Heat stress-induced NO enhanced perylenequinone biosynthesis of Shiraia sp. via calcium signaling pathway.

Perylenequinones (PQs) are natural photosensitizing compounds used as photodynamic therapy, and heat stress (HS) is the main limiting factor of mycelial growth and secondary metabolism of fungi. This study aimed to unravel the impact of HS-induced Ca2+ and the calcium signaling pathway on PQ biosynthesis of Shiraia sp. Slf14(w). Meanwhile, the intricate interplay between HS-induced NO and Ca2+ and the calcium signaling pathway was investigated. The outcomes disclosed that Ca2+ and the calcium signaling pathway activated by HS could effectively enhance the production of PQs in Shiraia sp. Slf14(w). Further investigations elucidated the specific mechanism through which NO signaling molecules induced by HS act upon the Ca2+/CaM (calmodulin) signaling pathway, thus propelling PQ biosynthesis in Shiraia sp. Slf14(w). This was substantiated by decoding the downstream positioning of the CaM/CaN (calcineurin) pathway in relation to NO through comprehensive analyses encompassing transcript levels, enzyme assays, and the introduction of chemical agents. Concurrently, the engagement of Ca2+ and the calcium signaling pathway in heat shock signaling was also evidenced. The implications of our study underscore the pivotal role of HS-induced Ca2+ and the calcium signaling pathway, which not only participate in heat shock signal transduction but also play an instrumental role in promoting PQ biosynthesis. Consequently, our study not only enriches our comprehension of the mechanisms driving HS signaling transduction in fungi but also offers novel insights into the PQ synthesis paradigm within Shiraia sp. Slf14(w). KEY POINTS: • The calcium signaling pathway was proposed to participate in PQ biosynthesis under HS. • HS-induced NO was revealed to act upon the calcium signaling pathway for the first time.

L-Arginine enhanced perylenequinone production in the endophytic fungus Shiraia sp. Slf14(w) via NO signaling pathway.

Perylenequinones (PQ) are natural polyketides used as anti-microbial, -cancers, and -viral photodynamic therapy agents. Herein, the effects of L-arginine (Arg) on PQ biosynthesis of Shiraia sp. Slf14(w) and the underlying molecular mechanism were investigated. The total content of PQ reached 817.64 ± 72.53 mg/L under optimal conditions of Arg addition, indicating a 30.52-fold improvement over controls. Comparative transcriptome analysis demonstrated that Arg supplement promoted PQ precursors biosynthesis of Slf14(w) by upregulating the expression of critical genes associated with the glycolysis pathway, and acetyl-CoA and malonyl-CoA synthesis. By downregulating the expression of genes related to the glyoxylate cycle pathway and succinate dehydrogenase, more acetyl-CoA flow into the formation of PQ. Arg supplement upregulated the putative biosynthetic gene clusters for PQ and activated the transporter proteins (MFS and ABC) for exudation of PQ. Further studies showed that Arg increased the gene transcription levels of nitric oxide synthase (NOS) and nitrate reductase (NR), and activated NOS and NR, thus promoting the formation of nitric oxide (NO). A supplement of NO donor sodium nitroprusside (SNP) also confirmed that NO triggered promoted biosynthesis and efflux of PQ. PQ production stimulated by Arg or/and SNP can be significantly inhibited upon the addition of NO scavenger carboxy-PTIO, NOS inhibitor Nω-nitro-L-arginine, or soluble guanylate cyclase inhibitor NS-2028. These results showed that Arg-derived NO, as a signaling molecule, is involved in the biosynthesis and regulation of PQ in Slf14(W) through the NO-cGMP-PKG signaling pathway. Our results provide a valuable strategy for large-scale PQ production and contribute to further understanding of NO signaling in the fungal metabolite biosynthesis. KEY POINTS: • PQ production of Shiraia sp. Slf14(w) was significantly improved by L-arginine addition. • Arginine-derived NO was firstly reported to be involved in the biosynthesis and regulation of PQ. • The NO-cGMP-PKG signaling pathway was proposed for the first time to participate in PQ biosynthesis.

Comparative Evaluation of Microbiota Dynamics and Metabolites Correlation Between Spontaneous and Inoculated Fermentations of Nanfeng Tangerine Wine.

Understanding the evolution of microorganisms and metabolites during wine fermentation is essential for controlling its production. The structural composition and functional capacity of the core microbiota determine the quality and quantity of fruit wine. Nanfeng tangerine wine fermentation involves a complex of various microorganisms and a wide variety of metabolites. However, the microbial succession and functional shift of the core microbiota in this product fermentation remain unclear. Therefore, high-throughput sequencing (HTS) and headspace-gas chromatography-mass spectrometry (HS/GC-MS) were employed to reveal the core functional microbiota for the production of volatile flavors during spontaneous fermentation (SF) and inoculated fermentation (IF) with Saccharomyces cerevisiae of Nanfeng tangerine wine. A total of 13 bacterial and 8 fungal genera were identified as the core microbiota; Lactobacillus and Acetobacter were the dominant bacteria in SF and IF, respectively. The main fungal genera in SF and IF were Hanseniaspora, Pichia, and Saccharomyces with a clear succession. In addition, the potential correlations analysis between microbiota succession and volatile flavor dynamics revealed that Lactobacillus, Acetobacter, Hanseniaspora, and Saccharomyces were the major contributors to the production of the volatile flavor of Nanfeng tangerine wine. The results of the present study provide insight into the effects of the core functional microbiota in Nanfeng tangerine wine and can be used to develop effective strategies for improving the quality of fruit wines.

New Helvolic Acid Derivatives with Antibacterial Activities from Sarocladium oryzae DX-THL3, an Endophytic Fungus from Dongxiang Wild Rice (Oryza rufipogon Griff.).

Three new helvolic acid derivatives (named sarocladilactone A (1), sarocladilactone B (2) and sarocladic acid A (3a)), together with five known compounds (6,16-diacetoxy-25-hy- droxy-3,7-dioxy-29-nordammara-1,17(20)-dien-21-oic acid (3b), helvolic acid (4), helvolinic acid (5), 6-desacetoxy-helvolic acid (6) and 1,2-dihydrohelvolic acid (7)), were isolated from the endophytic fungus DX-THL3, obtained from the leaf of Dongxiang wild rice (Oryza rufipogon Griff.). The structures of the new compounds were elucidated via HR-MS, extensive 1D and 2D NMR analysis and comparison with reported data. Compounds 1, 2, 4, 5, 6 and 7 exhibited potent antibacterial activities. In particular, sarocladilactone B (2), helvolinic acid (5) and 6-desacetoxy-helvolic acid (6) exhibited strongly Staphylococcus aureus inhibitory activity with minimum inhibitory concentration (MIC) values of 4, 1 and 4 µg/mL, respectively. The structure-activity relationship (SAR) of these compounds was primarily summarized.

Transcriptome analysis on fructose as the sole carbon source enhancing perylenequinones production of endophytic fungus Shiraia sp. Slf14.

Perylenequinones (PQ), a class of naturally occurring polypeptides, are widely used as a clinical drug for treating skin diseases and as a photodynamic therapy against cancers and viruses. In this study, the effects of different carbon sources on PQ biosynthesis by Shiraia sp. Slf14 were compared, and the underlying molecular mechanism of fructose as the sole carbon to enhance PQ production was investigated by transcriptome analysis. The results indicated that fructose enhanced PQ yield to 1753.64 mg/L, which was 1.73-fold higher than that obtained with glucose. Comparative transcriptome analysis demonstrated that most of the upregulated genes were related to transport systems, energy and central carbon metabolism in Shiraia sp. Slf14 cultured in fructose. The genes involved in glycolysis and pentose phosphate pathways, and encoding citrate synthase, ATP-citrate lyase, and acetyl-CoA carboxylase were substantially upregulated, resulting in increased overall acetyl-CoA and malonyl-CoA production. However, genes involved in gluconeogenesis, glyoxylate cycle pathway, and fatty acid synthesis were significantly downregulated, resulting in higher acetyl-CoA influx for PQ formation. In particular, the putative PQ biosynthetic cluster was upregulated in Shiraia sp. Slf14 cultured in fructose, leading to a significant increase in PQ production. The results of real-time qRT-PCR and related enzyme activities were also consistent with those of transcriptome analysis. These findings provide a remarkable insight into the underlying mechanism of PQ biosynthesis and pave the way for improvements in PQ production by Shiraia sp. Slf14.

The accumulation of rare-earth yttrium ions by Penicillium sp. ZD28.

To obtained fungal resources with excellent tolerance and accumulation capacity to rare earth yttrium ions (Y3+), rare earth ore samples were collected and used for microbial screening. A fungus hyper-resistant to Y3+ was obtained and the effects of the fungus in three physiological states (growth process, mycelial pellets with physiological activity and the fungus powder after being ground) on the Y3+ accumulation were investigated. The Y3+ resistant fungus was identified as Penicillium sp. ZD28, and its mycelium pellets (about 1 mm in diameter) showed poor ability to accumulate Y3+ with an adsorption capacity of less than 81 µmol/g. However, the fungus was able to remove 99% of Y3+ during the growth process, at an initial concentration of less than 600 µM. Bioaccumulation of Y was observed on the cell surface of the ZD28 strain by elemental mapping using scanning electron microscopy-energy dispersive X-ray spectroscopy. The adsorbent (the dry fungal powder) had a remarkable adsorption property for Y3+ that was greater than 455 µmol/g in conditions of 465 µM < [Y3+] < 6382 µM. Penicillium sp. ZD28 has major potential applications in the accumulation of yttrium group rare earth ions. This research has formed a theoretical foundation for the application of this biological method to extract rare earth ions in the mining and smelting of yttrium group rare earth elements.

Dimorphism of Trichosporon cutaneum and impact on its lipid production.

BACKGROUND: Compared to the oleaginous yeast Yarrowia lipolytica, Trichosporon cutaneum can metabolize pentose sugars more efficiently, and in the meantime is more tolerant to inhibitors, which is suitable for lipid production from lignocellulosic biomass. However, this species experiences dimorphic transition between yeast-form cells and hyphae during submerged fermentation, which consequently affects the rheology and mass transfer performance of the fermentation broth and its lipid production. RESULTS: The strain T. cutaneum B3 was cultured with medium composed of yeast extract, glucose and basic minerals. The experimental results indicated that yeast-form morphology was developed when yeast extract was supplemented at 1 g/L, but hyphae were observed when yeast extract supplementation was increased to 3 g/L and 5 g/L, respectively. We speculated that difference in nitrogen supply to the medium might be a major reason for the dimorphic transition, which was confirmed by the culture with media supplemented with yeast extract at 1 g/L and urea at 0.5 g/L and 1.0 g/L to maintain total nitrogen at same levels as that detected in the media with yeast extract supplemented at 3 g/L and 5 g/L. The morphological change of T. cutaneum B3 affected not only the content of intracellular lipids but also their composition, due to its impact on the rheology and oxygen mass transfer performance of the fermentation broth, and more lipids with less polyunsaturated fatty acids such as linoleic acid (C18:2) were produced by the yeast-form cells. When T. cutaneum B3 was cultured at an aeration rate of 1.5 vvm for 72 h with the medium composed of 60 g/L glucose, 3 g/L yeast extract and basic minerals, 27.1 g (dry cell weight)/L biomass was accumulated with the lipid content of 46.2%, and lipid productivity and yield were calculated to be 0.174 g/L/h and 0.21 g/g, respectively. Comparative transcriptomics analysis identified differently expressed genes for sugar metabolism and lipid synthesis as well as signal transduction for the dimorphic transition of T. cutaneum B3. CONCLUSIONS: Assimilable nitrogen was validated as one of the major reasons for the dimorphic transition between yeast-form morphology and hyphae with T. cutaneum, and the yeast-form morphology was more suitable for lipid production at high content with less polyunsaturated fatty acids as feedstock for biodiesel production.

Intrinsic kinetic model of photoautotrophic microalgae based on chlorophyll fluorescence analysis.

As photoautotrophic microorganisms, microalgae feature complex mechanisms of photosynthesis and light energy transfer and as such studying their intrinsic growth kinetics is fairly difficult. In this article, the quantum yield of photochemical reaction was introduced in a study of microalgal kinetics to establish an intrinsic kinetic model of photoautotrophic microalgal growth. The blue-green algae Synechococcus sp. PCC7942 was used to verify the kinetic model developed using chlorophyll fluorescence analysis and growth kinetics determination. Results indicate that the kinetic model can realistically reflect the light energy utilization efficiency of microalgae as well as their intrinsic growth kinetic characteristics. The model and method proposed in this article may be utilized in intrinsic kinetics studies of photoautotrophic microorganisms.

Improved production of antifungal angucycline Sch47554 by manipulating three regulatory genes in Streptomyces sp. SCC-2136.

Sch47554 and Sch47555 are two angucyclines with antifungal activities against various yeasts and dermatophytes from Streptomyces sp. SCC-2136. The sch gene cluster contains several putative regulatory genes. Both schA4 and schA21 were predicted as the TetR family transcriptional regulators, whereas schA16 shared significant similarity to the AraC family transcriptional regulators. Although Sch47554 is the major product of Streptomyces sp. SCC-2136, its titer is only 6.72 mg/L. This work aimed to increase the production of this promising antifungal compound by investigating and manipulating the regulatory genes in the Sch47554 biosynthetic pathway. Disruption of schA4 and schA16 led to a significant increase in the production of Sch47554, whereas the titer was dramatically decreased when schA21 was disrupted. Overexpression of these genes gave opposite results. The highest titer of Sch47554 was achieved in Streptomyces sp. SCC-2136/ΔschA4 (27.94 mg/L), which is significantly higher than the wild type. Our results indicate that SchA4 and SchA16 are repressors, whereas SchA21 acts as an activator. This work thus provides an initial understanding of functional roles of regulatory elements in the biosynthesis of Sch47554. Several efficient producing strains of Sch47554 were constructed by disrupting or overexpressing particular regulatory genes, which can be further engineered for industrial production of this medicinally important molecule.

Identification and heterologous reconstitution of a 5-alk(en)ylresorcinol synthase from endophytic fungus Shiraia sp. Slf14.

A new type III polyketide synthase gene (Ssars) was discovered from the genome of Shiraia sp. Slf14, an endophytic fungal strain from Huperzia serrata. The intron-free gene was cloned from the cDNA and ligated to two expression vectors pET28a and YEpADH2p-URA3 for expression in Escherichia coli BL21(DE3) and Saccharomyces cerevisiae BJ5464, respectively. SsARS was efficiently expressed in E. coli BL21(DE3), leading to the synthesis of a series of polyketide products. Six major products were isolated from the engineered E. coli and characterized as 1,3-dihydroxyphenyl-5-undecane, 1,3-dihydroxyphenyl-5-cis-6'-tridecene,1,3-dihydroxyphenyl-5-tridecane, 1,3-dihydroxyphenyl-5-cis-8'-pentadecene, 1,3-dihydroxyphenyl-5-pentadecane, and 1,3-dihydroxyphenyl-5-cis-10'-heptadecene, respectively, based on the spectral data and biosynthetic origin. Expression of SsARS in the yeast also led to the synthesis of the same polyketide products, indicating that this enzyme can be reconstituted in both heterologous hosts. Supplementation of soybean oil into the culture of E. coli BL21(DE3)/SsARS increased the production titers of 1-6 and led to the synthesis of an additional product, which was identified as 5-(8'Z,11'Z-heptadecadienyl) resorcinol. This work thus allowed the identification of SsARS as a 5-alk(en)ylresorcinol synthase with flexible substrate specificity toward endogenous and exogenous fatty acids. Desired resorcinol derivatives may be synthesized by supplying corresponding fatty acids into the culture medium.

New Insights into the Glycosylation Steps in the Biosynthesis of Sch47554 and Sch47555.

Sch47554 and Sch47555 are antifungal compounds from Streptomyces sp. SCC-2136. The availability of the biosynthetic gene cluster made it possible to track genes that encode biosynthetic enzymes responsible for the structural features of these two angucyclines. Sugar moieties play important roles in the biological activities of many natural products. An investigation into glycosyltransferases (GTs) might potentially help to diversify pharmaceutically significant drugs through combinatorial biosynthesis. Sequence analysis indicates that SchS7 is a putative C-GT, whereas SchS9 and SchS10 are proposed to be O-GTs. In this study, the roles of these three GTs in the biosynthesis of Sch47554 and Sch47555 are characterized. Coexpression of the aglycone and sugar biosynthetic genes with schS7 in Streptomyces lividans K4 resulted in the production of C-glycosylated rabelomycin, which revealed that SchS7 attached a d-amicetose moiety to the aglycone core structure at the C-9 position. Gene inactivation studies revealed that subsequent glycosylation steps took place in a sequential manner, in which SchS9 first attached either an l-aculose or l-amicetose moiety to 4'-OH of the C-glycosylated aglycone, then SchS10 transferred an l-aculose moiety to 3-OH of the angucycline core.

Enhanced production of perylenequinones in the endophytic fungus Shiraia sp. Slf14 by calcium/calmodulin signal transduction.

Perylenequinones (PQ) that notably produce reactive oxygen species upon exposure to visible light are a class of photoactivated polyketide mycotoxins produced by fungal plant pathogens such as Shiraia sp. The involvement of Ca2+/calmodulin (CaM) signalling in PQ biosynthesis was investigated by submerged culturing of Shiraia sp. Slf14, a species that produces hypocrellins HA and HB and elsinochromes EA, EB, and EC. Our results showed that the total content of PQ reached 1894.66 ± 21.93 mg/L under optimal conditions of Ca2+ addition, which represents a 5.8-fold improvement over controls. The addition of pharmacological Ca2+ sensor inhibitors strongly inhibited PQ production, which indicates that Ca2+/CaM signalling regulates PQ biosynthesis. The expression levels of Ca2+ sensor and PQ biosynthetic genes were downregulated following addition of inhibitors but were upregulated upon addition of Ca2+. Inhibition was partially released by external Ca2+ supplementation. Fluo-3/AM experiments revealed that similar cytosolic Ca2+ variation occurred under these conditions. These results demonstrated that Ca2+ signalling via the CaM transduction pathway plays a pivotal role in PQ biosynthesis.

Molecular cloning, expression, and functional analysis of the copper amine oxidase gene in the endophytic fungus Shiraia sp. Slf14 from Huperzia serrata.

Huperzine A (HupA) is a drug used for the treatment of Alzheimer's disease. However, the biosynthesis of this medicinally important compound is not well understood. The HupA biosynthetic pathway is thought to be initiated by the decarboxylation of lysine to form cadaverine, which is then converted to 5-aminopentanal by copper amine oxidase (CAO). In this study, we cloned and expressed an SsCAO gene from a HupA-producing endophytic fungus, Shiraia sp. Slf14. Analysis of the deduced protein amino acid sequence showed that it contained the Asp catalytic base, conserved motif Asn-Tyr-Asp/Glu, and three copper-binding histidines. The cDNA of SsCAO was amplified and expressed in Escherichia coli BL21(DE3), from which a 76 kDa protein was obtained. The activity of this enzyme was tested, which provided more information about the SsCAO gene in the endophytic fungus. Gas Chromatograph-Mass Spectrometry (GC-MS) revealed that this SsCAO could accept cadaverine as a substrate to produce 5-aminopentanal, the precursor of HupA. Phylogenetic tree analysis indicated that the SsCAO from Shiraia sp. Slf14 was closely related to Stemphylium lycopersici CAO. This is the first report on the cloning and expression of a CAO gene from HupA-producing endophytic fungi. Functional characterization of this enzyme provides new insights into the biosynthesis of the HupA an anti-Alzheimer's drug.

Isolation, diversity and acetylcholinesterase inhibitory activity of the culturable endophytic fungi harboured in Huperzia serrata from Jinggang Mountain, China.

Huperzia serrata has many important medicinal properties with proven pharmacological potential. Some of these properties may be mediated by its endophytic fungi. To test this hypothesis, in the present study, we provided a first insights into evaluating the species composition and acetylcholinesterase (AChE) inhibitory activity of the culturable endophytic fungi of H. serrata from the regional at Jinggang Mountain in southeastern China. A total number of 885 fungal isolates distributed across 44 genera and 118 putative species were obtained from 1422 fragments of fine H. serrata roots, stems and leaves base on ITS-rDNA sequences BLAST analysis. The endophytic fungi were phylogenetically diverse and species-rich, with high rate of colonization and isolation. The assemble of endophytic fungi consisted mainly of Ascomycota (97.15%), followed by Basidiomycota (1.92%) and unknown fungal species (0.90%). Colletotrichum (64.29%), Phyllosticta (3.39%), Hypoxylon (2.81%), Xylaria (2.25%) and Nigrospora (2.04%) were the most abundant genera, whereas the remaining genera were infrequent groups. Although, roots yielded low abundance strains, the diverse and species-rich were both higher than that of stems and leaves. In addition, out of the 247 endophytic fungi strains determinated, 221 fungal extracts showed AChE inhibition activities in vitro. Among them, 22 endophytic fungi strains achieved high inhibitory activity (≥50%) on AChE which belongs to 13 genera and five incertae sedis strains. Four endophytic fungi designated as JS4 (Colletotrichum spp.), FL14 (Ascomycota spp.), FL9 (Sarcosomataceae spp.) and FL7 (Dothideomycetes spp.) were displayed highly active (≥80%) against AChE, which the inhibition effects were even more intense than the positive control. Our findings highlight that H. serrata grown in Jinggang Mountain harbors a rich and fascinating endophytic fungus community with potential AChE inhibitory activity, which could further broaden the natural acetylcholinesterase inhibitors resources used for Alzheimer's disease treatment.

[Molecular cloning, expression and characterization of lysine decarboxylase gene of endophytic fungus Shiraia sp. Slf14 from Huperzia serrata].

Objective: Huperzine A (HupA) was approved as a drug for the treatment of Alzheimer's disease. The HupA biosynthetic pathway was started from lysine decarboxylase (LDC), which catalyzes lysine to cadaverine. In this study, we cloned and expressed an LDC gene from a HupA-producing endophytic fungus, and tested LDC activities. Methods: An endophytic fungus Shiraia sp. Slf14 from Huperzia serrata was used. LDC gene was obtained by RT-PCR, and cloned into pET-22b(+) and pET-32a(+) vectors to construct recombinant plasmids pET- 22b-LDC and pET-32a-LDC. These two recombinant plasmids were transformed into E. coli BL21, cultured for 8 h at 24 °C, 200 r/min with 1×10­3 mol/L IPTG into medium to express the LDC proteins, respectively. LDC proteins were purified by Ni2+ affinity chromatography. Catalytic activities were measured by Thin Layer Chromatography. At last, the physicochemical properties and structures of these two LDCs were obtained by bioinformatics software. Results: LDC and Trx-LDC were expressed in E. coli BL21 successfully. SDS-PAGE analysis shows that the molecular weight of LDC and Trx-LDC were 24.4 kDa and 42.7 kDa respectively, which are consistent with bioinformatics analysis. In addition, TLC analysis reveals that both LDC and Trx-LDC had catalytic abilities. Conclusion: This work can provide fundamental data for enriching LDC molecular information and reveal the HupA biosynthetic pathway in endophytic fungi.

Efficient strategy for maintaining and enhancing the huperzine A production of Shiraia sp. Slf14 through inducer elicitation.

Huperzine A (HupA), a naturally occurring lycopodium alkaloid, is a potent, highly specific and reversible inhibitor of acetylcholinesterase and is a potential treatment for Alzheimer's disease. However, isolating HupA from Huperziaceae plants is inefficient; thus, extracting this compound from endophytic fungi may be more controllable and sustainable. However, the large-scale production of this chemical from endophytes is limited by the innate instability of endophytic fungi. In this study, we maintained the stability and viability of the HupA-producing endophytic fungus Shiraia sp. Slf14 and enhanced the HupA titers during fermentation by adding Huperzia serrata extracts (HSE), L-lysine, and acetic acid into the culture as inducers. Adding trace amounts of HupA clearly improved the HupA production of Shiraia sp. Slf14, reaching a maximum content of approximately 40 µg g(-1). Moreover, the addition of HSE and L-lysine promoted HupA production in the flask fermentation. The aforementioned bioprocessing strategy may be potentially applied to other endophytic fungal culture systems for the efficient production of plant secondary metabolites.

Draft Genome Sequence of Streptomyces sp. Strain PRh5, a Novel Endophytic Actinomycete Isolated from Dongxiang Wild Rice Root.

Here, we report the draft genome sequence of Streptomyces sp. strain PRh5 (China Center for Type Culture Collection [CCTCC] number 2013487), which is used to produce nigericin and nocardamine. The genome sequence will allow for the characterization of the molecular mechanisms underlying its beneficial properties.

Whole-Genome Shotgun Assembly and Analysis of the Genome of Shiraia sp. Strain Slf14, a Novel Endophytic Fungus Producing Huperzine A and Hypocrellin A.

Here, we report the draft genome sequence of Shiraia sp. strain Slf14 (China Center for Type Culture Collection [CCTCC] no. 209294), which is used to produce huperzine A and hypocrellin A. The genome sequence will allow for the characterization of the molecular mechanisms underlying its beneficial properties.

[Improved production of microbial lipids in the two-liquid phase fermentation system].

In the present study, we developed a two-liquid phase fermentation system by adding 1% n-dodecane as oxygen-vector to enhance the microbial lipids productivity of Trichosporon fermentans using cassava starch hydrolysate. Results suggest that the oxygen-vector could alleviate the oxygen shortage in flask fermentation. The cell mass and lipids concentration were 101.2 g/L and 50.28 respectively in 2 L fermenter with the presence of 1% n-dodecane. Additionally, gas chromatography analysis also reveals that the microbial lipids produced by T. fermentans contained a higher percentage of saturated fatty acid in the oxygen-vector case.

[Microbial oil production by Trichosporon cutaneum B3 using cassava starch].

Microbial oil, as raw material for biodiesel, can be produced by Trichosporon cutaneum B3 using cassava starch hydrolysate. Batch cultures demonstrated that there was little inhibitory effect with the concentration of cassava starch hydrolysate up to 90 g/L. The favorable initial pH, C/N molar ratio, nitrogen source and its concentration were 6.0, 116, yeast extract and 3.0 g/L, respectively. Under the optimized conditions, dry biomass reached 15.2 g/L and lipid content reached 40.9% after culture for 144 h in flask. Batch cultures in a 2 L stirred-tank fermenter were run for 44 h and resulted in dry biomass, lipid content and lipid yield of 28.7 g/L, 42.8% and 12.27 g/L, respectively. The chemical compositions of biodiesel prepared from lipids of T cutaneum B3 mainly included palmitic acid methyl ester, stearic acid methyl ester, oleic acid methyl ester and linoleic acid methyl ester etc., and its main physicochemical properties were in compliance with relevant national diesel standards. Therefore, the biodiesel prepared from lipids of T cutaneum B3 can serve as a potential fossil fuel alternatives.