Full-length agave transcriptome reveals candidate glycosyltransferase genes involved in hemicellulose biosynthesis.

Agave species are typical crassulacean acid metabolism (CAM) plants commonly cultivated to produce beverages, fibers, and medicines. To date, few studies have examined hemicellulose biosynthesis in Agave H11648, which is the primary cultivar used for fiber production. We conducted PacBio sequencing to obtain full-length transcriptome of five agave tissues: leaves, shoots, roots, flowers, and fruits. A total of 41,807 genes were generated, with a mean length of 2394 bp and an annotation rate of 97.12 % using public databases. We identified 42 glycosyltransferase genes related to hemicellulose biosynthesis, including mixed-linkage glucan (1), glucomannan (5), xyloglucan (16), and xylan (20). Their expression patterns were examined during leaf development and fungal infection, together with hemicellulose content. The results revealed four candidate glycosyltransferase genes involved in xyloglucan and xylan biosynthesis, including glucan synthase (CSLC), xylosyl transferase (XXT), xylan glucuronyltransferase (GUX), and xylan α-1,3-arabinosyltransferase (XAT). These genes can be potential targets for manipulating xyloglucan and xylan traits in agaves, and can also be used as candidate enzymatic tools for enzyme engineering. We have provided the first full-length transcriptome of agave, which will be a useful resource for gene identification and characterization in agave species. We also elucidated the hemicellulose biosynthesis machinery, which will benefit future studies on hemicellulose traits in agave.

CURATE.AI-assisted dose titration for anti-hypertensive personalized therapy: study protocol for a multi-arm, randomized, pilot feasibility trial using CURATE.AI (CURATE.AI ADAPT trial).

Aims: Artificial intelligence-driven small data platforms such as CURATE.AI hold potential for personalized hypertension care by assisting physicians in identifying personalized anti-hypertensive doses for titration. This trial aims to assess the feasibility of a larger randomized controlled trial (RCT), evaluating the efficacy of CURATE.AI-assisted dose titration intervention. We will also collect preliminary efficacy and safety data and explore stakeholder feedback in the early design process. Methods and results: In this open-label, randomized, pilot feasibility trial, we aim to recruit 45 participants with primary hypertension. Participants will be randomized in 1:1:1 ratio into control (no intervention), home blood pressure monitoring (active control; HBPM), or CURATE.AI arms (intervention; HBPM and CURATE.AI-assisted dose titration). The home treatments include 1 month of two-drug anti-hypertensive regimens. Primary endpoints assess the logistical (e.g. dose adherence) and scientific (e.g. percentage of participants for which CURATE.AI profiles can be generated) feasibility, and define the progression criteria for the RCT in a 'traffic light system'. Secondary endpoints assess preliminary efficacy [e.g. mean change in office blood pressures (BPs)] and safety (e.g. hospitalization events) associated with each treatment protocol. Participants with both baseline and post-treatment BP measurements will form the intent-to-treat analysis. Following their involvement with the CURATE.AI intervention, feedback from CURATE.AI participants and healthcare providers will be collected via exit survey and interviews. Conclusion: Findings from this study will inform about potential refinements of the current treatment protocols before proceeding with a larger RCT, or potential expansion to collect additional information. Positive results may suggest the potential efficacy of CURATE.AI to improve BP control. Trial registration number: NCT05376683.

Comparing the Performance of Multiple Small-Data Personalized Tacrolimus Dosing Models for Pediatric Liver Transplant: A Retrospective Study.

Tacrolimus is a potent immunosuppressant used after pediatric liver transplant. However, tacrolimus's narrow therapeutic window, reliance on physicians' experience for the dose titration, and intra- and inter-patient variability result in liver transplant patients falling out of the target tacrolimus trough levels frequently. Existing personalized dosing models based on the area-under-the-concentration over time curves require a higher frequency of blood draws than the current standard of care and may not be practically feasible. We present a small-data artificial intelligence-derived platform, CURATE.AI, that uses data from individual patients obtained once daily to model the dose and response relationship and identify suitable doses dynamically. Retrospective optimization using 6 models of CURATE.AI and data from 16 patients demonstrated good predictive performance and identified a suitable model for further investigations.Clinical Relevance- This study established and compared the predictive performance of 6 personalized tacrolimus dosing models for pediatric liver transplant patients and identified a suitable model with consistently good predictive performance based on data from pediatric liver transplant patients.

Exploring cadmium stress responses in sisal roots: Insights from biochemical and transcriptome analysis.

Sisal is a leaf fiber crop with a high integrated value and a wide range of uses in the application of soil remediation of heavy metal contamination. This study provides a preliminary understanding of how sisal responds to Cd stress and presents a theoretical basis for exploring the potential of sisal in the remediation of Cd-contaminated soils. In this work, the activities of the antioxidant enzymes (SOD, POD, and CAT) of sisal were measured by hydroponics with the addition of CdCl2·2.5H2O and different concentrations of Cd stress. Whole transcriptome sequencing (RNA-Seq) analysis was performed with lllumina sequencing technology, and qRT-PCR was conducted to verify the differential genes. The results obtained were as follows: (1) Short-term low concentration of Cd stress (20 mg/kg) had a transient promotion effect on the growth of sisal roots, but Cd showed a significant inhibitory effect on the growth of sisal roots over time. (2) Under different concentrations of Cd stress, the Cd content in sisal root was greater than that in sisal leaf, and Cd accumulated mainly in sisal roots. (3) With the increase of Cd stress concentration, the antioxidant enzyme catalase activity increased, peroxidase activity showed a decreasing trend, and superoxide dismutase showed a trend of increasing and then decreasing. (4) Transcriptome sequencing analysis detected 123 differentially expressed genes (DEGs), among which 85 genes were up-regulated and 38 genes were down-regulated. The DEGs were mainly concentrated in flavonoid biosynthesis and glutathione metabolism, and both processes had some regulatory effects on the Cd tolerance characteristics of sisal. This study elucidated the physiological, biochemical and transcriptomic responses of sisal under cadmium stress, and provided a theoretical basis for the ecological restoration function of sisal.

Transcriptome Sequencing of Agave amaniensis Reveals Shoot-Related Expression Patterns of Expansin A Genes in Agave.

Agave species are widely planted for fiber production. However, the molecular basis of agave fiber development has not been well understood. In this study, we performed a transcriptomic analysis in A. amaniensi, a well-known variety with high-quality fiber production. Approximately 43.87 million clean reads were obtained using Illumina sequencing. The de novo assembly produced 66,746 unigrams, 54% of which were annotated in a public database. In the Nr database, 21,490 unigenes of A. amaniensis were shown to be most closely related to Asparagus officinalis. Nine expansin A orthologs with full coding regions were obtained, which were named EXP1a, EXP1b, EXP2, EXP3, EXP4a, EXP4b, EXP11, EXP12, and EXP13. The maximum likelihood phylogenetic tree revealed the species-specific expansion of expansin genes in Arabidopsis, rice and agave. The expression analysis suggested the negative correlation between the expression of expansin genes and the leaf growth rate, except AhEXP11. Moreover, expansin genes were differentially affected by abiotic and biotic stresses. Notably, AhEXP2 expression level was highly upgraded after the infection of Phytophthora nicotiana. Nutrient deficiency also influent expansin genes expression. Together, our research will benefit future studies related to fiber development, disease resistance and nutrient usage in agave.

Digital game-based interventions for cognitive training in healthy adults and adults with cognitive impairment: protocol for a two-part systematic review and meta-analysis.

INTRODUCTION: Digital game-based training interventions are scalable solutions that may improve cognitive function for many populations. This protocol for a two-part review aims to synthesise the effectiveness and key features of digital game-based interventions for cognitive training in healthy adults across the life span and adults with cognitive impairment, to update current knowledge and impact the development of future interventions for different adult subpopulations. METHODS AND ANALYSIS: This systematic review protocol follows the Preferred Reporting Items for Systematic Reviews and Meta-Analyses Protocols guidelines. A systematic search was performed in PubMed, Embase, CINAHL, Cochrane Library, Web of Science, PsycINFO and IEEE Explore on 31 July 2022 for relevant literature published in English from the previous 5 years. Experimental, observational, exploratory, correlational, qualitative and mixed methods studies will be eligible if they report at least one cognitive function outcome and include a digital game-based intervention intended to improve cognitive function. Reviews will be excluded but retained to search their reference lists for other relevant studies. All screening will be done by at least two independent reviewers. The appropriate Joanna Briggs Institute Critical Appraisal Tool, according to the study design, will be applied to perform the risk of bias assessment. Outcomes related to cognitive function and digital game-based intervention features will be extracted. Results will be categorised by adult life span stages in the healthy adult population for part 1 and by neurological disorder in part 2. Extracted data will be analysed quantitatively and qualitatively, according to study type. If a group of sufficiently comparable studies is identified, we will perform a meta-analysis applying the random effects model with consideration of the I2 statistic. ETHICS AND DISSEMINATION: Ethics approval is not applicable for this study since no original data will be collected. The results will be disseminated through peer-reviewed publications and conference presentations. PROSPERO REGISTRATION NUMBER: CRD42022351265.

Development of a Specific Nested PCR Assay for the Detection of 16SrI Group Phytoplasmas Associated with Sisal Purple Leafroll Disease in Sisal Plants and Mealybugs.

Sisal purple leafroll disease (SPLD) is currently the most destructive disease affecting sisal in China, yet its aetiology remains unclear. In our previous research, it was verified to be associated with phytoplasmas, and nested PCR based on the 16S rRNA gene using universal primers R16mF2/R16mR1 followed by R16F2n/R16R2 was confirmed as the most effective molecular method for the detection of phytoplasmas associated with SPLD (SPLDaP). However, the method has a shortcoming of inaccuracy, for it could produce false positive results. To further manage the disease, accurate detection is needed. In this study, we developed a specific nested PCR assay using universal primers R16F2n/R16R2, followed by a set of primers designed on 16Sr gene sequences amplified from SPLDaP, nontarget bacteria from sisal plants, and other phytoplasma subgroups or groups. This established method is accurate, specific, and effective for detection of 16SrI group phytoplasma in sisal, and its sensitivity is up to 10 fg/µL of total DNA. It also minimized the false positive problem of nested PCR using universal primers R16mF2/R16mR1 followed by R16F2n/R16R2. This method was further used to verify the presence of phytoplasma in Dysmicoccusneobrevipes, and the results showed that D. neobrevipes could be infected by SPLDaP and thus could be a candidate for vector transmission assays.

IDentif.AI-Omicron: Harnessing an AI-Derived and Disease-Agnostic Platform to Pinpoint Combinatorial Therapies for Clinically Actionable Anti-SARS-CoV-2 Intervention.

Nanomedicine-based and unmodified drug interventions to address COVID-19 have evolved over the course of the pandemic as more information is gleaned and virus variants continue to emerge. For example, some early therapies (e.g., antibodies) have experienced markedly decreased efficacy. Due to a growing concern of future drug resistant variants, current drug development strategies are seeking to find effective drug combinations. In this study, we used IDentif.AI, an artificial intelligence-derived platform, to investigate the drug-drug and drug-dose interaction space of six promising experimental or currently deployed therapies at various concentrations: EIDD-1931, YH-53, nirmatrelvir, AT-511, favipiravir, and auranofin. The drugs were tested in vitro against a live B.1.1.529 (Omicron) virus first in monotherapy and then in 50 strategic combinations designed to interrogate the interaction space of 729 possible combinations. Key findings and interactions were then further explored and validated in an additional experimental round using an expanded concentration range. Overall, we found that few of the tested drugs showed moderate efficacy as monotherapies in the actionable concentration range, but combinatorial drug testing revealed significant dose-dependent drug-drug interactions, specifically between EIDD-1931 and YH-53, as well as nirmatrelvir and YH-53. Checkerboard validation analysis confirmed these synergistic interactions and also identified an interaction between EIDD-1931 and favipiravir in an expanded range. Based on the platform nature of IDentif.AI, these findings may support further explorations of the dose-dependent drug interactions between different drug classes in further pre-clinical and clinical trials as possible combinatorial therapies consisting of unmodified and nanomedicine-enabled drugs, to combat current and future COVID-19 strains and other emerging pathogens.

The complete chloroplast genome of Agave amaniensis (Asparagales: Asparagaceae: Agavoideae).

Agave amaniensis Trel. & W. Nowell (1933) has long been used for phytosteroid production, which is also one of the parents of the famous Agave hybrid cultivar 11648 for sisal fiber production. However, its systematic position and phylogenetic relationship remains unknown at the chloroplast (cp) genome level. Therefore, we have sequenced and assembled the cp genome of A. amaniensis via Illumina sequencing. The cp genome is 157,282 bp in length with a GC content of 37.84%. A large single-copy region of 85,899 bp, a small single-copy region of 18,233 bp, and inverted repeat regions of 26,575 bp were found in the cp genome. Based on the annotation, 86 protein-coding genes, eight rRNAs, and 38 tRNAs were identified in the cp genome with total lengths of 78,981 bp, 9050 bp, and 2867 bp, respectively. The phylogenetic tree indicates that A. amaniensis is closely related with A. H11648, A. angustifolia, and A. americana.

Phylogeny and Expression Atlas of the NITRATE TRANSPORTER 1/PEPTIDE TRANSPORTER FAMILY in Agave.

Agave species are widely cultivated crassulacean acid metabolism (CAM) plants for alcoholic beverages, food and fiber production. Among these, the Agave hybrid H11648 ((A. amaniensis × A. angustifolia) × A. amaniensis) is the main cultivar for sisal fiber in the tropical areas of Brazil, China, and African countries. The plants of Agave hybrid H11648 have a long life cycle and large leaves, which require a huge amount of nitrogen nutrient. However, the molecular basis of nitrogen transport and allocation has not been well understood in agave. In this study, we identified 19 NITRATE TRANSPORTER 1/PEPTIDE TRANSPORTER FAMILY(NPF) genes (called AhNPFs) with full-length coding sequences in Agave hybrid H11648. Our analysis of gene expression in various types of tissues revealed the tissue-specific expression pattern of AhNPFs. We further examined their expression patterns at different leaf developmental stages, under abiotic/biotic stresses and nutrient deficiency. The results reveal several candidate regulators in the agave NPF family, including AhNPF4.3/5.2/7.1. We first characterized the NPF genes in agave based on published leaf transcriptome datasets and emphasized their potential functions. The study will benefit future studies related to nitrogen nutrient in agave.

The complete chloroplast genome of Agave sisalana.

Agave sisalana is one of the Agave cultivars for sisal fiber production around the tropical areas of the world. In the present study, we successfully sequenced and assembled its chloroplast genome. The full size of the genome is 157,268 bp with a GC content at 37.85%. The genome is constructed with a large single copy region (LSC, 85,894 bp), a pair of inverted repeat regions (IR, 26,573 bp) and a small single copy region (SSC, 18,228 bp). Besides, 86 protein-coding genes, 38 tRNAs and 8 rRNAs are annotated on the chloroplast genome. Phylogenetic result reveals that A. sisalana is closely related with A. americana and A. H11648.

First Report of Paramyrothecium breviseta Causing Leaf Spot Disease of Coffea canephora in China.

Coffee is a tropical plant with two widely cultivated species, namely Coffea arabica and Coffea canephora. A leaf spot disease causing brownish and necrotic lesions was broken out on the C. canephora coffee seedlings in a nursery in Ruili County, Yunnan Province, China, during 2018 to 2019. The incidence of the disease was 15% ~ 20%. Ten diseased leaf samples from five diseased plants were collected for pathogen isolation by tissue separation method. Leaf pieces were cut from the margin of the necrotic lesions (4 × 6 mm), surface-sterilized for 30 s in 75% ethanol, followed by 0.1% arsenic mercury solution for 15 s, then washed 3~4 times with sterilized distilled water and transferred onto potato dextrose agar (PDA) medium in petri plates. Four morphologically similar isolates were obtained from lesions and cultivated on PDA at 25°C. Initial colonies of isolates were round, neat edge, white, floccose mycelium and developed dark green-to-black concentric rings that were sporodochia bearing viscid spore masses after 5~7 days. Conidia were acetates, hyaline and cylindrical with both rounded ends and 4.8 to 6.4 µm long × 1.6 to 2.6 µm wide. Koch's test were conducted on three healthy plants leaves of original source variety C. canephora No.2 and C.arabica Catimor CIFC7963 (control plants) with spore suspension (1 × 106/mL), respectively. Meanwhile, equal numbers of healthy plants were inoculated with water as controls. After inoculation, the plants were transferred into an incubator at 25℃ with saturated humidity. After 10 days of inoculation, all the tested plants presented similar typical symptoms with the diseased leaves under natural conditions; whereas the controls remained healthy. Koch's postulates were performed by re-isolating the fungus from the inoculated leaves and verifying its colony and morphological characters. Two single spore isolates cultured on PDA medium were selected for DNA extraction. The ribosomal internal transcribed spacer (ITS) was PCR amplified by using primers ITS1 and ITS4 (White et al., 1990), ß-tubulin gene by Bt2a and Bt2b (Glass and Donaldson, 1995), the RNA polymerase II second largest subunit (rpb2) by RPB2-5F2 and RPB2-7cR (O'Donnell et al, 2007), calmodulin (cmda) gene by CAL-228F and CAL2Rd (Groenewald et al., 2013). The sequences of ITS (MT853067 ~ MT853068), ß-tubulin (MT897899 ~ MT897900), rpb2 (MW256264~ MW286265) and cmda (MT897897~ MT897898) were deposited in GenBank databases. BLAST analysis revealed that the representative isolates sequences shared 99.31%~99.65% similarities to the ITS sequence of Paramyrothecium breviseta (Accession Nos. NR_155670.1), 99.43% similarities to the ß-tubulin sequence of P. breviseta (Accession Nos. KU846406.1), 98.98% similarities to the rpb2 sequence of P. breviseta (Accession Nos. KU846351.1), and 98.54%~98.71% similarities to the cmda sequence of P. breviseta (Accession Nos. KU846262.1). As it shown in the phylogenetic tree derived from combined ITS, ß-tubulin, rpb2, and cmda gene sequences, the two representative isolates were clustered together with P. breviseta CBS 544.75 with 98% strong bootstrap support, which confirmed that P. breviseta is the causal agent of leaf spot of Coffea canephora. To our knowledge, this is the first report of a leaf spot disease caused by P. breviseta on C. canephora in China, which raised the caution that P. breviseta is also pathogenic to Coffea Arabica.

Effect of returning biochar from different pyrolysis temperatures and atmospheres on the growth of leaf-used lettuce.

The aim of this study is to evaluate the effects of biochar on the plant's growth. A pot experiment was carried out in our study. Rice straw-derived biochar were charred at two heating temperatures (400 °C/800 °C) and two oxygen-limited atmospheres (CO2/N2), respectively. The FESEM/EDS technique (field emission scanning electron microscopy with X-ray energy-dispersive spectroscopy) was used to study soils, biochar and plant samples. FESEM images indicated that the structure of the biochar was highly heterogeneous with larger macropores, which can enhance soil porosity. Fine soil mineral particles blocked the biochar inner pores and channels after returning biochar to soil. EDS analysis indicated that the Al and Fe contents increased on the surface of biochar after their returning, which reduced the toxicity of Al and Fe in the soil. The returning straw directly inhibited the growth of leaf-used lettuce. Four returning biochar all significantly improved leaf-used lettuce growth, and the effects of biochar prepared under 400 °C and a CO2 atmosphere were better than those prepared under 800 °C and a N2 atmosphere. Changes of nitrogen content in the biochar before and after their returning were consistent with the improvement of soil available nitrogen, and plant growth was positively correlated with the nitrogen content of biochar. This study explored the impact of biochar on soil nutrients and revealed the mechanism of biochar returning to the field to promote plant growth. It is of great significance in studying and improving the characteristics of soil nutrients.

Preparation of high porosity biochar materials by template method: a review.

Biochar plays an important role in soil improvement, pollutant removal, and nitrogen reduction. The excellent adsorption performance of biochar is closely related to its pore structure. Therefore, this paper combines a large amount of literatures to investigate the principle and method of preparing carbon materials by using the template method, and the idea of preparing high porosity biochar by template method was proposed. The results show that: (1) The specific surface area of the carbon materials prepared by the template method is more than 400 m2 g-1, and the total pore volume is more than 0.3 cm3 g-1, which is much higher than the biochar materials prepared under the traditional high temperature anoxic pyrolysis. (2) Compared with the hard template method, a soft template method with simple operation, low toxicity of the compound, and low cost is selected. (3) The lignin, which is also a hydrophilic carbon source similar to phenolic resin, can be used as an ideal carbon precursor. (4) In the selection of templating agents, the specific surface area and total pore volume of carbon materials prepared by using F127 as a template are relatively large, showing more excellent pore size performance. (5) Finally, the idea of using template method to prepare high porosity biochar is proposed: lignin extracted from straw material is used as precursor, block polymer F127 is used as template, an appropriate amount of a cross-linking agent and a solvent is added, and finally the target biochar material is prepared by pyrolysis carbonization.

Agrobacterium tumefaciens-mediated transformation of a hevein-like gene into asparagus leads to stem wilt resistance.

Asparagus stem wilt, is a significant and devastating disease, typically leading to extensive economic losses in the asparagus industry. To obtain transgenic plants resistant to stem wilt, the hevein-like gene, providing broad spectrum bacterial resistance was inserted into the asparagus genome through Agrobacterium tumefaciens-mediated transformation. The optimal genetic transformation system for asparagus was as follows: pre-culture of embryos for 2 days, inoculation using a bacterial titre of OD600 = 0.6, infection time 10 min and co-culturing for 4 days using an Acetosyringone concentration of 200 µmol/L. Highest transformation frequencies reached 21% and ten transgenic asparagus seedlings carrying the hevein-like gene were identified by polymerase chain reaction. Moreover, integration of the hevein-like gene in the T1 generation of transgenic plants was confirmed by southern blot hybridization. Analysis showed that resistance to stem wilt was enhanced significantly in the transgenic plants, in comparison to non- transgenic plants. The results provide additional data for genetic improvement and are of importance for the development of new disease-resistant asparagus varieties.

Identification and Expression of SAUR Genes in the CAM Plant Agave.

Agave species are important crassulacean acid metabolism (CAM) plants and widely cultivated in tropical areas for producing tequila spirit and fiber. The hybrid H11648 of Agave ((A. amaniensis × A. angustifolia) × A. amaniensis) is the main cultivar for fiber production in Brazil, China, and African countries. Small Auxin Up-regulated RNA (SAUR) genes have broad effect on auxin signaling-regulated plant growth and development, while only few SAUR genes have been reported in Agave species. In this study, we identified 43, 60, 24, and 21 SAUR genes with full-length coding regions in A. deserti, A. tequilana, A. H11648, and A. americana, respectively. Although phylogenetic analysis revealed that rice contained a species-specific expansion pattern of SAUR gene, no similar phenomena were observed in Agave species. The in silico expression indicated that SAUR genes had a distinct expression pattern in A. H11648 compared with other Agave species; and four SAUR genes were differentially expressed during CAM diel cycle in A. americana. Additionally, an expression analysis was conducted to estimate SAUR gene expression during different leaf developmental stages, abiotic and biotic stresses in A. H11648. Together, we first characterized the SAUR genes of Agave based on previously published transcriptome datasets and emphasized the potential functions of SAUR genes in Agave's leaf development and stress responses. The identification of which further expands our understanding on auxin signaling-regulated plant growth and development in Agave species.