First Report of Nigrospora oryzae Causing Leaf Spot Disease on Amorphophallus albus in Shaanxi Province of China.

Amorphophallus albus P. Y. Liu & J. F. Chen is a typical cash crop widely planted in southwest China (Gao et al., 2022). In early August of 2021, a peculiar leaf spot disease was first detected on A. albus in Ankang Academy of Agricultural Sciences manufacturing base (32°69'N, 109°02'E), Shaanxi, China. Small irregular yellow-brown spots (1 to 2 mm) were observed on the surface of A. albus leaf. Following infection of the leaf, it expanded (3 to 5 mm) and became necrotic. Nine planting bases were investigated, and approximately 75% of plants were symptomatic during the rapid expansion period of bulb growth in Hanyin, Langao and Hanbin counties, Ankang City, Shaanxi, China. Higher disease incidence was observed at temperatures above 30℃ and humidity above 80%. Twenty-seven symptomatic tissues of infected leaves were first surface sterilized by immersion in 75% ethanol for 1 minute, followed by rinsing three times in sterile distilled water. The tissues were then cut into 4-5 mm pieces, plated on 1.5% potato dextrose agar (PDA), and incubated at 28±2°C. The hyphal tip from the growing edge of colonies cultured for three days at 28±2℃ was transferred to PDA to obtain pure cultures. Fungal colonies were white, then grey to black with an unevenly distributed, fast-growing aerial mycelium covering the petri dish within five days at 28±2℃. The colony turned dark brown when maintained in the dark at 28±2℃ after seven days, then grayish brown upon sporulation after 15 days (Fig.1f-g). Conidia were brown or black, smooth, spherical to sub-spherical, single-celled (8-12 µm × 10-13µm, average 9-11.5 µm in diameter, n=5µm). The nutritional hyphae exhibited septa, and a portion of the aerial hyphae formed a long, rough conidium, giving rise to a nearly spherical apical sac (Fig.1h). The surface gave rise to several small peduncles bearing clusters of surfaced spherical conidia (Fig.1i). Surfaced spherical conidia were generated on the surface of the small peduncle (Fig.1j). These morphological features were consistent with Nigrospora oryzae (Li et al., 2017). Genomic DNA was extracted from mycelia of the pathogen using an Ezup column fungal genomic DNA extraction kit (Sangon Biotech, Shanghai, China). To confirm the identity of the pathogen, the genomic fragments for the internal transcribed spacer (ITS), LSU (28S) and BenA gene of the isolate were amplified by PCR (Wang et al., 2017) and sent for sequencing. The resultant sequence (GeneBank ID of gene ITS, LSU, BenA are OR723825, OR775345, OR277316, respectively) were compared with the voucher specimens. BLAST results showed >99% identity with those of N.oryzae (GeneBank ID of N.oryzae strain LC2707 ITS, LSU, BenA are KX985954, KY806242, KY019481, respectively). A neighbor joining phylogenetic tree with the concatenated sequences of these genes showed that A-pb169 had the closest match with N. oryzae (Fig. 2). For pathogenicity testing, fifty plants in a period of rapid expansion of bulb growth were selected. Four leaves per plant were inoculated by sprayed till runoff with a conidial suspension of the pathogen (50 µL, 1×106 conidia/ml sterile water), and incubated at 30±2℃ and 80 ± 5% humidity. Control plants received sterile water. On the third day after inoculation, a yellow-brown spot appeared on leave surfaces, the spot gradually expanded; the infection rate was 90 to 95%. Fifteen days after inoculation, infected leaves showed symptoms like those observed in the field, whereas 100 control leaves sprayed with sterile water remained symptomless (Fig.1 a-e). The pathogen was reisolated from infected leaves and confirmed as N. oryzae by morphology and molecular identification. To our knowledge, this is the first report of leaf spot disease of A. albus caused by N. oryzae in China. Since its one of the major cash crops of the southeastern China, further work is necessary to determine its spread and economic impact as well as developing sustainable disease management options.

A new endophytic Penicillium oxalicum with aphicidal activity and its infection mechanism.

BACKGROUND: Aphid infestation adversely affects the yield and quality of crops. Rapid reproduction and insecticidal resistance have made controlling aphids in the field challenging. Therefore, the present study investigated the insecticidal property of Penicillium oxalicum (QLhf-1) and its mechanism of action against aphids, Hyalopterus arundimis Fabricius. RESULTS: Bioassay revealed that the control efficacy of the spores against aphids (86.30% and 89.05% on the third day and fifth day after infection, respectively) were higher than other components, such as the mycelium. Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) showed that QLhf-1 invaded the aphid cuticle through spores and used the aphid tissues as a nutrient source for growth and reproduction, causing stiffness and atrophy and a final death. Three extracellular enzymes, lipase, protease, and chitinase had a synergistic effect with spores, and they acted together to complete the infection process by degrading the aphid body wall and accelerating the infection process. CONCLUSION: The newly discovered endophytic penicillin strain P. oxalicum 'QLhf-1' can effectively kill aphids. The results provided strong evidence for the biological control of aphids, and lay a foundation for the development and utilization of QLhf-1. © 2024 Society of Chemical Industry.

The Isolation, Identification and Immobilization Method of Three Novel Enzymes with Diosgenin-Producing Activity Derived from an Aspergillus flavus.

Diosgenin is an important raw material used in the synthesis of steroid drugs, and it is widely used in the pharmaceutical industry. The traditional method of producing diosgenin is through using raw materials provided via the plant Dioscorea zingiberensis C. H. Wright (DZW), which is subsequently industrially hydrolyzed using a high quantity of hydrochloric and sulfuric acids at temperatures ranging from 70 °C to 175 °C. This process results in a significant amount of unmanageable wastewater, creates issues of severe environmental pollution and consumes high quantities of energy. As an alternative, the enzymolysis of DZW to produce diosgenin is an environmentally and friendly method with wide-ranging prospects for its application. However, there are still only a few enzymes that are suitable for production on an industrial scale. In this study, three new key enzymes, E1, E2, and E3, with a high conversion stability of diosgenin, were isolated and identified using an enzyme-linked-substrate autography strategy. HPLC-MS/MS identification showed that E1, a 134.45 kDa protein with 1019 amino acids (AAs), is a zinc-dependent protein similar to the M16 family. E2, a 97.89 kDa protein with 910 AAs, is a type of endo-ß-1,3-glucanase. E3, a 51.6 kDa protein with 476 AAs, is a type of Xaa-Pro aminopeptidase. In addition, the method to immobilize these proteins was optimized, and stability was achieved. The results show that the optimal immobilization parameters are 3.5% sodium alginate, 3.45% calcium chloride concentration, 1.4 h fixed time, and pH 8.8; and the recovery rate of enzyme activity can reach 43.98%. A level of 70.3% relative enzyme activity can be obtained after employing six cycles of the optimized technology. Compared with free enzymes, immobilized enzymes have improved stability, acid and alkaline resistance and reusability, which are conducive to large-scale industrial production.

Discovery, Identification, and Insecticidal Activity of an Aspergillus flavus Strain Isolated from a Saline-Alkali Soil Sample.

Aphids are one of the most destructive pests in agricultural production. In addition, aphids are able to easily develop resistance to chemical insecticides due to their rapid reproduction and short generation periods. To explore an effective and environmentally friendly aphid control strategy, we isolated and examined a fungus with aphid-parasitizing activity. The strain (YJNfs21.11) was identified as Aspergillus flavus by ITS, 28S, and BenA gene sequence analysis. Scanning electron microscopy and transmission electron microscopy revealed that the infection hyphae of 'YJNfs21.11' colonized and penetrated the aphid epidermal layer and subsequently colonized the body cavity. Field experiments showed that 'YJNfs21.11' and its fermentation products exerted considerable control on aphids, with a corrected efficacy of 96.87%. The lipase, protease, and chitinase secreted by fungi help aphid cuticle degradation, thus assisting spores in completing the infection process. Additionally, changes were observed in the mobility and physical signs of aphids, with death occurring within 60 h of infection. Our results demonstrate that A. flavus 'YJNfs21.11' exhibits considerable control on Aphis gossypii Glover and Hyalopterus arundimis Fabricius, making it a suitable biological control agent.

Identification and a phased pH control strategy of diosgenin bio-synthesized by an endogenous Bacillus licheniformis Syt1 derived from Dioscorea zingiberensis C. H. Wright.

Diosgenin is widely used as one precursor of steroidal drugs in pharmaceutical industry. Currently, there is no choice but to traditionally extract diosgenin from Dioscorea zingiberensis C. H. Wright (DZW) or other plants. In this work, an environmentally friendly approach, in which diosgenin can be bio-synthesized by the endophytic bacterium Bacillus licheniformis Syt1 isolated from DZW, is proposed. Diosgenin produced by the strain was identified by high-performance liquid chromatography (HPLC), nuclear magnetic resonance (NMR), and Fourier transform infrared spectroscopy (FTIR). The thermal gravimetric analysis (TGA) showed that the melting point of the diosgenin product was 204 °C. The optical rotation measurement exhibited that the optical rotation was α20589 = - 126.1° ± 1.5° (chloroform, c = 1%): negative sign means that the product is left-handed, which is very important to further produce steroid hormone drugs. Cholesterol may be the intermediate product in the diosgenin biosynthesis pathway. In the batch fermentation process to produce diosgenin using the strain, pH values played an important role. A phased pH control strategy from 5.5 to 7.5 was proved to be more effective to improve production yield than any single pH control, which could get the highest diosgenin yield of 85 ± 8.6 mg L-1. The proposed method may replace phyto-chemistry extraction to produce diosgenin in the industry in the future.Key points• An endophytic Bacillus licheniformis Syt1 derived from host can produce diosgenin.• A dynamic pH industrial control strategy is better than any single pH control.• Proposed diosgenin-produced method hopefully replaces phyto-chemistry extraction.

Synthesis, crystal structure and antifungal activity of a divalent cobalt(II) complex with uniconazole.

Azole compounds have attracted commercial interest due to their high bactericidal and plant-growth-regulating activities. Uniconazole [or 1-(4-chlorophenyl)-4,4-dimethyl-2-(1H-1,2,4-triazol-1-yl)pent-1-en-3-ol] is a highly active 1,2,4-triazole fungicide and plant-growth regulator with low toxicity. The pharmacological and toxicological properties of many drugs are modified by the formation of their metal complexes. Therefore, there is much interest in exploiting the coordination chemistry of triazole pesticides and their potential application in agriculture. However, reports of complexes of uniconazole are rare. A new cobalt(II) complex of uniconazole, namely dichloridotetrakis[1-(4-chlorophenyl)-4,4-dimethyl-2-(1H-1,2,4-triazol-1-yl-κN(4))pent-1-en-3-ol]cobalt(II), [CoCl2(C15H18ClN3O)4], was synthesized and structurally characterized by element analysis, IR spectrometry and X-ray single-crystal diffraction. The crystal structural analysis shows that the Co(II) atom is located on the inversion centre and is coordinated by four uniconazole and two chloride ligands, forming a distorted octahedral geometry. The hydroxy groups of an uniconazole ligands of adjacent molecules form hydrogen bonds with the axial chloride ligands, resulting in one-dimensional chains parallel to the a axis. The complex was analysed for its antifungal activity by the mycelial growth rate method. It was revealed that the antifungal effect of the title complex is more pronounced than the effect of fungicide uniconazole for Botryosphaeria ribis, Wheat gibberellic and Grape anthracnose.

Flavitalea gansuensis sp. nov., isolated from soil from an arid area, and emended descriptions of the genus Flavitalea and Flavitalea populi.

A gram-reaction-negative, rod-shaped, gliding and bright-yellow-pigmented bacterial strain, designated JCN-23(T), was isolated from a soil sample collected from an arid area in Gansu Province in north-west China, and characterized by using a polyphasic taxonomic approach. This isolate grew optimally at 30 °C and in the absence of NaCl. The only respiratory quinone was menaquinone-7 and the major cellular fatty acids were iso-C(15 : 0), iso-C(17 : 0) 3-OH, summed feature 9 (iso-C(17 : 1)ω9c and/or C(16 : 0) 10-methyl) and summed feature 3 (C(16 : 1)ω7c and/or C(16 : 1)ω6c). The only polyamine was homospermidine and the major polar lipid was phosphatidylethanolamine. The DNA G+C content was 47.1 mol%. Comparative 16S rRNA gene sequence analysis showed that strain JCN-23(T) was a member of the phylum Bacteroidetes, exhibiting the highest 16S rRNA gene sequence similarity to Flavitalea populi CCTCC AB 208255(T) (97.6 %). No other recognized bacterial species showed more than 93.4 % 16S rRNA gene sequence similarity to the novel isolate. DNA-DNA hybridization experiments showed a low level (26 %) of DNA-DNA relatedness between strain JCN-23(T) and F. populi CCTCC AB 208255(T). On the basis of the phenotypic and genotypic data and phylogenetic inference, strain JCN-23(T) is considered to represent a novel species of the genus Flavitalea, for which the name Flavitalea gansuensis sp. nov. is proposed. The type strain is JCN-23(T) ( = ACCC 05418(T) = KCTC 23071(T)). Emended descriptions of the genus Flavitalea and Flavitalea populi are also proposed.

Isolation, identification and antimicrobial activities of two secondary metabolites of Talaromyces verruculosus.

From the ethyl acetate extract of the culture broth of Talaromyces verruculosus, a rhizosphere fungus of Stellera chamaejasme L., (-)-8-hydroxy-3-(4-hydroxypentyl)-3,4-dihydroisocoumarin (1) and (E)-3-(2,5-dioxo-3-(propan-2-ylidene)pyrrolidin-1-yl)acrylic acid (2) were isolated and evaluated for their antimicrobial activities. Their structures were elucidated by UV, IR, MS, 1H-NMR, 13C-NMR and 2D NMR spectra. Compound 1 exhibited the significant activities in vitro against two strains of bacteria and four strains of fungi. Compound 2 gave slight activities on the fungi at 100 µg mL(-1), but no activities on the bacteria. Compound 1 should be considered as a new lead or model compound to develop new isocoumarin antimicrobial agents.

In vitro antifungal activity of sanguinarine and chelerythrine derivatives against phytopathogenic fungi.

In order to understand the antifungal activity of some derivatives of sanguinarine (S) and chelerythrine (C) and their structure-activity relationships, sixteen derivatives of S and C were prepared and evaluated for in vitro antifungal activity against seven phytopathogenic fungi by the mycelial growth rate method. The results showed that S, C and their 6-alkoxy dihydro derivatives S1-S4, C1-C4 and 6-cyanodihydro derivatives S5, C5 showed significant antifungal activity at 100 µg/mL against all the tested fungi. For most tested fungi, the median effective concentrations of S, S1, C and C1 were in a range of 14-50 µg/mL. The structure-activity relationship showed that the C=N+ moiety was the determinant for the antifungal activity of S and C. S1-S5 and C1-C5 could be considered as the precursors of S and C, respectively. Thus, the present results strongly suggested that S and C or their derivatives S1-S5 and C1-C5 should be considered as good lead compounds or model molecules to develop new anti-phytopathogenic fungal agents. can't login to work station for 2hrs--took 2 hrs vacation

Deinococcus soli sp. nov., a gamma- and uv-radiation-resistant bacterium from north-west China.

An ionizing- and UV-radiation-resistant bacterial strain, designated ZLM-202T, was isolated from an arid soil sample collected from Xinjiang Province, north-west China. The soil sample was irradiated before serial dilution plating was performed using twofold-diluted marine agar. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain ZLM-202T was a member of the genus Deinococcus, exhibiting sequence similarities of 86.3-92.2% to the type strains of recognized Deinococcus species. Strain-ZLM-202 was strictly aerobic and showed optimum growth at 30-37 degrees C and pH 7.0. The major respiratory menaquinone was MK-8. The major fatty acids were 16:1 omega7c, 16:0, 15:1 omega6c, 15:0 iso and 16:1 omega5c. L-ornithine was detected in its peptidoglycan. The polar lipid profile consisted mainly of various unknown phosphoglycolipids, aminophospholipids, glycolipids and phospholipids. The DNA G + C content was 65.5 mol. %. The strain was shown to be extremely resistant to gamma radiation (> 10 kGy) and UV light (> 600 J m(-2)). On the basis of the phylogenetic, chemotaxonomic and phenotypic data, strain ZLM-202T represents a novel species of the genus Deinococcus, for which the name Deinococcus soli sp. nov. is proposed. The type strain is ZLM-202T (= CCTCC AB 208223T = KCTC 13419T).

(E)-3-[2,5-Dioxo-3-(propan-2-yl-idene)pyrrolidin-1-yl]acrylic acid.

The title compound, C(10)H(11)NO(4), was extracted from a culture broth of Penicillium verruculosum YL-52. The mol-ecular structure is essentially planar, with an r.m.s. deviation of 0.01342 (2) Šfor the non-H atoms. In the crystal structure, adjacent mol-ecules are connected into a centrosymmetric dimer through a pair of O-H⋯O hydrogen bonds. The dimers are further extended into a chain by weak C-H⋯O hydrogen bonds.