本文選題：噬菌體 + 產(chǎn)氣腸桿菌�。� 參考：《河南科技學院》2017年碩士論文

【摘要】：產(chǎn)氣腸桿菌是一種重要的條件致病菌,可引起肺炎、菌血癥、泌尿系統(tǒng)感染等,本研究以一株多重耐藥產(chǎn)氣腸桿菌為宿主,從污水中分離出一株噬菌體,并對該噬菌體的生物學特性和基因組進行分析,以期找到一種抗生素替代方案,為之后的噬菌體治療做鋪墊。主要內(nèi)容與結(jié)果如下:1.產(chǎn)氣腸桿菌噬菌體的分離從醫(yī)院獲得一株多重耐藥產(chǎn)氣腸桿菌3-SP,通過菌種鑒定,耐藥性分析等,確定菌種信息,并將其保存于菌庫。然后以該菌為宿主,以環(huán)境污水為樣品,分離出一株烈性噬菌體,命名為phiEap-2,并制備好噬菌體原種懸浮液,保存于本實驗室。2.產(chǎn)氣腸桿菌噬菌體生物學特性研究在透射電鏡下觀察發(fā)現(xiàn),phiEap-2為蝌蚪狀,蛋白質(zhì)衣殼的頭部直徑約55 nm,非收縮尾部長度約為117 nm,直徑10 nm,可判斷該噬菌體屬于長尾噬菌體科。一步生長曲線的繪制表明,噬菌體phiEap-2的潛伏期為20min,爆發(fā)期約40min。穩(wěn)定性研究表明,噬菌體phiEap-2對溫度較為敏感,在高溫(70-80℃)處理下會失去活性,而在人體體溫(37℃)、室溫(24℃)和低溫(4℃)下,會保持較高的活性;噬菌體phiEap-2在pH3或pH=14時存活率急劇下降,總體來看,在堿性條件下該噬菌體的存活率比在酸性條件下的存活率高,這表明該噬菌體更加耐堿而不耐酸。宿主范圍試驗表明,該株噬菌體具有很強的專一性。3.產(chǎn)氣腸桿菌噬菌體基因組分析經(jīng)過提取與純化,得到純度較高的的噬菌體phiEap-2基因組,對其進行測序,并將其基因組全序列及注釋信息上傳至Genbank,序列號為KT287080。phiEap-2基因組長度為40491bp,GC含量為51.95%,屬于線狀雙鏈DNA。對噬菌體的基因組進行預測之后,得到62個開放閱讀框,其中23個開放閱讀框的功能可以被注釋,39個開放閱讀框被注釋為假定蛋白。分析表明,phiEap-2結(jié)構(gòu)基因模塊與噬菌體FSL SP-031和粘質(zhì)沙雷氏菌噬菌體ETA有一定程度的序列保守性,雖然噬菌體之間存在著顯著差異,但是結(jié)構(gòu)基因模塊是在這三株噬菌體(phiEap-2,FSL SP-031和ETA)基因組相同位置發(fā)現(xiàn)的,這表明phi Eap-2與沙門氏菌噬菌體FSL SP-031和粘質(zhì)沙雷氏菌噬菌體ETA在進化上是相關(guān)的。
[Abstract]:Enterobacter aerogenes is an important conditional pathogen, which can cause pneumonia, bacteremia, urinary system infection, etc. In this study, a bacteriophage was isolated from sewage with a multidrug resistant Enterobacter aerogenes as host. The biological characteristics and genome of the bacteriophage were analyzed in order to find an alternative to antibiotics and pave the way for the later phage therapy. The main contents and results are as follows: 1. A multidrug resistant Enterobacter aerogenes 3-SP1 was isolated from the hospital. The information of the bacteria was determined by the identification of strains and the analysis of drug resistance, and stored in the library. Then a strong bacteriophage named phiEap-2 was isolated from environmental sewage and the suspension of bacteriophage was prepared and stored in our laboratory. The biological characteristics of Enterobacter aerogenes phage were observed under transmission electron microscope. It was found that phiEap-2 was tadpole shaped, the head diameter of protein capsid was about 55 nm, the length of non-contractile tail was about 117 nm, and the diameter was 10 nm. The one-step growth curve showed that the latent period of phage phiEap-2 was 20 min and the outbreak time was about 40 min. The stability study showed that the bacteriophage phiEap-2 was sensitive to temperature and could lose its activity under high temperature (70 ~ 80 鈩，

本文編號：1957996