中华鳖源按蚊伊丽莎白菌的分离鉴定及药敏特性研究

ISOLATION, IDENTIFICATION AND ANTIMICROBIAL SUSCEPTIBILITY OF ELIZABETHKINGIA ANOPHELIS DERIVED FROM TRIONYX SINENSIS

  • 摘要: 研究对从江西省抚州市南丰县某中华鳖养殖基地患病中华鳖(Trionyx sinensis)组织中分离培养得到的一株按蚊伊丽莎白菌(Elizabethkingia anophelis)疑似菌株进行鉴定和药敏特性研究。在无菌环境中解剖患病中华鳖得到肝脏、脾脏组织, 匀浆后接种在LB培养基上, 分离得到优势菌株EJX2021。对该菌株进行形态学观察、16S RNA基因序列测定和生理生化鉴定以鉴定菌株种类; 进行人工回归感染、组织病理学观察、毒力因子分析和抗生素药物敏感性试验研究其致病能力和药物敏感性特征。从患病中华鳖中分离出一株优势菌株EJX2021, 结合其表型特征、16S RNA基因序列测定和生理生化鉴定, 确定其为按蚊伊丽莎白菌。在人工回归感染后, 该菌株对中华鳖15d的LD50为1.62×108 CFU/mL, 95%置信区间为(0.78—3.37)×108 CFU/mL; 组织病理学观察肠道、肺脏、脾脏、肝脏等组织出现不同程度病变。毒力因子分析显示该菌株包含246个潜在毒力基因。药物敏感性试验显示该菌株为多重耐药菌株, 对多西环素、米诺环素、卡那霉素、哌拉西林、恩诺沙星、氟苯尼考敏感, 对头孢曲松、头孢哌酮中度敏感, 对新霉素等15种抗生素具有耐药性。研究证明了按蚊伊丽莎白菌是能够导致中华鳖病害的一种病原菌, 为该细菌导致中华鳖病害的检测、鉴别和防治提供科学依据。

     

    Abstract: Elizabethkingia anophelis has been recognized as an important opportunistic pathogen in human and animal infection in recent years. The study represents the first successful isolation of this bacterium from Trionyx sinensis. The objective of this study was to isolate and culture a suspected strain of Elizabethkingia anophelis from infected Trionyx sinensis, identified the strain, and analyze its antimicrobial susceptibility characteristics. In our study, liver and spleen tissues were obtained from diseased Trionyx sinensis in aseptic environment. These tissues were ground and cultured in LB medium to obtain the target strain EJX2021. Identification of the strain involved morphological observations, determination of the 16sRNA gene sequence, and physiological and biochemical identification. Subsequently, artificial regression infection, histopathological observation, virulence factor analysis and antibiotic susceptibility test were performed to study the pathogenic ability and antimicrobial susceptibility characteristics. We successfully isolated a dominant strain EJX2021, from the diseased Trionyx sinensis. It was identified as Elizabethkingia anophelis based on phenotypic characteristics, 16sRNA gene sequence, and physiological and biochemical identification. The results of artificial regression infection showed that LD50 of the strain against Trionyx sinensis at 15d was 1.62×108 CFU/mL, with a 95% confidence interval of 0.78—3.37 ×108 CFU/mL. Histopathology analysis revealed different degrees of pathological changes in the intestines, lungs, spleen, liver and other tissues. Virulence factor analysis showed that the strain contained 246 potential virulence genes in 16 categories. In terms of antimicrobial susceptibility, the strain exhibited multi-drug resistance. It was sensitive to doxycycline, minocycline, kanamycin, piperacillin, enrofloxacin and florfenicol, moderately sensitive to ceftriaxone and cefoperazone, and resistant to 15 antibiotics such as neomycin. Our study proved that Elizabethkingia anophelis can cause the disease of Trionyx sinensis, and provided scientific basis for the detection, identification and control of the Trionyx sinensis disease caused by the bacteria.

     

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