Andrea Bonisoli-Alquati

Chronic exposure to low-dose radiation at Chernobyl favours adaptation to oxidative stress in birds

Summary Ionizing radiation produces oxidative stress, but organisms can adapt to their exposure with physiological adaptive responses. However, the role of radioadaptive responses in wild populations remains poorly known. At Chernobyl, studies of birds and other taxa including humans show that chronic exposure to radiation depletes antioxidants and increases oxidative damage. Here, we present analyses of levels of the most important intracellular antioxidant (i.e. glutathione, GSH), its redox status, DNA damage and body condition in 16 species of birds exposed to radiation at Chernobyl. We use an approach that allows considering the individual bird as the sampling unit while controlling for phylogenetic effects, thus increasing the statistical power by avoiding the use of species means as done for most previous comparative studies. As a consequence, we found a pattern radically different from previous studies in wild populations, showing that GSH levels and body condition increased, and oxidative stress and DNA damage decreased, with increasing background radiation. Thus, when several species are considered, the overall pattern indicates that birds are not negatively affected by chronic exposure to radiation and may even obtain beneficial hormetic effects following an adaptive response. Analysis of the phylogenetic signal supports the existence of adaptation in the studied traits, particularly in GSH levels and DNA damage. We also show that, under equal levels of radiation, the birds that produce larger amounts of the pigment pheomelanin and lower amounts of eumelanin pay a cost in terms of decreased GSH levels, increased oxidative stress and DNA damage, and poorer body condition. Radiation, however, diminished another potential cost of pheomelanin, namely its tendency to produce free radicals when exposed to radiation, because it induced a change towards the production of less pro-oxidant forms of pheomelanin with higher benzothiazole-to-benzothiazine ratios, which may have facilitated the acclimation of birds to radiation exposure. Our findings represent the first evidence of adaptation to ionizing radiation in wild animals, and confirm that pheomelanin synthesis represents an evolutionary constraint under stressful environmental conditions because it requires GSH consumption.

Variation in sperm morphometry and sperm competition among barn swallow (Hirundo rustica) populations

Spermatozoa vary greatly in size and shape among species across the animal kingdom. Postcopulatory sexual selection is thought to be the major evolutionary force driving this diversity. In contrast, less is known about how sperm size varies among populations of the same species. Here, we investigate geographic variation in sperm size in barn swallows Hirundo rustica, a socially monogamous passerine with a wide Holarctic breeding distribution. We included samples from seven populations and three subspecies: five populations of ssp. rustica in Europe (Czech, Italy, Norway, Spain, and Ukraine), one population of ssp. transitiva in Israel, and one population of ssp. erythrogaster in Canada. All sperm traits (head length, midpiece length, tail length, and total length) varied significantly among populations. The variation among the European rustica populations was much lower than the differences among subspecies, indicating that sperm traits reflect phylogenetic distance. We also performed a test of the relationship between the coefficient of between-male variation in total sperm length and extrapair paternity levels across different populations within a species. Recent studies have found a strong negative relationship between sperm size variation and extrapair paternity among species. Here, we show a similar negative relationship among six barn swallow populations, which suggests that the variance in male sperm length in a population is shaped by the strength of stabilizing postcopulatory sexual selection.

Elevated mortality among birds in Chernobyl as judged from skewed age and sex ratios.

Background Radiation has negative effects on survival of animals including humans, although the generality of this claim is poorly documented under low-dose field conditions. Because females may suffer disproportionately from the effects of radiation on survival due to differences in sex roles during reproduction, radiation-induced mortality may result in male-skewed adult sex ratios. Methodology/Principal Finding We estimated the effects of low-dose radiation on adult survival rates in birds by determining age ratios of adults captured in mist nets during the breeding season in relation to background radiation levels around Chernobyl and in nearby uncontaminated control areas. Age ratios were skewed towards yearlings, especially in the most contaminated areas, implying that adult survival rates were reduced in contaminated areas, and that populations in such areas could only be maintained through immigration from nearby uncontaminated areas. Differential mortality in females resulted in a strongly male-skewed sex ratio in the most contaminated areas. In addition, males sang disproportionately commonly in the most contaminated areas where the sex ratio was male skewed presumably because males had difficulty finding and acquiring mates when females were rare. The results were not caused by permanent emigration by females from the most contaminated areas because none of the recaptured birds had changed breeding site, and the proportion of individuals with morphological abnormalities did not differ significantly between the sexes for areas with normal and higher levels of contamination. Conclusions/Significance These findings are consistent with the hypothesis that the adult survival rate of female birds is particularly susceptible to the effects of low-dose radiation, resulting in male skewed sex ratios at high levels of radiation. Such skewed age ratios towards yearlings in contaminated areas are consistent with the hypothesis that an area exceeding 30,000 km2 in Chernobyl’s surroundings constitutes an ecological trap that causes dramatic excess mortality.

Egg testosterone affects wattle color and trait covariation in the ring-necked pheasant

Transfer of maternal hormones to the eggs is a major source of offspring phenotypic variation. The developmental and organizational effects of egg hormones can extend into adulthood and affect behavioral and morphological traits involved in sexual and reproductive behavior, with important consequences for offspring fitness. In this study, we injected testosterone (T) in egg albumen of captive ring-necked pheasant (Phasianus colchicus) eggs. We then assessed the consequences for chick growth, cell-mediated immunity, and multiple male secondary sexual traits at maturity by comparison with a control group. We also compared the covariation between traits in the two experimental groups. We found that control males had redder wattles than males from T-injected eggs, suggesting that attractiveness and reproductive success of the offspring might vary depending on maternal transfer of T to the eggs. T treatment also modified the covariation between cell-mediated immunity and wattle coloration and between the area of the wattle and the expression of another secondary sexual trait, the ear tufts. These effects are likely to translate into fitness differences among the offspring if mate acquisition depends on the simultaneous expression of several traits that are differentially affected by the same maternal contribution. Maternal effects mediated by egg hormones might affect the fitness of the offspring not only by directional modification of phenotypic traits, but also by facilitating or inhibiting their covariation. This suggests the possibility that female choice based on the relative expression of multiple secondary sexual traits exerts a pressure on how maternal transfer of androgens contributes to developmental programs.

Ionizing radiation, antioxidant response and oxidative damage: A meta-analysis

One mechanism proposed as a link between exposure to ionizing radiation and detrimental effects on organisms is oxidative damage. To test this hypothesis, we surveyed the scientific literature on the effects of chronic low-dose ionizing radiation (LDIR) on antioxidant responses and oxidative damage. We found 40 publications and 212 effect sizes for antioxidant responses and 288 effect sizes for effects of oxidative damage. We performed a meta-analysis of signed and unsigned effect sizes. We found large unsigned effects for both categories (0.918 for oxidative damage; 0.973 for antioxidant response). Mean signed effect size weighted by sample size was 0.276 for oxidative damage and -0.350 for antioxidant defenses, with significant heterogeneity among effects for both categories, implying that ionizing radiation caused small to intermediate increases in oxidative damage and small to intermediate decreases in antioxidant defenses. Our estimates are robust, as shown by very high fail-safe numbers. Species, biological matrix (tissue, blood, sperm) and age predicted the magnitude of effects for oxidative damage as well as antioxidant response. Meta-regression models showed that effect sizes for oxidative damage varied among species and age classes, while effect sizes for antioxidant responses varied among species and biological matrices. Our results are consistent with the description of mechanisms underlying pathological effects of chronic exposure to LDIR. Our results also highlight the importance of resistance to oxidative stress as one possible mechanism associated with variation in species responses to LDIR-contaminated areas.

Abundance and genetic damage of barn swallows from Fukushima

A number of studies have assessed or modeled the distribution of the radionuclides released by the accident at the Fukushima-Daiichi Nuclear Power Plant (FDNPP). Few studies however have investigated its consequences for the local biota. We tested whether exposure of barn swallow (Hirundo rustica) nestlings to low dose ionizing radiation increased genetic damage to their peripheral erythrocytes. We estimated external radiation exposure by using thermoluminescent dosimeters, and by measuring radioactivity of the nest material. We then assessed DNA damage by means of the neutral comet assay. In addition, we conducted standard point-count censuses of barn swallows across environmental radiation levels, and estimated their abundance and local age ratio. Radioactivity of nest samples was in the range 479–143,349 Bq kg−1, while external exposure varied between 0.15 and 4.9 mGy. Exposure to radioactive contamination did not correlate with higher genetic damage in nestlings. However, at higher levels of radioactive contamination the number of barn swallows declined and the fraction of juveniles decreased, indicating lower survival and lower reproduction and/or fledging rate. Thus, genetic damage to nestlings does not explain the decline of barn swallows in contaminated areas, and a proximate mechanism for the demographic effects documented here remains to be clarified.

The effects of radiation on sperm swimming behavior depend on plasma oxidative status in the barn swallow (Hirundo rustica).

Sperm are highly susceptible to reactive oxygen species (ROS) that can damage sperm DNA and structure, resulting in reduced fertilizing capacity. Exposure to radioactive contamination can also impair sperm swimming behavior and fertilizing ability, both through a reduction of sperm DNA integrity and via an increased generation of reactive oxygen species (ROS). However, the relationship between individual oxidative status and sperm swimming behavior has never been investigated in any wild population of animals exposed to radioactive contamination. We studied the motility of sperm collected from barn swallows, Hirundo rustica, breeding under different levels of radioactive contamination following the Chernobyl accident in 1986, in relation to individual oxidative status. We tested the hypothesis that the degree of impairment of sperm swimming behavior by radioactive contamination depended on plasma antioxidant capacity, the level of reactive oxygen metabolites (ROMs) and oxidative stress (sensu Costantini et al. 2006), a better oxidative status being associated with higher sperm motility. Sperm behavior parameters were subjected to principal component (PC) analysis, which extracted four PCs explaining 86% of the variance in sperm motility. PC2, representing sperm with high track velocity and ample lateral head displacement, was significantly predicted by the interaction between radiation level and either oxidative damage or oxidative stress. Contrary to our predictions, the highest values of PC2 were associated with relatively high radiation levels, particularly for high levels of either ROMs or oxidative stress. In addition, there was a tendency for values of PC3 (representing the percent of motile sperm) and PC4 (representing slow sperm with high beat cross frequency) to depend on the interaction between radiation level and total plasma antioxidant protection. Our results confirm the importance of oxidative status in determining the genetic and physiological outcome of exposure to radioactive contamination, complementing previous studies relating sperm abnormality to circulating levels of specific antioxidants. Our results also complement previous evidence that oxidative damage of sperm was negatively related to sperm motility, thus indicating a possible trade-off in quenching pro-oxidant compounds in the plasma and the seminal fluid.

High frequency of albinism and tumours in free-living birds around Chernobyl

The effects of radioactive contamination on the phenotype of free-living organisms are poorly understood, mainly because of the difficulty of capturing the large numbers of individual specimens that are required to quantify rare events such as albinism and tumour formation. We hypothesized that the frequency of abnormalities like albinism and the frequency of radiation-induced diseases like cancer would increase with the level of background radiation, that the two markers of radiation would be positively correlated, and that the reduction in abundance of animals would be greater in species with a higher frequency of albinism and tumour formation, if these markers reliably reflected poor viability. Here we analyzed the frequency of albinistic feathers and tumours in a sample of 1669 birds captured during 2010-2012 at eight sites around Chernobyl that varied in level of background radiation from 0.02 to more than 200μSv/h. We recorded 111 cases of partial albinism and 25 cases of tumour formation. Nominal logistic models were used to partition the variance into components due to species and background radiation. Radiation was a strong predictor of the two markers in birds, with a small, but significant effect of species for albinism. The slope of the relationship between abundance and radiation in different bird species was significantly inversely correlated with the frequency of albinism and tumours, as was to be expected if a common underlying cause (i.e. radiation) affects both variables. These findings are consistent with the hypothesis that background radiation is a cause of albinism and tumours, that albinism and tumours are biomarkers of radiation exposure, and that high frequencies of albinism and tumours were present despite the low viability of birds with these conditions.

Nestling rearing is antioxidant demanding in female barn swallows (Hirundo rustica)

Reproduction is a demanding activity, since organisms must produce and, in some cases, protect and provision their progeny. Hence, a central tenet of life-history theory predicts that parents have to trade parental care against body maintenance. One physiological cost thought to be particularly important as a modulator of such trade-offs is oxidative stress. However, evidence in favour of the hypothesis of an oxidative cost of reproduction is contradictory. In this study, we manipulated the brood size of wild barn swallows Hirundo rustica soon after hatching of their nestlings to test whether an increase in nestling rearing effort translates into an increased oxidative damage and a decreased antioxidant protection at the end of the nestling rearing period. We found that, while plasma oxidative damage was unaffected by brood size enlargement, females rearing enlarged broods showed a decrease in plasma non-enzymatic antioxidants during the nestling rearing period. This was not the case among females rearing reduced broods and among males assigned to either treatment. Moreover, individuals with higher plasma oxidative damage soon after the brood size manipulation had lower plasma non-enzymatic antioxidants at the end of the nestling rearing period, suggesting that non-enzymatic antioxidants were depleted to buffer the negative effects of high oxidative damage. Our findings point to antioxidant depletion as a potential mechanism mediating the cost of reproduction among female birds.

MHC genotype predicts mate choice in the ring-necked pheasant Phasianus colchicus.

Females of several vertebrate species selectively mate with males on the basis of the major histocompatibility complex (MHC) genes. As androgen‐mediated maternal effects have long‐lasting consequences for the adult phenotype, both mating and reproductive success may depend on the combined effect of MHC genotype and exposure to androgens during early ontogeny. We studied how MHC‐based mate choice in ring‐necked pheasants (Phasianus colchicus) was influenced by an experimental in ovo testosterone (T) increase. There was no conclusive evidence of in ovo T treatment differentially affecting mate choice in relation to MHC genotype. However, females avoided mating with males with a wholly different MHC genotype compared with males sharing at least one MHC allele. Females also tended to avoid mating with MHC‐identical males, though not significantly so. These findings suggest that female pheasants preferred males with intermediate MHC dissimilarity. Male MHC heterozygosity or diversity did not predict the expression of ornaments or male dominance rank. Thus, MHC‐based mating preferences in the ring‐necked pheasant do not seem to be mediated by ornaments’ expression and may have evolved mainly to reduce the costs of high heterozygosity at MHC loci for the progeny, such as increased risk of autoimmune diseases or disruption of coadapted gene pools.