【摘要】：鴨疫里氏桿菌感染具有發(fā)病率高、死亡率高的特點,是目前引起世界范圍內水禽感染的重要疾病,同時也是造成養(yǎng)鴨業(yè)經濟損失的主要疾病之一。由于鴨疫里氏桿菌極易產生耐藥性,因此給臨床治療帶來了極大的困擾。目前關于鴨疫里氏桿菌耐藥性的研究報道還比較少,尤其是關于分子耐藥機制和傳播機制的研究鮮有報道。2013年2月至2015年7月,從廣東省9個地區(qū)收集具有典型鴨疫里氏桿菌病癥狀的病(死)鴨和鵝的腦、肝組織樣品,分離培養(yǎng)鴨疫里氏桿菌。采用特異性PCR方法進行分子學鑒定,同時采用全自動微生物鑒定儀來鑒定疑似菌株,共分離得到115株鴨疫里氏桿菌,其中鴨源54株,鵝源61株。用瓊脂二倍稀釋法測定32種抗菌藥物的敏感性。藥敏結果顯示,分離株有很高的耐藥性,有103株(89.6%)鴨疫里氏桿菌分離株存在多重耐藥現(xiàn)象。采用PCR和測序方法對115株分離菌進行了耐藥基因和整合子類型檢測和分析。結果顯示:共有5大類抗生素的25種耐藥基因被檢測出來。在鴨疫里氏桿菌分離株中首次檢測出三種氨基糖苷磷酸轉移酶基因aph(3’)-Ⅱ、aph(3’)-Ⅳ、aph(4’)-Ⅰa,氨基糖苷轉移酶基因aad B,紅霉素耐藥基因erm B、erm C和磺胺類抗菌藥物耐藥的sul3。9株鴨疫里氏桿菌株含有1類整合子,其中6株攜帶978bp大小的aad A2基因盒和3株攜帶2287bp大小的aac(6’)-Ⅱ-cat B3-aad A1基因盒。對分離得到的菌株進行Sma I-PFGE分子分型,共有58株分型成功,得到16種不同的Sma I-PFGE型(以相似度≥90%為判定標準),包括11個簇和5個單一型。本研究中的鴨疫里氏桿菌分離株存在廣泛的克隆傳播,且克隆傳播存在于不同種屬的宿主動物(鴨和鵝)間及相同、不同地區(qū)間分離到的菌株中。為探究抗生素耐藥性與生物膜的相關性,用結晶紫染色的方法定量測定生物膜形成能力。結果表明,有95(82.6%)株鴨疫里氏桿菌分離株具有產膜能力,其中40(34.8%)株是強產膜菌株,且對12~23種抗菌藥物具有抗藥性;25(21.7%)中等產膜株和30(26.1%)弱產膜菌株可對7~12種種抗菌藥物耐藥。20(17.4%)株沒有產膜能力,耐藥種數(shù)小于等于6種�？梢钥闯錾锬ば纬赡芰εc耐藥譜的種數(shù)之間存在一定的關系。此外,通過S1-PFGE和雜交的方法確定鴨疫里氏桿菌中flo R基因既位于染色體上也位于質粒上,發(fā)現(xiàn)在13株鴨疫里氏桿菌中存在一個新型攜帶flo R基因的質粒,其大小23kb,與之前的報道的質粒不同。通過接合轉移和質粒穩(wěn)定性實驗等方法證明這個新型質�？梢赞D移到與之不同種屬的大腸桿菌工程菌C600中,轉移頻率為10~(-6)~10~(-9),但是穩(wěn)定性很差。
[Abstract]:Richter Duck infection is an important disease that causes worldwide waterfowl infection with high incidence and high mortality. It is also one of the major diseases that cause economic losses in duck industry. It is very difficult for the clinical treatment because of the resistance of Ribrella canalicus to drug resistance. At present, there are few reports on drug resistance of Rim Duck, especially on the mechanism of molecular resistance and transmission. From February 2013 to July 2015, The brain and liver tissue samples of dead ducks and geese with typical symptoms of Ribrella duck were collected from 9 regions of Guangdong Province and isolated and cultured. The molecular identification was carried out by specific PCR, and the suspected strains were identified by automatic microbiological identification instrument. A total of 115 strains of Riemiella Duck were isolated, of which 54 were from ducks and 61 from geese. The sensitivity of 32 antimicrobial agents was determined by Agar double dilution method. The results of antimicrobial susceptibility showed that the isolates had high drug resistance and 103 strains (89.6%) had multiple drug resistance. The resistance genes and integron types of 115 isolates were detected and analyzed by PCR and sequencing. The results showed that a total of 25 resistant genes of 5 major antibiotics were detected. Three aminoglycoside phosphotransferase gene aph (3')-鈪，

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