High-quality genome assembly enables prediction of allele-specific gene expression in hybrid poplar.

Poplar (Populus) is a well-established model system for tree genomics and molecular breeding, and hybrid poplar is widely used in forest plantations. However, distinguishing its diploid homologous chromosomes is difficult, complicating advanced functional studies on specific alleles. In this study, we applied a trio-binning design and PacBio high-fidelity long-read sequencing to obtain haplotype-phased telomere-to-telomere genome assemblies for the 2 parents of the well-studied F1 hybrid "84K" (Populus alba × Populus tremula var. glandulosa). Almost all chromosomes, including the telomeres and centromeres, were completely assembled for each haplotype subgenome apart from 2 small gaps on one chromosome. By incorporating information from these haplotype assemblies and extensive RNA-seq data, we analyzed gene expression patterns between the 2 subgenomes and alleles. Transcription bias at the subgenome level was not uncovered, but extensive-expression differences were detected between alleles. We developed machine-learning (ML) models to predict allele-specific expression (ASE) with high accuracy and identified underlying genome features most highly influencing ASE. One of our models with 15 predictor variables achieved 77% accuracy on the training set and 74% accuracy on the testing set. ML models identified gene body CHG methylation, sequence divergence, and transposon occupancy both upstream and downstream of alleles as important factors for ASE. Our haplotype-phased genome assemblies and ML strategy highlight an avenue for functional studies in Populus and provide additional tools for studying ASE and heterosis in hybrids.

Evolutionary radiation of the Eurasian Pinus species under pervasive gene flow.

Evolutionary radiation, a pivotal aspect of macroevolution, offers valuable insights into evolutionary processes. The genus Pinus is the largest genus in conifers with c . 90% of the extant species emerged in the Miocene, which signifies a case of rapid diversification. Despite this remarkable history, our understanding of the mechanisms driving radiation within this expansive genus has remained limited. Using exome capture sequencing and a fossil-calibrated phylogeny, we investigated the divergence history, niche diversification, and introgression among 13 closely related Eurasian species spanning climate zones from the tropics to the boreal Arctic. We detected complex introgression among lineages in subsection Pinus at all stages of the phylogeny. Despite this widespread gene exchange, each species maintained its genetic identity and showed clear niche differentiation. Demographic analysis unveiled distinct population histories among these species, which further influenced the nucleotide diversity and efficacy of purifying and positive selection in each species. Our findings suggest that radiation in the Eurasian pines was likely fueled by interspecific recombination and further reinforced by their adaptation to distinct environments. Our study highlights the constraints and opportunities for evolutionary change, and the expectations of future adaptation in response to environmental changes in different lineages.

Scots pine - panmixia and the elusive signal of genetic adaptation.

Scots pine is the foundation species of diverse forested ecosystems across Eurasia and displays remarkable ecological breadth, occurring in environments ranging from temperate rainforests to arid tundra margins. Such expansive distributions can be favored by various demographic and adaptive processes and the interactions between them. To understand the impact of neutral and selective forces on genetic structure in Scots pine, we conducted range-wide population genetic analyses on 2321 trees from 202 populations using genotyping-by-sequencing, reconstructed the recent demography of the species and examined signals of genetic adaptation. We found a high and uniform genetic diversity across the entire range (global FST 0.048), no increased genetic load in expanding populations and minor impact of the last glacial maximum on historical population sizes. Genetic-environmental associations identified only a handful of single-nucleotide polymorphisms significantly linked to environmental gradients. The results suggest that extensive gene flow is predominantly responsible for the observed genetic patterns in Scots pine. The apparent missing signal of genetic adaptation is likely attributed to the intricate genetic architecture controlling adaptation to multi-dimensional environments. The panmixia metapopulation of Scots pine offers a good study system for further exploration into how genetic adaptation and plasticity evolve under gene flow and changing environment.

Evolutionary origin and establishment of a dioecious diploid-tetraploid complex.

Polyploids recurrently emerge in angiosperms, but most polyploids are likely to go extinct before establishment due to minority cytotype exclusion, which may be specifically a constraint for dioecious plants. Here we test the hypothesis that a stable sex-determination system and spatial/ecological isolation facilitate the establishment of dioecious polyploids. We determined the ploidy levels of 351 individuals from 28 populations of the dioecious species Salix polyclona, and resequenced 190 individuals of S. polyclona and related taxa for genomic diversity analyses. The ploidy survey revealed a frequency 52% of tetraploids in S. polyclona, and genomic k-mer spectra analyses suggested an autopolyploid origin for them. Comparisons of diploid male and female genomes identified a female heterogametic sex-determining factor on chromosome 15, which probably also acts in the dioecious tetraploids. Phylogenetic analyses revealed two diploid clades and a separate clade/grade of tetraploids with a distinct geographic distribution confined to western and central China, where complex mountain systems create higher levels of environmental heterogeneity. Fossil-calibrated phylogenies showed that the polyploids emerged during 7.6-2.3 million years ago, and population demographic histories largely matched the geological and climatic history of the region. Our results suggest that inheritance of the sex-determining system from the diploid progenitor as intrinsic factor and spatial isolation as extrinsic factor may have facilitated the preservation and establishment of polyploid dioecious populations.

Effects of landscapes and range expansion on population structure and local adaptation.

Understanding the origin and distribution of genetic diversity across landscapes is critical for predicting the future of organisms in changing climates. This study investigated how adaptive and demographic forces have shaped diversity and population structure in Pinus densata, a keystone species on Qinghai-Tibetan Plateau (QTP). We examined the distribution of genomic diversity across the range of P. densata using exome capture sequencing. We applied spatially explicit tests to dissect the impacts of allele surfing, geographic isolation and environmental gradients on population differentiation and forecasted how this genetic legacy may limit the persistence of P. densata in future climates. We found that allele surfing from range expansion could explain the distribution of 39% of the c. 48 000 genotyped single nucleotide polymorphisms (SNPs). Uncorrected, these allele frequency clines severely confounded inferences of selection. After controlling for demographic processes, isolation-by-environment explained 9.2-19.5% of the genetic structure, with c. 4.0% of loci being affected by selection. Allele surfing and genotype-environment associations resulted in genomic mismatch under projected climate scenarios. We illustrate that significant local adaptation, when coupled with reduced diversity as a result of demographic history, constrains potential evolutionary response to climate change. The strong signal of genomic vulnerability in P. densata may be representative for other QTP endemics.

Parentage and relatedness reconstruction in Pinus sylvestris using genotyping-by-sequencing.

Estimating kinship is fundamental for studies of evolution, conservation, and breeding. Genotyping-by-sequencing (GBS) and other restriction based genotyping methods have become widely applied in these applications in non-model organisms. However, sequencing errors, depth, and reproducibility between library preps could potentially hinder accurate genetic inferences. In this study, we tested different sets of parameters in data filtering, different reference populations and eight estimation methods to obtain a robust procedure for relatedness estimation in Scots pine (Pinus sylvestris L.). We used a seed orchard as our study system, where candidate parents are known and pedigree reconstruction can be compared with theoretical expectations. We found that relatedness estimates were lower than expected for all categories of kinship estimated if the proportion of shared SNPs was low. However, estimates reached expected values if loci showing an excess of heterozygotes were removed and genotyping error rates were considered. The genetic variance-covariance matrix (G-matrix) estimation, however, performed poorly in kinship estimation. The reduced relatedness estimates are likely due to false heterozygosity calls. We analyzed the mating structure in the seed orchard and identified a selfing rate of 3% (including crosses between clone mates) and external pollen contamination of 33.6%. Little genetic structure was observed in the sampled Scots pine natural populations, and the degree of inbreeding in the orchard seed crop is comparable to natural stands. We illustrate that under our optimized data processing procedure, relatedness, and genetic composition, including level of pollen contamination within a seed orchard crop, can be established consistently by different estimators.

Interspecific Plastome Recombination Reflects Ancient Reticulate Evolution in Picea (Pinaceae).

Plastid sequences are a cornerstone in plant systematic studies and key aspects of their evolution, such as uniparental inheritance and absent recombination, are often treated as axioms. While exceptions to these assumptions can profoundly influence evolutionary inference, detecting them can require extensive sampling, abundant sequence data, and detailed testing. Using advancements in high-throughput sequencing, we analyzed the whole plastomes of 65 accessions of Picea, a genus of ∼35 coniferous forest tree species, to test for deviations from canonical plastome evolution. Using complementary hypothesis and data-driven tests, we found evidence for chimeric plastomes generated by interspecific hybridization and recombination in the clade comprising Norway spruce (P. abies) and 10 other species. Support for interspecific recombination remained after controlling for sequence saturation, positive selection, and potential alignment artifacts. These results reconcile previous conflicting plastid-based phylogenies and strengthen the mounting evidence of reticulate evolution in Picea. Given the relatively high frequency of hybridization and biparental plastid inheritance in plants, we suggest interspecific plastome recombination may be more widespread than currently appreciated and could underlie reported cases of discordant plastid phylogenies.

Evolution and Function of the Populus SABATH Family Reveal That a Single Amino Acid Change Results in a Substrate Switch.

Evolutionary mechanisms of substrate specificities of enzyme families remain poorly understood. Plant SABATH methyltransferases catalyze methylation of the carboxyl group of various low molecular weight metabolites. Investigation of the functional diversification of the SABATH family in plants could shed light on the evolution of substrate specificities in this enzyme family. Previous studies identified 28 SABATH genes from the Populus trichocarpa genome. In this study, we re-annotated the Populus SABATH gene family, and performed molecular evolution, gene expression and biochemical analyses of this large gene family. Twenty-eight Populus SABATH genes were divided into three classes with distinct divergences in their gene structure, expression responses to abiotic stressors and enzymatic properties of encoded proteins. Populus class I SABATH proteins converted IAA to methyl-IAA, class II SABATH proteins converted benzoic acid (BA) and salicylic acid (SA) to methyl-BA and methyl-SA, while class III SABATH proteins converted farnesoic acid (FA) to methyl-FA. For Populus class II SABATH proteins, both forward and reverse mutagenesis studies showed that a single amino acid switch between PtSABATH4 and PtSABATH24 resulted in substrate switch. Our findings provide new insights into the evolution of substrate specificities of enzyme families.

The mortality risk factor of community acquired pneumonia patients with chronic obstructive pulmonary disease: a retrospective cohort study.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is one of the most common comorbidities in community acquired pneumonia (CAP) patients. We aimed to investigate the characteristics and mortality risk factors of COPD patients hospitalized with CAP. METHODS: A retrospective cohort study was conducted at Shanghai Pulmonary Hospital and Shanghai Dahua Hospital. Clinical and demographic data in patients diagnosed with CAP were collected between January 2015 and June 2016. Logistic regression analysis was performed to screen mortality risk factors of COPD patients hospitalized with CAP. RESULTS: Of the total 520 CAP patients, 230 (44.2%) patients had been diagnosed comorbid with COPD (COPD-CAP). CAP patients comorbid with COPD patients had higher rate of need for ICU admission (18.3% vs 13.1%) and need for NIMV (26.1% vs 1.4%) than without COPD (nCOPD-CAP). The PSI, CURB-65 and APACHE-II scores in COPD-CAP patients were higher than that in nCOPD-CAP patients (95 vs 79, P < 0.001; 1 vs 1, P < 0.001; 13 vs 8, P < 0.001, respectively). Logistic regression analysis indicated that aspiration, D-dimer > 2.0 µg/mL and CURB-65 ≥ 3 were risk factors associated with in-hospital mortality ((odd ratio) OR = 5.678, OR = 4.268, OR = 20.764, respectively) in COPD-CAP patients. The risk factors associated with 60-day mortality in COPD-CAP patients were comorbid with coronary heart disease, aspiration, need for NIMV (non-invasive mechanical ventilation) and CURB-65 ≥ 3 (OR = 5.206, OR = 7.921, OR = 3.974, OR = 18.002, respectively). CONCLUSIONS: COPD patients hospitalized with CAP had higher rate of need for NIMV, need for ICU admission and severity scores than those without COPD. Aspiration, D-dimer > 2.0 µg/mL, comorbid with coronary heart disease, need for NIMV and CURB-65 ≥ 3 were mortality risk factors in CAP patients comorbid with COPD.

Subcellular Relocalization and Positive Selection Play Key Roles in the Retention of Duplicate Genes of Populus Class III Peroxidase Family.

Gene duplication is the primary source of new genes and novel functions. Over the course of evolution, many duplicate genes lose their function and are eventually removed by deletion. However, some duplicates have persisted and evolved diverse functions. A particular challenge is to understand how this diversity arises and whether positive selection plays a role. In this study, we reconstructed the evolutionary history of the class III peroxidase (PRX) genes from the Populus trichocarpa genome. PRXs are plant-specific enzymes that play important roles in cell wall metabolism and in response to biotic and abiotic stresses. We found that two large tandem-arrayed clusters of PRXs evolved from an ancestral cell wall type PRX to vacuole type, followed by tandem duplications and subsequent functional specification. Substitution models identified seven positively selected sites in the vacuole PRXs. These positively selected sites showed significant effects on the biochemical functions of the enzymes. We also found that positive selection acts more frequently on residues adjacent to, rather than directly at, a critical active site of the enzyme, and on flexible regions rather than on rigid structural elements of the protein. Our study provides new insights into the adaptive molecular evolution of plant enzyme families.

Structural and functional evolution of positively selected sites in pine glutathione S-transferase enzyme family.

Phylogenetic analyses have identified positive selection as an important driver of protein evolution, both structural and functional. However, the lack of appropriate combined functional and structural assays has generally hindered attempts to elucidate patterns of positively selected sites and their effects on enzyme activity and substrate specificity. In this study we investigated the evolutionary divergence of the glutathione S-transferase (GST) family in Pinus tabuliformis, a pine that is widely distributed from northern to central China, including cold temperate and drought-stressed regions. GSTs play important roles in plant stress tolerance and detoxification. We cloned 44 GST genes from P. tabuliformis and found that 26 of the 44 belong to the largest (Tau) class of GSTs and are differentially expressed across tissues and developmental stages. Substitution models identified five positively selected sites in the Tau GSTs. To examine the functional significance of these positively selected sites, we applied protein structural modeling and site-directed mutagenesis. We found that four of the five positively selected sites significantly affect the enzyme activity and specificity; thus their variation broadens the GST family substrate spectrum. In addition, positive selection has mainly acted on secondary substrate binding sites or sites close to (but not directly at) the primary substrate binding site; thus their variation enables the acquisition of new catalytic functions without compromising the protein primary biochemical properties. Our study sheds light on selective aspects of the functional and structural divergence of the GST family in pine and other organisms.

Mitochondrial DNA capture and divergence in Pinus provide new insights into the evolution of the genus.

The evolution of the mitochondrial (mt) genome is far from being fully understood. Systematic investigations into the modes of inheritance, rates and patterns of recombination, nucleotide substitution, and structural changes in the mt genome are still lacking in many groups of plants. In this study, we sequenced >11kbp mtDNA segments from multiple accessions of 36 pine species to characterize the evolutionary patterns of mtDNA in the genus Pinus. We found extremely low substitution rates and complex repetitive sequences scattered across different genome regions, as well as chimeric structures that were probably generated by multiple intergenomic recombinations. The mtDNA-based phylogeny of the genus differed from that based on chloroplast and nuclear DNA in the placement of several groups of species. Such discordances suggest a series of mtDNA capture events during past range shifts of the pine species and that both vertical and horizontal inheritance are implicated in the evolution of mtDNA in Pinus. MtDNA dating revealed that most extant lineages of the genus originated during Oligocene-Miocene radiation and subgenus Strobus diversified earlier than subgenus Pinus. Our findings illustrate a reticular evolutionary pathway for the mt genome through capture and recombination in the genus Pinus, and provide new insights into the evolution of the genus.

[Determination of arsenic speciation in Scomberomorus niphonius by capillary electrophoresis-inductively coupled plasma mass spectrometry].

A method for the detection of arsenocholine (AsC), arsenobetaine (AsB), As(III), dimethylarsinic (DMA), monomethylarsonic (MMA) and As (V) by capillary electrophoresis-inductively coupled plasma mass spectrometry (CE-ICP-MS) was established. The results showed that the six species of arsenic were separated within 20 min under the optimized conditions. Good linearities of 6 arsenic species were observed in the range from 2 to 50 µg x L(-1) with the linear correlation greater than 0.996, the detection limits were 0.10-1.08 µg x L(-1) and the RSDs (n = 5) of the peak areas were smaller than 7%. The method was successfully adopted to the determination of the species in Scomberomorus niphonius. The recoveries were between 93% and 98%, and we found the arsenobetaine (AsB) was the main species in the sample. The method was suitable for the analysis of other biological samples with the advantages of good stability, less sample consumption, short analysis time and convenience.

[Study on fluorescence labeling and determination of polypeptide (PC2~PC6) by high performance liquid chromatography].

This study was based on the thiol groups (-SH) of PC2~PC6, which could be reacted with the Monobromobimane (mBBr), in order to get polypeptide derivatives with fluorescent signal. A new method was developed for measuring the Polypeptides by high performance liquid chromatography with fluorescence detector, then the chromatographic conditions of HPLC was optimized; meawhile the reaction proportion of PCs and mBBr was identified by Trap-MS. The results showed that, the reaction proportion of PCs and mBBr was 1:1, the polypeptide derivatives had good stability; the five compounds separation was better, and the peak time focused on the 16.6~22.0 min; the linear correlation coefficient of PC2, PC3, PC4, PC5 and PC6 was >0.9991, and the limits of quantification were 0.3, 0.05, 0.3, 0.5 and 0.8 mg · L(-1) respectively, the recovery rate was 83.0%-102.0%; the method was reproducible, RSD<2%, this method for measuring the peptide compounds was rapid and accurate.

Comprehensive analysis of metal(loid)s and associated metal(loid) resistance genes in atmospheric particulate matter.

Despite global concerns about metal(loid)s in atmospheric particulate matter (PM), the presence of metal(loid) resistance genes (MRGs) in PM remains unknown. Therefore, we conducted a comprehensive investigation of the metal(loid)s and associated MRGs in PMs in two seasons (summer and winter) in Xiamen, China. According to the geoaccumulation index (Igeo), most metal(loid)s, except for V and Mn, exhibited enrichment in PM, suggesting potential anthropogenic sources. By employing Positive Matrix Factorization (PMF) model, utilizing a dataset encompassing both total and bioaccessible metal(loid)s, along with backward trajectory simulations, traffic emissions were determined to be the primary potential contributor of metal(loid)s in summer, whereas coal combustion was observed to have a dominant contribution in winter. The major contributor to the carcinogenic risk of metal(loid)s in both summer and winter was predominantly attributed to coal combustion, which serves as the main source of bioaccessible Cr. Bacterial communities within PMs showed lower diversity and network complexity in summer than in winter, with Pseudomonadales being the dominant order. Abundant MRGs, including the As(III) S-adenosylmethionine methyltransferase gene (arsM), Cu(I)-translocating P-type ATPase gene (copA), Zn(II)/Cd(II)/Pb(II)-translocating P-type ATPase gene (zntA), and Zn(II)-translocating P-type ATPase gene (ziaA), were detected within the PMs. Seasonal variations were observed for the metal(loid) concentration, bacterial community structure, and MRG abundance. The bacterial community composition and MRG abundance within PMs were primarily influenced by temperature, rather than metal(loid)s. This research offers novel perspectives on the occurrence of metal(loid)s and MRGs in PMs, thereby contributing to the control of air pollution.

Molecular evolution and expression divergence of the Populus polygalacturonase supergene family shed light on the evolution of increasingly complex organs in plants.

Plant polygalacturonases (PGs) are involved in cell separation processes during many stages of plant development. Investigation into the diversification of this large gene family in land plants could shed light on the evolution of structural development. We conducted whole-genome annotation, molecular evolution and gene expression analyses of PG genes in five species of land plant: Populus, Arabidopsis, rice, Selaginella and Physcomitrella. We identified 75, 44, 16 and 11 PG genes from Populus, rice, Selaginella and Physcomitrella genomes, respectively, which were divided into three classes. We inferred rapid expansion of class I PG genes in Populus, Arabidopsis and rice, while copy numbers of classes II and III PG genes were relatively conserved in all five species. Populus, Arabidopsis and rice class I PG genes were under more relaxed selection constraints than class II PG genes, while this selective pressure divergence was not observed in Selaginella and Physcomitrella PG families. In addition, class I PG genes underwent marked expression divergence in Populus, rice and Selaginella. Our results suggest that PG gene expansion occurred after the divergence of the lycophytes and euphyllophytes, and this expansion was likely paralleled by the evolution of increasingly complex organs in land plants.

Antidepressant-like effects of essential oil and asarone, a major essential oil component from the rhizome of Acorus tatarinowii.

CONTEXT: Depression is one of the most common psychiatric diseases. Acorus tatarinowii Schott (Araceae) has shown many bioactivities in treatment of senile dementia and epilepsy. However, there is no report on antidepressant-like effects of the essential oil (EO) and its major components on animals under standardized experimental procedures. OBJECTIVE: This study was designed to investigate the antidepressant properties of EO and asarones from the rhizomes of A. tatarinowii. MATERIALS AND METHODS: Gas chromatography-mass spectrometry (GC/MS) was used to determine the composition of EO. The forced swimming test (FST), tail suspension test (TST) and open-field test (OFT) were used to evaluate the antidepressant-like effects of EO and asarones. EO [30, 60, 120 or 240 mg/kg, per os (p.o.)], asarones (α-asarone and ß-asarone) [5, 10 and 20 mg/kg, intraperitoneal (i.p.)] and imipramine (15 mg/kg, i.p.) were administered at 1 h, 30 min and 30 min before the test, respectively. RESULTS: From the results of GC/MS, it was found that the main components of the EO were α-asarone (9.18%) and ß-asarone (68.9%). From the results of FST and TST, the immobility time can be reduced to 166 ± 17 s (p < 0.01) and 146 ± 15 s (p < 0.05) by EO at the dose of 120 mg/kg. Moreover, significant antidepressant-like effects were shown by α-asarone with the immobility time of 178 ± 15 s (p < 0.05) and 159 ± 17 s (p < 0.01) in FST, or 147 ± 12 (p < 0.05) and 134 ± 12 s (p < 0.01) in TST at the dose of 10 and 20 mg/kg. ß-Asarone also displayed antidepressant-like effects with an immobility time of 179 ± 18 s (p < 0.05) in FST or 142 ± 14 (p < 0.05) in TST at 20 mg/kg. However, no change in ambulation was observed in the OFT. CONCLUSION: The results obtained indicate that the EO and asarones from the rhizomes of A. tatarinowii can be considered as a new therapeutic agent for curing depression.

Genomic clines across the species boundary between a hybrid pine and its progenitor in the eastern Tibetan Plateau.

Most species have clearly defined distribution ranges and ecological niches. The genetic and ecological causes of species differentiation and the mechanisms that maintain species boundaries between newly evolved taxa and their progenitors are, however, less clearly defined. This study investigated the genetic structure and clines in Pinus densata, a pine of hybrid origin on the southeastern Tibetan Plateau, to gain an understanding of the contemporary dynamics of species barriers. We analyzed genetic diversity in a range-wide collection of P. densata and representative populations of its progenitors, Pinus tabuliformis and Pinus yunnanensis, using exome capture sequencing. We detected four distinct genetic groups within P. densata that reflect its migration history and major gene-flow barriers across the landscape. The demographies of these genetic groups in the Pleistocene were associated with regional glaciation histories. Interestingly, population sizes rebounded rapidly during interglacial periods, suggesting persistence and resilience of the species during the Quaternary ice age. In the contact zone between P. densata and P. yunnanensis, 3.36% of the analyzed loci (57 849) showed exceptional patterns of introgression, suggesting their potential roles in either adaptive introgression or reproductive isolation. These outliers showed strong clines along critical climate gradients and enrichment in a number of biological processes relevant to high-altitude adaptation. This indicates that ecological selection played an important role in generating genomic heterogeneity and a genetic barrier across a zone of species transition. Our study highlights the forces that operate to maintain species boundaries and promote speciation in the Qinghai-Tibetan Plateau and other mountain systems.

Inference of the distribution of fitness effects of mutations is affected by single nucleotide polymorphism filtering methods, sample size and population structure.

The distribution of fitness effects (DFE) of new mutations has been of interest to evolutionary biologists since the concept of mutations arose. Modern population genomic data enable us to quantify the DFE empirically, but few studies have examined how data processing, sample size and cryptic population structure might affect the accuracy of DFE inference. We used simulated and empirical data (from Arabidopsis lyrata) to show the effects of missing data filtering, sample size, number of single nucleotide polymorphisms (SNPs) and population structure on the accuracy and variance of DFE estimates. Our analyses focus on three filtering methods-downsampling, imputation and subsampling-with sample sizes of 4-100 individuals. We show that (1) the choice of missing-data treatment directly affects the estimated DFE, with downsampling performing better than imputation and subsampling; (2) the estimated DFE is less reliable in small samples (<8 individuals), and becomes unpredictable with too few SNPs (<5000, the sum of 0- and 4-fold SNPs); and (3) population structure may skew the inferred DFE towards more strongly deleterious mutations. We suggest that future studies should consider downsampling for small data sets, and use samples larger than 4 (ideally larger than 8) individuals, with more than 5000 SNPs in order to improve the robustness of DFE inference and enable comparative analyses.

Eco-friendly approach for enhancing the floatability of non-metallic components in waste printed circuit boards: Adding gutter oil during dry grinding.

Waste printed circuit boards (WPCBs) are an attractive secondary resource that is challenging to dispose of due to its complexity. Reverse flotation is an effective method to remove non-metallic particles (NMPs) to obtain metals from WPCBs. Nevertheless, the removal of NMPs is usually inadequate in the present flotation practice. Thus, to provide a clean approach to improve the removal efficiency of NMPs, the method of adding gutter oil during dry grinding process was adopted to enhance the hydrophobic sites on the surface of NMPs to improve the floatability. The surface morphology of NMPs was analyzed by SEM, the results show that the rough morphology inhibited the adhesion of bubbles, while water occupied the cracks and pores, making it challenging for collector adsorption, which result in unstable particle-bubble adhesion. The results of FTIR indicate that both NMPs and gutter oil have -CH3, -CH2, -C = O, -C-O functional groups, which promotes the adsorption of gutter oil on the surface of NMPs. The contact angle (CA) results show that the adsorption of gutter oil on the particle surface is conducive to the formation of enhanced CA. Furthermore, the flotation enhancement effect was verified by flotation kinetic experiments. The accumulated floats yield of NMPs conditioned by gutter oil during grinding is increased from 67.05% (NMPs without conditioning) to 95.02%, and the resin recovery is increased by 31.10%. It is demonstrated that dry grinding with gutter oil can strengthen the floatability of NMPs, which provides a potential approach to increase the flotation efficiency of WPCBs.