Complete genome sequence of a novel mitovirus isolated from the fungus Fusarium oxysporum f. sp. ginseng causing ginseng root rot.

A novel mitovirus, tentatively designated as "Fusarium oxysporum mitovirus 2" (FoMV2), was isolated from the pathogenic Fusarium oxysporum f. sp. ginseng strain 0414 infecting Panax ginseng. The complete genome of FoMV2 is 2388 nt in length with a GC content of 30.57%. It contains a large open reading frame (ORF) encoding a putative RNA-dependent RNA polymerase (RdRp) of 713 amino acids with a molecular weight of 83.05 kDa. The sequence identity between FoMV2 and Botrytis cinerea mitovirus 8 and Fusarium verticillioides mitovirus 1 was 87.94% and 77.85%, respectively. Phylogenetic analysis showed that FoMV2 belongs to the genus Unuamitovirus in the family Mitoviridae. To the best of our knowledge, this is the first report of an unuamitovirus isolated from F. oxysporum f. sp. ginseng causing ginseng root rot.

Development of a Novel Real-Time Quantitative PCR Method for Detection of Ilyonectria robusta, the Predominant Species Causing Ginseng Rusty Root Rot.

Rusty root rot is the most destructive soilborne disease of ginseng caused by pathogenic Ilyonectria spp., predominantly Ilyonectria robusta, in China. However, there remains no effective strategy to control the disease. Current control of the disease requires that soil and ginseng seeds and seedlings infected with I. robusta are avoided during planting. Therefore, rapid and accurate detection of I. robusta would be indispensable in disease control programs. A one-step polymerase chain reaction (PCR) and quantitative real-time PCR (qPCR) assay was developed to detect I. robusta in ginseng seeds, roots, and soil. The species-specific primers HIS H3-F and HIS H3-R, designed based on a partial histone gene sequence of I. robusta, yielded a 268-bp product using the optimized PCR and qPCR protocol. DNA of I. robusta was detected by qPCR in all diseased soil and ginseng roots and seeds resulting from artificial inoculation and sampled from natural fields. I. robusta was detected at an abundance of 1.42 fg/µl at 12 h postinoculation and 191.31 fg/µl at 7 days postinoculation in ginseng roots that showed disease symptoms. In naturally infected soil sampled from ginseng fields, pathogen abundances ranging from 13.23 to 503.39 fg/µl were detected, which were 2.04 to 11.01 times higher than those in ginseng roots. The pathogen was first detected and was more abundant on the surface of the ginseng seed coat compared with that in the seed kernel. This study provides a high-efficiency detection technique for early diagnosis of I. robusta and real-time disease prevention and control strategies.

Antagonistic potential of Trichoderma as a biocontrol agent against Sclerotinia asari.

In the present study, the inhibitory potential of 14 Trichoderma strains (isolated from Asarum rhizosphere) was investigated against Sclerotinia asari using the plate dilution method. The activity of antioxidant enzymes viz; catalase (CAT), peroxidase (POD), superoxide dismutase (SOD), and malondialdehyde (MDA) in S. asari treated with the two Trichoderma strains was also evaluated. Untargeted metabolomic analysis by using LC/MS analysis was carried out to determine differential metabolites in T. hamatum (A26) and T. koningiopsis (B30) groups. Moreover, transcriptome analysis of S. asari during the inhibition of S. asari by B30, and A26 compared with the control (CK) was performed. Results indicated that inhibition rates of T. koningiopsis B30, and T. hamatum A26 were highest compared to other strains. Similarly, non-volatile metabolites extracted from the B30 strain showed a 100% inhibition of S. asari. The activity of CAT, SOD, and POD decreased after treatment with A26 and B30 strains while increasing MDA content of S. asari. Antifungal activity of differential metabolites like abamectin, eplerenone, behenic acid, lauric acid, josamycin, erythromycin, and minocycline exhibited the highest inhibition of S. asari. Transcriptome analysis showed that differentially expressed genes were involved in many metabolic pathways which subsequently contributed toward antifungal activity of Trichoderma. These findings suggested that both Trichoderma strains (B30 and A26) could be effectively used as biocontrol agents against Sclerotinia disease of Asarum.

Deciphering the transcriptomic response of Ilyonectria robusta in relation to ginsenoside Rg1 treatment and the development of ginseng rusty root rot.

Rusty root rot is a severe disease in ginseng (Panax ginseng C. A. Mey) production caused by Ilyonectria robusta. The severity of the disease may be related to the residual ginsenosides in soil. In order to elucidate the response mechanism between Rg1 treatment and the occurrence of ginseng rust, we performed growth, reproduction and transcriptome analysis on treated Rg1. The results showed that Rg1 significantly promoted the mycelial growth and sporulation compared with the control, and aggravated the disease symptoms of Panax ginseng. A total of 6708 transcripts out of 213 131 annotated genes identified from global transcriptomic analysis were differentially expressed in Ilyonectria robusta grown during the Rg1 treatment. These genes were found to be related to the carbon-nitrogen metabolism, transport and assimilation. Many of these genes were also associated with pathogenicity based on the Phi-base database. Several transcription factors were related to specific biological processes, such as nitrogen utilization. The current results revealed that Rg1 played a major role in the development of rusty root rot by promoting fungal cell growth and affected the expression of genes required for pathogenesis. Rg1 could aggravate the invasion of Ilyonectria robusta on ginseng root, which preliminarily revealed the reason for the aggravation of rusty root rot in ginseng soil-borne.

The periplasmic chaperone protein Psg_2795 contributes to the virulence of Pseudomonas savastanoi pv. glycinea: the causal agent of bacterial blight of soybean.

Pseudomonas savastanoi pv. glycinea (Psg also named P. syringae pv. glycinea and P. amygdali pv. glycinea) is the causative agent of bacterial blight in soybean. The identification of virulence factors is essential for understanding the pathogenesis of Psg. In this study, a mini-Tn5 transposon mutant library of Psg strain PsgNC12 was screened on soybean, and one low-virulent mini-Tn5 mutant, designated as 4573, was identified. Sequence analysis of the 4573-mutant revealed that the mini-Tn5 transposon was inserted in the Psg_2795 gene. Psg_2795 encodes a FimC-domain protein that is highly conserved in Pseudomonas. Further analysis revealed that the mutation and knockout of Psg_2795 results in a reduced virulence phenotype on soybean, decreased motility, weakened bacterial attachment to a glass surface and delayed the population growth within soybean leaves. The phenotype of the 4573-mutant could be complemented nearly to wild-type levels using an intact Psg_2795 gene. Collectively, our results demonstrate that Psg_2795 plays an important role in the virulence, motility, attachment and the population growth of PsgNC12 in soybean. This finding provides a new insight into the function of periplasmic chaperone proteins in a type I pilus and provides reference information for identifying Psg_2795 homologues in P. savastanoi and other bacteria.

A Novel Biocontrol Strain Bacillus amyloliquefaciens FS6 for Excellent Control of Gray Mold and Seedling Diseases of Ginseng.

The biocontrol efficacy of Bacillus amyloliquefaciens FS6 against seedling diseases and gray mold of ginseng (Panax ginseng), as well as application techniques, were evaluated in a series of field trials. FS6 fermentation broth showed a strong antagonistic effect against the ginseng fungal pathogens, and the inhibition rates on mycelial growth and spore germination were 84 to 88% and 71 to 72%, respectively. Field evaluation showed that combination of seed and soil treatments exhibited better protection than that of individual treatment alone. FS6 wettable powder soil treatment in combination with thiamethoxam plus metalaxyl-M plus fludioxonil for seed coating performed the best, with >83% overall control efficacy for seedling diseases. FS6 had a long-acting effect of >78% control efficacy on ginseng gray mold at 30 days after the last application, almost 2.5- and 2-fold better than that of B. amyloliquefaciens B7900 wettable powder and cyprodinil, respectively. In addition, FS6 reduced the diversity and relative abundance of fungi and affected the fungi and bacterial composition in the rhizosphere soil of ginseng. Therefore, FS6 can be used to effectively control seedling diseases and gray mold in ginseng.

The effect of Glomus intraradices on the physiological properties of Panax ginseng and on rhizospheric microbial diversity.

BACKGROUND: Glomus intraradices is a species of arbuscular mycorrhizal fungi that, as an obligate endomycorrhiza, can form mutually beneficial associations with plants. Panax ginseng is a popular traditional Chinese medicine; however, problems associated with ginseng planting, such as pesticide residues, reduce the ginseng quality. METHODS: In this experiment, we studied the effect of inoculating G. intraradices on several physiological properties and microbial communities of ginseng. UV-Visible Spectrum method was used to detect physical properties. Denaturing gradient gel electrophoresis method was used to analyze microbial communities. RESULTS: The results indicated that inoculation with G. intraradices can improve the colonization rate of lateral ginseng roots, increase the levels of monomeric and total ginsenosides, and improve root activity as well as polyphenol oxidase and catalase activities. We also studied the bacterial and fungal communities in ginseng rhizospheric soil. In our study, G. intraradices inoculation improved the abundance and Shannon diversity of bacteria, whereas fungi showed a reciprocal effect. Furthermore, we found that G. intraradices inoculation might increase some beneficial bacterial species and decreased pathogenic fungi in rhizospheric soil of ginseng. CONCLUSION: Our results showed that G. intraradices can benefit ginseng planting which may have some instructive and practical significance for planting ginseng in farmland.

[Study on etiology of retinoic acid-induced cleft palate in mouse].

OBJECTIVE: To investigate the morphologic changes of embryonic palatal development exposed to retinoic acid (RA) in mouse, and to detect the significance of the expression of TGFbeta1, TGFbeta3, EGF and BCL2. METHODS: The stage of palatal development was examined by light microscopy. S-P immunohistochemistry and in-situ hybridization was used to detect spatio-temporal patterns of expression of TGFbeta1, TGFbeta3, EGF and BCL2 in embryonic palate. RESULTS: The fetus exposed to RA resulted in formation of small palatal shelves without contact and fusion of each other to form and intact palate. RA can regulate the embryonic palatal expression of genes involved in RA-induced cleft palate. CONCLUSIONS: RA can inhibit the proliferation of MEPM cell to form small palatal shelves and induce abnormal differentiation of MEE cell causing the bi-palatal shelves no contact and fuse with each other, then induce the formation of cleft palate. RA can regulate the spatio-temporal patterns of expression of TGFbeta1, TGFbeta3 and EGF in embryonic palatal processes and the change of special expression of these genes in embryonic palatal processes are involved in RA-induced cleft palate.