本文選題：肺炎克雷伯菌 + β-內(nèi)酰胺酶��； 參考：《山東大學(xué)》2015年碩士論文

【摘要】：抗生素被認(rèn)為是二十一世紀(jì)最偉大的醫(yī)學(xué)發(fā)現(xiàn),對(duì)細(xì)菌、病毒、寄生蟲等具有抑制和殺滅作用。在畜牧養(yǎng)殖業(yè)中,它可以防治動(dòng)物疫病、促進(jìn)畜禽生長(zhǎng)。然而大量使用甚至濫用抗生素,會(huì)使動(dòng)物腸道微生物的生存環(huán)境遭到破壞,導(dǎo)致動(dòng)物機(jī)體抵御外界的能力下降,造成疾病繼發(fā)頻發(fā)。養(yǎng)殖產(chǎn)品的抗生素殘留,將直接影響到食品安全,通過從各條食物鏈源源不斷侵入人體,誘導(dǎo)人類出現(xiàn)原發(fā)耐藥性,威脅整個(gè)人類的健康。近年來(lái),越來(lái)越多的多重耐藥肺炎克雷伯菌隨著抗生素尤其是由p-內(nèi)酰胺類和喹諾酮類等廣譜抗生素的大量使用而出現(xiàn)。肺炎克雷伯菌是典型的條件致病菌,可經(jīng)過人或動(dòng)物的呼吸道進(jìn)入肺內(nèi)引發(fā)病理性突變,其耐藥機(jī)制主要是產(chǎn)生p-內(nèi)酰胺酶、喹諾酮類等耐藥基因。在分子水平上檢測(cè)耐藥基因型并對(duì)耐藥基因的傳播進(jìn)行相關(guān)研究,以便采取科學(xué)有效的防控措施顯得尤為重要。目前,對(duì)肺炎克雷伯菌耐藥性的研究多以醫(yī)院臨床分離菌株為主,畜牧業(yè)養(yǎng)殖中產(chǎn)生耐藥性菌株的研究較少。本文從山東地區(qū)某肉禽養(yǎng)殖廠的種雞、商品雞的身體表面以及屠宰場(chǎng)的污水中采集樣品,分離純化出75株肺炎克雷伯菌作為實(shí)驗(yàn)菌株,瓊脂紙片藥敏試驗(yàn)顯示肺炎克雷伯菌對(duì)臨床常用的頭孢菌素類氨芐西林(AMP)的耐藥率已經(jīng)達(dá)到了100%,對(duì)頭孢他啶(CAZ)、卡那霉素(KAN)、氯霉素(CHL)、環(huán)丙沙星(CIP)也具有很強(qiáng)耐藥性,耐藥率在80%以上。以75株肺炎克雷伯菌DNA為模板進(jìn)行PCR擴(kuò)增檢測(cè),共檢出產(chǎn)p-內(nèi)酰胺酶耐藥基因菌株72株,其中有66株blaSHV呈陽(yáng)性,55株blaTEM呈陽(yáng)性,有60株blaCTX-M呈陽(yáng)性；對(duì)blaSHV和blaCTX-M陽(yáng)性菌株進(jìn)行基因分型檢測(cè),以SHV-11、SHV-1型和CTX-M-1為主；此外還檢測(cè)了細(xì)菌質(zhì)粒介導(dǎo)的喹諾酮類耐藥基因,發(fā)現(xiàn)有68株耐藥基因陽(yáng)性菌株,其中有4株qnrA呈陽(yáng)性,56株qnrB呈陽(yáng)性,23株qnrS呈陽(yáng)性,檢出qepA為陽(yáng)性的菌株共有2株,aac(6')-1b-cr的陽(yáng)性菌株共31株,共有66株菌株同時(shí)攜帶β-內(nèi)酰胺酶與喹諾酮類耐藥基因。根據(jù)整合子基因序列intI1、intI2 和intI3設(shè)計(jì)引物,對(duì)75株肺炎克雷伯菌進(jìn)行整合子檢測(cè),結(jié)果顯示有70株Ⅰ型整合子檢測(cè)呈陽(yáng)性,有9株含有Ⅱ型整合子,沒有發(fā)現(xiàn)Ⅲ型整合子菌株。檢測(cè)中共發(fā)現(xiàn)3種基因盒陣列,分別為dfrA17-aadA5、dfrA12-orfF-aadA2和dfrAl-sat2-aadA1,這其中出現(xiàn)頻率最大的基因盒是dfrA12-orfF-aadA2,它的大小為1989bp,共出現(xiàn)19次。本文通過對(duì)肺炎克雷伯菌耐藥性及耐藥基因的分布情況進(jìn)行初步研究,了解耐藥性產(chǎn)生的原因和耐藥基因轉(zhuǎn)移機(jī)制,以此提高人們對(duì)抗菌藥物在養(yǎng)殖業(yè)中濫用情況的認(rèn)識(shí),同時(shí)加強(qiáng)養(yǎng)殖場(chǎng)周邊環(huán)境的監(jiān)測(cè),為預(yù)防和控制肺炎克雷伯菌的潛在傳播和發(fā)展,減緩細(xì)菌耐藥提供科學(xué)依據(jù)。
[Abstract]:Antibiotics are considered the greatest medical discoveries of the 21 century, inhibiting and killing bacteria, viruses, parasites, etc.In animal husbandry, it can prevent and control animal disease and promote the growth of livestock and poultry.However, the extensive use and even abuse of antibiotics will destroy the living environment of animal intestinal microbes, reduce the ability of animal body to resist the outside world, and lead to the frequent occurrence of diseases.Antibiotic residues in aquaculture products will directly affect food safety, through the continuous invasion of the human body from the various food chains, leading to the emergence of primary drug resistance, threatening the health of the whole human.In recent years, more and more multidrug resistant Klebsiella pneumoniae have emerged with the extensive use of antibiotics, such as p- lactams and quinolones.Klebsiella pneumoniae (Klebsiella pneumoniae) is a typical conditional pathogen, which can cause pathological mutation through the respiratory tract of human or animal. The drug resistance mechanism of Klebsiella pneumoniae is mainly the production of plactamases, quinolones and other drug resistant genes.It is very important to detect drug resistance genotypes at molecular level and to study the transmission of drug resistance genes in order to take scientific and effective prevention and control measures.At present, the drug resistance of Klebsiella pneumoniae is mainly studied by clinical isolates in hospitals.In this paper, 75 strains of Klebsiella pneumoniae were isolated and purified from the body surface of broiler, commercial chicken and the sewage of slaughterhouse in a meat and poultry breeding factory in Shandong province, and 75 strains of Klebsiella pneumoniae were isolated and purified as experimental strains.Agar disk drug sensitivity test showed that Klebsiella pneumoniae was resistant to the commonly used clinical cephalosporins ampicillin (AMP) and had a high resistance to ceftazidime, kanamycin, chloramphenicol, ciprofloxacin and CIPs, and also to ceftazidime, kanamycin, chloramphenicol and ciprofloxacin.The drug resistance rate was more than 80%.Using 75 strains of Klebsiella pneumoniae DNA as template, 72 strains of plactamase-resistant gene were detected by PCR amplification. Among them, 66 strains were blaSHV positive, 55 strains were blaTEM positive, and 60 strains were blaCTX-M positive.The genotypes of blaSHV and blaCTX-M positive strains were mainly SHV-11, SHV-1 and CTX-M-1, and the quinolone resistance genes mediated by bacterial plasmids were also detected, and 68 positive strains were found.Among them, there were 4 qnrA positive strains, 56 qnrB positive strains and 23 qnrS positive strains. There were 31 positive strains with qepA positive, 66 strains carrying 尾 -lactamases and quinolones resistance genes.75 strains of Klebsiella pneumoniae were tested for integron with primers designed according to the sequence of integron gene intI1tI2 and intI3. The results showed that 70 strains of type 鈪，

本文編號(hào)：1735539