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The radial growth of trees plays a crucial role in determining forest carbon sequestration capacity. Understanding the growth dynamics of trees and their response to environmental factors is essential for predicting forest's carbon sink potential under future climate change. Coniferous forest trees are particularly sensitive to climate change, with growth dynamics responding rapidly to environmental shifts. We collected and analyzed data from 99 papers published between 1975 and 2023, and examined the effects of exogenous factors (such as temperature, water, and photoperiod) and endogenous factors (including tree age and species) on cambial activity and radial growth in conifers. We further explored the mechanisms underlying these effects. The results showed that climate warming had the potential to advance the onset while delayed the end of xylem differentiation stages in conifers in temperate and boreal regions. Water availability played a crucial role in regulating the timing of cambial phenology and wood formation by influencing water potential and cell turgor. Additionally, the photoperiod not only participated in regulating the start and end times of growth, but also influenced the timing of maximum growth rate occurrence. Future climate warming was expected to extend the growing season, leading to increase in growth of conifers in boreal regions and expanding forests to higher altitudes or latitudes. However, changes in precipitation patterns and increased evapotranspiration resulting from temperature increases might advance the end of growing season and reduce growth rate in arid areas. To gain a more comprehensive understanding of the relationship between radial growth and climatic factors, it is necessary to develop process-based models to elucidate the physiological mechanisms underlying wood formation and the response of trees to climatic factors.

T cell-redirecting antibody for treatment of solid tumors via targeting mesothelin.

T cell engaging bispecific antibodies (TCBs) have recently become significant in cancer treatment. In this study we developed MSLN490, a novel TCB designed to target mesothelin (MSLN), a glycosylphosphatidylinositol (GPI)-linked glycoprotein highly expressed in various cancers, and evaluated its efficacy against solid tumors. CDR walking and phage display techniques were used to improve affinity of the parental antibody M912, resulting in a pool of antibodies with different affinities to MSLN. From this pool, various bispecific antibodies (BsAbs) were assembled. Notably, MSLN490 with its IgG-[L]-scFv structure displayed remarkable anti-tumor activity against MSLN-expressing tumors (EC50: 0.16 pM in HT-29-hMSLN cells). Furthermore, MSLN490 remained effective even in the presence of non-membrane-anchored MSLN (soluble MSLN). Moreover, the anti-tumor activity of MSLN490 was enhanced when combined with either Atezolizumab or TAA × CD28 BsAbs. Notably, a synergistic effect was observed between MSLN490 and paclitaxel, as paclitaxel disrupted the immunosuppressive microenvironment within solid tumors, enhancing immune cells infiltration and improved anti-tumor efficacy. Overall, MSLN490 exhibits robust anti-tumor activity, resilience to soluble MSLN interference, and enhanced anti-tumor effects when combined with other therapies, offering a promising future for the treatment of a variety of solid tumors. This study provides a strong foundation for further exploration of MSLN490's clinical potential.

A novel bispecific antibody drug conjugate targeting HER2 and HER3 with potent therapeutic efficacy against breast cancer.

Antibody drug conjugate (ADC) therapy has become one of the most promising approaches in cancer immunotherapy. Bispecific targeting could enhance the efficacy and safety of ADC by improving its specificity, affinity and internalization. In this study we constructed a HER2/HER3-targeting bispecific ADC (BsADC) and characterized its physiochemical properties, target specificity and internalization in vitro, and assessed its anti-tumor activities in breast cancer cell lines and in animal models. The HER2/HER3-targeting BsADC had a drug to antibody ratio (DAR) of 2.89, displayed a high selectivity against the target JIMT-1 breast cancer cells in vitro, as well as a slightly higher level of internalization than HER2- or HER3-monospecific ADCs. More importantly, the bispecific ADC potently inhibited the viability of MCF7, JIMT-1, BT474, BxPC-3 and SKOV-3 cancer cells in vitro. In JIMT-1 breast cancer xenograft mice, a single injection of bispecific ADC (3 mg/kg, i.v.) significantly inhibited the tumor growth with an efficacy comparable to that caused by combined injection of HER2 and HER3-monospecific ADCs (3 mg/kg for each). Our study demonstrates that the bispecific ADC concept can be applied to development of more potent new cancer therapeutics than the monospecific ADCs.

Genome-Wide Association Study Reveals Polygenic Architecture for Limber Pine Quantitative Disease Resistance to White Pine Blister Rust.

Development of durable resistance effective against a broad range of pathotypes is crucial for restoration of pathogen-damaged ecosystems. This study dissected the complex genetic architecture for limber pine quantitative disease resistance (QDR) to Cronartium ribicola using a genome-wide association study. Eighteen-month-old seedlings were inoculated for resistance screening under controlled conditions. Disease development was quantitatively assessed for QDR-related traits over 4 years postinoculation. To reveal the genomic architecture contributing to QDR-related traits, a set of genes related to disease resistance with genome-wide distribution was selected for targeted sequencing for genotyping of single-nucleotide polymorphisms (SNPs). The genome-wide association study revealed a set of SNPs significantly associated with quantitative traits for limber pine QDR to white pine blister rust, including number of needle spots and stem cankers, as well as survival 4 years postinoculation. The peaks of marker-trait associations displayed a polygenic pattern, with genomic regions as potential resistant quantitative trait loci, distributed over 10 of the 12 linkage groups (LGs) of Pinus. None of them was linked to the Cr4-controlled major gene resistance previously mapped on LG08. Both normal canker and bole infection were mapped on LG05, and the associated SNPs explained their phenotypic variance up to 52%, tagging a major resistant quantitative trait locus. Candidate genes containing phenotypically associated SNPs encoded putative nucleotide-binding site leucine-rich repeat proteins, leucine-rich repeat-receptor-like kinase, cytochrome P450 superfamily protein, heat shock cognate protein 70, glutamate receptor, RNA-binding family protein, and unknown protein. The confirmation of resistant quantitative trait loci broadens the genetic pool of limber pine resistance germplasm for resistance breeding.

Quality appraisal of clinical practice guidelines for motor neuron diseases or related disorders using the AGREE II instrument.

Objectives: This study aimed to systematically assess the quality of CPGs for motor neuron diseases (MNDs) or related disorders and identify the gaps that limit evidence-based practice. Methods: Four scientific databases and six guideline repositories were searched for eligible CPGs. Three researchers assessed the eligible CPGs using the Appraisal of Guidelines Research and Evaluation II instrument. The distribution of the level of evidence and strength of recommendation of these CPGs were determined. The univariate regression analysis was used to explore the characteristic factors affecting the quality of CPGs. Results: Fifteen CPGs met the eligibility criteria: 10 were for MND and 5 were for spinal muscular atrophy. The mean overall rating score was 44.5%, and only 3 of 15 CPGs were of high quality. The domains that achieved low mean scores were applicability (24.4%), rigor of development (39.9%), and stakeholder involvement (40.3%). Most recommendations were based on low-quality evidence and had a weak strength. The CPGs that were updated, meant for adults, and evidence based, and used a CPG quality tool and a grading system were associated with higher scores in certain specific domains and overall rating. Conclusion: The overall quality of CPGs for MNDs or related disorders was poor and recommendations were largely based on low-quality evidence. Many areas still need improvement to develop high-quality CPGs, and the use of CPG quality tools should be emphasized. A great deal of research on MNDs or related disorders is still needed to fill the large evidence gap.

Combination of vonoprazan and amoxicillin as the first-line Helicobacter pylori eradication therapy: a multicenter, prospective, randomized, parallel-controlled study.

The eradication rate of Helicobacter pylori (H. pylori) decreased gradually. This study aimed to analyze the efficacy and safety of a 14-day combination of vonoprazan and amoxicillin as the first-line eradication therapy for H. pylori infection and compared them with those of the bismuth quadruple therapy. A prospective randomized clinical trial (RCT) was designed, involving patients with H. pylori infection in 6 institutions who did not receive any treatment yet. They were randomly assigned into the VA-dual group (vonoprazan 20 mg b.i.d + amoxicillin 750 mg q.i.d) or EACP-quadruple group (esomeprazole 20 mg + amoxicillin 1000 mg + clarithromycin 500 mg + colloidal bismuth subcitrate 220 mg b.i.d) for 14 days in a ratio of 1:1. At least 28 days later, the eradication rate was detected by the 13C-urea breath test (UBT). A total of 562 patients from February 2022 to September 2022 were enrolled and 316 were random. In the ITT analysis, the eradication rates of H. pylori in the VA-dual group and EACP-quadruple group were 89.9% and 81.0%, respectively, p = 0.037. In the PP analysis were 97.9% and 90.8%, p = 0.009. The different eradication rate was 8.9% (95% CI 1.2-16.5%) and 7.2% (95% CI 1.8-12.4%) in ITT and PP analyses, both lower limit of the 95%CI was still higher than the prespecified margin. In addition, the incidence of adverse events in the VA-dual group was significantly lower than that in the EACP-quadruple group (19.0% vs. 43.0%, P < 0.001). The efficacy and safety of a 14-day combination therapy of vonoprazan and amoxicillin in eradicating H. pylori are superior to bismuth quadruple therapy, and this combination significantly reduces the use of antibiotics.

Long-term passaging of pseudo-typed SARS-CoV-2 reveals the breadth of monoclonal and bispecific antibody cocktails.

The continuous emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants poses challenges to the effectiveness of neutralizing antibodies. Rational design of antibody cocktails is a realizable approach addressing viral immune evasion. However, evaluating the breadth of antibody cocktails is essential for understanding the development potential. Here, based on a replication competent vesicular stomatitis virus model that incorporates the spike of SARS-CoV-2 (VSV-SARS-CoV-2), we evaluated the breadth of a number of antibody cocktails consisting of monoclonal antibodies and bispecific antibodies by long-term passaging the virus in the presence of the cocktails. Results from over two-month passaging of the virus showed that 9E12 + 10D4 + 2G1 and 7B9-9D11 + 2G1 from these cocktails were highly resistant to random mutation, and there was no breakthrough after 30 rounds of passaging. As a control, antibody REGN10933 was broken through in the third passage. Next generation sequencing was performed and several critical mutations related to viral evasion were identified. These mutations caused a decrease in neutralization efficiency, but the reduced replication rate and ACE2 susceptibility of the mutant virus suggested that they might not have the potential to become epidemic strains. The 9E12 + 10D4 + 2G1 and 7B9-9D11 + 2G1 cocktails that picked from the VSV-SARS-CoV-2 system efficiently neutralized all current variants of concern and variants of interest including the most recent variants Delta and Omicron, as well as SARS-CoV-1. Our results highlight the feasibility of using the VSV-SARS-CoV-2 system to develop SARS-CoV-2 antibody cocktails and provide a reference for the clinical selection of therapeutic strategies to address the mutational escape of SARS-CoV-2.

Prognostic value of preoperative enhanced computed tomography as a quantitative imaging biomarker in pancreatic cancer.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies with high mortality and short survival time. Computed tomography (CT) plays an important role in the diagnosis, staging and treatment of pancreatic tumour. Pancreatic cancer generally shows a low enhancement pattern compared with normal pancreatic tissue. AIM: To analyse whether preoperative enhanced CT could be used to predict postoperative overall survival in patients with PDAC. METHODS: Sixty-seven patients with PDAC undergoing pancreatic resection were enrolled retrospectively. All patients underwent preoperative unenhanced and enhanced CT examination, the CT values of which were measured. The ratio of the preoperative CT value increase from the nonenhancement phase to the portal venous phase between pancreatic tumour and normal pancreatic tissue was calculated. The cut-off value of ratios was obtained by the receiver operating characteristic (ROC) curve of the tumour relative enhancement ratio (TRER), according to which patients were divided into low- and high-enhancement groups. Univariate and multivariate analyses were performed using Cox regression based on TRER grouping. Finally, the correlation between TRER and clinicopathological characteristics was analysed. RESULTS: The area under the curve of the ROC curve was 0.768 (P < 0.05), and the cut-off value of the ROC curve was calculated as 0.7. TRER ≤ 0.7 was defined as the low-enhancement group, and TRER > 0.7 was defined as the high-enhancement group. According to the TRER grouping, the Kaplan-Meier survival curve analysis results showed that the median survival (10.0 mo) with TRER ≤ 0.7 was significantly shorter than that (22.0 mo) with TRER > 0.7 (P < 0.05). In the univariate and multivariate analyses, the prognosis of patients with TRER ≤ 0.7 was significantly worse than that of patients with TRER > 0.7 (P < 0.05). Our results demonstrated that patients in the low TRER group were more likely to have higher American Joint Committee on Cancer stage, tumour stage and lymph node stage (all P < 0.05), and TRER was significantly negatively correlated with tumour size (P < 0.05). CONCLUSION: TRER ≤ 0.7 in patients with PDAC may represent a tumour with higher clinical stage and result in a shorter overall survival.

Transcriptomic Reprogramming and Genetic Variations Contribute to Western Hemlock Defense and Resistance Against Annosus Root and Butt Rot Disease.

Western hemlock (Tsuga heterophylla) is highly susceptible to Annosus root and butt rot disease, caused by Heterobasidion occidentale across its native range in western North America. Understanding molecular mechanisms of tree defense and dissecting genetic components underlying disease resistance will facilitate forest breeding and disease control management. The aim of this study was to profile host transcriptome reprogramming in response to pathogen infection using RNA-seq analysis. Inoculated seedlings were clearly grouped into three types: quantitative resistant (QR), susceptible (Sus), and un-infected (Uif), based on profiles of H. occidentale genes expressed in host tissues. Following de novo assembly of a western hemlock reference transcriptome with more than 33,000 expressed genes, the defensive transcriptome reprogramming was characterized and a set of differentially expressed genes (DEGs) were identified with gene ontology (GO) annotation. The QR seedlings showed controlled and coordinated molecular defenses against biotic stressors with enhanced biosynthesis of terpenoids, cinnamic acids, and other secondary metabolites. The Sus seedlings showed defense responses to abiotic stimuli with a few biological processes enhanced (such as DNA replication and cell wall organization), while others were suppressed (such as killing of cells of other organism). Furthermore, non-synonymous single nucleotide polymorphisms (ns-SNPs) of the defense- and resistance-related genes were characterized with high genetic variability. Both phylogenetic analysis and principal coordinate analysis (PCoA) revealed distinct evolutionary distances among the samples. The QR and Sus seedlings were well separated and grouped into different phylogenetic clades. This study provides initial insight into molecular defense and genetic components of western hemlock resistance against the Annosus root and butt rot disease. Identification of a large number of genes and their DNA variations with annotated functions in plant resistance and defense promotes the development of genomics-based breeding strategies for improved western hemlock resistance to H. occidentale.

A new pair of cytotoxic enantiomeric isoprenylated chromone derivatives from Pestalotiopsis sp.

A new pair of enantiomeric isoprenylated chromone derivatives, (±)-pestaloficiol X [(±)-1], along with a known compound pestaloficiol J (2), were isolated from the plant endophytic fungus Pestalotiopsis sp. The racemic mixture 1 was separated through chiral HPLC. The structures of new compounds (±)-1 were elucidated on the basis of extensive spectroscopic data and their absolute configurations were further configured through computational analysis of their electronic circular dichroism (ECD) spectra. Compound (+)-1 showed significant inhibitory potency against HL-60 and HEP-3B cell lines, with IC50 values of 1.35 ± 0.15 and 3.70 ± 0.33 µM, respectively, while compound (-)-1 showed significant inhibitory potency against HL-60 and HEP-3B cell lines, with IC50 values of 2.39 ± 0.26 and 2.99 ± 0.35 µM, respectively.

Comparative Association Mapping Reveals Conservation of Major Gene Resistance to White Pine Blister Rust in Southwestern White Pine (Pinus strobiformis) and Limber Pine (P. flexilis).

All native North American white pines are highly susceptible to white pine blister rust (WPBR) caused by Cronartium ribicola. Understanding genomic diversity and molecular mechanisms underlying genetic resistance to WPBR remains one of the great challenges in improvement of white pines. To compare major gene resistance (MGR) present in two species, southwestern white pine (Pinus strobiformis) Cr3 and limber pine (P. flexilis) Cr4, we performed association analyses of Cr3-controlled resistant traits using single nucleotide polymorphism (SNP) assays designed with Cr4-linked polymorphic genes. We found that ∼70% of P. flexilis SNPs were transferable to P. strobiformis. Furthermore, several Cr4-linked SNPs were significantly associated with the Cr3-controlled traits in P. strobiformis families. The most significantly associated SNP (M326511_1126R) almost colocalized with Cr4 on the Pinus consensus linkage group 8, suggesting that Cr3 and Cr4 might be the same R locus, or have localizations very close to each other in the syntenic region of the P. strobiformis and P. flexilis genomes. M326511_1126R was identified as a nonsynonymous SNP, causing amino acid change (Val376Ile) in a putative pectin acetylesterase, with coding sequences identical between the two species. Moreover, top Cr3-associated SNPs were further developed as TaqMan genotyping assays, suggesting their usefulness as marker-assisted selection (MAS) tools to distinguish genotypes between quantitative resistance and MGR. This work demonstrates the successful transferability of SNP markers between two closely related white pine species in the hybrid zone, and the possibility for deployment of MAS tools to facilitate long-term WPBR management in P. strobiformis breeding and conservation.

Fine dissection of limber pine resistance to Cronartium ribicola using targeted sequencing of the NLR family.

BACKGROUND: Proteins with nucleotide binding site (NBS) and leucine-rich repeat (LRR) domains (NLR) make up one of most important resistance (R) families for plants to resist attacks from various pathogens and pests. The available transcriptomes of limber pine (Pinus flexilis) allow us to characterize NLR genes and related resistance gene analogs (RGAs) in host resistance against Cronartium ribicola, the causal fungal pathogen of white pine blister rust (WPBR) on five-needle pines throughout the world. We previously mapped a limber pine major gene locus (Cr4) that confers complete resistance to C. ribicola on the Pinus consensus linkage group 8 (LG-8). However, genetic distribution of NLR genes as well as their divergence between resistant and susceptible alleles are still unknown. RESULTS: To identify NLR genes at the Cr4 locus, the present study re-sequenced a total of 480 RGAs using targeted sequencing in a Cr4-segregated seed family. Following a call of single nucleotide polymorphisms (SNPs) and genetic mapping, a total of 541 SNPs from 155 genes were mapped across 12 LGs. Three putative NLR genes were newly mapped in the Cr4 region, including one that co-segregated with Cr4. The tight linkage of NLRs with Cr4-controlled phenotypes was further confirmed by bulked segregation analysis (BSA) using extreme-phenotype genome-wide association study (XP-GWAS) for significance test. Local tandem duplication in the Cr4 region was further supported by syntenic analysis using the sugar pine genome sequence. Significant gene divergences have been observed in the NLR family, revealing that diversifying selection pressures are relatively higher in local duplicated genes. Most genes showed similar expression patterns at low levels, but some were affected by genetic background related to disease resistance. Evidence from fine genetic dissection, evolutionary analysis, and expression profiling suggests that two NLR genes are the most promising candidates for Cr4 against WPBR. CONCLUSION: This study provides fundamental insights into genetic architecture of the Cr4 locus as well as a set of NLR variants for marker-assisted selection in limber pine breeding. Novel NLR genes were identified at the Cr4 locus and the Cr4 candidates will aid deployment of this R gene in combination with other major/minor genes in the limber pine breeding program.

Comparative Transcriptomics and RNA-Seq-Based Bulked Segregant Analysis Reveals Genomic Basis Underlying Cronartium ribicola vcr2 Virulence.

Breeding programs of five-needle pines have documented both major gene resistance (MGR) and quantitative disease resistance (QDR) to Cronartium ribicola (Cri), a non-native, invasive fungal pathogen causing white pine blister rust (WPBR). WPBR is one of the most deadly forest diseases in North America. However, Cri virulent pathotypes have evolved and can successfully infect and kill trees carrying resistance (R) genes, including vcr2 that overcomes MGR conferred by the western white pine (WWP, Pinus monticola) R gene (Cr2). In the absence of a reference genome, the present study generated a vcr2 reference transcriptome, consisting of about 20,000 transcripts with 1,014 being predicted to encode secreted proteins (SPs). Comparative profiling of transcriptomes and secretomes revealed vcr2 was significantly enriched for several gene ontology (GO) terms relating to oxidation-reduction processes and detoxification, suggesting that multiple molecular mechanisms contribute to pathogenicity of the vcr2 pathotype for its overcoming Cr2. RNA-seq-based bulked segregant analysis (BSR-Seq) revealed genome-wide DNA variations, including about 65,617 single nucleotide polymorphism (SNP) loci in 7,749 polymorphic genes shared by vcr2 and avirulent (Avcr2) pathotypes. An examination of the distribution of minor allele frequency (MAF) uncovered a high level of genomic divergence between vcr2 and Avcr2 pathotypes. By integration of extreme-phenotypic genome-wide association (XP-GWAS) analysis and allele frequency directional difference (AFDD) mapping, we identified a set of vcr2-associated SNPs within functional genes, involved in fungal virulence and other molecular functions. These included six SPs that were top candidate effectors with putative activities of reticuline oxidase, proteins with common in several fungal extracellular membrane (CFEM) domain or ferritin-like domain, polysaccharide lyase, rds1p-like stress responsive protein, and two Cri-specific proteins without annotation. Candidate effectors and vcr2-associated genes provide valuable resources for further deciphering molecular mechanisms of virulence and pathogenicity by functional analysis and the subsequent development of diagnostic tools for monitoring the virulence landscape in the WPBR pathosystems.

In-vitro anti-fungal assay and association analysis reveal a role for the Pinus monticola PR10 gene (PmPR10-3.1) in quantitative disease resistance to white pine blister rust.

Pathogenesis-related (PR) proteins play important roles in plant defense response. However, functional investigation of PR10 genes is still limited and their physiological roles have not been conclusively characterized in biological processes of conifer trees. Here, we identified multiple novel members in the western white pine (Pinus monticola) PmPR10 family by bioinformatic mining available transcriptomic data. Phylogenetic analysis of protein sequences revealed four PR10 and two PR10-like clusters with a high synteny across different species of five-needle pines. Of 10 PmPR10 genes, PmPR10-3.1 was selected and expressed in Escherichia coli. The purified recombinant protein exhibited inhibitory effects on spore hyphal growth of fungal pathogens Cronartium ribicola, Phoma exigua, and Phoma argillacea by in-vitro anti-fungal analysis. Genetic variation analysis detected a total of 21 single nucleotide polymorphisms (SNPs) within PmPR10-3.1 in a collection of P. monticola seed families. A nonsynonymous SNP (t178g) showed significant association with relative levels of quantitative disease resistance (QDR), explaining about 8.7% of phenotypic variation as the peak value across all SNPs. Our results provide valuable insight into the genetic architecture underlying P. monticola QDR and imply that PmPR10-3.1 may function as an important component in conifer basal immunity for non-specific resistance to a wide spectrum of pathogens.

Experimental and molecular dynamics study into the surfactant effect upon coal wettability.

In order to improve the wettability and permeability of coal seams, the water injection efficiency of coal seams has to be boosted, the amount of dust generation has to be reduced, and coal and gas outburst must be prevented, and a surfactant is used to modulate the coal surface wettability. In this work, taking coal samples from Pingdingshan mine in Henan as the research object, their surface chemistry was initially scrutinized and then coal surface engineering via surfactants was inspected by a contact angle test. The coal wettability was ameliorated with surfactants, particularly using the 1 wt% non-ionic surfactant Triton X-100, which elicited a 47% lower contact angle than the raw coal. The surface free energy of the coal sample modified by 1.0 wt% Triton X-100 was increased from 44.51 mN m-1 to 49.52 mN m-1. The microstructural characteristics of coal samples allowed leveraging the Wiser model to construct three kinds of surfactant-coal adsorption models to dissect the adsorption configuration of the system. The results indicate that the addition of surfactants increases both the interaction of water with the coal and the diffusion coefficient of water molecules, resulting in the coal surface transformation from hydrophobicity to hydrophilicity. Our current work can provide salutary guidance and reference for coal water injection and dust suppression.

Association Mapping and Development of Marker-Assisted Selection Tools for the Resistance to White Pine Blister Rust in the Alberta Limber Pine Populations.

Since its introduction to North America in the early 1900s, white pine blister rust (WPBR) caused by the fungal pathogen Cronartium ribicola has resulted in substantial economic losses and ecological damage to native North American five-needle pine species. The high susceptibility and mortality of these species, including limber pine (Pinus flexilis), creates an urgent need for the development and deployment of resistant germplasm to support recovery of impacted populations. Extensive screening for genetic resistance to WPBR has been underway for decades in some species but has only started recently in limber pine using seed families collected from wild parental trees in the USA and Canada. This study was conducted to characterize Alberta limber pine seed families for WPBR resistance and to develop reliable molecular tools for marker-assisted selection (MAS). Open-pollinated seed families were evaluated for host reaction following controlled infection using C. ribicola basidiospores. Phenotypic segregation for presence/absence of stem symptoms was observed in four seed families. The segregation ratios of these families were consistent with expression of major gene resistance (MGR) controlled by a dominant R locus. Based on linkage disequilibrium (LD)-based association mapping used to detect single nucleotide polymorphism (SNP) markers associated with MGR against C. ribicola, MGR in these seed families appears to be controlled by Cr4 or other R genes in very close proximity to Cr4. These associated SNPs were located in genes involved in multiple molecular mechanisms potentially underlying limber pine MGR to C. ribicola, including NBS-LRR genes for recognition of C. ribicola effectors, signaling components, and a large set of defense-responsive genes with potential functions in plant effector-triggered immunity (ETI). Interactions of associated loci were identified for MGR selection in trees with complex genetic backgrounds. SNPs with tight Cr4-linkage were further converted to TaqMan assays to confirm their effectiveness as MAS tools. This work demonstrates the successful translation and deployment of molecular genetic knowledge into specific MAS tools that can be easily applied in a selection or breeding program to efficiently screen MGR against WPBR in Alberta limber pine populations.

Erratum to: Kinetic Characterization of Tyrosinase-catalyzed Oxidation of Four Polyphenols.

The article "Kinetic Characterization of Tyrosinase-catalyzed Oxidation of Four Polyphenols", written by Wan-yu LIU, Congming ZOU, Jian-hua HU, Zi-jun XU, Lu-qin SI, Jun-jun LIU, Jian-geng HUANG, was originally published electronically on the publisher's internet portal on May 2020 without open access. With the author(s)' decision to opt for Open Choice, the copyright of the article is changed to © The Author(s) 2020 and the article is forthwith distributed under a Creative Commons Attribution 4.0 International License ( https://creativecommons.org/licenses/by/4.0/ ), which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.The original article has been corrected.

Kinetic Characterization of Tyrosinase-catalyzed Oxidation of Four Polyphenols.

Phenolic compounds such as chlorogenic acid, cryptochlorogenic acid, neochlorogenic acid and caffeic acid are widely distributed in fruits, vegetables and traditional Chinese medicines with a wide range of biological activities. Tyrosinase plays a critical role in the food industry, but recent studies have proposed unexplored aspects of clinical application. Tyrosinase-catalyzed oxidation of four polyphenols as well as its underlying mechanism remains unclear. In the current work, we investigated the kinetic properties of tyrosinase-catalyzed oxidation of the four polyphenols of interest. To measure the unstable o-quinone products, an analytical method using 3-methyl-2-benzothiazolinone hydrazone (MBTH) was established. The optimal incubation time, buffer pH, temperature and enzyme concentration for the enzyme activity in the presence of each polyphenol of interest were investigated. Under the final optimized conditions, the kinetics and substrate specificity of four polyphenols were examined. Kinetic data showed that tyrosinase had the greatest substrate affnity to chlorogenic acid compared with its isomers and caffeic acid. The catalytic effciency with chlorogenic acid was 8- to 15-fold higher than that with the other 3 polyphenols. Molecular docking study demonstrated that the tight binding of chlorogenic acid at the peripheral site should be the major reason for the specifcity to chlorogenic acid. In light of this, the rational design of high-affnity inhibitors against tyrosinase may focus on the binding of both the Cu site and peripheral site. This study will supply a basis for the selection of phenolic acids in food industry and health care.

Genetic diversity of strawberry mild yellow edge virus from eastern Canada.

Strawberry mild yellow edge virus (SMYEV) is a member of the genus Potexvirus, family Alphaflexiviridae. It is one of the most common pathogenic viruses infecting cultivated strawberries worldwide. In this study, we investigated the genetic diversity of SMYEV in strawberry fields that were severely affected by strawberry decline disease in the eastern Canadian provinces of New Brunswick, Nova Scotia, Prince Edward Island and Quebec. A total of 134 SMYEV coat protein (CP) gene sequences, representing 85 nucleic acid haplotypes, were identified in 56 field samples. A highly divergent SMYEV population was found in all four provinces, but there was little genetic differentiation among the populations, and moreover, the Canadian SMYEV isolates formed a unique dissimilar, genetically divergent population group when compared to those reported in other countries. Phylogenetic analysis revealed three new SMYEV subclades that consisted mainly of Canadian variants and were composed of 76 sequence haplotypes (76/85, 88%). Mixed infections by different SMYEV variants were observed in 38 samples (38/56, 68%). Evolutionary analysis suggested that the SMYEV strains in eastern Canada possibly originated outside of Canada but adapted to conditions in the region through genetic mutations.

Characterization of Cronartium ribicola dsRNAs reveals novel members of the family Totiviridae and viral association with fungal virulence.

BACKGROUND: Mycoviruses were recently discovered in the white pine blister rust (WPBR) fungus Cronartium ribicola (J.C. Fisch.). Detection and characterization of their double stranded RNA (dsRNA) would facilitate understanding of pathogen virulence and disease pathogenesis in WPBR systems. METHODS: Full-length cDNAs were cloned from the dsRNAs purified from viral-infected C. ribicola, and their cDNA sequences were determined by DNA sequencing. Evolutionary relationships of the dsRNAs with related mycoviruses were determined by phylogenetic analysis. Dynamic distributions of the viral RNAs within samples of their fungal host C. ribicola were investigated by measurement of viral genome prevalence and viral gene expression. RESULTS: In this study we identified and characterized five novel dsRNAs from C. ribicola, designated as Cronartium ribicola totivirus 1-5 (CrTV1 to CrTV5). These dsRNA sequences encode capsid protein and RNA-dependent RNA polymerase with significant homologies to dsRNA viruses of the family Totiviridae. Phylogenetic analysis showed that the CrTVs were grouped into two distinct clades. CrTV2 through CrTV5 clustered within the genus Totivirus. CrTV1 along with a few un-assigned dsRNAs constituted a distinct phyletic clade that is genetically distant from presently known genera in the Totiviridae family, indicating that CrTV1 represents a novel genus in the Totiviridae family. The CrTVs were prevalent in fungal samples obtained from infected western white pine, whitebark pine, and limber pines. Viral RNAs were generally expressed at higher levels during in planta mycelium growth than in aeciospores and urediniospores. CrTV4 was significantly associated with C. ribicola virulent pathotype and specific C. ribicola host tree species, suggesting dsRNAs as potential tools for dissection of pathogenic mechanisms of C. ribicola and diagnosis of C. ribicola pathotypes. CONCLUSION: Phylogenetic and expression analyses of viruses in the WPBR pathogen, C. ribicola, have enchanced our understanding of virus diversity in the family Totiviridae, and provided a potential strategy to utilize pathotype-associated mycoviruses to control fungal forest diseases.