4.1. Exposure to Pb: a threat for the Réunion harrier?
Considering the relatively low concentrations measured in both livers and bones compared to the toxicity thresholds, we show that the Réunion harrier is not abnormally exposed to Pb. Lead is an element occurring naturally in the environment, so it is often detected at varying concentrations in biological tissues even in uncontaminated areas. Lead concentrations above toxic thresholds have been found in several species of raptors in different continents (Pain et al. 2019; Taggart et al. 2020) and are commonly related to hunting or poaching activities (Taggart et al. 2020; Green et al. 2022). In the case of the Réunion harrier, the measured concentrations never exceed the threshold values defined for different tissues (Taggart et al. 2020). This finding suggests that the Réunion harrier is not threatened currently by an overexposure to Pb due to embedded shot or Pb ingestion from prey. Lead may be accumulated non-homogeneously in the different organs of vertebrates and the information given by the measured Pb concentrations in different tissues of the same individual is therefore complementary. Bones provide information on the long-term exposure of individuals because Pb can be accumulated in this tissue for several years as it is not remobilized, except in females for the formation of eggshells (Finley and Dieter 1978). Thus, bone concentrations represent relevant indicators for studying Pb exposure of animals over their entire lifespan (Krone 2018). In our study, no individual had concentration in humerus above the toxicity threshold (10 000 µg/kg dw) and we can conclude that we got no evidence that the studied individuals experienced excessive exposure to Pb during their lifetime. However, among the 30 individuals analysed, two harriers had Pb shots in the body, these shots being considered as the cause of death. This shows the current reality of poaching that still represents a threat for the conservation of this species due to its very small population size. In these 2 harriers, no abnormal contamination was detected, probably because the Pb contained in these munitions did not have time to be spread into the body.
4.2. Mercury and Se concentrations in the Réunion harrier
The Réunion harrier exhibit relatively high concentrations of Hg in liver, more than 50% of the individuals analysed having levels over 1 600 µg/kg dw. Above this threshold, Hg is susceptible to cause deleterious effects in the Réunion harrier at the cellular level, particularly through the induction of oxidative stress (Ackerman et al. 2016). In addition, 13% of individuals have liver residues greater than 6 000 µg Hg/kg dw, which has been related to reproductive disturbance in birds such as decrease in egg weight, hatching rates and/or an increase in unfertilized eggs (Shore et al. 2011). The Réunion harrier population could suffer troubles in breeding success (Augiron 2022). Nowadays, about 200 pairs live on the island and represent the entire breeding population in the world. Current knowledges suggest that both the number of fledging per pair and the hatching success would have decrease since the 1970’s. In 1975-76, the annual fecundity was estimated at 1.4 young per pair (Clouet 1978), it was 1.2 in 2000 (Grondin and Philippe 2011) while, between 2013 and 2019, it declined to 1 young (Augiron 2022). The hatching rate of eggs measured in the period 2013–2019 was 42% (Augiron 2022), which is low compared to other harrier species (Corbacho et al. 1997; Curtis et al. 2004). Moreover, the number of fledging per egg was 0.68 in 1975–1976 and decreased to 0.28 for the period 2013–2019 but the low sample size in 1975-76 and the absence of intermediate points prevent drawing robust conclusion (Augiron 2022). Several studies reported the impact of Hg on the embryonic development of birds. Moreover, (Heinz et al. 2009) showed experimentally that raptors are among the most sensitive group to Hg that at the embryonic stage. Thus, it is not excluded that the Hg exposure was responsible or at least contribute to the low reproductive success since the transfer to the egg usually occurred from the mother (Scheuhammer 1987; Ackerman et al. 2011). The origin of Hg measured in the harriers remains undetermined. Liver concentrations were correlated to urban areas within the area of 55.5 km2 around the individuals, which suggests possible urban and/or industrial origin of Hg present in the Réunion harrier. A major part of individuals with high residues levels have been found in the vicinity of agglomerations, such as Saint-Pierre and Saint-Denis, but to our knowledge, there is no known Hg contamination in these areas. Moreover, any site polluted by Hg was listed in the Réunion Island in the GEORISQUES database (Ministère en charge de l’Environnement 2022). In birds, Hg contamination levels are mostly related to their diet, particularly when they are carnivorous and related to aquatic environments, even indirectly (Ackerman et al. 2016). In aquatic ecosystems under anaerobic conditions, Hg may be methylated as methylmercury, which is highly lipophilic. It then has increased transfer capacity in food chains according to a biomagnification process. The Réunion harrier’s diet is mainly composed of terrestrial animals notably small mammals, reptiles, or small birds (Probst 2008; Augiron 2022). In addition, some opportunistic feeding behaviours may have been observed but it rarely feeds in aquatic environments (Probst 2008). Added to the fact that Hg concentrations in harriers were not correlated to the wetland area within the areas considered these trophic traits do not support the hypothesis of a transfer from aquatic ecosystems with the current data accessible. Soils in Réunion island have moderately high concentrations of Hg, due to the volcanic origin of the island (Dœlsch et al. 2006a). The median is 0.16 mg Hg/kg (min-max: 0.03–0.81 mg/kg, (Dœlsch et al. 2006a) while it is 0.041 mg Hg/kg (min-max: 0.005–1.370 mg/kg, (Marchant et al. 2017), 0.10 mg Hg/kg (min-max: 0.07–1.22) and 0.23 mg Hg/kg (min-max: 0.07–1.53) in rural and urban soils, respectively, in the United Kingdom (Ross, S.M. et al. 2007). Some of the Réunion soils have Hg contents above the threshold value of 0.5 mg/kg proposed by the Finnish Ministry of the Environment for contaminated soils. However, all the concentrations reported in (Dœlsch et al. 2006a) are below the guide value corresponding to an environmental or health risk, i.e., 2 mg/kg (Marchant et al. 2017). We cannot exclude that the relatively high exposure of the Réunion harrier we have highlighted has as main origin the Hg occurring naturally in the Réunion Island soils. In this case, this would mean that the harrier population is naturally exposed to Hg since a very long time and thus, it would not explain the recent decrease of breeding success suggested over the last 45 years. Investigations remain to be carried out to better understand the origin and pathways of Hg transfer to Réunion harrier and whether this toxic metal may threaten its population or not.
If Se is an essential element for animals that is known for its role in Hg detoxification, it may become toxic for birds in case of overexposure (Ohlendorf and Heinz 2011). Reproductive impairment is more sensitive to Se overexposure than are the health and survival of young and adult birds (Ohlendorf and Heinz 2011). Reproductive problems, primarily deformities of embryos and hatchling failure, may occur from liver concentration of about 10 000 µg/kg dw (3 000 µg/kg wet weight) in egg-laying females. The two highest Se concentrations measured in the Reunion harrier, 8 616 and 8 069 µg/kg dw, are closed but below this threshold. They were measured in two adult females found in May and July, so, during the breeding period. Sublethal effects other than reproductive disturbance may occur for liver concentration over 30 000 µg/kg dw (10 000 µg/kg wet weight) in nestlings and adults (Ohlendorf and Heinz 2011). This suggests that Réunion harrier is likely not exposed to toxic level of Se. Considering the protective role of Se against Hg toxicity, the molar ratio Se:Hg revealed a deficiency in Se only in the individual exposed to the highest Hg concentration. For all the others, molar ratios were higher than one, which lead to assume that the Réunion harrier is able to detoxify efficiently methylmercury and limits effects. However, we have to keep in mind that in some animals such as fish, the role of Se in Hg detoxification is still a controversial issue, notably the actual Se:Hg ratio protecting against Hg toxicity that remains unclear (Burger et al. 2012).
4.3. Cadmium concentrations and multi-exposure
The Réunion harrier is not severely exposed to Cd, as shown by the relatively low concentrations measured in livers which are below the threshold of 3 000 µg/kg dw except for one individual. We show that Cd concentrations in liver increased with those of Hg. This evidence a concomitant exposure within the surrounding landscape of an individual but, to our knowledge, the potential mixture effects of these two toxic metals have never been investigated in birds. If Cd concentrations remain below toxic level, such threshold is not defined by considering exposure to other chemicals that can modulate the overall response of organisms. From a toxicological point of view, a synergistic interaction between Hg and Cd, i.e., an increased effect of the mixture compared to equitoxic exposure to single metal, was reported in invertebrates (Mohan et al. 1986; Frías-Espericueta et al. 2009). In humans, exposure to a mixture of Hg and Cd significantly reduced their concentrations in the blood, liver, kidneys and brain compared to those measured when each element is administrated alone at the same dose (Orr et al. 2018). Overall, mixture toxicity remains poorly understood in the field of toxicology and ecotoxicology and more studies are needed to conclude whether co-exposure to Cd and Hg may impact or not the Réunion harrier into the wild.