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2.4. Chemical and microbial analyses Analysis of DM and CP concentration in the experimental

diets, excreta and probiotic products was done according to AOAC (1990) methods (930.05 and

976.05, respectively). The GE was measured by using the bomb calorimeter (model 1261, Parr

Instrument Co., Moline, IL), and chromium concentration was determined with an automated

spectrophotometer (Jasco V-650, Jasco Corp., Tokyo, Japan) according to the procedure of Fenton

and Fenton (1979). The microbiological assay of faecal samples (d 14 and 28) and intestinal

digesta (d 28) was conducted by culturing in different media for the determination of total

anaerobic bacteria (Tryptic soy agar), Bifidobacterium spp. (MRS agar), Lactobacillus spp. (MRS

agar+0.02% NaN3+0.05% L-cystine hydrochloride monohydrate), Clostridium spp. (TSC agar)

and coliforms (violet red bile agar). The microbiological assay of probiotic products was also

carried out by culturing technique. The L. acidophilus was enumerated using MRS agar+0.02%NaN3+0.05% L-cystine hydrochloride monohydrate, B. Subtilis by using plate count agar, S.

cerevisiae and A. oryzae by potato dextrose agar. The anaerobic conditions during the assay ofanaerobic were created by using gas pack anaerobic system (BBL, No. 260678; Difco, Detroit,

MI). The tryptic soy agar (No. 236950), MRS agar (No. 288130), violet red bile agar (No.

216695), plate count agar (No. 247940), and potato dextrose agar (No. 213400) used were

purchased from Difco Laboratories (Detroit,MI), and TSC agar (CM0589) was purchased fromOxoid (Hampshire, UK). The pH of probiotic products was determined by pH meter (Basic pH

Meter PB-11, Sartorius, Germany).2.5. Small intestine morphology Three cross-sections for each intestinal sample were prepared

after staining with azure A and eosin using standard paraffin embedding procedures. A total of 10

intact, welloriented crypt-villus units were selected in triplicate for each intestinal cross-section as

described previously (Jin et al., 2008). Villus height was measured from the tip of the villi to the

villus crypt junction, and crypt depth was defined as the depth of the invagination between

adjacent villi. All morphological measurements (villus height and crypt depth) were made in 10-μm increments by using an image processing and analysis system (Optimus software version 6.5,

Media Cybergenetics, North Reading, MA).2.6. Statistical analysesAll the data obtained in the current study were analyzed in accordance with a randomized complete block design using the GLM procedure of SAS (SAS Inst. Inc., Cary, NC). In Exp. 1, one-way analysis of variance test was used and when significant differences (Pb0.05) were determined among treatment means, they were separated by using Duncan's multiple range tests. In Exp. 2, the data were analyzed as

a 2×2 factorial arrangement of treatments in randomized complete block design. The main effects of probiotic products (LF or SF), antibiotic (colistin or lincomycin), and their interaction were determined by the Mixed procedures of SAS. However, as

the interaction (probiotic x antibiotic) was not statistically significant (Pb0.05), it was

removed from the final model. The pen was the experimental unit for all analysis in both experiments. The bacterial concentrations were transformed (log) before statistical analysis.3.1. Experiment 1

3.2. Experiment 2n compared with pigs fed NC and LF diets (Table 4).n (d 28) than pigs fed NC, PC and LF idium spp. and coliforms than pigs fed NC , ADFI and G:F were observed in pigs fed PC, LF and SF dietsctively) when compared with pigs fed LF diet (Table 7).pigs fed PC (d 14 and 28) and SF (d 28) diets had less (Pb0.05) faecalof DM and GE; however, pigs fed PC and SF diets had greater ATTD of CP whewhen compared with pigs fed NC diet. Moreover, pigs fed PC and SF diets had hiHowever, different antibiotics had no effect on the ATTD of DM, CP and y treatments had no effect on the performance of pigs during phase I (Table

Moreover, pigs fed SF diet had greater (Pb0.05) faecal Lactobacillus spp. populatioDuring phase I, pigs fed SF diet consumed more feed than pigs fed LF diet, wherer, different antibiotics had no effect on the performance of pigs. Pigs fed SF diet

had greater ATTD of DM and CP during phases I and II (Pb0.01 and 0.001, respe3.2.2. Bacterial population in intestinePigs fed SF diet had greater (Pb0.05) Lactobacillus spp. And less Clostridium spp. (Pb0.01) and

coliform (Pb0.05) population in the ileum than pigs fed LF diet (Table 8). Additionally,higher (Pb0.05) caecal Bifidobacterium spp. Population was observed in pigs fed SF diet.

Antibiotics had no effect on the ileal microbial population; however, pigs fed colistin diet had less

number of Bifidobacterium spp. (Pb0.05) and coliforms (Pb0.01) in the cecum, whereas, feeding

of lincomycin diet resulted in reduced (Pb0.05) caecal Clostridium tion.3.1.2. Bacterial population in faeces3.2.1. Growth performance and apparent total tract digestibilityeas the ADG and ADFI were similar between pigs fed LF and SF diets (Table 6).

During phase II and the overall experimental period, pigs fed SF diet showedverall experimentalperiod. The dietary treatments had no influence on the ATTDDietary treatments had no effect on the faecal total anaerobes and Bifidobacterium

gher (Pb0.05) ADG and better G:F than pigs fed LF diet during phase II and the ospp. population at d 14 and 28, and Lactobacillus spp. at d 14 (Table 5). However,

3). However, during phase II and the overall experimental period, improved (Pb0.05)

better ADG (Pb0.01), ADFI (Pb0.01) and G:F (Pb0.05) thanpigs fed LF diet. Howev3.2.3. Small intestinal morphology3.1.1. Growth performance and apparent total tract digestibility

2.4 化学和微生物分析 在试验日粮干物质和粗蛋白含量的分析中，排泄物和益生菌产品是根据AOAC（1990）方法（分别为930.05和976.05 ）分析。使用弹式量热计（model 1261, Parr

Instrument Co., Moline, IL）测定总能量，根据芬顿博士（1979）的使用自动化分光光度计确定铬浓度。 第14和28天排泄物样本和第28天的肠道消化物的微生物分析，将样品在不同培养基中培养，测定总厌氧菌（胰酶解物大豆琼脂),双歧杆菌（MRS培养基），乳杆菌种（MRS培养基＋0.02% NaN3+0.05%L胱氨酸盐酸盐水化合物），梭菌属某些种（TSC培养基）和大肠杆菌群（紫色红胆汁琼脂）。通过培养技术对"益生菌"产品进行了微生物分析。嗜酸乳杆菌用MRS培养基＋0.02% NaN3+0.05%L胱氨酸盐酸盐水化合物培养，枯草芽孢杆菌通过琼脂培养基平板计数，酵母菌和米曲霉菌通过土豆葡萄糖培养基计数。通过使用厌氧袋厌氧系统创造厌氧条件，(BBL, No. 260678; Difco, Detroit, MI)。大豆胰酶解物的琼脂(No.236950)，MRS琼脂培养基(No. 288130)，结晶紫中性红胆盐琼脂(No.216695)，平板计数琼脂(No. 247940)，和马铃薯葡萄糖培养基(No. 213400)的使用是从Difco实验室（(Detroit,MI)购买的，和TSC培养基是从Oxoid (Hampshire, UK)购买的。益生菌产品的pH值由pH计(Basic pH Meter PB-11, Sartorius,Germany).测定。2.5 小肠形态学 每个肠道样本取三横截面，使用标准的石蜡包埋程序与天青A和曙红染色。共有10个完好无损，良好的导向隐窝绒毛单位被选定为每个肠道截面，每个样品一式三份的 (Jin et

al.2008)。从绒毛尖端绒毛到隐窝交界处测量绒毛高度，这个被定义为相邻绒毛内陷深度和隐窝深度。所有形态测量（绒毛高度和隐窝深度）通过使用一个图像处理的分析系统（擎天柱软件版本6.5，网络媒体遗传学，北读，MA），增量为10-μm。4. DiscussionPrevious studies on probiotics lack information on the method of production used, however, the

preparation of probiotics by LF method is fairly common (Patel et al., 2004). The probiotic

products used in the present study differedfrom the previous reports in that harvested probiotic microbes were added directly to the diets. In

this study, the microbial biomass grown on the CB was directly sprayed onthe carrier (corn and soybean meal) to obtain LF probiotic product. In case of the SF probiotic

product, corn and soybean meal was used as a substrate during fermentation and as a carrier of

probiotic microbes. We have reported previously that multi-microbe probiotic product prepared by

SF method was better than the probiotic product prepared by submerged liquid fermentation in

improving performance, nutrient retention and reducing harmful intestinal bacteria in broilers(Shim et al., 2010). In the current study, LF and SF method was used and corn–soybean meal was

used as a substrate forthe growth of potential probiotic microbes under optimum conditions.2.6 统计分析The different probiotic products and antibiotics had no influence on the morphology of different

segments of the small intestine, except for the greater (Pb0.05) villus height:crypt depth at the

jejunum and ileum noticed in pigs fed lincomycin diet (Table 9).

3 结果3.2 实验23.1 实验1多的乳酸菌属。3.1.2 粪便中的细菌数量析中细菌数转化为log计数。3.2.1 生长性能和表观总消化道消化率3.1.1 生长性能和表观总消化道消化率总能量的总体表观消化率不同的抗生素没有影响。SF的猪有更多的粗蛋白总体表观消化率（表4）。表观消化率上不同的饮食处理没有影响，然而，与饲喂NC和LF的猪相比，饲喂PC和猪在第一阶段的饮食处理对性能没有影响（表3）。然而，在第二阶段和整个试验期间，饲喂LF的猪表现出更好的ADG（P<0.01），ADFI（P<0.01）和G：F（P<0.05）。然当与饲喂NC饮食的猪相比时，饲喂PC，LF和SF饮食的猪其ADG，ADFI和G：F显试验期间更好的G：F比饲喂SF饮食的猪有更高的平均日增重。在干物质和总能的总体法检验。在实验二中，对作为的2×2因子安排的完全随机区组设计的处理数据进行分析。而，在猪的性能上，不同的抗生素没有影响。在第一和第二阶段中，饲喂SF的猪比饲喂有较少的粪便梭菌属和大肠杆菌群。而且，饲喂SF的猪比饲喂NC，PC和LF的猪有更在第一阶段，饲喂SF的猪比饲喂LF的猪消耗更多的饲料，饲喂SF和饲喂LF的猪的总益生菌产品的主要成效（LF或SF），抗生素（粘菌素或洁霉素），和他们的相互作用由LF的猪有更多的干物质和粗蛋白的总体表观消化率（表7）。然而，对干物质，粗蛋白和它就要从最终的模型中剔除。在这两个实验中所有分析中都以畜舍为实验单位。在统计分有影响（表5）。然而，饲喂PC（第14天和28天）和SF（第28天）的猪比饲喂NC的猪体表观消化率和平均日增重是相同的（表6）。在第二阶段和整个试验期间饲喂SF的猪比 不同的饮食处理对粪便中的第14天和28天的厌氧菌和双歧杆菌属和第14天的乳酸菌属没SAS的混合程序计算。然而，当相互作用（益生菌x抗生素)没有统计学意义时(P<0.05)，采用单向方差分析测试,不同处理之间显着性差异水平定为（P<0.05），用邓肯多重比较方著提高(P<0.05)。而且，饲喂PC和SF饮食的猪有更高的平均日增重，在第二阶段和整个第3.2.2章。 在肠道内细菌种群与喂养LF饲料的猪相比(表8)，在喂养SF饲料的猪的回肠道中乳酸菌属的数量较多(Pb0.05)，梭菌属(Pb0.01)与大肠杆菌属(Pb0.05)的数量较少。此外,在喂养SF饲料的猪的盲肠中发现较多的双歧杆菌属菌种(Pb0.05)。抗生素对回肠中的微生物种群没有影响，然而，喂养含粘菌素饲料，会导致猪的盲肠中双歧杆菌属(Pb0.05)和大肠杆菌属(Pb0.01)的数量下降。相反的，喂养含洁霉素饲料，会导致猪的盲肠中梭菌属(Pb0.05)的数量减少。3.2.3。 小肠形态学除了喂养含洁霉素饲料会导致猪空肠和回肠的绒毛高度 （窝深度 ）增加(Pb0.05)外，不同的益生菌产品和抗生素不会对小肠的不同部位的形态产生影响。 在一个完全随机区组设计中对所有在当前的研究中获得的数据进行分析。在实验一中，

4。讨论虽然先前对益生菌的研究缺乏关于产品使用方法的信息,但采用LF方法制备益生菌还是相当普遍的(Patel et al., 2004)。在本研究中，益生菌产品的使用不同于先前的报道，而是直接将提取的益生菌添加到饲料中。在本研究中,在CB上生长的微生物被直接喷洒在介质上(玉米和大豆)，进而获得LF益生菌产品。益生菌产品（玉米和大豆）被用作发酵过程中的基质和益生菌的载体。我们以前有报告采用LF方法制备益生菌在改善性能,营养保留和减少有害肠道细菌方面要比用液态发酵的方法来得好(Shim et al., 2010)。在目前的研究中,采用LF和SF方法，以玉米和黄豆粉为基质，在最佳条件下培养益生菌。5.