Maolin Wang

Pilot‐scale comparison of four duckweed strains from different genera for potential application in nutrient recovery from wastewater and valuable biomass production

The application potential of four duckweed strains from four genera, Wolffia globosa 0222, Lemna japonica 0223, Landoltia punctata 0224 and Spirodela polyrhiza 0225, were compared in four parallel pilot-scale wastewater treatment systems for more than 1 year. The results indicated that each duckweed strain had unique potential advantages. Unlike L. japonica 0223 and La. punctata 0224, which grow throughout the year, S. polyrhiza 0225 and W. globosa 0222 do not survive cold weather. For year round performance, L. japonica 0223 was best not only in dry biomass production (6.10 g·m(-2) ·day(-1) ), but also in crude protein (35.50%), total amino acid (26.83%) and phosphorus (1.38%) content, plus recovery rates of total nitrogen (TN), total phosphorus (TP) and CO2 (0.31, 0.085 and 7.76 g·m(-2) ·day(-1) , respectively) and removal rates of TN and TP (0.66 and 0.089 g·m(-2) ·day(-1) , respectively). This strongly demonstrates that L. japonica 0223 performed best in wastewater treatment and protein biomass production. Under nutrient starvation conditions, La. punctata 0224 had the highest starch content (45.84%), dry biomass production (4.81 g·m(-2) ·day(-1) ) and starch accumulation (2.9 g·m(-2) ·day(-1) ), making it best for starch biomass production. W. globosa 0222 and S. polyrhiza 0225 showed increased flavonoid biomass production, with higher total flavonoid content (5.85% and 4.22%, respectively) and high dominant flavonoids (>60%). This study provides useful information for selecting the appropriate local duckweed strains for further application in wastewater treatment and valuable biomass production.

The influence of duckweed species diversity on biomass productivity and nutrient removal efficiency in swine wastewater

The effect of temperature, light intensity, nitrogen and phosphorus concentrations on the biomass and starch content of duckweed (Landoltia punctata OT, Lemna minor OT) in monoculture and mixture were assessed. Low light intensity promoted more starch accumulation in mixture than in monoculture. The duckweed in mixture had higher biomass and nutrient removal efficiency than those in monoculture in swine wastewater. Moreover, the ability of L. punctata C3, L. minor C2, Spirodela polyrhiza C1 and their mixtures to recovery nutrients and their biomass were analyzed. Results showed that L. minor C2 had the highest N and P content, while L. punctata C3 had the highest starch content, and the mixture of L. punctata C3 and L. minor C2 had the greatest nutrient removal rate and the highest biomass. Compared with L. punctata C3 and L. minor C2 in monoculture, their biomass in mixture increased by 17.0% and 39.8%, respectively.

The influence of duckweed species diversity on ecophysiological tolerance to copper exposure

In excess, copper is toxic to plants. In the plants, Landoltia punctata and Lemna minor grown in mixed and monoculture, the effects of exposure to varying concentrations of copper (0.01, 0.1, 0.5 and 1mgL(-1) Cu) for seven days were assessed by measuring changes in the chlorophyll, protein and malondialdehyde (MDA) content, catalase (CAT), superoxide dismutase (SOD) and ascorbate peroxidase (APX) activity. According to results, Cu levels in plants increased with increasing Cu concentration. The level of photosynthetic pigments and crude proteins decreased only upon exposure to high Cu concentrations. However, the starch and malondialdehyde (MDA) content increased. These results suggested a stress alleviation that was possibly the result of antioxidants such as CAT and SOD, the activities of which increased with increasing Cu levels. APX activity increased in L. punctata, but decreased in L. minor, under monoculture or mixed culture conditions. In addition, the duckweed in mixed culture exhibited increased antioxidant enzyme activities which provide increased resistance to copper in moderate copper concentrations. As the copper concentration increased, the duckweed in the mixed culture limited the uptake of copper to avoid toxicity.

Development and application of a novel screening method and experimental use of the mutant bacterial strain Clostridium beijerinckii NCIMB 8052 for production of butanol via fermentation of fresh cassava

Classic chemical mutagenesis has a demonstrated potential to create a strain of Clostridium with improved fermentation performance for obtaining high butanol yield. However, screening methods to select the desired bacterial phenotype from large populations are not currently feasible or readily available. In this paper, a simple, novel screening method to select target strains by using trypan blue dye as an indicator in solid starch media to detect the target mutant bacterium was devised. A well performing mutant strain Clostridium beijerinckii no. 15 was obtained using this screening method. It was derived from wild-type C. beijerinckii NCIMB 8052 by N-methyl-N′-nitro-N-nitrosoguanidine (NTG) mutagenesis, and its fermentative capability for producing butanol was investigated, using hydrolysate of fresh cassava and other substrates. The mutant strain C. beijerinckii no. 15 produced a maximum yield of 15.06 g L−1 butanol and 23.78 g L−1 total solvents, when the fresh cassava substrate was given a simple pretreatment at 121 °C for 1 h. For other substrates, the highest butanol yields: 20.67 g L−1 from glucose, 17.96 g L−1 from liquefied dextrin, and 13.59 g L−1 from liquefied corn powder (P ≤ 0.05) were also achieved. Experimental trials showed that the mutant strain had a significantly greater fermentation ability, and produced a high-yield butanol from fresh cassava hydrolysates. Further research showed that the highest overall yield of butanol resulted from the improvement of enzymatic activity of C. beijerinckii no. 15s α-amylase (P ≤ 0.05). Also, the protein structure of the α-amylase was predicted and its amino acid mutation sites were analyzed. Results revealed that these mutant amino acids may have somehow improved α-amylase activity and thus, led to producing more butanol. Our research will contribute to efforts to develop C. beijerinckii fermentation of fresh cassava to produce butanol.

Identification and assessment of the effects of yeast decarboxylases expressed in Escherichia coli for producing higher alcohols

To contribute to the improvement of methods for the regulation and production of higher alcohols using micro‐organisms, we assessed the yields achieved using 10 decarboxylase genes from three different yeast species (Saccharomyces cerevisiae, Candida tropicalis and Pichia pastoris) by cloning them into vectors and overexpressing them in Escherichia coli hosts of different genotypes. Genes that produced the greatest yields in higher alcohol production were further assessed for the catalytic effects of the decarboxylase enzymes in the different E. coli hosts.

Effect of nitrogen and phosphorus deficiency on transcriptional regulation of genes encoding key enzymes of starch metabolism in duckweed (Landoltia punctata).

The production of starch by plants influences their use as biofuels. Nitrogen (N) and phosphorus (P) regulate starch gene expression during plant growth and development, yet the role of key enzymes such as ADP-glucose pyrophosphorylase (E.C. 2.7.7.27 AGPase) in starch metabolism during N- and P-deficiency remains unknown. We investigated the effect of N- and P-deficiency on the expression of large (LeAPL1, LeAPL2, and LeAPL3) and small (LeAPS) subunits of AGPase in duckweed (Landoltia punctata) and their correlation with starch content. We first isolated the full-length cDNA encoding LeAPL1 (GenBank Accession No. KJ603244) and LeAPS (GenBank Accession No. KJ603243); they contained open reading frames of 1554 bp (57.7-kDa polypeptide of 517 amino acids) and 1578 bp (57.0 kDa polypeptide of 525 amino acids), respectively. Real-time PCR analysis revealed that LeAPL1 and LeAPL3 were highly expressed during early stages of N-deficiency, while LeAPL2 was only expressed during late stage. However, in response to P-deficiency, LeAPL1 and LeAPL2 were upregulated during early stages and LeAPL3 was primarily expressed in the late stage. Interestingly, LeAPS was highly expressed following N-deficiency during both stages, but was only upregulated in the early stage after P-deficiency. The activities of AGPase and soluble starch synthesis enzyme (SSS EC 2.4.1.21) were positively correlated with changes in starch content. Furthermore, LeAPL3 and LeSSS (SSS gene) were positively correlated with changes in starch content during N-deficiency, while LeAPS and LeSSS were correlated with starch content in response to P-deficiency. These results elevate current knowledge of the molecular mechanisms underlying starch synthesis.

Analysis of DNA methylation of Spirodela polyrhiza (Grater Duckweed) in response to abscisic acid using methylation-sensitive amplied polymorphism

Spirodela polyrhiza is an a energy feedstock, which can produce a kind of dormant fronds called turions to survive cold. ABA can induce S. polyrhiza to form turions and is the most important hormone for plants to resist abiotic stresses. DNA methylation plays an important role in plant development by regulating gene expression. In this study, we studied DNA methylation variation in the fronds of S. polyrhiza treated with ABA and compared DNA methylation variation between fronds and turions, using the methylation-sensitive amplified polymorphism procedure. We selected 60 pairs of selective amplification primers to assess the status and levels of cytosine methylation. The results showed that ABA triggered the hemimethylation or internally full methylation of cytosine. With the prolongation of ABA treatment, the methylation of the total DNA increased. The alteration analysis of cytosine methylation showed that the number of demethylation events were much lower than those of methylation, which indicates that methylation was predominant. In addition, the methylation level in turions was higher than in the fronds. Moreover, the sequences of 14 differentially amplified DNA fragments were analyzed. According to Blast analysis, most of the 14 fragments were identified as genes or DNA involved in the abiotic stress response. The fragment M11 is homologous to ATPase. ABA may affect the methylation status of ATPase gene to regulate its expression to resist chilling. Our study showed that ABA might affect gene expression via changing the methylation status of the cytosine nucleotide.

Metagenomic analysis of microbial community and function involved in cd-contaminated soil

BackgroundSoil contaminated with the heavy metal Cadmium (Cd) is a widespread problem in many parts of the world. Based on metagenomic analysis, we investigated the functional potential and structural diversity of the microbial community in Cd-contaminated and non-contaminated soil samples and we explored the associated metabolic pathway network in cluster of orthologous groups (COG) and Kyoto Encyclopedia of Genes and Genomes (KEGG).ResultsThe results showed that microorganisms in these soils were quite abundant, and many of them possessed numerous physiological functions. However, Cd-contamination has the potential to reduce the microbial diversity and further alter the community structure in the soil. Notably, function analysis of the crucial microorganisms (e. g. Proteobacteria, Sulfuricella and Thiobacillus) indicated that these bacteria and their corresponding physiological functions were important for the community to cope with Cd pollution. The COG annotation demonstrated that the predominant category was the microbial metabolism cluster in both soil samples, while the relative abundance of metabolic genes was increased in the Cd-contaminated soil. The KEGG annotation results exhibited that the non-contaminated soil had more genes, pathways, modules, orthologies and enzymes involved in metabolic pathways of microbial communities than the Cd-contaminated soil. The relative abundance of some dominant KEGG pathways increased in the Cd contaminated soil, and they were mostly enriched to the metabolism, biosynthesis and degradation of amino acids, fatty acids and nucleotides, which was related to Cd tolerance of the microorganisms.ConclusionsCd-contamination can decrease the taxonomic species of microbes in soil and change the soil microbial composition. The functional pathways involved in the soil change with microbial structure variation, many of which are related to the heavy metal tolerance of soil microbes. The Cd-contaminated soil microbes is a potential resource for exploring cadmium resistant or tolerant bacteria.