Lipopeptides from Bacillus velezensis induced apoptosis-like cell death in the pathogenic fungus Fusarium concentricum.

AIMS: Stem rot caused by Fusarium concentricum is a new disease of Paris polyphylla reported by our research group. The present study investigates the growth inhibitory and apoptotic effects of Bacillus velezensis FJAT-54560 lipopeptide against F. concentricum. METHODS AND RESULTS: HPLC preparation and LC-MS analysis results show that the crude lipopeptides secreted by Bacillus velezensis FJAT-54560 isolated from Jasminum sambac consist of C14-17 iturin A, C14 fengycin B, C16 fengycin A/A2, C18 fengycin A, C20 fengycin B2, C21 fengycin A2, C22-23 fengycin A, C12-16 surfactin A, and C15 surfactin A derivatives. The mass ratios (g/g) of iturin, fengycin, and surfactin in lipopeptides are 2.40, 67.51, and 30.08%, respectively. Through inhibition zone and inhibition rate experiments, we found that crude lipopeptides and purified fengycin exhibit strong antifungal activity against F. concentricum, including accumulation of reactive oxygen species, loss of mitochondrial membrane potential, DNA fragmentation, Ca2+ accumulation, chromatin condensation, and phosphatidylserine externalization. Transcriptomic analysis indicates that crude lipopeptide-induced apoptosis in F. concentricum cells may be mediated by apoptosis-inducing factors and apoptosis mediators and can serve as a metacaspase-independent model. CONCLUSION: Lipopeptides from Bacillus velezensis FJAT-54560 can control the pathogenic fungus F. concentricum by inducing apoptosis.

Efficacy of Platelet-Rich Plasma in the Treatment of Diabetic Foot Ulcers: A Systematic Review and Meta-Analysis.

BACKGROUND: The probiological healing effect of platelet-rich plasma (PRP) during tissue repair has recently gathered much attention. This study aimed to conduct a systematic review and meta-analysis of patients with diabetic foot ulcer (DFU) receiving PRP or conventional treatment to evaluate their efficacy. METHODS: PubMed, Excerpta Medica Database (EMBASE), Cochrane Library, and China National Knowledge Infrastructure (CNKI) databases were comprehensively searched by 2 independent reviewers following PRISMA guidelines for the inclusion of randomized controlled trials (RCTs) comparing PRP with conventional treatments for DFUs. The primary measurements of healing rate and healing time, the methodological quality and extracted data were assessed using Review Manager 5.3. Statistical significance was set at P < 0.05. RESULTS: A total of 10 RCTs involving 550 patients were included in this study, PRP was observed to significantly improve the healing rate (risk ratio [RR] = 1.38, 95% confidence interval [CI] 1.05-1.82, P = 0.02) and shorten the healing time (mean difference [MD] = -23.23, 95% CI -45.97 to -0.49, P = 0.05) of patients with DFU when compared to the conventional treatment. CONCLUSIONS: Compared to conventional treatment, PRP effectively promoted the healing of patients with DFU by evidently improving the healing rate and healing time.

Additive Manufacturing of Micro-Architected Copper based on an Ion-Exchangeable Hydrogel.

Additive manufacturing (AM) of copper through laser-based processes poses challenges, primarily attributed to the high thermal conductivity and low laser absorptivity of copper powder or wire as the feedstock. Although the use of copper salts in vat photopolymerization-based AM techniques has garnered recent attention, achieving micro-architected copper with high conductivity and density has remained elusive. In this study, we present a facile and efficient process to create complex 3D micro-architected copper structures with superior electrical conductivity and hardness. The process entails the formulation of an ion-exchangeable photoresin, followed by the utilization of digital light processing (DLP) printing to sculpt 3D hydrogel scaffolds, which were transformed into Cu2+-chelated polymer frameworks (Cu-CPFs) with a high loading of Cu2+ ions through ion exchange, followed by debinding and sintering, results in the transformation of Cu-CPFs into miniaturized copper architectures. This methodology represents an efficient pathway for the creation of intricate micro-architected 3D metal structures.

A General Strategy for Sustainable 3D Printing Based on A Multifunctional Photoinitiator.

A multifunctional photoinitiator is presented, offering precise control over light-induced polymerization initiation at 450 nm and material degradation at 365 nm. This is accomplished by covalently linking photoactive bis(acyl)phosphane oxide and photocleavable o-nitrobenzyl ether moieties onto the surface of γ-cyclodextrin. Upon degradation, the resulting linear polymers can be easily re-dissolved in their corresponding monomer and re-cured, exhibiting superior mechanical properties compared to the pristine material. Moreover, this photoinitiator enables the successful 3D printing of intricate and precise structures, representing a promising general strategy for developing recyclable photoresins for 3D printing applications.

First Report of Apiospora arundinis Causing Wet Root Rot of Pseudostellaria heterophylla in China.

Pseudostellaria heterophylla is one of the traditional medicines in China. From 2020 to 2022, postharvest wet root rot disease was observed with an incidence of 2~5% on the tuberous roots of the harvested P. heterophylla in Zherong county, Fujian province, China, which usually occurs under damp and unventilated conditions. The symptoms of the disease were as follows: white mycelia grew on the surface of tuberous root initially and gradually wrapped around the roots, the internal root tissue turned yellow and became wet decay finally. To identify the causal agent, a total of 20 samples with symptomatic tuberous roots were collected. Small pieces (3 mm2) were treated by surface disinfection with 75% ethanol and 1% NaOCl, then rinsed 3 times with sterile water. These treated pieces were transferred onto potato dextrose agar (PDA) and incubated at 25°C in the dark for 7 d. Ten pure cultures were obtained using single-spore isolation method. The fungus colonies initially produced white aerial mycelium, subsequently exhibited yellow pigmentation. Mycelia were consisted of smooth, hyaline, branched, and septate hyphae. The conidia were solitary or clustered, brown or dark brown, smooth, ellipsoidal to spherical, 6.66 (5.50-7.81)×5.65 (4.17-7.22) µm (n=50) in size. The conidiophores were hyaline or pale brown and produced conidiogenous cells, which were pale brown, smooth, ampulliform, and 10.14 (8.82-15.30) um long (n=50). Based on these morphological characteristics, the fungus was identified as the genus Apiospora (Arthrinium). The rDNA-ITS region and partial ß-tubulin gene (BenA) were amplified using the primers ITS1/ITS4 (White et al. 1990) and Bt2a/Bt2b (Glass and Donaldson 1995), respectively. The sequences of isolates FJAT-32563 and FJAT-32564 were deposited in GenBank (ITS, OM920984 and OM920985; BenA, OM953823 and OM953824). All sequences had more than 99% similarity with those of A. arundinis strain CBS:106.12 (ITS, KF144883; BenA, KF144973). In the multilocus phylogenetic analysis (ITS + BenA), the two isolates clustered together with other strains of A. arundinis with 100% bootstrap support. The isolates were therefore identified as A. arundinis based on both morphological and molecular characteristics. To confirm the pathogenicity, fresh tuberous roots were selected and surface disinfected, then the roots were immersed with a quarter length in the conidial suspension (106/mL) for 30 min, whereas the control roots were immersed with sterile water (n=30). They were placed in petri dish with wet filter paper at 25±2℃, maintaining 80% relative humidity in the dark. The white aerial mycelium appeared at 5 days after inoculation, and wet root rot decaying occurred after inoculation for 21 days. The symptoms were similar to those described above, whereas the control roots were asymptomatic. The same fungus was re-isolated from the infected roots, showing similar morphological characteristics and molecular traits. Koch's postulates were completed and the pathogenicity test for the isolates has been repeated thrice. Previously, A. arundinis was reported to infect peach and sugarcane (Ji et al. 2020; Liao et al. 2022). To our knowledge, this is the first report of A. arundinis causing wet root rot of P. heterophylla in China. The disease would be a potentially new threat to this medicinal plant.

Antifungal activities of Bacillus velezensis FJAT-52631 and its lipopeptides against anthracnose pathogen Colletotrichum acutatum.

This study was aim at investigating antifungal activities of Bacillus velezensis FJAT-52631 and its lipopeptides against Colletotrichum acutatum ex situ and in situ. The results showed that the strain FJAT-52631 and its crude lipopeptides (10 mg/ml) exhibited strong inhibitory effects on growth of C. acutatum FJAT-30256 with an inhibition rate of 75.3% and an inhibition zone diameter of 17.66 mm, respectively. Both the viable bacterial cultures and lipopeptides of FJAT-52631 could delay the onset of loquat anthracnose by 1 day and lower the incidence of loquat anthracnose in situ. The whole cultures of B. velezensis FJAT-52631 displayed a 50% biocontrol efficacy on loquat anthracnose at the fourth day after inoculation, but the crude lipopeptides not. The average lesion diameter of the whole-culture treated group was 5.62 mm, which was smaller than that of control group (6.81 mm). All the three types of lipopeptides including iturin A, fengycin, and surfactin A secreted from the strain FJAT-52631 exhibited antifungal activities. Among them, surfactin A displayed higher antifungal activity at a concentration of 1.25 mg/mL than other two lipopeptides even if at a concentration of 60 mg/mL. Thus, the results indicated that surfactin A produced by FJAT-52631 played a major role in the biocontrol of the loquat anthracnose. Scanning electron microscopy (SEM) observation revealed the structural deformities in the mycelia of C. acutatum. The above results suggested that the antifungal lipopeptides from B. velezensis FJAT-52631 would be potential in biocontrol against anthracnose disease of loquat caused by C. acutatum.

Effect of Bacillus subtilis and Lactobacillus plantarum on solid-state fermentation of soybean meal.

BACKGROUND: As feeding probiotics, Bacillus subtilis FJAT-4842 and Lactobacillus plantarum FJAT-13737 were employed to improve the nutritional value and reduce the risk of contamination in the solid-state fermentation of soybean meal (SBM). RESULTS: After fermentation with bacterium starters, the crude protein, free amino acid and lactic acid increased, with higher protease and cellulose activity. The addition of L. plantarum could increase the crude protein and lactic acid content by 50.1% and 94.9%, respectively. After fermentation, the contents of crude fiber and phytic acid decreased by 45.9% and 48.1%. Compared with control treatment, the addition of both B. subtilis FJAT-4842 and L. plantarum FJAT-13737 could effectively elevate the production of free amino acid and esters. Moreover, the addition of bacterium starter could prevent the production of mycotoxin and promote bacterial diversity of the fermented SBM. Especially, the addition of B. subtilis could reduce the relative content of Staphylococcus. After fermentation for 7 days, lactic acid bacteria including Pediococcus, Weissella and Lactobacillus became the main bacteria in the fermented SBM. CONCLUSION: The addition of bacterium starter is of benefit with respect to improving the nutritional value and reducing the risk of contamination in the solid-state fermentation of soybean. © 2023 Society of Chemical Industry.

Functional Polythioamides Derived from Thiocarbonyl Fluoride.

Polythioamide is a unique type of sulfur-containing polymer with advanced functionalities. Nonetheless, the elemental sulfur commonly used in their synthesis tends to react readily with unsaturated functional groups, thereby limiting the scope of eligible substrates. Inspired by the highly efficient sulfur-fluoride exchange (SuFEx) polymerization through discrete hubs, we present herein a pioneering and versatile approach to the synthesis of polythioamides from diboronic acids, secondary diamines, and thiocarbonyl fluoride as the central connective hub. Well-defined structures, including previously inaccessible unsaturated substrates, were realized. These newly devised polythioamides can efficiently and selectively bind to metal ions and were applied in precious-metal recovery. Further development resulted in PdII -crosslinked single-chain nanoparticles serving as recyclable homogeneous catalysts, thus demonstrating the vast potential of these unprecedented polythioamides. We anticipate that thiocarbonyl fluoride could emerge as a potent hub for facilitating the intricate synthesis of sulfur-containing polymers.

Effect of metal ions on lipopeptide secretion from Bacillus subtilis strain FJAT-4: Negative regulation by Ca2.

AIMS: This study aimed to investigate the effect of metal ions on lipopeptide production by Bacillus subtilis strain FJAT-4 and the mechanism of negative regulation by Ca2+ . METHODS AND RESULTS: The quantitative measurement of lipopeptides in response to K+ , Na+ , Mg2+ and Ca2+ addition was carried out by LC-MS. The contents of fengycin and surfactin varied within the range of 116.24-129.80 mg/L and 34.03-63.11 mg/L in the culture media containing K+ , Na+ and Mg2+ , while the levels were 0.86 and 0.63 mg/L in the media containing Ca2+ . Ca2+ at a high concentration (45 mM) did not adversely affect the growth of strain FJAT-4, but caused significant downregulation of lipopeptide synthesis-related gene expression, corresponding to a decrease in lipopeptide production. This inhibition by Ca2+ was further investigated by proteomic analysis. In total, 112 proteins were upregulated and 524 proteins were downregulated in the presence of additional Ca2+ (45 mM). Among these differentially expressed proteins (DEPs), 28 were related to phosphotransferase activity, and 42 were related to kinase activity. The proteomics results suggested that altered levels of three two-component signal-transduction systems (ResD/ResE, PhoP/PhoR and DegU/DegS) might be involved in the control of expression of the fen and srfA operons of FJAT-4 under high calcium stress. CONCLUSIONS: The Ca2+ at the high concentration (45 mM) triggers a decrease in lipopeptide production, which might be attributed to the regulation of three two-component signal-transduction systems ResD/ResE, PhoP/PhoR and DegU/DegS. SIGNIFICANCE AND IMPACT OF THE STUDY: The regulatory effect of calcium on the expression of genes encoding lipopeptide synthetases can be applied to optimize the production of lipopeptides.

First Report of Green Mold Disease Caused by Penicillium ochrochloron on Medicinal Plant Pseudostellaria heterophylla in China.

Pseudostellaria heterophylla is one of the Chinese herbal medicines with high medicinal and economic values. From 2019 to 2021, postharvest green mold disease was observed with an incidence of 2~5% on the tuberous roots of the harvested P. heterophylla at eight locations in Zherong county, Fujian Province, China. The symptoms were as follows: white mycelial growth on the tuberous roots surface initially, then green mold layers forming, and the tuberous roots decaying finally. To identify the causal agent, a total of 20 symptomatic tuberous roots were collected. Small pieces (5 mm×5 mm) were treated by surface disinfestion with 75% ethanol and 1% NaOCl, then rinsed 3 times with sterile distilled water. These treated pieces were transferred onto potato dextrose agar (PDA) and incubated at 25°C in the dark for 7 days. Twenty pure cultures were obtained by single-spore isolation method. Colonies on PDA medium initially appeared as white mycelium that developed grayish-green conidia with white margins. Mycelium was septate and colorless. Conidiophores were predominantly monoverticillate, occasionally biverticillate. Stipes was long and slender. Phialides were ampulliform to almost cylindrical with collula, 11.25 (7.80-23.50) µm long (n=50). Conidia were smooth walled and pale green, with globose to ellipsoidal shape, 2.75 (2.37-3.27)× 2.47 (2.18-3.13) µm (n=50). Based on these morphological characteristics, the isolates matched the description of the genus Penicillium. Genomic DNAs from two representative isolates (FJAT-32578 and FJAT-32579) were extracted with a fungal genomic DNA extraction kit. The rDNA ITS region and partial ß-tubulin gene (BenA) were amplified using the primers ITS1/ITS4 (White et al. 1990) and Bt2a/Bt2b (Glass and Donaldson 1995), respectively. The sequences of isolate FJAT-32578 and FJAT-32579 were deposited in GenBank (ITS, OM920986 and OM920987; BenA, OM953825 and OM953826). All sequences showed above 99% similarity to P. ochrochloron type strain CBS357.48 (ITS, NR111509; BenA, GU981672). In multilocus phylogenetic analysis (ITS + BenA), the two isolates from this study clustered together with other strains of P. ochrochloron with 100% bootstrap support. The two isolates were thus identified as P. ochrochloron based on both morphological and molecular characteristics. Pathogenicity tests were conducted in triplicate by inoculating the aseptic wounds with 10 µl of conidial suspension (1×106 conidia/ml) of the two isolates in the each healthy tuberous root (cv. Zheshen No.1). The experiment was conducted twice. All the inoculated tuberous roots were placed in sterilized Petri dishes with moistened filter paper, and incubated at 25 ± 2 °C. Fifteen days after inoculation, all inoculated tuberous roots demonstrated the same symptoms as those observed in the field conditions. The re-isolated fungi from the artificially infected tuberous roots were confirmed as P. ochrochloron using the method described above, while the control tuberous roots treated with sterile water did not develop symptoms, fulfilling Koch's postulates. To our knowledge, this is the first report of P. ochrochloron causing green mold disease on P. heterophylla in China, which would be a potentially new threat to the medicinal plant.

A Dpagt1 Missense Variant Causes Degenerative Retinopathy without Myasthenic Syndrome in Mice.

Congenital disorders of glycosylation (CDG) are a heterogenous group of primarily autosomal recessive mendelian diseases caused by disruptions in the synthesis of lipid-linked oligosaccharides and their transfer to proteins. CDGs usually affect multiple organ systems and vary in presentation, even within families. There is currently no cure, and treatment is aimed at ameliorating symptoms and improving quality of life. Here, we describe a chemically induced mouse mutant, tvrm76, with early-onset photoreceptor degeneration. The recessive mutation was mapped to Chromosome 9 and associated with a missense mutation in the Dpagt1 gene encoding UDP-N-acetyl-D-glucosamine:dolichyl-phosphate N-acetyl-D-glucosaminephosphotransferase (EC 2.7.8.15). The mutation is predicted to cause a substitution of aspartic acid with glycine at residue 166 of DPAGT1. This represents the first viable animal model of a Dpagt1 mutation and a novel phenotype for a CDG. The increased expression of Ddit3, and elevated levels of HSPA5 (BiP) suggest the presence of early-onset endoplasmic reticulum (ER) stress. These changes were associated with the induction of photoreceptor apoptosis in tvrm76 retinas. Mutations in human DPAGT1 cause myasthenic syndrome-13 and severe forms of a congenital disorder of glycosylation Type Ij. In contrast, Dpagt1tvrm76 homozygous mice present with congenital photoreceptor degeneration without overt muscle or muscular junction involvement. Our results suggest the possibility of DPAGT1 mutations in human patients that present primarily with retinitis pigmentosa, with little or no muscle disease. Variants in DPAGT1 should be considered when evaluating cases of non-syndromic retinal degeneration.

General Access to N-CF3 Secondary Amines and Their Transformation to N-CF3 Sulfonamides.

The synthesis of N-CF3 compounds through fluorination or trifluoromethylation of N-containing compounds has been extensively investigated. However, general access to N-CF3 compounds simply from N-CF3 secondary amines is hampered by the challenging preparation and instability of these amines, as well as a much lower reactivity due to the strong electron-withdrawing nature and steric bulk of the trifluoromethyl moiety. Herein, we report a general and highly efficient synthesis of N-CF3 secondary amines with excellent isolated yields via the addition of the in situ generated difluoromethyl imine (R-N=CF2 ) intermediates with hydrogen fluoride, which is mildly produced by triethylsilane and silver fluoride. N-CF3 sulfonamides, highly desirable but scarce at present, are easily accessible from these valuable building blocks through an unprecedented route. This study will bring new vitality to the synthesis of N-CF3 compounds.

Stress response mechanisms and description of three novel species Shewanella avicenniae sp. nov., Shewanella sedimentimangrovi sp. nov. and Shewanella yunxiaonensis sp. nov., isolated from mangrove ecosystem.

Three Gram-staining negative, facultatively anaerobic, rod-shaped and motile strains, FJAT-51800T, FJAT-52962T and FJAT-54481T were isolated from the sediment samples of Zhangjiang Estuary Mangrove National Nature Reserve in Fujian Province, China. The 16S rRNA gene sequencing results indicated they could be novel members of the genus Shewanella. The optimum temperature for growth was 30 °C. The respiratory quinones of the strains were ubiquinone Q-7 or Q-8, and menaquinone MK-7. Polar lipids of the strains FJAT-52962T and FJAT-51800T were phosphatidyl glycerol, phosphatidyl ethanolamine, and unidentified aminophospholipids while strain FJAT-54481 consist of phosphatidylglycerol, phosphatidylethanolamine, unidentified aminophospholipids, two unidentified aminolipids and four unidentified lipids. The major fatty acid of the three strains was iso-C15:0. The genomic DNA G + C contents of strains FJAT-51800T, FJAT-52962T and FJAT-54481T were 48.2, 55.3 and 48.1%, respectively. The average nucleotide identity and digital DNA-DNA hybridization values between strains FJAT-51800T, FJAT-52962T and FJAT-54481T and other closely related Shewanella members were below the cut-off level (95-96%) for species identification. Genome analysis showed that these strains encode genes for osmo-regulation. Based on the results of phenotypic, chemotaxonomic and genome analyses, strains FJAT-51800T, FJAT-52962T and FJAT-54481T represent three novel species of the genus Shewanella, for which the names Shewanella avicenniae sp. nov., Shewanella sedimentimangrovi sp. nov., and Shewanella yunxiaonensis sp. nov., are proposed. The type strains are FJAT-51800T (= GDMCC 1.2204T = KCTC 82448T), FJAT-52962T (= MCCC 1K05496T = KCTC 82445T) and FJAT-54481T (= GDMCC 1.2348T = KCTC 82646T).

Evidence that nano-TiO2 induces acute cytotoxicity to the agronomically beneficial nitrogen-fixing bacteria Sinorhizobium meliloti.

When nano-sized titanium dioxide (nano-TiO2) absorbs ultra-violet (UV-A) radiation, it produces reactive oxygen species that can be toxic to bacteria. We used the agronomically beneficial nitrogen-fixing bacterium Sinorhizobium meliloti strain 1021 as a model microorganism to detect nano-TiO2 toxicity. Sinorhizobium meliloti was exposed to aqueous dispersions of micrometer-sized TiO2 (micron-TiO2, 44 µm) or nanometer-sized TiO2 (nano-TiO2, 21 nm) at nominal concentrations of 0, 100, 300, 600, 900, and 1800 mg TiO2/L. There were fewer viable S. meliloti cells after exposure to nano-TiO2 under dark and UV-A light conditions. Nano-TiO2 was more toxic to S. meliloti with UV-A irradiation (100% mortality at 100 mg TiO2/L) than under dark conditions (100% mortality at 900 mg TiO2/L). Micron-TiO2 concentrations less than 300 mg TiO2/L had no effect on S. meliloti viability under dark or UV-A light conditions. Exposure to 600 mg/L or more of micron-TiO2 under UV-A light could also photo-kill S. meliloti cells (100% mortality). Further studies are needed to ascertain whether nano-TiO2 interferes with the growth of N2-fixing microorganisms in realistic agricultural environments.

Biocontrol of tomato bacterial wilt by the new strain Bacillus velezensis FJAT-46737 and its lipopeptides.

BACKGROUND: There is an urgent need to discover biocontrol agents to control bacterial wilt. This study reports on a new lipopeptide-producing biocontrol strain FJAT-46737 and explores its lipopeptidic compounds, and this study investigates the antagonistic effects of these compounds. RESULTS: Based on a whole genome sequence analysis, the new strain FJAT-46737 was identified as Bacillus velezensis, and seven gene clusters responsible for the synthesis of bioactive secondary metabolites in FJAT-46737 were predicted. The antimicrobial results demonstrated that FJAT-46737 exhibited broad-spectrum antimicrobial activities in vitro against three bacteria and three fungi. Pot experiments showed that the control efficiencies for tomato bacterial wilt of the whole cultures, the 2-fold diluted supernatants and the crude lipopeptide of FJAT-46737 were 66.2%, 82.0%, and 96.2%, respectively. The above results suggested that one of the antagonistic mechanisms of FJAT-46737 was the secretion of lipopeptides consisting of iturins, fengycins and surfactins. The crude lipopeptides had significant antagonistic activities against several pathogens (including Ralstonia solanacearum, Escherichia coli and Fusarium oxysporum) and fengycins were the major antibacterial components of the lipopeptides against R. solanacearum in vitro. Furthermore, the rich organic nitrogen sources (especially yeast extracts) in the media promoted the production of fengycin and surfactin by FJAT-46737. The secretion of these two lipopeptides was related to temperature fluctuations, with the fengycin content decreasing by 96.6% and the surfactins content increasing by 59.9% from 20 °C to 40 °C. The optimal temperature for lipopeptide production by FJAT-46737 varied between 20 °C and 25 °C. CONCLUSIONS: The B. velezensis strain FJAT-46737 and its secreted lipopeptides could be used as new sources of potential biocontrol agents against several plant pathogens, and especially the bacterial wilt pathogen R. solanacearum.

Bacillus dafuensis sp. Nov., Isolated from a Forest Soil in China.

A Gram-staining-positive, motile, rod-shaped, aerobic bacterial strain FJAT-25496 T was isolated from a forest soil sample collected from Dafu County, Chengdu City, Sichuan Province, China. Strain FJAT-25496 T grew at 15-50 °C (optimum 35 °C), pH 6.0-10.0 (optimum 8.0), and NaCl tolerance of 0-4.0% (w/v) (optimum 0%). The strain contained iso-C15:0 and anteiso-C15:0 as the predominant cellular fatty acids as well as MK-7 as the only menaquinone. The major polar lipids were diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE), and phosphatidyl glycerol (PG). Phylogenetic analyses based on 16S rRNA gene sequences showed that strain FJAT-25496 T was a member of the genus Bacillus, and most closely related to Bacillus solani FJAT-18043 T (97.8%), Bacillus praedii FJAT-25547 T (97.8%), Bacillus depressus BZ1T (97.7%), Bacillus oceanisediminis H2T (97.7%), and Bacillus ciccensis 5L6T (97.6%). Based on complete genome comparisons between strain FJAT-25496 T and the type strains of its closely related species, the average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values were 73.4-76.3% and 21.7-23.2%, respectively. The genomic DNA G + C content of strain FJAT-25496 T was 36.9%. On the basis of phenotypic, phylogenetic, chemotaxonomic, and genomic features, strain FJAT-25496 T is considered to represent a novel species of the genus Bacillus, for which the name Bacillus dafuensis sp. nov. is proposed. The type strain is FJAT-25496 T (= CCTCC AB 2019182 T = KCTC 43120 T).

Reclassification of Brevibacterium frigoritolerans DSM 8801T as Bacillus frigoritolerans comb. nov. Based on Genome Analysis.

16S rRNA gene sequence analysis showed that the type strain Brevibacterium frigoritolerans DSM 8801T had the highest similarity (99.7%) with the reference strain Bacillus simplex NBRC 15720T, followed by Bacillus muralis DSM 16288T (99.6%), Bacillus butanolivorans DSM 18926T (99.5%), and Bacillus loiseleuriae FJAT-27997T (97.9%). This relationship is confirmed by the phylogenetic analysis indicating that Bre. frigoritolerans DSM 8801T fell in the genus Bacillus group and formed a clade with the closely related Bacillus species. Average nucleotide identity and in silico DNA-DNA hybridization (isDDH) values between strain DSM 8801T and the most closed reference strain Bac. simplex NBRC 15720T were both much lower than species definition threshold values of 95% and 70%, respectively, which indicated strain DSM 8801T should not be affiliated to one of the validly named Bacillus species. The percentage of conserved protein (POCP) values between the strain DSM 8801T and the type strains of the above species were 80.7%, 69.2%, 72.2%, 53.6%, and 50.0%, respectively, higher than the genus definition threshold value of 50%. The main isoprenoid quinone of strain DSM 8801T was MK-7, the main polar lipids were diphosphatidylglycerol (DPG), phosphatidylglycerol (PG), and phosphatidyl ethanolamine (PE), and the major fatty acids were anteiso-C15:0 and iso-C15:0. On the basis of the phenotypic, chemotaxonomic, phylogenetic and genomic characteristics, Bre. frigoritolerans DSM 8801T should belong to the genus Bacillus, and be proposed to reclassify as Bacillus frigoritolerans comb. nov., with the type strain DSM 8801T (=ATCC 25097T  = CCUG 43489T  = CIP 67.20T  = JCM 11681T).

Strong inhibitory activities and action modes of lipopeptides on lipase.

Lipopeptides have been reported to exhibit anti-obesity effects. In this study, we obtained a Bacillus velezensis strain FJAT-52631 that could coproduce iturins, fengycins, and surfactins. Results showed that the FJAT-52631 crude lipopeptide, purified fengycin, iturin, and surfactin standards exhibited strong inhibition activities against lipase with dose-dependence manners (half maximal inhibitory concentration (IC50) = 0.011, 0.005, 0.056, and 0.005 mg/mL, respectively). Moreover, fengycin and surfactin had the comparable activities with orlistat, but iturin not. It was revealed that the inhibition mechanism and type of the lipopeptides were reversible and competitive. The quenching mechanism of lipase was static and only one binding site between lipase and lipopoeptide was inferred from the fluorescence analysis. The docking analysis displayed that fengycin and surfactin could directly interact with the active amino acid residues (Ser or Asp) of lipase, but not with iturin. Our work suggests that the B. velezensis lipopeptides would have great potential to act as lipase inhibitors.

Genomic insights into mite phylogeny, fitness, development, and reproduction.

BACKGROUND: Predatory mites (Acari: Phytoseiidae) are the most important beneficial arthropods used in augmentative biological pest control of protected crops around the world. However, the genomes of mites are far less well understood than those of insects and the evolutionary relationships among mite and other chelicerate orders are contested, with the enigmatic origin of mites at one of the centres in discussion of the evolution of Arachnida. RESULTS: We here report the 173 Mb nuclear genome (from 51.75 Gb pairs of Illumina reads) of the predatory mite, Neoseiulus cucumeris, a biocontrol agent against pests such as mites and thrips worldwide. We identified nearly 20.6 Mb (~ 11.93% of this genome) of repetitive sequences and annotated 18,735 protein-coding genes (a typical gene 2888 bp in size); the total length of protein-coding genes was about 50.55 Mb (29.2% of this assembly). About 37% (6981) of the genes are unique to N. cucumeris based on comparison with other arachnid genomes. Our phylogenomic analysis supported the monophyly of Acari, therefore rejecting the biphyletic origin of mites advocated by other studies based on limited gene fragments or few taxa in recent years. Our transcriptomic analyses of different life stages of N. cucumeris provide new insights into genes involved in its development. Putative genes involved in vitellogenesis, regulation of oviposition, sex determination, development of legs, signal perception, detoxification and stress-resistance, and innate immune systems are identified. CONCLUSIONS: Our genomics and developmental transcriptomics analyses of N. cucumeris provide invaluable resources for further research on the development, reproduction, and fitness of this economically important mite in particular and Arachnida in general.

Photoreceptor degeneration in a new Cacna1f mutant mouse model.

The Cacna1f gene encodes the α1F subunit of an L-type voltage-gated calcium channel, Cav1.4. In photoreceptor synaptic terminals, Cav1.4 channels mediate glutamate release and postsynaptic responses associated with visual signal transmission. We have discovered a new Cacna1f mutation in nob9 mice, which display more severe phenotypes than do nob2 mice. To characterize the nob9 phenotype at different ages, we examined the murine fundus, applied retinal optical coherence tomography, measured flash electroretinograms (ERGs) in vivo, and analyzed the retinal histology in vitro. After identifying the X-linked recessive inheritance trait, we sequenced Cacna1f as the candidate gene. Mutations in this gene were detected by polymerase chain reaction (PCR) and confirmed by restriction fragment length polymorphism. Morphologically, an early-onset of retinal disorder was detected, and the degeneration of the outer plexiform layers progressed rapidly. Moreover, the mutant mice showed drastically reduced scotopic ERGs with increasing age. In 14-month-old nob9 retinas, immunostaining of cone opsins demonstrated a reduction in the number of short-wavelength opsins (S-opsins) to 54% of wild-type levels, and almost no middle-wavelength opsins (M-opsins) were observed. No cone ERGs could be detected from residual cones, in which S-opsins abnormally migrated to inner segments of the photoreceptors. The mutations of the Cacna1f gene in nob9 mice involved both a single nucleotide G to A transition and a 10-nucleotide insertion, the latter resulting in a frame-shift mutation in exon 14.