溶藻弧菌群体基因组学研究

doi:10.16288/j.yczz.21-061

遗传 ›› 2021, Vol. 43 ›› Issue (4): 350-361.doi: 10.16288/j.yczz.21-061

溶藻弧菌群体基因组学研究

郑宏源¹, 闫琳², 杨超^1,³, 武雅蓉¹, 秦婧靓¹, 郝彤宇¹, 杨大进², 郭云昌², 裴晓燕², 赵彤言¹, 崔玉军¹()

^1. 军事科学院军事医学研究院微生物流行病研究所,北京 100071
^2. 国家食品安全风险评估中心,北京 100022
^3. 深圳市疾病预防控制中心,深圳 518055

收稿日期:2021-02-09 修回日期:2021-03-16 出版日期:2021-04-20 发布日期:2021-04-20
通讯作者: 崔玉军 E-mail:cuiyujun.new@gmail.com
作者简介:郑宏源,在读硕士研究生,专业方向：细菌基因组流行病学。E-mail: zhenghongyuan.chn@qq.com
基金资助:
国家重点研发计划编号(2018YFC1603902);国家重点研发计划编号(2017YFC1601500);中国博士后科学基金项目资助编号(2020M672836)

Population genomics study of Vibrio alginolyticus

Hongyuan Zheng¹, Lin Yan², Chao Yang^1,³, Yarong Wu¹, Jingliang Qin¹, Tongyu Hao¹, Dajin Yang², Yunchang Guo², Xiaoyan Pei², Tongyan Zhao¹, Yujun Cui¹()

^1. State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing 100071, China
^2. National Center for Food Safety Risk Assessment, Beijing 100022, China
^3. Shenzhen Center for Disease Control and Prevention, Shenzhen 518055, China

Received:2021-02-09 Revised:2021-03-16 Online:2021-04-20 Published:2021-04-20
Contact: Cui Yujun E-mail:cuiyujun.new@gmail.com
Supported by:
Supported by the National Key R&D Program Nos of China(2018YFC1603902);Supported by the National Key R&D Program Nos of China(2017YFC1601500);China Postdoctoral Science Foundation No(2020M672836)

摘要/Abstract

摘要：

溶藻弧菌(Vibrio alginolyticus)是一种能够对人类以及鱼、虾、贝类等水产品致病的弧菌,给人类健康带来威胁,也给水产养殖业造成巨大的经济损失。目前该物种基于全基因组的遗传多样性和重要遗传元件研究报道较少。本研究对采集自全国4个省份的68株溶藻弧菌进行高通量测序,获得全基因组序列,并结合113株公开发表的全球序列数据,利用fineSTRUCTURE软件、VFDB毒力因子库和CARD、ResFinder耐药数据库,对溶藻弧菌的种群结构和毒力、耐药因子分别进行解析。结果表明：溶藻弧菌可分为谱系1和谱系2。两个谱系在美洲和亚洲均有分布,但欧洲仅分离到谱系1菌株;共鉴定发现12个克隆群,其中一个克隆群内菌株存在跨洋传播现象。该物种携带tlh、OmpU、IlpA等多种不同功能的毒力因子;毒力因子在两个谱系间的分布无特异性,但存在地域间差异：其中欧洲菌株携带VP1611、vcrD、vopD和fleR/flrC的比率低于其他地区,而基因IlpA的携带率则明显高于其他地区,我国广西菌株中fleR/flrC基因携带率低于其他省份,且不携带IlpA。多个基因组携带blaCARB-42、tet(34)、tet(35)、parE、CRP、rsmA、TxR和fos等与多种抗生素耐受相关的基因,其中TxR和fos基因在谱系2中的出现频率远高于谱系1;此外,TxR基因在亚洲菌株中的携带率高于美洲和欧洲地区,而在我国四川菌株中的携带率则低于其他省份。在5个基因组中(VA24、VA28、2014V-1011、ZJ-T和Vb1833)观察到质粒或ICE等携带多种耐药基因的大片段。本研究通过群体基因组学的研究方法,揭示了溶藻弧菌的种群结构组成和毒力、耐药相关元件的分布,为进一步了解溶藻弧菌的遗传特征和致病机制提供必要基础,为该病原的监测、预防和控制工作提供科学支撑。

关键词: 溶藻弧菌, 全基因组测序, 种群结构, 毒力因子, 耐药基因

Abstract:

Vibrio alginolyticus is a Gram-negative bacillus that causes vibriosis to human and aquatic products, including fish, shrimp and shellfish. It poses a threat to public health and causes enormous economic losses to the aquaculture industry. However, research on genetic diversity and pathogenicity-related genetic elements based on whole genome is still lacking. In this study, sixty-eight strains of V. alginolyticus were collected from four provinces of China and the whole genome sequences were obtained. Combined with 113 publicly available genome sequences downloaded from NCBI, we inferred the population structure of V. alginolyticus by using fineSTRUCTURE software, and identified the virulence and antibiotic resistance factors using the VFDB, CARD and ResFinder database. The results indicated that V. alginolyticus included two main lineages, named Lineage 1 and Lineage 2. Both lineages distributed in America and Asia, but all the European genomes were classified into Lineage 1. A single cross-ocean transmission event was inferred from one of the 12 identified clonal groups in our dataset. V. alginolyticus genome contains a variety of virulence factors, such as tlh, OmpU, and IlpA, etc. The distribution of virulence factors revealed no lineage-specificity, but some of which revealed differences in their geographical distribution. A lower frequency of VP1611, vcrD, vopD, fleR/flrC and a higher frequency of IlpA were observed in genomes of Europe than other continents. In China, a lower frequency of fleR/flrC, and no IlpA were observed in genomes from Guangxi province. Among the identified antibiotic resistance genes, TxR and fos are significantly enriched in Lineage 2. In addition, TxR is more common in genomes from Asia, compared with the American and European genomes. But in China, the frequency of TxR in Sichuan genomes is much lower than in other provinces. We also found that large fragments of plasmids or ICEs that carried multiple drug resistance genes were present in five V. alginolyticus genomes (VA24, VA28, 2014V-1011, ZJ-T and Vb1833). Based on population genomics analysis, our study delineated the population structure, distribution of virulence and antibiotic resistance related factors of V. alginolyticus, which lays a foundation for future study of genetic characters and pathogenesis mechanism of this pathogen and will improve the works on monitoring, prevention and control of this pathogen.

Key words: Vibrio alginolyticus, whole genome sequencing, population structure, virulence factor, antimicrobial resistance gene

郑宏源, 闫琳, 杨超, 武雅蓉, 秦婧靓, 郝彤宇, 杨大进, 郭云昌, 裴晓燕, 赵彤言, 崔玉军. 溶藻弧菌群体基因组学研究[J]. 遗传, 2021, 43(4): 350-361.

Hongyuan Zheng, Lin Yan, Chao Yang, Yarong Wu, Jingliang Qin, Tongyu Hao, Dajin Yang, Yunchang Guo, Xiaoyan Pei, Tongyan Zhao, Yujun Cui. Population genomics study of Vibrio alginolyticus[J]. Hereditas(Beijing), 2021, 43(4): 350-361.

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	中国	美洲	亚洲其他国家	欧洲	其他	总数
谱系1	48^*(35%^#)	58^*(42%^#)	6^*(4%^#)	17^*(12%^#)	8^*(6%^#)	137
谱系2	28^*(64%^#)	10^*(23%^#)	3^*(7%^#)	0^*	3^*(7%^#)	44

相关毒力基因	毒力因子编号	毒力因子名称	毒力因子功能
flgD(100%) flgC(100%) flgB(100%) fliM(100%) fliG(100%) cheR(99%) cheW(99%) cheY(99%) flgF(99%) cheV(94%) flgE(84%) flgG(49%) fleR/flrC(45%) flhA(1%)	VF0519	Flagella	Adhesin factor
vcrH(100%) vopB(99%) vscI(99%) exsD(99%) exsA(99%) vopQ(99%) vxsC(98%) vscD(98%) tyeA(97%) vscO(97%) vscU(90%) vcrD(86%) vscQ(82%) vopD(76%) vscN(2%) vscC(1%)	VF0408	T3SS1	Rapid apoptosis, Cell rounding, Osmotic lysis
tlh(100%)	TX170	Thermolabile_hemolysin	Apoptosis, Membrane vesiculation, Necrosis
OmpU(98%)	VF0514	OmpU	Porin, Iron balance, Vaccine candidate
VP1611(87%)	VF0512	MAM7	Multivalent adhesion molecule MAM7
IlpA(26%)	VF0513	IlpA	Immunogenic lipoprotein A
htpB(9%)	VF0159	Hsp60	Heat shock protein HtpB
ugd(1%)	VF0560	Capsule	UDP-glucose 6-dehydrogenase
hcp-2(1%)	SS181	VAS	T6SS substrate Hcp-2

耐药基因	耐受抗生素	耐药机制
blaCARB-42(99%) blaVEB-1(1%)	Beta-lactam resistance	Antibiotic inactivation
E.coli_parE(100%) QnrVC1(6%) QnrA1(1%) QnrS5(1%) QnrVC5(1%)	Quinolone resistance	Antibiotic target alteration (parE) Antibiotic target protection (qnr)
tet(35)(100%) tet(34)(98%) TxR(34%) tet(59)(1%)	Tetracycline resistance	Antibiotic efflux
ANT(2'')-Ia(2%) APH(3'')-Ib(2%) APH(6)-Id(2%) ANT(3'')-IIa(1%) APH(3')-Ia(1%)	Aminoglycoside antibiotic	Antibiotic inactivation
fos(5%)	Fosfomycin resistance	Antibiotic inactivation
sul1(2%) sul2(2%)	Sulfonamide antibiotic	Antibiotic target replacement
cmlA5(1%) floR(1%) catA3(1%)	Phenicol antibiotic	Antibiotic efflux (cmlA5, floR) Antibiotic inactivation (catA3)
Ere(A)(1%) mph(A)(1%)	Macrolide antibiotic	Antibiotic inactivation
arr-2(1%)	Rifamycin antibiotic	Antibiotic inactivation
dfrA15(1%) dfrA31(1%)	Trimethoprim resistant	Antibiotic target replacement
BRP(MBL)(1%)	Glycopeptide antibiotic	Antibiotic inactivation
CRP(100%) rsmA(98%)	ARG_regulator	Antibiotic efflux
qacEdelta1(2%)	Quaternary ammonium compounds	Antibiotic efflux

溶藻弧菌群体基因组学研究

Population genomics study of Vibrio alginolyticus

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	VP1611 (%)	vcrD (%)	flgE (%)	vscQ (%)	vopD (%)	flgG (%)	fleR/flrC (%)	IlpA (%)
谱系1	85	82	89	77	77	45	45	29
谱系2	93	95	68	98	73	61	43	16
中国	92	89	87	89	78	53	53	20
美洲	93	91	88	79	81	47	44	29
亚洲其他国家	89	89	56	78	89	44	44	0
欧洲	47	41	88	88	35	65	18	65
山东	90	90	77	95	74	56	62	21
湖北	100	100	94	76	82	47	41	24
四川	71	86	100	86	100	43	86	14
广西	100	80	100	100	60	40	20	0