Identification of virulence genes in a pathogenic strain of Pseudomonas aeruginosa by representational difference analysis

Division of Infectious Diseases, Massachusetts General Hospital, Boston, Massachusetts 02114, USA.
Journal of Bacteriology (Impact Factor: 2.69). 03/2002; 184(4):952-61. DOI: 10.1128/jb.184.4.952-961.2002
Source: PubMed

ABSTRACT Pseudomonas aeruginosa is an opportunistic pathogen that may cause severe infections in humans and other vertebrates. In addition, a human clinical isolate of P. aeruginosa, strain PA14, also causes disease in a variety of nonvertebrate hosts, including plants, Caenorhabditis elegans, and the greater wax moth, Galleria mellonella. This has led to the development of a multihost pathogenesis system in which plants, nematodes, and insects have been used as adjuncts to animal models for the identification of P. aeruginosa virulence factors. Another approach to identifying virulence genes in bacteria is to take advantage of the natural differences in pathogenicity between isolates of the same species and to use a subtractive hybridization technique to recover relevant genomic differences. The sequenced strain of P. aeruginosa, strain PAO1, has substantial differences in virulence from strain PA14 in several of the multihost models of pathogenicity, and we have utilized the technique of representational difference analysis (RDA) to directly identify genomic differences between P. aeruginosa strains PA14 and PAO1. We have found that the pilC, pilA, and uvrD genes in strain PA14 differ substantially from their counterparts in strain PAO1. In addition, we have recovered a gene homologous to the ybtQ gene from Yersinia, which is specifically present in strain PA14 but absent in strain PAO1. Mutation of the ybtQ homolog in P. aeruginosa strain PA14 significantly attenuates the virulence of this strain in both G. mellonella and a burned mouse model of sepsis to levels comparable to those seen with PAO1. This suggests that the increased virulence of P. aeruginosa strain PA14 compared to PAO1 may relate to specific genomic differences identifiable by RDA.

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Available from: Laurence G Rahme, Aug 20, 2015
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    • "Here, we observed that the initial production phase of HCN/CN − by strain PA14 was more rapid than by strain MPAO1 which is widely used as a reference strain for studying P. aeruginosa PAO1 virulence (Fig. 3B). Clinical isolate PA14 is significantly more virulent than PAO1 in a wide range of hosts, including mice, nematodes, insects as Galleria mellonella, and plants as Arabidopsis thaliana (Choi et al., 2002; Rahme et al., 1995; Tan et al., 1999). Concerning , the well characterized host infection model, C. elegans, it has been shown that strain PA14 is able to kill these worms more efficiently than PAO1 (Tan et al., 1999 "
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    • "This is a molecular method by which genes whose presence differs between two strains can be isolated (Agron et al., 2002; Sambrook & Russell, 2001; Winstanley, 2002). As this method of gene isolation is not biased by prior assumptions about function, it provides a global approach to identification of virulence genes and has been utilized successfully in other pathogenic bacteria for this purpose (Adderson et al., 2003; Choi et al., 2002; Li et al., 2003; Winstanley, 2002). In the present study, we used suppressive subtractive hybridization between genomic DNA from an environmental isolate of B. cepacia (strain ATCC 25416 T ; Bcep25416) and the epidemic B. cenocepacia clinical isolate J2315 (BcenJ- 2315) to identify nucleotide sequences present only in the clinical isolate. "
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