Bt-Modified Transgenic Rice May Shift the Composition and Diversity of Rhizosphere Microbiota.

Plants significantly shape root-associated microbiota, making rhizosphere microbes useful environmental indicator organisms for safety assessment. Here, we report the pyrosequencing of the bacterial 16S ribosomal RNA in rhizosphere soil samples collected from transgenic cry1Ab/cry1Ac Bt rice Huahui No. 1 (GM crop) and its parental counterpart, Minghui63. We identified a total of 2579 quantifiable bacterial operational taxonomic units (OTUs). Many treatment-enriched microbial OTUs were identified, including 14 NonGM-enriched OTUs and 10 GM-enriched OTUs. OTUs belonging to the phyla Proteobacteria, Actinobacteria, Acidobacteria, Firmicutes, Nitrospirae, Chlorobi and GN04 were identified as statistically different in abundance between GM and the other two treatments. Compared with the different impacts of different rice varieties on microbiota, the impact of rice planting on microbiota is more obvious. Furthermore, Huahui No. 1 transgenic Bt rice had a greater impact on the rhizosphere bacterial communities than Minghui63. Early developmental stages of the transgenic Bt rice had a significant impact on many Bacillaceae communities. Soil chemical properties were not significantly altered by the presence of transgenic Bt rice. The peak concentration level of Bt protein products was detected during the seedling stage of transgenic Bt rice, which may be an intriguing factor for bacterial diversity variations. Based on these findings, we conclude that transgenic Bt rice has a significant impact on root-associated bacteria. This information may be leveraged in future environmental safety assessments of transgenic Bt rice varieties.

Expression of Cry1Ab/2Aj Protein in Genetically Engineered Maize Plants and Its Transfer in the Arthropod Food Web.

While transgenic Bacillus thuringiensis (Bt) maize provides pest resistance and a reduced application of chemical pesticides, a comprehensive environmental risk assessment is mandatory before its field release. This research determined the concentrations of Bt protein in plant tissue and in arthropods under field conditions in Gongzhuling City, northeastern China, to provide guidance for the selection of indicator species for non-target risk assessment studies. Bt maize expressing Cry1Ab/2Aj and non-transformed near-isoline were grown under identical environmental and agricultural conditions. Cry1Ab/2Aj was detected in plant tissues and arthropods collected from Bt maize plots during pre-flowering, flowering, and post-flowering. The expression of Cry1Ab/2Aj varied across growth stages and maize tissues, as well as in the collected arthropods at the three growth stages. Therefore, representative species should be chosen to cover the whole growing season and to represent different habitats and ecological functions. Dalbulus maidis (Hemiptera: Cicadellidae), Rhopalosiphum padi (Hemiptera: Aphididae), Heteronychus arator (Coleoptera: Scarabaeidae), and Somaticus angulatus (Coleoptera: Tenebrionidae) are suitable non-target herbivores. Propylea japonica (Coleoptera: Coccinellidae), Paederus fuscipes (Coleoptera: Staphylinidae), Chrysoperla nipponensis (Neuroptera: Chrysopidae), and spiders are suggested predators. Apis cerana and Apis mellifera ligustica (both Hymenoptera: Apidae) represent pollinators and Folsomia candida (Collembola: Isotomidae) decomposers.

Genomic and Transcriptomic Insights into the Evolution and Divergence of MIKC-Type MADS-Box Genes in Carica papaya.

MIKC-type MADS-box genes, also known as type II genes, play a crucial role in regulating the formation of floral organs and reproductive development in plants. However, the genome-wide identification and characterization of type II genes as well as a transcriptomic survey of their potential roles in Carica papaya remain unresolved. Here, we identified and characterized 24 type II genes in the C. papaya genome, and investigated their evolutional scenario and potential roles with a widespread expression profile. The type II genes were divided into thirteen subclades, and gene loss events likely occurred in papaya, as evidenced by the contracted member size of most subclades. Gene duplication mainly contributed to MIKC-type gene formation in papaya, and the duplicated gene pairs displayed prevalent expression divergence, implying the evolutionary significance of gene duplication in shaping the diversity of type II genes in papaya. A large-scale transcriptome analysis of 152 samples indicated that different subclasses of these genes showed distinct expression patterns in various tissues, biotic stress response, and abiotic stress response, reflecting their divergent functions. The hub-network of male and female flowers and qRT-PCR suggested that TT16-3 and AGL8 participated in male flower development and seed germination. Overall, this study provides valuable insights into the evolution and functions of MIKC-type genes in C. papaya.

Time-ordering japonica/geng genomes analysis indicates the importance of large structural variants in rice breeding.

Temperate japonica/geng (GJ) rice yield has significantly improved due to intensive breeding efforts, dramatically enhancing global food security. However, little is known about the underlying genomic structural variations (SVs) responsible for this improvement. We compared 58 long-read assemblies comprising cultivated and wild rice species in the present study, revealing 156 319 SVs. The phylogenomic analysis based on the SV dataset detected the putatively selected region of GJ sub-populations. A significant portion of the detected SVs overlapped with genic regions were found to influence the expression of involved genes inside GJ assemblies. Integrating the SVs and causal genetic variants underlying agronomic traits into the analysis enables the precise identification of breeding signatures resulting from complex breeding histories aimed at stress tolerance, yield potential and quality improvement. Further, the results demonstrated genomic and genetic evidence that the SV in the promoter of LTG1 is accounting for chilling sensitivity, and the increased copy numbers of GNP1 were associated with positive effects on grain number. In summary, the current study provides genomic resources for retracing the properties of SVs-shaped agronomic traits during previous breeding procedures, which will assist future genetic, genomic and breeding research on rice.

Reducing Seed Shattering in Weedy Rice by Editing SH4 and qSH1 Genes: Implications in Environmental Biosafety and Weed Control through Transgene Mitigation.

Mitigating the function of acquired transgenes in crop wild/weedy relatives can provide an ideal strategy to reduce the possible undesired environmental impacts of pollen-mediated transgene flow from genetically engineered (GE) crops. To explore a transgene mitigation system in rice, we edited the seed-shattering genes, SH4 and qSH1, using a weedy rice line ("C9") that originally had strong seed shattering. We also analyzed seed size-related traits, the total genomic transcriptomic data, and RT-qPCR expression of the SH4 or qSH1 gene-edited and SH4/qSH1 gene-edited weedy rice lines. Substantially reduced seed shattering was observed in all gene-edited weedy rice lines. The single gene-edited weedy rice lines, either the SH4 or qSH1 gene, did not show a consistent reduction in their seed size-related traits. In addition, reduced seed shattering was closely linked with the weakness and absence of abscission layers and reduced abscisic acid (ABA). Additionally, the genes closely associated with ABA biosynthesis and signaling transduction, as well as cell-wall hydrolysis, were downregulated in all gene-edited weedy rice lines. These findings facilitate our deep insights into the underlying mechanisms of reduced seed shattering in plants in the rice genus Oryza. In addition, such a mitigating technology also has practical applications for reducing the potential adverse environmental impacts caused by transgene flow and for managing the infestation of weedy rice by acquiring the mitigator from GE rice cultivars through natural gene flow.

Metabolomic and transcriptomic analyses reveal the mechanism of sweet-acidic taste formation during pineapple fruit development.

Pineapple (Ananas comosus L.) is one of the most valuable subtropical fruit crop in the world. The sweet-acidic taste of the pineapple fruits is a major contributor to the characteristic of fruit quality, but its formation mechanism remains elusive. Here, targeted metabolomic and transcriptomic analyses were performed during the fruit developmental stages in two pineapple cultivars ("Comte de Paris" and "MD-2") to gain a global view of the metabolism and transport pathways involved in sugar and organic acid accumulation. Assessment of the levels of different sugar and acid components during fruit development revealed that the predominant sugar and organic acid in mature fruits of both cultivars was sucrose and citric acid, respectively. Weighted gene coexpression network analysis of metabolic phenotypes and gene expression profiling enabled the identification of 21 genes associated with sucrose accumulation and 19 genes associated with citric acid accumulation. The coordinated interaction of the 21 genes correlated with sucrose irreversible hydrolysis, resynthesis, and transport could be responsible for sucrose accumulation in pineapple fruit. In addition, citric acid accumulation might be controlled by the coordinated interaction of the pyruvate-to-acetyl-CoA-to-citrate pathway, gamma-aminobutyric acid pathway, and tonoplast proton pumps in pineapple. These results provide deep insights into the metabolic regulation of sweetness and acidity in pineapple.

Genomic Analysis of LEA Genes in Carica papaya and Insight into Lineage-Specific Family Evolution in Brassicales.

Late embryogenesis abundant (LEA) proteins comprise a diverse superfamily involved in plant development and stress responses. This study presents a first genome-wide analysis of LEA genes in papaya (Carica papaya L., Caricaceae), an economically important tree fruit crop widely cultivated in the tropics and subtropics. A total of 28 members were identified from the papaya genome, which belong to eight families with defined Pfam domains, i.e., LEA_1 (3), LEA_2 (4), LEA_3 (5), LEA_4 (5), LEA_5 (2), LEA_6 (2), DHN (4), and SMP (3). The family numbers are comparable to those present in Ricinus communis (Euphorbiaceae, 28) and Moringa oleifera (Moringaceae, 29), but relatively less than that found in Moringa oleifera (Cleomaceae, 39) and Arabidopsis thaliana (Brassicaceae, 51), implying lineage-specific evolution in Brassicales. Indeed, best-reciprocal-hit-based sequence comparison and synteny analysis revealed the presence of 29 orthogroups, and significant gene expansion in Tarenaya and Arabidopsis was mainly contributed by whole-genome duplications that occurred sometime after their split with the papaya. Though a role of transposed duplication was also observed, tandem duplication was shown to be a key contributor in gene expansion of most species examined. Further comparative analyses of exon-intron structures and protein motifs supported fast evolution of this special superfamily, especially in Arabidopsis. Transcriptional profiling revealed diverse expression patterns of CpLEA genes over various tissues and different stages of developmental fruit. Moreover, the transcript level of most genes appeared to be significantly regulated by drought, cold, and salt stresses, corresponding to the presence of cis-acting elements associated with stress response in their promoter regions. These findings not only improve our knowledge on lineage-specific family evolution in Brassicales, but also provide valuable information for further functional analysis of LEA genes in papaya.

Cas12a-based on-site, rapid detection of genetically modified crops.

A CRISPR/LbCas12a-based nucleic acid detection method that uses crude leaf extracts as samples and is rapid (≤40 min for a full run) and highly sensitive (0.01%) can be used to monitor genetically modified organisms in the field.

Resequencing of 388 cassava accessions identifies valuable loci and selection for variation in heterozygosity.

BACKGROUND: Heterozygous genomes are widespread in outcrossing and clonally propagated crops. However, the variation in heterozygosity underlying key agronomic traits and crop domestication remains largely unknown. Cassava is a staple crop in Africa and other tropical regions and has a highly heterozygous genome. RESULTS: We describe a genomic variation map from 388 resequenced genomes of cassava cultivars and wild accessions. We identify 52 loci for 23 agronomic traits through a genome-wide association study. Eighteen allelic variations in heterozygosity for nine candidate genes are significantly associated with seven key agronomic traits. We detect 81 selective sweeps with decreasing heterozygosity and nucleotide diversity, harboring 548 genes, which are enriched in multiple biological processes including growth, development, hormone metabolisms and responses, and immune-related processes. Artificial selection for decreased heterozygosity has contributed to the domestication of the large starchy storage root of cassava. Selection for homozygous GG allele in MeTIR1 during domestication contributes to increased starch content. Selection of homozygous AA allele in MeAHL17 is associated with increased storage root weight and cassava bacterial blight (CBB) susceptibility. We have verified the positive roles of MeTIR1 in increasing starch content and MeAHL17 in resistance to CBB by transient overexpression and silencing analysis. The allelic combinations in MeTIR1 and MeAHL17 may result in high starch content and resistance to CBB. CONCLUSIONS: This study provides insights into allelic variation in heterozygosity associated with key agronomic traits and cassava domestication. It also offers valuable resources for the improvement of cassava and other highly heterozygous crops.

Genomic Analyses of Penicillium Species Have Revealed Patulin and Citrinin Gene Clusters and Novel Loci Involved in Oxylipin Production.

Blue mold of apple is caused by several different Penicillium species, among which P. expansum and P. solitum are the most frequently isolated. P. expansum is the most aggressive species, and P. solitum is very weak when infecting apple fruit during storage. In this study, we report complete genomic analyses of three different Penicillium species: P. expansum R21 and P. crustosum NJ1, isolated from stored apple fruit; and P. maximae 113, isolated in 2013 from a flooded home in New Jersey, USA, in the aftermath of Hurricane Sandy. Patulin and citrinin gene cluster analyses explained the lack of patulin production in NJ1 compared to R21 and lack of citrinin production in all three strains. A Drosophila bioassay demonstrated that volatiles emitted by P. solitum SA and P. polonicum RS1 were more toxic than those from P. expansum and P. crustosum strains (R27, R11, R21, G10, and R19). The toxicity was hypothesized to be related to production of eight-carbon oxylipins. Putative lipoxygenase genes were identified in P. expansum and P. maximae strains, but not in P. crustosum. Our data will provide a better understanding of Penicillium spp. complex secondary metabolic capabilities, especially concerning the genetic bases of mycotoxins and toxic VOCs.

Prolonged symptom onset to admission time is associated with severe Coronavirus disease: A meta combined propensity-adjusted analysis.

BACKGROUND: Patients with severe COVID-19 are more likely to develop adverse outcomes with a huge medical burden. We aimed to investigate whether a shorter symptom onset to admission time (SOAT) could improve outcomes of COVID-19 patients. METHODS: A single-center retrospective study combined with a meta-analysis was performed. The meta-analysis identified studies published between 1 December 2019 and 15 April 2020. Additionally, clinical data of COVID-19 patients diagnosed between January 20 and February 20, 2020, at the First Affiliated Hospital of the University of Science and Technology of China were retrospectively analyzed. SOAT and severity of illness in patients with COVID-19 were used as effect measures. The random-effects model was used to analyze the heterogeneity across studies. Propensity score matching was applied to adjust for confounding factors in the retrospective study. Categorical data were compared using Fisher's exact test. We compared the differences in laboratory characteristic varied times using a two-way nonparametric, Scheirer-Ray-Hare test. RESULTS: In a meta-analysis, we found that patients with adverse outcomes had a longer SOAT (I2 = 39%, mean difference 0.88, 95% confidence interval = 0.47-1.30). After adjusting for confounding factors, such as age, complications, and treatment options, the retrospective analysis results also showed that severe patients had longer SOAT (mean difference 1.13 [1.00, 1.27], p = 0.046). Besides, most biochemical marker levels improved as the hospitalization time lengthened without the effect of disease severity or associated treatment (p < 0.001). CONCLUSION: Shortening the SOAT may help reduce the possibility of mild patients with COVID-19 progressing to severe illness.

Design and synthesis of Grp94 selective inhibitors based on Phe199 induced fit mechanism and their anti-inflammatory effects.

Glucose-regulated protein 94 (Grp94), a member of the Heat shock protein 90 (Hsp90) family, is implicated in many human diseases, including cancer, neurodegeneration, inflammatory, and infectious diseases. Here, we describe our effort to design and develop a new series of Grp94 inhibitors based on Phe199 induced fit mechanism. Using an alkynyl-containing inhibitor as a starting point, we developed compound 4, which showed potent inhibitory activity toward Grp94 in a fluorescence polarization-based assay. With improved physicochemical properties and suitable pharmacokinetic properties, compound 4 was advanced into in vivo bioactivity evaluation. In a dextran sulfate sodium (DSS)-induced mouse model of ulcerative colitis (UC), compound 4 showed anti-inflammatory property and reduced the levels of pro-inflammatory cytokines (TNF-α and IL-6). Together, these findings provide evidence that this approach may be promising for further Grp94 drug development efforts.

Identification and Expression Analyses of the Special 14-3-3 Gene Family in Papaya and its Involvement in Fruit Development, Ripening, and Abiotic Stress Responses.

Plant 14-3-3 proteins play key roles in regulating growth, development, and stress responses. However, little is known about this gene family in papaya (Carica papaya L.). We characterized eight 14-3-3 genes from the papaya genome and designed them as CpGRF1-8. Based on phylogenetic, conserved motif, and gene structure analyses, papaya CpGRFs were divided into ε and non-ε groups. Expression analysis showed differential and class-specific transcription patterns in different organs. Quantitative real-time polymerase chain reaction analysis showed that most CpGRFs had large changes in expression during fruit development and ripening. This indicated that the CpGRFs were involved in regulating fruit development and ripening. Significant expression changes occurred after cold, salt, and drought treatments in papaya seedlings, indicating that CpGRFs were also involved in signaling responses to abiotic stress. These results provide a transcription profile of 14-3-3 genes in organs, during fruit development and ripening and in response to stress. Some highly expressed, fruit-specific, and stress-responsive candidate CpGRFs will be identified for further genetic improvement of papayas.

Risk factors on admission associated with hospital length of stay in patients with COVID-19: a retrospective cohort study.

Treating patients with COVID-19 is expensive, thus it is essential to identify factors on admission associated with hospital length of stay (LOS) and provide a risk assessment for clinical treatment. To address this, we conduct a retrospective study, which involved patients with laboratory-confirmed COVID-19 infection in Hefei, China and being discharged between January 20 2020 and March 16 2020. Demographic information, clinical treatment, and laboratory data for the participants were extracted from medical records. A prolonged LOS was defined as equal to or greater than the median length of hospitable stay. The median LOS for the 75 patients was 17 days (IQR 13-22). We used univariable and multivariable logistic regressions to explore the risk factors associated with a prolonged hospital LOS. Adjusted odds ratios (aORs) and 95% confidence intervals (CIs) were estimated. The median age of the 75 patients was 47 years. Approximately 75% of the patients had mild or general disease. The univariate logistic regression model showed that female sex and having a fever on admission were significantly associated with longer duration of hospitalization. The multivariate logistic regression model enhances these associations. Odds of a prolonged LOS were associated with male sex (aOR 0.19, 95% CI 0.05-0.63, p = 0.01), having fever on admission (aOR 8.27, 95% CI 1.47-72.16, p = 0.028) and pre-existing chronic kidney or liver disease (aOR 13.73 95% CI 1.95-145.4, p = 0.015) as well as each 1-unit increase in creatinine level (aOR 0.94, 95% CI 0.9-0.98, p = 0.007). We also found that a prolonged LOS was associated with increased creatinine levels in patients with chronic kidney or liver disease (p < 0.001). In conclusion, female sex, fever, chronic kidney or liver disease before admission and increasing creatinine levels were associated with prolonged LOS in patients with COVID-19.

Increases in Genetic Diversity of Weedy Rice Associated with Ambient Temperatures and Limited Gene Flow.

Hypotheses regarding the association of increased species or genetic diversity with gradually warmer regions as a global pattern have been proposed, but no direct and solid experimental data are available to approve the association between plant genetic diversity and ambient temperatures. To test the diversity-temperature hypothesis, we studied genetic diversity and genetic differentiation of weedy rice (Oryza sativa f. spontanea) populations occurring naturally in early- and late-season rice fields that share nearly the same ecological conditions but with slightly different temperatures. Data collected from 10-year historical climatic records indicated a ~2 ℃ higher average air temperature in the late rice-cultivation seasons than in the early seasons. Results based on molecular fingerprints of 27 SSR (simple sequence repeat) loci showed a higher level of genetic diversity in the late-season weedy rice populations than in the early-season populations. In addition, a positive correlation was detected between the increased proportion of genetic diversity (ΔHe ) and genetic differentiation among the weedy rice populations, suggesting limited gene flow. Therefore, we conclude from this study that increased genetic diversity in the late-season weedy rice populations is probably caused by the higher ambient temperatures. This finding provides evidence for the possible association between genetic diversity and ambient temperatures.

A new isolation device for shortening gene flow distance in small-scale transgenic maize breeding.

The transmission of pollen is the main cause of maize gene flow. Under the compulsory labeling system for genetically modified (GM) products in China, isolation measures are crucial. At present, there is no effective isolation device for preventing and controlling the short-range flow of GM maize pollen. The purposes of the present experiments were to overcome the deficiencies of existing technology and to demonstrate a new isolation device for decreasing the gene flow distance of GM maize. The isolation device we invented was shown to be more robust than traditional isolation methods, and it can be disassembled and repeatedly reused. The most important point was that the frequency of gene flow could be greatly reduced using this device. When the distance from the isolation device was more than 1 m, the gene flow rate could be decreased to less than 1%, and when the distance from the isolation device was more than 10 m, the gene flow rate could be reduced to less than 0.1%. When the isolation device was adopted to isolate GM maize in conjunction with bagging the tassels of GM maize at the pollination stage, the gene flow could be controlled to less than 0.1% when the distance from the isolation device was more than 1 m. This device was, however, only applicable for small plots and can shorten the isolation distance of GM maize planting and improve the purity of seeds, all while meeting the needs of close isolation breeding. The use of this device represents a feasible method for risk prevention and control of GM crops.

Genes encoding light-harvesting chlorophyll a/b-binding proteins in papaya (Carica papaya L.) and insight into lineage-specific evolution in Brassicaceae.

Light-harvesting chlorophyll a/b-binding (Lhc) proteins comprise a plant-specific superfamily involved in photosynthesis and stress responses. Despite their importance, little is known in papaya (Carica papaya), an economically important tree fruit crop as well as a species close to the model plant arabidopsis (Arabidopsis thaliana). This study reports a first genome-wide analysis of Lhc superfamily genes in papaya, and a total of 28 members that represent four defined families or 26 orthologous groups were identified from the papaya genome. The superfamily number is comparable to 28 or 27 reported in castor (Ricinus communis) and jatropha (Jatropha curcas), respectively, two Euphorbiaceous plants also without any recent whole-genome duplication (WGD), but relatively less than 35, 34, 32, 32, 37, 30 or 32 present in cassava (Manihot esculenta), arabidopsis, A. lyrata, A. halleri, Capsella rubella, C. grandiflora, and Eutrema salsugineum, respectively, representative species having experienced one or two recent WGDs. Local duplication was shown to play a predominant role in gene expansion in papaya, castor, and jatropha, which is only confined to the Lhcb1 group. By contrast, WGD plays a relatively more important role in cassava, arabidopsis, and other Brassicaceous plants. Further comparison of Brassicaceous plants revealed that loss of the SEP6 group in arabidopsis is lineage-specific, occurring sometime after papaya-arabidopsis divergence but before the radiation of Brassicaceous plants. Transcriptional profiling revealed a leaf-preferential expression pattern of most CpLhc superfamily genes and their transcript levels were markedly regulated by three abiotic stresses, i.e., mimicking drought, cold, and high salt. These findings not only facilitate further functional studies in papaya, but also improve our knowledge on lineage-specific evolution of this special gene superfamily in Brassicaceae.

Evaluation of genetic diversity within asparagus germplasm based on morphological traits and ISSR markers.

Asparagus officinalis L. is a dioecious perennial plant globally known for its fine flavor and high nutritional value. An evaluation of genetic diversity in 46 asparagus accessions was carried out based on morphological and inter-simple sequence repeat (ISSR) markers. The result show that the coefficient of variation for 20 morphological characteristics is between 12.45 and 62.22%. Factor analysis revealed that nine factors explained 83.37% of the total variance. At Euclidean distance of 135.7, 46 accessions were divided into two clusters. Genetic similarity coefficient (GSC) based on ISSR data ranged from 0.60 to 0.97, suggesting a relatively abundant genetic base. Furthermore, the 46 asparagus accessions could also be grouped into three major clusters at a GSC of 0.74. And there is no significant relation between the two marker systems using the Mantel test. Clustering based on morphological traits compared with that based on ISSR data was not consistent, however, some common groupings were observed between two dendrograms. Therefore the results elucidated asparagus germplasm genetic background and determined hybrid parents, which will facilitate optimal application of asparagus germplasm resources and provide additional data for genetic improvement.