Dietary DNA metabarcoding reveals a trophic niche partitioning among sympatric Iberian sandgrouses and bustards.

The study of trophic niche partitioning is of great importance for understanding community structure and species coexistence, particularly if these are threatened. Here DNA metabarcoding was used to assess the diet of four threatened steppe bird species (two bustards and two sandgrouses), with the aim of better understanding their dietary requirements, trophic interactions, and potential threats. The results showed seasonal and interspecific differences in their plant diet, with greater importance of cultivated plants during autumn and winter (around 50 % of their diet) than spring. Plants of the genus Convolvulus and of the family Brassicaceae were frequently consumed by all species. In spring, poppies (Papaver spp.) were a considerable part of their diet, and could be used as a source of carotenoids or for their anti-parasitic properties. Furthermore, results evidenced a trophic niche partitioning among species, with a marked segregation between bustard species and, to a lesser extent, between sandgrouse species. Diet similarity was generally higher between species from different orders that occur in mixed-species flocks (bustard - sandgrouse) than between species of the same order. This partitioning was probably related to a stratification in habitat use rather than to specialisation and might prevent competition to some extent. However, the homogenization of trophic resources resulting from agricultural intensification could pose an important threat, particularly during autumn, when weeds are scarcer and the most abundant trophic resource are sown seeds, which are often treated with pesticides.

Transgenerational effects of triazole fungicides on gene expression and egg compounds in non-exposed offspring: A case study using Red-Legged Partridges (Alectoris rufa).

Triazole fungicides are widely used to treat cereal seeds before sowing. Granivorous birds like the Red-legged Partridge (Alectoris rufa) have high exposure risk because they ingest treated seeds that remain on the field surface. As triazole fungicides can act as endocrine disruptors, affecting sterol synthesis and reproduction in birds several months after exposure, we hypothesized that these effects could also impact subsequent generations of exposed birds. To test this hypothesis, we exposed adult partridges (F0) to seeds treated at commercial doses with four different formulations containing triazoles as active ingredients (flutriafol, prothioconazole, tebuconazole, and a mixture of the latter two), simulating field exposure during late autumn sowing. During the subsequent reproductive season, two to four months after exposure, we examined compound allocation of steroid hormones, cholesterol, vitamins, and carotenoids in eggs laid by exposed birds (F1), as well as the expression of genes encoding enzymes involved in sterol biosynthesis in one-day-old chicks of this F1. One year later, F1 animals were paired again to investigate the expression of the same genes in the F2 chicks. We found changes in the expression of some genes for all treatments and both generations. Additionally, we observed an increase in estrone levels in eggs from partridges treated with flutriafol compared to controls, a decrease in tocopherol levels in partridges exposed to the mixture of tebuconazole and prothioconazole, and an increase in retinol levels in partridges exposed to prothioconazole. Despite sample size limitations, this study provides novel insights into the mechanisms of action of the previously observed effects of triazole fungicide-treated seeds on avian reproduction with evidence that the effects can persist beyond the exposure windows, affecting unexposed offspring of partridges fed with treated seeds. The results highlight the importance of considering long-term chronic effects when assessing pesticide risks to wild birds.

Water and mosquitoes as key components of the infective cycle of Francisella tularensis in Europe: a review.

Francisella tularensis is the pathogen of tularemia, a zoonotic disease that have a broad range of hosts. Its epidemiology is related to aquatic environments, particularly in the subspecies holarctica. In this review, we explore the role of water and mosquitoes in the epidemiology of Francisella in Europe. F. tularensis epidemiology has been linked to natural waters, where its persistence has been associated with biofilm and amebas. In Sweden and Finland, the European countries where most human cases have been reported, mosquito bites are a main route of transmission. F. tularensis is present in other European countries, but to date positive mosquitoes have not been found. Biofilm and amebas are potential sources of Francisella for mosquito larvae, however, mosquito vector capacity has not been demonstrated experimentally, with the need to be studied using local species to uncover a potential transmission adaptation. Transstadial, for persistence through life stages, and mechanical transmission, suggesting contaminated media as a source for infection, have been studied experimentally for mosquitoes, but their natural occurrence needs to be evaluated. It is important to clear up the role of different local mosquito species in the epidemiology of F. tularensis and their importance in all areas where tularemia is present.

Exploring the influence of density-dependence and weather on the spatial and temporal variation in common vole (Microtus arvalis) abundance in Castilla y León, NW Spain.

BACKGROUND: The common vole has invaded the agroecosystems of northwestern Spain, where outbreaks cause important crop damage and management costs. Little is yet known about the factors causing or modulating vole fluctuations. Here, we used 11 years of vole abundance monitoring data in 40 sites to study density-dependence and weather influence on vole dynamics. Our objective was to identify the population dynamics structure and determine whether there is direct or delayed density-dependence. An evaluation of climatic variables followed, to determine whether they influenced vole population peaks. RESULTS: First- and second-order outbreak dynamics were detected at 7 and 33 study sites, respectively, together with second-order variability in periodicity (2-3 to 4-5-year cycles). Vole population growth was explained by previous year abundance (mainly numbers in summer and spring) at 21 of the sites (52.5%), by weather variables at 11 sites (27.5%; precipitation or temperature in six and five sites, respectively), and by a combination of previous abundance and weather variables in eight sites (20%). CONCLUSIONS: We detected variability in vole spatiotemporal abundance dynamics, which differs in cyclicity and period. We also found regional variation in the relative importance of previous abundances and weather as factors modulating vole fluctuations. Most vole populations were cyclical, with variable periodicity across the region. Our study is a first step towards the development of predictive modeling, by disclosing relevant factors that might trigger vole outbreaks. It improves decision-making processes within integrated management dealing with mitigation of the agricultural impacts caused by voles. © 2023 Society of Chemical Industry.

Diet and Spatial Ecology Influence Red-Legged Partridge Exposure to Pesticides Used as Seed Treatment.

Seed treatment with pesticides is an extended agricultural practice with a high risk to granivorous birds that consume those seeds. To characterize that risk, it is necessary to understand the ecological factors that determine the exposure chances of birds to treated seeds. We investigated how pesticide uptake by red-legged partridges was related to cultivated plant ingestion and to the use of recently sown fields. We analyzed pesticide residues in 144 fecal samples from 32 flocks and determined the plant diet composition using DNA metabarcoding. Habitat use was studied through the monitoring of 15 GPS-tagged partridges. We confirmed, through the analysis of seeds, that >80% of cereal fields from the area had seeds treated with triazole fungicides. Tebuconazole was detected in 16.6% of partridges' feces. During the sowing season, cultivated plants accounted for half of the plant diet, but no association was found between cultivated plant consumption and pesticide intake. GPS tracking revealed that tebuconazole was detected in feces when partridges had recently used sown fields, whereas nonexposed partridges showed no overlap with recently sown areas. Our results highlight the need to incorporate field ecology into the characterization of pesticide exposure to improve the efficacy of environmental risk assessment.

Vole outbreaks may induce a tularemia disease pit that prevents Iberian hare population recovery in NW Spain.

Iberian hare populations have suffered severe declines during recent decades in Spain. Between 1970 and 1990s, a rapid increase in irrigation crop surface in NW Spain (Castilla-y-León region) was followed by a common vole massive range expansion and complete colonization of lowland irrigated agricultural landscapes from mountainous habitats. The subsequent large cyclic fluctuations in abundance of colonizing common voles have contributed to a periodic amplification of Francisella tularensis, the etiological agent that causes human tularemia outbreaks in the region. Tularemia is a fatal disease to lagomorphs, so we hypothesize that vole outbreaks would lead to disease spill over to Iberian hares, increasing prevalence of tularemia and declines among hare populations. Here we report on the possible effects that vole abundance fluctuations and concomitant tularemia outbreaks had on Iberian hare populations in NW Spain. We analysed hare hunting bag data for the region, which has been recurrently affected by vole outbreaks between 1996 and 2019. We also compiled data on F. tularensis prevalence in Iberian hares reported by the regional government between 2007 and 2016. Our results suggest that common vole outbreaks may limit the recovery of hare populations by amplifying and spreading tularemia in the environment. The recurrent rodent-driven outbreaks of tularemia in the region may result in a "disease pit" to Iberian hares: at low host densities, the rate of population growth in hares is lower than the rate at which disease-induced mortality increases with increased rodent host density, therefore, keeping hare populations on a low-density equilibrium. We highlight future research needs to clarify tularemia transmission pathways between voles and hares and confirm a disease pit process.

A non-invasive method to monitor farmland bird exposure to triazole fungicides.

The treatment of seeds with pesticides is an extended practice in current agriculture. There is a high risk of exposure in granivorous birds, such as the red-legged partridge (Alectoris rufa), that can consume those seeds remaining on the surface during sowing. Fungicide exposure could in turn affect bird reproductive capacity. To better understand to what extent triazole fungicides are a threat to granivorous birds, we need an easy and reliable method to quantify field exposure. In this study, we tested a novel non-invasive method to detect the presence of triazole fungicide residues in farmland bird faeces. We experimentally exposed captive red-legged partridges to validate the method, and then applied it in a real scenario to assess exposure of wild partridges. We exposed adult partridges to seeds treated with two formulations containing triazole fungicides as active ingredients: Vincit®Minima (flutriafol 2.5%) and Raxil®Plus (prothioconazole 25% and tebuconazole 15%). We collected two types of faeces (caecal and rectal samples) immediately after exposure and 7 days later and quantified the concentrations of the three triazoles and their common metabolite (1,2,4-triazole). The three active ingredients and 1,2,4-triazole were only detected in faeces collected immediately after exposure. Triazole fungicide detection rates in rectal stool were 28.6%, 73.3% and 80% for flutriafol, prothioconazole and tebuconazole, respectively. In caecal samples, detection rates were 40%, 93.3% and 33.3%, respectively. 1,2,4-triazole was detected in 53% of rectal samples. For an applied use of the method in the field, we collected 43 faecal samples from wild red-legged partridges during autumn cereal seed sowing and found detectable levels of tebuconazole in 18.6% of the analysed wild partridges. The results of the experiment were then used to estimate actual exposure levels from this prevalence value found in wild birds. Our study shows that faecal analysis can be a useful tool to assess farmland bird exposure to triazole fungicides, when samples are fresh and the method has been validated for the detection of target molecules.

Strengthening the genomic surveillance of Francisella tularensis by using culture-free whole-genome sequencing from biological samples.

Introduction: Francisella tularensis is a highly infectious bacterium that causes the zoonotic disease tularemia. The development of genotyping methods, especially those based on whole-genome sequencing (WGS), has recently increased the knowledge on the epidemiology of this disease. However, due to the difficulties associated with the growth and isolation of this fastidious pathogen in culture, the availability of strains and subsequently WGS data is still limited. Methods: To surpass these constraints, we aimed to implement a culture-free approach to capture and sequence F. tularensis genomes directly from complex samples. Biological samples obtained from 50 common voles and 13 Iberian hares collected in Spain were confirmed as positive for F. tularensis subsp. holarctica and subjected to a WGS target capture and enrichment protocol, using RNA oligonucleotide baits designed to cover F. tularensis genomic diversity. Results: We obtained full genome sequences of F. tularensis from 13 animals (20.6%), two of which had mixed infections with distinct genotypes, and achieved a higher success rate when compared with culture-dependent WGS (only successful for two animals). The new genomes belonged to different clades commonly identified in Europe (B.49, B.51 and B.262) and subclades. Despite being phylogenetically closely related to other genomes from Spain, the detected clusters were often found in other countries. A comprehensive phylogenetic analysis, integrating 599 F. tularensis subsp. holarctica genomes, showed that most (sub)clades are found in both humans and animals and that closely related strains are found in different, and often geographically distant, countries. Discussion: Overall, we show that the implemented culture-free WGS methodology yields timely, complete and high-quality genomic data of F. tularensis, being a highly valuable approach to promote and potentiate the genomic surveillance of F. tularensis and ultimately increase the knowledge on the genomics, ecology and epidemiology of this highly infectious pathogen.

Protect European green agricultural policies for future food security.

European green agricultural policies have been relaxed to allow cultivation of fallow land to produce animal feed and meet shortfalls in exports from Ukraine and Russia. However, conversion of semi-natural habitats will disproportionately impact long term biodiversity and food security.

From pattern to process? Dual travelling waves, with contrasting propagation speeds, best describe a self-organised spatio-temporal pattern in population growth of a cyclic rodent.

The dynamics of cyclic populations distributed in space result from the relative strength of synchronising influences and the limited dispersal of destabilising factors (activators and inhibitors), known to cause multi-annual population cycles. However, while each of these have been well studied in isolation, there is limited empirical evidence of how the processes of synchronisation and activation-inhibition act together, largely owing to the scarcity of datasets with sufficient spatial and temporal scale and resolution. We assessed a variety of models that could be underlying the spatio-temporal pattern, designed to capture both theoretical and empirical understandings of travelling waves using large-scale (>35,000 km2 ), multi-year (2011-2017) field monitoring data on abundances of common vole (Microtus arvalis), a cyclic agricultural rodent pest. We found most support for a pattern formed from the summation of two radial travelling waves with contrasting speeds that together describe population growth rates across the region.

Viral Zoonoses in Small Wild Mammals and Detection of Hantavirus, Spain.

We screened 526 wild small mammals for zoonotic viruses in northwest Spain and found hantavirus in common voles (Microtus arvalis) (1.5%) and high prevalence (48%) of orthopoxvirus among western Mediterranean mice (Mus spretus). We also detected arenavirus among small mammals. These findings suggest novel risks for viral transmission in the region.

Bird exposure to fungicides through the consumption of treated seeds: A study of wild red-legged partridges in central Spain.

Sown seeds are a key component of many farmland birds' diets due to natural food shortages in autumn and winter. Because these seeds are often treated with pesticides, their ingestion by birds can result in toxic effects. For risk assessment, data on treated seed toxicity should be combined with information about exposure risk for wild birds and the factors that modulate it. We characterized the exposure of red-legged partridges to pesticide-treated seeds through the analysis of digestive contents of birds shot by hunters (n = 194) in an agricultural region in central Spain. We measured the contribution of sown seeds to the partridges' diet and how it related to pesticide exposure. Moreover, we evaluated the influence of landscape composition on the intake of sown seeds and pesticides by partridges. During peak sowing time, seeds constituted half (50.7%) of the fresh biomass ingested by partridges, which consumed mostly winter cereal seeds (42.3% of biomass). Residues of seven fungicides and one insecticide (active ingredients) were detected in 33.0% of birds. The presence of pesticides in digestive contents was linked to the ingestion of cereal sown seeds. Moreover, dietary exposure of birds to pesticides was modulated by landscape characteristics, being lower in areas with heterogeneous landscapes, greater habitat mosaic and more natural vegetation. The estimated dietary intake of pesticides resulting from our field observations, in combination with experimental data on pesticide toxicity, raise concerns about the risks that pesticide-treated cereal seeds pose to granivorous bird populations. Our results highlight the importance of farming landscape composition and diversification, which should be considered as a priority in the agricultural policy to mitigate pesticide risks to farmland birds through the consumption of treated seeds.

Metabarcoding insights into the diet and trophic diversity of six declining farmland birds.

Knowledge of feeding ecology of declining species, such as farmland birds, is essential to address their conservation requirements, especially when their habitats are suffering important reductions of trophic resources. In this study, we apply a metabarcoding approach to describe the diet composition of six of the most significant farmland birds inhabiting European cereal pseudo-steppes: little bustard, great bustard, pin-tailed sandgrouse, black-bellied sandgrouse, red-legged partridge, and common quail. We further studied seasonal diet variations (autumn to spring) in all species but the common quail, whose diet was studied during spring and summer. We show that study species´ diets mostly consisted of plants, although in the case of little bustard and great bustard arthropods are also highly relevant. Among arthropods, we found high proportions of thrips, arachnids, and springtails, which were previously unreported in their diet, and some taxa that could be used as antiparasitic food. Moreover, we report that little bustard's diet is the least rich of that of all studied species, and that diet of all these species is less diverse in winter than in autumn and spring. Diet composition of these declining species supports the importance of natural and semi-natural vegetation and landscape mosaics that can provide a wide variety of arthropods, plants, and seeds all year-round.

Do human infrastructures shape nest distribution in the landscape depending on individual personality in a farmland bird of prey?

Individuals' distribution across habitats may depend on their personality. Human activities and infrastructures are critical elements of the landscape that may impact the habitat selection process. However, depending on their personality, individuals may respond differently to these unnatural elements. In the present study, we first investigated whether some human infrastructures (buildings, roads and paths) shaped Montagu's harrier nest spatial distribution in the landscape according to female personality (boldness). Second, we tested if the reproductive success of females depended on their boldness and nest location regarding infrastructures. Using a long-term (19 years) dataset, we calculated, for each infrastructure type, the distance from each nest to the nearest infrastructure and the infrastructure density around the nest. We tested the effects of female boldness (bold vs. shy) and its interaction with egg-laying date on these six metrics. Nest location in the landscape depended on female personality and on some human infrastructures: the building density was smaller around nests from shy females than from bold ones. Nest distribution related to other infrastructure metrics did not depend on female boldness. The pattern related to building density is consistent with some habitat choice hypotheses, which are discussed. Path density around nests negatively affected reproductive success regardless of female boldness, and late breeders nested further away from paths than early breeders. Human activities on paths (more common later in the season) could lead to disturbance and a decrease in parental care, reducing reproductive success. Increasing human presence in farmlands implies a need to better understand its impact on population composition, in terms of personality. Our results suggest that individual behavioural differences should be taken into account in studies assessing the effects of human disturbance on animal populations, to propose more appropriate conservation measures.

La distribution des individus au sein de l'habitat peut dépendre de leur personnalité. Les activités et infrastructures humaines sont des éléments importants du paysage qui peuvent impacter le processus de sélection des habitats. Cependant, en fonction de leur personnalité, les individus peuvent répondre différemment à ces éléments non naturels. Dans cette étude, nous avons d'abord examiné si certaines infrastructures humaines (bâtiments, routes et chemins) influençaient la répartition spatiale des nids de busard cendré en fonction de la personnalité des femelles (témérité). Ensuite, nous avons testé si le succès reproducteur des femelles dépendait de leur témérité et de la localisation du nid par rapport aux infrastructures. En utilisant des données à long terme (19 ans), nous avons calculé, pour chaque type d'infrastructure, la distance de chaque nid à la plus proche infrastructure et la densité en infrastructures autour de chaque nid. Nous avons testé les effets de la témérité des femelles (téméraire vs. timide) et son interaction avec la date de ponte sur les six métriques. La répartition spatiale des nids dépendait de la personnalité des femelles et de certaines infrastructures humaines: la densité en bâtiments était plus faible autour des nids de femelles timides qu'autour de ceux des téméraires. La distribution des nids par rapport aux autres métriques d'infrastructure n'était pas influencée par la témérité des femelles. Le patron lié à la densité en bâtiments est en accord avec certaines hypothèses de choix d'habitat qui sont discutées. La densité en chemins autour des nids affectait négativement le succès reproducteur indépendamment de la témérité des femelles, et les femelles tardives nichaient plus loin des chemins que les précoces. Les activités humaines sur les chemins (plus fréquentes tard dans la saison de reproduction) pourraient entrainer une perturbation et une diminution des soins parentaux, réduisant ainsi le succès reproducteur. L'augmentation de la présence humaine dans les zones agricoles implique la nécessité de mieux comprendre son impact sur la composition de la population, en termes de personnalité. Nos résultats suggèrent que les différences comportementales interindividuelles devraient être prises en compte dans les études évaluant les effets des perturbations humaines sur les populations animales afin de proposer des mesures de conservations plus appropriées.

Linking Zoonosis Emergence to Farmland Invasion by Fluctuating Herbivores: Common Vole Populations and Tularemia Outbreaks in NW Spain.

The expansion and intensification of agriculture are driving profound changes in ecosystems worldwide, favoring the (re)emergence of many human infectious diseases. Muroid rodents are a key host group for zoonotic infectious pathogens and frequently invade farming environments, promoting disease transmission and spillover. Understanding the role that fluctuating populations of farm dwelling rodents play in the epidemiology of zoonotic diseases is paramount to improve prevention schemes. Here, we review a decade of research on the colonization of farming environments in NW Spain by common voles (Microtus arvalis) and its public health impacts, specifically periodic tularemia outbreaks in humans. The spread of this colonizing rodent was analogous to an invasion process and was putatively triggered by the transformation and irrigation of agricultural habitats that created a novel terrestrial-aquatic interface. This irruptive rodent host is an effective amplifier for the Francisella tularensis bacterium during population outbreaks, and human tularemia episodes are tightly linked in time and space to periodic (cyclic) variations in vole abundance. Beyond the information accumulated to date, several key knowledge gaps about this pathogen-rodent epidemiological link remain unaddressed, namely (i) did colonizing vole introduce or amplified pre-existing F. tularensis? (ii) which features of the "Francisella-Microtus" relationship are crucial for the epidemiology of tularemia? (iii) how virulent and persistent F. tularensis infection is for voles under natural conditions? and (iv) where does the bacterium persist during inter-epizootics? Future research should focus on more integrated, community-based approaches in order to understand the details and dynamics of disease circulation in ecosystems colonized by highly fluctuating hosts.

Farmland composition and farming practices explain spatio-temporal variations in red-legged partridge density in central Spain.

Many farmland bird populations are declining, and their negative trends are often associated with changes in land-use or farming practices, including the use of agrochemicals. The red-legged partridge (RLP) is a Mediterranean farmland game species of high socio-economic importance whose populations are thought to have declined sharply since the mid-20th century associated with farmland changes. However, no large-scale studies have tested whether abundance or trends of RLP are related to farmland composition or management. We used hierarchical distance sampling models to estimate RLP abundance in 2010 in central Spain (Castilla-La Mancha), a main European population stronghold of this species. We studied associations between RLP density and land-uses (including variation in management: irrigated crops or organic farming). We also assessed regional abundance variation over seven years (2010-2017) and its relationship with changes in land-use. Our results show that RLP abundance increased with the availability of natural vegetation and traditional rain-fed vineyards, but decreased with increasing proportions of tree crops and irrigated vineyards; the latter association was less pronounced in areas sensitive to nitrate contamination in water, where the amount of fertilizers applied in farmland and use of certain farming practices is more strictly regulated. These results support the idea that increases in intensive vineyards are detrimental to the RLP. We also report a strong population decline of RLP in the region, with a 51% abundance reduction in seven years. This decline was steeper in areas where more natural vegetation had been lost and where ecological tree crops had increased. Overall, our results indicate that changes in land-use (type of crop, or the destruction of natural vegetation in farmland) and farming practices (e.g. use of irrigation in certain crops, use of nitrates) have important impacts on this farmland bird, affecting both spatial distribution and population dynamics.

Patterns of flea infestation in rodents and insectivores from intensified agro-ecosystems, Northwest Spain.

BACKGROUND: Fleas frequently infest small mammals and play important vectoring roles in the epidemiology of (re)emerging zoonotic disease. Rodent outbreaks in intensified agro-ecosystems of North-West Spain have been recently linked to periodic zoonotic diseases spillover to local human populations. Obtaining qualitative and quantitative information about the composition and structure of the whole flea and small mammal host coexisting communities is paramount to understand disease transmission cycles and to elucidate the disease-vectoring role of flea species. The aims of this research were to: (i) characterise and quantify the flea community parasiting a small mammal guild in intensive farmlands in North-West Spain; (ii) determine and evaluate patterns of co-infection and the variables that may influence parasitological parameters. METHODS: We conducted a large-scale survey stratified by season and habitat of fleas parasitizing the small mammal host guild. We report on the prevalence, mean intensity, and mean abundance of flea species parasitizing Microtus arvalis, Apodemus sylvaticus, Mus spretus and Crocidura russula. We also report on aggregation patterns (variance-to-mean ratio and discrepancy index) and co-infection of hosts by different flea species (Fager index) and used generalized linear mixed models to study flea parameter variation according to season, habitat and host sex. RESULTS: Three flea species dominated the system: Ctenophthalmus apertus gilcolladoi, Leptopsylla taschenbergi and Nosopsyllus fasciatus. Results showed a high aggregation pattern of fleas in all hosts. All host species in the guild shared C. a. gilcolladoi and N. fasciatus, but L. taschenbergi mainly parasitized mice (M. spretus and A. sylvaticus). We found significant male-biased infestation patterns in mice, seasonal variations in flea abundances for all rodent hosts (M. arvalis, M. spretus and A. sylvaticus), and relatively lower infestation values for voles inhabiting alfalfas. Simultaneous co-infections occurred in a third of all hosts, and N. fasciatus was the most common flea co-infecting small mammal hosts. CONCLUSIONS: The generalist N. fasciatus and C. a. gilcolladoi dominated the flea community, and a high percentage of co-infections with both species occurred within the small mammal guild. Nosopsyllus fasciatus may show higher competence of inter-specific transmission, and future research should unravel its role in the circulation of rodent-borne zoonoses.

Birds feeding on tebuconazole treated seeds have reduced breeding output.

Drilled seeds are an important food resource for many farmland birds but may pose a serious risk when treated with pesticides. Most compounds currently used as seed treatment in the EU have low acute toxicity but may still affect birds in a sub-chronic or chronic way, especially considering that the sowing season lasts several weeks or months, resulting in a long exposure period for birds. Tebuconazole is a triazole fungicide widely used in agriculture but its toxicity to birds remains largely unknown. Our aim was to test if a realistic scenario of exposure to tebuconazole treated seeds affected the survival and subsequent reproduction of the red-legged partridge (Alectoris rufa). We fed captive partridges with wheat seeds treated with 0%, 20% or 100% of tebuconazole application rate during 25 days in late winter (i.e. tebuconazole dietary doses were approximately 0.2 and 1.1 mg/kg bw/day). We studied treatment effects on the physiology (i.e. body weight, biochemistry, immunology, oxidative stress, coloration) and reproduction of partridges. Exposed birds did not reduce food consumption but presented reduced plasmatic concentrations of lipids (triglycerides at both exposure doses, cholesterol at high dose) and proteins (high dose). The coloration of the eye ring was also reduced in the low dose group. Exposure ended 60 days before the first egg was laid, but still affected reproductive output: hatching rate was reduced by 23% and brood size was 1.5 times smaller in the high dose group compared with controls. No significant reproductive effects were found in the low dose group. Our results point to the need to study the potential endocrine disruption mechanism of this fungicide with lagged effects on reproduction. Risk assessments for tebuconazole use as seed treatment should be revised in light of these reported effects on bird reproduction.

Egg Overspray with Herbicides and Fungicides Reduces Survival of Red-Legged Partridge Chicks.

Within the environmental risk assessment conducted for pesticide registration in the European Union (EU), avian reproductive toxicity is characterized after exposing adults. However, eggs of ground-nesting species can be exposed when pesticide applications occur during laying or incubation. We simulated environmentally realistic exposure of red-legged partridge (Alectoris rufa) eggs to an herbicide (2,4-D) and a fungicide (tebuconazole) applied to winter cereal crops during the breeding season of most farmland birds. We analyzed the effects on hatching success, offspring survival, and physiology. Exposure by overspray led to greater pesticide accumulation in the eggshell or content than exposure through contact with treated soil (3.1-13.7 times higher, depending on the pesticide and target sample). Egg overspray with tebuconazole significantly increased chick mortality, which was 26% higher than that of controls. 2,4-D caused a similar but a close to significant increase (chick mortality 24% higher than controls). Exposure to either pesticide through contact with treated soils did not affect chick survival but altered some biochemical parameters posthatching. Our experiment shows that egg spraying with pesticides should be considered as a relevant exposure scenario in risk assessment procedures, given its potential to affect the reproductive success of ground-nesting farmland birds.

Multi-level analysis of exposure to triazole fungicides through treated seed ingestion in the red-legged partridge.

Triazole fungicides are the most widely used products to treat cereal seeds. Granivorous birds, such as red-legged partridges (Alectoris rufa), which consume seeds left on the surface of fields after sowing, have a high risk of exposure. As triazole fungicides can affect sterol synthesis, we tested the hypothesis that treated seed consumption could alter the synthesis of sex hormones and reduce the reproductive capacity of partridges. We exposed adult partridges to seeds treated with four different formulations containing triazoles as active ingredients (flutriafol, prothioconazole, tebuconazole, and a mixture of the latter two) simulating a field exposure during the late autumn sowing season. All treatments produced biochemical changes and an overexpression of genes encoding for enzymes involved in the biosynthesis of sterols and steroid hormones, such as PMVK, ABCA1, MVD, PSCK9, DHCR7 and HSD17B7. Plasma levels of oestradiol were reduced in partridges exposed to tebuconazole. We also monitored reproduction 3 months after exposure (laying date, egg fertilization and hatching rates). We observed a 14-day delay in the laying onset of partridges that had been exposed to flutriafol as compared to controls. These results show that the consumption of seeds treated with triazole fungicides has the potential to affect granivorous bird reproduction. We recommend the evaluation of lagged reproductive effects as part of the protocols of environmental risk assessment of pesticides in wild birds in light of the effects resulting from the exposure to triazole-treated seeds.