Conjugative Plasmid Transfer and Adhesion Dynamics in an Escherichia coli Biofilm

School of Chemistry and Molecular Biosciences, University of Queensland, Brisbane, QLD 4072, Australia.
Applied and Environmental Microbiology (Impact Factor: 3.67). 09/2009; 75(21):6783-91. DOI: 10.1128/AEM.00974-09
Source: PubMed


A conjugative plasmid from the catheter-associated urinary tract infection strain Escherichia coli MS2027 was sequenced and annotated. This 42,644-bp plasmid, designated pMAS2027, contains 58 putative genes and is most closely related to plasmids belonging to incompatibility group X (IncX1). Plasmid pMAS2027 encodes two important virulence factors: type 3 fimbriae and a type IV secretion (T4S) system. Type 3 fimbriae, recently found to be functionally expressed in E. coli, played an important role in biofilm formation. Biofilm formation by E. coli MS2027 was specifically due to expression of type 3 fimbriae and not the T4S system. The T4S system, however, accounted for the conjugative ability of pMAS2027 and enabled a non-biofilm-forming strain to grow as part of a mixed biofilm following acquisition of this plasmid. Thus, the importance of conjugation as a mechanism to spread biofilm determinants was demonstrated. Conjugation may represent an important mechanism by which type 3 fimbria genes are transferred among the Enterobacteriaceae that cause device-related infections in nosocomial settings.

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Available from: Cheryl-lynn Y Ong, Feb 27, 2015
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    • "In particular, they cause problems with keeping abiotic surfaces sterile in hospital settings, leading to conditions such as urinary tract infections (UTI) [3]. As surface structures like fimbriae and conjugal pili are essential in the establishment of these kinds of bacterial plaques or biofilms, the occurrence of biofilm promoting genes on mobile genetic elements [4], [5] represent an alarming development in need of close monitoring. "
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    ABSTRACT: Plasmids are a highly effective means with which genetic traits that influence human health, such as virulence and antibiotic resistance, are disseminated through bacterial populations. The IncX-family is a hitherto sparsely populated group of plasmids that are able to thrive within Enterobacteriaceae. In this study, a replicon-centric screening method was used to locate strains from wastewater sludge containing plasmids belonging to the IncX-family. A transposon aided plasmid capture method was then employed to transport IncX-plasmids from their original hosts (and co-hosted plasmids) into a laboratory strain (Escherichia coli Genehogs®) for further study. The nucleotide sequences of the three newly isolated IncX-plasmids (pLN126_33, pMO17_54, pMO440_54) and the hitherto un-sequenced type-plasmid R485 revealed a remarkable occurrence of whole or partial gene cassettes that promote biofilm-formation in Klebsiella pneumonia or E. coli, in all four instances. Two of the plasmids (R485 and pLN126_33) were shown to directly induce biofilm formation in a crystal violet retention assay in E. coli. Sequence comparison revealed that all plasmid-borne forms of the type 3 fimbriae encoding gene cassette mrkABCDF were variations of a composite transposon Tn6011 first described in the E. coli IncX plasmid pOLA52. In conclusion, IncX-plasmids isolated from Enterobacteriaceae over almost 40 years and on three different continents have all been shown to carry a type 3 fimbriae gene cassette mrkABCDF stemming from pathogenic K. pneumoniae. Apart from contributing general knowledge about IncX-plasmids, this study also suggests an apparent ubiquity of a mobile form of an important virulence factor and is an illuminating example of the recruitment, evolution and dissemination of genetic traits through plasmid-mediated horizontal gene transfer.
    PLoS ONE 08/2012; 7(7):e41259. DOI:10.1371/journal.pone.0041259 · 3.23 Impact Factor
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    • "In this way, fate of plasmids in a bacterial community can be monitored in situ non-destructively. Using this approach, spread of different IncP-1 and IncP-9 plasmids was monitored in a variety of environments including agar-surface grown colonies (Christensen et al., 1996; Fox et al., 2008; Krone et al., 2007), biofilm model systems (Christensen et al., 1998; Haagensen et al., 2002; Hausner and Wuertz, 1999; Krol et al., 2011; Ong et al., 2009; Seoane et al., 2011), freshwater microcosms (Dahlberg et al., 1998) or plant leaves (Normander et al., 1998). Interestingly, spatial analysis of green fluorescence revealed that invasive spread of IncP plasmids was neither detectable in recipient colonies on agar surfaces nor in recipient microcolonies in flow-chamber biofilms suggesting that local factors limit plasmid transfer (Christensen et al., 1998, 1996; Fox et al., 2008). "
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    ABSTRACT: Most natural conjugative IncF plasmids encode a fertility inhibition system that represses transfer gene expression in the majority of plasmid-carrying cells. The successful spread of these plasmids in clinically relevant bacteria has been suggested to be supported by a transitory derepression of transfer gene expression in newly formed transconjugants. In this study, we aimed to monitor the extent of transitory derepression during agar surface matings in situ by comparing plasmid spread of the IncF plasmid R1 and its derepressed mutant R1drd19 at low initial cell densities. A zygotic induction strategy was used to visualize the spatial distribution of fluorescent transconjugants within the heterogeneous environment. Epifluorescence and confocal microscopy revealed different transfer patterns for both plasmids, however, spread beyond the first five recipient cell layers adjacent to the donor cells was not observed. Similar results were observed for other prototypical conjugative plasmids. These results cannot rule out that transitory derepression contributes to the limited R1 plasmid invasion, but other factors like nutrient availability or spatial structure seem to limit plasmid spread.
    Plasmid 01/2012; 67(2):155-61. DOI:10.1016/j.plasmid.2012.01.001 · 1.58 Impact Factor
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    • "Type 3 fimbriae are 2-4 nm wide and 0.5-2 μm long surface organelles that are characterised by their ability to mediate agglutination of tannic acid-treated human RBC (MR/K agglutination) [29]. Several studies have clearly demonstrated a role for type 3 fimbriae in biofilm formation [17,28,30-33]. Type 3 fimbriae also mediate various adherence functions such as binding to epithelial cells (from the respiratory and urinary tracts) and extracellular matrix proteins (e.g. "
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    ABSTRACT: Catheter-associated urinary tract infection (CAUTI) is the most common nosocomial infection in the United States and is caused by a range of uropathogens. Biofilm formation by uropathogens that cause CAUTI is often mediated by cell surface structures such as fimbriae. In this study, we characterised the genes encoding type 3 fimbriae from CAUTI strains of Escherichia coli, Klebsiella pneumoniae, Klebsiella oxytoca, Citrobacter koseri and Citrobacter freundii. Phylogenetic analysis of the type 3 fimbrial genes (mrkABCD) from 39 strains revealed they clustered into five distinct clades (A-E) ranging from one to twenty-three members. The majority of sequences grouped in clade A, which was represented by the mrk gene cluster from the genome sequenced K. pneumoniae MGH78578. The E. coli and K. pneumoniae mrkABCD gene sequences clustered together in two distinct clades, supporting previous evidence for the occurrence of inter-genera lateral gene transfer. All of the strains examined caused type 3 fimbriae mediated agglutination of tannic acid treated human erythrocytes despite sequence variation in the mrkD-encoding adhesin gene. Type 3 fimbriae deletion mutants were constructed in 13 representative strains and were used to demonstrate a direct role for type 3 fimbriae in biofilm formation. The expression of functional type 3 fimbriae is common to many Gram-negative pathogens that cause CAUTI and is strongly associated with biofilm growth. Our data provides additional evidence for the spread of type 3 fimbrial genes by lateral gene transfer. Further work is now required to substantiate the clade structure reported here by examining more strains as well as other bacterial genera that make type 3 fimbriae and cause CAUTI.
    BMC Microbiology 06/2010; 10(1):183. DOI:10.1186/1471-2180-10-183 · 2.73 Impact Factor
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