Biorefining process of agricultural onions to functional vinegar.

Biorefinery of onion vinegar (OV) is attractive as a method for producing functional foods from onions or onion by-products. In this study, a two-stage fermentation of OV using Saccharomyces cerevisiae ATCC9763 and Acetobacter pasteurianus CICC20001 was carried out at 28 °C, the titratable acidity reached 4.01%, and the YA/E was 69.64% at 72 h. Based on this, semi-continuous fermentation was performed, proceeded to charge-discharge consisting of three cycles, and the yield, productivity, and specific production rate were 76.71%, 17.73 g/(L·d), and 20.51 h-1, respectively, which was higher than fed-batch fermentation. The in vivo antioxidant experiments showed that OV significantly increased GSH-Px, SOD, and CAT enzyme activities of Caenorhabditis elegans at 271.57, 129.26, and 314.68%, respectively. Nutritional analysis revealed that the total flavonoids and polyphenols were 3.01 mg/mL and 976.76 µg/mL, respectively. It was also shown that the acetic acid to total organic acid (A/T) ratio of OV was 79.02%, and the total free amino acid content was 262.30 mg/100 mL, 1.78-7.44 times higher than other fruit vinegar. The OV prepared in this study showed higher quality than the commercial vinegar.

Preparation of Dendrobium officinale Flower Anthocyanin and Extended Lifespan in Caenorhabditis elegans.

The Dendrobium officinale flower is a non-medicinal part of the plant, rich in a variety of nutrients and bioactive ingredients. The purpose of this article was to explore the preparation conditions of anthocyanins (ACNs) from the D. officinale flower. Subsequently, its anti-aging effects were evaluated with Caenorhabditis elegans. Results showed that the ACNs had antioxidant activities on scavenging free radicals (DPPH· and ABTS+·), and the clearance rate was positively correlated with the dose. Additionally, ACNs significantly increased the activity of superoxide dismutase (SOD) in C. elegans, which was 2.068-fold higher than that of the control. Treatment with ACNs at 150 µL extended the lifespan of C. elegans by 56.25%, and treatment with ACNs at 50 µL promoted fecundity in C. elegans. Finally, the protective effect of ACNs enhanced stress resistance, thereby increasing the survival numbers of C. elegans, which provided insights for the development and practical application of functional products.

Onion Vinegar Quality Evaluation and its Alleviate Oxidative Stress Mechanism in Caenorhabditis elegans Via SKN-1.

Recently, there has been renewed interest in biorefining of agricultural onion into functional products. In this study, onion vinegar (OV) are prepared by a two-stage semi-continuous fermentation method, and its content of total flavonoids (3.01 mg/mL) and polyphenols (976.76 µg/mL) is superior to other commercial vinegars. OV possesses a high radical scavenging activity and enhances the antioxidant enzyme activities in vivo, alleviating intracellular oxidative stress in Caenorhabditis elegans. Treated by OV, the 1,1-diphenyl-2-picryl-hydrazyl radical (DPPH·), diammonium 2,2'-azino-bis (3-ethylbenzo thiazoline-6-sulfonic acid) (ABTS+·) and 2-phenyl-4,4,5,5- tetramethylimidazoline-1-oxyl 3-Oxide (PTIO·) free radicals clearance rates are 88.76, 98.76 and 90.54%, respectively in vitro. Whereas the glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and catalase (CAT) enzyme activities in C. elegans reach 271.57, 129.26, and 314.68%, respectively. Using RNAi and RT-PCR, it has been further confirmed that OV modulates transcription factor SKN-1, the nuclear factor erythroid 2-related factor 2 (Nrf2) homologous, in C. elegans, enhancing the resistance of C. elegans against sodium arsenite stress. Lifespan analysis reveals that 1 mL OV extends the maximum lifespan of the nematode to 26 days. Evidence is presented which shows that OV increases the lifespan of C. elegans by activating the SKN-1 signaling pathway. Overall, the OV is a well functional condiment, enhancing the value-added of onion.

Identification and genomic analysis of two novel duck-origin GPV-related parvovirus in China.

BACKGROUND: Since early 2015, mule duck and Cherry Valley duck flocks have been suffering from short beak and dwarfism syndrome. This widely spreading infectious disease is characterized by growth retardation, smaller beak and tarsus with high morbidity and low mortality rate. For better understanding, we identified and characterized virus isolates named AH and GD from diseased Cherry Valley duck and mule duck flocks and investigated the damage caused by novel parvovirus-related virus (NGPV) to tissues and organs, including kidney, brain, pancreas, liver, spleen, bursa of fabricius and myocardial tissues. RESULTS: AH and GD isolates shared high nucleotide identity with goose parvovirus (GPV). Alignment studies of AH and GD isolates showed 94.5-99.2% identity with novel parvovirus-related virus (NGPV), 98.7-91.5% identity with GPV and 79.9-83.7% with muscovy duck parvovirus (MDPV). Compared with other NGPV, classical GPV and MDPV sequences, a four 14-nucleotide-pair insertion in GD isolate was found in left open reading frame (ORF) (87-100 nt and 350-363 nt) and in right ORF (4847-4861 nt and 5122-5135 nt). However, in AH isolate, a five 14-nucleotide-pair deletions similar to other NGPV were found. The complete genome sequence comparison of eleven NGPV isolates from mule ducks and cherry valley ducks revealed no remarkable difference between them. Notably, the myocardium and bursa of fabricius of both disease and healthy animals are perfectly normal while other tissues have inflammatory cells exudation. CONCLUSIONS: The AH and GD strains are novel parvovirus-related virus that isolates from mule ducks or cherry valley ducks which DNA sequence has no remarkable difference. The histopathology of tissues and organs such as kidney, brain etc. revealed non-significant changes in experimental and control animals. Overall, this study has contributed better understanding of molecular biology of NGPV strains and will help to develop the candidate strain for vaccine preparation to get better protection against these viral infections.

The nutritive value of cassava starch extraction residue for growing ducks.

The cassava root meal (CRM) has been utilized as a cheap energy alternative to replace maize in poultry diets. Recently, the CRM in turn has an increasing demand for starch extraction industry, which renders large amounts of residues. This study evaluated the nutrient composition, amino acid profile, and feeding value of cassava starch extraction residue meal (CReM) for growing ducks. A total of 960, 11-day-old, ducklings were housed in 24 floor pens and allocated randomly into four dietary treatments: (i) 0CReM (control), (ii) 50 g CReM/kg, (iii) 100 g CReM/kg, and (iv) 150 g CReM/kg. The analyses (/kg) of CReM showed high gross energy (3306.88 kcal), ME (2109.54 kcal), and starch (514.0 g), with poor crude protein (20.9 g) and moderate crude fiber (140.0 g) and ash (60.0 g) contents. The total amino acid (AA) content amounted to 19.9 g/kg of CReM DM, in which the methionine, lysine, cystine, and isoleucine were present in low levels. The dietary inclusion of CReM up to 150 g/kg, between 11 and 42 days of age, had no significant effects (P > 0.05) on duck growth parameters, mortality, dressed weight, internal organs, or abdominal fat. Besides, the tested CReM levels did not show any significant effect on the blood proteins or liver enzymes. The results, therefore, revealed that the CReM contains a considerable amount of energy and could be incorporated successfully up to 150 g/kg in the diets of growing ducks.

Combinatorial metabolic engineering of industrial Gluconobacter oxydans DSM2343 for boosting 5-keto-D-gluconic acid accumulation.

BACKGROUND: L-(+)-tartaric acid (L-TA) is an important organic acid, which is produced from the cream of tartar or stereospecific hydrolysis of the cis-epoxysuccinate. The former method is limited by the availability of raw material and the latter is dependent on the petrochemical material. Thus, new processes for the economical preparation of L-TA from carbohydrate or renewable resource would be much more attractive. Production of 5-keto-D-gluconate (5-KGA) from glucose by Gluconobacter oxydans is the first step to produce L-TA. The aim of this work is to enhance 5-KGA accumulation using combinatorial metabolic engineering strategies in G. oxydans. The sldAB gene, encoding sorbitol dehydrogenase, was overexpressed in an industrial strain G. oxydans ZJU2 under a carefully selected promoter, P0169. To enhance the efficiency of the oxidation by sldAB, the coenzyme pyrroloquinoline quinone (PQQ) and respiratory chain were engineered. Besides, the role in sldAB overexpression, coenzyme and respiratory chain engineering and their subsequent effects on 5-KGA production were investigated. RESULTS: An efficient, stable recombinant strain was constructed, whereas the 5-KGA production could be enhanced. By self-overexpressing the sldAB gene in G. oxydans ZJU2 under the constitutive promoter P0169, the resulting strain, G. oxydans ZJU3, produced 122.48 ± 0.41 g/L of 5-KGA. Furthermore, through the coenzyme and respiratory chain engineering, the titer and productivity of 5-KGA reached 144.52 ± 2.94 g/L and 2.26 g/(L · h), respectively, in a 15 L fermenter. It could be further improved the 5-KGA titer by 12.10 % through the fed-batch fermentation under the pH shift and dissolved oxygen tension (DOT) control condition, obtained 162 ± 2.12 g/L with the productivity of 2.53 g/(L · h) within 64 h. CONCLUSIONS: The 5-KGA production could be significantly enhanced with the combinatorial metabolic engineering strategy in Gluconobacter strain, including sldAB overexpression, coenzyme and respiratory chain engineering. Fed-batch fermentation could further enlarge the positive effect and increase the 5-KGA production. All of these demonstrated that the robust recombinant strain can efficiently produce 5-KGA in larger scale to fulfill the industrial production of L-TA from 5-KGA.

Global phylogeny of Shigella sonnei strains from limited single nucleotide polymorphisms (SNPs) and development of a rapid and cost-effective SNP-typing scheme for strain identification by high-resolution melting analysis.

The current Shigella sonnei pandemic involves geographically associated, multidrug-resistant clones. This study has demonstrated that S. sonnei phylogeny can be accurately defined with limited single nucleotide polymorphisms (SNPs). By typing 6 informative SNPs using a high-resolution melting (HRM) assay, major S. sonnei lineages/sublineages can be identified as defined by whole-genome variation.

Identification and genomic analysis of two duck-origin Tembusu virus strains in southern China.

Egg-laying duck flocks in Guangdong province, southern China, have been suffering a widely spreading infectious disease with abrupt egg drops and death since the winter of 2010. However, the causative pathogen was not known. We obtained two independent virus isolates named FS and JM from the diseased layer duck flocks and identified them as duck-origin Tembusu virus by PCR detection, sequencing the entire length of the open reading frames (ORFs). The two isolates FS and JM shared high sequence similarity to the isolates of duck-origin Tembusu virus that was first emerged in eastern China in April 2010. Blast analysis shows that the whole ORF sequences of FS and JM have the highest similarity (>99 %) to BYD-1(the first reported duck-origin Tembusu virus) and JS804 (the first reported goose-origin Tembusu virus), indicating that the full-length genomes were highly conserved in waterfowl-origin Tembusu viruses. The present study suggests that duck-origin Tembusu viruses have spread fast from eastern China to southern China, causing widely spreading infections. The high conservation of duck-origin Tembusu virus strains provides the genomic basis for choosing the strains for vaccine preparation for better protection against this new virus infection.

[Molecular engineering of transketolase from Escherichia coli and tartaric semialdehyde biosynthesis].

Transketolase (EC 2.2.1.1, TK) is a thiamine diphosphate-dependent enzyme that catalyzes the transfer of a two-carbon hydroxyacetyl unit with reversible C-C bond cleavage and formation. It is widely used in the production of chemicals, drug precursors, and asymmetric synthesis by cascade enzyme catalysis. In this paper, the activity of transketolase TKTA from Escherichia coli K12 on non-phosphorylated substrates was enhanced through site-directed saturation mutation and combined mutation. On this basis, the synthesis of tartaric semialdehyde was explored. The results showed that the optimal reaction temperature and pH of TKTA_M (R358I/H461S/R520Q) were 32 ℃ and 7.0, respectively. The specific activity on d-glyceraldehyde was (6.57±0.14) U/mg, which was 9.25 times higher than that of the wild type ((0.71±0.02) U/mg). Based on the characterization of TKTA_M, tartaric acid semialdehyde was synthesized with 50 mmol/L 5-keto-d-gluconate and 50 mmol/L non-phosphorylated ethanolaldehyde. The final yield of tartaric acid semialdehyde was 3.71 g with a molar conversion rate of 55.34%. Hence, the results may facilitate the preparation of l-(+)-tartaric acid from biomass, and provide an example for transketolase-catalyzed non-phosphorylated substrates.

A Peptidomic Approach to Identify Novel Antigen Biomarkers for the Diagnosis of Tuberculosis.

Background: Here, we conducted a peptidomic study in murine model to identify novel antigen biomarkers for the diagnosis of tuberculosis (TB) with improved performance. Methods: Four recombinant proteins, including Mycobacterium tuberculosis protein 32 (MPT32), Mycobacterium tuberculosis protein 64 (MPT64), culture filtrate protein 10 (CFP10), and phosphate ABC transporter substrate-binding lipoprotein (PstS1) were expressed and intravenously injected into BALB/c mice. The serum were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The concentrations of candidate peptides in serum of suspected TB patients were determined using competitive enzyme-linked immunosorbent assay. Results: A total of 65 peptides from 4 MTB precursor recombinant proteins were identified in mouse serum by LC-MS/MS, of which 5 peptides were selected as candidates for serological analysis. The concentrations of peptides MPT64-2, CFP10-2 and PstS1-2 in TB patients were significantly higher than those in non-TB patients. MPT64-2 exhibited the most promising sensitivity (81.4%), followed by PstS1-2 and CFP10-2. In addition, PstS1-2 had the highest specificity (93.3%), followed by CFP10-2 and MPT64-2. According to the area under the curve (AUC), MPT64-2 (AUC = 0.863), PstS1-2 (AUC = 0.812) and CFP10-2 (AUC = 0.809) exhibited better diagnostic validity. Conclusion: We develop an effective approach to identify new antigen biomarkers via LC-MS/MS-based peptidomics. Multiple peptides exhibit promising efficacy in diagnosis of active TB patients.

Complete genome sequence of the pathogenic bacterium Riemerella anatipestifer strain RA-GD.

Riemerella anatipestifer is a well-described pathogen of waterfowl and other avian species which can cause a great loss to the poultry industry. Here we obtained the complete genome sequence of R. anatipestifer strain RA-GD, which was isolated from an infected duck in Guangzhou, China, and was cultivated in our laboratory.

[Synthesis of pyrroloquinoline quinone by recombinant Gluconobacter oxydans].

Pyrroloquinoline quinone (PQQ), an important redox enzyme cofactor, has many physiological and biochemical functions, and is widely used in food, medicine, health and agriculture industry. In this study, PQQ production by recombinant Gluconobacter oxydans was investigated. First, to reduce the by-product of acetic acid, the recombinant strain G. oxydans T1 was constructed, in which the pyruvate decarboxylase (GOX1081) was knocked out. Then the pqqABCDE gene cluster and tldD gene were fused under the control of endogenous constitutive promoter P0169, to generate the recombinant strain G. oxydans T2. Finally, the medium composition and fermentation conditions were optimized. The biomass of G. oxydans T1 and G. oxydans T2 were increased by 43.02% and 38.76% respectively, and the PQQ production was 4.82 and 20.5 times higher than that of the wild strain, respectively. Furthermore, the carbon sources and culture conditions of G. oxydans T2 were optimized, resulting in a final PQQ yield of (51.32±0.899 7 mg/L), 345.6 times higher than that of the wild strain. In all, the biomass of G. oxydans and the yield of PQQ can be effectively increased by genetic engineering.

[Design and application of guide device for femoral neck section].

OBJECTIVE: To explore the application effect of a new guide device designed according to the anatomical characteristics of the femoral neck cross section to assist the internal fixation of multiple screw of the femoral neck. METHODS: From October to December 2016, 10 adult dry femur specimens, including 7 males and 3 females, aged 37 to 58(47.5±7.5) years old, were selected. Three hollow screws were implanted to simulate the treatment of femoral neck fracture with the new guider technology and the traditional guider technology as the control. The screw location accuracy parameters, screw puncture times, screw parallelism, operation time and fluoroscopy times of the two methods were recorded and compared. RESULTS: Sixty screws were successfully implanted in 20 specimens. There was no significant difference in screw parallelism, operation time and fluoroscopy times between the two methods(P>0.05). There were significant differences in the distance between screw and cortex, the distance between screw and femoral neck, the area ratio between screw and femoral neck, the distance between screws and the number of screw punctures between two methods(P<0.05). It showed that the new type of guide had more advantages than the traditional method in locating screw accuracy and reducing the number of punctures. CONCLUSIONS: Compared with the traditional technique, the new guider and percutaneous puncture technique under two-dimensional fluoroscopy have better localization of internal fixation screw for femoral neck fracture.

Obtuse triangle screw configuration for optimal internal fixation of femoral neck fracture: an anatomical analysis.

OBJECTIVES:: To identify the optimal screw configuration for internal fixation of femoral neck fractures based on anatomic analysis on radiologic imaging. METHODS:: 30 proximal femurs of 15 adults were constructed by CT. 3 femoral neck sections (FNS), the subcapital, medial, and the fundus, were projected on to the lateral femoral trochanteric wall. The simulated 3 screw configurations in the projection of FNS include: 2 inverted equilateral triangles symmetrised to the axis of the FNS (IET-FNS group) or the coronal axis of the proximal femur (IET-PR group) and an obtuse triangle (OT group). The distance between the screws, the distance between the centre of the FNS and the screws, and the area ratio of the triangle/FNS were calculated. RESULTS:: The projection of the FNS on to the lateral femoral trochanteric wall is displayed as a rotating forward ellipse. Measurements of distance between screws, distance between the centre of the FNS to the screws, and the area ratio of triangle/FNS were significantly larger in the OT group than in the IET-FNS and IET-PF groups ( p < 0.05). The values of the 3 parameters in the IET-FNS group were also larger than those in the IET-PF group ( p < 0.05). CONCLUSIONS:: The obtuse triangle screw configuration displayed advantages with respect to the parameters of distance between screws, distance between the centre of FNS to screws, and the triangle area. Therefore, the obtuse triangle screw configuration may be the ideal pattern for internal fixation of femoral neck fractures (Pauwels I and II). This needs to be corroborated with biomechanics testing.

Enhancement of 5-keto-d-gluconate production by a recombinant Gluconobacter oxydans using a dissolved oxygen control strategy.

The rapid and incomplete oxidation of sugars, alcohols, and polyols by the gram-negative bacterium Gluconobacter oxydans facilitates a wide variety of biological applications. For the conversion of glucose to 5-keto-d-gluconate (5-KGA), a promising precursor of the industrial substance L-(+)-tartaric acid, G. oxydans DSM2343 was genetically engineered to strain ZJU2, in which the GOX1231 and GOX1081 genes were knocked out in a markerless fashion. Then, a secondary alcohol dehydrogenase (GCD) from Xanthomonas campestris DSM3586 was heterologously expressed in G. oxydans ZJU2. The 5-KGA production and cell yield were increased by 10% and 24.5%, respectively. The specific activity of GCD towards gluconate was 1.75±0.02 U/mg protein, which was 7-fold higher than that of the sldAB in G. oxydans. Based on the analysis of kinetic parameters including specific cell growth rate (µ), specific glucose consumption rate (qs) and specific 5-KGA production rate (qp), a dissolved oxygen (DO) control strategy was proposed. Finally, batch fermentation was carried out in a 15-L bioreactor using an initial agitation speed of 600 rpm to obtain a high µ for cell growth. Subsequently, DO was continuously maintained above 20% to achieve a high qp to ensure a high accumulation of 5-KGA. Under these conditions, the maximum concentration of 5-KGA reached 117.75 g/L with a productivity of 2.10 g/(L·h).

Genome Sequence of Avirulent Riemerella anatipestifer Strain RA-SG.

Riemerella anatipestifer is a pathogenic bacterium that has spread all over the world and is associated with epizootic infections in waterfowl and other avian species. R. anatipestifer RA-SG is an avirulent strain, isolated from an infected duck in Guangdong province, China. The genome sequence of this species is presented herein.

Complete genome sequence of a newly emerging newcastle disease virus isolated in china.

Goose/GD/2010 is a newly emerging Newcastle disease virus (NDV) isolated from a sick goose flock in southern China. Here, we report the complete genome sequence of this isolate, which belongs to NDV subgenotype VIIb. This is the first report about the complete genome information of an isolate of subgenotype VIIb in China.

[Discerning the femoral neck anteversion (FNA) from the torsion angle on 3D CT].

OBJECTIVE: To discern the differences between femoral neck anteversion (FNA) and torsion angle through 3D CT reconstruction. METHODS: From March 2010 to October 2010,30 healthy adult volunteers' femur were reconstructed by 3D CT, included 15 males and 15 females with an average age of (43.66 +/- 7.57) years old ranging from 25 to 65 years. Display the FNA and the torsion angle by image post-processing, measuring torsion angle by "Center way" and direct measurement of FNA. RESULTS: FNA was the angle between the axle wire of femoral neck and the shape face of femoral,the angle were (13.326 +/- 6.085) degrees. Torsion angle was the angle between the macropinacoid of cross section of femoral neck and the shape face of femoral, the angle were(31.335 +/- 2.079) degrees. There was no significant difference in left and right femur. CONCLUSION: FNA is different from torsion angle. FNA is the angle between the line and the surface with the sharp angle towards the lower outside. The torsion angle is the angle between the two surfaces with the sharp angle towards the lower back.

Expression of Yes-associated protein in non-small cell lung cancer and its relationship with clinical pathological factors.

BACKGROUND: Yes-associated protein (YAP) plays an important role in signal transduction and gene transcription regulation in normal cells, with elevated and over-expressed YAP levels observed in various malignant tumors. The aim of this study was to investigate the expression of YAP in non-small cell lung cancer (NSCLC), and to study the possible relationship of YAP expression with the occurrence and development of NSCLC. METHODS: YAP expression was assessed in 40 cases of NSCLC tumor tissues by immunohistochemistry, and their protein and mRNA levels were evaluated through Western blotting and reverse transcription-polymerase chain reaction (PCR), respectively. Normal lung tissues obtained from the same patient were used as control. Statistical analysis was performed to correlate the YAP expression to clinical pathological factors, such as tumor type, stage and grade. RESULTS: YAP-positive expression was found in 28 (70%) of the 40 cases of NSCLC, which included 10 cases of squamous cell carcinoma (25%), 17 cases of adenocarcinoma (42.5%) and 1 case of squamous adenocarcinoma (2.5%). In the 28 YAP-positive cases, 19 cases showed lymph node metastasis and were classified in TNM stage II + III (47.5%); the other nine cases showed no lymph node metastasis (22.5%) and were classified in the TNM stage I. There was no relationship between YAP expression and patients' age, gender or tumor histological grades. However, YAP showed significant over expression in late period of T stage (P = 0.012), TNM stage (P = 0.039), and lymph node metastasis (P = 0.013), respectively. Notably, YAP-positive expression was significantly higher in adenocarcinoma than that in squamous cell carcinoma (P = 0.041). CONCLUSIONS: Over-expression of YAP was associated with NSCLC, especially lung adenocarcinoma. The high YAP expression in late period of tumor stage and lymph node metastasis may indicate that YAP expression could be an early marker for NSCLC tumorigenesis.

Micropatterning of organic semiconductor microcrystalline materials and OFET fabrication by "hot lift off".

Considerable progress toward the development of electronic devices that rely on organic semiconductors as the active material component has been made in recent years. The key step for realization of the advanced organic electronic, or optical, device is the ability to micropattern different kinds of electronic materials, such as organic semiconductor/conducting materials, over large areas with micrometer-sized resolution. Here we demonstrate a simple and direct method for micropatterning small-molecule microcrystalline films using an epoxy stamp. The "hot lift off" method is highly selective, creating patterns with high resolution and relies on straightforwardly tailoring the adhesive properties between the epoxy and the film. This process is well suited for patterning many types of materials.