First identification of potato tuber rot caused by Penicillium solitum, its silver nanoparticles synthesis, characterization and use against harmful pathogens.

Potato is one of the highly consumed vegetable crop grown in different regions across Pakistan that is affected by fungal diseases. The current research was conducted to identify fungal pathogen causing mold-like disease of potato in Khyber Pakhtunkhwa (KP), Pakistan. For molecular identification and characterization of the fungal disease; potato tuber samples were collected followed by culturing on potato dextrose agar (PDA). Based on morphological features, the pathogen was identified as a Penicillium species. This result was obtained in 45 different isolates from potato tubers. Molecular identification was done using ß-tubulin primers and ITS5 sequencing of 13 different isolates that releveled 98% homology with BLAST (GenBank accession no. KX958076) as Penicillium solitum (GenBank accession nos. ON307317; ON307475 and ON310801). Phylogenetic tree was constructed that showed Penicillium solitum prevalence along with Penicillium polonicum and Penicillium citrinum on potato tubers. Based on this, Penicillium solitum based silver nanoparticles (Ag NPs) were synthesized and characterized using UV-visible spectroscopy, Fourier transform infrared (FTIR) spectroscopy, X-ray diffraction (XRD), energy dispersive X-ray (EDX) and field emission scanning electron microscopy (FE SEM). UV-analysis showed a characteristic peak at 410 nm confirming synthesis of Penicillium solitum based Ag NPs. This was further confirmed by XRD followed by EDX and SEM that showed face cubic crystal structure with Ag as major constituent of 18 nm formed spherical Ag NPs. FTIR showed band stretching of O-H, N-O and C-H of biological origin. Similarly, Penicillium solitum based Ag NPs presented strong anti-bacterial and anti-fungal activity at 0.5 level of significance LSD. According to our knowledge, this is the first report of Penicillium solitum identification in Pakistan, its Ag NPs synthesis and characterization to be used against pathogens of agricultural significance.

Complete Genome Sequence of Pythium oligandrum, Isolated from Rhizosphere Soils of Chinese Angelica sinensis.

Most of the Pythium species are pathogenic to a wide range of economically important crops and, sometimes, can even cause diseases in animals and humans. An exception is that the soil-inhabiting P. oligandrum is an effective biocontrol agent against a diverse suite of pathogens and promotes plant growth. In this work, we sequenced the whole genome of P. oligandrum PO-1, isolated from rhizosphere soils of Chinese Angelica sinensis, using a combination of long-read single-molecule real-time sequencing technology (Pacific Biosciences [PacBio]) and Illumina sequencing. The 2.5-Gb and 5.2-Gb bases were generated respectively. The sequencing depths were 93× with PacBio and 145× with Illumina sequencing. With the PacBio sequencing results further corrected by Illumina sequencing, the genome was assembled into 71 scaffolds with a total size of 39.10 Mb (N50 = 1.45 Mb; L50 = 9)and the longest scaffold is 3.49 Mb. Genome annotation identifies 15,632 protein-coding genes and 0.47 Mb of transposable elements. Our genomic assembly and annotation have been greatly improved compared with the already released three genomes of P. oligandrum. This genomic data will provide valuable information to understand the mechanism underlying its biocontrol potentials and will also facilitate the dissection of genome evolution and environmental adaptation within the genus Pythium. [Formula: see text] The author(s) have dedicated the work to the public domain under the Creative Commons CC0 "No Rights Reserved" license by waiving all of his or her rights to the work worldwide under copyright law, including all related and neighboring rights, to the extent allowed by law, 2022.

Genomic landscape of a relict fir-associated fungus reveals rapid convergent adaptation towards endophytism.

Comparative and pan-genomic analyses of the endophytic fungus Pezicula neosporulosa (Helotiales, Ascomycota) from needles of the relict fir, Abies beshanzuensis, showed expansions of carbohydrate metabolism and secondary metabolite biosynthetic genes characteristic for unrelated plant-beneficial helotialean, such as dark septate endophytes and ericoid mycorrhizal fungi. The current species within the relatively young Pliocene genus Pezicula are predominantly saprotrophic, while P. neosporulosa lacks such features. To understand the genomic background of this putatively convergent evolution, we performed population analyses of 77 P. neosporulosa isolates. This revealed a mosaic structure of a dozen non-recombining and highly genetically polymorphic subpopulations with a unique mating system structure. We found that one idiomorph of a probably duplicated mat1-2 gene was found in putatively heterothallic isolates, while the other co-occurred with mat1-1 locus suggesting homothallic reproduction for these strains. Moreover, 24 and 81 genes implicated in plant cell-wall degradation and secondary metabolite biosynthesis, respectively, showed signatures of the balancing selection. These findings highlight the evolutionary pattern of the two gene families for allowing the fungus a rapid adaptation towards endophytism and facilitating diverse symbiotic interactions.

Facultative symbiosis with a saprotrophic soil fungus promotes potassium uptake in American sweetgum trees.

Several species of soil free-living saprotrophs can sometimes establish biotrophic symbiosis with plants, but the basic biology of this association remains largely unknown. Here, we investigate the symbiotic interaction between a common soil saprotroph, Clitopilus hobsonii (Agaricomycetes), and the American sweetgum (Liquidambar styraciflua). The colonized root cortical cells were found to contain numerous microsclerotia-like structures. Fungal colonization led to increased plant growth and facilitated potassium uptake, particularly under potassium limitation (0.05 mM K+ ). The expression of plant genes related to potassium uptake was not altered by the symbiosis, but colonized roots contained the transcripts of three fungal genes with homology to K+ transporters (ACU and HAK) and channel (SKC). Heterologously expressed ChACU and ChSKC restored the growth of a yeast K+ -uptake-defective mutant. Upregulation of ChACU transcript under low K+ conditions (0 and 0.05 mM K+ ) compared to control (5 mM K+ ) was demonstrated in planta and in vitro. Colonized plants displayed a larger accumulation of soluble sugars under 0.05 mM K+ than non-colonized plants. The present study suggests reciprocal benefits of this novel tree-fungus symbiosis under potassium limitation mainly through an exchange of additional carbon and potassium between both partners.

Hybrid Genome Assembly and Gene Repertoire of the Root Endophyte Clitopilus hobsonii QYL-10 (Entolomataceae, Agaricales, Basidiomycetes).

Clitopilus hobsonii (Entolomataceae, Agaricales, Basidiomycetes) is a common soil saprotroph. There is also evidence that C. hobsonii can act as a root endophyte benefitting tree growth. Here, we report the genome assembly of C. hobsonii QYL-10, isolated from ectomycorrhizal root tips of Quercus lyrata. The genome size is 36.93 Mb, consisting of 13 contigs (N50 = 3.3 Mb) with 49.2% GC content. Of them, 10 contigs approached the length of intact chromosomes, and three had telomeres at one end only. BUSCO analysis reported a completeness score of 98.4%, using Basidiomycota_odb10 lineage data. Combining ab-initio, RNA-seq data, and homology-based predictions, we identified 12,710 protein-coding genes. Approximately, 1.43 Mb of transposable elements (3.88% of the assembly), 36 secondary metabolite biosynthetic gene clusters, and 361 genes encoding putative carbohydrate-active enzymes were identified. This genomic resource will allow functional studies aimed to characterize the symbiotic interactions between C. hobsonii and its host trees and will also provide a valuable foundation for further research on comparative genomics of the Entolomataceae.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

The Chromosome-Scale Genome Resource for Two Endophytic Fusarium species, F. culmorum and F. pseudograminearum.

Genus Fusarium (Ascomycota, Hypocreales, Nectriaceae) includes many economically important plant pathogens that cause devastating diseases of a wide range of crops and trees. Interestingly, there is increasing evidence that some Fusarium species also live as endophytes and benefit plant growth and stress tolerance. In this work, we sequence the whole genomes of endophytic F. culmorum and F. pseudograminearum, isolated from a coastal dunegrass (Leymus mollis), using long-read single-molecule real-time sequencing technology. Their genomes are assembled into four chromosomes and a mitochondrial genome with a total assembly size of 40.05 and 42.90 M, respectively. This resource should not only facilitate functional studies designed to better understand what makes the two Fusarium species such successful plant-beneficial fungi but should also reveal their genome evolution and adaptation.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Epigenetic silencing of SFRP1 and SFRP5 by hepatitis B virus X protein enhances hepatoma cell tumorigenicity through Wnt signaling pathway.

Secreted frizzled-related proteins (SFRPs) are antagonists of the Wnt signaling pathway whose epigenetic downregulation have been shown to be involved in hepatocarcinogenesis. However, dysregulation of SFRPs induced by hepatitis B virus (HBV) X protein (HBx) has never been studied in HBV-related hepatocellular carcinoma (HBV-HCC). In this study, we sought to determine the clinical significance and underlying mechanism of HBx-induced SFRPs dysregulation in hepatoma cells and HBV-HCC patients. Our results showed that SFRP1 and SFRP5 expression were dramatically decreased by HBx in hepatoma cells. The repressed expression in hepatoma cells was partially rescued by a DNA methylation inhibitor and synergistically increased by a combination treatment with a histone deacetyltransferases inhibitor. In addition, we identified that SFRP1 and SFRP5 promoters were hypermethylated in both HBx-expressing hepatoma cells and HBV-HCC tissues. Downregulation of SFRP1 and SFRP5 in HBV-HCC tissues was significantly correlated with overexpression of DNA methyltransferase 1 (DNMT1) and poor tumor differentiation. HBx facilitated the binding of DNMT1 and DNMT3A to SFRP1 and SFRP5 promoters, and resulted in epigenetic silencing of SFRP1 and SFRP5. Moreover, overexpression of SFRP1, SFRP5 or RNA interference mediated silencing of DNMT1 inactivated the Wnt signaling pathway and decreased the expression levels of Wnt target genes c-Myc and CyclinD1, thus impeding HCC growth in vitro and in vivo, and regressing HBx-induced epithelial-mesenchymal transition (EMT). Our findings strongly suggest that epigenetic silencing of SFRP1 and SFRP5 by HBx allows constitutive activation of Wnt signaling pathway and hence contributes to hepatocarcinogenesis.

[Hepatic lineage differentiation of hepatic progenitor cells by bone morphogenetic protein or leukemia inhibitory factor].

OBJECTIVE: To search for the optimal approach for hepatocyte-directed differentiation of hepatic progenitor cells and investigate the molecular mechanism of the hepatic differentiation. METHODS: Hepatic progenitor cells were infected with recombinant adenovirus which containing human LIF, BMP2 or BMP9 gene. The maturation and differentiation of progenitor cells were examined by PAS staining and ICG uptake methods at 4, 7 and 10 days post infection. The production of Albumin (Alb) was measured by luciferase activity at day 4, 7, 10 and 14. RESULTS: PAS staining assay revealed that BMP2 and BMP9 enhanced glycogen storage in hepatic progenitor cells most obviously at day 7. The percentages of positive cells were 30% and 45% respectively at 7 days post-infection. Meanwhile, 40% and 30% cells were positive by ICG uptake assay after BMP2 and BMP9 induction. Luciferase activity indicated that BMP9 induced ALB-Luc activity most significantly at day 7. However, less inductive activity was found in LIF-treated group. CONCLUSION: These results indicated tuat hepatic progenitor cells were differentiated into hepatocyte-like cells by BMPs and LIF induction.

Enhancement of canonical Wnt/ß-catenin signaling activity by HCV core protein promotes cell growth of hepatocellular carcinoma cells.

BACKGROUND: The Hepatitis C virus (HCV) core protein has been implicated as a potential oncogene or a cofactor in HCV-related hepatocellular carcinoma (HCC), but the underlying mechanisms are unknown. Overactivation of the Wnt/ß-catenin signaling is a major factor in oncogenesis of HCC. However, the pathogenesis of HCV core-associated Wnt/ß-catenin activation remains to be further characterized. Therefore, we attempted to determine whether HCV core protein plays an important role in regulating Wnt/ß-catenin signaling in HCC cells. METHODOLOGY: Wnt/ß-catenin signaling activity was investigated in core-expressing hepatoma cells. Protein and gene expression were examined by Western blot, immunofluorescence staining, RT-qPCR, and reporter assay. PRINCIPAL FINDINGS: HCV core protein significantly enhances Tcf-dependent transcriptional activity induced by Wnt3A in HCC cell lines. Additionally, core protein increases and stabilizes ß-catenin levels in hepatoma cell line Huh7 through inactivation of GSK-3ß, which contributes to the up-regulation of downstream target genes, such as c-Myc, cyclin D1, WISP2 and CTGF. Also, core protein increases cell proliferation rate and promotes Wnt3A-induced tumor growth in the xenograft tumor model of human HCC. CONCLUSIONS/SIGNIFICANCE: HCV core protein enhances Wnt/ß-catenin signaling activity, hence playing an important role in HCV-associated carcinogenesis.

Hepatitis C virus core protein activates Wnt/ß-catenin signaling through multiple regulation of upstream molecules in the SMMC-7721 cell line.

The core protein of hepatitis C virus (HCV) has been implicated in HCV-induced liver pathogenesis. Previous data have shown that the HCV core protein has pleiotropic functions, including transcriptional regulation of a number of cellular genes, although the mechanism of gene regulation remains unclear. Wnt/ß-catenin signaling is also involved in hepatocellular carcinoma (HCC) tumorigenesis. To elucidate the molecular mechanism of HCV pathogenesis, we examined whether HCV core protein activates Wnt/ß-catenin signaling in the hepatoma cell line SMMC-7721. The effects of core protein on Wnt/ß-catenin signaling cascades were investigated by luciferase reporter gene assay, immunofluorescence, western blot and RT-PCR analysis. Here, we demonstrate that HCV core protein plays an essential role in activating ß-catenin/Tcf-4-dependent transcriptional activity and increases active ß-catenin expression and nuclear accumulation in SMMC-7721 cells. An RT-PCR assay indicated that core protein upregulates gene expression of canonical Wnt ligands, such as Wnt2, Wnt3, Wnt3a, Wnt8b, Wnt10a, Wnt10b, frizzled receptors Fzd1, 2, 5, 6, 7, 9, and LRP5/6 co-receptors. However, Wnt antagonists SFRP3, 5 and Dkk1 were moderately repressed. Furthermore, ectopic expression of core protein markedly promoted cell proliferation. The soluble Fzd molecule FrzB or the ß-catenin inhibitor siBC efficiently blocked cell growth stimulation by the core gene. Our present findings demonstrate that the HCV core protein activates canonical Wnt signaling through tight regulation of several important molecules upstream of ß-catenin and presumably results in promotion of cell proliferation in the SMMC-7721 cell line. Taken together, these data suggested that the core protein may be directly involved in Wnt/ß-catenin-mediated liver pathogenesis.

Ectopic expression in the giant fiber system of Drosophila reveals distinct roles for roundabout (Robo), Robo2, and Robo3 in dendritic guidance and synaptic connectivity.

The Roundabout (Robo) receptors have been intensively studied for their role in regulating axon guidance in the embryonic nervous system, whereas a role in dendritic guidance has not been explored. In the adult giant fiber system of Drosophila, we have revealed that ectopic Robo expression can regulate the growth and guidance of specific motor neuron dendrites, whereas Robo2 and Robo3 have no effect. We also show that the effect of Robo on dendritic guidance can be suppressed by Commissureless coexpression. Although we confirmed a role for all three Robo receptors in giant fiber axon guidance, the strong axon guidance alterations caused by overexpression of Robo2 or Robo3 have no effect on synaptic connectivity. In contrast, Robo overexpression in the giant fiber seems to directly interfere with synaptic function. We conclude that axon guidance, dendritic guidance, and synaptogenesis are separable processes and that the different Robo family members affect them distinctly.