杂交鲟维氏气单胞菌的分离鉴定及其耐药性分析
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摘要
试验旨在对贵州省某养鱼场发病杂交鲟体内分离的优势菌株进行鉴定、致病力探究及耐药性分析。本试验通过革兰染色、生化鉴定、细菌16S rDNA基因分析、动物回归试验、药敏试验、部分毒力基因检测与耐药基因检测。结果显示:分离菌为革兰阴性短杆菌,细菌16S rDNA扩增测序与比对,得大小为1 477 bp的片段,其片段基因序列与GenBank中的维氏气单胞菌序列相似度达99.00%,药敏试验显示:分离菌对环丙沙星、氟苯尼考、头孢哌酮等药物敏感,对四环素、磺胺异恶唑、苯唑西林药物出现耐药,耐药基因检测显示:分离菌含有耐磺胺基因(Sul1,sul2)、Ⅰ类整合子(Intl1)3种耐药基因,毒力基因检测显示:分离菌含有气溶素(Aer)和粘附素(Aha)两种毒力基因。综合本试验结果,分离菌为维氏气单胞菌(A.veronii),含有毒力基因,并对环丙沙星、氟苯尼考、头孢哌酮等药物敏感。本试验对贵州地区鲟发病流行的病因做出准确诊断,为该地区细菌性鱼病预防与合理用药提供依据。
The purpose of the experiment was to identify the dominant strains isolated from the diseased sturgeon in a fish farm in Guizhou Province, explore the pathogenicity and analyze the drug resistance. The results showed that the isolates were Gram-negative Brevibacterium spp. and 16 S rDNA amplification, sequencing and alignment of bacteria were performed. The size of the 16 S rDNA fragment was 1477 bp. The gene sequence of the fragment was similar to that of Aeromonas Virginis in GenBank. The similarity was 99.00%. Drug susceptibility test showed that the isolates were sensitive to ciprofloxacin, florfenicol and cefoperazone, and resistant to tetracycline, sulfamidoxazole and oxacillin. Drug resistance gene detection showed that the isolates contained sulfonamide-resistant genes(Sul1, sul2) and class I integron(Intl1). Virulence gene detection showed that the isolates contained aerosol. Aer and Aha were two virulence genes. According to the results of this experiment, the isolated strain is Aeromonas veronii, which contains virulence genes and is sensitive to ciprofloxacin, florfenicol and cefoperazone. This experiment did an accurate diagnosis of the causes of the occurrence and epidemic of sturgeon in Guizhou, and provided a basis for the prevention and rational use of bacterial fish diseases in Guizhou
The purpose of the experiment was to identify the dominant strains isolated from the diseased sturgeon in a fish farm in Guizhou Province, explore the pathogenicity and analyze the drug resistance. The results showed that the isolates were Gram-negative Brevibacterium spp. and 16 S rDNA amplification, sequencing and alignment of bacteria were performed. The size of the 16 S rDNA fragment was 1477 bp. The gene sequence of the fragment was similar to that of Aeromonas Virginis in GenBank. The similarity was 99.00%. Drug susceptibility test showed that the isolates were sensitive to ciprofloxacin, florfenicol and cefoperazone, and resistant to tetracycline, sulfamidoxazole and oxacillin. Drug resistance gene detection showed that the isolates contained sulfonamide-resistant genes(Sul1, sul2) and class I integron(Intl1). Virulence gene detection showed that the isolates contained aerosol. Aer and Aha were two virulence genes. According to the results of this experiment, the isolated strain is Aeromonas veronii, which contains virulence genes and is sensitive to ciprofloxacin, florfenicol and cefoperazone. This experiment did an accurate diagnosis of the causes of the occurrence and epidemic of sturgeon in Guizhou, and provided a basis for the prevention and rational use of bacterial fish diseases in Guizhou
引文
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[16] 李忱炜.肺炎链球菌溶血素对细菌毒力的影响及诱导RAW264.7细胞凋亡的研究[D].重庆:重庆医科大学,2011.
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[19] 曹恒源,乔晔,陈小娟,等.圆口铜鱼维氏气单孢菌的分离鉴定及药敏试验[J].水生态学杂志,2016,37(4):95-100.
[20] 胡天野,吴同垒,孟庆峰,等.框镜鲤维氏气单胞菌的生物学特性[J].中国兽医杂志,2012,48(8):18-21.
[2] 董靖,丁浩,胥宁,等.杂交鲶维氏气单胞菌鉴定及联合药敏试验研究[J].中国预防兽医学报,2017,39(11):897-901.
[3] 黎炯,叶星,卢迈新,等.罗非鱼维氏气单胞菌的分离鉴定和药敏试验[J].水生态学杂志,2011,32(3):132-136.
[4] 杨移斌,夏永涛,邱军强,等.鲟源维氏气单胞菌分离鉴定及药敏特性研究[J].四川动物,2014,33(3):414-418.
[5] ZHU C K,LIU G J,ZHANG Z S,et al,Identificationof the pat hogenic Aeromonas veronii isolated from rock car,Procyp is rabaudi[J].Chinese Journal of Zoonoses,2017,33(6):526-534.
[6] WOO P C,LAU S K,WONG S S,et al.Two cases of continuous ambulatory peritoneal dialysis-associated peritonitis due to Plesiomonas shigelloides[J].Jcour-nal of Clinical Microbiology,2004,42(2):933-935.
[7] WANG X L,XU L P,CAO H,et al.Identification and drug sensitivity of a Plesiomonas shigelloides isolated from diseased sturgeons[J].Acta Microbiologica Sin-ica,2013,53(7):723-729.
[8] HU Q D,LIN Q,SHI C B,et al.Isolation and identi-fication of pathogenic Plesiomonas shigelloides from diseased Grass carp[J].Acta Microbiologica Sinica,2014,54(2):229-235.
[9] Hickman-Brenner F.MacDonald K.Steigerwalt A.et al,Aer-omonas veronii,a new ornithine decarboxylase-positive species.that may cause diarrhea[J].J Clin Microbiol,1987,25(5):900-906.
[10] 崔树玉,李景学,孙启华,等,自淡水鱼中分离出一株维隆氏气单胞菌[J].中国卫生检验杂志,1994,4(3):133.
[11] 房海,陈翠珍,张晓君,等.中华绒螯蟹病原维氏气单胞菌的检验[J].中国人兽共患病学报,2008,24(1):45-49.
[12] 黄小丽,汪开毓.斑点叉尾鮰维氏气单胞菌病的诊断与防治[J].水产科技情报,2009,36(5):240-241.
[13] 杨求华.养殖鳗鲡致病性气单胞菌的分离与鉴定[D].厦门:集美大学,2012.
[14] 张明洋,胡安东,程振涛,等.杂交鲟类志贺邻单胞菌的分离鉴定及耐药性分析[J].中国畜牧兽医,2019,46(1):264-270.
[15] 王兴丽,汪开毓,陈德芳,等.一株虹鳟源维氏气单胞菌的分离鉴定及组织病理学观察[J].中国预防兽医学报,2016,38(11):879-883.
[16] 李忱炜.肺炎链球菌溶血素对细菌毒力的影响及诱导RAW264.7细胞凋亡的研究[D].重庆:重庆医科大学,2011.
[17] 邱军强,杨先乐,程训佳.嗜水气单胞菌毒力因子特性及作用机理研究进展[J].中国病原生物学杂志,2009,4(8):616-619.
[18] Zepeda-Velazquez A P,Vega-Sanchez V,Salgado-Miranda C,et al.Histopathological findings in farmed rainbow trout(Oncor-hynchus mykiss) naturally infected with 3 different Aeromonasspecies [J].Can J Vet Res,2015,79(3):250-254.
[19] 曹恒源,乔晔,陈小娟,等.圆口铜鱼维氏气单孢菌的分离鉴定及药敏试验[J].水生态学杂志,2016,37(4):95-100.
[20] 胡天野,吴同垒,孟庆峰,等.框镜鲤维氏气单胞菌的生物学特性[J].中国兽医杂志,2012,48(8):18-21.