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Prophages mediate defense against phage infection through diverse mechanisms - PubMed

Prophages mediate defense against phage infection through diverse mechanisms

Joseph Bondy-Denomy et al. ISME J. 2016 Dec.

Abstract

The activity of bacteriophages poses a major threat to bacterial survival. Upon infection, a temperate phage can either kill the host cell or be maintained as a prophage. In this state, the bacteria carrying the prophage is at risk of superinfection, where another phage injects its genetic material and competes for host cell resources. To avoid this, many phages have evolved mechanisms that alter the bacteria and make it resistant to phage superinfection. The mechanisms underlying these phentoypic conversions and the fitness consequences for the host are poorly understood, and systematic studies of superinfection exclusion mechanisms are lacking. In this study, we examined a wide range of Pseudomonas aeruginosa phages and found that they mediate superinfection exclusion through a variety of mechanisms, some of which affected the type IV pilus and O-antigen, and others that functioned inside the cell. The strongest resistance mechanism was a surface modification that we showed is cost-free for the bacterial host in a natural soil environment and in a Caenorhabditis. elegans infection model. This study represents the first systematic approach to address how a population of prophages influences phage resistance and bacterial behavior in P. aeruginosa.

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Figures

Figure 1
Figure 1

Phage sensitivity profiles of strains in the PA14 prophage collection. Wild-type PA14, mutant strains lacking the O-antigen (wbpL) or pilus (pilA), and PA14 lysogenized with 30 different phages (x axis) were infected with a group of 30 phages (y axis). The outcome of the infection is shown in the color scale in the legend. Double (2x, MP29 and JBD44a) and triple (3x, MP29, JBD44a and JBD24) lysogens were made to assess the additivity of resistance. Phage groupings along the y axis represent homoimmune repressor families, and repressor-mediated resistance is shown in navy blue. Sequenced phage genomes are highlighted in red.

Figure 2
Figure 2

Phage plaque formation assays. Eight different phage lysates were applied in 10-fold serial dilutions to lawns of wild-type PA14, or the indicated PA14 lysogens and plaque formation was assessed. The resistance group that a given lysogen belongs to (see Figure 1) is shown. See Supplementary Figure S5 for more plaque assays.

Figure 3
Figure 3

Alignment of the phage genomes highlights the presence of a highly variable accessory genome. Each box represents a single open reading frame, with gray boxes denoting conserved genes, and colored boxes denoting non-conserved 'accessory genes'. Different vertical positions and colors for the accessory genes indicate that they encode proteins from distinct sequence families. Previously characterized gene Tip is accessory gene 1. See Supplementary Table 3 for more details on conserved gene functions.

Figure 4
Figure 4

(a) PA14 or a PA14(JBD26) lysogen was infected with phage JBD26, JBD88a or JBD93 or (b) PA14 or PA14 lysogens of JBD23 or JBD30 were infected with phage JBD88a (at time 0) and the efflux of K+ was measured over time. A PA14▵pilA mutant was also infected with JBD88a as a negative control. Efflux is represented as a percentage of the total K+ efflux detected when wild-type PA14 was infected with a given phage (see Materials and methods).

Figure 5
Figure 5

Prophages that impart strong phage resistance frequently alter T4P function. (a) The diameter of twitching motility zones for indicated PA14 lysogens is shown as a percentage, relative to wild-type PA14. Average value of three replicate experiments is shown and the error bars denote the standard deviations. (b) Representative images of twitching motility zones for PA14 lysogens visualized with crystal violet.

Figure 6
Figure 6

The JBD26 lysogen modifies pilus function with no discernable evolutionary cost. (a) PA14, PA14(JBD26) or a pilA mutant were competed with a lacZ-tagged PA14 strain in minimal media in autoclaved compost. The relative abundance of each strain was enumerated after 1 week of incubation. Error bars show the 95% confidence interval. (b) 80–100 C. elegans nematodes were plated on lawns of E. coli OP50, wild-type PA14 or PA14(JBD26) and live and dead worms were scored over the course of 130 h.

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