
Isolation, Identification and Virulence Gene Analysis of an Aeromonas hydrophila from Diseased Pelodiscus sinensis
Xue Mingyang, Zhou Yong, Liang Hongwei, Li Xiang, Fan Yuding, Zeng Lingbing, Qu Chunjuan, Meng Yan
Isolation, Identification and Virulence Gene Analysis of an Aeromonas hydrophila from Diseased Pelodiscus sinensis
The study aims to determine the pathogen causing disease in a Chinese soft-shelled turtle farm in Anhui Province, and provide reference for effective prevention and control of the disease in future production. The pathogenic bacteria were isolated from the moribund turtle with typical symptoms under aseptic condition, and then were identified by the morphological examination, biochemical analysis, and sequence analysis of bacterial 16S rDNA. The drug susceptibility test was conducted by Kirby-Bauer agar diffusion method, and the virulence genes of the pathogen were analyzed. The results showed that the pathogen was identified as Aeromonas hydrophila, called AM-1. Artificial infection showed that the typical symptoms of the disease could be replicated as the ones naturally occurred and this bacterium was of high pathogenicity to Chinese soft-shelled turtle. The drug resistance tests showed that the isolated pathogen was highly sensitive to enrofloxacin and levofloxacin, while extremely resistant to cotrimoxazole, ampicillin and tetracycline. The amplification results of 6 virulence genes showed that the isolated pathogen carried 5 virulence genes including aerolysin, heat-labile enterotoxin, heat-stable enterotoxin, hemolysin and elastase. This study indicates that Aeromonas hydrophila AM-1 is the pathogenic bacteria which could cause the deadly disease outbreak of the Chinese turtle. Enrofloxacin and levofloxacin can be used for treatment.
Pelodiscus sinensis / Aeromonas hydrophila / virulence gene / pathogenicity {{custom_keyword}} /
[1] |
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[2] |
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[3] |
陆承平. 致病性嗜水气单胞菌及其所致鱼病综述[J]. 水产学报, 1992, 16(3):282-288.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[4] |
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[5] |
尹梦雅, 陈静涛, 黄松钱, 等. 中华鳖3种细菌的鉴定及致病性研究[J]. 中国农学通报, 2019, 35(7):145-152.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[6] |
虞蕴如, 储卫华, 李克敏, 等. 中华鳖红脖子红底板病的病原分离鉴定[J]. 动物医学进展, 2004, 25(2):102-105.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[7] |
郑杰夫, 张一柳, 钟蕾, 等. 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.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[8] |
林锋. 患红底板症的中华鳖幼鳖的病原菌分离与鉴定[J]. 安徽农业科学, 2013, 426(29):11706-11708.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[9] |
赵小平, 汪官余, 郑永华, 等. 中华鳖"烂颈病"病因学的初步研究[J]. 淡水渔业, 2005, 6(13):44-46.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[10] |
钱明明, 宋杰, 武斌, 等. 中华鳖常见疾病的诊断及其综合防治[J]. 动物医学进展, 2009, 30(4):103-107.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[11] |
Aeromonas hydrophila is a gram-negative opportunistic pathogen of animals and humans. The pathogenesis of A. hydrophila is multifactorial. Genomic subtraction and markers of genomic islands (GIs) were used to identify putative virulence genes in A. hydrophila PPD134/91. Two rounds of genomic subtraction led to the identification of 22 unique DNA fragments encoding 19 putative virulence factors and seven new open reading frames, which are commonly present in the eight virulence strains examined. In addition, four GIs were found, including O-antigen, capsule, phage-associated, and type III secretion system (TTSS) gene clusters. These putative virulence genes and gene clusters were positioned on a physical map of A. hydrophila PPD134/91 to determine their genetic organization in this bacterium. Further in vivo study of insertion and deletion mutants showed that the TTSS may be one of the important virulence factors in A. hydrophila pathogenesis. Furthermore, deletions of multiple virulence factors such as S-layer, serine protease, and metalloprotease also increased the 50% lethal dose to the same level as the TTSS mutation (about 1 log) in a blue gourami infection model. This observation sheds light on the multifactorial and concerted nature of pathogenicity in A. hydrophila. The large number of putative virulence genes identified in this study will form the basis for further investigation of this emerging pathogen and help to develop effective vaccines, diagnostics, and novel therapeutics.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[12] |
付乔芳, 邱军强, 胡鲲, 等. 嗜水气单胞菌国内分离株的毒力因子分布与致病性相关性分析[J]. 生物学杂志, 2011, 28(6):53-57.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[13] |
赵增连. 甲鱼虹彩病毒(STIV)病原学研究及检测方法的建立[D]. 武汉:华中农业大学, 2008.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[14] |
A set of oligonucleotide primers capable of initiating enzymatic amplification (polymerase chain reaction) on a phylogenetically and taxonomically wide range of bacteria is described along with methods for their use and examples. One pair of primers is capable of amplifying nearly full-length 16S ribosomal DNA (rDNA) from many bacterial genera; the additional primers are useful for various exceptional sequences. Methods for purification of amplified material, direct sequencing, cloning, sequencing, and transcription are outlined. An obligate intracellular parasite of bovine erythrocytes, Anaplasma marginale, is used as an example; its 16S rDNA was amplified, cloned, sequenced, and phylogenetically placed. Anaplasmas are related to the genera Rickettsia and Ehrlichia. In addition, 16S rDNAs from several species were readily amplified from material found in lyophilized ampoules from the American Type Culture Collection. By use of this method, the phylogenetic study of extremely fastidious or highly pathogenic bacterial species can be carried out without the need to culture them. In theory, any gene segment for which polymerase chain reaction primer design is possible can be derived from a readily obtainable lyophilized bacterial culture.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[15] |
王艺, 刘文珍, 傅松哲, 等. 一株从草鱼中分离的嗜水气单胞菌的病原学及基因组特征[J]. 水产学报, 2019, 43(4):480-490.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[16] |
徐阳, 姚洪, 乔帼, 等. 养殖大菱鲆感染嗜水气单胞菌的分离,毒力分析及ERIC-PCR分型[J]. 大连海洋大学学报, 2018, 33(4):430-434.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[17] |
李梦茜, 张振旺. 罗非鱼细菌性败血症病原菌嗜水气单胞菌拮抗菌株的筛选和鉴定[J]. 江苏农业科学, 2017, 45(3):129-131.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[18] |
孙浩, 郑礼洲, 魏伟, 等. 大鲵水肿病病原的分离鉴定及生物学特性研究[J]. 江西农业大学学报, 2014, 1(30):193-198.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[19] |
朱越雄, 曹广力, 薛仁宇, 等. 中华绒螯蟹致病性嗜水气单胞菌的鉴定及特性[J]. 苏州大学学报:自然科学版, 2002. 18(3):100-106.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[20] |
赵明军, 朱芝秀, 王小英, 等. 中华鳖嗜水气单胞菌所致疾病研究[J]. 中国饲料, 2003. 7(15):26-27.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[21] |
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[22] |
邱德全, 何建国, 钟英长, 等. 嗜水气单胞菌的致病性研究[J]. 中山大学学报论丛, 1996, S1(1):83-90.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[23] |
赵宇亮, 吕梦溪, 苏欣, 等. 中华鳖病原维氏气单胞菌的分子鉴定及分子分类学研究[J]. 河北师范大学学报:自然科学版, 2011, 35(4):417-422.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[24] |
李文艳, 孙霞. 环境因子对温室养殖中华鳖嗜水气单胞菌病发生的影响[J]. 科技信息, 2011, 1(10):128-128.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[25] |
孟庆珍, 尹飞, 傅超英, 等. 养殖中华鳖蜡样芽孢杆菌的分离鉴定和致病性研究[J]. 水生生物学报, 2019, 43(3):571-578.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[26] |
康绍珠, 朱芝秀, 黄志琴, 等. 甲鱼嗜水气单胞菌的分离鉴定及体外抑菌试验[J]. 养殖与饲料, 2019, 1(5):11-15.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[27] |
王帅兵, 熊良伟, 朱善元, 等. 嗜水气单胞菌分离鉴定与药敏试验[J]. 江苏农业科学, 2017, 45(24):165-168.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[28] |
邱军强, 杨先乐, 程训佳, 等. 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.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
[29] |
李巧玲, 张召兴, 吴同垒, 等. 貉源嗜水气单胞菌的分离及其生物学特性鉴定[J]. 中国兽医学报, 2018, 38(9):1740-1744.
{{custom_citation.content}}
{{custom_citation.annotation}}
|
{{custom_ref.label}} |
{{custom_citation.content}}
{{custom_citation.annotation}}
|
/
〈 |
|
〉 |