FK506, a widely used immunosuppressant, is produced by industrial fermentation processes using various Streptomyces species. However, the low titer becomes a bottleneck for its application and industrialization. It urgently required a full understanding of the biological mechanisms for FK506 overproduction. Towards this end, comparative metabolomics approach was employed to analyze metabolite concentrations difference of Streptomyces tsukubaensis cultivated in two media with low and high productivities. Initially, 98 intracellular metabolites were identified and 13 metabolites involved in five pathways were determined to be directly correlated with FK506 biosynthesis. Then in-depth analysis elucidated how those key factors exerted influence on FK506 biosynthesis. Many previously unreported metabolites were shown to play an important role in FK506 biosynthesis and provided potential regulation points for external manipulation. Based on such key information, rationally designed feeding strategy was carried out. Results showed that the FK506 yield increased from 251 to 405 mg/L, whereas, by-products FK520 and 37,38-dihydro-FK506 decreased by 31% and 39%, respectively, compared with the values of control. To our knowledge, it is the first study to apply the comparative metabolomics method to identify key metabolites to promote the FK506 production. The strategies developed here can easily be extended to titer improvement of other important microbial natural products and process optimization.

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There are some arguments between the use of hydroxyapatite and porous coating. Some studies have shown that there is no difference between these two coatings in total hip arthroplasty (THA), while several other studies have shown that hydroxyapatite has advantages over the porous one. We have collected the studies in Pubmed, MEDLINE, EMBASE, and the Cochrane library from the earliest possible years to present, with the search strategy of “(HA OR hydroxyapatite) AND ((total hip arthroplasty) OR (total hip replacement)) AND (RCT* OR randomiz* OR control* OR compar* OR trial*)”. The randomized controlled trials and comparative observation trials that evaluated the clinical and radiographic effects between hydroxyapatite coating and porous coating were included. Our main outcome measurements were Harris hip score (HHS) and survival, while the secondary outcome measurements were osteolysis, radiolucent lines, and polyethylene wear. Twelve RCTs and 9 comparative observation trials were included. Hydroxyapatite coating could improve the HHS (p < 0.01), reduce the incidence of thigh pain (p = 0.01), and reduce the incidence of femoral osteolysis (p = 0.01), but hydroxyapatite coating had no advantages on survival (p = 0.32), polyethylene wear (p = 0.08), and radiolucent lines (p = 0.78). Hydroxyapatite coating has shown to have an advantage over porous coating. The HHS and survival was duration-dependent—if given the sufficient duration of follow-up, hydroxyapatite coating would be better than porous coating for the survival. The properties of hydroxyapatite and the implant design had influence on thigh pain incidence, femoral osteolysis, and polyethylene wear. Thickness of 50 to 80 μm and purity larger than 90% increased the thigh pain incidence. Anatomic design had less polyethylene wear.

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In silico metabolic network models are valuable tools for strain improvement with desired properties. In this work, based on the comparisons of each pathway flux under two different objective functions for the reconstructed metabolic network of Streptomyces roseosporus, three potential targets of zwf2 (code for glucose-6-phosphate hydrogenase), dptI (code for α-ketoglutarate methyltransferase), and dptJ (code for tryptophan oxygenase) were identified and selected for the genetic modifications. Overexpression of zwf2, dptI, and dptJ genes increased the daptomycin concentration up to 473.2, 452.5, and 489.1 mg/L, respectively. Furthermore, co-overexpression of three genes in series resulted in a 34.4% higher daptomycin concentration compared with the parental strain, which ascribed to the synergistic effect of the enzymes responsible for daptomycin biosynthesis. Finally, the engineered strain enhanced the yield of daptomycin up to 581.5 mg/L in the fed-batch culture, which was approximately 43.2% higher than that of the parental strain. These results demonstrated that the metabolic network based on in silico prediction would be accurate, reasonable, and practical for target gene identification and strain improvement.