(2005) have shown that phage mediate changes in the bacterial population, controlling the emergence of different variants. In contrast, we have repeatedly observed the conversion of the Pf4 into the superinfective form at the time when SCVs become detectable in the biofilm supernatant. Infect Immun 71: 29932999. The medium flow was resumed at a flow rate of 6mlh1. Bacteriophage have been suggested to be one of the most abundant biological agents on the planet, estimated at a total of 1 1031 (Rohwer and Edwards, 2002), compared with 26 1030 bacterial cells (Whitman et al., 1998), outnumbering prokaryotes by approximately 10:1. Bethesda, MD 20894, Web Policies This work was supported by RFBR grant 07-04-12100-ofi, 09-04-01546-a, 07-04-92168-NCNI_a, NATO SFPP 982833, and the "Fundamental science for medicine - 2008" program of the presidium of the Russian Academy of Sciences. Pseudomonas aeruginosa gene products Pilt and pilU are required for cytotoxicity in vitro and virulence in a mouse model of acute pneumonia. Proc Natl Acad Sci U S A 91(6):21912195, Woolford JL Jr, Cashman JS, Webster RE (1974) F1 coat protein synthesis and altered phospholipid metabolism in f1 infected Escherichia coli. Complete genome sequence of Pseudomonas aeruginosa PAO1, an opportunistic pathogen. In contrast, 5 of the 15 mice infected with the Pf4 mutant survived for the duration of the experiment (Figure 6). Present address: 6Current address: Budde Schou A/S, Vester Soegade 10, DK-1901 Copenhagen V, Denmark. (2005). CSLM images (XZ plane) of the treated and untreated biofilms are shown in (b). Nature 472(7344):499503, Feng JN, Russel M, Model P (1997) A permeabilized cell system that assembles filamentous bacteriophage. observed. PubMed Please enable it to take advantage of the complete set of features! Chembiochem 10(16):25992606, Zhang Y (2008) I-TASSER server for protein 3D structure prediction. Virology 500:275284, Smilowitz H (1974) Bacteriophage f1 infection: fate of the parental major coat protein. Epub 2015 Dec 9. The phage titre of P. aeruginosa PAO1 biofilm effluent that were determined on the PAO1 lawn or the Pf4 mutant lawn are shown as black and grey bars, respectively. Small colony variants were also quantified in the biofilm effluents for both strains. Moreover, the data suggest that microcolony expansion in the WT requires the presence of the phage for the microcolonies to expand beyond 50m in diameter. Both clinical and environmental isolates of P. aeruginosa show mutator phenotypes (Kenna et al., 2007). FEMS Microbiol Rev. Filamentous phage binds to the F pilus of a host E. coli cell through pIII. They also have a longer elimination half-life and thus better pharmacokinetic and pharmacodynamic . The .gov means its official. It has been assumed that the role of the C-terminal domain of pIII in the infection is that of a tether for the receptor-binding domains N1N2 to the main body of the virion. While most phages are pathogens that kill their bacterial hosts, the filamentous phages of the sub-class Inoviridae live in cooperative relationships with their bacterial hosts, akin to the principal Filamentous bacteriophage is a family of viruses ( Inoviridae) that infect bacteria. Filamentous Ff-like phages are capable of persistent infection while their DNA replicates independently as an episome in Escherichia coli [].In contrast, the life cycle of some Inoviridae phages does not mirror the behavior of the coliphages. One clone was identified and selected for further verification by colony PCR. Extracellular DNA required for bacterial biofilm formation. Unable to load your collection due to an error, Unable to load your delegates due to an error, (A) Bacterial and Archaeal virus subfamilies are represented and grouped based on their Baltimore classification. (B) Protein sequence similarity network plot of all predicted openreading frames from 56 filamentous phage genomes. For instance, the low concentration of the original antigen-specific lymphocytes in the population can be a problem while constructing immune libraries. mBio. (2007). These data therefore suggest that the Pf4 phage plays an essential role in the formation of hollow colonies and the process of cell death within the microcolonies. Filamentous phage are released from the bacterial membrane by a two-step mechanism involving a short C-terminal fragment of pIII. Curr Issues Mol Biol 7(2):135149, Das B (2014) Mechanistic insights into filamentous phage integration in Vibrio cholerae. This study shows that at the time when cell death occurs in biofilms of the WT, the Pf4 phage converts into a superinfective form, which correlates with the appearance of variants in the dispersal population. The flow cell was sterilized in 10% (v/v) bleach for 4h, thoroughly rinsed in sterile milliQ water and connected to sterile silicon tubing (Silastic laboratory tubings). Furthermore, the strong selective pressure of phage-mediated lysis can drive evolution of the target population, so that infection-resistant mutants may become dominant in the population. As expected, when cell-free effluent from the WT was serially diluted and spotted onto the WT lawn, no plaque formation was observed for first 3 days of biofilm development and plaques were observed from day 4 onwards (Figure 5). Phage-mediated variation has significant medical implications, where it has been shown that phage can mediate conversion of P. aeruginosa into a mucoid phenotype, which has been associated with poor outcomes for CF patients (Miller and Rubero, 1984; Hoiby et al., 2001). A similar approach was used for peptide libraries [88]. Biofilm differentiation and dispersal in mucoid Pseudomonas aeruginosa isolates from patients with cystic fibrosis. and transmitted securely. Comolli JC, Hauser AR, Waite L, Whitchurch CB, Mattick JS, Engel JN . J Exp Med 195: 14551462. Simultaneous positive and negative selection creates competitive conditions: a small number of antigen-positive (target) cells and an excess of antigen-negative adsorbing cells, which bind the unspecific antibodies from the phage library. Olszak T, Latka A, Roszniowski B, Valvano MA, Drulis-Kawa Z. Curr Med Chem. The M13 particle consists of a single-stranded DNA (ssDNA) . Nature 540:597601, Yan Z, Yin M, Xu D, Zhu Y, Li X (2017) Structural insights into the secretin translocation channel in the type II secretion system. Tax calculation will be finalised at checkout, Arnold K, Bordoli L, Kopp J, Schwede T (2006) The SWISS-MODEL workspace: a web-based environment for protein structure homology modelling. A hallmark of the differentiation process typical of P. aeruginosa biofilms is the generation of phage and the simultaneous appearance of variants, such as SCVs, in the dispersal population. Recently, virus-neutralizing scFv fragments specific to the flu virus H5N1 [26, 71] and virus-neutralizing Fab-fragments specific to the rabies virus [72] were extracted from immune libraries. (2005). This difference cannot be fully explained by cell transformation efficiency. This is supported by the observation that the Pf4 genes are some of the most strongly induced genes in the biofilms of P. aeruginosa (Whiteley et al., 2001). The unbound phage antibodies are removed by washing the culture dishes in the case of a monolayer and by centrifugation in the case of a suspension. Biopanning with immobilized antigens. Varying the elution and screening conditions during selection can help to select high-affinity antibodies. For example, virulence can be transmitted between strains of V. cholerae by the CTX phage that carries the genes coding for cholera toxin (CT) and the RTX toxin (Faruque et al., 2003; McLeod et al., 2005). FOIA Mol Microbiol 63(4):10081025, Goldbourt A, Gross BJ, Day LA, McDermott AE (2007) Filamentous phage studied by magic-angle spinning NMR: resonance assignment and secondary structure of the coat protein in Pf1. The pEX2.5 suicide vector was purified using the Wizard Plus Minipreps DNA purification systems (Promega Inc., Sydney, NSW, Australia) and electroporated into P. aeruginosa PAO1 as described (Choi and Schweizer, 2005). Biofilm formation by the small colony variant phenotype of Pseudomonas aeruginosa. Present address: 5Current address: Expression Engineering Group, Bioprocessing Technology Institute, 20 Biopolis Way, No. Briefly, bacteria were grown for 17h in MINS medium (Nicas and Iglewski, 1984) at 37C with shaking, diluted 1:100 in MINS medium 3h before infection, concentrated by centrifugation and then re-suspended in phosphate-buffered saline. Libraries have also been constructed against human autoantigens [59-61] and against antigens that cause allergic reactions [62]. Mol Microbiol 59: 11141128. (2003). Special thanks to H. Schweizer for providing the Gateway vectors and advice on their application. This procedure is called "biopanning" or affinity enrichment. Kenna DT, Doherty CJ, Foweraker J, Macaskill L, Barcus VA, Govan JRW . In this case, selection is performed under conditions that stimulate active endocytosis. Biophys J 99(5):14651474, Petrenko VA, Smith GP, Gong X, Quinn T (1996) A library of organic landscapes on filamentous phage. Conners R, Len-Quezada RI, McLaren M, Bennett NJ, Daum B, Rakonjac J, Gold VAM. J Mol Biol 228(3):720724, Stopar D, Spruijt RB, Wolfs CJ, Hemminga MA (2002) Structural characterization of bacteriophage M13 solubilization by amphiphiles. Some phages can only reproduce via a lytic lifecycle, in which they burst and kill their host cells. The process of triggering the signals for production and release of new phages is fascinating and include a variety of factors such as a drop in pH or in temperature (Singleton et al., 1982; Wolf et al., 1987; Gardel and Mekalanos, 1994; Wang et al., 2007 ). (Webb et al., 2003). J Mol Biol 293(5):10171027, Heilpern AJ, Waldor MK (2000) CTX infection of Vibrio cholerae requires the tolQRA gene products. Federal government websites often end in .gov or .mil. Nat Commun 7:13858, Cao BR, Yang MY, Mao CB (2016) Phage as a genetically modifiable supramacromolecule in chemistry, materials and medicine. Science 284: 13181322. These large microcolonies were not entirely empty, where a number of viable cells could be observed in the centres along with dead cells (Figure 3e), whereas the mutant microcolonies were not significantly larger, 52.25.5m, than those observed on day 5 (Figure 3f) and no centralized cell death or hollowing was observed. (2007). J Bacteriol 135(3):883887, CAS Proc Natl Acad Sci U S A 86(11):40024006, Model P, Jovanovic G, Dworkin J (1997) The Escherichia coli phage shock protein operon. The culture was concentrated to 100l and plated onto LB agar supplemented with 30gml1 of gentamicin (LB+Gm30) and incubated at 37C for 24h. Transformant colonies were patched onto LB+Gm30 and LB+Cb200 (LB agar supplemented with 200gml1 of carbenicillin) plates to differentiate single-crossover mutants from double-crossover mutants. Virol J. The infection starts by binding of the N-terminal N2 and N1 domains to the primary and secondary host receptors, F pilus and TolA protein, respectively, whereas the life cycle terminates by the C-terminal domain-mediated release of the membrane-anchored virion from the cell. (1990). J Bacteriol 62: 293300. . Therefore, we opted to exogenously add back the phage to confirm its effects on the biofilm. Unexpectedly, mice infected with the Pf4 mutant survived significantly longer than those infected with its isogenic WT strain, showing that Pf4 contributes to the virulence of P. aeruginosa.
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life cycle of filamentous phage
