菲律宾蛤仔养殖水体中大肠菌群安全限量的研究

在不同大肠菌群含量的养殖水体中,通过菲律宾蛤仔对大肠菌群的富集研究,发现菲律宾蛤仔对大肠菌群的富集量随着大肠菌群在养殖水体中含量的升高而增大,推导出的富集量公式为y=0.9254x2+17.366x-123.77。参照CAC、91/492/EEC和NSSP的标准,通过相关换算及与国内标准对比,建议菲律宾蛤仔养殖水体中大肠菌群的安全限量定为≤70cfu/100ml,生食菲律宾蛤仔养殖水体中大肠菌群的安全限量定为≤14cfu/100ml。     

硝化细菌浓度及滤料对养殖水中氨氮处理效果的影响

分别研究了不同硝化细菌浓度(0、20、60、120 mL/100 L)和不同微生物滤料(珊瑚石、锅炉煤渣、牡蛎壳)对养殖水中氨氮处理效果的影响。结果显示,添加硝化细菌后,水体中的氨氮浓度呈现下降趋势,在8～12 h出现极低值后,开始上升,但上升速度较慢;随着水体中硝化细菌添加量的增加,水体中的氨氮浓度下降速度加快;水体中亚硝酸氮浓度呈现先上升后下降的趋势,并在4～6 h出现极高值,然后迅速下降,且硝化细菌添加量越高,下降速度越快。硝化细菌对以珊瑚石和锅炉煤渣为滤料的养殖水体中氨氮和亚硝酸氮的处理效果显著优于牡蛎壳,但珊瑚石和锅炉煤渣之间无显著差异。综合试验结果,应急水质处理时,硝化细菌菌剂的添加量以一次60 mL/100 L(或以活菌计数为1.2×109个/100 L)、间隔24 h添加1次为宜;经过脱硫筛选之后的锅炉煤渣可以作为循环水养殖用滤料。     

褐藻酸降解菌引起海带内皮层细胞储藏颗粒变化的初步研究

实验对正常海带用褐藻酸降解菌进行了人工感染，经培养使其产生烂斑，通过组织显微切片进行比较观察，发现褐藻酸降解菌首先侵入内皮层，然后再逐渐侵入到外皮层、髓部，最终导致海带病烂。切片中发现在侵染海带切片内皮层的细胞中，有大量储藏颗粒，形成椭圆或圆形。其直径范围在1．5－4．0μm之间，平均直径为2．7μm。该颗粒主要存在内皮层，从其产生到消亡的过程分析，与褐藻酸降解菌的侵入有关，并作为营养起到促进菌增殖的作用。     

鲤细胞因子多克隆抗体的制备及检测

为从蛋白质水平研究细胞因子在鲤体内免疫应答过程中的合成变化,本研究采用PCR技术克隆TNF-α、IL-1β、IL-6、IL-12、IL-10和TGF-β基因含有部分抗原决定簇的片段,引入双酶切位点Bam HⅠ和HindⅢ后连接至p ET-32a/21a,构建相应的表达载体,制备多克隆抗体。采用ELISA检测抗体效价,并以此作为实验工具,检测经嗜水气单胞菌感染后鲤血清中炎性细胞因子的合成变化。结果显示,基因TNF-α、IL-1β、IL-6、IL-12、IL-10和TGF-β融合蛋白分子量分别约为31.8、31.7、35.3、32.5、18.0和33.6 ku;抗体效价达到2.4×106;在病原菌感染后的不同阶段,促炎细胞因子TNF-α、IL-1β、IL-6、IL-12和抗炎细胞因子IL-10、TGF-β呈现出不同的合成变化。研究表明,制备的抗体具有较高的效价、亲和力和特异性,可用于鲤细胞因子的定量研究,该抗体的获得为鲤免疫应答与细胞因子合成的系统研究奠定了基础。同时,获取的鲤细胞因子TNF-α、IL-1β、IL-6、IL-12、IL-10和TGF-β的抗体亦可用于其他鱼类细胞因子蛋白质水平的定量研究。     

Effects of different dietary copper sources on the growth and intestinal microbial communities of Pacific white shrimp (Litopenaeus vannamei)

An 8‐week feeding trial was conducted to evaluate the effects of dietary copper sources on growth performance and intestinal microbial communities of juvenile Pacific white shrimp (Litopenaeus vannamei). Four isonitrogenous (420 g/kg crude protein) and isolipidic (70 g/kg crude lipid) experimental diets were formulated to contain different copper sources, Diet NSC (no‐supplemented copper), Diet CS (copper sulphate), Diet Availa (copper amino acid complex from Availa^®Cu100) and Diet M (1:1, copper sulphate + copper amino acid complex). A total of 360 Pacific white shrimp juveniles (initial weight: 1.86 ± 0.03 g) were randomly allocated into four groups, with three replicates per group and 30 shrimps each replicate. An Illumina‐based sequencing method was used to analyse the intestinal bacterial composition of shrimp juveniles. The results revealed that shrimps fed Diet M had significantly higher percent weight gain (PWG) than that fed Diet NSC and Diet CS, while there were no significant differences between Diet M and Diet Availa. Survival was not affected by the dietary copper sources. Proteobacteria and Bacteroidetes were dominant in the intestines of Pacific white shrimp. Shrimps fed Diet M and Diet Availa had higher bacterial richness and diversity than those fed Diet NSC and Diet CS with no significant differences among all treatments. Furthermore, Diet M and Diet Availa which contained copper amino acid complex reduced the relative abundance of Vibrionaceae in shrimp intestinal tract. This study indicated that dietary copper sources could affect the intestinal microbial communities and diets supplemented with Availa^®Cu100 could have a positive impact on the growth performance and decrease the potential risk of disease in Pacific white shrimp.     

乳酸菌培养液中活菌数与吸光度的关系研究

通过平板计数法测定脱脂乳为培养基的发酵液中乳酸菌的活菌数,并测其对应的吸光度,研究两者之间的相关关系。结果表明:在发酵过程中以脱脂乳为培养基的发酵液中乳酸菌的活菌数与吸光度之间呈线性相关关系。因此,通过测定发酵液的吸光度可以评定发酵乳制品中乳酸菌的活菌数。     

耐胃肠道环境及肠道病原菌拮抗的猪源乳酸菌的分离与筛选

从健康猪粪便筛选分离到对肠道致病菌有较强抑制特性的3株乳酸菌,分别鉴定为屎肠球菌(Enterococcus faecium)、粪肠球菌(Enterococcus faecalis)和干酪乳杆菌(Lactobacillus casei)。体外益生特性研究表明:3株乳酸菌能耐受胃肠道环境,能黏附猪小肠表面粘液,对猪源产肠毒素型大肠杆菌(Escherichia coli O139,E. coli O139)、E. coli C83905、E. coli C83524、E. coli C83529、伤寒沙门氏菌(Salmonella typhimurium O4Hi,ST O4 Hi)、金黄色葡萄球菌(Staphylococcus aureus,SA)均有较强的抑制特性。对小白鼠肠道大肠杆菌的数量进行研究,结果证明对照组大肠杆菌数量没有明显降低,而添加乳酸菌组大肠杆菌数量明显降低。对小白鼠腹腔注射E. coliC83905,乳酸菌组死亡率明显小于对照组。结果证明3株乳酸菌在猪饲料添加剂方面有较好的应用前景。     

罗非鱼"突眼"病细菌性病原的分离与鉴定

从患"突眼"症状的罗非鱼体内分离到多株细菌,经人工感染证实其中TL60829NA菌株为病原菌,经生理、生化和API细菌鉴定系统以及分子生物学试验,鉴定其为痢疾志贺氏菌(Shigella dysenteriae)。其主要生理生化特征为革兰氏阴性短杆菌,不具动力,氧化酶、过氧化氢酶、V-P反应、吲哚试验、苯丙氨酸、甘露醇、KCN均为阴性;对氨苄西林、羧苄西林、红霉素、多西环素、氟哌酸、卡那霉素敏感,对呋喃唑酮、复方新诺明、磺胺和庆大霉素不敏感。     

红壤溶磷菌的筛选及溶磷机制

采用以磷酸铝为磷源的蒙金娜(PVK)液体培养基研究了从红壤土中筛选出的4种溶磷菌的溶磷效果,选出其中的优势菌株B1,并对其溶磷机理做出初步探讨。结果表明,所筛选出的4株溶磷菌在液体培养条件下均有显著的溶磷效果,其中菌株B1在培养4 d后有效溶磷量最大,达到292.8 mg L-1。各处理培养液pH在培养期间均有显著下降,pH从7.0下降至3.2~4.7。高效液相色谱测定发现,各菌株培养液中有机酸的种类与含量随培养时间变化而不同,其中菌株B1主要分泌草酸和苹果酸,培养1 d后有机酸总量可达到5 mmol L-1;通过添加有机酸对磷酸铝活化的试验表明,分泌有机酸溶磷仅是菌株B1溶磷机制之一,可能还存在其他溶磷机制。菌株B1生长的适宜pH范围为5~9,最适培养温度为30℃,100 ml三角瓶的最适装液量为30~40 ml。经鉴定,菌株B1与苏云金芽孢杆菌有99.9%的相似性。     

Fungal endophyte Phomopsis liquidambari B3 enriches the diversity of nodular culturable endophytic bacteria associated with continuous cropping of peanut

Previously, the application of the fungal endophyte Phomopsis liquidambari B3 exhibited notable effect in alleviating soil obstacles caused by continuous cropping of peanut. It was supposed that P. liquidambari B3 could enhance the efficiency of nodulation and nitrogen fixation by enriching the diversity of nodular endophytic bacteria under continous monocropping system. To verify this hypothesis, a pot experiment was conducted under natural conditions. Four treatments were set as follow: discontinuous cropping soil (NS), continuous cropping soil (CK), continuous cropping soil inoculate with actived P. liquidambari B3 (CSB3), and continuous cropping soil inoculate with sterilized P. liquidambari B3 (CSIB3). As a result, a total of 120 isolates were obtained. Our study clearly declared that the addition of fungal endophyte effectively enriched the genetic diversity and the community composition in CSB3. In contrast, a low genetic diversity level and simplex community structure were exhibited in CK and CSIB3. Meanwhile, the positive percentage rate and corresponding community composition of the plant growth-promoting isolates in NS and CSB3 were larger and more abundant than the other. This is the first time to describe the effects of fungal endophytes on the diversity of the nodular culturable endophytic bacteria associated with continuous cropping of peanut.