一株鲈鱼源致病性鰤鱼诺卡氏菌XXLX2的分离鉴定及比较基因组分析

Isolation and identification of a pathogenic Nocardia seriolae strain XXLX2 from seabass and comparative genomic analysis

  • 摘要: 分离、鉴定河南新乡某养殖场大口黑鲈 (Micropterus salmoides) 患病的病原菌,了解分离菌致病性,结合耐药基因注释分析药敏结果,对其基因组结构、毒力因子与近缘菌株进行比较分析,以期寻找共同的免疫保护抗原功能基因。结合菌落形态特征、理化特性分析,并基于16S rRNA基因序列比对鉴定分离菌,对其进行溶血试验、人工回归感染试验、药敏试验及全基因组测序分析。分离菌XXLX2鉴定为鰤鱼诺卡氏菌 (Nocardia seriolae),其在血平板上无溶血圈现象,经回归感染试验计算出其对大口黑鲈的半数致死量 (LD50) 为1.49×105 CFU·mL−1,且被感染鱼与自然患病鱼的症状相符。XXLX2菌株对多粘菌素B、红霉素和β-内酰胺类抗生素显示出抗性,耐药基因和药敏试验结果基本一致。通过基因组比较分析,XXLX2菌株与EM150506、NK201610020和UTF1等3株不同来源的鰤鱼诺卡氏菌具有较近的亲缘关系,且具有较好的共线性;XXLX2菌株与3株鰤鱼诺卡氏菌在毒力因子的比对中存在一定差异,但整体具有较高的保守性。通过对XXLX2菌株进行基因组注释与比较分析,为进一步探索鰤鱼诺卡氏菌致病机制及基因工程疫苗研究提供了基础数据支撑。

     

    Abstract: We isolated and identified the pathogen causing disease in largemouth bass (Micropterus salmoides) from a farm in Xinxiang, and investigated its pathogenicity. We combined the annotation of drug-resistant genes to analyze drug sensitivity results, and compared its genome structure, virulence factors and closely related strains to search for the common immune protective antigen functional genes. By the analyses of colony morphology and physicochemical properties, and based on the identification of isolated bacteria through 16S rRNA gene sequence alignment, we conducted hemolysis test, artificial regression infection test, drug sensitivity test, and whole genome sequencing analysis, then identified the isolated bacteria XXLX2 which was identified as Nocardia seriolae, without hemolysis circle on blood agar plates. The median lethal dose (LD50) for largemouth bass was 1.49×105 CFU·mL−1 through regression infection tests, and the symptoms of infected bass were consistent with those of naturally diseased bass. The XXLX2 strain exhibited resistance to polymyxin B, erythromycin, and β-lactam antibiotics, and the results of drug resistance gene and drug sensitivity test were generally consistent. Through genome comparison analysis, the XXLX2 strain showed close phylogenetic relationship and good collinearity with three strains of N. seriolae from different sources, namely EM150506, NK201610020, and UTF1. There were certain differences in the comparison of virulence factors between the XXLX2 strain and the three strains of N. seriolae, but generally, they exhibited high conservation. Genomic annotation and comparative analysis of the XXLX2 strain provide basic data support for further exploration of the pathogenic mechanism of N. seriolae and research on genetically engineered vaccines.

     

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