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    A bottom-up view of antimicrobial resistance transmission in developing countries

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    Visible-NIR hyperspectral classification of grass based on multivariate smooth mapping and extreme active learning approach

    Study areaGrassland herbage samples are from Shaerqin base, institute of grassland research of CAAS (Chinese Academy of Agricultural Sciences). We obtained the permission of the institution to take HSI of the grassland sample. Our work did not cause damage to grassland. Researcher Weihong Yan of the institute provided us with relevant information about grassland. The land use type in the study area is mainly grassland, which is composed of forage species, most of which are representative species of typical grassland. We take this area as an example to conduct research on grass classification. By enriching the relevant recognition technology, it can also be used as a reference for the pastures of other grasslands. The grass species Grass1 for the experiment is shown in Table 1. The official introduction of plant materials is detailed in the flora of China15.Table 1 Samples information for Grass1 dataset.Full size tableThe field hyperspectral platformWe assemble a system for collecting HSI in the field: HyperSpec©PTU-D48E HSI instrument, high-precision scanning PTZ, tripod, data analysis software Hyperspec, etc. The light source is natural light. The imaging instrument is in line scanning mode. Table 2 shows the technical parameters.Table 2 Technical parameters of hyperspectral instrument.Full size tableData collectionIn July 2021, the data was collected during the lush grass growth period. Collect data from 11:00 a.m. to 2:00 p.m. every day. At this time, it is sunny, cloudless and the wind force does not exceed level 2. So as to ensure the consistency of the acquisition time line and avoid the influence of different degrees of light on the reflectivity as far as possible. The measuring points are arranged facing the sun and the opposite direction of the shadow. We collect data from different angles of the grassland, which is based on the growth of various types of forages, and selects relatively concentrated places within the study area. Each shot is a single category of grass. The image resolution is 1166 × 1004 pixels (Fig. 1). The imaging spectrometer is fixed with scanning head when shooting. Data acquisition and transmission are executed on Hyperspec software. Then save it as a BIL file. The ENVI5.3 software was used to extract the forage spectrum to establish the dataset Grass1. Well balanced regions with a clear image, uniform spectral distribution are selected for further segmentation. The average value of spectral reflectance of grass pixels was taken as the reflectance spectrum of a single type of grass.Figure 1True color map of grass samples.Full size imageMethodologyIn Fig. 2, we present the framework of visible-NIR hyperspectral classification of grass based on multivariate smooth mapping and extreme active learning (MSM–EAL). Specifically, we first introduce the proposed MSM algorithm for global enhanced spectral reconstruction, which utilizes smooth manifold projection technology to alleviate the problems of difficult feature selection and redundant data. Then, the EAL framework is proposed to address the matter of hyperspectral labeled samples and spectral classification. In the following, each step of this method will be presented in detail.Figure 2Proposed MSM–EAL framework for grass HSI classification.Full size imageThe proposed MSM algorithmIn the process of field HSI acquisition, on the one hand, the surface distribution of grass is uneven and the plant height is different, causing certain scattering effect and coverage spectrum change. On the other hand, HSI is easy to be disturbed by external natural factors such as light, wind and shadow, resulting in a certain degree of distortion. Multiplicative scatter correction (MSC) is a scattering correction effect, which helps to eliminate the scattering effect caused by the above reasons and enhance the spectral variability. The moving window smooth spectral matrix (Nirmaf) belongs to the smooth effect, which improve the signal-to-noise ratio of the spectrum and reduce the influence of random noise16,17. Preprocessing methods are different and related to each other. We design an enhanced preprocessing multivariate smooth (MS) method that fusing MSC and smooth Nirmaf to target grass spectral signal features. In the follow-up, a model will be established to verify the validity of MS.Most of the high-dimensional spatial data have the characteristics of being embedded in a manifold body, so the manifold learning isometric feature mapping (Isomap) based on spectral theory is adopted. Isomap preserves the global geometric features of the initial data and extracts features by reconstructing the underlying smooth manifold of HSI. It is nonlinear dimensionality reduction based on linear and multidimensional scaling transformation18. Isomap has been applied in image and HSI classification19,20, but there is no report on visible-NIR hyperspectral classification of grass.In view of the above, we proposed the multivariate smooth mapping (MSM) spectral reconstruction algorithm, which can be represented as follows:$$ MSM_{z} { } = { }frac{{left( {P_{j} – b_{j} } right)left( {2n + 1} right) + n_{j} cdot mathop sum nolimits_{j = – n}^{n} C_{j} P_{k + j} }}{{n_{j} left( {2n + 1} right)}} + V_{Z} F_{Z}^{frac{1}{2}} { } $$
    (1)
    where Pj, bj, and Cj represent the raw reflectance value of spectrum j, baseline shift amount, and weight factor, respectively, k and nj represent the polynomial degree and offset, respectively. MSMz is the feature cube reconstructed to Z dimension from the spectrum calculated by 2n + 1 moving window width, V eigenvector matrix and F eigenvalue matrix.In Isomap equidistant mapping, the shortest path of edge Pi Pj needs to be solved, and the representation matrix is:$$ D_{G} = [d_{G}^{2} (P_{i} ,P_{j} )]_{i,j = 1}^{n} $$
    (2)
    where d (Pi, Pj) is the weight of the edge Pi Pj calculated from the neighborhood graph G and its side Pi Pj.The proposed EAL frameworkLabeling hyperspectral samples is expensive in terms of time and cost, at the same time, the lower spatial resolution and more bands increase the difficulty of labeling. Active learning (AL) provides an efficient labeling strategy, which only needs to label a relatively small number of samples to learn a more accurate model21. The pool-based AL selects the most informative samples according to the query strategy for limited labeling through iteration, so as to facilitate model improvement. Commonly used query strategies are uncertainty criteria, such as least confidence22, the bayesian active learning disagreement (BALD), the entropy sampling23, etc.Due to there is still an over-fitting problem, different strategies such as hybrid prediction and regularization need to be used for non-recursive datasets24. The research25 proposed that extreme gradient boosting algorithm (XGBoost) based on gradient boosting. As a classification method, XGBoost has been successfully applied in Kaggle competition and other fields. Its most important feature for visible-NIR hyperspectral classification is that can easily and directly classify according to features, and the physical interpretation of features can help understand the electronic nature behind spectral classification. XGBoost is a machine learning algorithm based tree structure that integrates multiple weak classifiers to achieve flexible and high-precision classification. It is an upgraded version of gradient boosting decision tree. The optimization process of XGBoost entailed: (1) Expanding the objective function to the second order, and finds a new objective function for the new base model to improve the calculation accuracy. (2) L2 regularization term is added to the loss function to prevent over-fitting. (3) Using blocks storage structure realize automatic parallel computing26,27. The algorithm steps are as follows:The objective function:$$ Lleft( Phi right) = mathop sum limits_{i} lleft( {y^{i} ,widehat{{y^{i} }}} right) + mathop sum limits_{k} Omega left( {f_{k} } right) $$
    (3)
    In formula (3), the first and second terms are the loss function term and the regularization term, respectively. Where,$$ Omega left( {f_{k} } right) =upgamma {text{T}} + frac{1}{2}lambda left| w right|^{2} $$
    (4)
    γ and λ are regularization parameters which are used to adjust complexity of the tree.Next, second derivative Taylor expansion of the objective function. Where (g_{i}) and (h_{i}) are the first derivative and second derivative, respectively.$$ L^{left( t right)} = mathop sum limits_{i = 1}^{n} lleft( {y_{i} ,widehat{{y_{i}^{t – 1} }} + f_{t} left( {x_{i} } right)} right) + Omega left( {f_{t} } right) $$
    (5)
    $$ g_{i} = partial_{{hat{y}_{i} (t – 1)}} lleft( {y_{i} ,widehat{{y_{i}^{t – 1} }}} right) $$
    (6)
    $$ h_{i} = partial_{{widehat{{y_{i} }}(t – 1)}}^{2} lleft( {y_{i} ,widehat{{y_{i}^{t – 1} }}} right) $$
    (7)
    $$ {text{L}}^{left( t right)} approx mathop sum limits_{i = 1}^{n} left[ {lleft( {y_{i} ,widehat{{y_{i}^{t – 1} }}} right) + g_{i} f_{i} left( {x_{i} } right) + frac{1}{2}h_{i} f_{t}^{2} left( {x_{i} } right)} right] + Omega left( {f_{t} } right) $$
    (8)
    Final objective function:$$ {hat{text{L}}}^{ i} left( q right) = – frac{1}{2}mathop sum limits_{j = 1}^{T} frac{{(mathop sum nolimits_{{i in I_{j} }} g_{i} )^{2} }}{{mathop sum nolimits_{{i in I_{j} }} h_{i} + lambda }} + gamma T $$
    (9)
    Equation (9) can be used as the fraction of tree cotyledons, and the tree structure is directly proportional to the fraction. If the result after splitting is less than the maximum value of the given parameter, the cotyledon depth stops growing24,28.AL solves the problems of limited number and high cost of grass hyperspectral labeling samples. The default model of traditional AL is logistic regression, which is mostly studied on the ideal public dataset. However, the actual data has more uncertain noise, which still poses a certain challenge to AL. Consequently, we propose the extreme active learning (EAL) framework to minimize the classification cost of visible-NIR hyperspectral. The framework replaces the logistic regression model with XGBoost. Taking advantage of AL, XGBoost can improve performance with less training marker samples. By jointing of XGBoost and AL, EAL provides significantly better results than AL in field Grassl dataset recognition. Additionally, based on the characteristics of XGBoost, EAL more intuitively enhances the physical essence behind spectral classification than AL. Algorithm 1 summarizes the workflow of EAL framework.Random forest (RF) and decision tree (DT) were used to compare with EAL. RF and DT are frequently used in the field of grassland remote sensing9,29. Furthermore, RF, DT and XGBoost have the same point is that are learning algorithms based on tree structure. DT determines the direction by judging the conditions of the decision node12. RF is an integrated learning of multiple decision trees30. More

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    Author Correction: Associations between carabid beetles and fungi in the light of 200 years of published literature

    These authors contributed equally: Gábor Pozsgai, Ibtissem Ben Fekih.State Key Laboratory of Ecological Pest Control for Fujian and Taiwan Crops, Institute of Applied Ecology, Fujian Agriculture and Forestry University, Fuzhou, 350002, ChinaGábor Pozsgai, Ibtissem Ben Fekih, Jie Zhang & Minsheng YouJoint international Research Laboratory of Ecological Pest Control, Ministry of Education, Fuzhou, 350002, ChinaGábor Pozsgai, Gábor L. Lövei & Minsheng YouCE3C – Centre for Ecology, Evolution and Environmental Changes, Azorean Biodiversity Group and Universidade dos Açores, Angra do Heroísmo, 9700-042, Azores, PortugalGábor PozsgaiInstitute of Environmental Microbiology, College of Resources and Environment, Fujian Agriculture and Forestry University, Fuzhou, 350002, ChinaIbtissem Ben Fekih & Christopher RensingBasic Forestry and Proteomics Research Center, College of Life Science, Fujian Provincial Key Laboratory of Haixia Applied Plant Systems Biology, Fujian Agriculture and Forestry University, Fuzhou, 350002, ChinaMarkus V. KohnenLaboratoire de Biologie et de Physiologie des Organismes, Faculté des Sciences Biologiques, Université des Sciences et de la Technologie Houari Boumediène, BP 32 El Alia, Alger, 16111, AlgeriaSaid AmraniDuna-Ipoly National Park Directorate, Költő u. 21, H-1121, Budapest, HungarySándor BércesJuhász-Nagy Pál Doctoral School, University of Debrecen, Egyetem tér 1, H-4032, Debrecen, HungarySándor BércesDepartment of Zoology, Plant Protection Institute, Centre for Agricultural Research, Nagykovácsi út 26-30, H-1029, Budapest, HungaryDávid FülöpFujian University Key Laboratory for Plant-Microbe Interaction, College of Plant Protection, Fujian Agriculture and Forestry University, Fuzhou, 350002, ChinaMohammed Y. M. JaberDepartment of Plant and Environmental Sciences, University of Copenhagen, Thorvaldsensvej 40, 1871, Frederiksberg C, DenmarkNicolai Vitt MeylingDepartment of Algology and Mycology Faculty of Biology and Environmental Protection, University of Łódź, Banacha 12/16, PL-90-237, Łódź, PolandMalgorzata Ruszkiewicz-MichalskaDepartment of Molecular Biotechnology and Microbiology, University of Debrecen, Egyetem tér 1, Debrecen, H-4032, HungaryWalter P. PflieglerFujian Provincial Key Laboratory of Insect Ecology, College of Plant Protection, Fujian Agriculture and Forestry University, Fuzhou, 350002, ChinaFrancisco Javier Sánchez-GarcíaÁrea de Biología Animal, Departamento de Zoología y Antropología Física, Facultad de Veterinaria, Universidad de Murcia, Murcia, 30100, SpainFrancisco Javier Sánchez-GarcíaDepartment of Agroecology, Aarhus University, Flakkebjerg Research Centre, Forsøgsvej 1, DK-4200, Slagelse, DenmarkGábor L. Lövei More

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    The skilled ecosystem engineers with big teeth and paddle tails

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    Changes in plant biodiversity facets of rocky outcrops and their surrounding rangelands across precipitation and soil gradients

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    Municipal biowaste treatment plants contribute to the contamination of the environment with residues of biodegradable plastics with putative higher persistence potential

    Choice of biowaste treatment plants and sample identifiersCompost samples were collected from four central municipal biowaste treatment plants (denominated as #1 to #4) in Baden-Wurttemberg, Germany (Table 1). All plants used a state-of-the-art two-stage biowaste treatment process comprising of (a) anaerobic digestion/biogas production and (b) subsequent composting of the solid digestate to produce a high-quality mature compost sold for direct use as fertilizer in agriculture. The composts were regularly analyzed by an independent laboratory for quality and residual contamination and consistently fulfilled the quality requirements of the label RAL-GZ 251 Gütezeichen Kompost of the German Bundesgütegemeinschaft Kompost e.V. (www.gz-kompost.de). Plants #1 and #3 produce in addition a liquid fertilizer, which is separated from the solid digestate at the end of stage a) by press filtration and which is also intended for direct use on agricultural soil (replacement of liquid manure). In case of plants #1, #3, and #4 up to 25 wt% of shrub/tree cuttings were added to the solid digestate for composting. All plants used sieving (typically with a 12 or a 20 mm mesh) at the end of the process to assure the necessary purity of their finished composts. Whenever technically possible, we as well took samples of the pre-compost immediately before this final sieving step to evaluate its contribution to the removal of residual BPD fragments. For analysis, composts were passed consecutively through two sieves with mesh sizes of 5 mm and 1 mm, yielding two fragment preparations for IR-analysis namely a > 5 mm fraction corresponding to the contamination by residual “macroplastic” (5 mm is a commonly used upper size limit for “microplastic”, anything larger is macroplastic) and a 1–5 mm fraction corresponding to the regulatory relevant residual contamination by microplastic. The lower limit of 1 mm rather than 2 mm was chosen in anticipation of the expected changes in regulation, where the replacement of the 2 mm limit by a 1 mm limit is imminent.Table 1 Technical data of the investigated plants and incidence of BDP fragments in the sampled composts.Full size tableOccurrence of plastic fragments  > 1 mm in the sampled compostsComposting times of 5–9 weeks were used in the investigated plants (Table 1), which is shorter than the 12 weeks indicated in EN 13432 for the 90% disintegration of a compostable plastic material, but a realistic time span for state-of-the-art technical waste treatment. Since we were not in a position to estimate the quantity of BDP entering the plants, since for technical reasons we were unable to obtain a representative sample, we cannot say, whether any residual BDP detected by us in the finished composts was due to a yet incomplete disintegration process or whether it corresponds to the 10% material still permissible by EN 13432 even after the full composting step. However, in 7 out of the 12 sampled composts and pre-composts fragments with chemical signatures corresponding to the BDPs poly (lactic acid) (PLA) and poly (butylene-adipate-co-terephthalate) (PBAT) were identified in the > 5 mm and/or the 1–5 mm sieving fractions using FTIR analysis3 (Fig. 1; Table 1). All recovered fragments appeared to stem from foils, bags or packaging, since they were thin compared to their length and width (see Suppl Figure S1 for typical examples). Fragments with overlapping signatures, most likely PBAT/PLA mixtures or blends, were also found (see Suppl Figure S2 for the interpretation of the spectra). In addition, the recorded BDP fragment spectra (Fig. 1A) showed high similarity to the FTIR spectra of commercial compostable bags sold in the vicinity of the biowaste treatment plants (Fig. 1B), which together with the geometry of the recovered fragments led us to assuming that the majority of the BDP entered the biowaste in the form of such bags.Figure 1FTIR spectra of BDP fragments from composts and commercial bags. (A) BDP fragments recovered from the composts and (B) the commercial compostable bags. Fragments were coded as follows: p or f for pre-compost or finished compost, followed by the plant number (#1 to #4), an indication of the size fraction ( > 5 mm or 1–5 mm) in which the fragment was found, and finally, the fragment number. Fragment F#1_5mm_4 therefore represents the 4th fragment collected in the  > 5 mm size fraction from the finished compost of plant number 1. Bags were arbitrarily numbered 1–10, see Suppl Table S1 for supplier information. The spectra (in grey) of the reference materials for PLA and PBAT are given as basis for the interpretation. Spectra in red refer to test samples consisting only of PBAT, while those in blue indicate samples composed of PBAT/PLA mixtures.Full size imageThe BDP fragments were found alongside fragments of commodity plastics (mostly PE) in all cases. Finished composts tended to contain fewer and smaller fragments than the corresponding pre-composts. The final sieving of the pre-composts to prepare the finished composts hence appears to be quite effective in removing such fragments, in particular those from the > 5 mm size fraction (Table 1) and for that reason has become state-of-the-art in preparing quality composts (contamination by plastic fragments > 2 mm of less than 0.1 wt%). Given that the size of the fragments is a crucial factor regarding ecological risk, we analyzed the sizes (length Î width) of the BDP fragments in comparison to that of the plastic fragments with signatures of commodity plastics such as PE (Fig. 2). BDP fragments found in a given compost sample tended to be smaller than the fragments stemming from non-BDP materials, which may indicate that BDPs degrade faster or tend to disintegrate into tinier particles than commodity plastics. This may also explain why in the compost from plant #2, no BDP fragments were found in the particle fraction retained by the 5 mm sieve ( > 5 mm fraction), while 19 such particles were found in the fraction then retained by the 1 mm sieve (1–5 mm fraction). Interestingly, plant #2 is the only one included in our study that uses no mechanical breakdown of the incoming biowaste. This reduces the mechanical stress on the incoming material. Mechanical stress can alter the properties of plastic foils such as the crystallinity whereby crystallinity has been shown to influence the biological degradation of BDP such as PLA7.Figure 2Size distribution of plastic fragments  > 1 mm. (A) Fragments found in the finished compost from plant #1, (B) in the finished compost from plant #2, and (C) in the pre-compost from plant #3. For reasons of statistical relevance, only samples containing more than 20 BDP fragments per kg of compost were included in the analysis.Full size imageMaterial characteristics of BDP fragments in comparison to those of commercial biodegradable bagsIn order to verify whether the BDP fragments recovered from the composts differed from the compostable bags in any parameter with possible relevance for biodegradation and environmental impact16, the physico-chemical properties of bags and fragments were studied in detail. Since we wanted to have a maximum of information of the BDP fragments, size/weight was a limiting factor in selecting fragments for analysis. Fragments of at least 1 mg were required for the FT-IR analysis. 5 mg-fragments could be analyzed in addition by 1H-NMR, while the full set of analytics (FT-IR, 1H-NMR, and DSC) required at least 10 mg of sample.For insight into the chemical composition, 1H-NMR spectra of the commercial bags and all suitable BDP fragments were compared (Fig. 3). In case of material mixtures and blends, the 1H-NMR analysis allows quantification of the PBAT/PLA weight ratio in the materials and also of the ratio of the butylene terephthalate (BT) and butylene adipate (BA) units in the involved PBAT polyesters.Figure 31H NMR spectra of BDP fragments from composts and commercial bags. (A) BDP fragments recovered from the composts and (B) the commercial compostable bags. Fragments were coded as follows: p or f for pre-compost or finished compost, followed by the plant number (#1 to #4), an indication of the size fraction ( > 5 mm or 1–5 mm) in which the fragment was found, and finally, the fragment number. Bags were arbitrarily numbered 1–10, see Suppl Table S1 for supplier information. The spectra (in grey) of the reference materials for PLA and PBAT are given as basis for the interpretation. Spectra in red refer to test samples consisting only of PBAT, while those in blue indicate samples composed of PBAT/PLA mixtures. (C) Chemical structures of PLA and PBAT, chemical shifts of the protons are assigned as indicated in the reference spectra in (B).Full size imageThe 1H-NMR spectra corroborate the FTIR measurements in that all investigated commercial bags were made from PBAT/PLA mixtures of varied composition (Table 2). By comparison, some of the fragments, for instance, f#1_5mm_4, appeared to consist of only PBAT. Other fragments, e.g., f#1_1mm_9, were mixtures of PLA and PBAT (Table 2). However, even in the case of PBAT/PLA mixtures, the average PBAT content tended to be higher in the fragments than in the bags, while the BT/BA monomer ratio in the respective PBATs, was also significantly higher in the fragments than in the bags. If we assume the fragments to stem from similar compostable bags as the ones included in our comparison, this would mean that during composting of such a bag, the PLA degrades more quickly than the PBAT, whereas within a given PBAT polyester, the BA unit is more easily degraded than the BT unit. Evidence can indeed be found in the pertinent literature that PLA has faster biodegradation kinetics than PBAT, while BT is more resistant to mineralization than BA17,18.Table 2 Composition of commercial compostable bags and BDP fragments recovered from the composts as analyzed by 1H-NMR.Full size tableNext, differential scanning calorimetry (DSC) was used to analyze fragments compared to commercial bags in regard to the presence of amorphous vs. crystalline domains, a parameter expected to affect biodegradation kinetics and therefore the putative environmental impact of the produced microplastic16 upon release into the environment with the composts. Whereas amorphous domains show glass transition, crystalline domains show melting, both of which can be discerned by the respective phase transition enthalpy in the DSC curves (Fig. 4).Figure 4DSC curves of BDP fragments and compostable bags #1 and #7. Curves for the reference materials (in grey) for PLA and PBAT are given for comparison. Curves were recorded during the first heating run (temperature range: − 50 °C to 200 °C, heating rate: 10 °C min−1). (A) and (B) curves in red refer to test samples consisting only of PBAT, while those in blue indicate samples composed of PBAT/PLA mixtures. Fragments were coded as follows: p or f for pre-compost or finished compost, followed by the plant number (#1 to #4), an indication of the size fraction ( > 5 mm or 1–5 mm) in which the fragment was found, and finally, the fragment number.Full size imageThe curve for the reference PBAT shows a glass transition temperature (Tg) of − 29 °C and a broad melting range between 100 and 140 °C for the crystalline domains, while that of the PLA reference shows a glass transition temperature of 58 °C and a narrower melting peak between 144 °C and 162 °C. The curve for commercial bag #1, which had a comparatively high PLA content, shows a pronounced melting peak in the expected range; the same is the case for fragment p#3_5mm_1 and to a lesser extent for fragment p#3_5mm_9, two fragments, which also have high PLA contents. The DSC curves of the other fragments and bag #1 are undefined in comparison, which is due to their high PBAT content. According to the DSC curves, most of the investigated materials are semicrystalline, i.e., contain both amorphous (glass transition) and crystalline (melting) domains. However, the DCS data alone allow only a qualitative discussion of the differences between fragments and bags.To obtain quantitative data on the crystallinity differences, wide angle X-ray scattering (WAXS) spectra were recorded. WAXS requires fragments at least 3 cm long, which restricted the number of fragment samples to three, all of which were found in pre-compost samples. The corresponding curves are shown in Fig. 5A–C. The spectra of the commercial biodegradable bags are shown in Suppl Figure S3. Foils were in addition prepared by heat pressing from the reference materials for PLA and PBAT in order to include them into the WAXS measurements (Fig. 5D). While the foils produced from the PBAT reference material produced crystallinity peaks at 16.2°, 17.3°, 20.4°, 23.2°, and 24.8°, the foil prepared from the PLA reference material showed only an amorphous halo at 15.5° and 31.5°, which is in accordance with values published in the literature19. A more pronounced crystallinity peak was obtained in the case of an additionally annealed PLA foil.Figure 5WAXS curves with Lorenz fitting for (A) fragment p#3_5mm_1, (B) fragment p#3_5mm_9, and (C) fragment p#4_5mm_2. (D) WAXS curves for foils produced from the PBAT and PLA reference materials; the percent values indicate the crystallinity. The dash lines are the fitting peak curves for the XRD spectrum. Crystallinity can be obtained by dividing the integration area of the fitted peaks by the integration area of the entire spectrum. Fragments were coded as follows: p or f for pre-compost or finished compost, followed by the plant number (#1 to #4), an indication of the size fraction ( > 5 mm or 1–5 mm) in which the fragment was found, and finally, the fragment number.Full size imageIn case of the fragments and bags, the peaks of PLA and PBAT overlapped to some extent in the WAXS spectra, but by conducting Lorenz fitting using Origin software, the overall crystallinity could be calculated as follows:$$chi = { 1}00% , *{text{ Aa}}/left( {{text{Aa }} + {text{ Ac}}} right)$$where χ is the crystallinity and Aa and Ac represent the areas of the amorphous and crystalline peaks.Using this equation, crystallinities of 55% (fragments p#3_5mm_1), 34% (p#3_5mm_9), and 34% (p#4_5mm_2) were calculated for the fragments. The foils prepared in house for the reference materials had similar crystallinities (43% in case of the annealed PLA foil and 26% of the PBAT foil), while the simple PLA foil was amorphous. By comparison, for eight of the commercial bags, crystallinities in the range from 1% to 7% were calculated, whereas these values were 14% and 15% for the remaining two bag types (Suppl Figure S3).The high crystallinity of the larger fragments recovered from the pre-compost samples suggests that crystalline domains of BDP materials may indeed disintegrate more slowly than the amorphous ones, as prior studies on microbial biodegradation have suggested7,8. Admittedly, such large fragments per se would not enter the environment, since the final sieving step used to prepare the finished composts is quite efficient at removing them. However, it is tempting to extrapolate that residual BDP in general are remnants of the more crystal domains of the original material, even though experimental proof of this assumption is at present not possible. 10 wt% of a BDP bag is allowed to remain after standard composting. It is usually assumed that any such residues continue to degrade with comparable speed. However, should these residues correspond to the more crystalline domains, rather than degrading with similar speed as the bulk material, the more crystalline fragments can be expected to persist for a much longer and at present unpredictable length of time in the environment, e.g. when applied to the soil with the composts; in particular, when they are also enriched in PBAT and BT units as suggested by our analysis of the chemical composition. Data from the use of biodegradable foils in agriculture show that the degradation in the environment may take years20. Altogether this may have unforeseen economic and environmental consequences, especially when considering the high fraction of BDP fragments < 5 mm. Putative consequences include changes in soil properties, the soil microbiome and therefore in plant performance21, a factor indispensable for worldwide nutrition.Residues of BDP fragments  1 mm were found in the collected LF samples. This is hardly surprising, given that the LF is produced by press filtration of the digestate after the anaerobic stage. Such a filtration step can be expected to retain fragments > 1 mm in the produced filter cake, which goes into the composting step, leaving the filtrate, i.e. the LF, essentially free of such particles. Anaerobic digestion is currently not assumed to contribute significantly to the degradation of BDP17,22, but the process conditions (mixing, pumping) may promote breakdown of larger fragments, particularly when additives such as plasticizers23 leach out of the material.Since the residual solids content of the LF is low (plant #1: 8.6 wt%, plant #3: 5.8 wt%), a combination of enzymatic-oxidative treatment and µFTIR imaging originally developed for environmental samples from aqueous systems24,25 could be adapted for the analysis (size and chemical signature) of particles in the LF down to a size of 10 µm. The corresponding data are compiled in Table 3. In all cases, residual fragments from PBAT-based polymers represented the dominant plastic fraction in the investigated samples; i.e. approximately 53% of all plastic particles in the LF from plant #1 (11,520 BDP particles per liter) and 65% in the case of plant #3 (12,480 BDP particles per liter). Liquid manure is applied several times a year to fields at a concentration of 2–3 L m−2. According to our analysis > 20,000 BDP microparticles of a size ranging from 10 µm to 500 µm enter each m2 of agricultural soil whenever LF is applied on agricultural surfaces.Table 3 Microplastic fragments (BDP/all) found per liter of liquid fertilizer.Full size tableDue to the complexity of the matrix, a similar analysis of individual plastic fragments  1 mm. Six compost samples representing the more contaminated ones based on the content of fragments > 1 mm, namely, f#1, f#2, p#3, f#3, p#4 and f#4 (nomenclature: f or p for finished or pre-compost, followed by plant number), were extracted with a 90/10 vol% chloroform/methanol mixture. The amounts of PBAT and PLA in the obtained extracts were then quantified via 1H-NMR (Table 4). Briefly, the intensity of characteristic signals in the extract spectra of the compost samples (see Suppl Figure S4) were compared to peak intensities produced by calibration standards of the pure polymer dissolved at a known concentration in the chloroform/methanol. All samples and standards were normalized using the 1,2-dichloroethan signal at 3.73 ppm as internal standard. See also Suppl Figure S5 for an exemplification of the quantification of the PBAT/PLA ratios. Based on the amounts of PBAT and PLA extracted from a known amount of compost, the total mass concentration (wt% dry weight) of these polymers in the composts was calculated.Table 4 Evidence of PBAT and PLA residues caused by fragments  2 mm. Moreover, residues of PBAT and PLA were found in all investigated compost samples, including the finished compost from plant #4, which had shown no contamination by larger BPD fragments (Table 1). The pre-compost from that plant had shown a few contaminating BDP fragments in the > 5 mm fraction. However, in regard to the fragments More