本文選題：副溶血性弧菌 + 生化分型��； 參考：《青島大學(xué)》2017年碩士論文

【摘要】：副溶血性弧菌(V.parahaemolyticus,VP)是嗜鹽性革蘭氏陰性菌,主要存在于我國沿海區(qū)域,該菌能夠引起食源性食物中毒及疾病,從而導(dǎo)致地方性流行的胃腸炎暴發(fā),其發(fā)生規(guī)模在我國沿海和部分內(nèi)陸區(qū)域已經(jīng)表現(xiàn)出明顯的上升趨勢,高居細(xì)菌性食物中毒的首要位置。大量研究發(fā)現(xiàn),耐熱性直接溶血素(thermostable direct hemolysin,TDH)或TDH相關(guān)溶血素(TDH-related hemolysin,TRH)存在于能夠致病的副溶血弧菌中。這兩種溶血素是導(dǎo)致副溶血性弧菌致病的主要毒力因子,與該菌的致病能力密切相關(guān)。副溶血性弧菌實(shí)驗(yàn)室的常用分型方法主要有生化分型和血清學(xué)分型,這兩種方法不能有效地對目標(biāo)菌的同源性進(jìn)行分析,但是脈沖場凝膠電泳技術(shù)(Pulsed field gel electrophoresis,PFGE)方法具有分型力好、分辨力強(qiáng)、可重復(fù)性的優(yōu)點(diǎn),常被用于微生物的鑒定分型研究。本研究對淄博市2013年-2016年自水產(chǎn)品分離出的副溶血性弧菌菌株毒力相關(guān)基因的分布情況進(jìn)行了分析,同時(shí)應(yīng)用PFGE技術(shù)對VP分離菌株進(jìn)行分子分型,研究其同源性,為副溶血性弧菌導(dǎo)致食源性疾病的防控提供實(shí)驗(yàn)依據(jù)。目的:利用血清分型方法、毒力基因分子檢測和PFGE技術(shù)對淄博市2013年-2016年市售水產(chǎn)品中分離得到的副溶血性弧菌進(jìn)行比對分析,了解近4年該目標(biāo)菌之間存在的關(guān)系,分析其流行規(guī)律,為有效預(yù)防和控制食源性疾病的發(fā)生提供科學(xué)依據(jù)。方法:對淄博市2013年-2016年分離自市售新鮮水產(chǎn)品的副溶血性弧菌菌株進(jìn)行復(fù)蘇培養(yǎng)和復(fù)核鑒定。對上述目標(biāo)分離菌株進(jìn)行血清分型,探討血清型的分布特點(diǎn);用PCR對tdh和trh基因進(jìn)行檢測;PFGE技術(shù)進(jìn)行副溶血性弧菌的分子分型,探討菌株的同源性。結(jié)果:(1)85株分離株經(jīng)復(fù)核鑒定,復(fù)核結(jié)果與初始鑒定一致。共分出7個(gè)O抗原血清型,以O(shè)2群為優(yōu)勢血清型,占29.41%。K抗原鑒定出型別較少,只有5種,且分布較分散,無明顯趨勢。(2)多重?zé)晒釶CR檢測結(jié)果表明,85株分離菌株毒力相關(guān)基因tdh攜帶率為3.53%,trh攜帶率為5.88%,trh高于tdh。(3)PFGE分型結(jié)果顯示,血清型同為O2:K28的2株分離株P(guān)FGE圖譜不盡相同,其他分離菌株血清型各不相同且PFGE圖譜各自成型,相似度極低。攜帶毒力基因的分離株間親緣關(guān)系較遠(yuǎn),沒有明顯優(yōu)勢帶型。結(jié)論:(1)淄博市副溶血性弧菌分離菌株O抗原分布以O(shè)2群居多,K抗原型別較少且分布較分散,無明顯趨勢。(2)市售水產(chǎn)品分離菌株中,毒力基因tdh和trh檢出率較低,且均只攜帶單一毒力基因。(3)本次實(shí)驗(yàn)水產(chǎn)品中副溶血性弧菌的血清型和PFGE圖譜類別相關(guān)性不明顯,PFGE帶型分散,其親緣關(guān)系較遠(yuǎn)。
[Abstract]:Vibrio parahaemolyticus (V. parahaemolyticus) is a halophilic gram-negative bacteria, which mainly exists in coastal areas of China. It can cause foodborne food poisoning and diseases, thus leading to the outbreak of endemic gastroenteritis. Its occurrence scale has shown an obvious upward trend in coastal and some inland areas of China and occupies the primary position of bacterial food poisoning. A large number of studies have found that thermostable direct hemolysin (TDH) or TDH-related hemolysin (TRH) are present in vibrio parahaemolyticus which can cause the disease. These two hemolysins are the main virulence factors leading to virulence of Vibrio parahaemolyticus and are closely related to the pathogenicity of Vibrio parahaemolyticus. The commonly used typing methods of Vibrio parahaemolyticus laboratory mainly include biochemical typing and serological typing. These two methods can not effectively analyze the homology of the target bacteria, but pulsed field gel electrophoresisPFGE has good typing power. The advantages of strong resolution and repeatability are often used in the identification and typing of microorganisms. In this study, the distribution of virulence related genes of Vibrio parahaemolyticus isolated from aquatic products in Zibo City from 2013 to 2016 was analyzed. At the same time, the virulence genes of Vibrio parahaemolyticus strains isolated from aquatic products from 2013 to 2016 were genotyped by PFGE to study their homology. To provide experimental basis for the prevention and control of foodborne diseases caused by Vibrio parahaemolyticus. Objective: to compare and analyze Vibrio parahaemolyticus isolated from aquatic products sold in Zibo city from 2013 to 2016 by using serotyping method, virulence gene molecular detection and PFGE technique, and to understand the relationship between Vibrio parahaemolyticus and Vibrio parahaemolyticus in recent 4 years. The analysis of its epidemic law provides scientific basis for effective prevention and control of foodborne diseases. Methods: Vibrio parahaemolyticus strains isolated from fresh aquatic products from 2013 to 2016 in Zibo City were resuscitated and identified. The serotyping of the target isolates was carried out, and the distribution of the serotypes was discussed. The molecular typing of Vibrio parahaemolyticus was carried out by using PCR to detect the tdh and trh genes, and the homology of the strains was discussed. Results the results were consistent with the initial identification. A total of 7 O antigen serotypes were identified, with O2 group as the dominant serotype, accounting for 29.41.K antigens, only 5 types of O antigen serotypes were identified, and their distribution was scattered. The results of multiplex PCR analysis showed that the carrying rate of virulence related gene tdh was 3.53 and 5.88trh was higher than that of tdh.(3)PFGE typing. The results showed that the PFGE patterns of the two isolates with the same serotype of O2:K28 were different. The serotypes of other isolated strains were different and the PFGE profiles were formed respectively, and the similarity was very low. The isolates carrying virulence genes had a long genetic relationship, and there was no obvious dominant band type. Conclusion the distribution of O antigen of Vibrio parahaemolyticus isolates in Zibo city is less and more scattered than that of O 2 colony, and there is no obvious trend. The detection rate of virulence gene tdh and trh is lower in the isolated strains of aquatic products on the market. The serotype of Vibrio parahaemolyticus in aquatic products was not significantly correlated with the type of PFGE profile.
【學(xué)位授予單位】：青島大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R155.5

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