Natural Competence Is a Major Mechanism for Horizontal DNA Transfer in the Oral Pathogen Porphyromonas gingivalis

Porphyromonas gingivalis is a Gram-negative anaerobe that resides exclusively in the human oral cavity. Long-term colonization by P. gingivalis requires the bacteria to evade host immune responses while adapting to the changing host physiology and alterations in the composition of the oral microflora. The genetic diversity of P. gingivalis appears to reflect the variability of its habitat; however, little is known about the molecular mechanisms generating this diversity. Previously, our research group established that chromosomal DNA transfer occurs between P. gingivalis strains. In this study, we examine the role of putative DNA transfer genes in conjugation and transformation and demonstrate that natural competence mediated by comF is the dominant form of chromosomal DNA transfer, with transfer by a conjugation-like mechanism playing a minor role. Our results reveal that natural competence mechanisms are present in multiple strains of P. gingivalis, and DNA uptake is not sensitive to DNA source or modification status. Furthermore, extracellular DNA was observed for the first time in P. gingivalis biofilms and is predicted to be the major DNA source for horizontal transfer and allelic exchange between strains. We propose that exchange of DNA in plaque biofilms by a transformation-like process is of major ecological importance in the survival and persistence of P. gingivalis in the challenging oral environment

The DNA Transfer Region of Bacteroides Conjugative Transposons Tc(r)Em(r)DOT and Tc(r)ERL

157 p.Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 1999.This dissertation presents the first detailed study of the conjugal transfer genes found on a conjugative transposon. The complete sequence of a functional transfer region is presented and comparative analysis is made between the transfer gene sequences of several conjugative transposons. Two of the newly discovered transfer genes possess sequence homology with genes from other transfer systems and the significance of this relationship is discussed. Additionally, the first nucleotide and amino acid level homology between the Bacteroides conjugative transposons and the NBUs is revealed. Transfer of both NBUs and conjugative transposons depends on the regulatory protein RteB, but little is known about how RteB mediates its effect on both elements. This homology may provide a clue about this mechanism of regulation. Mutational analysis of the transfer region has so far revealed genes necessary for self-transfer of the conjugative transposon, but not plasmid mobilization. Antibodies for four of the transfer proteins have been used in protein localization studies to show that these gene products are localized to the cell membrane. The antibodies have also helped to demonstrate that a mutation in one of the transfer proteins results in increased production of the other transfer proteins. This mutation provides new insights into the complex regulatory system that controls transfer as mediated by the conjugative transposons. The combination of sequence data and a working transfer model will allow us to better compare and contrast the Bacteroides conjugative transposons with other systems for DNA transfer and protein export while also providing new insights into the process of DNA transfer in general.U of I OnlyRestricted to the U of I community idenfinitely during batch ingest of legacy ETD

Microbial Transport at Yellowstone: By Land, Sea, or Air?

Researcher George Bonheyo and his colleagues of the University of Illinois are featured in this web article that focuses on the means of microbial transport at Mammoth Hot Springs in Yellowstone National Park. Included within the site are numerous links to resources, other science topics, further readings involving microbes and Yellowstone National Park, and the University Of Illinois At Urbana-Champaign home page

DNA Persistence in a Sink Drain Environment.

Biofilms are organized structures composed mainly of cells and extracellular polymeric substances produced by the constituent microorganisms. Ubiquitous in nature, biofilms have an innate ability to capture and retain passing material and may therefore act as natural collectors of contaminants or signatures of upstream activities. To determine the persistence and detectability of DNA passing through a sink drain environment, Bacillus anthracis strain Ames35 was cultured (6.35 x 107 CFU/mL), sterilized, and disposed of by addition to a sink drain apparatus with an established biofilm. The sink drain apparatus was sampled before and for several days after the addition of the sterilized B. anthracis culture to detect the presence of B. anthracis DNA. Multiple PCR primer pairs were used to screen for chromosomal and plasmid DNA with primers targeting shorter sequences showing greater amplification efficiency and success. PCR amplification and detection of target sequences indicate persistence of chromosomal DNA and plasmid DNA in the biofilm for 5 or more and 14 or more days, respectively

Identification of Differential Gene Expression in Bacteria Associated with Coral Black Band Disease by Using RNA-Arbitrarily Primed PCR

RNA-arbitrarily primed PCR techniques have been applied for the first time to identify differential gene expression in black band disease (BBD), a virulent coral infection that affects reef ecosystems worldwide. The gene activity for the BBD mat on infected surfaces of the brain coral Diploria strigosa was compared with that for portions of the BBD mat that were removed from the coral and suspended nearby in the seawater column. The results obtained indicate that three genes (DD 95-2, DD 95-4, and DD 99-9) were up-regulated in the BBD bacterial mat on the coral surface compared to the transcript base levels observed in the BBD mat suspended in seawater. Clone DD 95-4 has homology with known amino acid ABC transporter systems in bacteria, while clone DD 99-9 exhibits homology with chlorophyll A apoprotein A1 in cyanobacteria. This protein is essential in the final conformation of photosystem I P700. DD 95-2, the only gene that was fully repressed in the BBD mat samples suspended in seawater, exhibited homology with the AraC-type DNA binding domain-containing proteins. These transcriptional activators coordinate the expression of genes essential for virulence in many species of gram-negative bacteria

Agarose gel images for <i>Bacillus anthracis</i> PCR samples.

<p>Depicted in A, from left to right, is 100 bp ladder, no-template control (NTC) and 3 reactions utilizing BA-5449 primer set, NTC and 3 reactions utilizing <i>cya</i>-1 primer set, NTC and 3 reactions utilizing <i>PA</i>7/6 primer set and 100 bp ladder. Depicted in B, from left to right, are duplicate reactions with bleached <i>Bacillus anthracis</i> template material and duplicate reactions with autoclaved <i>Bacillus anthracis</i> template material, with all 4 reactions utilizing the ITSeub primer pair.</p

Primers utilized for <i>Bacillus anthracis</i> & eubacteria DNA amplification and the expected amplicons.

<p>Primers utilized for <i>Bacillus anthracis</i> & eubacteria DNA amplification and the expected amplicons.</p