Unlocking the mystery of the hard-to-sequence phage genome: PaP1 methylome and bacterial immunity

Lu, Shuguang; Le, Shuai; Tan, Yinling; Li, Ming; Liu, Chang; Zhang, Kebin; Huang, Jianjun; Chen, Haimei; Rao, Xiancai; Zhu, Jing; Zou, Lingyun; Ni, Qingshan; Li, Shu; Wang, Jing; Jin, Xiaolin; Hu, Qiwen; Yao, Xinyue; Zhao, Xia; Zhang, Lin; Huang, Guangtao; Hu, Fuquan

doi:10.1186/1471-2164-15-803

Cited by 10 publications

(15 citation statements)

References 64 publications

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“…BlastN analysis revealed that the genome sequences of phages PaP1 4 18 19 20 , K8, C11, JG004 21 , vB_PaeM_C2-10_Ab1 14 22 , vB_PaeM_C2-10_Ab02 22 , vB_PaeM_C2-10_Ab08 22 , vB_PaeM_C2-10_Ab10 22 , vB_PaeM_C2-10_Ab15 22 , PAK_P1 10 15 16 , PAK_P2 10 16 , and PAK_P4 10 16 share an identity of above 90% and query coverage also above 90% with the PaoP5 genome ( Table S2 and Fig. S3 ).…”

Section: Resultsmentioning

confidence: 99%

Characterization and Comparative Genomic Analyses of Pseudomonas aeruginosa Phage PaoP5: New Members Assigned to PAK_P1-like Viruses

Shen

Jin

et al. 2016

Sci Rep

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As a potential alternative to antibiotics, phages can be used to treat multi-drug resistant bacteria. As such, the biological characteristics of phages should be investigated to utilize them as effective antimicrobial agents. In this study, phage PaoP5, a lytic virus that infects Pseudomonas aeruginosa PAO1, was isolated and genomically characterized. PaoP5 comprises an icosahedral head with an apex diameter of 69 nm and a contractile tail with a length of 120 nm. The PaoP5 genome is a linear dsDNA molecule containing 93,464 base pairs (bp) with 49.51% G + C content of 11 tRNA genes and a 1,200 bp terminal redundancy. A total of 176 protein-coding genes were predicted in the PaoP5 genome. Nine PaoP5 structural proteins were identified. Three hypothetical proteins were determined as structural. Comparative genomic analyses revealed that seven new Pseudomonas phages, namely, PaoP5, K8, C11, vB_PaeM_C2-10_Ab02, vB_PaeM_C2-10_Ab08, vB_PaeM_C2-10_Ab10, and vB_PaeM_C2-10_Ab15, were similar to PAK_P1-like viruses. Phylogenetic and pan-genome analyses suggested that the new phages should be assigned to PAK_P1-like viruses, which possess approximately 100 core genes and 150 accessory genes. This work presents a detailed and comparative analysis of PaoP5 to enhance our understanding of phage biology.

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Section: Resultsmentioning

confidence: 99%

Characterization and Comparative Genomic Analyses of Pseudomonas aeruginosa Phage PaoP5: New Members Assigned to PAK_P1-like Viruses

Shen

Jin

et al. 2016

Sci Rep

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“…This, coupled with resistance of phage DNA to restriction enzymes digestion, points to high level of DNA modifications. Phage DNA modification is not an unknown phenomenon, with the most commonly described being the adenine and cytosine methylation ( 47 , 48 ). However, it should be noted that ISTD and NOVI DNAs were not digested with the Dpn I restriction enzyme that cleaves only when its recognition site, GATC, is methylated.…”

Section: Discussionmentioning

confidence: 99%

Characterization, Antibiofilm, and Depolymerizing Activity of Two Phages Active on Carbapenem-Resistant Acinetobacter baumannii

et al. 2020

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Acinetobacter baumannii is a leading cause of healthcare-associated infections worldwide. Its various intrinsic and acquired mechanisms of antibiotic resistance make the therapeutic challenge even more serious. One of the promising alternative treatments that is increasingly highlighted is phage therapy, the therapeutic use of bacteriophages to treat bacterial infections. Two phages active against nosocomial carbapenem-resistant A. baumannii strain 6077/12, vB_AbaM_ISTD, and vB_AbaM_NOVI, were isolated from Belgrade wastewaters, purified, and concentrated using CsCl gradient ultracentrifugation. The phages were screened against 103 clinical isolates of A. baumannii from a laboratory collection and characterized based on plaque and virion morphology, host range, adsorption rate, and one-step growth curve. Given that phage ISTD showed a broader host range, better adsorption rate, shorter latent period, and larger burst size, its ability to lyse planktonic and biofilm-embedded cells was tested in detail. Phage ISTD yielded a 3.5-and 2-log reduction in planktonic and biofilm-associated viable bacterial cell count, respectively, but the effect was time-dependent. Both phages produced growing turbid halos around plaques indicating the synthesis of depolymerases, enzymes capable of degrading bacterial exopolysaccharides. Halos tested positive for presence of phages in the proximity of the plaque, but not further from the plaque, which indicates that the observed halo enlargement is a consequence of enzyme diffusion through the agar, independently of the phages. This notion was also supported by the growing halos induced by phage preparations applied on pregrown bacterial lawns, indicating that depolymerizing effect was achieved also on non-dividing sensitive cells. Overall, good rates of growth, fast adsorption rate, broad host range, and high depolymerizing activity, as well as antibacterial effectiveness against planktonic and biofilm-associated bacteria, make these phages good candidates for potential application in combating A. baumannii infections.

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“…Bio-spin columns were obtained from Bio-Rad (Hercules, CA, USA). Phage PaP1 was propagated and extracted, and its genomic DNA was extracted and purified as described previously [ 21 , 22 ]. Other commercially available reagents were of the highest quality.…”

Section: Methodsmentioning

confidence: 99%

Error-Free Bypass of 7,8-dihydro-8-oxo-2′-deoxyguanosine by DNA Polymerase of Pseudomonas aeruginosa Phage PaP1

Xue

Liu

et al. 2017

Genes

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As one of the most common forms of oxidative DNA damage, 7,8-dihydro-8-oxo-2′-deoxyguanosine (8-oxoG) generally leads to G:C to T:A mutagenesis. To study DNA replication encountering 8-oxoG by the sole DNA polymerase (Gp90) of Pseudomonas aeruginosa phage PaP1, we performed steady-state and pre-steady-state kinetic analyses of nucleotide incorporation opposite 8-oxoG by Gp90 D234A that lacks exonuclease activities on ssDNA and dsDNA substrates. Gp90 D234A could bypass 8-oxoG in an error-free manner, preferentially incorporate dCTP opposite 8-oxoG, and yield similar misincorporation frequency to unmodified G. Gp90 D234A could extend beyond C:8-oxoG or A:8-oxoG base pairs with the same efficiency. dCTP incorporation opposite G and dCTP or dATP incorporation opposite 8-oxoG showed fast burst phases. The burst of incorporation efficiency (kpol/Kd,dNTP) is decreased as dCTP:G > dCTP:8-oxoG > dATP:8-oxoG. The presence of 8-oxoG in DNA does not affect its binding to Gp90 D234A in a binary complex but it does affect it in a ternary complex with dNTP and Mg2+, and dATP misincorporation opposite 8-oxoG further weakens the binding of Gp90 D234A to DNA. This study reveals Gp90 D234A can bypass 8-oxoG in an error-free manner, providing further understanding in DNA replication encountering oxidation lesion for P.aeruginosa phage PaP1.

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Unlocking the mystery of the hard-to-sequence phage genome: PaP1 methylome and bacterial immunity

Cited by 10 publications

References 64 publications

Characterization and Comparative Genomic Analyses of Pseudomonas aeruginosa Phage PaoP5: New Members Assigned to PAK_P1-like Viruses

Characterization and Comparative Genomic Analyses of Pseudomonas aeruginosa Phage PaoP5: New Members Assigned to PAK_P1-like Viruses

Characterization, Antibiofilm, and Depolymerizing Activity of Two Phages Active on Carbapenem-Resistant Acinetobacter baumannii

Error-Free Bypass of 7,8-dihydro-8-oxo-2′-deoxyguanosine by DNA Polymerase of Pseudomonas aeruginosa Phage PaP1

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