# A Novel Singleton Giant Phage Yong-XC31 Lytic to the _Pyropia_ Pathogen _Vibrio mediterranean_
## 3 Results
### Genome Analysis of Yong-XC31  
Yong-XC31 is a typical giant phage at genome level. Complete genome sequencing was conducted using next-generation sequencing (NGS) with an average read length of 284.6 bases. 81.66% of the sequencing reads were matched to the complete genome (133,281 out of 163,212 reads) with an average sequencing depth of 130.6-fold. The complete genome of Yong-XC31 is 290,532 bp long with a G + C content of 45.87%. Yong-XC31 genome is unique sharing the highest BLASTn homology with the most related phage BONAISHI (sequence ID: MH595538, query cover as low as 1%, identity 70.62%). 25.2% predicted Yong-XC31 ORFs might encode novel proteins as they have no similarity to protein sequences in current public databases (26 January 2021).  
No tRNA gene was found in the genome. Among the 318 ORFs of Yong-XC31, 17.3% were predicted functions, 82.7% predicted as hypothetical proteins. No ORFs associated with virulence factors, toxins or antibiotic resistance genes were found among the annotated ones within Yong-XC31 genome. The predicted ORFs could be classified into five functional categories, including DNA replication/regulation, bacteriophage packaging, bacteriophage structure, lysis and hypothetical protein (Figure 6, Supplement Table S1).  
6 ORFs were annotated as structural and assembly genes (Supplement Table S1). 15 ORFs were annotated to be involved in DNA replication, recombination, and repair. 2 ORFs encode homing endonucleases of HNH family (ORF68 and ORF141), one of which is an intron-encoded homing endonuclease (ORF141) located between genes encoding subunits of RNAP.  
As mentioned in the introduction, giant bacteriophages commonly contain many genes that do not exist in the small genome bacteriophages. Notably, Yong-XC31 harbours multiple enzyme genes related to bacteriolysis. At least 5 ORFs predicted to play a role in host cell lysis, including a peptidase (family M48, ORF29), a permuted papain-like amidase  
Figure 6: Genome map of _Vibrio_ phage Yong-XC31. The arrow indicates the coding protein sequence, the protein function is distinguished by color, red stands for lysis, yellow for packaging, blue for DNA replication/regulation and green for structure, gray indicates hypothetical protein.  
enzyme (ORF175), a glycosyl hydrolases (family 18, ORF295), a peptidoglycan hydrolase LytM (ORF143) and a lysin (ORF302). Remarkably, the lysin has a modular structure comprising an N-terminal glycoside hydrolase 19 domain, and a C-terminal peptidoglycan binding domain (PGBD).  
As introduction described, some giant phages contain additional genes which may replace the function of the host proteins, thereby reducing the dependence of giant phages on their bacterial host. Noticeably, Yong-XC31 harbours eight paralogous genes (ORF34, ORF46, ORF47, ORF69, ORF136, ORF140, ORF142, ORF153) for RNAP. In particular, Yong-XC31 harbours genes (ORF265 and ORF271) encoding thymidylulate synthase (dTMP synthase) and thymidylulate kinase (TdR kinase) responsible for the de novo biosynthesis of thymidylate (dTMP) and as the salvage enzyme which leads to the production of dTMP even in presence of dTMP synthase inhibition [42]. Yong-XC31 also harbours a gene (ORF278) encoding tRNA nucleotides/poly(A) polymerase participating translation, ribosomal structure and biogenesis [43].