【摘要】：瓜實(shí)蠅Bactrocera cucurbitae(Coquillett)是重要的農(nóng)業(yè)害蟲(chóng),生物防治實(shí)蠅技術(shù)—昆蟲(chóng)不育技術(shù)[Sterile Insect Technique(SIT)]已越來(lái)越受到國(guó)內(nèi)外農(nóng)業(yè)專(zhuān)家的青睞,而SIT中的輻照過(guò)程削弱了靶標(biāo)實(shí)蠅的體質(zhì)健康,關(guān)于這一方面的研究還鮮少有詳細(xì)的報(bào)道,本文研究了輻照對(duì)瓜實(shí)蠅遺傳區(qū)性品系[B.cucurbitae genetic sexing strain(GSS)]腸道菌的多樣性和相對(duì)豐度的影響,并分離、鑒定和篩選優(yōu)勢(shì)菌屬飼喂蟲(chóng)源,在一定程度上降低了輻照的傷害。研究結(jié)果如下:1、以100Gy137Cs輻照瓜實(shí)蠅遺傳區(qū)性品系的5、8、11和13日齡的雌、雄成蟲(chóng),并采用基于16SrDNA的IlluminaMiSeq測(cè)序技術(shù)分別對(duì)其腸道菌群進(jìn)行測(cè)序分析。結(jié)果表明從瓜實(shí)蠅雌、雄成蟲(chóng)腸道中共檢測(cè)到4個(gè)菌門(mén)和19個(gè)主要菌屬,其中變形菌門(mén)Proteobacteria(占總量的50%～98%)、厚壁菌門(mén)Firmicutes(占總量的2%～47%)和普羅威登斯菌屬Providencia sp.、腸桿菌屬Enterobacter sp.分別是瓜實(shí)蠅GSS腸道中的優(yōu)勢(shì)菌門(mén)和菌屬;輻照顯著的減少了腸道中的微生物多樣性,降低了優(yōu)勢(shì)菌屬的相對(duì)豐度,這與瓜實(shí)蠅的性別和成蟲(chóng)日齡無(wú)關(guān)。2、利用傳統(tǒng)的微生物分離技術(shù)從瓜實(shí)蠅雌、雄5日齡成蟲(chóng)腸道中共分離得到7種菌屬,分別為:腸桿菌屬(37%),普羅威登斯菌屬(18.5%),克雷伯氏菌屬Klebsiella sp.(1.9%),芽孢桿菌屬Lysinibacillus sp.(1.9%),西地西菌屬 Cedecea sp.(24%),Bacterium sp.(11.1%)和未知屬名的其他菌屬other(5.6%)。3、利用腸桿菌屬作為瓜實(shí)蠅GSS幼蟲(chóng)飼料的添加劑,用活菌(PI)、死菌(API)分別作為兩個(gè)處理組,用無(wú)菌(NPI)作為對(duì)照,飼喂寄主至化蛹后,分別測(cè)定瓜實(shí)蠅GSS的生物學(xué)和形態(tài)學(xué)指標(biāo),結(jié)果發(fā)現(xiàn)活菌組能顯著的增加瓜實(shí)蠅GSS的蛹重、頭寬和胸長(zhǎng),而死菌和活菌組都能顯著的提高雌、雄成蟲(chóng)的耐壓存活率,它們提高的平均百分比分別是18%和4.72%(雌蟲(chóng)),15%和11%(雄蟲(chóng))。因腸桿菌屬易分離和培養(yǎng),在所分離的菌屬中所占比例最大,而應(yīng)用腸桿菌屬優(yōu)化幼蟲(chóng)飼料有顯著的效果,結(jié)果證明腸桿菌屬是瓜實(shí)蠅GSS腸道中的優(yōu)勢(shì)菌屬,也是強(qiáng)化幼蟲(chóng)飼料的益生菌屬。本研究成果將為未來(lái)開(kāi)發(fā)利用營(yíng)養(yǎng)菌作為瓜實(shí)蠅GSS幼蟲(chóng)飼料的添加劑,從而在一定程度上修復(fù)輻照的損傷提供理論依據(jù),以期提高不育成蟲(chóng)的質(zhì)量,為能高效的應(yīng)用SIT技術(shù)打下基礎(chǔ)。
[Abstract]:Bactrocera cucurbitae (Coquillett) is an important agricultural pest. Biological control technology of fly, insect sterility technique [Sterile Insect Technique (SIT)] has been more and more favored by agricultural experts at home and abroad. However, the irradiation process in SIT weakens the physical health of the target fly, and there are few detailed reports on this aspect. The effects of irradiation on the diversity and relative abundance of intestinal bacteria in the genetic line [B.cucurbitae genetic sexing strain (GSS)] of melon fruit fly were studied, and the dominant bacteria were isolated, identified and screened, which reduced the damage caused by irradiation to some extent. The results were as follows: 1. 100Gy137Cs was used to irradiate the female, male and male adults of 5, 811 and 13 day old female and male adults of genetic region of fruit fly, and the intestinal flora was sequenced by IlluminaMiSeq sequencing technique based on 16SrDNA. The results showed that there were 4 phylum and 19 main genera in the intestinal tract of female and male adults of fruit fly, among which Proteobacteria was 50%. Firmicutes (2 / 47% of total) and Providencia sp., (Enterobacter sp.) It is the dominant phylum and genus in GSS intestinal tract of fruit fly, respectively. Irradiation significantly reduced the microbial diversity in the intestine and the relative abundance of the dominant genus, which had nothing to do with the sex of the fruit fly and the age of the adult. 2. The traditional microbiological isolation technique was used to isolate the female from the fruit fly. Seven species of bacteria were isolated from the intestinal tract of male adults at 5 days of age. They were Enterobacter (37%), Providence (18.5%), Klebsiella (1.9%) and Bacillus (Lysinibacillus sp. (1.9%). Cedecea sp. (24%), Bacterium sp. (11. 1%) and other unknown genus other (5. 6%). 3. Enterobacter was used as additive for GSS larvae of fruit fly and (PI), was used as a living bacterium. Dead bacteria (API) was used as two treatment groups, and sterile (NPI) was used as control. After feeding host to pupa, the biological and morphological indexes of GSS were measured respectively. The results showed that the live bacteria group could significantly increase the pupae weight of GSS. The head width and chest length were significantly increased in both dead and living bacteria groups, and the average percentage of increase was 18% and 4.72% (female), 15% and 11% (male), respectively. Because Enterobacterium is easy to isolate and culture, it accounts for the largest proportion in the isolated genus, and the application of Enterobacter to optimize larval feed has remarkable effect. The results show that Enterobacter is the dominant genus in the intestinal tract of GSS. It is also a probiotic genus that strengthens larval feed. The results of this study will provide a theoretical basis for the future development and utilization of nutritious bacteria as a feed additive for GSS larvae of fruit flies, thus repairing the damage caused by irradiation to a certain extent, in order to improve the quality of sterile adults. It lays the foundation for the efficient application of SIT technology.
【學(xué)位授予單位】：福建農(nóng)林大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類(lèi)號(hào)】：S433

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