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    Early-season plant-to-plant spatial uniformity can affect soybean yields

    Sites description and field operationsA total of six field studies were conducted in two different regions over two seasons. Four studies (two dryland and two irrigated) were in Kansas, United States (dryland: 39°4′30″ N, − 96°44′43″ W, irrigated: 39°4′25″N, − 96°43′12″ W) during the 2019 and 2020 growing seasons (hereafter referred to as USDry19, USIrr19, USDry20, and USIrr20 studies). The remaining two studies (dryland) were in Entre Rios, Argentina (31°50′49″ S; 60°32′16″ W) during the 2018/2019 and 2019/2020 growing seasons (hereafter referred to as Arg19 and Arg20 studies). The soils were Fluventic Hapludolls [silt loam, 40% sand, 13% clay, 47% silt, organic matter (OM) 1.7%, 7.7 pH, 31.1 ppm P (Bray−1)] at the US dryland studies, and Pachic Argiudolls [silty clay loam, 10.1% sand, 30.6% clay and 59.3% silt, OM 3.2%, 6.8 pH, 34.7 ppm P (Bray−1)] at the US irrigated studies. At the Argentinian studies soil was a Vertic Argiudoll in 2019 [silty clay loam to clay loam, 3.9% sand, 27.6% clay, 67.9% silt, OM 2.65%, 7.2 pH, 12.5 ppm P (Bray−1)] and an Acuic Argiudoll in 2020 [silt loam to silty-clay-loam, 5.6% sand, 28.6% clay, 65.8% silt, OM 3.33%].The US dryland and irrigated studies were sown on June 4, 2019, and May 20, 2020. In 2019, the dryland study was replanted on June 29 due to poor emergence after the first sowing. The studies in Argentina were sown on December 5 in 2018 and November 20 in 2019. At all six studies, plots were kept free of weeds, pests, and diseases through recommended chemical control.The genotypes used in the US were P40A47X (MG 4.0) and P39A58X (MG 3.9) (Corteva Agriscience, Johnston, IA, USA) in 2019 and 2020, respectively. Both varieties are tolerant to glyphosate and dicamba herbicides (RR2X) and have low lodging probability. For the northeast region of Kansas, recommended sowing dates range from May 15 to June 15 along with MG 421. In addition, recommended seeding rates are between 270 and 355 thousand seeds ha−1 for low-yielding environments and 190 to 285 thousand seeds ha−1 for medium- and high-yielding environments13. In Argentina, the genotype AW5815IPRO (MG 5.8, Bayer, Leverkusen, Germany) was used both in 2020 and 2021, it is tolerant to glyphosate and sulfonylureas, and has low lodging probability. Recommended sowing dates for Entre Rios considering soybeans as a single crop range from October 20 to December 10, and MG usually range from 4 to 6; lastly, seeding rate recommendations are between 200 and 250 thousand seeds ha−1 in the region22.Study designThe studies carried out in the US were arranged as a split plot design with three replicates in both 2019 and 2020. In 2019, the main plot treatment factor was planter type with two levels [John Deere (Moline, Illinois, US) Max Emerge planter (ME, 12 rows), and John Deere Exact Emerge Planter (EE, 16 rows)], and the split-plot treatment factor was seeding rate with two levels (160 and 321 thousand seeds ha−1). In 2020 the main plot treatment factor was also planter type with two levels (ME and EE), and the split-plot treatment factor was seeding rate with four levels (160, 215, 270 and 321 thousand seeds ha−1). Planting speed was 7 km h−1 in both studies and years, plots were 24 and 32 rows wide when planted with ME and EE, respectively, with 0.76 m row spacing. Plot length was 80 m in the dryland studies and 160 m in the irrigated studies. The studies in Argentina were arranged as a single replicate of each seeding rate (100, 230, 360 and 550 thousand seeds ha−1) in both years. Planting speed was 5.5 km h−1 in both years, and plots were 10 rows wide with 0.52 m row spacing and 350 m in length.All treatment factors in US studies were evaluated with the overall goal of producing substantial variation in the variable of interest, plant-to-plant spatial uniformity, rather than to make an inference of their effect on yield. The Argentinian studies were only used for selection of stand uniformity variables due to the single replicate. Plant spatial uniformity variables were first fitted using the data from US studies (details below), and then the best explanatory metrics were selected to re-fit the relationships combining both data sets from US and Argentina. Finally, sowing dates, maturity groups, and seeding rates evaluated in this study at both locations (Arg and US) were aligned with those recommended for each region.Data collection and spacing uniformity variablesTwo segments of 2 m in length were established early in the season inside each plot. At the V5 (US studies) and R1 (Arg studies) soybean development stage23, the cumulative distance of the plants within each segment was measured and then used to calculate multiple derived variables. Plant spacing (cm) was calculated as the average distance between neighboring plants. In addition, the distance from a plant to each neighboring plant was classified as shorter or longer than the plant spacing (named nearest and farthest neighbor distance, respectively). Achieved versus Target Evenness Index (ATEI, dimensionless) was calculated as the ratio between the observed plant spacing and the theoretical plant spacing (TPS, cm), where TPS is the expected plant spacing derived from a specific seeding rate and row width (Eq. 1).$$ATEI = frac{Spacing;(cm) }{{TPS;(cm)}}$$
    (1)
    The ATEI index was designed to account for the proximity of the observed plant spacing to the TPS. Values closer to 1 indicate that the plant spacing is close to the TPS and values that are below or above 1 indicate that the plant spacing is lower or higher than the TPS, respectively; thereby departing from an ideal plant spacing. Hence, ATEI values greater than 1 depict both (i) non-uniform plant-to-plant spacing distribution and (ii) plant densities below the target (seeding rate). To further understand the meaning of ATEI, the relative density (rd) was calculated as the ratio between plant density (based on the number of plants in the 2 m segment) and seeding rate.To account for the unevenness of distance from a plant to both neighboring plants within the row, we used the Evenness Index (EI, dimensionless), calculated as the ratio between the distance to the nearest neighbor (cm) and the plant spacing (cm) of a given plant (Eq. 2). The Evenness Index values range from 0 to 1, a value closer to 1 indicates that a plant is equidistantly spaced to both of its neighboring plants within the row, if zero then those plants are occupying the same position (as doubles). It is important to note that EI does not provide information on the spacing (in distance, cm) or how close the spacing is compared to the TPS, but only describes the unevenness distance of a plant to its neighboring plants within a row.$$Evenness ;Index; (EI) = frac{nearest; neighbor ;(cm)}{{Spacing; (cm)}}$$
    (2)
    In addition, the distance from a plant to its preceding neighboring plant, and the TPS were used to classify the position of each plant into one of eight classes (Fig. 1). Plants were classified in classes ranging from “double” (preceding plant distance  Double-skip) as a function of seeding rate, planter type and their interaction (fixed effects), and block nested in site-year (random effect) (Tables 1 and 2). Independent models for each of the 4 US studies were built assessing the effects of planter type, seeding rate, and their interaction (fixed effects), and seeding rate nested in planter type, and in block (random effects) on the same variables previously mentioned (Supplementary Table 1). The models were run using the lmer function from lme4 package in R (R Core Team, 2021). In addition, the US and Arg studies were combined to evaluate the effect of site-year on yield, plant density, and all stand uniformity variables (Supplementary Fig. 1) using the lm function from package stats. Means separation were performed using Fisher’s LSD (Least Significance Difference) test (alpha = 0.05) with emmeans function from package emmeans.Table 1 Effect of planter type, seeding rate, and their interaction on variables from plant position classification for all US studies. References: percentage of perfectly spaced plants (Perfect), percentage of plants misplaced by 66% (Mis 66), percentage of plants misplaced by 33% (Mis 33), percentage of double plants (Double), percentage of short skips plants (Short-skip), percentage of long skip plants (Long-skip), percentage of double skips plants (Double-skip), and percentage of greater than double skip plants ( > Double-skip).Full size tableTable 2 Effect of planter type, seeding rate, and their interaction on yield and stand uniformity variables for all US studies. References: Spacing between plants standard deviation (Spacing sd), achieved versus targeted evenness index mean and standard deviation (ATEI and ATEI sd, respectively), and evenness index mean and standard deviation (EI and EI sd, respectively).Full size tableCommunity-scale data from the four US studies were combined and fitted to bivariate linear regression models with yield as the response variable and each of the stand spatial uniformity variables as the explanatory variable. Significant models (alpha = 0.05) were further evaluated by calculating the coefficient of determination (R2) and root mean squared error (RMSE) (Fig. 2). Models with the lower RMSE and higher R2 were selected as those that best captured the effect of non-uniform stands on soybean yield. After variables were selected, both US and Arg data sets were combined and the linear regressions between the selected variables and yield were re-fitted to assess the consistency of the relationships when an independent data set was included. Community-scale yield from US and Arg studies was modelled as a function of the selected stand uniformity variable, country (US and Arg), and their interaction (fixed effects) (Fig. 3). The spatial uniformity metric showing the most consistent relationship for both US and Arg studies (i.e., non-significant interaction between stand uniformity metric and country), was selected to continue the analysis. The bivariate linear regression models were run with function lm.Figure 2Relationship between stand uniformity variables and soybean yield for US studies. ATEI mean and sd achieved versus targeted evenness index mean and standard deviation, EI mean and sd evenness index mean and standard deviation, Perfect percentage of perfectly spaced plants, R2 coefficient of determination, RMSE root mean square error. All stand uniformity variables presented a significant slope at alpha = 0.05.Full size imageFigure 3Relationship of spacing standard deviation (Spacing sd, cm) and achieved versus targeted evenness index standard deviation (ATEI sd) to soybean yield. Different colors and line types denote different countries (Argentina, Arg—full line, red points; United States, US—dashed line, blue points). R2 coefficient of determination, RMSE root mean square error.Full size imageDifferent environmental conditions and seeding rate levels may modify the effect of plant spatial uniformity on yield. To explore this, each of the studies from Arg and US were separated into low- (USDry19 and ArgDry20, mean of 2.7 Mg ha−1), medium- (USIrr19, USDry20 and ArgDry19, mean of 3.0 Mg ha−1), and high- (USIrr20, mean of 4.3 Mg ha−1) yield environments based on the effect of site-year on yield (Supplementary Fig. 1). Additionally, the tested seeding rates were separated in low ( 300 thousand seeds ha−1) levels based on the current optimal seeding rate for medium yielding environments (235 thousand seeds ha−1, 4 Mg ha−1)13 and the extreme values proposed by Suhre et al.11 (148 and 445 thousand seeds ha−1). This classification was used to model yield as a function of (i) the selected stand uniformity metric, yield environment, and their interaction, and (ii) the selected stand uniformity metric, seeding rate levels, and their interaction. These models were tested to obtain a robust conclusion on the overall effect of yield environment and seeding rate levels, and their interactions (all treated as fixed effects) with plant-to-plant spatial uniformity relative to the response variable, soybean yield. The Akaike information criteria (AIC) was used to compare the full (with interactions) relative to the reduced models (single effects).Ethics declarationsExperimental research and field studies on plants including the collection of plant material, complied with relevant institutional, national, and international guidelines and legislation. More

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    Coral community data Heron Island Great Barrier Reef 1962–2016

    Study site and field data collectionPermanent 1 m2 photoquadrats were established on Heron Reef in 1962/63, using 9 mm diameter mild steel (rebar) pegs, which were replaced over time. From the 1990’s, replacement pegs were stainless steel for greater longevity. Four sites were established, the protected (south) crest, inner flat, exposed (north) crest and exposed pools. Co-ordinates for each site are presented in Table 1, the layout shown in Fig. 2, and sites have been well described previously5,6. At each census, a 1 m2 frame divided into a 5 × 5 grid using string was placed over the pegs, and the quadrat photographed from directly above at low tide. From 1963 until 2003, a 35 mm camera and colour slide film were used. The camera was attached to a tripod affixed to the 1 m2 frame, and captured around 2/3 of the quadrat. The frame (and camera) were then rotated 180 degrees to capture the remainder of the quadrat. After 2003, a hand-held digital camera was used, with the entire quadrat being captured in a single image. Concurrent with each census, mud maps of each quadrat were hand drawn in the field, and all colonies identified in situ by someone with expertise in coral taxonomy.Table 1 Coordinates of the study sites on Heron Island Reef (WGS84).Full size tableFig. 2Quadrat layouts for each of the four sites respectively, noting that the north crest and north ridge have been treated as a single north crest site in previous publications. Underlining indicates original 1962/63 quadrats. Other quadrats were added in or after 2008, as indicated in the text. Contiguous quadrats are pictured bordering each other. Spacing between separate quadrats or groups of quadrats is not shown to scale. Note that up until 2005, NRNW was known as NR. The acronyms in each quadrat represent its name.Full size imageAt the protected (south) crest, a set of six contiguous quadrats were established in 1963 in a 2 × 3 arrangement parallel to the waterline, and about 420 m southeast of the island. This site is exposed at low tide, and was photographed once all water had drained off it. Images of quadrats A, C & E (the shoreward row) from 1963 to 2012 have been fully processed, and the data have been through QA/QC. Data after 2012 exist as images only. These quadrats form the basis of previous analyses1,4,5,6 for this site. Photographs are available for quadrats B, D & F, but apart from 2003–2010, have not been processed. In 2010, an additional two quadrats were established either side of the original six, leading to a 2 × 5 arrangement. Again, only imagery is available for these additional quadrats.At the inner flat, two pairs of contiguous quadrats were established in 1962, 44 m apart, about 70 m south of the island. This site is covered by ~10 cm of water at low tide, so could only be photographed on a still day. Imagery for this site is only available to 2012, after which the marker stakes appear to have been removed in a cleanup of the area. Images for one quadrat in each pair have been processed, but have not been subject to full QA/QC.At the exposed (north) crest main site, a set of four contiguous quadrats was established about 1100 m northeast of the island in 1963. An additional single quadrat (north ridge) was established 326 m to the east. Images from 1963 to 2012 have been fully processed, and the data have been through QA/QC. Data after 2012 exist as images only. In 2005, the single north ridge quadrat was expanded to 4 m2, and in 2008, both subsites were expanded to six quadrats in a 2 × 3 arrangement. These additional quadrats have been digitised up to 2012, but have not been through full QA/QC.The exposed pools are two individual quadrats about 5 m apart about 30 m north of the eastern (north ridge) exposed crest site. These are on the edge of a natural pool, and range from ~5–50 cm deep at low tide, and so could only be photographed on a calm day. Imagery for this site is only available until 2005, after which the marker stakes could not be relocated. Images from 1963 to 1998 have been processed, but have not been through full QA/QC.Retrieval of coral composition data from the photoquadratsProcessing of the images involved scanning the colour slides to produce digital images, and then orthorectifying each image to a 1 m2 basemap in ArcGIS (ESRI Ltd). The corners of the frame, and the holes for the string grid, were used as control points for the orthorectification. For images that originated as colour slides, each half of the quadrat was individually orthorectified to the same basemap, producing a single image of the entire quadrat (see Fig. 3). While contiguous quadrats were orthorectified individually, they were done so against a basemap containing all quadrats in the group, meaning that the resulting images can be easily merged to create a single image of the group. The outlines of all visible coral colonies ( >~1 cm2), and other benthic organisms such as algae and clams, were then digitised in ArcGIS to create a single shapefile for each quadrat for each year. Each colony was represented as an individual feature within the shapefile, and was assigned a unique colony number and species based on the mud maps drawn in the field. Colony numbers were consistent across years, allowing individual colonies to be tracked over time. If a colony underwent fission, the original colony number was retained for each, with the addition of a unique identifier after a decimal point. For example, if colony 35 split in two, the resultant colonies were identified as 35.1 and 35.2. If 35.2 later split again, the resultant colonies were identified as 35.2.1 and 35.2.2. If the colony overlapped the edge of the quadrat, only the area within the quadrat was digitised, and a flag was applied to indicate that only part of the colony was included (edgestatus = 1 in the data). Upon completion of digitisation, ArcGIS was used to calculate the area and perimeter of all colonies. While multiple census were conducted in 1963, 1971 and 1983, only a single census in each year has been processed. There are currently no plans to undertake further digitisation or QA/QC of this data set.Fig. 3Example orthorectified and stitched (prior to 2001) images from the NCNE quadrat, showing the effects of a cyclone that removed all colonies in 1972, and slow recovery over subsequent decades.Full size image More

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    Global soil map pinpoints key sites for conservation

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    Towards an absolute light pollution indicator

    DefinitionWe present here a new statistical approach to measure and characterize light pollution. The objective is to define an indicator which is not limited to clear sky measurements and does not require a precise calibration of a photometer. The key attributes of the indicator are the following:

    It requires the automated acquisition of a large number of zenithal NSB measures when the Sun is below (-18^{circ }) and the Moon below (-5^{circ });

    The acquisitions must at least cover a period of 6 months in order to record a wide range of possible weather conditions from perfectly clear to totally overcast skies. The objective is to obtain a significant sample of every type of cloud conditions (e.g. cloud density and ceiling altitude) as well as a good characterization of the average clear sky ;

    It is based on the analysis of the zenithal NSB measure dispersion which is directly linked to the level of light pollution a site experiences.

    As presented above in Fig. 2, the NSB density histograms, which are assembled from a large number of NSB measures, display a higher density zone which denotes a characteristic clear sky level that we name nominal NSB in the scope of the indicator calculation. On both sides of the clear sky level (above and below), NSB measures are distributed in a way that reflect the zenithal night sky luminance in cloudy conditions: NSB measures above the clear sky level mean that the light pollution is amplified by clouds while those below the clear sky level indicate a darker environment where clouds mask light pollution from distant sources as well as natural light sources. The calculation of the indicator is based on the evaluation of the NSB measure dispersion on both sides of the nominal NSB (i.e. characteristic clear sky level). Since there can have strong variations of artificial light emitted into the environment at the beginning and end of the night (decrease then increase of human activity, extinction of public lighting, etc.), the range of NSB measures retained for calculating the indicator is restricted to a portion in the middle of the night, typically 2 h.Figure 8 shows a typical NSB density histogram for a site which is quite severely impacted by light pollution. It covers a 2 h time range between 23:00 UTC and 01:00 UTC and one can easily see that the zone above the nominal NSB is much higher and denser than the one below, i.e. cloud conditions create more often a brighter environment than a darker one and with a greater amplitude.Figure 8NSB density histogram where the nominal NSB that represents the most common clear sky conditions is identified. It delimits two areas, the NSB bright dispersion above the nominal NSB and the NSB dark dispersion below.Full size imageBased on the determination of the nominal NSB, a quantitative indicator, called NDR for NSB Dispersion Ratio, is calculated in the following way:$$begin{aligned} NDR = (N_b cdot MAD_b) / (N_d cdot MAD_d) end{aligned}$$where (N_b) is the number of measures above the nominal NSB (brighter sky), (N_d) is the number of measures below the nominal NSB (darker sky), (MAD_b) is the median absolute deviation of the measures in the bright dispersion zone (above the nominal NSB) and (MAD_d) is the median absolute deviation of the measures in the dark dispersion zone (below the nominal NSB). The median absolute deviation is a statistical tool used to measure the variability of a data set, which is exactly what we try to achieve with the two NSB extensions above and below the nominal NSB. It is formally defined as (MAD = median(|X_i – tilde{{mathbf {X}}}|)) where (X_i) in our case represents an NSB value and (tilde{{mathbf {X}}}) is (median(X_i)). The median absolute deviation is a better choice than the usual standard deviation to measure the spread of NSB measures since the data does not follow a normal distribution.In order to make the determination of the NSB Dispersion Ratio stronger from a statistical standpoint, we use a bootstrapping with replacement resampling method on the set of night portions used to compute the indicator. Assuming we have N night portions at our disposal, we randomly select a sample of N items in this set of night portions knowing that a given item can appear multiple times in the sample (hence the bootstrapping with replacement). The NDR value is then computed for the considered sample. This process is repeated 1000 times and the average NDR value if eventually computed. This average value represents the actual NDR indicator of the considered site.The NDR indicator takes into account both the number of NSB values on each side of the nominal NSB and the dispersion of these values. This is what makes it relevant as an indicator of light pollution which encompasses all kinds of meteorological conditions experienced at a particular site. On that aspect, it is therefore not an astronomical light pollution indicator since it is not focused on clear sky conditions. On the opposite, it requires to have a significant number of NSB measures in all sorts of cloudy conditions so that a valid NDR indicator can be derived.A key aspect of the NDR calculation methodology is to determine the level of the nominal NSB, i.e. the typical clear sky level, since it will be used to differentiate the NSB measures that go in each of the two sets to calculate the bright and dark dispersions. As we have seen earlier in the article, such a determination can be biased by natural light sources that raise or lower clear sky NSB at different times of the night. This can result into a “blurry” high density zone which makes the determination of the nominal NSB difficult or even impossible depending on the observation period. Based on the quantitative estimate of the different natural light sources presented above, the most important bias to address is the contribution of the Galactic plane. This contribution must be eliminated for all the NSB measures which are used to calculate the NDR indicator. In order to do that, Noxi, the Ninox processing software developed by DarkSkyLab, calculates for each NSB measure the corresponding Galactic plane and star fields contribution using the galactic coordinates of the zenith and integrating the combined flux of all stars in the field of view using the UCAC4 astrometry and photometry star catalogue. It is not possible to cancel the contribution of the airglow due to its unpredictable nature, but since it only appears in rare occasions, it is not seen as a problem and is ignored. Regarding the contribution of the zodiacal light, it is considered as minimal at the zenith and it is also ignored.As an example, Fig. 9 shows on the left an NSB density histogram where the Galactic plane bias has not been corrected in the data, and on the right the same data but with the Galactic plane bias corrected. It is easy to see that in the latter the nominal NSB is much easier to determine, providing a more accurate reference level to calculate the NDR indicator. Once the Galactic plane bias has been corrected, the nominal NSB is determined as the highest density zone of the NSB histogram. It must be noted that, as of today, all the NSB measures are corrected from the Galactic plane bias without regards to the presence of clouds or high levels of light pollution. This results into an additional source of inaccuracy that will be addressed in the future through the implementation of two heuristics within the Noxi software:

    1.

    A first heuristic will determine if a night portion is considered as having a clear sky or not so that the Galactic bias correction is applied only if the sky is clear. In order to do that, we have developed an indicator called the NSS (for Night Sky Stability). To determine the NSS for a full night of measures or just a night portion, we fit the NSB curve with a degree 10 polynomial and we then compute the difference between each NSB measure and it polynomial counterpart. As a result, we obtain a set of residuals. The variance of all the residuals defines the NSS for the considered NSB dataset. Below a given value, the sky is considered as clear knowing that the NSS indicator has been calibrated on several NSB data sets for which the corresponding weather conditions are known;

    2.

    A second heuristic will allow us to weight the Galactic bias correction to be applied to NSB measures according to their value. For non-polluted skies with high values of NSB, the full Galactic bias correction will be applied while below a certain NSB threshold (for instance 21 mag(_{mathrm{SQM}})/arcsec(^{2}) which corresponds roughly to the brightest parts of the Milky Way) no correction will be applied.

    Figure 9NSB density histograms of the same data set with no correction of the Galactic plane bias applied on the left and a full correction applied on the right.Full size imageThe NDR indicator is unitless since it is the ratio of two quantities with the same unit (mag(_{mathrm{SQM}})/arcsec(^{2})). For the data set presented in Fig. 9, the NDR value which is obtained is 25 (which is justified by the fact that the bright extension in the density histogram is much higher and denser than the dark extension). This denotes a quite high level of light pollution despite the fact that the nominal NSB is at a level of 21.6 mag(_{mathrm{SQM}})/arcsec(^{2}). This highlights the fact that there is not always a strict correlation between the typical clear sky NSB obtained for a given site and its NDR indicator, i.e. the presence of clouds decreases the NSB more than we could have expected just by knowing the clear sky NSB. On that respect, the NDR ratio brings more information that the clear sky NSB alone.In addition to provide an indicator which is representative of light pollution in all possible atmospheric conditions, the NDR provides a tool to compare locations in a more meaningful way than just using a set of standalone NSB evaluations. First it is not dependent of an inter-calibration between different systems and second its statistical nature makes it more robust when it comes to perform comparisons.NDR into practiceThe NDR indicator has been calculated for several different sites by DarkSkyLab during various projects in France that involved NSB measuring sessions in the field. To demonstrate some of the results that have been obtained, Fig. 10 provides the density histograms of 4 different sites which have quite different light pollution profiles.Figure 10NSB density histograms of 4 different sites used to compute the NDR indicator. The nominal NSB (which corresponds to the most common clear sky conditions) is noted with a white tick mark next to the vertical axis. Relative levels of the bright and dark dispersion terms (((N_b cdot MAD_b)) and ((N_d cdot MAD_d))) are noted respectively with an orange tick mark and a green tick mark. The computed values of the NDR indicator and nominal NSB are provided in the top-left corner of each figure.Full size imageTo build these diagrams, only the measures acquired during a few hours in the middle of the nights have been used to ensure the maximum stability of the NSB curves and avoid lighting extinctions that create large gaps in NSB profiles. The Galactic plane bias is corrected on all plots and the same NSB scale is used in order to perform comparisons between the 4 sites. One can notice that the number of measures and nights for the 4 sites are quite different. However, they are all sufficient to derive a meaningful value of the NDR indicator using the bootstrapping with replacement resampling method described above, but it is clear that the more NSB measures used, the more accurate the NDR indicator.Table 2 summarizes the NDR indicators as well as the nominal NSB for the 4 sites which are sorted in the order of decreasing NDR indicator values.Table 2 Summary of the nominal NSB and NDR indicators of the 4 different sites.Full size tableOne can see that the NDR indicator values are not strictly correlated to the nominal NSB values, e.g. despite the fact that the nominal NSB of site (a) is slightly better than the one of site (b), the NDR indicator value is much larger for site (a) than for site (b). This can be explained if we consider the specificities of each site:

    Cervières (a) is a small village in the Haut-Forez area, France, which is surrounded by large cities (Lyon, Saint-Etienne and Clermont-Ferrand at a distance between 50 to 80 km) and a closer mid-size city (Roanne at 30 km). At the top of that, the town of Noirétable and a large highway rest area are just 2 km away without any nocturnal extinction applied (as opposed to the village of Cervières itself for which public lighting is turned off from 23:00 to 05:00 local time). These conditions are favourable to the presence of a constant light pollution background which has a negative impact on the zenithal NSB measures in most cloudy conditions (distant large cities for high elevation clouds and Noirétable and the highway rest area for lower elevation clouds). Only rare cloud conditions actually protect the site from the effect of mid-distance light sources. In clear sky conditions, however, the fact that there is no close light sources provides reasonably good NSB levels;

    The Copernic Association Observatory (b) is located 6 km from the large town of Gap in the mountain area of Hautes-Alpes in the south of France. There is no significant short distant light sources but in many cloud conditions the contribution of Gap has a very negative impact on the zenithal luminance. However, due to the fact that the observatory is at a higher altitude on the hills surrounding the city of Gap, there are cloud conditions that make the site darker. In clear sky conditions, the proximity of Gap does not permit a quality better than that of a rural sky;

    The Astrièves Observatory (c) is located near the center of the small town of Gresse-en-Vercors in the Parc Naturel Régional du Vercors. There is a full nocturnal extinction of the village for a large part of the night resulting in a good sky quality in clear sky conditions. The large city of Grenoble is at a distance of 30 km in a valley at the north-east, and the two locations are separated by a few mountains which efficiently help masking the light pollution as soon as the cloud ceiling is below a certain altitude, resulting into a dark environment. On the opposite, high elevation clouds reflect the light from Grenoble and increase the zenithal luminance;

    Eourres (d) is a small and isolated village located 20 km west of Sisteron in the department of Hautes-Alpes, France, which is surrounded by mountains. There is no significant light sources closer than those of Sisteron and this results into a very good night sky quality with, most of the times, a very dark environment in cloudy conditions.

    Figure 11 provides a graphical representation of the NDR indicator values for the 4 sites. On the NDR scale, the value 1 indicates that the bright and dark dispersion terms (respectively ((N_b cdot MAD_b)) and ((N_d cdot MAD_d))) are equal, which means there is a balance between dark and bright conditions at the zenith on the considered site with reference to the most common clear sky level.Figure 11Summary of the NDR indicators obtained for the 4 sites. The diagram uses 1 as the pivotal value to delineate sites according to the two bright and dark dispersion terms ((N_b cdot MAD_b)) and ((N_d cdot MAD_d)).Full size imageThe NDR can theoretically vary between 0 (totally dark site) and several hundreds (extremely bright site) but in practice the best sites can reach NDR indicator values down to 0.3 in the best preserved locations and up to 200 for very large and polluted cities.Robustness of the NDR indicatorIt is important to evaluate how the NDR indicator is dependant on the number of measures used to compute it and to figure out what would be the minimum number of night sessions required to obtain a meaningful NDR indicator value at a given site. To achieve that, we have used the data from two of the four sites presented above (the two which have the largest number or recorded nights: Cervières with 424 nights and the Astrièves Observatory with 373 nights). The 1000-step bootstrapping procedure has been repeatedly executed on each data set with a regularly decreasing sample of nights: starting from the full number of nights, a decrement of 10 nights is applied at each step until only 20 nights are remaining. At every bootstrap step, each sample is composed of n nights randomly chosen among the N available ones knowing that any night can be selected several times.Figure 12 shows the NDR indicator values that have been obtained for each of the two sites as a function of the night sample considered. The 95% confidence interval is plotted against each NDR indicator value (it is preferred to the standard deviation since the NSB distribution in the data sets is not normal). In the right plot of Fig. 12, the last confidence interval for the 24 night sample is too wide to fit in the y-axis NDR range (the top value is 195).Figure 12Results of the NDR resampling on the two data sets of Cervières and Astrièves Observatory. The horizontal axis is the number of nights considered into the night sample and the vertical axis provides the NDR indicator obtained for each sampling set through a 1000-iteration bootstrapping with replacement calculation.Full size imageDiscussion on the required number of nightsWe can see in Fig. 12 that the NDR indicator and the confidence interval remain stable down to 200 nights. Below this threshold, the NDR starts to become unstable with growing confidence intervals. Based on this data, we can estimate that the minimum number of nights required to compute a robust NDR indicator is 200 (therefore between 7 and 8 months since there are periods around the full moons where there is no night portions recorded).However, depending on the measuring session objectives, the NDR indicator can be considered as accurate enough even when using a smaller number of nights. If the goal is simply to get a first estimate of the light pollution level at a given site, we can consider that 90 nights (a little more than 3 months of measures) are enough. On the opposite, if we want to perform a comparison between several sites for evaluating the impact of light pollution on a particular species, we might want to perform at least 200 nights of measurement to get a better accuracy for the NDR indicator. The experience from DarkSkyLab through many NSB measuring sessions is that 3 to 4 months of measures are required to get a meaningful density histogram, hence an accurate enough NDR indicator, so that a site can be sufficiently characterized from a light pollution perspective. Such a measuring period usually guarantees that the clear sky nominal NSB is well defined and that various cloud conditions have been observed. This estimate is sustained by the results obtained in Fig. 12.Value of the NDR indicator for ecological researchThe study of the impact of light pollution on biodiversity is currently in full expansion, amplifying a political and citizen demand for the reclamation of the night2,32,33.We identify three main contributions of the NDR indicator for ecological research. First, it overcomes the limits of an old problem of communication in terms of measurement units between disciplines and potentially limits the use of units without real meaning from a biodiversity point of view34,35. Secondly, the use of the NDR indicator limits the common biases linked to a characterization of the effects of anthropogenic light which is too limited in time and space35. Indeed, the life history traits of species are not only shaped by the intensity of light emitted into the nocturnal environment but also by its variation over time34,35,36. Currently, the characterization of light pollution is too often limited in time and space, which can lead to misinterpretation37. Thirdly, the NDR indicator provides ecological researchers with a unit of measurement that integrates a sufficiently long time step to study the impact of light pollution on the evolutionary processes at work in the life of species and particularly on population dynamics and animal behavior36,38,39.Limitations and future improvements of the NDR indicatorThe main limitation of the NDR indicator resides in the possible difficulty to identify a well defined value for the nominal NSB, i.e. the NSB value that represents the most common clear sky conditions of a given site. For the most part, this is due to the contribution of the Galactic plane to the zenithal sky brightness and, to a lower extent, to the contribution of other natural light sources (dense star fields, airglow and zodiacal light). The residual spread of NSB measures is due to changing atmospheric conditions at various time scales, but, for this particular contribution, we can expect a statistical compensation to eliminate a systematic associated bias.At the moment, the contribution of the Galactic plane and star fields is canceled into the NSB measures by calculating in the Noxi software the integrated flux of all the stars that belong to the field of view (using the UCAC4 star catalogue). However, this approach has proven some limitations, especially in the southern hemisphere where the Galactic center goes through the zenith and is particularly bright. A probable explanation for that lack of predictability is the fact that the Galactic plane contains diffuse sources such as nebulae which are not accounted for into the star catalogues and which actually cannot be ignored. To address this issue, DarkSkyLab has the project to create a brightness map of the Galactic plane with a square degree resolution or better so that the contribution of all sources can be correctly accounted for.In addition to better correcting the Galactic plane bias, an improvement must be made with regards to the NSB measures that need to be corrected. At the moment, all NSB measures are corrected from the Galactic plane bias without regards to the presence of clouds or high levels of light pollution. So a first heuristic must be implemented to only apply the bias correction to clear sky NSB measures. An other heuristic must also be developed to reduce the correction applied as a function of the NSB level.A third limitation of the NDR indicator is related to a possible lack of cloudy conditions at some sites (e.g. in the Atacama desert in Chile with more than 320 clear nights per year), the reason simply being that the NDR indicator requires the presence of clouds to differentiate the bright and dark extensions into the NSB density histograms. This means that the NDR indicator can hardly be used for such astronomy-oriented sites which experience rare cloudy conditions. More