穿心莲内酯影响猪链球菌生物被膜形成

doi:10.11843/j.issn.0366-6964.2018.03.021

摘要/Abstract

摘要：

旨在探究穿心莲内酯对猪链球菌生物被膜形成的抑制作用，为猪链球菌病的耐药性提供替代疗法。试验采用微量稀释法测定穿心莲内酯对猪链球菌的最小抑菌浓度（MIC）和最小杀菌浓度（MBC）；利用结晶紫染色（CV）和扫描电镜（SEM）检测穿心莲内酯对猪链球菌生物被膜形成的影响。结果表明：1）穿心莲内酯对猪链球菌的MIC和MBC分别为0.25和0.5 mg·mL^-1；2）扫描电镜观察可见阴性对照组细菌镶嵌于生物被膜中，而穿心莲内酯在亚抑菌浓度下（1/2MIC、1/4MIC、1/8MIC）能使细菌生物被膜中的细菌数量和生物被膜形成量均减少，并呈剂量依赖性；3）luxS基因和毒力基因的RT-PCR检测显示亚抑菌浓度（1/2MIC、1/4MIC）的穿心莲内酯明显降低gapdh、sly、fbps、ef和luxS基因的转录，显著增加cps2J、mrp和gdh基因的转录；4）哈维氏弧菌生物发光法测定细菌培养液显示穿心莲内酯在亚抑菌浓度下（1/2MIC、1/4MIC、1/8MIC）对猪链球菌AI-2信号分子的产生有抑制作用，呈剂量依赖性。穿心莲内酯亚抑菌浓度的干预可抑制猪链球菌生物被膜的形成，并且可致毒力基因表达水平差异。

Abstract:

The aim of this study was to evaluate the inhibitory effect of Andrographolide on the Streptococcus suis. The dilution method was conducted to determine the minimum inhibitory biofilm formation of S. suis and to provide an alternative therapy for the drug resistance of concentration (MIC) and minimum bactericidal concentration (MBC) of Andrographolide on S. suis; The effects of Andrographolide on the biofilm formation of S. suis were detected by crystal violet staining (CV) and scanning electron microscopy (SEM). Result were as follows:1)The MIC and MBC of Andrographolide on S. suis were 0.25 and 0.5 mg·mL^-1, respectively. 2)Scanning electron microscopy showed that the bacteria were embedded in the biofilm in the blank control group. Andrographolide(1/2MIC, 1/4MIC, 1/8MIC)can make the number of bacteria in the bacterial biofilm and the morphology of the biofilm changed greatly. According to the dose of the trend changed, bacterial biofilm gradually reduced. The results showed that Andrographolide had inhibitory effect on the formation of S. suis biofilm in a dose-dependent manner. 3)Through quantitatively detected the expression of luxS gene and virulence gene in S. suis by RT-PCR, we can see the sub-MICs (1/2MIC, 1/4MIC) of Andrographolide can respectively reduce the expression of gapdh, sly, fbps, ef and luxS genes and significantly increase the expression of cps2J, mrp, and gdh genes. 4)Detected the bacterial culture medium by Bioluminescence of Vibrio harveyi showed that Andrographolide (1/2MIC, 1/4MIC, 1/8MIC) had inhibitory effect on the production of AI-2 signal molecule in S. suis, in a dose-dependent manner. Intervention of Andrographolide can inhibit the formation of biofilm in S. suis, and can cause the difference of virulence gene expression levels.

中图分类号:

S853.7

王金玉, 姜晓文, 王娜, 王婕然, 于苗苗, 钟秋, 于文会. 穿心莲内酯影响猪链球菌生物被膜形成[J]. 畜牧兽医学报, 2018, 49(3): 629-636.

WANG Jin-yu, JIANG Xiao-wen, WANG Na, WANG Jie-ran, YU Miao-miao, ZHONG Qiu, YU Wen-hui. Effect of Andrographolide on the Biofilm Formation of Streptococcus suis[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2018, 49(3): 629-636.

0
/ / 推荐

导出引用管理器 EndNote|Reference Manager|ProCite|BibTeX|RefWorks

链接本文: https://www.xmsyxb.com/CN/10.11843/j.issn.0366-6964.2018.03.021

https://www.xmsyxb.com/CN/Y2018/V49/I3/629

参考文献

[1] LUN Z R, WANG Q P, CHEN X G, et al. Streptococcus suis:an emerging zoonotic pathogen[J]. Lancet Infect Dis, 2007, 7(3):201-209.
[2] ZHANG Y Y, DING D D, LIU M L, et al. Effect of the glycosyltransferases on the capsular polysaccharide synthesis of Streptococcus suis serotype 2[J]. Microbiol Res, 2016, 185:45-54.
[3] MOHAMMADI Z, PALAZZI F, GIARDINO L, et al. Microbial biofilms in endodontic infections:an update review[J]. Biomed J, 2013, 36(2):59-70.
[4] 尤理想, 赵青, 周敏, 等. 细菌生物膜检测方法改进与应用[J]. 实验技术与管理, 2015, 32(3):72-76.
YOU L X, ZHAO Q, ZHOU M, et al. Improved method of bacterial biofilm detection and its application[J]. Experimental Technology and Management, 2015, 32(3):72-76. (in Chinese)
[5] MAN N K, DEGREMONT A, DARBORD J C, et al. Evidence of bacterial biofilm in tubing from hydraulic pathway of hemodialysis system[J]. Artif Organs, 1998, 22(7):596-600.
[6] SERRA D O, RICHTER A M, KLAUCK G, et al. Microanatomy at cellular resolution and spatial order of physiological differentiation in a bacterial biofilm[J]. mBio, 2013, 4(2):00103-13.
[7] DE LA FUENTE-NÚÑEZ C, REFFUVEILLE F, FERNÁNDEZ L, et al. Bacterial biofilm development as a multicellular adaptation:antibiotic resistance and new therapeutic strategies[J]. Curr Opin Microbiol, 2013, 16(5):580-589.
[8] CASTELO-BRANCO D S C M, RIELLO G B, VASCONCELOS D C, et al. Farnesol increases the susceptibility of Burkholderia pseudomallei biofilm to antimicrobials used to treat melioidosis[J]. J Appl Microbiol, 2016, 120(3):600-606.
[9] WANG S, YANG Y B, ZHAO Y L, et al. Sub-MIC tylosin inhibits Streptococcus suis biofilm formation and results in differential protein expression[J]. Front Microbiol, 2016, 7:384.
[10] CALZAS C, LEMIRE P, AURAY G, et al. Antibody response specific to the capsular polysaccharide is impaired in Streptococcus suis serotype 2-infected animals[J]. Infect Immun, 2015, 83(1):441-453.
[11] SANTHAKUMARI S, KANNAPPAN A, PANDIAN S K, et al. Inhibitory effect of marine cyanobacterial extract on biofilm formation and virulence factor production of bacterial pathogens causing vibriosis in aquaculture[J]. J Appl Phycol, 2016, 28(1):313-324.
[12] HUSAIN F M, AHMAD I. Doxycycline interferes with quorum sensing-mediated virulence factors and biofilm formation in Gram-negative bacteria[J]. World J Microbiol Biotechnol, 2013, 29(6):949-957.
[13] ALASIL S M, OMAR R, ISMAIL S, et al. Inhibition of quorum sensing-controlled virulence factors and biofilm formation in Pseudomonas aeruginosa by culture extract from novel bacterial species of Paenibacillus using a rat model of chronic lung infection[J]. Int J Bacteriol, 2015, 2015:671562.
[14] KIM B S, PARK S J, KIM M K, et al. Inhibitory effects of Chrysanthemum boreale essential oil on biofilm formation and virulence factor expression of Streptococcus mutans[J]. Evid Based Complement Alternat Med, 2015, 2015:616309.
[15] ZIUZINA D, BOEHM D, PATIL S, et al. Cold plasma inactivation of bacterial biofilms and reduction of quorum sensing regulated virulence factors[J]. PLoS One, 2015, 10(9):e0138209.
[16] KASIMANICKAM V R, OWEN K, KASIMANICKAM R K. Detection of genes encoding multidrug resistance and biofilm virulence factor in uterine pathogenic bacteria in postpartum dairy cows[J]. Theriogenology, 2015, 85(2):173-179.
[17] CHOI N Y, KANG S Y, KIM K J. Artemisia princeps inhibits biofilm formation and virulence-factor expression of antibiotic-resistant bacteria[J]. Biomed Res Int, 2015, 2015:239519.
[18] AVIN S, AVINASH A, PATIL N, et al. Recent advances in the development of antibacterial agents[J]. Res J Pharm Biol Chem Sci, 2015, 6(5):1220-1228.
[19] HSU Y H, HSU Y L, LIU S H, et al. Development of a bifunctional andrographolide-based chemical probe for pharmacological study[J]. PLoS One, 2016, 11(4):e0152770.
[20] 于文会, 许晶, 魏庆微, 等. 蒲公英水提物对猪链球菌生物被膜体外干预作用[J]. 畜牧兽医学报, 2015, 46(10):1875-1881.
YU W H, XU J, WEI Q W, et al. Dandelion aqueous extract affects biofilm formation by S. suis[J]. Acta Veterinaria et Zootechnica Sinica, 2015, 46(10):1875-1881. (in Chinese)
[21] CHOI S C. Effects of bacterial biofilms and iron oxide on copper distribution in a simulated marine environment:an implication to the sediments in the Pearl River estuary[D]. Hong Kong, China:The Hong Kong Polytechnic University, 2003:189-230.
[22] 张文元, 杨亚冬, 唐靓. 十四元环大环内酯类对细菌生物被膜形成的影响及对环丙沙星的增效作用[J]. 中国卫生检验杂志, 2005, 15(11):1288-1289.
ZHANG W Y, YANG Y D, TANG L. Effects of 14-membered macrolides on the formation of Pseudomonas aeruginosa biofilms and synergism between macrolide and ciprofloxacin[J]. Chinese Journal of Health Laboratory Technology, 2005, 15(11):1288-1289. (in Chinese)
[23] VAN CALSTEREN M R, GAGNON F, LACOUTURE S, et al. Structure determination of Streptococcus suis serotype 2 capsular polysaccharide[J]. Biochem Cell Biol, 2010, 88(3):513-525.
[24] DOBRETSOV S, QIAN P Y. The role of epibotic bacteria from the surface of the soft coral Dendronephthya sp. in the inhibition of larval settlement[J]. J Exp Mar Biol Ecol, 2004, 299(1):35-50.
[25] 吴恋, 王春艳, 吕维, 等. 藏药大车前草中大车前苷的定性和定量分析[J]. 成都中医药大学学报, 2014, 37(1):48-50, 59.
WU L, WANG C Y, LV W, et al. Qualitative and quantitative analysis of plantamajoside in Plantago major L.[J]. Journal of Chengdu University of TCM, 2014, 37(1):48-50, 59. (in Chinese)
[26] Vaccine composition against Streptococcus suis infection:EP, 2949340[P]. 2015-12-02.
[27] HUANG Y H, CAI T G, XIA X, et al. Research advances in the intervention of inflammation and cancer by active ingredients of traditional Chinese medicine[J]. J Pharm Pharm Sci, 2016, 19(1):114-126.
[28] CALLEJO R, PRIETO M, ROCCA F, et al. Molecular characterization of Streptococcus suis serotype 2 strains isolated from humans and diseased pigs in Argentina[C/OL]//Proceedings of Xix Lancefield International Symposium on Streptococci and Streptococcal Diseases. Buenos Aires, Argentina, 2014.[2017-12-26].https://www.researchgate.net/publication/268222687_MOLECULAR_CHARACTERIZATION_OF_STREPTOCOCCUS_SUIS_SEROTYPE_2_STRAINS_ISOLATED_FROM_HUMANS_AND_DISEASED_PIGS_IN_ARGENTINA
[29] 汪洋. 猪链球菌生物被膜形成及致病机理研究[D]. 南京:南京农业大学, 2011:1-10.
WANG Y. Study on the mechanism of biofilm formation and molecular pathogenesis of Streptococcus suis[D]. Nanjing:Nanjing Agricultural University, 2011:1-10. (in Chinese)
[30] SAHA M, WAHL M. Seasonal variation in the antifouling defence of the temperate brown alga Fucus vesiculosus[J]. Biofouling, 2013, 29(6):661-668.
[31] LUTZ L, PEREIRA D C, PAIVA R M, et al. Macrolides decrease the minimal inhibitory concentration of anti-pseudomonal agents against Pseudomonas aeruginosa from cystic fibrosis patients in biofilm[J]. BMC Microbiol, 2012, 12:196.