菊花绿变病植原体在中国的发生及分子鉴定

对内蒙古农业大学校园内表现花器绿变症状的菊花样品进行采集和DNA提取,应用植原体16S rRNA基因和rp基因的引物进行巢式PCR扩增,从感病样品中分别扩增得到了长度均约为1.2 kb的片段。序列一致性分析表明,菊花绿变植原体16S rRNA基因与翠菊黄化植原体匈牙利风信子株系(GenBank登录号MN080271)、印度玉米株系(KY565571)、印度繁缕株系(KC623537)和印度马铃薯株系(KC312703)的核酸一致性最高,为99.9%,rp基因序列与翠菊黄化植原体立陶宛洋葱株系(GU228514)的核酸一致性最高,为99.8%。基于16S rRNA基因和rp基因构建系统进化树时发现,菊花绿变植原体均与16SrI-B亚组成员聚为一起。16S rRNA基因相似性系数分析表明,菊花绿变植原体与洋葱黄化植原体(AP006628)的相似性系数最高为1.00,洋葱黄化植原体(AP006628)在分类上属于16SrI-B亚组。因此,我们可以确定该菊花绿变植原体属于16SrI-B亚组。这是我国首次报道菊花绿变病的发生。     

坛紫菜叶状体的抗生素联合除菌效果

为研究坛紫菜及其外生菌群对氯霉素、竹桃霉素和头孢噻肟3种抗生素的敏感性,通过将其与氨苄青霉素、卡那霉素和庆大霉素的组合,优化坛紫菜叶状体的除菌方法,并利用涂布平板、qRT-PCR及16S rRNA测序分析其除菌效果。结果显示,当坛紫菜先以氨苄青霉素(300 mg/L)、卡那霉素(100 mg/L)和庆大霉素(100 mg/L)混合处理18 h,再以氯霉素(50 mg/L)、头孢噻肟(200 mg/L)和竹桃霉素(50 mg/L)混合处理4 d,叶状体的健康率保持在96.3%以上,而对可培养细菌的抑制率达到99.9%。根据qRT-PCR和16S rRNA的结果可知,该种组合方法较前3种抗生素的组合,其菌总数下降41.5%,微生物的丰度和多样性指数明显下降。其中,假单胞菌、交替赤杆菌、交替单胞菌以及海杆菌等得到针对性的抑制。研究表明,多种抗生素的优化组合能够对坛紫菜叶片的菌群结构产生显著影响,使其受到抑制,但仍无法达到绝对除菌的效果。     

Construction and Partial Biological Characteristics Analysis of Listeriolysin S llsB Gene Deletion Strain

In order to analyze the effect of listeriolysin S (LLS) llsB gene deletion on the biological characteristics of Listeria monocytogenes (LM),this study used homologous recombination to construct the llsB gene deletion strain LM90-ΔllsB,and the biological characteristics of growth characteristics,median lethal dose (LD₅₀) and organ-borne bacteria were studied in healthy Kunming male mice at 8 weeks old and weighing 40 g±5 g.The llsB gene deletion strain was successfully constructed,and the deletion strain had good genetic stability through continuous passage to 20 generations in vitro.Based on its growth curve examination,we found that the growth rate of the mutant strain was slightly higher than that of the parent strain.The results of mice infection test showed that the LD₅₀ of the parent strain and the deletion strain were 10^6.17 and 10^6.50 CFU,respectively.Compared with the parent strain,the amount of bacteria load of the deletion strain in the liver and spleen of the mice was extremely significantly decreased (P<0.01),the results showed that the infection ability of the mutant strain on mice was obviously weakened.No Listeria monocytogenes was detected in the brain.The results suggested that llsB gene might have direct or indirect regulatory effect on some biological characteristics of LM90,and it would provide a theoretical basis for further understanding of the pathogenic mechanism of LLS,prevention and control of listeriosis.     

规模化畜禽养殖场粪便中多重耐药菌分离鉴定及其耐药特征

为了解禽畜粪便中多重耐药菌的污染特征,本研究对鸡粪、牛粪、猪粪和有机肥这4种不同样品中多重耐药菌进行了计数分析,并对不同来源的多重耐药菌株进行了分离和纯化,进一步开展了基于16S rDNA序列比对的分子生物学鉴定以确定其种属地位,以及基于药敏试验的耐药性特征分析。结果表明:不同养殖动物粪便中四环素、恩诺沙星、磺胺甲恶唑和泰乐菌素4种抗生素多重耐药菌的绝对数量和相对数量排序均为鸡粪>猪粪>牛粪,粪源有机肥中可培养的多重耐药菌的绝对数量和相对数量在堆肥后均有所下降。通过对耐药菌株鉴定和分类学分析,发现禽畜粪便中多耐药菌的菌门集中分布在Proteobacteria、Firmicutes和Actinobacteria,多重耐药菌的优势菌属为Escherichia、Corynebacterium、Kurthia;而有机肥样品中多重耐药菌的优势菌属为Staphylococcus、Glutamicibacter。菌株的药敏试验表明,3类动物粪污中的多重耐药菌都对红霉素、四环素有着较高的耐药率,均高于80%,而对阿米卡星这种抗生素表现出较好的敏感性,其耐药率低于30%。通过对Ⅰ、Ⅱ类整合子基因盒的扩增,发现PCR产物的大小从0.8 kb到1.8 kb不等,基因盒ad A2、dfr A17、dfr A1及sat2在养殖场粪污中检出率均较高。     

四种短体线虫的形态和分子生物学鉴定

 采用形态学和分子生物学方法对来自荷兰的藏橐吾和铃兰、泰国的高山榕、缅甸的香蕉等种苗(球)分别鉴定出玻利维亚短体线虫Pratylenchus bolivianus、铃兰短体线虫P. convallariae、咖啡短体线虫P. coffeae和斯佩奇短体线虫 P. speijeri。对这4种线虫的形态特征进行较为详细的描述后,认为头环、口针、侧区、生殖系统和尾形等形态特征是种类鉴定的重要依据;进一步利用引物28S-D2A/28S-D3Br扩增测序得到上述4种线虫的28S rRNA基因D2/D3区序列,序列分析发现玻利维亚短体线虫从欧洲到美洲的不同种群之间的遗传距离变异较小,仅为0~0.007;铃兰短体线虫同一种群不同个体之间的遗传变异较大,为0.006~0.029;咖啡短体线虫不同种群之间的平均遗传距离为0.013;斯佩奇短体线虫不同种群之间的平均遗传距离为0.010;咖啡短体线虫P. coffeae和斯佩奇短体线虫P. speijeri亲缘关系很近,二者种间平均遗传距离仅为0.026。本研究再次证明线虫28S rRNA基因D2/D3区基因序列可作为短体线虫种间鉴定的依据。     

Effects of oral administration of Spirulina platensis and probiotics on serum immunity indexes,colonic immune factors,fecal odor,and fecal flora in mice

To evaluate the effects of Spirulina platensis and probiotics on growth, immunity indexes, fecal flora, and fecal odor in mice, 40 mice were randomly allotted to four groups, and each was administrated with nothing, S. platensis, probiotics, or both for 28 days, respectively. Then, many indexes were measured. The results showed that S. platensis was more effective (P < 0.001) than probiotics in improving mice's feed conversion ration (FCR). In immunity, probiotics administration increased (P < 0.042) serum IgE, IgM, IFN-γ, colonic AHR, TLR4, and NF-κB protein expression and decreased (P < 0.039) serum IL-1α, IL-21, IL-22, and colonic ARNT gene expression. However, the S. platensis showed weaker effect, which increased (P < 0.025) the serum IgE, IgM, TNF-α, and the colonic AHR and NF-κB protein expression, and decreased (P < 0.01) serum IL-21. Probiotics consumption decreased the fecal odor by decreasing (P < 0.02) fecal Escherichia coli, indole-3-acetic acid (IAA), and skatole contents, and the S. platensis decreased (P = 0.04) the IAA. These results indicated that oral administration of probiotics, S. platensis, or both of them in mice probably benefited body's immunity and reduced fecal odor. However, their mechanisms were still unclear and need further study.     

Using genetic methods for analysis of fish meals and feeds employed in Greek mariculture

Large quantities of high protein fish meals are needed to sustain cultured species and thus the impact to marine ecosystem has been highly discussed. The aim of this study was to apply a PCR‐cloning methodology for a robust insight into the composition of commercial fish meals and feeds for farmed species of the Greek mariculture, assessing the risk posed by aquaculture to marine ecosystems but also the risk posed by commercial fish feeds to the increase in trophic level of species farmed in Greece. 89% of the sequences were identified to species level and only 11% to genus/family level. Overall, a total of 49 taxa were identified (44 fish species/taxon, five non‐fish species/taxon). Even though small pelagic fish like Engraulis sp. were the main portion, a wide range of species constituted the fish meals and feeds. Plant and animal species were also detected as an alternative protein source. Feed products employed in Greek mariculture still contain large portions of fish meals which increase the mean trophic level of farmed species causing a farming up trend. The results emphasize that such molecular methodologies are needed to certify aquafeeds allowing fish feed producers to demonstrate their commitment to sustainable aquaculture.     

Outer membrane protein A of Acinetobacter baumannii induces autophagy via Akt/mTOR/p70S6K signaling pathway in RAW264.7 cells

AIM:To analyze the effects of outer membrane protein A (OmpA) from Acinetobacter baumannii ATCC 19606 on the autophagy of RAW264.7 cells. METHODS:The RAW264.7 cell model stimulated by OmpA was established. The effects of OmpA on the autophagy of RAW264.7 cells were detected by immunofluorescence, Western blot and transmission electron microscopy. RESULTS:The OmpA increased the expression of LC3B-Ⅱ and reduced the phosphorylation levels of Akt, mTOR and p70S6K. Rapamycin further reduced the phosphorylation levels of mTOR and p-70S6K, and increased the expression of LC3B-Ⅱ induced by OmpA. CONCLUSION:The OmpA of Acinetobacter baumannii induces autophagy via Akt/mTOR/p70S6K signaling pathway in the RAW264.7 cells. This work provides a basis for further research on the molecular mechanism of autophagy induced by Acinetobacter baumannii to find a new method against the infection of Acinetobacter baumannii.     

基于B/S的农业气象自动化观测系统国家级平台设计与应用

农业气象自动化观测系统国家级平台是依托中国气象局组织的农业气象自动化观测试点工作而设计开发,平台的搭建采用B/S架构实现多级用户、跨平台登录,采用XML及JSON等数据传输格式实现国家级、省级、台站级用户信息交互,实现农业气象观测XML数据传输状态的监控、台站端设备状态的评估以及数据内容的实时查询。