AN ABSTRACT OF THE THESIS OF
David A. Hermann for the degree of Master of Science in Radiation Health Physics presented on April 24, 2020.
Title: Butterfly Puddling: An Alternative Cs-137 Pathway to the Pale Grass Blue Butterfly Following the Fukushima Nuclear Accident
Abstract approved: ______Kathryn A. Higley
Environmental modeling as a result of the Fukushima accident has been at the forefront for present day health physics. The accident released radionuclides in the environment and the fate and transport of these radionuclides are of interest when considering doses to non-human biota. The Pale Grass Blue Butterfly was studied following the accident due to previous research and its abundance in rural and urban areas. The most studied pathways of exposure for the Pale
Grass Blue Butterfly were the traditional ingestion and external pathways, but another potential exposure pathway is through the behavior of butterfly puddling. Butterfly puddling is a behavior of adult male butterflies to congregate around puddles to drink large amounts of water in order to concentrate the puddles dissolved nutrients, primarily sodium. The fluid passes through the males to adjust for low diet concentrations of sodium from plants. The concentrated sodium is then transferred to females as gifts during copulation and distributed to their eggs. As cesium and potassium have similar chemical properties because both are alkali metals, radiocesium may permeate biological sodium channels due to the same properties. This paper will entail a literature review of this concept, with a proposed butterfly egg dose to determine if it may have caused the researchers unconventional results.
©Copyright by David A. Hermann April 24, 2020 All Rights Reserved Butterfly Puddling: An Alternative Cs-137 Pathway to the Pale Grass Blue Butterfly Following the Fukushima Nuclear Accident
by David A. Hermann
A THESIS
submitted to
Oregon State University
in partial fulfillment of the requirements for the degree of
Master of Science
Presented April 24, 2020 Commencement June 2020 Master of Science thesis of David A. Hermann presented on April 24, 2020
APPROVED:
Major Professor, representing Radiation Health Physics
School of Nuclear Science and Engineering
Dean of the Graduate School
I understand that my thesis will become part of the permanent collection of Oregon State University libraries. My signature below authorizes release of my thesis to any reader upon request.
David A. Hermann, Author ACKNOWLEDGEMENTS
The author expresses sincere appreciation to the United States Army for the opportunity to attend
Oregon State University as part of the Army Medical Department’s Long-Term Health
Education and Training program. My sincere appreciation to Dr Kathryn A. Higley for providing exceptional academic advisement, and the Radioecology research group for providing additional guidance and research input.
CONTRIBUTION OF AUTHORS
Contributions include the OSU NSE Radioecology Research group, with Dr. Higley serving as academic advisor and group leader.
TABLE OF CONTENTS
Page
1 Introduction...……………………………………………………………………………………1
2 Literature Review.……………….………………………………………………………………3
2.1 Fukushima Nuclear Accident & Cs-137 Deposition……………………..……………….…3
2.2 Cesium-137………..………………………………………..…………………..……………5
2.2.1 Cesium-137 Radioactive Transformation and Decay…………...….…..…………..….5
2.2.2 Cs-137 Dose………………………….……………….……………..………..………11
2.2.3 Cs-137 Uptake………………………………………...……...... ….……….…………20
2.3 Pale Grass Blue Butterfly………...………………...... ………………...………25
2.3.1 Species…………….……………….…….…………………………………………...25
2.3.2 Anatomy and Lifecycle…………………….…………………………………………25
2.3.3 Lepidoptera Anatomy and Lifecycle to Irradiation…………………………………...29
2.3.4 Reproduction…………...…………………….……………………………………….30
2.3.5 Habitat ………………………………………………………….…………………….31
2.3.6 Butterfly Puddling Behavior………………………………………………………….32
2.3.7 Environmental Studies…………...……………………...……….………..………….37
2.3.7.1 Environmental Indicator Species.…………………………………….………37
2.3.7.2 Environmental Study Complexities……………………………………….….39
3 Materials & Methods.……….…………………..…...…………………………………………47
3.1 Internal Absorbed Dose……………………………………………………………………..47
3.2 External Absorbed Dose…………………………...………………………………………..48
TABLE OF CONTENTS (Continued)
Page
4 Results …………………………………………………………………………………………51
4.1 Internal Absorbed Dose Due to Puddling Behavior………………………………………..51
4.2 External Absorbed Dose due to Cs-137 Dispersion………………………………………..53
4.3 Internal + External Absorbed Dose………………………………………………………...56
5 Discussion ……………………………………………………………………………………..57
6 Conclusion.……………………………...….…………………………………………………..65
References.………………………………………………………………………...……………..66
Appendices.………………………………………………………………………………………78
LIST OF FIGURES
Figure Page
2.1 Cs-137 Decay ……………...………………………………………..…….…...……………...9
2.2 Oxalis corniculata Plant………………………………………………………………………32
2.3 Scanning Electron Microscope of Probiscus………………………………………….……...33
5.1 ICRP 30 GI Model vs Proposed Lepidoptera Cs/Na Ratio Biokinetic Uptake Model……….58
LIST OF TABLES
Table Page
2.1 Biological and Environmental Half-Life of 137Cs………………………..…………………..7
2.2 Cs-137 Average and Maximum Energy……………………………………………………….8
2.3 Absorbed Fractions for Uniform Distribution of Activity in Small Spheres and Thick Ellipsoids………………………………………………………………………….14
2.4 Absorbed Fractions for Energies and Sphere Sizes for Electrons……………………………15
2.5 Decay Data for for 137Cs…………………...…………………………………………...…....17
2.6 Decay Data for 137mBa…………………………………………………….………………….17
2.7 MIRD vs ICRP Methods for Dose…………………………………..……………………….19
2.8 Atomic and Ionic Radii of Alkali Metals and Monovalent Cations…………………………22
2.9 Permeability Ratios for Metal Cations in Sodium Channels…………………………….…...23
2.10 Radiotolerances of different stages of Lepidoptera………………………………………....29
2.11 Biological impacts of radiation conducted since the Fukushima accident……..…………...41
2.12 Typical attributes of male lepidopteran insects (and their progeny) receiving substerilizing doses of radiation………………………………………………………………….42
4.1 Egg Mass Calculation………………………..………………………….…………………....51
4.2 Lycaena hippothoe Egg Density Calculation………..……………………………………...... 51
4.3 Sodium (Na), Cs-137 and Nuclear Transformation per Egg…………………………………51
4.4 Internal MeV/g (x, γ, beta, ce, and auger)……..…..………………………..………..……...51
4.5 Sum of Cs-137Decay Yield, Energy and Absorbed Fraction…………………………………52
4.6 Absorbed Fraction Interpolation Calculations………………………………………………..52
4.7 Internal Absorbed Dose per Egg per Day…………………………………………………….53
4.8 Internal Absorbed Dose per Egg for 4 Days…………………………………………………53
LIST OF TABLES (Continued)
Table Page
4.9 Egg Volume, height and Cs-137 Dispersion Activity (Bq/cm2)………………….………….53
4.10 Ground External Flux (Photons and Betas)……..…………………………………………..54
4.11 Ground Contamination Absorbed Dose (Photons and Betas)……………………………....54
4.12 Leaf Top Contamination External Flux (Photons and Beta)………………………………..55
4.13 Leaf Top Contamination Shielding and Half-life Reduction………………………………..55
4.14 Leaf Top Contamination Absorbed Dose (Photons and Betas)……………………………..55
4.15 Internal + External Absorbed Dose Per Day……………..………………………….………56
4.16 Internal + External Absorbed Dose 4 Days…..………………………………….…………..56
LIST OF APPENDICES
Appendix Page
A. Abbreviations………. …………………………………………………………….…...….....78
DEDICATION
This thesis is dedicated to my family, for all their dedication and support.
1
1. INTRODUCTION
The Fukushima Daiichi nuclear power plant (FDNPP) nuclear accident (described in
Section 2.1) brought much research and discussion to the nuclear field. The World was watching as agencies came together to describe what was happening and to minimize effects. A major factor that contributed to the accident was the widespread assumption in Japan that its nuclear power plants were so safe that an accident of this magnitude was simply unthinkable (Amano &
IAEA, 2015). The questions which evolved placed researchers in the spotlight with environmental radiation modeling and effects. Because of the large-scale nature of the accident, many research questions have been developed for studies on the biological consequences of the accident at the ecological, organismal, and molecular levels (Otaki, 2018).
The Pale Grass Blue Butterfly, with its relatively short life cycle of 30 days and dependence on a single plant, allowed researchers the opportunity to reduce confounding factors and evaluate it in the laboratory. However, Hyama et al. and Nohara et. al. concluded low-dose radiation effects seen in the butterfly are not compatible with the conventional understanding of
Lepidoptera’s high radioresistence, with their studies focused on external exposure and laboratory studies on radioactive cesium consumption. This paper will provide another mechanism of radioactive cesium uptake through puddling behavior to determine if this may have caused their unconventional results. Butterfly puddling (described in Section 2.4), is the behavior which male butterflies suck water from puddles to concentrate sodium. As the
Malpighian tubules secrete urine (much like kidneys), water and ions are taken up by the hindgut. Sodium is then transferred to the female during copulation, and subsequently to the egg.
Cesium-137 is known to be an analog to potassium; could there be similar properties due to 2 cesium and sodium being alkali metals (described in Section 2.2.3)? The next question is then, is the egg the most radiosensitive life-stage of the butterfly? The literature review will provide information on cesium deposition after the FNPP accident, cesium basics, uptake, and radiation dose. The literature review will then focus on the Pale Grass Blue butterfly anatomy, lifecycle, reproduction, habitat, puddling behavior and previous environmental research studies. This will provide the concept, methods and references to determine a proposed internal and external absorbed dose to the Pale Grass Blue Butterfly egg due to puddling behavior, and likely effects, if any, to the butterfly.
3
2. LITERATURE REVIEW 2.1 Fukushima Nuclear Accident & Cs-137 Deposition
Following a major earthquake, a 15-metre tsunami disabled the power supply and cooling of three Fukushima Daiichi reactors, causing a nuclear accident on 11 March 2011 (World
Nuclear Association, 2018). The fuel overheated causing a water-metal reaction that oxidized fuel cladding releasing hydrogen that built up in containment; consequently, an explosive concentration built up and hydrogen explosions occurred on March 14 that blew off the roof and walls at units 1 and 3, followed by releases of airborne radioactive material (J. E. Martin, 2013).
The earthquake and subsequent tsunami, which flooded over 500 square kilometres of land, resulted in the loss of more than 20,000 lives and destroyed property, infrastructure and natural resources (UN, 2013). While the Dai-ichi NPP releases must be considered “significant” relative to prior sources off Japan, we should not assume that dose effects on humans or marine biota are necessarily harmful (Buesseler, Aoyama, & Fukasawa, 2011). Wild organisms inhabiting areas near the FDNPP were exposed to maximum radiation dose rates ranging from 1 to 3.9 mGy/day in terrestrial organisms (Tamaoki, 2016). In comparison to radiation from naturally occurring sources, the Japanese people receive an effective dose of on about 2.1 millisieverts (mSv) annually and a total of about 170 mSv over their lifetimes (UNSCEAR, 2014). Cytogenetic damages will certainly be measurable; however, changes in reproduction in plants and animals
(considered to be the most sensitive endpoint affecting populations) will be difficult to discern against high natural variability (Garnier-Laplace, Beaugelin-Seiller, & Hinton, 2011).
Several different kinds of large-scale environmental monitoring have been repeatedly implemented since the accident, and the accumulated monitoring data have revealed the 4 characteristics of environmental radiological conditions around the Fukushima NPP (Saito et al.,
2016). Unlike Chernobyl, there was no large explosive release of core reactor material, so most of the isotopes reported to have spread thus far via atmospheric fallout are primarily the radioactive gases plus fission products such as cesium, which are volatilized at the high temperatures in the reactor core, or during explosions and fires (Buesseler et al., 2011). Iodine-
131 and cesium-137 were found to be the two most important radionuclides for dose assessment
(UNSCEAR, 2014). Cs-137 is one of the most important radionuclides in terms of exposure to humans and the environment, because of the large amounts released and long half-life of 30.1 years (Imaizumi, 2016). The boiling point of cesium is very low, 671°C, thus, cesium released in nuclear explosions completely evaporates, and, as the cloud cools, the cesium binds with atmospheric aerosol particles (Lehto J. & Huo X., 2011).
Cs-137 was deposited on the ground (UNSCEAR, 2014). Cs-137 deposition in the 80 km zone and in East Japan overall were estimated to be 1.6 and 2.0 PBq, respectively; with about 70 percent of radiocesium was estimated to be deposited in forested regions, 20 percent in agricultural regions and 5 percent in urban regions. (Saito et al., 2016). A maximum value of
15.5 MBq/m2 was measured in a grid square from the municipality of Okuma located just north of the Fukushima Daiichi NPP, with an average ambient dose equivalent rate of 54.8 μSv/h
(IAEA, 2015). The dose effect due to forest soils on land ecosystems were small, 2-6 mSv/d
(converted here from Gy using a relative biological effectiveness factor of 1, appropriate for doses due to I and Cs isotopes, so 1 Sv = 1 Gy) (Buesseler et al., 2011). This radiocesium deposition on the ground has gradually penetrated into the ground, with general slow movement, and decreased air dose rates (Saito et al., 2016). The maximum concentration levels in samples 5 of river water and well water were 2.0Bq/kg for Cs-137, which were far lower than the values relating to Limits on Food and Drink Ingestion (200Bq/kg) (NRA Japan, 2012).
2.2 Cesium-137
2.2.1 Cesium-137 Radioactive Transformation and Decay
Atoms undergo transformation because constituents in the nucleus are not arrayed in the lowest potential energy states possible; therefore, a rearrangement of the nucleus occurs in such a way that this excess energy is emitted and the nucleus is transformed to an atom of a new element (J. E. Martin, 2013). The rate of decay, or transformation, of a radionuclide is described by its activity, that is, by the number of atoms that decay per unit time (Turner, 2007), with curies (Ci) or becquerels (Bq) units defining the quantity of a radionuclide undergoing radioactive transformation (J. E. Martin, 2013). The old unit of activity was the curie (Ci), originally defined as the number of disintegrations per second occurring in a mass of 1 g of
226Ra, later was simply set equal to 3.7 X 1010 s-1, where 1Ci=3.7 X 1010 Bq (Attix, 1986). It must be emphasized that although the becquerel is defined in terms of a number of atoms transformed per second, it is not a measure of the rate of transformation, only quantity of radioactive material, as 60Co releases one beta particle and two gamma rays per transformation
(Cember & Johnson, 2009).
The number of known different atoms, each with a distinct combination of Z and A, is large, numbering over 3200 nuclides, of these, 266 are stable (i.e., nonradioactive) (Shultis &
Faw, 2002). As Z increases, the line of stability moves from N = Z to N / Z ∼ 1.5 due to the influence of the Coulomb force. (Loveland, Morrissey, & Seaborg, 2006). The long-range
Coulomb repulsion between protons is balanced by the presence of additional neutrons providing 6 additional short-range attractive nuclear forces (KAPL, 2009). These can be grouped into three major categories that will determine how they must undergo transformation to become stable: neutron-rich nuclei, which lie below the zigzag line of stable elements, proton-rich nuclei, which are above the line, heavy nuclei with Z > 83 (J. E. Martin, 2013). Nuclei having an excess of neutrons tend to emit an electron (β- particle), thus leaving the nucleus with one less neutron and one more proton, i.e., the atomic number Z is increased by 1; and one or more γ-rays are then emitted to reach the ground state. (Attix, 1986). The general β- decay reaction is thus written as:
+ (Eq 2.1) ט + P → [ D] + e (Shultis & Faw, 2002)
Somewhat more complex transformations of neutron-rich radionuclides are 60Co and
137Cs. These neutron-rich nuclei fall below the line of stability (J. E. Martin, 2013). The nuclide
Cs137 is an isotope of the element cesium (atomic number 55, chemical symbol Cs). There are
137 nucleons in the nucleus consisting of 55 protons and 82 neutrons (Nucleonica, 2018).
Cesium133 is the only stable isotope of cesium, although this element can exist in over 20 isotopic forms, including the complete series from Cs123 to Cs144, with the exception of 124Cs
(Ashraf, Akib, Maah, Yusoff, & Balkhair, 2015). Among the numerous isotopes of cesium that are also produced by nuclear fission are 134Cs (T1/2 = 2.1 y) and 135Cs, but the former is not as persistent as 137Cs and the activity level of the latter is much lower than that of 137Cs (NCRP,
2007).
Each radionuclide has its own characteristic decay constant (λ), which is the probability a radionuclide decays in a unit time for an infinitesimal time interval. (Shultis & Faw, 2002). The decay constant (λ), a characteristic of each nucleus, is related to the half-life, t1/2, by λ = ln 2 / t1/2
(Loveland et al., 2006). The isotope of most concern for Department of Energy (DOE) 7 environmental management sites and other areas is cesium-137 which has a half-life of 30 years
(Peterson, MacDonell, Haroun, & Monette, 2007). It should be noted that the time-scale of
137Cs retention in higher organisms (biological half-lives of generally less than one year; Table
2.1) dictates that 137Cs accumulation in these organisms (Ashraf et al., 2015). Additionally, cesium depositing on plant surfaces is retained to the same extent as other particulate debris. A removal half-time of l4 days due to weathering is generally assumed (Komarov & Bennett,
1983).
The radioactive decay (half-life) equation develops with the compartment model and the differential equation method. The lecture by Reese (2019) described this as: q(t) k
Where rate of change = rate in – rate out; k = removal constant = 1/t.
dq(t)/dt = −kq(t) ln q(t) = -kt + ln q(0) (Eq 2.2) ∫dq(t)/q(t)=∫-kdt ln (q) – ln q(0) = -kt lnq(t) = −kt+c ln (q(t)/q(0)) = -kt Boundary conditions: @t=0 q(t)=0 elnq(t)/q(0) = e-kt ln q(0) = -k(0)+c q(t)/q(0) = e-kt c = ln q(0) q(t) = q(0) e-kt
The final radioactive decay equation is q(t)=q(0)e-kt (Eq 2.3)
TABLE 2.1 Biological and environmental half-life of 137Cs (Ashraf et al., 2015): Organism Half-life Organism Half-life Moss 4.5 years Hen 1.5 days Lichen 5-8 years Cow 3 days Plant surface 14 days Fish 70–300 days Flowing river 1.4 years Child 57 days Monomictic lakea 1.2 years Woman 84 days Meromictic lakeb 6.7 years Man 105 days Flowing river 1.4 years aMix from top to bottom during one period each year. bHas layers of water that do not intermix.
Cs-137 disintegrates by beta minus emission to the ground state of Ba-137 (5.6%) and via the 661 keV isomeric level of Ba-137 (94.4%) which has a half-life of 2.55 min (Chu, Ekstrom, 8
& Firestone, 1999). The lifetimes of nuclear excited states vary, but ∼10–10 s can be regarded as typical; in some cases, however, selection rules prevent photon emission for an extended period.
Such a long-lived nuclear state is termed metastable and is designated by the symbol m:137mBa
(Turner, 2007). The excited nuclei of 137mBa emit gamma rays in 90.1% of emissions, 8.11% by internal conversion of K shell electrons (αK = 0.0811), and 1.72% by electron conversion from other shells; i.e., for each 100 transformations of 137Cs, 85.1 gamma photons, 7.66 internally converted electrons from the K shell, and 1.64 electrons from other shells will occur through relief of the excitation energy retained in 137mBa (J. E. Martin, 2013).
Table 2.2 Cs-137 Average and Maximum Energy (Missouri University EHS, 2005): Major Betas: Major Gammas: Max E (MeV) Avg E # per 100 dis E (MeV) # per 100 dis 0.512 0.157 95 0.662 90 1.173 0.415 5
For those radionuclides where the gamma-ray emission is delayed, as in the case of
99mTc and 137Cs, the gamma-ray emission is called an isomeric transition, where the atomic number and the atomic mass number of the radionuclide is not changed (Cember & Johnson,
2009). As the outer electrons cascade down in energy to fill the innershell vacancy, x rays and
Auger electrons are also emitted (Shultis & Faw, 2002). An interesting example of internal conversion is given by 137Cs; the 137Cs isotope is transformed by beta emission to 137mBa, which is an excited metastable state of 137Ba (Cember & Johnson, 2009). An excited nucleus, instead of emitting a γ-ray of energy hv, can impart the same amount of energy directly to one of its own atomic electrons, which then escapes the atom with a net kinetic energy of hv - Eb, where
Eb is the binding energy of the electron's original shell (Attix, 1986). 9
Fig. 2.1: Cs-137 Decay (Nucleonica, 2018). Copyright Nucleonica Gmbh, www.nucleonica.com. Reprinted with permission
Since the half-life of 137mBa is 2.6 minutes, it is, for practical purposes, in secular equilibrium with its 137Cs parent (Cember & Johnson, 2009). Secular equilibrium is the extreme case that the daughter decays much more rapidly than the parent (i.e., A1<< A2) (Shultis & Faw,
2002). Activity of daughter builds up to that of the parent in about seven half-lives (∼7 T1/2)
(Turner, 2007). If the period of observation (or calculation) is such that the activity of the parent nuclide remains essentially unchanged, then the activity of the radioactive product (commonly referred to as the radioactive daughter) in the equation for the second member of a radioactive series can be simplified as follows:
λ t A(t) = A1 (1 - e ¯ 2 ) (Eq. 2.4) (J. E. Martin, 2013)
In general, charged particles lose considerable energy by ionization, whereas photons and neutrons give up energy by scattering and absorption reactions (J. E. Martin, 2013). The principal mechanisms of energy deposition by photons in matter are photoelectric absorption,
Compton scattering, pair production, and photonuclear reactions (Turner, 2007). Photoelectric 10 absorption and Compton scattering, which involve interactions only with the orbital electrons of the absorber, predominate in the case where the quantum energy of the photons does not greatly exceed 1.02 MeV, the energy equivalent of the rest mass of two electrons (Cember & Johnson,
2009). The Cs-137 661 keV gamma ray creates compton effect (MARLAP, 2004), with compton scattering interactions especially important for gamma rays of medium energy (0.5–1.0 MeV) (J.
E. Martin, 2013). Compton scattering is an elastic collision between a photon and a “free” electron, an electron whose binding energy to an atom is very much less than the energy of the photon (Cember & Johnson, 2009). After scattering, the photon has a longer wavelength λ' and the electron recoils with an energy Te and momentum pe (Shultis & Faw, 2002). The scattered photon likely continues to interact with other atoms until the photon is absorbed by material
(Rumanek & Kudlas, 2018).
A beta particle is a high-speed electron that is so labeled because it originates from the nucleus of a radioactive atom and lose energy due to direct ionization, delta rays from electrons ejected by ionization, production of bremsstrahlung, and Cerenkov radiation (J. E. Martin, 2013).
High-energy beta particles (i.e.,in the MeV range) can emit bremsstrahlung (Turner, 2007).
Bremsstrahlung, (German: “braking radiation”), is electromagnetic radiation produced by a sudden slowing down or deflection of charged particles (especially electrons) passing through matter in the vicinity of the strong electric fields of atomic nuclei (Britannica, 2019). Since beta particles and orbital electrons are about the same size, beta particles are deflected through a rather tortuous path (J. E. Martin, 2013). The Cs-137 beta range in air is 144 cm, water/tissue is
0.20 cm and 0.16 cm in plastic (Cincinnati, 2019).
11
2.2.2 Cesium-137 Dose
The basic quantity for calculating exposures to ionizing radiation is the absorbed dose, which is defined as the amount of energy absorbed per unit mass of tissue of an organ or organism (ICRP, 2008), with the unit J kg–1 called the Gray (Gy) (Turner, 2007). Whereas the roentgen (R) is a unit of exposure, the rad is defined in terms of the energy - 100 ergs - absorbed/g of absorbing material, and thus is a unit of dose received (Ducoff, 1971). Photons produce secondary electrons in air, for which the average energy needed to make an ion pair is
W = 34 eV ip–1 = 34 JC–1; one finds: 1R = 2.58x10.4C/kg x 33.97J/C = 8.76x10-3Jkg-3 (0.876rad) , with 1 R producing a dose of 9.5 × 10–3 Gy (0.95 rad) in soft tissue (Turner, 2007). Since
1 J = 107 ergs and since 1 kg = 1000 g, then 1Gy = 100 rads (Cember & Johnson, 2009).
Radiation damage increases with the linear energy transfer (LET) of the radiation; thus, for the same absorbed dose, the biological damage from high-LET radiation (e.g., alpha particles, neutrons, etc.) is much greater than from low-LET radiation (beta particles, gamma rays, x-rays, etc.) (J. E. Martin, 2013). The ICRP calls its unit of radiation-weighted dose the equivalent dose, with Sievert, and the U.S. NRC weighted dose unit, the rem, dose equivalent; symbolized by the letter H, and has named it the Sievert (Sv) (Cember & Johnson, 2009). The dose equivalent, denoted by H, is defined as the product of the absorbed dose and a factor Q, the quality factor, that characterizes the damage associated with each type of radiation:
H (dose equivalent) = D (absorbed dose) x Q (quality factor) (Eq.2.5) (J. E. Martin, 2013)
The definition of dose equivalent is necessary because different radiations produce different amounts of biological damage even though the deposited energy may be the same (J. E. Martin,
2013). According to the equation and the values for Q, an absorbed dose of 1 mGy of X-, beta, or 12 gamma radiation corresponds to an equivalent dose of 1 mSv, while an absorbed dose of 1 mGy of 5 MeV neutrons gives an equivalent dose of 10 mSv (Cember & Johnson, 2009). Like absorbed dose, dose equivalent is a point function, when dose is expressed in Gy, the (SI) unit of dose equivalent is the sievert (Sv) since 1 Gy = 100 rad, 1 Sv = 100 rem (Turner, 2007). Given
1 R = 0.96 rad, the radiation factor (quality factor) of 1 for x and gamma radiation, 1 R is also approximately equivalent to 1 rem dose equivalent in tissue (Lowe, 2000). To avoid unnecessary words when the context is used is clear, we frequently use the generic term dose when we refer to the radiation-weighted dose (Cember & Johnson, 2009).
The radiation absorbed dose concept implies that the absorbed energy is uniformly distributed throughout the entire mass of the tissue of interest, with biomedical effects on the tissue, organ, and organism levels and thus is appropriate for radiation safety measurements
(Cember & Johnson, 2009). The average kinetic energy of the β- or β+ particles in a β-ray spectrum is found to be roughly 0.3-0.4 times Emax, depending on the individual spectral shape, where estimating the absorbed dose is Ē=1/3Eβmax if more accurate information is not available
(Attix, 1986). If A denotes the average concentration, in Bq g–1, of the radionuclide in the tissue and Ē denotes the average alpha- or beta-particle energy [Ē=1/3Eβmax (J. E. Martin, 2013)], in
MeV per disintegration, then the rate of energy absorption per gram of tissue, otherwise known as absorbed dose rate beta radiation is (Turner, 2007):
D = AĒ x 1.60×10–13 × 103 (Eq. 2.6) ∙ D = 1.60×10–10AĒ (Eq. 2.7)
The internal dose equation can be generalized in terms of the total number of transformations that occur in a source organ, which is a function of q0 and λeff, and three key factors, mT, ΣYiĒi, 13 and AF(TS)i, which are constants for any given radionuclide and a selected source–target organ pair (J. E. Martin, 2013). If an internally deposited radionuclide emits particles that have a short range, then their energies will be absorbed in the tissue that contains them. Such is the case for an alpha or low-energy beta emitter (Turner, 2007). The accumulated radiation dose is obtained by integrating the instantaneous dose rate equation from the time of initial deposition
(t=0) in the tissue to a later time (J. E. Martin, 2013):
-10 -λt D = 1.6 X 10 ΣYiĒiAF(TS)i qo ∫ e (Eq. 2.8) Mt
To understand what “short distances” mean in the present context, we recall that the macroscopic cross section μ is the probability per unit distance of travel that an electronic collision takes place
(J. E. Martin, 2013). This average travel distance before an interaction, 1/µ, is called the mean- free-path length (Shultis & Faw, 2002). The mean free path can also be thought of as the average thickness of a medium in which an interaction is likely to occur (J. E. Martin, 2013). Attix
(2004) gives two limiting cases the dose at P will then equal the sum of the energy per unit mass of medium that is given to charged particles plus γ-rays in radioactive decay:
In a small radioactive object V (i.e., having a mean radius not much greater than the maximum charged-particle range d), CPE is well approximated at any internal point P that is at least a distance d from the boundary of V. Then, if d << 1 / µ for the γ-rays, the absorbed dose D at P approximately equals the energy per unit mass of medium that is given to the charged particles in radioactive decay (less their radiative losses), * since the photons practically all escape from the object and are assumed not to be scattered back by its surroundings. In low-Z media radiative losses are ≈ 1 % or less for β rays, and nil for α’s, so they are ordinarily ignored. (Attix, 1986)
For internal exposure to gamma emitters, dose conversion factors increase in proportion to the mass of the organism due to the higher absorbed fractions, with the dependence being more pronounced for high-energy photon emitters (e.g. 137Cs/137mBa) (ICRP, 2008). Whereas both alpha and beta radiations have definite ranges in matter and therefore can be completely 14 stopped, gamma radiation can only be reduced in intensity by increasingly thicker absorbers; only a fraction of the energy will be absorbed (Cember & Johnson, 2009). For very small radioactive objects (V dʋ) this absorbed fraction approaches zero (Attix, 1986).
Table 2.3: Absorbed Fractions for Uniform Distribution of Activity in Small Spheres and Thick Ellipsoids (Cember & Johnson, 2009): Mass (kg) AF (0.662 MeV) 0.3 0.096 0.4 0.108 0.5 0.117 0.6 0.124 1.0 0.144 2.0 0.173
It is also useful to express the exponential attenuation of photons in terms of a half- thickness, X1/2, or half-value layer (HVL) (J. E. Martin, 2013). The half value layer (HVL) is defined as the thickness of a shield or an absorber that reduces the radiation level by a factor of
2, that is to half the initial level (Cember & Johnson, 2009). Which can be solved for x1/2 to yield:
( ) -µx[½] = = e x1/2 = HVL = (Eq. 2.9) µ (J. E. Martin, 2013)
For alpha and beta emitters, the dose conversion factors for internal exposure are, to some extent, independent of size if it is assumed that they are evenly distributed within an organism
(ICRP, 2008). Also it is normally assumed that all the beta energy is absorbed, since the range in tissue of the betas is very small (Cember & Johnson, 2009). When, instead, this condition is not fulfilled, as in small target volumes the absorbed fraction may be significantly lower and its dependence on target dimension and β− energy cannot be neglected (Amato, Lizio, & Baldari,
2009). The range of electrons in living tissue increases from 160 µm for 100-keV electrons to 5 15 mm for 1-MeV electrons (ICRP, 2008). Absorbed fractions for electrons in small spheres are given in Table 2.4.
Table 2.4: Absorbed Fractions for Energies and Sphere Sizes for Electrons (Stabin & Konijnenberg, 2000): Energy (MeV) Sphere Mass (g) Sphere Radius (cm) Absorbed Fraction 0.2 0.01 0.13 0.864 0.2 0.1 0.29 0.937 0.4 0.01 0.13 0.644 0.4 0.1 0.29 0.832
Detailed analyses of experimental data show that the ability to absorb energy from beta particles depends mainly on the number of absorbing electrons in the path of the beta—that is, on the areal density (electrons/cm2) of electrons in the absorber, and, to a much lesser degree, on the atomic number of the absorber (Cember & Johnson, 2009). It is, therefore, desirable to have a simple expression to show the relative abundances of the radioisotope and the stable isotopes, accomplished by using the concept of specific activity (Loveland et al., 2006). If the radionuclide under consideration weighs 1 g, then, the number of atoms is given by, where A is the atomic weight of the nuclide (Cember & Johnson, 2009):
N = 6.02 × 1023 [ ] × W [ g ] (Eq.2.10) A [ ]
Since the number (N) of radioactive atoms in a radioactive element and its half-life T1/2 are specific for each and every isotope of a radioactive element, the activity per unit mass of a pure radionuclide is a constant that is defined as the specific activity of the element (J. E. Martin,
2013). Specific activity can also be expressed in terms of the disintegration rate (Bq or dpm), or counting rate (counts/min, cpm, or counts/s, cps), or curies (or mCi, mCi) of the specific radionuclide per unit mass of the sample (Loveland et al., 2006). It is more appropriate to refer to measured values of activity per unit mass in a nonpure sample as having a concentration of so 16 many Bq or Ci per gram of a sample or source, otherwise the impression may be given that the sample is pure when in fact it is a mixture (J. E. Martin, 2013).
SA= λN = 0.693 × 6.02*1023 [Bq/g] (Eq. 2.11) T A (Cember & Johnson, 2009)
The MIRD schema is a general approach for medical internal radiation dosimetry.
Although the schema has traditionally been used for organ dosimetry, it is also applicable to dosimetry at the suborgan, voxel, multicellular and cellular levels. (Howell, Wessels, &
Loevinger, 1999). The Medical Internal Radiation Dosimetry (MIRD) Committee of the Society of Nuclear Medicine and Report Number 38 of the International Commission on Radiological
Protection (ICRP-38) were compiled for internal dosimetry calculations and contain all the radiation information necessary for such calculations (J. E. Martin, 2013). In all models, radiation transport is simulated for mono-energetic electrons or photons with energies in the range of approximately 0.01–5 MeV (ICRP, 2008). Evaluated Nuclear Structure Data File
(ENSDF) is built around the properties of nuclear energy levels in individual isotopes (Croff,
Haese, & Gove, 1979), which provides recommended nuclear structure and decay information in a standard format (M. J. Martin, 2007). ICRP 108 (2008), Environmental Protection: The
Concept and Use of Reference Animals and Plants, utilizes the nuclide-specific decay data on energies and yields of radiations emitted by the radionuclides from ICRP Publication 38 (ICRP,
2008), shown in Tables 2.5 and 2.6.
17
Table 2.5: Decay Data for 137Cs (ICRP, 1983): Radiation Type Yield Energy (MeV) β- 1 9.460E-01 1.734E-01 β- 2 5.400E-02 3.347E-01 Listed β ce and Auger Radiations 1.87E-01
Table 2.6: Decay Data for 137mBa (ICRP, 1983): Radiation Type Yield Energy (MeV) γ 1 8.980E-01 6.616E-01 ce-K, γ 1 8.320E-02 6.242E-01 ce-L1, γ 1 1.190E-02 6.557E-01 ce-L2, γ 1 1.690E-03 6.560E-01 ce-L3, γ 1 1.390E-03 6.564E-01 ce-M, γ 1 3.180E-03 6.606E-01 ce-N+, γ 1 9.020E-04 6.616E-01 Kα1 X-ray 3.920E-02 3.219E-02 Kα2 X-ray 2.130E-02 3.182E-02 Auger-KLL 5.350E-03 2.618E-02 Auger-KLX 2.530E-03 3.075E-02 Auger-LMM 4.760E-02 3.577E-03 Auger-LMX 2.780E-02 4.531E-03 Auger-LMX 1.49E-01 9.28E-04 Listed β ce and Auger Radiations 6.51E-02 Listed X and γ Radiations 5.96E-01
The initial dose rates from the uniformly distributed 137Cs may be calculated by using the appropriate value for the specific effective energy (SEE) of the betas plus the gammas
(Cember & Johnson, 2009). Expressing the energies in MeV, the ICRP defines the specific effective energy (SEE) imparted per gram of tissue in a target organ T from the emission of a specified radiation R in a source organ S per transformation as follows:
-1 SEE(TS)R≡AF((TS)R YRERWR [MeVg ] (Eq.2.12) MT (Turner, 2007)
18
To express this contribution in sieverts, we multiply the SEE by the factor (1.60 x 10–13 JMeV–1)
/(10–3 kg g–1) =1.60x 10–10 Sv (MeV g–1)–1 to obtain (Turner, 2007):
-10 H(TS) [Sv] = 1.6x10 Us x SEE(TS) (Eq. 2.13) (J. E. Martin, 2013)
Following the approach introduced by the Medical Internal Radiation Dose (MIRD) committee, the mean radiation absorbed dose from beta radiation to the target volume is below, where A is the cumulated activity (Bq s), β is the average beta energy emitted per disintegration and ϕ is the absorbed fraction: Dβ=A Δβ ϕ/m (Amato et al., 2009). The absorbed dose for gamma ray emitter written in the MIRD notation is:
Dγ(v)=As Δi ϕi(vs) mv (Eq. 2.14) (Howell et al., 1999)
The various ICRP and MIRD internal dosimetry models are similar in terms of their assumptions and defining equations (Bevelacqua, 2005). There are four main assumptions for internal dosimetry: 1st order kinetics, blood is the transfer compartment, no cycling, and daughter products follow their parent (Hamby, 2019). The dosimetric model is based on the biological model where the GI tract is taken to consist of the 4 sections (ICRP, 1979), with a fractional absorption in the gastrointestinal tract (f1) which is also ‘alimentary tract transfer factor’(ICRP,
2015). The ICRP 30 GI model consists of ingestion to stomach, small intestine, upper large intestine, lower large intestine to excretion; f1 consists of transfer from small intestine to body fluids. First-order kinetics is a considerable simplification of the complex processes involved in transfer of material through the lumen of the alimentary tract, but is expected to provide a reasonably accurate representation of the mean residence time of a radionuclide in each segment of the tract (ICRP, 2015). Additionally, it is usually assumed that daughter radionuclides 19 produced from their parent within the body stay with and behave metabolically like their parent
(ICRP, 1979). The ICRP and MIRD methods are the same, namely to calculate a dose from an internal emitter with the MIRD formulation calculates the dose in gray by integrating the dose rate over an infinitely long time after intake of the radionuclide, while the ICRP formulation calculates the equivalent dose in sievert accumulated during 50 years after the intake (Cember &
Johnson, 2009).
Table 2.7: MIRD vs ICRP Methods for Dose (Cember & Johnson, 2009). MIRD ICRP ______D(rkrh), Total lifetime dose H50,T(TS) 50-yr equivalent dose, Sv Gy Ẫ, Cumulated activity, Bq-d Total number of transformations in 50 yrs S(rkrh), Sv/Bq-d SEE(TS), Sv/trans
Frequently the health physicist may find it necessary to know the quantitative relationship between dose rate and distance from a plane radiation source (Cember & Johnson, 2009). If the activity is uniformly spread over the area such that gamma rays are emitted isotropically as SA
γ/cm2 s, and the area source is very much larger than the distance x, the differential flux contributed by each ring at a point P a distance x away from the center of the disc is :
ϕ u(P)A = × ln( ) (Eq. 2.15) (J. E. Martin, 2013)
The radiation absorbed dose rate (Gy/s) from exposure of photons is then calculated according to
Cember & Johnson (2009) with the equation below. The linear attenuation coefficient, µ, is the probability of interaction per unit distance in an absorbing medium (J. E. Martin, 2013).
-13 -1 D = ϕ[ ] x E [ ] x 1.6 x 10 [ ] x µm [ cm ] (eq. 2.16) ∙ / ρm [ ] x 1 [ ] (Cember & Johnson, 2009)
20
Radiation dose calculations for beta particles are based on the number traversing a medium (usually tissue) per unit area, their range, which is energy dependent, and the energy deposition fraction per unit mass µβ. (J. E. Martin, 2013). If the surface concentration is Ca Bq per cm2, then we may assume that 50% of the betas go up from the surface and 50% go down into the surface, furthermore, some of the downward directed betas are backscattered to give the equation for beta flux:
-10 / φo = 3.6x10 x Ca x Ē [ ] (Eq. 2.17) (Cember & Johnson, 2009)
Beta absorption coefficients for air, tissue (or water), and solid materials as a function of Eβmax are given as:
-1.4 µβ(air)=18.6(Eβmax-0.036) [ ] (Eq. 2.18.1) -1.37 µβ(tissue)=18.6(Eβmax -0.036) (Eq. 2.18.2) -1.14 µβ(other)=17(Eβmax) (Eq. 2.18.3) (J. E. Martin, 2013)
The dose rate above a contaminated area is then calculated by:
/ Dβ [mGy/h]= φo [ ] x µβ,t [ ] (Eq 2.19) 10-6[J/g/mGy] (Cember & Johnson, 2009)
2.2.3 Cs-137 Uptake
The transport and fate of anthropogenic Cs-137 is related to the chemical properties of ionic Cs (Cs+), particularly the biologically essential K+, which generally dictates a high degree of mobility and bioavailability of this radionuclide (Ashraf et al., 2015). This is due to Cesium being an alkali metal, along with lithium, sodium, potassium, and rubidium (Ross, 2016). In the periodic table the alkali metals in the group IA are all characterized by having a single s electron 21 in their outer shell (Turner, 2007). Since chemical reactions involve the outer electrons, atoms with similar outer electronic structures have similar chemical properties (Cember & Johnson,
2009).
Intake refers to the activity that enters the respiratory tract or gastrointestinal tract from the environment, while uptake refers to the activity that enters body fluids from the respiratory or alimentary tract or through the skin (ICRP, 2012). Evidence indicates that caesium chloride and other commonly occurring compounds of caesium are rapidly and almost completely absorbed from the gastrointestinal tract (ICRP, 1979). Cs+ uptake is generally mediated by monovalent cation transport systems located on the plasma membrane (Avery, 1995). The ion pumps responsible for maintaining gradients of ions across the plasma membrane provide important examples of active transport, maintained by the Na+-K+ pump (Cooper, 2000). For example, Cs chemical similarity to the biologically essential alkali cation K+ facilitates high levels of metabolism-dependent intracellular accumulation (Avery, 1995). Observations by
Beaugé & Sjodin (1968) show that cesium ions are actively transported into muscle cells and that cesium inward movement is to a large extent chemically coupled to sodium outward movement; with about an 80% uptake curve. Accumulation is also influenced by external pH, with increasing Cs+ accumulation at higher alkaline pH values; hyperpolarization of the membrane occurs when the pH of the external medium is increased, which results in increased K+ fluxes
(Avery, Codd, & Gadd, 1990).
It is well-established that the biouptake of 137Cs is constituted by two major processes with different predominance depending on the trophic position of the organism, i.e. the direct uptake from the water (bioconcentration) and the uptake from contaminated food (Hagstrom, 22
2002). Cell membranes possess ion channels and pumps that are specific to ion size and charge
(NCRP, 2007). For example, K+ channels are selectively permeable to K+ over Na+ ions, typically at a ratio of 1000:1 (Furutani, 2018). Also, the ionic radii of K + and Cs + (133 and 165 pm, respectively) are sufficiently similar, such that Cs+ appears to have an equal or greater affinity than K+ for transport (Avery, 1995). When a potassium ion binds to the selectivity filter, eight water molecules are released from a hydrated potassium ion. The ion is then bound to eight carbonyl oxygens (C=O) at each of the four sites in the selectivity filter (Weston, 2019). The selectivity sequence of K+ channels for alkali metal cations (K+ ≈ Rb+ > Cs+ > Na+ > Li+) indicates that K+ channel permeation is biased against larger hydration energy and larger size, as expected for a relatively “low-field-strength” site (Naranjo, Moldenhauer, Pincuntureo, & Díaz-
Franulic, 2016). Despite its chemical relativeness to the essential cations K+ and Na+, there exists no known biological function for Cs+ (Hagstrom, 2002).
Table 2.8: Atomic and ionic radii of alkali metals and monovalent cations (Avery, 1995) Metal/cation Atomic radius (pm) Ionic radius (pm) Li 152 78 Na 154 98 K 227 133 Rb 248 149 Cs 265 165
Sodium channels are ion channels that transport sodium ions across cellular membranes
(Nature, 2019). The detailed structure of the narrow ‘selectivity filter’ is, however, significantly different to that of potassium channels, being both wider and shorter as well as being partly lined by amino acid side chains (Corry & Thomas, 2011). The selectivity filter (SF) determines the channel’s Na+ permeability over other cations (Naylor et al., 2016). Several studies performed shortly after the cloning of the three subunits of the channel (α, β, and γ EnaC) identified amino 23 acids presumed to be near the extracellular interface of its transmembrane pore as a likely site for the filter, as mutations at this site often reduced cation selectivity (Yang & Palmer, 2018). But, unlike the clear multi-ion picture now available for Kv channels in which backbone carbonyls craft the selectivity filter, a comparable structure of the eukaryotic Na+ selectivity filter and the chemical basis for this process remain unresolved (Ahern, Payandeh, Bosmans, & Chanda,
2015). Although a clear mechanistic picture has not emerged yet, several factors are believed to contribute to Na+ selectivity, including the coordination number of the ion and its partial solvation/desolvation, the electron-donating ability of the residues of the protein pore, and the rigidity and size of the selectivity filter (Nogueira & Corry, 2019). The selectivity ratio of the biologically important alkali cations is high (Grant, 2009). The proposed pore model of the channel accommodates the permeant metal cations in a partly hydrated form (unlike the K channel) following the sequence: Li+ ≈ Na+ > K+ > Rb+ > Cs+ (Hille, 1972), with the
Na+:K+ selectivity of sodium channels 10:1 (Grant, 2009). The similarity between the permeability ratios of sodium channels in squids and amphibians implies that the selectivity filter has little change (Hille, 1972).
Table 2.9: Permeability Ratios For Metal Cations In Sodium Channels (Hille, 1972) Genus Li K Rb Cs Tl Loligo 1.1 0.083 0.025 0.017 - Rana 0.93 0.086 0.012 0.013 0.33
Generally sodium channels are either ligand-gated or voltage-gated (Nature, 2019).
“Gated” refers to the response to a specific stimulus, such as a change in membrane potential
(voltage-gated ion channels) or the binding of a neurotransmitter (ligand-gated ion channels)
(Wilson, 2018). When the Na+ voltage-gated channel opens, membrane potential goes from -70 mV to less negative values. This is because a positive ion is moving inward, making the inside of 24 the membrane more positive (Cummings, 2019). Opening and closing rates for sodium channels are relatively insensitive to the ionic composition of the bathing medium, implying that gating is a structural property of the channel rather than a result of the movement or accumulation of particular ions around the channel (Hille, 1972). Physiological studies suggest that batrachotoxin opens the sodium channel by shifting its activation curve far toward hyperpolarizing voltages and by eliminating sodium inactivation; based on an upper limit estimate of the half-time for Na influx, was 1:0.14:0.02: 0.005 for Na+, K+, Rb+, and Cs+, respectively (Tanaka, Ecclestol, &
Barchi, 1982).
The release of Cs-137 into the aquatic ecosystem generally occurs through association with suspended soil particles of different sizes and mineralogical composition that enter aquatic ecosystems, which considerably affects their transport and bioavailability (Ashraf et al., 2015).
This is due to the dominant aqueous species in most soil and groundwater systems is the uncomplexed Cs+ ion with little tendency for cesium to form aqueous complexes in the soil/water environment (NCRP, 2007). The uncomplexed Cs+ ion forms extremely weak aqueous complexes with sulfate, chloride, and nitrate (EPA, 1999). Unlike many other radionuclides, sorption of cesium to sediments is highly dependent on the mineralogy of the sediment (NCRP, 2007). Cs-137-contaminated soil binds strongly to clay, and the migration rate of clay-bound Cs-137 shows low mobility, less than 1 cm per year, suggesting that the major portion of Cs-137 is distributed in the upper layer of the soil column, within 10 centimeters of the surface (Tamaoki, 2016). The extent to which adsorption will occur will depend on: (1) the concentration of mica-like clays in the soil, and (2) the concentration of major cations, such as
K+ which has a small ionic radius as Cs+, that can effectively compete with Cs+ for adsorption 25 sites (EPA, 1999). Mica, illite, vermiculite and smectite carry net negative charges because not all of the cations in the interstices have sufficient positive charge to balance the charge of the oxygen or hydroxyl anions associated with the mineral structures (NCRP, 2007). The concentration associated with sandy soil particles is estimated to be 280 times higher than in interstitial water (water in the pore space between soil particles); concentration ratios are much higher (about 2,000 to more than 4,000) in clay and loam soils (Peterson et al., 2007).
Accordingly, cations compete for the available binding sites, displacing each other in the following manner: Cs+ > Rb+ > K+ > Na+ (NCRP, 2007).
2.3 Pale Grass Blue Butterfly
2.3.1 Species
Butterflies are insects from the order Lepidoptera, which also includes moths
(“Butterfly,” n.d.). Belonging to the Lycaenidae family, Polyommatinae subfamily (blues); the
Pseudozizeeria genus comprises of a single species maha (Hoskins, 2018). The pale grass blue butterfly Zizeeria maha, is also known as Pseudozizeeria maha (Atsuki Hiyama, 2010). This species in Japan is divided into two subspecies: the mainland subspecies Z. maha argia, distributed in the Japanese mainlands to the north of Nakano-shima Island in the Tokara
Archipelago, and the Okinawa subspecies Z. maha okinawana distributed in the Ryukyu
Archipelago to the south of Takara-jima Island in the Tokara Archipelago (Hiyama et al., 2010).
2.3.2 Pale Grass Blue Butterfly Anatomy and Lifecycle
Some butterflies and moths live only a few days (like the coppers and small blues), others live for about 6-12 months (migrating Monarchs, mourning cloaks, and some moths) (Col,
2018). The Pale Grass Blue Butterfly life cycle is completed in about a month (Hiyama et al., 26
2010), with four stages of metamorphosis: egg, larva (first to fourth or fifth instars (Sakauchi,
Taira, Toki, Iraha, & Otaki, 2019)), pupa, and adult (Drexel University, 2018). Individuals carry genes which govern development at each stage of the lifecycle but different genes come into play at each stage. For example, adults carry caterpillar genes that are “switched off”; mutant genes control features in that stage, i.e. adult wings, probiscus, etc. (Hoskins, 2019). It takes about 4 days for the egg to hatch (Tan, 2014), larval period about 13–15 days (Hiyama et al., 2010), 6 days as pupa (Tan, 2014) and the adult lifespan about a week (Hiyama et al., 2010). The eggs are laid singly on the underside of a leaf of the host plant (Tan, 2014), with 104 +/- 67 (mean +/-
SD) larvae per pair (Hiyama et al., 2010), ranging from 60-200 eggs (Tamil Nadu Agricultural
University, 2019). Eggs are small but visible to the naked eye, because they are conspicuous as white dots on green leaves. (Atsuki Hiyama, 2010). The egg is about 0.4mm in diameter, discoid-shaped with a depressed micropylar at the center of the upper surface (Tan, 2014). The egg height of the closely related Lycaenidae Polyommatinae Cupido is 0.2mm (Eeles, 2019).
Estimated egg volume for Z. maha is 0.04 mm3, and related L. hippothoe is 0.08 mm3 (García-
Barros, 1996). Of the same family, Lycaena hippothoe egg weight is 0.086 +/- 0.013 mg (Fischer
& Fiedler, 2001).
Although species-specific differences are present, radial complexity of the eggshell demonstrates fundamental identity throughout the ditrysian Lepidoptera (Fehrenbach, Dittrich, &
Zissler, 1987). The vitelline envelope is the outer proteinaceous layer outside the oocyte in an insect egg and lies inside the outer shell of the egg which is commonly referred to as the chorion
(Wikipedia, 2017). Observations by Fehrenbach, Dittrich, & Zissler (1987) on three Lepidoptera show the vitelline envelope, bilayered and several µm thick undergoes a marked structural 27 changed, shortly after fertilization and deposition, supplanted by a homogenous layer of 10 nm.
After completing the vitelline envelope the follicle cells synthesize and secrete a set of proteins that wrap the oocyte in a chorion comprising, in Hyalophora, approximately 50% of the dry weight of the mature egg; when deposition is complete is about 60 µm thick. (Telfer, 2009). The egg shell also is peppered with thousands of microscopic pores called aeropyles, and act as breathing tubes for the developing larva (Hoskins, 2019). Radial grooves covers all over the egg on top and on its sides (Easai, 2019) with a finely reticulated outer surface (Hoskins, 2019).
The hatching behavior of Zizeeria maha argia and Zizeeria maha okinawana is slightly different from that of the other subspecies; the 360° turn completed with continuous eating of the top of the eggshell, and after escaping from the egg, these larvae eat the side wall of the eggshell for approximately 15 min (Gurung et al., 2016). An average larva has a body weight of 0.035 g and consumes 0.388 g of leaves throughout its life on average (Nohara, Taira, et al., 2014). The young caterpillar feeds by nibblying away a layer of the leaf lamina, causing thin stripes of whitish marks to appear on the leaf (Tan, 2014); and, because their skins do not expand to accommodate growth, must shed their skins several times (Bailowitz & Sitter, 2019). After about
2-3 days of growth in the first instar, and reaching a length of about 1.8mm, the caterpillar moults to the next instar (Tan, 2014). Each stage between molts is called an instar; each instar is larger than the previous one (Bailowitz & Sitter, 2019). The 2nd instar caterpillar reaches a length of about 2.8mm, and after about 3 days in this stage, it moults again. The 3rd instar takes about 3 to 4 days to complete with the body length reaching about 5.5-6mm (Tan, 2014). The fourth instar caterpillar is the same species as the previous three, even though it looks very different
(Missouri Botanical Garden, 2016) with a more distinctive appearance, featuring a dense coat of 28 short whitish setae all over the body surface (Tan, 2014). After about 4 days of feeding in the 4th instar (Tan, 2014), the fifth instar eats, grows, and becomes too big for its skin. When the caterpillar molts for the fifth and final time, the new skin underneath forms the outer shell of the
Chrysalis (Missouri Botanical Garden, 2016). Six days later, the pupa turns black, and the pupal stage comes to an end with the emergence of the adult butterfly (Tan, 2014).
The adult lifespan is about a week (Hiyama et al., 2010). The adults which emerged from pupae were observed to complete expanding their wings in about 5 minutes, and were able to fly
30 to 40 minutes after emergence (Wago & Unno, 1976). On the Honshu mainland, including the
Fukushima area, the adults emerge mainly from late April to late October. During this six-month period, the generations repeat five to six times continuously (Hiyama et al., 2013). The wingspan of Adult Butterfly is 20-25mm (Tan, 2014). Its forewing size, from the base to the apex, is a little more than 1.0 cm. (Taira, Nohara, Hiyama, & Otaki, 2014). The color of the upper wing surface of the male is quite different from that of the female, that of the male is light blue while that of the female is pitch black; thus able to differentiate conspecific mates (Wago & Unno, 1976).
This species overwinters mostly as third and fourth (final) instar larvae (Otaki, Hiyama,
Iwata, & Tadashi, 2010), although there are no specific overwintering stages in this butterfly
(Sakauchi et al., 2019). Many of these larvae die in winter due to cold temperatures; pupation and molting are problematic below 12°C and no pupation can be observed after the end of
November (Otaki et al., 2010). The overwintering larvae have been suggested to be primarily fourth instar, and they become fifth instar in the spring (Taira, Hiyama, Nohara, Sakauchi, &
Otaki, 2015). Observations by Sakauchi, Taira, Toki, Iraha, & Otaki (2019) showed many overwintering larvae of various instars but not as the fifth instar and not as other stages. 29
2.3.3 Lepidoptera Anatomy and Lifecycle to Irradiation
Anatomy and lifecycle are important when considering irradiation to Lepidoptera.
Regulatory authorities and scientists from many internationally recognized institutions have generated research data on the effectiveness of irradiation as a quarantine treatment against a range of insect pests, including Lepidoptera, infesting various fruits and vegetables (IAEA,
2009). Lepidoptera eggs and larvae comprise the key group of quarantine pests of fruits after
Tephritidae fruit flies (Hallman, 2011). Families of quarantine concern in the Lepidoptera include Crambidae, Gelechiidae, Geometridae, Gracillaridae, Lycaenidae, Lymantriidae,
Metarbilidae, Noctuidae, Oecophoridae, Pyralidae, Sesiidae, and Tortricidae (G Hallman, Valter,
Blackburn, & Parker, 2013). In 2006, the U.S. Animal and Plant Health Inspection Service
(APHIS) accepted a generic dose of 400 Gy for all Insecta minus pupae and adults of
Lepidoptera, which may require higher doses (G Hallman, Levang-Brilz, Zettler, & Winborne,
2010). The dose-response tests, with the target radiation dose of 20 (late eggs), 40, 60, 80, 100,
120, 140, and 160 Gy (late fifth instars in vitro) respectively applied to all stages, showed that the tolerance to radiation increased with increasing age and developmental stage (Zhan et al.,
2014). Table 2.10 presents relative radiotolerances of stages of Lepidoptera for measures of efficacy used in phytosanitary irradiation (PI), which supports the hypothesis that radiotolerance increases as insects develop (G Hallman et al., 2010).
Table 2.10: Radiotolerances of different stages of Lepidoptera (G Hallman et al., 2010). Order Prevention of first (min. dose [Gy] to achieve near 100% efficacy)_____ Gelechiidae Adult Larva 1-1.5 d (100), 6-6.5 d (100), 12-12.5 d (125) Adult Instar 4 (120); pupa 1-3 d (400), 6-8 d (700) Adult Egg (175); larva (175); pupa (>1000) Noctuidae Pupation Egg 3 d (<200), 4 d (200); instar 3 (200), 5 (400) Pyralidae F1 Egg Production Egg (200); larva 7 d (200); pupa 5 d (>350); adult (>550) Tortricidae Adult Instar 1 (90), 2-3 (120), 4-5 (>150); pupa (>>150) 30
2.3.4 Reproduction
Female polyommatine butterflies are thought to mate only once (Gurung et al., 2016), with mating behavior generally taking place soon after emergence of the female (Wago & Unno,
1976). Lepidopterans produce two types of sperm cells, nucleated eupyrene and smaller, anucleated apyrene sperm (Koudelova & Cook, 2001). The role of apyrene sperm is not fully understood, although it has been suggested that they aid the transfer of eupyrene sperm to the female, have a nutritive function, or may be involved in sperm competition (Carpenter & Marec,
2005). There is good evidence that apyrene sperm protect a males reproductive investment by delaying female remating (Cook & Wedell 1999). Thus, apyrene sperm, as well as eupyrene sperm, must be considered when investigating male reproductive success (Koudelova & Cook,
2001). The percentage eupyrene sperm found in the spermatheca of females that had mated with male progeny of irradiated males were significantly lower than the percentage eupyrene sperm found in the spermatheca of females that had mated with untreated or irradiated males (Ocampo,
2001). Thus, even if females have received enough sperm to fertilize the eggs, they might be able to perceive that insufficient sperm quantities were transferred during mating, with these females more likely to remate (Koudelova & Cook, 2001). This increases mating ability and superior sperm competitiveness (Carpenter & Marec, 2005)
Both types of sperm are transferred to the female during copulation via the spermatophore and both reach the site of sperm storage, the spermatheca (Koudelova & Cook,
2001). Due to spermatophore not transferred intact to the female, most of it forms during mating within an organ in the female called the bursa copulatrix (University of Minnesota, 2019). The bursa copulatrix is a sac-like organ in female Lepidoptera in which the spermatophore is stored 31 immediately after mating which secretes enzymes that break the spermatophore down into nutrients that can be used by the female (University of Minnesota, 2019). One nutrient nutrient of interest is sodium. Sodium is gathered by male butterflies and moths during mud-puddling
(discussed in Section 2.3.5: Butterfly Puddling) is transferred to their female mates through the spermatophore, and this nuptial gift increases reproductive success. (Beck, Muhlenberg, &
Fiedler, 1999). Sodium enhances sperm motility, or facilitate amino acid uptake in the insect gut and thus make more amino acids available for growth and reproduction (Molleman, 2010). When the sperm reach the egg, they move through the micropyle and fertilization occurs (University of
Minnesota, 2019). Internal fertilization of the eggs takes from a few seconds to many minutes or hours (Col, 2018). Once the egg is fertilized, the female deposits the egg (oviposits) on an appropriate hostplant. (University of Minnesota, 2019). Lepidoptera utilize glands, which secrete the cement that glues eggs to the oviposition substrate (Chew & Robbins, 1984).
2.3.5 Habitat
The pale grass blue butterfly lives on or just above the surface of the ground throughout its life stages because its host plant, Oxalis corniculata (Taira et al., 2014). The larvae of the pale grass blue butterfly feed on Oxalis corniculata (Taira, Hiyama, et al., 2015), an exceedingly common little plant with a bright yellow flower, found in all temperate and tropical regions
(Kershaw, 1907). Leaves resemble those of clover, each consisting of three heart-shaped leaflets that are 1/4 to 2/5 of an inch (0.6–1 cm) long and 1/6 to 1 inch (0.4–2.5 cm) wide. In comparison to the leaves, leaf stalks are long, almost 3 inches (7 cm) in length (CA, 2019). The midrib is
110µm thick and 70µm wide, and the lamina is about 90µm thick (Karunanithi, Rajkishore, &
Pol, 2016). The host plant is often found in rice and vegetable fields, grasslands and gardens, 32 therefore, Z. maha is often found in man-made environments, including urban areas (Hiyama et al., 2013). It is most abundant on the plains, but also commonly occurs on savannah/woodland mosaics, Acacia scrub, and in forest glades and clearings (Hoskins, 2018).
Fig. 2.2 Oxalis corniculata Plant (Starr & Starr, 2018). Reprinted with permission.
The adult butterflies have a weak fluttering flight, and are usually spotted in the vicinity of its host plant, Yellow Wood Sorrel (Oxalis corniculata) (Tan, 2014). It is seen from 8:00 to
18:00 (Wago & Unno, 1976), mostly at a height of less than 1 m, partly because its host plant is usually less than 10 cm high (Tan, 2014). This species does not require large space to live even in the natural environment. (Atsuki Hiyama, 2010), as it disperses over only a limited distance, likely a few kilometers at most (Hiyama et al., 2013). In the warm east coast area of Japan facing the Pacific Ocean, adults are observed from late March to early December, with adults emerging five to six times a year (Otaki et al., 2010).
2.3.6 Butterfly Puddling Behavior
Naturalists have long been fascinated by the sight of butterflies, aggregated often by the hundreds, drinking at water sources (S. R. Smedley & Eisner, 1996). Puddling refers to the 33 process in which butterflies sip from puddles of water, mud, dung or carrion on the ground with their proboscis unfurled and probing into the ground to take in water and nutrients (Khew,
Wong, Ong, & Wong, 2008). Puddling behavior is well known among lepidopterologists and naturalists (Inoue et al., 2012), and is a conspicuous feature of the family Papilionidae,
Lycaenidae and Nymphalidae (Hasan, Neha, Baki, & Quamruzzaman, 2018). Blue butterflies
(Lycanidae, mostly tribe Polyommatini) regularly aggregate at puddles, with the bulk of puddling observations (76.2%) in a forest butterfly community (Beck et al., 1999). The water is rich in mineral salts and other essential nutrients (mostly sodium chloride and nitrogen-rich solutions) that have leached from the surrounding soil and rocks (Col, 2018). Experiments show that moderate concentrations of Na+ solutions are preferred and that the proboscis sensilla housed Na+ receptor neurons to detect various concentrations of Na+ (Inoue et al., 2012). Rivers contain only 9 ppm, while seawater contains approximately 11,000 ppm sodium (Lenntech,
2020). Therefore, adult Lepidoptera rarely use seawater as a source for sodium (Pola & García-
París, 2005) due to high Na+ concentrations (Inoue et al., 2012).
Fig. 2.3: Scanning Electron Microscope of Proboscis (Li, 2019). Reprinted with permission.
34
Almost all butterflies and moths are dependent on their host plants as food in the larval stage and on flowers in the adult stage, and previous research has shown that plant leaves contain higher amounts of K+ than Na+ (Inoue et al., 2015). Mud-puddling behaviors by insects serve to compensate for shortages in the larval diet, especially for sodium (Shen et al., 2008), due to the low sodium content of land plants (Beck et al., 1999). The diet of adults (primarily nectar and fruits) lack sodium/salt, and the hostplants that they feed on in their younger larval stages do not provide enough sodium either (Khew et al., 2008). This exchange primarily consists of the excretion of K+ that the butterflies acquired from host plant leaves during the larval stage, and the absorption of Na+ (Inoue et al., 2015). Observations by Smedley & Eisner (1996) showed that potassium acquisition and transfer do not appear to be concomitants of puddling, as the ion is in plentiful supply in land plants, and insect herbivores generally show no lack of it. When performing this behavior, butterflies and moths simultaneously sip and excrete water solutions as urine (Inoue et al., 2012). Fluid ejection results from a need to pass great quantities of soil moisture through the gut in order to extract sufficient useful, but dilute, chemicals (Khew et al.,
2008). Excretion is a two-step process, with much of the fluid that is taken up by the Malpighian tubules being reabsorbed by the hindgut before it passes out of the body (Klowden, 2013).
The insect digestive system can be divided into three different parts: foregut, midgut and hindgut (Wikipedia, 2020). Starting in the foregut food triggers the salivary glands, the crop acts as a ‘pre-stomach’ to store/partially digest, then to the midgut food is broken down and then moves through a valve called the pyloric valve to enter the Malpighian tubules and hindgut to be absorbed (Fahmy, 2007). The functional kidney in insects consists of the Malpighian tubules and hindgut (Dennis Kolosov, Piermarini, & O’Donnell, 2018). Malpighian tubules are the primary 35 excretion organ of insects but operate in a different manner from kidneys, which base their filtration on hydrostatic pressure (Klowden, 2013). The Malpighian tubules actively transport ions from the hemolymph into the tubule lumen, which osmotically drags water to produce an isosmotic primary urine (Yerushalmi, Misyura, MacMillan, & Donini, 2018). Hemolymph is the circulating fluid or “blood” of insects which moves through the open circulatory system, directly bathing the organs and tissues (Kanost, 2009).The rest of the hindgut plays a major role in homeostasis by regulating the absorption of water and salts from waste products in the alimentary canal (NC-State, 2019).
The segment where control of the sodium cation occurs (Phillips & Audsley, 1995) apparently takes place in the hindgut (S. R. Smedley & Eisner, 1996). Although it has been observed that the ilium (anterior hindgut) of the male is 1.65 times as long as and 2.00 times as wide as that of the female and bears dense packing of villi (S. Smedley & Eisner, 1995), the ilium is unlikely to be involved with transepithelial ion transport. Cells showing the characteristic of a transporting epithelium are not present, with ultrastructural studies support proposal of active ion transport across the rectal epithelium (D Kolosov & O’Donnell, 2019).
Observations by Inoue et al. (2015) show the butterfly alimentary tract selectively absorbs or excretes each ion independently according to the concentration of ions in the absorbed water and in their hemolymph, such as in the Malpighian tubule (MT), assuming the concentration of each substance in the absorbed water is adequate. In plant-feeding insects that ingest K+-rich diets, fluid secretion relies primarily on the transport of K+ by the epithelial cells in the secretory portion of the MT with a primary role for a vacuolar-type H+-ATPase proton pump coupled to a
K+/H+ exchanger to drive K+ from the cell to the tubule lumen (Dennis Kolosov et al., 2018). It 36 is through the selective rectal reabsorption of desirable constituents from Malpighian tubular fluid and gut contents that the insect achieves osmotic and ionic regulation (Irvine, 1969). In most insects, the epithelium of the rectal sac is thickened to form structures known as rectal pads or papillae (Jarial, 1993). Changes in fluid reabsorption from the rectal lumen alter hydrostatic pressure in the lumen of the ileac plexus, and that resultant changes in the opening of the Piezo and Cav channels lead to the opening of voltage-dependent channels that are involved in reabsorption of Na+ and K+ (D Kolosov & O’Donnell, 2019).
A butterfly can puddle from anything between a few seconds to an hour or more, depending on a variety of factors (Khew et al., 2008). Time spend puddling is inversely related to the Na+ content in the external fluid source (D Kolosov & O’Donnell, 2019). In extreme cases a butterfly may imbibe an amount of fluid 600 times its own body weight in a single puddling session, leaving the excess water as it drinks and retaining only the needed minerals (Ghosh &
Mukherjee, 2016). A single male Gluphisia, weighing about 80 mg, may puddle uninterruptedly for hours, during which it pumps an astounding 10-50 ml of fluid through its gut (S. R. Smedley
& Eisner, 1996).
Puddling is associated with both the insects’ nutritional ecology and their reproductive biology (S. R. Smedley, 2009), as sodium is vital for many physiological functions, including digestion, excretion, reproduction and flight. (Khew et al., 2008). Quantitatively, the male sequesters in the order of 17 µg of sodium by puddling, and relinquishes at mating about 10 µg of this supply, of which the female then passes 5 µg to the eggs. (S. R. Smedley & Eisner, 1996).
It is usually the younger males that puddle; by not needing to puddle, females can focus their energies on obtaining more of the other kinds of nutrients, hunting out better oviposition sites, 37 and on laying healthier, more fertile eggs (Khew et al., 2008). Occasionally, however, female butterflies visit mud puddles (Beck et al., 1999), presumably because their male-derived sodium reserves had been depleted (Boggs & Dau, 2004). Sex differentials in puddling behavior can usually be explained by transfers of nutrients from males to females during mating (Molleman,
2010). Sodium gathered by male butterflies and moths during mud-puddling is transferred, sometimes in large amounts, to their female mates through the spermatophore, and this nuptial gift increases reproductive success (Beck et al., 1999). Apportionment of received sodium appears to occur promptly: within the day after mating eggs already contain a significant allocation of sodium (S. R. Smedley & Eisner, 1996). These extra salts and minerals improve the viability of the female’s eggs, increasing the couple’s chances of passing on their genes to another generation. (Khew et al., 2008).
2.3.7 Environmental Studies
At the time of the nuclear accident researchers were studying this butterfly (Taira et al.,
2014), on the establishment of a lycaenid model system for butterfly physiology and genetics, and to establish a laboratory rearing method (Hiyama et al., 2010). Thus, researchers were able to study the effects of the accident on this species promptly (Taira et al., 2014).
2.3.7.1 Environmental Indicator Species
While much is discussed on the butterfly it should be noted the United Nations Scientific
Committee on the Effects of Atomic Radiation (UNSCEAR) (2008, 2013) followed the reference animals and plants (RAPs) concept endorsed by the International Commission on Radiological
Protection (ICRP) (ICRP 2008). Among reference animals, a representative above ground 38 invertebrate [insect] to be studied for environmental protection is bees (Otaki & Taira, 2017).
ICRP 108 gives the following points:
Collectively, therefore, in selecting a small but practical set of Reference Animals and Plants, the following points were considered: that a reasonable amount of radiobiological information is already available on them, including data on probable radiation effects; that they are amenable to future research, in order to obtain the necessary missing or imprecise data, particularly with regard to radiation effects; that they are considered to be typical representative fauna or flora of particular ecosystems and have a wide geographic variation; that they are likely to be exposed to radiation from a range of radionuclides in a given situation, both as a result of bio-accumulation and the nature of their surroundings, and because of their overall life span, life cycle, and general biology; that their life cycles are likely to be of some relevance for evaluating total dose or dose rate, and for producing different types of dose–effect responses; that their exposure to radiation can be modelled using relatively simple geometries; that there is a reasonable chance of being able to identify any effects at the level of the individual organism that could be related to radiation exposure (bacteria and unicellular organisms were excluded because of their high resistance to radiation); and that they have some form of public or political resonance, so that both decision makers and the general public at large are likely to know what these organisms actually are, in common language. (ICRP, 2008).
The researchers proposed 6 criteria called “the postulates of pollutant-induced biological impacts” in analogy with Koch’s postulates of infectious diseases (Otaki & Taira, 2017). Koch formulated a set of criteria that could be used to identify the pathogen responsible for a specific disease: 1) The organism must be regularly associated with the disease and its characteristic lesions; 2) The organism must be isolated from the diseased host and grown in culture; 3) The disease must be reproduced when a pure culture of the organism is introduced into a healthy, susceptible host; 4) The same organism must be reisolated from the experimentally infected host
(Racaniello, 2010). Although there are some points that must be improved in the future, the researchers maintain studies on the blue butterfly largely satisfy all 6 postulate criteria: 1) spatial relationship, 2) temporal relationship, 3) direct exposure, 4) phenotypic variability or spectrum,
5) experimental reproduction (external exposure), and 6) experimental reproduction (internal exposure) (Nohara, Hiyama, Taira, & Otaki, 2018). 39
Currently, butterflies and moths are used for multiple disciplines, such as developmental biology, evolutionary biology, ecology, behavioral biology, and environmental sciences; in the past, this species was used to evaluate the ecological risk associated with transgenic maize pollen
(Hiyama et al., 2012). To be a useful indicator species for environmental pollution, the primary requirements are that many individuals can be efficiently collected from the field without damaging the wild populations or ecosystems and that morphological abnormalities can be visually determined without ambiguity (Gurung et al., 2016). Advantages of the pale grass blue butterfly, is that it can be studied without any specific permission in Japan, is the most common and abundant butterfly in Japan and is distributed in both rural and urban regions (Taira, Iwasaki,
& Otaki, 2015). The pale grass blue butterfly’s small size, low flying ability, and short life cycle
(approximately 1 month) allowed researchers to breed hundreds or thousands of individuals in the laboratory to perform rigorous statistical analyses. (Taira et al., 2014). Additionally, it eats only a single species of plant, which simplifies the field work, the rearing method, and the interpretation of field and experimental results (Hiyama et al. 2010). Thus, the field work and laboratory experiments were bridged to understand the state of this butterfly in an environment
(Hiyama, Taira, Sakauchi, & Otaki, 2018).
2.3.7.2 Environmental Study Complexities
For any postaccident ecological impact assessment of the Fukushima accident, great care is needed in the quantification of radiation dose to biota, consideration of confounding effects
(e.g., from the tsunami, complex mixture of toxicants), and careful sampling designs if meaningful results are to be obtained (Garnier-Laplace et al., 2011). Since the Fukushima accident, large-scale environmental monitoring activities have been implemented and enormous 40 amounts of environmental monitoring data have been accumulated (Miyahara, 2016). The United
Nations Scientific Committee on the effects of Atomic Radiation (UNSCEAR) report was among the most comprehensive international scientific analyses of the levels and effects of exposure to radiation following the accident at Fukushima-Daiichi Nuclear Power Station (UNSCEAR,
2016a). The overall results were largely consistent, although the IAEA was more definite on whether population-level effects observed in field studies may be linked to radiation exposure, concluding that no impacts on populations and the ecosystems (both terrestrial and marine environments) were expected (UNSCEAR, 2016b). To keep abreast of new scientific information that has emerged since the launch of its 2013 report, UNSCEAR conducted reviews of relevant scientific literature published in the 2014 and 2015 to guide the Committee’s future programme of work (UNSCEAR, 2016a). Several studies are listed in Table 2.11 on biological impacts of radiation conducted since the Fukushima accident, on genetic damage, abnormality rates and population abundance (Tamaoki, 2016). The Committee had expressed reservations about these observations, noting that uncertainties with regard to dosimetry and possible confounding factors made it difficult to substantiate firm conclusions from the cited field studies
(UNSCEAR, 2017). The Committee had noted that the substantial impacts reported for populations of wild organisms from these studies were inconsistent with the main findings of the
Committee’s theoretical assessment (UNSCEAR, 2016b). It identified a need for multidisciplinary field studies tailored to analyze the impacts of ionizing radiation on populations of wild organisms interacting under the conditions prevalent within ecosystems in areas with enhanced levels of radioactive material (UNSCEAR, 2017).
41
Table 2.11: Biological impacts of radiation conducted since the Fukushima accident (Tamaoki, 2016). Reduced male forewing size and more severe abnormalities Hiyama et al., 2012 found in F1 offspring from female first-voltine than their parents.
More severe abnormalities found in adult butterflies collected in the fall Hiyama et al., 2013 of 2011 than in the spring of 2011.
Recovery of abnormality rate to normal levels after the fall of 2012. Hiyama et al., 2015
Increased mortality and abnormality rates in larva when fed Nohara et al., 2014a Oxalis corniculate collected from the Fukushima area.
Some abnormal phenotypes considered due to internal exposure inherited Nohara et al., 2014b by the next generation
The authors of this suite of publications maintained that exposures due to releases from the FDNPS accident would have led to mortality and abnormalities in the studies butterfly species, that mutations would have been passed on to the progeny and that populations would have decreased considerably in areas close to FDNPS (UNSCEAR, 2015). At the time of the nuclear accident, the researchers were studying this butterfly (Taira et al., 2014) and provided evidence to suggest that the high abnormality rates observed in the pale grass blue butterfly were induced by “anthropogenic radioactive mutagens” (UNSCEAR, 2017). The meltdown and explosion of the Fukushima Dai-ichi NPP occurred on 12 March 2011, when Z. maha was overwintering as larvae; on that date and thereafter, these larvae were exposed to artificial radiation not only externally but also internally from ingested food (Taira, Hiyama, et al., 2015).
Adult butterflies collected in the fall of 2011 showed more severe abnormalities than butterflies collected in the spring of 2011 (Hiyama et al., 2013). Abnormalities of the F1 generation were observed that were not necessarily seen in the first-voltine parents, and these
F1 abnormalities were inherited by the F2 generation, suggesting that genetic damage was introduced in germ-line cells of this butterfly (Hiyama et al., 2012). In contrast, it has been 42 studied the lower dose of radiation used to induce F1 sterility increases the quality and competitiveness of the released insects as measured by improved dispersal after release, increased mating ability, and superior sperm competitiveness (Carpenter & Marec, 2005).
Additionally, critics frequently argued that the data were unusable because the background abnormality rate was too high to detect the influence of the radiation; F1 generation background rate varied from 1.4% to 19.8% with laboratory-reared adults scoring higher abnormality rates of adults than the field-collected ones (Hiyama et al., 2013). When partially sterile males mate with fertile females the radiation-induced deleterious effects are inherited by the F1 generation.
As a result egg hatch is reduced and the resulting (F1) offspring are both highly sterile and predominately male, as shown in Table 2.12 (Carpenter & Marec, 2005).
Table 2.12: Typical attributes of male lepidopteran insects (and their progeny) receiving substerilizing doses of radiation (Carpenter & Marec, 2005) Dose Applied to P1, Gy Egg Hatch % Larval Mortality Egg Hatch %(F)(M) 0 71.8 20.0 82.5 / 76.8 100 46.1 51.1 10.8 / 13.9 200 30.8 69.5 0.9 / 7.5 250 19.1 75.1 0.8 / 6.1
In insects, germline cells are allocated at the earliest stage of embryogenesis (Hiyama et al., 2012). At the later instar stages, germline cells undergo active division, growth and differentiation (Taira, Hiyama, et al., 2015). When germ-line cells are damaged, even if radiation-exposed individuals do not demonstrate pathological traits, their progeny could exhibit abnormal traits, because the somatic cells produced from the sperm and oocytes after fertilization contained mutations and expressed pathological traits (Hiyama et al., 2013). Thus, theoretically, internal exposure through ingestion could cause germline DNA damage (Taira, Hiyama, et al.,
2015). 43
The authors agree that the original study (Hiyama et al. 2012) should be confirmed by additional studies because the results are not compatible with the conventional understanding of UNSCEAR and radiation biology (Otaki & Taira, 2017). Lepidopteran cells are more resistant than human cells to short-term high-dose exposures but in the study, the larvae and pupae were vulnerable to the long-term low-dose exposures at the organismal level (Hiyama et al., 2013). The mortality and abnormality rate in larva of the pale grass blue butterfly fed O. corniculata leaves collected from areas in Fukushima increased sharply at the lower dose range of the ingested radioactive Cs, and some of the abnormalities found in first generation of the butterfly succeeded in the next generation (Nohara, Hiyama, Taira, Tanahara, & Otaki, 2014).
Noting some technical errors, doses were wrongly specified in units of becquerels and reference was made to dose-response models that were inappropriate for the end points being studied (UNSCEAR, 2015). However, this is not an error but rather reflects the research philosophy. The researchers maintain that the gray (Gy) or sievert (Sv) cannot be used accurately, at least in their systems, because to obtain these values, calculations are required based on many unreliable assumptions (Otaki & Taira, 2017). Leaf samples were measured with a germanium semiconductor radiation well type detector, Canberra GCW-4023 (Nohara,
Hiyama, et al., 2014). Semiconductor detectors are very efficient in detecting photons, with proportional counters useful for particle counting (J. E. Martin, 2013). Additionally, the researchers used scintillation survey meter measurements with dose values shown in the paper not considering β-ray doses (Hiyama et al., 2013). The calculated half mortality dose (the dose required to cause death in 50% of the individuals in the treated group; fundamentally equivalent to the median lethal dose, LD50) was 1.9 Bq/body (1900 mBq/body), whereas the calculated half 44 abnormality dose (the dose required to cause death or morphological abnormality in 50% of the individuals in the treated group, which is fundamentally equivalent to the median toxic dose,
TD50) was 0.76 Bq/body (760 mBq/body) (Taira, Hiyama, et al., 2015). Because insects, especially Lepidoptera (butterflies and moths), are thought to be highly resistant to radiation exposure, the dose-dependent decline in the survival rates of adults obtained may have been unexpected (Hiyama et al., 2013), with reviews indicating that reported observations of LD50 values for adult insects vary from 20 to 3000 Gy (ICRP, 2008).
Furthermore, Yoshioka (2015) investigated flying insects, including pollinators (bees, wasps, butterflies, and hoverflies) were shown to be abundant; with no marked loss of ecosystem functions and services by flying insects in the evacuation zone. And, Otaki synthesized the results from several studies of the effects on the same species of butterfly following the FDNPS accident, and reported that ionizing radiation was unlikely to be the exclusive source of the environmental disturbances observed (UNSCEAR, 2017). In addition, impact of the tsunami itself, may cause the environmental effects observed(Otaki & Taira, 2017) It has been hypothesized that not only radiation, which has often been the focus of previous studies in
Fukushima and Chernobyl, but also the cessation of the disturbances caused by anthropogenic activities such as farming and gardening seriously affect the biodiversity and ecosystem services in evacuation zones (Yoshioka, Mishima, & Fukasawa, 2015).
Lepidopteran insect cells are known to exhibit very high radioresistance (Chandna et al.,
2009). It takes an average dose of 10,000 mSv to kill a Lepidoptera cell, and it requires an average dose of 1,300 mSv to Lepidoptera eggs to reduce their hatch rate by 50% (Jorgensen,
2012). The radiosensitivity in eggs of tasar silkworm, Antheraea proylei (Lepidoptera) was 45 studied by analyzing the frequency of embryonic deaths, larval survivability, moulting, larval durations, stage lethalities, growth of hatched larvae, green and dry cocoon weights; embryonic lethalities expressed in LD50 value was found to be 13 Gy at 12 hours after oviposition (Chanu
& Ibotombi, 2011).
The intrinsic nature of radioresistance in insect cells, especially in those of lepidopteran insects, provides a useful working model to understand how organisms like insects can tolerate such high dosages of irradiation (Cheng, Lee, & Wang, 2009). The Law of Bergoni &
Tribondeau (1906), which was based on histopathological criteria of radiation damage, stated that the radiosensitivity of cells was related directly to their proliferative activity and mitotic future, and inversely to their degree of differentiation (Ducoff, 1971). The mitotically cells of this line are approximately 50-100 times more radioresistant than mammalian cells (Koval,
1983b). It is possible that superior x-ray repair confers a certain amount of radioresistance in all insect cells and that this resistance is magnified in the Lepidoptera through unique chromosomal features (Koval, 1983a), such as very small size (Koval, 1983b). It should be noted, chromosome number does not appear to correlate with radiosensitivity in insects (Koval, 1983a). All of the lepidopteran lines are highly heteroploidy and have approximate chromosome numbers near or above 100, with Diptera having a small number of relatively large monokinetic chromosomes
(Koval, 1983b). In another view, the radioresistance of lepidopteran cells has been attributed to be a consequence of their possession of holokinetic chromosome (Chanu & Ibotombi, 2011) and to the fate of the radiation-induced chromosome fragments during mitotic cell cycles. (Carpenter
& Marec, 2005). The basis is explained with chromosomal breakage due to radiation damage could be expected to result in the loss of chromosome parts and subsequent cell death in 46 monokinetic species and because chromosome fragments could be retained in chromosomally holokinetic species, thereby decreasing the amount of cell killing (Koval, 1983a). Lepidopteran chromosomes possess a localized, large kinetochore plate and cover a significant portion of the chromosome length, ensuring that most radiation-induced breaks will not lead to the loss of chromosome fragments as is typical in species with monocentric chromosomes (Carpenter &
Marec, 2005). Previous studies on a cell line isolated from a lepidopteran, Trichoplusia ni,
TN368, concluded from the results of unscheduled DNA synthesis experiments that the higher
DNA repair efficiency might account for its superior radioresistance over mammalian cells
(Cheng et al., 2009). Significant DNA damage was detected only at 20 Gy and higher doses, in contrast with human cells that showed similar damage at 2 Gy, and display very low induction of apoptosis at doses up to 200 Gy (Chandna et al., 2009). For highly radioresistant Lepidoptera, multiple chromosome rearrangements must be induced in lepidopteran males to be manifested as
DLMs, explaining why lepidopteran males require very high radiation doses (350–500 Gy) to be fully sterilized. (Carpenter & Marec, 2005).
The lethal exposure for most mammals lies between 200 R and 1000 R (2-10 Gy), and for insects it lies between 1000 R and 100,000 R (10-1000 Gy) (Chanu & Ibotombi, 2011).
Considering the Do of x-ray survival curves for most mammalian cells to be between 1.0 and 1.5
Gy, the dipteran cells demonstrate radioresistance 3 to 9 times that of mammalian cells, whereas the lepidopteran cells are 52 to 104 times more radioresistant (Koval, 1983a). This high radioresistance in Lepidoptera also applies to germ cells, and in particular to mature sperm
(Carpenter & Marec, 2005).
47
3. MATERIALS & METHODS
This paper started with a theses for an alternative Cs-137 pathway to the Pale Grass Blue
Butterfly due to puddling, expanding to radiation effects. This provided the egg as the most radiosensitive life-stage. The literature review developed the necessary information and parameters for the proposed egg absorbed dose due to butterfly puddling, which were organized and calculated in Excel (shown in Chapter 4. Results). The methods used to calculate dose to the
Pale Grass Blue Butterfly egg applied health physics textbook concepts and international standards (ICRP). Although the butterfly is known to overwinter as larvae with emergence in
April, a maximum dose was calculated as if the butterfly was exposed directly after the accident with the maximum recorded activities in this thesis. Information from other Lepidoptera studies and organisms were utilized when information on the butterfly was unavailable. A calculation was performed over computer code, in order to demonstrate an understanding of dosimetry calculations, units and assumptions. This is unlike the research philosophy of Hiyama et. al. and
Nohara et. al., leaving measurement values for interpretation. Otaki & Taira (2017) describe the researchers’ philosophy for leaving original values (µSv/h) for their external research, saying they ‘understand’ that Sv is used only for human doses, and there are too many assumptions to convert Bq to Gy. This chapter will provide the methods used to calculate internal and external absorbed dose to the egg due to butterfly puddling, which are given in Chapter 4 Results.
3.1 INTERNAL ABSORBED DOSE
The egg was chosen as it is the most radiosensitive stage in the lifecycle. The male Pale
Grass Blue Butterfly drinks from puddles to bioconcentrate sodium through voltage-gated sodium channels in the rear hindgut. Due to sodium channel permeation, Cs-137 uptake occurs. 48
The cesium transfer from spermatophore to the female and then the egg, develops the proposed calculation. The internal absorbed dose for photons (xray, gamma) and beta (including ce,
Auger) was calculated for 1 day and 4-days to account for the number of days for egg development.
The internal absorbed dose calculation utilized equations 2.8 and 2.13 to give:
-10 ∙ D = 1.6x10 [ ] x Us x ΣSEE x 86,400 [ ] (Eq. 3.1) ∙ where the D is expressed in Gy, 1.6x10–10 converts MeV/g to J/kg (J. E. Martin, 2013), where
1 J/kg = 1 Gy; Us is the number of nuclear transitions, where only the physical radioactive decay
(no biological decay) is used due to a ‘sink model’ for the 4-day egg stage; SEE is the specific effective energy expressed in MeV/g; and 86,400 s/d converts days to seconds when the decay constant is expressed in days (Turner, 2007). The activity per egg (E), expressed in Bq, was determined by the amount of intake (Y) and the f1 uptake (Z) with:
Intake (Y) [Bq] = C x D (Eq. 3.2.1) E [Bq] = Y x Z (Eq. 3.2.2)
Where C is the concentration of Cs-137 in water expressed in Bq/L, D is intake of water expressed in liters, and Z is the uptake, or fraction (f1) of ingested radioactivity that enters the hemolymph. The specific effective energy is calculated with equations 2.5 and 2.12 without the radiation weighting factor, to calculate Gy instead of Sv by:
SEE = [ ] (Eq. 3.3)
Y is the yield and E is the radiation energy expressed in MeV, given in Tables 2.5 and 2.6. AF is the absorbed fraction which is interpolated from Tables 2.3 and 2.4, and M is the mass of the exposed material (egg) expressed in grams.
49
3.2 EXTERNAL ABSORBED DOSE
The Literature Review provided information on the Fukushima accident, Cs-137 radiation decay and dose, and applicable information on the Pale Grass Blue Butterfly. These concepts and parameters were then assigned to appropriate textbook calculations, with an assumed homogenous surface contamination on the ground and the top of the Oxalis leaf. The calculation then applied the concept as if the egg was laid in the middle of the 3 Oxalis leaves. It was then broken down to account for photon and beta radiation absorbed dose per day, and a
4-day absorbed dose to account for the number of days for egg development.
The external calculation for betas utilized eq. 2.3, 2.17 and 2.19, where φo is the energy
2 fluence rate, Ca is the surface concentration expressed in Bq per cm and Ē is the average beta energy expressed in MeV, with the calculation given by:
-10 -(µ*d) / φo = 3.6x10 x Ca x Ē x e [ ] (Eq. 3.4) (Cember & Johnson, 2009)
Beta absorption coefficients for air and tissue as a function of Eβmax is given as:
-1.37 µβ(tissue)=18.6(Eβmax -0.036) [ ] (Eq. 3.5.1) -1.4 µβ(air)=18.6(Eβmax-0.036) (Eq. 3.5.2) -1.14 µβ(other)=17(Eβmax) (Eq. 3.5.3) (J. E. Martin, 2013)
The beta dose rate above the contaminated area is then calculated by:
/ Dβ = φo [ ] x µβ,t [ ] [ ] (Eq. 3.6) / 10-6[ ] (Cember & Johnson, 2009)
The external absorbed dose for photons utilized eq. 2.15 and 2.16, where flux is described below. E is interpolated with Tables 2.5 and 2.6, the linear attenuation coefficient (µm) is 50 interpolated in Table 4.10 with assumption of material composition of the reference organisms taken to be the same as for skeletal muscle (Prohl, 2004). Each parameter is given in the calculation with:
-13 -1 D = ϕ [ ] x E [ ] x 1.6 x 10 [ ] x µm [cm ] [ ] (eq. 3.7) ∙ / ρm [ ] x [ 1 ] (Cember & Johnson, 2009)
The external flux (ϕ) applied eq. 2.15, where S is the surface activity expressed in Bq per cm2,
R is the radius of the source expressed in cm, and x is the distance from the source to the egg expressed in cm:
ϕ u = × ln( ) (Eq. 3.8)
The physical half-life (30 years) is much greater than the time of exposure (4 days), thus it was assumed there was little to no decrease in activity during the exposure timeframe. On the other hand, the biological half-life shown in Table 2.1 is much shorter than the physical half-life.
Eq. 2.3 provides the calculation to account for decrease in biological (plant surface) half-life activity, where λ is the decay constant, t is the time, and Ao is the original activity with:
-λt A(t) = Ao e (Eq. 3.9)
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4. RESULTS
4.1 Internal Absorbed Dose Due to Puddling Behavior
Table 4.1: Egg Mass Calculation
egg diameter,mm egg volume, cm3 density (ρ), g/cm3 (M) egg mass,g 0.4 0.00004 1.075 0.000043 Tan, 2014 García-Barros, 1996 L. hippothoe (same family) Egg vol*Density Table 4.2: Lycaena hippothoe Egg Density Calculation
Lycaena hippothoe egg mass (g) egg volume mm3 egg vol cm3 egg density (g/cm3) 0.000086 0.08 0.00008 1.075 Fischer & Fiedler, 2001 García-Barros, 1996 Egg vol*0.001 Egg mass(g)/vol(cm3) Table 4.3: Sodium (Na), Cs-137, and Nuclear Transformations (nt) per Egg
Na to all eggs (mg) Total eggs per pair mated Na Each egg (mg) Uptake (f1) 0.005 104 4.8077E-05 0.017 Smedley, 1996 (Hiyama, 2010) Na to Eggs/Total Eggs Hille, 1972
River Na (9ppm=9mg/L) Water intake (L) River Cs-137 (Bq/kg=Bq/L) Activity Intake (Bq) 9 5.34E-06 2 1.07E-05 Lenntech, 2020 Na each egg (mg) / (9mg Na/L water) NRA Japan, 2012 2 Bq/L * L water
Activity per Egg (Bq) λ (day-1) nt 1 day each egg nt 4 days each egg 1.82E-07 6.32E-05 1.82E-07 7.26E-07 Intake * Uptake Ln(2) / (30.04 y * 365 d/y) ∫ Ae^-λt → A/λ(1-e^-λt) (Martin, 2013) Table 4.4: Internal MeV/g (x and γ; beta, ce auger)
Σ Y*E*AF (x and γ) [MeV/nt] MeV/g x and γ radiation 3.62E-02 8.41E+02 See notes: Sum off all gamma and x ray (Y*E*AF) (Y*E*AF)/M
Σ Y*E*AF (Beta ce auger) [MeV/nt] MeV/g beta, ce and Auger 0.1568 3.65E+03 See notes: Sum off all beta ce and auger (Y*E*AF) (Y*E*AF)/M
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Table 4.5: Sum of Cs-137 Decay Yield, Energy, and Absorbed Fraction
Yield Energy (MeV) Absorbed Fraction Y∙E∙AF β- 1 0.946 0.1734 0.73 0.119747 β- 2 0.054 0.4246 0.48 0.011006 Gamma 1 0.898 0.6616 0.06 0.035647 ce-K, gamma 1 0.0832 0.6242 0.4 0.020773 ce-L1, gamma 1 0.0119 0.6557 0.37 0.002887 ce-L2, gamma 1 0.00169 0.656 0.37 0.000410 ce-L3, gamma 1 0.00139 0.6564 0.37 0.000338 ce-M, gamma 1 0.00318 0.6606 0.36 0.000756 ce-N+, gamma 1 0.000902 0.6616 0.36 0.000215 K α1 x ray 0.0392 0.03219 0.26 0.000328 K α2 x ray 0.0213 0.03182 0.26 0.000176 Auger KLL 0.00535 0.02618 1 0.000140 Auger KLX 0.00253 0.03075 1 0.000078 Auger LMM 0.0476 0.003577 1 0.000170 Auger LMX 0.0278 0.004531 1 0.000126 Auger MXT 0.149 0.0009276 1 0.000138 ICRP 38; AF β electron by interpolation Siegel & Stabin (1994) tables; Photon AF with Martin (Table 9-5)
Table 4.6: Absorbed Fraction Interpolation calculations
electron at 0.4 MeV (given in Table) electron at 0.7 MeV (given in Table) mass g AF mass g AF 0.000043 0.65 0.000043 0.32 0.01 0.67 0.01 0.36 0.1 0.85 0.1 0.67 Beta at 0.183 MeV Beta at 0.385 MeV mass g AF mass g AF 0.000043 0.72 0.000043 0.52 0.01 0.77 0.01 0.57 0.1 0.89 0.1 0.78 Material composition of the ref organisms is taken to be the same as for skeletal muscle (Prohl, 2004). AF Tables (Siegel & Stabin, 1994).
Photon at 0.662 MeV Photon at 0.03 MeV kg AF kg AF 0.000043 0.06 0.000043 0.26 0.3 0.096 0.3 0.357 0.4 0.108 0.4 0.388 Photon AF according to Martin (Table 9-5) 53
Table 4.7: Internal Absorbed Dose Per Egg per day
Absorbed Dose (Gy) (1 day) x and γ beta ce Auger All Radiation 2.11E-09 9.15E-09 1.13E-08 D(Gy)=A*MeV/g*(1.6^-10)*(8.64^4 s/d)
Table 4.8: Internal Absorbed Dose per Egg for 4 Days
Absorbed Dose (Gy) (4 days) x and γ beta ce Auger All Radiation 8.44E-09 3.66E-08 4.51E-08
D(Gy)=A*MeV/g*(1.6^-10)*(8.64^4 s/d)
4.2 External Absorbed Dose due to Cs-137 Dispersion
Table 4.9: Egg Volume, height and Cs-137 Dispersion Activity (Bq/cm2)
Egg mass g Egg Diameter,cm Leaf Stalk/Egg to ground-cm Egg Height, cm 0.000043 0.04 7 0.02 Egg Density*Vol Tan, 2014 Ca, 2019 Eeles, 2019 Density (ρ) kg/cm3 Egg Vol cm3 Dispersion Activity Bq/cm2 0.001075 0.00004 1550 L. hippothoe García-Barros, 1996 15.5 MBq/m2 IAEA, 2015 Lycaena hippothoe egg mass (g) egg volume mm3 egg vol cm3 egg density (g/cm3) 0.000086 0.08 0.00008 1.075
Fischer & Fiedler, 2001 García-Barros, 1996 Egg vol*0.001 Egg mass(g)/vol(cm3)
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Table 4.10: Ground External Flux (Photons and Betas)
Energy Fluence Rate β (J/cm2 /h): y(i) E(i) MeV per dis, betas, ce µ(β,tissue) cm2/g 3.6x10E-10*C(Bq/cm2)*E(MeV/t)*e^-(µβ,air*d) Auger rad 0.00 2.52E-01 15.6 Energy Fluence Rate β, equation: Cember (2009) ICRP 38 µ=18.6(Em-.036)^-1.37
E(max) MeV µ(β,air) cm2/g Reduction due to air 1.173 1.34E+01 0.00 Missouri, 2005 µ=16(Em-0.036)^-1.37 e^-(µ(β,air) * d height)
MeV cm2/g Interpolation: 0.5 0.03269 NIST (2004) (photon MeV) 0.596 0.032546 egg 0.6 0.03254 MeV cm-1 Interpolation: 0.5 0.000105 (photon MeV) 0.596 0.0000992 Martin (2013) air 0.6 0.0000971 The material composition of the reference organisms is taken to be the same as for skeletal muscle (Prohl, 2004). Interpolation at 0.596 MeV, µen/ρ gives 0.032546 cm2/g. 0.032546cm2/g*1.04g/cm3=0.0338cm-1 (µ-photons)
Table 4.11: Ground Contamination Absorbed Dose (Photons and Betas)
Abs Dose x and γ Abs Dose x and γ radiation Total Absorbed dose 4d: radiation (Gy/s) (Gy/d) x and γ radiation (Gy) 6.18E-09 5.34E-04 2.14E-03 D(Gy/s)=[φ(photon/cm2s) x E(MeV/photon) x1.6*10E-13 x µ(cm-1)] / ρ(kg/cm3)
Abs Dose betas, ce and Abs Dose betas, ce and Total Absorbed Dose 4d: betas, Auger (mGy/h) Auger per day (Gy/d) ce and Auger (Gy) 0.00 0.00 0.00 Dβ(mGy/h)=Energy Fluence Rate x µβ,t [cm2/g] / 10^-6 [J/g/mGy]
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Table 4.12: Leaf Top Contamination External Flux (Photons and Betas)
Flux (photons/cm2 s): Y E MeV (x and γ) µ cm-1 (photons) Disp Act/2*ln((1)/(0.009+0.006)) 3255 0.596 0.0338 Flux Equation for plane source (Martin) ICRP 38 Interpolation - see notes
β (J/cm2 /h): 3.6x10E-10 * C(Bq/cm2) * E(MeV/t) * e^- Y E MeV (beta, ce, Auger) µ(β,t) cm2/g (µβ,leaf*d) * e^-(µβ,leaf*d) 3.021E-08 0.252 15.6 Energy Fluence Rate β (J/cm2 /h), Cember (2009) ICRP 38 µ=18.6(Em-.036)^-1.37
Table 4.13: Leaf Top Contamination Shielding and Biological Half-life Reduction
E(βmax) MeV µ(β,leaf) cm2/g Leaf thickness (cm) β Reduction due to leaf 1.173 14.17 0.009 0.880 Missouri, 2005 µ=17(Em)^-1.14 Karunanithi, 2016 (90 µm) e^-(µ(β,leaf) * d leaf thickness)
E(βmax) MeV µ(β,shell) cm2/g Egg Shell Thickness (cm) β Reduction due to egg shell 1.173 15.6 0.006 0.911 Missouri, 2005 µ=18.6(Em-.036)^-1.37 Telfer, 2009 (60 µm) e^-(µ(β,shell) * d egg thickness)
E(βmax) MeV µ(β,larvae) cm2/g Egg Height β to larvae 1.173 15.6 0.02 0.268 Missouri, 2005 µ=18.6(Em-.036)^-1.37 Eeles, 2019 1-e^-(µ(β,shell) * d egg height)
leaf t1/2 (days) λ (d^-1) 1 day Reduction from t 1/2 4 day Reduction from t 1/2 14 0.0495 0.952 0.820 Ashraf et al., 2015 λ=0.693/t1/2 e^-λt e^-λt
Table 4.14: Leaf Top Contamination Absorbed Dose (Photons and Betas)
Abs Dose x and γ Abs Dose x and γ radiation Total Absorbed dose 4d: radiation (Gy/s) (Gy) 1 day x and γ radiation (Gy) 9.76E-09 8.02E-04 2.77E-03 D(Gy/s)=φ(photon/cm2s) x E(MeV/photon)x1.6*10^-13 x µ(cm-1)/ρ(kg/cm3)
Abs Dose betas, ce and Abs Dose betas, ce and Total Absorbed Dose 4d: Auger mGy/hr Auger (Gy) 1 day betas, ce and Auger (Gy) 4.71E-01 1.08E-02 3.71E-02 Dβ(mGy/h)=Energy Fluence Rate x µβ,t [cm2/g] / 10^-6 [J/g/mGy]
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4.3 Internal + External Absorbed Dose
Table 4.15: Internal + External Absorbed Dose 1 Day
Internal Absorbed Dose 1 Day x γ radiation x γ radiation beta, ce and beta, ce and All radiation Gy All radiation mGy Gy mGy Auger Gy Auger mGy 2.11E-09 2.11E-06 9.15E-09 9.15E-06 1.13E-08 1.13E-05
External Absorbed Dose 1 Day x and γ x and γ radiation beta, ce and beta, ce and External: All All radiation mGy radiation Gy mGy Auger Gy Auger mGy radiations Gy 0.001 1.336 0.011 10.764 0.012 12.101
Internal + External Absorbed Dose 1 Day x and γ x and γ radiation betas, ce, auger betas, ce, All rad Gy All rad mGy radiation Gy mGy Gy auger mGy 0.001 1.336 0.011 10.764 0.012 12.101 Table 4.16: Internal + External Absorbed Dose 4 Days
Internal Absorbed Dose for 4 day Egg Development
x γ radiation (Gy) beta, ce and Auger (Gy) All rad (Gy) All rad (mGy)
8.44E-09 3.66E-08 4.51E-08 4.51E-05
External Absorbed Dose for 4 day Egg Development All radiation x and γ radiation (Gy) beta, ce and Auger (Gy) All radiation (Gy) (mGy) 2.77E-03 0.037 0.040 39.882
Internal + External Absorbed Dose 4 day egg development All radiation 4 All radiation x and γ radiation (Gy) betas, ce, and auger (Gy) days (Gy) (mGy) 0.003 0.037 0.040 39.882
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5. DISCUSSION
A Literature Review was performed to find a method to calculate dose to the Pale Grass
Blue Butterfly egg. Alternative reasons for the internal exposure results are rather difficult, although not impossible, to explain (Hiyama et al., 2013). One explanation can be described by puddling behavior, with Cs137 permeation through voltage-gated sodium channels in the rear hindgut, and subsequent transfer to eggs from spermatophore. Although direct measurements of absorbed dose and dose distributions in vivo would be preferable (Jeifry A. Siegel et al., 1999), radiation dose from internal emitters cannot be measured directly; it can only be calculated
(Cember & Johnson, 2009). Accepted methods for calculations and units are used to quantify probable effects. Thus, a theoretical absorbed dose calculation was conducted for the most radiosensitive life-stage of a Lepidoptera, the egg.
Recent approaches to estimating exposures to biota from radionuclides in the environment have been based on a number of simplifying assumptions that potentially lead to overestimation (IAEA, 1992). For internal exposure, it is complete energy absorption; for external exposure, it is homogeneity and infinite extent of the surrounding contaminated medium
(Taranenko, Prohl, & Gomez-Ros, 2004). Despite such uncertainties, it is believed that the consistent application of conservative assumptions and parameter choices will ensure a high degree of confidence in the conclusions yield conservative (i.e. maximum) estimates of dose rates (IAEA, 1992). For the external absorbed dose calculation, it was assumed the highest recorded grid square (15.5 MBq/m2) was homogenous. With the internal absorbed dose calculation, assumptions were based using first-order kinetics, instantaneous transfer to the blood compartment (hemolymph in insects), and no cycling. If cycling was considered Cs-137 egg 58 activity would approach zero as the Malpighian tubules excrete potassium, which is a cesium analog. Only one permeation ratio in the hindgut was taken into account, although other
‘barriers’ may exist. The f1 permeation ratio is described in ICRP 30, as a transfer from the small intestine to the body fluids. This is unlike the research conducted in this paper, where the permeation of cesium was found to be through the lower hindgut, rectal pads. As other permeation or barriers may exist beyond the hindgut, further research is necessary to study sodium transfer due to puddling. S. R. Smedley & Eisner (1996) demonstrates why further barriers may exist, as male Lepidoptera’s relinquish 10 µg of sodium at mating, of which the female then passes 5 µg to the eggs. Further barriers, or transfer ratios, may include sodium transfer from the hemolymph to spermatophore and others.
Ingestion Intake
Stomach Foregut
Small Intestine f1 Blood Midgut
Anterior Hindgut Upper Large Intestine Rear Hindgut f1 Hemolymph Upper Large Intestine Excretion Excretion
Fig 5.1: ICRP 30 GI Model vs Proposed Lepidoptera Puddling Biokinetic Model
The type of radiation played a significant role in the internal absorbed dose. In health physics literature it is assumed that all internal beta radiation is absorbed - due to its limited travel distance. Jeffry A. Siegel & Stabin (1994) demonstrated this in their research to calculate absorbed fractions on small structures containing moderate to high energy electron emitters. 59
Thus, an absorbed fraction was included in the calculation for the butterfly egg due to its extremely small volume. The internal beta dose was calculated as 9.15x10-6 mGy/d, while the gamma radiation dose was 2.11x10-6 mGy/d. Gamma radiation is known to travel long distances and interact with matter of higher density, thus absorbed fractions are easily located in literature.
As shown, beta radiation contributed the most. This in in agreement that Cesium-137 is often referred to in terms of its danger as an external gamma radiation source; but because cesium-137 is also a beta emitter, it is, however, a hazard as an internal source of radiation (Nelson, Ullberg,
Kristoffersson, & Ronnback, 1960) if enough activity is consumed. This demonstrates that when using radiation detectors and calculations, all radiation types in a radionuclide’s decay scheme with correct units (in this case Gy) should be used. The dose calculation is then applied to determine effects on the species. This is unlike the Nohara et al. (2014, 2018) studies, which focused on larvae consumption of leaves using a germanium semiconductor detector to measure
(gamma) radiation on leaves, leaving their final LD50 results in Bq/body. Instead, this paper applied units in Gy to determine likely effects to the species. The researchers may have misunderstood the difference between activity, dose equivalent, and effective dose. Otaki &
Taira (2017) responded to UNSCEAR (2015) critical comments by saying the researchers used the original values (µSv/h) obtained from a NaI(Tl) scintillation survey meter for their external research, and they ‘understand’ that Sv is used only for human doses. As shown in eq. 2.5 this may not be the case, as dose equivalent (Sv) can be calculated by the absorbed dose times the radiation quality factor (QF), with beta and photons having a QF of 1. Effective dose is also expressed in sieverts, but based on the tissue weighting factor as it relates to stochastic risk. 60
As the literature review provided information, there were additional variations to consider. Cs-137 was the focus of this paper due its description as being the radionuclide of most concern. This relates to its long half-life. The 30-year physical half-life is normally given, although as shown in Table 2.1, Cs137 has a much lower environmental and biological half-life ranging from 14 days for plant surfaces to 105 days for man. As the plant surface half-life approaches the 4-day egg development timeframe, decrease in activity was taken into account.
Thus, the external dose was more appropriately calculated for exposure from the top of the
Oxalis leaf, and from the ground surface. The ground surface exposure resulted in no beta absorbed dose to the egg due to beta’s limited travel distance, showing gamma as the most concern for external absorbed dose when the distance surpasses beta’s limited travel distance.
The Oxalis plant has 3 heart shaped leaves for each stalk with maximum measurements observed by Ca (2019) as 2.5 cm wide x 1 cm length. Thus, a conservative (maximum) approach applied the disk-shaped source equation and as if the egg is laid in the middle of the 3 leaves with R as 1 cm. Due to the egg being laid on the underside of the leaf, consideration was warranted for shielding effect by the leaf and shielding by the eggshell. Although the egg has multiple layers, only the chorion was used to calculate absorbed dose to the developing larvae. This is due to the chorion being µm thick versus other segments only nm thick. Additionally, due to proximity of the egg to the contamination, external beta radiation exposure yielded the highest absorbed dose to the egg, being 10.7 mGy/d of the total (beta + photon) 12.1 mGy/d.
There were additional variations for the butterfly’s lifecycle and previous research on its radioresistence. The approximate 6-day egg development period observed by Hiyama et. al.
(2010) was not utilized as the researchers were not able to record the exact number of days in 61 each developmental stage except the pupal stage. Detailed observations by Tan (2014) provided the 4-day egg development. In either case, external and internal radiation dose can be compared, along with any probable effects. To ensure units were consistent with literature and keep dose consistent with deterministic effects, the absorbed dose, expressed in units of Gy, was thus utilized. Based on all concepts and findings in this paper, the proposed butterfly egg absorbed dose (internal + external) due to puddling behavior is 12.1 mGy for 1-day, with the 4-day egg development total dose of 39.8 mGy. This calculated dose is higher than literature, with
Buesseler et al. (2011) showing 2-6 mSv/d on land ecosystems. This may be due to assumptions based on probability interactions, homogenous tissue assumptions, use of one layer of the eggshell, and conservative (maximum) exposure from the middle of the 3 leaves instead of observations by Tan (2014) on the underside of one leaf. Due to the high natural abnormality rate of the Pale Grass Blue butterfly and previous research on lepidoptera radioresistance, there are likely no effects to the population due to puddling and external exposure. According to
Jorgensen (2012) and Chanu & Ibotombi (2011) it takes 1,300 mSv to reduce Lepidoptera egg hatch rate by 50%, and observations by Koval (1983a) show lepidopteran cells are 52 to 104 times more radioresistant than mammal cells. The results are also comparable to Japan’s normal background radiation (2.1 mSv/year) as reported by UNSCEAR (2014).
When conducting laboratory studies to bridge the gap and better understand field studies, it is necessary to understand all behaviors of the species. Otherwise the results may not reflect those from the field. A multidisciplinary approach is essential to achieve this (UNSCEAR,
2016b). Butterfly puddling was described in this paper as one of those ‘difficult to explain internal exposures’ with research on cesium uptake due to sodium permeation. The proposed 62 internal absorbed dose due to puddling behavior is 9.56x10-6 mGy/day with an overall 4-day dose of 3.83x10-5 mGy. This is much lower than the total external dose of 39.8 mGy. The negligible internal dose is likely due to Cs-137’s low river concentration as cesium bonds strongly to soil and sodium channel’s minimal uptake, unlike potassium’s analog behavior. It is thus necessary to consider the radionuclides environmental and biological behaviors when evaluating radiological effects for a species. The strong bond to sediment was evident in the literature review findings. Keeping these two key points in mind is likely the reasoning such behaviors are not included in the total dose, with radiological studies more focused on potassium analog mechanisms. This is also the reasoning Cs134 was not considered in this paper’s calculation with little to no uptake. However, such studies may need to be included when researching long term low dose effects, such as those studies by Hiyama et. al. and Nohara et. al..
Although, there will likely be no adverse effects to the population with Lepidoptera’s high, regular abnormality rates of 20%. If the absorbed dose was more significant it would need to impact the population and not just a few. For example, in organisms whose reproductive rates are very high and on which selective pressures are strong, the value of one or even many thousands of individuals to the population may be rather insignificant (IAEA, 1992).
Research observations by Hille (1972) show a relatively low Cs permeation for sodium channels with a sequence of: Li+ ≈ Na+ > K+ > Rb+ > Cs+. Also observations by Avery (1995) show Cs+ appears to have an equal or greater affinity than K+ for transport. These observations show the mechanisms for uptake needed to be accounted, as organism behaviors and diets rely on certain nutrients. The radiation absorbed dose due to Cs-137 will be larger when biological mechanisms and behaviors relate to potassium. Absorbed dose will be minimal or negligible for 63 sodium mechanisms. Different mechanisms of uptake also pertain to plants. For example, it has been estimated that a 570-1500-fold greater accumulation of 137Cs occurs when plants are grown in water as opposed to soil, with the variability in this estimate attributed to differences in the moisture content of the soils (Ashraf et al., 2015).
Research is still ongoing for behaviors like puddling, insect digestion/excretory systems, and ion channel mechanisms, with an environmental focus starting in the late 1970’s. The ICRP put out its first recommendation for protection of the environment in 1977, updated in 1990 and broadened approach in 2007: to one that includes protection of the environment itself against the detrimental effects of radiation exposure, without unduly limiting the desirable human actions that may be associated with such exposure (ICRP, 2008). This brought activities needed to meet this challenge by identifying where the most significant gaps are in the data base and undertake research programmes to obtain the needed data (IAEA, 2002). While many environmental radiation activities and studies are ongoing, most are due to accidents. These include the
Chernobyl Nuclear Power Plant explosion and the Fukushima Nuclear Power Plant accident.
Observations and studies were thus conducted by Hiyama et. al. on the Pale Grass Blue butterfly following the FNPP accident. Additionally, while research was performed to quantify sodium channel permeation in 1972, the mechanism for sodium channel permeation is still ongoing.
Reconciling how a channel can be exquisitely selective yet let ions pass at such large rates is an unresolved intellectual challenge that has fascinated scientists ever since these channels were proposed more than 60 years ago (Nogueira & Corry, 2019). This is unlike the postassium channel, which has detailed mechanism for permeation. Additionally, the recent research by
Kolosov & O’Donnell (2019) provided the necessary parameters on Lepidoptera puddling 64 behavior and digestive/excretory system for this paper, with an understanding how the
Malpighian tubules and hindgut are part of the 2-part excretory system. It is with these ongoing, combined, and mutidisciplinary research efforts that ecological radiation effects will be undertood, whether direct or indirect.
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6. CONCLUSION
This paper analyzed literature in order to describe an alternative Cs-137 pathway to the
Pale Grass Blue Butterfly following the Fukushima Nuclear Power Plant accident. Butterfly puddling behavior provides an alternative approach as most radiological studies focus on potassium analog pathways. To better understand the process and find a method the thesis was broken down into radiological and biological portions for Cs-137 and the butterfly. The literature review finally focused on previous radiological studies leading to the egg as the most radiosensitive life-stage. After bridging the biological and radiation aspects, this paper found a method to calculate internal and external absorbed dose to the egg with probable effects. Future research will need to bridge the understanding between environmental, biological and radiological concepts to better understand how behaviors and biological mechanisms affect radiation dose to species.
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APPENDIX
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Appendix A: Abbreviations
Abbreviations: Abbreviations used in this paper; definitions are discussed, as necessary.
β Symbol for Beta (radiation)
γ Symbol for gamma (radiation)
A Mass Number ce conversion electron
Cs Cesium (Multiple ways to display isotope such as Cs-137, 137Cs, 137Cs)
ENSDF Evaluated Nuclear Structure Data File
FDNPP Fukushima Daiichi nuclear power plant
ICRP International Commission on Radiological Protection
K Potassium
LD50 Lethal Dose of 50% of population
Li Lithium
MIRD Medical Internal Radiation Dosimetry
Na Sodium
P. Pseudozizeeria (Genus of Pale Grass Blue Butterfly)
RAP Reference Animals and Plant
Rb Rubidium
T1/2 Half-life
UNSCEAR United Nations Scientific Committee on the effects of Atomic Radiation
Z Atomic Number (also the proton number)
Z. Zizeeria (Abbreviated Genus of Pale Grass Blue Butterfly; see P.)
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Abrreviations Cont. (Measurements)
µ micro- Bq Becquerel s second eV electron volt m milli- Ci Curie d day g gram c centi- Gy Gray y year k kilo- Sv Sievert
M Mega-