<Emphasis Type="Italic">Narcissus tazetta</Emphasis> lectin shows strong inhibitory effects against respiratory syncytial virus,influenza A (H1N1, H3N2, H5N1) and B viruses

A mannose-binding lectin (Narcissus tazetta lectin [NTL]) with potent antiviral activity was isolated and purified from the bulbs of the Chinese daffodil Narcissus tazetta var. chinensis, using ion exchange chromatography on diethylaminoethyl (DEAE)-cellulose, affinity chromatography on mannose-agarose and fast protein liquid chromatography (FPLC)-gel filtration on Superose 12. The purified lectin was shown to have an apparent molecular mass of 26 kDa by gel filtration and 13 kDa by SDS-PAGE, indicating that it is probably a dimer with two identical subunits. The cDNA-derived amino acid sequence of NTL as determined by molecular cloning also reveals that NTL protein contains a mature polypeptide consisting of 105 amino acids and a C-terminal peptide extension. Three-dimensional modelling study demonstrated that the NTL primary polypeptide contains three subdomains, each with a conserved mannose-binding site. It shows a high homology of about 60%–80% similarity with the existing monocot mannose-binding lectins. NTL could significantly inhibit plaque formation by the human respiratory syncytial virus (RSV) with an IC₅₀ of 2.30 μg/ml and exhibit strong antiviral properties against influenza A (H1N1, H3N2, H5N1) and influenza B viruses with IC₅₀ values ranging from 0.20 μg/ml to 1.33 μg/ml in a dose-dependent manner. It is worth noting that the modes of antiviral action of NTL against RSV and influenza A virus are significantly different. NTL is effective in the inhibition of RSV during the whole viral infection cycle, but the antiviral activity of NTL is mainly expressed at the early stage of the viral cycle of influenza A (H1N1) virus. NTL with a high selective index (SI=CC₅₀/IC₅₀≥141) resulting from its potent antiviral activity and low cytotoxicity demonstrates a potential for biotechnological development as an antiviral agent.     

利用pSOS系统筛选靶向HBV病毒X基因的siRNA

构建靶向乙肝病毒(HBV)X基因的真核表达载体pSOS-X-siRNA和pSOS-siRNA,转染肝癌细胞系HepG2和HepG2.2.15,筛选和验证高效siRNA。设计4个靶向X基因siRNA,将siRNA和HBx基因插入载体pSOS得重组质粒pSOS-X-siRNA;pSOS-X-siRNA经PacI酶切去除目的片段HBx后得pSOS-siRNA。将质粒pSOS-X-siRNA和pSOS-siRNA分别转HepG2和HepG2.2.15肝癌细胞株。荧光显微镜下观察HepG2细胞绿色荧光(GFP)减弱程度预估干扰效率。ELISA检测HepG2.2.15细胞上清HBsAg、HBeAg表达,Western blotting检测胞内蛋白HBsAg、HBcAg表达,Real time PCR检测胞内HBsmRNA、HBx mRNA的转录。转染4d后,siRNA4使HepG2细胞的GFP信号表达程度最低,为阴性对照的9%,预测其干扰效果最强。siRNA4使HepG2.2.15细胞转染后4d上清的HBsAg蛋白表达为对照的(13.92±1.14)%(P0.05)、HBeAg为(21.69±4.92)%(P0.05),胞内的HBsAg、HBcAg蛋白表达量灰度比值为0.175±0.025、0.0825±0.028,均为各处理组中最低(P0.01),HBs mRNA和HBx mRNA分别降为对照的0.237±0.028(P0.01)、0.110±0.022(P0.01),差异有统计学意义,证实siRNA4为高效干扰位点。利用pSOS成功筛选并验证构建靶向HBV X基因的siRNA靶点。     

Multidisciplinary approach to diagnosis and management of osteosarcoma – a review of the St Vincent's Hospital experience

Background

Osteosarcoma is the most common primary malignant bone tumour in children and young adults. Despite advances in the diagnosis and management of osteosarcoma, there have been few recent studies describing the experiences of tertiary referral centres. This paper aims to describe and discuss the clinical features, pre-operative work-up, management and outcomes of these patients at St Vincent's Hospital (Melbourne, Australia).

Methods

Retrospective study of fifty-nine consecutive patients managed for osteosarcoma at St Vincent's Hospital between 1995 and 2005.

Results

Median age at diagnosis was 21 (range, 11–84) years. Gender distribution was similar, with thirty-one male and twenty-eight female patients.Twenty-five patients had osteosarcoma in the femur, eleven each were located in the humerus and tibia, six were identified in the pelvis, and one each in the clavicle, maxilla, fibula, sacrum, ulna and radius.Pre-operative tissue diagnosis of osteosarcoma was obtained through computed tomography-guided percutaneous biopsy in over ninety percent of patients.Following initial therapy, over fifty percent of patients remained relapse-free during the follow-up period, with twelve percent and twenty-seven percent of patients documented as having local and distant disease recurrence, respectively. Of patients with recurrent disease, sixty-two percent remained disease-free following subsequent surgical intervention (most commonly, pulmonary metastatectomy).

Conclusion

Patient outcomes can be optimised through a multidisciplinary approach in a tertiary referral centre. At St Vincent's Hospital, survival and relapse rates of patients managed for osteosarcoma compare favourably with the published literature.

     

Apoptosis,cytokine and chemokine induction by non-structural 1 (NS1) proteins encoded by different influenza subtypes

Background

Influenza pandemic remains a serious threat to human health. Viruses of avian origin, H5N1, H7N7 and H9N2, have repeatedly crossed the species barrier to infect humans. Recently, a novel strain originated from swine has evolved to a pandemic. This study aims at improving our understanding on the pathogenic mechanism of influenza viruses, in particular the role of non-structural (NS1) protein in inducing pro-inflammatory and apoptotic responses.

Methods

Human lung epithelial cells (NCI-H292) was used as an in-vitro model to study cytokine/chemokine production and apoptosis induced by transfection of NS1 mRNA encoded by seven infleunza subtypes (seasonal and pandemic H1, H2, H3, H5, H7, and H9), respectively.

Results

The results showed that CXCL-10/IP10 was most prominently induced (> 1000 folds) and IL-6 was slightly induced (< 10 folds) by all subtypes. A subtype-dependent pattern was observed for CCL-2/MCP-1, CCL3/MIP-1α, CCL-5/RANTES and CXCL-9/MIG; where induction by H5N1 was much higher than all other subtypes examined. All subtypes induced a similar temporal profile of apoptosis following transfection. The level of apoptosis induced by H5N1 was remarkably higher than all others. The cytokine/chemokine and apoptosis inducing ability of the 2009 pandemic H1N1 was similar to previous seasonal strains.

Conclusions

In conclusion, the NS1 protein encoded by H5N1 carries a remarkably different property as compared to other avian and human subtypes, and is one of the keys to its high pathogenicity. NCI-H292 cells system proves to be a good in-vitro model to delineate the property of NS1 proteins.

     

野化培训大熊猫采食和人为砍伐对拐棍竹无性系种群更新的影响

亚高山竹类占据着野生大熊猫(Ailuropoda melanoleuca)食物组分的99%,竹子的生命周期与大熊猫的生活史密切相关,竹子的更新和生长直接影响着大熊猫的生存与保护.为了弄清大熊猫的采食利用和人为砍伐是否促进或制约竹子的繁殖更新,应用样方法、定位观察法,连续研究了卧龙自然保护区大熊猫栖息地内野化培训大熊猫采食、人为砍伐和对照样方中拐棍竹无性系种群的更新动态.结果表明,在相同种群数量和环境条件的基础上(p>0.05),被大熊猫采食竹子的比例为67.07%,致死率29.07%;人为砍伐竹子的比例为65.67%,致死率46.68%,可见与大熊猫采食相比,砍伐更为影响拐棍竹种群的生存.从出笋数量来看,不同处理方式有利于拐棍竹无性系种群的更新(p<0.05),尤其是人为砍伐措施大大提高了竹子的出笋率,但人为砍伐样方的竹笋质量(地径和株高生长)却远低于大熊猫采食和对照样方,未能达到大熊猫觅食的选择利用标准.不同年份之间,各种处理方式下的拐棍竹出笋数量波动较大(p=0.006～0.035),并随着恢复时间的延长,逐渐趋于相似(2007,p=0 825).不同处理方式之间,拐棍竹无性系种群的年补充率,各年份均具有显著性差异(p<0.05,除2007年外),年死亡率仅2003年和2004年有明显的差异(p<0.05),2005～2007年均不显著(p>0.05).拐棍竹无性系种群的年补充率与年死亡率之间表现出年补充率﹥年死亡率的格局(p<0.05),唯有大熊猫采食样方的2004年和2005年、人为砍伐和对照样方的2005年的年死亡率略高于年补充率(p>0.05),这表明拐棍竹无性系种群对大熊猫采食和人为砍伐具有无性系整合的补偿效应.     

Peanut gene expression profiling in developing seeds at different reproduction stages during <Emphasis Type="Italic">Aspergillus parasiticus</Emphasis>infection

Background  

Peanut (Arachis hypogaea L.) is an important crop economically and nutritionally, and is one of the most susceptible host crops to colonization of Aspergillus parasiticus and subsequent aflatoxin contamination. Knowledge from molecular genetic studies could help to devise strategies in alleviating this problem; however, few peanut DNA sequences are available in the public database. In order to understand the molecular basis of host resistance to aflatoxin contamination, a large-scale project was conducted to generate expressed sequence tags (ESTs) from developing seeds to identify resistance-related genes involved in defense response against Aspergillus infection and subsequent aflatoxin contamination.     

Sequence context affects the rate of short insertions and deletions in flies and primates

Background  

Insertions and deletions (indels) are an important evolutionary force, making the evolutionary process more efficient and flexible by copying and removing genomic fragments of various lengths instead of rediscovering them by point mutations. As a mutational process, indels are known to be more active in specific sequences (like micro-satellites) but not much is known about the more general and mechanistic effect of sequence context on the insertion and deletion susceptibility of genomic loci.     

Blood Selenium Associated with Health and Fertility in Norwegian Dairy Herds

A survey of blood selenium (Se) concentrations in Norwegian Red heifers and dry period cows was conducted to reveal possible association to management, feeding, health and fertility. Selenium contents were determined in 254 herd blood samples consisting of pooled samples from individual non-lactating animals from herds in 5 counties. The Se concentrations showed a normal distribution with mean 0.09 μg Se/g blood, with a standard deviation (SD) of 0.05, and ranged from 0.02 to 0.23 μg/g, with 50 % of the samples being between 0.06 and 0.11 μg/g. The herds with Se concentrations below 0.06 μg/g were smaller (21.4 ± 8.7 cow-years) than those with Se levels above 0.11 μg/g (27.5 ± 14.1 cow-years) (P < 0.01), but there were no differences in milk yield, incidence of replacement, proportion of animal culling, amount of concentrate or grass silage as percentage of energy consumption between the groups. Treatment registration records showed a tendency that more animals in the low Se herds were treated for all the diseases included in this investigation (64.8 animals per 100 cow-years) than those in the high Se herds (57.5 per 100 cow-years), while no such differences were revealed for individual disorders. There was, however, a significant difference in bulk milk somatic cell counts (BMSCC) between low and high Se herds, their values being 137 000 and 155 000 cells/ml, respectively. This difference was significantly influenced by herd size. Furthermore, a total of 4 916 lactations were analyzed from individual health and fertility recordings, including 2 934 first lactations and 1 982 later lactations. The present study revealed a reduced incidence of disease treatment with increased Se concentrations from 0.02 to 0.23 μg Se/g blood. In this regard, there seemed to be an optimum of 0.10 to 0.15 μg Se/g for all types of mastitis treatments summarized, and for treatment of retained placenta. Thus, herd Se concentrations below and above these values was connected with increased probability for sum mastitis and retained placenta, reflecting the effect of the quadratic term of Se. The cow (composite) milk somatic cell count (SCC) was lower in lactations from low Se herds than in high Se herds with a marked SCC increase in the Se concentration interval from 0.11–0.13 μg/g blood. In conclusion, heifers and dry period cows in Norway are low in blood Se content and there seems to be a positive association between increased blood Se concentration pre partum and decreased incidence of mastitis, ovarian cysts and anoestrus/silent oestrus post partum.     

Landscape fragmentation generates spatial variation of diet composition and quality in a generalist herbivore

Forest fragmentation may benefit generalist herbivores by increasing access to various substitutable food resources, with potential consequences for their population dynamics. We studied a European roe deer (Capreolus capreolus) population living in an agricultural mosaic of forest, woodlots, meadows and cultivated crops. We tested whether diet composition and quality varied spatially across the landscape using botanical analyses of rumen contents and chemical analyses of the plants consumed in relation to landscape metrics. In summer and non-mast winters, roe deer ate more cultivated seeds and less native forest browse with increasing availability of crops in the local landscape. This spatial variation resulted in contrasting diet quality, with more cell content and lower lignin and hemicellulose content (high quality) for individuals living in more open habitats. The pattern was less marked in the other seasons when diet composition, but not diet quality, was only weakly related to landscape structure. In mast autumns and winters, the consumption of acorns across the entire landscape resulted in a low level of differentiation in diet composition and quality. Our results reflect the ability of generalist species, such as roe deer, to adapt to the fragmentation of their forest habitat by exhibiting a plastic feeding behavior, enabling them to use supplementary resources available in the agricultural matrix. This flexibility confers nutritional advantages to individuals with access to cultivated fields when their native food resources are depleted or decline in quality (e.g. during non-mast years) and may explain local heterogeneities in individual phenotypic quality.     

营养盐初始组合浓度对类波氏真眼点藻生长和油脂积累的影响

为了研究培养基中主要营养元素氮、磷、硫初始组合浓度对类波氏真眼点藻(Eustigmatos cf.polyphem D.J.Hibberd)生长、油脂积累和脂肪酸组成的影响,分别以NaNO_3、K_2HPO_4、MgSO_4为氮、磷、硫源,设置不同的营养盐初始组合浓度对其进行培养,采用干重法、重量法、气相色谱分析法、元素分析仪等依次对生物量、总脂、脂肪酸组成和细胞内元素含量进行测定。结果显示,氮、磷、硫三者的初始组合浓度变化对类波氏真眼点藻的生长和油脂积累具有明显的影响,9 mmol/L NaNO_3、0.92 mmol/L K_2HPO_4、0.3 mmol/L MgSO_4营养条件下其生物量达到最大值,为9.19 g/L;低氮、低磷胁迫或二者共同胁迫均促进其油脂积累,但低硫胁迫对油脂的积累影响不大,最大油脂含量出现在3 mmol/L NaNO_3、0.029 mmol/L K_2HPO_4、0.3 mmol/L MgSO_4条件下,为68.7%(DW)。此外,类波氏真眼点藻富含棕榈油酸,占总脂肪酸含量的最大值达60.88%。选择合理的氮、磷、硫初始组合浓度能够有效促进其生长或油脂及棕榈油酸的积累。