Application of pectin hydrolyzing bacteria in tobacco to improve flue-cured tobacco quality.

To study the relationship between the diversity of the surface microbial community and tobacco flavor, and to improve tobacco quality using microorganisms. The microbial community composition and diversity of 14 samples of flue-cured tobacco from tobacco-producing areas in Yunnan with varying grades were analyzed by high-throughput sequencing. PICRUSt was used for predicting microbial functions. A strain of Bacillus amyloliquefaciens W6-2 with the ability to degrade pectin was screened from the surface of flued-cured tobacco leaves from Yunnan reroasted tobacco leave. The enzyme preparation was prepared through fermentation and then applied for treating flue-cured tobacco. The improvement effect was evaluated by measuring the content of macromolecule and the changes in volatile components, combined with sensory evaluations. The bacterial communities on the surface of flue-cured tobacco exhibited functional diversity, consisting primarily of Variovorax, Pseudomonas, Sphingomonas, Burkholderia, and Bacillus. These bacterial strains played a role in the aging process of flue-cured tobacco leaves by participating in amino acid metabolism and carbohydrate metabolism. These metabolic activity converted complex macromolecules into smaller molecular compounds, ultimately influence the smoking quality and burning characteristics of flue-cured tobacco. The pectinase preparation produced through fermentation using W6-2 has been found to enhance the aroma and sweetness of flue-cured tobacco, leading to improved aroma, reduced impurities, and enhanced smoothness. Additionally, the levels of pectin, cellulose, and hemicellulose decreased, while the levels of water-soluble sugar and reducing sugar increased, and the contents of esters, ketones, and aldehydes increased, and the contents of benzoic acid decreased. The study revealed the correlation between surface microorganisms and volatile components of Yunnan tobacco leaves, and the enzyme produced by the pectin-degrading bacteria W6-2 effectively improved the quality of flue-cured tobacco.

Endoplasmic reticulum membrane protein MoScs2 is important for asexual development and pathogenesis of Magnaporthe oryzae.

Most secretory proteins are folded and modified in the endoplasmic reticulum (ER). In Saccharomyces cerevisiae, the absence of Scs2 protein will lead to the separation of the endoplasmic reticulum and plasma membrane, resulting in endoplasmic reticulum dysfunction, but its function is not clear in rice blast fungus or even filamentous fungus. In this study, we report the identification and characterization of MoSCS2 in the pathogenesis of the rice blast fungus Magnaporthe oryzae. Protein subcellular localization showed that MoSCS2 is mainly localized in the endoplasmic reticulum. Compared to the wild-type strain Guy11, the deletion mutant ΔMoscs2 showed a significant reduction in growth and conidiation. MoSCS2 deficiency also resulted in abnormal conidial morphology and septum formation. The ΔMoscs2 mutant shows delayed appressorium formation, and the appressorium of ΔMoscs2 mutant could not form huge turgor pressure to penetrate the host epidermal cell wall. Pathogenicity and plant leave infection assays showed that knockout of MoSCS2 significantly inhibited the expansion of the invasive hyphae in host cells, ultimately leading to the decline of pathogenicity. Moreover, MoSCS2 gene is also involved in the regulation of cell wall and endoplasmic reticulum stress response. In conclusion, MoSCS2 plays an important role in the growth, asexual production, conidia morphogenesis, infection-related morphogenesis and pathogenicity of M. oryzae.

De novo purine nucleotide biosynthesis mediated by MoAde4 is required for conidiation, host colonization and pathogenicity in Magnaporthe oryzae.

Amidophosphoribosyltransferase catalyzes the conversion of 5-phosphoribosyl-1-pyrophosphate into 5-phosphoribosyl-1-amine in the de novo purine biosynthetic pathway. Herein, we identified and characterized the functions of MoAde4, an orthologue of yeast Ade4 in Magnaporthe oryzae. MoAde4 is a 537-amino acid protein containing GATase_6 and pribosyltran domains. MoADE4 transcripts were highly expressed during the conidiation, early-infection, and late-infection stages of the fungus. Disruption of the MoADE4 gene resulted in ΔMoade4 exhibiting adenine, adenosine, and hypoxanthine auxotrophy on minimal medium. Conidia quantification assays showed that sporulation was significantly reduced in the ΔMoade4 mutant. The conidia of ΔMoade4 could still form appressoria but mostly failed to penetrate the rice cuticle. Pathogenicity tests showed that ΔMoade4 was completely nonpathogenic on rice and barley leaves, which was attributed to restricted infectious hyphal growth within the primary cells. The ΔMoade4 mutant was defective in the induction of strong host immunity. Exogenous adenine partially rescued conidiation, infectious hyphal growth, and the pathogenicity defects of the ΔMoade4 mutant on barley and rice leaves. Taken together, our results demonstrated that purine nucleotide biosynthesis orchestrated by MoAde4 is required for fungal development and pathogenicity in M. oryzae. These findings therefore act as a suitable target for antifungal development against recalcitrant plant fungal pathogens. KEY POINTS: • MoAde4 is crucial for de novo purine nucleotide biosynthesis. • MoAde4 is pivotal for conidiogenesis and appressorium development of M. oryzae. • MoAde4 is involoved in the pathogenicity of M. oryzae.