Projects / Programmes
Molecular Biology of Microorganisms
January 1, 2009
- December 31, 2012
| Code |
Science |
Field |
Subfield |
| 1.05.00 |
Natural sciences and mathematics |
Biochemistry and molecular biology |
|
| 3.01.00 |
Medical sciences |
Microbiology and immunology |
|
| 4.06.00 |
Biotechnical sciences |
Biotechnology |
|
| Code |
Science |
Field |
| B230 |
Biomedical sciences |
Microbiology, bacteriology, virology, mycology |
| Code |
Science |
Field |
| 1.07 |
Natural Sciences |
Other natural sciences |
Bacteria, Archeae, Fungi, resistance, virulence, genetical mechanisms, antibiotics, halocins, membrane, transport, plasmids, regulation of gene expression, biosynthesis, transformation, conjugation, evolution, halophiles, psychrophiles
Researchers (12)
Organisations (1)
Abstract
Microorganisms are the most successful of organisms due to their exceptional metabolic diversity and successful competition for nutrients and space. For this purpose they: 1. synthesize antibiotics, 2. develop defence against antibiotics and 3. inhabit extreme environments.
1. Microorganisms are the causative agents of numerous diseases however, only if they can overcome the defence mechanisms of the host, successfully compete with other microorganisms and spread within the host. By interfering with the synthesis of virulence factors the infective process can be interrupted however, only if the mechanisms of their synthesis are understood.
Colicins are bacteriocins synthesized by Escherichia coli strains and act specifically against cells of the same or related species. They are thus antibiotics of a narrow range of activity which can destroy enteropathogens but not the remaining microbial flora. The goals and expected results of the proposed research are: understanding expression of colicin K as a basis for more efficient production of bacteriocins which would be used for therapeutic purposes, as well as a model for the elucidation of mechanisms which regulate virulence determinants of bacteria within a population of bacterial cells.
2. Microorganisms are successful also due to the synthesis of antimicrobial compounds. An example is the Bcr transporter from Bacillus licheniformis which mediates resistance against the peptide antibiotic bacitracin. Bcr is a member of the family of ABC transporters which can be either specific for a certain compound, for example an antibiotic or, nonspecific such as "multidrug resistance" proteins. It is interesting that both bacterial cells resistant against bacitracin due to the bacitracin transporter, as well as human cells with the P-glycoprotein exhibit sensitivity to detergents. Among the proteins that compose the Bcr transporter, the key protein is the transmembrane BcrC. We have found that BcrC is related to the PAP2 superfamily of proteins as well as that, it exhibits phosphatase activity. Our results indicate that BcrC is an essential bacterial protein. The aim of our planned research work is to elucidate BcrC's essential role.
3. Among the extreme environments are those with a low temperature and a high concentration of NaCl. In such environments fungi have until recently been unknown. We were first to describe extremophilic fungi in the waters of the Sečovlje salterns and in waters with extremely high NaCl content from three continents. Recently, studies of fungi in the extremely cold environment of the Arctic as well as extremely halophile Archaea have been commenced. Halophile and psychrophile fungi belong to the rare extremophile eukaryotes. As such they will be a valuable source of new biotechnological products with special characteristics and unique models for the studies of adaptation of cellular biomolecules to extreme conditions. The isolated fungi will be identified using classical and molecular techniques, with emphasis on determination of new species. On the level of physiological studies we will focus on adaptation of cell wall components, gene regulation of melanin synthesis, lipid composition of the membrane, its fluidity, participation of rafts, production of compatible solutes at high salt conditions and low temperatures, as well as ion concentration in the cytoplasm in relation to function of membrane ion pumps.
Halophile Archaea are important producers of biologically active substances, halocins. Different halocins will be studied at the protein and genetic level. Following isolation of halophile Archaea, biochemical and phylogenetic analysis will be used to identify the isolated strains and determine the production of halocins. These will be biochemically characterized. By studying the interaction of halocins with lipid membranes we will try to elucidate the mechanism of halocin action.
Significance for science
Our studies on the source of ESBL and metalo ß-lactamses represent an important contribution to the broader understanding of the evolution and ecology of antibiotic resistance determinants. Besides resistant bacteria, significant are also antibiotic tolerant bacteria as they represent a reservoir for chronic infections. We have found that there is no direct correlation between the SOS response, provoked by DNA damage, and ß-lactam antibiotics which are the most frequently prescribed antimicrobials. On the other hand, the SOS response could be involved in persister formation, presumably due to metabolic inactivity of a small part of the bacterial population rendering them antibiotic tolerant. Even though the first report on persisters was published in 1944, their formation has remained unresolved. At least two hypothesis have attempted to elucidate persister formation: 1) the SOS system, as part of the bacterial stress response, induces synthesis of the tisB gene product, the Tis toxin/antitoxin, which targets the membrane to reduce the proton motive force and ATP levels, promoting cell dormancy; and 2) accumulation of indol, which as a signal molecule in the stationary phase, induces synthesis of secreted proteins, toxic substances and proteins involved the bacterial response to oxidative stres. Neither hypothesis clearly elucidates persister formation. Our results investigating additional gene products involved in persister formation reveal new potential targets for development of novel antimicrobial agents. Using metagenomics as a tool to gain insight into the extent and nature of species' pangenome we found that the diversity of prokaryotic populations is preserved by phage predation. A combination of metagenomic and single-cell genomic approach on metagenomic DNA originating from saturated brine, revealed the presence of three new and abundant microbes belonging to recently the described Nanohaloarchaea and Gammaproteobacteria. Comparisons of macroscopic microbial communities found in limestone caves, based on 16S rRNA, showed that, these microorganisms are likely true cave dwellers and are not imported from the surface as all inspected caves supported communities composed mostly of phylotypes common to all sampling sites.
Significance for the country
In Slovenia and on a global scale antibiotic resistances represent one of the greatest threats to human and animal health. To successfully treat bacterial infections and limit the dissemination of antibiotic resistances careful surveillance of resistance determinants is essential. As antibiotic treatment is generally initiated emperically, the consecutive application of several antibiotics or combinations of antibiotics will be required, increasing costs as well as the possibility of further dissemination of antibiotic resistances. Hosptilization will be more frequent increasing treatment costs, influencing the quality of life and endangering other hospitalized patients. Novel antimicrobial substances with a more narrow range of activity should be used, targeting only the disease causing pathogens without disturbing the microbiota essential for health. Marine sponges are a rich source of antimicrobial agents with great potential for industrial as well as biomedical use. Additionally, using strains that produce antimicrobials, synthesized only in the pathogen, resistance evolution could be prevented. Persisters are likely the main cause of persistent chronic infections, particularly associated with hospital settings. Elucidation of the mechanisms involved in appearance of persisters would enhance the efficacy of traditional antibiotics and contribute to reduction in length of hospitilazation and health care costs. Bioresource is Slovenia's most valuable resource. The economic and strategic benefits that arise from our studies on extremely halophilic and subterranean environments include the discovery of microorganisms that produce novel antibiotics and therapeutic agents, enzymes and polymers for use in industrial and scientific applications. Throughout the years we have developed a network of international collaborators which enables us to use new sequencing technologies and single-cell genomics in our research, making it competitive at an international level.
Most important scientific results
Annual report
2009,
2010,
2011,
final report,
complete report on dLib.si
Most important socioeconomically and culturally relevant results
Annual report
2009,
2010,
2011,
final report,
complete report on dLib.si
