广西罗非鱼源无乳链球菌分离鉴定及耐药性分析

doi:10.16431/j.cnki.1671-7236.2025.06.041

摘要/Abstract

摘要： 【目的】掌握广西地区无乳链球菌流行规律和耐药特征，为罗非鱼健康养殖提供科学依据。【方法】采集广西贵港市、北海市、玉林市罗非鱼养殖场病鱼肝脏、脾脏、脑等组织作为样本进行细菌分离培养，通过对分离菌株形态观察及16S rRNA测序分析进行病原鉴定；利用PCR方法检测各分离菌株血清型、主要毒力基因，并采用K-B法检测其耐药性。【结果】共分离38株链球菌，分离菌株在血平板上形成乳白色圆形、具备溶血环的小菌落；革兰染色为阳性，呈典型短链状，经鉴定均为Ⅰa型无乳链球菌；分离菌株均能扩增出毒力基因sodA、cylE、gapC、bac和bca，但未能扩增出毒力基因scpB；38株分离株对氨基糖苷类药物敏感性较低，对阿米卡星耐药率达到85%以上；分离菌株对β-内酰胺类药物敏感性较高，对青霉素G、头孢哌酮敏感性达100%。38株分离菌中有23株分离菌株呈多重耐药性，耐药数量为3～10重，其中4重及以上为13株，占总分离菌株的34.21%。【结论】广西地区罗非鱼源无乳链球菌临床菌株主要血清型为Ⅰa型，其主要毒力基因为sodA、cylE、gapC、bac和bca；分离菌株对氨基糖苷类药物耐药，提示避免使用该类药物进行防治。

关键词: 罗非鱼; 无乳链球菌; 血清型; 毒力基因; 耐药性

Abstract: 【Objective】 The aim of this experiment was to understand the epidemic regularity and drug resistance characteristics of Streptococcus agalactiae in Guangxi,and provide scientific basis for healthy tilapia breeding.【Method】 The diseased fish from tilapia farm in Yulin,Beihai,Guigang in Guangxi were collected for bacterial isolation and culture.The isolated strains were identified by morphological observation and 16S rRNA sequencing analysis.The serotypes and main virulence genes of the isolates were identified by PCR,and their drug resistance was determined by K-B method.【Result】 A total of 38 Streptococcus strains were isolated,and the isolated strains formed milky white round small colonies with hemolytic rings on the blood plate.Gram staining was positive with typical short chain,and all were identified as Ⅰa Streptococcus agalactiae.Virulence genes sodA,cylE,gapC,bac and bca could be detected in all isolates,but virulence gene scpB could not be detected.The sensitivity of 38 isolates to aminoglycosides was low,and the resistance rate to amikacin was more than 85%.The isolates were highly sensitive to β-lactam drugs,among which penicillin G and cefoperazone were 100% sensitive.Among 38 strains of isolates,23 isolates showed multiple drug resistance,the number of drug resistance was 3-10,and 13 isolates were more than 4,accounting for 34.21% of the total isolates.【Conclusion】 The main serotype of Streptococcus agalactis derived from tilapia in Guangxi was type Ⅰa.The main virulence genes were sodA, cylE,gapC, bac and bca.The isolates were resistant to aminoglycosides,suggesting that the use of these drugs should be avoided.

Key words: tilapia; Streptococcus agalactiae; serotype; virulence gene; drug resistance

中图分类号:

全琛宇, 周英宁, 蒲婵娟, 陈婷婷, 许心婷, 卢冰霞, 许艺兰, 赵硕, 杨讯业, 段群棚, 秦毅斌, 李斌, 陈忠伟, 何颖. 广西罗非鱼源无乳链球菌分离鉴定及耐药性分析[J]. 中国畜牧兽医, 2025, 52(6): 2893-2903.

QUAN Chenyu, ZHOU Yingning, PU Chanjuan, CHEN Tingting, XU Xinting, LU Bingxia, XU Yilan, ZHAO Shuo, YANG Xunye, DUAN Qunpeng, QIN Yibin, LI Bin, CHEN Zhongwei, HE Ying. Isolation,Identification and Drug Resistance Analysis of Streptococcus agalactiae from Tilapia in Guangxi[J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(6): 2893-2903.

参考文献

[1] JAFAR Q A,SAMEER A Z,SALWA A M,et al.Molecular investigation of Streptococcus agalactiae isolates from environmental samples and fish specimens during a massive fish kill in Kuwait Bay[J].Pakistan Journal of Biological Sciences,2008,11(21):2500-2504.
[2] AMAL M N A,ZAMRI-SAAD M,IFTIKHAR A R,et al.An outbreak of Streptococcus agalactiae infection in cage-cultured golden pompano,Trachinotus blochii (Lacépède),in Malaysia[J].Journal of Fish Diseases,2012,35(11):849-852.
[3] WANG Y T,HUANG H Y,TSAI M A,et al.Phosphoglycerate kinase enhanced immunity of the whole cell of Streptococcus agalactiae in tilapia,Oreochromis niloticus[J].Fish & Shellfish Immunology,2014,41(2):250-259.
[4] YE X,LI J,LU M X,et al.Identification and molecular typing of Streptococcus agalactiae isolated from pond-cultured tilapia in China[J].Fisheries Science,2011,77(4):623-632.
[5] 吴南,郭佳,熊丽,等.罗非鱼链球菌病疫苗的研究进展[J].南方水产科学,2013,9(6):90-96.WU N,GUO J,XIONG L,et al.Research progress on vaccine of streptococcosis in tilapia[J].South China Fisheries Science,2013,9(6):90-96.(in Chinese)
[6] CHEN M,LI L P,WANG R,et al.PCR detection and PFGE genotype analyses of streptococcal clinical isolates from tilapia in China[J]. Veterinary Microbiology,2012,159(3/4):526-530.
[7] 胡大胜,黄钧,黄艳华,等.12种药物对罗非鱼无乳链球菌的抑菌试验[J].中国水产,2012,8:60-63.HU D S,HUANG J,HUANG Y H,et al.Bacteriostatic test of 12 kinds of drugs against Streptococcus agalactis of tilapia[J]. China Fisheries,2012,8:60-63.(in Chinese)
[8] 黄艳华，马沙，韩书煜，等.2011～2016年广西罗非鱼源无乳链球菌的耐药谱型分析[J].南方农业学报，2018,49(5):1023-1031.HUANG Y H,MA S,HAN S Y,et al.Antibiogram types of Streptococcus agalactiae strains isolated from Oreochromis spp.in Guangxi during 2011－2016[J].Journal of Southern Agriculture,2018,49(5):1023-1031.(in Chinese)
[9] 梁静真,黄立春,韦慕兰,等.广西罗非鱼源无乳链球菌耐药性及其四环素耐药基因检测[J].南方农业学报,2018,49(10):2077-2086.LIANG J Z,HUANG L C,WEI M L,et al.Detection of antibiotics resistance and tetracycline resistance genes in Streptococcus agalactiae from tilapia in Guangxi[J].Journal of Southern Agriculture,2018,49(10):2077-2086.(in Chinese)
[10] 鲁晶娣,韦盘秋,曾令杰,等.柳州地区罗非鱼无乳链球菌与海豚链球菌分离株药敏性分析[J].广西畜牧兽医,2019,35(2):57-60.LU J D,WEI P Q,ZENG L J,et al.Drug sensitivity analysis of Streptococcus agalactiae and Streptococcus porposus isolates from tilapia in Liuzhou area[J].Guangxi Journal of Animal Husbandry & Veterinary Medicine,2019,35(2):57-60.(in Chinese)
[11] BOSSHARD P P,SANTINI Y,DOMINIQUE G,et al.Bacterial diversity and community composition in the chemocline of the meromictic alpine Lake Cadagno as revealed by 16S rDNA analysis[J].FEMS Microbiology Ecology,2000,31(2):173-182.
[12] IMPERI M,PATARCCHIA M,ALFARONE G,et al.A multiplex PCR assay for the direct identification of the capsular type (Ⅰa to Ⅸ) of Streptococcus agalactiae[J].Journal of Microbiological Methods,2010,80(2):212-214.
[13] 陈怀君,郭长明,朱善元,等.海南罗非鱼源无乳链球菌的分离鉴定及生物学特性研究[J].中国畜牧兽医,2021,48(8):3058-3068.CHEN H J,GUO C M,ZHU S Y,et al.Isolation,identification and biological characteristics of Streptococcus agalactiae from tilapia in Hainan[J].China Animal Husbandry & Veterinary Medicine,2021,48(8):3058-3068.(in Chinese)
[14] 袁伟.中国罗非鱼源无乳链球菌流行特征及耐药性研究[D].上海:上海海洋大学,2017.YUAN W.Epidemiological characteristics and drug resistance of Streptococcus agalactiae isolated from tilapia in China[D].Shanghai:Shanghai Ocean University,2017.(in Chinese)
[15] WEI T,SIMKO V,LEVY M,et al.Package 'corrplot’[J].Statistician,2017,56(316):e24.
[16] VILLANUEVA R A M， CHEN Z J. ggplot2:Elegant graphics for data analysis (2nd ed.)[J].Measurement:Interdisciplinary Research and Perspectives,2019,17(3):160-167.
[17] 袁橙,陈怀君,袁圣,等.福建省漳州市罗非鱼无乳链球菌分离鉴定及毒力基因和耐药性研究[J].中国畜牧兽医,2022,49(9):3655-3664.YUAN C,CHEN H J,YUAN S,et al.Isolation,identification and study on virulence gene and drug resistance of Streptococcus agalactiae from tilapia in Zhangzhou city,Fujian province[J].China Animal Husbandry & Veterinary Medicine,2022,49(9):3655-3664.(in Chinese)
[18] 郭长明,陈怀君,袁橙,等.广西罗非鱼无乳链球菌的分离鉴定、毒力基因及耐药性研究[J].中国畜牧兽医,2021,48(7):2607-2616.GUO C M,CHEN H J,YUAN C,et al.Isolation,identification,virulence genes and antibiotic resistance analysis of Streptococcus agalactiae from tilapia in Guangxi[J].China Animal Husbandry & Veterinary Medicine,2021,48(7):2607-2616.(in Chinese)
[19] 刘婵.广东省罗非鱼无乳链球菌分子流行病学及耐药性研究[D].天津:天津农学院,2018.LIU C.Molecular epidemiological and drug resistance of Streptococcus agalactiae isolated in tilapia from Guangdong province[D].Tianjin:Tianjin Agricultural University,2018.(in Chinese)
[20] 邝伟键,黄良宗,刘明杰,等.罗非鱼源无乳链球菌的分离鉴定及药敏试验[J].畜牧与兽医,2017,49(9):62-66.KUANG W J,HUANG L Z,LIU M J,et al.Isolation,identification and drug sensitivity of Streptococcus agalactiae from Nile tilapiai[J].Animal Husbandry & Veterinary Medicine,2017,49(9):62-66.(in Chinese)
[21] 刘丹宁,吴昌旭,冀禹彤,等.百色地区罗非鱼源无乳链球菌的分离鉴定及耐药性研究[J].广西畜牧兽医,2023,39(6):272-276.LIU D N,WU C X,JI Y T,et al.Isolation,identification and drug resistance of Streptococcus agalactiae derived from tilapia in Baise area[J].Guangxi Journal of Animal Husbandry & Veterinary Medicine,2023,39(6):272-276.(in Chinese)
[22] 方丽华.猪链球菌2型超氧化物歧化酶SodA与丝氨酸/苏氨酸去磷酸酶Stp1影响细菌毒力研究[D].杭州:浙江大学,2016.FANG L H.Superoxide dismutase SodA and a serine threonine/phosphatase Stpl of Streptococcus suis type 2 are critical for its virulence[D].Hangzhou:Zhejiang University,2016.(in Chinese)
[23] 邓永强,汪开毓.鱼类无乳链球菌病的研究进展[J].中国畜牧兽医,2016,43(9):2490-2495.DENG Y Q,WANG K Y.Research progress on fish Streptococcus agalactiae disease[J].China Animal Husbandry & Veterinary Medicine,2016,43(9):2490-2495.(in Chinese)
[24] 杜苏兰,雷连成,王玲玲,等.湖北吉富罗非鱼源Ⅰa无乳链球菌毒力分析及药敏特性[J].淡水渔业,2023,53(6):58-68.DU S L,LEI L C,WANG L L,et al.Virulence analysis and drug sensitivity of type Ⅰa Streptococcus agalactiae isolated from the genetic improvement of farmed tilapia in Hubei[J].Freshwater Fisheries,2023,53(6):58-68.(in Chinese)