Genitic information and biological characteristics of mycobacteriophage Chy5
JIANG Li-sha1, LIU Ping3, WU Ting-ting4, GUO Shu-liang2
1. Department of Respiratory Medicine, The First Branch, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400010,China; 2. Department of Respiratory Medicine, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016,China; 3. Department of Respiratory Medicine, People’s Hospital of Changshou District, Chongqing 401220,China; 4. Department of Respiratory Medicine, The Third People’s Hospital of Chengdu, Chengdu 610031,China;
Abstract：To select the phage infecting Mycobacterium tuberculosis, and understand its biological characteristics and genetic information, mycobacteriophage was isolated and purified from soil. The morphological characteristics were observed by electron-microscopy, the host range was then examined by single-spot examination. Mycobacteriophage DNA was extracted. Shutgun sequences were assembled, then the gaps were filled with PCR fragments. DNAStar software Packages was used to analyze the general characteristic of genome, Glimmer 3.0 and GeneMark were applied to predict the function of the coding gene. In addition, the collinearity analysis were conducted. Lysogens was isolated to conduct an Ultraviolet induction test and super infection immunity. In results, a novel mycobacteriophage Chy5 was isolated. The head diameter was (61.5±1.4 nm) and the tail length was (114.1±2.1 nm). It could infect different clinical isolates of Mycobacterium tuberculosis, which was classified into Siphoviridae. The full length of Chy5 genome was 51 214 bp with a G+C content of 63.60%. Chy5 genome was most similar with D29, which were 88 predicted genes encoding integrase and repressor as well as attP site. The Chy5 lysogen could release free phage particles to form plaques after UV induction. Chy5 could not clear lawn of Chy5 lysogens. So Chy5 is a novel lysogenic mycobacteriophage, which could lyse mycobacterium tuberculosis and be applied to construct Luciferase reporter phages(LRP).
[1] Kumar V, Loganathan P, Sivaramakrishnan G, et al.Characterization of temperate phage Che12 and construction of a new tool for diagnosis of tuberculosis[J].Tuberculosis, 2008, 88(6): 616-623. DOI: 10.1016/j.tube.2008.02.007 [2] Sivaramakrishnan G, Subramanyam BCP, Kumar V.Luciferase reporter phage phAE85 for rapid detection of rifampicin resistance in clinical isolates of Mycobacterium tuberculosis[J]. Asian Pac J Trop Med,2013,6(9):728-731. DOI: 10.1016/S1995-7645(13)60127-3 [3] O’Donnell MR, Pym A, Jain PA, et al. novel reporter phage to detect tuberculosis and rifampin resistance in a high-HIV-burden population[J].J Clin Microbiol,2015,53(7):2188-2194. DOI: 10.1128/JCM.03530-14 [4] Guo S,Ao Z.Phage in the diagnosis and treatment of tuberculosis[J].Front Bioaci(Landmark Ed),2012,17:2691-2697. DOI: 10.2741/4080 [5] RybnikerJ,Kramme S, Small PL. Host range of 14 mycobacteriophages in Mycobacterium ulcerans and seven other Mycobacteria including Mycobacterium tuberculosis-application for identification and susceptibility testing[J].J Med Microbiology,2006, 55: 37-42. DOI: 10.1099/jmm.0.46238-0 [6] Hatfull GF.The secret lives of mycobacteriophages[J]. Adv Virus Res, 2012,82:179-288. DOI: 10.1016/B978-0-12-394621-8.00015-7 [7] Lewis JA, Hatfull GF.Identifcation and characterization of mycobacteriophage L5 excisionase[J]. Mol Microbiol, 2000, 35(2): 350-360. DOI: 10.1046/j.1365-2958.2000.01695.x [8] Lee MH, Hatfull GF.Mycobacteriophage L5 integrase-mediated site-specifc integration in vitro[J]. J Bacteriol, 1993, 175(21): 6836-6841. DOI: 10.1128/jb.175.21.6836-6841.1993 [9] Donnelly-Wu MK, Jacobs WR Jr, Hatfull GF.Superinfection immunity of mycobacteriophage L5: Applications for genetic transformation of Mycobacteria[J]. Mol Microbiol, 1993, 7(3):407-417. DOI: 10.1111/j.1365-2958. 1993.tb01132.x [10] Broussard GW, Oldfield LM, Villanueva VM,et al.Integration-dependent bacteriophage immunity provides insights into the evolution of genetic switches[J]. Mol Cell, 2013, 49(2):237-248. DOI: 10.1016/j.molcel. 2012.11.012 [11] Pham TT, Jacobs-Sera D, Pedulla ML, et al.Comparative genomic analysis of mycobacteriophageTweety: evolutionary insights and construction of compatible site-specific integration vectors for mycobacteria[J]. Microbiol, 2007, 153(Pt 8):2711-2723. DOI: 10.1099/mic.0.2007/008904-0 [12] Jacobs-seca D, Marinelli LJ, Bowman C, et al. On the nature of mycobacteriophage diversity and host preference[J].Virology, 2012,434(2):187-201. DOI: 10.1016/j.virol.2012.09.026 [13] Ford ME, Sarkis GJ, Belanger AE, et al.Genome structure of mycobacteriophage D29:implications for phage evolution[J]. J Mol Biol, 1998, 279(1): 143-164. DOI: 10.1006/jmbi.1997.1610 [14] Sampson T, Broussard GW, Marinelli LJ, et al.Mycobacteriophages BPs, Angel and Halo:comparative genomics reveals a novel class of ultra-small mobile genetic elements[J].Microbiol, 2009, 155(Pt 9):2962-2977. DOI: 10.1099/mic.0.030486-0 [15] Pope WH, Jacobs-Sera D, Russell DA, et al.Expanding the diversity of mycobacteriophages: insights into genome architecture and evolution[J]. PLoS One, 2011, 6(1):e16329. DOI: 10.1371/journal.pone.0016329
2020, Vol. 36 
