中华鳖(Pelodiscus sinensis)迟缓爱德华氏菌(Edwardsiella tarda)的分离鉴定与药物敏感性分析
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摘要
目前,细菌性疾病是中华鳖养殖中的常见疾病。实验采用常规方法从呈白底板症状的中华鳖(Pelodiscus sinensis)肝脏、脾脏、肾脏、胃积液等中分离纯化病原菌,经形态观察、生理生化鉴定和16S r RNA序列分析确定其种属,通过回归感染实验判断分离菌株致病性,并以纸片扩散法测定其药物敏感性。结果显示:自病鳖中分离可得到一种优势菌落,其表面湿润光滑、微隆起、半透明,呈灰黄色;基于VITEK 2的64个生化反应的分析提示,分离菌理化特性与迟缓爱德华氏菌(Edwardsiella tarda)基本一致;16S rRNA序列分析显示,其与迟缓爱德华氏菌同源性达98%以上。腹腔注射回归感染实验引起稚鳖死亡,且细菌分离能再次得到相同分离株,表明分离到的迟缓爱德华氏菌具有致病性。药敏试验表明该菌对庆大霉素、阿米卡星、新霉素、头孢哌酮、氨曲南等5种抗生素较为敏感。本研究可为中华鳖病害的正确诊断和科学防治提供参考。
The disease affected by bacteria is regular in culture of Pelodiscus Sinensis. The pathogenic bacteria were isolated from liver,kidney,spleen and gastric fluid of the diseased Pelodiscus sinensis with white abdominal shell. The species of the isolated strain was identified according to the morphological observation,physiological and biochemical characteristics identification and 16 S rRNA sequences analysis. Then,an artificial infection experiment was carried out to identify its pathogenicity. Kirby-Bauer method was used for antibiotic susceptibility test. The results showed that the bacterial strain isolated,its surface being evidenced by moist,smooth and slightly raised,translucent and grayish-yellow color,was finally identified as Edwardsiella tarda( E. tarda) according to 64 biochemical reaction of VITEK 2 and more than 98% homologous 16 S rRNA gene sequences. In artificial infection test by injection,the isolate led to the death of juvenile turtles,and the bacterium was recovered from these artificially infected animals. The antibiotic susceptibility test revealed that the E. tarda was sensitive to gentamicin,amikacin,neomycin,cefoperazone and aztreonam. The study will provide references for correct diagnosis and scientific control of the disease in Pelodiscus sinensis.
The disease affected by bacteria is regular in culture of Pelodiscus Sinensis. The pathogenic bacteria were isolated from liver,kidney,spleen and gastric fluid of the diseased Pelodiscus sinensis with white abdominal shell. The species of the isolated strain was identified according to the morphological observation,physiological and biochemical characteristics identification and 16 S rRNA sequences analysis. Then,an artificial infection experiment was carried out to identify its pathogenicity. Kirby-Bauer method was used for antibiotic susceptibility test. The results showed that the bacterial strain isolated,its surface being evidenced by moist,smooth and slightly raised,translucent and grayish-yellow color,was finally identified as Edwardsiella tarda( E. tarda) according to 64 biochemical reaction of VITEK 2 and more than 98% homologous 16 S rRNA gene sequences. In artificial infection test by injection,the isolate led to the death of juvenile turtles,and the bacterium was recovered from these artificially infected animals. The antibiotic susceptibility test revealed that the E. tarda was sensitive to gentamicin,amikacin,neomycin,cefoperazone and aztreonam. The study will provide references for correct diagnosis and scientific control of the disease in Pelodiscus sinensis.
引文
[1]吴惠仙.中华鳖六株病原菌的分类鉴定及其中草药拮抗研究[J].中国预防兽医学报,2004,26(5):372-375.
[2]沈锦玉,尹文林,钱冬,等.中华鳖细菌性疾病免疫防治的研究[J].浙江大学学报(农业与生命科学版),2000,26(3):325-328.
[3]郑杰夫,张一柳,钟蕾.中华鳖源嗜水气单胞菌的研究进展[J].水产科学,2016,35(2):191-198.
[4]林峰,刘莉,曹铮,等.鳖源致病性嗜水气单胞菌的分离与鉴定[J].湖北农业科学,2014,53(22):5487-5489.
[5]马有智,舒妙安.中华鳖温和气单胞菌病的病原研究[J].浙江大学学报(理学版),2000,27(3):329-333.
[6]蔡完其,孙佩芳,刘至治.中华鳖爱德华氏菌病病原和组织病理研究[J].水产学报,1997,21(4):428-433.
[7]潘晓艺,郝贵杰,姚嘉赘,等.中华鳖爱德华菌病病原菌的分离鉴定及致病因子研究[J].淡水渔业,2010,40(6):40-45.
[8]胡广洲,李登峰,苏秀榕,等.患暴发性败血症中华鳖体内细菌的分离与鉴定[J].中国水产科学,2010,17(4):859-868.
[9]林启存,朱丽敏,李忠全,等.中华鳖弗氏柠檬酸杆菌败血症病原分离鉴定与药敏试验[J].水产科学,2008,27(1):42-43.
[10]沈锦玉,潘晓艺,余旭平,等.中华鳖白底板病病原的分析[J].中国水产科学,2007,14(5):815-822.
[11]Bauer A W,Kirby W M,Sherris J C,et al.Antibiotic susceptibility testing by a standardized single disk method.[J].Am J Clin Pathol,1966,45:493-496.
[12]Janda J M,Abhott S L.Infections associated with the genus Edwardsiella:the role of Edwardsiella tarda in human disease[J].Clinical infectious Dis,1993,17(4):742-748.
[13]马勋,欧阳志明,陈怀青,等.迟缓爱德华氏菌对Hep-2细胞的侵袭特性[J].微生物学报,1998,38(5):336-340.
[14]陈强,龚晖,杨金先.欧洲鳗迟钝爱德华氏菌的分离鉴定[J].中国人兽共患病学报,2011,27(1):7-10.
[15]陈言峰,周爱国,邹记兴,等.乌鳢溃烂病病原分析[J].水产科技情报,2015,42(4):213-216.
[16]邓威,徐赟霞,赵良炜,等.大菱鲆造血器官肿大病的病原分离、鉴定及药物敏感性[J].天津农业科学,2015,21(6):45-49.
[17]朱成科,叶华,郑曙明,等.中华倒刺鲃迟缓爱德华菌的分离鉴定[J].淡水渔业,2013,43(3):61-65.
[18]程俊茗,万明月,周晋扬,等.鲫源迟缓爱德华氏菌的分离鉴定及其毒力基因的检测[J].微生物学通报,2017,44(10):2380-2390.
[19]江为民,王宇,康惠,等.中华鳖爱德华氏菌病的病原与防治研究[J].内陆水产,2000(11):11-13.
[20]陈晓凤,周常义,青新.鳖“白板病”致病细菌的研究[J].水产学报,1997,21(3):309-315.
[21]王鸿泰,何力,张俊.饲料质量与中华鳖白底板病的关系[J].淡水渔业,2002,32(2):32-33.
[22]杨先乐.中华鳖疾病的防治[J].科学养鱼,1999(5):20-22.
[23]孔蕾,朱凝瑜,贝亦江,等.中华鳖摩氏摩根菌的鉴定及致病性研究[J].海洋与湖沼,2013,44(3):722-727.
[2]沈锦玉,尹文林,钱冬,等.中华鳖细菌性疾病免疫防治的研究[J].浙江大学学报(农业与生命科学版),2000,26(3):325-328.
[3]郑杰夫,张一柳,钟蕾.中华鳖源嗜水气单胞菌的研究进展[J].水产科学,2016,35(2):191-198.
[4]林峰,刘莉,曹铮,等.鳖源致病性嗜水气单胞菌的分离与鉴定[J].湖北农业科学,2014,53(22):5487-5489.
[5]马有智,舒妙安.中华鳖温和气单胞菌病的病原研究[J].浙江大学学报(理学版),2000,27(3):329-333.
[6]蔡完其,孙佩芳,刘至治.中华鳖爱德华氏菌病病原和组织病理研究[J].水产学报,1997,21(4):428-433.
[7]潘晓艺,郝贵杰,姚嘉赘,等.中华鳖爱德华菌病病原菌的分离鉴定及致病因子研究[J].淡水渔业,2010,40(6):40-45.
[8]胡广洲,李登峰,苏秀榕,等.患暴发性败血症中华鳖体内细菌的分离与鉴定[J].中国水产科学,2010,17(4):859-868.
[9]林启存,朱丽敏,李忠全,等.中华鳖弗氏柠檬酸杆菌败血症病原分离鉴定与药敏试验[J].水产科学,2008,27(1):42-43.
[10]沈锦玉,潘晓艺,余旭平,等.中华鳖白底板病病原的分析[J].中国水产科学,2007,14(5):815-822.
[11]Bauer A W,Kirby W M,Sherris J C,et al.Antibiotic susceptibility testing by a standardized single disk method.[J].Am J Clin Pathol,1966,45:493-496.
[12]Janda J M,Abhott S L.Infections associated with the genus Edwardsiella:the role of Edwardsiella tarda in human disease[J].Clinical infectious Dis,1993,17(4):742-748.
[13]马勋,欧阳志明,陈怀青,等.迟缓爱德华氏菌对Hep-2细胞的侵袭特性[J].微生物学报,1998,38(5):336-340.
[14]陈强,龚晖,杨金先.欧洲鳗迟钝爱德华氏菌的分离鉴定[J].中国人兽共患病学报,2011,27(1):7-10.
[15]陈言峰,周爱国,邹记兴,等.乌鳢溃烂病病原分析[J].水产科技情报,2015,42(4):213-216.
[16]邓威,徐赟霞,赵良炜,等.大菱鲆造血器官肿大病的病原分离、鉴定及药物敏感性[J].天津农业科学,2015,21(6):45-49.
[17]朱成科,叶华,郑曙明,等.中华倒刺鲃迟缓爱德华菌的分离鉴定[J].淡水渔业,2013,43(3):61-65.
[18]程俊茗,万明月,周晋扬,等.鲫源迟缓爱德华氏菌的分离鉴定及其毒力基因的检测[J].微生物学通报,2017,44(10):2380-2390.
[19]江为民,王宇,康惠,等.中华鳖爱德华氏菌病的病原与防治研究[J].内陆水产,2000(11):11-13.
[20]陈晓凤,周常义,青新.鳖“白板病”致病细菌的研究[J].水产学报,1997,21(3):309-315.
[21]王鸿泰,何力,张俊.饲料质量与中华鳖白底板病的关系[J].淡水渔业,2002,32(2):32-33.
[22]杨先乐.中华鳖疾病的防治[J].科学养鱼,1999(5):20-22.
[23]孔蕾,朱凝瑜,贝亦江,等.中华鳖摩氏摩根菌的鉴定及致病性研究[J].海洋与湖沼,2013,44(3):722-727.