一株裂解性奶牛乳房炎源屎肠球菌噬菌体的分离及其生物学特性分析

doi:10.11843/j.issn.0366-6964.2017.04.014

Abstract

Abstract:

The aim of the present study was to isolate and characterize the phage that could lyse Enterococcus faecium from fecal samples of dairy cattle in Shihezi, Xinjiang, and provide the basic data for the phage therapy as well as prevention. The phage was isolated and purified from dairy cattle fecal samples by double-layer agar plate method. And its plaque characteristics was observed. The purified phage was concentrated, negatively stained with phosphotungstic acid and observed by transmission electron microscopy. The genome of the isolated phage was sequenced. Its genetic and evolutional history were analyzed. The host range, optimal multiplicity of infection (MOI), one-step growth curve, temperature, and pH stability of the phage were investigated by double-layer agar plate methods. A lytic phage of Enterococcus faecium was isolated and purified, and named vB_EfaM_XJ3. The phage plaques were circular, clear and transparent with smooth edge, and its diameter was 1.2-1.5 mm. The electron microscope observation showed that the phage particles, whose head was twenty-sided, head length was about 53.15 nm, tail length was about 14.42 nm×199.62 nm. The genome of the phage was comprised of double strand DNA, the size was 157.189 kb with GC content of 50.27%. The homology of amino acid sequence of phage vB_EfaM_XJ3 with Salmonella phage SKML-39 was 98%. The study of biological characteristics showed that the vB_EfaM_XJ3 only crack its host bacteria at present. The optimal MOI was 0.001, the incubation period was 15 min, the burst period was 175 min and the burst size was 84. The phage could withstand the temperature up to 50℃ and keep stable titer under pH 5.0-11.0. A lytic phage of Enterococcus faecium was isolated and identified. Its biological characteristics and genetic were analyzed. The results laid the foundation for the further research and application of the antibacterial agent for Salmonella and Enterococcus faecium.

CLC Number:

S852.61

ZHANG Qian, YU Hui-ju, SUN Yao-qiang, ZHANGXIANG Li-lan, ZHANG Pei-sheng, LIU Ge, QU Yong-gang, TONG Yi-gang, LI Yan. Isolation and Characterization of a Lytic Phage Infecting Enterococcus faecium of Bovine Mastitis[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2017, 48(4): 706-713.

0
/ / Recommend

Add to citation manager EndNote|Reference Manager|ProCite|BibTeX|RefWorks

URL: https://www.xmsyxb.com/EN/10.11843/j.issn.0366-6964.2017.04.014

https://www.xmsyxb.com/EN/Y2017/V48/I4/706

References

[1] 阚威. 奶牛乳房炎链球菌和肠球菌的分离鉴定及毒力基因的检测[D]. 兰州:甘肃农业大学, 2014.
KAN W. Isolation, characerization and virulence gene analysis of Streptococcus and Enterococcus collected from bovine mastitis[D]. Lanzhou:Gansu Agricultural University, 2014. (in Chinese)
[2] 董翔宸. 奶牛乳房炎肠球菌ESP基因真核重组质粒的构建与表达[D]. 兰州:甘肃农业大学, 2015.
DONG X C. Construction and expression of eukaryotic vector of ESP gene from bovine Enterococcus[D]. Lanzhou:Gansu Agricultural University, 2015. (in Chinese)
[3] KLIMIEN? I, RU?AUSKAS M, ŠPAKAUSKAS V, et al. Prevalence of gram positive bacteria in cow mastitis and their susceptibility to beta-lactam antibiotics[J]. Vet Med Zoot, 2011, 56(78):65-72.
[4] 齐亚银, 剡根强, 王静梅, 等. 致羔羊脑炎型粪肠球菌的分离及鉴定[J]. 石河子大学学报:自然科学版, 2005, 23(2):200-202.
QI Y Y, YAN G Q, WANG J M, et al. Isolation and identification of Enterococcus faecalis in lambs with meningitis[J]. Journal of Shihezi University (Natural Science), 2005, 23(2):200-202. (in Chinese)
[5] WALL S K, ZHANG J, ROSTAGNO M H, et al. Phage therapy to reduce preprocessing Salmonella infections in market-weight swine[J].Appl Environ Microbiol, 2010, 76(1):48-53.
[6] 张哲. 两株裂解性粪肠球菌噬菌体的分离鉴定及生物学特性分析[D]. 长春:吉林大学, 2015.
ZHANG Z. Isolation and biological characteristic of two lytic Enteroocus bacteriophages active against Enterococcus faecalis[D]. Changchun:Jilin University, 2015. (in Chinese)
[7] BISWAS B, ADHYA S, WASHART P, et al. Bacteriophage therapy rescues mice bacteremic from a clinical isolate of vancomycin-resistant Enterococcus faecium[J]. Infect Immun, 2002, 70(1):204-210.
[8] 宫鹏娟, 张蕾, 夏翡翡, 等. 屎肠球菌噬菌体m5的分离鉴定及其对小鼠菌血症的治疗研究[J]. 中国兽医学报, 2016, 36(5):746-749,762.
GONG P J,ZHANG L, XIA F F, et al. Bacteriophage m5 therapy rescues mice bacteremic from a clinical isolate of vancomycin-resistant Enterococcus faecium[J]. Chinese Journal of Veterinary Science, 2016, 36(5):746-749,762.(in Chinese)
[9] JAMALLUDEEN N,JOHNSON R P,FRIENDSHIP R, et al. Isolation and characterization of nine bacteriophages that lyse O149 enterotoxigenic Escherichia coli[J]. Vet Microbiol, 2007, 124:47-57.
[10] 萨姆布鲁克J, 拉塞尔D W. 分子克隆实验指南[M]. 3版.黄培堂,译.北京:科学出版社, 2002.
SAMBROOK J, RUSSELL D W. Molecular cloning:A laboratory manual[M]. 3rd ed. HUANG P T, trans. Beijing:Science Press, 2002. (in Chinese)
[11] 余静丹. 鲍曼不动杆菌噬菌体的分离鉴定及生物学特性的研究[D]. 长春:吉林大学, 2013.
YU J D. Isolation, characterization and biological features of lytic bacteriophages infecting Acinetobacter baumannii clinical isolates[D]. Changchun:Jilin University, 2013(in Chinese).
[12] NASU H, ⅡDA T, SUGAHARA T, et al. A filamentous phage associated with recent pandemic Vibrio parahaemolyticus O3:K6 strains[J]. J Clin Microbiol, 2000, 38(6):2156-2161.
[13] 冯烨, 刘军, 孙洋, 等. 噬菌体最新分类与命名[J]. 中国兽医学报, 2013, 33(12):1954-1958.
FENG Y, LIU J, SUN Y, et al. An introduction to current classification and nemenclature of bacterial viruses[J]. Chinese Journal of Veterinary Science, 2013, 33(12):1954-1958.(in Chinese)
[14] 包红朵, 张鹏禹, 周艳, 等. 沙门氏菌裂解性噬菌体的分离鉴定及其生物学特性[J]. 食品科学, 2015, 36(5):131-136.
BAO H D, ZHANG P Y, ZHOU Y, et al. Isolation and biological characteristics of lytic Salmonella bacteriophage[J]. Food Science, 2015, 36(5):131-136.(in Chinese)
[15] BUENO E, GARCÍA P, MARTÍNEZ B, et al. Phage inactivation of Staphylococcus aureus in fresh and hard-type cheeses[J]. Int J Food Microbiol, 2012, 158(1):23-27.
[16] FAUQUET C M, MAYO M A, MANILOFF J, et al. Virus taxonomy:Ⅷth report of the international committee on taxonomy of viruses[M]. Amsterdam:Elsevier Academic Press, 2005.
[17] KING A M Q, ADAMS M J, CARSTENS E B, et al. Virus taxonomy:ninth report of the international committee on taxonomy of viruses[M]. Amsterdam:Elsevier Academic Press, 2011.
[18] ANANY H, LINGOHR E J, VILLEGAS A, et al. A Shigella boydii bacteriophage which resembles Salmonella phage ViI[J]. Virol J, 2011, 8(1):242.
[19] 张文惠, 安小平, 范航, 等. 一株粪肠球菌噬菌体的分离及其生物学特性研究[J]. 生物技术通讯, 2013,24(4):484-487.
ZHANG W H, AN X P, FAN H, et al. Isolation and characterization of an Enterococcus faecalis phage[J]Letters in Biotechnology, 2013, 24(4):484-487. (in Chinese)
[20] 邢少贞, 张湘莉兰, 舒鹏, 等. 粪肠球菌噬菌体vB_E. faecalis_IME196的生物学特性及其全基因组分析[J]. 微生物学通报,2016, 43(9):2040-2048.
XING S Z, ZHANGXIANG L L, SHU P, et al. Characterization and complete genome sequence analysis of Enterococcus faecalis bacteriophage vB_E. faecalis_IME196[J]. Microbiology China, 2016, 43(9):2040-2048. (in Chinese)

Isolation and Characterization of a Lytic Phage Infecting Enterococcus faecium of Bovine Mastitis

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

References

Related Articles 15

Recommended Articles

Metrics

Comments

[1]	CHONG Qian, ZHAO Xuehui, CAO Qing, XUE Huiwen, GOU Huitian. Research Progress of Applied Research on Listeria monocytogenes Phage [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(5): 1819-1826.
[2]	XU Zhenyu, DENG Xiaoyu, WANG Yueli, SUN Can, WU Aodi, CAO Jian, YI Jihai, WANG Yong, WANG Zhen, CHEN Chuangfu. Biological Characteristics of Brucella abortus A19ΔBtpA Deletion Strain and Its Immunogenicity Study [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(5): 2135-2145.
[3]	ZHAO Canqi, FENG Yu, LÜ Lang, LI Yanjun, WEI Yulei, DING Jiabo, CHEN Xiang, JIANG Hui. Study on Purification of Bovine Brucellosis by Competitive ELISA and Indirect ELISA [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(5): 2146-2153.
[4]	GAO Jie, LI Xiaocheng, MU Yang, ZHANG Hui, WEI Rong, LI Jie. Biological Characteristics and Immune Effect Evaluation of Outer Membrane Vesicles of Capsular Type B Pasteurella multocida [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(5): 2168-2175.
[5]	FEI Guoqing, NING Zhiyuan, ZHAO Zefang, LIU Yanqiu, LIU Tengfei, LI Xian, CONG Rihua, CHEN Hong, CHEN Shulin. Isolation, Identification of Luteal Cells from Cows during Pregnancy and Investigation of Their Culture Characteristics [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(5): 2214-2225.
[6]	HE Xiaolan, ZHAO Yankun, MENG Lu, LIU Huimin, GAO Jiaojiao, ZHENG Nan. Research Progress in Heteroresistance of Staphylococcus aureus [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(4): 1432-1445.
[7]	DAI Fan, LIU Zhanyou, ZHANG Xuyang, LI Wu. Aconitate Decarboxylase 1 Could Regulate the Inflammatory Response Caused by BCG [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(4): 1696-1706.
[8]	TIAN Rui, XU Sixiang, XIE Feng, LIU Guangjin, WANG Gang, LI Qingxia, DAI Lei, XIE Guoxin, ZHANG Qiongwen, LU Yajing, WANG Guangwen, WANG Jinxiu, ZHANG Wei. Bioinformatics Analysis of the Genome of Clostridium perfringens Isolated from Cattle [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(4): 1707-1715.
[9]	JIN Ruiyan, ZHANG Yang, HUANG Zhongshun, RAN Wei, DING Honglei. Phenotype Analysis of Antimicrobial Resistance and β-lactamase, PMQR Genes Detection of Escherichia coli Isolated from Mawang Ducks [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(4): 1738-1746.
[10]	GAO Yuanji, LIU Chang, CHEN Miao, CHEN Songbiao, ZHANG Junfeng, LI Jing, JIA Yanyan, LIAO Chengshui, GUO Rongxian, DING Ke, YU Zuhua, SHANG Ke. Structure, Secretory Characteristics, and Pathogenic Mechanism of Bacterial Outer Membrane Vesicles [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(3): 971-983.
[11]	LI Pengxu, LI Shijing, SUN Jun, XIANG Wei, ZHAO Miaomiao, HOU Tianmu, LI Huaming, GUANG Min, CHEN Ruige, XU Mengran, WU Xiaomin, JIANG Hexiang, LEI Liancheng, ZHANG Fuxian. Molecular Subtyping and Identification of Streptococcus suis Meningitidis Type 2 and Its Biological Characteristics [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(3): 1192-1207.
[12]	SU Wennan, LIU Jiaqi, ZHONG Jiacheng, CHEN Jidang, ZHU Wanjun, ZHANG Yishan, ZHANG Jipei. Complete Genome Re-sequence and Comparative Genomic Analysis of Avibacterium paragallinarum from Geese [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(3): 1208-1216.
[13]	FU Xuezhen, LI Xincan, QIAN Hongyu, LÜ Hong, WU Chanyu, WANG Xiaohan, WANG Xiaohua, WANG Zhiying, ZHOU Zuoyong. Antibacterial Effect and Mechanism of Bergamot Essential Oil on Corynebacterium pseudotuberculosis [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(3): 1217-1227.
[14]	QIN Yi, HU Wenjie, FANG Xiaowei, GUO Qian, TIAN Lanxin, LIU Fang, FANG Chun. Effect of Deletion of the Lipoteichoic Acid Synthase ltaS Gene on the Pathogenicity of Listeria monocytogenes [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(2): 670-679.
[15]	SONG Yan, YUAN Yongfeng, QIAN Hongyu, LI Xincan, LUO Hongyan, WANG Zhiying, ZHOU Zuoyong. Identification and Partial Biological Characteristics Analysis of Corynebacterium pseudotuberculosis Isolated from Goats [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(2): 680-687.