[Prediction of Soil Bacterial Community Structure and Function in Minqin Desert-oasis Ecotone Artificial Haloxylon ammodendron Forest].

Exploring the effects of artificial Haloxylon ammodendron forest planting on the structure and function of a desert soil bacterial community provides data reference for soil micro-ecological restoration and land quality improvement in desert oasis transition zones. Illumina high-throughput sequencing technology and PICRUSt2 functional prediction analysis were used to identify and analyze the structure and function of soil bacterial communities, and the Mantel correlation test and RDA analysis were used to explain the physicochemical factors affecting the structure and function of soil bacterial communities. The results showed that:① the soil bacterial OTU number, Chao1 index, and Shannon index were significantly higher in the H. ammodendron forest than in the mobile dune soil, and the PCoA analysis and Adonis test showed significant differences in the soil bacterial community structure between H. ammodendron and mobile dune soil (P=0.001). ② A total of 34 phyla, 89 classes, 174 orders, 262 families, and 432 genera of bacteria were detected in all samples, and the phyla Proteobacteria, Actinobacteria, Cyanobacteria, and Chloroflexi accounted for 76.05% of the relative abundance of soil bacteria, which belonged to the dominant soil bacteria, among which the relative abundance of Actinobacteria in H. ammodendron forest soil was extremely significantly higher than that in mobile dune soil (P < 0.01). ③PICRUSt2 function prediction revealed that the soil bacterial community of H. ammodendron forest included six categories of primary functions and 28 categories of secondary functions, among which the metabolism of carbohydrates, metabolism of amino acids, and metabolism of cofactors and vitamins were all greater than 10% in relative abundance and were the main metabolic functions of H. ammodendron forest soil bacteria. ④ The planting of H. ammodendron forest significantly improved the nutrient content of soil organic matter and other nutrients. Soil pH, organic matter, total nitrogen, and fast-acting phosphorus were the main physicochemical factors affecting the bacterial community, with soil organic matter significantly affecting the soil bacterial community structure (P < 0.05) and metabolic function (P < 0.05). In conclusion, the artificial H. ammodendron forest helped to increase desert soil microbial diversity, increase the relative abundance of soil bacterial metabolic function genes, and improve the desert soil microenvironment.

Epstein-Barr virus infection with non-tumor-associated Anti-N-Methyl-D-Aspartate receptor encephalitis: a case report and review of literature.

To study a case of a middle-aged male with a non-tumor-associated Epstein-Barr virus (EBV) infection associated with Anti-N-methyl-D-aspartate receptor encephalitis (NMDARE), to explore the role of EBV in the pathogenesis of anti-NMDARE. The patient was diagnosed with "Anti-NMDARE, EBV infection" by using Cerebrospinal fluid (CSF) autoimmune encephalitis profile, and Metagenomics Next-Generation Sequencing (mNGS) pathogenic microbial assays, we discuss the relationship between EBV and NMDARE by reviewed literature. EBV infection may trigger and enhance anti-NMDARE, and the higher the titer of NMDAR antibody, the more severe the clinical presentation.

Structure characterization of an exopolysaccharide from a Shiraia-associated bacterium and its strong eliciting activity on the fungal hypocrellin production.

Hypocrellins are fungal perylenequinones (PQs) from Shiraia fruiting bodies and potential photosensitizers for cancer photodynamic therapy. Shiraia fruiting bodies harbor diverse bacterial communities dominated by Pseudomonas. The present study was to characterize the exopolysaccharide (EPS) of P. fulva SB1 which acted as an elicitor to stimulate the PQ accumulation of the host Shiraia. A bacterial EPS named EPS-1 was purified from the culture broth of P. fulva SB1, which consisted of mannose (Man) and glucose (Glc) with an average molecular weight of 9.213 × 104 Da. EPS-1 had (1 â†’ 2)-linked α-mannopyranose (Manp) backbone and side chains of α-D-Manp-(1→ and α-D-Manp-(1 â†’ 6)-ß-D-Glcp-(1 â†’ 6)-α-D-Manp(1 â†’ group attached to the O-6 positions of (1 â†’ 2)-α-D-Manp. EPS-1 at 30 mg/L stimulated both intracellular and extracellular hypocrellin A (HA) by about 3-fold of the control group. The EPS-1 treatment up-regulated the expression of key genes for HA biosynthesis. The elicitation of HA biosynthesis by EPS-1 was strongly dependent on the induced reactive oxygen species (ROS) generation. The results may provide new insights on the role of bacterial EPS in bacterium-fungus interactions and effective elicitation strategy for hypocrellin production in mycelial cultures.

Identification and profiling of the community structure and potential function of bacteria from the fruiting bodies of Sanghuangporus vaninii.

Sanghuangporus sp., a medicinal and edible homologous macrofungus known as 'forest gold', which has good effects on antitumor, hypolipidemia and the treatment of gynecological diseases. However, the natural resources of fruiting body are on the verge of depletion due to its long growth cycle and over exploitation. The growth and metabolism of macrofungi are known to depend on the diverse bacterial community. Here, we characterized the diversity and potential function of bacteria inhabiting in the fruiting body of the most widely applied S. vaninii using a combination method of high-throughput sequencing with pure culturing for the first time, and tested the biological activities of bacterial isolates, of which Illumina NovaSeq provided a more comprehensive results on the bacterial community structure. Total 33 phyla, 82 classes, 195 orders, 355 families, 601 genera and 679 species were identified in the fruiting body, and our results revealed that the community was predominated by the common Proteobacteria, Gammaproteobacteria, Burkholderiales, Methylophilaceae (partly consistent with pure-culturing findings), and was dominated by the genera of distinctive Methylotenera and Methylomonas (yet-uncultured taxa). Simultaneously, the functional analysis showed that companion bacteria were involved in the pathways of carbohydrate transport and metabolism, metabolism of terpenoids and polyketides, cell wall/membrane/envelope biogenesis, etc. Hence, it was inferred that bacteria associated with fruiting body may have the potential to adjust the growth, development and active metabolite production of host S. vaninii combined with the tested results of indole-3-acetic acid and total antioxidant capacity. Altogether, this report first provided new findings which can be inspiring for further in-depth studies to exploit bioactive microbial resources for increased production of Sanghuangporus, as well as to explore the relationship between medicinal macrofungi and their associated endophytes.

Association between coronary artery calcification and cognitive function in a Chinese community-based population.

BACKGROUND: Coronary atherosclerosis and cognitive impairment are both age-related diseases, with similar risk factors. Coronary artery calcium (CAC), a marker of coronary atherosclerosis, may play a role in early detection of individuals prone to cognitive decline. This study aimed to investigate the relationship between CAC and cognitive function, and the capability of CAC to identify participants with a high risk of dementia in a Chinese community-based population. METHODS: A total of 1332 participants, aged 40-80 years and free of dementia from a community located in Beijing were included. All participants completed neurocognitive questionnaires and noncontrast CT examinations. Cognitive performance tests (including verbal memory, semantic fluency, executive function, and global cognitive function tests), the Cardiovascular Risk Factors, Aging, and Incidence of Dementia (CIDE) risk score, and the CAC score (CACS) were evaluated by questionnaires and CT. A CAIDE score ≥ 10 was considered to indicate a high risk of dementia in late-life. Participants were divided into three groups according to CACS (0, 1-399, ≥ 400). RESULTS: After adjusting for risk factors, CACS was significantly associated with verbal memory (r = -0.083, P = 0.003) and global cognitive function (r = -0.070, P = 0.012). The prevalence of a high risk of dementia in the subgroups of CACS = 0, 1-399, and ≥ 400 was 4.67%, 13.66%, and 24.79%, respectively (P < 0.001). Individuals with CACS ≥ 400 had a higher risk of CAIDE score ≥ 10 [OR = 2.30 (1.56, 4.56), P = 0.014] than those with CACS = 0. The receiver-operating characteristic curves showed that the capability of CACS to identify participants with a high risk of dementia was moderate (AUC = 0.70, 95% CI: 0.67-0.72,P < 0.001). CONCLUSIONS: CAC, a marker of subclinical atherosclerosis, was significantly associated with cognitive performance in verbal memory and global cognitive function. CAC had a moderate capability to identify participants with a high risk of dementia, independent of age, education, and other risk factors.

Nitric oxide donor sodium nitroprusside-induced transcriptional changes and hypocrellin biosynthesis of Shiraia sp. S9.

BACKGROUND: Nitric oxide (NO) is a ubiquitous signaling mediator in various physiological processes. However, there are less reports concerning the effects of NO on fungal secondary metabolites. Hypocrellins are effective anticancer photodynamic therapy (PDT) agents from fungal perylenequinone pigments of Shiraia. NO donor sodium nitroprusside (SNP) was used as a chemical elicitor to promote hypocrellin biosynthesis in Shiraia mycelium cultures. RESULTS: SNP application at 0.01-0.20 mM was found to stimulate significantly fungal production of perylenequinones including hypocrellin A (HA) and elsinochrome A (EA). SNP application could not only enhance HA content by 178.96% in mycelia, but also stimulate its efflux to the medium. After 4 days of SNP application at 0.02 mM, the highest total production (110.34 mg/L) of HA was achieved without any growth suppression. SNP released NO in mycelia and acted as a pro-oxidant, thereby up-regulating the gene expression and activity of reactive oxygen species (ROS) generating NADPH oxidase (NOX) and antioxidant enzymes, leading to the increased levels of superoxide anion (O2-) and hydrogen peroxide (H2O2). Gene ontology (GO) analysis revealed that SNP treatment could up-regulate biosynthetic genes for hypocrellins and activate the transporter protein major facilitator superfamily (MFS) for the exudation. Moreover, SNP treatment increased the proportion of total unsaturated fatty acids in the hypha membranes and enhanced membrane permeability. Our results indicated both cellular biosynthesis of HA and its secretion could contribute to HA production induced by SNP. CONCLUSIONS: The results of this study provide a valuable strategy for large-scale hypocrellin production and can facilitate further understanding and exploration of NO signaling in the biosynthesis of the important fungal metabolites.

Nitric Oxide and Hydrogen Peroxide Signaling in Extractive Shiraia Fermentation by Triton X-100 for Hypocrellin A Production.

Shiraia mycelial culture is a promising biotechnological alternative for the production of hypocrellin A (HA), a new photosensitizer for anticancer photodynamic therapy (PDT). The extractive fermentation of intracellular HA in the nonionic surfactant Triton X-100 (TX100) aqueous solution was studied in the present work. The addition of 25 g/L TX100 at 36 h of the fermentation not only enhanced HA exudation to the broth by 15.6-fold, but stimulated HA content in mycelia by 5.1-fold, leading to the higher production 206.2 mg/L, a 5.4-fold of the control on day 9. After the induced cell membrane permeabilization by TX100 addition, a rapid generation of nitric oxide (NO) and hydrogen peroxide (H2O2) was observed. The increase of NO level was suppressed by the scavenger vitamin C (VC) of reactive oxygen species (ROS), whereas the induced H2O2 production could not be prevented by the NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (PTIO), suggesting that NO production may occur downstream of ROS in the extractive fermentation. Both NO and H2O2 were proved to be involved in the expressions of HA biosynthetic genes (Mono, PKS and Omef) and HA production. NO was found to be able to up-regulate the expression of transporter genes (MFS and ABC) for HA exudation. Our results indicated the integrated role of NO and ROS in the extractive fermentation and provided a practical biotechnological process for HA production.

[Altitudinal differentiation in the radial growth of Betula platyphylla and its response to climate in cold temperate forest: A case of Oakley Mountain, Northeast China].

Climate change has profound impacts on the structure and function of forest ecosystem. Under the background of climate warming, the growth patterns and climate responses of different species in different environments determine the development and stability of ecosystems. Using the method of tree chronology with ring width data of Betula platyphylla in Oakley Mountain of Daxing'an Mountains, we examined the relationship between the growth climate response of the pioneer B. platyphylla in larch forest and temperature rise and altitudinal changes. The results showed that climate warming resulted in the differentiation on B. platyphylla in different altitude. The radial growth of B. platyphylla increased significantly at the low altitude area (1050 m), while there were little changes at the relatively high altitude area (1250 m). Before the time of rapid warming (1980), the radial growth of B. platyphylla on two altitude areas were significantly stressed by low temperature in winter (October to February). During the period of rapid warming (1981-2010), low temperature stress in winter did not affect tree growth, with the growing season (May to July) temperature becoming the main limiting factors to the radial growth of B. platyphylla at the relatively high altitude areas. The growth of B. platyphylla increased in the low altitude stands with favorable water and heat conditions. The distribution of B. platyphylla may generally spread to high altitude sites in the study area under climate warming.

Bacteria Associated With Shiraia Fruiting Bodies Influence Fungal Production of Hypocrellin A.

Hypocrellin A (HA) is a natural red perylenequinone pigment from Shiraia fruiting body, which was used clinically on various skin diseases and developed as a photodynamic therapy agent against cancers. The fruiting bodies may harbor a diverse but poorly understood microbial community. In this study, we characterized the bacterial community of Shiraia fruiting body using a combination of culture-based method and Illumina high-throughput sequencing, and tested the involvement of some companion bacteria in fungal HA production using the fungal-bacterial confrontation assay. Our results revealed that the bacterial community in the fruiting body was dominated by Bacillus and Pseudomonas. Some Pseudomonas isolates such as P. fulva, P. putida, and P. parafulva could stimulate fungal HA accumulation by Shiraia sp. S9. The bacterial treatment of P. fulva SB1 up-regulated the expression of polyketide synthase (PKS) for HA biosynthesis and transporter genes including ATP-binding cassette (ABC) and major facilitator superfamily transporter (MFS) for HA exudation. After the addition of live P. fulva SB1, the mycelium cultures of Shiraia sp. S9 presented a higher HA production (225.34 mg/L), about 3.25-fold over the mono-culture. On the other hand, B. cereus was capable of alleviating fungal self-toxicity from HA via down-regulation of HA biosynthetic genes or possible biodegradation on HA. To our knowledge, this is the first report on the diversified species of bacteria associated with Shiraia fruiting bodies and the regulation roles of the companion bacteria on fungal HA biosynthesis. Furthermore, the bacterial co-culture provided a good strategy for the enhanced HA production by Shiraia.

Inducing perylenequinone production from a bambusicolous fungus Shiraia sp. S9 through co-culture with a fruiting body-associated bacterium Pseudomonas fulva SB1.

BACKGROUND: Fungal perylenequinonoid (PQ) pigments from Shiraia fruiting body have been well known as excellent photosensitizers for medical and agricultural uses. The fruiting bodies are colonized by a diverse bacterial community of unknown function. We screened the companion bacteria from the fruiting body of Shiraia sp. S9 and explored the bacterial elicitation on fungal PQ production. RESULTS: A bacterium Pseudomonas fulva SB1 isolated from the fruiting body was found to stimulate the production of fungal PQs including hypocrellins A, C (HA and HC), and elsinochromes A-C (EA, EB and EC). After 2 days of co-cultures, Shiraia mycelium cultures presented the highest production of HA (325.87 mg/L), about 3.20-fold of that in axenic culture. The co-culture resulted in the induction of fungal conidiation and the formation of more compact fungal pellets. Furthermore, the bacterial treatment up-regulated the expression of polyketide synthase gene (PKS), and activated transporter genes of ATP-binding cassette (ABC) and major facilitator superfamily transporter (MFS) for PQ exudation. CONCLUSIONS: We have established a bacterial co-culture with a host Shiraia fungus to induce PQ biosynthesis. Our results provide a basis for understanding bacterial-fungal interaction in fruiting bodies and a practical co-culture process to enhance PQ production for photodynamic therapy medicine.

Enhanced Production of Hypocrellin A in Submerged Cultures of Shiraia bambusicola by Red Light.

Hypocrellin A (HA), a promising photosensitizer for anticancer photodynamic therapy (PDT), is a fungal perylenequinone pigment from the fruiting body of Shiraia bambusicola, a traditional Chinese medicine for treating skin diseases. The mycelial cultures are becoming a biotechnological alternative for HA production. In this study, light of different wavelengths was investigated to develop an effective eliciting strategy for HA production in the cultures. Under red LED light (627 nm) at 200 lux, the maximum HA production (175.53 mg L-1 ) in mycelium cultures was reached after 8 days, about 3.82-fold of the dark control. Red light not only promoted HA biosynthesis in mycelia (intracellular HA), but also stimulated HA secretion into the medium (extracellular HA). We found 14 of 310 transcripts differentially expressed under red light treatment were possible candidate genes for HA biosynthetic pathway. Gene ontology (GO) analysis revealed that red light treatment could change the gene expressions responsible for HA biosynthesis and the transmembrane activity, suggesting both intracellular HA and its secretion could contribute to the enhancement of total HA production in the cultures. The results provided new insights of red light elicitation and effective strategy for HA production in mycelium cultures.

Transcriptomic analysis of Lycium ruthenicum Murr. during fruit ripening provides insight into structural and regulatory genes in the anthocyanin biosynthetic pathway.

Fruit development in Lycium ruthenicum Murr. involves a succession of physiological and biochemical changes reflecting the transcriptional modulation of thousands of genes. Although recent studies have investigated the dynamic transcriptomic responses during fruit ripening in L. ruthenicum, most have been limited in scope, and thus systematic data representing the structural genes and transcription factors involved in anthocyanin biosynthesis are lacking. In this study, the transcriptomes of three ripening stages associated with anthocyanin accumulation, including S1 (green ripeness stage), S2 (skin color change) and S3 (complete ripeness stage) in L. ruthenicum were investigated using Illumina sequencing. Of a total of 43,573 assembled unigenes, 12,734 were differentially expressed during fruit ripening in L. ruthenicum. Twenty-five significantly differentially expressed structural genes (including PAL, C4H, 4CL, CHS, CHI, F3H, F3'H, F3'5'H, DFR, ANS and UFGT) were identified that might be associated with anthocyanin biosynthesis. Additionally, several transcription factors, including MYB, bHLH, WD40, NAC, WRKY, bZIP and MADS, were correlated with the structural genes, implying their important interaction with anthocyanin biosynthesis-related genes. Our findings provide insight into anthocyanin biosynthesis and regulation patterns in L. ruthenicum and offer a systematic basis for elucidating the molecular mechanisms governing anthocyanin biosynthesis in L. ruthenicum.

Effects of 5-Azacytidine on Growth and Hypocrellin Production of Shiraia bambusicola.

Hypocrellins, fungal perylenequinones of Shiraia bambusicola are developed as important photodynamic therapy agents against cancers and viruses. Due to the limitation of the wild resources, the mycelium culture is a promising alternative for hypocrellin production. As DNA methylation has profound effects on fungal growth, development and secondary metabolism, we used both McrBC cleavage and HPLC analysis to reveal the status of DNA methylation of S. bambusicola mycelium. We found that DNA methylation is absent in mycelia, but DNA methylation inhibitor 5-azacytidine (5-AC) still induced the fluffy phenotype and decreased hypocrellin contents significantly. Simultaneously, a total of 4,046 differentially expressed genes were induced by 5-AC, including up-regulated 2,392 unigenes (59.12%) and down-regulated 1,654 unigenes (40.88%). Gene ontology analysis showed 5-AC treatment changed expression of genes involved in membrane composition and oxidation-reduction process. The fluffy phenotype in 5-AC-treated S. bambusicola was closely related to strong promotion of developmental regulator WetA and the repression of the sexual developmental actor VeA and LaeA. It was a surprise finding that 5-AC reduced reactive oxygen species (ROS) production significantly in the mycelia via the inhibition of NADPH oxidase gene (NOX) expression and NOX activity. With the treatment of vitamin C and H2O2, we found that the reduced ROS generation was involved in the down-regulated expression of key genes for hypocrellin biosynthesis and the decreased hypocrellin production. To our knowledge, this is the first attempt to examine DNA methylation level in S. bambusicola. Our results suggested that the mediation of ROS generation could not be ignored in the study using 5-AC as a specific DNA methylation inhibitor.

Deciphering transcriptome profiles of tetraploid Artemisia annua plants with high artemisinin content.

To investigate on the effects of autopolyploidization on growth and artemisinin biosynthesis in Artemisia annua, we performed a comprehensive transcriptomic characterization of diploid and induced autotetraploid A. annua. The polyploidization treatment not only enhanced photosynthetic capacity and endogenous contents of indole-3-acetic acid (IAA), abscisic acid (ABA) and jasmonic acid (JA), oxidative stress, but increased the average level of artemisinin in tetraploids from 42.0 to 63.6%. The obvious phenotypic alterations in tetraploids were observed including shorter stems, larger size of stomata and glandular secretory trichomes (GSTs), larger leaves, more branches and roots. A total of 8763 (8.85%) differentially expressed genes (DEGs) were identified in autotetraploids and mainly involved in carbohydrate metabolic processes, cell wall organization and defense responses. Both the up-regulated expression of DNA methylation unigenes and enhanced level of DNA methylation in autotetraploids indicated a possible role of DNA methylation on transcriptomic remodeling and phenotypic alteration. The up-regulated genes were enriched in response to extracellular protein biosynthesis, photosynthesis and hormone stimulus for cell enlargement and phenotypic alteration. The genomic shock induced by chromosome duplication stimulated the expression of transcripts related to oxidative stress, biosynthesis and signal transduction of ABA and JA, and key enzymes in artemisinin biosynthetic pathway, leading to the increased accumulation of artemisinin. This is the first transcriptomic research that identifies DEGs involved in the polyploidization of A. annua. The results provide novel information for understanding the complexity of polyploidization and for further identification of the factors and genes involve in artemisinin biosynthesis.

Improved hypocrellin A production in Shiraia bambusicola by light-dark shift.

Hypocrellin A (HA) is a major bioactive perylenequinone from the fruiting body of Shiraia bambusicola used for the treatment of skin diseases and developed as a photodynamic therapy (PDT) agent against cancers and viruses. The mycelial culture of S. bambusicola under dark is a biotechnological alternative for HA production but with low yield. In this study, light and dark conditions were investigated to develop effective elicitation on HA production in the cultures. Our results showed the constant light at 200 lx stimulated HA production without any growth retardation of mycelia. A light/dark shift (24: 24 h) not only increased HA content in mycelia by 65%, but stimulated HA release into the medium with the highest total HA production 181.67 mg/L on day 8, about 73% increase over the dark control. Moreover, light/dark shifting induced the formation of smaller and more compact fungal pellets, suggesting a new effective strategy for large-scale production of HA in mycelium cultures. The light/dark shift up-regulated the expression levels of two reactive oxygen species (ROS) related genes including superoxide-generating NADPH oxidase (Nox) and cytochrome c peroxidase (CCP), and induced the generation of ROS. With the treatment of vitamin C, we found that ROS was involved in the up-regulated expression of key biosynthetical genes for hypocrellins and improved HA production. These results provide a basis for understanding the influence of light/dark shift on fungal metabolism and the application of a novel strategy for enhancing HA production in submerged Shiraia cultures.

Efficient degradation of triclosan by an endophytic fungus Penicillium oxalicum B4.

Triclosan (TCS), a widely used antimicrobial and preservative agent, is an emerging contaminant in aqueous and soil environment. Microbial degradation of TCS has not been reported frequently because of its inhibition of microbe growth. To explore the new microbial resources for TCS biodegradation, fungal endophytes were isolated and screened for the degradation potential. The endophytic strain B4 isolated from Artemisia annua L. showed higher degradation efficiency and was identified as Penicillium oxalicum based on its morphology and ITS sequences of ribosomal DNA. In both medium and synthetic wastewater, TCS (5 mg/L) was almost completely degraded within 2 h by the strain B4. The high capacity of TCS uptake (127.60 ± 8.57 mg/g dry weight, DW) of fungal mycelium was observed during the first 10 min after TCS addition. B4 rapidly reduced initial content (5.00 mg/L) of TCS to 0.41 mg/L in medium in 10 min. Then, the accumulation of TCS in mycelium was degraded from 0.45 to 0.05 mg/g DW after 1-h treatment. The degradation metabolites including 2-chlorohydroquinone, 2, 4-dichloropheno, and hydroquinone were found to be restrained in mycelia. The end products of the biodegradation in medium showed no toxicity to Escherichia coli. The new characteristics of high adsorption, fast degradation, and low residual toxicity highlight the potential of endophytic P. oxalicum B4 in TCS bioremediation.

Transcriptomic responses involved in enhanced production of hypocrellin A by addition of Triton X-100 in submerged cultures of Shiraia bambusicola.

The addition of surfactant is a useful strategy to enhance the product yield in submerged fermentation process. In this study, we sought to explore the mechanism for the elicitation of Triton X-100 on production of hypocrellin A (HA) in cultures of Shiraia bambusicola through transcriptomic analysis. Triton X-100 at 2.5% (w/v) not only induced HA biosynthesis in mycelia, but also stimulated the release of HA into the medium. We found 23 of 2463 transcripts, possible candidate genes for HA biosynthesis under Triton X-100 induction. Gene ontology (GO) analysis showed Triton X-100 treatment changed expression of genes involved in transmembrane transport and oxidation-reduction process, indicating that enhanced HA production was mainly due to both elicited biosynthesis in mycelium and the increased membrane permeability for HA release. These data provided new insights into elicitation of surfactants in submerged cultures of fungi.

Enhanced production of hypocrellin A by ultrasound stimulation in submerged cultures of Shiraia bambusicola.

Hypocrellin A (HA), a naturally occurring fungal perylenequinone, is widely used in clinic to treat skin diseases and developed as a photodynamic therapy (PDT) agent against cancers. In this study, a low intensity ultrasound (US, 0.28W/cm2 at 40kHz) was conducted thrice of repeated US exposure (5-min) with an interval of 12h to stimulate HA production of Shiraia bambusicola after 72h of the initial submerged cultures. US not only increased the content of HA by 177.2% in mycelia, but stimulated the release of HA into the medium with the highest total production of HA (247.67mg/L) on day 8. US could result in the decreased pellet diameter, the enhanced membrane permeability, the alternation of membrane compounds and reactive oxygen species (ROS) generation. Furthermore, the ultrasonic treatment up-regulated the expression of some HA biosynthetic genes including polyketide synthase gene (PKS), O-methyltransferase gene (Omef), O-methyltransferase/FAD-dependent monooxygenase (Mono) and FAD/FMN-dependent oxidoreductase gene (FAD), and activated major facilitator superfamily transporter gene (MFS) for HA exudation. The enhancement of HA production was mainly due to both the stimulated cellular biosynthesis and the enhanced fungal exudation of HA. These results provide a basis for understanding the US elicitation and a valuable strategy for enhancing HA production in submerged Shiraia cultures.

Cytoprotective role of nitric oxide in HepG2 cell apoptosis induced by hypocrellin B photodynamic treatment.

Hypocrellin B (HB), a natural perylenequinone pigment, has been successfully employed in the photodynamic therapy (PDT) in a variety of human cancer cells due to its high singlet oxygen yield. To investigate the generation of nitric oxide (NO) and its role on cancer cell death induced by PDT, we used human hepatocellular carcinoma (HepG2) cells and HB as a photosensitizer. HB/light treatment decreased the growth of HepG2 cells in a dose-dependent manner with an IC50 of 3.10µM, activated caspase-3, -9 and induced apoptosis in HepG2 cells. It was found that exposure of the cells to HB/light resulted in inducible nitric oxide synthase (iNOS) activation and followed by significant increase in NO generation. Incubating cells with a NOS inhibitor N(ω)-monomethyl-l-arginine (l-NMMA) and an NO scavenger 2-(4-carboxyphenyl)-4, 4, 5, 5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO) enhanced HB/light-induced caspase-3, -9 activation and apoptosis significantly while decreasing DAF fluorescence-assessed NO generation substantially. Cells could be rescued from HB/light-induced apoptosis by an exogenous NO donor, sodium nitroprusside (SNP). Our findings suggested that induced NO was acting cytoprotectively and PDT efficacy of HB could be improved by using pharmacological modulators of NO or NOS.

Antioxidant and DNA Damage Protecting Activity of Exopolysaccharides from the Endophytic Bacterium Bacillus cereus SZ1.

An endophytic bacterium was isolated from the Chinese medicinal plant Artemisia annua L. The phylogenetic and physiological characterization indicated that the isolate, strain SZ-1, was Bacillus cereus. The endophyte could produce an exopolysaccharide (EPS) at 46 mg/L. The 1,1-diphenyl-2-picrylhydracyl (DPPH) radical scavenging activity of the EPS reached more than 50% at 3-5 mg/mL. The EPS was also effective in scavenging superoxide radical in a concentration dependent fashion with an EC50 value of 2.6 mg/mL. The corresponding EC50 for scavenging hydroxyl radical was 3.1 mg/mL. Moreover, phenanthroline-copper complex-mediated chemiluminescent emission of DNA damage was both inhibited and delayed by EPS. The EPS at 0.7-1.7 mg/mL also protected supercoiled DNA strands in plasmid pBR322 against scission induced by Fenton-mediated hydroxyl radical. The preincubation of PC12 cells with the EPS prior to H2O2 exposure increased the cell survival and glutathione (GSH) level and catalase (CAT) activities, and decreased the level of malondialdehyde (MDA) and lactate dehydrogenase (LDH) activity in a dose-dependent manner, suggesting a pronounced protective effect against H2O2-induced cytotoxicity. Our study indicated that the EPS could be useful for preventing oxidative DNA damage and cellular oxidation in pharmaceutical and food industries.