秀丽隐杆线虫益生菌动物模型的建立及应用研究
|
摘要
由于畜禽肠道菌群复杂、无菌动物的费用昂贵和高技术要求、限制了乳酸菌等益生菌作用效果及机理的研究,致使乳酸菌等益生菌制剂(微生态制剂)的质量难以控制、合理的开发应用受到了很大的影响。因此,本研究选用肠道结构简单、细胞内部基因调控研究比较深入、在病原菌激活机制、感染过程及免疫反应等诸多方面和哺乳动物存在高度相似性的秀丽隐杆线虫(Caenorhabditis elegans)作为实验动物,以沙门氏菌作为病原指示菌,以乳酸菌为研究益生菌,建立了线虫益生菌实验动物模型,并应用于乳酸菌的研究。
本研究首先采用改良线虫液体培养法,成功孵化了秀丽隐杆线虫突变株SS104;在确定了沙门氏菌的最适攻毒量及乳酸菌的最佳保护剂量基础上,建立了秀丽隐杆线虫益生菌动物模型;以线虫的存活率为判定标准,从17株乳酸菌中筛选出了抑菌效果较好的S64株和较差的CL11株菌,并通过对线虫存活率及肠道菌数量检测分析,比较了两株的活菌、灭活菌和培养上清液作用效果的差异;探讨了线虫动物模型筛选的乳酸菌对断奶仔猪和雏鸡生产性能、腹泻率、免疫性能及肠道菌群的影响,从而验证线虫动物模型筛选的乳酸菌对线虫的作用效果与对畜禽作用效果的相似性。
秀丽隐杆线虫益生菌动物模型的成功建立为乳酸菌等益生菌的筛选、作用效果及机理研究提供了技术支撑,为微生态制剂的质量控制、开发利用提供了保障。本研究内容共分四部分。
试验一秀丽隐杆线虫益生菌动物模型的建立
应用改良线虫液体培养法,将同期化的秀丽隐杆线虫(SS104)L4期幼虫孵化、分装于24孔培养板,再将24孔随机分为不同处理组,每组4孔,每孔15-20条线虫;然后以GFP标记沙门氏菌(DT104)作为线虫的致病菌、以乳酸菌(S64株)作为线虫的有益保护菌,探索建立线虫动物模型的条件及效果。结果显示,改良线虫液体培养法操作简单、条件易控制;沙门氏菌浓度在107cfu/ml、108cfu/ml和109cfu/ml均能致死线虫,但以109cfu/ml时致死速度快、致死率最高,于第6d,所有线虫全部死亡(致死率100%);第10d时,浓度为108cfu/ml乳酸菌对线虫存活率达到87.9%,较对照组提高5.1%,而浓度为109cfu/ml的乳酸菌对线虫的存活率仅为63.2%,较对照组下降19.2%;保护实验第9d,攻毒前加入乳酸菌和攻毒后加入乳酸菌对线虫的保护率分别为44.9%和58.1%;说明乳酸菌能够有效保护线虫免受GFP标记沙门氏菌的感染。结果表明,本研究成功建立了秀丽隐杆线虫益生菌动物模型。
试验二利用线虫动物模型对乳酸菌的筛选研究
首先应用16S rRNA序列分析法对17株乳酸菌株进行了鉴定,在此基础上,以沙门氏菌DT104为病原指示菌,进行了乳酸菌体外抑菌和线虫体内抑菌实验。结果显示,17株乳酸菌株对沙门氏菌的抑菌圈直径在19mm-25mm之间,培养液上清也均能有效抑制沙门氏菌的生长(OD600值在0.058-0.074之间);17株乳酸菌对线虫的保护率为-2.6%-56%不等,其中CL9、L3、LB1、LB4、S64株的保护率分别为59%、53.2%,51.1%、58.7%和55.7%(P﹤0.05),而CL10、CL11、CL12、LB2四株菌株保护率仅为-3.2%、-6.2%,2.2%和2.9%(P﹤0.01)。该研究结果表明,秀丽隐杆线虫动物模型对乳酸菌的筛选效果优于体外抑菌实验筛选法。
试验三线虫益生菌动物模型的应用研究
——筛选乳酸菌S64株和CL11株作用效果比较研究
通过对线虫存活率和肠道菌数的检测,对线虫动物模型筛选的有效乳酸菌株S64和无效菌株CL11的作用效果进行了比较试验,结果显示:(1)S64乳酸菌株组和CL11株组线虫存活率分别为72.2%和25.7%,对线虫的保护率分别为51.7%和2.2%,差异显著(P﹤0.05);灭活S64乳酸菌株组和CL11株组线虫存活率分别为57.4%和25.8%,对线虫的保护率分别为33.9和2.3%,差异显著(P﹤0.05); S64和CL11株乳酸菌培养上清液组线虫存活率分别为36.7%和35.8%,对线虫的保护率分别为13.2%和12.3%,差异不显著(P>0.05)。(2)S64株活菌组线虫肠道内乳酸菌数高于活菌CL11株组,线虫肠道内沙门氏菌数低于活菌CL11株组和对照组,差异显著(P﹤0.05);灭活菌S64株组线虫肠道内沙门氏菌数低于灭活菌CL11株组和对照组,差异显著(P﹤0.05);S64和CL11株培养上清液组线虫肠道内沙门氏菌数均与对照组差异不显著(P>0.05);虫体荧光结果显示,S64株乳酸菌活菌和灭活菌均可有效抑制沙门氏菌在线虫肠道内的生长。
试验四线虫益生菌动物模型的应用研究
——筛选乳酸菌对畜禽作用效果的研究
为探讨线虫动物模型筛选的乳酸菌对畜禽作用的有效性,本研究以断奶仔猪和雏鸡作为实验动物进行了饲喂实验。
(1)选用大三元断奶仔猪96头,随机分为8组,分别饲喂乳酸菌S64、LB1、LB2、ALB2、L3和CL11株发酵液体饲料,对照1组饲喂基础颗粒料,对照2组饲喂湿拌料,实验至21d经检测料肉比、腹泻率和肠道菌数,结果显示,乳酸菌ALB2、S64、LB1和L3株发酵饲料组料肉比较对照2组分别下降了49%、47%、36%和24% (P﹤0.05),腹泻指数较对照2组分别下降了50.89%、64.29%、93.51%和32.5%(P﹤0.05),腹泻率较对照2组分别下降了50.01%、81.82%、63.65%和23.72%;CL11和LB2株组料肉比分别提高了4%和12%(P<0.05),腹泻指数分别提高了15%和27.27%(P<0.05),腹泻率分别提高了21.78%和29.95%(P<0.05);对照1组较对照2组料肉比、腹泻指数和腹泻率分别下降了15%、7.14%和16.63%(P>0.05);LB1株发酵组较对照组和LB2发酵组肠道乳酸菌数量显著增加( P < 0. 05),肠杆菌数量显著减少( P < 0. 05)。
(2)选用200羽1日龄海兰褐蛋公鸡,随机分成4组,第1、2、3组分别为乳酸菌S64、L3和CL11株饮水组,第4组为空白对照组,对各组鸡抗体效价、免疫器官指数和肠道菌群数进行测定,结果显示,35日龄时,第1、2组血凝抑制效价显著高于对照组和第3组(P﹤0.05),第3组较对照组差异不显著(P>0.05);第1、2组鸡肠道乳酸菌数量较第3组和对照组增加显著(P﹤0.05),肠杆菌数量显著减少(P﹤0.05);第1组胸腺、法氏囊和脾脏指数均显著高于对照组(P﹤0.05),第3组较对照组差异不显著(P>0.05)。ERIC-PCR结果显示,S64株组肠道菌群指纹条带较L3株和CL11株组明显增多,表明S64株更能有效调整鸡肠道菌群平衡。
本试验结果显示,线虫动物模型筛选的有效乳酸菌(S64和LB1株)能够明显提高断奶仔猪的生产性能和雏鸡的免疫等功能,表明线虫动物模型筛选的乳酸菌对线虫的作用效果与对畜禽作用效果是相似的。
The study concerning effection and mechanism of Lactobacillus has been restricted by complicated intestinal microflora of mammal animals and expensive cost on germ-free animals. Caenorhabditis elegans has been used as a laboratory animal model for the fact that there is a broad overlap between mammals and C. elegans about infection mechanism, immune reaction and so on. In this study, Salmonella used for pathogen and Lactobacillus used for probiotic. Caenorhabditis elegans has been used for estabilished a laboratory animal model for Conduct research on probiotic.
The C.elegans strain SS104 has been hatched firstly by fluild incubation method, and proper dose of S. typhimurium and Llactobacillus has been confirmed. We have evaluated 17 Lactobacillus isolates from chicken and pig intestine for their ability to protect the worms from Salmonella-induced death in vitro; When tested on C. elegans infected with S. typhimurium DT104, the isolates were, however, differentiated in their ability to protect the nematode from Salmonella-induced death. Two Lactobacillus isolates S64 and CL11 (S64 with full, but CL11 with no protection) were further studied. The study on difference culture fractions (the extracellular culture fluid, live and heat-killed cells) of isolates CL11 and S64 has been designed; We observed the effects of Lactobacillus isolates on performance , diarrhea incidence, immune activities and intestinal microflora of wean piglets and layer chicken in order to evaluate similarity between livestock and C.elegans.
The study consists of four parts:
1 Establishment of the assay with C. elegans used as an animal moder for study probiotics
Synchronizated L4 larvae has been hatched, resuspension in 24-well titre plates with each well containing 2 ml of S medium and 10– 15 worms, and incubated at 25°C during the assay. Each treatment had four wells. To explore the experimental condition of C.elegans used as animal model for evaluate the effect of a Lactobacillus isolate on protecting the nematode from Salmonella-induced death. The result showed : the fluid incubation method of C.elegans was more easy to operate; C.elegans was infected and died with S. typhimurium at 10~7cfu/ml、10~8cfu/ml and 10~9cfu/ml, however, all the worms died in 6~(th) days at 10~9cfu/ml;The survival rate (87.9%) of Lactobacillus-treated worms at 108cfu/ml was higher than that of C. elegans fed E. coli OP50 (82.8%) only on 10~(th) day, but survival rate of C.elegans was 63.2% at 10~9cfu/ml; The protective rate of Lactobacillus for C.elegans was 50.3% . meanwhile, the green fluorescence was emitted by the GFP- Salmonella of Lactobacillus-treated worms was more diminish than positive control in the digestive tract of the worms.
2 Presreening of Lactobacillus isolates using a laboratory animal model of Caenorhabditis elegans
17 Lactobacillus isolates has been confirmed by 16S rRNA analysis method. The spot-on-the-lawn assay demonstrated that all 17 tested Lactobacillus isolates were inhibitive to S. typhimurium . The size of growth inhibition haloes ranged from 19 to 25 mm in diameter and the extracellular culture fluid (ECF) recovered from tested Lactobacillus cultures all had a similar degree of inhibition towards S. typhimurium; When tested on C. elegans infected with S. typhimurium, the isolates were, however, differentiated in their ability to protect the nematode from Salmonella-induced death. On the last day (9~(th)) of the assays, the protective rate of Lactobacillus ranged from -2.6% to -56%. Among them, five isolates (CL9, L3, LB1, LB4, and S64) provided maximum protection(59%、53.2%,51.1%、58.7% and 55.7% respectively ) (P﹤0.05) to the infected worms that had a similar level of longevity compared to the negative control. The protective rate of two isolates CL10 and CL11 was -3.2% and -6.2%(P﹤0.01) and demonstrated negative effective on C.elegans . Our observations suggest that C. elegans can be used as an animal model for pre-screening probiotics.
3 Application research on Caenorhabditis elegans as an animal model for study probaotics
——Effection of Lactobacillus isolates S64 and CL11 screened by C.elegans To explore possible mechanisms underlying the protection offered by Lactobacillus different culture fractions, including the ECF, live and heat-killed cells, of Lactobacillus isolate (S64 and CL11) were investigated for their effect on the survival rate C. elegans infected with Salmonella and the colonization of the pathogen in the digestive tract of the nematode. The result demonstrated: (1)On the 9th day in the assay, the survival rate of C.elegans treated with lactobacillus isolate S64 and CL11 live cells was 72.2% and 25.7%(P<0.05), the protetive rate was 51.7% and 2.2% respectively; The survival rate of C.elegans treated with lactobacillus isolate S64 and CL11 heat-killed cells was 57.4% and 25.8%(P<0.05),the protetive rate was 33.9% and 2.3% respectively; The survival rate of C.elegans treated with ECF of S64 and CL11 was 36.7% and 35.8%(P>0.05),the protetive rate was 13.2% and 12.3% respectively. (2) On the 8~(th) day in the assay , the amount of Lactobacillus in the digestive tract treated with live Lactobacillus isolate S64 was increase distinctly compare to that of treated with live isolate CL11(P<0.05); The amount of Salmonella treated with live and heat-killed isolate S64 was significantly reduced than that of treated with isolate CL11 and positive control(P<0.05); The amount of Salmonella treated with ECF of isolate S64 and CL11 was at similar degree(P>0.05); Meanwhile, the green fluorescence was emitted by the GFP-salmonella of live isolate S64-treated worms was significantly diminished than positive control and isolate CL11-treated worms.
4 Application research on Caenorhabditis elegans as an animal model for study probaotics
——Effection of Lactobacillus isolates screened by C.elegans on livestock
To explore the effction of effection of Lactobacillus isolates screened by C.elegans on livestock , the feeding Trial had beedn designed using wean piglet and chicken as animal modle.
(1) 96 weaned piglets were divided into 8 groups randomly and fed them with ferment liquid feed (FLF) which fermented by Lactobacillus isolates S64、LB1、LB2、ALB2、L3、CL11、dry feed (control 1) and liquid feed (control 2, no lactobacillus) respectively. The result showed: The ratio of feed to meat was reduced 49%、47%、36% and 24%(P<0.05), diarrhea index was reduced 50.89%、64.29%、93.51% and 32.5%(P<0.05),Diarrhea incidence was dropped by 50.01%、81.82%、63.65% and 23.72% (P<0.05), when fed them with FLF fermented by Lactobacillus isolates ALB2、S64、LB1 and L3 respectively; The ratio of feed to meat was increased 4% and12% (P < 0.05),diarrhea index was increased 15% and 27.27%(P<0.05),diarrhea incidence increased 21.78% and 29.95%(P<0.05), when fed them with FLF fermented by Lactobacillus isolates CL11 and LB2 respectively; Lactobacillus isolates LB1 was beneficial to the balance of gut flora than Lactobacillus isolates LB2.
(2)A total of 200 newly hatched Layer Chicken were randomly allotted into 4 dietary treatments fed them with Lactobacillus isolates S64, L3 and CL11. On 35d in the assay, the hemagglutination inhibition antibody potency of Newcastle disease vaccine immunized of Lactobaccilus isolates S64 and L3 increased distinctly compare to negative control and isolate CL11(P<0.05); The amounts of Lactobaccilus in the Cecal contents significantly increased (P<0.05), and the amounts of E.coli and and Salmonella significantly reduced (P<0.05) when fed them with Lactobacillus isolate S64 and L3. Immune organ index of layer chicken when fed them with Lactobacillus isolate S64 improved significantly(P<0.05). The data of the treatment fed with Lactobacillus isolate CL11 was similar to negative control(P>0.05).The intestinal flora of the chickens fed Lactobacillus isolates S64 was changed clearly using ERIC-PCR based fingerprints
These data demonstrated Lactobacillus isolates S64 and LB1 which screened by C.elegans was beneficial to production performance of wean piglets and to immunologic function of chickens. The result showed that the effection of Lactobacillus is similar between C.elegans and livestocks, and C.elegans can be used for study probiotic as a laboratory animal model.
本研究首先采用改良线虫液体培养法,成功孵化了秀丽隐杆线虫突变株SS104;在确定了沙门氏菌的最适攻毒量及乳酸菌的最佳保护剂量基础上,建立了秀丽隐杆线虫益生菌动物模型;以线虫的存活率为判定标准,从17株乳酸菌中筛选出了抑菌效果较好的S64株和较差的CL11株菌,并通过对线虫存活率及肠道菌数量检测分析,比较了两株的活菌、灭活菌和培养上清液作用效果的差异;探讨了线虫动物模型筛选的乳酸菌对断奶仔猪和雏鸡生产性能、腹泻率、免疫性能及肠道菌群的影响,从而验证线虫动物模型筛选的乳酸菌对线虫的作用效果与对畜禽作用效果的相似性。
秀丽隐杆线虫益生菌动物模型的成功建立为乳酸菌等益生菌的筛选、作用效果及机理研究提供了技术支撑,为微生态制剂的质量控制、开发利用提供了保障。本研究内容共分四部分。
试验一秀丽隐杆线虫益生菌动物模型的建立
应用改良线虫液体培养法,将同期化的秀丽隐杆线虫(SS104)L4期幼虫孵化、分装于24孔培养板,再将24孔随机分为不同处理组,每组4孔,每孔15-20条线虫;然后以GFP标记沙门氏菌(DT104)作为线虫的致病菌、以乳酸菌(S64株)作为线虫的有益保护菌,探索建立线虫动物模型的条件及效果。结果显示,改良线虫液体培养法操作简单、条件易控制;沙门氏菌浓度在107cfu/ml、108cfu/ml和109cfu/ml均能致死线虫,但以109cfu/ml时致死速度快、致死率最高,于第6d,所有线虫全部死亡(致死率100%);第10d时,浓度为108cfu/ml乳酸菌对线虫存活率达到87.9%,较对照组提高5.1%,而浓度为109cfu/ml的乳酸菌对线虫的存活率仅为63.2%,较对照组下降19.2%;保护实验第9d,攻毒前加入乳酸菌和攻毒后加入乳酸菌对线虫的保护率分别为44.9%和58.1%;说明乳酸菌能够有效保护线虫免受GFP标记沙门氏菌的感染。结果表明,本研究成功建立了秀丽隐杆线虫益生菌动物模型。
试验二利用线虫动物模型对乳酸菌的筛选研究
首先应用16S rRNA序列分析法对17株乳酸菌株进行了鉴定,在此基础上,以沙门氏菌DT104为病原指示菌,进行了乳酸菌体外抑菌和线虫体内抑菌实验。结果显示,17株乳酸菌株对沙门氏菌的抑菌圈直径在19mm-25mm之间,培养液上清也均能有效抑制沙门氏菌的生长(OD600值在0.058-0.074之间);17株乳酸菌对线虫的保护率为-2.6%-56%不等,其中CL9、L3、LB1、LB4、S64株的保护率分别为59%、53.2%,51.1%、58.7%和55.7%(P﹤0.05),而CL10、CL11、CL12、LB2四株菌株保护率仅为-3.2%、-6.2%,2.2%和2.9%(P﹤0.01)。该研究结果表明,秀丽隐杆线虫动物模型对乳酸菌的筛选效果优于体外抑菌实验筛选法。
试验三线虫益生菌动物模型的应用研究
——筛选乳酸菌S64株和CL11株作用效果比较研究
通过对线虫存活率和肠道菌数的检测,对线虫动物模型筛选的有效乳酸菌株S64和无效菌株CL11的作用效果进行了比较试验,结果显示:(1)S64乳酸菌株组和CL11株组线虫存活率分别为72.2%和25.7%,对线虫的保护率分别为51.7%和2.2%,差异显著(P﹤0.05);灭活S64乳酸菌株组和CL11株组线虫存活率分别为57.4%和25.8%,对线虫的保护率分别为33.9和2.3%,差异显著(P﹤0.05); S64和CL11株乳酸菌培养上清液组线虫存活率分别为36.7%和35.8%,对线虫的保护率分别为13.2%和12.3%,差异不显著(P>0.05)。(2)S64株活菌组线虫肠道内乳酸菌数高于活菌CL11株组,线虫肠道内沙门氏菌数低于活菌CL11株组和对照组,差异显著(P﹤0.05);灭活菌S64株组线虫肠道内沙门氏菌数低于灭活菌CL11株组和对照组,差异显著(P﹤0.05);S64和CL11株培养上清液组线虫肠道内沙门氏菌数均与对照组差异不显著(P>0.05);虫体荧光结果显示,S64株乳酸菌活菌和灭活菌均可有效抑制沙门氏菌在线虫肠道内的生长。
试验四线虫益生菌动物模型的应用研究
——筛选乳酸菌对畜禽作用效果的研究
为探讨线虫动物模型筛选的乳酸菌对畜禽作用的有效性,本研究以断奶仔猪和雏鸡作为实验动物进行了饲喂实验。
(1)选用大三元断奶仔猪96头,随机分为8组,分别饲喂乳酸菌S64、LB1、LB2、ALB2、L3和CL11株发酵液体饲料,对照1组饲喂基础颗粒料,对照2组饲喂湿拌料,实验至21d经检测料肉比、腹泻率和肠道菌数,结果显示,乳酸菌ALB2、S64、LB1和L3株发酵饲料组料肉比较对照2组分别下降了49%、47%、36%和24% (P﹤0.05),腹泻指数较对照2组分别下降了50.89%、64.29%、93.51%和32.5%(P﹤0.05),腹泻率较对照2组分别下降了50.01%、81.82%、63.65%和23.72%;CL11和LB2株组料肉比分别提高了4%和12%(P<0.05),腹泻指数分别提高了15%和27.27%(P<0.05),腹泻率分别提高了21.78%和29.95%(P<0.05);对照1组较对照2组料肉比、腹泻指数和腹泻率分别下降了15%、7.14%和16.63%(P>0.05);LB1株发酵组较对照组和LB2发酵组肠道乳酸菌数量显著增加( P < 0. 05),肠杆菌数量显著减少( P < 0. 05)。
(2)选用200羽1日龄海兰褐蛋公鸡,随机分成4组,第1、2、3组分别为乳酸菌S64、L3和CL11株饮水组,第4组为空白对照组,对各组鸡抗体效价、免疫器官指数和肠道菌群数进行测定,结果显示,35日龄时,第1、2组血凝抑制效价显著高于对照组和第3组(P﹤0.05),第3组较对照组差异不显著(P>0.05);第1、2组鸡肠道乳酸菌数量较第3组和对照组增加显著(P﹤0.05),肠杆菌数量显著减少(P﹤0.05);第1组胸腺、法氏囊和脾脏指数均显著高于对照组(P﹤0.05),第3组较对照组差异不显著(P>0.05)。ERIC-PCR结果显示,S64株组肠道菌群指纹条带较L3株和CL11株组明显增多,表明S64株更能有效调整鸡肠道菌群平衡。
本试验结果显示,线虫动物模型筛选的有效乳酸菌(S64和LB1株)能够明显提高断奶仔猪的生产性能和雏鸡的免疫等功能,表明线虫动物模型筛选的乳酸菌对线虫的作用效果与对畜禽作用效果是相似的。
The study concerning effection and mechanism of Lactobacillus has been restricted by complicated intestinal microflora of mammal animals and expensive cost on germ-free animals. Caenorhabditis elegans has been used as a laboratory animal model for the fact that there is a broad overlap between mammals and C. elegans about infection mechanism, immune reaction and so on. In this study, Salmonella used for pathogen and Lactobacillus used for probiotic. Caenorhabditis elegans has been used for estabilished a laboratory animal model for Conduct research on probiotic.
The C.elegans strain SS104 has been hatched firstly by fluild incubation method, and proper dose of S. typhimurium and Llactobacillus has been confirmed. We have evaluated 17 Lactobacillus isolates from chicken and pig intestine for their ability to protect the worms from Salmonella-induced death in vitro; When tested on C. elegans infected with S. typhimurium DT104, the isolates were, however, differentiated in their ability to protect the nematode from Salmonella-induced death. Two Lactobacillus isolates S64 and CL11 (S64 with full, but CL11 with no protection) were further studied. The study on difference culture fractions (the extracellular culture fluid, live and heat-killed cells) of isolates CL11 and S64 has been designed; We observed the effects of Lactobacillus isolates on performance , diarrhea incidence, immune activities and intestinal microflora of wean piglets and layer chicken in order to evaluate similarity between livestock and C.elegans.
The study consists of four parts:
1 Establishment of the assay with C. elegans used as an animal moder for study probiotics
Synchronizated L4 larvae has been hatched, resuspension in 24-well titre plates with each well containing 2 ml of S medium and 10– 15 worms, and incubated at 25°C during the assay. Each treatment had four wells. To explore the experimental condition of C.elegans used as animal model for evaluate the effect of a Lactobacillus isolate on protecting the nematode from Salmonella-induced death. The result showed : the fluid incubation method of C.elegans was more easy to operate; C.elegans was infected and died with S. typhimurium at 10~7cfu/ml、10~8cfu/ml and 10~9cfu/ml, however, all the worms died in 6~(th) days at 10~9cfu/ml;The survival rate (87.9%) of Lactobacillus-treated worms at 108cfu/ml was higher than that of C. elegans fed E. coli OP50 (82.8%) only on 10~(th) day, but survival rate of C.elegans was 63.2% at 10~9cfu/ml; The protective rate of Lactobacillus for C.elegans was 50.3% . meanwhile, the green fluorescence was emitted by the GFP- Salmonella of Lactobacillus-treated worms was more diminish than positive control in the digestive tract of the worms.
2 Presreening of Lactobacillus isolates using a laboratory animal model of Caenorhabditis elegans
17 Lactobacillus isolates has been confirmed by 16S rRNA analysis method. The spot-on-the-lawn assay demonstrated that all 17 tested Lactobacillus isolates were inhibitive to S. typhimurium . The size of growth inhibition haloes ranged from 19 to 25 mm in diameter and the extracellular culture fluid (ECF) recovered from tested Lactobacillus cultures all had a similar degree of inhibition towards S. typhimurium; When tested on C. elegans infected with S. typhimurium, the isolates were, however, differentiated in their ability to protect the nematode from Salmonella-induced death. On the last day (9~(th)) of the assays, the protective rate of Lactobacillus ranged from -2.6% to -56%. Among them, five isolates (CL9, L3, LB1, LB4, and S64) provided maximum protection(59%、53.2%,51.1%、58.7% and 55.7% respectively ) (P﹤0.05) to the infected worms that had a similar level of longevity compared to the negative control. The protective rate of two isolates CL10 and CL11 was -3.2% and -6.2%(P﹤0.01) and demonstrated negative effective on C.elegans . Our observations suggest that C. elegans can be used as an animal model for pre-screening probiotics.
3 Application research on Caenorhabditis elegans as an animal model for study probaotics
——Effection of Lactobacillus isolates S64 and CL11 screened by C.elegans To explore possible mechanisms underlying the protection offered by Lactobacillus different culture fractions, including the ECF, live and heat-killed cells, of Lactobacillus isolate (S64 and CL11) were investigated for their effect on the survival rate C. elegans infected with Salmonella and the colonization of the pathogen in the digestive tract of the nematode. The result demonstrated: (1)On the 9th day in the assay, the survival rate of C.elegans treated with lactobacillus isolate S64 and CL11 live cells was 72.2% and 25.7%(P<0.05), the protetive rate was 51.7% and 2.2% respectively; The survival rate of C.elegans treated with lactobacillus isolate S64 and CL11 heat-killed cells was 57.4% and 25.8%(P<0.05),the protetive rate was 33.9% and 2.3% respectively; The survival rate of C.elegans treated with ECF of S64 and CL11 was 36.7% and 35.8%(P>0.05),the protetive rate was 13.2% and 12.3% respectively. (2) On the 8~(th) day in the assay , the amount of Lactobacillus in the digestive tract treated with live Lactobacillus isolate S64 was increase distinctly compare to that of treated with live isolate CL11(P<0.05); The amount of Salmonella treated with live and heat-killed isolate S64 was significantly reduced than that of treated with isolate CL11 and positive control(P<0.05); The amount of Salmonella treated with ECF of isolate S64 and CL11 was at similar degree(P>0.05); Meanwhile, the green fluorescence was emitted by the GFP-salmonella of live isolate S64-treated worms was significantly diminished than positive control and isolate CL11-treated worms.
4 Application research on Caenorhabditis elegans as an animal model for study probaotics
——Effection of Lactobacillus isolates screened by C.elegans on livestock
To explore the effction of effection of Lactobacillus isolates screened by C.elegans on livestock , the feeding Trial had beedn designed using wean piglet and chicken as animal modle.
(1) 96 weaned piglets were divided into 8 groups randomly and fed them with ferment liquid feed (FLF) which fermented by Lactobacillus isolates S64、LB1、LB2、ALB2、L3、CL11、dry feed (control 1) and liquid feed (control 2, no lactobacillus) respectively. The result showed: The ratio of feed to meat was reduced 49%、47%、36% and 24%(P<0.05), diarrhea index was reduced 50.89%、64.29%、93.51% and 32.5%(P<0.05),Diarrhea incidence was dropped by 50.01%、81.82%、63.65% and 23.72% (P<0.05), when fed them with FLF fermented by Lactobacillus isolates ALB2、S64、LB1 and L3 respectively; The ratio of feed to meat was increased 4% and12% (P < 0.05),diarrhea index was increased 15% and 27.27%(P<0.05),diarrhea incidence increased 21.78% and 29.95%(P<0.05), when fed them with FLF fermented by Lactobacillus isolates CL11 and LB2 respectively; Lactobacillus isolates LB1 was beneficial to the balance of gut flora than Lactobacillus isolates LB2.
(2)A total of 200 newly hatched Layer Chicken were randomly allotted into 4 dietary treatments fed them with Lactobacillus isolates S64, L3 and CL11. On 35d in the assay, the hemagglutination inhibition antibody potency of Newcastle disease vaccine immunized of Lactobaccilus isolates S64 and L3 increased distinctly compare to negative control and isolate CL11(P<0.05); The amounts of Lactobaccilus in the Cecal contents significantly increased (P<0.05), and the amounts of E.coli and and Salmonella significantly reduced (P<0.05) when fed them with Lactobacillus isolate S64 and L3. Immune organ index of layer chicken when fed them with Lactobacillus isolate S64 improved significantly(P<0.05). The data of the treatment fed with Lactobacillus isolate CL11 was similar to negative control(P>0.05).The intestinal flora of the chickens fed Lactobacillus isolates S64 was changed clearly using ERIC-PCR based fingerprints
These data demonstrated Lactobacillus isolates S64 and LB1 which screened by C.elegans was beneficial to production performance of wean piglets and to immunologic function of chickens. The result showed that the effection of Lactobacillus is similar between C.elegans and livestocks, and C.elegans can be used for study probiotic as a laboratory animal model.
引文
陈德富,陈喜文.现代分子生物学实验原理与技术[M].北京:科学出版社,2006,2
陈琼,孔繁德,彭海滨,等.51株沙门氏菌的耐药性分析[J].畜牧与兽医,2008,40(8):77-78
陈润生.猪生产学[M] .北京:中国农业出版社,1995
陈杖榴,刘健华.食品动物源细菌耐药性与公共卫生[J].兽医导刊, 2007,9:45-47
丁德忠,庞小燕,华修国,等. SPF仔猪肠道菌群结构的分子分析[J].上海交通大学学报(农业科学版),2005,25(3):256-260
房春红,刘杰,许修宏.乳酸菌素的研究现状和发展趋势[J].中国乳品工业,2006,34(2):53-55
高巍,孟庆翔,肖训军,等.生长猪胃肠道乳酸菌、双歧杆菌和大肠埃希菌的数量和分布规律[J] .中国农业大学学报,2001,6(5):81-86
高巍,孟庆翔.生长育肥猪胃肠道正常厌氧菌群的数量和分区[J].中国农业大学学报,2000,5(5):88-93
顾瑞霞,伊萌.乳酸菌抗肿瘤特性的研究进展.中国微生态学杂志[J].1999, 11 (4): 253-255
郭兴华.益生菌基础与应用[M].北京:北京科学技术出版社,2001,10
何昭阳,王增辉,吴延春.雏鸡消化道主要正常茵群定植规律的研究[J].畜牧兽医学报,2000,31(1):41-48
何昭阳,徐凤宇,管清华,等.犊牛腹泻与肠道菌群的变化[J].中国预防兽医学报,2000,22(5):345-346
何谦,郭进超,李岩,等.发酵液体饲料在断奶仔猪的应用进展[J].养猪,2007,5:5-7
胡东良.乳酸菌的抗肿瘤、抗变异原及免疫增强作用.中国乳品工业[J],1997,25 (6): 11-14
黄怡,王士长,崔艳红,等.多菌种复合益生素对三黄鸡的生产性能及肠道主要菌群的影响[J].安徽农业科学,2006,34(3):485-486
贾熙华,曹诚.秀丽隐杆线虫在医药学领域的应用和进展[J].药学学报, 2009, 44 (7): 687?694
李国平,周伦江,邵良平,等.日粮中添加复合菌制剂对雏鸡盲肠大肠杆菌数、肠道pH 值、血液SOD和GSH- Px影响的研究[J].中国兽医杂志, 1999, 25(8) : 7-8
李路胜,周响艳,冯定远.乳酸菌对麻羽肉鸡生长和免疫机能的影响研究[J].饲料工业,2006,27 (20):39 - 41
李晓晖.微生态饲料添加剂研究进展[J].饲料博览,2002,3:40～41
林雪彦,王中华,李福昌,等.乳酸杆菌黏附鸡消化道上皮细胞超微结构的观察[J].中国兽医学报,2008,28(12):1367-1373
李晓霞.新城疫病毒气溶胶发生与传染机制及其在生产鸡舍环境中的监测(D).山东农业大学,2009
李犹平.采用ERIC-PCR技术研究鸡白痢、新陈疫感染雏鸡及健康鸡肠道菌群结构(D).四川农业大学,2007
林雪彦,牛钟相.乳酸杆菌高黏附性的适宜条件研究.中华微生物学和免疫学杂志[J]. 2005, 25(12):102
凌代文.乳酸细菌分类鉴定及实验方法[M].北京:中国轻工业出版社. 1999, 6-40
刘恩岐,仓林让.模型动物-秀丽隐杆线虫研究进展[J].动物科学与医学. 2003,20(10):23-24
刘永杰,赵艳兵,崔玢陶,等.给雏鸡饲喂乳酸菌对其盲肠主要正常菌群定植的影响[J].中国兽医杂技,1999,29(5):29-30
柳洪洁,成连贵,牛钟相,等.SPF鸡与普通鸡消化道主要正常菌群数量的比较[J].生态学杂志,2005,24(7):795-798
鲁海峰,魏桂芳,李仲逵,等. ERIC-PCR分子杂交技术分析大熊猫肠道菌群结构[J].中国微生态杂志,2005,17(2):81-84
罗晓花,孙新文.益生素作用机理及目前应用情况[J].饲料博览,2007,9:16-18
马雪云,王红妹,杨玉华.乳酸杆菌活菌制剂对大肠杆菌和鸡白痢沙门氏菌体外颉颃试验[J].山东畜牧兽医,2006,(3):4-5
那淑敏,贾士芳,陈秀珠,等.嗜酸乳酸菌产生的广谱抗菌肤AP311的分离和鉴定[J].微生物学报,2001,41:494-498
潘莉,杜惠敏,黄海东,等.腹泻儿童肠道菌群结构特征的ERIC-PCR指纹图分析[J].中国微生态学杂志,2003, 15 (3):141-143
庞林海,杜爱芳,李孝军,等.秀丽隐杆线虫培养特性与保存方法研究[J].浙江农业学报,2007,19(1) :34-36
宋凡,曹国文,戴荣国,等.复合微生态添加剂对断奶仔猪生产性能与血液生理生化指标的影响.饲料工业,2006, 27(8):36-38
沈露露,王大勇.秀丽线虫衰老调控的生理与分子机制[J].生理科学进展, 2009,40(1):75-79
沈叙庄.关注对动物使用抗生素与细菌耐药的问题[J].中华儿科杂志,2002,40(8):452-453
田召芳,常维山,唐珂心,等.产细菌素乳酸菌的筛选及体外抑菌试验[J].中国微生态学杂志,2003, 15 (2 ):87-88
万荣峰,江善祥. 3株乳酸菌体外颉颃致病性大肠埃希菌试验[J].畜牧与兽医,2007,39(3):50-52.
温彩霞,许建华,黄秀旺,等.复合型菌剂对鸡白痢菌的抑制作用及其各菌在鸡体内定居情况[J].中微生态学杂志,2003,15(1):28-32
王爱萍,张改平,张存信,等.不同日龄雏鸡消化道内乳酸菌自然分布状况研究[J].中国兽医杂志,2001,37(6):16-17
王坚镪,汤瑾,蒋燕群.益生菌对大鼠肠道菌群紊乱和细菌易位的影响[J].上海交通大学学报,2006,26(2):163-165.
王金全,周岩华,蔡辉益.国际猪液体饲喂研究进展[J].养猪,2009,4:11-13
王海泽,杨建成,卢占军.抗生素添加剂替代品的研究进展[J].饲料博览,2002,8:13-14
王鲁溪,宋树川,杨韶宇.一起B群沙门氏菌食物中毒的调查分析[J].中国公共卫生管理,2005,21(5):433
王茂起,冉陆,王竹天. 2001年中国食源性致病菌及其耐药性主动监测研究[J].卫生研究,2004,33(1):49-54
吴向军,袁杰利,梁红春.乳酸菌细胞裂解物对金黄色葡萄球菌、大肠埃希菌的抑制作用[J].中国生态学杂志,2007,19(5):422-423
许树林,李艳冰,卢向东.健康雉鸡与白痢雉鸡肠道菌群的检测比较[J].黑龙江畜牧兽医,2001,(9):7-8
徐卉芳,牛钟相,肉牛粪便正常菌群研究[J].黑龙江畜牧兽医,1999,(5):3-4
义刚,崔丽春,赵丽丽,等.重组干酪乳酸菌在模拟消化环境中生存性能的研究[J].中国微生态学杂志,2006,18(6):424-427
禹慧明,廖玲,陈平洁,等.断奶仔猪肠道菌群的研究[J].中国微生态学杂志,2000,12(2):81-82
袁杰利,文姝,康白,等.三种益生菌对种鸡肠内环境及生产性能的影响[J].中国微生态学杂志,1998,10(2):124-126
袁改玲,朱瑞英,才学鹏,等.秀丽新杆线虫的研究进展及其应用[J].中国兽医科技,2004,34(10):45-47
袁铁铮,姚斌.分子水平上益生菌研究进展[J].中国生物工程杂志,2004,24 (10):27-32
张德珍,潘道东,戴传超.一株降胆固醇乳酸菌的鉴定及其在模拟胃肠环境中抗性的研究1[J].食品科学,2004, 25( 11 ):281-283
张辉华,曹永长,毕英佐,等. 6株乳酸菌体外抑菌实验[J].中国兽医杂志,2001,37 (7 ):8-11
张辉华,曹永长,毕英佐,等.乳酸菌体外对大肠埃希菌O-78颉颃作用试验[J].中国微生态学杂志,2002,14(6):322-323
张美玲,周志华,赵立平.粪便样品中大肠杆菌多态性分子研究[J].微生物学通报, 2005 , 32(2) :5 - 9
张文鑫,龚伟.四种活菌剂对仔猪黄痢病原菌的体外抑菌试验[J].云南农业大学学报,1997,2(2):133-136
赵桂英,杨亮宇,段纲,等.断奶仔猪胃肠道正常菌群的数量和分区[J].黑龙江畜牧兽医,2003,(9):27-28.
赵桂英,段纲,杨亮宇等.仔猪消化道茵群变化与仔猪腹泻的关系[J].家畜生态,2004,25(4):44
Aballay, A., Yorgey, P., Ausubel, F.M.. Salmonella typhimurium proliferates and establishes a persistent infection in the intestine of Caenorhabditis elegans. Curr. Biol. 2000,10: 1539–1542
Aballay, A., Ausubel, F.M.. Programmed cell death mediated by ced-3 and ced-4 protects Caenorhabditis elegans form Salmonella typhimurium-mediated killing. PNAS. 2001, 98: 2735-2739
Aballay,A., Ausubel, F.M.. Caenorhabditis elegans as a host for the study of host–pathogen interactions. Curr. Opin. Micro. 2002, 5: 97–101
Aballay, A., Drenkard, E., Hilbun,L.R., et al. Caenorhabditis elegans Innate Immune Response Triggered by Salmonella enterica Requires Intact LPS and Is Mediated by a MAPK Signaling Pathway. Curr. Biol.. 2003,13: 47–52
Alakomi H.L.,Skytta E., Saarela M.,etal. Lactic acid permeabilizes gram negative bacteria by disrupt the outer membrane. Appl Environ Microbiol. 2000, 66 (5): 2001-2005
Alegado, R.A., Campbell, M.C., Chen, et al. Characterization of mediators of microbial virulence and innate immunity using the Caenorhabditis elegans host–pathogen model. Cell. Microbiol. 2003, 5: 435–444
Alcedo J , Kenyon C. Regulation of C. elegans longevity by specific gustatory and olfactory neurons. Neuron, 2004, 41 : 45-55
Alvarez Olmos M.I., Oberhelman R.A.. Probiotic agents and infectious disease:a modern perspective traditional therapy. Clin Infect Dis.2001,32(11):1567-1576
Amit-Romach D., Sklan Z.U. Microflora ecology of the chicken intestine using 16S ribosomal D primers. Poultry Science. 2004,83(7):1093-1098
Apfeld J., Kenyon C. Regulation of lifespan by sensory perception in Caenorhabditis elegans. Nature, 1999, 402 : 804～809
Bemet-Camard M.F.,Lievin V.,Brassart D.,et al. The human Lactobacillus acidophilus strain LA1 secretes a non bacteriocin antibacterial substance(s) active in vitro and in vivo. Applied and Environmental Microbiology.1997,63:2747-2753
Berdichevsky A., Viswanathan M., Horvitz H.R., et al. C. elegans SIR-2. interacts with 14-3-3 proteins to activate DAF-16 and extend life span. Cell. 2006, 125 : 1165 -1177
Berends B.R., Van Knapen F., Mossel D.A., et al. Impact on human health of Salmonella spp.on pork in the Netherlands and the anticipated effects of some currently proposed control strategies. Int. J. Food Microbiol. 1998, 44: 219–229
Berman1 J.R., Kenyon1 C. Germ- cell loss extends C. elegans life span through regulation of DAF-16 by kri21 and lipophilic hormone signaling. Cell. 2006:124 : 1055-1068
Braungart E., Gerlach M., Riederer P., et al. Caenorhabditis elegans MPP+ model of Parkinson's disease for high-throughput drug screenings. Neurodegener Dis. 2004, 1: 175-183.
Brenner S. The genetics of Caenorhabditis elegans. Genetics.1974, 77(1): 71~94
Borch E., Nesbakken T., Christensen H.. Hazard identification in swine slaughter with respect to foodborne bacteria. Int. J. Food Microbiol. 1996, 30: 9–25
Coconnier M.H,Lievin V.,Hemery E.,et al. Antagonistic activity against Helicobacter infection in vitro and in vivo by the human Lactobacillus acidophilus strain LB. Applied and environmental Microbiology. 1998,64:4573-4580
Collado, M.C., Isolauri, E., Salminen, S., et al. The impact of probiotic on gut health. Curr Drug Metab. 2009, 10: 68-78
Collins J.J., Evason K., Kornfeld K.. Pharmacology of delayed aging and extended lifespan of Caenorhabditis elegans .Exp Gerontol. 2006, 41: 1032-1039
Conway P.L.,Gorbach S.L., Goldin B. R.. Survival of Lactic acid Bacteria in the human stomachand adhesion to intestinal cells. Journal of Diary Science. 1987,70:1-12
Couillault, C., Pujol, N., Reboul, J., et al. TLR-independent control of innate immunity in Caenorhabditis elegans by the TIR domain adaptor protein TIR-1, an ortholog of human SARM. Nat. Immunol. 2004, 5: 488–494
Daba H.,Lacroix C. Influence of Growth Conditionson the Production and Activity of Mesenterocin 5 by a Strain of Leuconostoc measenteroides. Appl Microbiol Biotechnol. 1993,39:166
Daba H.,Lacroix C. Simple Method of Purification and Sequencing of a Bacteriocin Produced by Pediococcus acidilactici UL 45 .J Appl. Bateriol.1994,77:6822
Dalloul R.A.,Lillehoj H.S.,Shellem T.A., et al. Enhanced mucosal immunity against Eimeria acervulin broilers fed a Lactobacillus-based probiotic. Poultry Science. 2003,82 (1):62-66
Darby C., Cosma C.L., Thomas J.H., et al. Lethal paralysis of Caenorhabditis elegans by Pseudomonas aeruginosa. Proc Natl.Acad.Sci.USA.1999, 96:15202-15207
Di Giovanni G.D., Watrud L.S., Seidler R.J. ,et al. Fingerprinting of mixed bacterial strains and BIOLOG gram-negative(GN) substrate communities by Enterobacterial Repetitive Intergenic Consensus Sequence PCR ( ERIC-PCR) [J ]. Current Microbiolo,1999 ,38(4) :217 - 223
Donnet-Hughs A.,Rochat F.,Serrant P.,et al. Modulation of nonspecific mechanisms of defense by lactic acid bacteria:effective dose. Journal of Diary Science.1999,82:863-869
Francis, M.M., Maricq, A.V.. Electrophysiological analysis of neuronal and muscle function in C. elegans. In: Strange, K. (Eds), C. elegans: Methods and Applications. Humana Press. New Jersey. pp. 2006, 175-192
Gallagher L.A Manoil C. Pseudomonas aeruginosa PAO1 kills Caenorhabditis elegans by cyanide poisoning. J. Bacteriol. 2001, 183:6207-6214
Galan J.E., Bliska J.B. Cross-talk between bacterial pathogens and their host cells. Annu. Rev. Cell. Dev. Biol. 1996, 12:221-255
Galan J.E, Collmer A. Type III secretion machines: bacterial devices for protein delivery into host cells. Science. 1999, 284:1322-1328
Garsin, D.A., Sifri, C.D., Mylonakis, E., et al. A simple model host for identifying Gram-positive virulence factors. PNAS. 2001, 98: 10892–10897
Garsin, D.A., Villanueva, J.M., Begun, J., et al. 2003. Long-lived C. elegans daf-2 mutants are resistant to bacterial pathogens. Science. 2003, 300: 1921
Gill H.S. Stimulation of the immune system by lactic cultures. Int Dairy J,1998,8:535-544
Gill H.S.,Rutherfurd K.J.,Prasad J.,et al. Enhancement of natural and acquired immunity by Lactobacillus rhamnosus (HN001), Lactobacillus acidophilus (HN017) and Bifidobacterium lactis (HN019). Br J Nutr,2000,83:167-176
Gill H.S,Cross M.L,Rutherfurd K.J .Dietary probiotic supplementation enhances natural killer cell activity in the elderly:an investigation of age-related immunological changes.J.Clin. Immunol.2001,21:264-271
Gong, J., Forster, R. J., Yu, H., et al. Molecular analysis of bacterial populations in the ileum of broiler chickens and comparison with bacteria in the cecum. FEMS Microbiol. Ecol. 2002, 41:171-179
Gong, J., Si, W., Forster, R.J., et al. 16S rDNA-based analysis of mucosa-associated bacterial community and phylogeny in the chicken gastrointestinal tracts: from crops to ceca. FEMS Microbiol. Ecol. 2007, 59:147-157
Gopal P.K.,Prasad J.,Smart J.,et al. In Vitro adherence properties of Lactobacillus rhamnosus DR20 and Bifidobacterium lactis DR10 strains and their antagonistic activity against an enterotoxigenic Escherichia col. International Journal of Food Microbiology. 2001,67:207-216
Herich R., Revajova V., Levkut M. The effect of Lactobacillus paracasei and raftilose P95 Upon the non～specific immune response of piglets. Food and Agricultural Immunology. 2002,14: 171-179
Hodgkin J., Kuwabara P.E, Corneliussen B. A novel bacterial pathogen, Microbacterium nematophilum, induces morphological change in the nematode C. elegans. Curr Biol. 2000, 10:1615-1618
Hoffmann, J.A., Kafatos, F.C., Janeway J., et al. 1999. Phylogenetic perspectives in innate immunity. Science. 1999, 284: 1313–1318
Hoffmann, J.A. The immune response of Drosophila. Nature. 2003, 426: 33–38.
Holo H,Ivind Nilssen. Lactococcin A. A New Bacteriocin from Lactococcus lactis subsp. cremois: Isolation and Characterization of the Protein and Its Gene. J Bacteriol,1991:173(12):38793
Hsu A.L, Murphy C.T, Kenyon C. Regulation of aging and age-related disease by DAF-16 and heat- shock factor. Science, 2003, 300 : 1142-1145
Ikeda,T., Yasui,C., Hoshino,K., et al. Influence of Lactic Acid Bacteria on Longevity of Caenorhabditis elegans and Host Defense against Salmonella enterica Serovar Enteritidis. Appl. Environ. Microbiol. 2007, 73: 6404-6409
Jianhua G., Robert J. F., Hai Y,et al. Diversity and phylogenetic analysis of bacteria in the mucosa of chicken ceca and comparison with bacteria in the cecal lumen. FEMS Microbiology Letters.2002, 208: 1-7
Jianhua G., Weiduo S., Robert J. F. et al. 16S rRNA gene-based analysis of mucosa-associated bacterial community and phylogeny in the chicken gastrointestinal tracts: from crops to ceca. FEMS. Microbiol. Ecol. 2007, 59 (1): 147-157
Jin,L.Z., Ho Y.W., Abdullah N. , et al. Effects of adherent Lactobacillus cultures on growth, weight of or and intestinal microflora and volatile fatty acids in broilers. 1998, 70(3):197-209
Jin L. Z,Marquardt R., Baidoo S.K.. Inhibition of Enterotoxigenic Escherichia coli K88, K99 and 987P by the Lactobacillus isolates from Porcine Intestine[J]. Journal of the Science of food and Agriculture.2000,80:619-624
Jyoti B.D.,Suresh A.K.,Venkatesh K.V. Diacetyl production and growth of Lactobacillus rhamnosus on multiple substrates. World Journal of Microbiology and Biotechnology.2003,19(5):509-514
Kim, D.H., Feinbaum, R., Alloing G., et al.. A conserved p38 MAP kinase pathway in Caenorhabditis elegans innate immunity. Science. 2002, 297: 623–626
Kim, D.H., Amusable, F.M.. 2005. Evolutionary perspectives on innate immunity from the study of Caenorhabditis elegans. Curr. Opin. Immunol. 2005,17: 4-10
Kligler, B., Cohrssen, A., Probiotics. Am Fam Physician. 2008, 78: 1073-1078
Kurz,C.L., Ewbank, J.J. 2003. Caenorhabditis elegans: an emerging genetic model for the study of innate immunity. Nat. Rev. Genet. 2003, 4 : 380-390
Kato T.,Matsuda S.. Plantaricin-149. A Bacteriocin Produced by Lactobacillus plantarum NRIC 149. J Ferment & Bioengin. 1994, 77(3):277
Kurz C.L, Ewbank J.J. Caenorhabditis elegans for the study of host–pathogen interactions. Trends Microbiol. 2000, 8: 142-144
Labrousse A., Chauvet S., Couillault C., et al. Caenorhabditis elegans is a model host for Salmonella typhimurium. Curr Biol. 2000, 10:1543-1545
Lai CH, Chou CY, Chang LY, et al. Identification of novel human genes evolutionarily conserved in Caenorhabditis elegans by comparative proteomics [J]. Genome Res, 2000, 10: 703-713
Lewus, C.B., Montville, T.J. Detection of bacteriocins produced by lactic acid bacteria. J. Microbiol. Methods. 1991,13:145–150
Ljungh A., Wadstrom, T. 2006. Lactic acid bacteria as probiotics. Curr. Issues Intest. Microbiol. 2006, 7: 73-89
Li, M., Gong, J., Cotrill, M., et al. 2003. Evaluation of QIAamp? DNA Mini Stoll Kit for microbial ecological studies. J. Microbiol. Methods. 2003, 54:13-20
Liberati, N.T. Fitzgerald, K.A., Kim, D.H., et al. Requirement for a conserved Toll/interleukin-1 resistance domain protein in the Caenorhabditis elegans immune response. Proc. Natl. Acad. Sci. U. S. A. 2004, 101: 6593–6598
Libina N, Berman J.R, Kenyon C.. Tissue specific activities of C. elegans DAF-16 in the regulation of lifespan. Cell. 2003, 115 : 489-502
MacDougall L., Fyfe M., McIntyre L., et al. 2004. Frozen Chicken Nuggets and Strips - A Newly Identified Risk Factor for Salmonella Heidelberg Infection in British Columbia, Canada. J. Food Prot. 2004,67:1111-1115
Mahajan-Miklos S., Tan M.W, Rahme L.G., et al. Molecular mechanisms of bacterial virulence elucidated using a Pseudomonas aeruginosa–Caenorhabditis elegans pathogenesis model. Cell. 1999, 96:47-56
Mahajan-Miklos S., Rahme L.G, Ausubel F.M. Elucidating the molecular mechanisms of bacterial virulence using non-mammalian hosts. Mol Microbiol. 2000, 37: 981-988
Majowicz S.E., McNab W.B., Sockett P., et al. Burden and Cost of Gastroenteritis in a Canadian Community. J. Food Prot. 2006, 69: 651-659
Mallo, G.V. Mallo, G.V., Kurz, C.L., et al. Inducible antibacterial defense system in C. elegans. Curr. Biol. 2002, 12: 1209–1214
Matsuzaki T.,Yamazaki R.,Hashimoto S.,et al. The effect of oral feeding of Lactobacillus casei strain shirota on immunoglobulin E production in mice. Journal of Diary Science. 1998, 81: 48-53
Mikkelsen L.L,Jensen B.B. Performance and microbial activity in the gastrointestinal tract of piglets fed fermented liquid feed at weaning. Journal of Animal Feed Science, 1998,7:211-215
Miller S.I., Kudral A.M., Mekalanos J.J. A two-component regulatory system (phoP phoQ) controls Salmonella typhimurium virulence. Proc. Natl. Acad. Sci. 1989, 86: 5054-5058
Mousing J., Jensen P.T., Halgaard C., et al. Nation-wide Salmonella enterica surveillance and control in Danish slaughter swine herds. Prev. Vet. Med. 1997, 29: 247–261
Moy T., Ball A.R., ZafiaA., et al. Identification of novel antimicrobials using a live-animal infection model. Proc. Natl. Acad. Sci. U. S. A. 2006, 103: 10414-10419
Naidu S., Bidlack W.R., Clemens R.A.. Probiotic Spectra of Lactic Acid Bacteria (LAB). Crit. Rev. Food Sci. Nutr. 1999, 38: 13–126
Ngo T H ,Loredana B ,Ashley H ,et al . Characterization of Bacillus species used for oral bacteriotherapy and bacterioprophylaxis of gastrointestinal disorders. Applied and Environmental Microbiology ,2000 ,66 (12) : 5241-5247
Ogawa M.,Shimizu K.,Nomoto K.,et al. Inhibition of in vitro growth of Shiga toxin-producing Escherichia coli O157:H7 by probiotic Lactobacillus strains due to production of lactic acid.International Journal of food Microbiology.2001,68:135-140
Olsen S.J., Bishop R., Brenner F.W., et al.. The Changing Epidemiology of Salmonella: Trends in Serotypes Isolated from Humans in the United States, 1987–1997. J. Infect Dis. 2001, 183: 753-761
O’Quinn A. L., Wiegand E. M., Jeddeloh J. A. Burkholderia pseudomallei kills the nematode Caenorhabditis elegans using an endotoxinmediated paralysis. Cell Microbiol. 2001, 3:381-394
Pascual M.,Hugas M.,Badiola J. I.,et al. Lactobacillus salivarius CTC2197 prevents Salmonella enteritidis colonization in chickens. Applied and Environmental Microbiology. 1999, 65: 4981-4986
Peng G.C., Hsu C.H. The efficacy and safety of heat-killed Lactobacillus paracasei for treatment of perennial allergic rhinitis induced by house-dust mite. Pediatr. Allergy. Immunol. 2005, 16:433-438
Perdigon G.,Vintini E.,Alvarez S.,et al. Study of the possible mechanisms involved in the mucosal immune system activation by Lactic Acid Bacteria. Journal of Diary Science.1999,82:1108-1114
Poppe C., Smart N., Khakhria R., et al. Salmonella typhimurium DT104: A virulent and drug-resistant pathogen. 1998, Can Vet J. 39, 559-565
Poppe C., Ziebell K., Martin L., et al. Diversity in antimicrobial resistance and other characteristics among Salmonella Typhimurium DT104 isolates. Microb. Drug Resist. 2002, 8:107-121
Pujol N., Link E.M., Liu L.X., et al. A reverse genetic analysis of components of the Toll signaling pathway in Caenorhabditis elegans. Curr Biol. 2001, 11: 809–821
Riddle D.L., Rlumenthal T., Meyer B.J., et al. Introduce to C.elegans, In: C.elegans II. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York. 1997, 1-22
Salminen M.K., Tynkkynen S., Rautelin H., et al. Lactobacillus bacteremia during a rapid increase in probiotic use of Lactobacillus rhamnosus GG in Finland. Clin Infect Dis. 2002,35:1155-1160
Sarwar H,Ghazala H,Randy G. Genetic Transformationof an Entomopathogenic Nematode by Microinjection. Journal of Invertebrate Pathology. 1995 ,66: 293 - 296
Schulenburg H., Kurz C.L., Ewbank J.J. Evolution of the innate immune system: the worm perspective. Immunol. Rev. 2004, 198: 36–58
Shaham S. Methods in cell biology. In: Ambros, V.(Eds) .Wormbook. The C. elegans Research Community, WormBook. 2006, 1-75
Shiffrin E J.Barassart D,Servin A L,Rochat F,et al. Immune modulation of blood leukocytes in humans by lactic acid bacteria:criteria for strain selection[J]. Am J Clin Nutr,1997,66(2):515S～520S
Sifri1 C.D., Begun J., Ausubel F.M. The worm has turned– microbial virulence modeled in Caenorhabditis elegans. Trends.Micro. 2005, 13:119-127
Suau A.A, Bonnet R.G, Sutren M.N., et al. Direct Analysis of Genes Encoding 16S rRNA from Complex Communities Reveals Many Novel Molecular Species within the Human Gut. Appl.Environ.Microbiol. 1999, 65(11): 4799–4807
Sulston J.E., Horvitz, H.R. Post-embryonic cell lineages of the nematode, Caenorhabditis elegans. Dev Biol. 1977, 56
Sulston J , Hodgkin J.Methods in the nematode Caenorhabditis elegans [M]. New York:Cold Spring HarborLaboratory Press, 1988, 587 - 606
Sulston J.E. Neuronal cell lineages in the nematode, Caenorhabditis elegans. Cold Spring Harb Symp Quant Biol. 1983, 48(2): 443-452
Stoffels G.,Nissen-meyer J. Purification and Characterization of a New Bateriocin Isolated from a Carnobacterium sp. Appol. Env Microbiol. 1992,58(5):1417
Stiernagle T. Maintenance of C. elegans. In: Fay, D. (Eds), WormBook. The C. elegans Research Community, WormBook. pp. 2006, 1-11
Tannock G. W.,Munro K.,Harmsen H.J.,et al. Analysis of the fecal microflora of human subjects consuming a probiotic product containing Lactobacillus rhamnosus DR20. Appl Environ Microbiol,2000,66:2578-2588
Tan M.W., Mahajan-Miklos S., Ausubel F.M. Killing of Caenorhabditis elegans by Pseudomonas aeruginosa used to model mammalian bacterial pathogenesis. Proc Natl Acad Sci USA. 1999, 96:715-720
Tan M.W Rahme L.G., Sternberg J.A., et al. Pseudomonas aeruginosa killing of Caenorhabditis elegans used to identify P. aeruginosa virulence factors. Proc Natl Acad Sci USA. 1999, 96: 2408-2413
Tan M.W. Ausubel F.M. Caenorhabditis elegans: a model genetic host to study Pseudomonas aeruginosa pathogenesis. Curr Opin Microbiol .2000, 3:29-34
Tejada-Simon M.V,Lee J.H,Ustunol Z.,et al. Ingestion of yogurt containing Lactobacillus acidophilus and Bifidobacterium to potentiate immunoglobuhn a responses to cholera toxin in mice. Journal of Diary Science,1999,82:649-660
Tenor, J.L., McCormick, B.A., Ausubel F. M., et al. Caenorhabditis elegans-based screen identifies Salmonella virulence factors required for conserved host–pathogen interactions. Curr. Biol. 2004, 14: 1018–1024
Tissenbaum H.A., Guarente L. Increased dosage of a sir-2 gene extends lifespan in Caenorhabditis elegans. Nature. 2001, 410 : 227-230
Troemel E.R., Chu S.W., Lee S.S., et al. p38 MAPK Regulates Expression of Immune Response Genes and Contributes to Longevity in C. elegans. PLoS Genetics. 2006, 2:1725-1739
Tynkkynen S., Satokari R., Saarela M., et al. Comparison of ribotyping, randomly amplified polymorphic DNA analysis, and pulsed-field gel electrophoresis in typingof Lactobacillus rhamnosus and L. casei strains. Appl Environ Microbiol .1999,65:3908-3914
van Winsen RL, LipmanL J.A, Biesterveld S, et al. Mechanism of Salmonella reduction in fermented pig feed. Journal of the Science of Food and Agriculture, 2001,81:342-346
Voisine C., Varma H., Walker N., et al. Identification of potential therapeutic drugs for Huntington’s disease using Caenorhabditis elegans. PLoS One. 2007, 2: 504
Weisburg, W.G., Barns S.M., Pelletier D.A., et al. 16S Ribosomal DNA Amplification for Phylogenetic Study. J. BACTERIOL.1991,173(2):697-703
Zhou J.S,Shu Q.,Rutherfurd K.J.,et al. Safety assessment of potential probiotic lactic acid bacteria strains Lactobacillus rhamnosus HN001 , L.acidophilus HN017 , and Bifidobacterium lactis HN019 in BALB/c mice. Int J Food Microbiol. 2000,56:87-96
陈琼,孔繁德,彭海滨,等.51株沙门氏菌的耐药性分析[J].畜牧与兽医,2008,40(8):77-78
陈润生.猪生产学[M] .北京:中国农业出版社,1995
陈杖榴,刘健华.食品动物源细菌耐药性与公共卫生[J].兽医导刊, 2007,9:45-47
丁德忠,庞小燕,华修国,等. SPF仔猪肠道菌群结构的分子分析[J].上海交通大学学报(农业科学版),2005,25(3):256-260
房春红,刘杰,许修宏.乳酸菌素的研究现状和发展趋势[J].中国乳品工业,2006,34(2):53-55
高巍,孟庆翔,肖训军,等.生长猪胃肠道乳酸菌、双歧杆菌和大肠埃希菌的数量和分布规律[J] .中国农业大学学报,2001,6(5):81-86
高巍,孟庆翔.生长育肥猪胃肠道正常厌氧菌群的数量和分区[J].中国农业大学学报,2000,5(5):88-93
顾瑞霞,伊萌.乳酸菌抗肿瘤特性的研究进展.中国微生态学杂志[J].1999, 11 (4): 253-255
郭兴华.益生菌基础与应用[M].北京:北京科学技术出版社,2001,10
何昭阳,王增辉,吴延春.雏鸡消化道主要正常茵群定植规律的研究[J].畜牧兽医学报,2000,31(1):41-48
何昭阳,徐凤宇,管清华,等.犊牛腹泻与肠道菌群的变化[J].中国预防兽医学报,2000,22(5):345-346
何谦,郭进超,李岩,等.发酵液体饲料在断奶仔猪的应用进展[J].养猪,2007,5:5-7
胡东良.乳酸菌的抗肿瘤、抗变异原及免疫增强作用.中国乳品工业[J],1997,25 (6): 11-14
黄怡,王士长,崔艳红,等.多菌种复合益生素对三黄鸡的生产性能及肠道主要菌群的影响[J].安徽农业科学,2006,34(3):485-486
贾熙华,曹诚.秀丽隐杆线虫在医药学领域的应用和进展[J].药学学报, 2009, 44 (7): 687?694
李国平,周伦江,邵良平,等.日粮中添加复合菌制剂对雏鸡盲肠大肠杆菌数、肠道pH 值、血液SOD和GSH- Px影响的研究[J].中国兽医杂志, 1999, 25(8) : 7-8
李路胜,周响艳,冯定远.乳酸菌对麻羽肉鸡生长和免疫机能的影响研究[J].饲料工业,2006,27 (20):39 - 41
李晓晖.微生态饲料添加剂研究进展[J].饲料博览,2002,3:40～41
林雪彦,王中华,李福昌,等.乳酸杆菌黏附鸡消化道上皮细胞超微结构的观察[J].中国兽医学报,2008,28(12):1367-1373
李晓霞.新城疫病毒气溶胶发生与传染机制及其在生产鸡舍环境中的监测(D).山东农业大学,2009
李犹平.采用ERIC-PCR技术研究鸡白痢、新陈疫感染雏鸡及健康鸡肠道菌群结构(D).四川农业大学,2007
林雪彦,牛钟相.乳酸杆菌高黏附性的适宜条件研究.中华微生物学和免疫学杂志[J]. 2005, 25(12):102
凌代文.乳酸细菌分类鉴定及实验方法[M].北京:中国轻工业出版社. 1999, 6-40
刘恩岐,仓林让.模型动物-秀丽隐杆线虫研究进展[J].动物科学与医学. 2003,20(10):23-24
刘永杰,赵艳兵,崔玢陶,等.给雏鸡饲喂乳酸菌对其盲肠主要正常菌群定植的影响[J].中国兽医杂技,1999,29(5):29-30
柳洪洁,成连贵,牛钟相,等.SPF鸡与普通鸡消化道主要正常菌群数量的比较[J].生态学杂志,2005,24(7):795-798
鲁海峰,魏桂芳,李仲逵,等. ERIC-PCR分子杂交技术分析大熊猫肠道菌群结构[J].中国微生态杂志,2005,17(2):81-84
罗晓花,孙新文.益生素作用机理及目前应用情况[J].饲料博览,2007,9:16-18
马雪云,王红妹,杨玉华.乳酸杆菌活菌制剂对大肠杆菌和鸡白痢沙门氏菌体外颉颃试验[J].山东畜牧兽医,2006,(3):4-5
那淑敏,贾士芳,陈秀珠,等.嗜酸乳酸菌产生的广谱抗菌肤AP311的分离和鉴定[J].微生物学报,2001,41:494-498
潘莉,杜惠敏,黄海东,等.腹泻儿童肠道菌群结构特征的ERIC-PCR指纹图分析[J].中国微生态学杂志,2003, 15 (3):141-143
庞林海,杜爱芳,李孝军,等.秀丽隐杆线虫培养特性与保存方法研究[J].浙江农业学报,2007,19(1) :34-36
宋凡,曹国文,戴荣国,等.复合微生态添加剂对断奶仔猪生产性能与血液生理生化指标的影响.饲料工业,2006, 27(8):36-38
沈露露,王大勇.秀丽线虫衰老调控的生理与分子机制[J].生理科学进展, 2009,40(1):75-79
沈叙庄.关注对动物使用抗生素与细菌耐药的问题[J].中华儿科杂志,2002,40(8):452-453
田召芳,常维山,唐珂心,等.产细菌素乳酸菌的筛选及体外抑菌试验[J].中国微生态学杂志,2003, 15 (2 ):87-88
万荣峰,江善祥. 3株乳酸菌体外颉颃致病性大肠埃希菌试验[J].畜牧与兽医,2007,39(3):50-52.
温彩霞,许建华,黄秀旺,等.复合型菌剂对鸡白痢菌的抑制作用及其各菌在鸡体内定居情况[J].中微生态学杂志,2003,15(1):28-32
王爱萍,张改平,张存信,等.不同日龄雏鸡消化道内乳酸菌自然分布状况研究[J].中国兽医杂志,2001,37(6):16-17
王坚镪,汤瑾,蒋燕群.益生菌对大鼠肠道菌群紊乱和细菌易位的影响[J].上海交通大学学报,2006,26(2):163-165.
王金全,周岩华,蔡辉益.国际猪液体饲喂研究进展[J].养猪,2009,4:11-13
王海泽,杨建成,卢占军.抗生素添加剂替代品的研究进展[J].饲料博览,2002,8:13-14
王鲁溪,宋树川,杨韶宇.一起B群沙门氏菌食物中毒的调查分析[J].中国公共卫生管理,2005,21(5):433
王茂起,冉陆,王竹天. 2001年中国食源性致病菌及其耐药性主动监测研究[J].卫生研究,2004,33(1):49-54
吴向军,袁杰利,梁红春.乳酸菌细胞裂解物对金黄色葡萄球菌、大肠埃希菌的抑制作用[J].中国生态学杂志,2007,19(5):422-423
许树林,李艳冰,卢向东.健康雉鸡与白痢雉鸡肠道菌群的检测比较[J].黑龙江畜牧兽医,2001,(9):7-8
徐卉芳,牛钟相,肉牛粪便正常菌群研究[J].黑龙江畜牧兽医,1999,(5):3-4
义刚,崔丽春,赵丽丽,等.重组干酪乳酸菌在模拟消化环境中生存性能的研究[J].中国微生态学杂志,2006,18(6):424-427
禹慧明,廖玲,陈平洁,等.断奶仔猪肠道菌群的研究[J].中国微生态学杂志,2000,12(2):81-82
袁杰利,文姝,康白,等.三种益生菌对种鸡肠内环境及生产性能的影响[J].中国微生态学杂志,1998,10(2):124-126
袁改玲,朱瑞英,才学鹏,等.秀丽新杆线虫的研究进展及其应用[J].中国兽医科技,2004,34(10):45-47
袁铁铮,姚斌.分子水平上益生菌研究进展[J].中国生物工程杂志,2004,24 (10):27-32
张德珍,潘道东,戴传超.一株降胆固醇乳酸菌的鉴定及其在模拟胃肠环境中抗性的研究1[J].食品科学,2004, 25( 11 ):281-283
张辉华,曹永长,毕英佐,等. 6株乳酸菌体外抑菌实验[J].中国兽医杂志,2001,37 (7 ):8-11
张辉华,曹永长,毕英佐,等.乳酸菌体外对大肠埃希菌O-78颉颃作用试验[J].中国微生态学杂志,2002,14(6):322-323
张美玲,周志华,赵立平.粪便样品中大肠杆菌多态性分子研究[J].微生物学通报, 2005 , 32(2) :5 - 9
张文鑫,龚伟.四种活菌剂对仔猪黄痢病原菌的体外抑菌试验[J].云南农业大学学报,1997,2(2):133-136
赵桂英,杨亮宇,段纲,等.断奶仔猪胃肠道正常菌群的数量和分区[J].黑龙江畜牧兽医,2003,(9):27-28.
赵桂英,段纲,杨亮宇等.仔猪消化道茵群变化与仔猪腹泻的关系[J].家畜生态,2004,25(4):44
Aballay, A., Yorgey, P., Ausubel, F.M.. Salmonella typhimurium proliferates and establishes a persistent infection in the intestine of Caenorhabditis elegans. Curr. Biol. 2000,10: 1539–1542
Aballay, A., Ausubel, F.M.. Programmed cell death mediated by ced-3 and ced-4 protects Caenorhabditis elegans form Salmonella typhimurium-mediated killing. PNAS. 2001, 98: 2735-2739
Aballay,A., Ausubel, F.M.. Caenorhabditis elegans as a host for the study of host–pathogen interactions. Curr. Opin. Micro. 2002, 5: 97–101
Aballay, A., Drenkard, E., Hilbun,L.R., et al. Caenorhabditis elegans Innate Immune Response Triggered by Salmonella enterica Requires Intact LPS and Is Mediated by a MAPK Signaling Pathway. Curr. Biol.. 2003,13: 47–52
Alakomi H.L.,Skytta E., Saarela M.,etal. Lactic acid permeabilizes gram negative bacteria by disrupt the outer membrane. Appl Environ Microbiol. 2000, 66 (5): 2001-2005
Alegado, R.A., Campbell, M.C., Chen, et al. Characterization of mediators of microbial virulence and innate immunity using the Caenorhabditis elegans host–pathogen model. Cell. Microbiol. 2003, 5: 435–444
Alcedo J , Kenyon C. Regulation of C. elegans longevity by specific gustatory and olfactory neurons. Neuron, 2004, 41 : 45-55
Alvarez Olmos M.I., Oberhelman R.A.. Probiotic agents and infectious disease:a modern perspective traditional therapy. Clin Infect Dis.2001,32(11):1567-1576
Amit-Romach D., Sklan Z.U. Microflora ecology of the chicken intestine using 16S ribosomal D primers. Poultry Science. 2004,83(7):1093-1098
Apfeld J., Kenyon C. Regulation of lifespan by sensory perception in Caenorhabditis elegans. Nature, 1999, 402 : 804～809
Bemet-Camard M.F.,Lievin V.,Brassart D.,et al. The human Lactobacillus acidophilus strain LA1 secretes a non bacteriocin antibacterial substance(s) active in vitro and in vivo. Applied and Environmental Microbiology.1997,63:2747-2753
Berdichevsky A., Viswanathan M., Horvitz H.R., et al. C. elegans SIR-2. interacts with 14-3-3 proteins to activate DAF-16 and extend life span. Cell. 2006, 125 : 1165 -1177
Berends B.R., Van Knapen F., Mossel D.A., et al. Impact on human health of Salmonella spp.on pork in the Netherlands and the anticipated effects of some currently proposed control strategies. Int. J. Food Microbiol. 1998, 44: 219–229
Berman1 J.R., Kenyon1 C. Germ- cell loss extends C. elegans life span through regulation of DAF-16 by kri21 and lipophilic hormone signaling. Cell. 2006:124 : 1055-1068
Braungart E., Gerlach M., Riederer P., et al. Caenorhabditis elegans MPP+ model of Parkinson's disease for high-throughput drug screenings. Neurodegener Dis. 2004, 1: 175-183.
Brenner S. The genetics of Caenorhabditis elegans. Genetics.1974, 77(1): 71~94
Borch E., Nesbakken T., Christensen H.. Hazard identification in swine slaughter with respect to foodborne bacteria. Int. J. Food Microbiol. 1996, 30: 9–25
Coconnier M.H,Lievin V.,Hemery E.,et al. Antagonistic activity against Helicobacter infection in vitro and in vivo by the human Lactobacillus acidophilus strain LB. Applied and environmental Microbiology. 1998,64:4573-4580
Collado, M.C., Isolauri, E., Salminen, S., et al. The impact of probiotic on gut health. Curr Drug Metab. 2009, 10: 68-78
Collins J.J., Evason K., Kornfeld K.. Pharmacology of delayed aging and extended lifespan of Caenorhabditis elegans .Exp Gerontol. 2006, 41: 1032-1039
Conway P.L.,Gorbach S.L., Goldin B. R.. Survival of Lactic acid Bacteria in the human stomachand adhesion to intestinal cells. Journal of Diary Science. 1987,70:1-12
Couillault, C., Pujol, N., Reboul, J., et al. TLR-independent control of innate immunity in Caenorhabditis elegans by the TIR domain adaptor protein TIR-1, an ortholog of human SARM. Nat. Immunol. 2004, 5: 488–494
Daba H.,Lacroix C. Influence of Growth Conditionson the Production and Activity of Mesenterocin 5 by a Strain of Leuconostoc measenteroides. Appl Microbiol Biotechnol. 1993,39:166
Daba H.,Lacroix C. Simple Method of Purification and Sequencing of a Bacteriocin Produced by Pediococcus acidilactici UL 45 .J Appl. Bateriol.1994,77:6822
Dalloul R.A.,Lillehoj H.S.,Shellem T.A., et al. Enhanced mucosal immunity against Eimeria acervulin broilers fed a Lactobacillus-based probiotic. Poultry Science. 2003,82 (1):62-66
Darby C., Cosma C.L., Thomas J.H., et al. Lethal paralysis of Caenorhabditis elegans by Pseudomonas aeruginosa. Proc Natl.Acad.Sci.USA.1999, 96:15202-15207
Di Giovanni G.D., Watrud L.S., Seidler R.J. ,et al. Fingerprinting of mixed bacterial strains and BIOLOG gram-negative(GN) substrate communities by Enterobacterial Repetitive Intergenic Consensus Sequence PCR ( ERIC-PCR) [J ]. Current Microbiolo,1999 ,38(4) :217 - 223
Donnet-Hughs A.,Rochat F.,Serrant P.,et al. Modulation of nonspecific mechanisms of defense by lactic acid bacteria:effective dose. Journal of Diary Science.1999,82:863-869
Francis, M.M., Maricq, A.V.. Electrophysiological analysis of neuronal and muscle function in C. elegans. In: Strange, K. (Eds), C. elegans: Methods and Applications. Humana Press. New Jersey. pp. 2006, 175-192
Gallagher L.A Manoil C. Pseudomonas aeruginosa PAO1 kills Caenorhabditis elegans by cyanide poisoning. J. Bacteriol. 2001, 183:6207-6214
Galan J.E., Bliska J.B. Cross-talk between bacterial pathogens and their host cells. Annu. Rev. Cell. Dev. Biol. 1996, 12:221-255
Galan J.E, Collmer A. Type III secretion machines: bacterial devices for protein delivery into host cells. Science. 1999, 284:1322-1328
Garsin, D.A., Sifri, C.D., Mylonakis, E., et al. A simple model host for identifying Gram-positive virulence factors. PNAS. 2001, 98: 10892–10897
Garsin, D.A., Villanueva, J.M., Begun, J., et al. 2003. Long-lived C. elegans daf-2 mutants are resistant to bacterial pathogens. Science. 2003, 300: 1921
Gill H.S. Stimulation of the immune system by lactic cultures. Int Dairy J,1998,8:535-544
Gill H.S.,Rutherfurd K.J.,Prasad J.,et al. Enhancement of natural and acquired immunity by Lactobacillus rhamnosus (HN001), Lactobacillus acidophilus (HN017) and Bifidobacterium lactis (HN019). Br J Nutr,2000,83:167-176
Gill H.S,Cross M.L,Rutherfurd K.J .Dietary probiotic supplementation enhances natural killer cell activity in the elderly:an investigation of age-related immunological changes.J.Clin. Immunol.2001,21:264-271
Gong, J., Forster, R. J., Yu, H., et al. Molecular analysis of bacterial populations in the ileum of broiler chickens and comparison with bacteria in the cecum. FEMS Microbiol. Ecol. 2002, 41:171-179
Gong, J., Si, W., Forster, R.J., et al. 16S rDNA-based analysis of mucosa-associated bacterial community and phylogeny in the chicken gastrointestinal tracts: from crops to ceca. FEMS Microbiol. Ecol. 2007, 59:147-157
Gopal P.K.,Prasad J.,Smart J.,et al. In Vitro adherence properties of Lactobacillus rhamnosus DR20 and Bifidobacterium lactis DR10 strains and their antagonistic activity against an enterotoxigenic Escherichia col. International Journal of Food Microbiology. 2001,67:207-216
Herich R., Revajova V., Levkut M. The effect of Lactobacillus paracasei and raftilose P95 Upon the non～specific immune response of piglets. Food and Agricultural Immunology. 2002,14: 171-179
Hodgkin J., Kuwabara P.E, Corneliussen B. A novel bacterial pathogen, Microbacterium nematophilum, induces morphological change in the nematode C. elegans. Curr Biol. 2000, 10:1615-1618
Hoffmann, J.A., Kafatos, F.C., Janeway J., et al. 1999. Phylogenetic perspectives in innate immunity. Science. 1999, 284: 1313–1318
Hoffmann, J.A. The immune response of Drosophila. Nature. 2003, 426: 33–38.
Holo H,Ivind Nilssen. Lactococcin A. A New Bacteriocin from Lactococcus lactis subsp. cremois: Isolation and Characterization of the Protein and Its Gene. J Bacteriol,1991:173(12):38793
Hsu A.L, Murphy C.T, Kenyon C. Regulation of aging and age-related disease by DAF-16 and heat- shock factor. Science, 2003, 300 : 1142-1145
Ikeda,T., Yasui,C., Hoshino,K., et al. Influence of Lactic Acid Bacteria on Longevity of Caenorhabditis elegans and Host Defense against Salmonella enterica Serovar Enteritidis. Appl. Environ. Microbiol. 2007, 73: 6404-6409
Jianhua G., Robert J. F., Hai Y,et al. Diversity and phylogenetic analysis of bacteria in the mucosa of chicken ceca and comparison with bacteria in the cecal lumen. FEMS Microbiology Letters.2002, 208: 1-7
Jianhua G., Weiduo S., Robert J. F. et al. 16S rRNA gene-based analysis of mucosa-associated bacterial community and phylogeny in the chicken gastrointestinal tracts: from crops to ceca. FEMS. Microbiol. Ecol. 2007, 59 (1): 147-157
Jin,L.Z., Ho Y.W., Abdullah N. , et al. Effects of adherent Lactobacillus cultures on growth, weight of or and intestinal microflora and volatile fatty acids in broilers. 1998, 70(3):197-209
Jin L. Z,Marquardt R., Baidoo S.K.. Inhibition of Enterotoxigenic Escherichia coli K88, K99 and 987P by the Lactobacillus isolates from Porcine Intestine[J]. Journal of the Science of food and Agriculture.2000,80:619-624
Jyoti B.D.,Suresh A.K.,Venkatesh K.V. Diacetyl production and growth of Lactobacillus rhamnosus on multiple substrates. World Journal of Microbiology and Biotechnology.2003,19(5):509-514
Kim, D.H., Feinbaum, R., Alloing G., et al.. A conserved p38 MAP kinase pathway in Caenorhabditis elegans innate immunity. Science. 2002, 297: 623–626
Kim, D.H., Amusable, F.M.. 2005. Evolutionary perspectives on innate immunity from the study of Caenorhabditis elegans. Curr. Opin. Immunol. 2005,17: 4-10
Kligler, B., Cohrssen, A., Probiotics. Am Fam Physician. 2008, 78: 1073-1078
Kurz,C.L., Ewbank, J.J. 2003. Caenorhabditis elegans: an emerging genetic model for the study of innate immunity. Nat. Rev. Genet. 2003, 4 : 380-390
Kato T.,Matsuda S.. Plantaricin-149. A Bacteriocin Produced by Lactobacillus plantarum NRIC 149. J Ferment & Bioengin. 1994, 77(3):277
Kurz C.L, Ewbank J.J. Caenorhabditis elegans for the study of host–pathogen interactions. Trends Microbiol. 2000, 8: 142-144
Labrousse A., Chauvet S., Couillault C., et al. Caenorhabditis elegans is a model host for Salmonella typhimurium. Curr Biol. 2000, 10:1543-1545
Lai CH, Chou CY, Chang LY, et al. Identification of novel human genes evolutionarily conserved in Caenorhabditis elegans by comparative proteomics [J]. Genome Res, 2000, 10: 703-713
Lewus, C.B., Montville, T.J. Detection of bacteriocins produced by lactic acid bacteria. J. Microbiol. Methods. 1991,13:145–150
Ljungh A., Wadstrom, T. 2006. Lactic acid bacteria as probiotics. Curr. Issues Intest. Microbiol. 2006, 7: 73-89
Li, M., Gong, J., Cotrill, M., et al. 2003. Evaluation of QIAamp? DNA Mini Stoll Kit for microbial ecological studies. J. Microbiol. Methods. 2003, 54:13-20
Liberati, N.T. Fitzgerald, K.A., Kim, D.H., et al. Requirement for a conserved Toll/interleukin-1 resistance domain protein in the Caenorhabditis elegans immune response. Proc. Natl. Acad. Sci. U. S. A. 2004, 101: 6593–6598
Libina N, Berman J.R, Kenyon C.. Tissue specific activities of C. elegans DAF-16 in the regulation of lifespan. Cell. 2003, 115 : 489-502
MacDougall L., Fyfe M., McIntyre L., et al. 2004. Frozen Chicken Nuggets and Strips - A Newly Identified Risk Factor for Salmonella Heidelberg Infection in British Columbia, Canada. J. Food Prot. 2004,67:1111-1115
Mahajan-Miklos S., Tan M.W, Rahme L.G., et al. Molecular mechanisms of bacterial virulence elucidated using a Pseudomonas aeruginosa–Caenorhabditis elegans pathogenesis model. Cell. 1999, 96:47-56
Mahajan-Miklos S., Rahme L.G, Ausubel F.M. Elucidating the molecular mechanisms of bacterial virulence using non-mammalian hosts. Mol Microbiol. 2000, 37: 981-988
Majowicz S.E., McNab W.B., Sockett P., et al. Burden and Cost of Gastroenteritis in a Canadian Community. J. Food Prot. 2006, 69: 651-659
Mallo, G.V. Mallo, G.V., Kurz, C.L., et al. Inducible antibacterial defense system in C. elegans. Curr. Biol. 2002, 12: 1209–1214
Matsuzaki T.,Yamazaki R.,Hashimoto S.,et al. The effect of oral feeding of Lactobacillus casei strain shirota on immunoglobulin E production in mice. Journal of Diary Science. 1998, 81: 48-53
Mikkelsen L.L,Jensen B.B. Performance and microbial activity in the gastrointestinal tract of piglets fed fermented liquid feed at weaning. Journal of Animal Feed Science, 1998,7:211-215
Miller S.I., Kudral A.M., Mekalanos J.J. A two-component regulatory system (phoP phoQ) controls Salmonella typhimurium virulence. Proc. Natl. Acad. Sci. 1989, 86: 5054-5058
Mousing J., Jensen P.T., Halgaard C., et al. Nation-wide Salmonella enterica surveillance and control in Danish slaughter swine herds. Prev. Vet. Med. 1997, 29: 247–261
Moy T., Ball A.R., ZafiaA., et al. Identification of novel antimicrobials using a live-animal infection model. Proc. Natl. Acad. Sci. U. S. A. 2006, 103: 10414-10419
Naidu S., Bidlack W.R., Clemens R.A.. Probiotic Spectra of Lactic Acid Bacteria (LAB). Crit. Rev. Food Sci. Nutr. 1999, 38: 13–126
Ngo T H ,Loredana B ,Ashley H ,et al . Characterization of Bacillus species used for oral bacteriotherapy and bacterioprophylaxis of gastrointestinal disorders. Applied and Environmental Microbiology ,2000 ,66 (12) : 5241-5247
Ogawa M.,Shimizu K.,Nomoto K.,et al. Inhibition of in vitro growth of Shiga toxin-producing Escherichia coli O157:H7 by probiotic Lactobacillus strains due to production of lactic acid.International Journal of food Microbiology.2001,68:135-140
Olsen S.J., Bishop R., Brenner F.W., et al.. The Changing Epidemiology of Salmonella: Trends in Serotypes Isolated from Humans in the United States, 1987–1997. J. Infect Dis. 2001, 183: 753-761
O’Quinn A. L., Wiegand E. M., Jeddeloh J. A. Burkholderia pseudomallei kills the nematode Caenorhabditis elegans using an endotoxinmediated paralysis. Cell Microbiol. 2001, 3:381-394
Pascual M.,Hugas M.,Badiola J. I.,et al. Lactobacillus salivarius CTC2197 prevents Salmonella enteritidis colonization in chickens. Applied and Environmental Microbiology. 1999, 65: 4981-4986
Peng G.C., Hsu C.H. The efficacy and safety of heat-killed Lactobacillus paracasei for treatment of perennial allergic rhinitis induced by house-dust mite. Pediatr. Allergy. Immunol. 2005, 16:433-438
Perdigon G.,Vintini E.,Alvarez S.,et al. Study of the possible mechanisms involved in the mucosal immune system activation by Lactic Acid Bacteria. Journal of Diary Science.1999,82:1108-1114
Poppe C., Smart N., Khakhria R., et al. Salmonella typhimurium DT104: A virulent and drug-resistant pathogen. 1998, Can Vet J. 39, 559-565
Poppe C., Ziebell K., Martin L., et al. Diversity in antimicrobial resistance and other characteristics among Salmonella Typhimurium DT104 isolates. Microb. Drug Resist. 2002, 8:107-121
Pujol N., Link E.M., Liu L.X., et al. A reverse genetic analysis of components of the Toll signaling pathway in Caenorhabditis elegans. Curr Biol. 2001, 11: 809–821
Riddle D.L., Rlumenthal T., Meyer B.J., et al. Introduce to C.elegans, In: C.elegans II. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York. 1997, 1-22
Salminen M.K., Tynkkynen S., Rautelin H., et al. Lactobacillus bacteremia during a rapid increase in probiotic use of Lactobacillus rhamnosus GG in Finland. Clin Infect Dis. 2002,35:1155-1160
Sarwar H,Ghazala H,Randy G. Genetic Transformationof an Entomopathogenic Nematode by Microinjection. Journal of Invertebrate Pathology. 1995 ,66: 293 - 296
Schulenburg H., Kurz C.L., Ewbank J.J. Evolution of the innate immune system: the worm perspective. Immunol. Rev. 2004, 198: 36–58
Shaham S. Methods in cell biology. In: Ambros, V.(Eds) .Wormbook. The C. elegans Research Community, WormBook. 2006, 1-75
Shiffrin E J.Barassart D,Servin A L,Rochat F,et al. Immune modulation of blood leukocytes in humans by lactic acid bacteria:criteria for strain selection[J]. Am J Clin Nutr,1997,66(2):515S～520S
Sifri1 C.D., Begun J., Ausubel F.M. The worm has turned– microbial virulence modeled in Caenorhabditis elegans. Trends.Micro. 2005, 13:119-127
Suau A.A, Bonnet R.G, Sutren M.N., et al. Direct Analysis of Genes Encoding 16S rRNA from Complex Communities Reveals Many Novel Molecular Species within the Human Gut. Appl.Environ.Microbiol. 1999, 65(11): 4799–4807
Sulston J.E., Horvitz, H.R. Post-embryonic cell lineages of the nematode, Caenorhabditis elegans. Dev Biol. 1977, 56
Sulston J , Hodgkin J.Methods in the nematode Caenorhabditis elegans [M]. New York:Cold Spring HarborLaboratory Press, 1988, 587 - 606
Sulston J.E. Neuronal cell lineages in the nematode, Caenorhabditis elegans. Cold Spring Harb Symp Quant Biol. 1983, 48(2): 443-452
Stoffels G.,Nissen-meyer J. Purification and Characterization of a New Bateriocin Isolated from a Carnobacterium sp. Appol. Env Microbiol. 1992,58(5):1417
Stiernagle T. Maintenance of C. elegans. In: Fay, D. (Eds), WormBook. The C. elegans Research Community, WormBook. pp. 2006, 1-11
Tannock G. W.,Munro K.,Harmsen H.J.,et al. Analysis of the fecal microflora of human subjects consuming a probiotic product containing Lactobacillus rhamnosus DR20. Appl Environ Microbiol,2000,66:2578-2588
Tan M.W., Mahajan-Miklos S., Ausubel F.M. Killing of Caenorhabditis elegans by Pseudomonas aeruginosa used to model mammalian bacterial pathogenesis. Proc Natl Acad Sci USA. 1999, 96:715-720
Tan M.W Rahme L.G., Sternberg J.A., et al. Pseudomonas aeruginosa killing of Caenorhabditis elegans used to identify P. aeruginosa virulence factors. Proc Natl Acad Sci USA. 1999, 96: 2408-2413
Tan M.W. Ausubel F.M. Caenorhabditis elegans: a model genetic host to study Pseudomonas aeruginosa pathogenesis. Curr Opin Microbiol .2000, 3:29-34
Tejada-Simon M.V,Lee J.H,Ustunol Z.,et al. Ingestion of yogurt containing Lactobacillus acidophilus and Bifidobacterium to potentiate immunoglobuhn a responses to cholera toxin in mice. Journal of Diary Science,1999,82:649-660
Tenor, J.L., McCormick, B.A., Ausubel F. M., et al. Caenorhabditis elegans-based screen identifies Salmonella virulence factors required for conserved host–pathogen interactions. Curr. Biol. 2004, 14: 1018–1024
Tissenbaum H.A., Guarente L. Increased dosage of a sir-2 gene extends lifespan in Caenorhabditis elegans. Nature. 2001, 410 : 227-230
Troemel E.R., Chu S.W., Lee S.S., et al. p38 MAPK Regulates Expression of Immune Response Genes and Contributes to Longevity in C. elegans. PLoS Genetics. 2006, 2:1725-1739
Tynkkynen S., Satokari R., Saarela M., et al. Comparison of ribotyping, randomly amplified polymorphic DNA analysis, and pulsed-field gel electrophoresis in typingof Lactobacillus rhamnosus and L. casei strains. Appl Environ Microbiol .1999,65:3908-3914
van Winsen RL, LipmanL J.A, Biesterveld S, et al. Mechanism of Salmonella reduction in fermented pig feed. Journal of the Science of Food and Agriculture, 2001,81:342-346
Voisine C., Varma H., Walker N., et al. Identification of potential therapeutic drugs for Huntington’s disease using Caenorhabditis elegans. PLoS One. 2007, 2: 504
Weisburg, W.G., Barns S.M., Pelletier D.A., et al. 16S Ribosomal DNA Amplification for Phylogenetic Study. J. BACTERIOL.1991,173(2):697-703
Zhou J.S,Shu Q.,Rutherfurd K.J.,et al. Safety assessment of potential probiotic lactic acid bacteria strains Lactobacillus rhamnosus HN001 , L.acidophilus HN017 , and Bifidobacterium lactis HN019 in BALB/c mice. Int J Food Microbiol. 2000,56:87-96