Putative Secondary Metabolite Research Articles

Antibacterial Compounds and Draft Genome Sequence of Streptomyces sp. PA5.6 Isolated from Deciduous Dipterocarp Forest Soils, University of Phayao, Thailand

Antibiotic resistance is a major problem in the control of infectious diseases. To overcome this obstacle, a common approach is to search for novel antimicrobial drugs from natural sources, especially microorganisms such as Streptomyces. Streptomyces has long been demonstrated to produce bioactive secondary metabolites with antimicrobial activities. The objectives of this research were to isolate and characterize Streptomyces strains from the deciduous dipterocarp forest soils in the University of Phayao, and to evaluate their metabolites for antibacterial activities. The results show that the morphological and physiological characteristics of PA5.6 are consistent with those of the genus Streptomyces. Furthermore, a comparative analysis of the whole genome sequence verified that PA5.6 represents a novel species within the genus Streptomyces. The genome sequence analysis of Streptomyces sp. PA5.6 revealed a total length of 9,146,340 bp, a G + C content of 71.1 %, and an N50 scaffold of 320,345 bp. The antiSMASH analysis revealed putative secondary metabolite Biosynthetic Gene Clusters (BGCs) involved in the biosynthesis of antimicrobial metabolites, including terpene, Type I PKS, NRPS and lassopeptide gene clusters. Functional prediction of these gene clusters indicates that they are involved in the biosynthesis of several antimicrobial-associated metabolites, such as albaflavenone, monensin and citrulassin D. Streptomyces sp. PA5.6 culture supernatant and its crude ethyl acetate extract exhibited antibacterial activity against several drug-resistant, bacterial pathogens, including S. aureus, E. faecalis, E. coli, P. aeruginosa, A. baumannii, MRSA, VREF, ESBL-E. coli and CRPA. The GC-MS analysis of the crude ethyl acetate extracts revealed benzeneacetic acid, benzeneacetic acid, 4-hydroxy and benzeneacetamide as major active compounds metabolites. In conclusion, Streptomyces sp. PA5.6 exhibited promising potential for producing bioactive compounds against bacterial pathogens. HIGHLIGHTSThe Streptomyces sp. PA5.6 isolated from the soil in the deciduous dipterocarp forest and was identified as a new species within the genus Streptomyces by comparative analysis of the whole genome sequence. The crude ethyl acetate extract of Streptomyces sp. PA5.6 exhibited antibacterial activity against bacterial pathogens. GC-MS analysis of crude ethyl acetate extract showed major and minor compounds that exhibit antimicrobial activity. GRAPHICAL ABSTRACT

Exploring the Role of a Putative Secondary Metabolite Biosynthesis Pathway in Mycobacterium abscessus Pathogenesis Using a Xenopus laevis Tadpole Model.

Mycobacterium abscessus (Mab) is an emerging human pathogen that has a high rate of incidence in immunocompromised individuals. We have found a putative secondary metabolite pathway within Mab, which may be a key factor in its pathogenesis. This novel pathway is encoded in a gene cluster spanning MAB_0284c to 0305 and is related to Streptomyces pathways, producing the secondary metabolites streptonigrin and nybomycin. We constructed an in-frame deletion of the MAB_0295 (phzC) gene and tested it in our Xenopus laevis animal model. We have previously shown that X. laevis tadpoles, which have functional lungs and T cells, can serve as a reliable comparative model for persistent Mab infection and pathogenesis. Here, we report that tadpoles intraperitoneally infected with the ∆phzC mutant exhibit early decreased bacterial loads and significantly increased survival compared with those infected with WT Mab. ∆phzC mutant Mab also induced lower transcript levels of several pro-inflammatory cytokines (IL-1β, TNF-α, iNOS, IFN-γ) than those of WT Mab in the liver and lungs. In addition, there was impaired macrophage recruitment and decreased macrophage infection in tadpoles infected with the ∆phzC mutant, by tail wound inoculation, compared to those infected with the WT bacteria, as assayed by intravital confocal microscopy. These data underline the relevance and usefulness of X. laevis tadpoles as a novel comparative animal model to identify genetic determinants of Mab immunopathogenesis, suggesting a role for this novel and uncharacterized pathway in Mab pathogenesis and macrophage recruitment.

Molecular Phylogenetic and Comparative Genomic Analysis of Pleurocordyceps fusiformispora sp. nov. and Perennicordyceps elaphomyceticola in the Family Polycephalomycetaceae.

Several Pleurocordyceps species have been reported as hyperparasitic fungi. A new species, Pleurocordyceps fusiformispora, and a known species, Perennicordyceps elaphomyceticola, are described here based on morphology and phylogenetic evidence from six genes (ITS, SSU, LSU, TET1-α, RPB1, and RPB2). Pl. fusiformispora differed from the other Pleurocordyceps species by producing flaky colonies, ovoid or elliptic α-conidia, and fusiform or long fusiform β-conidia. Both full genomes of Pe. elaphomyceticola and Pl. fusiformispora were sequenced, annotated, and compared. The antiSMASH and local BLAST analyses revealed significant differences in the number and types of putative secondary metabolite biosynthetic gene clusters, i.e., NPPS, PKS, and hybrid PKS-NRPS domains, between the two species. In addition, the putative BGCs of six compounds, namely ε-poly lysine, 4-epi-15-epi-brefeldin A, Monorden D/monocillin IV/monocillin VII/pochonin M/monocillin V/monocillin II, Tolypyridone, Piperazine, and Triticone DABFC, were excavated in the present study. This study motivates the use of heterologous expression and gene knockout methods to discover novel biologically active SMs from Polycephalomycetaceae.

Metabolic re-programming in confrontations of Colletotrichum graminicola and Aspergillus nidulans with Bacillus biocontrol agents

We established confrontations between two different fungi, i.e., the maize anthracnose and stalk rot pathogen Colletotrichum graminicola, and the ubiquitous fungus Aspergillus nidulans, and different biocontrol species, i.e., Bacillus subtilis, Bacillus velezensis, and Bacillus amyloliquefaciens. In all fungus–bacterium confrontations tested, growth arrest and, thus, distance inhibition was observed on solid substrata. LC–MS/MS analyses of culture filtrates suggested formation of several metabolites only synthesized in confrontations. Interestingly, microscopy of fungal hyphae grown in liquid medium showed protrusions and color changes occurred only in media harboring fungus-bacterium confrontations. These observations indicate metabolic re-programming and suggest formation of putative secondary metabolites in interactions involving microbial biocontrol agents.

Genome Sequence Analysis of Native Xenorhabdus Strains Isolated from Entomopathogenic Nematodes in Argentina.

Entomopathogenic nematodes from the genus Steinernema (Nematoda: Steinernematidae) are capable of causing the rapid killing of insect hosts, facilitated by their association with symbiotic Gram-negative bacteria in the genus Xenorhabdus (Enterobacterales: Morganellaceae), positioning them as interesting candidate tools for the control of insect pests. In spite of this, only a limited number of species from this bacterial genus have been identified from their nematode hosts and their insecticidal properties documented. This study aimed to perform the genome sequence analysis of fourteen Xenorhabdus strains that were isolated from Steinernema nematodes in Argentina. All of the strains were found to be able of killing 7th instar larvae of Galleria mellonella (L.) (Lepidoptera: Pyralidae). Their sequenced genomes harbour 110 putative insecticidal proteins including Tc, Txp, Mcf, Pra/Prb and App homologs, plus other virulence factors such as putative nematocidal proteins, chitinases and secondary metabolite gene clusters for the synthesis of different bioactive compounds. Maximum-likelihood phylogenetic analysis plus average nucleotide identity calculations strongly suggested that three strains should be considered novel species. The species name for strains PSL and Reich (same species according to % ANI) is proposed as Xenorhabdus littoralis sp. nov., whereas strain 12 is proposed as Xenorhabdus santafensis sp. nov. In this work, we present a dual insight into the biocidal potential and diversity of the Xenorhabdus genus, demonstrated by different numbers of putative insecticidal genes and biosynthetic gene clusters, along with a fresh exploration of the species within this genus.

Genomic Comparison of Two Species of Samsoniella with Other Genera in the Family Cordycipitaceae.

Whole genomes of Samsoniella hepiali ICMM 82-2 and S. yunnanensis YFCC 1527 were sequenced and annotated, as well as compared with whole genome sequences of other species in the family Cordycipitaceae. S. hepiali ICMM 82-2, S. hepiali FENG and S. yunnanensis YFCC 1527 had 54, 57 and 58 putative secondary metabolite biosynthetic gene clusters, respectively. S. hepiali had one unique domain and S. yunnanensis YFCC 1527 six. Both S. hepiali and S. yunnanensis YFCC 1527 had curvupallide-B, fumosorinone and fujikurin putative biosynthetic gene clusters. C. javanica had biosynthetic gene clusters for fumonisin. The 14 genomes had common domains, namely A-P-C-P-C and KS-AT-DH-ER-KR-ACP. The A-P-C-P-C domain may be involved in the biosynthesis of dimethylcoprogen. The maximum likelihood and the Bayesian inference trees of KS-AT-DH-ER-KR-ACP were highly consistent with the multigene phylogenetic tree for the 13 species of Cordycipitaceae. This study facilitates the discovery of novel biologically active SMs from Cordycipitaceae using heterologous expression and gene knockdown methods.

Rhizospheric Bacillus spp. Exhibit Miticidal Efficacy against Oligonychus coffeae (Acari: Tetranychidae) of Tea.

Oligonychus coffeae (Acari: Tetranychidae), popularly known as red spider mite (RSM) is one of the major pests of commercial tea (Camellia sinensis (L.) O. Kuntze) plantation world over. Many attempts have been made in the past to control this devastating pest using a variety of microbial bioagents, however, area-wise field success is very limited. We carried out an in vitro study to explore the potential of rhizospheric Bacillus spp. (B. amyloliquefaciens BAC1, B. subtilis LB22, and B. velezensis AB22) against O. coffeae through adulticidal and ovicidal activity. The 100% adult and egg mortality was observed with bacterial suspension (1 × 109 CFU/mL) by B. velezensis AB22, showing the lowest LC50 values for both adults and eggs of O. coffeae, i.e., 0.28 × 105 and 0.29 × 105, respectively. The study also throws some insights into the underlying mechanism through electron microscopy study and identification of some putative pesticidal metabolites from all the species. The three Bacillus species were observed to have four commonly secreted putative bioactive secondary metabolites, brevianamide A, heptadecanoic acid, thiolutin, and versimide responsible for their bio-efficacy against O. coffeae. The outcome of our study provides a strong possibility of introducing Bacillus spp. as a biological miticide and developing synthetic metabolites mimicking the mechanistic pathway involved in microbial bioefficacy.

Taxonomy-guided selection of Paraburkholderia busanensis sp. nov.: a versatile biocontrol agent with mycophagy against Colletotrichum scovillei causing pepper anthracnose.

This study introduces a novel approach based on the taxonomy-guided selection of bacterial biocontrol agents from a known beneficial taxonomic group. Following 16S rRNA screening, we focused on the genus Paraburkholderia, which harbors strains with large genomes and versatile benefits to plants. A strain designated P39 was selected, identified, and characterized for its biocontrol activity against Colletotrichum scovillei. Strain P39 exhibited antagonism against C. scovillei by producing compounds, including volatiles, with antifungal activity, both in vitro and on pepper fruits. Genomic, physiological, and biochemical analyses revealed that the selected strain represents a novel species, named Paraburkholderia busanensis. Genomic analyses provided insights into the fitness and biocontrol activities of the selected strains. Moreover, P39 displays mycophagy, consuming fungal mycelia and transforming them into bacterial biomass, particularly in nutrient-poor media supplemented with fungal mycelia. The genome harbored chitin and N-acetylglucosamine utilization genes, suggesting a proposed pathway for the utilization of fungal cells as a nutrient source. Microscopic observations further supported the ability of this strain to rupture and damage fungal hyphae, depriving them of their cellular constituents. This study successfully demonstrated the implementation of a taxonomy-guided approach for the selection of bacterial strains for biocontrol. These findings contribute to our understanding of biocontrol strategies, bacteria-fungi interactions, and the identification of Paraburkholderia busanensis sp. nov. as a potential candidate for the biocontrol of pepper anthracnose. Additionally, this strain serves as a valuable resource for antifungal compounds and volatiles, and for the study of bacteria-fungal interactions and mycophagy. IMPORTANCE Traditional control methods for postharvest diseases rely on fungicides, which cause human health and environmental concerns. This study introduces a taxonomy-guided strategy for selecting biocontrol agents. By focusing on Paraburkholderia group, which harbors diverse plant-beneficial strains, the inadvertent selection of harmful strains was circumvented, thereby obviating the need for laborious in vitro screening assays. A highly promising candidate, strain P39, has been identified, exhibiting remarkable biocontrol activity against Colletotrichum scovillei. Through comprehensive genomic, physiological, and biochemical analyses, P39 was characterized as a novel species within the Paraburkholderia genus and designated Paraburkholderia busanensis. Moreover, these findings deepen our understanding of bacterial-fungal interactions, as they elucidate a potential pathway for the utilization of fungal chitin, thereby enhancing our understanding of bacterial mycophagy. P. busanensis is a promising source of antifungal volatiles and putative novel secondary metabolites.

Integrated Human Skin Bacteria Genome Catalog Reveals Extensive Unexplored Habitat-Specific Microbiome Diversity and Function.

The skin is the largest organ in the human body. Various skin environments on its surface constitutes a complex ecosystem. One of the characteristics of the skin micro-ecosystem is low biomass, which greatly limits a comprehensive identification of the microbial species through sequencing. In this study, deep-shotgun sequencing (average 21.5Gigabyte (Gb)) from 450 facial samples and publicly available skin metagenomic datasets of 2069 samples to assemble a Unified Human Skin Genome (UHSG) catalog is integrated. The UHSG encompasses 813 prokaryotic species derived from 5779 metagenome-assembled genomes, among which 470 are novel species covering 20 phyla with 1385 novel assembled genomes. Based on the UHSG, the core functions of the skin microbiome are described and the differences in amino acid metabolism, carbohydrate metabolism, and drug resistance functions among different phyla are identified. Furthermore, analysis of secondary metabolites of the near-complete genomes further find 1220 putative novel secondary metabolites, several of which are found in previously unknown genomes. Single nucleotide variant (SNV) reveals a possible skin protection mechanism: the negative selection process of the skin environment to conditional pathogens. UHSG offers a convenient reference database that will facilitate a more in-depth understanding of the role of skin microorganisms in the skin.

Genomic Analysis of a Novel Antarctic Bacterium, Cyrobacterium sp.SO2 Provides Insights into its Genomic Potential for Production of Antimicrobial Compounds

A novel strain of Cryobacterium designated as SO2, was isolated from the Antarctic. Hence, this study was undertaken to gain further insight into the antimicrobial compounds and secondary metabolites produced by Cryobacterium sp. SO2. It was found that strain SO2 is a Gram-positive that exhibits an irregular rod shape, which formed yellow to orange pigmented colonies on semi-solid media. Strain SO2 grows at temperatures ranging from 4 to 25 ºC. It has a complete genomic size of 4.097 Mb. SO2 has a DNA G+C content of 68.43%, and genomic annotation showed that the genome contained 3,862 CDS, 10 rRNA, 55 tRNA and 1 tm-RNA. Phylogenetic and OrthoANI analysis suggested Cryobacterium sp. strains SO1, N22, TMB1-8, LW097, TMN39-1, C. zongtaiiTMN-42, C. arcticumPAMC27867 and C. soli GCJ02 as its closest phylogenetic neighbour. Genome annotation shows that strain SO2 confers β-lactamase class A, cephalosporin-C deacetylases, and 27 drug-resistance encoding genes, and allows resistance to ceftazidime. Functional annotation identifies 28.74% of predicted genes are of unknown functions. Genome mining indicates that there are six putative secondary metabolite gene clusters in strain SO2. They are made up of RRE-containing, terpene, beta-lactone, T3PKS, NAPAA, and 2dos. This finding shows strain SO2 harbours genes that may be involved in the production of compounds with antibacterial and antioxidant activities.

Identification and characterization of Brevibacillus halotolerans B-4359: a potential antagonistic bacterium against red pepper anthracnose in Korea.

Our study aimed to identify potential biocontrol agents (BCAs) against major phytopathogens under in vitro conditions by screening the Freshwater Bioresources Culture Collection (FBCC), Korea. Of the identified 856 strains, only 65 exhibited antagonistic activity, among which only one representative isolation, Brevibacillus halotolerans B-4359 was selected based on its in vitro antagonistic activity and enzyme production. Cell-free culture filtrate (CF) and volatile organic compounds (VOCs) of B-4359 were shown to be effective against the mycelial growth of Colletotrichum acutatum. Notably, B-4359 was found to promote spore germination in C. acutatum instead of exhibiting a suppressive effect when the bacterial suspension was mixed with the spore suspension of C. acutatum. However, B-4359 showed an excellent biological control effect on the anthracnose of red pepper fruits. Compared to other treatments and untreated control, B-4359 played a more effective role in controlling anthracnose disease under field conditions. The strain was identified as B. halotolerans using BIOLOG and 16S rDNA sequencing analyses. The genetic mechanism underlying the biocontrol traits of B-4359 was characterized using the whole-genome sequence of B-4359, which was closely compared with related strains. The whole-genome sequence of B-4359 consisted of 5,761,776 bp with a GC content of 41.0%, including 5,118 coding sequences, 117 tRNA, and 36 rRNA genes. The genomic analysis identified 23 putative secondary metabolite biosynthetic gene clusters. Our results provide a deep understanding of B-4359 as an effective biocontrol agent against red pepper anthracnose for sustainable agriculture.

Comparative genome features and secondary metabolite biosynthetic potential of Kutzneria chonburiensis and other species of the genus Kutzneria

Actinobacteria are well known as a rich source of diversity of bioactive secondary metabolites. Kutzneria, a rare actinobacteria belonging to the family Pseudonocardiaceae has abundance of secondary metabolite biosynthetic gene clusters (BGCs) and is one of important source of natural products and worthy of priority investigation. Currently, Kutzneria chonburiensis SMC256T has been the latest type-strain of the genus and its genome sequence has not been reported yet. Therefore, we present the first report of new complete genome sequence of SMC256T (genome size of 10.4 Mbp) with genome annotation and feature comparison between SMC256T and other publicly available Kutzneria species. The results from comparative and functional genomic analyses regarding the phylogenomic and the clusters of orthologous groups of proteins (COGs) analyses indicated that SMC256T is most closely related to Kutzneria sp. 744, Kutzneria kofuensis, Kutzneria sp. CA-103260 and Kutzneria buriramensis. Furthermore, a total of 322 BGCs were also detected and showed diversity among the Kutzneria genomes. Out of which, 38 clusters showing the best hit to the most known BGCs were predicted in the SMC256Tgenome. We observed that six clusters responsible for biosynthesis of antimicrobials/antitumor metabolites were strain-specific in Kutzneria chonburiensis. These putative metabolites include virginiamycin S1, lysolipin I, esmeraldin, rakicidin, aclacinomycin and streptoseomycin. Based on these findings, the genome of Kutzneria chonburiensis contains distinct and unidentified BGCs different from other members of the genus, and the use of integrative genomic-based approach would be a useful alternative effort to target, isolate and identify putative and undiscovered secondary metabolites suspected to have new and/or specific bioactivity in the Kutzneria.

Genomic and phenotypic characterization of a red-pigmented strain of Massilia frigida isolated from an Antarctic microbial mat.

The McMurdo Dry Valleys of Antarctica experience a range of selective pressures, including extreme seasonal variation in temperature, water and nutrient availability, and UV radiation. Microbial mats in this ecosystem harbor dense concentrations of biomass in an otherwise desolate environment. Microbial inhabitants must mitigate these selective pressures via specialized enzymes, changes to the cellular envelope, and the production of secondary metabolites, such as pigments and osmoprotectants. Here, we describe the isolation and characterization of a Gram-negative, rod-shaped, motile, red-pigmented bacterium, strain DJPM01, from a microbial mat within the Don Juan Pond Basin of Wright Valley. Analysis of strain DJMP01's genome indicates it can be classified as a member of the Massilia frigida species. The genome contains several genes associated with cold and salt tolerance, including multiple RNA helicases, protein chaperones, and cation/proton antiporters. In addition, we identified 17 putative secondary metabolite gene clusters, including a number of nonribosomal peptides and ribosomally synthesized and post-translationally modified peptides (RiPPs), among others, and the biosynthesis pathway for the antimicrobial pigment prodigiosin. When cultivated on complex agar, multiple prodiginines, including the antibiotic prodigiosin, 2-methyl-3-propyl-prodiginine, 2-methyl-3-butyl-prodiginine, 2-methyl-3-heptyl-prodiginine, and cycloprodigiosin, were detected by LC-MS. Genome analyses of sequenced members of the Massilia genus indicates prodigiosin production is unique to Antarctic strains. UV-A radiation, an ecological stressor in the Antarctic, was found to significantly decrease the abundance of prodiginines produced by strain DJPM01. Genomic and phenotypic evidence indicates strain DJPM01 can respond to the ecological conditions of the DJP microbial mat, with prodiginines produced under a range of conditions, including extreme UV radiation.

Genome Sequence Resources for Verticillium dahliae-Inhibiting Streptomyces spp. Isolated from Agricultural Soil

Gram-positive Streptomyces bacteria are common in soil, where they can inhibit soilborne plant pathogens by producing antimicrobial secondary metabolites. This study generated genome assemblies of Streptomyces isolates from soil that inhibit the soilborne plant pathogen Verticillium dahliae. Five bacterial isolates were cultured from soil collected from the Salinas Valley region of California. In vitro tests showed that three bacterial isolates inhibited V. dahliae growth when applied at low and high spore concentrations, whereas two isolates only inhibited V. dahliae when applied at high spore concentrations. Generation of hybrid Illumina-Nanopore assemblies showed that isolate genomes ranged from 7.45 to 8.95 Mbs. The contig number per assembly ranged from 1 to 54. All assemblies had 100% completeness values based on a standard set of conserved orthologs found in all bacteria. The number of predicted genes ranged from 6,594 to 8,098. Over 25 putative secondary metabolite biosynthetic clusters were identified in each assembly. Calculation of digital DNA:DNA hybridization and average nucleotide identity values using additional genome assemblies from type specimens of Streptomyces showed that the five isolates are likely novel Streptomyces spp. The results and data generated from this study will provide a basis for future research on the use of Streptomyces for managing common soilborne pathogens such as V. dahliae. [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, 2023.

Genomic insights into a Pseudomonas amygdali isolate from Hibiscus rosa-sinensis

The taxonomy of Pseudomonas has been extensively studied, yet the determination of species is currently difficult because of recent taxonomic changes and the lack of complete genomic sequence data. We isolated a bacterium causing a leaf spot disease on hibiscus (Hibiscus rosa-sinensis). Whole genome sequencing revealed similarity to Pseudomonas amygdali pv. tabaci and pv. lachrymans. The genome of this isolate (referred to as P. amygdali 35–1) shared 4987 genes with P. amygdali pv. hibisci, but possessed 204 unique genes and contained gene clusters encoding putative secondary metabolites and copper resistance determinants. We predicted this isolate's type III secretion effector (T3SE) repertoire and identified 64 putative T3SEs, some of which are present in other P. amygdali pv. hibisci strains. Assays showed that the isolate was resistant to copper at a concentration of 1.6 mM. This study provides an improved understanding of the genomic relatedness and diversity of the P. amygdali species.

A putative terpene cyclase gene (CcPtc1) is required for fungal development and virulence in Cytospora chrysosperma.

Cytospora chrysosperma is a destructive plant pathogenic fungus, which causes canker disease on numerous woody plants. However, knowledge concerning the interaction between C. chrysosperma and its host remains limited. Secondary metabolites produced by phytopathogens often play important roles in their virulence. Terpene cyclases (TC), polyketide synthases (PKS) and non-ribosomal peptide synthetases (NRPS) are the key components for the synthesis of secondary metabolites. Here, we characterized the functions of a putative terpene type secondary metabolite biosynthetic core gene CcPtc1 in C. chrysosperma, which was significantly up-regulated in the early stages of infection. Importantly, deletion of CcPtc1 greatly reduced fungal virulence to the poplar twigs and they also showed significantly reduced fungal growth and conidiation compared with the wild-type (WT) strain. Furthermore, toxicity test of the crude extraction from each strain showed that the toxicity of crude extraction secreted by ΔCcPtc1 were strongly compromised in comparison with the WT strain. Subsequently, the untargeted metabolomics analyses between ΔCcPtc1 mutant and WT strain were conducted, which revealed 193 significantly different abundant metabolites (DAMs) inΔCcPtc1 mutant compared to the WT strain, including 90 significantly downregulated metabolites and 103 significantly up-regulated metabolites, respectively. Among them, four key metabolic pathways that reported to be important for fungal virulence were enriched, including pantothenate and coenzyme A (CoA) biosynthesis. Moreover, we also detected significant alterations in a series of terpenoids, among which (+)-ar-turmerone, pulegone, ethyl chrysanthemumate, and genipin were significantly down-regulated, while cuminaldehyde and (±)-abscisic acid were significantly up-regulated. In conclusion, our results demonstrated that CcPtc1 acts as a virulence-related secondary metabolism factor and provides new insights into the pathogenesis of C. chrysosperma.

Mucilaginibacter straminoryzae sp. nov., isolated from rice straw used for growing periphyton.

A rod-shaped, non-motile, Gram-negative bacterium, strain RS28T, was isolated from rice straw used as material for periphyton growth. Phylogenetic analysis of the 16S rRNA gene sequence revealed that strain RS28T was affiliated with the genus Mucilaginibacter and had the highest sequence similarity to Mucilaginibacter ginkgonis HMF7856T (96.47 %) and Mucilaginibacter polytrichastri DSM 26907T (96.12 %). Strain RS28T was found to grow at pH 5.5-8.0, 17-40 °C and in the presence of 0-1.5 % (w/v) NaCl. Strain RS28T contained summed feature 3 (comprising C16 : 1 ω7c and/or C16 : 1 ω6c), iso-C15 : 0 and iso-C17 : 0 3-OH as the major fatty acids (> 10.0 %). The major polar lipids were phosphatidylethanolamine, two unidentified phospholipids, two unidentified aminophospholipids, three unidentified aminolipids and one unidentified lipid. The respiratory quinone was menaquinone 7. The genomic DNA G+C content was 44.7 mol%. Strain RS28T possessed six putative secondary metabolite gene clusters involved in the synthesis of resorcinol, NRPS-like, terpene, lassopeptide, T3PKS and arylpolyene. On the basis of the phenotypic, chemotaxonomic, and phylogenetic characteristics, strain RS28T represents a novel species of the genus Mucilaginibacter, for which the name Mucilaginibacter straminoryzae sp. nov. is proposed. The type strain is RS28T (=KCTC 92039T=LMG 32424T).

Mining of novel secondary metabolite biosynthetic gene clusters from acid mine drainage.

Acid mine drainage (AMD) is usually acidic (pH < 4) and contains high concentrations of dissolved metals and metalloids, making AMD a typical representative of extreme environments. Recent studies have shown that microbes play a key role in AMD bioremediation, and secondary metabolite biosynthetic gene clusters (smBGCs) from AMD microbes are important resources for the synthesis of antibacterial and anticancer drugs. Here, 179 samples from 13 mineral types were used to analyze the putative novel microorganisms and secondary metabolites in AMD environments. Among 7,007 qualified metagenome-assembled genomes (MAGs) mined from these datasets, 6,340 MAGs could not be assigned to any GTDB species representative. Overall, 11,856 smBGCs in eight categories were obtained from 7,007 qualified MAGs, and 10,899 smBGCs were identified as putative novel smBGCs. We anticipate that these datasets will accelerate research in the field of AMD bioremediation, aid in the discovery of novel secondary metabolites, and facilitate investigation into gene functions, metabolic pathways, and CNPS cycles in AMD.

Effects of trichothecene production by Trichoderma arundinaceum isolates from bean-field soils on the defense response, growth and development of bean plants (Phaseolus vulgaris).

The trichothecene toxin-producing fungus Trichoderma arundinaceum has potential as a biological control agent. However, most biocontrol studies have focused only on one strain, IBT 40837. In the current study, three Trichoderma isolates recovered from bean-field soils produced the trichothecene harzianum A (HA) and trichodermol, the latter being an intermediate in the HA biosynthesis. Based on phylogenetic analysis, the three isolates were assigned to the species T. arundinaceum. Their genome sequences had a high degree of similarity to the reference IBT 40837 strain, in terms of total genome size, number of predicted genes, and diversity of putative secondary metabolite biosynthetic gene clusters. HA production by these bean-field isolates conferred significant in vitro antifungal activity against Rhizoctonia solani and Sclerotinia sclerotiorum, which are some of the most important bean pathogens. Furthermore, the bean-field isolates stimulated germination of bean seeds and subsequent growth of above ground parts of the bean plant. Transcriptomic analysis of bean plants inoculated with these T. arundinaceum bean-field soil isolates indicated that HA production significantly affected expression of plant defense-related genes; this effect was particularly significant in the expression of chitinase-encoding genes. Together, these results indicate that Trichoderma species producing non-phytotoxic trichothecenes can induce defenses in plants without negatively affecting germination and development.

Genomic Comparative Analysis of Cordyceps pseudotenuipes with Other Species from Cordyceps.

The whole genome of Cordyceps pseudotenuipes was sequenced, annotated, and compared with three related species to characterize the genome. The antibiotics and Secondary Metabolites Analysis Shell (antiSMASH) and local BLAST analysis were used to explore the secondary metabolites (SMs) and biosynthesis gene clusters (BGCs) of the genus Cordyceps. The genome-wide basic characteristics of C. pseudotenuipes, C. tenuipes, C. cicadae, and C. militaris revealed unequal genome size, with C. cicadae as the largest (34.11 Mb), followed by C. militaris (32.27 Mb). However, the total gene lengths of C. pseudotenuipes and C. tenuipes were similar (30.1 Mb and 30.06 Mb). The GC contents of C. pseudotenuipes, C. tenuipes, C. cicadae, and C. militaris genomes differed slightly (51.40% to 54.11%). AntiSMASH and local BLAST analysis showed that C. pseudotenuipes, C. tenuipes, C. cicadae, and C. militaris had 31, 28, 31, and 29 putative SM BGCs, respectively. The SM BGCs contained different quantities of polyketide synthetase (PKS), nonribosomal peptide synthetase (NRPS), terpene, hybrid PKS + NRPS, and hybrid NRPS + Other. Moreover, C. pseudotenuipes, C. tenuipes, C. cicadae, and C. militaris had BGCs for the synthesis of dimethylcoprogen. C. pseudotenuipes, C. tenuipes, and C. cicadae had BGCs for the synthesis of leucinostatin A/B, neosartorin, dimethylcoprogen, wortmanamide A/B, and beauvericin. In addition, the SM BGCs unique to C. pseudotenuipes were clavaric acid, communesin, and deoxynivalenol. Synteny analysis indicated that the scaffolds where the SM BGC was located were divided into more than 70 collinear blocks, and there might be rearrangements. Altogether, these findings improved our understanding of the molecular biology of the genus Cordyceps and will facilitate the discovery of new biologically active SMs from the genus Cordyceps using heterologous expression and gene knockdown methods.