禽源产色葡萄球菌的分离鉴定及其耐药性分析

doi:10.16431/j.cnki.1671-7236.2020.07.027

摘要/Abstract

摘要： 为了解禽源产色葡萄球菌的耐药性，本研究从发病金陵花鸡中分离到1株产色葡萄球菌GZQX2019，并进行细菌分离培养、革兰氏染色镜检、生化试验、16S rDNA序列分析、药敏试验、部分耐药与肠毒素基因检测和动物回归试验。结果显示，分离菌在鲜血琼脂培养基上长出表面凸起、边缘整齐的溶血现象白色菌落；革兰氏染色镜检呈葡萄串状球菌。生化试验结果显示，分离菌蔗糖、精氨酸水解酶、木糖等反应阳性，尿素、木糖醇、麦芽糖等反应阴性。系统进化树显示，分离菌与产色葡萄球菌处于同一分支；药敏试验结果显示，分离菌对红霉素、头孢他啶、多西环素等6种抗菌药耐药，对头孢呋辛、头孢哌酮、新霉素等11种抗菌药敏感。耐药与肠毒素基因检测结果显示，分离菌能检测出热敏感细胞肠毒素基因Alt，耐大环内酯类基因ermB，β-内酰胺类基因VIM、TEM，四环素类基因tetB。动物回归试验结果显示，分离菌对小鼠和雏鸡致病能力不强或者没有致病力。试验成功分离到1株禽源产色葡萄球菌，丰富了禽源产色葡萄球菌的认知。

关键词: 金陵花鸡; 产色葡萄球菌; 耐药基因; 毒力基因

Abstract: In order to understand the resistance of avian-origin Staphylococcus chromogenes,a Staphylococcus chromogenes strain named GZQX2019 was isolated from Jinling chicken.The bacteria were isolated and cultured,Gram staining microscopy,biochemical test,16S rDNA sequence analysis,drug sensitivity test,partial resistance and enterotoxin gene detection and animal regression test.The bacteria separation results showed that the white colony with convex surface and neat edge appeared on the blood agar medium,and the Staphylococcus was detected by Gram staining.The results of biochemical tests showed that the reactions of sucrose,arginine hydrolase and xylose were positive,while urea,xylitol and maltose were negative.The phylogenetic tree showed that the isolation was in the same branch with Staphylococcus chromogenes.The drug sensitivity test showed the isolation was resistant to 6 kinds of antibacterial such as erythromycin,ceftazidime and doxycycline,and sensitive to 11 kinds of antibacterial such as cefuroxime,cefoperazone and neomycin.The results of drug resistance and enterotoxin gene detection showed that Alt,ermB,VIM,TEM and tetB could be detected.The results of animal regression tests showed that the isolates were not pathogenic or non-pathogenic to mice and chicks.The experiment successfully isolated one avian-origin Staphylococcus chromogenes,which enriched the cognition of avian-origin Staphylococcus chromogenes.

Key words: Jinling chicken; Staphylococcus chromogenes; drug resistance gene; virulence gene

中图分类号:

S855.1⁺1

陈广, 温贵兰, 张喜懿, 田浪, 杨佰启. 禽源产色葡萄球菌的分离鉴定及其耐药性分析[J]. 中国畜牧兽医, 2020, 47(7): 2215-2222.

CHEN Guang, WEN Guilan, ZHANG Xiyi, TIAN Lang, YANG Baiqi. Isolation and Identification of Avian-origin Staphylococcus chromogenes and Its Drug Resistance Analysis[J]. China Animal Husbandry and Veterinary Medicine, 2020, 47(7): 2215-2222.

参考文献

[1] 马玲,谭义俊,邵帅,等.可形成生物膜的禽源产色葡萄球菌的分离鉴定及耐药性研究[J].西南农业学报,2012,25(2):703-707. MA L,TAN Y J,SHAO S,et al.Identification of biofilm-forming Staphylococcus chromogenes isolated from poultry and its drug-resistance experiment[J].Southwest China Journal of Agricultural Sciences,2012,25(2):703-707.(in Chinese)
[2] KOKSAL F,YASAR H,SAMASTI M.Antibiotic resistance patterns of coagulase-negative Staphylococcus strains isolated from blood cultures of septicemic patients in Turkey[J].Microbiological Research,2009,164(4):404-410.
[3] SAWANT A A,GILLESPIE B E,OLIVER S P.Antimicrobial susceptibility of coagulase-negative Staphylococcus species isolated from bovine milk[J].Veterinary Microbiology,2009,134(1-2):73-81.
[4] WALLER K P,ASPAN A,NYMAN A,et al.CNS species and antimicrobial resistance in clinical and subclinical bovine mastitis[J].Veterinary Microbiology,2011,152(1-2):112-116.
[5] PIESSENS V,VAN COILLIE E,VERBIST B,et al.Distribution of coagulase-negative Staphylococcus species from milk and environment of dairy cows differs between herds[J].Journal of Dairy Science,2011,94(6):2933-2944.
[6] VANDERHAEGHEN W,PIEPERS S,LEROY F,et al.Invited review:Effect,persistence,and virulence of coagulase-negative Staphylococcus species associated with ruminant udder health[J].Journal of Dairy Science,2014,97(9):5275-5293.
[7] ANDRESEN L O,AHRENS P,DAUGAARD L,et al.Exudative epidermitis in pigs caused by toxigenic Staphylococcus chromogenes[J].Veterinary Microbiology,2005,105(3-4):291-300.
[8] SAITO K,HIGUCHI T,KURATA A,et al.Characterization of non-pigmented Staphylococcus chromogenes[J].Journal of Dairy Science,1996,58(7):711-713.
[9] XU J,TAN X,ZHANG X,et al.The diversities of Staphylococcal species,virulence and antibiotic resistance genes in the subclinical mastitis milk from a single Chinese cow herd[J].Microbial Pathogenesis,2015,88(1):29-38.
[10] MARTINS K B,FACCIOLI P Y,BONESSO M F,et al.Characteristics of resistance and virulence factors in different species of coagulase-negative Staphylococci isolated from milk of healthy sheep and animals with subclinical mastitis[J].Journal of Dairy Science,2017,100(3):2184-2195.
[11] PARK J Y,FOX L K,SEO K S,et al.Detection of classical and newly described Staphylococcal superantigen genes in coagulase negative Staphylococci isolated from bovine intramammary infections[J].Veterinary Microbiology,2011,147(1-2):149-154.
[12] STĘPIEŃ-PYŚNIAK D,WILCZYŃSKI J,MAREK A,et al.Staphylococcus simulans associated with endocarditis in broiler chickens[J].Avian pathology,2017;46(1):44-51.
[13] PYZIK E,MAREK A,STĘPIEŃ-PYŚNIAK D,et al.Detection of antibiotic resistance and classical enterotoxin genes in coagulase-negative Staphylococci isolated from poultry in Poland[J].Journal of Veterinary Research,2019,63(2):183-190.
[14] 李巧玲,张召兴,吴同垒,等.貉源嗜水气单胞菌的分离及其生物学特性鉴定[J].中国兽医学报,2018,38(9):1740-1744. LI Q L,ZHANG Z X,WU T L,et al.Isolation and identification of the biological characteristics of Aeromonas hydrophila from raccoon dog[J].Chinese Journal of Veterinary Medicine,2018,38(9):1740-1744.(in Chinese)
[15] 张炳亮,董发明,王文文,等.河南地区猪源性大肠杆菌耐药性及耐药基因型分析[J].西北农林科技大学学报(自然科学版),2019,47(4):1-6. ZHANG B L,DONG F M,WANG W W,et al.Drug resistance and genotype of Escherichia coli in Henan[J].Journal of Northwest A&F University (Natural Science Edition),2019,47(4):1-6.(in Chinese)
[16] 陈伟,张永正,汪雪雁,等.动物源性大肠杆菌ESBLs类耐药基因与HPI基因的双重PCR检测及相关性分析[J].中国兽医科学,2013,43(8):800-805. CHEN W,ZHANG Y Z,WANG X Y,et al.Detection of extended spectrum β-lactamases and high pathogenicity island genes of animal-origin Escherichia coli by duplex PCR and correlation analysis[J].China Veterinary Science,2013,43(8):800-805.(in Chinese)
[17] 张立平,张忠,曾瑾,等.宁夏地区162株大肠杆菌临床分离株对四环素类药物的耐药性分析[J].广东医学,2013,34(23):3580-3583. ZHANG L P,ZHANG Z,ZENG J,et al.Resistance analysis of tetracyclines to 162 clinical isolates of E.coli in Ningxia[J].Guangdong Medical Journal,2013,34(23):3580-3583.(in Chinese)
[18] 裘鹏,郭玉茹,张泽辉,等.基于转录组学技术分析细菌耐药机制的研究进展[J].中国家禽,2019,41(4):45-49. QIU P,GUO Y R,ZHANG Z H,et al.Progress in drug resistance mechanism of bacteria based on transcriptome technology[J].Chinese Poultry,2019,41(4):45-49.(in Chinese)
[19] 李文杰.奶牛乳房炎凝固酶阴性葡萄球菌的分离鉴定与耐药性分析[D].杨凌:西北农林科技大学,2017. LI W J.The separation and identification of dairy cow mastitis and drug resistance analysis in coagulase negative Staphylococcus[D].Yangling:Northwest A&F University,2017.(in Chinese)
[20] 赵华男,瞿玥,韩博,等.奶牛乳房炎源性产色葡萄球菌的肠毒素基因检测[J].中国兽医杂志,2018,54(5):24-27. ZHAO H N,QU Y,HAN B,et al.Detection of enterotoxin genes in Staphylococcus chromogenes isolated from bovine mastitis[J].China Veterinary Journal,2018,54(5):24-27.(in Chinese)
[21] TAPONEN S,PYÖRÄLÄ S.Coagulase-negative Staphylococci as cause of bovine mastitis——Not so different from Staphylococcus aureus?[J]. Veterinary Microbiology,2009,134(1):29-36.
[22] 逯杏花,潘玲.禽源金黄色葡萄球菌的分离鉴定及耐药性分析[J].中国家禽,2014,36(20):63-64. LU X H,PAN L.Isolation,identification and drug resistance analysis of avian-derived Staphylococcus aureus[J].Chinese Poultry,2014,36(20):63-64.(in Chinese)
[23] 千国胜,余维根,舒鑫标,等.种鸭化脓性关节炎病原分析[J].中国家禽,2018,40(3):66-69. QIAN G S,YU W G,SHU X B,et al.Analysis of the pathogen of purulent arthritis in breeding ducks[J].Chinese Poultry,2018,40(3):66-69.(in Chinese)
[24] 戈胜强,柴同杰,秦梅.奶牛乳房炎葡萄球菌的分离鉴定及耐药性检测[J].西北农林科技大学学报(自然科学版),2008,36(11):40-46. GE S Q,CHAI T J,QIN M.Isolation identification,medicine sensitivity test of Staphylococcus of dairy cow mastitis[J].Journal of Northwest A&F University (Natural Science Edition),2008,36(11):40-46.(in Chinese)
[25] LASAGNO M,ORTIZ M,VISSIO C,et al.Pathogenesis and inflammatory response in experimental caprine mastitis due to Staphylococcus chromogenes[J].Microbial Pathogenesis,2018,116:146-152.
[26] DOS SANTOS D C,LANGE C C,AVELLAR-COSTA P,et al. Staphylococcus chromogenes,a coagulase-negative Staphylococcus species that can clot plasma[J].Journal of Clinical Microbiology,2016,54(5):1372-1375.