添加嗜酸乳杆菌对凡纳滨对虾生长、酶活性及相关酶mRNA表达的影响

本实验旨在研究饲料中添加嗜酸乳杆菌(Lactobacillus acidophilus)对凡纳滨对虾(Litopenaeus vannamei)幼虾生长、非特异性免疫酶活性、抗病力和相关酶m RNA表达的影响。选取840尾初始均重为(0.58±0.01)g的凡纳滨对虾幼虾为研究对象,分为7个处理,每个处理3个重复。投喂添加不同比例(0%,0.1%,0.2%,0.4%,0.6%,0.8%,1.0%)嗜酸乳杆菌的饲料,养殖期8周。结果显示:添加不同比例嗜酸乳杆菌对凡纳滨对虾的成活率无显著影响(P0.05)。增重率与特定生长率随添加量的增加呈先上升后下降的趋势,但均显著高于对照组(P0.05),在0.4%组达到最大值;当添加0.2%时,饲料系数最低,其他各组显著低于对照组(P0.05),而蛋白质效率的变化规律则与饲料系数相反。血清酚氧化物酶(phenoloxidase,PO)、碱性磷酸酶(alkaline phosphatase,AKP)和酸性磷酸酶(acid phosphatase,ACP)活性随添加量的增加先升高后降低,分别为0.4%、0.1%和0.2%时,达到最大值。过氧化氢酶(catalase,CAT)、溶菌酶(lysozyme,LZM)和超氧化物歧化酶(superoxide dismutase,SOD)活性,实验组均显著高于对照组(P0.05)。实验组过氧化氢酶(CAT)m RNA表达量和溶菌酶(LZM)m RNA表达量均显著高于对照组(P0.05)。哈维弧菌(Vibrio harveyi)攻毒96 h,对虾存活率随添加量的增加显著升高(P0.05)。以增重率(weight gain rate,WGR)为判断依据,根据折线模型得出,饲料中添加0.23%嗜酸乳杆菌可显著促进凡纳滨对虾生长,并提高非特异性免疫酶活性。     

Lactic acid bacteria strains for enhancing the fermentation quality and aerobic stability of Leymus chinensis silage

Leymus chinensis is an important grass in China and Russia. Six lactic acid bacteria (LAB) strains (LB, LPL1, LPL2, LPL3, LCL and WH) from L. chinensis silage were screened and identified and their effects on fermentation quality were investigated. All six strains were grown at 6·5% NaCl and pH 4·00. Strains LPL1, LPL2 and LPL3 were identified as Lactobacillus plantarum, and LB, WH and LCL were classified as Lactobacillus brevis, Weissella hellenica and Lactobacillus casei respectively. The six isolated strains and a commercial inoculant (Lactobacillus buchneri) were added to L. chinensis for ensiling at densities of 500 and 600 kg m^?3. The control was sprayed with the same volume of distilled water. The effects of the strains on fermentation quality after 45 d ensiling and aerobic stability during 8 d of exposure to air were evaluated. The 600 kg m^?3 silage had lower pH, butyric acid, ammonia nitrogen content and coliform bacteria counts than the 500 kg m^?3 density silage (P < 0·05). The six isolated strains decreased pH, butyric acid content and increased lactic acid content, and all inoculants increased L. chinensis silage aerobic stability except LCL (P < 0·05). The fermentation quality of L. chinensis silage increased with higher ensiling density. The LAB strains improved the fermentation quality, and high‐quality silage could be obtained at low ensiling density with the addition of the LAB strains. The strains improved the aerobic stability; Lb. buchneri and Lb. brevis showed the best performance.     

Effects of ethanol,molasses and Lactobacillus plantarum on fermentation characteristics and aerobic stability of total mixed ration silages

This study was conducted to evaluate the effects of Lactobacillus plantarum, molasses and/or ethanol on fermentation quality and aerobic stability of total mixed ration (TMR) silage, which is widely used in dairy cow diets at mid‐to‐late lactation in Tibet. TMR was treated with ethanol (E), molasses (M), Lactobacillus plantarum(L), ethanol+molasses (EM), ethanol+Lactobacillus plantarum (EL) plus control. After 45 d of ensiling, inoculant significantly (P < 0·05) increased lactic acid (LA) concentration and decreased pH, ammonia nitrogen (AN) concentration, and aerobic bacterial and yeast counts, compared to control. After the first 3 d of aerobic exposure, LA for silages without ethanol started to decrease, while LA for E silages almost remained unchanged till the end of the aerobic exposure period. The pH in TMR silages without ethanol gradually increased, while that for E and EL remained about 4·60 and 4·00, respectively, and EL showed the lowest pH among six silages over the course of aerobic exposure. Aerobic bacterial counts in control, M and EM silages were significantly higher (P < 0·05) than those in E, L and EL, and yeast counts in E and EL silages were significantly lower (P < 0·05) than those in other silages after 9 d of aerobic exposure. The results suggest that inoculation with L. plantarum was more effective in altering fermentation characteristics than adding molasses, while ethanol showed a potential to protect TMR silages from pH increase and delayed the growth of aerobic bacteria and yeast either alone or in combination with L. plantarum.     

Aerobic stability of sugar-cane silage inoculated with tropical strains of lactic acid bacteria

Aerobic stability is an important feature in the evaluation of silages. The aims were to investigate the chemical and microbiological changes that occur in sugar-cane (Saccharum spp.) silage after aerobic exposure, to identify the major species of yeasts associated with the aerobic deterioration process and to select lactic acid bacteria (LAB) strains that can improve the aerobic stability of this silage. Fourteen wild LAB strains belonging to Lactobacillus plantarum, L. brevis and L. hilgardii were evaluated using experimental silos. Silage samples were collected at 0, 96 and 216 h after aerobic exposure to determinate the DM, WSC, pH, products of fermentation, to evaluate the silage temperatures and to identify yeast species associated with the aerobic deterioration of silage. The strains tested were able to modify the fermentative and chemical parameters and the diversity of yeasts species of silage after aerobic exposure. There was no association between the facultative or obligatory heterofermentative fermentation patterns and the increased aerobic stability of silage. Aerobic stability of sugar-cane silages was associated with high acetic acid and 1,2-propanediol concentrations. L. hilgardii UFLA SIL51 and UFLA SIL52 strains promoted an increase in aerobic stability of silage.     

重组植物乳杆菌生物学特性比较研究

试验对植物乳杆菌(Lactobacillus plantarum,Lp)出发菌与携带绿色荧光蛋白(green fluorescent protein,GFP)标记基因的重组植物乳杆菌(Lp-GFP)生物学特性进行了详细的比较研究,以便于深入开展益生菌植物乳杆菌在动物肠道内分布和定植规律的研究。结果显示,Lp和Lp-GFP生长曲线的变化趋势基本相同,其中Lp-GFP的生长速度略有滞后;Lp和Lp-GFP最适培养温度均为37 ℃,最适初始pH均为6.5;Lp和Lp-GFP耐高温、耐人工胃液、耐人工肠液和耐胆盐的特性趋势一致;Lp和Lp-GFP的抑菌特性无显著差异(P>0.05);除了红霉素,Lp和Lp-GFP对其他药物的敏感性基本没有发生变化。综上所述,植物乳杆菌的重组对其生物学特性没有显著的影响,这为重组标记菌株的后续培养和机理研究奠定了重要基础。     

植物乳杆菌P158的生长曲线及其细菌素的特性

对源于醪糟的植物乳杆菌P158生长曲线分析、药敏实验检测及其细菌素特性的初步研究。结果表明：植物乳杆菌P158培养36h时生物量达到最大值,约为1011 cfu/ml,且培养40h的发酵上清液对藤黄微球菌和铜绿假单胞杆菌的抑菌效果最好,细菌素效价约为1145IU/ml,而发酵上清液pH值降至最低（pH3.53）,显示了菌株生长与细菌素的形成密切相关。该菌株对多粘菌素B、多数氨基糖苷类耐药,对青霉素类、四环素类、大环内酯类等药物敏感。细菌素对酸、热稳定,对木瓜蛋白酶、胃蛋白酶、胰蛋白酶、蛋白酶K敏感;对多数的革兰氏阴性菌和革兰氏阳性菌及一些真菌具有较强抑制作用,却对近源乳酸菌几乎无抑制效果。该细菌素可望作为食品生物防腐剂。     

微生物发酵及菜粕混合发酵对甘蔗尾叶营养价值的影响

为了有效合理地开发甘蔗尾叶在饲料行业中的应用价值,并在生产中推广应用。本试验以新鲜的甘蔗尾叶和菜粕为原料,进行微生物发酵及菜粕混合发酵对甘蔗尾叶营养价值的影响研究。试验分 6个组：对照组、试验Ⅰ组、试验Ⅱ组、试验Ⅲ组、试验IV组、试验V组,每组设 3 个重复。40d 后测定各组青贮常规营养成分含量和现场感官评定。结果表明：（1）试验Ⅱ、Ⅲ、IV、V粗蛋白均高于对照组和试验Ⅰ组。试验Ⅱ组、Ⅲ组较对照组分别提高75.04%、153.95%,试验Ⅲ组较试验1组提高95.70%,试验IV组较试验Ⅱ组提高78.65%,试验IV组较试验Ⅲ组提高33.94%。（2）粗纤维、酸性洗涤纤维、中性洗涤纤维Ⅲ组较对照组分别降低12.97%,11.66%,11.49%。（3）生物制剂组的pH 值在3.85-3.97,自然发酵组pH 值在3.95-4.12。（4）添加发酵菌剂和不同梯度的菜粕,都不同程度的提高甘蔗尾叶的 CP,降低甘蔗尾叶的 CF。其中以试验Ⅲ组效果最好,甘蔗尾叶发酵后粗蛋白提高了153.95%,粗纤维降低了12.97%。     

以乳酸杆菌为载体的猪传染性胃肠炎病毒S基因A、D抗原位点DNA疫苗的构建

应用pRc/CMV2真核质粒骨架,先后插入猪传染性胃肠炎病毒(TGEV)S基因AD抗原位点片段和乳酸杆菌复制子基因Rep.8014,构建了pRc/CMV2-SAD-Rep.8014真核表达穿梭质粒。将其转染PK-15细胞,RT-PCR检测到S基因A D抗原位点的mRNA表达。随后,将pRc/CMV2-SAD-Rep.8014质粒电转化猪源的Lactobacillus acidophilusSW1株,成功构建了以乳酸杆菌为载体的带有TGEV S基因AD抗原位点的DNA疫苗株L.acidophilusTGES-1。本研究结合了乳酸杆菌胃肠道常在共生菌的优势和TGE呈现肠道局部发病的特点,达到益生性与免疫预防的双重功效,对乳酸杆菌口服DNA疫苗进行了有益的探索。     

瑞士乳酸杆菌HZ521株的分子鉴定及其部分生物学特性研究

根据GenBank中乳酸杆菌16 S～23 S rRNA基因间沉默区序列设计引物,进一步鏊定猪源乳酸杆菌分离株HZ521;并通过体外试验,以嗜酸乳酸杆菌ATCC 4356为参考菌株,探讨HZ521株的益生素作用.部分16 S～23 S rRNA基因序列同源性分析结果表明,该分离株属于瑞士乳酸杆菌.HZ521株具有较强的产酸能力,能耐受强酸,对Hela细胞的黏附率为87.2%,显著高于ATCC 4356株(P<0.01).表明瑞士乳酸杆菌HZ521株具有益生素特性,可作为肠道益生素的候选菌株.     

副干酪乳杆菌HD1.7遗传转化体系的初步建立

优化副干酪乳杆菌(Lactobacillus paracasei)HD1.7的电转化条件,为构建L .paracasei HD1.7prcR基因敲除突变株奠定基础。通过电转化方法将自杀质粒pUC18-prcR-tet导入L. paracasei HD1.7内,对影响L .paracasei HD1.7电转化的几个主要因素进行了初步研究。当L. paracasei HD1.7生长8 h时,加入青霉素使终浓度为12.5 μg/mL,再培养1 h。经AEB1电转缓冲液(蔗糖浓度为0.3 mol/L)处理后,在1.8?2.0 KV电压下完成电击,并迅速加入MRS培养基中,复苏5 h,成功地将自杀质粒pUC18-prcR-tet转入L. paracasei HD1.7内,并发生了部分同源重组。本研究得到了副干酪乳杆菌（Lactobacillus paracasei）HD1.7较优电转化条件,为继续研究L.paracasei HD1.7的群体感应调控因子prcR奠定了基础。