[Functional Genomics Analysis of Nitrogen and Phosphorus Transformation in Maize Rhizosphere Microorganisms].

Fertilizer reduction and efficiency improvement is an important basis for ensuring the safety of the agricultural ecological environment. Microorganisms are the key driving force for regulating the soil nitrogen and phosphorus cycle. Studying the nitrogen and phosphorus transformation function of rhizosphere microorganisms can provide a microbiological regulation approach for further improving the use efficiency of soil nitrogen and phosphorus. Based on the field micro-plot experiments of three typical farmland soils(phaeozem, cambisol, and acrisol), metagenomic sequencing technology was used to study the differences in functional genes and regulatory factors of maize rhizosphere microorganisms during soil nitrogen and phosphorus transformation. The results showed that the functional diversity of maize rhizosphere microorganisms was affected by soil type. The functional diversity of rhizosphere microorganisms in phaeozem and cambisol was mainly affected by water content and nutrient content, and that in acrisol was affected by total phosphorus(TP) and available phosphorus(AP). For soil nitrogen transformation, the gene abundance of related enzymes in the pathway of nitrogen transformation was the highest in the urease gene(ureC) and glucose dehydrogenase gene(gdh), which were 7.25×10-5-12.88×10-5 and 4.47×10-5-7.49×10-5, respectively. The total abundance of assimilatory nitrate reduction functional genes in acrisol was higher than that in phaeozem and cambisol, and the total abundance of functional genes related to other processes was the highest in cambisol. The abundance of functional genes encoding enzymes related to nitrogen metabolism was mainly driven by soil bacterial richness, total potassium(TK), and TP. For soil phosphorus transformation, the number of alkaline phosphatase genes(phoD) catalyzing organic phosphorus mineralization was 1093, and the number of acid phosphatase genes(PHO) was 42. The abundance of phoD was two orders of magnitude higher than that of PHO. In addition, fertilization had no significant effect on the abundance of phoD and PHO in the same soil type. Random forest analysis showed that the abundances of phoD and PHO were significantly affected by soil moisture, organic matter(OM), and total nitrogen(TN), but AP content had the greatest impact on PHO abundance. These results clarified the nitrogen and phosphorus transformation characteristics of maize rhizosphere microorganisms at the functional genomic level and enriched the molecular biological mechanism of the microbial nitrogen and phosphorus transformation function.

Study on the Antipyretic and Anti-inflammatory Mechanism of Shuanghuanglian Oral Liquid Based on Gut Microbiota-Host Metabolism.

Nowadays, there has been increased awareness that the therapeutic effects of natural medicines on inflammatory diseases may be achieved by regulating the gut microbiota. Shuanghuanglian oral liquid (SHL), the traditional Chinese medicine preparation, has been shown to be effective in clearing heat-toxin, which is widely used in the clinical treatment of respiratory tract infection, mild pneumonia, and common cold with the wind-heat syndrome. Yet the role of gut microbiota in the antipyretic and anti-inflammatory effects is unclear. In this study, a new strategy of the 16S rRNA gene sequencing and serum metabolomics that aims to explore the role of SHL in a rat model of the systemic inflammatory response induced by lipopolysaccharide would be a major advancement. Our results showed that the gut microbiota structure was restored in rats with inflammation after oral administration of SHL, thereby reducing inflammation. Specifically, SHL increased the relative abundance of Bacteroides and Faecalibacterium and decreased the abundance of Bifidobacterium, Olsenella, Aerococcus, Enterococcus, and Clostridium in the rat model of inflammatory disease. Serum metabolomic profile obtained by the orbitrap-based high-resolution mass spectrometry revealed significant differences in the levels of 39 endogenous metabolites in the inflammatory model groups, eight metabolites of which almost returned to normal levels after SHL treatment. Correlation analysis between metabolite, gut microbiota, and inflammatory factors showed that the antipyretic and anti-inflammatory effects of SHL were related to the recovery of the abnormal levels of the endogenous metabolites (N-acetylserotonin and 1-methylxanthine) in the tryptophan metabolism and caffeine metabolism pathway. Taken together, these findings suggest that the structural changes in the gut microbiota are closely related to host metabolism. The regulation of gut microbiota structure and function is of great significance for exploring the potential mechanism in the treatment of lipopolysaccharide-induced inflammatory diseases with SHL.

[Molecular identification of Armillaria gallica by PCR-RFLP analysis].

Armillaria gallica is a symbiotic fungus in the cultivation process of Gastrodia elata and Polyporus.The rhizomorph of A. gallica invades the stalk of the G. elata or the Sclerotium of the Polyporus,and is digested and utilized by the latter,becoming their important source of nutrition. Different nature of A. gallica affects the growth of G. elata and Polyporus. The authors collected A. gallica from 13 commercially available regions and screened two A. gallica,A and B,at the genetic and metabolic levels,in order to distinguish between the two A. gallica market. We have established convenient and effective DNA molecular identification method.By comparing the sequence differences between the A. gallica type A and type B invertase genes,PCR-RFLP primers were designed based on differential fragment. Primer ZTM.F/ZTM.R can amplified A. gallica type A and B,producing a band of about 304 bp in length. The restriction endonuclease EcoR V could recognize the difference sequence of A and B types of A. gallica. The type B was digested to form two fragments,thereby specifically identifying the A. gallica as type B. The established methods of PCR-RFLP is an accurate identification method for A. gallica. Therefore,in the cultivation process of G. elata and Polyporus,suitable strains can be selected according to different needs of variety,growth stage and ecological environment,and the yield and quality can be improved according to local conditions.

[Analysis of monosaccharide composition of twelve polysaccharides by pre-column derivatization procedure UPLC-MS/MS].

This study is to establish a pre-column derivatization procedure with 1-phenyl-3-methyl-5-pyrazolone (PMP) UPLC-MS/MS method for the determination of the monosaccharide composition of 12 polysaccharides. At the same time, the monosaccharide components of polysaccharides in Armillaria gallica were analyzed. The separation was performed on a ACQUITY ZORBAX RRHD Eclipse Plus C18 column(2.1 mm×100 mm, 1.8 µm),using 95% acetonitrile (A) and ammonium acetate-5% acetonitrile-water (B) as mobile phase with gradient elution. The target components were detected in multiple-reaction monitoring (MRM) mode by mass spectrometry with electrospray ionization (ESI) source operated in ionization mode. The results showed that based on the monosaccharides detection method established by UPLC-MS/MS, the linearity of the 12 monosaccharides components were linear in their linear range (R²>0.990), and the recovery rate were 92.30%-105.6%. 11 monosaccharides such as fructose, mannose, and glucose were detected in A. gallica samples. The method established in this experiment is robust, highly reproducible and accurate, and is suitable for the determination of monosaccharide components such as A. gallica.

A CqFerritin protein inhibits white spot syndrome virus infection via regulating iron ions in red claw crayfish Cherax quadricarinatus.

It is well known that iron is an essential element for all living organism. The intracellular iron availability is also important for the host's innate immune response to various pathogens, in which the iron homeostasis can be regulated by ferritin due to its iron storage property. In this study, a full-length cDNA sequence of ferritin (named as CqFerritin) was identified with 1410 bp from red claw crayfish Cherax quadricarinatus, which contained an open reading frame of 513 bp, encoding 170 amino acids with a conserved ferritin domain. Tissue distribution analysis demonstrated that CqFerritin was widely expressed in various tissues with high presence in haemocyte, haematopoietic tissue (Hpt) and heart, while lowest expression in hepatopancreas. In addition, loss-of-function of CqFerritin by gene silencing resulted in significantly higher expression of an envelope protein VP28 of white spot syndrome virus (WSSV) in red claw crayfish Hpt cell cultures, indicating the potential antiviral response of CqFerritin. To further explore the effect on WSSV replication by CqFerritin, recombinant CqFerritin protein (rCqFerritin) was transfected into Hpt cells followed by WSSV infection. Importantly, the replication of WSSV was obviously decreased in Hpt cells if transfected with rCqFerritin protein, suggesting that CqFerritin had clearly negative effect on WSSV infection. Furthermore, intracellular accumulation of iron ions was found to promote the WSSV replication in a dose-dependent manner, illustrating that the iron level regulated by CqFerritin was likely to be vital for WSSV infection in red claw crayfish. Taken together, these data suggest that CqFerritin plays an important role in immune defense against WSSV infection in a crustacean C. quadricarinatus.