Genome analysis of Rossellomorea sp. y25, a deep sea bacterium isolated from the sediments of South China Sea.

Rossellomorea sp. y25, a putative new species of yellow pigment-producing, aerobic and chemoheterotrophic bacterium belonging to the family Bacillaceae, was isolated from the sediments at the depth of 1829 m in the South China Sea. In this study, we present the complete genome sequences of strain y25, which consisted of only one circular chromosome with 4,633,006 bp and the content of G + C was 41.76%. A total of 4466 CDSs, 106 tRNA, 33 rRNA, and 101 sRNA genes were obtained. Genomic analysis of strain y25 showed that it has the ability to produce antioxidant carotenoids and a large number of heavy metal resistance genes, such as arsenic, cadmium and zinc. In addition, strain y25 contains a prophage that may contribute to host protection against lysis by related Bacillus-like phages. This is the first report of genome-wide information on a bacterium of the genus Rossellomorea isolated from the deep sea, providing insights into how microorganisms of this genus adapt to deep-sea environments.

Application of Acetate as a Substrate for the Production of Value-Added Chemicals in Escherichia coli.

At present, the production of the majority of valuable chemicals is dependent on the microbial fermentation of carbohydrate substrates. However, direct competition is a potential problem for microbial feedstocks that are also used within the food/feed industries. The use of alternative carbon sources, such as acetate, has therefore become a research focus. As a common organic acid, acetate can be generated from lignocellulosic biomass and C1 gases, as well as being a major byproduct in microbial fermentation, especially in the presence of an excess carbon source. As a model microorganism, Escherichia coli has been widely applied in the production of valuable chemicals using different carbon sources. Recently, several valuable chemicals (e.g., succinic acid, itaconic acid, isobutanol, and mevalonic acid) have been investigated for synthesis in E. coli using acetate as the sole carbon source. In this review, we summarize the acetate metabolic pathway in E. coli and recent research into the microbial production of chemical compounds in E. coli using acetate as the carbon source. Although microbial synthetic pathways for different compounds have been developed in E. coli, the production titer and yield are insufficient for commercial applications. Finally, we discuss the development prospects and challenges of using acetate for microbial fermentation.

Activating autophagy promotes skin regeneration induced by mechanical stretch during tissue expansion.

Background: Tissue expansion, a technique in which skin regeneration is induced by mechanical stretch stimuli, is commonly used for tissue repair and reconstruction. In this study, we aimed to monitor the autophagy levels of expanded skin after the application of expansion stimuli and explore the effect of autophagy modulation on skin regeneration. Methods: A rat scalp expansion model was established to provide a stable expanded skin response to mechanical stretch. Autophagy levels at different time points (6, 12, 24, 48 and 72 h after the last expansion) were detected via western blotting. The effect of autophagy regulation on skin regeneration during tissue expansion was evaluated via skin expansion efficiency assessment, western blotting, immunofluorescence staining, TUNEL staining and laser Doppler blood flow imaging. Results: The autophagic flux reached its highest level 48 h after tissue expansion. Activating autophagy by rapamycin increased the area of expanded skin as well as the thicknesses of epidermis and dermis. Furthermore, activating autophagy accelerated skin regeneration during tissue expansion by enhancing the proliferation of cells and the number of epidermal basal and hair follicle stem cells, reducing apoptosis, improving angiogenesis, and promoting collagen synthesis and growth factor secretion. Conversely, the regenerative effects were reversed when autophagy was blocked. Conclusions: Autophagy modulation may be a promising therapeutic strategy for improving the efficiency of tissue expansion and preventing the incidence of the complication of skin necrosis.

S100 calcium-binding protein A9 promotes skin regeneration through toll-like receptor 4 during tissue expansion.

Background: In plastic surgery, tissue expansion is widely used for repairing skin defects. However, low expansion efficiency and skin rupture caused by thin, expanded skin remain significant challenges in promoting skin regeneration during expansion. S100 calcium-binding protein A9 (S100A9) is essential in promoting wound healing; however, its effects on skin regeneration during tissue expansion remain unclear. The aim of the present study was to explore the role of S100A9 in skin regeneration, particularly collagen production to investigate its importance in skin regeneration during tissue expansion. Methods: The expression and distribution of S100A9 and its receptors-toll-like receptor 4 (TLR-4) and receptor for advanced glycation end products were studied in expanded skin. These characteristics were investigated in skin samples of rats and patients. Moreover, the expression of S100A9 was investigated in stretched keratinocytes in vitro. The effects of S100A9 on the proliferation and migration of skin fibroblasts were also observed. TAK-242 was used to inhibit the binding of S100A9 to TLR-4; the levels of collagen I (COL I), transforming growth factor beta (TGF-ß), TLR-4 and phospho-extracellular signal-related kinase 1/2 (p-ERK1/2) in fibroblasts were determined. Furthermore, fibroblasts were co-cultured with stretched S100A9-knockout keratinocytes by siRNA transfection and the levels of COL I, TGF-ß, TLR-4 and p-ERK1/2 in fibroblasts were investigated. Additionally, the area of expanded skin, thickness of the dermis, and synthesis of COL I, TGF-ß, TLR-4 and p-ERK1/2 were analysed to determine the effects of S100A9 on expanded skin. Results: Increased expression of S100A9 and TLR-4 was associated with decreased extracellular matrix (ECM) in the expanded dermis. Furthermore, S100A9 facilitated the proliferation and migration of human skin fibroblasts as well as the expression of COL I and TGF-ß in fibroblasts via the TLR-4/ERK1/2 pathway. We found that mechanical stretch-induced S100A9 expression and secretion of keratinocytes stimulated COL I, TGF-ß, TLR-4 and p-ERK1/2 expression in skin fibroblasts. Recombined S100A9 protein aided expanded skin regeneration and rescued dermal thinning in rats in vivo as well as increasing ECM deposition during expansion. Conclusions: These findings demonstrate that mechanical stretch promoted expanded skin regeneration by upregulating S100A9 expression. Our study laid the foundation for clinically improving tissue expansion using S100A9.

Engineering Escherichia coli for Isobutanol Production from Xylose or Glucose-Xylose Mixture.

Aiming to overcome the depletion of fossil fuels and serious environmental pollution, biofuels such as isobutanol have garnered increased attention. Among different synthesis methods, the microbial fermentation of isobutanol from raw substrate is a promising strategy due to its low cost and environmentally friendly and optically pure products. As an important component of lignocellulosics and the second most common sugar in nature, xylose has become a promising renewable resource for microbial production. However, bottlenecks in xylose utilization limit its wide application as substrates. In this work, an isobutanol synthetic pathway from xylose was first constructed in E. coli MG1655 through the combination of the Ehrlich and Dahms pathways. The engineering of xylose transport and electron transport chain complexes further improved xylose assimilation and isobutanol production. By optimizing xylose supplement concentration, the recombinant E. coli strain BWL4 could produce 485.35 mg/L isobutanol from 20 g/L of xylose. To our knowledge, this is the first report related to isobutanol production using xylose as a sole carbon source in E. coli. Additionally, a glucose-xylose mixture was utilized as the carbon source. The Entner-Doudorof pathway was used to assimilate glucose, and the Ehrlich pathway was applied for isobutanol production. After carefully engineering the recombinant E. coli, strain BWL9 could produce 528.72 mg/L isobutanol from a mixture of 20 g/L glucose and 10 g/L xylose. The engineering strategies applied in this work provide a useful reference for the microbial production of isobutanol from xylose or glucose-xylose mixture.

Response surface optimization of poly-ß-hydroxybutyrate synthesized by Bacillus cereus L17 using acetic acid as carbon source.

A strain of Bacillus that can tolerate 10 g/L acetic acid and use the volatile fatty acids produced by the hydrolysis and acidification of activated sludge to produce polyhydroxyalkanoate was screened from the activated sludge of propylene oxide saponification wastewater. The strain was identified by 16S rRNA sequencing and phylogenetic tree analysis and was named Bacillus cereus L17. Various characterization methods showed that the polymer synthesized by strain L17 is poly-ß-hydroxybutyrate, which has low crystallinity, good ductility and toughness, high thermal stability and a low polydispersity coefficient. It has wide thermoplastic material operating space as well as industrial and medicinal applications. The optimal fermentation conditions were determined by single factor optimization. Then, Plackett-Burman and Box-Behnken design experiments were carried out according to the single factor optimization results, and the response surface optimization was completed. The final results were: initial pH 6.7, temperature 25 °C, and loading volume 124 mL. The verification experiment showed that the yield of poly-ß-hydroxybutyrate after optimization increased by 35.2 % compared to that before optimization.

Corrigendum: Identification of common Hub genes in human dermal fibroblasts stimulated by mechanical stretch at both the early and late stages.

[This corrects the article DOI: 10.3389/fsurg.2022.846161.].

Macrophage contribution to the survival of transferred expanded skin flap through angiogenesis.

Background: Despite the application of tissue expansion in the reconstruction of significant tissue defects, complications with expanded random-pattern skin flaps remain a major challenge. Insufficient angiogenesis is one of the keys factors in flap ischemia and dysfunction. Macrophages play a key role in promoting tissue angiogenesis, but their effects on expanded flap angiogenesis and the survival of the transferred skin flap are still unknown. Methods: A rat scalp expansion model was established to evaluate the dynamic changes of macrophages in expanded skin. Clodronate liposomes (Clo-lipo) were injected into the expanded scalps to deplete the macrophages, and the expanded scalp flaps with macrophage depletion were orthotopically transferred. The remaining expanded rat scalp flaps were treated with either a macrophage-colony stimulating factor (M-CSF) alone or M-CSF in combination with Clo-lipo and transferred. The number of macrophages, blood perfusion, microvascular densities (MVDs), flap survival, histological changes, and gene expression related to macrophage polarization and angiogenesis were determined with immunofluorescence (IF) staining, full-field laser perfusion imager, hematoxylin and eosin (HE) staining, and quantitative real-time polymerase chain reaction. Results: The number of pan-macrophages significantly increased in the expanded scalp on days 14 and 21 after expander placement. The depletion rate after treatment with Clo-lipo was 29.06%, and the number of macrophages was significantly reduced in the group that underwent Clo-lipo treatment on day 14 before flap transfer (P<0.05). Macrophage depletion resulted in decreased blood perfusion, reduced MVDs, lower expression of factors, and poor survival rate. The recruitment of macrophages with a M-CSF led to higher blood perfusion, increased MVDs, greater expression of angiogenic factors, and better flap survival after flap transfer. Conclusions: Alternatively activated macrophages in the expanded flap could significantly promote angiogenesis, improve blood perfusion, and ultimately increase the flap survival rate. Modulating alternatively activated macrophages may provide a key therapeutic strategy to promote expanded skin flap survival. Our study has provided a basis for clinically improving random-pattern skin flap survival.

Montelukast Attenuates Retraction of Expanded Flap by Inhibiting Capsule Formation around Silicone Expander through TGF-ß1 Signaling.

BACKGROUND: Tissue expansion has tremendous applications in plastic surgery, but flap retraction provides insufficient tissue for use. Inspired by the use of montelukast to suppress capsular contracture, the authors investigated the effects of montelukast on capsule formation around the expander and retraction of the expanded scalp of the rat. METHODS: Thirty-six male Sprague-Dawley rats were randomly divided into control and montelukast groups. In each group, 12 expanded flaps with or without capsules were harvested for histologic and molecular analysis; the six remaining expanded flaps were transferred to repair defects. Myofibroblast and transforming growth factor-ß1 expression in the capsule was determined using immunofluorescence. Capsule ultrastructure was observed using transmission electron microscopy. Related protein expression in the capsules was detected using Western blot analysis. RESULTS: A comparison of control and montelukast groups revealed that areas of the harvested expanded flaps with capsules were greater (2.04 ± 0.11 cm 2 versus 2.42 ± 0.12 cm 2 , respectively; P = 0.04); the retraction rate decreased (41.3% ± 2.16% versus 28.13% ± 2.17%, respectively; P < 0.01). However, the increased areas and decreased retraction disappeared after capsule removal. The number of myofibroblasts declined. Thin, sparse collagen fibers were observed in the capsules. The expression of COL1, COL3, TGF-ß1, EGR1, and phosphorylated ERK1/2 in the capsules decreased. Furthermore, the recipient area repaired by the transferred expanded flap was increased from 4.25 ± 0.39 cm 2 to 6.58 ± 0.31 cm 2 ( P < 0.01). CONCLUSION: Montelukast attenuates retraction of the expanded flap by inhibiting capsule formation through suppressing transforming growth factor-ß1 signaling. CLINICAL RELEVANCE STATEMENT: This study provides novel insights into a method for increasing the area of the expanded flap.

Production of polyhydroxyalkanoate by mixed cultivation of Brevundimonas diminuta R79 and Pseudomonas balearica R90.

The microflora in the activated sludge of propylene oxide saponification wastewater is characterized by a clear succession after enrichment and domestication, and the specifically enriched strains can significantly increase the yield of polyhydroxyalkanoate. In this study, Pseudomonas balearica R90 and Brevundimonas diminuta R79, which are dominant strain after domestication, were selected as models to examine the interactive mechanisms associated with the synthesis of polyhydroxyalkanoate by co-cultured strains. RNA-Seq analysis revealed the up-regulated expression of the acs and phaA genes of strains R79 and R90 in the co-culture group, which enhanced their utilization of acetic acid and synthesis of poly-ß-hydroxybutyrate. Cell dry weight and the yield of poly-ß-hydroxybutyrate in the co-culture group were accordingly considerably higher than those in the respective pure culture groups. In addition, two-component system, quorum-sensing, flagellar synthesis-related, and chemotaxis-related genes were enriched in strain R90, thereby indicating that compared with the R79 strain, R90 can adapt more rapidly to a domesticated environment. Expression of the acs gene was higher in R79 than in R90, and consequently, strain R79 could more efficiently assimilate acetate in the domesticated environment, and thus predominated in the culture population at the end of the fermentation period.

Microbial production of valuable chemicals by modular co-culture strategy.

Nowadays, microbial synthesis has become a common way for producing valuable chemicals. Traditionally, microbial production of valuable chemicals is accomplished by a single strain. For the purpose of increasing the production titer and yield of a recombinant strain, complicated pathways and regulation layers should be fine-tuned, which also brings a heavy metabolic burden to the host. In addition, utilization of various complex and mixed substrates further interferes with the normal growth of the host strain and increases the complexity of strain engineering. As a result, modular co-culture technology, which aims to divide a target complex pathway into separate modules located at different single strains, poses an alternative solution for microbial production. Recently, modular co-culture strategy has been employed for the synthesis of different natural products. Therefore, in this review, various chemicals produced with application of co-cultivation technology are summarized, including co-culture with same species or different species, and regulation of population composition between the co-culture members. In addition, development prospects and challenges of this promising field are also addressed, and possible solution for these issues were also provided.

Metformin Promotes Mechanical Stretch-Induced Skin Regeneration by Improving the Proliferative Activity of Skin-Derived Stem Cells.

Background: Skin expansion by mechanical stretch is an essential and widely used treatment for tissue defects in plastic and reconstructive surgery; however, the regenerative capacity of mechanically stretched skin limits clinical treatment results. Here, we propose a strategy to enhance the regenerative ability of mechanically stretched skin by topical application of metformin. Methods: We established a mechanically stretched scalp model in male rats (n = 20), followed by their random division into two groups: metformin-treated (n = 10) and control (n = 10) groups. We measured skin thickness, collagen volume fraction, cell proliferation, and angiogenesis to analyze the effects of topical metformin on mechanically stretched skin, and immunofluorescence staining was performed to determine the contents of epidermal stem cells and hair follicle bulge stem cells in mechanically stretched skin. Western blot was performed to detect the protein expression of skin-derived stem cell markers. Results: Compared with the control group, metformin treatment was beneficial to mechanical stretch-induced skin regeneration by increasing the thicknesses of epidermis (57.27 ± 10.24 vs. 31.07 ± 9.06 µm, p < 0.01) and dermis (620.2 ± 86.17 vs. 402.1 ± 22.46 µm, p < 0.01), number of blood vessels (38.30 ± 6.90 vs. 17.00 ± 3.10, p < 0.01), dermal collagen volume fraction (60.48 ± 4.47% vs. 41.28 ± 4.14%, p < 0.01), and number of PCNA+, Aurora B+, and pH3+ cells. Additionally, we observed significant elevations in the number of proliferating hair follicle bulge stem cells [cytokeratin (CK)15+/proliferating cell nuclear antigen (PCNA)+] (193.40 ± 35.31 vs. 98.25 ± 23.47, p < 0.01) and epidermal stem cells (CK14+/PCNA+) (83.00 ± 2.38 vs. 36.38 ± 8.96, p < 0.01) in the metformin-treated group, and western blot results confirmed significant increases in CK14 and CK15 expression following metformin treatment. Conclusion: Topical application of metformin enhanced the regenerative capacity of mechanically stretched skin, with the underlying mechanism possibly attributed to improvements in the proliferative activity of skin-derived stem cells.

Identification of Common Hub Genes in Human Dermal Fibroblasts Stimulated by Mechanical Stretch at Both the Early and Late Stages.

Background: Mechanical stretch is vital for soft tissue regeneration and development and is utilized by plastic surgeons for tissue expansion. Identifying the common hub genes in human dermal fibroblasts (HDFs) stimulated by mechanical stretch at different stages will help elucidate the mechanisms involved and improve the efficiency of tissue expansion. Methods: A gene expression dataset (GSE58389) was downloaded from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) in HDFs between cyclic mechanical stretching and static samples were identified at 5 and 24 h. Common DEGs overlapped in both the 5 h and 24 h groups. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed to determine the functions of the DEGs. Protein-protein interaction networks were constructed using the STRING database. The top 10 hub genes were selected using the plug-in Cytohubba within Cytoscape. The regulatory network of hub genes was predicted using NetworkAnalyst. Results: A total of 669 and 249 DEGs were identified at the early (5 h) and late stages (24 h), respectively. Of these, 152 were present at both stages and were designated as common DEGs. The top enriched GO terms were "regulation of autophagy" at the early stage, and "sterol biosynthetic processes" at the late stage. The top KEGG terms were "pyrimidine metabolism" and "synaptic vesicle cycle" at the early and late stages, respectively. Seven common DEGs [DEAD-box helicase 17 (DDX17), exocyst complex component 7 (EXOC7), CASK interacting protein 1 (CASKIN1), ribonucleoprotein PTB-binding 1 (RAVER1), late cornified envelope 1D (LCE1D), LCE1C, and polycystin 1, transient receptor potential channel interacting (PKD1)] and three common DEGs [5'-3' exoribonuclease 2 (XRN2), T-complex protein 1 (TCP1), and syntaxin 3 (STX3)] were shown to be upregulated and downregulated hub genes, respectively. The GO terms of the common hub genes were "skin development" and "mRNA processing." After constructing the regulatory network, hsa-mir-92a-3p, hsa-mir-193b-3p, RNA polymerase II subunit A (POLR2A), SMAD family member 5 (SMAD5), and MYC-associated zinc finger protein (MAZ) were predicted as potential targets in both stages. Conclusion: At the early stage, there were clear changes in gene expression related to DNA and chromatin alterations; at late stages, gene expression associated with cholesterol metabolism was suppressed. Common DEGs related to skin development, transcriptional regulation, and cytoskeleton rearrangement identified in both stages were found to be potential targets for promoting HDF growth and alignment under mechanical stretch.

Identification and Characterization of a Novel Endo-ß-1,4-Xylanase from Streptomyces sp. T7 and Its Application in Xylo-Oligosaccharide Production.

A xylanase-producing strain, identified as Streptomyces sp. T7, was isolated from soil by our lab. The endo-ß-1,4-xylanase (xynST7) gene was found in the genome sequence of strain T7, which was cloned and expressed in Escherichia coli. XynST7 belonged to the glycoside hydrolase family 10, with a molecular mass of approximately 47 kDa. The optimum pH and temperature of XynST7 were pH 6.0 and 60 °C, respectively, and it showed wide pH and temperature adaptability and stability, retaining more than half of its enzyme activity between pH 5.0 and 11.0 below 80 °C. XynST7 showed only endo-ß-1,4-xylanase activity without cellulase- or ß-xylosidase activity, and it showed maximal hydrolysis for corncob xylan in all the test substrates. Then, XynST7 was used for the production of xylo-oligosaccharides (XOSs) by hydrolyzing xylan extracted from raw corncobs. The maximum yield of the XOS was 8.61 ± 0.13 mg/mL using 15 U/mL of XynST7 and 1.5% corncob xylan after 10 h of incubation at 60 °C. The resulting hydrolysate products mainly consisted of xylobiose and xylotriose. These data indicated that XynST7 might by a promising tool for various industrial applications.

Deciphering Key Foreign Body Reaction-Related Transcription Factors and Genes Through Transcriptome Analysis.

Background: Silicone implants are widely used in the field of plastic surgery for wound repair and cosmetic augmentation. However, molecular mechanisms and signaling pathways underlying the foreign body reaction (FBR) of a host tissue to the silicone require further elucidation. The purpose of this study was to identify key FBR-related transcription factors (TFs) and genes through transcriptome analysis. Methods: We used a rat model with a subcutaneous silicone implant in the scalp and performed high throughput sequencing to determine the transcriptional profiles involved in the FBR. The function was analyzed by Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway-enrichment analysis. A protein-protein interaction (PPI) network of differentially expressed mRNAs (DEmRNAs) was constructed to identify the hub genes and key modules and to determine the regulatory TF-mRNA relationships. In addition, the hub gene and transcript expression levels were determined by Quantitative Reverse Transcription polymerase Chain Reaction (qRT-PCR). Myofibroblasts differentiation and macrophage recruitment were identified by immunofluorescence. The protein expression of MMP9 was detected by immunohistochemistry and Western blot. Results: We identified ten hub genes (Fos, Spp1, Fn1, Ctgf, Tlr2, Itgb2, Itgax, Ccl2, Mmp9, and Serpine1) and 3 TFs (FOS, IRF4, and SPI1) that may be crucial (particularly FOS) for the FBR. Furthermore, we identified multiple differentially expressed genes involved in several important biological processes, including leukocyte migration, cytokine‒ cytokine receptor interaction, phagocytosis, extracellular matrix (ECM) organization, and angiogenesis. We also identified potentially significant signaling pathways, including cytokine‒cytokine receptor interaction, phagosome, ECM‒receptor interaction, complement and coagulation cascades, the IL-17 signaling pathway, and the PI3K‒Akt signaling pathway. In addition, qRT-PCR confirmed the expression patterns of the TFs and hub genes, Western blot and immunohistochemistry validated the expression patterns of MMP9. Conclusion: We generated a comprehensive overview of the gene networks underlying the FBR evoked by silicone implants. Moreover, we identified specific molecular and signaling pathways that may perform key functions in the silicone implant-induced FBR. Our results provide significant insights into the molecular mechanisms underlying silicone-induced FBR and determine novel therapeutic targets to reduce complications related to silicone implantation.

Metabolic engineering for biosynthesis of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) from glucose and propionic acid in recombinant Escherichia coli.

In this study, novel polyhydroxyalkanoate (PHA)-associated genes (phaCp and phaABp) cloned from Propylenella binzhouense L72T were expressed in Escherichiacoli cells for PHA production, and the recombinant strains were used to analyze PHA yields with various substrates. The highest poly (3-hydroxybutyrate-co-3-hydroxy-valerate) (PHBV) yield (1.06 g/L) and cell dry weight (3.31 g/L) in E. coli DH5α/ΔptsG-CpABp were achieved by using glucose and propionicacid as substrates. Structural verification of PHBV produced by E. coli DH5α/ΔptsG-CpABp was performed to evaluate the characteristics of the polymers using Fourier transform infrared spectroscopy and nuclear magnetic resonance analysis. In addition, the X-ray diffraction results showed improved crystallinity of PHBV, and thermogravimetric analysis showed good thermal stability of 298 °C. The above findings indicated that the expression of phaCp and phaABp genes resulted in improved PHBV synthesis activity, and the polymer had better performance at higher processing temperatures.

Biosynthesis of vanillin by different microorganisms: a review.

Vanillin is a popular flavoring agent widely used around the world. Vanillin is generated by natural extraction, chemical synthesis, or tissue culture technology, but these production methods no longer meet the increasing worldwide demand for vanillin. Accordingly, a biotechnological approach may provide an effective replacement route to obtaining vanillin. Processes for environmentally friendly production of vanillin in microorganisms from different carbon sources, such as eugenol, isoeugenol, lignin, ferulic acid, sugars, and waste residues, with high productivity and yield have been developed. However, challenges remain for optimizing the vanillin biosynthesis process and further improving production titer and yield. In this review, successful and applicable strategies for increasing vanillin titer and yield in different microorganisms are summarized. Additionally, perspectives for further optimizing the production of vanillin are discussed.

Paracoccus shandongensis sp. nov., Isolated from Activated Sludge.

A novel strain, wg2T, was isolated from activated sludge obtained from wastewater treatment plant in Shandong province, China. The bacterium was Gram-strain-negative, aerobic, rod-shaped, non-flagellated and non-gliding. This bacterium was characterized to determine its taxonomic position using the polyphasic approach. Strain wg2T grew at 25-45 °C (optimum, 30 °C), at salinities of 0-7.0% (w/v) NaCl (optimum, 0-2.0%) and at pH 7-9 (optimum, pH 7.0). Phylogenetic analysis based on 16S rRNA gene sequence showed that strain wg2T clustered with species of genus Paracoccus and shares high similarities with Paracoccus sediminis DSM 26170 T (98.1%) and Paracoccus fontiphilus MVW-1 T (97.7%), respectively. The genome size of strain wg2T was 3.93 Mbp and the DNA G + C content was 66.05%. The dDDH values and ANI between strain wg2T and each of reference strains P. sediminis DSM 26170 T, P. fontiphilus MVW-1 T and P. denitrificans DSM 413 T were 18.3, 12.5, 24.5% and 85.3, 87.0, 78.4%, respectively. The major respiratory quinone was found to be Q-10 and the major fatty acid was C18:1 ω7c. The polar lipids consisted of aminoglycolipid (AGL), phosphatidylcholine (PC), glycolipid (GL), phosphatidylserine (PS), phosphatidylglycerol phosphate (PGP), aminophospholipids (APL). Combining above descriptions, strain wg2T should represent a novel species of genus Paracoccus, for which the name Paracoccus shandongensis sp. nov., is proposed. The type strain is wg2T (= KCTC 72862 T = CCTCC AB 2019401 T).

Mesorhizobium xinjiangense sp. nov., isolated from rhizosphere soil of Alhagi sparsifolia.

A beige-pigmented, Gram-strain-negative, aerobic, rod-shaped, non-flagellated and non-gliding bacterium, designated strain lm94T, was isolated from rhizosphere soil of Alhagi sparsifolia obtained from Alar city, located in Xinjiang province, China. Growth occurred at 20-45 °C (optimum, 37 °C), in the presence of 0-6% (w/v) NaCl (optimum, 0-1%) and at pH 6.0-9.5 (optimum, pH 7.0-7.5). Phylogenetic analysis based on 16S rRNA gene sequence showed that strain lm94T belonged to the genus Mesorhizobium, with highest sequence similarity to Mesorhizobium wenxiniae WYCCWR 10195T (96.6%). Genome sequencing revealed a genome size of 5 256 375 bp and a G + C content of 63.6 mol%. The average nucleotide identity value and the digital DNA-DNA hybridization value between strain lm94T and M. wenxiniae LMG 30254T were 75.0% and 20.0%, respectively. The major respiratory quinone was Q-10. The major fatty acids were C19:0 cyclo ω8c and Summed Feature 8 (C18:1 ω6c and/or C18:1 ω7c) and its polar lipids consisted of phosphatidylethanolamine (PE), phosphatidylglycerol (PG), unidentified phospholipid (PL), phosphatidylcholine (PC), diphosphatidylglycerol (DPG), unidentified aminolipid (AL), unknown glycolipid (GL), unidentified aminophospholipid (APL2) and unidentified polar lipid (L1 and L2). On the basis of these data, strain lm94T is considered to represent a novel species of the genus Mesorhizobium, for which the name Mesorhizobium xinjiangense sp. nov. is proposed. The type strain is lm94T (=KCTC 72863T=CCTCC AB2019377T).

Biosynthesis of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) in metabolically recombinant Escherichia coli.

In this study, a phaCR gene encoding PHA synthase was identified in Rhodoligotrophos defluvii which was adjacent to ß-ketothiolase encoded by phaAR gene and acetoacetyl-CoA reductase encoded by phaBR gene. Amino acid comparison of PhaCR showed the highest homology of 65.98% with PhaC of R. appendicifer, while its homology with typical class I PHA synthase in Cupriavidus necator was only 42.54%. PHA synthesis genes were then transformed into E. coli harboring phaCABR and phaCRABC which were cultured with 15 g/L glucose respectively, and 20.46 wt% and 16.95 wt% of CDW for poly(3-hydroxybutyrate) (PHB) were accumulated respectively. To further explore the effect of substrate specificity for PHA production, the ptsG gene was then deleted and 15 g/L glucose and 1.5 g/L propionate were co-employed as carbon sources, which enabled the synthesis of poly(3HB-co-3HV) copolymer. As a result, poly(3HB-co-3HV) was accumulated up to 24.74 wt% of CDW, and the highest content of 3-hydroxyvalerate (3HV) was 10.86 mol%. The Td5 was 260 °C, which implied that it possessed good thermal stability, and the Mw of GPC in recombinant strains were between 22 and 26 × 104 g/mol, and the highest PDI was 3.771. The structure of poly (3HB-co-3HV) copolymer was determined through 1H NMR analysis.