大蒜鳞茎提取液对黄瓜2种种传病害的抑制及化感作用研究

【目的】明确大蒜鳞茎提取液在抑制种传病害过程中对寄主植物黄瓜的化感效应,为开发利用大蒜鳞茎提取液作为新生物源农药提供理论依据。【方法】以吉木萨尔白皮蒜为研究材料,按照ISTA种传病害研究方法,以不同质量分数(2.5%,1.25%,0.625%和0.312 5%)大蒜鳞茎水提液,分别活体接种2种种传病害(黄瓜镰刀菌枯萎病和猝倒病)病原菌孢子,用菌落数计算防治效果;测定种子萌发和幼苗生长时期的发芽指数、活力指数、发芽率、根鲜质量、根干质量、根系活力、α-淀粉酶活力、出苗率、叶绿素含量、硝酸还原酶活性等指标,以化感效应指数为依据,确定不同质量分数大蒜鳞茎提取液对寄主黄瓜幼苗的化感作用。【结果】大蒜鳞茎提取液对黄瓜2种种传病害均表现出较强的抑制作用,以质量分数0.625%的大蒜鳞茎提取液处理黄瓜种子4 h,黄瓜枯萎病和猝倒病均未发生,防治效果达到100%。与对照相比,质量分数1.25%大蒜鳞茎提取液处理黄瓜种子的活力指数提高3.74%,根系活力提高7.44%,α-淀粉酶活力增加50.35%,幼苗的硝酸还原酶活性增加66.67%;质量分数2.5%大蒜鳞茎提取液处理黄瓜种子的活力指数提高0.14%,根系活力提高19.72%,α-淀粉酶活力增加6.34%,幼苗的硝酸还原酶活性增加130.31%。【结论】一定质量分数的大蒜鳞茎水提液在抑制黄瓜2种种传病害的同时,还存在着对寄主植物的化感作用,对种子发芽和幼苗生长总体表现为促进作用。

重寄生菌对花棒锈病的抑制作用

花棒（Hedysarum scoparium Fisch. et Mey）是优良的固沙先锋树种，花棒锈病（Uromyces onobrychidis （Desm．）Lév．）是花棒的常见病害之一，在一些地区严重制约了花棒的先锋固沙作用。本文对花棒锈病菌上的重寄生菌菌种来源、温度和碳源对重寄生菌菌丝生长及产孢的影响、室内接种试验、花棒锈病菌的重寄生现象及对花棒锈病的自然抑制力进行了研究。研究发现，花棒锈病菌的重寄生菌寄生于花棒锈病菌的夏孢子堆和冬孢子堆上，被重寄生菌寄生后，夏孢子堆南橙黄色变为灰白色，冬孢子堆南棕褐色变为灰色，后期夏孢子失去萌发力，孢子堆原基处只存有重寄生菌的分生孢子器。重寄生菌在20～25℃ PDA培养基上产孢量大。重寄生菌为Sphaerellopsis sp．，是一种自然重寄生现象，并对花棒锈病有很强的抑制作用，如果充分利用这一生物链，人工干预这一现象的早发和多发，可以有效提高花棒固沙能力的发挥，其潜质意义很大，是以菌治病、以菌哺林的新资源。

ZSB生物种衣剂对水稻细菌性条斑病抑制作用的研究

室内抑菌试验表明，ＺＳＢ生物种衣剂对水稻细菌性条斑病（以下简称细条病）菌有较强的拮抗作用。经种衣剂包衣后。

Antifungal Activity of Extracts from Clerodendrum Bungei Leaves against Two Species of Phytopathogens

The test was undertaken to reveal the antifungal activity of extracts from Clerodendrum bungei leaves against Pestalotia and Rhizoctonia solani, the results showed that optimal condition for best antifungal activity of extracts against pestalotia and Rhizoctonia solani are as follows： material-liquid ratio of 1：6,75% ethanol as extracting solvent, reflux at 90℃ for 1.5 h. The substances with good dissolubility in ethanol and water solution such as organic acid, bioflavonoid and alkaloid are main antifungal bioactive substances in Clerodendrun bungei.

Inhibitory Effects of Original Nano-Cu_2O Drug and Nano-Cu_2O Suspension on Snake Melon Botrytis cinerea

The drug-containing culture medium method for the test of toxicity was adopted to compare inhibitive effects of original nano-Cu2O drug and nano-Cu2O suspension, and nano-Cu2O drug has better inhibitive effects on snake melon Botry- tis cinerea than original nano-Cu2O drug with the same mass concentration, and inhibitory effects are positively correlated with concentration. Correlation coefficients of the toxicity regression equation are 0.892 2 and 0.996 1, effective concentration EC50 of original nano-Cu2O drug and that of nano-Cu2O suspension are 3 948.9 and 167.9 mg/kg. Original nano-Cu2O drug has an inhibitive effect on snake melon Botrytis cinerea, but the inhibition of nano-Cu2O suspension is more obvious.

Antifungal Effect of Streptomyces 702 Antifungal Monomer Component DZP8 on Rhizoctonia solani and Magnaporthe grisea

The aim of this study was to investigate the in vitro antifungal effects of antifungal monomer component DZP8 isolated from Streptomyces 702 on the mycelium growth, sclerotium formation and germination of Rhizoctonia solani and on the mycelium growth, conidial formation, germination, appressorium formation of Magnaporthe grisea. The results showed that the antifungal monomer component DZP8 has strong antifungal effect on both the R. solani and M. grisea. The EC50 and EC90 of DZP8 were 1.81 and 3.35 μg/ml on Ft. solani respectively, and 37.01 and 136.21 μg/ml on M. grisea respectively. Under the treatment of 48.01 μg/ml DZP8, the sclerotium formation rate of R. solani was just 39.21%, the formation time delayed by 216 h and the dry weight decreased by 81.37% in comparison the con- trol; and 33.51 μg/ml DZP8 significantly inhibited the sclerotium germination. In the presence of 160.08 μg/ml DZP8, the sporulation of M. grisea was just 9.29% of control sample; 20.14 μg/ml DZP8 inhibited the conidial germination suppression rate by 95.16%, and the appressorium formation by 100%.

Inhibitory Effects of Some Plant Extracts on 5 Bacterial Strains

[Objective] This study was conducted to develop potential natural plant products for controlling walnut blight pathogen and other bacteria. [Method] Inhibitory effects of extracts obtained from 15 plants with 3 solvents on bacteria were investi- gated by disk diffusion method. [Results] Except the extracts from Magnolia grandi- flora and Typha orientalis, extracts of 13 plant leaves presented inhibitory effects on 5 bacteria strains to certain degrees. Among them, the effect of water extract of Aesculu schinensis on Bacillus sp. XHE8 was the strongest, with inhibition zone di- ameter reaching （31.3＋3.9） mm and the ratio to control above 5.0. Four of the 5 tested strains were sensitive to the extracts of Sambucus chinensis, and 3 of them were inhibited by Ophiopegon japonicas extracts and Reineckia camea extracts, with ratios of treatment to control large than 1.5 in all. Leaf extract of A. chinensis had significant anti-bacteria ability, and could be used as a potential plant source for bactericide. [Conclusion] The results laid a foundation for exploring active com- pounds and elucidating the mechanism in it.

Effective regularity in modulation on gastric motility induced by different acupoint stimulation

AIM： To investigate whether manual acupuncture at representative acupoints in different parts of the body can modulate responses of gastric motility in rats and regular effects in different acupoint stimulation. METHODS： The gastric motor activity of rats was recorded by the intrapyloric balloon. The changes of gastric motility induced by the stimulation were compared with the background activity in intragastric pressure and/or waves of gastric contraction recorded before any stimulation. Morphological study was also conducted by observing the Evans dye extravasation in the skin after mustard oil injection into the intragastric mucous membrane to certify cutaneous innervations of blue dots related to gastric segmental innervations. RESULTS： In all six rats that received mustard oil injections into intragastric mucosa, small blue dots appeared in the skin over the whole abdomen, but mainly in peri-midline upper- and middle- abdomen and middle-back, a few in thigh and groin. It may speculate that cutaneous innervations of blue dots have the same distribution as gastric segmental innervations. Acustimulation in acupoints of head-neck, four limbs, upper chest-dorsum and Iower-dorsum induced markedly augmentation of gastric motility （acupoints on headneck such as St-2： n = 16, 105.19 ± 1.36 vs 112.25 ± 2.02 and St-3： n = 14, 101.5 ± 1.75 vs 109.36 ± 1.8; acupoints on limbs such as Sp-6： n = 19, 100.74 ± 1.54 vs 110.26 ± 3.88; St-32： n = 17, 103.59 ± 1.64 vs 108.24 ± 2.41; St-36： n = 16, 104.81 ± 1.72 vs 110.81 ± 2.74 and U-11： n = 17, 106.47 ± 2.61 vs 114.77 ± 3.77, P 〈 0.05-0.001）. Vigorous inhibitory regulations of gastric motility induced by acu-stimulation applied in acupoints on whole abdomen and middle-dorsum were significantly different as compared with the controls before acu-stimulation （abdomen acupoints such as Cv-12： n = 11, 109.36 ± 2.09 vs 101 ± 2.21; Cv-6： n = 18, 104.39 ± 1.42 vs 91.83 ± 3.22 and St-21： n = 12, 107 ± 2.97 vs 98.58 ± 2.81; acupoints on middledorsum such as BI-17： n = 19, 100.63 ± 1.4 vs 92.21 ± 2.07 and BI-21： n = 19, 103.84 ± 1.48 vs 97.58 ± 2.16, P 〈 0.05-0.001）. CONCLUSION： Regular regulatory effects of facilitation and inhibition on gastric motility appear to be somatotopically organized in the acupoints of whole body, and the effective regularity of site-special acupoints on gastric motility is involved in segmental innervations between stomach and acupoints.

Inhibitory effect of emodin and Astragalus polysaccharide on the replication of HBV

AIM: To evaluate the anti-viral effect of emodin plus Astragalus polysaccharide (APS) in hepatitis B virus (HBV) transgenic mice.METHODS: Sixty HBV transgenic mice (HBV TGM) whose weight varied between 18 and 24 g were randomly divided into 3 groups, with 20 mice in each group. Group A was the normal control, where the mice were treated with physiological saline; group B was the positive control where the mice were treated with lamivudine solution (100 mL/kg per day). Group C was the experimental group where the mice were treated with physiological saline containing emodin and APS (57.59 mg/kg per day and 287.95 mg/kg per day, respectively). The mice were treated daily for 3 wk. After 1 wk recovery time, the mice were sacrifi ced and serum as well as liver tissues were collected for ELISA and histological examination.RESULTS: After 21 d treatment, HBV DNA levels in group B and group C significantly declined when compared with group A (P < 0.05). However, a signif icant increase in HBV DNA content was observed in group B, whereas this phenomenon was not observed in group C. A reduction in the contents of HBsAg, HBeAg and HBcAg in the mice from group B and C was observed when compared with group A.CONCLUSION: Emodin and APS have a weak but persistent inhibitory effect on HBV replication in vivo, which may function as a supplementary modality in the treatment of hepatitis B infection.

Inhibition of genes expression of SARS coronavirus by synthetic small interfering RNAs

RNA interference (RNAi) is triggered by the presence of a double-stranded RNA (dsRNA), and results in the silencing of homologous gene expression through the specific degradation of an mRNA containing the same sequence. dsRNAmediated RNAi can be used in a wide variety of eucaryotes to induce the sequence-specific inhibition of gene expression.Synthetic 21-23 nucleotide (nt) small interfering RNA (siRNA) with 2 nt 3' overhangs was recently found to mediate efficient sequence-specific mRNA degradation in mammalian cells. Here, we studied the effects of synthetic siRNA duplexes targeted to SARS coronavirus structural proteins E, M, and N in a cell culture system. Among total 26 siRNA duplexes, we obtained 3 siRNA duplexes which could sequence-specifically reduce target genes expression over 80％ at the concentration of 60 nM in Vero E6 cells. The downregulation effect was in correlation with the concentrations of the siRNA duplexes in a range of 0～60 nM. Our results also showed that many inactive siRNA duplexes may be brought to life simply by unpairing the 5' end of the antisense strands. Results suggest that siRNA is capable of inhibiting SARS coronavirus genes expression and thus may be a new therapeutic strategy for treatment of SARS.

Inhibition of hepatitis B virus surface antigen expression by small hairpin RNA in vitro

AIM: To explore the anti-hepatitis B virus effect of RNA interference (RNAi) using small hairpin RNA (shRNA)expression vector.METHODS: Hepatitis B virus surface antigen green fluorescent protein (HBs-GFP) fusion vector and shRNA expression vectors were constructed and cotransfected transiently into HepG2 cells. mRNAs extracted from HepG2 cells were detected by real-time PCR. Fluorescence of HBs-GFP protein was detected by fluorescence-activated cell sorting (FACS). The effective shRNA expression vector was transfected into HepG2.2.15 cells. HBsAg and HBeAg in HepG2.2.15 cells were analyzed by radioimmunoassay (RIA) method.RESULTS: FACS revealed that shRNA targeting at HBsAg reduced the GFP signal by 56% compared to the control.Real-time PCR showed that HBs-GFP mRNA extracted from HepG2 cells cotransfected with pAVU6+27 and HBs-GFP expression plasmids decreased by 90% compared to the empty vector control. The expressions of HBsAg and HBeAg were also inhibited by 43% and 64%, respectively.CONCLUSION: RNAi using shRNA expression vector can inhibit the expression of HBsAg, providing a fresh approach to screening the efficient small interfering RNAs (siRNAs).

Down-regulation of HIV-1 Infection by Inhibition of the MAPK Signaling Pathway

The human immunodeficiency virus type 1 （HIV-1） can interact with and exploit the host cellular machinery to replicate and propagate itself. Numerous studies have shown that the Mitogen-activated protein kinase （MAPK） signal pathway can positively regulate the replication of HIV-1, but exactly how each MAPK pathway affects HIV-1 infection and replication is not understood. In this study, we used the Extracellular signal-regulated kinase （ERK） pathway inhibitor, PD98059, the Jun N-terminal kinase （JNK） pathway inhibitor, SP600125, and the p38 pathway inhibitor, SB203580, to investigate the roles of these pathways in HIV-1 replication. We found that application of PD98059 results in a strong VSV-G pseudotyped HIV-1NL4-3 luciferase reporter virus and HIV-1NL4-3 virus inhibition activity. In addition, SB203580 and SP600125 also elicited marked VSV-G pseudotyped HIV-INL4-3 luciferase reporter virus inhibition activity but no HIV-1NL4-3 virus inhibition activity. We also found that SB203580 and SP600125 can enhance the HIV-1 inhibition activity of PD98059 when cells were treated with all three MAPK pathway inhibitors in combination. Finally, we show that HIV-1 virus inhibition activity of the MAPK pathway inhibitors was the result of the negative regulation of HIV-1 LTR promoter activity.

Neutralizing antibodies in hepatitis C virus infection

Hepatitis C virus (HCV) is a major cause of hepatitis world-wide. The majority of infected individuals develop chronic hepatitis which can then progress to liver cirrhosis and hepatocellular carcinoma. Spontaneous viral clearance occurs in about 20%-30% of acutely infected individuals and results in resolution of infection without sequaelae. Both viral and host factors appear to play an important role for resolution of acute infection. A large body of evidence suggests that a strong, multispecific and long-lasting cellular immune response appears to be important for control of viral infection in acute hepatitis C. Due too the lack of convenient neutralization assays, the impact of neutralizing responses for control of viral infection had been less defined. In recent years, the development of robust tissue culture model systems for HCV entry and infection has finally allowed study of antibody-mediated neutralization and to gain further insights into viral targets of host neutralizing responses. In addition, detailed analysis of antibody-mediated neutralization in individual patients as well as cohorts with well defined viral isolates has enabled the study of neutralizing responses in the course of HCV infection and characterization of the impact of neutralizing antibodiesfor control of viral infection. This review will summarize recent progress in the understanding of the molecular mechanisms of antibody-mediated neutralization and its impact for HCV pathogenesis.

Combination of small interfering RNA and lamivudine on inhibition of human B virus replication in HepG2.2.15 cells

AIM: To observe the inhibition of hepatitis B virus (HBV) replication and expression by combination of siRNA and lamivudine in HepG2.2.15 cells. METHODS: Recombinant plasmid psil-HBV was constructed and transfected into HepG2.2.15 cells. The transfected cells were cultured in lamivudine-containing medium (0.05 μmol/L) and harvested at 48, 72 and 96 h. The concentration of HBeAg and HBsAg was determined using ELISA. HBV DNA replication was examined by real- time PCR and the level of HBV mRNA was measured by RT-PCR. RESULTS: In HepG2.2.15 cells treated with combination of siRNA and lamivudine, the secretion of HBeAg and HBsAg into the supernatant was found to be inhibited by 91.80% and 82.40% (2.89 ± 0.48 vs 11.73 ± 0.38, P < 0.05; 4.59 ± 0.57 vs 16.25 ± 0.48, P < 0.05) at 96 h, respectively; the number of HBV DNA copies within culture medium was also significantly decreased at 96 h (1.04 ± 0.26 vs 8.35 ± 0.33, P < 0.05). Moreover, mRNA concentration in HepG2.2.15 cells treated with combination of siRNA and lamivudine was obviously lower compared to those treated either with siRNA or lamivudine (19.44 ± 0.17 vs 33.27 ± 0.21 or 79.9 ± 0.13, P < 0.05). CONCLUSION: Combination of siRNA and lamivudine is more effective in inhibiting HBV replication as compared to the single use of siRNA or lamivudine in HepG2.2.15 cells.

Modulation of MMP-2 and TIMP-2 by low dose radiation in mice bearing S180 sarcoma

Objective To investigate the inhibition of low dose radiation （LDR） on S180 sarcomas and its modulation of MMP-2 and TIMP-2 in mice. Methods $180 subcutaneously implanted tumor model mice were randomly divided into two groups： control （N） and low dose radiation （LDR） groups. N mice were sacrificed after 12 h, whereas LDR mice were sacrificed after 12 （LDR-12 h）, 24 （LDR-24 h）, 48 （LDR-48 h）, and 72 （LDR-72 h） h. Thereafter, we measured the tumor volumes. Histopathology was performed, and P-V immunohistochemistry was applied to assess MMP-2 and TIMP-2 expression. Results Compared with the control group, the tumor growth was significantly inhibited in the LDR groups （P 〈 0.05）. MMP-2 expression was considerably reduced in LDR-24h （P 〈 0.05） and LDR-48h （P 〈 0.05）, whereas the change of TIMP-2 was not obvious in the LDR groups （P 〉 0.05） in contrast to that of the control group. Conclusion LDR can effectively suppress the growth of S180 implanted tumors by reducing MMP-2, which is associated with invasion and metastasis.

In vitro porcine brain tubulin assembly inhibition by water extract from a Chinese medicinal herb,Tripterygium hypoglaucum Hutch

AIM： To investigate the effect of Tripterygium hyp-oglaucum Hutch （THH） on the assembly and disassembly process of tubulin and its possible mode of action.METHODS： In vitro porcine brain tubulin assembly assay was employed to analyze the inhibitory effects of THH at different concentrations （0.05 μg/L, 0.07 μg/L, 0.09μg/L）. Colchicine （0.0025 mmol/L, 0.0050 mmol/L, 0.0075 mmol/L） was used as a positive control.RESULTS： THH could significantly inhibit the assembly of isolated porcine brain tubulin at all tested concentrations.CONCLUSION： THH is capable of inducing aneuploidy in mammals via tubulin polymerization inhibition pathway and may pose a genetic risk to human beings.

INHIBITORY ROLE OF TRANSCRIPTION FACTOR COUP-TFII IN EXPRESSION OF HTERT IN HELA CELLS

To clone and identify the proteins involved in regulating the transcription of hTERT and study the role of genes in both hTERT transcription and telomerase activity. Methods The full cDNA of COUP-TFII was cloned from HeLa cDNA library by hTERT promoter-based yeast one-hybrid assay and then in-frame inserted into His-tag fusion expression vector pEK318. The His-tag COUP-TFII fusion proteins were purified by Ni-NTA chromatography. The interaction of COUP-TFII with hTERT promoter in vitro was identified by electrophoretic mobility shift assay and Footprint. The role of COUP-TFII in both hTERT transcription and telomerase activity were probed through Luciferase reporter assay, Northern blot, and TRAP-PCR ELISA. Results COUP-TFII could firmly bind to the downstream E-box and the other two binding sites in hTERT promoter. Luciferase reporter assay indicated COUP-TFII could suppress hTERT promoter activity and stable introduction of COUP-TFII into HeLa cells also decreased both endogenous hTERT transcription and telomerase activity. Conclusion The human COUP-TFII can firmly bind to hTERT promoter, and inhibit telomerase activity through decreasing hTERT transcription. It will greatly facilitate understanding of telomerase regulation in normal and cancer cells

Inhibition of lung cancer stem cells self-renewal and tumorigenicity by lentivirus-delivered Bmi1 shRNA

Objective： The aim of the study was to observe the effect of Bmil reduction on the self-renewal and tumorigenicity ability of lung cancer stem cells （LCSCs） in human lung adenocarcinoma. Methods： Human lung adenocarcinoma cells A549 were consecutively passaged in NOD/SCID mice treated with Paclitaxel weekly. The proportions of LCSCs in A549 cells and the cells from the third passage （A549-3rd） were compared. The expression of Bmil in LCSCs was silenced by intratumoral injection with lentivirus-delivered Broil small hairpin RNA （shRNA）. RT-PCR and Western blot were used to test the mRNA and protein expressions of Broil in LCSCs. The protein level of p16INK4A was analyzed by Western blotting. The self- renewal and tumorigenicity ability of LCSCs were evaluated by counting the sphere formation rate in serum-free medium and the tumor formation rate in NOD/SCID mice. Results： In vivo passaging ofA549 cells under chemotherapy pressure enriched for LCSCs. The expression of Broil in LCSCs increased. Down-regulation of Bmil by RNA interference resulted in reduced self-renewal and tumorigenicity ability of LCSCs and paralleled the increased expression of p16INK4A, a Bmil target. Conclu- sion： Broil regulates self-renewal and tumorigenicity of LCSCs by silencing some target genes, including p16INK4A.

Suppressive Effect of Brassica juncea on the Root-Lesion Nematode （ Pratylenchus penetrans）

Incorporation of sulfur-rich crucifer tissues into soil is known to suppress a variety of soil-bome plant pathogens and pests. The potentials of using Brassica juncea as green manure to kill the root lesion nematode Pratylenchus penetrans m soil and to decrease damages to subsequent crops were assessed in field and pot experiments. In the first trial, green manures containing B. juncea were grown and incorporated during spring to summer 2009. Japanese radish was then cultivated in each plot. In the second trial in spring 2010, green manure was grown and incorporated during summer to autumn 2009, and greater burdock was cultivated in pots containing soil sampled from each plot. Neither trial showed clear effects on nematode populations in the soil. However, in the first trial, Japanese radish grown following a B. juncea breeding line with a high content of sinigrin had a lower root lesion index and a higher number of marketable taproots than grown in the fallow soil. In the second trail, greater burdock cultivated in pots following incorporation ofB. juncea had a lower root lesion index with the incorporation of white mustard, which is widely used as a landscape plant. These findings suggest that B. juncea used as green manure can potentially decrease damage to subsequent crops caused by the root-lesion nematode, although it had no positive effect on decreasing populations of the root-lesion nematode in the soil.

Inhibitory effect of adenoviral vector-mediated AT2R gene transfection on neointimal hyperplasia

Objective: To investigate the effect of adenoviral vector-mediated AT2R gene transfection on neointimal hyperplasia after rat carotid artery balloon injury. Methods :AT2R gene was transferred into rat carotid arteries by recombinant adenovirus pAd-AT2R after the establishment of rat carotid balloon injury restenosis model. The arteries were harvested on the 14th day after gene transfer. The efficiency of trans-gene delivery was measured by the expression of adenovirus-encoding green fluorescent protein (GFP) under fluorescent microscope. The expression of AT2R and PCNA (proliferating cell nuclear antigen) was e-valuated by RT-PCR, immunocytochemistry, immunofluorescence staining, confocal microscopy, respectively. The ratio of intimal to medial area (I/M) was quantified with images and determined by an image analysis system. Results: GFP-positive area in adventitia, media and the forming neointima was about 40%. Adenoviral delivery of rat AT2R gene up-regulated AT2R expression in balloon-injured rat carotid and reduced PCNA expression and I/M significantly in neointima(P<0. 01). Double immunofluorescence labeling of AT2R and PCNA also showed that AT2R gene transfer inhibited VSMCs proliferation in neointima. Conclusion: AT2R gene transfer may be a novel promising therapy to limit neointimal hyperplasia.