Jérôme Moreau

Karine Monceau

Denis Thiery
email: thiery@bordeaux.inra.fr

Denis Thiéry

Larval food influences temporal oviposition and egg quality traits in females of Lobesia botrana

Keywords: Lobesia botrana, fecundity, egg size, percent egg hatch, cultivars, female age

Many phytophagous insects are agricultural pests, and control methods require accurate monitoring and decisions based on the determination of population age structure. The reproductive output (fecundity, egg size and percent egg hatch) is a central life history trait because it determines the offspring number, and temporal oviposition patterns are of primary importance in conditioning larval hatching and the occurrence of later larval instars in time. In turn, these phenomena determine the window for natural enemy attack and thus impact the context of biological control programs. In addition, for most phytophagous insects, the quality of the host plants that larvae consume determines the insects' reproductive output. The purpose of the present study was to determine whether the number of eggs laid, egg size and egg hatch percentage vary with female age and the cultivar on which females develop as larvae, as well as the temporal effects of these parameters. This determination was performed in laboratory experiments where larvae were reared on artificial diets based on dried fruits of seven cultivars. Our results showed that the cultivars had a significant effect on female temporal oviposition. Independent of the food tested, the numbers of oviposited eggs, their size and percent egg hatch decreased with daily oviposition rank. Such temporal patterns must be incorporated in age-structured mathematical models used in the design of control strategies. Temporal oviposition and variation in egg quality traits will also be useful in biological control programs, especially when based on egg or larval parasitoids, which is thus discussed.

Introduction

The abundance of phytophagous insects and the optimal regulation of their interactions with their trophic resources are determined by numerous interacting biotic and abiotic factors.

Consequently, substantial literature on phytophagous insects has been devoted to understanding the factors that govern female reproductive output because this output determines the potential number of offspring produced. The female reproductive output depends on at least three determinant life history traits: fecundity, egg size and percent egg hatch. The number of eggs laid by a female is clearly important to determine their reproductive potential, but egg size (often correlated to percent egg hatch) is also considered a crucial reproductive parameter. Numerous studies have examined the relationship between egg size and fitness components of the progeny. Such studies often demonstrate that within a species, small eggs are less likely to hatch (Fox and Czesak 2000 and references therein), and hatching individuals from larger eggs have higher fitness than those from smaller eggs [START_REF] Karlsson | Fecundity in butterflies: adaptations and constraints[END_REF][START_REF] Fox | Evolutionary ecology of progeny size in arthropods[END_REF][START_REF] Roff | Host suitability and preference studies of Trichogramma cordubensis (Hymenoptera: Trichogrammatidae)[END_REF]Torres-Vila and Rodriguez-Molina 2002).

Many insect species are pests and are responsible for huge annual losses in global crop production (Thacker 2002). Most of the control methods currently used to increase crop production rely on accurate monitoring and decisions, which require precise information on the biology and the ecology of the pest [START_REF] Tammaru | Responses of ovipositing moths (Lepidoptera: Geometridae) to host plant deprivation: Life-history aspects and implications for population dynamics[END_REF]. In capital breeder insects (whose individual reproductive potential is limited by the nutrition ingested during the larval stages), larval food quality is one of the most important factors that determine female reproductive output [START_REF] Awmack | Host plant quality and fecundity in herbivorous insects[END_REF]. Several studies have shown the influence of plant quality on larval development, larval survival, and female reproductive output [START_REF] Awmack | Host plant quality and fecundity in herbivorous insects[END_REF]Thiéry and Moreau 2005;Moreau et al. 2006a;2006b;[START_REF] Moreau | Reproductive output of females and males from natural populations of European grapevine moth (Lobesia botrana) occurring on different grape varieties[END_REF]. However, most studies have considered the effect of the host plant on fecundity, egg size and percent egg hatch without considering the temporal effect on these life history traits. This information is lacking because for the same fecundity, the temporal oviposition pattern could be dramatically different. For instance, all eggs could be laid in a single session, or an equal number of eggs could be laid each day throughout the life of the female. These different temporal oviposition patterns can lead to differences in population growth rate and the timing of further larval instars. They can also influence biological control management by modifying the optimal windows of attack for natural enemies (e.g., egg parasitoids or predators and larval parasitoids). Information on temporal oviposition is needed for progress in the study of pest population age structure determination and the construction of basic life tables [START_REF] Carey | Insect Biodemography[END_REF] for a review; [START_REF] Ainseba | An innovative multistage, physiologically structured, population model to understand the European grapevine moth dynamics[END_REF][START_REF] Farahani | Life table of Spodoptera exigua (Lepidoptera: Noctuidae) on five soybean cultivars[END_REF]). To our knowledge, only a few studies have examined the effect of the host plant on temporal oviposition (see [START_REF] Hafiz | Use of life tables to assess host plant resistance in cowpea to Aphis craccivora Koch (Homoptera: Aphididae)[END_REF][START_REF] Samih | Age specific reproduction parameters of cotton whitefly (Bemisia tabaci) and silverleaf whitefly (B. argentifolii) on cotton and rapeseed[END_REF], and no studies have examined the egg size and percent egg hatch throughout the oviposition period, which are necessary for a complete understanding of the variation in insect oviposition and integration into life history tables.

The European grapevine moth, Lobesia botrana (Denis and Schiffermueller, Lepidoptera: Tortricidae), is certainly the most harmful grape pest in Europe, north Africa and west Asia [START_REF] Bovey | Super-famille des Tortricoidea[END_REF][START_REF] Roehrich | Tortricids in vineyards[END_REF]Iorati et al. 2011;Thiéry et al. 2014;), and its recent introduction to Chilean and California vineyards highlights the problem of pest management [START_REF] Gutierrez | Prospective analysis of the invasive potential of the European grapevine moth Lobesia botrana (Den. and Schiff.) in California[END_REF][START_REF] Varela | Can European grapevine moth, Lobesia botrana (Lepidoptera: Tortricidae) be eradicated from California?[END_REF]. Lobesia botrana may cause serious damage to the grape directly by consuming flower clusters and fruits, or by facilitating infection by pathogenic fungi such as grey mould disease, Botrytis cinerea (Persoon, Helotiales: Sclerotiniaceae), or black mould, Aspergillus spp. (Micheli, Eurotiales: Trichocomaceae) [START_REF] Cozzi | Effect of Lobesia botrana damages on black aspergilli's rot and ochratoxin A content in grapes[END_REF]Thiéry 2008;[START_REF] Delbac | Grape flowers and berries damage by Lobesia botrana larvae (Denis & Schiffernüller) (Lepidoptera: Tortricidae) and relation to larval age[END_REF]. It may also facilitate attack by secondary pests, including fruit flies [START_REF] Barata | New insights into the ecological interaction between grape berry microorganisms and drosophila flies during the development of sour rot[END_REF]) such as Drosophila melanogaster (Meigen, Diptera: Drosophilidae) [START_REF] Gravot | La pourriture acide. I -Aetiologie: recherche de causes de cette pourriture dans le vignoble bordelais[END_REF]) and D. suzukii (Matsumura, Diptera: Drosophilidae) [START_REF] Rouzes | The spotted wing drosophila (Drosophila suzukii) entered the drosophila communities in the French Sauternes Vineyard[END_REF]. Therefore, L. borana is a highly problematic pest in vineyards and requires permanent monitoring and control (Thiéry 2008;2011;[START_REF] Ortega-Lopez | Male flight phenology of the European grapevine moth Lobesia botrana (Lepidoptera: Tortricidae) in different wine-growing regions in Spain[END_REF]. This pest is an ideal candidate for testing the effect of the host plant on temporal oviposition because (i) previous studies have shown strong effects of cultivars where larvae feed on both larval developmental and reproductive life history traits (Moreau et al. 2006a;2006b;2006c;[START_REF] Moreau | Reproductive output of females and males from natural populations of European grapevine moth (Lobesia botrana) occurring on different grape varieties[END_REF]Thiéry et al. 2014) and (ii) information on temporal oviposition is needed for advancements in grape pest mathematical models [START_REF] Ainseba | An innovative multistage, physiologically structured, population model to understand the European grapevine moth dynamics[END_REF].

The present work focuses on the temporal oviposition of the main pest of European vineyards and on how the host plant can affect this temporal pattern. Therefore, we determine whether the cultivar on which the females feed as larvae affect the number of eggs laid, egg size, and egg hatch percentage, as well as the temporal patterns of these three life history traits. To examine this dependence, we conducted laboratory experiments by rearing larvae on artificial diets derived from seven different cultivars (Chardonnay, Chasselas, Gewurztraminer, Grenache, Merlot, Pinot and Riesling). We then measured individual female oviposition based on time, egg size and percent egg hatch.

Materials and Methods

Study system, origin and maintenance of moths

The strain of L. botrana (INRA Bordeaux) used for this study originated from individuals collected in a French Sauternes vineyard (cultivar Semillon) in 1997, to which wild adults are periodically added. This rearing line is maintained with a substantial number of caged adults (several thousand a week) to avoid genetic drift. This laboratory strain has conserved genetic variability because considerable variation is found in the larval and adult behaviours and in larval immune parameters [START_REF] Vogelweith | Host plant variation plastically impacts different traits of the immune system of a phytophagous insect[END_REF]. The stock colony is maintained without diapause on a semi-artificial diet (as described in Thiéry and Moreau 2005), with the following composition: 150 ml water, 3 g agar, 9 g maize flour, 11 g wheat germ, 9 g yeast, 0.9 g ascorbic acid, 0.3 g benzoic acid, 0.3 ml maize oil, 0.3 g nipagin, and 0.2 g iprodione, at 24 ± 1 °C, 60 ± 10% RH with a photoperiod 15: 8 h light/dark + 1 of dusk. The first 15photophase hours were at 1000-lux luminosity, and the last hour (dusk) was at 25 lux. All tests were performed under these conditions.

Larval diet treatments and general procedure

The influence of different grape cultivars on L. botrana was tested using a standardized procedure (Thiéry and Moreau 2005;Moreau et al. 2006a;2006b). Compared to direct feeding on bunches in the laboratory or in the field, this procedure has at least three main advantages: a) feeding isolated larvae prevents competition and subsequent food deprivation; b) it prevents differences in grape bunch compactness, which impact larval feeding behaviour (our unpublished observations) and the climatic environment (temperature and insolation) of the larvae [START_REF] Pieri | Effects of defoliation on temperature and wetness of grapevine berries[END_REF]; and c) it prevents infections by fungi on grapes, which may affect larval fitness as shown by Savoupoulou-Soultani and Tzanakakis (1988) and [START_REF] Mondy | Response to dietary phytopathogenic fungus (Botrytis cinerea) in grape berry moth (Lobesia botrana): the significance of fungus sterols[END_REF].

To avoid immature competition, larvae were reared individually to pupation in Eppendorf tubes filled with 1.5 ml of a medium containing the following (for 100 Eppendorfs): 150 ml water, 5 g agar, 6 g cellulose powder, 4 g vitamin-free casein, 3.5 g glucose, 2 g mineral salt, 0.12 g cholesterol, 0.12 g maize oil, 0.25 g benzoic acid, 0.1 g nipagin and 12 g freeze-dried grape fruit powder. Freeze dried material was obtained from freshly collected grape flower clusters within 12 h of collection using a Christ alpha 1-4 LD plus device. This classic procedure preserves fresh foods (Ratti 2001) and is typically used for secondary metabolites in grapes (see [START_REF] Michalczyk | The effect of air-drying, freeze-drying and storage on the quality and antioxidant activity of some selected berries[END_REF]). The grape cultivar dried powders were obtained from grape flower clusters of V. vinifera cv. Chardonnay, Chasselas, Gewurztraminer, Grenache, Merlot, Pinot and Riesling, all of which were harvested from our gene collection of grape plants "Domaine de la Grande Ferrade", INRA-Bordeaux. The insecticide-free grape flower clusters were collected at the beginning of the growing season (beginning of May 2003) at phenological stages 23-27 (Eichhorn and Lorenz 1977), which correspond to the grape phenology on which the first annual generation of L. botrana larvae feeds.

The Eppendorf lids were pierced to allow air circulation. Using a fine brush, newly hatched larvae (age < 8 h) were transferred individually to the diets in each Eppendorf, with 100 larvae per diet (cultivar). Neonate larvae from eggs produced by thousands of caged females were randomly chosen and assigned to the different diets. Eppendorf tubes were randomized in the Eppendorf racks, which were moved within the climatic chamber every three days to minimize the effect of possible climatic gradients.

The larvae from each diet were monitored daily until pupation. Two-day-old pupae were then carefully removed from the diet and weighed to the nearest 0.1 mg. Because it is difficult to weigh adult moths with sufficient accuracy, we used the mass of living pupae as an index of adult body size. Pupae were then placed individually in glass tubes (70 mm x 9 mm diameter) covered with cotton plugs and stored in the test room until emergence under the same conditions previously described. Adults were sexed after emergence by checking their ventral abdominal extremity (Thiéry, 2008).

All newly emerged female adults resulting from the eight larval diets were used to evaluate the temporal effect of (1) egg laying (2) egg size and (3) larval hatching. Because it has been reported that the cultivars on which larvae develop can modify the female oviposition preferences [START_REF] Moreau | Larval host plant origin modifies the adult oviposition preference of the female European grapevine moth Lobesia botrana[END_REF], we decided not to run the oviposition experiments on one specific cultivar, which may have interfered with the temporal oviposition. Thus, we used an inert substrate (i.e., considered equal for all females) where females could deposit their eggs. Newly emerged females (less than 1 day old) were individually confined to 0.5 litre transparent cellophane bags as mating and oviposition chambers, and they were provided with water ad libitum through a soaked cotton dental wick.

One-or two-day-old virgin males originating from the same diet were added to each caged virgin female 1 h before dusk, which is just before their sexual activity [START_REF] Bovey | Super-famille des Tortricoidea[END_REF]. Only one male was randomly assigned to each female. Pairs were caged in these bags until the death of both sexes.

Females could behave and oviposit freely inside the cellophane bag until death. Each morning, the cellophane bags were checked, and new eggs laid during the previous night were marked with a specific colour outside the bags. For the analyses, only females that laid a sufficient number of eggs (> 7 eggs) during their lifetime (because non-mated females can lay only a few eggs) and that began to lay at the beginning of her life were considered to obtain a representative picture of the temporal oviposition in this species. We thus obtained the temporal oviposition for a variable number of females that depended on the sex ratio, larval At the end of the experiment (i.e., when the females were found dead in the bags), we randomly selected a sample of females that had laid a sufficient number of eggs each day in each cultivar to assess egg size and percent egg hatch: Chardonnay n = 14, Chasselas n = 9, Gewurztraminer n = 12, Grenache n = 9, Merlot n = 17, Pinot n = 15 and Riesling n = 10. For each selected female, three eggs from the walls of the cellophane bags were randomly selected per oviposition day, when possible. Indeed, some females sometimes laid less than three eggs per day, particularly at the end of life. Previous studies we have done (results not shown in this paper) showed that measuring three eggs per oviposition day and per female gives the same results than if we measured more eggs whatever the oviposition rank. Each egg was then measured with an ocular micrometre. The egg surface (estimated as an elliptic surface, S = π x a x b in mm 2 , where a and b were the ellipse semi axes) was used as an index of egg size. The mean egg size per day was estimated for each female from this sample. To estimate daily fertility (i.e., hatching success), the measured eggs were incubated at 22°C for 10 days until hatching.

Statistical Analysis

For all analyses, mixed effects models were performed, including the identity of the females as a random effect. The pupal mass of the females was also included to control for its potential effect. The number of eggs laid by day and cultivar was compared using negative binomial generalized linear mixed effects model (GLMM) accounting for zero-inflated data.

The statistical significance of each parameter was tested with χ 2 -statistics for unbalanced design [START_REF] Fox | An R Companion to Applied Regression, Second Edition[END_REF]. The proportion of hatching eggs was arcsine square root transformed to normalize. However, the transformed variable was not normally distributed and did not meet the assumption of homoscedasticity, as was the case for the size of eggs laid.

Therefore, the proportion of hatched eggs and their size were compared among cultivars and days using a linear mixed effects model based on rank transformation (as a more powerful alternative to the classical nonparametric Friedman test, Baguley 2012). The statistical significance of each parameter was tested with Wald χ 2 -statistics for unbalanced design. In each case, pairwise Wilcoxon multiple comparison post hoc tests were performed (P-values were adjusted for multiple comparisons using the Benjamini-Yekutieli step-up procedure, [START_REF] Benjamini | The control of the false discovery rate in multiple testing under dependency[END_REF]. The same procedure was used to analyse differences in egg size.

All statistical analyses were performed using R software (v. 3.1.1, R Development Core Team 2014) and implemented using the following packages: lme4 (linear mixed effects models), glmmADMB (negative binomial generalized linear mixed effects model with zeroinflated data) and car (deviance analysis for unbalanced design).

Results

1) Number of eggs laid throughout the oviposition period

The total number of eggs laid did not differ among females fed on the different cultivar diets (GLMM: χ 2 = 4.07, df = 6, P = 0.67) but varied positively according to the mass of the pupae (χ 2 = 18.77, df = 1, P < 0.0001). The number of eggs laid decreased with oviposition day (female age) with the maximum of eggs laid in the first day of oviposition and with only few eggs laid at the end of the oviposition period (χ 2 = 231.57, df = 6, P < 0.0001, Fig. 1). A significant interaction between cultivar and day was found (interaction cultivar x day, χ 2 = 62.51, df = 36, P < 0.01, Fig. 2), indicating that larval diets with different grape cultivars had an effect on female temporal oviposition. For some cultivars, such as Gewürztraminer and Pinot noir, the number of eggs laid remained stable for the first four days of oviposition and decreased for the remaining oviposition days, whereas for other cultivars, such as Chardonnay, the numbers of eggs laid decreased linearly with oviposition days (Fig. 2).

2) Egg size variation throughout the oviposition period

The size of the eggs laid by females fed with different cultivars during their larval period did not differ (linear mixed effect model: χ 2 = 11.45, df = 6, P = 0.08) and did not vary with female pupal mass (χ 2 = 0.62, df = 1, P = 0.43). However, the size of eggs decreased with increasing days (χ 2 = 340.92, df = 6, P < 0.0001, Fig. 3). There was also a significant interaction between day and cultivar (χ 2 = 132.26, df = 36, P < 0.0001): for some cultivars, such as Chardonnay or Riesling, the size of eggs laid remained stable for the first four days of oviposition and then decreased, whereas for other cultivars, such as Pinot noir, the size of eggs laid decreased progressively (Fig. 4).

3) Percent egg hatch throughout the oviposition period

The percent of hatching eggs did not vary according to the female mass (linear mixed effects model: χ 2 = 0.40, df = 1, P = 0.53) or among cultivars (χ 2 = 10.44, df = 6, P = 0.11). However, the percent of hatched eggs decreased with increasing oviposition days (χ 2 = 93.77, df = 6, P < 0.0001, Fig. 5), with no interaction with cultivar (interaction cultivar x day, χ 2 = 38.80, df = 35, P = 0.30). At the beginning of the oviposition period, approximately 90% of eggs hatched, whereas only approximately 60% of eggs hatched at the end of the oviposition period.

Discussion

One of the goals of the present study was to assess the temporal effect on three major life history reproductive traits of L. botrana (fecundity, egg size and percent egg hatch). Our results show that all three parameters decreased over time. The second goal of this study was to determine whether larval feeding on a diet containing different cultivars had an effect on the temporal oviposition patterns and on egg size and percent egg hatch throughout the oviposition period. Our results clearly showed that the cultivar on which females developed as larvae had a significant effect on the temporal oviposition and egg size. However, no effect on percent egg hatch was detected. This result indicates that food characteristics associated with the immature stages affect how adult females distribute eggs over time. This phenomenon, combined with the effect of consuming different cultivars on the larval growth rates, may explain how the distribution of the next adult generation varies over time (Thiéry et al, 2014).

In the context of our study, the distinction between income and capital breeding insects is important. Income breeder females mature their eggs throughout their adult life, whereas capital breeders emerge with a nearly fixed number of ovocytes [START_REF] Papaj | Ovarian dynamics and host use[END_REF].

Lobesia botrana is considered a capital breeder (species with non-feeding adults); its female reproductive potential is thus limited by the resources accumulated during the larval stage [START_REF] Slansky | Nutritional ecology of insects, mites, spiders, and related invertebrates: An overview[END_REF][START_REF] Awmack | Host plant quality and fecundity in herbivorous insects[END_REF]. In our experimental design, females lacked access to any additional food except clear water, so the resources mobilized for the egg production were derived from reserves accumulated during the larval stage [START_REF] Awmack | Host plant quality and fecundity in herbivorous insects[END_REF]. Previous studies showed that pupal mass is a good predictor of fecundity in L. botrana [START_REF] Moreau | Reproductive output of females and males from natural populations of European grapevine moth (Lobesia botrana) occurring on different grape varieties[END_REF]). Egg-laying activity is under physiological pressure from the oogenesis process and oocyte formation [START_REF] Mcdonald | The relationship of age and ovarian development to mating of Delia antiqua (Diptera: Anthomyiidae)[END_REF][START_REF] Chapman | The insects: Structure and function[END_REF]. Females used in the present study had an average egg load of 144.6 ± 37.8 eggs and laid approximately 60% of its eggs in the two first days of the laying period.

Subsequently, the number of eggs laid decreased daily until the death of the female. The large number of eggs laid during the first night could be adaptive because under natural conditions L. botrana adults have a relatively short life expectancy (ca. one week according to the climatic conditions) (Moreau et al. 2006a;Moreau et al. 2006b;Thiéry 2008). In addition, the predation risk for a moth is high because nocturnal insectivores often prey on moths; these predators include bats, species of owls and other species of birds [START_REF] Arlettaz | Competition for food by expanding pipistrelle bat populations (Pipistrellus pipistrellus) might contribute to the decline of lesser horseshoe bats (Rhinolophus hipposideros)[END_REF]. Given that there is a delay between encountering a mate and the first egg laid of approximately four days for a female from our rearing strain (Moreau et al. 2006a) or in the field [START_REF] Moreau | Reproductive output of females and males from natural populations of European grapevine moth (Lobesia botrana) occurring on different grape varieties[END_REF], females have only a few days to lay eggs. In that case, laying a maximum number of eggs as quickly as possible could be adaptive. This phenomenon was observed, for example, in Dryas iulia (Fabricius, Lepidoptera: Nymphalidae), whose females laid the greatest number of eggs at the beginning of the laying sequence [START_REF] Dunlap-Pianka | Ovarian dynamics in heliconiine butterflies: programmed senescence versus eternal youth[END_REF].

We showed that eggs laid on the first day of oviposition are larger than eggs laid on the subsequent days. This size decrease is consistent with previous results performed in the same species [START_REF] Moreau | Host plant cultivar of the grapevine moth Lobesia botrana affects the life history traits of an egg parasitoid[END_REF]. Lepidoptera ovaries structure and morphological aspects of oogenesis are well known (see Swevers et al. 2005 for a review or Chapman et al. 2012 for a book). Briefly, oocytes are produced in the germarium and begin to move down the ovariole, enlarging as they pass through the vitellarium, where yolk containing both protein and lipids is deposited on them (vitellogenesis). The fat body is the principal site of production of the major yolk protein precursor (YPP), which is vitellogenin (Vg) in most insects (Swevers et al. 2005). In many butterflies and moths, ovaries contain only previtellogenic oocytes, and vitellogenesis starts at eclosion. Therefore, their ovarioles usually contain a series of oocytes in successive stages of development. In L. botrana, we found a delay of approximately 4 days between encountering a mate and the first egg laid (Moreau et al. 2006a;[START_REF] Moreau | Reproductive output of females and males from natural populations of European grapevine moth (Lobesia botrana) occurring on different grape varieties[END_REF]. During this time, the first eggs have time to mature and enlarge with yolk. The only reason for the differential size is thus the amount of yolk, which is related to the decreased synthesis by the fat body or its depletion.

The proportion of larvae that hatched from eggs was strongly dependent on the oviposition days; fewer eggs hatched at the end of the life of females independently of the cultivars, which is consistent with the amount of yolk in the egg. This decline can be interpreted as a result of the egg size decrease, which was clearly associated with the egg size in that species (Moreau et al. 2006;[START_REF] Moreau | Reproductive output of females and males from natural populations of European grapevine moth (Lobesia botrana) occurring on different grape varieties[END_REF], most likely because larger eggs have the largest nutritional provisions inside, as already shown in other species [START_REF] Berrigan | The allometry of egg size and number in insects[END_REF][START_REF] Fox | Evolutionary ecology of progeny size in arthropods[END_REF]. This decrease is not due to a decrease in sperm numbers because we showed that in L. botrana, one male ejaculate contains much more spermatozoids than necessary to fertilize all female eggs [START_REF] Muller | Male larval nutrition affects adult reproductive success in wild European grapevine moth (Lobesia botrana)[END_REF].

Our results show that the effect of the cultivar on which females developed as larvae had a significant effect on the temporal oviposition and egg size. For some cultivars, the number and the size of eggs laid remained stable for the first four days of oviposition and decreased for the remaining oviposition days, whereas for other cultivars, the numbers of eggs laid or the size of eggs decreased through oviposition days. However, no effect of cultivar was detected for percent egg hatch. This effect is very weak in comparison to the profound effect of cultivar on total number of eggs laid, total mean egg size and mean percent egg hatch that we have previously demonstrated (Moreau et al. 2006a;2006b;[START_REF] Moreau | Reproductive output of females and males from natural populations of European grapevine moth (Lobesia botrana) occurring on different grape varieties[END_REF].

We found here that time within the oviposition period affects the number of eggs laid and the probability that an egg hatches. These individual temporal effects also influence the population scale. The data provided here on time-dependent offspring production in L. botrana could also be useful for the development of age-structured mathematical models of vineyard infestation such as that developed by [START_REF] Ainseba | An innovative multistage, physiologically structured, population model to understand the European grapevine moth dynamics[END_REF]. Therefore, we also believe that this feature needs to be incorporated in future L. botrana population agestructured mathematical models. These effects of time would also influence biological control management based on egg parasitoids, predators and larval parasitoids to determine the optimal windows in which natural enemies should be released. As an example, several species of Trichogramma can be used in inundative biological control programs in a variety of crops against numerous pests [START_REF] Morrison | Trichogramma spp[END_REF][START_REF] Abd El-Monsef | Biological control of grape berry moths Eupoecilia ambiguella HB. and Lobesia botrana Den. and Schiff. (Lepidoptera Tortricidae) by using egg parasitoids of the genus Trichogramma[END_REF][START_REF] Agamy | Field evaluation of the egg parasitoid, Trichogramma evanescens West. against the olive moth Prays oleae (Bern.) in Egypt[END_REF][START_REF] Andrade | Performance of four Trichogramma species (Hymenoptera: Trichogrammatidae) as biocontrol agents of Heliothis virescens (Lepidoptera: Noctuidae) under various temperature regimes[END_REF] including L. botrana with irregular results [START_REF] Barnay | Dynamique des populations et relation hôte-parasitoïde chez le couple Lobesia botrana Den. and Schiff. Trichogramma cacoeciae Marchal, dans le cadre de la lutte biologique en vignoble[END_REF][START_REF] Hommay | Comparison between the control efficacy of Trichogramma evanescens Westwood and of two Trichogramma cacoeciae Marchal strains against vine moth (Lobesia botrana Den. & Schiff.) depending on their release density[END_REF][START_REF] El-Wakeil | Efficacy of Trichogramma evanescens in controlling the grape berry moth Lobesia botrana in grape farms in Egypt[END_REF]. Parasitism efficiency in Trichogramma parasitoids is, of course, influenced by environmental factors [START_REF] Calvin | Impact of environmental factors on Trichogramma pretiosum reared on Southwestern corn borer eggs[END_REF][START_REF] Pizzol | Temperature dependent differences in biological traits between two strains of Trichogramma cacoeciae (Hym., Trichogrammatidae)[END_REF] and by intrinsic factors [START_REF] Schmidt | Host recognition and acceptance by Trichogramma[END_REF][START_REF] Pizzol | Parasitoid and host egg ages have independent impact on various biological traits in a Trichogramma species[END_REF]) such as egg size, quality and age [START_REF] Roff | Host suitability and preference studies of Trichogramma cordubensis (Hymenoptera: Trichogrammatidae)[END_REF][START_REF] Moreau | Host plant cultivar of the grapevine moth Lobesia botrana affects the life history traits of an egg parasitoid[END_REF][START_REF] Pizzol | Parasitoid and host egg ages have independent impact on various biological traits in a Trichogramma species[END_REF]. Indeed, large and numerous eggs are often more conspicuous for the searching females and more suitable for the subsequent development of their progeny (Van Huis and De Rooy 1998;[START_REF] Moreau | Host plant cultivar of the grapevine moth Lobesia botrana affects the life history traits of an egg parasitoid[END_REF][START_REF] Pizzol | Parasitoid and host egg ages have independent impact on various biological traits in a Trichogramma species[END_REF]). With our results, the optimal window for parasitoids or predator release must occur when the first eggs are laid (bigger eggs and more eggs). This optimal window could be calculated because we showed that for each grape variety, the emergence of L. botrana females occurred as a single wave after approximately ten days (Thiéry et al. 2014). In addition, considering the cultivar effect on temporal oviposition could be useful when monitoring eggs in vineyards.

Insecticides against the larvae (growth regulators or Bt) are mostly applied at a typical egg stage called 'black head', which corresponds to 2/3 of the egg incubation period. Shorter or longer oviposition periods as a function of previous generation food (i.e., the cultivar on which the female fed as larva) would thus cause, at the population and the vineyard scales, important variation in these typically monitored stages. We strongly believe that this information concerning temporal oviposition is needed to advance our knowledge of pest ecology with the aim of better biological control.

Author contribution

  survival and mating success: Chardonnay n = 33, Chasselas n = 24, Gewurztraminer n = 24, Grenache n = 20, Merlot n = 35, Pinot n = 34 and Riesling n = 29.

Figure legends Fig. 1

 legends1 Figure legends

Fig. 2

 2 Fig. 2 Number of eggs laid by Lobesia botrana female according to the day of oviposition and

Fig. 3

 3 Fig. 3 Egg size according to the day of oviposition for all cultivars. Bold line: median; box:

Fig. 4

 4 Fig. 4 Egg size according to the day of oviposition and for each cultivar where the female

Fig. 5

 5 Fig. 5 Proportion of hatching eggs according to the day of oviposition for all cultivars. Bold

  Figure 1

  Figure 2

Figure 4

 4 Figure 4

Acknowledgements

This research was supported by INRA, University of Bourgogne and a grant from the Regional Council of Bourgogne. The authors thank Marc-Etienne Toulouse for experimental contributions in rearing Lobesia botrana.

Conflict interest

The authors declare that they have no conflict of interest.

Ethical approval

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed.

Informed consent

This article does not contain any studies with human participants performed by any of the authors.


Sofiane Bouzid

Magali Chaudey

Muriel Fadairo

Frédéric Perdreau

Strategic Groups in the French Franchising Sector

Keywords: Franchising, Strategic groups, Statistical models, Performance. JEL Classification: L21, C18

niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

HAL

  is a multi-disciplinary open access archive for the deposit and dissemination of scientific research documents, whether they are published or not. The documents may come    Strategic Groups in the French Franchising SectorSofiane Bouzid, Magali Chaudey, Muriel Fadairo, Frédéric Perdreau To cite this version: Sofiane Bouzid, Magali Chaudey, Muriel Fadairo, Frédéric Perdreau. Strategic Groups in the French Franchising Sector. 30th Annual International Society of Franchising Conference University of Groningen, Faculty of Economics and Business, Jun 2016, Groningen, Netherlands. halshs-01420796


Elva Fuentes
email: elva.fts@gmail.com

Jérôme Moreau

Maria Teixeira

Vincent Bretagnolle

Karine Monceau

Effects of conventional vs. organic farming practices on raptor nestling health: Neither black nor white

Keywords: Behaviour, carotenoid, Circus pygargus, physiology, stress, trade-off

come   L'archive ouverte pluridisciplinaire

Introduction

The biodiversity crisis, faced nowadays, spares no species. Among the most affected taxa, bird numbers decreased by 25% in Europe between 1980 and 2016 with farmland birds showing the steepest decline [START_REF] Heldbjerg | Continuous population declines for specialist farmland birds 1987-2014 in Denmark indicates no halt in biodiversity loss in agricultural habitats[END_REF][START_REF] Reif | Continent-wide gradients in open-habitat insectivorous bird declines track spatial patterns in agricultural intensity across Europe[END_REF][START_REF] Rigal | Farmland practices are driving bird population decline across Europe[END_REF]. At the same time, agricultural practices changed to increase yields, leading ecologists to hypothesise a causal link between agricultural intensification and farmland bird decline [START_REF] Fuller | Population declines and range contractions among lowland farmland birds in Britain[END_REF][START_REF] Krebs | The second silent spring?[END_REF]Chamberlain et al. 2000). Among the features of this intensification, mechanisation appeared to have a direct negative impact on ground-nesting birds [START_REF] Crick | Causes of nest failure among buntings in the UK[END_REF][START_REF] Tews | Estimated mortality of selected migratory bird species from mowing and other mechanical operations in Canadian agriculture[END_REF][START_REF] Santangeli | Identifying effective actions to guide volunteer-based and nationwide conservation efforts for a ground-nesting farmland bird[END_REF]. Landscape homogenisation and the use of pesticides also raised concerns because they can have insidious effects on farmland species [START_REF] Matson | Agricultural intensification and ecosystem properties[END_REF][START_REF] Benton | Farmland biodiversity: is habitat heterogeneity the key?[END_REF][START_REF] Mason | Immune suppression by neonicotinoid insecticides at the root of global wildlife declines[END_REF][START_REF] Sumasgutner | Landscape homogenization due to agricultural intensification disrupts the relationship between reproductive success and main prey abundance in an avian predator[END_REF]Moreau et al. 2022a). To counter the negative impacts of conventional farming (CF) involving intensive agricultural practices, organic farming (OF) emerged as an alternative in the late 1970s [START_REF] Lockeretz | Organic Farming: An International History[END_REF]. OF is defined as a production system excluding synthetic pesticides and fertilisers that uses nature-based solutions [START_REF] Keesstra | The superior effect of nature based solutions in land management for enhancing ecosystem services[END_REF] to promote and enhance agro-ecosystem health [START_REF] Lockeretz | Organic Farming: An International History[END_REF].

Three main interrelated characteristics distinguish OF from CF: agricultural practices, natural and semi-natural habitat proportion and farm inputs. Agricultural practices can be harmful to farmland birds either directly or indirectly. A direct impact of agricultural machines used under intensive practices is the death of ground-nesting birds [START_REF] Crick | Causes of nest failure among buntings in the UK[END_REF][START_REF] Vickery | The management of lowland neutral grasslands in Britain: effects of agricultural practices on birds and their food resources[END_REF][START_REF] Tews | Estimated mortality of selected migratory bird species from mowing and other mechanical operations in Canadian agriculture[END_REF]. Incubating adults, broods and chicks are frequently destroyed during mechanical mowing, tilling and harvesting activities [START_REF] Schekkerman | Mortality of black-tailed godwit Limosa limosa and Northern lapwing Vanellus vanellus chicks in wet grasslands: influence of predation and agriculture[END_REF][START_REF] Santangeli | Identifying effective actions to guide volunteer-based and nationwide conservation efforts for a ground-nesting farmland bird[END_REF]Bretagnolle et al. 2018a). Agricultural practices can also have indirect effects on birds, by increasing disturbance and decreasing available resources. For instance, manure spreading, manual and mechanical weed and pest control, tillage and multiple cultivations all together generally increase labour in OF systems [START_REF] Karlen | Nutrient, labor, energy and economic evaluations of two farming systems in Iowa[END_REF][START_REF] Jansen | Labour, livelihoods and the quality of life in organic agriculture in Europe[END_REF][START_REF] Orsini | Labour use on organic farms: a review of research since 2000[END_REF], and can be identified as a potential source of disturbance for farmland birds. Human presence and activity are known to affect bird feeding rates [START_REF] Exposito-Granados | Physiological stress and behavioural responses of European Rollers and Eurasian Scops Owls to human disturbance differ in farming habitats in the south of Spain[END_REF], induce physiological stress (Mougeot & Arroyo, 2017;[START_REF] Rabdeau | Does anthropization affect the physiology, the behaviour and the life-history traits of Montagu's harrier chicks?[END_REF], and alter behaviour including parental care [START_REF] Fernández | Human disturbance affects parental care of marsh harriers and nutritional status of nestlings[END_REF], sometimes leading to nest failures [START_REF] Strasser | Reproductive failure of a human-tolerant species, the American kestrel, is associated with stress and human disturbance[END_REF]. Farming activity has even been shown to increase nest abandonment in passerine birds and to cause up to 32% nest failure in river lapwing, Vanellus duvaucelii [START_REF] Assandri | Nest density, nestsite selection, and breeding success of birds in vineyards: management implications for conservation in a highly intensive farming system[END_REF][START_REF] Mishra | Factors influencing nesting success of the river lapwing, Vanellus duvaucelii (Lesson, 1826)[END_REF]. Moreover, intensive agricultural practices that involve clearing of ditches, removal of hedgerows, and intensification of grassland management are associated with lower territory and breeding densities in yellowhammer, Emberiza citrinella (Bradburry et al. 2000) and barn owl, Tyto alba [START_REF] De Bruijn | Population ecology and conservation of the Barn Owl Tyto alba in farmland habitats in Liemers and Achterhoek (The Netherlands)[END_REF]. These intensive land-uses commonly found in CF lead to simplified landscapes where important habitats for birds are no longer available [START_REF] Bradbury | Habitat associations and breeding success of yellowhammers on lowland farmland[END_REF][START_REF] Sumasgutner | Landscape homogenization due to agricultural intensification disrupts the relationship between reproductive success and main prey abundance in an avian predator[END_REF][START_REF] Tarjuelo | Living in seasonally dynamic farmland: The role of natural and semi-natural habitats in the movements and habitat selection of a declining bird[END_REF]) and where their food supplies are reduced through fewer weed seeds and natural insect reservoirs [START_REF] Matson | Agricultural intensification and ecosystem properties[END_REF][START_REF] Chamberlain | The contribution of hedgerow structure to the value of organic farms to birds[END_REF][START_REF] Benton | Linking agricultural practice to insect and bird populations: a historical study over three decades[END_REF]. Insects rely on the availability of non-uniform patches of grass, weeds and hedgerows for food and shelter [START_REF] Benton | Linking agricultural practice to insect and bird populations: a historical study over three decades[END_REF]. These features are thus associated with higher insect diversity [START_REF] Holland | Effect of woody borders on insect density and diversity in crop fields: a landscape-scale analysis[END_REF] and higher bird diversity and abundance [START_REF] Chamberlain | A comparison of bird populations on organic and conventional farm systems in southern Britain[END_REF][START_REF] Chamberlain | The contribution of hedgerow structure to the value of organic farms to birds[END_REF][START_REF] Freemark | Birds on organic and conventional farms in Ontario: partitioning effects of habitat and practices on species composition and abundance[END_REF].

Hedgerows and landscape complexity also maintain small mammal populations [START_REF] Butet | Seasonal changes in small mammal assemblages from field boundaries in an agricultural landscape of western France[END_REF][START_REF] Gelling | Are hedgerows the route to increased farmland small mammal density? Use of hedgerows in British pastoral habitats[END_REF]) and consequently have a positive effect on populations of birds of prey [START_REF] Koks | Do voles make agricultural habitat attractive to Montagu's Harrier Circus pygargus[END_REF][START_REF] Butet | Responses of common buzzard (Buteo buteo) and Eurasian kestrel (Falco tinnunculus) to land use changes in agricultural landscapes of Western France[END_REF], even if some find such a link only under favourable food conditions [START_REF] Sumasgutner | Landscape homogenization due to agricultural intensification disrupts the relationship between reproductive success and main prey abundance in an avian predator[END_REF]. OF has been shown to have a positive effect on biodiversity, an effect which is even greater in simplified landscapes [START_REF] Roschewitz | The effects of landscape complexity on arable weed species diversity in organic and conventional farming[END_REF][START_REF] Fischer | Mixed effects of landscape structure and farming practice on bird diversity[END_REF][START_REF] Henckel | Organic fields sustain weed metacommunity dynamics in farmland landscapes[END_REF]. In fact, OF practices that might imply non-cropped habitat management such as the establishment of hedgerows and field margins benefit insects and therefore insectivorous birds [START_REF] Hole | Does organic farming benefit biodiversity?[END_REF][START_REF] Lockeretz | Organic Farming: An International History[END_REF]. Under OF agricultural systems, natural and semi-natural areas are typically higher than under CF systems [START_REF] Gibson | Plant diversity and land use under organic and conventional agriculture: a whole-farm approach[END_REF][START_REF] Norton | Consequences of organic and non-organic farming practices for field, farm and landscape complexity[END_REF], hence OF promotes resources for birds, especially food abundance [START_REF] Fischer | Mixed effects of landscape structure and farming practice on bird diversity[END_REF]. Different meta-analyses support that view, showing that OF promotes higher overall biodiversity, from flora to insects and bird species richness and abundance [START_REF] Tuck | Land-use intensity and the effects of organic farming on biodiversity: a hierarchical meta-analysis[END_REF][START_REF] Stein-Bachinger | To what extent does organic farming promote species richness and abundance in temperate climates? A review[END_REF]. Lastly, farm inputs are the main characteristic contrasting OF from CF. Although organic plant protection products can be used, no synthetic pesticides or fertilisers are employed for crop production in OF systems. Indeed, in Europe for farms to be considered organic the prohibition of use of synthetic pesticides and fertilisers is the only mandatory practice, other practices such as natural or semi-natural areas management being generally recommended but not systematic in OF systems (European regulation (EU) 2018/848). Recent reviews highlighted that synthetic pesticides are the main factor negatively affecting farmland birds [START_REF] Stanton | Analysis of trends and agricultural drivers of farmland bird declines in North America: A review[END_REF][START_REF] Rigal | Farmland practices are driving bird population decline across Europe[END_REF]. Pesticides can have indirect effects on birds through the elimination of crop pests and weeds, reducing food and habitat resources [START_REF] Hole | Does organic farming benefit biodiversity?[END_REF][START_REF] Gibbons | A review of the direct and indirect effects of neonicotinoids and fipronil on vertebrate wildlife[END_REF]. Furthermore, pesticides lack specificity and can affect non-target species [START_REF] Sánchez-Bayo | Pesticide residues and bees-A risk assessment[END_REF][START_REF] Vyas | Rodenticide incidents of exposure and adverse effects on non-raptor birds[END_REF].

The use of pesticides can thus have direct effects on birds, either inducing lethal effects (immediate death after exposure) or delayed sublethal effects altering life history traits. In fact, different functions of a bird's organism can be affected by pesticides, from the cellular level to higher physiological functions and behaviours [START_REF] Mitra | Synthetic chemical pesticides and their effects on birds[END_REF][START_REF] Moreau | Feeding partridges with organic or conventional grain triggers cascading effects in life-history traits[END_REF]Moreau et al. , 2022a;;[START_REF] Gaffard | Feeding on grains containing pesticide residues is detrimental to offspring development through parental effects in grey partridge[END_REF]. Many experimental studies have revealed altered life history traits due to pesticide toxicity, which may compromise bird survival, and consequently populations dynamics (Lopez-Antia et al. 2015a, 2015b;[START_REF] Ortiz-Santaliestra | Egg overspray with herbicides and fungicides reduces survival of red-legged partridge chicks[END_REF]see Moreau et al. 2022a for a recent review). In natura, birds are exposed to numerous molecules as multiple pesticides are usually applied to fields either at the same time or sequentially, inducing cocktail effects [START_REF] Gaffard | Feeding on grains containing pesticide residues is detrimental to offspring development through parental effects in grey partridge[END_REF]. Moreover, the persistence of some molecules in soil and water can lead to contamination without direct contact with the applied chemicals [START_REF] Jagannath | Eggshell pigmentation indicates pesticide contamination[END_REF][START_REF] Goulson | An overview of the environmental risks posed by neonicotinoid insecticides[END_REF][START_REF] Fuentes | Neonicotinoids: Still present in farmland birds despite their ban[END_REF]. A recent study on wild passerines showed that birds nesting in hedges surrounded by CF were less vigorous than those nesting in hedges surrounded by OF, and that this might be due to pesticide contamination under CF (Moreau et al. 2022b).

Montagu's harriers, Circus pygargus, a migratory raptor species inhabiting agricultural lands in Europe, nest on the ground in cereal crops. In France, populations have declined by 10-30% over the past 30 years [START_REF] Millon | Les populations nicheuses de rapaces en France : analyse des résultats de l'enquête Rapaces[END_REF][START_REF] Comolet-Tirman | Statuts et tendances des populations d'oiseaux nicheurs de France. Bilan simplifié du premier rapportage national au titre de la Directive Oiseaux[END_REF] and the principal cause is believed to be agricultural intensification [START_REF] Butet | Effects of agriculture development on vole dynamics and conservation of Montagu's harrier in western French wetlands[END_REF][START_REF] Arroyo | Conservation of the Montagu's harrier (Circus pygargus) in agricultural areas[END_REF][START_REF] Arroyo | Montagu's harrier. -BWP update[END_REF][START_REF] Santangeli | Identifying effective actions to guide volunteer-based and nationwide conservation efforts for a ground-nesting farmland bird[END_REF]). Montagu's harriers mostly feed on common voles, Microtus arvalis, which display cyclic population dynamics in Western Europe, hence food availability and diet vary from year to year (Millon & Bretagnolle, 2008). In years of low vole abundance, Montagu's harriers compensate by feeding on Orthopterans (grasshoppers and crickets) [START_REF] Salamolard | Responses of an avian predator to variations in prey density at a temperate latitude[END_REF]. For this farmland specialist bird, the type of agricultural system at a local scale might be of great importance during the breeding season. Indeed, eggs and chicks are located on the ground in crop fields, and the altricial chicks stay in the nest during the whole rearing period (~35 days). Consequently, at all developmental stages, Montagu's harrier offspring might be affected by the composition and management of the habitats in the close surroundings. If the agricultural system in the vicinity leads to altered life history traits, survival may be compromised, reducing future recruitment of fledglings into the population of this declining species.

In the present study, we explored the patterns of life history traits variation in nestlings along a gradient of OF near their nests. Accordingly, we investigated the relative impacts of OF vs. CF on multiple life history traits of Montagu's harrier chicks at a local scale (up to 2000 m) around nests. Compared with OF, CF may decrease food availability (due to both lower quality habitats and intensive management practices) and increase exposure to pesticides (through contact or prey consumption). Therefore, we hypothesised that a higher percentage of CF around the nest may adversely affect fledglings. Since all life history traits involve complex interconnected functions, expectations for a single trait are complicated to raise. A multi-trait approach is therefore crucial to elucidate the effects of agricultural systems (OF vs. CF) on birds, and eventually to highlight potential trade-offs between life history traits. We expect chicks reared in nests surrounded by a lower percentage of OF (i.e., a higher percentage of CF) would have a poorer overall health status that might be reflected by inferior body condition, more markers (physical and biochemical) of chronic stress, a depressed immune system, and lower vigour than chicks surrounded by a higher percentage of OF.

Material and methods

Ethics statement

All experiments conformed to the French guidelines for the ethical use of animals in research (APAFIS#18557-2019010822312199v2). Bird handling was allowed and licensed by the Centre de Recherches sur la Biologie des Populations d'Oiseaux (CRBPO; Museum National d'Histoire Naturelle; licence #1308).

Study area and model species

The study site is located in southwestern France (4611′N, 028′W), in the Long-Term Social-Ecological Research 'Zone Atelier Plaine & Val de Sèvre' (LTSER ZAPVS), a 450 km² area where annual crop identification is carried out, allowing mapping of organic crop plots in detail. In France, OF is regulated by the European legislation (European regulation (EU) 2018/848) on organic production, which details all practices to respect for farms to be considered organic, the main one being the complete ban on the use of synthetic pesticides and fertilizers. Other farming practices, including weeding or non-cropped habitat management, being at the discretion of each farmer. Nonetheless, [START_REF] Catarino | Experimental and empirical evidence shows that reducing weed control in winter cereal fields is a viable strategy for farmers[END_REF] report general information on CF and OF systems of the study area; the main differences being smaller farm areas, more diversified crops and a lower nitrogen supply in OF systems. In this intensive agricultural area, cereal crops are the predominant cultures (average coverage of 41.5% between 2009 and 2016), followed by corn, sunflowers and oilseed rape (Bretagnolle et al. 2018b). Natural (or semi-natural) habitats and urbanized areas represent only a small proportion of the study area (average coverage of 13.5% and 9.8% respectively between 2009-2016; Bretagnolle et al. 2018b) thus only organic crop plots were used for analyses (see section 2.7 for further information). Montagu's harrier monitoring has taken place in this site since 1994 (Bretagnolle et al. 2018b). This species nests on the ground, mainly in cereal crops, and lays up to eight eggs [START_REF] Arroyo | Patterns of egg and clutch size variation in the Montagu's Harrier[END_REF]Millon et al. 2008). The incubation period lasts 29 days and the rearing period between 30 and 35 days [START_REF] Arroyo | Interactive effects of food and age on breeding in the Montagu's Harrier Circus pygargus[END_REF]). The mean productivity in this site is 2.05 fledglings per breeding attempt [START_REF] Arroyo | Montagu's harrier. -BWP update[END_REF], breeding success depending mainly on the availability of common voles, their main prey in this area [START_REF] Salamolard | Responses of an avian predator to variations in prey density at a temperate latitude[END_REF]. Foraging range varies with location; in the study area males use home ranges of ~14 km² [START_REF] Salamolard | Utilisation de l'espace par le Busard Cendré Circus pygargus, superficie et distribution des zones de chasse[END_REF], but home ranges during the breeding season can stretch to 100 km² [START_REF] Guixé | Appropriateness of Special Protection Areas for wide-ranging species: the importance of scale and protecting foraging, not just nesting habitats[END_REF].

Overall experimental design

From 2016 to 2021, the reproduction of Montagu's harrier was monitored from late April until late July. All nests were searched by professional ornithologists, and once located, visited and mapped using a global positioning system (GPS) receiver, they were stored on a geographical information system (GIS; QUANTUMGIS 3.16.5; QGIS Development Team, 2021, see Fig. 1).

Nests were visited twice during the incubation period and every week after hatching (lag time between visits 7 ± 2 days). At 15 days old, chicks were sexed and banded with a unique coded aluminium ring from the Museum National d'Histoire Naturelle. Sex was determined by the colour of the iris (brown for females and grey for males) [START_REF] Leroux | Sex ratio variations in broods of Montagu's Harriers Circus pygargus[END_REF].

For the present study, we focused on data collected during the last visit before fledgling, when nestlings were 26 ± 2 days old. Briefly (see details below), chicks were caught in the nest and carefully handled to take morphometric measurements and breath rate and collect blood samples (Fig. 2). A phytohemagglutinin (PHA) skin injection was performed, and behaviour during handling was also recorded (Fig. 2). Chicks were handled and released at the nest, and the handling time was minimised to reduce potential stress. The following day of the visit, nests were again visited to record chicks' inflammatory response to the PHA skin injection (see details below). A total of 380 chicks were monitored (N2016 = 40, N2017 = 32, N2018 = 81, N2019 = 84, N2020 = 85, N2021 = 58) from 137 nests (N2016 = 15, N2017 = 16, N2018 = 24, N2019 = 31, N2020 = 29, N2021 = 22). No sex ratio bias was observed for the entire period (187 females and 193 males; binomial test p = 0.80).

Morphometric measurements

Morphometrics for body condition assessment

Body condition is a morphometric estimate reflecting the overall health status of individuals, notably involved in post-fledgling survival of birds [START_REF] Blums | Individual quality, survival variation and patterns of phenotypic selection on body condition and timing of nesting in birds[END_REF][START_REF] Vitz | Influence of condition and habitat use on survival of postfledging songbirds[END_REF][START_REF] Duijns | Body condition explains migratory performance of a long-distance migrant[END_REF]. Body condition for each nestling was estimated from the scale mass index following [START_REF] Peig | New perspectives for estimating body condition from mass/length data: the scaled mass index as an alternative method[END_REF]:

𝑆𝑀𝐼 𝑖 = 𝑀𝑎𝑠𝑠 𝑖 × ( 𝑀𝑒𝑎𝑛 𝑡𝑎𝑟𝑠𝑢𝑠 𝑙𝑒𝑛𝑔𝑡ℎ 𝑜𝑓 𝑡ℎ𝑒 𝑔𝑟𝑜𝑢𝑝 𝑀𝑒𝑎𝑛 𝑡𝑎𝑟𝑠𝑢𝑠 𝑙𝑒𝑛𝑔𝑡ℎ 𝑖 ) 𝑏 𝑠𝑚𝑎
where 𝑏 𝑠𝑚𝑎 is the slope of the major axis regression of log (body mass) on log (mean of the two measures of the left and right tarsus length) with the standard major axis method (package 'smatr') [START_REF] Warton | smatr 3 -an R package for estimation and inference about allometric lines[END_REF] for each chick i. The mass was obtained by weighing chicks with a Pesola 500 g spring scale (accuracy ± 5 g) (Schindellegi, Switzerland) and the left and right tarsus were measured twice with a digital calliper (accuracy ± 0.1 mm).

Fault bars

Fault bars are malformations in feathers induced by punctual stressors during chick growth, which may result in feather and even rachis breaks [START_REF] Jovani | Fault bars in bird feathers: mechanisms, and ecological and evolutionary causes and consequences[END_REF]; see [START_REF] Rabdeau | Does anthropization affect the physiology, the behaviour and the life-history traits of Montagu's harrier chicks?[END_REF] for an example of recent usage). Chick tails were photographed and the number of fault bars (all breaks in barbs and barbules) was counted on the most affected rectrix feather by the same experimenter to avoid observer bias. The number of fault bars on each chick ranged between 0 and 10.

Colouration

Colouration of nestling bare parts, a carotenoid-related trait, can be limited through the diet and mediated by different physiological effects [START_REF] Garcia-Heras | Pollutants and diet influence carotenoid levels and integument coloration in nestlings of an endangered raptor[END_REF]. The colour of the cere and tarsus (bare parts) were scored by direct comparison with a colorimetric chart from Yolk Colour Fan (Roche, Neuilly-sur-Seine, France). Scores ranged from 0 (almost white / pale yellow) to 15 (orange) and were highly correlated with colour parameters obtained using a spectroradiometer [START_REF] Faivre | Breeding strategy and morphological characters in an urban population of blackbirds, Turdus merula[END_REF]. Cere scores ranged from 0 to 5 and tarsi ranged from 0 to 6. Both scores were highly correlated (Spearman's correlation test, ρ >0.77, p <0.0001, 95% confidence interval CI = [0.72, 0.81], n = 380), and only cere scores were used for statistical analyses.

Physiological measures

Breath rate score

Breath rate usually refers to a proxy of physiological stress [START_REF] Carere | Shy and bold great tits (Parus major): body temperature and breath rate in response to handling stress[END_REF][START_REF] Torné-Noguera | Great Tit (Parus major) breath rate in response to handling stress: urban and forest birds differ[END_REF][START_REF] Rabdeau | To change or not to change experimenters: caveats for repeated behavioural and physiological measures in Montagu's harrier[END_REF]. Breath rate was measured using chicks handled in back position, and corresponded to the number of thoracic movements counted during 15s

for three consecutive replicates. The breath rate score was then obtained by averaging the three replicates.

2.5.2. Physiological parameters assessed from blood

Blood collection

Blood sampling was made on the brachial vein through a puncture using a sterile needle and heparinised capillaries. A single microcapillary of 10 µL was used for both sedimentation rate and haematocrit, a droplet of blood was used for blood smear (see below), and two capillaries of 100 µL each were placed in Eppendorf tubes and kept refrigerated (0-5C) for 2-3 h while transporting to the lab, where they were stored at -20C until further analyses (carotenoid titration and acetylcholinesterase activity; see below).

Sedimentation rate and haematocrit

Erythrocyte sedimentation rate (ESR) corresponds to the proportion of red blood cells sedimented per hour, and it increases in response to various inflammatory or infectious processes [START_REF] Saino | Sexual ornamentation and immunocompetence in the barn swallow[END_REF]. Microcapillaries were kept vertical for 5 h in a refrigerated container (0-5C), lengths were measured with an electronic calliper to the nearest 0.1 mm, and ESR in millimetres per h (mm/h) was estimated as follows:

𝐸𝑆𝑅 = 𝑣𝑜𝑙𝑢𝑚𝑒 𝑛𝑜𝑡 𝑓𝑖𝑙𝑙𝑒𝑑 𝑏𝑦 𝑒𝑟𝑦𝑡ℎ𝑟𝑜𝑐𝑦𝑡𝑒𝑠 𝑡𝑜𝑡𝑎𝑙 𝑏𝑙𝑜𝑜𝑑 𝑣𝑜𝑙𝑢𝑚𝑒 𝑖𝑛 𝑡ℎ𝑒 𝑡𝑢𝑏𝑒 × 𝑡𝑖𝑚𝑒 𝑜𝑓 𝑠𝑒𝑑𝑖𝑚𝑒𝑛𝑡𝑎𝑡𝑖𝑜𝑛 Once ESR was measured, microcapillary tubes were centrifuged at 5000 rpm for 5 min to measure haematocrit. Haematocrit is used as an index of anaemia and corresponds to the proportion of the tube filled with erythrocytes divided by the total volume of blood sample in the tube, given as a percentage [START_REF] Saino | Sexual ornamentation and immunocompetence in the barn swallow[END_REF]; see also [START_REF] Biard | Interpreting immunological indices: the importance of taking parasite community into account. An example in blackbirds Turdus merula[END_REF][START_REF] Moreau | Feeding partridges with organic or conventional grain triggers cascading effects in life-history traits[END_REF].

Carotenoid titration

Carotenoids are involved in different functions of birds, including as antioxidants, and a high concentration in blood can reflect physiological stress [START_REF] Mcgraw | Carotenoids, immunocompetence, and the information content of sexual colors: an experimental test[END_REF]. Eppendorf tubes containing ~150 µL of blood were centrifuged (10 min at 9000 rpm) and plasma was collected and kept at -20C until laboratory analyses. Plasma carotenoid concentration was subsequently obtained in micrograms per millilitre (µg/mL) using a standardised colorimetric technique (see [START_REF] Biard | Carotenoid-based bill colour is an integrative signal of multiple parasite infection in blackbird[END_REF] for additional information).

Acetylcholinesterase activity

Acetylcholinesterase (AChE) activity is negatively affected by different pesticide molecules altering neural functions, which in turn has disruptive consequences on diverse behaviours and complex functions [START_REF] Grue | Neurophysiological and behavioral changes in non-target wildlife exposed to organophosphate and carbamate pesticides: thermoregulation, food consumption, and reproduction[END_REF][START_REF] Eng | Imidacloprid and chlorpyrifos insecticides impair migratory ability in a seed-eating songbird[END_REF]Moreau et al. 2022a). AChE activity in milliunits per minute (mU/min) was obtained by a method derived from [START_REF] Ellman | A new and rapid colorimetric determination of acetylcholinesterase activity[END_REF] using 5 µL of blood diluted in 100 µL of a 1% Triton X-100 solution and measured using a SpectraMax iD3 microplate reader (Molecular Devices Inc., Sunnyvale, CA, USA) over 40 min at 324 nm and 37C. AChE activity was calculated using SoftMax Pro7 software (Molecular Devices, San Jose, CA, USA) as the maximum change in absorbance per minute during kinetics analysis.

Blood smear

For each bird, a blood smear was performed with a droplet of blood sample. Slides were airdried and fixed in absolute methanol before being stained with Giemsa solution (Sigma Chemical, St. Louis, MO, USA). Blood smears were examined with an oil immersion microscope under 1000x magnification. Total white blood cells (WBC) count was estimated from the number of leucocytes found among 8000 erythrocytes. A high number of leucocytes reflects stress and inflammatory processes [START_REF] Ots | Haematological health state indices of reproducing great tits: methodology and sources of natural variation[END_REF]. Different types of leucocytes were counted and used to estimate the relative proportion of heterophils to lymphocytes (H/L ratio). The H/L ratio is used as an index of physiological stress, as it is supposed to increase in response to various stressors [START_REF] Maxwell | Avian blood leucocyte responses to stress[END_REF][START_REF] Ots | Haematological health state indices of reproducing great tits: methodology and sources of natural variation[END_REF][START_REF] Moreau | Feeding partridges with organic or conventional grain triggers cascading effects in life-history traits[END_REF]see Biard et al. 2015 for details).

Immune challenge

Before release into the nest, the immune system of nestlings was challenged using phytohemagglutinin (PHA) skin tests [START_REF] Smits | Simplifying the phytohaemagglutinin skin-testing technique in studies of avian immunocompetence[END_REF]. Briefly, 0.5 ± 0.1 mg of PHA (Sigma Chemical, St. Louis, MO, USA) diluted into 0.1 mL of phosphate-buffered saline (PBS) was injected subcutaneously in the wing patagium as described by Sternalski et al. (2012a). The thickness of the patagium was measured three times before injection using a pressuresensitive dial thickness gauge (spessimeter accuracy ± 0.01 mm) and again 24 h after injection.

The response to the PHA skin test corresponds to the change in average thickness (in mm) of the injection site, and thus the magnitude of this change reflects the immune potential of nestlings (Sternalski et al. 2012a; see also [START_REF] Biard | Interpreting immunological indices: the importance of taking parasite community into account. An example in blackbirds Turdus merula[END_REF].

Behavioural measures

Behaviour is considered an indicator of individual health that can be altered by multiple chemicals and is often regarded as a more sensitive response to stress or toxicity levels than other physiological measures [START_REF] Grue | Neurophysiological and behavioral changes in non-target wildlife exposed to organophosphate and carbamate pesticides: thermoregulation, food consumption, and reproduction[END_REF][START_REF] Clotflelter | The role of animal behaviour in the study of endocrine-disrupting chemicals[END_REF][START_REF] Hellou | Behavioural ecotoxicology, an "early warning" signal to assess environmental quality[END_REF]Moreau et al. 2022a). When handling chicks (i.e., from capture in the nest to release), the behaviour of bird was scored using four behavioural traits as described by [START_REF] Rabdeau | To change or not to change experimenters: caveats for repeated behavioural and physiological measures in Montagu's harrier[END_REF]: (i) activity during nest approach (act-nest); (ii) total activity during handling (act-tot); (iii) rate of beak attacks (beak); (iv) rate of claw attacks (claws). Act-nest was assessed when the experimenter approached the nest to catch the chicks and scored as follows: stayed motionless = 0, chicks retreated = 1, chicks ran away = 2. Act-tot was measured throughout handling and scored as follows: chicks stayed motionless = 0, chicks rarely moved = 1, chicks moved regularly = 2, chicks moved continuously = 3. The number of beak and claw attacks were counted during handling and divided by the duration of handling, resulting in a rate (number of attacks per minute).

Statistical analyses

A principal component analyses (PCA) was performed on the four behavioural scores using singular value decomposition of a standardised (centred and scaled) data matrix [START_REF] Crawley | The R Book[END_REF]. The first axis of the PCA reflecting global activity and vigour during handling (PC1activity) was retained as its eigenvalue was >1, and it accounted for 48.69% of the variance. PC1-activity was positively correlated with both rates of beak (r = 0.56) and claw (r = 0.57) attacks, act-tot (r = 0.55) and act-nest (r = 0.24) scores.

Normality and homoscedasticity of all response variables were tested using the 'fitdistrplus' package prior to model implementation, and when these assumptions were not met, distribution with the best fit was selected using the same package (Delignette-Muller & Dutang, 2015). Model residuals and spatial autocorrelation (using Moran's I) were also checked using 'DHARMa' package [START_REF] Hartig | _DHARMa: Residual Diagnostics for Hierarchical (Multi-Level / Mixed) Regression Models_. R package version 0[END_REF]. PC1-activity, body condition, ESR, breath rate score, AChE activity and PHA response were used as response variables in Linear Mixed-effects Models (LMMs). Carotenoid concentration was square root transformed, haematocrit and H/L ratio were log-transformed prior to LMM execution, since the distributions of these variables did not meet normality. Generalised LMMs fitted with a Poisson distribution (PGLMMs) were implemented for fault bars and colouration scores and a Generalised LMM fitted with a Negative Binomial distribution (NB-GLMM) was used for the total WBC count. For all models (LMMs, PGLMMs and NB-GLMM), the percentage of OF around the nest (log-transformed, hereafter OF effect) and the year as factor were included as fixed effects, and the nest identity as a random effect (to avoid pseudo-replication as chicks can belong to the same nest) (Bolker et al. 2009). The year was considered a fixed effect because we wanted to control for interannual variability of environmental conditions (phenology, food availability and weather for instance). Because year could have been considered a random factor, we ran all models changing only the year fixed effect into a random effect. This approach did not change the results obtained with year as a fixed effect, hence we used the results of the first analyses. All models were run with the 'lme4' package [START_REF] Bates | Fitting linear mixed-effects models using lme4[END_REF].

As we had no a priori information on the scale of the OF effect on life history traits for each chick, we estimated the percentage OF around the nest for 20 buffer sizes (i.e., radii every 100 m, from 100 m to 2000 m). To this end, the GPS coordinates of nests and a database of the LTSER ZAPVS, which georeferenced the agricultural plots using OF practices, were used with the 'sf' package [START_REF] Pebesma | Simple features for R: Standardized support for spatial vector data[END_REF]. Since OF agricultural plots varied over time, the percentage of OF around nests for a given year was calculated using the georeferenced plots of the corresponding year. We ran 20 models per life history trait (one per buffer size) and extracted an Akaike Information Criterion corrected for small sample size (AICc) using the 'AICcmodavg' package [START_REF] Mazerolle | AICcmodavg: Model selection and multimodel inference based on (Q)AIC(c). R package version 2[END_REF]. From a set of candidate models, AICc allows selection of the model with the most support, which has the lowest AICc, assuming that models with ΔAICc <2 (i.e., a difference in AICc between candidate models <2) are similarly supported and cannot be distinguished from each other [START_REF] Burnham | Multimodel inference: understanding AIC and BIC in model selection[END_REF]. For every life history trait, a plot was generated representing the AICc of the 20 models (one for each buffer size), with a red dotted line illustrating the model selection criterion (lowest AICc of the candidate models + 2), with the shape and colour reflecting the significance and direction of the OF effect. To avoid type II errors, we chose to present our results based on effect sizes (estimates of the model) assorted with their 95% confidence intervals (CI) (following [START_REF] Garamszegi | Comparing effect sizes across variables: generalization without the need for Bonferroni correction[END_REF][START_REF] Nakagawa | A farewell to Bonferroni: the problems of low statistical power and publication bias[END_REF][START_REF] Nakagawa | Effect size, confidence interval and statistical significance: a practical guide for biologists[END_REF][START_REF] Cumming | The new statistics: Why and how[END_REF].

The OF effect direction corresponded to the sign of the estimate and was considered significant when 95% CI did not include 0. Calculation of the bootstrapped CI was obtained for 2000 iterations and using the package 'boot' [START_REF] Canty | boot: Bootstrap R (S-Plus) Functions[END_REF]. We discuss effect sizes assorted with their 95% CI which allow to quantify the biological effect we tested and provide comparable estimates for future research and meta-analyses, still, we provide the results from model comparison using likelihood ratio-based χ 2 -statistics in supplementary materials [START_REF] Fox | Multivariate linear models in R. An R Companion to Applied Regression[END_REF].

To assess potential links between variables, Spearman's rank correlation tests were performed. All statistics were conducted with R v4.0.5 software (R Core Team, 2021), implemented using the aforementioned packages.

Results

Summary statistics and Spearman's rank correlation coefficients of the 12 life history traits are presented in the Supplementary Material (see Table S1 and Fig. S2). Only ESR and haematocrit (Spearman's correlation test, ρ > -0.42, p <0.0001, 95% CI = [-0.51; -0.32], n = 343) were correlated (Supplementary Material Fig. S2). The set of candidate models for body condition, fault bars, breath rate score, ESR, haematocrit, carotenoid concentration, AChE activity, WBC count, and PHA skin test did not yield any specific buffer size with substantial support (i.e., AICc <2 with a significant OF effect) for an influence of OF percentage around nests (Fig. S2 in supplementary materials). For cere colouration score and H/L ratio, the best models (i.e., those with ΔAICc <2) identified a positive effect for OF percentage at a range of radii (Fig. 3).

The cere colouration score increased with OF percentage around the nest for buffer sizes >500 m (Fig. 3a). For H/L ratio, a positive relationship was detected between 300 and 900 m (Fig. 3b). Concerning the PC1-activity, a positive relationship with the percentage of OF was detected at 200 m around the nests.

We then selected the model with the lowest AICc to probe the relationships between the trait and the OF percentage around the nest (Fig. 4). Chicks displayed a significantly higher cere colouration score when surrounded by a higher percentage of OF at 1500 m (effect size = 0.14; 95% CI = [0.04; 0.22]), the lowest AICc for this life history trait (Fig. 4a). The lowest AICc for the H/L ratio was at a radius of 600 m around nests; at this distance the H/L ratio significantly increased with increasing OF percentage (effect size = 0.11; 95% CI = [0.04; 0.18]; Fig. 4b). For PC1-activity, the lowest AICc was at 200 m, an increase in OF at this distance from nests significantly increased the global activity of chicks (effect size = 0.20; 95% CI = [0.06; 0.40]; Fig. 4c).

Discussion

The aim of the present study was to explore life history traits variation of Montagu's harrier chicks linked to agricultural systems around nests. The results revealed an effect on two of the 12 traits investigated, although moderate. Cere colouration and H/L ratio were positively linked to OF at different radii, suggesting that the effects of the agricultural system operate at different distances, but not necessarily at the same range. The cere colouration score increased with increasing OF percentage at distances >500 m with the best model selected at 1500 m. The H/L ratio increased significantly with OF at distances from 300 to 900 m, the best model being at 600 m. Global activity of chicks increased with increasing OF percentage at 200 m only. For all other life history traits, namely body condition, fault bars, breath rate score, ESR, haematocrit, carotenoid concentration, AChE activity, WBC count, and PHA skin test, no effect of OF was detected at any of the buffer sizes.

In the present study, we found evidence for the impacts of farming practices (CF and OF) on Montagu's harrier chicks at a low spatial scale (<2 000 m) around nests. First, OF was found to be associated with a higher H/L ratio. This trait is indicative of stress undergone by chicks during their rearing period. Indeed, H/L ratio, is considered a reliable indicator of mild to moderate stress, and to reflect chronic stress levels rather than short-term stress responses [START_REF] Maxwell | Avian blood leucocyte responses to stress[END_REF][START_REF] O'dell | A comparison of techniques measuring stress in birds[END_REF]. This result suggests that chicks in areas with a higher OF percentage may suffer more disturbance. Indeed, compared with CF, OF practices involve more repetitive passages [START_REF] Karlen | Nutrient, labor, energy and economic evaluations of two farming systems in Iowa[END_REF][START_REF] Reissig | Workload on organic and conventional family farms in Switzerland[END_REF][START_REF] Orsini | Labour use on organic farms: a review of research since 2000[END_REF], which may equate to more stress experienced by chicks. Furthermore, areas with more OF fields are generally better appreciated than areas with CF fields by urban inhabitants attracted to naturalistic features and elements [START_REF] Zasada | Multifunctional peri-urban agriculture-A review of societal demands and the provision of goods and services by farming[END_REF]. Diversified habitats offer more opportunities for recreation and leisure than uniform ones [START_REF] Hansen | Nature and nature values in organic agriculture. An analysis of contested concepts and values among different actors in organic farming[END_REF]. Anthropic disturbances can induce stress in Montagu's harrier chicks, leading to physiological responses to this stress and even to the formation of fault bars (see [START_REF] Rabdeau | Does anthropization affect the physiology, the behaviour and the life-history traits of Montagu's harrier chicks?[END_REF]. Repeated human activities, whether related to OF practices or recreation, could also induce prolonged absence of adults, resulting in reduced parental care, and thus more stress. Additionally, a higher presence of conspecifics or other raptor species around OF crops is also possible.

Competition between Montagu's harriers nesting in semi-colonies is presumably low but could increase in years of low vole abundance. The presence of other raptor species foraging in the same area would also increase competition for food [START_REF] Garcia | Intra-and interspecific agonistic behaviour in sympatric harriers during the breeding season[END_REF].

Competition for food resources might increase the time parents spend foraging, leaving chicks alone for longer periods. The presence of more raptors may increase the risk of predation of Montagu's harrier nests, which are particularly vulnerable and attractive in low vole years [START_REF] Arroyo | Colonial breeding and nest defence in Montagu's harrier (Circus pygargus)[END_REF]. Pairs are often seen mobbing common buzzards, Buteo buteo, or black kites, Milvus migrans, to protect their nests (authors' personal observations), and this might result in more frequent parental absences. Consequently, an increase in intra-and interspecific negative interactions surrounding OF crop plots would increase the time chicks are unattended. An alternative explanation for the higher H/L ratio observed for chicks in nests surrounded by a high OF percentage is the presence of a higher number and/or diversity of pathogens. Indeed, H/L ratio is known to increase in response to infectious diseases as well as psychological disturbances [START_REF] Ots | Haematological health state indices of reproducing great tits: methodology and sources of natural variation[END_REF]), hence we cannot exclude the potential effect of increased pathogen exposure in OF systems. Pesticides may reduce pathogen diversity in CF systems compared with areas in which OF is more prevalent [START_REF] Goutte | Benefits of Parasitism in Polluted Environments: A Review and Perspectives[END_REF].

Therefore, chicks from nests located in these zones might be more prone to parasite exposure, which could, in turn, explain why they exhibit a higher number of heterophil leucocytes, and thus a higher H/L ratio. Moreover, several studies reported that H/L ratio is species-specific, and that it may vary between captive and wild individuals, depending on sex, age, nutritional and infectious status, and even brood size [START_REF] Ruiz | Hematological parameters and stress index in rufous-collared sparrows dwelling in urban environments[END_REF][START_REF] Limiñana | Blood chemistry and hematologic values in free-living nestlings of Montagu's Harriers (Circus pygargus) in a natural habitat[END_REF][START_REF] Ortego | Physiological response to stress in fledgling Lesser Kestrels Falco naumanni: the role of physical condition, sex and individual genetic diversity[END_REF][START_REF] Parejo | Immunity and fitness in a wild population of Eurasian kestrels Falco tinnunculus[END_REF][START_REF] Banbura | Spatial and temporal variation in heterophil-tolymphocyte ratios of nestling passerine birds: comparison of blue tits and great tits[END_REF][START_REF] Catitti | Hungry or angry? Experimental evidence for the effects of food availability on two measures of stress in developing wild raptor nestlings[END_REF]. Previous studies on Montagu's harriers attempted to set reference interval values for haematological parameters of captive adults and free-living nestlings, but reliable comparisons were difficult to make with the present work, and further investigation is required [START_REF] Lavin | Hematological values of Montagu's harrier (Circus pygargus)[END_REF][START_REF] Limiñana | Blood chemistry and hematologic values in free-living nestlings of Montagu's Harriers (Circus pygargus) in a natural habitat[END_REF].

Additionally, even if chicks in OF and CF areas face similar parasite diversity, their investment in different life history traits might vary. For instance, chicks from nests mainly surrounded by CF may face a trade-off between investment for immune parameters and other life history traits such as growth [START_REF] Sheldon | Ecological immunology: costly parasite defences and tradeoffs in evolutionary ecology[END_REF][START_REF] Norris | Ecological immunology: life history trade-offs and immune defense in birds[END_REF]. A recent study on grey partridge showed that hatchlings exposed to pesticides were of lower body size compared with those not exposed, but they underwent compensatory growth to compensate for the initial differences [START_REF] Gaffard | Feeding on grains containing pesticide residues is detrimental to offspring development through parental effects in grey partridge[END_REF]). In the present work, we did not observe any difference in body condition, but growth trajectories should be properly investigated to exclude this hypothesis.

Although we did not detect any effect of the agricultural system on the immune responses of nestlings assessed by PHA skin tests at a local scale, colouration of the cere, a sexual trait indicative of chick quality [START_REF] Sternalski | Carotenoid limitation and allocation priorities in asynchronous raptor nestlings[END_REF], was lower in chicks mainly surrounded by CF. Various studies highlighted the covariation between colouration and immunocompetence in bird nestlings, due to the role of carotenoids in both immune functions and colouration [START_REF] Saino | Better red than dead: carotenoid-based mouth coloration reveals infection in barn swallow nestlings[END_REF][START_REF] Saino | Gape coloration reliably reflects immunocompetence of barn swallow (Hirundo rustica) nestlings[END_REF][START_REF] Baeta | Carotenoid trade-off between parasitic resistance and sexual display: an experimental study in the blackbird (Turdus merula)[END_REF]. Thus, competing demand for carotenoids for colouration and the immune system might occur. At a local scale, a large percentage of CF is presumed to reduce available resources; carotenoids are provided by the diet, which may explain why chick colouration is adversely affected compared with chicks surrounded by OF. Another potential trade-off leading to lower colouration scores in chicks surrounded by CF is the allocation of carotenoids to detoxification processes [START_REF] Edge | The carotenoids as antioxidants -a review[END_REF][START_REF] Krinsky | The antioxidant and biological properties of the carotenoids[END_REF] see [START_REF] Bustnes | Immune status, carotenoid coloration, and wing feather growth in relation to organochlorine pollutants in great black-backed gulls[END_REF]. Indeed, exposure of birds to pollutants is known to induce oxidative stress, leading to a higher demand for antioxidants (including carotenoids) for redox processes [START_REF] Ortiz-Santaliestra | Pollutant accumulation patterns in nestlings of an avian top predator: biochemical and metabolic effects[END_REF][START_REF] Moreau | Feeding partridges with organic or conventional grain triggers cascading effects in life-history traits[END_REF], reviewed in Moreau et al. 2022a). Some studies on wild raptors found compensatory responses between different functions, linked to low concentrations of pesticides altering bird health [START_REF] Bustnes | Immune function and organochlorine pollutants in arctic breeding glaucous gulls[END_REF][START_REF] Rivera-Rodríguez | Incidence of organochlorine pesticides and the health condition of nestling ospreys (Pandion haliaetus) at Laguna San Ignacio, a pristine area of Baja California Sur, Mexico[END_REF][START_REF] Ortiz-Santaliestra | Pollutant accumulation patterns in nestlings of an avian top predator: biochemical and metabolic effects[END_REF][START_REF] Garcia-Heras | Blood concentrations of PCBs and DDTs in an avian predator endemic to southern Africa: Associations with habitat, electrical transformers and diet[END_REF]. Despite the absence of a significant effect of the agricultural system on plasma carotenoid concentrations in the present work, we can suppose that pesticides used in CF might affect carotenoid allocation in Montagu's harrier nestlings, leading to trade-offs involving other physiological functions such as detoxification and oxidative stress, not measured here. Nevertheless, even if CF and OF provide an equivalent amount of food resources, it is possible that OF provides a greater diversity of food supply. Indeed, as mentioned above, OF promotes insect and passerine bird diversity, both of which are alternative prey for Montagu's harriers. Nestling colouration mainly depends on the proportion of insects and birds in their diet since these are richer in carotenoids [START_REF] Sternalski | Carotenoids in nestling Montagu's harriers: variations according to age, sex, body condition and evidence for diet-related limitations[END_REF]. Thus, it is likely that chicks surrounded by OF benefit from higher carotenoid levels in their diet, resulting in more intense colouration of secondary sexual traits.

The global activity level of chicks has been found to be higher when the surrounding OF at 200 m increases. This result matches with previous findings on 6 passerine birds (Moreau et al. 2022b), which reported a higher vigour of individuals when they originated from organic hedgerows compared to those captured in conventional ones. The main hypothesis raised by the authors is a lower exposition of birds to pesticides in OF systems and not an increased food abundance as birds did not show variation in body condition (Moreau et al. 2022b). In our case, the same pattern arises as we did not detect an effect of OF on body condition of Montagu's harrier chicks. Still, as we detected a beneficial effect of OF on chicks' activity only at 200 m around nests, we can question the potential role of pesticides in their behavioural differences. The increased chick activity when there is more OF at 200 m could be alternatively explained by human disturbances. In fact, after crops harvests, nests are exposed to curious walkers who can visit nests untimely, generating aggressive reactions of chicks and eventually sensitization to humans [START_REF] Rabdeau | To change or not to change experimenters: caveats for repeated behavioural and physiological measures in Montagu's harrier[END_REF][START_REF] Rabdeau | Does anthropization affect the physiology, the behaviour and the life-history traits of Montagu's harrier chicks?[END_REF]. However, even if the effect size seems to indicate a medium effect of OF on the vitality of chicks, the assorted CI is relatively large suggestive of a large variability among chicks [START_REF] Nakagawa | Effect size, confidence interval and statistical significance: a practical guide for biologists[END_REF][START_REF] Cumming | The new statistics: Why and how[END_REF], interpretation on this result should thus be considered with caution.

Conclusion

In the current context of overall biodiversity loss, it is crucial to understand the relative impacts of CF and OF practices which are often opposed one to the other. However, as evidenced by our results, the picture is quite more complex than just black and white. The multi-trait approach used here revealed that CF practices can have detrimental effects on chick quality, and that OF can also exert adverse effects on their physiological stress. In fact, studying only the colouration of secondary sexual traits would have led to partial conclusions about the relative effects of CF on chick quality, whereas studying additionally morphometric and physiological traits that are indicative of stressors, such as H/L ratio, facilitated complete conclusions. In fact, both agricultural systems may impact Montagu's harrier chicks, with potential consequences for immediate survival during migration and later during adulthood.

Multi-trait approaches are important because population dynamics reflect complex physiological processes of individuals that shape life history traits (development, reproduction, and survival), which in turn impact populations.

However, the present study is preliminary, as based on a rough cost/benefit analysis of these agricultural systems. One may also consider that we did not expect large effect size for the impact of agricultural systems on chicks since the relationships among the different life history traits and how they are shaped are quite complex. For example, this study did not consider that chicks are fed by their father, who may hunt up to several kilometres away, potentially masking/compensating for low habitat quality at the local scale [START_REF] Guixé | Appropriateness of Special Protection Areas for wide-ranging species: the importance of scale and protecting foraging, not just nesting habitats[END_REF][START_REF] Krupiński | Ranging behaviour and habitat use in Montagu's Harrier Circus pygargus in extensive farmland of Eastern Poland[END_REF]. Our study only considered the habitats in the vicinity of the nest from a binary point of view (OF vs. CF), and therefore did not capture all parameters such as the proportion of natural, semi-natural or urban habitats in the landscape potentially influencing chick health. Human activities related to OF and CF should be investigated in more detail to further determine the effects of different types and frequencies of disturbances on ground-nesting farmland birds. Finally, OF and CF contrast in their use of pesticides, which may partly explain some of our results (see Moreau et al. 2022a for an overview), especially knowing that pesticide use is the main driver of birds' decline in Europe [START_REF] Rigal | Farmland practices are driving bird population decline across Europe[END_REF].

Therefore, in future work, titration of pesticides in blood should be performed to explore their impact on chick health.

Figure legends

Fig. 1 :

 1 Fig. 1: Spatial distribution of Montagu's harrier nests across the Long-Term Socio-Ecological

Fig. 2 :

 2 Fig. 2: Summary of the experimental design measuring different morphological, physiological

Fig. 3 :

 3 Fig. 3: Model selection using AICc to assess the responses of life history traits (life history

Fig. 4 :

 4 Fig. 4: Organic farming effect on (a) cere colouration score, (b) H/L ratio, and (c) PC1-activity

Acknowledgements

We would like to thank everyone who contributed to fieldwork and data collection during the 6 years of this study: Romain Bonnet, Pierre de Bouët du Portal, Landry Boussac, Guillaume Brouard, Mélissa Desbonnes, Agathe Gaffard, Mazzerine Laboureau, Gildas Lemonnier, Camille Mangelinck, Mona Moreau, Laure Pellicier, Guillaume Peplinski, Jean-Samuel Perroteau, Baptiste Perrotin, Juliette Rabdeau, Margaux Ruiz, Anna Terras, Alexis Veldeman and Alexandre Villers. We are also grateful to all farmers who allowed us to access their fields.

A special thanks to Thomas Schneider-Bruchon for his qualitative help on statistical analyses.

Funding

This work was supported by the French National Research Agency (ANR JCJC PestiStress, grant #19-CE34-0003-01), the French National Centre of Scientific Research (CNRS), the French National Research Institute for Agriculture, Food and the Environment (INRAE), the University of La Rochelle, and the region Nouvelle-Aquitaine, with partial funding through the BioBird project.

Declaration of competing interest

The authors declare that they have no competing financial interests or personal relationships that could have appeared to influence the work reported in the present study.