本文選題：納米氧化鋅　切入點(diǎn)：斷奶仔豬　出處：《中國農(nóng)業(yè)大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：高劑量氧化鋅(2000 mg Zn/kg)被證實(shí)具有良好的抑制仔豬斷奶腹瀉的效果,但其會(huì)造成嚴(yán)重的環(huán)境鋅污染。為降低使用量,減少污染,納米氧化鋅被開發(fā)出來并應(yīng)用于仔豬飼料。納米氧化鋅相比于傳統(tǒng)氧化鋅具有較小的粒子結(jié)構(gòu)與較大的比表面積,但是否能夠在低劑量下(500 mg Zn/kg)有效地抑制仔豬腹瀉,及其作用機(jī)理有待研究。本試驗(yàn)擬通過動(dòng)物實(shí)驗(yàn),比較低劑量納米氧化鋅和傳統(tǒng)高劑量氧化鋅抑制仔豬腹瀉的效果差別。為研究納米氧化鋅對(duì)仔豬腸道菌群結(jié)構(gòu)及黏膜屏障功能的影響,通過運(yùn)用高通量測(cè)序技術(shù),研究納米氧化鋅對(duì)斷奶仔豬前腸和后腸菌群結(jié)構(gòu)的影響,以及納米氧化鋅對(duì)腸道通透性、腸道形態(tài)、腸道微生物代謝產(chǎn)物、腸道緊密連接蛋白與腸道干細(xì)胞標(biāo)志蛋白的調(diào)控規(guī)律和機(jī)制。本試驗(yàn)共分為兩個(gè)部分:試驗(yàn)一:研究不同類型氧化鋅對(duì)斷奶仔豬腹瀉率、生產(chǎn)性能和腸道屏障的影響。選用120只28日齡初始體重為6.5 ± 0.5 kg杜×長×大三元雜交斷奶仔豬,隨機(jī)分為4個(gè)處理組,每個(gè)處理6個(gè)重復(fù),分別飼喂基礎(chǔ)日糧、添加2000 mg Zn/kg普通氧化鋅組、添加500 mg Zn/kg包被氧化鋅組、添加500 mg Zn/kg納米氧化鋅組。飼養(yǎng)42 d,試驗(yàn)開始和結(jié)束時(shí)對(duì)仔豬稱重,每天記錄仔豬的生長性能和腹瀉發(fā)生率。飼養(yǎng)試驗(yàn)結(jié)束后,進(jìn)行仔豬腸道通透性試驗(yàn),并屠宰取樣,取回腸和結(jié)腸腸段組織,進(jìn)行緊密連接蛋白和腸道干細(xì)胞標(biāo)志蛋白檢測(cè)。實(shí)驗(yàn)結(jié)果顯示,與對(duì)照組相比,在不影響采食量的前提下,納米氧化鋅組仔豬的飼料轉(zhuǎn)化率顯著提高(P0.05),且納米氧化鋅與傳統(tǒng)氧化鋅和包被氧化鋅的改善效果接近(P0.05)。添加納米氧化鋅在第1-3周和第4-6周的腹瀉發(fā)生率相比對(duì)照組顯著降低(P0.05),添加納米氧化鋅組與傳統(tǒng)氧化鋅和包被氧化鋅相比沒有差異。與對(duì)照組相比,納米氧化鋅處理組能夠有效降低腸道通透性(P0.05),能夠差異性的增加回腸和結(jié)腸絨毛高度,從而顯著增加絨毛高度與隱窩深度的比值(P0.05)。蛋白免疫印跡結(jié)果顯示,與對(duì)照組和傳統(tǒng)氧化鋅處理組相比,回腸緊密連接蛋白Claudin-1、Claudin-3和Occludin的表達(dá)量在包被氧化鋅和納米氧化鋅組顯著增加(P0.05),Claudin-5的表達(dá)量無顯著性差異(P0.05);結(jié)腸中緊密連接蛋白Claudin-5的表達(dá)量在包被氧化鋅和納米氧化鋅組顯著高于對(duì)照組和傳統(tǒng)氧化鋅組(P0.05),Claudin-3和Occludin的表達(dá)量在傳統(tǒng)氧化鋅、包被氧化鋅和納米氧化鋅處理組中均增加顯著(P0.05)�；啬c干細(xì)胞標(biāo)志蛋白Lgr5和Bmil的表達(dá)量顯著上升(P0.05),結(jié)腸中Bmil的表達(dá)量顯著上升(P0.05),但Lgr5的表達(dá)量則無顯著變化(P0.05)。試驗(yàn)二:研究納米氧化鋅對(duì)斷奶仔豬腸道微生物區(qū)系的影響。在試驗(yàn)一的基礎(chǔ)上,收集回腸和結(jié)腸內(nèi)容物,運(yùn)用Illumina二代測(cè)序技術(shù),檢測(cè)微生物菌群結(jié)構(gòu)以及微生物代謝產(chǎn)物的濃度;取回腸和結(jié)腸腸段進(jìn)行腸道短鏈脂肪酸受體表達(dá)量的檢測(cè)。微生物測(cè)序結(jié)果表明,納米氧化鋅添加到日糧中,回腸微生物的多樣性顯著上升,其中回腸厚壁菌門的毛螺菌科、乳酸菌科、韋榮球菌科和紅蝽菌科比例顯著上升(P0.05)。變形菌門的腸桿菌科的比例明顯降低(P0.05)。結(jié)腸菌群多樣性雖然沒有顯著的變化,但其有益菌和致病菌的比例發(fā)生顯著性變化,有益菌毛螺菌科、瘤胃菌科、韋榮球菌科和乳酸桿科比例提高,鏈球菌科、普雷沃氏菌科和理研菌科等比例下降(P0.05)。微生物代謝產(chǎn)物結(jié)果表明,飼喂納米氧化鋅后,回腸乙酸濃度顯著下降,丁酸濃度顯著上升(P0.05)。結(jié)腸中,乙酸、丙酸和丁酸濃度均顯著上升(P0.05)。同時(shí),脂肪酸受體檢測(cè)結(jié)果顯示,納米氧化鋅處理后,脂肪酸受體GPR41的表達(dá)量相比于對(duì)照組顯著上升,與傳統(tǒng)氧化鋅和包被氧化鋅處理組類似。綜上所述,低劑量納米氧化鋅(500 mg Zn/kg)具有與高劑量傳統(tǒng)氧化鋅(2000 mg Zn/kg)類似,能夠顯著降低仔豬的腹瀉發(fā)生率,提高生長性能,其作用機(jī)制可能是通過改善斷奶仔豬腸道微生物菌群結(jié)構(gòu)、提高腸道黏膜屏障功能和局部黏膜免疫來實(shí)現(xiàn)的。
[Abstract]:Zinc Oxide high dose (2000 mg Zn/kg) has been shown to have good inhibition of post weaning diarrhea effect, but it will cause serious environmental pollution. In order to reduce zinc consumption, reduce pollution, nano Zinc Oxide have been developed and applied to the piglet feed. Compared with Zinc Oxide nano particle structure a larger surface area than the traditional Zinc Oxide has a small, but whether it in a low dose (500 mg Zn/kg) effectively inhibited the diarrhea of piglets, and its mechanism remains to be studied. This paper through animal experiment, compared with low dose and high dose of traditional nano Zinc Oxide Zinc Oxide inhibit piglet diarrhea effect. In order to study the effect of nano Zinc Oxide on intestinal bacteria in piglets group structure and the mucosal barrier function, through the use of high-throughput sequencing technology, influence of nano Zinc Oxide on Weaned Piglets foregut and hindgut microflora, and nano Zinc Oxide Intestinal permeability, intestinal morphology and intestinal microbial metabolites, intestinal tight junction protein and intestinal stem cell marker protein regulation and mechanism. The test is divided into two parts: Test: a study of different types of Zinc Oxide on Weaned Piglets diarrhea rate, affect the production performance and intestinal barrier. A total of 120 28 day old initial the weight of 6.5 + 0.5 kg Duroc * Landrace * Yorkshire crossbred weaned piglets were randomly divided into 4 groups with 6 replicates of each treatment were fed basal diet supplemented with 2000 mg Zn/kg, Zinc Oxide group, adding 500 mg Zn/kg package by Zinc Oxide group, adding 500 mg nano Zn/kg Zinc Oxide group. After 42 d. And at the end of the piglets weighing the start of the test, the incidence rate of every record on growth performance and diarrhea. After the end of the feeding trial, of intestinal permeability in piglets, and slaughtered, retrieve the intestines and colon segment into the organization. For the tight junction protein and intestinal stem cell marker protein detection. The experimental results show that compared with the control group, without affecting the food intake, Zinc Oxide group nano piglet feed conversion rate increased significantly (P0.05), and Zinc Oxide and Zinc Oxide and traditional nano package is close to Zinc Oxide (P0.05) to improve the effect of adding nano. Zinc Oxide in the 1-3 week and the 4-6 week incidence of diarrhea was significantly decreased compared to the control group (P0.05), adding nano Zinc Oxide group and traditional Zinc Oxide and Zinc Oxide package showed no difference compared with control group, treatment group of nano Zinc Oxide can effectively reduce the intestinal permeability (P0.05), can increase the difference of the ileum and colon villi the height, and increase the ratio of villus height and crypt depth (P0.05). Western blot results showed that compared with the control group and the traditional Zinc Oxide group, the ileum is closely connected to the egg White Claudin-1, expression of Claudin-3 and Occludin by Zinc Oxide and Zinc Oxide group increased significantly in the nano package (P0.05), no significant difference in the expression of Claudin-5 (P0.05); the expression of tight junction protein Claudin-5 in the colon by Zinc Oxide and Zinc Oxide nano group significantly higher than that of the control group and the traditional group of Zinc Oxide in the package (P0.05) the expression of Claudin-3 and Occludin, in the traditional Zinc Oxide, Zinc Oxide and Zinc Oxide nano coated treatment group were increased significantly (P0.05). The ileum stem cell marker protein expression of Lgr5 and Bmil increased significantly (P0.05), significantly increased the expression of Bmil in colon (P0.05), but the expression of Lgr5 was not significant change (P0.05). Experiment two: the influence of nano Zinc Oxide on the intestinal microflora of weaned piglets. On the basis of Experiment 1, collecting ileum and colon contents, using the two generation Illumina sequencing technology, detection of microorganisms Microbial community structure and microbial metabolites concentration; retrieve the intestines and colon segment of intestinal short chain fatty acid receptor expression of microbial detection. The sequencing results showed that the nano Zinc Oxide was added to the diets, significantly increased the diversity of microorganisms in the ileum, ileum Firmicutes hair spirillaceae, lactic acid bacteria, Veillonella, increased significantly, and the proportion of red bug bacteria (P0.05). The proportion of Enterobacteriaceae Proteobacteria decreased significantly (P0.05). The colonic microflora diversity although no significant changes, but the proportion of bacteria and pathogens remarkable changes in bacteria, Mao spirillaceae, rumen bacteria families, Veillonella, and Lactobacillu, increase the proportion of streptococcaceae, and Poulet Was was a decline in the proportion of RIKEN bacteria (P0.05). The results showed that microbial metabolites, with nano Zinc Oxide after the ileum decreased the concentration of acetic acid, butyric acid concentration Increased significantly (P0.05). In the colon, acetic acid, propionic acid and butyric acid concentration increased significantly (P0.05). At the same time, fatty acid receptor assay showed that nano Zinc Oxide after the treatment, the expression of fatty acid receptor GPR41 compared with the control group increased significantly, and the traditional Zinc Oxide and Zinc Oxide group was similar. In conclusion, low the dose of nano Zinc Oxide (500 mg Zn/kg) with high doses of traditional Zinc Oxide (2000 mg Zn/kg), can significantly reduce the incidence of diarrhea, improve the growth performance and its mechanism may be through improving piglet intestinal microflora structure, improve intestinal mucosal barrier function and local mucosal immunity to achieve.

【學(xué)位授予單位】：中國農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：S828.5

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