations of all PCB classes with four or more chlorine atoms, as well as total PCBs, were significantly higher in fish collected downstream of the FUD site, suggesting that these congeners originated at the site (Mann-Whitney U tests, U = 0–4, p = 0.001–0.004; p values Bonferroni corrected; all statistics presented in Suppl. Table 1). For stickleback, concentration differences were significant only for PCBs containing 5, 9 or 10 chlorines (Mann-Whitney U tests, U = 7–27.5, p = 0.013 – 0.046; p values Bonferroni corrected; all statistics presented in Suppl. Table 1). Heavy PCB congeners, especially those containing 8 or more chlorines, are not readily transported in the atmosphere (Wania and Mackay, 1996) and therefore their presence indicates a local source.

Figure 2

Difference in [PCB] for each PCB class (based on the number of chlorines in the molecule, and arranged along the horizontal axis from one to ten chlorines) between fish collected downstream vs. upstream of the FUD site at Northeast Cape, St. Lawrence Island for both ninespine stickleback (Panel A; n = 5 upstream fish and 42 downstream fish) and Alaska blackfish (Panel B; n = 10 upstream fish and 29 downstream fish). The concentration of lighter PCBs (those containing three or fewer chlorines) were not significantly higher or lower in upstream vs. downstream sites, a finding consistent with atmospheric deposition of these congeners. For blackfish, concentrations of all PCB classes with four or more chlorine atoms, as well as total PCBs, were significantly higher in fish collected downstream of the FUD site, suggesting that these congeners originated at the site (Mann-Whitney U tests, U = 0–4, p = 0.001–0.004; p values Bonferroni corrected; all statistics presented in Suppl. Table 1). For stickleback, concentration differences were significant only for PCBs containing 5, 9 or 10 chlorines (Mann-Whitney U tests, U = 7–27.5, p = 0.013 – 0.046; p values Bonferroni corrected; all statistics presented in Suppl. Table 1). Heavy PCB congeners, especially those containing 8 or more chlorines, are not readily transported in the atmosphere (Wania and Mackay, 1996) and therefore their presence indicates a local source.

Figure 3

Mean vitellogenin (VTG) concentrations of male ninespine stickleback collected from Suqitughneq River (Suqi) sites 2, 4, 5, 6, 7, 11 and 13, from the Tapisaggak River (Tapi; TP) located east of the Suqitughneq River in a separate drainage, and from Troutman Lake (TL) sites 1, 2 and 3 in the village of Gambell on the western side of St. Lawrence Island. Suqitughneq sites 2–11 are located downstream of the FUD site, while Suqitughneq site 13 is located upstream. Genotypic sex was confirmed using sex-specific primers in a PCR assay. Many males in the Suqitughneq River are abnormally induced for VTG production, indicating exposure to estrogenic contaminants. The frequency of VTG induction in Suqitughneq River males (induced vs. not induced) is significantly higher than in males from the Tapisaggak River or Troutman Lake (chi-square test, χ² = 7.01, df = 2, p < 0.05). Comparing only males with non-zero values of VTG (only induced males), concentrations are significantly higher in Suqitughneq River males than in combined males from the Tapisaggak River and Troutman Lake (Welch-corrected t = 3.368, df = 38.53, p = 0.0017). Given a lack of homogeneity of variance, we also compared VTG concentrations in induced Suqitughneq River males with induced males from the other two sites using the non-parametric Mann-Whitney U test, and results were similar (U = 109, p = 0.0246). Variation in male VTG concentration is high in the Suqitughneq River, likely because fish move around between more and less contaminated reaches. To the best of our knowledge, these are the first VTG data ever collected for ninespine stickleback in the wild.

Figure 4

Thyroid hormone (thyroxine; T₄) concentrations in whole-body homogenates of ninespine stickleback collected in the Suqitughneq River sites 2, 4, 5, 6, 7, 11 and 13, in the Tapisaggak River (Tapi, TP) located east of the Suqitughneq River in a separate drainage, and in Troutman Lake (TL) in the village of Gambell on the western side of St. Lawrence Island. T₄ concentration significantly differed by site (ANOVA, F_8,275 = 18.29, p < 0.001), with stickleback collected at Suqitughneq River sites 2, 4 and 7 having significantly lower T₄ concentrations than stickleback collected at other Suqitughneq River sites or in the Tapisaggak River or Troutman Lake (Tukey’s post hoc test, indicated by letters over SE error bars showing which groups significantly differ from each other).

Figure 5

Percent total methylation of gonad, kidney, liver, spleen and thyroid of ninespine stickleback collected upstream (n = 12) vs. downstream (n = 12) of the FUD site in the Suqitughneq River at Northeast Cape, St. Lawrence Island (sample size exceptions: downstream gonad n = 9, upstream spleen n = 11, downstream spleen n = 9, downstream thyroid n = 11). Only gonads differed significantly between upstream and downstream fish (Mann Whitney U test, U = 23, p = 0.0278). Males and females showed the same pattern for gonads, but were analyzed together because of low sample size.