β-Glucan Synthase Gene Overexpression and β-Glucans Overproduction in <i>Pleurotus ostreatus</i> Using Promoter Swapping

<div><p>Mushroom β-glucans are potent immunological stimulators in medicine, but their productivities are very low. In this study, we successfully improved its production by promoter engineering in <i>Pleurotus ostreatus</i>. The promoter for β-1,3-glucan synthase gene (<i>GLS</i>) was replaced by the promoter of glyceraldehyde-3-phosphate dehydrogenase gene of <i>Aspergillus nidulans</i>. The homologous recombination fragment for swapping <i>GLS</i> promoter comprised five segments, which were fused by two rounds of combined touchdown PCR and overlap extension PCR (TD-OE PCR), and was introduced into <i>P. ostreatus</i> through PEG/CaCl₂-mediated protoplast transformation. The transformants exhibited one to three fold higher transcription of <i>GLS</i> gene and produced 32% to 131% higher yield of β-glucans than the wild type. The polysaccharide yields had a significant positive correlation to the <i>GLS</i> gene expression. The infrared spectra of the polysaccharides all displayed the typical absorption peaks of β-glucans. This is the first report of successful swapping of promoters in filamentous fungi.</p></div

Comparison of predicted GLS amino acid sequences among <i>P. ostreatus</i> TD300, <i>Laccaria bicolor</i> S238N-H82, and <i>P. ostreatus</i> PC15 v2.0.

<p><b>Lb</b>: GLS from <i>Laccaria bicolor</i> S238N-H82, GeneBank accession numbers is XM_001875351. <b>Po PC</b>: GLS from <i>P. ostreatus</i> PC15 v2.0, collected by visual inspection using the JGI Genome Portal for the <i>P. ostreatus</i> PC15 v2.0 genome (<a href="http://genome.jgi-psf.org" target="_blank">http://genome.jgi-psf.org</a>); <b>Po TD</b>: GLS from <i>P. ostreatus</i> TD300, GeneBank accession numbers is JX889617.</p

Semi-quantitative RT-PCR analysis of <i>GLS</i> mRNA in the fifth generation transformants.

<p>The amount of <i>GLS</i> mRNA, expressed as the ratio of densitometric measurement of the sample to the corresponding internal standard (β-<i>actin</i>), is shown in the upper panels. * <i>p</i><0.05 comparing to WT; ** <i>p</i><0.01 comparing to WT.</p

The infrared spectroscopy of the β-glucans produced by the fifth generation transformants.

<p>The infrared spectroscopy of the β-glucans produced by the fifth generation transformants.</p

Primers used in this study.

<p>Primers used in this study.</p

The polysaccharide yield of the fifth generation transformants.

<p>* <i>p</i><0.05 comparing to WT; ** <i>p</i><0.01 comparing to WT.</p

Monitoring impacts of ecological engineering on ecosystem services with geospatial techniques in karst areas of SW China

Ecosystem services (ESs) synergies have been broadly studied, whose synergy control areas and driving mechanisms were still not fully understood. Here, We analyzed soil conservation (SC), water yield (WY), net primary productivity (NPP) and food supply (FS) on the basis of geospatial analysis techniques from 1992 to 2015. The results showed that the SW karst areas contributed 23.7% of SC and 14.56% of NPP with only 8.4% of China's continent. 49.28% of China’s land was synergy control areas, while accounted for 82.23% in the SW karst areas. The cause of this phenomenon was twofold, one was the significant increase in NDVI was mainly due to human activities (HA), and the coincidence rates between the HA dominated areas and the synergies control areas up to 90.7%. These results emphasized that ecological engineering was a key factor of the synergy among multi-ESs, the synergy control areas should be priority for ecological restoration

Agarose gel analysis of TD-OE PCR products for the construction of the homologous recombination fragment for promoter swapping.

<p>hPG: the fusion product of <i>hph</i>, <i>P_gpd</i>, and <i>GLS</i>₁₀₂₅; HRF: the homologous recombination fragment which was the fusion product of UH, <i>P_gpd</i>₁₀₃₅, and hPG.</p

Climate change enhances the positive contribution of human activities to vegetation restoration in China

Ecological projects have huge impacts on vegetation restoration (VR) in China. However, are all of the effects due to human activities (HA) without the contribution of climate change (CC)? It is unclear what role CC plays in VR. Here, we quantitatively evaluate the contributions of climate change and human activities (CC_con and HA_con) to the net primary productivity (NPP) by the method of partial derivative and six different scenarios. HA contributed 61.19% to the NPP trend and controlled 32.84% of the NPP growth in China. Among the climate factors, temperature made the largest contribution to the NPP (45.89%) and had the largest control area (36.36%). CC promoted the positive HA_con in 54% of China, and in 68.35% of the area, the enhancement effect reached more than 50%. Our results answer a longstanding question and emphasize the important role of CC in enhancing the positive HA_con to VR

Biodegradation of Quinoline by a Newly Isolated Salt-Tolerating Bacterium <i>Rhodococcus gordoniae</i> Strain JH145

Quinoline is a typical nitrogen-heterocyclic compound with high toxicity and carcinogenicity which exists ubiquitously in industrial wastewater. In this study, a new quinoline-degrading bacterial strain Rhodococcus sp. JH145 was isolated from oil-contaminated soil. Strain JH145 could grow with quinoline as the sole carbon source. The optimum growth temperature, pH, and salt concentration were 30 °C, 8.0, and 1%, respectively. 100 mg/L quinoline could be completely removed within 28 h. Particularly, strain JH145 showed excellent quinoline biodegradation ability under a high-salt concentration of 7.5%. Two different quinoline degradation pathways, a typical 8-hydroxycoumarin pathway, and a unique anthranilate pathway were proposed based on the intermediates identified by liquid chromatography–time of flight mass spectrometry. Our present results provided new candidates for industrial application in quinoline-contaminated wastewater treatment even under high-salt conditions