三種基因分型技術在病原微生物溯源方面的應用
發(fā)布時間：2018-10-12 21:40
【摘要】： 各種細菌性疾病一直危害著人類健康和畜牧業(yè)的健康發(fā)展。分型技術能夠根據(jù)細菌的生化特征或基因組成分析不同來源菌株間的關系,從而對某次疾病暴發(fā)追蹤溯源。過去的幾十年里,大量基于DNA的基因分型技術發(fā)展起來,表現(xiàn)出傳統(tǒng)分型技術無法比擬的優(yōu)越性。基因分型技術要滿足一些標準才能廣泛推廣,如分辨力、穩(wěn)定性、簡便快速、易于解釋等。本研究主要比較脈沖場凝膠電泳PFGE、Sau-PCR和ERIC-PCR三種方法在微生物溯源中的應用,來探討適于廣泛推廣的技術。 首先用三種方法對一起由福氏志賀菌引起的食物中毒事件進行溯源分析,分型結果高度一致,推論此次菌痢疫情的罪魁禍首是該餐館中污染了福氏志賀菌的食物。據(jù)此建議相關食品監(jiān)督部門和衛(wèi)生檢疫部門加強對其監(jiān)測和管理。三種技術所用時間長短和成本花費多少的比較依次為PFGESau-PCRERIC-PCR。因此建議基層單位可首選便宜快速的ERIC-PCR或Sau-PCR進行初期的溯源分析。 其次利用三種方法對傳代培養(yǎng)30代和室溫放置30天的志賀菌進行基因分型分析,驗證這三種方法帶型的穩(wěn)定性。PFGE帶型未發(fā)生可檢測的變化,具有完美的重復性。而ERIC-PCR發(fā)生了較大變化,容易提供錯誤的流行病學信息。發(fā)現(xiàn)Sau-PCR對于傳代培養(yǎng)和室溫放置的志賀菌分型結果都非常穩(wěn)定,使實驗室間的數(shù)據(jù)交流和建立數(shù)據(jù)庫成為可能。進一步論證了Sau-PCR推廣使用的可行性。 最后比較了Sau-PCR和ERIC-PCR技術對9株耐甲氧西林金黃色葡萄球菌(MRSA)的分型研究。結果顯示MRSA的Sau-PCR基因型與SCCmec(staphylococcal cassette chromosome mec)基因型一致。而Sau-PCR技術無需了解菌株的基因信息,不需要特殊設備,具有操作簡便快捷的優(yōu)點,為MRSA的快速檢測和溯源分析提供了有力的工具。 本論文依次比較了三種分型技術對病原微生物溯源的簡便快捷性、穩(wěn)定性和對革蘭氏陽性菌MRSA的分辨力,發(fā)現(xiàn)了新發(fā)展的Sau-PCR技術比PFGE簡便易行,比ERIC-PCR穩(wěn)定、分辨力更強,為新方法Sau-PCR在基層單位的推廣使用提供了進一步的實驗和理論依據(jù)。
[Abstract]:A variety of bacterial diseases have been harmful to human health and the healthy development of animal husbandry. Typing technique can analyze the relationship between different strains according to the biochemical characteristics or gene composition of bacteria, so as to trace the source of a disease outbreak. In the past few decades, a large number of genotyping techniques based on DNA have been developed, showing unparalleled superiority compared with traditional genotyping techniques. Genotyping techniques must meet some criteria to be widely used, such as resolution, stability, simplicity and rapidity, easy to explain, and so on. In this study, the application of pulsed field gel electrophoresis (PFGE,Sau-PCR) and ERIC-PCR in microbial traceability was compared. Three methods were used to trace the food poisoning caused by Shigella flexneri, and the results were highly consistent. It was concluded that the culprit of the disease was the food contaminated with Shigella flexneri in the restaurant. Accordingly, the relevant food supervision departments and health and quarantine departments should strengthen their monitoring and management. The length of time and cost of the three technologies are compared in order of PFGESau-PCRERIC-PCR. Therefore, it is suggested that grassroots units should choose cheap and fast ERIC-PCR or Sau-PCR for initial traceability analysis. Then three methods were used to analyze the genotyping of Shigella after 30 generations of passage culture and 30 days at room temperature to verify the stability of the three methods. The PFGE band type was not detectable and had perfect reproducibility. However, ERIC-PCR has changed greatly and it is easy to provide wrong epidemiological information. It was found that Sau-PCR was very stable for subculture and room temperature storage of Shigella, which made it possible to exchange data between laboratories and establish databases. The feasibility of popularizing Sau-PCR is further demonstrated. Finally, Sau-PCR and ERIC-PCR techniques were compared in the typing of 9 strains of methicillin-resistant Staphylococcus aureus (MRSA). The results showed that the Sau-PCR genotype of MRSA was consistent with that of SCCmec (staphylococcal cassette chromosome mec) genotype. Sau-PCR technology does not need to know the genetic information of the strain and does not need special equipment. It has the advantages of simple and fast operation. It provides a powerful tool for rapid detection and traceability analysis of MRSA. In this paper, we have compared the convenience, stability and resolution of three typing techniques to trace pathogenic microorganisms to MRSA. It is found that the newly developed Sau-PCR technique is easier than PFGE, more stable than ERIC-PCR, and has stronger resolution. It provides further experimental and theoretical basis for the popularization and application of the new method Sau-PCR in basic units.
【學位授予單位】：華南理工大學
【學位級別】：碩士
【學位授予年份】：2010
【分類號】：R37;S852.6
本文編號：2267694
[Abstract]:A variety of bacterial diseases have been harmful to human health and the healthy development of animal husbandry. Typing technique can analyze the relationship between different strains according to the biochemical characteristics or gene composition of bacteria, so as to trace the source of a disease outbreak. In the past few decades, a large number of genotyping techniques based on DNA have been developed, showing unparalleled superiority compared with traditional genotyping techniques. Genotyping techniques must meet some criteria to be widely used, such as resolution, stability, simplicity and rapidity, easy to explain, and so on. In this study, the application of pulsed field gel electrophoresis (PFGE,Sau-PCR) and ERIC-PCR in microbial traceability was compared. Three methods were used to trace the food poisoning caused by Shigella flexneri, and the results were highly consistent. It was concluded that the culprit of the disease was the food contaminated with Shigella flexneri in the restaurant. Accordingly, the relevant food supervision departments and health and quarantine departments should strengthen their monitoring and management. The length of time and cost of the three technologies are compared in order of PFGESau-PCRERIC-PCR. Therefore, it is suggested that grassroots units should choose cheap and fast ERIC-PCR or Sau-PCR for initial traceability analysis. Then three methods were used to analyze the genotyping of Shigella after 30 generations of passage culture and 30 days at room temperature to verify the stability of the three methods. The PFGE band type was not detectable and had perfect reproducibility. However, ERIC-PCR has changed greatly and it is easy to provide wrong epidemiological information. It was found that Sau-PCR was very stable for subculture and room temperature storage of Shigella, which made it possible to exchange data between laboratories and establish databases. The feasibility of popularizing Sau-PCR is further demonstrated. Finally, Sau-PCR and ERIC-PCR techniques were compared in the typing of 9 strains of methicillin-resistant Staphylococcus aureus (MRSA). The results showed that the Sau-PCR genotype of MRSA was consistent with that of SCCmec (staphylococcal cassette chromosome mec) genotype. Sau-PCR technology does not need to know the genetic information of the strain and does not need special equipment. It has the advantages of simple and fast operation. It provides a powerful tool for rapid detection and traceability analysis of MRSA. In this paper, we have compared the convenience, stability and resolution of three typing techniques to trace pathogenic microorganisms to MRSA. It is found that the newly developed Sau-PCR technique is easier than PFGE, more stable than ERIC-PCR, and has stronger resolution. It provides further experimental and theoretical basis for the popularization and application of the new method Sau-PCR in basic units.
【學位授予單位】：華南理工大學
【學位級別】：碩士
【學位授予年份】：2010
【分類號】：R37;S852.6
【引證文獻】
相關期刊論文 前1條
1 張淑紅;吳清平;徐曉可;張菊梅;郭偉鵬;;Sau-PCR分子分型技術及應用[J];中國食品衛(wèi)生雜志;2012年02期
，本文編號：2267694
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