Vietnam Journal of Biotechnology

The yeast Pichia anomala VTCC Y0787 was optimized for maximum lipase production. Ihe pH optimum for this secretion was 9.0; this lipase was active in an alkaline pH range (pH > 9) and had a potential application in detergent industry. We used the design of optimum multifactorial experiments Plackett-Burman to estimate level effect of physical and chemical factors on lipase production. As the result, yeast (%), inoculum's size (%) and vegetable oil (%) were identified as significant factors. After screening, these factors were subsequently optimized using the response surface methodology (RSM) - Central Composite Designs (CCD). These optimal levels were found out yeast extract (3.0%), inoculum's size (2.37%; 10^ cells/ml) and vegetable oil (4.19%) in which the lipase yield was the highest. The regression equations (model) obtained and predicted maximum lipase yield 18.8078 (U/ml). We also verified model in shaking flasks (100 ml) as well as automatic fermentor (3 litres). Lipase production for 100 ml and 3 litres recorded from supernatant was 18.95 (U/ml) and 22.52 (U/ml), respectively.

The Phen-den plant (Phyllanthus reticulatus Poir.) has been widely used in Vietnamese tradional medicine for detoxification, antiseptic, diuretic, anti-inflammation treatments…However, there is still lack of studies on this plant’s antioxidant and in vitro hepatoprotective activities in Vietnam. Recently, the use of liver enzymes such as CYP450 reductase (CPR) in determining the in vitro hepatoprotective activities of potential compounds, which are originated from herbal plants, is increasing based on its high efficiency and short duration. Furthermore, the other in vitro assays such as DPPH (2,2-diphenyl-1-picryl-hydrazylhydrate) scavenging, inhibition of lipid peroxidation (MDA) are popularly employed for determination of antioxidant activities. With the success in isolation of CPR from Saccharomyces cerevisiae WR1A2 strain (5 litres/batch), we set up the assay for screening potential samples which could be able to induce CPR for in vitro hepatoprotective action. The obtained results from this CPR screening assay showed the similarity to those of other popular antioxidant assays such as DPPH or MDA. The resulted from our assay showed that the water extract of Phen-den plant had the highest ability to induce CPR supporting the hepatoprotective activities. Also, the water extract, EtOAc extract and total extract exhibited very strong antioxidant activities through DPPH scavenging assay and inhibition of lipid peroxidation with the SC50 values ranging at 13.84 µg/ml, 37.64 µg/ml, 28.31 µg/ml and the IC50 values ranging at 5.99 µg/ml, 4.77 µg/ml, 27.58 µg/ml, respectively.

Kappaphycus striatus is growing in some central coastal provinces as a source of carrageenan extract. It is mainly propagated in the form of vegetative reproduction and sporulation. However, this method still has some limitations. At present, there is no report on the micropropagation of this species through callus induction. In this study, effect of plant growth regulators (PGRs) (Naphthalene acetic acid [NAA] and 6-bezyl amino purine [BAP]), intensity of light and concentration of agar on callus induction derived from branches of Kappaphycus striatus maintaned in laboratory for 1 month were tested. After 2 month culture, the results showed that branches of Kappaphycus striatus cultured on PES (Provasoli enriched seawater) medium (without PGRs) gave the best callus induction rate (75.7%) and survival rate (77.3%) compared to those on PES medium supplemented PGRs. The optimal conditions for callus induction were PES solidified medium supplemented with 1.5 – 2.0% agar in 5 µmol.m-2.s-1 of light intensities. Callus induction rates (66.7% – 67%), survival rate (63.7% – 64.3%) of explants from branches incubating on PES medium with 1.5 – 2.0% agar were higher than differrent agar concentrations. Callus induction rates (67%), survival rate (77.7%) of explants from branches incubating on PES medium under 5 µmol.m-2.s-1 were higher than differrent light intensities. The results showed that there were three different types of calluses observed namely white filamentous callus, brown filamentous callus and compact callus. These calli that were big and had filamentous type, will be a good material for the next production stage of embryonic callus production and seedling regeneration from micropropagules.

Paramignya trimera is a valuable traditional medicinal plant, typically distributing in Khanh Hoa, Vietnam. In recent years, P. trimera has been exploited to treat liver diseases such as hepatitis, cirrhosis and liver cancers. The recorded during the investigation, in Khanh Hoa currently appears a number of species with similar phenotypes known collectively as P. trimera and appeared some morphology with differences in leaf morphology and thorns. Therefore, this study was carried out for identification, breeding and propagation conservation.This study has been described, characterized morphological characteristics and analyzed DNA barcodes based on ITS, matK and rbcL gene markers. About the morphology, P. trimera is distinguished from other species of the genus Paramignya with the following characteristics: elongated, single leaf, leaf-shaped lobe end, leaf veins shaped like a feather. Petiole with big, pointed and straight spines. Corolla 3, 6 stamens, upper election, with three sepals and two ovule leaves, the seed has a  seed coat. The analysis of DNA barcodes shows that the results combination of the analysis of the three regions of ITS, matK and rbcL can be used to determine the genetic relationship between five samples of P. trimera. The nucleotide sequence lengths of the samples for the three regions of the ITS, matK and rbcL genes are 850 bp, 850 bp and 500 bp, respectively. On the basis of phenotypic tree analysis, the samples (X1, X2, X3, X4, X5) have the same evolutionary origin in ITS, matK and rbcL. Base on to identified the five morphological forms of P. trimera distributed in Khanh Hoa as the same species and expanded the distribution area as well as in-depth research on P. trimera morphologies in Khanh Hoa, Vietnam.

Lipase A tái tổ hợp (rLipA) từ Bacillus subtilis FS2 được xác định là có đồng thời cả hai hoạt tính lipase và gelatinase, trong đó gelatinase là hoạt tính phụ của enzyme này. Các đột biến trong trung tâm hoạt động của lipase là S77C, D133N và H156N đã được tạo ra bằng phương pháp Mega-primer. Cả ba dòng đột biến đều đã được tách dòng và biểu hiện thành công trong tế bào E. coli BL21 (DE3). Tuy nhiên chỉ có hai dòng đột biến S77C và H156N được tinh sạch còn đột biến D133N bị biểu hiện dưới dạng không tan. Các thông số động học Km và Vmax đối với hai hoạt tính lipase và gelatinase của dòng tái tổ hợp đã được xác định và so sánh với các dòng đột biến. Hai đột biến S77C và H156N đã làm mất hoàn toàn hoạt tính lipase nhưng đột biến S77C lại làm tăng hoạt tính phụ gelatinase lên khoảng 20 lần. Các đột biến tại trung tâm hoạt động của lipase đã làm thay đổi cấu hình không gian và mạng lưới liên kết hydro dẫn đến thay đổi khả năng liên kết giữa enzyme với cơ chất, do vậy lipase có thể dễ dàng kết hợp với cơ chất của gelatinase và thúc đẩy phản ứng phân cắt liên kết gelatin. Hoạt tính gelatinase của rLipA có thể được quyết định bởi một số các gốc amino acid khác trong trung tâm hoạt động của enzyme này.

Recently, LED lighting technology has been developed fast and strongly. It has been widely used in many new fields. LEDs have been tested as an artificial light source during micropagation and tissue culture of various plants species as an alternative to traditional light sources to save energy and improve the efficiency of the culture process. In this study, effects of LED light at different wavelengths and luminous intensities on growth of Anoectochilus roxburghii's buds in vitro were investigated. Anoectochilus's buds were cultured under the different light conditions, such as LED in monochromatic red (R), LED in monochromatic green (B) , LED in the green and red light combining with warm white light (W) in different ratios (BRW 1, BRW2, BRW3 and BR). After 3 months of culture, the results showed LED B at strong luminous intensity (79 ± 3 μmol.m-2.s-1) inhibited the growth and development of Anoectochilus roxburghii. In contrast, BR LED with low luminous intensity (30 ± 1μmol.m-2.s-1) had made a positive effect on growth and development of Anoectochilus roxburghii. The plant height (5,88 cm), root length (1,33 cm), fresh plant weight (0,169 g/plant), leaf area (0,82 cm2), fresh leaf weight (18,33 mg/leaves) of plants in vitro were found higher than those of plants grown under T5 fluorescent lighting conditions (as control). Besides, chlorophyll a, chlorophyll b and total chlorophyll content of leaves (285,40 µg/g, 196,40 µg/g, 481,80 µg/g respectively) were higher than in the control and other versions of LED combine. The results also showed that the LED light with combined ratio of BR = 1: 4 at luminous intensity of 30 μmol.m-2.s-1 were suitable for the growth of Anoectochilus roxburghii. Therefore, BR LED (1:4) light condition should be replaced fluorescent light sources in vitro culture of Anoectochilus roxburghii in future.

Various members of NAC transcription factor family have been shown to play important roles in regulating plant responses to abiotic stresses, such as drought, cold and salinity. Our previous research on differential expression patterns of twenty three soybean NAC genes (GmNACs) by realtime quantitative PCR suggested a correlation between inducible expression of GmNAC085 and the drought tolerance degree in DT51 and MTD720 soybean cultivars, which presented for the drought-tolerant and the drought-sensitive, respectively. Therefore, the gene has been proposed as a potential candidate for engineering in order to produce new varieties with better drought stress tolerance. However, functional studies of GmNAC085 should be carried out to identify how this transcriptional factor can contribute in the plant stress-responsive pathway. Herein, this paper presented that we have successfully developed a recombinant binary vector carrying full-length cDNA of GmNAC085 and expression of this gene is placed under the control of constitutive promoter CaMV 35S. The generated construct was firstly transformed into E.coli for sequencing the target gene and then the identified genuine construct was transformed into Agrobacterium tumefaciens. This can be used for plant transformation mediated by Agrobacterium in serving for GmNAC085 - related studies using in planta system such as the model plant Arabidopsis and also serving for the development of drought-tolerant crops by genetic engineering. Additionally, results from sequence alignment analysis revealed that GmNAC085s of DT51 and MTD720 had identical nucleotide sequence, thus supporting our hypothesis that difference in GmNAC085 gene expression levels, not the variation in GmNAC085 protein sequence or structure, might cause the difference in plant resistance degree to drought stress in these two soybean varieties.

Enzyme AGPase là một trong những enzyme quan trọng, xúc tác cho bước đầu tiên trong quá trình tổng hợp tinh bột và đã được chứng minh là enzyme điều hòa, điều chỉnh tốc độ phản ứng của toàn bộ chu trình sinh tổng hợp glycogen ở vi khuẩn và tinh bột ở thực vật. Ở thực vật bậc cao, AGPase được xác định là enzyme dị lập thể, được cấu tạo bởi hai tiểu phần lớn (AGPL) và hai tiểu phần nhỏ (AGPS) do hai gen tương tứng mã hóa. Trong nghiên cứu này, chúng tôi đã phân lập hai gen mã hóa cho tiểu phần lớn và tiểu phần nhỏ của AGPase từ giống sắn KM140. Hai gen được nối với nhau bằng trình tự P2a và được biểu hiện đồng thời trên một khung đọc mở dưới sự điều khiển của promoter CaMV 35S. Cấu trúc này được chèn vào vector pBI121 và được biến nạp vào cây thuốc lá bằng phương pháp chuyển gen thông qua A. tumefaciens. Cây chuyển gen được kiểm tra bằng phương pháp PCR, Western blot và định lượng hàm lượng tinh bột bằng phương pháp Anthrone. Kết quả đã cho thấy hàm lượng tinh bột tích lũy trong lá cây chuyển gen cao hơn các cây đối chứng từ 13-116% trong cùng điều kiện nuôi dưỡng. Nghiên cứu của chúng tôi tạo ra thêm một hướng tiếp cận trong việc tạo cây trồng biến đổi gen tăng cường khả năng tích lũy tinh bột.

Streptococcus agalactiae is one of the microbial pathogens causing the dark-body disease on snakeskin gourami fish (Trichogaster pectoralis) that affects the growth and quality of fish. This research aimed to isolate and select bacteria inhibiting S. agalactiae which are able to use for controling pathogenic bacteria instead of antibiotics. Fourteen bacteria strains were isolated and screened from healthy fishes, sediment and water samples at fish ponds in Dong Thap province. Among these strains, L7 strain showed the highest inhibition ability with the clear zone diameter was 9,3 mm. The results of the 16S rRNA sequence analysis indicated that the L7 strain belonged to Bacillus subtilis. The experiment to evaluate the inhibition capacity and fish disease control of selected B. subtilis in experimental conditions was conducted by challenging fish with S. agalactiae. Fishes in the control treatment was infected with S. agalactiae at 106 CFU/mL had survival rate 41,7%. The experimental treatments NT1, NT2, NT3 which were treated with B. subtilis at concentrations of 105 CFU/mL, 106 CFU/mL, and 107 CFU/mL gave higher survival rates compared with the non-treated control, with the rates of 60%, 76,7%, and 81,7%, respectively. These results revealed that the isolated B. subtilis is potential used in control dark-body disease on snakeskin gourami fish.

Salmonella resistance to antimicrobials is a major health problem in the world. Thus, we conducted a cross-sectional study to determine the prevalence of Salmonella serovars isolates from retail meats in Ha Noi and their susceptibility to 8 antimicrobials commonly used in the treatment and growth promotion in domestic livestocks in Vietnam. Salmonella was detected in 25/90 (27.8%) samples. Nine different serovars were identified, including S. Typhimurium (44%), S. Derby (16%), S. Warragul, S. Indiana, S. Rissen (8%), and S. London, Meleagridis, Give, Assine (4%). S. Typhimurium (44 %) is the most prevalent types. Resistance to at least one antibiotic was showed in 13 strains (52%). All isolates were 44% (11/25) resistant to streptomycin and tetracycline, 32% (8/25) resistant to chloramphenicol. The multiple antimicrobial resistance accounted for 69.2% of isolates (9/13). All strains were susceptible to ceftazidime. This data indicated that the retail meats could constitute a source of human exposure to multidrug-resistant Salmonella and therefore could be considered a potential vehicle of resistant Salmonella foodborne diseases. There is an urgent need for surveillance and control programmes on Salmonella and use of antimicrobials in Vietnam to protect the health of consumers.