一株高效抑藻放线菌的分离筛选及鉴定

从福建云霄国家红树林自然保护区滩涂沉积物样品中,共分离获得521株纯培养物.通过检测球形棕囊藻(Pha-eocystis globosa)荧光强度计算抑藻率,从521株菌中筛选到27株具有抑藻活性的菌株.在27株抑藻菌中,菌株O3-26对球形棕囊藻具有最高的抑藻率(高达96.71%).菌株O3-26的抑藻谱实验显示,该菌株抑藻活性表现出一定的种属特异性,对硅藻门(Bacillariophyta)的三角褐指藻(Phaeodactylum tricornutum)和中肋骨条藻(Skeletonema costatum)2株测试藻株没有抑制作用,而对绿藻门(Chlorophyta)盐生杜氏藻(Dunaliella salina)和自养小球藻(Chlorella autotrophi-ca)2株藻株具有较强抑藻作用.扫描电镜观察显示,该菌株孢子丝直至螺旋状且孢子表面带刺.生理生化实验显示,该菌株在所得到的大多数培养基上生长良好,在营养琼脂培养基中可以产生水溶性色素;不能在棉子糖作为唯一碳源的培养基上生长.16SrRNA基因相似性分析表明,菌株O3-26属于链霉菌属(Streptomyces),并与灭癌素链霉菌(Streptomycesgancidicus)15412菌株具有最高的同源性(99%).生理生化实验表明,二者之间生理特征存在一定差异.综合形态特征、生理特征以及系统发育分析的结果,鉴定该菌株为灭癌素链霉菌

嗜虫耶尔森氏菌HlyA及HasA外分泌表达系统的构建

【目的】构建一株具备外分泌蛋白功能的工程菌,解决杀虫毒素无法由胞内分泌至胞外,无法直接作用于虫体等问题,为松墨天牛防治提供新思路。【方法】本研究先测定从松墨天牛肠道及其生境中分离出的嗜虫耶尔森氏菌(CSLH88)的生长特性及抗性,进而对其进行分子改造。构建HlyA (pGHKW2)以及HasA (pGHKW4)外分泌表达载体,利用电穿孔法将其转入CSLH88菌株,获得能够表达绿色荧光蛋白的工程菌。利用稀释涂板及荧光体式镜检测技术对两个质粒进行遗传稳定性检测,并采用SDS-PAGE及Western blotting技术验证蛋白外分泌功能。【结果】CSLH88菌株培养2–4 h能够进入对数生长期,并对卡那霉素(Kan)敏感。成功构建了含有Kan抗性基因的pGHKW2(GenBank:MK562405)和pGHKW4(GenBank:MK562404)两个外分泌表达载体的CSLH88工程菌株。其中,发现pGHKW4质粒更加适合在嗜虫耶尔森氏菌中稳定遗传。SDS-PAGE及Western blotting检测结果表明HlyA系统无法在CSLH88菌株中将目的蛋白分泌到胞外,而HasA系统则可以有效地发挥外分泌表达功能。【结论】通过对HlyA及HasA两个外分泌表达系统进行研究,从中筛选出HasA型血红素转运系统作为CSLH88菌株的外分泌表达系统,为后续外分泌杀虫毒素蛋白菌株构建以及CSLH88菌株的致病性研究奠定基础。国家重点研发计划课题(2017YFD0600105);;\n国家自然科学基金(31601905);;\n福建省科技厅自然科学基金(2016J01097);;\n福建省科技计划项目(2018N5002);;\n福建农林大学科研基金(xjq201614);福建农林大学林学院林学高峰学科项目(71201800720,71201800753,71201800779);;\n福建省大学生创新创业训练计划项目(201810389102);;\n福建省林业科学研究项目[Minlinke(2017)03]~

STUDIES ON THE CHEMICAL CONSTITUENT FROM SEA CUCUMBER MENSAMARIA INTERCEDENS──Ⅱ.PMI-1, A NOVEL FUCOSAN

利用乙醇沉淀分级技术从二色桌片参双酶水解液里分离到一种纯的活性多糖，定名PMI-1。用高效薄层层析法鉴定其组成的单糖，发现PMI-1只含有L-岩藻糖。根据其分析结果和理化特性，PMI－1应是由若藻糖基和岩藻糖-4硫酸酯基组成的均多糖。研究PMI－1的核磁共振谱，可认定它是一种具有直链的和α1－α联接的均多糖。因此它是一种新的岩藻聚糖。Using double enzymes digestion and fractionations by precipitation with alcohol, a pure active polysaccharide PMI-1 was obtained from the dried body wall of Mensamaria intercedens. Monosaccharide composition was identified by thin-layer chromatography, the results indicate that PMI-1 only consisits of L-fucose. According to analytical and chemicophysical data, PMI-1 seemed to be homosaccharide consisted of fucose and fllcose-4sulfate with the molar ratio of 1: 1. Studied on the 1 HNMR and 13MNMR data of PMI-1 showed that PMI-1 had a straightsaccharide chain and α1-α linkage.福建省重中之重项

Studies on the Chemical Composition of Sea Cucumber Mensamaria Intercedens Ⅲ. Immunomodulative Effects of PMI-1

研究了二色桌片参多糖－１（ＰＭＩ－１）在体内、体外的免疫调节作用．结果表明，二色桌片参多糖－１体外能促进小鼠脾淋巴细胞增殖，促进小鼠脾淋巴细胞分泌ＩＬ－２；体内能显著促进迟发型超敏反应，增加脾指数、胸腺指数；具有显著的免疫增强作用。The immunoactivities of PMI-1 are studied in vivo and in vitro. PMI-1(5. 50. 100u g/mL ) in vitro could promote the proliferation of mice lymphocyte, and increase IL-2 production as well. In vivo, PMI-1 could significantly improve the delayed-type hypersensitivity (DTH) degree and the weights of spleen and thymus, although it has little effect on the production of hemolysin antibodies. The results indicate that PMI-1 could remarkedly enhance the immunofunction in mice

Isolation and Identification of a High Algicidal Actinomycete

通讯作者：[email protected][中文文摘]从福建云霄国家红树林自然保护区滩涂沉积物样品中，共分离获得521株纯培养物．通过检测球形棕囊藻(Phaeocystisglobosa)荧光强度计算抑藻率，从521株菌中筛选到27株具有抑藻活性的菌株．在27株抑藻菌中，菌株03-26对球形棕囊藻具有最高的抑藻率(高达96.71%).菌株03-26的抑藻谱实验显示，该菌株抑藻活性表现出一定的种属特异性，对硅藻门(Bacillariophyta)的三角褐指藻(Phaeodactylum tricornutum)和中肋骨条藻(Skeletonema costatum)2株测试藻株没有抑制作用，而对绿藻门(Chlorophyta）盐生杜氏藻(Dunaliella salina)和自养小球藻(Chlorella autotrophica)2株藻株具有较强抑藻作用．扫描电镜观察显示，该菌株孢子丝直至螺旋状且孢子表面带刺．生理生化实验显示，该菌株在所得到的大多数培养基上生长良好，在营养琼脂培养基中可以产生水溶性色素；不能在棉子糖作为唯一碳源的培养基上生长．１６ＳｒＲＮＡ基因相似性分析表明，菌株Ｏ３－２６属于链霉菌属（Ｓｔｒｅｐｔｏｍｙｃｅｓ），并与灭癌素链霉菌（Ｓｔｒｅｐｔｏｍｙｃｅｓｇａｎｃｉｄｉｃｕｓ）１５４１２菌株具有最高的同源性（９９％）．生理生化实验表明，二者之间生理特征存在一定差异．综合形态特征、生理特征以及系统发育分析的结果，鉴定该菌株为灭癌素链霉菌．[英文文摘]521 strains were isolated from mangrove sediments in the Yunxiao Mangrove National Nature Reserve,Fujian,China.27 algicidal isolates were screened based on the analysis of fluorescence intensity of Phaeocystis globosa.The strain named O3-26,which showed the highest algcidal activity(96.71%) against P.globosa among these 27 algicidal isolates,was studied in this article.The test of algicidal spectrum illustrated O3-26 showing a algicidal characteristic of algae species specificity,with high algicidal effects on Dunaliella salina and Chlorella autotrophica,no algicidal effects on phaeodactylum tricornutum and Skeletonema costatum.Observed by scanning electron microscope,03-26 has spiral sporot hrix and barbed spores.This strain can grow well in most of the medium in this article,and produce soluble pigment in nutrient agar.The physiological tests demonstrated that 03-26 owns a wide range of carbon sources,but can be grown on raffinose medium as sole carbon source.On the basis of 16 SrRNA gene sequence similarity,the strain,03-26 was shown to be related most closely to Streptomyces gancidicus 154152(99%).Although some differences fromS.gancidicus were tested from physiological and biochemical experiments,03-26 was still indentificated as S.gancidicus by comparison on morphological,physiogical characteristics and phylogenetic analysis.国家高技术研究发展计划(863)项目(2008AA09Z408); 国家自然科学基金项目(40930847,31070442);近海海洋环境科学国家重点实验室(厦门大学)访问学者基金项目(MELRS1124