羊源产酸克雷伯菌的分离鉴定及感染小鼠的转录组学研究

doi:10.16431/j.cnki.1671-7236.2023.10.004

摘要/Abstract

摘要： 【目的】分离鉴定羊源产酸克雷伯菌,探讨产酸克雷伯菌对动物的致病机制。【方法】采集海南黑山羊肺脏组织样品进行细菌分离培养,通过生化试验及16S rRNA基因检测方法对分离株进行鉴定,并对分离菌进行药敏试验、小鼠致病性试验、RNA-Seq分析和差异表达基因(DEGs)分析。【结果】分离出1株革兰阴性短杆状细菌,胰蛋白胨大豆琼脂培养基上为光滑圆形的乳白色菌落。分离菌能分解葡萄糖、乳糖、蔗糖等几种碳水化合物,靛基质试验阳性。16S rRNA基因检测结果显示,分离株与已公布序列的相似性＞99%,鉴定该分离菌为产酸克雷伯菌,命名为KOHN1。药敏试验结果显示,分离菌KOHN1对青霉素、苯唑青霉素、氨苄青霉素、羧苄青霉素、头孢氨苄、麦迪霉素、万古霉素和克林霉素具有耐药性,而对于其余21种抗菌药敏感。分离菌KOHN1攻毒小鼠后,肺脏组织病理学切片观察到小鼠肺部充血、水肿和渗出等急性炎症。DEGs富集分析显示,炎症细胞因子和趋化因子的DEGs在小鼠肺脏中大量表达。【结论】本试验成功分离出1株产酸克雷伯菌,分离株对小鼠致病性较强,免疫反应通路在病菌侵染过程中发挥着重要作用。该研究结果为后续产酸克雷伯菌的临床预防、诊疗及药物研发提供了参考依据。

关键词: 海南黑山羊; 产酸克雷伯菌; 分离鉴定; RNA-Seq; 免疫机制

Abstract: 【Objective】 The aim of this study was to isolate and identify a strain of Klebsiella oxytoca from goat and investigate the pathogenic mechanism of Klebsiella oxytoca in animals.【Method】 The lung tissue samples were collected from Hainan Black goats for bacterial isolation and culture,and the isolates were identified by biochemical test and 16S rRNA gene detection methods,and drug sensitivity test,mouse pathogenicity test, RNA-Seq analysis,and differentially expressed genes (DEGs) analysis on the isolated bacteria were performed.【Result】 One Gram-negative short rod-shaped bacterium was isolated with a smooth round milky colony on tryptic soybean agar medium.The isolated bacteria could decompose glucose,lactose,sucrose and other carbohydrates,and the indole test was positive.16S rRNA genetic testing results showed that the similarity between the isolate and the published sequence was ＞99%, and the isolated strain was identified as Klebsiella oxytoca,which was named KOHN1.The drug susceptibility test results showed that KOHN1 strain was resistant to penicillin,oxacillin,ampicillin,carbenicillin,cephalexin,midecamycin,vancomycin and clindamycin,while it was susceptible to the other 21 antimicrobial agents tested.Acute inflammation such as pulmonary congestion,edema and exudation were observed in histopathological sections of the lungs of mice infected with the isolated strain KOHN1.DEGs enrichment analysis showed that DEGs of inflammatory cytokines and chemokines were highly expressed in lung of mice.【Conclusion】 In this experiment,a strain of Klebsiella oxytoca was successfully isolated,the isolate was highly pathogenic to mice,and the immune response pathway played an important role in the process of bacterial infection,which provided a reference for the subsequent clinical prevention,diagnosis and treatment and drug development of Klebsiella oxytoca.

Key words: Hainan Black goats; Klebsiella oxytoca; isolation and identification; RNA-Seq; immune mechanism

中图分类号:

S852.61

李旭博, 蒋俊明, 何美荣, 李昊洋, 李虹, 陈思, 陈巧玲, 满初日嘎, 杜丽, 王凤阳, 高宏岩. 羊源产酸克雷伯菌的分离鉴定及感染小鼠的转录组学研究[J]. 中国畜牧兽医, 2023, 50(10): 3906-3916.

LI Xubo, JIANG Junming, HE Meirong, LI Haoyang, LI Hong, CHEN Si, CHEN Qiaoling, MAN Churiga, DU Li, WANG Fengyang, GAO Hongyan. Isolation,Identification of Klebsiella oxytoca from Goat and Transcriptomics Study on Infected Mice[J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(10): 3906-3916.

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