Jingping Ge

Statistical optimization of xylitol production from corncob hemicellulose hydrolysate by Candida tropicalis HDY-02.

The statistical experimental designs were adopted to optimize the culture medium in xylitol production by Candida tropicalis HDY-02 with corncob hemicellulose hydrolysate as substrate. In the first step, Plackett-Burman design was used for screening the important variables. KH(2)PO(4), yeast extract, (NH(4))(2)SO(4) and MgSO(4)·7H(2)O were found to significantly affect xylitol yield. In the second step, central composite design (CCD) was used to determine the optimum level of each of the significant variables. A second-order polynomial was determined by the multiple regression analysis of the experimental data. The interactive effects of yeast extract and MgSO(4)·7H(2)O on xylitol yield of C. tropicalis HDY-02 were determined to be significant. The validation experimental was consistent with the prediction model. The optimum combinations for xylitol yield were 5 gl(-1) (NH(4))(2)SO(4), 1.3 gl(-1) KH(2)PO(4), 4.6 gl(-1) yeast extract and 0.6 gl(-1) MgSO(4)·7H(2)O. Under these optimal conditions, the continuous fed-batch experiments could produce xylitol of 58 gl(-1) with a yield of 0.73 g g(-1) xylose.

Purification and Partial Characterization of a Novel Bacteriocin Synthesized by Lactobacillus paracasei HD1-7 Isolated from Chinese Sauerkraut Juice.

Bacteriocins have antimicrobial activities against food-spoiling bacteria and food-borne pathogens. Paracin 1.7, a bacteriocin synthesized by Lactobacillus paracasei HD1-7 isolated from Chinese sauerkraut juice, was studied. Following partial purification with ammonium sulfate precipitation, CM Sepharose Fast Flow, and Sephadex G-10 chromatography, the molecular weight of Paracin 1.7 was about 10 kDa based on Tricine-SDS-PAGE results. A 2.87 fold purified bacteriocin was produced, reaching a final yield of 39.93% and the specific activity of 1.56 × 103 AU/mg. The N-terminal amino acid sequence of Paracin 1.7 was VSNTFFA, and the LC/LTQ results revealed that the N-terminal amino acid sequence was similar to that of ABC-type oligopeptide transport system protein and N-acetylmuramoyl-L-alanine amidase. Paracin 1.7 was sensitive to protease K, had antimicrobial activities at a broad pH range (3.0–8.0), and was heat resistant (121 °C for 20 min). Paracin 1.7 from Lactobacillus paracasei HD1-7 is a novel bacteriocin that has potential applications in food preservation.

Strain Isolation and Study on Process Parameters for Xylose-to-Xylitol Bioconversion

ABSTRACT A xylitol producing yeast was isolated from soil and identified as Candida tropicalis W103 by 18S r DNA gene sequence analysis and physiological characteristics. The optimal fermentation conditions for Candida tropicalis W103 were: 35°C, pH 4.5 and 120 g l−1 initial xylose concentration. However, the maximum yield and productivity of xylitol were obtained at KLa 16.5 h−1 and 18.3 h−1, respectively. A two-stage aeration strategy (0–24 h, KLa 18.3 h−1; after 24 h, shift KLa to 16.5 min−1) was applied in the fermentation to get higher xylitol yield and productivity. After 60 h in batch fermentations, both the xylitol concentration and xylose consumption reached the maximum, obtaining 87.1 g l−1 of xylitol with 1.45 g l−1 h−1 productivity and 0.72 g g−1 xylose yield. Fed-batch fermentation with 120 g l−1 initial xylose and regulating xylose concentration to 40–55 g l−1 during 24–96 h was performed to reach a higher productivity of 1.82 g l−1 h−1, and xylitol concentration of 218.7 g l−1.

Bacterial diversity and community structure during fermentation of Chinese sauerkraut with Lactobacillus casei 11MZ-5-1 by Illumina Miseq sequencing

The bacterial diversity and community structure involved in Chinese sauerkraut is one of the most important factors shaping the final characteristics of traditional foods. In this research, Lactobacillus casei 11MZ‐5‐1 was applied in Chinese sauerkraut fermentation as a starter culture. Illumina Miseq sequencing analysis was used to reveal the bacterial diversity and community structure during Chinese sauerkraut fermentation. A total of 177 283 high‐quality reads of 16S rRNA V4 regions were obtained. The inoculation of L. casei 11MZ‐5‐1 decreased considerably the bacterial richness and bacterial diversity. This inoculum led to the replacement of Lactococcus by Lactobacillus. The levels of Pseudomonas and Enterobacter bacteria decreased. These findings reveal the evolution of important bacterial groups that are involved in fermentation and will facilitate improvements in the Chinese sauerkraut fermentation process.

Bacterial succession and metabolite changes during flax (Linum usitatissimum L.) retting with Bacillus cereus HDYM-02.

High-throughput sequencing and GC-MS (gas chromatography-mass spectrometry) were jointly used to reveal the bacterial succession and metabolite changes during flax (Linum usitatissimum L.) retting. The inoculation of Bacillus cereus HDYM-02 decreased bacterial richness and diversity. This inoculum led to the replacement of Enterobacteriaceae by Bacillaceae. The level of aerobic Pseudomonadaceae (mainly Azotobacter) and anaerobic Clostridiaceae_1 gradually increased and decreased, respectively. Following the addition of B. cereus HDYM-02, the dominant groups were all degumming enzyme producers or have been proven to be involved in microbial retting throughout the entire retting period. These results could be verified by the metabolite changes, either degumming enzymes or their catalytic products galacturonic acid and reducing sugars. The GC-MS data showed a clear separation between flax retting with and without B. cereus HDYM-02, particularly within the first 72 h. These findings reveal the important bacterial groups that are involved in fiber retting and will facilitate improvements in the retting process.

Construction of recombinant baculoviruses expressing hemagglutinin of H5N1 avian influenza and research on the immunogenicity

Recombinant baculoviruses with different promoter and regulatory elements were constructed to enhance the expression of target protein and boost the efficacies of avian influenza vaccine. Hemagglutinin gene was cloned into the baculovirus transfer vectors driven by cytomegaloviru (CMV) and White spot syndrome virus immediate-early promoter one (WSSV ie1) promoter respectively, with different regulatory elements. The recombinant baculoviruses were directly used as vaccines to immunize specific pathogen-free chickens. The protein expression levels of recombinant baculoviruses BV-S-HA and BV-S-ITRs-HA were respectively 2.43 and 2.67 times than that of BV-S-con-HA, while the protein expression levels of BV-A-HA and BV-A-ITRs-HA were respectively 2.44 and 2.69 times than that of BV-S-con-HA. Immunoglobulin G (IgG) antibody levels induced by BV-A and BV-S series recombinant baculovirus were significantly higher than the commercialized vaccine group (P < 0.05). Among the groups with same promoter, the IgG antibody levels induced by the baculovirus containing regulatory elements were significantly higher than control group. Additionally, the immune effects induced by BV-A series recombinant baculoviruses with WSSV ie1 promoter were significantly stronger than the BV-S series recombinant baculoviruses with CMV promoter. The avian influenza vaccine prepared based on baculovirus vector can simultaneously stimulate the humoral and cellular immune responses.

Bio-chemical characterization of a β-mannanase from Bacillus licheniformis HDYM-04 isolated from flax water-retting liquid and its decolorization ability of dyes

A β-mannanase was purified from the bacteria, Bacillus licheniformis HDYM-04, which was a high β-mannanase-producing strain (576.16 U mL−1 at 48 h during fermentation). The optimal temperature and pH for this enzyme activity were 60 °C and 8.0, respectively. The enzyme activity was significantly enhanced by Co2+, Mg2+, K+, Ba2+ and Na+ while inhibited by β-mercaptoethanol, DTT, SDS, Tween and EDTA. It was also noticed that the enzyme did not lose its activity in a wide interval such as high NaCl concentration (5 mol L−1) and high urea concentration (5 mol L−1). When amino acid residues were modified with NEM, N-AI and NBS, the enzyme activities significantly decreased. CD spectra demonstrated that the secondary structure of β-mannanase consists of 0% α-helix, 27.9% β-sheet, 24.4% turn and 47.7% random coil. CD spectra also supported that β-mannanase was stable under temperature and pH variation. Ten structurally different dyes could be effectively decolorized by β-mannanase within 24 h, in which Congo red (100%), malachite green (100%), titan yellow (98.6%) and phenol red (91%) had high decolorization rates. This property showed the compounds potential application in textile and paper-making industries to decolorize redundant dyes on them.

Flax retting by degumming composite enzyme produced by Bacillus licheniformis HDYM-04 and effect on fiber properties

Abstract Flax enzymatic retting with composite enzyme produced by microbes with inexpensive substrates is widely researched due to less contamination and lower cost. Bacillus licheniformis HDYM-04, isolated from a liquid sample of flax retting pool, efficiently produced degumming enzymes after 48 h of fermentation with inexpensive konjaku flour, consisted of 587.5 U/mL pectinase, 365.2 U/mL mannanase, and 140.1 U/mL xylanase. Almost half the maximum activity of three above-mentioned degumming enzymes was maintained at pH 4.0–6.0 which demonstrated its stability in pH condition of flax retting. After 120 h of retting with this composite enzyme, scanning electronic microscopy showed more significant reduction in gummy components on the fiber surface than those of water retting. The fiber strength was 182.4 ± 9.3 N, 14.3% higher than water-retted samples. The long fiber rate and fiber yield also verified higher fiber productivity. The results permitted this degumming composite enzyme an applicable potential in flax retting.

Construction of recombinant baculovirus vaccines for Newcastle disease virus and an assessment of their immunogenicity

Newcastle disease (ND) is a lethal avian infectious disease caused by Newcastle disease virus (NDV) which poses a substantial threat to Chinas poultry industry. Conventional live vaccines against NDV are available, but they can revert to virulent strains and do not protect against mutant strains of the virus. Therefore, there is a critical unmet need for a novel vaccine that is safe, efficacious, and cost effective. Here, we designed novel recombinant baculovirus vaccines expressing the NDV F or HN genes. To optimize antigen expression, we tested the incorporation of multiple regulatory elements including: (1) truncated vesicular stomatitis virus G protein (VSV-GED), (2) woodchuck hepatitis virus post-transcriptional regulatory element (WPRE), (3) inverted terminal repeats (ITRs) of adeno-associated virus (AAV Serotype II), and (4) the cytomegalovirus (CMV) promoter. To test the in vivo efficacy of the viruses, we vaccinated chickens with each construct and characterized the cellular and humoral immune response to challenge with virulent NDV (F48E9). All of the vaccine constructs provided some level of protection (62.5-100% protection). The F-series of vaccines provided a greater degree of protection (87.5-100%) than the HN-series (62.5-87.5%). While all of the vaccines elicited a robust cellular and humoral response subtle differences in efficacy were observed. The combination of the WPRE and VSV-GED regulatory elements enhanced the immune response and increased antigen expression. The ITRs effectively increased the length of time IFN-γ, IL-2, and IL-4 were expressed in the plasma. The F-series elicited higher titers of neutralizing antibody and NDV-specific IgG. The baculovirus system is a promising platform for NDV vaccine development that combines the immunostimulatory benefits of a recombinant virus vector with the non-replicating benefits of a DNA vaccine.

Construction of Recombinant Baculoviruses Expressing Infectious Bursal Disease Virus Main Protective Antigen and Their Immune Effects on Chickens.

In order to overcome the limitations of conventional vaccines for infectious bursal disease virus (IBDV), we constructed recombinant dual expression system baculoviruses with VP2 and VP2/4/3, the main protective antigens of IBDV. We compared the immune effects of the baculoviruses in avian cells and detected their control effects on chickens with infectious bursal disease. We used Western blot analysis to measure VP2 protein and VP2/4/3 polyprotein expression in avian cells infected using the Bac-to-Bac baculovirus expression system. The recombinant baculoviruses were used to vaccinate specific pathogen-free chickens, which produced specific protective antibodies and strong cellular immune responses. The results of the virus challenge experiment revealed that the protective efficiency of VP2 and VP2/4/3 virus vaccines were 95.8% and 100%, respectively, both of which were higher than the vaccine group (87.5%), and significantly higher than the control group (50%). The results demonstrated that the immune effect of BV-S-ITRs-VP2/4/3 was superior to that of BV-S-ITRs-VP2. Compared with traditional attenuated vaccine and genetically engineered live vector vaccine, the dual expression viral vector vaccine has good bio-safety. The results of this study provide a foundation for the further development of poultry vaccines, in addition to providing a useful reference for developing non-replicating live vaccines against other viral diseases.