Taxonomic identification and antagonistic activity of Streptomyces luomodiensis sp. nov. against phytopathogenic fungi.

Banana wilt caused by Fusarium oxysporum f. sp. cubense tropical race 4 (Foc TR4) is a devastating fungal disease. Biocontrol strategies hold immense potential for inhibiting the spread of Foc TR4. Here, 30 actinobacteria were isolated from soils and screened for their antagonistic activity against Foc TR4. Strain SCA4-21T was selected due to its strongest antagonistic activity against Foc TR4. Strain SCA4-21T also exhibited strong antagonistic activity against the other eight phytopathogenic fungi. The strain was identified as the genus Streptomyces according to its physiological, biochemical, and phenotypic characteristics. The phylogenetic trees of 16S rRNA sequences demonstrated that strain SCA4-21T formed a subclade with S. iranensis HM 35T and/or S. rapamycinicus NRRL B-5491T with low bootstrap values. Considering that 16S rRNAs did not provide sufficient resolution for species-level identification, the whole genome of strain SCA4-21T was sequenced. Multilocus sequence analysis (MLSA) based on five housekeeping gene alleles (atpD, gyrB, recA, rpoB, and trpB) revealed that strain SCA4-21T clustered into S. hygroscopicus subsp. hygroscopicus NBRC 13472T with 100% of bootstrap value. The analysis of the genome-based phylogeny also approved the results. Average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) were 91.26 and 44.30%, respectively, with values below the respective species level threshold of 95 and 70%. Hence, strain SCA 4-21T represented a novel species within the genus Streptomyces, named Streptomyces luomodiensis sp. nov. The type strain is SCA4-21T (=GDMCC4.340T = JCM36555T). By the CAZymes analysis, 348 carbohydrate-active enzymes (CAZymes) were detected, including 15 chitinases and eight ß-1,3-glucanases. The fermentation broth of strain SCA4-21T, exhibiting strong antagonistic activity against Foc TR4, demonstrated high activities of chitinase and ß-1,3-glucanase, which might be involved in antifungal activity. Our results showed an innovative potential biocontrol agent for managing plant fungal diseases, specifically banana fusarium wilt.

Biocontrol mechanism of Bacillus siamensis sp. QN2MO-1 against tomato fusarium wilt disease during fruit postharvest and planting.

Tomato fusarium wilt caused by Fusarium oxysporum f. sp. lycopersici (Fol) is a highly destructive disease, resulting in severe economic losses of global tomato production annually. An eco-friendly alternative to chemical fungicide using biological control agents (BCAs) is urgently needed. Here, Bacillus siamensis QN2MO-1 was isolated from Noli fruit and had a strong antagonistic activity against Fol in vitro and in vivo. Strain QN2MO-1 also exhibited a broad-spectrum antifungal activity against the selected 14 phytopathogenic fungi. The crude protein produced by strain QN2MO-1 could inhibit the spore germination of Fol and destroy the spore structure. It was closely related with the generation of chitinase and ß-1,3-glucanase secreted by strain QN2MO-1. In a pot experiment, the application of B. siamensis QN2MO-1 effectively alleviated the yellowing and wilting symptoms of tomato plants. The disease index and incidence rate were decreased by 72.72% and 80.96%, respectively. The rhizospheric soil in tomato plants owed a high abundance of microbial community. Moreover, strain QN2MO-1 also enhanced the plant growth and improved the fruit quality of tomato. Therefore, B. siamensis QN2MO-1 will be explored as a potential biocontrol agent and biofertilizer.

A newly isolated Trichoderma Parareesei N4-3 exhibiting a biocontrol potential for banana fusarium wilt by Hyperparasitism.

Banana Fusarium wilt caused by Fusarium oxysporum f. sp. cubense tropical race4 (Foc TR4) is one of the most destructive soil-borne fungal diseases and currently threatens banana production around the world. Until now, there is lack of an effective method to control banana Fusarium wilt. Therefore, it is urgent to find an effective and eco-friendly strategy against the fungal disease. In this study, a strain of Trichoderma sp. N4-3 was isolated newly from the rhizosphere soil of banana plants. The isolate was identified as Trichoderma parareesei through analysis of TEF1 and RPB2 genes as well as morphological characterization. In vitro antagonistic assay demonstrated that strain N4-3 had a broad-spectrum antifungal activity against ten selected phytopathogenic fungi. Especially, it demonstrated a strong antifungal activity against Foc TR4. The results of the dual culture assay indicated that strain N4-3 could grow rapidly during the pre-growth period, occupy the growth space, and secrete a series of cell wall-degrading enzymes upon interaction with Foc TR4. These enzymes contributed to the mycelial and spore destruction of the pathogenic fungus by hyperparasitism. Additionally, the sequenced genome proved that strain N4-3 contained 21 genes encoding chitinase and 26 genes encoding ß-1,3-glucanase. The electron microscopy results showed that theses cell wall-degrading enzymes disrupted the mycelial, spore, and cell ultrastructure of Foc TR4. A pot experiment revealed that addition of strain N4-3 significantly reduced the amount of Foc TR4 in the rhizosphere soil of bananas at 60 days post inoculation. The disease index was decreased by 45.00% and the fresh weight was increased by 63.74% in comparison to the control. Hence, Trichoderma parareesei N4-3 will be a promising biological control agents for the management of plant fungal diseases.

Antimicrobial mechanisms and secondary metabolite profiles of Streptomyces hygroscopicus subsp. hygroscopicus 5-4 against banana fusarium wilt disease using metabolomics.

Fusarium wilt of bananas (FWB) is seriously affecting the sustainable development of the banana industry and is caused by the devastating soil-borne fungus Fusarium oxysporum f. sp. cubense tropical race 4 (Foc TR4). Biological control is a promising strategy for controlling Fusarium wilt in bananas. We previously identified Streptomyces hygroscopicus subsp. hygroscopicus 5-4 with strong antifungal activity against the FWB. The most possible antimicrobial mechanism of strain 5-4 was explored using the metabolomics approach, light microscopy imaging, and transmission electron microscopy (TEM). The membrane integrity and ultrastructure of Foc TR4 was damaged after extract treatment, which was supported by the degradation of mycelium, soluble protein content, extracellular reducing sugar content, NADH oxidase activity, malondialdehyde content, mitochondrial membrane potential, and mitochondrial respiratory chain complex enzyme activity. The extracts of strain 5-4 cultivated at different times were characterized by a liquid chromatography-mass spectrometer (LC-MS). 647 known metabolites were detected in the extracts of strains 5-4. Hygromycin B, gluten exorphin B4, torvoside G, (z)-8-tetradecenal, piperitoside, sarmentosin, pubescenol, and other compounds were the main differential metabolites on fermentation culture for 7 days. Compared with strain 5-4 extracts, hygromycin B inhibited the mycelial growth of Foc TR4, and the EC50 concentration was 7.4 µg/mL. These results showed that strain 5-4 could destroy the cell membrane of Foc TR4 to inhibit the mycelial growth, and hygromycin B may be the key antimicrobial active metabolite. Streptomyces hygroscopicus subsp. hygroscopicus 5-4 might be a promising candidate strain to control the FWB and provide a scientific basis for the practical application of hygromycin B as a biological control agent.

Discovery of Niphimycin C from Streptomyces yongxingensis sp. nov. as a Promising Agrochemical Fungicide for Controlling Banana Fusarium Wilt by Destroying the Mitochondrial Structure and Function.

Banana Fusarium wilt caused by Fusarium oxysporum f. sp. cubense (Foc) is the most destructive soil-borne fungal disease. Tropical race 4 (Foc TR4), one of the strains of Foc, can infect many commercial cultivars, which represents a threat to global banana production. Currently, there are hardly any effective chemical fungicides to control the disease. To search for natural product-based fungicides for controlling banana Fusarium wilt, we identified a novel strain Streptomyces yongxingensis sp. nov. (JCM 34965) from a marine soft coral, from which a bioactive compound, niphimycin C, was isolated using an activity-guided method. Niphimycin C exhibited a strong antifungal activity against Foc TR4 with a value of 1.20 µg/mL for EC50 and obviously inhibited the mycelial growth and spore germination of Foc TR4. It caused the functional loss of mitochondria and the disorder of metabolism of Foc TR4 cells. Further study showed that niphimycin C reduced key enzyme activities of the tricarboxylic acid (TCA) cycle and the electron transport chain (ETC). It displayed broad-spectrum antifungal activities against the selected 12 phytopathogenic fungi. In pot experiments, niphimycin C reduced the disease indexes in banana plantlets and inhibited the infection of Foc TR4 in roots. Hence, niphimycin C could be a promising agrochemical fungicide for the management of fungal diseases.

Potential Biological Control of Endophytic Streptomyces sp. 5-4 Against Fusarium Wilt of Banana Caused by Fusarium oxysporum f. sp. cubense Tropical Race 4.

Fusarium wilt of banana caused by Fusarium oxysporum f. sp. cubense tropical race 4 (Foc TR4) is one of the most disastrous fungal diseases. Biological control is a promising strategy for controlling Fusarium wilt of banana. To explore endophytic actinomycetes as biocontrol resources against Foc TR4, antagonistic strains were isolated from different tissues of medicinal plants. Here, a total of 144 actinomycetes were isolated and belonged to Nonomuraea, Kitasatospora, and Streptomyces. Forty-three isolates exhibited antifungal activities against Foc TR4. The strain labeled with 5-4 isolated from roots of Piper austrosinense had a broad-spectrum antifungal activity by the production of chitinase and ß-1,3-glucanase and was identified as Streptomyces hygroscopicus subsp. hygroscopicus 5-4. Furthermore, disease index of banana wilt was significantly reduced by application of strain 5-4 in comparison with application of Foc TR4 alone. Exogenous application of strain 5-4 increased the expression levels of defense genes such as (PAL), peroxidase (POD), pathogenesis-related protein 1 (PR-1), hydrolytic enzymes (ß-1,3-glucanase), lysin motif receptor kinase 1 (LYK-1), and mitogen-activated protein kinase 1 (MPK-1). The antifungal mechanism assay demonstrated that extracts of strain 5-4 inhibited spore gemination and hyphal growth of Foc TR4, and caused abnormally swollen, deformity, and rupture of Foc TR4 hypha. Thus, S. hygroscopicus subsp. hygroscopicus 5-4 could be used as a potential biological agent for controlling Fusarium wilt of banana.

Biocontrol potential and antifungal mechanism of a novel Streptomyces sichuanensis against Fusarium oxysporum f. sp. cubense tropical race 4 in vitro and in vivo.

Most commercial banana cultivars are highly susceptible to Fusarium wilt caused by soilborne fungus Fusarium oxysporum f. sp. cubense (Foc), especially tropical race 4 (TR4). Biological control using antagonistic microorganism has been considered as an alternative method to fungicide. Our previous study showed that Streptomyces sp. SCA3-4 T had a broad-spectrum antifungal activity from the rhizosphere soil of Opuntia stricta in a dry hot valley. Here, the sequenced genome of strain SCA3-4 T contained 6614 predicted genes with 72.38% of G + C content. A polymorphic tree was constructed using the multilocus sequence analysis (MLSA) of five house-keeping gene alleles (atpD, gyrB, recA, rpoB, and trpB). Strain SCA3-4 T formed a distinct clade with Streptomyces mobaraensis NBRC 13819 T with 71% of bootstrap. Average nucleotide identity (ANI) values between genomes of strain SCA3-4 T and S. mobaraensis NBRC 13819 T was 85.83% below 95-96% of the novel species threshold, and named after Streptomyces sichuanensis sp. nov. The type strain is SCA3-4 T (= GDMCC 4.214 T = JCM 34964 T). Genomic analysis revealed that strain SCA3-4 T contained 36 known biosynthetic gene clusters of secondary metabolites. Antifungal activity of strain SCA3-4 T was closely associated with the production of siderophore and its extracts induced the apoptosis of Foc TR4 cells. A total of 12 potential antifungal metabolites including terpenoids, esters, acid, macrolides etc. were obtained by the gas chromatography-mass spectrometry (GC-MS). Greenhouse experiment indicated that strain SCA3-4 T could significantly inhibit infection of Foc TR4 in the roots and corms of banana seedlings and reduce disease index. Therefore, strain SCA3-4 T is an important microbial resource for exploring novel natural compounds and developing biopesticides to manage Foc TR4. KEY POINTS: • Strain SCA3-4 T was identified as a novel species of Streptomyces. • Siderophore participates in the antifungal regulation. • Secondary metabolites of strain SCA3-4 T improves the plant resistance to Foc TR4.

Identification and Antifungal Mechanism of a Novel Actinobacterium Streptomyces huiliensis sp. nov. Against Fusarium oxysporum f. sp. cubense Tropical Race 4 of Banana.

Banana is an important fruit crop. Fusarium wilt caused by Fusarium oxysporum f. sp. cubense tropical race 4 (Foc TR4) seriously threatens the global banana industry. It is difficult to control the disease spread using chemical measures. In addition, commercial resistant cultivars are also lacking. Biological control is considered as a promising strategy using antagonistic microbes. Actinomycetes, especially Streptomyces, are potential sources of producing novel bioactive secondary metabolites. Here, strain SCA2-4 T with strong antifungal activity against Foc TR4 was isolated from the rhizospheric soil of Opuntia stricta in a dry hot valley. The morphological, physiological and chemotaxonomic characteristics of the strain were consistent with the genus Streptomyces. Based on the homology alignment and phylogenetic trees of 16S rRNA gene, the taxonomic status of strain SCA2-4 T exhibited a paradoxical result and low bootstrap value using different algorithms in the MEGA software. It prompted us to further discriminate this strain from the closely related species by the multilocus sequence analysis (MLSA) using five house-keeping gene alleles (atpD, gyrB, recA, rpoB, and trpB). The MLSA trees calculated by three algorithms demonstrated that strain SCA2-4 T formed a distinct clade with Streptomyces mobaraensis NBRC 13819 T . The MLSA distance was above 0.007 of the species cut-off. Average nucleotide identity (ANI) values between strain SCA2-4 T genome and two standard strain genomes were below 95-96% of the novel species threshold. Strain SCA2-4 T was assigned to a novel species of the genus Streptomyces and named as Streptomyces huiliensis sp. nov. The sequenced complete genome of SCA2-4 T encoded 51 putative biosynthetic gene clusters of secondary metabolites. Genome alignment revealed that ten gene clusters were involved in the biosynthesis of antimicrobial metabolites. It was supported that strain SCA2-4 T showed strong antifungal activities against the pathogens of banana fungal diseases. Extracts abstracted from the culture filtrate of strain SCA2-4 T seriously destroyed cell structure of Foc TR4 and inhibited mycelial growth and spore germination. These results implied that strain SCA2-4 T could be a promising candidate for biological control of banana Fusarium wilt.

Biocontrol Ability and Mechanism of a Broad-Spectrum Antifungal Strain Bacillus safensis sp. QN1NO-4 Against Strawberry Anthracnose Caused by Colletotrichum fragariae.

Strawberry is a very popular fruit with a special taste, color, and nutritional value. Anthracnose caused by Colletotrichum fragariae severely limits fruit shelf life during post-harvest storage. Use of traditional chemical fungicides leads to serious environment pollution and threatens food safety. Biocontrol is considered as a promising strategy to manage the post-harvest fruit diseases. Here, strain QN1NO-4 isolated from noni (Morinda citrifolia L.) fruit exhibited a high antifungal activity against C. fragariae. Based on its physicochemical profiles and phylogenetic tree of the 16S rRNA sequence, strain QN1NO-4 belonged to the genus Bacillus. The average nucleotide identity (ANI) calculated by comparing two standard strain genomes was below 95-96%, suggesting that the strain might be a novel species of the genus Bacillus and named as Bacillus safensis sp. QN1NO-4. Its extract effectively reduced the incidence of strawberry anthracnose of harvested fruit. Fruit weight and TSS contents were also maintained significantly. The antifungal mechanism assays indicated that the extract of the test antagonist inhibited mycelial growth and spore germination of C. fragariae in vitro. Cells of strain QN1NO-4 demonstrated the cytoplasmic heterogeneity, disappeared organelles, and ruptured ultrastructure. Notably, the strain extract also had a broad-spectrum antifungal activity. Compared with the whole genome of strain QN1NO-4, several functional gene clusters involved in the biosynthesis of active secondary metabolites were observed. Fifteen compounds were identified by gas chromatography-mass spectrometry (GC-MS). Hence, the fruit endophyte B. safensis sp. QN1NO-4 is a potential bio-agent identified for the management of post-harvest disease of strawberry fruit.

A Novel Antifungal Actinomycete Streptomyces sp. Strain H3-2 Effectively Controls Banana Fusarium Wilt.

Banana Fusarium wilt disease caused by Fusarium oxyspoum f. sp. cubense (Foc) seriously threatens the banana industry. Foc tropical race 4 (Foc TR4) can infect almost all banana cultivars. Compared with traditional physical and chemical practices, biocontrol strategy using beneficial microbes is considered as an environmentally sound option to manage fungal disease. In this study, a strain, H3-2, isolated from a non-infected banana orchard, exhibited high antifungal activity against Foc TR4. According to its morphological, physiological, and biochemical characteristics, the strain H3-2 was identified as Streptomyces sp. and convinced by the polymorphic phylogenic analysis of 16S rRNA sequences. Extracts of the strain H3-2 suppressed the growth and spore germination of Foc TR4 in vitro by destroying cell membrane integrity and mycelial ultrastructure. Notably, the strain and its extracts showed broad-spectrum antifungal activity against the selected seven fungal phytopathogens. Fourteen chemical compounds in the extracts were identified by gas chromatography-mass spectrometer (GC-MS), primarily phenolic compounds. Additional pot inoculation experiment demonstrated that the fermentation broth of the strain H3-2 promoted the growth of banana seedlings by efficiently inhibiting the spread of banana Fusarium wilt disease. This study demonstrated the potential application of the novel Streptomyces sp. H3-2 for the management of banana Fusarium wilt.

Acetobacter orientalis XJC-C with a high lignocellulosic biomass-degrading ability improves significantly composting efficiency of banana residues by increasing metabolic activity and functional diversity of bacterial community.

Banana residues are an important energy resource after fruit harvesting. The optionally dumping and burning causes severely environmental problems. Traditional compost efficiency was limited by lignocellulosic composition of banana residues. Inoculation with cellulase-producing microbes provides an efficient strategy for improving degradation of lignocellulosic materials. In our study, a newly isolated cellulolytic bacterium Acetobacter orientalis XJC-C with a salt and high temperature resistance was identified from a marine soft coral. By contrast, the strain can biodegrade different lignocellulosic agricultural residues, especially banana straw. The highest cellulolytic and ligninolytic enzyme activities were detected during composting at 40 days. Compared with the negative and positive control groups, the lignin degradation rate reached 76.24% in the A. orientalis XJC-C group, increased by 47.08% and 21.85%, respectively. Moreover, the strain improved significantly the metabolic activity and functional diversity of bacterial community. Hence, A. orientalis XJC-C will be a promising candidate for degrading lignocellulosic agricultural residues.

Newly Isolated Streptomyces sp. JBS5-6 as a Potential Biocontrol Agent to Control Banana Fusarium Wilt: Genome Sequencing and Secondary Metabolite Cluster Profiles.

Banana is a key staple food and fruit in countries all over the world. However, the development of the global banana industry is seriously threatened by Fusarium wilt disease, which is caused by Fusarium oxysporum f. sp. cubense (Foc). In particular, Foc tropical race 4 (Foc TR4) could infect more than 80% of global banana and plantain crops. Until now, there were no commercial chemicals or resistant cultivars available to control the disease. Biological control using actinomycetes is considered a promising strategy. In this study, 88 actinomycetes were isolated from a banana orchard without symptoms of Fusarium wilt disease for more than 10 years. An actinobacterial strain labeled as JBS5-6 has exhibited strong antifungal activities against Foc TR4 and other selected 10 phytopathogenic fungi. Based on phenotypic and biochemical traits as well as complete genome analysis, strain JBS5-6 was assigned to Streptomyces violaceusniger. Extracts of the strain inhibited the mycelial growth and spore germination of Foc TR4 by destroying membrane integrity and the ultrastructure of cells. The complete genome of strain JBS5-6 was sequenced and revealed a number of key function gene clusters that contribute to the biosynthesis of active secondary metabolites. Sixteen chemical compounds were further identified by gas chromatography-mass spectrometry (GC-MS). 5-hydroxymethyl-2-furancarboxaldehyde was one of the dominant components in strain JBS5-6 extracts. Moreover, fermentation broth of strain JBS5-6 significantly reduced the disease index of banana seedlings by inhibiting the infection of Foc TR4 in a pot experiment. Hence, strain JBS5-6 is a potential biocontrol agent for the management of disease and the exploitation of biofertilizer.

A Newly Isolated Streptomyces sp. YYS-7 With a Broad-Spectrum Antifungal Activity Improves the Banana Plant Resistance to Fusarium oxysporum f. sp. cubense Tropical Race 4.

Fusarium wilt of banana caused by Fusarium oxysporum f. sp. cubense (Foc) is one of the most destructive diseases, severely limiting the development of banana industry. Especially, Foc tropical race 4 (Foc TR4) can infect and destroy almost all banana cultivars. Until now, there is still a lack of an effective method for controlling fusarium wilt. A biocontrol strategy using Actinobacteria is considered as a promising method for management of disease and pest. In this study, 229 Actinobacteria were isolated from rhizosphere soil samples of a primitive ecological mountain. An actinobacterium strain marked with YYS-7 exhibited a high antifungal activity against Foc TR4. Combining the physiological and biochemical characteristics as well as alignment of the 16S rRNA sequence, the strain YYS-7 was assigned to Streptomyces sp. The crude extracts of Streptomyces sp. YYS-7 obviously inhibited the mycelial growth of Foc TR4. The cell integrity and ultrastructure were seriously destroyed. In addition, Streptomyces sp. YYS-7 and crude extracts also showed a broad-spectrum antifungal activity against the selected seven phytopathogenic fungi. A gas chromatography-mass spectrometry (GC-MS) was used to predict the antifungal metabolites. A total of eleven different compounds were identified, including phenolic compounds, hydrocarbons, esters and acids. In the pot experiment, the crude extracts can significantly improve the banana plant's resistance to Foc TR4. Hence, Streptomyces sp. YYS-7 will be a potential biocontrol agent for the biofertilizer exploitation and the discovery of new bioactive substances.

Biodegradation of lignocellulosic agricultural residues by a newly isolated Fictibacillus sp. YS-26 improving carbon metabolic properties and functional diversity of the rhizosphere microbial community.

A new isolated cellulolytic bacterium from a soft coral was named as Fictibacillus sp YS-26 based on the morphologic and molecular characteristics. It can degrade different lignocellulosic agricultural residues by producing cellulolytic enzymes, α-amylase, protease, pectinase and xylanase. Especially, Fictibacillus sp. YS-26 exhibited the highest cellulolytic activities in the soybean meal medium. By contrast, the fermentation broth of Fictibacillus sp. YS-26 significantly enhanced utilization efficiency of carboxylic acids and polymers by soil microorganisms as well as the microbial metabolism function and community diversity in rhizosphere soil of banana plantlets. The fermentation broth also improved soil characters and increased the growth of banana plantlets. We found that soil total nitrogen and electrical conductivity had a positive relationship with the increase of microbial diversity. Hence, Fictibacillus sp. YS-26 will be a promising candidate for biodegradating lignocellulosic biomass and improving the soil microbial diversity.

Anti-Foc RT4 Activity of a Newly Isolated Streptomyces sp. 5-10 From a Medicinal Plant (Curculigo capitulata).

Fusarium wilt of banana caused by Fusarium oxysporum f. sp. cubense (Foc) is a disastrous soil-borne fungal disease. Foc tropical race 4 (Foc TR4) can infect almost all banana cultivars. Until now, there is a shortage of safety and effective control methods and commercial banana cultivars with a resistance against Foc TR4. Biocontrol using environmentally friendly microbes is a promising strategy for the management of Foc TR4. Here, a strain 5-10, newly isolated from a medicinal plant (Curculigo capitulata), exhibited a high antifungal activity against Foc TR4. Combing the morphological characteristics and molecular identification, strain 5-10 was classified as a Streptomyces genus. The sequenced genome revealed that more than 39 gene clusters were involved in the biosynthesis of secondary metabolites. Some multidrug resistance gene clusters were also identified such as mdtD, vatB, and vgaE. To improve the anti-Foc TR4 activity of the strain 5-10 extracts, an optimization method of fermentation broth was established. Antifungal activity increased by 72.13% under the fermentation system containing 2.86 g/L of NaCl and 11.57% of inoculation amount. After being treated with the strain 5-10 extracts, the Foc TR4 hyphae shrinked, deformed, and ruptured. The membrane integrity and cell ultrastructure incurred irreversible damage. Streptomyces sp. 5-10 extracts play a fungicidal role in Foc TR4. Hence, Streptomyces sp. 5-10 will be a potential biocontrol agent to manage fungal diseases by exploring the microbial fertilizer.

Involvement of abscisic acid-responsive element-binding factors in cassava (Manihot esculenta) dehydration stress response.

Cassava (Manihot esculenta) is a major staple food, animal feed and energy crop in the tropics and subtropics. It is one of the most drought-tolerant crops, however, the mechanisms of cassava drought tolerance remain unclear. Abscisic acid (ABA)-responsive element (ABRE)-binding factors (ABFs) are transcription factors that regulate expression of target genes involved in plant tolerance to drought, high salinity, and osmotic stress by binding ABRE cis-elements in the promoter regions of these genes. However, there is little information about ABF genes in cassava. A comprehensive analysis of Manihot esculenta ABFs (MeABFs) described the phylogeny, genome location, cis-acting elements, expression profiles, and regulatory relationship between these factors and Manihot esculenta betaine aldehyde dehydrogenase genes (MeBADHs). Here we conducted genome-wide searches and subsequent molecular cloning to identify seven MeABFs that are distributed unevenly across six chromosomes in cassava. These MeABFs can be clustered into three groups according to their phylogenetic relationships to their Arabidopsis (Arabidopsis thaliana) counterparts. Analysis of the 5'-upstream region of MeABFs revealed putative cis-acting elements related to hormone signaling, stress, light, and circadian clock. MeABF expression profiles displayed clear differences among leaf, stem, root, and tuberous root tissues under non-stress and drought, osmotic, or salt stress conditions. Drought stress in cassava leaves and roots, osmotic stress in tuberous roots, and salt stress in stems induced expression of the highest number of MeABFs showing significantly elevated expression. The glycine betaine (GB) content of cassava leaves also was elevated after drought, osmotic, or salt stress treatments. BADH1 is involved in GB synthesis. We show that MeBADH1 promoter sequences contained ABREs and that MeBADH1 expression correlated with MeABF expression profiles in cassava leaves after the three stress treatments. Taken together, these results suggest that in response to various dehydration stresses, MeABFs in cassava may activate transcriptional expression of MeBADH1 by binding the MeBADH1 promoter that in turn promotes GB biosynthesis and accumulation via an increase in MeBADH1 gene expression levels and MeBADH1 enzymatic activity. These responses protect cells against dehydration stresses by preserving an osmotic balance that enhances cassava tolerance to dehydration stresses.

Taxonomy and Broad-Spectrum Antifungal Activity of Streptomyces sp. SCA3-4 Isolated From Rhizosphere Soil of Opuntia stricta.

Actinobacteria are important producers of bioactive compounds. Extreme ecosystems cause evolution of novel secondary metabolic pathways of Actinobacteria and increase the possible discovery of new biological functions of bioactive compounds. Here, we isolated 65 Actinobacteria from rhizosphere soil samples of Opuntia stricta. An Actinobacteria strain (named SCA3-4) was screened against Fusarium oxysporum f. sp. cubense Tropical Race 4 (Foc TR4, ATCC 76255). The strain produced pink-white aerial mycelia and brown substrate mycelium on Gause No. 1 agar. Biverticillate chains of cylindrical spores were observed by scanning electron microscopy (SEM). Based on alignment of 16S rRNA sequences, a constructed phylogenetic tree showed that strain SCA3-4 shared a 99.54% similarity with Streptomyces lilacinus NRRL B-1968T. The morphological, biochemical, physiological, and molecular characteristics further indicated that strain SCA3-4 belongs to the Streptomyces sp. It can grow well on medium with the following antibiotics chloramphenicol, streptomycin, penicillin-G, gentamicin, erythromycin, nystatin or neomycin sulfate. The polymerase chain reaction (PCR) amplification of types I and II polyketide synthase genes (PKS-I and PKS-II) suggested its bioactive potential. Under treatment with 100 µg/ml of ethyl acetate extracts isolated from Streptomyces sp. SCA3-4, growth of Foc TR4 was inhibited and cell membrane was destroyed. Crude extracts also showed a broad-spectrum antifungal activity against 13 phytopathogenic fungi including Foc TR4 and displayed the lowest minimum inhibitory concentration (MIC) (0.781 µg/ml) against Colletotrichum fragariae (ATCC 58718). A total of 21 different compounds identified by gas chromatography-mass spectrometry (GC-MS) were composed of phenolic compound, pyrrolizidine, hydrocarbons, esters, and acids. Besides the known active compounds, Streptomyces sp. SCA3-4 possesses antimicrobial or other biological activities. Further attention will be paid on other compounds with no functional annotation, aiming at the discovery of new bioactive substances.

Deciphering microbial diversity associated with Fusarium wilt-diseased and disease-free banana rhizosphere soil.

BACKGROUND: Fusarium wilt of banana (Musa spp.) caused by the fungal pathogen Fusarium oxysporum f. sp. cubense (Foc) is a typical soilborne disease, that severely devastates the banana industry worldwide, and soil microbial diversity is closely related to the spread of Fusarium wilt. To understand the relationship between microbial species and Fusarium wilt, it is important to understand the microbial diversity of the Fusarium wilt-diseased and disease-free soils from banana fields. RESULTS: Based on sequencing analysis of the bacterial 16S rRNA genes and fungal internal transcribed spacer (ITS) sequences, Foc abundance, fungal or bacterial richness and diversity were higher in the diseased soils than in the disease-free soils. Although Ascomycota and Zygomycota were the most abundant fungi phyla in all soil samples, Ascomycota abundance was significantly reduced in the disease-free soils. Mortierella (36.64%) was predominant in the disease-free soils. Regarding bacterial phyla, Proteobacteria, Acidobacteria, Chloroflexi, Firmicutes, Actinobacteria, Gemmatimonadetes, Bacteroidetes, Nitrospirae, Verrucomicrobia and Planctomycetes were dominant phyla in all soil samples. In particular, Firmicutes contributed 16.20% of the total abundance of disease-free soils. At the bacterial genus level, Bacillus, Lactococcus and Pseudomonas were abundant in disease-free soils with abundances of 8.20, 5.81 and 2.71%, respectively; lower abundances, of 4.12, 2.35 and 1.36%, respectively, were found in diseased soils. The distribution characteristics of fungal and bacterial genera may contribute to the abundance decrease of Foc in the disease-free soils. CONCLUSION: Unique distributions of bacteria and fungi were observed in the diseased and disease-free soil samples from banana fields. These specific genera are useful for constructing a healthy microbial community structure of soil.

Genome-wide characterization of a SRO gene family involved in response to biotic and abiotic stresses in banana (Musa spp.).

BACKGROUND: Banana (Musa spp.) is one of the world's most important fruits and its production is largely limited by diverse stress conditions. SROs (SIMILAR TO RCD-ONE) have important functions in abiotic stress resistance and development of plants. They contain a catalytic core of the poly(ADP-ribose) polymerase (PARP) domain and a C-terminal RST (RCD-SRO-TAF4) domain. In addition, partial SROs also include an N-terminal WWE domain. Although a few of SROs have been characterized in some model plants, little is known about their functions in banana, especially in response to biotic stress. RESULTS: Six MaSRO genes in banana genome were identified using the PARP and RST models as a query. Phylogenetic analysis showed that 77 SROs from 15 species were divided into two structurally distinct groups. The SROs in the group I possessed three central regions of the WWE, PARP and RST domains. The WWE domain was lacking in the group II SROs. In the selected monocots, only MaSROs of banana were present in the group II. Most of MaSROs expressed in more than one banana tissue. The stress- and hormone-related cis-regulatory elements (CREs) in the promoter regions of MaSROs supported differential transcripts of MaSROs in banana roots treated with abiotic and biotic stresses. Moreover, expression profiles of MaSROs in the group I were clearly distinct with those observed in the group II after hormone treatment. Notably, the expression of MaSRO4 was significantly upregulated by the multiple stresses and hormones. The MaSRO4 protein could directly interact with MaNAC6 and MaMYB4, and the PARP domain was required for the protein-protein interaction. CONCLUSIONS: Six MaSROs in banana genome were divided into two main groups based on the characteristics of conserved domains. Comprehensive expression analysis indicated that MaSROs had positive responses to biotic and abiotic stresses via a complex interaction network with hormones. MaSRO4 could interact directly with MaNAC6 and MaMYB4 through the PARP domain to regulate downstream signaling pathway.

Quantitative proteomics analysis reveals resistance differences of banana cultivar 'Brazilian' to Fusarium oxysporum f. sp. cubense races 1 and 4.

Banana Fusarium wilt, caused by Fusarium oxysporum f. sp. cubense (Foc), is one of the most devastating diseases in banana production. Foc is classified into three physiological races. However, the resistance mechanisms of banana against different Foc races are poorly understood. In this study, we performed a comparative proteomics analysis to investigate the resistance mechanisms of 'Brazilian' against Foc1 and Foc4. The proteomes of 'Brazilian' roots inoculated with Foc1 and Foc4 and mock inoculated control at 48 h were analyzed using TMT based quantitative analysis technique. A total of 7325 unique protein species were identified, of which 689, 744, and 1222 protein species were differentially accumulated in Foc1 vs. CK, Foc4 vs. CK, and Foc1 vs. Foc4, respectively. The differential accumulations of candidate protein species were further confirmed by RT-qPCR, PRM, and physiological and biochemical assays. Bioinformatics analysis revealed that the differentially abundance protein species (DAPS) related to pattern recognition receptors, plant cell wall modification, redox homeostasis, and defense responses were differentially accumulated after Foc1 and Foc4 infection, suggesting that 'Brazilian' differed in resistance to the two Foc races. Our study lay the foundation for an in-depth understanding of the interaction between bananas and Foc at the proteome level. SIGNIFICANCE: The banana fusarium wilt disease is one of the most destructive disease of banana and is caused by Fusarium oxysporum f. sp. cubense (Foc). Foc is classified into three physiological races, namely, Foc1, Foc2, and Foc4. Among these races, Foc1 and Foc4 are widely distributed in south China and significantly lose yield. Although both physiological races (Foc1 and Foc4) can invade the Cavendish banana cultivar 'Brazilian', they have significant pathogenicity differences. Unfortunately, how the resistance differences are produced between two races is still largely unclear to date. In this study, we addressed this issue by performing TMT-based comparative quantitative proteomics analysis of 'Brazilian' roots after inoculation with Foc1 and Foc4 as well as sterile water as the control. We revealed that the series of protein species associated with pattern recognition receptors, plant cell wall modification, redox homeostasis, pathogenesis, phytohormones and signal transduction, plant secondary metabolites and programmed cell death etc. were involved in the response to Foc infection. Notably, the potential role of lipid signaling in banana defense against Foc are not reported previously but rather unveiled for the first time in this study. The current study represents the most extensive analysis of the protein profile of 'Brazilian' in response to Foc inoculation and includes for the first time the results from comparison quantitative proteomics analysis between plants inoculated with a pathogenic strain Foc4 and a nonpathogenic strain Foc1 of 'Brazilian', which will lay the foundation for an in-depth understanding of the interaction between bananas and Foc at the proteome level.