Extraction Methods Determine the Quality of Soil Microbiota Acquisition.

The soil microbiome plays a key role in plant health. Native soil microbiome inoculation, metagenomic profiling, and high-throughput cultivation require efficient microbe extraction. Sonication and oscillation are the most common methods used to extract soil microbiomes. However, the extraction efficiency of these methods has not been investigated in full. In this study, we compared the culturable microbe numbers, community structures, and alpha diversities among the different methods, including sonication, oscillation, and centrifugation, and their processing times. The study results showed that sonication significantly increases the culturable colony number compared with oscillation and centrifugation. Furthermore, the sonication strategy was found to be the main factor influencing extraction efficiency, but increased sonication time can aid in recovery from this impact. Finally, the extraction processing times were found to have a significant negative relationship with α-diversity among the extracted microbiota. In conclusion, sonication is the main factor for enriching in situ microbiota, and increased extraction time significantly decreases the α-diversity of the extracted microbiota. The results of this study provide insights into the isolation and utilization of different microorganism sources.

Integrative transcriptomic, proteomic, and phosphoproteomic analysis on the defense response to Magnaporthe oryzae reveals different expression patterns at the molecular level of durably resistant rice cultivar Mowanggu.

Rice blast, caused by Magnaporthe oryzae is one of the most destructive diseases of rice (Oryza sativa L.) in most rice-cultivated areas worldwide. Mowanggu (MWG) is a traditional landrace rice variety in Yunnan with broad-spectrum and durable blast resistance against rice blast fungus. However, the underlying disease-resistance mechanisms remain unknown. An integrative transcriptomic, proteomic, and phosphoproteomic analysis of MWG was performed after inoculation with M. oryzae in this study. The transcriptomic and proteomic results revealed that MWG was moderately correlated at the transcriptional and protein levels. Differentially expressed genes and proteins were up-regulated and significantly enriched in protein phosphorylation, peroxisome, plant-pathogen interactions, phenylpropanoid metabolism and phenylalanine biosynthesis pathways. The phosphoproteomic profile and phosphorylated-protein-interaction network revealed that the altered phosphoproteins were primarily associated with reactive oxygen species (ROS), glycolysis, MAPK signaling pathways, and amino acid biosynthesis. In addition, a series of physiological and biochemical parameters, including ROS, soluble sugars, soluble protein and callus accumulation and defense-related enzyme activities, were used to validate the possible blast resistance mechanisms of MWG. The integrative transcriptomic, proteomic, and phosphoproteomic analysis revealed the different expression patterns at the molecular level of the durably resistant rice cultivar MWG after inoculation with M. oryzae, which provides insight into the molecular mechanisms of rice blast resistance.

Identification and characterization of pathogenicity-related genes of Rhizoctonia solani AG3 during tobacco infection.

Tobacco target spot disease is caused by a ubiquitous soil-borne phytopathogen Rhizoctonia solani; the pathogenic mechanisms underlying the effects of R. solani remain unclear. Deeper understanding of the functional responses to R. solani during host plant infection would help identify the molecular mechanisms essential for successful host invasion. In this study, we performed global transcriptional analysis of R. solani during various stages (12, 24, 48, 72, 96, and 120 h) of tobacco infection via an RNA sequencing method, while utilizing the pathosystem model R. solani AG3-tobacco (Nicotiana tabacum L.). After R. solani inoculation, the number of differentially expressed genes of R. solani differed at the various time points. Moreover, several gene ontology and Kyoto encyclopedia of genes and genomes pathways were unique in different infection stages, especially with respect to the genes involved in plant cell wall degradation and catalysis of biotransformation reactions, such as the pectin metabolic process and pectin catabolic process. The overexpressing-PD8 N. benthamiana plants enhanced the susceptibility to R. solani. In addition, we found that large amounts of reactive oxygen species (ROS) were generated in tobacco after infected by R. solani. R. solani encoding FAD/NAD binding oxidoreductase and peroxidase gene family to eliminating ROS and counteract oxidative stress. Moreover, Perox3 was validated that can enhance the ability of scavenging ROS by co-injecting. Overall, our findings show that pectin-degrading enzymes and cytochrome P450 genes are critical for plant infection. These results provide comprehensive insights into R. solani AG3 transcriptome responses during tobacco invasion.

Integrated Analysis of MicroRNA and Target Genes in Brachypodium distachyon Infected by Magnaporthe oryzae by Small RNA and Degradome Sequencing.

Rice blast caused by Magnaporthe oryzae is one of the most important diseases that seriously threaten rice production. Brachypodium distachyon is a grass species closely related to grain crops, such as rice, barley, and wheat, and has become a new model plant of Gramineae. In this study, 15 small RNA samples were sequenced to examine the dynamic changes in microRNA (miRNA) expression in B. distachyon infected by M. oryzae at 0, 24, and 48 h after inoculation. We identified 432 conserved miRNAs and 288 predicted candidate miRNAs in B. distachyon. Additionally, there were 7 and 19 differentially expressed miRNAs at 24 and 48 h post-inoculation, respectively. Furthermore, using degradome sequencing, we identified 2,126 genes as targets for 308 miRNAs; using quantitative real-time PCR (qRT-PCR), we validated five miRNA/target regulatory units involved in B. distachyon-M. oryzae interactions. Moreover, using co-transformation technology, we demonstrated that BdNAC21 was negatively regulated by miR164c. This study provides a new approach for identifying resistance genes in B. distachyon by mining the miRNA regulatory network of host-pathogen interactions.

Genome-Wide Characterization and Expression Analysis of the SBP-Box Gene Family in Sweet Orange (Citrus sinensis).

SBP-box is an important plant-specific transcription factor family and is involved in diverse biological processes. Here, we identified a total of 15 SBP-BOX genes in the important fruit crop sweet orange (Citrus sinensis) and characterized their gene structures, conserved domain and motif, chromosomal location, and cis-acting regulatory elements. SBP genes were classified into four subfamilies based on the amino acid sequence homology, and the classification is equally strongly supported by the gene and protein structures. Our analysis revealed that segmental duplication events were the main driving force in the evolution of CsSBP genes, and gene pairs might undergo extensive purifying selection. Further synteny analysis of the SBP members among sweet orange and other plant species provides valuable information for clarifying the CsSBP family evolutionary relationship. According to publicly available RNA-seq data and qRT-PCR analysis from various sweet orange tissues, CsSBP genes may be expressed in different tissues and developmental stages. Gene expression analysis showed variable expression profiles of CsSBP genes under various abiotic stresses, such as high and low-temperature, salt, and wound treatments, demonstrating the potential role of SBP members in sweet orange response to abiotic stress. Noticeably, all CsSBP genes were also downregulated in sweet orange upon the infection of an important fungal pathogen Diaporthe citri. Our results provide valuable information for exploring the role of SBP-Box in sweet orange.

Genome-Wide Characterization, Evolution, and Expression Profile Analysis of GATA Transcription Factors in Brachypodium distachyon.

The GATA proteins, functioning as transcription factors (TFs), are involved in multiple plant physiological and biochemical processes. In this study, 28 GATA TFs of Brachypodium distachyon (BdGATA) were systematically characterized via whole-genome analysis. BdGATA genes unevenly distribute on five chromosomes of B. distachyon and undergo purifying selection during the evolution process. The putative cis-acting regulatory elements and gene interaction network of BdGATA were found to be associated with hormones and defense responses. Noticeably, the expression profiles measured by quantitative real-time PCR indicated that BdGATA genes were sensitive to methyl jasmonate (MeJA) and salicylic acid (SA) treatment, and 10 of them responded to invasion of the fungal pathogen Magnaporthe oryzae, which causes rice blast disease. Genome-wide characterization, evolution, and expression profile analysis of BdGATA genes can open new avenues for uncovering the functions of the GATA genes family in plants and further improve the knowledge of cellular signaling in plant defense.

TALE Transcription Factors in Sweet Orange (Citrus sinensis): Genome-Wide Identification, Characterization, and Expression in Response to Biotic and Abiotic Stresses.

Three-amino-acid-loop-extension (TALE) transcription factors comprise one of the largest gene families in plants, in which they contribute to regulation of a wide variety of biological processes, including plant growth and development, as well as governing stress responses. Although sweet orange (Citrus sinensis) is among the most commercially important fruit crops cultivated worldwide, there have been relatively few functional studies on TALE genes in this species. In this study, we investigated 18 CsTALE gene family members with respect to their phylogeny, physicochemical properties, conserved motif/domain sequences, gene structures, chromosomal location, cis-acting regulatory elements, and protein-protein interactions (PPIs). These CsTALE genes were classified into two subfamilies based on sequence homology and phylogenetic analyses, and the classification was equally strongly supported by the highly conserved gene structures and motif/domain compositions. CsTALEs were found to be unevenly distributed on the chromosomes, and duplication analysis revealed that segmental duplication and purifying selection have been major driving force in the evolution of these genes. Expression profile analysis indicated that CsTALE genes exhibit a discernible spatial expression pattern in different tissues and differing expression patterns in response to different biotic/abiotic stresses. Of the 18 CsTALE genes examined, 10 were found to be responsive to high temperature, four to low temperature, eight to salt, and four to wounding. Moreover, the expression of CsTALE3/8/12/16 was induced in response to infection with the fungal pathogen Diaporthe citri and bacterial pathogen Candidatus Liberibacter asiaticus, whereas the expression of CsTALE15/17 was strongly suppressed. The transcriptional activity of CsTALE proteins was also verified in yeast, with yeast two-hybrid assays indicating that CsTALE3/CsTALE8, CsTALE3/CsTALE11, CsTALE10/CsTALE12, CsTALE14/CsTALE8, CsTALE14/CsTALE11 can form respective heterodimers. The findings of this study could lay the foundations for elucidating the biological functions of the TALE family genes in sweet orange and contribute to the breeding of stress-tolerant plants.