Molecular analysis of probiotic strains
antimicrobial activity mechanisms against urogenital infectious agents
Financing institution: National Council of Scientific
Research in Higher Education
Coordinating institution: University of Bucharest, Faculty of
Biology, Department of Genetics
Project
director: Lecturer dr. Diana Pelinescu (maiden name
Smarandache)
Project
code: TE 124
Duration of contract: 36 months
Project budget: 680000 lei
Abstract
Urogenital
infections frequency, high degree of recurrence and the extent of microorganism’s
antimicrobial resistance to antimicrobial compounds imply searching for some
alternative strategies for the prevention and treatment of the infections. The important progress made in molecular biology field
determine during last decades obtaining valuable information on microbial
ecosystems, including gastrointestinal and urogenital tract and on molecular
mechanisms involved in complex interaction between microorganisms and body
host. These studies imply understanding
of pathogenic microorganism’s action and also the beneficial effect of
commensal microorganism on host health, such as lactic bacteria. During the
last decades probiotic therapy was used for prevention and treatment of some
diseases. Although, there has been significant progress in knowledge the
probiotic effect, though the mechanisms involved in probiotic effect are not
fully understood and that especially because complex interaction occurring in
vivo.
Main objective:
The major
objective of the project consists in the
selection of probiotic lactic acid bacteria with antimicrobial activity,
represented either by the microbicides effect or by the ability to inhibit or
reduce expression of virulence and pathogenicity factors in bacteria and
yeasts, pathogenic strains isolated from urogenital tract.
Working plan (objectives)
Year 2010
1. Isolation of new strains of lactic acid bacteria from various ecosystems - primary collection
2. Isolation and identification of pathogenic strains by conventional taxonomy techniques
1. Isolation of new strains of lactic acid bacteria from various ecosystems - primary collection
2. Isolation and identification of pathogenic strains by conventional taxonomy techniques
Year 2011
3. Screening to identify lactic
acid bacteria strains with antimicrobial activity – microbicides effect
4. Evaluation of virulence and pathogenicity of pathogenic strains and selection of strains with high pathogenicity and virulence
5. Screening to identify lactic acid bacteria strains with the ability to reduce the virulence and pathogenicity of pathogenic strains
4. Evaluation of virulence and pathogenicity of pathogenic strains and selection of strains with high pathogenicity and virulence
5. Screening to identify lactic acid bacteria strains with the ability to reduce the virulence and pathogenicity of pathogenic strains
Year 2012
6. Identification of pathogenic
strains by molecular biology techniques
7. Identification of selected lactic acid bacteria strains by polyphasic taxonomy techniques
8. Studies on the mechanisms involved in antimicrobial activity
7. Identification of selected lactic acid bacteria strains by polyphasic taxonomy techniques
8. Studies on the mechanisms involved in antimicrobial activity
Year 2013
9. Studies on gene expression of virulence and pathogenicity
factors
The Team
Project director
Lecturer dr. Diana Roxana Pelinescu (NEE Smarandache)
Dr. D.
Pelinescu started her research activity in
1998 in the research center affiliated with the Department of Genetics - Faculty
of Biology, and since 2002 was admitted as laboratory assistant in the same
departments. Between 2004 and present she was Assistant professor (4 years) and
Lecturer (2008 - present). The main research topics addressed in those over 10
years of activity have included: a) studies on the molecular taxonomy of
microorganisms with biotechnological potential (lactic bacteria and yeasts); b)
optimization methods as genomic DNA and plasmidial isolation; c) analysis of
microbial biodiversity from human and animal gastrointestinal tract and
respectively in fermented foods; d) testing for potential probiotic strains of
lactic bacteria from indigenous products, the production of compounds with
antimicrobial activity (organic acids, peroxide, bacteriocin); e) molecular
analysis and mechanistic elucidation of the functionality of pro – and
prebiotics in the inhibition of pathogenic micro-organisms to combat
gastrointestinal disorders and to improve human health, f) biotechnological
studies on obtaining microbial biomass and bio-active compounds in the case of
strains of lactic bacteria used in the product type porobiotic / nutraceutic
and inoculums for silage; g) improvement of strains of lactic bacteria through
electrotransformation techniques, h) obtaining the mutant yeast strains with
application in waste water with high concentration of methanol; j) organization
and maintenance of a microbial culture collection
Project Members
CS Dr. Ionela Sarbu (NEE Avram)
Researcher
Sarbu Ionela works within University from Bucharest, Faculty of Biology -
Department of Genetics from 2007 and in 2012 was presented her PhD thesis
entitled " Study of cellular and molecular mechanisms involved in the
probiotic effect of lactic bacteria strains ". Mrs Sarbu research activity was focused on: studies regarding the
biodiversity of lactic acid bacteria strains with biotechnological applications
using molecular biology techniques, analysis of mechanisms involved in the
beneficial effect of probiotic, prebiotic, nutraceutical and sinbyotics
products (antimicrobial activity, hypocholesterolemic activity, s.o.).
Associated Professor Dr. Carmen Mariana Chifiriuc
(NEE Balotescu)
Dr.
Carmen Chifiriuc, Assoc. Prof in the Faculty of
Biology - Department of Microbiology and Systematic Botany received a
research grant (9 months -1999) to Network of Pasteur Institutes and Associated
Institutes at the Department of opportunistic infections of the Pasteur
Institute of Bangui . The results
obtained were part of her doctoral thesis titled: Parasite-host
relationship. Opportunistic infections in HIV patients, sustained in 2005
with magna cum laude. In 2001 she was selected for the national data
manager system EARSS (European Antimicrobial Resistance Surveillance System),
being actively involved in setting up the network in Romania and the collection
of samples and data for antibiotic resistance. The fields of research include:
analysis of pathogenic microorganisms microbiology, resistance mechanisms of
viral isolates from clinical and environmental, epidemiological surveillance of
antibiotic resistance, investigation of new alternative anti-infective,
including new environmental alternative to treat biofilm infections associated
or resistant.
CS Dr. Adriana Robertina Ionescu
Research
activity of Dr. Robertina Ionescu in 2003 in University of Bucharest,
Faculty of Biology – Department of Genetics has focused on studies regarding
polyphasic taxonomy on microorganisms with biotechnological applications
(bacterial strains, yeast and fungal strains), biochemical and genetic analysis
of microorganisms with biodegradative capabilities, biotechnological
studies for improving biomass formation and biosynthesis of and
biologically active compounds (biosurfactants) and analysis of microbial
biodiversity in various ecosystems.
Results
The main
results obtained in this research project were:
Ø
isolation and characterization of 75 lactic acid bacteria strains
from specific ecological niches (newborn feces, fermented dairy products,
women urogenital tract without clinical manifestations, breast milk, s.o.);
Ø
obtaining a small collection of pathogens
(bacteria and yeast) newly isolated from urogenital infections (20
Gram-positive bacterial strains from urine, 14 Gram-negative bacterial strains
and 13 yeast strains from vaginal secretions), characterized in terms of
morpho-physiologic, biochemical and genetic traits;
Ø
characterization of microbial strains in terms of phenotypic expression
of virulence factors and pathogenicity (enzymes biosynthesis, substrate
adherence capacity, resistance to antimicrobial compounds - antibiotics and
antifungal compounds) and selection of strains with high virulence and
pathogenicity;
Ø
determination of antimicrobial activity of lactic acid bacteria strains against
selected pathogenic microbial strains and correlation with organic acids
biosynthesis and/or bacteriocins;
Ø
phenotypic and genotypic analysis of the virulence and pathogenicity factors
expression in the presence of lactic acid bacteria strains and selection of
lactic acid bacteria strains which cause a decrease in virulence and
pathogenicity factors of selected pathogenic microbial strains.
Studies
have allowed the selection of lactic bacteria strains which can be used in treatment or prophylaxis of urogenital
infections by administering them either individually due to the antimicrobial
activity or together with antimicrobial compounds (antibiotics and antifungal
compunds).
The project contributed to the formation of young
researchers as Mrs. Ionela Sarbu, who completed her doctoral thesis in 2012 by
integrating the results obtained in the project. Optimization techniques
applied in the project has allowed their use in practical work with students of
Biology, Ecology and Biochemistry and admitted students in the Master of
Genetics, Microbiology and Biotechnology, Medical Biology and Molecular
Biology, held at the Faculty of Biology, University of Bucharest.
Scientific
publications:
- Ditu L.M., Chifiriuc M.C., Bezitzoglu E., Voltsi C., Bleotu C., Pelinescu D., Mihaescu G., Lazar V., 2010, Modulation of virulence and antibiotic susceptibility of enteropathogenic E. coli strains by Enterococcus faecium probiotic strain culture fractions, Anaerobe 17(6), p. 448-451
- Cotar A.I., Chifiriuc M.C., Dinu S., Pelinescu D., Banu O., Lazar V., 2010, Quantitative real-time PCR study of the influence of probiotic culture soluble fraction on the expression of Pseudomonas aeruginosa quorum sensing genes, Roum. Arch. Microbiol. Immunol. 69 (4), p. 213 -223
- Chifiriuc M.C., Badea Cioaca A., Lazar V., 2011, In vitro assay of the antimicrobial activity of kephir against bacterial and fungal strains, Anaerobe 17 (6), p. 433-435
- Sarbu I., Vassu T., Stoica I., Vamanu E., Pelinescu D.R., 2011, Selection of lactic acid bacteria strains producing exopolysaccharides, European Biotechnology Congress 2011, 28 sept – 1 oct,+ Istanbul, Turcia, publicat in Current Opinion in Biotechnology, vol. 22 supplement 1, p. S96 – S97
- Sarbu I., Vassu T., Sasarman E., Stoica I., Chifiriuc C.M., Ionescu R.A., Pelinescu D.R., 2011, Antimicrobial activity assessment of some lactic acid bacteria strains, FEMS 10th, Symposium on Lactic Acid Bacteria, 28 aug-1 sept., Egmond Aan Zee, Olanda , E067
- Ionescu R., Tanase A-M, Sarbu I., Pelinescu D., Vassu T., Csutak O., Chiciudean I., Stoica I., 2012, Taxonomic identification of Pseudomonas strains producing biosurfactants, 5th Croatian Congress of Microbiology with International Participation 26 – 30 sept, Croatia, P5, p.60.
- Sarbu I., Pelinescu D., Stoica I., Ionescu R., Marutescu L., Popa M., Chifiriuc C., Vassu T., 2012, Phenotpic virulence profile and taxonomic identification of microbial strains isolated from urogenital infections, 5th Croatian Congress of Microbiology with International Participation 26 – 30 sept , Croatia, P18, p.73.
- Sarbu I., Vassu T., Stoica I., Ionescu R., Dascalu L., Popa M., Chifiriuc C., Pelinescu D., 2012, Antimicrobial activity of some lactic acid bacteria strains against pathogenic strains isolated from urogenital infections, Central European Symposium on Antimicrobials and Antimicrobial Resistance - CESAR 2012, 23 -26 sept, Croatia, p. 66
- Cotar A.I., Ionescu B., Pelinescu D., Voidarou D., Lazar V., Bezirtzoglou E., Chifiriuc M.C., 2013, Current solution for the interception of quorum sensing in Staphylococcus aureus, Current Organic Chemistry 17, p. 97-104.
- Rusu E., Enache-Soare S., Cristescu C., Cristescu V., Pelinescu D., Sarbu I., Vassu T., 2013, Identification of species of the genus Candida by analysis of 5.8S rRNA gene, Indian Journal of Medical Microbiology (submited)
- Sarbu I., Pelinescu D., Stoica I., Ionescu R., Dascalu L., Alexandru I., Chifiriuc C., Rusu E., Nedelcu I., Vassu T., 2013, Influence of non steroidal anti-inflammatory drugs on antifungal resistance of Candida strains isolated from vulvo-vaginal infections, Farmacia (in press)
