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J Virol. 2018 Jan 3. pii: JVI.01064-17. doi: 10.1128/JVI.01064-17. [Epub ahead of print]

Genomic and Biochemical Characterization of Acinetobacter Podophage Petty Reveals a Novel Lysis Mechanism and Tail-Associated Depolymerase Activity.

Author information

1
Department of Biochemistry and Biophysics, Texas A&M University, College Station, Texas, USA.
2
Center for Phage Technology, Texas A&M University, College Station, Texas, USA.
3
Department of Chemistry, Texas A&M University, College Station, Texas, USA.
4
Center for Phage Technology, Texas A&M University, College Station, Texas, USA jason.gill@tamu.edu.
5
Department of Animal Science, Texas A&M University, College Station, Texas, USA.

Abstract

The increased prevalence of drug-resistant, nosocomial Acinetobacter infections, particularly from pathogenic members of the Acinetobacter calcoaceticus-baumannii complex, necessitates the exploration of novel treatments such as phage therapy. In the present study, we characterize phage Petty, a novel podophage that infects multidrug-resistant Acinetobacter nosocomialis and Acinetobacter baumannii Genome analysis reveals that phage Petty is a 40,431bp ϕKMV-like phage, with a coding density of 92.2% and a G+C content of 42.3%. Interestingly, the lysis cassette encodes a class I holin and a single subunit endolysin, but lacks canonical spanins to disrupt the outer membrane. Analysis of other ϕKMV-like genomes revealed that spanin-less lysis cassettes are a feature of phages infecting Acinetobacter within this subfamily of bacteriophages. The observed halo surrounding Petty's large clear plaques indicated the presence of a phage-encoded depolymerase capable of degrading capsular exopolysaccharides (EPS). Gene 39, a putative tail fiber, was hypothesized to possess depolymerase activity based on weak homology to previously reported phage tail fibers. The 101.4 kDa protein gp39 was cloned and expressed, and its activity against Acinetobacter EPS in solution was determined. The enzyme degraded purified EPS from its host strain A. nosocomialis AU0783, reducing its viscosity, and generated reducing ends in solution, indicative of hydrolase activity. Given that the accessibility to cells within a biofilm is enhanced by degradation of EPS, phages with depolymerases may have enhanced diagnostic and therapeutic potential against drug-resistant Acinetobacter strains.Importance Bacteriophage therapy is being revisited as a treatment for difficult-to-treat infections. This is especially true for Acinetobacter infections, which are notorious for being resistant to antimicrobials. Thus, sufficient data needs to be generated with regard to phages with therapeutic potential, if they are to be successfully employed clinically. In this study, we describe the isolation and characterization of phage Petty, a novel lytic podophage, and its depolymerase. To our knowledge, it is the first phage reported able to infect both A. baumannii and A. nosocomialis The lytic phage has potential as an alternative therapeutic agent, and the depolymerase could be used for modulating EPS both during infections and in biofilms on medical equipment, as well as for capsular typing. We also highlight the lack of predicted canonical spanins in the phage genome, and confirm that, unlike the rounding of λ lysogens lacking functional spanin genes, A. nosocomialis cells infected with phage Petty lyse by bursting. This suggests phages like Petty employ a different mechanism to disrupt the outer membrane of Acinetobacter hosts during lysis.

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