Imaging Plate Autoradiography for Ingested Anthropogenic Cesium-137 in Butterfly Bodies: Implications for the Biological Impacts of the Fukushima Nuclear Accident

Abstract

The Fukushima nuclear accident in March 2011 caused biological impacts on the pale grass blue butterfly Zizeeria maha. At least some of the impacts are likely mediated by the host plant, resulting in "field effects". However, to obtain the whole picture of the impacts, direct exposure effects should also be evaluated. Here, we examined the distribution of experimentally ingested anthropogenic cesium-137 (¹³⁷Cs) in adult butterfly bodies using imaging plate autoradiography. We showed that ¹³⁷Cs ingested by larvae was incorporated into adult bodies and was biased to females, although the majority of ingested ¹³⁷Cs was excreted in the pupal cuticle and excretory material during eclosion. ¹³⁷Cs accumulation in adult bodies was the highest in the abdomen, followed by the thorax and other organs. These results suggest that ¹³⁷Cs accumulation in reproductive organs may cause adverse transgenerational or maternal effects mediated by reactive oxygen species (ROS) on germ cells. ¹³⁷Cs accumulation was detected in field individuals collected in September 2011 and September 2016 but not in May 2011, which is consistent with the abnormality dynamics known from previous studies. Taken together, these results contribute to an integrative understanding of the multifaceted biological effects of the Fukushima nuclear accident in the field.

Keywords: Fukushima nuclear accident; Zizeeria maha; cesium-137; exposure effect; field effect; imaging plate autoradiography; pale grass blue butterfly; radiation dose; transgenerational effect.

Figure 1

Imaging plate autoradiography of the pale grass blue butterfly Zizeeria maha. Larvae were fed a ¹³⁷Cs-containing artificial diet, and whole-body adults, pupal cuticle cases, whole-body pupae, and adult body parts were subjected to this analysis. (a) Examples of layout on an imaging plate. In the three panels of adults, pupal cuticle, and pupae, samples were aligned according to the ¹³⁷Cs radioactivity groups (the ingestion levels at the larval stage), H0–H6. All samples except pupae shown here are females. Pupal sex is unknown. Scale bars, 10 mm. (b) Examples of the imaging plate images of H6 samples of a whole-body adult, a pupal cuticle, a pupa, and adult body parts. In these examples, the pupal cuticle and pupa were exposed for 2 d (two days), and others were exposed for 6 d. Image contrast was adjusted to clarify the ¹³⁷Cs gradient. Red areas indicate the highest PSL value.

Figure 2

Radioactivity levels of adult whole-body samples (n = 3 at each exposure). (a) Radioactivity after 1-d or 2-d exposure of H6 female samples. (b) Radioactivity of 1-d to 8-d exposures of H6 male and female samples. The mean values of three measurements at each exposure time are plotted. Both male and female samples show a Spearman correlation coefficient ρ = 1 (p < 0.001). (c) Radioactivity of exposure of H0–H6 samples. Mean values of three measurements at each ¹³⁷Cs radioactivity group (ingestion level at the larval stage) are plotted. Note that both axes are logarithmic scales. Both male and female samples show a Spearman correlation coefficient ρ = 1 (p = 0.02).

Figure 3

Weight-adjusted radioactivity levels in adult whole-body samples. Blue and pink bars indicate males and females, respectively. Asterisks indicate statistical significance: **, p < 0.01 (Welch’s t-test). Error bars indicate standard deviation. (a) Single logarithmic expression. Note that the x-axis is logarithmic due to tenfold changes in ingestion levels at the larval stage. Inset shows H0–H4 at different scales of the y-axis. (b) Double logarithmic expression. Data are identical to (a). Only H4 was significantly different between sexes. (c) Virtual samples using H3–H6 samples (excluding H0) adjusted to the H3 levels (n = 12 for each sex).

Figure 4

Weight-adjusted radioactivity levels in pupal cuticle (exuviae) samples. Dark blue and orange bars indicate males and females, respectively. (a) Single logarithmic expression. Note that the x-axis is logarithmic due to tenfold changes in ingestion levels at the larval stage. Inset shows H0–H4 at different scales of the y-axis. Error bars indicate standard deviation. (b) Double logarithmic expression. Data are identical to (a). H4 (having a minimum number of samples required for t-test) was not significantly different between sexes. (c) Virtual samples using H3–H6 samples (excluding H0) adjusted to the H3 levels (n = 5 for males and n = 11 for females; one female H5 sample was discarded as an outlier). For comparison, adult virtual samples are also shown. Asterisks indicate statistical significance: **, p < 0.01 (Welch’s t-test). (d) Radioactivity ratio (pupal cuticle divided by adult whole body) using virtual samples.

Figure 5

Weight-adjusted radioactivity levels in pupal whole-body samples. (a) Single logarithmic expression. Note that the x-axis is logarithmic due to tenfold changes in ingestion levels at the larval stage. Inset shows H0–H4 at different scales of the y-axis. Error bars indicate standard deviation. (b) Double logarithmic expression. Data are identical to (a). (c) Comparison among the adult whole body, pupal cuticle, and pupa. Mean values of male and female mean values of adjusted data at the H3 level were used for this comparison. (d) ¹³⁷Cs distribution in a pupa at different radiation levels (H3–H5). Mean values of male and female mean values were used for this graph. H6 was excluded because of a lack of male pupal cuticle samples.

Figure 6

Radioactivity levels in six body parts of the H6 samples. (a) PSL values. Males (left) and females (right). (b) Wings. Four dots (two rectangular and two triangular dots) are shaded together in females, indicating that they are from the same individual. (c) Weight-adjusted PSL values. Males (left) and females (right). (d) Ratios of the weight-adjusted female PSL values to the weight-adjusted male PSL values. (e) Relative levels among the six body parts examined in the present study. These three samples had all body parts.

Figure 7

Comparison of field samples. The percentages of positive individuals with significantly larger PSL values than those of the control group are shown. Dark blue and orange bars indicate males and females, respectively. Asterisks indicate statistical significance: *, p < 0.05; **, p < 0.01 (Fisher’s exact test). (a) Comparisons between sampling dates using all samples (males and females together). (b) Comparisons between sampling dates using male (left) or female (right) samples. (c) Comparisons between male and female samples on the same sampling dates.