Exploring Oxidoreductases from Extremophiles for Biosynthesis in a Non-Aqueous System.

Organic solvent tolerant oxidoreductases are significant for both scientific research and biomanufacturing. However, it is really challenging to obtain oxidoreductases due to the shortages of natural resources and the difficulty to obtained it via protein modification. This review summarizes the recent advances in gene mining and structure-functional study of oxidoreductases from extremophiles for non-aqueous reaction systems. First, new strategies combining genome mining with bioinformatics provide new insights to the discovery and identification of novel extreme oxidoreductases. Second, analysis from the perspectives of amino acid interaction networks explain the organic solvent tolerant mechanism, which regulate the discrete structure-functional properties of extreme oxidoreductases. Third, further study by conservation and co-evolution analysis of extreme oxidoreductases provides new perspectives and strategies for designing robust enzymes for an organic media reaction system. Furthermore, the challenges and opportunities in designing biocatalysis non-aqueous systems are highlighted.

The Requirement and Protective Effects of Dietary Protein against Chronic Ammonia Exposure in Juvenile Genetically Improved Farmed Tilapia (Oreochromis niloticus).

Ammonia is a key risk factor in intensive aquaculture systems. This experiment is aimed at investigating the influence of dietary protein levels on genetically improved farmed tilapia (GIFT, Oreochromis niloticus) under chronic ammonia stress. GIFT juveniles of 4.00 ± 0.55 g were exposed to high ammonia level at 0.88 mg/L and fed with six diets comprising graded protein levels at 22.64%, 27.26%, 31.04%, 35.63%, 38.47%, and 42.66% for 8 weeks. The fish in negative control was fed the diet with 31.04% protein in normal water (0.02 mg ammonia/L water). Our results showed that high ammonia exposure (0.88 mg/L) caused significant decrease in fish growth performance, hematological parameters, liver antioxidant enzymes (catalase and glutathione peroxidase), and gill Na+- and K+-dependent adenosine triphosphatase (Na+/K+-ATP) activity. When fish were under high ammonia exposure, the weight gain rate, special growth rate, feed efficiency, and survival rate elevated significantly with dietary protein supplementation increase to 35.63%, whereas protein efficiency ratio, hepatosomatic index, and viscerosomatic index showed a decreased tendency. Dietary protein administration significantly enhanced crude protein but reduced crude lipid contents in the whole fish. Fish fed diets with 35.63%-42.66% protein had higher red blood cell counts and hematocrit percentage than fish fed 22.64% protein diet. The values of serum biochemical indices (lactate dehydrogenase, aspartate aminotransferase, and alanine aminotransferase), hepatic antioxidant enzymes (superoxide dismutase, catalase, and glutathione peroxidase), and gill Na+/K+-ATP activity were all elevated with the increment of dietary protein. Moreover, histological analysis indicated that dietary protein administration could prevent the ammonia-induced damages in fish gill, kidney, and liver tissues. Based on weight gain rate as a response criterion, the optimal dietary protein requirement for GIFT juveniles under chronic ammonia stress was 37.9%.

Integrating larval zebrafish model and network pharmacology for screening and identification of edible herbs with therapeutic potential for MAFLD: A promising drug Smilax glabra Roxb.

Metabolic-associated fatty liver disease (MAFLD) is becoming a prevalent chronic liver disease. Many medicinal and edible herbs exhibit remarkable biological activities in ameliorating MAFLD but lack a comprehensive assessment of their therapeutic efficacy. This study determined total phenolic and flavonoid contents and in vitro antioxidant properties of 34 edible herbs. Smilax glabra Roxb. (SGR), Coreopsis tinctoria Nutt., and Smilax china L. were obtained with the best bioactivity and antioxidant capacity. The high-cholesterol diet-induced larval zebrafish model was established to compare the anti-MAFLD activity of the three herb extracts mentioned above. In vivo experiments revealed that SGR intervention significantly decreased lipid accumulation, alleviated oxidative stress, and modulated intestinal microbiota composition in zebrafish. Furthermore, three potential active components in SGR and their possible mechanisms were explored through network pharmacology and molecular docking. Our study suggested that SGR is a potential candidate for developing new drugs or dietary supplements for MAFLD therapy.

Dietary host-associated Bacillus subtilis supplementation improves intestinal microbiota, health and disease resistance in Chinese perch (Sinipercachuatsi).

The purpose of this study was to evaluate the potential of a host-associated Bacillus subtilis 1-C-7 as a probiotic for Chinese perch (Siniperca chuatsi). Four test diets were formulated to contain graded levels of B. subtilis 1-C-7 at 0 (CY), 0.85 × 108 (Y1), 0.95 × 109 (Y2) and 0.91 × 1010 (Y3) CFU/kg diet. The test fish with initial weight 30.0 ± 1.2 g were fed the 4 test diets with 3 replicates in an indoor water-flow aquaculture system with 12 net cages (40 fish/cage) for 10 wk. At the conclusion of the feeding trial, the probiotic effects of B. subtilis on Chinese perch were analyzed based on growth performance, serum biochemical indices, histologic morphology of liver and gut, gut microbiota and the resistance to Aeromonas hydrophila. The results showed that the percentage of weight gain had no significant change in the Y1 and Y2 groups (P > 0.05) but decreased in the Y3 group compared to that in the CY group (P < 0.05). The fish in the Y3 group displayed the highest activity of serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) among these 4 groups (P < 0.05). The fish in the CY group had the highest value of malondialdehyde in the liver (P < 0.05) and showed severe nuclear migration and vacuolization of hepatocytes. The morphology indicated that all test fish had poor intestinal health. However, the fish in the Y1 group had a relatively normal intestinal histologic structure. The mid gut microbial diversity analysis showed that dietary B. subtilis supplementation increased the abundance of probiotics such as Tenericutes and Bacteroides, whereas it reduced the abundance of pernicious bacteria such as Proteobacteria, Actinobacteria, Thermophilia and Spirochaetes. The challenge test showed that dietary B. subtilis supplementation increased the resistance to A. hydrophila in Chinese perch. In conclusion, dietary supplementation of 0.85 × 108 CFU/kg B. subtilis 1-C-7 could improve the intestinal microbiota, intestinal health and disease resistance in Chinese perch, but more or excessive supplementation could reduce growth performance and have negative effects on health.

The probiotic effects of host-associated Bacillus velezensis in diets for hybrid yellow catfish (Pelteobagrus fulvidraco ♀ × Pelteobagrus vachelli ♂).

This study was to evaluate the potential of a host-associated Bacillus velezensis as a probiotic for hybrid yellow catfish (Pelteobagrus fulvidraco ♀ × Pelteobagrus vachelli ♂). Diets (B0 to B5) containing 0, 0.90 × 108, 0.80 × 109, 0.85 × 1010, 0.90 × 1011, 0.83 × 1012 CFU/kg B. velezensis YFI-E109 were fed to the fish with initial weight (3.07 ± 0.08 g) in a recirculating aquaculture system for six weeks with three replicates, respectively. Probiotic effects were analyzed based on growth, body composition, liver and gut morphology, gut microbiome, and liver metabolome. Analysis of the bacterial genome has shown that the most abundant genes in B. velezensis YFI-E109 were distributed in carbohydrate and amino acid metabolism. Fish in groups B3 and B4 had better growth performance, and higher intestinal amylase (AMS) and lipase (LPS) activities compared with other groups (P < 0.05). Fish in groups B0 and B5 showed significant liver damage, while this status improved in group B3. The liver malondialdehyde (MDA) content in group B3 was lower than that in other groups (P < 0.05). The abundance of Mycoplasma, Ralstonia and Acinetobacter was significantly reduced in B3 and B5 compared to B0. The amino acid and carbohydrate metabolism pathways were enriched in group B3 compared with group B0. In conclusion, dietary B. velezensis YFI-E109 supplementation has the potential to improve growth, liver metabolism, and liver and gut health, and reshape the gut microbiome of hybrid yellow catfish. Excessive B. velezensis YFI-E109 reduced the prebiotic effects. The recommended dietary supplementation of B. velezensis YFI-E109 is 0.31 × 1010 to 0.77 × 1011 CFU/kg for hybrid yellow catfish according to the quadratic regression method by plotting specific growth rate (SGR), feed conversion ratio (FCR), MDA and activities of AMS against dietary B. velezensis YFI-E109 levels.

Preliminary Study to Assess the Impact of Dietary Rutin on Growth, Antioxidant Capacity, and Intestinal Health of Yellow Catfish, Pelteobagrus fulvidraco.

This research aimed to examine the effects of dietary rutin supplementation on growth, body composition, serum biochemical indexes, liver enzyme activities and antioxidant-related genes expression, intestinal morphology, and microbiota composition of juvenile yellow catfish (Pelteobagrus fulvidraco). Rutin was added to the basal diets at doses of 0 (control), 100 mg/kg, and 500 mg/kg. Each diet was fed randomly into three tanks, each tank containing 30 fish with an initial body mass of (10.27 ± 0.62) g. The feeding trial was conducted in an indoor recirculating aquiculture system at 28 °C for 56 days. According to the findings, the inclusion of 100 mg/kg rutin significantly improved the growth performance of yellow catfish and reduced the feed conversion ratio; however, the growth promotion effect was diminished when the diet was supplemented with 500 mg/kg of rutin. The inclusion of 500 mg/kg rutin in the diet significantly reduced the level of crude lipid and protein of the whole fish. Serum activities of alkaline phosphatase, albumin, and total protein were all significantly increased when fish were fed the diet supplemented with 500 mg/kg rutin, while serum glucose was significantly lower compared to the control group. Meanwhile, dietary rutin at a concentration of 500 mg/kg significantly induced the hepatic mRNA expressions of antioxidant-related genes (including Cu/Zn-SOD, Mn-SOD, CAT, GPx) and inflammatory-associated genes (including TNFα, IL-10, LYZ). Incorporating rutin at doses of 100 mg/kg and 500 mg/kg into the diets resulted in a notable increase in superoxide dismutase (SOD) activity, while simultaneously reducing malondiadehyde (MDA) content in the liver and intestine. Intestinal villus height, villus width, muscular thickness, and lumen diameter were significantly increased with the administration of 500 mg/kg of dietary rutin. Gut microbial diversity analysis indicated that supplementing diets with 100 mg/kg and 500 mg/kg rutin significantly enhanced the abundance of Cetobacterium while decreasing Plesiomonas richness. In conclusion, dietary rutin levels at 100 mg/kg could enhance the growth, antioxidant capability, and intestinal health of yellow catfish under present experimental conditions.

One-Pot Purification and Immobilization of Phenylalanine Dehydrogenase from Bacillus nanhaiensi by Functional Reduced Graphene Oxide.

The one-pot immobilization of halophilic phenylalanine dehydrogenase from marine microorganism with metal ions modified reduced graphene oxide (CRGO) material was studied. Phenylalanine dehydrogenase was from Bacillus nanhaiensi and expressed with a C-terminal His-tag. Investigation of CRGO, CRGO-PEI, CRCO-Mn, and CRGO-PEI-Mn for one-pot purification and immobilization of phenylalanine dehydrogenase from crude enzyme solution was carried out. Enzyme activity yield rate achieved 80.0% by immobilization with CRCO-Mn, and the loading capacity was 6.7 mg/mg. Manganese ion coordination greatly improved the selectivity of the CRGO for the target His-tagged enzyme. Furthermore, the effect of NaCl concentration on the immobilization was investigated, which the loading capacity of CRGO-PEI and CRGO-Mn-PEI was increased by 10.7% and 30.6% with 1 M NaCl, respectively. The adsorption curves of crude enzyme one-pot immobilized by CRGO-Mn and purified enzyme immobilized by CRGO-Mn were similar. Therefore, one-pot immobilization strategy is promising for industrial application with advantages such as high efficiency and low cost, which shorten the pipelines for enzyme discovery towards industrial applications through the establishing of marine enzyme collections.