Screening and Analysis of Antifungal Strains Bacillus subtilis JF-4 and B. amylum JF-5 for the Biological Control of Fusarium Wilt of Banana.

PURPOSE: This study aimed to identify the antagonistic bacteria from the rhizosphere of healthy bananas that can effectively suppress the Fusarium wilt of banana, and to further investigate the inhibitory mechanism. METHOD: The primary and secondary screening techniques were implemented using the double-plate and fermentation antagonism methods. The strain was identified based on physiological and biochemical tests, 16S rRNA gene sequencing, and specific gene amplification. The effects of crude extract on the protein content, lipid peroxidation, and pectinase activity of mycelia were determined from the identified isolates. RESULTS: Two antagonistic bacteria, JF-4 and JF-5, were screened and initially identified as Bacillus subtilis (GenBank: OR125631) and B. amylum (GenBank: OR125632). The greenhouse experiment showed that the biological control efficiency of the two antagonists against the Fusarium wilt of banana was 48.3% and 40.3%, respectively. The catalase content produced by lipid peroxidation increased significantly after treatment with the crude extracts of JF-4 and JF-5 at concentrations of 0.69 µmol/L and 0.59 µmol/L, respectively. The protein and ergosterol content and pectinase activity decreased significantly. The two antagonistic bacteria might inhibit the growth of pathogens by enhancing lipid peroxidation and decreasing the synthesis of cell metabolites. Twenty compounds were identified by gas chromatography-mass spectrometry (GC-MS). B. subtilis JF-4 was further sequenced and assembled to obtain a complete circular chromosome genome of 681,804,824 bp. The genome consisted of a 4,310,825-bp-long scaffold. CONCLUSION: The findings of this study may help elucidate the mechanism behind this biocontrol isolate.

Analyses of key gene networks controlling carotenoid metabolism in Xiangfen 1 banana.

BACKGROUND: Banana fruits are rich in various high-value metabolites and play a key role in the human diet. Of these components, carotenoids have attracted considerable attention due to their physiological role and human health care functions. However, the accumulation patterns of carotenoids and genome-wide analysis of gene expression during banana fruit development have not been comprehensively evaluated. RESULTS: In the present study, an integrative analysis of metabolites and transcriptome profiles in banana fruit with three different development stages was performed. A total of 11 carotenoid compounds were identified, and most of these compounds showed markedly higher abundances in mature green and/or mature fruit than in young fruit. Results were linked to the high expression of carotenoid synthesis and regulatory genes in the middle and late stages of fruit development. Co-expression network analysis revealed that 79 differentially expressed transcription factor genes may be responsible for the regulation of LCYB (lycopene ß-cyclase), a key enzyme catalyzing the biosynthesis of α- and ß-carotene. CONCLUSIONS: Collectively, the study provided new insights into the understanding of dynamic changes in carotenoid content and gene expression level during banana fruit development.

Stimulation of photosynthesis and enhancement of growth and yield in Arabidopsis thaliana treated with amine-functionalized mesoporous silica nanoparticles.

Mesoporous silica nanoparticles (MSNs) of 50 nm diameter particle size with a pore size of approximately 14.7 nm were functionalized with amino groups (Am-MSNs) and the effects of exposure to these positively charged Am-MSNs on each of the life cycle stages of Arabidopsis thaliana were investigated. After growth in half strength MS medium amended with Am-MSNs (0-100 µg/mL) for 7 and 14 days, seed germination rate and seedling growth were significantly increased compared with untreated controls. The seedlings were then transferred to soil and irrigated with Am-MSNs solutions every 3 days until seed harvesting. After four weeks growth in soil, Am-MSNs treated plants showed up-regulation of chlorophyll and carotenoid synthesis-related genes, an increase in the content of photosynthetic pigments and an amplification of plant photosynthetic capacity. All these changes in plants were closely correlated with greater vegetative growth and higher seed yield. In all the experiments, 20 and 50 µg/mL of Am-MSNs were found to be more effective with respect to other treatments, while Am-MSNs at the highest level of 100 µg/mL did not result in oxidative stress or cell membrane damage in the exposed plants. To the best of our knowledge, this is the first report evaluating both physiological and molecular responses following exposure to plants of these specific Am-MSNs throughout their whole life cycle. Overall, these findings indicate that following exposure Am-MSNs play a major role in the increase in seed germination, biomass, photosynthetic pigments, photosynthetic capacity and seed yield in A. thaliana.

Nutritional component changes in Xiangfen 1 banana at different developmental stages.

Banana is an essential food resource in many tropical and subtropical countries. Metabolites in banana greatly influence its nutritional value and flavor. However, metabolic changes that occur in different developmental stages have not been comprehensively evaluated. In this study, widely targeted metabolomics based on multiple reaction monitoring was used in investigating dynamic changes in metabolites at three stages of fruit development. A total of 655 metabolites were identified in all the stages. A hierarchical cluster analysis of metabolites showed six clear expression patterns at the three developmental stages, and 69 up-regulated differential metabolites were identified in mature fruits compared with young and mature green fruits. A metabolic pathway analysis of differential metabolites showed significant enrichment of the flavonoid biosynthesis pathway and the phenylalanine, tyrosine, and tryptophan biosynthesis pathways. These results may serve as a reference for the isolation and identification of functional compounds from banana and for their sufficient utilization in the future.

Fermentation optimization and disease suppression ability of a Streptomyces ma. FS-4 from banana rhizosphere soil.

BACKGROUND: Fusarium wilt of banana is one of the most destructive diseases in banana-growing regions worldwide. Soil-borne diseases and soil microbial communities are closely related. The screening of antagonistic bacteria from soil microorganisms in areas with Fusarium wilt of banana is of great practical significance for controlling this disease. RESULTS: A strain designated FS-4 was isolated from healthy banana rhizosphere soil in an area affected by Fusarium wilt. This strain exhibited a significant antagonistic effect on the pathogen. Pot experiments revealed that the fermentation broth of strain FS-4 not only decreased the incidence of banana Fusarium wilt, but also promoted the growth of banana seedlings. The strain was identified as Streptomyces ma. by its morphological, physiological, and biochemical characteristics and 16S rRNA gene sequence analysis. The culture and fermentation conditions for this strain were optimized by single-factor and response surface experiments. The optimum culture conditions for Streptomyces ma. FS-4 were as follows: peptone 0.5%, saccharose 2.4, 0.05% K2HPO4, 0.05% MgCl2, and 0.05% NaCl at an initial pH of 7.0; 180 g at 28 °C; and inoculation size of 6% for 62 h. The diameter of bacteriostasis circle for Bacillus subtilis reached 26.7 mm. CONCLUSION: Streptomyces ma. FS-4 is an important microbial resource as a biological agent for the control of plant pathogenic fungi and can be used to promote banana growth.

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.

Transcriptome analysis of atemoya pericarp elucidates the role of polysaccharide metabolism in fruit ripening and cracking after harvest.

BACKGROUND: Mature fruit cracking during the normal season in African Pride (AP) atemoya is a major problem in postharvest storage. Our current understanding of the molecular mechanism underlying fruit cracking is limited. The aim of this study was to unravel the role starch degradation and cell wall polysaccharide metabolism in fruit ripening and cracking after harvest through transcriptome analysis. RESULTS: Transcriptome analysis of AP atemoya pericarp from cracking fruits of ethylene treatments and controls was performed. KEGG pathway analysis revealed that the starch and sucrose metabolism pathway was significantly enriched, and approximately 39 DEGs could be functionally annotated, which included starch, cellulose, pectin, and other sugar metabolism-related genes. Starch, protopectin, and soluble pectin contents among the different cracking stages after ethylene treatment and the controls were monitored. The results revealed that ethylene accelerated starch degradation, inhibited protopectin synthesis, and enhanced the soluble pectin content, compared to the control, which coincides with the phenotype of ethylene-induced fruit cracking. Key genes implicated in the starch, pectin, and cellulose degradation were further investigated using RT-qPCR analysis. The results revealed that alpha-amylase 1 (AMY1), alpha-amylase 3 (AMY3), beta-amylase 1 (BAM1), beta-amylase 3 (BAM3), beta-amylase 9 (BAM9), pullulanase (PUL), and glycogen debranching enzyme (glgX), were the major genes involved in starch degradation. AMY1, BAM3, BAM9, PUL, and glgX all were upregulated and had higher expression levels with ethylene treatment compared to the controls, suggesting that ethylene treatment may be responsible for accelerating starch degradation. The expression profile of alpha-1,4-galacturonosyltransferase (GAUT) and granule-bound starch synthase (GBSS) coincided with protopectin content changes and could involve protopectin synthesis. Pectinesterase (PE), polygalacturonase (PG), and pectate lyase (PEL) all involved in pectin degradation; PE was significantly upregulated by ethylene and was the key enzyme implicated pectin degradation. CONCLUSION: Both KEGG pathway enrichment analysis of DEGs and material content analysis confirmed that starch decomposition into soluble sugars and cell wall polysaccharides metabolism are closely related to the ripening and cracking of AP atemoya. A link between gene up- or downregulation during different cracking stages of atemoya fruits and how their expression affects starch and pectin contents were established by RT-qPCR analysis.

Polysaccharides from pineapple pomace: new insight into ultrasonic-cellulase synergistic extraction and hypoglycemic activities.

This study determined the optimal extraction conditions for ultrasonic-cellulase synergistic extraction of polysaccharides from pineapple pomace (PPP) using Plackett-Burman design and response surface methodology. The optimal hydrolysis temperature, ratio of material to water, pH value, hydrolysis time, ultrasonic power and the additive quantity of cellulase were 50 °C, 1:45 g/mL, 6.0, 100 min, 160 W and 2.0%, respectively, giving a extraction yield of 1.10 ±â€¯0.03%. PPP was further isolated and purified by DEAE-52 cellulose and Sephadex G-100 chromatography columns, revealing four main elution peaks, named PPF0, PPF1, PPP2 and PPF3, were obtained. The molecular weight, monosaccharide compositions, structural features and appearance morphology of polysaccharide fractions (PPFs) were analyzed by high-performance liquid chromatography (HPLC), gel permeation chromatography (GPC), UV spectroscopy, fourier transform infrared spectroscopy (FT-IR), and scanning electron microscopy (SEM). Furthermore, the hypoglycemic activities of PPFs with different concentrations were also investigated by insulin resistance HepG2 cells model in vitro. Results showed that PPF0, PPF1, PPF2 and PPF3 were composed of mannose, rhamnose, glucuronic acid, galacturonic acid, glucose, galactose, xylose, arabinose and fucose with molecular weight of 6.71 × 104, 1.11 × 104, 2.22 × 104 and 5.1 × 103 Da, respectively. All of them could alleviate the development of insulin resistance HepG2 cells with a dose-dependent relationship. The glucose consumption increased 46.4%, 50.5%, 82.1% and 53.6%; 86.8%, 81.6%, 86.8% and 84.2% at the concentration of 20 µg/mL, respectively, without or with insulin. These results suggested that PPFs can be explored as a potential hypoglycemic agent in biomedical and functional food.

Genome-Wide Identification and Analysis of U-Box E3 Ubiquitin⁻Protein Ligase Gene Family in Banana.

The U-box gene family is a family of genes which encode U-box domain-containing proteins. However, little is known about U-box genes in banana (Musa acuminata). In this study, 91 U-box genes were identified in banana based on its genome sequence. The banana U-box genes were distributed across all 12 chromosomes at different densities. Phylogenetic analysis of U-box genes from banana, Arabidopsis, and rice suggested that they can be clustered into seven subgroups (I⁻VII), and most U-box genes had a closer relationship between banana and rice relative to Arabidopsis. Typical U-box domains were found in all identified MaU-box genes through the analysis of conserved motifs. Four conserved domains were found in major banana U-box proteins. The MaU-box gene family had the highest expression in the roots at the initial fruit developmental stage. The MaU-box genes exhibited stronger response to drought than to salt and low temperatures. To the best of our knowledge, this report is the first to perform genome-wide identification and analysis of the U-box gene family in banana, and the results should provide valuable information for better understanding of the function of U-box in banana.

Optimization extraction, characterization and anticancer activities of polysaccharides from mango pomace.

Response surface methodology was used to optimize the extraction conditions for ultrasonic-assisted extraction of polysaccharides from mango pomace. The Optimum extraction conditions consisted of extraction temperature of 74 °C, ultrasonic power of 170 W, extraction time of 100 min, and raw material-to-water ratio of 1:40 g/mL. Under these conditions, the extraction yield was 3.71 ±â€¯0.07%. Three novel polysaccharide fractions, MG-1, MG-2 and MG-3 were purified from the crude polysaccharides by using DEAE-52 cellulose and Sephadex G-100 column chromatography. The molecular weight and monosaccharide composition of polysaccharide fractions (MPFs) were analyzed by high performance liquid gel permeation chromatography (HPGPC) and HPLC analysis, respectively. The characterizations of MPFs were conducted with FT-IR, 1H NMR and SEM. Furthermore, the anticancer activities of the polysaccharide fractions were also investigated in vitro. Results showed that MG-1, MG-2 and MG-3 exhibited significant anticancer activities against HepG2, MCF-7, A549, HeLa, A2780, HCT-116 and BGC-823 cells in a dose-dependent manner. MPFs were also showed to promote apoptosis as seen in the nuclear morphological examination study using calcein acetyl methoxy methyl easter (calcein-AM) and propidium iodide (PI) staining. This research could serve as a theoretical reference for the efficient utilization of MPFs in biomedical and functional food.

Optimization extraction and functional properties of soluble dietary fiber from pineapple pomace obtained by shear homogenization-assisted extraction.

Response surface methodology (RSM) was used to optimize the extraction conditions for shear homogenization-assisted extraction of soluble dietary fiber from pineapple pomace (s-SDF), and the absorption capacities and antioxidant activities of the obtained s-SDF were also investigated. The optimum extraction conditions consisted of a cutting speed of 9000 rpm, a cutting time of 20 min, a cellulase content of 5.0%, a hydrolysis time of 120 min, a pH value of 4.5, a hydrolysis temperature of 50 °C, and a raw material to water ratio of 1 : 45 g mL-1. Under these conditions, the theoretical and actual extraction yields of s-SDF were 8.80% and 8.76%, respectively. An absorption capacity analysis indicated that s-SDF exhibited higher absorption abilities to sodium cholate, cholesterol and fat. In addition, s-SDF possessed higher antioxidant activities, showing a positive concentration effect relationship for DPPH˙, ABTS+, ·OH and O2 -˙. The concentration of 1.0 mg mL-1 scavenged 76.72% DPPH˙, 58.40% ABTS+, 23.47% ·OH and 48.47% O2 -˙, respectively, and the reduction power was 0.70. These results indicated that pineapple pomace is a potential source of natural dietary fiber and a potential functional food ingredient.

Genome-wide analysis of the DNA-binding with one zinc finger (Dof) transcription factor family in bananas.

DNA-binding with one finger (Dof) domain proteins are a multigene family of plant-specific transcription factors involved in numerous aspects of plant growth and development. In this study, we report a genome-wide search for Musa acuminata Dof (MaDof) genes and their expression profiles at different developmental stages and in response to various abiotic stresses. In addition, a complete overview of the Dof gene family in bananas is presented, including the gene structures, chromosomal locations, cis-regulatory elements, conserved protein domains, and phylogenetic inferences. Based on the genome-wide analysis, we identified 74 full-length protein-coding MaDof genes unevenly distributed on 11 chromosomes. Phylogenetic analysis with Dof members from diverse plant species showed that MaDof genes can be classified into four subgroups (StDof I, II, III, and IV). The detailed genomic information of the MaDof gene homologs in the present study provides opportunities for functional analyses to unravel the exact role of the genes in plant growth and development.

Metabolism of Flavonoids in Novel Banana Germplasm during Fruit Development.

Banana is a commercially important fruit, but its flavonoid composition and characteristics has not been well studied in detail. In the present study, the metabolism of flavonoids was investigated in banana pulp during the entire developmental period of fruit. 'Xiangfen 1,' a novel flavonoid-rich banana germplasm, was studied with 'Brazil' serving as a control. In both varieties, flavonoids were found to exist mainly in free soluble form and quercetin was the predominant flavonoid. The most abundant free soluble flavonoid was cyanidin-3-O-glucoside chloride, and quercetin was the major conjugated soluble and bound flavonoid. Higher content of soluble flavonoids was associated with stronger antioxidant activity compared with the bound flavonoids. Strong correlation was observed between antioxidant activity and cyanidin-3-O-glucoside chloride content, suggesting that cyanidin-3-O-glucoside chloride is one of the major antioxidants in banana. In addition, compared with 'Brazil,' 'Xiangfen 1' fruit exhibited higher antioxidant activity and had more total flavonoids. These results indicate that soluble flavonoids play a key role in the antioxidant activity of banana, and 'Xiangfen 1' banana can be a rich source of natural antioxidants in human diets.

Genome-wide identification and expression profiling reveal tissue-specific expression and differentially-regulated genes involved in gibberellin metabolism between Williams banana and its dwarf mutant.

BACKGROUND: Dwarfism is one of the most valuable traits in banana breeding because semi-dwarf cultivars show good resistance to damage by wind and rain. Moreover, these cultivars present advantages of convenient cultivation, management, and so on. We obtained a dwarf mutant '8818-1' through EMS (ethyl methane sulphonate) mutagenesis of Williams banana 8818 (Musa spp. AAA group). Our research have shown that gibberellins (GAs) content in 8818-1 false stems was significantly lower than that in its parent 8818 and the dwarf type of 8818-1 could be restored by application of exogenous GA3. Although GA exerts important impacts on the 8818-1 dwarf type, our understanding of the regulation of GA metabolism during banana dwarf mutant development remains limited. RESULTS: Genome-wide screening revealed 36 candidate GA metabolism genes were systematically identified for the first time; these genes included 3 MaCPS, 2 MaKS, 1 MaKO, 2 MaKAO, 10 MaGA20ox, 4 MaGA3ox, and 14 MaGA2ox genes. Phylogenetic tree and conserved protein domain analyses showed sequence conservation and divergence. GA metabolism genes exhibited tissue-specific expression patterns. Early GA biosynthesis genes were constitutively expressed but presented differential regulation in different tissues in Williams banana. GA oxidase family genes were mainly transcribed in young fruits, thus suggesting that young fruits were the most active tissue involved in GA metabolism, followed by leaves, bracts, and finally approximately mature fruits. Expression patterns between 8818 and 8818-1 revealed that MaGA20ox4, MaGA20ox5, and MaGA20ox7 of the MaGA20ox gene family and MaGA2ox7, MaGA2ox12, and MaGA2ox14 of the MaGA2ox gene family exhibited significant differential expression and high-expression levels in false stems. These genes are likely to be responsible for the regulation of GAs content in 8818-1 false stems. CONCLUSION: Overall, phylogenetic evolution, tissue specificity and differential expression analyses of GA metabolism genes can provide a better understanding of GA-regulated development in banana. The present results revealed that MaGA20ox4, MaGA20ox5, MaGA20ox7, MaGA2ox7, MaGA2ox12, and MaGA2ox14 were the main genes regulating GA content difference between 8818 and 8818-1. All of these genes may perform important functions in the developmental processes of banana, but each gene may perform different functions in different tissues or during different developmental stages.

The banana E2 gene family: Genomic identification, characterization, expression profiling analysis.

The E2 is at the center of a cascade of Ub1 transfers, and it links activation of the Ub1 by E1 to its eventual E3-catalyzed attachment to substrate. Although the genome-wide analysis of this family has been performed in some species, little is known about analysis of E2 genes in banana. In this study, 74 E2 genes of banana were identified and phylogenetically clustered into thirteen subgroups. The predicted banana E2 genes were distributed across all 11 chromosomes at different densities. Additionally, the E2 domain, gene structure and motif compositions were analyzed. The expression of all of the banana E2 genes was analyzed in the root, stem, leaf, flower organs, five stages of fruit development and under abiotic stresses. All of the banana E2 genes, with the exception of few genes in each group, were expressed in at least one of the organs and fruit developments, which indicated that the E2 genes might involve in various aspects of the physiological and developmental processes of the banana. Quantitative RT-PCR (qRT-PCR) analysis identified that 45 E2s under drought and 33 E2s under salt were induced. To the best of our knowledge, this report describes the first genome-wide analysis of the banana E2 gene family, and the results should provide valuable information for understanding the classification, cloning and putative functions of this family.

Transcriptome characterization and expression profiles of the related defense genes in postharvest mango fruit against Colletotrichum gloeosporioides.

Anthracnose, caused by Colletotrichum gloeosporioides, is a major disease of the postharvest mango (Mangifera indica L.) fruit. However, a lack of transcriptomic and genomic information hinders our understanding of the molecular mechanisms underlying the mango fruit defense response. Here, we studied the host responses of the mango fruit against C. gloeosporioides using Illumina paired-end sequencing technology, and expression profiles of 35 defense-related genes were further analyzed by qRT-PCR. The results indicated that 5.9Gigabase pair clean reads were assembled into a total of 131,750 unigenes, of which 89,050 unigenes found to be homologous to genes in the NCBI GenBank database and 61,694 unigenes annonated in the Swiss-Prot database. Orthologous analyses showed that 47,770 unigenes were assigned with one or more Gene Ontology terms and 44,145 unigenes were classified into 256 Kyoto Encyclopedia of Genes and Genomes pathways. Moreover, qRT-PCR of 35 defense-related unigenes, including 17 ethylene response factors (ERFs), 6 nucleotide binding site-leucine-rich repeats (NBS-LRRs), 6 nonexpressor of pathogenesis-related genes (NPRs) and 6 pathogenesis-related protein (PRs), revealed that most of these defense-related genes were up-regulated after C. gloeosporioides infection. Taken together, our study provides a platform to discover new candidate genes in mango fruit in relation to pathogen resistance.

Effects of wax treatment on the physiology and cellular structure of harvested pineapple during cold storage.

Pineapple (Ananas comosus (L.) Merr. cv. 'Paris') is sensitive to low temperature and highly susceptible to blackheart during cold storage, which causes serious fruit decay. This work investigated the effect of wax treatment (Sta-Fresh 2952, 60 g/L) on blackheart of pineapple under chilling stress. Wax treatment significantly reduced blackheart symptoms after 14 days of storage and markedly delayed changes in firmness and flesh color during the whole period of storage. The weight loss of wax-treated fruit (2.6%) was less than the control (3.1%) at the 24th day of storage. The treatment decreased the activities of PG and EGase for maintaining cell wall stability during the later period of storage. In the control fruit, the structure of flesh cells was significantly damaged under chilling stress, with looser cell wall, absence of middle lamella, loss of membrane integrity, and many cells near the vascular tissue collapsed. The subcellular elements could be barely observed in the control after storage. These destructive symptoms were significantly alleviated in the wax-treated fruit. The results suggest that wax treatment could reduce blackheart of pineapple under chilling stress via maintenance of cell integrity.