Dataset Information

Genomic, Morphological and Functional Characterization of Virulent Bacteriophage IME-JL8 Targeting Citrobacter freundii.

ABSTRACT: Citrobacter freundii refers to a fish pathogen extensively reported to be able to cause injury and high mortality. Phage therapy is considered a process to alternatively control bacterial infections and contaminations. In the present study, the isolation of a virulent bacteriophage IME-JL8 isolated from sewage was presented, and such bacteriophage was characterized to be able to infect Citrobacter freundii specifically. Phage IME-JL8 has been classified as the member of the Siphoviridae family, which exhibits the latent period of 30-40 min. The pH and thermal stability of phage IME-JL8 demonstrated that this bacteriophage achieved a pH range of 4-10 as well as a temperature range of 4, 25, and 37°C. As revealed from the results of whole genomic sequence analysis, IME-JL8 covers a double-stranded genome of 49,838 bp (exhibiting 47.96% G+C content), with 80 putative coding sequences contained. No bacterial virulence- or lysogenesis-related ORF was identified in the IME-JL8 genome, so it could be applicable to phage therapy. As indicated by the in vitro experiments, phage IME-JL8 is capable of effectively removing bacteria (the colony count decreased by 6.8 log units at 20 min), and biofilm can be formed in 24 h. According to the in vivo experiments, administrating IME-JL8 (1 × 10⁷ PFU) was demonstrated to effectively protect the fish exhibiting a double median lethal dose (2 × 10⁹ CFU/carp). Moreover, the phage treatment led to the decline of pro-inflammatory cytokines in carp with lethal infections. IME-JL8 was reported to induce efficient lysis of Citrobacter freundii both in vitro and in vivo, thereby demonstrating its potential as an alternative treatment strategy for infections attributed to Citrobacter freundii.

SUBMITTER: Jia K

PROVIDER: S-EPMC7717962 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Genomic, Morphological and Functional Characterization of Virulent Bacteriophage IME-JL8 Targeting Citrobacter freundii.

Jia Kaixiang K Yang Nuo N Zhang Xiuwen X Cai Ruopeng R Zhang Yang Y Tian Jiaxin J Raza Sayed Haidar Abbas SHA Kang Yuanhuan Y Qian Aidong A Li Ying Y Sun Wuwen W Shen Jinyu J Yao Jiayun J Shan Xiaofeng X Zhang Lei L Wang Guiqin G

Frontiers in microbiology 20201119

Citrobacter freundii refers to a fish pathogen extensively reported to be able to cause injury and high mortality. Phage therapy is considered a process to alternatively control bacterial infections and contaminations. In the present study, the isolation of a virulent bacteriophage IME-JL8 isolated from sewage was presented, and such bacteriophage was characterized to be able to infect Citrobacter freundii specifically. Phage IME-JL8 has been classified as the member of the Siph ...[more]

PMID: 33329451

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Project description:BACKGROUND: With the use of broad-spectrum antibiotics, immunosuppressive drugs, and glucocorticoids, multidrug-resistant Acinetobacter baumannii (MDR-AB) has become a major nosocomial pathogen species. The recent renaissance of bacteriophage therapy may provide new treatment strategies for combatting drug-resistant bacterial infections. In this study, we isolated a lytic bacteriophage vB_AbaM-IME-AB2 has a short latent period and a small burst size, which clear its host's suspension quickly, was selected for characterization and a complete genomic comparative study. RESULTS: The isolated bacteriophage vB_AbaM-IME-AB2 has an icosahedral head and displays morphology resembling Myoviridae family. Gel separation assays showed that the phage particle contains at least nine protein bands with molecular weights ranging 15-100 kDa. vB_AbaM-IME-AB2 could adsorb its host cells in 9 min with an adsorption rate more than 99% and showed a short latent period (20 min) and a small burst size (62 pfu/cell). It could form clear plaques in the double-layer assay and clear its host's suspension in just 4 hours. Whole genome of vB_AbaM-IME-AB2 was sequenced and annotated and the results showed that its genome is a double-stranded DNA molecule consisting of 43,665 nucleotides. The genome has a G?+?C content of 37.5% and 82 putative coding sequences (CDSs). We compared the characteristics and complete genome sequence of all known Acinetobacter baumannii bacteriophages. There are only three that have been sequenced Acinetobacter baumannii phages AB1, AP22, and phiAC-1, which have a relatively high similarity and own a coverage of 65%, 50%, 8% respectively when compared with our phage vB_AbaM-IME-AB2. A nucleotide alignment of the four Acinetobacter baumannii phages showed that some CDSs are similar, with no significant rearrangements observed. Yet some sections of these strains of phage are nonhomologous. CONCLUSION: vB_AbaM-IME-AB2 was a novel and unique A. baumannii bacteriophage. These findings suggest a common ancestry and microbial diversity and evolution. A clear understanding of its characteristics and genes is conducive to the treatment of multidrug-resistant A. baumannii in the future.

Project description:BackgroundCitrobacter freundii is the most common class of pathogens in the genus Citrobacter and is an important pathogen associated with certain underlying diseases or immune dysfunction. The aim of this study was to elucidate the resistance mechanism of clinically derived carbapenem-resistant C. freundii isolate and to characterize the genetic environment and delivery pattern of the IncN1 plasmid carrying the bla IMP-4 gene from C. freundii isolate.Materials and methodsWe identified a clinical isolate of C. freundii L91 carrying bla IMP-4 and performed phylogenetic analysis by whole-genome sequencing. The complete genomic sequence of L91 was obtained using the Illumina HiSeq 4000-PE150 and PacBio RS II platforms. Antimicrobial susceptibility testing was determined by the VITEK 2 system. Plasmid characteristics were presented by S1-pulsed-field gel electrophoresis (PFGE), Southern blotting and conjugation experiments.ResultsS1-PFGE, Southern blot and conjugation assay confirmed the presence of bla IMP-4 genes on a conjugative plasmid in this isolate. C. freundii L91 and transconjugant L91-E. coli 600 strains both showed resistance to carbapenems. In silico analysis further showed that pIMP-4-L91 is an IncN1 plasmid with a length of 51,042 bp. Furthermore, bla IMP-4 gene was found encoded in the bla IMP-4-qacG2-aacA4-catB3 cassette array within a class 1 integron. A conserved structure sequence (ΔISKpn27-bla IMP-4-ΔISSen2-hp-hp-IS6100) was found in the upstream and downstream of the bla IMP-4.ConclusionWe performed a comprehensive phylogenetic analysis of carbapenemase-resistant C. freundii and elucidated the resistance mechanism of clinically derived C. freundii L91. Not only that, we also found that the bla IMP-4 gene is located on the IncN1 plasmid and has a horizontal transfer function and a certain ability to spread. To lower the risk of the dissemination of such C. freundii isolates in clinical settings, more surveillance is needed in the future.

Dataset Information

Genomic, Morphological and Functional Characterization of Virulent Bacteriophage IME-JL8 Targeting Citrobacter freundii.

Publications

Genomic, Morphological and Functional Characterization of Virulent Bacteriophage IME-JL8 Targeting <i>Citrobacter freundii</i>.

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