Attachment to and formation of biofilms on abiotic and biotic surfaces are important steps in the process by which many pathogens survive and proliferate under the hostile conditions they normally find in their ecological niches. Acquisition of essential nutrients such as iron is another condition that must be fulfilled by bacterial cells in order to proliferate under these adverse circumstances. Bacteria have responded to these challenges by expressing highly efficient systems that allow them to acquire iron from different sources and form cell communities on abiotic and biotic surfaces that proliferate successfully even under the harsh conditions found in clinical environments and the human host. Many of these properties are associated with the virulence phenotype of different pathogens and can be encoded by genetic traits that can be subjected to lateral gene transfer. The latter is a mechanism that plays a key role in bacterial evolution and adaptation a swell as in the emergence of bacterial pathogens with different virulence attributes.

Current Projects:

One of the main research projects in my laboratory focuses on the genetic and molecular characterization of iron acquisition systems in pathogenic bacteria. We have found that clinical isolates of Acinetobacter baumannii, an opportunistic pathogen that causes severe respiratory infections, express a siderophore-mediated iron acquisition system similar to the anguibactin system found in the fish pathogen Vibrio anguillarum. This finding suggests that the anguibactin siderophore-mediated iron acquisition system is part of a mobile genetic element that can be transferred among bacterial cells belonging to different genera and species. As part of these studies, we also became interested in histidine degradation and histamine biosynthesis and secretion by bacteria. Throughout our research we were the first to show that this biogenic amine is involved in siderophore biosynthesis and that it may play a role in the pathogenesis of the infections caused by these two unrelated disease-causing bacteria. We have also identified within a siderophore biosynthesis gene cluster located in the chromosome of A. baumannii the presence of genes encoding proteins belonging to the efflux pump protein family. We are analyzing these genes and their translation products with the purpose of elucidating the poorly characterized secretion process of bacterial siderophores. More recently, we became interested in the study of the genetic and molecular mechanisms that allow A. baumannii clinical isolates to form biofilms on abiotic surfaces. Classical molecular biology methods together with a novel insertion mutagenesis system and microscopy experiments led us to the identification of genes encoding structural proteins and uncharacterized regulatory factors that are essential for the formation of pili-like structures and cell attachment to plastic surfaces.

We are also interested in the characterization of siderophore-independent iron uptake systems such as those expressed by Actinobacillus actinomycetemcomitans. Our research demonstrated that this dental pathogen expresses at least two membrane transport systems potentially involved in iron acquisition and uses hemin as an iron source to grow under iron-deficient conditions. Recent studies showed that this dental pathogen harbors iron- and Fur-regulated genes, which may play a role in the physiology and virulence phenotype of this dental pathogen. Currently, we are in the process of generating isogenic derivatives affected in their ability to acquire this metal from iron-binding proteins with the purpose to elucidate the role of these acquisition systems in the pathogenesis of the dental disease caused by this bacterium. In addition, we are studying the effect of iron and the role of the iron repressor protein Fur on gene expression with a genome-wide approach that includes classical genetics as well as genomics and bioinformatics.

In addition, we are interested in the isolation and characterization of unique DNA fragments and genes involved in the pathogenesis of Brazilian purpuric fever, which is caused by an unusually invasive clone of Haemophilus influenzae biogroup aegyptius. Our results, which were obtained using a novel approach based on a PCR-based genome subtraction hybridization system, showed that the invasive clones of this pathogen contain unique DNA fragments that do not share significant similarity with genes deposited in the GenBank database. These novel sequences most likely represent fragments of DNA present in invasive strains that encode uncharacterized bacterial virulence factors. This possibility will be tested using genome-wide analysis and comparative genomics in collaboration with colleagues who have a long-standing interest in pathogens that belong to this bacterial family. These studies should provide information regarding the mechanisms responsible for the emergence of related bacteria expressing different virulence attributes.

Selected Publications:

• Tomaras, A. P., C. W. Dorsey, R. E. Edelmann, and L. A. Actis. Attachment to and biofilm formation on abiotic surfaces by Acinetobacter baumannii: involvement of a novel chaperone-usher pili assembly system. Microbiology, 149, 3461-3484, 2003.
• Dorsey, C. W., A. P. Tomaras, P. L. Connerly, M. E. Tolmasky, J. H. Crosa, and L. A. Actis. The siderophore-mediated iron acquisition systems of Acinetobacter baumannii ATCC 19606 and Vibrio anguillarum 775 are structurally and functionally related. Microbiology, 150, 3657-3667, 2004.
• McGillivary, G., A. P. Tomaras, E. R. Rhodes, and L. A. Actis. Cloning and sequencing of a genomic island found in the Brazilian purpuric fever clone of Haemophilus influenzae biogroup aegyptius, 73, 1927-1938, 2005.
• Rhodes, E. R., A. P. Tomaras, G. McGillivary, P. L. Connerly,and L. A. Actis. Genetic and functional analysis of the Actinobacillus actinomycetemcomitans AfeABCD siderophore independent iron-acquisition system. Infection and Immunity, 73, 3758-3763, 2005.
• McGillivary, G., L. M. Smoot, and L. A. Actis. Characterization of the IgA1 protease from the Brazilian purpuric fever strain F3031 of Haemophilus influenzae biogroup aegyptius. FEMS Microbiology Letters, 250, 229-236, 2005.
• Dorsey, C. W., A. P. Tomaras, and L. A. Actis. Sequence and organization of pMAC, an Acinetobacter baumannii plasmid harboring genes involved in organic peroxide resistance. Accepted for publication in Plasmid, 2006.