title: A novel termini analysis theory using HTS data alone for the identification of Enterococcus phage EF4-like genome termini.
abstract: BACKGROUND
  Enterococcus faecalis and Enterococcus faecium are typical enterococcal bacterial pathogens. Antibiotic resistance means that the identification of novel E. faecalis and E. faecium phages against antibiotic-resistant Enterococcus have an important impact on public health. In this study, the E. faecalis phage IME-EF4, E. faecium phage IME-EFm1, and both their hosts were antibiotic resistant. To characterize the genome termini of these two phages, a termini analysis theory was developed to provide a wealth of terminal sequence information directly, using only high-throughput sequencing (HTS) read frequency statistics.
  
  
  RESULTS
  The complete genome sequences of phages IME-EF4 and IME-EFm1 were determined, and our termini analysis theory was used to determine the genome termini of these two phages. Results showed 9 bp 3' protruding cohesive ends in both IME-EF4 and IME-EFm1 genomes by analyzing frequencies of HTS reads. For the positive strands of their genomes, the 9 nt 3' protruding cohesive ends are 5'-TCATCACCG-3' (IME-EF4) and 5'-GGGTCAGCG-3' (IME-EFm1). Further experiments confirmed these results. These experiments included mega-primer polymerase chain reaction sequencing, terminal run-off sequencing, and adaptor ligation followed by run-off sequencing.
  
  
  CONCLUSIONS
  Using this termini analysis theory, the termini of two newly isolated antibiotic-resistant Enterococcus phages, IME-EF4 and IME-EFm1, were identified as the byproduct of HTS. Molecular biology experiments confirmed the identification. Because it does not require time-consuming wet lab termini analysis experiments, the termini analysis theory is a fast and easy means of identifying phage DNA genome termini using HTS read frequency statistics alone. It may aid understanding of phage DNA packaging.
authors.affiliation: State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, P.R. China. zhangxianglilan@gmail.com.
authors.forename: Xianglilan
authors.initials: X
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, P.R. China. 18252093512@163.com.|School of Life Science & Technology, China Pharmaceutical University, 24 Tong Jia Xiang, Nanjing, 210009, P.R. China. 18252093512@163.com.
authors.forename: Yahui
authors.initials: Y
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, P.R. China. 824518891@qq.com.
authors.forename: Shasha
authors.initials: S
authors.lastname: Li
authors.affiliation: State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, P.R. China. anxiaoping2008@gmail.com.
authors.forename: Xiaoping
authors.initials: X
authors.lastname: An
authors.affiliation: State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, P.R. China. pgqd2008@163.com.
authors.forename: Guangqian
authors.initials: G
authors.lastname: Pei
authors.affiliation: State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, P.R. China. yong.huang1987@gmail.com.
authors.forename: Yong
authors.initials: Y
authors.lastname: Huang
authors.affiliation: State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, P.R. China. fanhang11@gmail.com.
authors.forename: Hang
authors.initials: H
authors.lastname: Fan
authors.affiliation: State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, P.R. China. zhiqiangmi.ime@gmail.com.
authors.forename: Zhiqiang
authors.initials: Z
authors.lastname: Mi
authors.affiliation: State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, P.R. China. zhang.zhiyi.cu@gmail.com.
authors.forename: Zhiyi
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, P.R. China. ww995936007ww@yeah.net.
authors.forename: Wei
authors.initials: W
authors.lastname: Wang
authors.affiliation: Beijing Computing Center, Beijing, 100094, P.R. China. allexchen@126.com.
authors.forename: Yubao
authors.initials: Y
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, P.R. China. tong.yigang@gmail.com.
authors.forename: Yigang
authors.initials: Y
authors.lastname: Tong
mesh_terms: D001435:Bacteriophages / Q000235:genetics*; D024881:Drug Resistance, Bacterial; D016983:Enterococcus / Q000302:isolation & purification / Q000821:virology*; D013293:Enterococcus faecalis / Q000302:isolation & purification / Q000821:virology; D016984:Enterococcus faecium / Q000302:isolation & purification / Q000821:virology; D016679:Genome, Viral; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D008957:Models, Genetic; D015233:Models, Statistical; D017422:Sequence Analysis, DNA / Q000379:methods*; D020079:Terminal Repeat Sequences