In Vitro and In Vivo Evaluation of Antitumor Activity of Ligustrum robustum, A Chinese Herbal Tea / 中国结合医学杂志

OBJECTIVE@#To examine the effect of the aqueous extract of Ligustrum robustum on tumor growth in vitro and in vivo and explore the possible molecular mechanisms.@*METHODS@#In in vitro study, cell viabilities of human cervical carcinoma cells (HeLa), human breast cancer cells (MCF-7), human prostate cancer cells (PC-3), human hepatoma cells (7721) and human colon carcinoma cells (SW480) were evaluated with cell counting kit-8. For L. robustum-treated Hela cells, early or late apoptosis were evaluated by annexin V/PI staining. Mitochondrial membrane potential was measured by staining cells with JC-1. Apoptosis was monitored by nuclear morphology based on chromatin condensation and fragmentation by 4',6-diamidino-2-phenylinole (DAPI) staining. Caspase-3 and -8 activity levels were measured by a colorimetric assay. In vivo, to evaluate the possible mechanism of L. robustum-mediated antitumor effect, nude mouse xenograft study was also conducted.@*RESULTS@#In in vitro study, L. robustum was found to be toxic to HeLa, MCF-7, PC-3, 7721, SW480, with an half maximal inhibitory concentration value of 2-5 mg/mL (P<0.05). Moreover, externalization of phosphatidylserine, loss of mitochondrial membrane potential, DNA fragmentation and activation of caspase-3 and -8 were detected in L. robustum-treated Hela cells. Using a nude mouse model bearing Hela xenografts, we found that L. robustum reduced tumor volume and tumor weight (P<0.05), but had no effect on body weight and histological damage of important organs. Intraperitoneal injection of L. robustum caused a significant reduction in serum aspartate transaminase and alanine transaminase levels (P<0.05). Furthermore, cleaved caspase-3-positive and terminal nucleotidyl transferase-mediated nick end labeling (TUNEL)-positive cells were observed in L. robustum-treated tumor tissues.@*CONCLUSIONS@#L. robustum inhibits tumor cell growth both in vitro and in vivo by inducing apoptosis in a caspase-dependent way without apparent hepatic toxicity and histological damage, which may offer partial scientific support for the ethnopharmacological claims of L. robustum as a herbal tea for its antitumor activity.

Non-modified magnetic beads coupled with multiple real-time PCR for detection and quantification of mycotoxigenic fungi in paprika samples / 南方医科大学学报

<p><b>OBJECTIVE</b>To establish a method for detecting 3 common toxigenic molds (Aspergillus, Penicillium, and Fusarium) based on non-modified magnetic beads coupled with multiple real-time PCR (NMB-multiple qPCR).</p><p><b>METHODS</b>The primers and genus-specific probes were designed based on the rDNA sequences to develop a multiple real-time PCR using non-modified magnetic bead to enrichment of fungal spores. The sensitivity, specificity and repeatability of this assay were evaluated.</p><p><b>RESULTS</b>The detection limit of this assay for spiked samples was 10(4) CFU/g, demonstrating a 10-fold greater detection sensitivity of this assay than that of real-time PCR. The NMB-multiple qPCR assay also showed good specificity and reproducibility and yielded comparable results with those by traditional colony counting method for spiked samples (P>0.05).</p><p><b>CONCLUSION</b>NMB-multiple qPCR assay we established allows rapid and sensitive detection of common mycotoxigenic fungi in paprika.</p>

Construction and secretory expression of beta-galactosidase gene from Lactobacillus bulgaricus in Lactococcus lactis / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>This study is to examine the secretion effects of beta-galactosidase in Lactococcus lactis.</p><p><b>METHODS</b>The usp45 and beta-galactosidase genes were cloned and inserted into plasmid pMG36e to obtain the recombinant plasmid pMG36e-usp-lacZ. This recombinant plasmid was transformed into both Escherichia coli DH5alpha and L. lactis MG1363. The enzyme activity, gene sequencing, SDS-PAGE and hereditary stability were assessed and studied.</p><p><b>RESULTS</b>The lacZ gene inserted into plasmids pMG36e-usp-lacZ was 99.37% similar to the GenBank sequence, and SDS-PAGE revealed an evident idio-strap at 116 KDa between L. lactis MG1363/pMG36e-usp-lacZ in both supernatant and cell samples. Beta-Galactosidase activity measured 0.225 U/mL in L. lactis pMG36e-usp-lacZ transformants, and its secretion rate was 10%. The plasmid pMG36e-usp-lacZ appeared more stable in MG1363.</p><p><b>CONCLUSION</b>The authors concluded that these new recombinant bacteria well expressed and secreted beta-galactosidase, indicating that the beta-galactosidase expression system was successfully constructed, and this might provide a new solution for management of lactose intolerance specifically and promote the use of gene-modified organisms as part of the food-grade plasmid in general.</p>

Preliminary study on bacteroides as the potential fecal contamination indicator bacteria / 中华预防医学杂志

<p><b>OBJECTIVE</b>To explore the possibility of Bacteroides spp. as fecal contamination indicator bacteria with real-time quantitative PCR (RT-PCR) assay through analyzing the correlation between Bacteroides spp. and coliform group in external environment.</p><p><b>METHODS</b>Quantity of coliform group and Bacteroides in water samples were detected by most-probable-number method (MPN) and RT-PCR, respectively, and their detection correlation was evaluated with linear correlation analysis. Both methods were also applied to detect the contaminated time limits and river water samples collected at four sampling sites in three different times.</p><p><b>RESULTS</b>Seventy two hours were needed for the numeration of coliform group with MPN method, while RT-PCR could detect Bacteroides within 3 hours. The contaminated time limit of indoor and outdoor water samples of coliform group was more than 40 days and 9 days, and Bacteroides 13 days and 5 days, respectively. Also, the positive correlation between the quantity of Bacteroides and coliform group in outdoor water samples was obtained, the quantity of Bacteroides was from 8.3 × 10(6) copies/ml to less than 10(4) copies/ml during the first day to the fifth day, while coliform group was 4.3 × 10(6) MPN/100 ml to 2.4 × 10(3) MPN/100 ml. A 100% coincidence rate of the detection results with both methods was also observed. These results indicated that the detection results of both methods had perfect consistency.</p><p><b>CONCLUSION</b>Bacteroides spp. can be potentially used as fecal contamination indicator bacteria with RT-PCR rapid detection.</p>

Preparation and evaluation of novel solid chlorine dioxide-based disinfectant powder in single-pack / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To prepare and evaluate novel chlorine dioxide-based disinfectant powder in single-pack that is more convenient for use and transportation.</p><p><b>METHODS</b>Orthogonal experiment was performed to determine the recipe of the disinfectant powder. Stability test, suspension quantitative bactericidal test, simulation field trial, and animal toxicity test were carried out to observe its bactericidal and toxicological effects.</p><p><b>RESULTS</b>The orthogonal experiment showed that the type of water solution had no effect on the disinfectant powder and the best ratio of sodium chlorite to solid acid was 1:3. Ten grams of the disinfectant powder was fully dissolved in 20 mL water for 2 min, and diluted to 500 mL in water. After 5-10 min, the concentration of chlorine dioxide (ClO2) solution was 266 mg/L to 276 mg/L. After stored at 54 degrees C for 14 d, the average concentration of ClO2 was decreased by 5.03%. Suspension quantitative bactericidal test showed that the average killing logarithm (KL) value for both Staphylococcus aureus and Escherichia coli in 100 mg/L ClO2 solution for 2 min was over 5.00. in simulation field trial, the average descending KL value for Escherichia coli in the solution containing 100 mg/L ClO2 for 5 min was over 3.00. The mouse acute LD50 in the solution 5 times exceeded 5000 mg/kg. The disinfectant powder was not toxic and irritative to rabbit skin and had no mutagenic effect on mouse marrow polychromatic erythrocytes (PCE).</p><p><b>CONCLUSION</b>The stability and bactericidal efficacy of solid chlorine dioxide-based disinfectant powder in single-pack are good. The solution containing 100 mg/L ClO2 can kill vegetative forms of bacteria. The concentration of ClO2 on the disinfecting surface of objects is 100 mg/L. The disinfectant powder is not toxic and irritative.</p>

Non-fusion and fusion expression of beta-galactosidase from Lactobacillus bulgaricus in Lactococcus lactis / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To construct four recombinant Lactococcus lactis strains exhibiting high beta-galactosidase activity in fusion or non-fusion ways, and to study the influence factors for their protein expression and secretion.</p><p><b>METHODS</b>The gene fragments encoding beta-galactosidase from two strains of Lactobacillus bulgaricus, wch9901 isolated from yogurt and 1.1480 purchased from the Chinese Academy of Sciences, were amplified and inserted into lactococcal expression vector pMG36e. For fusion expression, the open reading frame of the beta-galactosidase gene was amplified, while for non-fusion expression, the open reading frame of the beta-galactosidase gene was amplified with its native Shine-Dalgarno sequence upstream. The start codon of the beta-galactosidase gene partially overlapped with the stop codon of vector origin open reading frame. Then, the recombinant plasmids were transformed into Escherichia coli DH5 alpha and Lactococcus lactis subsp. lactis MG1363 and confirmed by determining beta-galactosidase activities.</p><p><b>RESULTS</b>The non-fusion expression plasmids showed a significantly higher beta-galactosidase activity in transformed strains than the fusion expression plasmids. The highest enzyme activity was observed in Lactococcus lactis transformed with the non-fusion expression plasmids which were inserted into the beta-galactosidase gene from Lactobacillus bulgaricus wch9901. The beta-galactosidase activity was 2.75 times as high as that of the native counterpart. In addition, beta-galactosidase expressed by recombinant plasmids in Lactococcus lactis could be secreted into the culture medium. The highest secretion rate (27.1%) was observed when the culture medium contained 20 g/L of lactose.</p><p><b>CONCLUSION</b>Different properties of the native bacteria may have some effects on the protein expression of recombinant plasmids. Non-fusion expression shows a higher enzyme activity in host bacteria. There may be a host-related weak secretion signal peptide gene within the structure gene of Lb. bulgaricus beta-galactosidase, and its translation product may introduce the enzyme secretion out of cells in special hosts.</p>

A primary study on the relationship between amino acid mutations in clinical isolates of Neisseria gonorrhoeae and their resistance to antibiotics / 中华流行病学杂志

<p><b>OBJECTIVE</b>To identify the relationship between amino acid mutations in Neisseria gonorrhoeae isolates and their antibiotic resistance.</p><p><b>METHODS</b>PI gene fragments of Neisseria gonorrhoeae from 17 clinical isolates were obtained with PCR amplification. They were cloned into the PCR cloning vector pBS-T to form pBS-T-PI and sequenced. The sequences of PI genes were analyzed. At the same time, minimum inhibitory concentration (MIC) of penicillin and tetracycline to these 17 isolates were measured and contrasted with the corresponding PI sequence.</p><p><b>RESULTS</b>The recombinants of PI gene from 17 clinical isolates of Neisseria gonorrhoeae were successfully constructed and sequenced. They were divided into PIA and PIB subtypes according to the results from blastn software by comparing the sequences with the GenBank. Mutations were found at the sites of 120 and 121. There were only some of the sequences having an aspartic acid (D) mutation on 120 and 121 sites, which was not the same as reported. On the other hand,there were two PI sequences,5-9 and 6-1, whose mutations on No. 120 were lysine, similar to those documented.</p><p><b>CONCLUSION</b>Some relationship between PI amino acids mutations at sites 120 and 121 in Neisseria gonorrhoeae isolates from Chengdu, China and their resistance to penicillin and tetracycline were found. However,further studies need to be done in the future to confirm this hypothesis.</p>