【摘要】：藥用植物內生真菌是一種新型的藥用微生物資源,其產(chǎn)生的真菌胞外多糖是一種新型綠色飼料添加劑,也具有真菌多糖的多種生物學特性,如高效、無毒、無殘留,能提高機體免疫力、抗病能力、提高畜禽的生產(chǎn)性能等。本研究以藥用植物絞股藍內生真菌JY25發(fā)酵液中提取的的胞外多糖(以下簡稱JY25P)為實驗材料,通過體內外試驗,觀察JY25P對肉雞生長、免疫特性的影響,并通過對體外培養(yǎng)T、B淋巴細胞和腸黏膜免疫細胞分泌細胞因子的影響,探索藥用植物內生真菌多糖增強肉雞腸道免疫性能的機制,為其實際生產(chǎn)應用提供理論依據(jù)和基礎數(shù)據(jù)。試驗一JY25P對肉雞生長性能和腸道結構的影響在肉雞日糧中全程添加不同濃度的JY25P,通過測定肉雞平均日增重(ADG)、平均日采食量(ADFI)、料重比(FGR)研究其對肉雞生長性能的影響;檢測不同試驗組肉雞十二指腸和空腸的絨毛高度、隱窩深度、絨毛高度/隱窩深度,以分析促進腸道營養(yǎng)物質吸收的機制。結果顯示,添加0.4%的JY25P平均末重提高了6.40%(P0.05),ADG提高了9.89%(P0.05),ADFI和FGR分別降低了12.63%(P0.05)和20.50%(P0.05),證實日糧中添加JY25P可以提高肉雞生長性能,從而提高經(jīng)濟效益;從飼喂JY25P的腸道結構分析發(fā)現(xiàn):添加0.4%的JY25P在42日齡可以提高肉雞十二指腸絨毛高度和V/C值分別提高了19.73%(P0.05)和24.08%(P0.05),空腸絨毛高度和V/C值分別提高了16.03%(P0.05)和25.35%(P0.05),證實JY25P可以改善肉雞小腸腸道結構,從而可以增強機體消化吸收功能。綜合考慮JY25P對肉雞生長性能和腸道結構的影響,日糧中JY25P的添加量為0.4%時生產(chǎn)效益最好。試驗二JY25P提高肉雞免疫性能的機制研究通過檢測全程飼喂JY25P肉雞的新城疫抗體效價和腸道黏膜SIgA、IL-4、IFN-γ免疫因子的變化以分析JY25P提高肉雞免疫能力的機制。結果顯示,添加0.6%的JY25P,在21、28、35、42日齡分別提高新城疫抗體效價35.77%、35.14%、25.00%、33.50%,差異顯著(P0.05),增強了機體的特異性免疫能力;JY25P能夠增加腸道黏膜分泌SIgA、IL-4和IFN-γ(P0.05),提高機體體液免疫和細胞免疫水平,同時維持腸道黏膜免疫平衡,促進機體的非特異性免疫。由此可見,JY25P的添加可增強特異性免疫能力、提高腸黏膜非特異性免疫性能達到增強機體免疫、抗病的作用。試驗三JY25P對體外培養(yǎng)淋巴細胞增殖和腸黏膜淋巴細胞因子的影響采用MTT法檢測JY25P對體外培養(yǎng)肉雞外周血及不同免疫器官的T、B淋巴細胞增殖的影響;以不同劑量的JY25P刺激肉雞腸道派伊爾淋巴細胞(PPL)、腸上皮內淋巴細胞(IEL)、固有層淋巴細胞(LPL)分泌IL-2、IL-10、TNF-α免疫因子。試驗結果顯示,添加80μg/mL的JY25P可刺激體外培養(yǎng)的外周血、胸腺、脾臟T淋巴細胞的增殖,分別提高142.23%(P0.01)、42.05%(P0.05)、35.09%(P0.01),添加80μg/mL的JY25P在體外直接刺激外周血、脾臟、法氏囊B淋巴細胞的增殖,分別提高67.99%(P0.01)、68.47%(P0.01)、48.73%(P0.05),說明JY25P可以增強淋巴細胞的增殖;添加160μg/mL的JY25P,可顯著提高淋巴細胞抗炎細胞因子(IL-10)的表達(P0.05),抑制炎性細胞因子(IL-2、TNF-α)的表達(P0.05),在一定程度上調節(jié)黏膜免疫狀態(tài)、促進組織修復,達到治療和緩解黏膜免疫紊亂的作用。綜上所述,日糧中添加0.4%~0.6%的JY25P可以促進肉雞的生長,通過改善腸道絨毛和隱窩結構促進營養(yǎng)物質的消化吸收,提高生產(chǎn)效益;同時,提高血清中新城疫抗體效價、促進腸黏膜分泌免疫因子,增強機體特異性和非特異性免疫能力;通過體外試驗證實,一定濃度的JY25P能夠刺激肉雞T、B淋巴細胞的增殖,調整腸黏膜免疫細胞分泌淋巴細胞因子,從而調節(jié)腸黏膜的免疫狀態(tài),增強肉雞腸道免疫和抗病能力,從而達到綜合提高生產(chǎn)性能的效果。
[Abstract]:Endophytic fungi of medicinal plants are a new kind of medicinal microbial resources. The extracellular polysaccharide produced by fungi is a new kind of green feed additive, and it also has many biological characteristics of fungal polysaccharides, such as high efficiency, non-toxic and no residue. It can improve the immunity of the body, resistance to disease, and improve the production performance of livestock and poultry. This study is a medicinal plant. Extracellular polysaccharide (hereinafter referred to as JY25P) extracted from the Gynostemma pentaphyllum endophyte JY25 fermentation broth was used as the experimental material. The effects of JY25P on the growth and immune characteristics of broilers were observed in vitro and in vitro. The effects of T, B lymphocytes and intestinal mucosal immune cells on the secretion of fine cell factors in vitro were investigated to explore the increase of Polysaccharides from endophytic fungi in medicinal plants. The mechanism of intestinal immune performance of strong broilers provides theoretical basis and basic data for its practical application. The effects of JY25P on growth performance and intestinal structure of broilers are added with different concentrations of JY25P in broiler diet, by measuring average daily weight gain (ADG), average daily feed intake (ADFI) and feed weight ratio (FGR) for meat in broilers. The effects of chicken growth performance; detection of the villus height of duodenum and jejunum in different test groups, the depth of the recess, the height of the villi / recess depth to analyze the mechanism of promoting the absorption of intestinal nutrients. The results showed that the average end weight of 0.4% JY25P increased by 6.40% (P0.05), ADG increased by 9.89% (P0.05), and ADFI and FGR decreased by 12.63% respectively. P0.05) and 20.50% (P0.05), confirmed that adding JY25P in the diet could improve the growth performance of Broilers and improve economic benefits. From the intestinal structure analysis of feeding JY25P, it was found that adding 0.4% JY25P at 42 days of age could increase the height of duodenum villus and V/C value of broilers to increase 19.73% (P0.05) and 24.08% (P0.05), and the height and V/C of jejunum villus. 16.03% (P0.05) and 25.35% (P0.05) were improved respectively. It was proved that JY25P could improve the intestinal intestinal structure of Broilers and enhance the body's digestive and absorption function. The effect of JY25P on growth performance and intestinal structure of broilers was considered. The production benefit was best when the dosage of JY25P was 0.4% in the diet. Test two JY25P to improve the immune performance of broilers. By detecting the antibody titer of Newcastle disease in JY25P broilers and the changes of SIgA, IL-4, and IFN- gamma immune factors in the intestinal mucosa to analyze the mechanism of JY25P to improve the immune ability of broilers, the results showed that the antibody titer of Newcastle disease was increased by 35.77%, 35.14%, 25%, 33.50% at 21,28,35,42 days (P0.05), respectively (P0.05). It can enhance the specific immunity of the body; JY25P can increase the secretion of SIgA, IL-4 and IFN- gamma (P0.05) in the intestinal mucosa, improve the body humoral immunity and cellular immunity level, and maintain the intestinal mucosal immune balance and promote the non specific immunity of the body. Thus, the addition of JY25P can enhance the specific immunity and improve the intestinal mucosa nonspecific. The effect of heterosexual immunity can enhance the immune and disease resistance. Experiment three JY25P on the proliferation of lymphocyte and the influence of intestinal mucosal lymphocyte factor in vitro, the effect of JY25P on the proliferation of T and B lymphocyte in peripheral blood and different immune organs of broilers in vitro was detected by MTT, and the intestinal pies of broiler were stimulated with different doses of JY25P. IL lymphocyte (PPL), intestinal intraepithelial lymphocyte (IEL) and lamina propria lymphocyte (LPL) secrete IL-2, IL-10, TNF- alpha immune factors. Experimental results showed that the addition of 80 mu JY25P could stimulate the proliferation of peripheral blood, thymus and spleen T lymphocytes in vitro, increased by 142.23% (P0.01), 42.05% (P0.05), 35.09% (P0.01), and 80 micron. The proliferation of peripheral blood, spleen, and bursal B lymphocyte proliferation by JY25P in vitro increased by 67.99% (P0.01), 68.47% (P0.01), 48.73% (P0.05), indicating that JY25P could enhance the proliferation of lymphocyte, and the expression of IL-10 (P0.05) was significantly increased by the addition of 160 mu JY25P (P0.05), and inflammatory cytokines were inhibited (IL-2, TN). The expression of F- alpha (P0.05), to some extent, regulates the mucosal immune state, promotes tissue repair, and achieves the effect of treating and alleviating the mucosal immune disorder. To sum up, the addition of 0.4%~0.6% JY25P in the diet can promote the growth of broiler, and improve the digestion and absorption of nutrients by improving the intestinal villi and recess structure, and improve the production efficiency. At the same time, the antibody titer of Newcastle disease in the serum was enhanced, the intestinal mucosa secreted the immune factors and the body specific and non specific immune ability was enhanced. Through in vitro experiments, it was proved that a certain concentration of JY25P could stimulate the proliferation of T and B lymphocytes in Broilers and adjust the intestinal mucosal immune cell secreting lymphocyte factors, thus regulating the immunity of intestinal mucosa. The state enhances the intestinal immunity and disease resistance of the broiler, so as to achieve the comprehensive improvement of the production performance.
【學位授予單位】：河南工業(yè)大學
【學位級別】：碩士
【學位授予年份】：2016
【分類號】：S831.5

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