1株致仔猪脑膜炎的猪链球菌的分离鉴定及致病性分析

doi:10.16431/j.cnki.1671-7236.2024.05.035

中国畜牧兽医 ›› 2024, Vol. 51 ›› Issue (5): 2143-2153.doi: 10.16431/j.cnki.1671-7236.2024.05.035

1株致仔猪脑膜炎的猪链球菌的分离鉴定及致病性分析

刘丁语^1,2, 刘宝玲¹, 何振文¹, 罗琴¹, 乔常宏¹, 陈翔宇¹, 王晓虎¹, 向华¹, 王刚¹, 谭臣², 蔡汝健¹

1. 广东省农业科学院动物卫生研究所, 广东省畜禽疫病防治研究重点实验室, 农业农村部兽用药物与诊断技术广东科学观测实验站, 广州 510640;
2. 华中农业大学动物医学院, 生猪健康养殖协同创新中心, 武汉 430000

收稿日期:2023-10-08 出版日期:2024-05-05 发布日期:2024-04-28
通讯作者: 蔡汝健 E-mail:466866569@qq.com
作者简介:刘丁语,E-mail:ytdl@webmail.hzau.edu.cn;刘宝玲,E-mail:745292679@qq.com。
基金资助:
广东省省级科技计划项目(2023B0202010014);广州市农村科技特派员项目(20212100015);2021年英德市科技计划项目

Isolation,Identification and Pathogenicity Analysis of a Strain of Streptococcus suis Causing Meningitis in Piglets

LIU Dingyu^1,2, LIU Baoling¹, HE Zhenwen¹, LUO Qin¹, QIAO Changhong¹, CHEN Xiangyu¹, WANG Xiaohu¹, XIANG Hua¹, WANG Gang¹, TAN Chen², CAI Rujian¹

1. Scientific Observation and Experiment Station of Veterinary Drugs and Diagnostic Techniques of Guangdong Province of Ministry of Agriculture and Rural Affairs, Key Laboratory of Livestock Disease Prevention of Guangdong Province, Institute of Animal Health, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, China;
2. The Cooperative Innovation Center for Sustainable Pig Production, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430000, China

Received:2023-10-08 Online:2024-05-05 Published:2024-04-28

摘要/Abstract

摘要： 【目的】确定广东某猪场送检的疑似猪链球菌引起仔猪脑膜炎的病猪组织中的病原,并测定其药物敏感性和致病性,为猪场用药及探索该病原引起脑膜炎的机制奠定基础。【方法】对疑似猪链球菌引起的仔猪脑膜炎送检病料进行细菌分离培养、形态观察、生化试验及分子学鉴定,并对其开展药物敏感性试验、毒力基因测定、小鼠感染试验及病理组织学诊断。【结果】仔猪脑组织分离株在血平板上出现溶血现象,革兰染色镜检细菌被染成紫色,形态为圆形,部分呈链状排列。生化鉴定结果显示,分离株能发酵棉子糖、乳糖、菊糖、七叶苷,不能发酵山梨醇、甘露醇、阿拉伯糖、松三糖、D-核糖,马尿酸盐还原试验为阴性,与猪链球菌相符。测序分析后确定分离株为猪链球菌。血清型鉴定结果显示,分离株为9 型猪链球菌,命名为GDS209。药物敏感性试验结果显示,分离株GDS209对头孢噻吩(先锋Ⅰ)、头孢氨苄(先锋Ⅳ)和氨苄西林/舒巴坦等10种抗菌药耐药,对替卡西林/克拉维酸、氟苯尼考、阿莫西林3种抗菌药敏感。毒力基因检测结果显示,分离株GDS209携带gdh、gapdh、fbps、orf2、srtA、ccpA毒力基因,不携带mrp、sly、luxS、epf毒力基因。小鼠感染试验结果显示,分离株GDS209对小鼠的半数致死量(LD₅₀)为1×10⁹ CFU。组织病理学观察显示,分离株GDS209能引起昆明小鼠心脏、肝脏、脾脏、肺脏、脑不同程度的淤血和炎症。【结论】本研究分离到1株9型猪链球菌,该分离菌对大部分猪场常用抗菌药耐药,对替卡西林/克拉维酸、氟苯尼考、阿莫西林3种抗菌药敏感,本研究结果可为9型猪链球菌的临床诊断和药物治疗提供一定的参考依据。

关键词: 仔猪脑膜炎; 猪链球菌; 分离鉴定; 耐药性; 致病性

Abstract: 【Objective】 The purpose of this experiment was to determine the pathogen in the tissue samples of pigs with suspected piglet meningitis caused by Streptococcus suis in a pig farm in Guangdong,and to determine its drug sensitivity and pathogenicity,so as to provide medication for pig farms and explore the mechanism of meningitis caused by this pathgen.【Method】 Bacterial isolation and culture,morphological observation,biochemical testing,and molecular identification were performed on the samples of piglets suspected to have meningitis caused by Streptococcus suis.Drug sensitivity testing,virulence gene testing,mouse infection testing,and histopathological diagnosis were also conducted.【Result】 The isolate from the piglet brain tissue showed hemolysis on the blood plate.Gram staining microscopy showed that bacteria were stained purple,with a circular shape and some arranged in a chain shape.Biochemical characteristics showed that the isolate could ferment raffinose,lactose,inulin and esculin,but could not ferment sorbitol,mannitol,arabinose,melezitose and D-ribose.The hippurate reduction test was negative,which were consistent with Streptococcus suis,and was determined to be Streptococcus suis after sequencing analysis.The serotype identification results showed that the isolate was Streptococcus suis type 9,named GDS209.The results of drug sensitivity test showed that GDS209 was resistant to 10 antibacterials such as cefalotin (pioneer Ⅰ),cefalexin (pioneer Ⅳ) and ampicillin/sulbactam,and was sensitive to ticarcillin/clavulanic acid,florfenicol and amoxicillin.The detection results of virulence genes showed that GDS209 carried gdh,gapdh,fbps,orf2,srtA and ccpA virulence genes,but did not carry mrp,sly,luxS and epf virulence genes.The mouse infection test showed that the LD₅₀ of GDS209 was 1×10⁹ CFU.Histopathological observation showed that GDS209 could cause congestion and inflammation in Kunming mice’s heart,liver,spleen,lung and brain to varying degrees.【Conclusion】 In this study,a strain of Streptococcus suis serotype 9 was isolated,which was resistant to commonly used antimicrobials in most pig farms and sensitive to ticarcillin/clavulanic acid,florfenicol and amoxicillin.The results of this study could provide some reference for the clinical diagnosis and drug treatment of Streptococcus suis serotype 9.

Key words: piglet meningitis; Streptococcus suis; isolation and identification; drug resistance; pathogenicity

中图分类号:

S852.61^＋1

刘丁语, 刘宝玲, 何振文, 罗琴, 乔常宏, 陈翔宇, 王晓虎, 向华, 王刚, 谭臣, 蔡汝健. 1株致仔猪脑膜炎的猪链球菌的分离鉴定及致病性分析[J]. 中国畜牧兽医, 2024, 51(5): 2143-2153.

LIU Dingyu, LIU Baoling, HE Zhenwen, LUO Qin, QIAO Changhong, CHEN Xiangyu, WANG Xiaohu, XIANG Hua, WANG Gang, TAN Chen, CAI Rujian. Isolation,Identification and Pathogenicity Analysis of a Strain of Streptococcus suis Causing Meningitis in Piglets[J]. China Animal Husbandry and Veterinary Medicine, 2024, 51(5): 2143-2153.

参考文献

[1] 韦冰梅,韦巧珍,陈玉君.新生儿细菌性脑膜炎常见病原菌跨膜机制的研究进展[J].广西医学,2021,43(2):228-230. WEI B M,WEI Q Z,CHEN Y J.Review on transmembrane mechanism of common pathogenic bacteria in neonatal bacteria meningitis[J].Guangxi Medical Journal,2021,43(2):228-230.(in Chinese)
[2] 陈明杰,曹梦园,曹芳,等.miRNA对细菌性脑膜炎中血脑屏障影响的研究进展[J].现代畜牧兽医,2021,12:83-86. CHEN M J,CAO M Y,CAO F,et al.Research progress on the effect of miRNA on the blood-brain barrier in bacterial meningitis[J].Modern Journal of Animal Husbandry and Veterinary Medicine,2021,12:83-86.(in Chinese)
[3] JIANG F,GUO J,CHENG C,et al.Human infection caused by Streptococcus suis serotype 2 in China:Report of two cases and epidemic distribution based on sequence type[J].BMC Infectious Diseases,2020,20(1):223.
[4] VAN SAMKAR A,BROUWER M C,SCHULTSZ C,et al.Streptococcus suis meningitis:A systematic review and Meta-analysis[J].PLoS Neglected Tropical Diseases,2015,9(10):e4191.
[5] PAN Z,MA J,DONG W,et al.Novel variant serotype of Streptococcus suis isolated from piglets with meningitis[J].Applied and Environmental Microbiology,2015,81(3):976-985.
[6] WU C F,CHEN S H,CHOU C C,et al.Serotype and multilocus sequence typing of Streptococcus suis from diseased pigs in Taiwan[J].Scientific Reports,2023,13(1):8263.
[7] SUSILAWATHI N M,TARINI N M A,FATMAWATI N N D,et al.Streptococcus suis-associated meningitis,Bali,Indonesia,2014-2017[J].Emerging Infectious Diseases,2019,25(12):2235-2242.
[8] 姜轩,刘梦月,孙英峰,等.天津地区猪7型链球菌的分离鉴定与小鼠致病性研究[J].黑龙江畜牧兽医,2019,7:29-32. JIANG X,LIU M Y,SUN X F,et al.Isolation and identification of Streptococcus suis serotype 7 in Tianjin and study on its pathogenicity in mice[J].Heilongjiang Animal Science and Veterinary Medicine,2019,7:29-32.(in Chinese)
[9] 陈申申,马家乐,张新锋,等.致仔猪脑膜炎猪链球菌的分离鉴定[J].畜牧兽医学报,2019,50(1):143-150. CHEN S S,MA J L,ZHANG X F,et al.Isolation and identification of Streptococcus suis causing meningitis in piglets[J].Acta Veterinaria et Zootechnica Sinica,2019,50(1):143-150.(in Chinese)
[10] 陈良荣,徐翠,陈慧婷,等.猪链球菌9型引起的猪脑膜脑炎型链球菌病病例报告[J].江西畜牧兽医杂志,2018,2:40-41. CHEN L R,XU C,CHEN H T,et al.A case report of porcine meningoencephalitis streptococcal disease caused by Streptococcus suis type 9[J].Jiangxi Journal of Animal Husbandry & Veterinary Medicine,2018,2:40-41.(in Chinese)
[11] 萧松建,周奕,张楚东,等.一例14型人感染猪链球菌病例流行病学调查[J].疾病监测,2019,34(9):861-864. XIAO S J,ZHOU Y,ZHANG C D,et al.Epidemiological investigation of a case of human infection with Streptococcus suis serotype 14[J].Disease Surveillance,2019,34(9):861-864.(in Chinese)
[12] WILLEMSE N,VAN DER ARK K,STOCKHOFE-ZURWIEDEN N,et al.Clonal expansion of a virulent Streptococcus suis serotype 9 lineage distinguishable from carriage subpopulations[J].Scientific Reports,2019,9(1):15429.
[13] 字嘉鑫,连英杰,陈敏仪,等.4株广东地区2型猪链球菌耐药性及致病性研究[J].中国畜牧兽医,2023,50(9):3740-3751. ZI J X,LIAN Y J,CHEN M Y,et al.Drug resistance and pathogenicity of 4 strains of Streptococcus suis type 2 in Guangdong[J]. China Animal Husbandry & Veterinary Medicine,2023,50(9):3740-3751.(in Chinese)
[14] 张欣,潘晨浩,赵瑞利,等.抗菌肽Temprine-La(FS)抑制2型猪链球菌生物被膜形成的研究[J].中国畜牧兽医,2021,48(1):324-337. ZHANG X,PAN C H,ZHAO R L,et al.Inhibitory effect of antimicrobial peptide Temprine-La(FS) on biofilm formation of Streptococcus suis type 2[J]. China Animal Husbandry & Veterinary Medicine,2021,48(1):324-337.(in Chinese)
[15] 郭长明,陈怀君,朱善元,等.广西某猪场猪链球菌分离鉴定及其生物学特性研究[J].中国畜牧兽医,2021,48(2):736-746. GUO C M,CHEN H J,ZHU S Y,et al.Isolation and identification of Streptococcus suis and biological characteristics research on a swine farm in Guangxi[J].China Animal Husbandry & Veterinary Medicine,2021,48(2):736-746.(in Chinese)
[16] WANG Y,WANG Y,LIU B,et al.pdh modulate virulence through reducing stress tolerance and biofilm formation of Streptococcus suis serotype 2[J].Virulence,2019,10(1):588-599.
[17] 邓智丹,黄良宗,白挨泉.广东省1株猪2型链球菌的分离鉴定及部分生物学特性研究[J].中国畜牧兽医,2021,48(2):717-725. DENG Z D,HUANG L Z,BAI A Q.Isolation,identification and biological characteristics of Streptococcus suis serotype 2 strain from Guangdong province[J]. China Animal Husbandry & Veterinary Medicine,2021,48(2):717-725.(in Chinese)
[18] 王治方,徐引弟,张青娴,等.2型猪链球菌河南株的生物学特性研究[J].养猪,2020,3:97-102. WANG Z F,XU Y D,ZHANG Q X,et al.Study on biological characteristics of Streptococcus suis serotype 2 Henan strain[J].Swine Production,2020,3:97-102.(in Chinese)
[19] 冯建东,李雨来.猪链球菌病的流行病学、临床症状、实验室诊断及防控[J].现代畜牧科技,2021,7:143-144. FENG J D,LI Y L.Epidemiology,clinical symptoms,laboratory diagnosis and prevention and control of Streptococcus suis[J].Modern Animal Husbandry Science & Technology,2021,7:143-144.(in Chinese)
[20] AMASS S F,SANMIGUEL P,CLARK L K.Demonstration of vertical transmission of Streptococcus suis in swine by genomic fingerprinting[J].Journal of Clinical Microbiology,1997,35(6):1595-1596.
[21] HILL J E,GOTTSCHALK M,BROUSSEAU R,et al.Biochemical analysis,cpn60 and 16S rDNA sequence data indicate that Streptococcus suis serotypes 32 and 34,isolated from pigs,are Streptococcus orisratti[J].Veterinary Microbiology,2005,107(1-2):63-69.
[22] 金梅林,贝为成,陈焕春,等.猪链球菌致病分子基础与防控技术[M].北京:科学出版社,2018. JIN M L,BEI W C,CHEN H C,et al.Molecular Basis and Prevention and Control Technology of Streptococcus suis Pathogenicity[M].Beijing:Science Press,2018.(in Chinese)
[23] DENICH L C,FARZAN A,FRIENDSHIP R,et al.A case-control study to investigate the serotypes of S. suis isolates by multiplex PCR in nursery pigs in Ontario,Canada[J].Pathogens,2020,9(1):44.
[24] 程建伟.四川部分地区猪链球菌分离鉴定及主要生物学特征研究[D].成都:西南民族大学,2022. CHENG J W.Isolation,identification and main biological characteristics of Streptococcus suis in some areas of Sichuan province[D].Chengdu:Southwest Minzu University,2022.(in Chinese)
[25] 魏述亮,邱家章,张恒彬.猪链球菌病研究进展[J].山东畜牧兽医,2023,44(1):86-89. WEI S L,QIU J Z,ZHANG H B.Advances in research on swine streptococcosis[J].Shandong Journal of Animal Science and Veterinary Medicine,2023,44(1):86-89.(in Chinese)
[26] TRAM G,JENNINGS M P,BLACKALL P J,et al.Streptococcus suis pathogenesis-A diverse array of virulence factors for a zoonotic lifestyle[J].Advances in Microbial Physiology,2021,78:217-257.
[27] GOTTSCHALK M,XU J,CALZAS C,et al.Streptococcus suis:A new emerging or an old neglected zoonotic pathogen?[J].Future Microbiology,2010,5(3):371-391.
[28] MAIREY E,GENOVESIO A,DONNADIEU E,et al.Cerebral microcirculation shear stress levels determine Neisseria meningitidis attachment sites along the blood-brain barrier[J].Journal of Experimental Medicine,2006,203(8):1939-1950.
[29] 王涛.猪链球菌穿血脑屏障毒力因子的筛选及鉴定[D].合肥:安徽医科大学,2013. WANG T.Screening and identification of virulence factors involving in the traversal of Streptococcus suis across the human blood-brain barrier[D].Hefei:Anhui Medical University,2013.(in Chinese)
[30] 郝淮杰.猪链球菌与人脑微血管内皮细胞相互作用研究[D].北京:中国人民解放军军事医学科学院,2011. HAO H J.Interaction between Streptococcus suis and human brain microvascular endothelial cells[D].Beijing:Academy of Military Medical Sciences,2011.(in Chinese)
[31] SEGURA M,GOTTSCHALK M.Streptococcus suis interactions with the murine macrophage cell line J774:Adhesion and cytotoxicity[J].Infection and Immunity,2002,70(8):4312-4322.
[32] BENGA L,FULDE M,NEIS C,et al.Polysaccharide capsule and suilysin contribute to extracellular survival of Streptococcus suis co-cultivated with primary porcine phagocytes[J].Veterinary Microbiology,2008,132(1-2):211-219.
[33] LUN S,PEREZ-CASAL J,CONNOR W,et al.Role of suilysin in pathogenesis of Streptococcus suis capsular serotype 2[J].Microbial Pathogenesis,2003,34(1):27-37.
[34] VANIER G,SEKIZAKI T,DOMINGUEZ-PUNARO M C,et al.Disruption of srtA gene in Streptococcus suis results in decreased interactions with endothelial cells and extracellular matrix proteins[J].Veterinary Microbiology,2008,127(3-4):417-424.
[35] 刘宝玲,楚品品,李春玲,等.猪链球菌临床分离株对四环素类抗生素的耐药性和耐药基因分析[J].中国畜牧兽医,2022,49(7):2796-2804. LIU B L,CHU P P,LI C L,et al.Analysis of resistance and resistance genes of Streptococcus suis clinical isolates to tetracyclines[J].China Animal Husbandry & Veterinary Medicine,2022,49(7):2796-2804.(in Chinese)
[36] YU Y,FANG J T,ZHENG M,et al.Combination therapy strategies against multiple-resistant Streptococcus suis[J].Frontiers in Pharmacology,2018,15(9):489.
[37] 王冬经,苏中华,元振杰,等.藏羊源链球菌的分离鉴定及生物学特性研究[J].中国畜牧兽医,2023,50(9):3823-3832. WANG D J,SU Z H,YUAN Z J,et al.Isolation,identification and biological characteristics of Streptococcus isolated from Tibetan sheep[J]. China Animal Husbandry & Veterinary Medicine,2023,50(9):3823-3832.(in Chinese)
[38] 王冰艺,马弘财,邹明昊,等.牦牛源链球菌的分离鉴定及耐药性分析[J].中国畜牧兽医,2023,50(2):745-753. WANG B Y,MA H C,ZOU M H,et al.Isolation,identification and drug resistance analysis of Streptococcus from yak[J]. China Animal Husbandry & Veterinary Medicine,2023,50(2):745-753.(in Chinese)

1株致仔猪脑膜炎的猪链球菌的分离鉴定及致病性分析

Isolation,Identification and Pathogenicity Analysis of a Strain of Streptococcus suis Causing Meningitis in Piglets

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