Haplotype-resolved genome assembly and resequencing analysis provide insights into genome evolution and allelic imbalance in Pinus densiflora.

Haplotype-level allelic characterization facilitates research on the functional, evolutionary and breeding-related features of extremely large and complex plant genomes. We report a 21.7-Gb chromosome-level haplotype-resolved assembly in Pinus densiflora. We found genome rearrangements involving translocations and inversions between chromosomes 1 and 3 of Pinus species and a proliferation of specific long terminal repeat (LTR) retrotransposons (LTR-RTs) in P. densiflora. Evolutionary analyses illustrated that tandem and LTR-RT-mediated duplications led to an increment of transcription factor (TF) genes in P. densiflora. The haplotype sequence comparison showed allelic imbalances, including presence-absence variations of genes (PAV genes) and their functional contributions to flowering and abiotic stress-related traits in P. densiflora. Allele-aware resequencing analysis revealed PAV gene diversity across P. densiflora accessions. Our study provides insights into key mechanisms underlying the evolution of genome structure, LTR-RTs and TFs within the Pinus lineage as well as allelic imbalances and diversity across P. densiflora.

SARS-CoV-2 spike-specific nasal-resident CD49a+CD8+ memory T cells exert immediate effector functions with enhanced IFN-γ production.

Virus-specific nasal resident T cells are important for protection against subsequent infection with a similar virus. Here we examine the phenotypes and functions of SARS-CoV-2-specific T cells in the nasal mucosa of vaccinated individuals with breakthrough infection (BTI) or without infection. Nasal tissues are obtained from participants during sinus surgery. Analysis of activation-induced markers implicates that a considerable proportion of spike (S)-reactive nasal CD8+ T cells express CD103, a tissue-resident marker. MHC-I multimer staining is performed to analyze the ex vivo phenotype and function of SARS-CoV-2 S-specific CD8+ T cells. We detect multimer+CD8+ T cells with tissue-resident phenotypes in nasal tissue samples from vaccinees without infection as well as vaccinees with BTI. Multimer+CD8+ T cells remain present in nasal tissues over one year after the last exposure to S antigen, although the frequency decreases. Upon direct ex vivo stimulation with epitope peptides, nasal multimer+CD8+ T cells-particularly the CD49a+ subset-exhibit immediate effector functions, including IFN-γ production. CITE-seq analysis of S-reactive AIM+CD8+ T cells confirms the enhanced effector function of the CD49a+ subset. These findings indicate that among individuals previously exposed to S antigen by vaccination or BTI, S-specific nasal-resident CD49a+CD8+ memory T cells can rapidly respond to SARS-CoV-2 during infection or reinfection.

Climate change impacts on Allium crop production: Insights from long-term observations in South Korea.

Climate change is an imminent threat, particularly affecting agricultural productivity, which relies heavily on weather conditions. Understanding the specific impacts of climate change on key crops is crucial for developing effective adaptation strategies to ensure food security. The growth patterns of onions and garlic were observed at over ten different locations in South Korea, and the yield data from the past 40 years were analyzed. The yield was significantly correlated with temperature and strongly affected by the frequent and unexpected patterns of precipitation. The increase in mean temperature during winter and the spatial and temporal concentration of precipitation are expected to be the most influential factors for Allium crop production in the future. In addition, the yields of onions and garlic can serve as good indicators for predicting the impacts of weather on agricultural productivity, given their extended cultivation periods and significant correlations with temperature and precipitation. As climate change scenarios become available, the results of this study can serve as a basis for predicting changes in agricultural production in the future and identifying opportunities to adapt cultivation systems for food security.

Correlation between reproductive performance and sow body weight change during gestation.

This study investigated the correlation between piglet performance and sow body weight change (BWC) during two gestational periods: 35-70, 70-105, and 35-105 days. A cohort of 70 sows was evaluated for BWC, backfat thickness change (BFC), caliper score change (CALC), feed intake, and weaning-to-estrus interval (WEI). The collected data were then analyzed according to the two specified periods. Our findings highlighted that piglet birth weight, weaning weight, and average daily weight gain (ADG) correlated with sow body characteristics, including BFC and CALC. The strongest correlation was observed with BWC. Piglet mortality was intimately associated with BFC. Piglet birth weight, weaning weight, and ADG showed a positive correlation with sow BWC, particularly during the 35-70 day period. Furthermore, sows displaying a higher BWC during the 70-105 day period, and also exhibiting a higher BW gain from 35-70 days, registered greater piglet weight gains and higher weaning weights. These trends became more apparent as the sow's BWC increased during the 70-105 day period. Piglet mortality increased when the sow exhibited a lower BWC during both the 35-70 and 70-105 day periods. No significant observations were found concerning the number of stillborn piglets, live-born piglets, or weaned piglets, and no interaction effects were detected between these periods. In conclusion, our findings underscore the significance of sow BWC during the early stages of gestation (d 35-70) for enhancing piglet performance from birth to weaning.

Pathogenicity, Host Resistance, and Genetic Diversity of Fusarium Species under Controlled Conditions from Soybean in Canada.

Fusarium spp. are commonly associated with the root rot complex of soybean (Glycine max). Previous surveys identified six common Fusarium species from Manitoba, including F. oxysporum, F. redolens, F. graminearum, F. solani, F. avenaceum, and F. acuminatum. This study aimed to determine their pathogenicity, assess host resistance, and evaluate the genetic diversity of Fusarium spp. isolated from Canada. The pathogenicity of these species was tested on two soybean cultivars, 'Akras' (moderately resistant) and 'B150Y1' (susceptible), under greenhouse conditions. The aggressiveness of the fungal isolates varied, with root rot severities ranging from 1.5 to 3.3 on a 0-4 scale. Subsequently, the six species were used to screen a panel of 20 Canadian soybean cultivars for resistance in a greenhouse. Cluster and principal component analyses were conducted based on the same traits used in the pathogenicity study. Two cultivars, 'P15T46R2' and 'B150Y1', were consistently found to be tolerant to F. oxysporum, F. redolens, F. graminearum, and F. solani. To investigate the incidence and prevalence of Fusarium spp. in Canada, fungi were isolated from 106 soybean fields surveyed across Manitoba, Saskatchewan, Ontario, and Quebec. Eighty-three Fusarium isolates were evaluated based on morphology and with multiple PCR primers, and phylogenetic analyses indicated their diversity across the major soybean production regions of Canada. Overall, this study contributes valuable insights into host resistance and the pathogenicity and genetic diversity of Fusarium spp. in Canadian soybean fields.

Development of a virtual reality system usability questionnaire (VRSUQ).

Virtual reality (VR) has gained significant attention as a technology that provides immersive experiences similar to the real world. In order for a VR system to be accepted, usability needs to be guaranteed. Accordingly, VR-related researchers are continuing their efforts to improve VR systems by conducting usability evaluations. However, existing studies have limitations in that they cannot comprehensively evaluate the detailed properties of VR systems by using questionnaires developed for general product usability evaluation or focusing only on some usability aspects of VR systems. This suggests it may be difficult to fully capture usability issues in a VR system, and that it is necessary to develop a usability evaluation tool that reflects the specific characteristics of the VR system. Therefore, this study develops and proposes the Virtual Reality System Usability Questionnaire (VRSUQ). In the development of the questionnaire, items were structured based on a literature review and discussions with experts. To account for the diverse characteristics of VR systems, the validity of the questionnaire was verified through both exploratory and confirmatory factor analysis, utilizing data obtained from three distinct experimental studies that employed different VR systems. In addition, by comparing the results of VRSUQ with the results from the System Usability Scale, which is widely used for perceived usability evaluation, alternative possibilities for using VRSUQ are presented. Further testing on various VR platforms is needed to ensure the reliability and validity of VRSUQ, and as results from using VRSUQ are accumulated, it is expected to be widely used as a more powerful and robust VR-specific perceived usability evaluation tool.

High-quality chromosome-level genome assembly of Nicotiana benthamiana.

Nicotiana benthamiana is a fundamental model organism in plant research. Recent advancements in genomic sequencing have revealed significant intraspecific genetic variations. This study addresses the pressing need for a precise genome sequence specific to its geographic origin by presenting a comprehensive genome assembly of the N. benthamiana LAB strain from the Republic of Korea (NbKLAB). We compare this assembly with the widely used NbLAB360 strain, shedding light on essential genomic differences between them. The outcome is a high-quality, chromosome-level genome assembly comprising 19 chromosomes, spanning 2,762 Mb, with an N50 of 142.6 Mb. Comparative analyses revealed notable variations, including 46,215 protein-coding genes, with an impressive 99.5% BUSCO completeness score. Furthermore, the NbKLAB assembly substantially improved the QV from 33% for NbLAB360 to 49%. This refined chromosomal genome assembly for N. benthamiana, in conjunction with comparative insights, provides a valuable resource for genomics research and molecular biology. This accomplishment forms a strong foundation for in-depth exploration into the intricacies of plant genetics and genomics, improved precision, and a comparative framework.

Multislice computed tomography demonstrating mental nerve paresthesia caused by periapical infection: A case report.

Components derived from an infected lesion within the bone can spread through various passages in the mandible, particularly via the mental foramen. Radiologically, the spread of infection is typically nonspecific and challenging to characterize; however, multislice computed tomography (MSCT) can effectively detect pathological changes in soft tissues and the bone marrow space. This report describes the case of a 55-year-old woman who experienced mental nerve paresthesia due to a periapical infection of the right mandibular second premolar. MSCT imaging revealed increased attenuation around the periapical lesion extending into the mandibular canal and loss of the juxtamental foraminal fat pad. Following endodontic treatment of the tooth suspected to be the source of the infection, the patient's symptoms resolved, and the previous MSCT imaging findings were no longer present. Increased bone marrow attenuation and obliteration of the fat plane in the buccal aspect of the mental foramen may serve as radiologic indicators of inflammation spreading from the bone marrow space.

Deep learning algorithm for the automated detection and classification of nasal cavity mass in nasal endoscopic images.

Nasal endoscopy is routinely performed to distinguish the pathological types of masses. There is a lack of studies on deep learning algorithms for discriminating a wide range of endoscopic nasal cavity mass lesions. Therefore, we aimed to develop an endoscopic-examination-based deep learning model to detect and classify nasal cavity mass lesions, including nasal polyps (NPs), benign tumors, and malignant tumors. The clinical feasibility of the model was evaluated by comparing the results to those of manual assessment. Biopsy-confirmed nasal endoscopic images were obtained from 17 hospitals in South Korea. Here, 400 images were used for the test set. The training and validation datasets consisted of 149,043 normal nasal cavity, 311,043 NP, 9,271 benign tumor, and 5,323 malignant tumor lesion images. The proposed Xception architecture achieved an overall accuracy of 0.792 with the following class accuracies on the test set: normal = 0.978 ± 0.016, NP = 0.790 ± 0.016, benign = 0.708 ± 0.100, and malignant = 0.698 ± 0.116. With an average area under the receiver operating characteristic curve (AUC) of 0.947, the AUC values and F1 score were highest in the order of normal, NP, malignant tumor, and benign tumor classes. The classification performances of the proposed model were comparable with those of manual assessment in the normal and NP classes. The proposed model outperformed manual assessment in the benign and malignant tumor classes (sensitivities of 0.708 ± 0.100 vs. 0.549 ± 0.172, 0.698 ± 0.116 vs. 0.518 ± 0.153, respectively). In urgent (malignant) versus nonurgent binary predictions, the deep learning model achieved superior diagnostic accuracy. The developed model based on endoscopic images achieved satisfactory performance in classifying four classes of nasal cavity mass lesions, namely normal, NP, benign tumor, and malignant tumor. The developed model can therefore be used to screen nasal cavity lesions accurately and rapidly.

Pan-chloroplast genomes for accession-specific marker development in Hibiscus syriacus.

Hibiscus syriacus L. is a renowned ornamental plant. We constructed 95 chloroplast genomes of H. syriacus L. cultivars using a short-read sequencing platform (Illumina) and a long-read sequencing platform (Oxford Nanopore Technology). The following genome assembly, we delineate quadripartite structures encompassing large single-copy, small single-copy, and inverted repeat (IRa and IRb) regions, from 160,231 bp to 161,041 bp. Our comprehensive analyses confirmed the presence of 79 protein-coding genes, 30 tRNA genes, and 4 rRNA genes in the pan-chloroplast genome, consistent with prior research on the H. syriacus chloroplast genome. Subsequent pangenome analysis unveiled widespread genome sequence conservation alongside unique cultivar-specific variant patterns consisting of 193 single-nucleotide polymorphisms and 61 insertions or deletions. The region containing intra-species variant patterns, as identified in this study, has the potential to develop accession-specific molecular markers, enhancing precision in cultivar classification. These findings are anticipated to drive advancements in breeding strategies, augment biodiversity, and unlock the agricultural potential inherent in H. syriacus.

Respiratory Epithelial Adenomatoid Hamartoma at an Unusual Location: A Case Report and Literature Review.

Respiratory epithelial adenomatoid hamartoma (REAH) in the head and neck is a rare benign lesion containing glandular tissue covered with ciliated respiratory epithelium. In the head and neck, REAH of the nasal cavity, paranasal sinuses, and nasopharynx have been reported in literature. Due to rareness of REAH and insufficient knowledge of its imaging features, the diagnosis can be challenging when we encounter a non-specific cystic mass at an uncommon site in the head or neck. Here, we report the case of a pathologically confirmed REAH showing a cystic mass centered at the buccal space (retromaxillary fat pad) with CT and MRI findings.

Application of deep learning with bivariate models for genomic prediction of sow lifetime productivity-related traits.

OBJECTIVE: Pig breeders cannot obtain phenotypic information at the time of selection for sow lifetime productivity (SLP). They would benefit from obtaining genetic information of candidate sows. Genomic data interpreted using deep learning (DL) techniques could contribute to the genetic improvement of SLP to maximize farm profitability because DL models capture nonlinear genetic effects such as dominance and epistasis more efficiently than conventional genomic prediction methods based on linear models. This study aimed to investigate the usefulness of DL for the genomic prediction of two SLP-related traits; lifetime number of litters (LNL) and lifetime pig production (LPP). METHODS: Two bivariate DL models, convolutional neural network (CNN) and local convolutional neural network (LCNN), were compared with conventional bivariate linear models (i.e., genomic best linear unbiased prediction, Bayesian ridge regression, Bayes A, and Bayes B). Phenotype and pedigree data were collected from 40,011 sows that had husbandry records. Among these, 3,652 pigs were genotyped using the PorcineSNP60K BeadChip. RESULTS: The best predictive correlation for LNL was obtained with CNN (0.28), followed by LCNN (0.26) and conventional linear models (approximately 0.21). For LPP, the best predictive correlation was also obtained with CNN (0.29), followed by LCNN (0.27) and conventional linear models (approximately 0.25). A similar trend was observed with the mean squared error of prediction for the SLP traits. CONCLUSION: This study provides an example of a CNN that can outperform against the linear model-based genomic prediction approaches when the nonlinear interaction components are important because LNL and LPP exhibited strong epistatic interaction components. Additionally, our results suggest that applying bivariate DL models could also contribute to the prediction accuracy by utilizing the genetic correlation between LNL and LPP.

A study of anti-wrinkle functions and improvement of cream with Phaseolus angularis.

Chronic exposure to ultraviolet (UV) radiation from sunlight accelerates skin ageing, which is followed by harsh, thick, dry and loose conditions. One of the most demonstrative symptoms is deep wrinkles induced by skin barrier disruption. Our previous research showed that Phaseolus angularis seed extract (PASE) effectively inhibits skin ageing through UVB protection in HaCaT cells by suppressing skin damage. However, its efficacy has not been evaluated in clinical trials so far. PASE cream's effectiveness was initially tested on the artificial skin model, revealing an increase in filaggrin and defence against skin damage. Based on these results, in this single-centred, randomized, double-blind study, we investigated the anti-ageing effect of PASE in human eye wrinkle areas. For these 21 healthy adult women aged 30 to 59, a PASE cream was applied to the right eye wrinkle area and a placebo to the left eye wrinkle area twice a day (morning and evening) for 12 weeks. The change in thick, deep crease wrinkles around the eyes was confirmed by visual evaluation, skin measurements and a questionnaire. As a result, the surface roughness (R1), maximum roughness (R2), average roughness (R3), smoothness depth (R4) and arithmetic mean roughness (R5) values in the group using the PASE cream all decreased. Particularly, R1, R4 and R5 significantly decreased by 18.1%, 18.6% and 25.0%, respectively. Subjects who applied PASE cream also experienced an improvement in skin moisture nearly twice the time compared to the placebo group. In addition, no participants reported side effects. Our study showed that PASE cream led to clinically significant levels of wrinkle improvement. In conclusion, as PASE is a natural, safe food with no side effects, it can be a good resource for natural anti-wrinkle functional cosmetics in the future.

L'exposition chronique aux rayons ultraviolets (UV) du soleil accélère le vieillissement cutané, qui provoque un épaississement et un assèchement de la peau et la rend plus lâche. La présence de rides profondes induites par la rupture de la barrière cutanée en constitue l'un des symptômes les plus manifestes. Lors d'études précédentes, nous nous sommes rendu compte que l'extrait de graines de Phaseolus angularis (PASE) inhibait efficacement le vieillissement de la peau en assurant la protection anti­UVB des cellules HaCaT grâce à la suppression des lésions cutanées. Cependant, son efficacité n'a pas été évaluée lors d'essais cliniques à ce jour. L'efficacité de la crème PASE a d'abord été testée sur le modèle de peau artificielle, sur laquelle elle a fait augmenter les taux de filaggrine et assuré une défense contre les lésions cutanées. Sur la base de ces résultats, dans cette étude unicentrique, randomisée et en double aveugle, nous avons étudié l'effet anti­âge de la PASE chez l'humain au niveau des rides proches de l'œil. Pour ces 21 femmes adultes en bonne santé âgées de 30 à 59 ans, une crème PASE a été appliquée sur la zone de rides de l'œil droit et un placebo sur la zone de rides de l'œil gauche deux fois par jour (matin et soir) pendant 12 semaines. La modification des rides profondes et épaisses autour des yeux a été confirmée par une évaluation visuelle, des mesures cutanées et un questionnaire. Il a été découvert que les valeurs de rugosité de surface (R1), de rugosité maximale (R2), de rugosité moyenne (R3), de profondeur de douceur (R4) et de moyenne arithmétique (R5) dans le groupe à l'aide de la crème PASE avaient toutes diminué. En particulier, R1, R4 et R5 ont significativement diminué de 18,1 %, de 18,6 % et de 25,0 %, respectivement. Les patients qui ont appliqué la crème PASE ont également présenté une amélioration de l'hydratation de la peau presque deux fois supérieure à celle du groupe placebo. En outre, aucun participant n'a signalé d'effets secondaires. Notre étude a montré que la crème PASE entraînait des niveaux cliniquement significatifs d'amélioration des rides. En conclusion, comme le PASE est un aliment naturel, sûr et dépourvu d'effets secondaires, elle peut constituer une bonne ressource pour les cosmétiques fonctionnels naturels antirides à l'avenir.

Nasal instillation of povidone-iodine ameliorates ongoing mucosal inflammation in a pre-sensitized murine model of Der p1-induced allergic rhinitis.

BACKGROUND: Interleukin (IL)-33, when cleaved into smaller fragments by proteases, becomes hyperactive, contributing to allergic inflammation. Povidone-iodine (PVP-I) is an iodine-based compound that exhibits antimicrobial properties and inhibits proteases. This study aimed to investigate whether PVP-I treatment inhibits IL-33 cleavage, improves allergic rhinitis (AR) symptoms, and suppresses allergic inflammation in a mouse model. METHODS: In vitro experiments using full-length recombinant human IL-33 and allergens, including house dust mites or Dermatophagoides pteronyssinus 1, were conducted using western blotting. Fifty BALB/c mice were divided into five groups: control (CON), AR with phosphate-buffered saline treatment (AR), PVP-I treatment (AR + PVP), trans-epoxysuccinyl-L-leucylamido(4-guanidino)butane (E64) treatment (AR + E64), and dexamethasone treatment (AR + Dexa). Nasal symptom scores, including rubbing and sneezing, were measured. The cytokine levels in the nasal lavage fluid (NLF) and the concentration of immunoglobulins in the blood serum were assessed. Nasal mucosa from each group was used for reverse transcriptase-polymerase chain reaction (RT-PCR) and histological analyses were conducted. RESULTS: PVP-I treatment reduced nasal symptoms, suppressed allergic inflammation, and decreased the levels of IL-33, IL-5, and IL-13 in the NLF and total immunoglobulin E (IgE) and specific IgE in the serum. Histopathological analysis revealed a reduction in the number of eosinophils and goblet cells in the nasal mucosa of the AR + PVP group when compared to the AR group. RT-PCR and immunofluorescence staining confirmed the downregulation of genes and proteins associated with allergic inflammation. CONCLUSIONS: These findings suggest that nasal irrigation with PVP-I may be a promising therapeutic option for managing AR by inhibiting IL-33 activation and suppressing allergic inflammation.

First Report of Charcoal Rot Caused by Macrophomina phaseolina on Dry Bean (Phaseolus vulgaris L.) in Western Canada.

In 2021, several dry bean (Phaseolus vulgaris L.) plants at the mid-seed-fill growth stage displaying wilting, chlorosis of the leaves and reduced vigor were collected near the Pembina - Emerson Border of Manitoba, Canada and North Dakota, USA. When symptomatic plants were examined, gray to dark brown discoloration was observed on the lower stem and the roots. Afterwards, brown to black discoloration was noticed on stem and root sections. Root and lower stem pieces (1 to 2 cm) from affected plants were surface sterilized with 70 % ethanol, followed by 1% NaOCl, rinsed twice in sterilized water, air dried on sterilized filter papers, and placed on potato dextrose agar (PDA) amended with 1 mg/mL of streptomycin sulfate. The PDA plates were incubated at 28°C with 12 h light/12 h dark for 10 days. The growing hyphae were transferred using the hyphal tip method to new PDA plates. Growing cultures were initially hyaline and turned from light gray to dark brown or black with age. Abundant dark and spherical to oblong shaped sclerotia with an average diameter of 97.9 µm (range: 66.8 to 143.5 µm, n =30) formed on the pure cultures 7 days after incubation. Additional pure culture was obtained through an isolation of a single microsclerotium followed by a single hyphal tip transfer. One isolate was identified as Macrophomina phaseolina based on morphological characteristics (Smith and Wyllie 1999). The morphological identity was confirmed by sequencing the rDNA internal transcribed spacer (ITS) region with universal primers ITS1/ITS4 (White et al. 1990) and calmodulin (CAL) and translation elongation factor-1 alpha (TEF-1α) genes with MpTefF/MpTefR, and MpCalF/MpCalR primer sets (Santos et al. 2020), respectively. The online resource Basic Local Alignment Search Tool (BLAST; https://www.ncbi.nlm.nih.gov/BLAST) confirmed the fungus identity as 100% M. phaseolina. The sequences of the original isolate BF21-25 were deposited in GenBank with accession numbers OQ615297 (ITS), OR357630 (CAL), and OR363106 (TEF-1α). To confirm pathogenicity, bioassays were conducted under controlled conditions. Four seeds of cultivar 'Etna' were sown per pot, and five pots were used for inoculated (approx. 4 × 105 microsclerotia/pot) and control (mock-inoculated with sterile PDA medium) treatments. For the inoculum, 20 g of macerated 10 to 14-day old M. phaseolina culture grown on PDA medium was applied to each pot using an inoculum layering technique. Pots were kept in the greenhouse with 28/17°C day/night, 13 h light/11 h dark cycle, and 70% relative humidity and watered weekly. Disease symptoms similar to those observed in the field were visible on all inoculated plants at the mid-seed-fill growth stage. Mock-inoculated control plants didn't show any symptoms. The experiment was repeated twice with similar results. The pathogen was re-isolated from infected plants to confirm Koch's postulates and identified as M. phaseolina based on the morphology and sequences of ITS, CAL and TEF-1α regions. To our knowledge, this is the first report of charcoal rot caused by M. phaseolina on dry bean in Western Canada.

First Report of Charcoal Rot on Soybean Caused by Macrophomina phaseolina in Manitoba, Canada.

Macrophomina phaseolina (Tassi) Goid. is a soilborne necrotrophic fungal pathogen causing charcoal rot on approximately 500 plant species worldwide (Mengistu et al. 2015). Charcoal rot occurs in eastern Canada and many regions of the USA, causing substantial yield losses in soybean [Glycine max (L.) Merr.] (Allen et al. 2017; Bradley et al. 2021; Wrather et al. 2001). However, it has not been reported in soybean in western Canada. Manitoba is the second largest soybean producer in Canada, comprising 31% of total seeded areas with 2.29 M acres in 2017 (Statistics Canada 2022). Still, soybean is a relatively new crop to Manitoba and annual surveys of soybean root diseases began in 2012. In August 2020, randomly selected soybean fields were surveyed for root diseases at 63 different locations in south-central and southwest Manitoba. A total of thirty diseased plants were sampled in a zigzag pattern at three random sites in each field and all samples were brought to the laboratory and rated for disease severity. All plants showed symptoms of root rot, and some samples exhibited wilting with yellowing-brown leaves attached to the stems by the petioles; when the taproot was sectioned longitudinally, black streaking could be observed. In the laboratory, 600 roots from 40 selected fields were processed for pathogen isolation and identification. A 1 cm section from each root was surface-sterilized in a 95% EtOH:5.25% NaOCl solution for 30 sec, rinsed in sterile water for 60 sec, and air-dried on sterilized filter paper in a laminar flow hood. Root tissues with two replicates were placed on potato dextrose agar (PDA) plates amended with streptomycin sulfate (2 mg/mL) and incubated at room temperature. Black microsclerotia were observed in cultures from three different fields and three individual fungal isolates were obtained from each field through isolation of a single microsclerotium and subsequent hyphal tip transfer. The mycelia were initially hyaline and turned gray to dark brown or black, forming numerous microsclerotia ranging in size from 13 to 61 µm long and 12 to 32 µm wide, based on measurements of approximately 100 microsclerotia per isolate using a Zeiss Axio Imager A2 microscope equipped with an AxioCam HRc (Carl Zeiss, Jena, Germany) and AxioVision software. The color of the microsclerotia was jet black and the shape was round to oblong or irregular, as described by Mengistu et al. (2015). Based on morphological characteristics and microscopic examination, three fungal isolates were identified as M. phaseolina (Mengistu et al. 2015). For molecular identification, genomic DNA was extracted from 10 to 14-day old mycelia and microsclerotia of each isolate using a ZymoBIOMICS™ DNA Miniprep Kit (Zymo Research Corp., Irvine, CA, USA) according to the manufacturer's instructions. The internal transcribed spacer (ITS) region, translation elongation factor-1α (TEF-1α), and calmodulin (CAL) genes were amplified using the primer sets ITS1/ITS4 (White et al. 1990), MpTefF/MpTefR, and MpCalF/MpCalR (Santos et al. 2020), respectively, according to the original reaction conditions. Subsequently, PCR products were sequenced at Eurofins Genomics (Louisville, KY, USA). BLASTn analysis in GenBank showed that the nucleotide sequences of these regions of the three isolates (NSRR20-MB-24, NSRR20-MB-34, and NSRR20-MB-40) matched multiple isolates of M. phaseolina with 100% query cover and 100% identity. Sequences were deposited in GenBank for the ITS (OK127887, OK142725, OK128266), TEF-1α (OR363103, OR363104, OR363105), and CAL (OR357627, OR357628, OR357629) regions. In addition, the ITS and TEF-1α sequences of the three novel isolates were further aligned with multiple previously reported isolates of M. phaseolina, M. pseudophaseolina, and M. euphorbiicola (Chen et al. 2013; Machado et al. 2019; Sarr et al. 2014) using Muscle and trimmed (Edgar 2004). Alignments were concatenated to generate a maximum likelihood tree. Once concatenated, sequences were re-aligned. The obtained alignments were employed to construct a phylogenetic tree using the max likelihood method and Tamura-Nei model (Tamura and Nei 1993) with 10,000 bootstrap replicates using MEGA 11 (Tamura et al. 2021). The ITS and TEF-1α analysis indicated that the isolates were grouped in three differentiated clades (Figure 1). Macrophomina phaseolina isolates clustered in the same clade at 98% similarity, with the three novel soybean isolates NSRR20-MB-24, NSRR20-MB-34, and NSRR20-MB-40 grouped closely in the cluster at 98% similarity and identified as M. phaseolina. In contrast, isolates of M. euphorbiicola formed another clade at 87% similarity and M. pseudophaseolina isolates grouped in a clade at 99%. The pathogenicity of the three isolates was evaluated under controlled conditions. Given that no information on charcoal rot resistance in soybean has been reported in Canada, one of the commonly grown varieties in Manitoba, "TH 32004", was selected for the pathogenicity test. Surface-sterilized soybean seeds, which had been pre-germinated for three days, were sown in a sterilized soilless growing mix (Sunshine #5) together with 5 g (approx. 1 × 105 microsclerotia) of macerated 10 to 14-day old inoculum grown on PDA-streptomycin agar medium at room temperature and applied using an inoculum layering technique. For the non-inoculated control, macerated PDA-streptomycin agar without mycelia was used. Twenty plants per treatment were maintained in a walk-in plant growth chamber with a 16 h photoperiod at 25/20 °C ± 1 °C (day/night) and 50% relative humidity. Plants were watered weekly but were subjected to water stress. Symptoms of charcoal rot were observed in the root systems of all inoculated soybean plants after 28 days, while no symptoms were observed in the control plants (Figure S1). There was production of microsclerotia on the roots inoculated with each isolate (data not shown). Three isolates of M. phaseolina were re-isolated from the inoculated plants and found to be identical to the inoculated isolates with respect to morphological characteristics in culture, as well as with respect to the ITS, TEF-1α and CAL DNA sequences. For each isolate and non-inoculated control, five seeds of 'TH 32004' were seeded per pot, and four pots were used for the inoculated and control treatments. The experiment was repeated twice in a randomized complete block design with similar results, fulfilling Koch's postulates. To our knowledge, this is the first report of charcoal rot caused by M. phaseolina on soybean in Manitoba, Canada.

Two long read-based genome assembly and annotation of polyploidy woody plants, Hibiscus syriacus L. using PacBio and Nanopore platforms.

Improvements in long read DNA sequencing and related techniques facilitated the generation of complex eukaryotic genomes. Despite these advances, the quality of constructed plant reference genomes remains relatively poor due to the large size of genomes, high content of repetitive sequences, and wide variety of ploidy. Here, we developed the de novo sequencing and assembly of high polyploid plant genome, Hibiscus syriacus, a flowering plant species of the Malvaceae family, using the Oxford Nanopore Technologies and Pacific Biosciences Sequel sequencing platforms. We investigated an efficient combination of high-quality and high-molecular-weight DNA isolation procedure and suitable assembler to achieve optimal results using long read sequencing data. We found that abundant ultra-long reads allow for large and complex polyploid plant genome assemblies with great recovery of repetitive sequences and error correction even at relatively low depth Nanopore sequencing data and polishing compared to previous studies. Collectively, our combination provides cost effective methods to improve genome continuity and quality compared to the previously reported reference genome by accessing highly repetitive regions. The application of this combination may enable genetic research and breeding of polyploid crops, thus leading to improvements in crop production.

NADH elevation during chronic hypoxia leads to VHL-mediated HIF-1α degradation via SIRT1 inhibition.

BACKGROUND: Under conditions of hypoxia, cancer cells with hypoxia inducible factor-1α (HIF-1α) from heterogeneous tumor cells show greater aggression and progression in an effort to compensate for harsh environmental conditions. Extensive study on the stability of HIF-1α under conditions of acute hypoxia in cancer progression has been conducted, however, understanding of its involvement during the chronic phase is limited. METHODS: In this study, we investigated the effect of SIRT1 on HIF1 stability in a typical chronic hypoxic conditon that maintains cells for 24 h under hypoxia using Western blotting, co-IP, measurement of intracellular NAD + and NADH levels, semi-quantitative RT-PCR analysis, invasion assay, gene knockdown. RESULTS: Here we demonstrated that the high concentration of pyruvate in the medium, which can be easily overlooked, has an effect on the stability of HIF-1α. We also demonstrated that NADH functions as a signal for conveyance of HIF-1α degradation via the SIRT1 and VHL signaling pathway under conditions of chronic hypoxia, which in turn leads to attenuation of hypoxically strengthened invasion and angiogenic activities. A steep increase in the level of NADH occurs during chronic hypoxia, leading to upregulation of acetylation and degradation of HIF-1α via inactivation of SIRT1. Of particular interest, p300-mediated acetylation at lysine 709 of HIF-1α is recogonized by VHL, which leads to degradation of HIF-1α via ubiquitin/proteasome machinary under conditions of chronic hypoxia. In addition, we demonstrated that NADH-elevation-induced acetylation and subsequent degradation of HIF-1α was independent of proline hydroxylation. CONCLUSIONS: Our findings suggest a critical role of SIRT1 as a metabolic sensor in coordination of hypoxic status via regulation of HIF-1α stability. These results also demonstrate the involvement of VHL in degradation of HIF-1α through recognition of PHD-mediated hydroxylation in normoxia and p300-mediated HIF-1α acetylation in hypoxia.

A global-temporal analysis on Phytophthora sojae resistance-gene efficacy.

Plant disease resistance genes are widely used in agriculture to reduce disease outbreaks and epidemics and ensure global food security. In soybean, Rps (Resistance to Phytophthora sojae) genes are used to manage Phytophthora sojae, a major oomycete pathogen that causes Phytophthora stem and root rot (PRR) worldwide. This study aims to identify temporal changes in P. sojae pathotype complexity, diversity, and Rps gene efficacy. Pathotype data was collected from 5121 isolates of P. sojae, derived from 29 surveys conducted between 1990 and 2019 across the United States, Argentina, Canada, and China. This systematic review shows a loss of efficacy of specific Rps genes utilized for disease management and a significant increase in the pathotype diversity of isolates over time. This study finds that the most widely deployed Rps genes used to manage PRR globally, Rps1a, Rps1c and Rps1k, are no longer effective for PRR management in the United States, Argentina, and Canada. This systematic review emphasizes the need to widely introduce new sources of resistance to P. sojae, such as Rps3a, Rps6, or Rps11, into commercial cultivars to effectively manage PRR going forward.

Occurrence of Ascospores and White Mold Caused by Sclerotinia sclerotiorum in Dry Bean Fields in Alberta, Canada.

White mold caused by the fungal pathogen Sclerotinia sclerotiorum (Lib.) de Bary is one of the most important biological constraints to dry bean (Phaseolus vulgaris L.) production in Canada. Disease forecasting is one tool that could help growers manage the disease while reducing fungicide use. However, predicting white mold epidemics has remained difficult due to their sporadic occurrence. In this study, over the course of four growing seasons (2018 to 2021), we surveyed dry bean fields in Alberta and collected daily in-field weather data and daily in-field ascospore counts. White mold levels were variable and generally high in all years, confirming that the disease is ubiquitous and a constant threat to dry bean production. Ascospores were present throughout the growing season, and mean ascospore levels varied by field, month, and year. Models based on in-field weather and ascospore levels were not highly predictive of final disease incidence in a field, suggesting that environment and pathogen presence were not limiting factors to disease development. Rather, significant effects of market class on disease were found, with pinto beans, on average, having the highest disease incidence (33%) followed by great northern (15%), black (10%), red (6%), and yellow (5%). When incidence of these market classes was modeled separately, different environmental variables were important in each model; however, average wind speed was a significant variable in all models. Taken together, these findings suggest that white mold management in dry bean should focus on fungicide use, plant genetics, irrigation management, and other agronomic factors.