Chuan-Chao Dai

Degradation of N-heterocyclic indole by a novel endophytic fungus Phomopsis liquidambari.

A broad-spectrum endophytic Phomopsis liquidambari, was used to degrade environmental pollutant indole. In the condition of using indole as sole carbon and nitrogen source, the optimum concentration of indole supplied was determined to be 100 mg L(-1), with 41.7% ratio of indole degradation within 120 h. Exogenous addition of plant litter significantly increased indole degradation to 99.1% within 60 h. Indole oxidation to oxindole and isatin were the key steps limiting indole degradation. Plant litter addition induced fungus to produce laccase and LiP to non-specific oxidize indole. The results of fungal metabolites pathway through HPLC-MS and NMR analysis showed that indole was firstly oxidized to oxindole and isatin, and deoxidated to indolenie-2-dione, then hydroxylated to 2-dioxindole, which pyridine ring were cleaved through C-N position and changed to 2-aminobenzoic acid. Such metabolic pathway was similar with bacterial degradation of indole-3-acetic acid in plant.

Involvement of abscisic acid and salicylic acid in signal cascade regulating bacterial endophyte-induced volatile oil biosynthesis in plantlets of Atractylodes lancea

The enormous biological diversity of endophytes, coupled with their potential to enhance the production of bioactive metabolites in plants, has driven research efforts focusing on endophytes. However, limited information is available on the impacts of bacterial endophytes on plant secondary metabolism and signaling pathways involved. This work showed that an endophytic Acinetobacter sp. ALEB16, capable of activating accumulation of plant volatile oils, also induced abscisic acid (ABA) and salicylic acid (SA) production in Atractylodes lancea. Pre-treatment of plantlets with biosynthetic inhibitors of ABA or SA blocked the bacterium-induced volatile production. ABA inhibitors suppressed not only the bacterium-induced volatile accumulation but also the induced ABA and SA generation; nevertheless, SA inhibitors did not significantly inhibit the induced ABA biosynthesis, implying that SA acted downstream of ABA production. These results were confirmed by observations that exogenous ABA and SA reversed the inhibition of bacterium-induced volatile accumulation by inhibitors. Transcriptional activities of genes in sesquiterpenoid biosynthesis also increased significantly with bacterium, ABA and SA treatments. Mevalonate pathway proved to be the main source of isopentenyldiphosphate for bacterium-induced sesquiterpenoids, as assessed in experiments using specific terpene biosynthesis inhibitors. These results suggest that Acinetobacter sp. acts as an endophytic elicitor to stimulate volatile biosynthesis of A. lancea via an ABA/SA-dependent pathway, thereby yielding additional insight into the interconnection between ABA and SA in biosynthesis-related signaling pathways.

Biocatalysis and biotransformation of resveratrol in microorganisms

Resveratrol, a major stilbene phytoalexin, is a valuable polyphenol that has been recognized for its benefits to human health. Resveratrol has antioxidant and antitumor effects and promotes longevity. It is used in medicine, health care products, cosmetics, and other industries. Therefore, a sustainable source for resveratrol production is required. This review describes the metabolic engineering of microorganisms, the biotransformation and biosynthesis of endophytes and the oxidation or degradation of resveratrol. We compare various available methods for resveratrol production, and summarize the practical challenges facing the microbial production of resveratrol. The future research direction for resveratrol is also discussed.

Degradation of a model pollutant ferulic acid by the endophytic fungus Phomopsis liquidambari.

Biodegradation of ferulic acid, by an endophytic fungus called Phomopsis liquidambari was investigated in this study. This strain can use ferulic acid as the sole carbon for growth. Both in mineral salt medium and in soil, more than 97% of added ferulic acid was degraded within 48 h. The metabolites were identified and quantified using GC-MS and HPLC-MS. Ferulic acid was first decarboxylated to 4-vinyl guaiacol and then oxidized to vanillin and vanillic acid, followed by demethylation to protocatechuic acid, which was further degraded through the β-ketoadipate pathway. During degradation, ferulic acid decarboxylase, laccase and protocatechuate 3,4-dioxygenase activities and their gene transcription levels were significantly affected by the variation of substrate and product concentrations. Moreover, ferulic acid degradation was determined to some extent by P. liquidambari laccase. This study is the first report of an endophytic fungus that has a great potential for practical application in ferulic acid-contaminated environments.

Variation of soil enzyme activities and microbial community structure in peanut monocropping system in subtropical China

The changes in soil microbial community are supposed to be one of key factors for peanut yield decline in long-term continuous monocropping systems. A series of peanut fields, where peanut were continuously monocropped for 3, 6, 10 and 15 years in subtropical China, were selected to investigate the effect of continuous monocropping on peanut yield, soil microbial community structure and enzyme activity. Peanut yield, urease and invertase activities decreased with time, but the activity of polyphenol oxidase decreased in the first several years and then increased. The results of both culture dependent methods and phospholipid fatty acids (PLFA) analysis showed that bacteria (Gram-positive and Gramnegative bacteria) and actinomycetes decreased, while fungal colony-forming units (CFUs) increased with time. Canonical Correspondence Analysis (CCA) showed that with increasing peanut monocropping years, soil microbial structure became more correlative with fungi compositions. The proportion of bacteria in total PLFA decreased from 67.4% to 53.0%, meanwhile the proportion of fungi was increased from 16.9% to 32.8%. Denaturing gradient gel electrophoresis (DGGE) analysis indicated that bacteria diversity decreased and fungi diversity increased with time, and changes in fungi diversity were much greater than those of bacteria.

Consequences of antagonistic interactions between endophytic fungus and bacterium on plant growth and defense responses in Atractylodes lancea.

Many studies have examined pair‐wise interactions between plants and endophytes, while overlooking the interplays among multiple endosymbionts and their combined impacts on hosts. In this study, Atractylodes lancea plantlets were inoculated with endophytic fungus Acremonium strictum AL16, or endophytic bacterium Acinetobacter sp., or both, to investigate the impacts of the three‐way symbiosis on the host. Our results showed that defense‐related responses of the co‐inoculated plantlets were delayed and weakened relative to plantlets with single inoculants, but no detrimental effects on phyto‐physiology (growth, photosynthesis) were observed after combined inoculations. Quantitative PCR analysis verified a decrease in AL16 colonization density within plants after co‐inoculation with the endobacteria. An in vitro assay was then performed to elucidate the suppressed plant defense responses and reduced fungal colonization by dual inoculation. The data showed that the presence of Acinetobacter sp. reduced AL16 colony diameter and spore germination rate without negatively affecting fungal morphology. Additionally, direct hyphal attachment of the bacterium to AL16 in vitro was visualized by scanning electronic microscopy. Therefore, we propose that a balanced and compatible symbiosis might require constraints conferred by the antagonistic endophyte Acinetobacter sp. on the fungus AL16 in the tripartite endophytic bacterium–fungus–plant system.

Plant symbionts: keys to the phytosphere

The exterior and interior of plants, aboveground and belowground, comprise a complex plant micro-ecosystem, known in recent years as the phytosphere. There are three components: the phyllosphere, endosphere, and rhizosphere. Although in comparison with other ecosystems the phytosphere is small, it similarly includes a great variety of functional microbes. Among these are certain microbes that live in symbiotic relationships with plants; these microbes are known as plant symbionts. Recent research has shown that these symbionts have tremendous effects on plant growth, confer resistance to abiotic stresses and pathogens, aid in the accumulation of metabolites, and have crucial relationships with other plant-associated microbes in the phytosphere. We review the ecological effects of plant symbionts on other microbes, and interactions between plant symbionts in the phytosphere. In addition, we discuss internal mechanisms and suggest future hot spots for research.

Survival of a novel endophytic fungus Phomopsis liquidambari B3 in the indole-contaminated soil detected by real-time PCR and its effects on the indigenous microbial community.

The recently isolated fungal strain Phomopsis liquidambari B3 can degrade high concentrations of indole, indicating its potential for the bioremediation of indole-contaminated soil. In this study, a specific real-time PCR was developed to detect the survival of P. liquidambari B3 in soil. Subsequently, degradation activity of strain B3 and its effects on indigenous microbial community were analyzed. Results showed the amount of P. liquidambari B3 genomic DNA increased to a maximum 5.67 log (pgg(-1) dry soil) 10 days after inoculation of 5.04 log (pgg(-1) dry soil), and then gradually decreased with time and after 40 days it was below the detection limit. By the end of the experiment (day 40), bioaugmented microsoms showed a 93.7% decrease in indole, while the values for biostimulated and control microcosms were much lower. Higher microbial biomass and enzyme activities were observed in bioaugmented soil. Denaturing gradient gel electrophoresis analysis showed bioaugmentation increased richness of resident microbial community. These results indicate that P. liquidambari B3 is effective for the remediation of indole-contaminated soil and also provides valuable information about the behavior of the inoculant population during bioremediation, which could be directly used in the risk assessment of inoculant population and optimization of bioremediation process.

Cross-Talk Between Calcium–Calmodulin and Brassinolide for Fungal Endophyte-Induced Volatile Oil Accumulation of Atractylodes lancea Plantlets

Using pharmacological and biochemical approaches, the signalling pathways between calcium (Ca2+)–calmodulin (CaM), brassinolide (BL), and nitric oxide (NO) for fungal endophyte-induced volatile oil accumulation were investigated in Atractylodes lancea plantlets. Gilmaniella sp. AL12 inoculation elevated the concentrations of BL, CaM, and [Ca2+]cyt, expression of the calmodulin 1 (CaM1) gene, and the levels of volatile oils. Treatment with AL12 or exogenous BL led to significant increases in the levels of cytosolic Ca2+ and CaM and CaM1 expression in plantlets. However, the upregulation of BL was almost completely blocked by pretreatments with CaM antagonists and Ca2+ channel blockers. Pretreatment with a BL inhibitor, brassinazole (BRz), did not influence the increase in levels of CaM induced by the endophyte. CaCl2-induced increases in NO generation, CaM antagonists, and Ca2+ channel blockers were able to suppress NO production, and the NO-specific scavenger was not able to suppress the generation of [Ca2+]cyt in plantlets. Exogenous BL was not able to induce NO generation, and BRz had no effect on NO generation. Our results suggest that Ca2+–CaM induced by this endophyte mediates NO generation and BL concentration, and also functions downstream of BL signalling, resulting in the upregulation of volatile oil accumulation in A. lancea plantlets.

Endophytic fungus Phomopsis liquidambari and different doses of N-fertilizer alter microbial community structure and function in rhizosphere of rice.

Microbial community structure and functions of rhizosphere soil of rice were investigated after applying low and high doses of nitrogenous fertilizer and Phomopsis liquidambari. Average well color development, substrate richness, catabolic diversity and soil enzymes activities varied after applying N-fertilizer and P. liquidambari and were greater in P. liquidambari treated soil than only N-fertilization. Multivariate analysis distinctly separated the catabolic and enzymes activity profile which statistically proved alteration of microbial functional diversity. Nitrogen fertilizer altered microbial community structure revealed by the increased content of total PLFAs, specific subgroup marker PLFAs except fungal PLFAs and by the decreased ratio of G+/G−, sat/monunsat, iso/anteiso, F/B except trans/cis while P. liquidambari inoculation enhanced N-fertilization effect except increased fungal PLFA and decreased trans/cis. PCA using identified marker PLFAs revealed definite discrimination among the treatments which further statistically confirmed structural changed of microbial community. Nitrogenase activity representative of N-fixing community decreased in N-fertilizer treatment while P. liquidambari inoculation increased. In short, application of P. liquidambari with low doses of N-fertilizer improved rice growth and reduced N-fertilizer requirement by increasing enzymes activities involved in C, N and P cycling, structural and functional diversity of microbes, nitrogenase activity involved in N2 fixation and accumulation of total-N.