猪链球菌选择性培养基优化
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摘要
目的对原有的猪链球菌选择性培养基进行优化改良,用于猪源样本中猪链球菌的分离。方法比较猪链球菌在其他高营养培养基上的生长适宜性;进一步通过药物敏感实验筛选对猪链球菌2型不敏感的抗菌药物,继而用琼脂平板稀释法测定候选抗菌药物的最小抑菌浓度,组方后用现场标本考核抑菌效果,用模拟样本和猪源标本考核分离效果。结果使用特殊蛋白胨替代原培养基中的酵母粉,可改善猪链球菌生长状态,使菌落更易于挑取。筛选出含有氨曲南50 mg/L,多粘菌素B 10 mg/L,链霉素40 000 U/L,结晶紫0.5 mg/L,呋喃妥因6 mg/L的改良选择性培养基,该培养基对猪源标本中常见的非目标菌抑制作用更强。在实验室模拟样本中,使用直接涂布法,检测猪链球菌灵敏度为1×103CFU/ml;在猪源样本中,经选择性脑心肉汤增菌后,改良选择性培养基可使猪链球菌检出率达到17.1%。结论改良选择性培养基对猪链球菌具有更强的选择作用,使用该培养基可以提高检测灵敏度,更适合动物源性标本中猪链球菌的分离培养。
Objective The original selective medium from pig-derived samples was modified for the separation of Streptococcus suis(S. suis). Methods S. suis was inoculated on different high nutrient culture media,and the colony morphology and the growth rate were observed. We further screened the antibacterial agents that were not sensitive to S. suis type 2 by drug sensitivity analysis and determined the minimum inhibitory concentration of the candidates by plate dilution method. After we had compounded the appropriate antibacterial agents,the clinical samples were used to evaluate the antibacterial effect, while the simulation and pig-derived samples were used to assess the separation efficiency. Results The yeast powder in the original medium was replaced by peptone special, it can improve the growth statue of S. suis, and the bacterial colony was easier to picking,aztreonam(50 mg/L),polymyxin B(10 mg/L),streptomycin(40 000 U/L),crystal violet(0.5 mg/L) and nitrofurantoin(6 mg/L) were screened out from modified selective medium,which had a stronger inhibitory effect on nontarget bacteria. In the simulation experiments,the sensitivity of modified medium was 1 × 103 CFU/ml by direct spreading. After being incubated in selective brain heart infusion broth, the pigderived samples were spread on the modified selective medium, and the positive rate of S. suis raised to 17.1%. Conclusion The modified selective medium was strongly selective to S. suis,which increased the detection sensitivity significantly. Therefore,the modified selective medium was more suitable for the separation of S. suis from animal-derived samples.
Objective The original selective medium from pig-derived samples was modified for the separation of Streptococcus suis(S. suis). Methods S. suis was inoculated on different high nutrient culture media,and the colony morphology and the growth rate were observed. We further screened the antibacterial agents that were not sensitive to S. suis type 2 by drug sensitivity analysis and determined the minimum inhibitory concentration of the candidates by plate dilution method. After we had compounded the appropriate antibacterial agents,the clinical samples were used to evaluate the antibacterial effect, while the simulation and pig-derived samples were used to assess the separation efficiency. Results The yeast powder in the original medium was replaced by peptone special, it can improve the growth statue of S. suis, and the bacterial colony was easier to picking,aztreonam(50 mg/L),polymyxin B(10 mg/L),streptomycin(40 000 U/L),crystal violet(0.5 mg/L) and nitrofurantoin(6 mg/L) were screened out from modified selective medium,which had a stronger inhibitory effect on nontarget bacteria. In the simulation experiments,the sensitivity of modified medium was 1 × 103 CFU/ml by direct spreading. After being incubated in selective brain heart infusion broth, the pigderived samples were spread on the modified selective medium, and the positive rate of S. suis raised to 17.1%. Conclusion The modified selective medium was strongly selective to S. suis,which increased the detection sensitivity significantly. Therefore,the modified selective medium was more suitable for the separation of S. suis from animal-derived samples.
引文
[1]Vecht U,Van Leengoed LA,Verheijen ER. Streptococcus suis infections in pigs in the Netherlands(Part I)[J].Vet Q,1985,7(4):315-321.
[2]罗隆泽,刘莉,李燕春,等.四川省人感染猪链球菌病的病原分离与鉴定[J].预防医学情报杂志,2005,21(4):386-387.
[3]Kataoka Y,Sugimoto C,Nakazawa M,et al. Detection of Streptococcus suis type 2 in tonsils of slaughtered pigs using improved selective and differential media[J].Veterinary Microbiology,1991,28(4):335-342.
[4]罗隆泽,李燕春,郭宗琪,等.猪链球菌选择性培养基研究[J].预防医学情报杂志,2007,23(5):511-514.
[5]Performance standards for antimicrobial susceptibility testing. M100-S20[S]. Clinical and Laboratory Standards Institute,2010.
[6]Gottschalk M, Segura M. The pathogenesis of the meningitis caused by Streptococcus suis:the unresolved questions[J]. Veterinary Microbiology,2000,76(3):259-272.
[7]Huang YT, Teng LJ, Ho SW, et al. Streptococcus suis infection[J]. Journal of microbiology,immunology,and infection,2005,38(5):306-313.
[8]Staats JJ,Feder I,Okwumabua O,et al. Streptococcus Suis:Past and Present[J]. Veterinary Research Communications,1997,21(6):381-407.
[9]Changyun Y, Xuemei B, Ji Z, et al. Spread of Streptococcus suis sequence type 7, China[J].Emerging Infectious Diseases,2008,14(5):787-791.
[10]Perch B, Kristjansen P, Skadhauge K. Group R streptococci pathogenic for man. Two cases of meningitis and one fatal case of sepsis[J]. Acta Pathol Microbiol Scand,1968,74(1):69-76.
[11]黄毓茂,黄引贤.2型猪链球菌的血清学鉴定[J].中国兽医学报,1995(1):63-65.
[12]廖虹瑜,罗隆泽,曾林子,等.2006-2016年四川省猪链球菌血清2型多位点序列分型分析[J].疾病监测,2017,32(7):544-547.
[13]Tang J,Wang C,Feng Y,et al. Streptococcal toxic shock syndrome caused by Streptococcus suis serotype 2[J]. Plos Medicine,2006,3(5):151-151.
[14]Rosendal S,Breton J,Henrichsen J,et al. Isolation of Streptococcus suis using a selective medium[J].Canadian journal of veterinary research,1986,50(4):537-539.
[15]Palmieri C,Varaldo PE, Facinelli B. Streptococcus suis, an Emerging Drug-Resistant Animal and Human Pathogen[J]. Frontiers in Microbiology,2011,2(1):235.
[2]罗隆泽,刘莉,李燕春,等.四川省人感染猪链球菌病的病原分离与鉴定[J].预防医学情报杂志,2005,21(4):386-387.
[3]Kataoka Y,Sugimoto C,Nakazawa M,et al. Detection of Streptococcus suis type 2 in tonsils of slaughtered pigs using improved selective and differential media[J].Veterinary Microbiology,1991,28(4):335-342.
[4]罗隆泽,李燕春,郭宗琪,等.猪链球菌选择性培养基研究[J].预防医学情报杂志,2007,23(5):511-514.
[5]Performance standards for antimicrobial susceptibility testing. M100-S20[S]. Clinical and Laboratory Standards Institute,2010.
[6]Gottschalk M, Segura M. The pathogenesis of the meningitis caused by Streptococcus suis:the unresolved questions[J]. Veterinary Microbiology,2000,76(3):259-272.
[7]Huang YT, Teng LJ, Ho SW, et al. Streptococcus suis infection[J]. Journal of microbiology,immunology,and infection,2005,38(5):306-313.
[8]Staats JJ,Feder I,Okwumabua O,et al. Streptococcus Suis:Past and Present[J]. Veterinary Research Communications,1997,21(6):381-407.
[9]Changyun Y, Xuemei B, Ji Z, et al. Spread of Streptococcus suis sequence type 7, China[J].Emerging Infectious Diseases,2008,14(5):787-791.
[10]Perch B, Kristjansen P, Skadhauge K. Group R streptococci pathogenic for man. Two cases of meningitis and one fatal case of sepsis[J]. Acta Pathol Microbiol Scand,1968,74(1):69-76.
[11]黄毓茂,黄引贤.2型猪链球菌的血清学鉴定[J].中国兽医学报,1995(1):63-65.
[12]廖虹瑜,罗隆泽,曾林子,等.2006-2016年四川省猪链球菌血清2型多位点序列分型分析[J].疾病监测,2017,32(7):544-547.
[13]Tang J,Wang C,Feng Y,et al. Streptococcal toxic shock syndrome caused by Streptococcus suis serotype 2[J]. Plos Medicine,2006,3(5):151-151.
[14]Rosendal S,Breton J,Henrichsen J,et al. Isolation of Streptococcus suis using a selective medium[J].Canadian journal of veterinary research,1986,50(4):537-539.
[15]Palmieri C,Varaldo PE, Facinelli B. Streptococcus suis, an Emerging Drug-Resistant Animal and Human Pathogen[J]. Frontiers in Microbiology,2011,2(1):235.
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