2017年安徽省马鞍山市环境来源气单胞菌的分子特征和耐药分析

汪永禄; 黄振洲; 王利; 王蓉; 代航; 蔡红艳; 杜小莉; 王多春

doi:10.3784/j.issn.1003-9961.2019.09.008

2017年安徽省马鞍山市环境来源气单胞菌的分子特征和耐药分析

doi: 10.3784/j.issn.1003-9961.2019.09.008

1.
马鞍山市疾病预防控制中心，安徽马鞍山 243000
2.
中国疾病预防控制中心传染病预防控制所，北京 102206

基金项目: 马鞍山市卫计委科教专项资金（No.2016-19）

详细信息

作者简介:
汪永禄，男，安徽省马鞍山市人，副主任技师，主要从事病原菌检测相关工作，Email：mas_wyl@163.com

通讯作者:
王多春，Tel：010–58900744，Email：wangduochun@icdc.cn

中图分类号: R378
计量
- 文章访问数: 11140
- HTML全文浏览量: 2578
- PDF下载量: 37
- 被引次数: 0
出版历程
- 收稿日期: 2019-05-06
- 网络出版日期: 2019-08-13
- 刊出日期: 2019-09-01

Molecular characteristics and drug resistance of Aeromonas from environmental Sources in Ma'anshan, Anhui

1.
Maanshan Center for Disease Control and Prevention, Maanshan 243000, Anhui, China
2.
National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention. Beijing 102206, China

Funds: This study was supported by the Special Fund for Science and Education of Health and Family Planning Commission of Maanshan City (No.2016-19)

More Information

Corresponding author: Wang Duochun, Email: wangduochun@icdc.cn

摘要

摘要: 目的了解2017年安徽省马鞍山市环境来源气单胞菌的流行特征、毒力基因特征及耐药状况。方法2017年对来自马鞍山市监测的海水产品、包装食品、生活用水、游泳池水样、食物中毒样本等进行气单胞菌分离检测，采用管家基因rpoD序列分析进行种水平鉴定，PCR检测气单胞菌6种毒力基因（Fla、Elastase、hlyA、ast、act和ascF-G）、脉冲场凝胶电泳（PFGE）分型分析分子型别，药敏试验检测药物敏感性。结果共检测样品869份，气单胞菌检出率为7.6%（66/869），检出率最高的是农村生活用水、食物中毒样品和海水产品。 66株气单胞菌分为7个种，最多的种是维氏气单胞菌（36.4%，24/869）、嗜水气单胞菌（19.7%，13/869）和简氏气单胞菌（19.7%，13/869）。 64株气单胞菌携带至少1种毒力基因，Fla和act总检出率最高（53.1%），Fla在豚鼠气单胞菌中检出率最高（83.3%），act在嗜水气单胞菌检出率最高（69.2%）。 66株气单胞菌分为64个PFGE带型，表现为多样性。药敏实验显示，气单胞菌对头孢替坦等7种抗生素100%敏感，对头孢唑啉等5种抗生素部分敏感（耐药率1.5% ~ 21.2%），而对氨苄西林、氨苄西林/舒巴坦的耐药率均高达90.0%。结论2017年马鞍山地区气单胞菌污染较为严重，携带较高比例的多种毒力基因，并且对多种抗生素耐药性较高，应该加强对气单胞菌及其耐药性的监测。
- 气单胞菌 /
- 分子特征 /
- 药物敏感性
Abstract: ObjectiveTo understand the epidemiological characteristics, virulence gene characteristics and drug resistance of Aeromonas from environmental sources in Ma’anshan of Anhui province.MethodsFrom January to December, 2017, Aeromonas strains isolated from seawater products, packaged foods, domestic water, swimming pool water and food poisoning samples in Ma'anshan were detected. The species of Aeromonas were identified through housekeeping gene rpoD sequence analysis. PCR was used to detect six virulence genes (Fla, Elastase, hlyA, ast, act and ascF-G), the molecular types of the strains were detected by using pulsed-field gel electrophoresis (PFGE), and drug susceptibility test of the strains was also conducted.ResultsA total of 66 Aeromonas strains were isolated from 869 samples (7.6%), and the isolated rates were higher in rural domestic water, food poisoning samples and seafood products respectively. Sixty-six strains of Aeromonas belonged to seven species, mainly including A. veronii (24, 36.4%), A. hydrophila (13, 19.7%) and A. jandaei (13, 19.7%) respectively. 64 strains of Aeromonas (97.0%) carried one or more virulence genes, with the highest detection rate of Fla and act (53.1%), Fla was mainly detected in A. caviae (83.3%) and act was mainly detected in A. hydrophila (69.2%). Sixty six strains of Aeromonas were divided into 64 PFGE types, indicating the diversity of molecular biotypes. Drug susceptibility test showed that Aeromonas was sensitive to seven antibiotics including cefotetan, and partially sensitive to five antibiotics (1.5%–21.2%) such as cefazolin, and highly resistant to ampicillin and ampicillin/sulbactam (90.0%).ConclusionThe contamination of Aeromonas, which carries multiple virulence genes and has high resistances to several antibiotics, is severe in Ma'anshan. It is necessary to strengthen the surveillance for Aeromonas and its drug resistance.
- Aeromonas /
- Molecular characteristic /
- Drug sensitivity

HTML全文

图 1 气单胞菌种水平rpoD序列鉴定结果

注：黑色三角形代表该种单胞菌的集合，三角形的大小反映为菌株数量的多少，括号内为菌株数量

Figure 1. Identification of Aeromonas strains at species level based on rpoD sequences.

下载: 全尺寸图片幻灯片

图 2 64株气单胞菌脉冲场凝胶电泳分型结果

Figure 2. PFGE results of 64 Aeromonas strains

下载: 全尺寸图片幻灯片

表 1 气单胞菌毒力基因检测引物

Table 1. Primers used for detection of virulence genes of Aeromonas

毒力基因	GenBank 序列号	引物序列（5′ ~ 3′）	产物长度（bp）
Fla	FlaF	TCCAACCGTYTGACCTC	608
Fla	FlaR	GMYTGGTTGCRATGGT	608
Elastase	ElaF	ACACGGTCAAGGAGATCAAC	513
Elastase	ElaR	CGCTGGTGTTGGCCAGCAGG	513
hlyA	hlyAF	GGCCGGTGGCCCGAAGATACGGG	597
hlyA	hlyAR	GGCGGCGCCGGACGAGACGGG	597
ast	astF	TCTCCATGCTTCCCTTCCACT	260
ast	astR	GTGTAGGGATTGAAGAAGCCG	260
act	actF	AGAAGGTGACCACCAAGAACA	232
act	actR	AACTGACATCGGCCTTGAACTC	232
Ascf-G	ASCF-GF	ATGAGGTCATCTGCTCGCGC	900
Ascf-G	ASCF-GR	GGAGCACAACCATGGCTGAT	900

下载: 导出CSV

表 2 不同样本中气单胞菌检出情况

Table 2. Detection of Aeromonas in different samples

样品名称	样品数（份）	分离阳性数（份）	阳性率（%）
农村生活用水	127	35	27.6
食物中毒样品	82	10	12.2
海水产品	100	9	9.0
游泳池水	121	7	5.8
城区生活用水	278	5	1.8
包装食品	161	0	0.0
合计	869	66	7.6

下载: 导出CSV

表 3 不同样品中气单胞菌的种类分布

Table 3. Species distribution of Aeromonas in different samples

样品名称	维氏气单胞菌（株）	嗜水气单胞菌（株）	简氏气单胞菌（株）	水生气单胞菌（株）	豚鼠气单胞菌（株）	疱氏气单胞菌（株）	舒氏气单胞菌（株）	总计（株）
农村生活用水	18	4	9	3	0	0	1	35
食物中毒样品	3	1	2	0	4	0	0	10
海水产品	0	2	0	5	2	0	0	9
游泳池水	1	5	1	0	0	0	0	7
城区生活用水	2	1	1	0	0	1	0	5
合计	24	13	13	8	6	1	1	66

下载: 导出CSV

表 4 66株气单胞菌毒力基因检测情况

Table 4. Detections of virulence genes of Aeromonas strains

毒力基因	维氏气单胞菌（n=24）		嗜水气单胞菌（n=13）		简氏气单胞菌（n=13）		水生气单胞菌（n=8）		豚鼠气单胞菌（n=6）		合计
毒力基因	检出数（株）	检出率（%）	检出数（株）	检出率（%）	检出数（株）	检出率（%）	检出数（株）	检出率（%）	检出数（株）	检出率（%）	检出数（株）	检出率（%）
Fla	13	54.2	6	46.2	5	38.5	5	62.5	5	83.3	34	53.1
act	12	50.0	9	69.2	6	46.2	3	37.5	4	66.7	34	53.1
Elastase	6	25.0	8	61.5	7	53.8	4	50.0	3	50.0	28	43.8
hlyA	8	33.3	6	46.2	4	30.8	4	50.0	2	33.3	24	37.5
Ast	9	37.5	6	46.3	4	30.8	3	37.5	3	50.0	23	35.9
Ascf-G	6	25.0	6	46.4	3	23.1	2	25.0	2	33.3	18	28.1

下载: 导出CSV

表 5 66株气单胞菌菌的药敏结果情况

Table 5. Drug susceptibility results of Aeromonas strains

药物名称	敏感		中度敏感		耐药
药物名称	菌株数量（株）	敏感率（%）	菌株数量（株）	敏感率（%）	菌株数量（株）	耐药率（%）
氨苄西林	1	1.5	3	4.5	62	94.0
氨苄西林/舒巴坦	3	4.5	3	4.5	60	90.9
头孢唑啉	33	50.0	18	27.3	15	22.7
头孢替坦	66	100.0	0	0.0	0	0.0
头孢他啶	64	97.0	0	0.0	2	3.0
头孢曲松	65	98.5	0	0.0	1	1.5
氨曲南	64	96.7	0	0.0	2	3.3
亚胺培南	66	100.0	0	0.0	0	0.0
阿米卡星	65	98.5	1	1.5	0	0.0
庆大霉素	66	100.0	0	0.0	0	0.0
妥布霉素	65	98.5	1	1.5	0	0.0
环丙沙星	66	100.0	0	0.0	0	0.0
左旋氧氟沙星	66	100.0	0	0.0	0	0.0
呋喃妥因	66	100.0	0	0.0	0	0.0
甲氨苄啶/磺胺甲恶唑	65	98.5	0	0.0	1	1.5
哌拉西林/他唑巴坦	66	100.0	0	0.0	0	0.0

下载: 导出CSV

参考文献(14)

[1]	Janda JM, Abbott SL. The genus Aeromonas: taxonomy, pathogenicity, and infection[J]. Clin Microbiol Rev, 2010,23(1):35–73. DOI: 10.1128/CMR.00039−09.
[2]	Parker JL, Shaw JG. Aeromonas spp. clinical microbiology and disease[J]. J Infect, 2011,62(2):109–118. DOI: 10.1016/j.jinf.2010.12.003.
[3]	Pu W, Guo G, Yang N, et al. Three species of Aeromonas (A. dhakensis, A. hydrophila and A. jandaei) isolated from freshwater crocodiles (Crocodylus siamensis) with pneumonia and septicemia[J]. Lett Appl Microbiol, 2019,68(3):212–218. DOI: 10.1111/lam.13112.
[4]	中华人民共和国国家质量监督检验检疫总局. GB/T 18652－2002　致病性嗜水气单胞菌检验方法［S］. 北京: 中国标准出版社, 2002. General Administration of Quality Supervision, Inspection and Quarantine of the People's Republic of China. GB/T 18652－2002 Methods for detection of pathogenic aeromonas hydrophila［S］. Beijing: China Standard Press, 2002.
[5]	Persson S, Al-Shuweli S, Yapici S, et al. Identification of clinical aeromonas species by rpoB and gyrB sequencing and development of a multiplex PCR method for detection of Aeromonas hydrophila, A. caviae, A. veronii, and A. media[J]. J Clin Microbiol, 2015,53(2):653–656. DOI: 10.1128/JCM.01963−14.
[6]	李欣悦, 李凤娟, 杜鹏程, 等. rpoD和gyrA基因对气单胞菌鉴定的比较分析[J]. 疾病监测,2016,31(3):200–204. DOI: 10.3784/j.issn.1003−9961.2016.03.006. Li XY, Li FJ, Du PC, et al. Comparative analysis of rpoD and gyrA gene in identification of Aeromonas[J]. Dis Surveill, 2016,31(3):200–204. DOI: 10.3784/j.issn.1003−9961.2016.03.006.
[7]	Senderovich Y, Ken-Dror S, Vainblat I, et al. A molecular study on the prevalence and virulence potential of Aeromonas spp. recovered from patients suffering from diarrhea in Israel[J]. PLoS One, 2012,7(2):e30070. DOI: 10.1371/journal.pone.0030070.
[8]	Rasmussen-Ivey CR, Figueras MJ, McGarey D, et al. Virulence factors of Aeromonas hydrophila: In the wake of reclassification[J]. Front Microbiol, 2016,7:1337. DOI: 10.3389/fmicb.2016.01337.
[9]	Nolla-Salas J, Codina-Calero J, Vallés-Angulo S, et al. Clinical significance and outcome of Aeromonas spp. infections among 204 adult patients[J]. Eur J Clin Microbiol Infect Dis, 2017,36(8):1393–1403. DOI: 10.1007/s10096−017−2945−4.
[10]	Navarro A, Martínez-Murcia A. Phylogenetic analyses of the genus Aeromonas based on housekeeping gene sequencing and its influence on systematics[J]. J Appl Microbiol, 2018,125(3):622–631. DOI: 10.1111/jam.13887.
[11]	Li FJ, Wang WQ, Zhu ZQ, et al. Distribution, virulence-associated genes and antimicrobial resistance of Aeromonas isolates from diarrheal patients and water, China[J]. J Infect, 2015,70(6):600–608. DOI: 10.1016/j.jinf.2014.11.004.
[12]	Zhou YY, Yu L, Nan Z, et al. Taxonomy, virulence genes and antimicrobial resistance of Aeromonas isolated from extra-intestinal and intestinal infections[J]. BMC Infect Dis, 2019,19(1):158. DOI: 10.1186/s12879−019−3766−0.
[13]	Hossain S, De Silva BCJ, Dahanayake PS, et al. Characterization of virulence properties and multi-drug resistance profiles in motile Aeromonas spp. isolated from zebrafish (Danio rerio)[J]. Lett Appl Microbiol, 2018,67(6):598–605. DOI: 10.1111/lam.13075.
[14]	Vincent AT, Emond-Rheault JG, Barbeau X, et al. Antibiotic resistance due to an unusual ColE1-type replicon plasmid in Aeromonas salmonicida[J]. Microbiology, 2016,162(6):942–953. DOI: 10.1099/mic.0.000286.