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Two Lineages of Pseudomonas aeruginosa Filamentous Phages: Structural Uniformity over Integration Preferences.

ABSTRACT: Pseudomonas aeruginosa filamentous (Pf) bacteriophages are important factors contributing to the pathogenicity of this opportunistic bacterium, including biofilm formation and suppression of bacterial phagocytosis by macrophages. In addition, the capacity of Pf phages to form liquid crystal structures and their high negative charge density makes them potent sequesters of cationic antibacterial agents, such as aminoglycoside antibiotics or host antimicrobial peptides. Therefore, Pf phages have been proposed as a potential biomarker for risk of antibiotic resistance development. The majority of studies describing biological functions of Pf viruses have been performed with only three of them: Pf1, Pf4, and Pf5. However, our analysis revealed that Pf phages exist as two evolutionary lineages (I and II), characterized by substantially different structural/morphogenesis properties, despite sharing the same integration sites in the host chromosomes. All aforementioned model Pf phages are members of the lineage I. Hence, it is reasonable to speculate that their interactions with P. aeruginosa and impact on its pathogenicity may be not completely extrapolated to the lineage II members. Furthermore, in order to organize the present numerical nomenclature of Pf phages, we propose a more informative approach based on the insertion sites, that is, Pf-tRNA-Gly, -Met, -Sec, -tmRNA, and -DR (direct repeats), which are fully compatible with one of five types of tyrosine integrases/recombinases XerC/D carried by these viruses. Finally, we discuss possible evolutionary mechanisms behind this division and consequences from the perspective of virus-virus, virus-bacterium, and virus-human interactions.

SUBMITTER: Fiedoruk K

PROVIDER: S-EPMC7549136 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature

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Two Lineages of Pseudomonas aeruginosa Filamentous Phages: Structural Uniformity over Integration Preferences.

Fiedoruk Krzysztof K Zakrzewska Magdalena M Daniluk Tamara T Piktel Ewelina E Chmielewska Sylwia S Bucki Robert R

Genome biology and evolution 20201001 10

Pseudomonas aeruginosa filamentous (Pf) bacteriophages are important factors contributing to the pathogenicity of this opportunistic bacterium, including biofilm formation and suppression of bacterial phagocytosis by macrophages. In addition, the capacity of Pf phages to form liquid crystal structures and their high negative charge density makes them potent sequesters of cationic antibacterial agents, such as aminoglycoside antibiotics or host antimicrobial peptides. Therefore, Pf phages have be ...[more]

PMID: 32658245

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Project description:Modern sequencing technologies have provided insight into the genetic diversity of numerous species, including the human pathogen Pseudomonas aeruginosa. Bacterial genomes often harbor bacteriophage genomes (prophages), which can account for upwards of 20% of the genome. Prior studies have found P. aeruginosa prophages that contribute to their host's pathogenicity and fitness. These advantages come in many different forms, including the production of toxins, promotion of biofilm formation, and displacement of other P. aeruginosa strains. While several different genera and species of P. aeruginosa prophages have been studied, there has not been a comprehensive study of the overall diversity of P. aeruginosa-infecting prophages. Here, we present the results of just such an analysis. A total of 6,852 high-confidence prophages were identified from 5,383 P. aeruginosa genomes from strains isolated from the human body and other environments. In total, 3,201 unique prophage sequences were identified. While 53.1% of these prophage sequences displayed sequence similarity to publicly available phage genomes, novel and highly mosaic prophages were discovered. Among these prophages, there is extensive diversity, including diversity within the functionally conserved integrase and C repressor coding regions, two genes responsible for prophage entering and persisting through the lysogenic life cycle. Analysis of integrase, C repressor, and terminase coding regions revealed extensive reassortment among P. aeruginosa prophages. This catalog of P. aeruginosa prophages provides a resource for future studies into the evolution of the species. IMPORTANCE Prophages play a critical role in the evolution of their host species and can also contribute to the virulence and fitness of pathogenic species. Here, we conducted a comprehensive investigation of prophage sequences from 5,383 publicly available Pseudomonas aeruginosa genomes from human as well as environmental isolates. We identified a diverse population of prophages, including tailed phages, inoviruses, and microviruses; 46.9% of the prophage sequences found share no significant sequence similarity with characterized phages, representing a vast array of novel P. aeruginosa-infecting phages. Our investigation into these prophages found substantial evidence of reassortment. In producing this, the first catalog of P. aeruginosa prophages, we uncovered both novel prophages as well as genetic content that have yet to be explored.

Project description:Filamentous bacteriophages frequently infect Pseudomonas aeruginosa and alter its phenotypic traits, including virulence factors. The first step in examination of these phages is to obtain suspensions with high virus titer, but as there are no methods for integrative filamentous phage multiplication, the aim was to design, describe, and compare two methods for this purpose. As models, three strains of Pseudomonas aeruginosa, containing (pro)phages Pf4, Pf5, and PfLES were used (PAO1, UCBPP-PA14, and LESB58, respectively). Method 1 comprised propagation of phages in 6 L of bacterial culture for 48 h, and method 2 applied 600 mL culture and incubation for 6 days with centrifugation and addition of new medium and inoculum at 2-day intervals. In method 1, phages were propagated by culture agitation, followed by centrifugation and filtration (0.45 and 0.22 μm), and in method 2, cultures were agitated and centrifuged several times to remove bacteria without filtration. Regardless of the propagation method, supernatants were subjected to concentration by PEG8000 and CsCl equilibrium density gradient centrifugation, and phage bands were removed after ultracentrifugation and dialyzed. In the obtained suspensions, phage titer was determined, and concentration of isolated ssDNA from virions was measured. When propagation method 2 was compared with method 1, the phage bands in CsCl were much thicker, phage number was 3.5-7.4 logs greater, and concentration of ssDNA was 7.6-22.4 times higher. When phage count was monitored from days 2 to 6, virion numbers increased for 1.8-5.6 logs, depending on phage. We also observed that filamentous phage plaques faded after 8 h of incubation when the double layer agar spot method was applied, whereas the plaques were visible for 24 h on single-layer agar. Finally, for the first time, we confirmed existence of replicative form and virions of PfLES (pro)phage as well as its ability to produce plaques. Similarly, for the first time, we confirmed plaque production of Pf5 (pro)phage present in P. aeruginosa strain UCBPP-PA14. The described method 2 has many advantages and can be further improved and adopted for filamentous phages of other hosts.

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Two Lineages of Pseudomonas aeruginosa Filamentous Phages: Structural Uniformity over Integration Preferences.

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Two Lineages of Pseudomonas aeruginosa Filamentous Phages: Structural Uniformity over Integration Preferences.

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