ABSTRACT
Corynespora cassiicola is a highly diverse fungal pathogen that commonly occurs in tropical, subtropical, and greenhouse environments worldwide. In this study, the isolates were identified as C. cassiicola, and the optimum growth and sporulation were studied. The phenotypic characteristics of C. cassiicola, concerning 950 different growth conditions, were tested using Biolog PM plates 1-10. In addition, the strain of C. cassiicola DWZ from tobacco hosts was sequenced for the using Illumina PE150 and Pacbio technologies. The host resistance of tobacco Yunyan 87 with different maturity levels was investigated. In addition, the resistance evaluation of 10 common tobacco varieties was investigated. The results showed that C. cassiicola metabolized 89.47% of the tested carbon source, 100% of the nitrogen source, 100% of the phosphorus source, and 97.14% of the sulfur source. It can adapt to a variety of different osmotic pressure and pH environments, and has good decarboxylase and deaminase activities. The optimum conditions for pathogen growth and sporulation were 25-30 °C, and the growth was better on AEA and OA medium. The total length of the genome was 45.9 Mbp, the GC content was 51.23%, and a total of 13,061 protein-coding genes, 202 non-coding RNAs and 2801 and repeat sequences were predicted. Mature leaves were more susceptible than proper mature and immature leaves, and the average diameter of diseased spots reached 17.74 mm at 12 days. None of the tested ten cultivars exhibited obvious resistance to Corynespora leaf spot of tobacco, whereby all disease spot diameters reached > 10 mm and > 30 mm when at 5 and 10 days after inoculation, respectively. The phenotypic characteristics, genomic analysis of C. cassiicola and the cultivar resistance assessment of this pathogen have increased our understanding of Corynespora leaf spot of tobacco.
Subject(s)
Ascomycota , Nicotiana , Plant Diseases , Nicotiana/microbiology , Nicotiana/genetics , Ascomycota/genetics , Ascomycota/pathogenicity , Plant Diseases/microbiology , Plant Leaves/microbiology , Genomics/methods , Disease Resistance/genetics , Genome, Fungal , PhenotypeABSTRACT
The present study aimed to investigate the impact of Flammulina velutipes polysaccharide (FVSP) on the rheological properties and structural alterations of myofibrillar protein (MP) and oxidized MP (OMP), utilizing techniques such as rhehometer, fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). In the unoxidized system, the addition of 5.00% FVSP significantly improved (p < 0.05) the storage and loss moduli of the composite gel and promoted the α-helix to ß-sheet transformation. These effects enhanced the protein's gel strength and water-holding capacity (WHC). In the oxidation system, 5.00% FVSP had significant effects (p < 0.05) on repair and improvement of the oxidized MP. These effects inhibited the cross-linking aggregation and degradation of the protein. In addition, the addition of FVSP significantly improved the gel properties of MPs after oxidation (p < 0.05), hindered fracture of the protein gel network structure. In summary, polysaccharides have a substantial effect on the functional characteristics of MP, and FVSP could potentially be applied in meat products.
Subject(s)
Flammulina , Muscle Proteins , Oxidation-Reduction , Polysaccharides , Flammulina/chemistry , Polysaccharides/chemistry , Animals , Muscle Proteins/chemistry , Swine , Gels/chemistry , Meat Products/analysis , Rheology , Myofibrils/chemistryABSTRACT
Epicoccum latusicollum is a fungus that causes a severe foliar disease on flue-cured tobacco in southwest China, resulting in significant losses in tobacco yield and quality. To better understand the organism, researchers investigated its optimal growth conditions and metabolic versatility using a combination of traditional methods and the Biolog Phenotype MicroArray technique. The study found that E. latusicollum exhibited impressive metabolic versatility, being able to metabolize a majority of carbon, nitrogen, sulfur, and phosphorus sources tested, as well as adapt to different environmental conditions, including broad pH ranges and various osmolytes. The optimal medium for mycelial growth was alkyl ester agar medium, while oatmeal agar medium was optimal for sporulation, and the optimum temperature for mycelial growth was 25°C. The lethal temperature was 40°C. The study also identified arbutin and amygdalin as optimal carbon sources and Ala-Asp and Ala-Glu as optimal nitrogen sources for E. latusicollum. Furthermore, the genome of E. latusicollum strain T41 was sequenced using Illumina HiSeq and Pacific Biosciences technologies, with 10,821 genes predicted using Nonredundant, Gene Ontology, Clusters of Orthologous Groups, Kyoto Encyclopedia of Genes and Genomes, and SWISS-PROT databases. Analysis of the metabolic functions of phyllosphere microorganisms on diseased tobacco leaves affected by E. latusicollum using the Biolog Eco microplate revealed an inability to efficiently metabolize a total of 29 carbon sources, with only tween 40 showing some metabolizing ability. The study provides new insights into the structure and function of phyllosphere microbiota and highlights important challenges for future research, as well as a theoretical basis for the integrated control and breeding for disease resistance of tobacco Epicoccus leaf spot. This information can be useful in developing new strategies for disease control and management, as well as enhancing crop productivity and quality.
ABSTRACT
Validamycin, is classified as an environmentally friendly fungicide. It has high efficacy with little associated pollution risk, and it has been used in China on tobacco for many years especially during leaf spot season. To understand changes in microbial communities and functional aspects of the tobacco phyllosphere after exposure to validamycin, the chemical was sprayed on tobacco leaves during brown spot epidemic periods caused by Alternaria alternata, and asymptomatic and symptomatic leaves of tobacco were sampled at different times (0 day before, 5, 10, and 15 days after application). The fungal and bacterial population diversity and structure were revealed using Illumina NovaSeq PE250 high-throughput sequencing technology, and Biolog-ECO technology which analyzes the metabolic differences between samples by using different carbon sources as the sole energy source. The results showed that the microbial community structure of both asymptomatic and symptomatic tobacco leaves changed after the application of valproate, with the microbial community structure of the asymptomatic tobacco leaves being more strongly affected than that of the symptomatic leaves, and the diversity of bacteria being greater than that of fungi. Phyllosphere fungal diversity in asymptomatic leaves increased significantly after application, and bacterial abundance and diversity in both asymptomatic and symptomatic leaves first increased and then decreased. Validamycin treatment effectively reduced the relative abundance of Alternaria, Cladosporium, Kosakonia, and Sphingomonas in leaves showing symptoms of tobacco brown spot, while the relative abundance of Thanatephorus, Pseudomonas, and Massilia increased significantly after application. Furthermore, the ability to metabolize a variety of carbon sources was significantly reduced in both types of leaves after validamycin application, and both types had a weaker ability to metabolize α-Ketobutyric Acid after application. This study reveals phyllosphere micro-ecological changes in symptomatic and asymptomatic tobacco leaves during different periods after validamycin application and the effects on the metabolic capacity of phyllosphere microorganisms. It can provide some basis for exploring the effect of validamycin on the control of tobacco brown spot.
ABSTRACT
Piriformospora indica is an important endophytic fungus with broad potential for alleviating biotic and abiotic stress on host plants. This study monitored the growth dynamics of P. indica on five commonly used artificial media for microorganisms and analyzed its metabolic characteristics using Biolog Phenotype Microarray (PM) technology. The results showed that P. indica grew fastest on Potato Dextrose Agar (PDA), followed by Kidney Bean Agar (KBA), Alkyl Ester Agar (AEA), Oatmeal Agar (OA), and Luria-Bertani Agar (LB), and the most suitable medium for spore production was OA. Using Biolog PM1-10, 950 metabolic phenotypes of P. indica were obtained. P. indica could metabolize 87.89% of the tested carbon sources, 87.63% of the tested nitrogen sources, 96.61% of the tested phosphorus sources, and 100% of the tested sulfur sources. P. indica displayed 92 kinds of tested biosynthetic pathways, and it could grow under 92 kinds of tested osmotic pressures and 88 kinds of tested pH conditions. PM plates 1-2 revealed 43 efficient carbon sources, including M-Hydroxyphenyl acid, N-Acetyl-D-Glucosamine, Tyramine, Maltotrios, α-D-Glucosine, I-Erythritol, L-Valine, D-Melezitose, D-Tagatose, and Turanose. PM plates 3,6-8 indicated 170 efficient nitrogen sources, including Adenosine, Inosine Allantoin, D, L-Lactamide, Arg-Met, lle-Trp, Ala-Arg, Thr-Arg, Trp-Tyr, Val-Asn, Gly-Gly-D-Leu, Gly-Gly-Phe, and Leu-Leu-Leu. This study demonstrates that P. indica can metabolize a variety of substrates, such as carbon and nitrogen sources, and has a wide range of environmental adaptability. The growth dynamics on artificial culture media and metabolic phenotypes of P. indica can be used to investigate its biological characteristics, screen for more suitable growth and sporulation conditions, and elucidate the physiological mechanisms that enhance the stress resistance of host plants. This study provides a theoretical basis for its better application in agriculture.
ABSTRACT
This study investigated the joint effect of the enzymatic and high-intensity ultrasonic (HIU) treatments on the emulsifying and structural properties of egg yolk (EY). The evaluation was performed under different HIU powers (i.e., 0, 240, and 480 W) and for different durations (10, 20, and 30 min). Results reveal that the emulsification activity and emulsification capacity at 240 W for 10 min significantly increase than that of the samples without HIU treatment (P < 0.05). The thermal stability reached a maximum of 90.33% in 480 W + 30 min. The HIU treatment under low power and for a short time (240 W + 10 min) reduced the average particle size of the EY to 224.77 nm. The combined treatment produced high zeta potential and total sulfhydryl contents. The highest hydrophobicity was 111.49 µg at 240 W for 20 min. The joint treatment enhanced the hardness and cohesiveness of the EY. Moreover, the α-helix content significantly decreased from 37.13% to 25.02% after the enzymatic treatment, while the ß-sheet content increased with time in the 240 W treatment. The G' and Gâ³ curves show an increasing tendency, and the highest G' and Gâ³ were obtained in the 240 W + 30 min group, which is consistent with the dense microstructure. These results reveal that the combined treatment can improve the emulsifying characteristics, which could influence the structural properties of EY, expanding its application in the food industry.
Subject(s)
Egg Yolk , Ultrasonics , Egg Yolk/chemistry , Hydrophobic and Hydrophilic Interactions , Particle Size , Sulfhydryl Compounds/chemistryABSTRACT
This research aims to investigate the effects of Staphylococcus xylosus YCC3 (Sx YCC3) and Lactobacillus plantarum MSZ2 (Lp MSZ2) on lipid hydrolysis and oxidation, the bacterial community's composition, and the volatile flavor compounds in fermented sausage. The bacterial community was examined by plate counting and high-throughput sequencing. Differential flavor compounds in non-inoculated and inoculated sausages were identified by principal component analysis (PCA) and orthogonal partial least squares discrimination analysis (OPLS-DA). The results showed that the free fatty acid (FFA) content was increased after inoculating with Sx YCC3 or Lp MSZ2. The pH, peroxide value (POV), thiobarbituric acid reactive substances (TBARS) value, lipoxygenase activity, and the counts of Enterobacteriaceae were lower in the inoculated sausage than in the non-inoculated sausage. The bacterial inoculation enhanced the competitiveness of Staphylococcus and Lactobacillus and restricted the growth of unwanted bacteria. The OPLS-DA revealed that (Z)-hept-2-enal, (E)-2-octenal, 1-nonanal, octanal, and 1-octen-3-ol were common differential flavor compounds that were found in the inoculated sausages but were not found in the non-inoculated sausages. A positive correlation was observed between the differential flavor compounds and the relative abundance of Staphylococcus or Lactobacillus, or the FFA content. Our results indicated that inoculation with Sx YCC3 or Lp MSZ2 can improve fermented sausages' flavor by enhancing their bacterial quality and increasing their FFA content.
ABSTRACT
Bradysia odoriphaga is an agricultural pest in China's vegetable industry. In this study, pupae and adults were exposed to various non-lethal high-temperatures. The results demonstrated a decreased rate of eclosion once the pupae were exposed to temperatures exceeding 37 °C for 1 h. No effect on the lifespan of unmated female adults was observed after exposure to temperature stress, while unmated male adult lifespan decreased (>37 °C for 2 h). The size of the testis and ovaries for unmated male and female adults decreased, as did the fecundity and egg hatching rate for mated females. Compared with the control group (25 °C), the testis size of unmated male adults decreased after high-temperature stress followed by recovery at 25 °C for 1 h, though the size of the ovaries of female adults did not change. Additionally, the size of the testis and ovaries for unmated male and female adults decreased following high-temperature stress and 24 h of recovery at 25 °C. High temperatures affected males more than females; 37 °C is the critical temperature to control the population of B. odoriphaga. These results lay the foundation for the future development of environmentally friendly high-temperature prevention and pest-control strategies.
ABSTRACT
Microglia-mediated neuroinflammation in response to injurious self and non-self-stimuli exerts detrimental effects on neurons, which may lead to cognitive impairment. Luteolin, a typical kind of natural flavonoid in honeysuckle, chrysanthemum, and Herba Schizonepetae, is widely recognized to be anti-inflammatory and antioxidant against peripheral inflammation. However, its protective effect against inflammation-induced cognitive impairment is currently unknown. In this paper, we investigated the relief potential of luteolin against lipopolysaccharide (LPS)-induced cognitive impairment and neuroinflammation and its possible anti-inflammatory mechanisms in lipopolysaccharide-stimulated BV2 microglia cells. In this study, luteolin ameliorated LPS-induced cognitive impairments, indicated by behavioral performance of neuroinflammatory model mice in Morris water maze tests. Protein analyses and histological examination also revealed protective effect of luteolin against neuronal damage, through inhibiting overproduction of inflammatory cytokines in both hippocampus and cortex of mice. We also observed luteolin in vitro significantly suppressed the levels of pro-inflammatory cytokines, such as tumor necrosis factor alpha (TNF-α) and interleukin-1 ß (IL-1ß), and inflammatory mediators like nitric oxide. Taken together, these results demonstrated luteolin was effective in alleviating cognitive impairment and limited neuronal damage via inhibiting the release of inflammatory mediators, suggesting luteolin is potential for further therapeutic research of neuroinflammation-related neurodegenerative diseases.
Subject(s)
Cognitive Dysfunction/drug therapy , Luteolin/therapeutic use , Microglia/drug effects , Neuroinflammatory Diseases/drug therapy , Animals , Blotting, Western , Cell Line , Cognitive Dysfunction/etiology , Enzyme-Linked Immunosorbent Assay , Female , Male , Mice , Mice, Inbred C57BL , Morris Water Maze Test , Neuroinflammatory Diseases/complicationsABSTRACT
Psoriasis is considered as a common chronic immune-mediated skin disorder characterized by abnormal keratinocyte proliferation. Luteolin, an anti-inflammatory natural flavonoid with well-accepted inhibition effect against keratinocyte proliferation, was hypothesized to have a potential therapeutic effect for psoriasis. In this paper, we investigated the relieving effect of luteolin against imiquimod-induced psoriasis-like lesions on BALB/c mice and its possible anti-inflammatory mechanisms in lipopolysaccharide-stimulated macrophages (RAW264.7 cells). We found that luteolin ameliorated psoriasis-like skin lesions, suppressed the cutaneous infiltration of macrophages, T cells and neutrophils, and downregulated the expression of cytokines like IL-6, IL-1ß, TNF-α, IL-17A and IL-23 in both skin lesions and eyeball blood of model mice. In vitro, we observed luteolin significantly suppressed the levels of psoriasis-related pro-inflammatory cytokines, such as IL-17A, IL-6, TNF-α and IL-23, and inflammatory mediators like nitric oxide NO, inducible NOS, COX-2 in RAW264.7 cells. The anti-inflammatory activity was accomplished by inhibiting NF-κB expression and activation. This study demonstrates luteolin is effective in alleviating psoriasis-like skin lesions and downregulating inflammatory response via NF-κB pathway, suggesting luteolin as a potential molecule for further therapeutic research of inflammation-related skin diseases like psoriasis.
Subject(s)
Imiquimod/toxicity , Luteolin/therapeutic use , Psoriasis/drug therapy , Animals , Anti-Inflammatory Agents/pharmacology , Cytokines/biosynthesis , Female , Macrophages/drug effects , Mice , Mice, Inbred BALB C , NF-kappa B/antagonists & inhibitors , NF-kappa B/physiology , Nitric Oxide/biosynthesis , Psoriasis/chemically induced , Psoriasis/immunology , RAW 264.7 CellsABSTRACT
The production of Chinese chives is reduced throughout China due to a root-feeding dipteran pest Bradysia odoriphaga Yang et Zhang (Diptera: Sciaridae), therefore deciphering the conditions influencing its growth and development are important in developing ecological control strategies. A study was conducted from 2014 to 2017 to determine the relationship between the abundance of B. odoriphaga and temperature (atmospheric and soil), soil water content, and atmospheric humidity in a Chinese chive field in Beijing City, China. Numbers of adults peaked in March and October to November and were lowest in July to August and December to next February; numbers of larvae were highest in December to next February and lowest in July to August. From 2014 to 2017, the numbers of adults and larvae were significantly correlated with monthly mean atmospheric temperatures and soil temperatures, but were not significantly correlated with monthly mean atmospheric relative humidity and soil water content. However, for both adults and larvae, numbers were significantly greater with high soil water contents compared with drought treatment. The results of this study suggest that the very low soil water contents, high atmospheric temperatures, and high soil temperatures were critical for regulating field populations of B. odoriphaga.
Subject(s)
Diptera , Insecticides , Animals , China , Humidity , TemperatureABSTRACT
Throughout China, the dipteran pest Bradysia odoriphaga significantly reduces Chinese chive production; therefore, identifying conditions that influence its growth and development is crucial for developing ecological regulation strategies. In this study, different non-lethal high temperatures and treatment durations were used to stress the third-instar larvae of B. odoriphaga, and the effects of this treatment on their growth and offspring were recorded and analyzed. The results showed that the average larval mortality increased with increased temperature and prolonged exposure times. After stress treatment at 40 °C for 2 h, 100% of larvae died within 5 days, which was not significantly different from the 5-day average larval mortality (90.66%) after stress at 37 °C for 4 h, but significantly higher than the 5-day average larval mortality (72.00%) after stress at 40 °C for 1 h. After 5 days, all still-living larvae could pupate, and there was no significant difference in average pupal period after pupation. However, the eclosion rate of subsequent pupae decreased with increased temperature and prolonged exposure times, and were only 43.00% and 42.73% after larvae were stressed at 37 °C for 4 h and 40 °C for 1 h, respectively. After eclosion into adults, there was no significant difference in the lifespan of unmated female adults, while the lifespan of unmated male adults was significantly reduced to 1.67 d and 2 d after larvae were stressed at 37 °C for 4 h and 40 °C for 1 h, respectively. However, there was no significant difference in male and female adult longevity after mating. There was no significant difference in oviposition or egg hatchability. This indicates that non-lethal high temperature at 37 °C for 4 h can hinder development and allow control of B. odoriphaga. There is great potential for non-lethal high temperature to be applied in the field to control agricultural pests.
ABSTRACT
Bradysia odoriphaga Yang & Zhang (Diptera: Sciaridae) is an important pest of Chinese chives. Information on the effects of biotic and abiotic factors on the flight performance of B. odoriphaga is crucial for understanding the pest's ability to disperse and migrate. In this study, the effects of sex and air temperature on the flight performance of B. odoriphaga imagoes were assessed by tethering individual imagoes to computerized flight mills for a 10-h experiment. The results showed that the percentage of imagoes that flew a particular distance gradually decreased as flight distance increased. The percentage of imagoes was significantly higher for males than females when the flight distance was <300 m. Sex and air temperature significantly affected average flight time (which ranged from 14.6 to 68.3 min) and average flight distance (which ranged from 10.4 to 107.2 m), but did not significantly affect average flight speed (which ranged from 3.8 to 6.4 m/min). For both females and males, the average flight distance and flight time were shortest at 18°C and longest at 22°C; the interaction between air temperature and sex was not significant. The results suggest that B. odoriphaga has a poor potential for long-distance migration. These findings will be helpful for developing forecasting and management systems for B. odoriphaga.
Subject(s)
Chive , Diptera , Animals , Female , Flight, Animal , Larva , Male , TemperatureABSTRACT
Magnetic dodecyl chitosan/silica composite was synthesized and characterized for the enrichment of emerging bisphenol contaminants. The result demonstrated that bispehnol A, bisphenol AF, bisphenol F, and bisphenol S can be efficiently extracted via the resulting composite from aqueous solution. The adsorption rate of four investigated bisphenols on the resulting composite was achieved in the range of 87â»99% within 15 min. Bispehnol A was taken as a representative adsorbate to investigate the adsorption studies in detail. The hydrophobic interaction was proposed as the principal mechanism for the adsorption of BPs. The satisfactory reusability of the resulting composite can be quickly achieved by magnetic separation technologies. Magnetic dodecyl chitosan/silica composite has a potential to be applied as a type of efficient and easily recyclable sorbent for the removal of trace organic pollutants from aqueous sample.
ABSTRACT
Sturgeon is an important fresh water-culture fish in China. A problem with sturgeon is its high susceptibility to spoilage. Food spoilage is reported to be regulated by quorum sensing (QS). To identify the QS signals acetylated homoserine lactones (AHLs) in sturgeon and test whether QS plays a role in the spoilage of sturgeon, we investigated the specific spoilage organisms (SSOs) in vacuum packaged sturgeon stored at 4°C and the production of AHLs by sturgeon SSOs. 16S rDNA sequencing and spoilage capabilities analysis revealed that Aeromonas veronii LP-11, Citrobacter freundii LPJ-2, and Raoultella ornithinolytica LPC-3 were the SSOs in sturgeon. Among the three SSOs, only A. veronii LP-11 induced the QS biosensors Agrobacterium tumefaciens KYC55 and Chromobacterium violaceum CV026, suggesting that it produced AHLs. Analysis by thin layer chromatography, high-performance liquid chromatography-triple quadrupole tandem mass spectrometry, and high-performance liquid chromatography quadrupole time-of-flight mass spectrometry (HPLC/qTOF-MS) identified that the AHLs produced by A. veronii were C6-SHL, C8-HSL, 3-oxo-C8-HSL, and 3-OH-C8-HSL. Our study revealed that QS system was probably involved in the regulation of sturgeon spoilage and for the first time reported the production of C8-HSL and 3-OH-C8-HSL by genus Aeromonas. As only HPLC/qTOF-MS effectively and accurately identified all the four AHLs produced by A. veronii LP-11, this study also showed that HPLC/qTOF-MS was the most efficient method for rapid analysis of AHLs in complex microbial sample. The study provides new insight into the microbiology of sturgeon spoilage which may be helpful for better sturgeon preservation.
ABSTRACT
The botanical compound allyl isothiocyanate (AITC) is toxic to many microorganisms and insects. The aim of this study was to assess the effects of AITC on the Bradysia odoriphaga Yang et Zhang (Diptera: Sciaridae) and the seeds and seedlings of the Chinese chive. Allyl isothiocyanate was toxic to all four developmental stages of B. odoriphaga. The adult was significantly more sensitive to AITC than the other three stages, which exhibited no significant differences to one another in sensitivity to the chemical. The control efficacy of AITC against B. odoriphaga was far superior in the greenhouse than the field. In addition, seedling survival was higher in the greenhouse compared with that in the field. In the absence of B. odoriphaga, seed germination and seedling growth of Chinese chives were inhibited by 16 µl/liter of AITC, and significant inhibition occurred under higher doses of AITC. These results indicate that AITC could be used to control B. odoriphaga during cultivation of Chinese chives.
Subject(s)
Chive/drug effects , Diptera , Insecticides , Isothiocyanates , Animals , Diptera/growth & development , Insecticides/toxicity , Isothiocyanates/toxicity , Larva/growth & development , Ovum/growth & development , Pupa/growth & development , Seedlings/drug effects , Seeds/drug effectsABSTRACT
BACKGROUND: The Pythium splendens is a potentially useful organism for the synthesis of large amounts of eicosapentaenoic acid. Peak biomass and lipid accumulation do not occur at the same time and growth temperature has an effect on the fatty acid composition. Little is known about the pathway or the genes involved in growth, lipid synthesis or temperature resistance in P. splendens. Analysis of the transcriptome and expression profile data for P.splendensRBB-5 were used to extend genetic information for this strain and to contribute to a comprehensive understanding of the molecular mechanisms involved in specific biological processes. METHODOLOGY/PRINCIPAL FINDINGS: This study used transcriptome assembly and gene expression analysis with short-read sequencing technology combined with a tag-based digital gene expression (DGE) system. Assembled sequences were annotated with gene descriptions, such as gene ontology (GO), clusters of orthologous group (COG) terms and KEGG orthology (KO) to generate 23,796 unigenes. In addition, we obtained a larger number of genes at different stages of fermentation (48, 100 and 148 h). The genes related to growth characteristics and lipid biosynthesis were analyzed in detail. Some genes associated with lipid and fatty acid biosynthesis were selected to confirm the digital gene expression (DGE) results by quantitative real-time PCR (qRT-PCR). CONCLUSION/SIGNIFICANCE: The transcriptome improves our genetic understanding of P.splendensRBB-5 greatly and makes a large number of gene sequences available for further study. Notably, the transcriptome and DGE profiling data of P.splendensRBB-5 provide a comprehensive insight into gene expression profiles at different stages of fermentation and lay the foundation for the study of optimizing lipid content and growth speed at the molecular level.
Subject(s)
Fermentation , Gene Expression Profiling , Pythium/genetics , Transcriptome , Gas Chromatography-Mass Spectrometry , Lipids/analysis , Real-Time Polymerase Chain Reaction , Sequence Analysis, DNAABSTRACT
A new full-length cDNA (PsD6) putatively encoding a ∆(6)-desaturase was cloned from the eicosapentaenoic acid-producing fungus Pythium splendens RBB-5. PsD6 contained an open reading frame of 1380 bp encoding a protein of 459 amino acids. The deduced amino acid sequence showed high similarity to those of other ∆(6)-desaturases. A recombinant vector expressing PsD6 (pPIC3.5K-D6) was constructed and transformed into Pichia pastoris GS115. The heterologous expressed PsD6 in P. pastoris desaturated linoleic acid to γ-linolenic acid but not desaturated α-linolenic acid to stearidonic acid. The results indicated that PsD6 was a fatty acid ∆(6)-desaturase and it had a substrate specificity for linoleic acid.
Subject(s)
Eicosapentaenoic Acid/metabolism , Linoleoyl-CoA Desaturase/genetics , Linoleoyl-CoA Desaturase/metabolism , Pythium/enzymology , Pythium/metabolism , Biotransformation , Cloning, Molecular , DNA, Fungal/chemistry , DNA, Fungal/genetics , Gene Expression , Genetic Vectors , Linoleic Acids/metabolism , Molecular Sequence Data , Open Reading Frames , Pichia/genetics , Pythium/genetics , Sequence Analysis, DNA , Sequence Homology, Amino AcidABSTRACT
A method was developed for rapid screening and isolation of eicosapentaenoic acid (EPA)-producing soil fungi through polymerase chain reaction (PCR) amplification. Genes coding for delta6 fatty acid desaturase and delta5 fatty acid desaturase were used as molecular markers for screening these EPA-producing fungi from soil. Three out of 65 soil fungi gave positive results through PCR amplification. Two out of these three strains were found to produce EPA when they had grown in 80 ml potato/dextrose liquid medium at (25 +/- 1) degrees C for 144 h. The EPA yields were 215.81 mg 1(-1) and 263.80 mg 1(-1), respectively. The other positive strain was detected to produce arachidonic acid (AA). This study indicates that molecular detection of genes encoding delta6 and delta5 desaturases is an efficient method for primary screening of EPA- or its related polyunsaturated fatty acids (PuFAs)-producing fungi, which can improve the screening efficiency prominently.