Alain Recking

Sebastien Zanker

François Lauters

M Regazzoni

Thomas Geay

B Camenen

L Brunel

F Fontaine

Measurement of sand transport with a submerged pump: presentation of the results of a test campaign carried out on the Isère River in July 2019

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

INTRODUCTION

Rivers transport sediments from the mountain to the sea, as bedload or suspension. Bedload comprises coarse sediments transported in contact with the bed, whereas suspension comprises fine sediments transported in the main flow. These definitions may appear simple, but it is no longer the case as soon as one questions what is 'coarse' and what is 'fine'? The sand fraction (often defined in the range [63m-2mm]) is usually used as a limit. But, obviously there is no strictly speaking a size limit between the two mode of transport, and sand can be present both in bedload and suspended load. Because sand does not clearly be associated with one or the other mode of transport it is usually excluded from sampling operations: bedload is often measured for the coarse fraction (considering sizes > 2mm for avoiding samplers clogging) whereas suspension is usually measured for the finest fraction (excluding coarse sands). As a result sediment budgets do not include a confident estimate of the sand fraction, which is clearly a problem, because anyone could observe that sand is far to be negligible in deposition zones (dams..). Sand is present in all parts of the river system. It controls the mobility of the bed, is a support for ecosystems and sand carried by rivers plays a key role in coastal morphodynamics [Bendixen et al., 2019].

To overcome this problem, samplers were specifically designed for measuring sand transport. Some bedload samplers are adapted for bedload measurement in presence of small bedforms (the Arnhem BTMA, the Nile sampler [L C Van Rijn and Gaweesh, 1992]), but they are not adapted for measuring sand transported in the water column. Other samplers such as the P61 or P72 [Davis, 2005] are more adapted for the water column, but cannot measure very close to the bed. The Delft bottle sampler [Beverage and Williams, 1989] was designed for measuring both close to the bed and in the water column. These samplers were specifically designed for not per-Measurement of sand transport with a submerged pump: presentation of the results of a test campaign carried out on the Isère River in July 2019 A.Recking*, F.Lauters**, S.Zanker**, M.Regazzoni*, T.Geay****, B.Camenen***, L.Brunet**, F.Fontaine* ABSTRACT: In alpine and piedmont gravel bed Rivers, sand can be transported as bed load or as suspended load over a coarser bed. This explains why most of the monitoring tools, which have been developed for bed load or suspended load, do not adequately measure the sand fraction, which is mainly transported at the transition between these two modes. This difficulty in measuring the sand fraction is a real problem because sand is a major component of the total load transferred from the mountain to the sea. Some techniques have been specially designed for sand measurement; they can be effective but not always easy to deploy. That is why we propose here to test a new method, based on a submerged pump initially developed for hostile environments such as drilling operations, and capable of withstanding high loads of fine sediments. The pumping tests were carried out in the Isère River during July 2019, and compared to a isokinetic flux sampler: the Delft bottle. We present the results and limitations of the method.

turbing the flow: they are supposed to be isokinetic, which means that their presence is transparent for the local flow and that they sample at the same velocity than ambient flow velocity.

However all these samplers are heavy devices designed to be stable in highly turbulent flows and must be handled with a winched cable from a truck or a boat. As a consequence measurements are time and money consuming and sampling a large river section can take several hours (usually one day for the whole section). This can induce a real bias because in the same time discharge and sediment rate can vary greatly. In addition, such devices are not adapted to sand transport measurement in small energetic alpine rivers (with low flow depth, but important sand load). As an alternative, other technics such as pump-samplers have been tested in the past [L C Van Rijn and Schaafsma, 1986]. These approaches were however limited by the available technology limiting the pumping height and velocities. In this paper we present new tests performed with a submerged pump specifically developed for large pumping height in drilling operations.

The tests were done in the Isère River during July 2019 in comparison with the Delft bottle. We present the results and limitations of the method.

MATERIAL AND METHOD

The pumping device

After a review of available pumping systems, we finally chose to test a submerged pump which allows, in comparison with other solutions (such as peristaltic pumps), higher intake velocities and pumping height. We used the 12 Volt Stainless Steel Monsoon XL distributed by Proactive Environmental Products. It is a centrifugal, single-cell, submerged pump initially developed for hostile environments such as drilling operations, and capable of withstanding high loads of fine sediments up to 35m. The pump is controlled by a separate servo amplifier, allowing to adapt the intake velocity to the ambient flow velocity. A series of trial and error tests were required to adapt the pump to our objectives, and we finally designed the PVC support shown in middle panel of Figure 1: after entering the device through the intake nozzle (Figure 1D), the water is filtered with a 1 mm mesh (Figure 1B) in order to protect the pump (Figure 1A) from the coarsest grains which settle in a little reservoir (Figure 1C). We used an intake nozzle of 7 mm in order to increase the intake velocities up to 2.5m/s.

Method

For comparison we also used a Delft Bottle sampler which permits a direct measurement of the local average sand transport. The bottle geometry causes a strong reduction of the flow velocity inside the bottle and settling of the sand particles larger than about 50 m. Because the capture and settling are not fully efficient, a calibration coefficient must be applied to the measured mass depending on the nozzle intake, the local flow velocity and the mean sand diameter.

For the tests the instruments (pump and Delft Bottle) were mounted on the Delft Bottle metallic support, which was handled with a winched cable from a truck positioned on a bridge. Some tests were done with both the Delft Bottle and the pump, and other tests were done with the pump only (Figure 2). The Delft Bottle can measure at a height of 5cm above the bed. The pump was installed above the bottle nozzle which means that the first measurement was possible at 35 cm above the bed only. When the pump was considered alone it could be positioned closer to the bed.

The water was pumped to the surface through a 8 mm flexible hose. Because we could observe partial obstruction of the pump nozzle with small particle of vegetation during the preliminary tests, we regularly stopped very shortly the pump during sampling which permits to clean the filter and intake nozzle by a backwash effect (due to the water column pressure). After each sampling, the pumped sediment was mixed with the sediments collected in the settling reservoir.

All samples (from the pump and the Delft Bottle) were sieved with a 50 m size mesh, oven dried, weighted and analyzed in the laboratory. Because the objective was to be closer to the isokinetic conditions, we also measured the ambient mean flow velocity using an ADCP. This permitted to adapt the intake nozzle for both Delft Bottle and the pump.

MEASUREMENT CAMPAIGN

Study site

A two days field campaign was done in the Isere river in 2019 (June 27 and July 4), at a location called the Campus bridge. At this location, the watershed is about 5700km², the river width is 60m and the slope is approximately 0.0005 m/m. The morphology shows alternate bars, where a Wolman count indicates a gravel bed with D 50 =1.4cm and D 84 =2.5cm. The Isere river has a pluvionival type hydrological regime, and it transports a lot of sand both as bedload and suspension. We measured from a bridge located approximately 10 m above the water surface.

Measurements

Measurements were done on July 4 (a first set not presented here was done on June 27 and allowed us to adjust the protocol and to do a first comparison between the DB ad the pump). During the day the discharge varied in the range 280-295 m3/s. The vertical profile was considered at 13m from the left bank where the ADCP indicate a surface flow velocity of approximately 1.5 m/s and a near bed flow velocity of 0.7 m/s. The average flow velocity was 1.25 m/s and the flow depth was approximately 2.5m.

We performed 3 runs summarized in Table1. In Run1 we measured simultaneously with both the Delft Bottle (used with the small straight nozzle) and the pump for comparison. In run 2 and 3 the pump was used alone. In Run 2 we stopped the run between each sample on the vertical to collect the coarse sand settled in the reservoir (Figure 1C). In Run 3, we did a continuous meas-urement on the vertical and the coarse sand collected at the end of the run in the settling reservoir (Figure 1C) was considered representative for the whole vertical.

RESULTS

Considering that we used the small straight nozzle, that the median sand diameter was coarse (>130 m) and that the average velocity was 1.25 m/s, a coefficient of 0.7 was deduced from the abacus provided with the sampler and applied to each Delft Bottle measurements. Concerning the pump, all measurements were done with an intake velocity of 2-2.5 m/s considering preliminary results with the ADCP. Actually, after analysis of the ADCP data, the flow velocity on the vertical was rather around 1.25 m/s, which means that we did not pump with isokinetic conditions.

Figure 3 presents the sand concentrations measured for the 3 runs. In Figure3a fluxes measured with the Delft Bottle in Run1 are converted to concentrations using the ADCP flow velocities, and are compared to the concentrations measured with the pump. The results are consistent (with slightly larger values observed for the pump), keeping in mind the large uncertainties associated with the abacus coefficient and with the non-isokinetic conditions for the pump.

Figure 3b compares the concentration profiles measured for each runs. An uncertainty exists for run3 because the coarse sand fraction collected in the settling reservoir was affected proportionally to each sample over the profile. However, we observe that the general shape of the concentration profile is conserved, and also that concentrations increase and decrease during the day, and information which was confirmed by independent measurements with a turbidimeter (supposing it can be used as a proxy for the transport dynamics) measuring on the left bank not far downstream.

The median diameters D 50 measured for samples of Run1 are compared in Figure 4. The diameters are consistent, and indicate relatively coarse sand. The figure also shows that sediments collected with the Delft Bottle are coarser than sediments collected with the pump. This can be explained by the fact that the pump collects integrality of the sand present in the flow, whereas the Delft Bottle collects only the part that actually settles in the bottle, with a capture efficiency which is less for the finest fraction <100µm [Beverage and Williams, 1989]. The results of these preliminary tests are very encouraging because we found a good consistency between the pump and the Delft Bottle. However it must be acknowledged that these results are associated with large uncertainties. The first uncertainty concerns the Delft Bottle measurements we used here as a reference. This system does not measure a concentration but a mass settled in the bottle, which supposes that what leaves the sampler is negligible. Several studies (recalled in [Beverage and Williams, 1989;L C Van Rijn and Schaafsma, 1986]) have compared the Delft Bottle with other samplers (such as the P61 measuring directly a concentration instead of a mass) and concluded that it was inefficient to capture diameters smaller than 100m, and as a consequence underestimate sand transport [63m -2mm] in a ratio of approximately 1:2. Uncertainties also exist with the correction coefficients we applied to the Delft Bottle measurements [J Dijkman, 1978;J. Dijkman and Milisic, 1982], and with additional sampling during raising and lowering of the instrument [L Van Rijn and Roberti, 2019].

Another source of uncertainties concerns the hydraulics conditions. Despite the ADCP measurements, we could not respect isokinetic conditions. How the sampling efficiency is impacted when the velocity inside the intake nozzle is larger or smaller than the ambient flow velocity is largely unknown. Available studies [Gray and Landers, 2014;Starosolszky, 1981] suggest that relative sampling rates (ratio of intake velocity to ambient flow velocity) larger than 1 leads to underestimation of the actual sand concentration and that inversely relative sampling rate smaller than 1 leads to overestimation; this error increases with coarse sands. But flume analysis [Nelson and Benedict, 1950] suggest that deviating intake velocity does not results in large error (<20%) provided that sampling rates are in the range [0.8-2] [L C Van Rijn and Schaafsma, 1986].

Uncertainties also exist for the distance to the bed: the Delft Bottle support was handled with a winched cable from a truck, from the surface to the bed, and we considered the sampler was positioned on the bed when the tension exerted on the cable was relaxed; which is very imprecise and subjective.

It will probably be impossible to have an exact reference for estimating the pump efficiency (this is actually the problem encountered for all sediment sampler validation). However, because the pump allows long sampling with a perfect timing control, and gives direct access to the local concentration, it seems to be a promising unbiased technic for measuring sand transport. In addition because samples are collected in real time at the surface it could represent a real gain of time compared to heavy standard instruments requiring removing the apparatus from the water between each sample. Several questions are still challenging and will motivate new field campaigns as for instance how to measure the exact flow velocity at the nozzle entrance, or how to sample the coarsest grains (captured in the entrance reservoir) during a continuous measurement?

Figure 1 .

 1 Figure 1. The pump (left photo) and its support (middle photo): A) emplacement for the pump B) filter C) settling reservoir and D) intake nozzle with diameters 10/7/6 mm (right photo)

Figure 2 :

 2 Figure 2: The pump mounted on the Delft Bottle (left); the pump mounted on the Delft Bottle support (middle) and sieving at 50 (right)

Figure 3 :

 3 Figure 3: Comparison between concentrations measured on the vertical profile of Run1 with the DB and the pump (left panel); concentration profiles measured for each run (right panel)

REFRENCES

Bendixen, M., J. Best, C. Hackney, and L. Iversen (2019), Time is running out for sand, Nature, 571, 29-31. Beverage, J. P., and D. T. Williams (1989), Comparison : US P-61 and Delft sediment samplers, Journal of Hydraulic Engineering, 115(12), 1702-1706. Davis, B. E. (2005), A Guide to the Proper Selection and Use of Federally Approved Sediment and Water-Quality SamplersRep., 20 pp. Dijkman, J. (1978), Some characteristics of USP-61 and Delft Bottle suspended sediment samplers, 211 pp, Delft University of Technology. Dijkman, J., and V. Milisic (1982), Investigations on suspended sediment samplersRep., Delft hydraulics laboratory and Jarolslav Cerni institute, Delft, Netherland. Gray, J. R., and M. Landers (2014), Measuring suspended sediment., Comprehensive water Quality and Purification, 159-204. Nelson, M. E., and P. G. Benedict (1950), Measurement and analysis of suspended sediment load in streams, ASCE proceedings, USA. Starosolszky, O. (1981), Measurement of River SedimentsRep., World Meteorological Organization. Van Rijn, L., and H. Roberti (2019), Manual Sediment Transport Measurements in Rivers, Estuaries and Coastal Seas . Available from http://www.coastalwiki.org/wiki/Manual_Sediment_Transport_Measurements_in_Rivers,_Estu aries_and_Coastal_Seas [accessed on 10-12-2019] edited. Van Rijn, L. C., and A. S. Schaafsma (1986), Evaluation of Measuring Instruments for Suspended Sediment, paper presented at International Conference Measuring Techniques BHRA, London, England. Van Rijn, L. C., and M. Gaweesh (1992), A New Total Load Sampler, Journal of Hydraulic Engineering, 118(12).


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

Greek and Phoenician sequences from Cyprus are compared across three periods (see table ). This demonstrates an increase in diversity and complexity over time, as reflected by:

• an increase in the length of the sequences, in the numbers of different symbols, the different sequences, and the nodes in the structure.

• a decrease in the centralisation value, meaning that the edges are less concentrated on some particular elements. However, the diversity of the elements composing the sequences (nouns, adjectives, etc.) is virtually unchanging, reflecting stability.

Δηώ Κόρη

Ἐλευσίνιος, α, ον Polytheisms" project to:

+ # # -500/-
• represent sequences and

• describe semantic relationships between the parts of the names of gods (called "elements" in the MAP terminology). The focus is on the structure of the sequences, not their content. These structures are modelled as graphs (a mathematical object composed by a set of nodes and a set of edges), so that they can be visualised, characterised, and compared using quantitative measurements.

  MAP syntax diagrams: an online tool to visualise, describe, and compare Ancient divine onomastic sequences sebastien.plutniak@posteo.net 0000-0002-6674-3806 TRACES, Université de Toulouse, France Summary MAP syntax diagrams is an online R Shiny application for studying Ancient divine onomastic sequences. It uses the format defined by the "Mapping Ancient

•

  Selection of subsets of sequences for comparison • Summary statistics: diversity of the elements (Shannon-Weaver entropy) and structural properties (network metrics) • Measurement of the difference between the subsets graph (Euclidean and "graph diffusion kernel" distances); display of a hierarchical clustering of the distances (for 4 and more subsets) • Generation of diagrams: 1) to graphically summarise a set of sequences and 2) to highlight the differences between two subsets • Export of statistics (in csv format) and diagrams (in svg format) Structural change through time Comparing gods: Apollo & Aphrodite Application main features From sequences to graphs In the MAP format, the sequence Δηώ καὶ Κόρη Ἐλευσινίαι is represented as [ Δηώ + Κόρη ] # Ἐλευσίνιος, α, ον and coded as [ 1 + 2 ] # 3". In MAP syntax diagrams, this sequence is modelled by a graph. The [ and ] symbols enclose a syntagma, + indicates a relation of coordination between two elements, # indicates a qualification.

  • MAP project: hps://map-polytheisms.huma-num.fr • Application: hps://analytics.huma-num.fr/Sebastien.Plutniak/map-syntax-diagrams • Code: hps://github.com/sebastien-plutniak/map-syntax-diagrams • Manual: Plutniak. S. 2020.

	Greek sequences (up to 11 elements) from Cyprus related to Aphrodite (n=92) and Apollo (n=46) are compared. The statistics indicate that Apollo's sequences have more diverse elements (diversity index: 1.81 vs 1.19) and subsequences (median transition rate: 0.11 vs 0.01). However, using the diagrams to study the

MAP Syntax diagrams : visualiser, décrire et comparer les séquences onomastiques divines de l'Antiquité. hps://hal.archives-ouvertes.fr/hal-02532617 differences (coloured in red) we can see that, for example, the complex nested structures ("[[]]") observed in Aphrodite's sequences are absent in Apollo's sequences.

1 hps://base-map-polytheisms.huma-num.fr


Javaid Akhter Bhat

Bernadette Julier
email: bernadette.julier@inrae.fr

Wagdi Ghaleb

Lina Qadir

Abraham J Escobar-Gutiérrez

The History of Domestication and Selection of Lucerne: A New Perspective From the Genetic Diversity for Seed Germination in Response to Temperature and Scarification

Keywords: alfalfa, breeding, genetic variation, Medicago sativa, seed dormancy, adaptation

Lucerne (Medicago sativa), a major perennial pasture legume, belongs to a species complex that includes several subspecies with wild and cultivated populations. Stand establishment may be compromised by poor germination. Seed scarification, deterioration and temperature have an impact on germination. The objective of this study was to analyse the genetic diversity of lucerne germination in response to three factors: (1) temperature, with seven constant temperatures ranging from 5 to 40 • C, was tested on 38 accessions, (2) seed scarification was tested on the same accessions at 5 and 22 • C, (3) seed deterioration was tested on two accessions and two seed lots at the seven temperatures. The germination dynamics of seed lots over time was modelled and three parameters were analysed: germinability (germination capacity), maximum germination rate (maximum% of seeds germinating per time unit), and lag time before the first seed germinates. Seed scarification enhanced germinability at both temperatures and its effect was much higher on falcata and wild sativa accessions. Incomplete loss of the hardseededness trait during domestication and selection is hypothesised, indicating that the introduction of wild material in breeding programmes should be followed by the selection for germinability without scarification. Seed lots with altered germinability had low germination at extreme temperatures, both cold and hot, suggesting that mild temperatures are required to promote germination of damaged seed lots. A large genetic diversity was revealed for germination (both capacity and rate) in response to temperature. All accessions had an optimal germination at 15 or 22 • C and a poor germination at 40 • C. The sativa varieties and landraces had a high germination from 5 to 34 • C while the germination of falcata and the wild sativa accessions were weakened at 5 or 34 • C, respectively. These differences are interpreted in terms of adaptation to the climate of their geographical origin regions in order to

INTRODUCTION

Lucerne (Medicago sativa L.) is a perennial forage legume that holds a major place in cultivated grasslands worldwide. It is used for ruminant feeding, given its high yield and nutritional properties. The numerous ecosystem services it provides [START_REF] Julier | Alfalfa (lucerne) in European cropping systems[END_REF] are important in the present phase of agroecological transition [START_REF] Lamichhane | Abiotic and biotic factors affecting crop seed germination and seedling emergence: a conceptual framework[END_REF], and mainly result from its ability to fix atmospheric nitrogen. M. sativa, which was first cultivated in its centre of origin (Near-East to Middle-East) 9,000 years ago, is the oldest cultivated forage crop and one of the oldest crops in the world. It spread several times to North Africa and Europe through invaders (Medes, Romans and Moors) and was introduced to Asia (India and China) more than 2,000 years ago [START_REF] Basigalup | Perspectives of alfalfa in australia, china, africa and latin america[END_REF]. From the 15th Century, lucerne became a popular crop in France and other European countries (De Serres, 1600) from where it accompanied explorers and migrants to America from the 16th Century onwards and to austral countries by the 19th Century [START_REF] Prosperi | Alfalfa domestication history, genetic diversity and genetic resources[END_REF]. Lucerne is now grown in temperate regions worldwide and has recently became a cornerstone crop in organic agriculture. Its direct annual world value is expected to grow significant over the coming years.

Medicago sativa is a complex of several diploid or autotetraploid perennial and allogamous subspecies (ssp), wild or cultivated, that freely intercross at the same ploidy level [START_REF] Small | A synopsis of the genus Medicago (Leguminosae)[END_REF]. Ssp sativa with coiled, non-spiny pods, and purple or purple yellow flowers, includes the vigorous cultivated autotetraploid varieties, some wild populations in Spain but also the diploid wild form coerulea. The diploid ssp glomerata exhibits coiled pods and yellow flowers. Ssp falcata with yellow flowers and slicked-shaped pods, originates from Northern Eurasia and is cold tolerant. The populations that result from hybridisation between ssp sativa or coerulea and ssp falcata are sometimes described as ssp xvaria [START_REF] Small | A numerical taxonomic analysis of the Medicago littoralis -M. truncatula complex[END_REF]. As shown by their variegated flower colour, most European varieties exhibit introgression from ssp falcata that confers cold tolerance and has enabled the selection of highly productive varieties adapted to continental or subarctic climates.

As many other annual and perennial legumes, in particular of the Papilionoideae subfamily [START_REF] Baskin | Seeds: ecology, biogeography, and evolution of dormancy and germination[END_REF][START_REF] Acosta | Cryo-exposure of Neonotonia wightii Wigth and Am seeds enhances field performance of plants[END_REF], it has been well documented that physical dormancy due to hardseededness is present in lucerne to various degrees [START_REF] Acharya | Cultivar and storage effects on germination and hard seed content of alfalfa[END_REF]. Hardseededness is characterised by a water-impermeable seed coat that ensures a maintenance of the population in the soil and prevents its germination up to several years [START_REF] Baskin | A classification system for seed dormancy[END_REF][START_REF] Finch-Savage | Seed dormancy and the control of germination[END_REF]. Despite the uncontested ecological advantages associated with the different types of seed dormancy [START_REF] Baskin | Seeds: ecology, biogeography, and evolution of dormancy and germination[END_REF], it hampers crop establishment due to low percentage of germinating seeds and slow rates of seedling emergence [START_REF] Acosta | Cryo-exposure of Neonotonia wightii Wigth and Am seeds enhances field performance of plants[END_REF]. This type of physical dormancy is broken by different treatments to the seeds, including scarification, which makes imbibition possible by overcoming coat's impermeability to water [START_REF] Baskin | Seeds: ecology, biogeography, and evolution of dormancy and germination[END_REF]. Even in cultivated varieties, the presence of hard seeds is common in commercial lots, although regulations and the market impose a minimum percentage of such seeds.

Seed deterioration, often generated by ageing, induces a reduction of germination capacity, or germinability, [START_REF] Mcdonald | Seed deterioration: physiology, repair and assessment[END_REF]. Evaluation of lot germinability along seed ageing is performed by the seed industry [START_REF] Corbineau | Markers of seed quality: from present to future[END_REF] and the institutions in charge of ex situ seed conservation (Genetic Resources Centres;[START_REF] Fu | Towards a better monitoring of seed ageing under ex situ seed conservation[END_REF]. Seed deterioration is regulated by seed storage conditions (temperature, humidity and oxygen partial pressure) but also by endogenous factors [START_REF] Zhou | A matter of life and death: molecular, physiological, and environmental regulation of seed longevity[END_REF]. Little information is available on the impact of temperature on germination of seed lots whose germination capacity is deteriorated.

Temperature is the most important environmental factor regulating both timing and rates of seed germination. The role of temperature in seed germination is related to enzyme activity and membrane permeability, which impact respiratory metabolism, and thus the rate of germination [START_REF] Bewley | Seeds physiology of development and germination[END_REF]. Plant species have different critical temperature ranges and basic thermal requirement to complete specific phenophases or the whole life cycle [START_REF] Luo | Temperature thresholds and crop production: a review[END_REF]. Seed germination in response to temperature can be summarised by the cardinal temperatures (minimum, optimum and maximum temperatures; [START_REF] Bewley | Seeds physiology of development and germination[END_REF][START_REF] Shafii | Estimation of cardinal temperatures in germination data analysis[END_REF][START_REF] Durr | Ranges of critical temperature and water potential values for the germination of species worldwide: contribution to a seed trait database[END_REF]. The response of different species to temperature during germination has been a topical issue for a long time [START_REF] Bewley | Seeds physiology of development and germination[END_REF][START_REF] Baskin | Seeds: ecology, biogeography, and evolution of dormancy and germination[END_REF][START_REF] Durr | Ranges of critical temperature and water potential values for the germination of species worldwide: contribution to a seed trait database[END_REF][START_REF] Tribouillois | Determination of germination response to temperature and water potential for a wide range of cover crop species and related functional groups[END_REF]. Major differences among species depend on the climatic conditions where the species grow or originate. Cultivated species germinate faster and under a wider range of temperatures than wild species, likely as a result of human selection [START_REF] Durr | Ranges of critical temperature and water potential values for the germination of species worldwide: contribution to a seed trait database[END_REF]. Information on genetic variation for germination capacity is useful in breeding programmes especially if wild accessions are used to adapt varieties to new climatic conditions or practices related to ecosystem services, or for the preservation of genetic diversity in genetic resources centres. Studies at the intra-specific level are not frequent in pasture species [START_REF] Moot | Base temperature and thermal time requirements for germination and emergence of temperate pasture species[END_REF][START_REF] Mcgraw | Effects of temperature on germination of 10 native legume species[END_REF][START_REF] Monks | Cardinal temperatures and thermal time requirements for germination of annual and perennial temperate pasture species[END_REF][START_REF] Ahmed | Analyse de la variabilité inter-et intra-spécifique de cinq espèces prairiales en réponse à la température pendant la germination et la phase hétérotrophe initiale[END_REF][START_REF] Ahmed | Genetic diversity of alfalfa (Medicago sativa) in response to temperature during germination[END_REF]. In the model legume species Medicago truncatula, a significant diversity for germination was evidenced among accessions [START_REF] Brunel | Using a model-based framework for analysing genetic diversity during germination and heterotrophic growth of Medicago truncatula[END_REF] and within a mapping population [START_REF] Dias | QTL analysis of seed germination and pre-emergence growth at extreme temperatures in Medicago truncatula[END_REF]. Knowledge of physical dormancy, seed deterioration and genetic diversity in response to temperature, is of paramount importance in the management of lucerne breeding programmes.

In this paper, the objective was to demonstrate a genetic diversity of Medicago sativa for seed germination in response to constant temperatures (from 5 to 40 • C). We performed experiments in controlled conditions with thirty-eight accessions of various geographical origins, from the subspecies sativa, falcata or glomerata, wild or cultivated. The effects of scarification and deterioration level of seed lots on germination at different temperatures were also tested. All the results were analysed and interpreted in terms of domestication, selection and breeding.

MATERIALS AND METHODS

Plant Material

In this study, seeds of thirty-eight accessions of wild populations, landraces and varieties of lucerne were evaluated as follows: 16 sativa varieties, 11 sativa landraces, four sativa wild accessions, one falcata variety, five falcata wild populations and one glomerata wild population (Table 1). In addition to the spp. membership, the status (wild, landrace or variety) that traces the breeding effort possibly applied to germination traits was used to define five groups of accessions: 'falcata + glomerata wild' , 'sativa wild' , 'falcata variety' , 'sativa landraces' and 'sativa varieties'. Seeds were obtained from INRAE Genetic Resources Centre in Lusignan, France, or from breeders. They were conserved at 5 • C and 30% relative humidity until they were used. For two landraces, Flamande from France and Gabès from Tunisia, deteriorated seed lots, characterised by poor germinability, were also included. Only two wild falcata, the falcata variety and the wild glomerata were diploid accessions and all the others were tetraploid accessions, so the effect of ploidy was not studied here.

The material was used to test three aspects of seed germination. (1) The effect of scarification, in interaction with genetic diversity, was tested on the whole set of 38 accessions at two temperatures (5 and 22 • C). Scarification consisted in scrubbing the seeds between two sheets of sandpaper (grade 150) for 30 s. (2) The effect of the deterioration level of seed lots on germination response to seven constant temperatures [START_REF] Monks | Cardinal temperatures and thermal time requirements for germination of annual and perennial temperate pasture species[END_REF]10,15,22,28,34 or 40 • C) was analysed on two accessions by comparing, within each accession, seed lots with high or low germinability.

(3) The genetic diversity for germination in response to these seven constant temperatures was evaluated on scarified seeds of the 38 accessions.

Germination Tests

For each accession and treatment, four repetitions of 100 seeds were prepared. For temperatures between 5 and 34 • C, the 100 seeds of each repetition were put in 90 mm diameter Petri dishes containing two sheets of autoclave-sterilised Whatman paper (ref.

3645 Whatman, France) humidified with 5 ml of deionised and autoclave-sterilised water. For the experiment at 40 • C, which was conducted in a growth chamber with 80% relative humidity, seeds were placed on two sheets of Whatman papers in transparent crystal polystyrene boxes (180 mm × 120 mm × 50 mm; GEVES trademark, Loire Plastic, France; [START_REF] Brunel | Using a model-based framework for analysing genetic diversity during germination and heterotrophic growth of Medicago truncatula[END_REF] with 15 ml of sterilised water.

For each temperature, the design was a random complete block design (RCBD) with four blocks. The dishes of each block were placed in a vented plastic box. The experiment was conducted in walk-in growth chambers at constant temperature in the dark and with vapour pressure deficit <1 kPa. Temperature of the chambers were checked by six thermocouples placed at different positions within the useful volume, logged every 20 s. Temperature and relative humidity in the chambers were recorded every minute during the germination experiment.

Germination counting was carried out at variable time intervals that depended on temperature treatments: once a day at 5 and 40 • C and twice a day for temperatures from 10 to 34 • C. Seeds were considered as germinated when the radicle or the cotyledon leaves had protruded out of the seed and was at least 2 mm long [START_REF] Bewley | Seeds physiology of development and germination[END_REF]. After each counting, if needed, deionised and autoclave-sterilised water was added to ensure non-limiting moisture. Counting was stopped when no new germination was observed during five consecutive visits. At 5 • C, the experiment lasted more than 3 months. Whenever needed, counting was performed in the growth chamber in order to keep the seeds at a constant temperature and a high air humidity.

Modelling of the Dynamics of Seed Lot Germination

For both the effect of seed deterioration and the genetic diversity, the cumulative percentages of germination values over time were fitted to a modified non-rectangular hyperbole [START_REF] Thornley | Mathematical models in plant physiology[END_REF], whose parameters have an eco-physiological meaning:

y = 1 2θ • α • (t -t c ) + y max - α. (t -t c ) + y max 2 -4.θ.α. (t -t c ) .y max .
Where y is the cumulated seed germination (%); θ is a unitless parameter that determines the sharpness of the knee of the curve; α is the maximum germination rate (% of germinating seeds per hour); t is the time (hour); t c is the lag to start germination (hour) during which no seed germinates; and y max is the asymptotic maximum seed germination (%). In order to limit the number of parameters to be estimated, and after several tests, θ was fixed to 0.97.

Statistical Analyses

For the effect of scarification in each of the two temperatures, the observed maximum seed germination (hereafter named as germinability) was submitted to an analysis of variance with the effects of scarification, group, accession within group, repetition, scarification × group and scarification × accession within group. The function aov of R was used (R Core Team, 2019). The subspecies, population type (variety, landrace or wild population), name of accession and country of origin are given. For the wild populations, the geographic coordinates of the collection site are indicated. Autumn dormancy is given when known. *Two seed lots of these accessions were tested.

For the germinability of the two seed lots with high and low germination percentages, a non-linear mixed model [START_REF] Pinheiro | Mixed-effects models in S and S-PLUS[END_REF] was used to estimate the three parameters (α, t c and y max ) and to test the seed lot factor, in Flamande and Gabès separately, at each temperature. The functions nlsList, nlme and update of package nlme were sequentially used [START_REF] Pinheiro | nlme: Linear and nonlinear mixed effects model[END_REF] in R.

For the 38 accessions, the non-linear mixed model was similarly used to test the effects of group and accession. A principal component analysis (PCA) was conducted to assess the variability in the three germination parameters of the model in response to temperature (function PCA of R).

RESULTS

Scarification Effect on the Percentage of Seed Germination

On average for the 38 accessions, scarification had a significant positive effect on germinability at both temperatures 5 and 22 (Table 2, Figure 1, and Supplementary Table 1). Germinability of non-scarified seeds varied from 6 to 94% at 5 An analysis of variance showed that the effects of group, accession within group, scarification, and their interactions were significant at both temperatures (Table 2 and Figure 1). The scarification effect was positive for the wild accessions, either for falcata + glomerata (+34 points of percentage) or sativa (+30 points of percentage) at 22 • C. At 5 • C, the figures for the same groups were +23 and +35 points of percentage. For the falcata variety, the effect of scarification was large at 22 • C (+48 points of percentage) but nil at 5 • C, where the germination was very poor without scarification but also with scarification. For the sativa landraces and varieties, scarification had a small but significant effect at both temperatures (+7 and +6 points of percentage at 5 • C and 22 • C, respectively).

Effect of Seed Lot Deterioration on the Germination Capacity in Response to Temperature

The seed lots confirmed their expected germinability, either high or low, at all temperatures (Table 3 andFigure 2). The two seed lots of the population Flamande had higher lags to start germination (t c ) and lower germination rates (α) than the two seed lots of the population Gabès. The low germinability of deteriorated seed lots were associated with low germination rates (α; Table 3). At 34 • C, the germinability of the deteriorated seed lot, compared to the highly germinating seed lot, was much lower for Flamande than for Gabès.

Genetic Diversity for Germination in Response to Temperature

A wide diversity for the three parameters of temperature response (germinability, maximum germination rate and lag to start germination) was evidenced among the 38 lucerne accessions (Supplementary Table 2 and Supplementary Figure 1, Figure 3). All parameters at all temperatures shown a significance among accessions (not shown). The effect of group of accessions were also significant except for the maximum germination rates at 5 and 28 • C (Table 4).

In the PCA analysis performed with the values of germinability (y max ) and the estimated values of maximum germination rate (α) and lag to start germination (t c ) estimated at seven temperatures [START_REF] Monks | Cardinal temperatures and thermal time requirements for germination of annual and perennial temperate pasture species[END_REF]10,15,22,28,34,40 • C; Figure 3), the first two axes were significant (eigenvalue > 1) and contributed for 75.5% of total variance (Supplementary Table 3 and Figure 3A). Axis 1 explained 65.7% of the total variability and was mainly correlated to germination rates at 10, 5 and 15 • C and germinability at 10 and 34 • C (Figures 3A,B and Supplementary Table 2). Axis 2, which explained 9.8% of the total variability, was mainly correlated to the germination rates at 40 and 34 • C (Supplementary Table 3). Germinability at all temperatures was positively correlated with maximum germination rate at all temperatures (Figure 3A). Lag to start germination from 10 to 34 • C was negatively correlated with both germinability and maximum germination rate (Figure 3A). As a whole, the accessions that had high germinability germinated shortly and quickly after seed imbibition.

All germination parameters were optimum at 15 or 22 • C for all groups of accessions (Table 4 and Supplementary Figure 1). With progressing colder temperatures, the lag to start germination increased and the germination rate decreased for all groups of accessions, while germinability decreased for the falcata and glomerata accessions only. With progressing higher temperatures, the germinability of the cultivated sativa accessions remained high up to 34 • C, while their lag to start germination increased and their germination rate decreased. At 40 • C, all germination parameters of the cultivated sativa accessions were altered. For the wild accessions and the falcata variety, germinability was still high at 28 • C, but not at 34 • C nor at 40 • C, and lag to start germination and germination rate were strongly modified. Figure 3B demonstrates that the groups behaved in a different way in the PCA. The accessions of both ssp sativa landrace and ssp sativa variety groups showed a high germinability as well as a high germination rate and a small lag to start germination. This means that these cultivated accessions are able to germinate shortly after the conditions are adequate and that all the seeds germinate at once. Moreover, it is to be noted that, as shown from their position on axis 2 of the PCA, the ssp sativa accessions with an autumn dormancy between 3 and 7 had a lower germination rate at 34 • C than the non-dormant accessions with dormancy 9 which originate from Tunisia, Morocco or Iran (Figure 3B). Compared to the cultivated sativa accessions, accessions of the wild falcata and glomerata group showed a low germinability and very low germination rate as well as a high lag to start germination. The falcata variety (Anik, no 34) also showed a low germinability, a low germination rate and a long lag to start germination. However, the ssp falcata accession Krasnokutskaya (no 38) from Russia showed a higher germinability, a higher germination rate and a lower lag to start germination than the other accessions of the ssp falcata wild group (Figure 3B). The wild sativa accessions that originated from Spain had a higher germinability and a higher germination rate than the wild falcata accessions. Interestingly, the wild accessions that originated from Spain had only slightly lower germination rate and lag to start germination than cultivated sativa accessions from 5 to 22 • C, but all their germination parameters were strongly altered at 34 • C, with a low germinability, a long lag to start germination and a slow germination rate (Table 3). At 5 • C, the germinability of the sativa accessions, whether wild or cultivated, were higher than that of the falcata and glomerata accessions.

DISCUSSION

This study aims at demonstrating that lucerne seed germination is influenced by physical treatment (scarification), seed lot deterioration and genetic diversity in response to temperature. It also illustrates the interaction between these factors and gives insights about the history of domestication and selection.

Scarification Effect on the Percentage of Seed Germination

Physical seed dormancy caused by the water-impermeable seed coat that inhibits water absorption is well described in many species (Baskin andBaskin, 2004, 2014), including legumes [START_REF] Taylor | Hardseededness in mediterranean annual pasture legumes in australia: a review[END_REF][START_REF] Smykal | The role of the testa during development and in establishment of dormancy of the legume seed[END_REF]. Scarification, by a physical or chemical alteration of the seed coat, increases seed germination. Our results on 38 accessions of lucerne indicate that, on average, non-scarified seeds of lucerne had a lower germinability than scarified seeds at both temperatures (5 and 22 • C). This scarification effect on germination response depended on accessions. It was higher for the wild accessions, either falcata or sativa ssp. than for the cultivated accessions. A large effect of mechanical scarification had previously been reported on one naturalised American yellow-flowered population [START_REF] Narem | Effects of stratification, warm treatment, and mechanical and acid scarification on the emergence of yellow-flowered alfalfa (Medicago sativa subsp. falcata) seeds[END_REF]. This could mean that the domestication syndrome which includes germination, well described on legumes [START_REF] Smykal | The role of the testa during development and in establishment of dormancy of the legume seed[END_REF], also applies to lucerne. Our results suggest that the seed dormancy mechanisms carried by wild accessions have been lost during domestication and breeding. This loss enables the seeds of cultivated accessions to germinate as soon as they are For each accession, the significance of the difference between seed lots is given. ***, **, *: significant at P < 0.001, P < 0.01, P < 0.05, respectively. NS: non-significant (P > 0.05). Repartition of the accessions on the first two axes. The codes of the varieties are given in Table 1. *y max for germinability, t c for the lag time to start germination, a for the rate of germination. y max , t c and a are followed by the experimental temperature. The F value is for the test of difference among groups. ***significant at P < 0.001, NS: non-significant at P = 0.05.

sown, even if severe stress conditions (frost, drought) could occur shortly afterwards. It is thus the responsibility of the farmer to choose an adequate sowing season to ensure good establishment and favourable growing conditions. In the wild, germination is submitted to the random occurrence of seed coat wound, thereby spreading germination over a long time period after the seeds reach the soil. As other domestication traits whose inheritance is usually simple, a small number of genes or a single gene probably govern the susceptibility of lucerne seed germination to scarification [START_REF] Smykal | The role of the testa during development and in establishment of dormancy of the legume seed[END_REF]. This domestication trait comes in addition to the domestication syndrome already described in lucerne: plant vigour caused by polyploidy and erect growth habit to increase forage production and facilitate harvesting, and coiled-shape pods to limit seed shattering [START_REF] Annicchiarico | Achievements and challenges in improving temperate perennial forage legumes[END_REF]. The early farmers have probably selected plants that do not require scarification, by sampling the first emerged seedlings after sowing, and modern breeders proceed similarly. However, in our study as in others [START_REF] Acharya | Cultivar and storage effects on germination and hard seed content of alfalfa[END_REF][START_REF] Palfi | Alfalfa (Medicago sativa L.) seed scarification by acid [HR][END_REF][START_REF] Kandil | Dormancy overcoming of some alfalfa varieties[END_REF], the germination of cultivated varieties was enhanced by scarification, which means that the loss of the hardseededness trait is not yet completed. Either the genetics of lucerne (autotetraploidy and allogamy) make the removal of this trait from the breeding populations difficult, or the hardseededness is under a quantitative genetic control that implies many genes [START_REF] Finkelstein | Molecular aspects of seed dormancy[END_REF]. The germination of the single falcata variety studied here was strongly enhanced by scarification. This variety has been bred for adaptation to Northern Canada from a wild Russian falcata population [START_REF] Pankiw | Anik alfalfa[END_REF] but germination without scarification has probably not been taken into account. In all situations, either with elite breeding pools or in breeding programs that include wild material, further selection devoted to this trait should improve germination and thus lucerne establishment.

Effect of Seed Deterioration

For both landraces, Flamande and Gabès, the low germination seed lot germinated less, with lower germination rate and longer lag to start germination than the high germination seed lot, at all temperatures. The differences between the two seed lots were higher at extreme temperatures than at average temperatures (15-22 • C). This effect of temperature on the germination of deteriorated seed lots has little been studied. On pea (Pisum sativum) and bean (Phaseolus vulgaris), for example, all the seed lots of all genotypes had a high germinability, even at extreme temperatures, except for two bean seed lots at 10 • C [START_REF] Raveneau | Pea and bean germination and seedling responses to temperature and water potential[END_REF]. For pea seeds produced under accelerated desiccation conditions, germination at a cold temperature (5 • C) was only reduced by 10% compared to a control seed lot [START_REF] Raveneau | Effet des vitesses de dessiccation de la graine et des basses températures sur la germination du pois protéagineux[END_REF]. The results obtained here suggest that the seeds with altered germinability should be sown in optimal temperature conditions, either for culture at the farm level or for multiplication in Genetic Resources Centres.

Genetic Diversity

The impact of temperature on germination is well established and cardinal temperatures have been determined on many species, such as weeds [START_REF] Long | The ecophysiology of seed persistence: a mechanistic view of the journey to germination or demise[END_REF], crops [START_REF] Durr | Ranges of critical temperature and water potential values for the germination of species worldwide: contribution to a seed trait database[END_REF] and cover crops [START_REF] Tribouillois | Determination of germination response to temperature and water potential for a wide range of cover crop species and related functional groups[END_REF]. All accessions evaluated in our work exhibited the highest germinability and rate, as well as the shortest lag to start germination at 15 • C or 22 • C. The optimal constant temperature for lucerne germination is thus within this range. As discussed below, the germination was not as good at colder or warmer temperatures but the effect of extreme temperatures depended on the accessions.

The within-species diversity for germination in response to temperature has received little attention either in the number of accessions or in the ranges of temperatures that have been tested. In our study, we analysed 38 accessions from the whole M. sativa complex, with diploid and tetraploid accessions of wild and cultivated sub-species. This range of diversity is interesting because subspecies freely intercross at a ploidy level. Wild populations that are under-represented in breeding pools [START_REF] Muller | Domestication history in the Medicago sativa species complex: inferences from nuclear sequence polymorphism[END_REF] could be used to integrate some traits for adaptation to new constraints, such as climate change, or new practices, such as cover crops [START_REF] Labreuche | Impacts de couverts permanents sur le blé tendre d'hiver[END_REF][START_REF] Moore | Regenerating agricultural landscapes with perennial groundcover for intensive crop production[END_REF]. These accessions were studied in seven temperatures ranging from 5 • C, a temperature which mimics cold autumns or springs, to a temperature of 40 • C, which could occur with late spring sowings under warm climates. In our study, germinability was very low, on average 12% only, at 40 • C. In contrast, germinability at 40 • C ranged from 60 to 80% in a preliminary study on a subset of seven accessions among the ones studied here [START_REF] Ahmed | Genetic diversity of alfalfa (Medicago sativa) in response to temperature during germination[END_REF]. We suppose that the high germinability in the latter study originates both from the temperature during counting (counting in the laboratory at 20 • C) and/or from the natural light received by the seeds during counting (L. Ahmed, personal observation).

Within cultivated sativa accessions, germinability varied little at temperatures from 5 to 34 • C. Breeding for stable germination is probably conducted, even unconsciously, when the first emerged plantlets are selected to constitute the pool of plants to be studied or crossed. In the field too, the first emerged plants gain a competitive advantage on the later emerged ones because they capture resources such as light. Indeed, there is no difference between landraces and varieties, meaning that this selection has been operating for a long time. Other studies have already established this low range of genetic diversity among varieties, but with experiments most frequently conducted under temperatures between 10 and 30 • C only [START_REF] Mcelgunn | Germination response of forage legumes to constant and alternating temperatures[END_REF][START_REF] Hampton | Temperature effects on the germination of herbage legumes in New Zealand[END_REF][START_REF] Brar | Germination of 20 forage legumes as influenced by temperature[END_REF][START_REF] Butler | Temperature affects the germination of forage legume seeds[END_REF]. Anyway, the non-dormant cultivated accessions (dormancy 9, from Tunisia, Morocco and Iran) had higher germination rates at 34 • C than the more dormant accessions (dormancy between 3 and 7). Non-dormant accessions are bred and used under warm climates in which hot periods may occur in the sowing time, either autumn or spring. This inability to germinate at high temperature has probably been selected as a global adaptation to local conditions.

The wild accessions, on average, had a lower germinability, a lower germination rate and a longer lag to start germination than cultivated accessions. Moreover, wild accessions adapted to cold conditions (falcata ssp) had a low germinability at 5 • C and wild accessions from Spain (sativa ssp) had a low germination rate at 34 • C but a high germination rate at 5 • C. This counterintuitive result can easily be explained: if the falcata accessions germinated at 5 • C (in the autumn), they would have a large risk of encountering cold conditions in the following weeks. It is an adaptation for populations that their seeds germinate when temperatures reach at least 10 • C. Reversely, if wild accessions from Spain germinated at 34 • C (in the spring), they would suffer from associated dry conditions. In the case where they germinate at 5 • C, this cold temperature is statistically not followed by colder temperatures, so establishment is preserved. These adaptations are ways to escape harsh (either cold or dry) conditions in the wild. In our study, the falcata variety had germination characteristics close to those of wild falcata. Its breeding, from a wild falcata accession [START_REF] Pankiw | Anik alfalfa[END_REF], probably did not include a selection for germination at low temperatures, as seen above for scarification. Krasnokutskaya, from Russia, received as a wild falcata accession by the Genetic Resources Centre of INRAE, Lusignan, France, showed germination characteristics close to those of cultivated sativa accessions, but its lag to start germination at 34 • C was longer. This germination ability suggests that Krasnokutskaya is probably a cultivated accession selected for improved establishment. Another study [START_REF] Julier | Genetic variability for morphology, growth and forage yield among perennial diploid and tetraploid lucerne populations (Medicago sativa L)[END_REF] revealed that this accession exhibited a relative erect growth habit, long stems and large seeds, which are relevant breeding traits.

This study has been conducted with seed lots that have been stored for different lengths of time in cold chambers. With this allogamous insect-pollinated species, the seed production of many accessions in the same location and year is challenging. The storage duration may impact seed germination response to temperature but we suppose that this impact is smaller than the genetic difference among accessions. Indeed, these results show that natural selection for germination in response to temperature has shaped species diversity. The mean value of this trait, which can be considered as an adaptative trait, has changed during the evolution of the Medicago sativa complex in several subspecies adapted to different regions and climates. In addition, human selection has been efficient to alter the effect of temperature on germination, as also demonstrated in experimental work [START_REF] Klos | Response of six alfalfa populations to selection under laboratory conditions for germination and seedling vigor at low temperature[END_REF]. In breeding programmes, a good germination in a large range of temperatures is required. When the plant material originates from cultivated pools, no more selection is needed. When wild accessions are introduced in a breeding pool, attention must be paid to reach a high germination, especially at extreme temperatures.

  FIGURE 1 | Germinability (observed data, average of 4 biological repetitions) of 38 lucerne populations from 5 different groups with or without scarification at 5 • C (A) and 22 • C (B).

  5 and 22 • C, for two other accessions at 5 • C only and for three other accessions at 22 • C only. This difference reached a maximum of 46 points of percentage at 5 • C (Villanueva de Jara, sativa wild) and 50 points of percentage at 22 • C (Anik, falcata variety).

FIGURE 2 |

 2 FIGURE 2 | Boxplot showing average germinability (y max parameter of the model) of two seed lots for two accessions (Flamande and Gabès) in response to temperature.

TABLE 1 |

 1 Description of the 38 accessions of Medicago sativa.

	Geographic coordinates

TABLE 2 |

 2 Mean observed germinability over 4 biological repetitions, with or without scarification at 5 and 22 • C for the five groups of accessions.

	Group

TABLE 3 |

 3 Germinability (y max in • C), lag to start germination (t c in h) and maximum germination rate (α in% of germinating seeds per h) for the two seed lots (high and low germination) of two lucerne accessions (Flamande and Gabès) at 7 temperatures.

	Temperature ( • C)	Parameter	Flamande high germ.	Flamande low germ.	Gabès high germ.	Gabès low germ.
	5	α	0.793	0.330 NS	0.806	0.609 NS
		t c	81.5	90.2**	79.6	83.6 NS
		y max	91.2	74.3***	85.6	72.2***
	10	α	1.485	0.687*	1.506	0.988**
		t c	28.4	29.2 NS	27.7	26.8 NS
		y max	95.5	72.9***	91.1	68.8***
	15	α	3.613	1.516***	4.672	2.188***
		t c	16.0	14.5 NS	13.1	11.1*
		y max	100.3	69.3***	89.6	73.3***
	22	α	6.246	2.042**	7.560	4.913*
		t c	12.3	12.3 NS	12.3	11.5*
		y max	94.5	76.2***	85.9	67.6***
	28	α	3.591	1.532 NS	7.506	3.035***
		t c	14.0	14.3 NS	12.0	11.0***
		y max	96.7	65.2***	84.1	61.5***
	34	α	1.886	0.616 NS	7.421	3.083***
		t c	16.6	16.7 NS	13.3	11.8***
		y max	90.5	33.0***	83.1	66.2***
	40	α	0.067	0.090 NS	0.120	0.052***
		t c	51.6	20.1**	4.0	10.6 NS
		y max	11.4	3.7*	17.5	5.7**

TABLE 4 |

 4 Germinability

	Sativa variety	F Value

(y max in%), lag to start germination (t c in h) and maximum germination rate (α in% of germinating seeds per h) for the 5 groups of lucerne accessions at 7 temperatures.

Temperature ( • C)

Parameter Falcata + glomerata wild Sativa wild Falcata variety Sativa landrace

Frontiers in Plant Science | www.frontiersin.org

January 2021 | Volume 11 | Article 578121

This study was carried out on two landrace accessions, with two seed lots per accession, one with a high germinability and the other with a poor germinability.Frontiers in Plant Science | www.frontiersin.org

ACKNOWLEDGMENTS

We thank the seed companies and institutes that provided us with seeds of their accessions: AgriObtentions, Alforex, ART, Barenbrug, CGS Sementi, CREA, DLF Seeds, Forage Genetics, Foratec Argentina, IFVCNS, Lantmännen, RAGT-2n. Cédric Perrot is thanked for the maintenance of growth chambers.

FUNDING

This project has received funding from the European Union's Horizon 2020 Programme for Research & Innovation under grant agreement no. 727312 (project EUCLEG).

DATA AVAILABILITY STATEMENT

The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found below: https://doi.org/10. 15454/YIECZS.

AUTHOR CONTRIBUTIONS

AE-G and BJ conceived the study. AE-G, LA, and BJ designed the experiment. LA collected the data with contributions of BJ, AE-G, and WG. BJ, WG, and LA carried out the data analysis. All co-authors contributed to wrote the manuscript. All authors contributed to the article and approved the submitted version.

SUPPLEMENTARY MATERIAL

The Supplementary Material for this article can be found online at: https://www.frontiersin.org/articles/10.3389/fpls.2020. 578121/full#supplementary-material

Conflict of Interest:

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.