TY  - JOUR
AU  - Ćurčić, Jovana
AU  - Dinić, Miroslav
AU  - Novović, Katarina
AU  - Vasiljević, Zorica
AU  - Kojić, Milan
AU  - Jovčić, Branko
AU  - Malešević, Milka
PY  - 2024
UR  - http://intor.torlakinstitut.com/handle/123456789/864
AB  - Infections caused by multidrug-resistant pathogens are one of the biggest challenges facing the healthcare system today. Quorum quenching (QQ) enzymes have the potential to be used as innovative enzyme-based antivirulence therapeutics to combat infections caused by multidrug-resistant pathogens. The main objective of this research was to describe the novel YtnP lactonase derived from the clinical isolate Stenotrophomonas maltophilia and to investigate its antivirulence potential against multidrug-resistant Pseudomonas aeruginosa MMA83. YtnP lactonase, the QQ enzyme, belongs to the family of metallo-β-lactamases. The recombinant enzyme has several advantageous biotechnological properties, such as high thermostability, activity in a wide pH range, and no cytotoxic effect. High-performance liquid chromatography analysis revealed the activity of recombinant YtnP lactonase toward a wide range of N-acyl-homoserine lactones (AHLs), quorum sensing signaling molecules, with a higher preference for long-chain AHLs. Recombinant YtnP lactonase was shown to inhibit P. aeruginosa MMA83 biofilm formation, induce biofilm decomposition, and reduce extracellular virulence factors production. Moreover, the lifespan of MMA83-infected Caenorhabditis elegans was prolonged with YtnP lactonase treatment. YtnP lactonase showed synergistic inhibitory activity in combination with gentamicin and acted additively with meropenem against MMA83. The described properties make YtnP lactonase a promising therapeutic candidate for the development of next-generation antivirulence agents.
T2  - International Journal of Biological Macromolecules
T1  - A novel thermostable YtnP lactonase from Stenotrophomonas maltophilia inhibits Pseudomonas aeruginosa virulence in vitro and in vivo
SP  - 130421
DO  - 10.1016/j.ijbiomac.2024.130421
ER  - 

@article{
author = "Ćurčić, Jovana and Dinić, Miroslav and Novović, Katarina and Vasiljević, Zorica and Kojić, Milan and Jovčić, Branko and Malešević, Milka",
year = "2024",
abstract = "Infections caused by multidrug-resistant pathogens are one of the biggest challenges facing the healthcare system today. Quorum quenching (QQ) enzymes have the potential to be used as innovative enzyme-based antivirulence therapeutics to combat infections caused by multidrug-resistant pathogens. The main objective of this research was to describe the novel YtnP lactonase derived from the clinical isolate Stenotrophomonas maltophilia and to investigate its antivirulence potential against multidrug-resistant Pseudomonas aeruginosa MMA83. YtnP lactonase, the QQ enzyme, belongs to the family of metallo-β-lactamases. The recombinant enzyme has several advantageous biotechnological properties, such as high thermostability, activity in a wide pH range, and no cytotoxic effect. High-performance liquid chromatography analysis revealed the activity of recombinant YtnP lactonase toward a wide range of N-acyl-homoserine lactones (AHLs), quorum sensing signaling molecules, with a higher preference for long-chain AHLs. Recombinant YtnP lactonase was shown to inhibit P. aeruginosa MMA83 biofilm formation, induce biofilm decomposition, and reduce extracellular virulence factors production. Moreover, the lifespan of MMA83-infected Caenorhabditis elegans was prolonged with YtnP lactonase treatment. YtnP lactonase showed synergistic inhibitory activity in combination with gentamicin and acted additively with meropenem against MMA83. The described properties make YtnP lactonase a promising therapeutic candidate for the development of next-generation antivirulence agents.",
journal = "International Journal of Biological Macromolecules",
title = "A novel thermostable YtnP lactonase from Stenotrophomonas maltophilia inhibits Pseudomonas aeruginosa virulence in vitro and in vivo",
pages = "130421",
doi = "10.1016/j.ijbiomac.2024.130421"
}

Ćurčić, J., Dinić, M., Novović, K., Vasiljević, Z., Kojić, M., Jovčić, B.,& Malešević, M.. (2024). A novel thermostable YtnP lactonase from Stenotrophomonas maltophilia inhibits Pseudomonas aeruginosa virulence in vitro and in vivo. in International Journal of Biological Macromolecules, 130421.
https://doi.org/10.1016/j.ijbiomac.2024.130421

Ćurčić J, Dinić M, Novović K, Vasiljević Z, Kojić M, Jovčić B, Malešević M. A novel thermostable YtnP lactonase from Stenotrophomonas maltophilia inhibits Pseudomonas aeruginosa virulence in vitro and in vivo. in International Journal of Biological Macromolecules. 2024;:130421.
doi:10.1016/j.ijbiomac.2024.130421 .

Ćurčić, Jovana, Dinić, Miroslav, Novović, Katarina, Vasiljević, Zorica, Kojić, Milan, Jovčić, Branko, Malešević, Milka, "A novel thermostable YtnP lactonase from Stenotrophomonas maltophilia inhibits Pseudomonas aeruginosa virulence in vitro and in vivo" in International Journal of Biological Macromolecules (2024):130421,
https://doi.org/10.1016/j.ijbiomac.2024.130421 . .

TY  - DATA
AU  - Ljubić, Verica
AU  - Milošević, Milena
AU  - Cvetković, Slobodan
AU  - Stojanović, Marijana
AU  - Novović, Katarina
AU  - Dinić, Miroslav
AU  - Popović, Mina
PY  - 2022
UR  - http://intor.torlakinstitut.com/handle/123456789/644
AB  - Using the universal primers (UNI16Sfw and UNI16Srev) [1], the representative isolates were identified by 16S rDNA sequencing. Amplification was carried out in a thermal cycler (Applied Biosystems, ThermoFisher Scientific) and DNA fragments were amplified as follows: initial denaturation at 94 °C for 5 min, followed by 30 cycles consisting of denaturation at 94 °C for 1 min, annealing at 55°C for 1 min, and polymerization at 72°C for 1 min, and a final extension at 72°C for 7 min. The expected length was 1549 bp. Aliquots (5 µl) of the amplified products were subjected to electrophoresis in 1% agarose gel (ThermoFisher Scientific) in TAE buffer (40 mM Tris acetate, 1 mM EDTA, pH 8.2). Gels were stained with ethidium bromide (500 ng/mL) and visualized under UV light (BioDoc Analyze). All amplicons were eluted and purified using GeneJet PCR Purification Kit (ThermoScientific) by following the manufacturer`s protocol. The PCR products that we obtained were sequenced by the Macrogen Sequencing Service (Macrogen, Amsterdam, The Netherlands) and analyzed by using BLAST algorithm (http://www.ncbi.nlm.nih.gov/index.html). Selected isolates were identified as follows: isolate B2 - Lacotbacillus reuteri, isolate H10 - Lactobacillus murinus, and isolate J7 - Klebsiella oxytoca [2]. The most numerous colonies belong to isolate B2, hence it was chosen for further characterization as a potential source for exopolysaccharide (EPS) production.
PB  - Springer
T2  - Biomass Conversion and Biorefinery
T1  - Supplementary information for the article: Ljubic, V.; Milosevic, M.; Cvetkovic, S.; Stojanovic, M.; Novovic, K.; Dinic, M.; Popovic, M. The New Exopolysaccharide Produced by the Probiotic Strain L. Reuteri B2: Extraction, Biological Properties, and Possible Application for Ni2+ Ion Removal from the Contaminated Water. Biomass Conversion and Biorefinery 2022. https://doi.org/10.1007/s13399-022-03292-5.
UR  - https://hdl.handle.net/21.15107/rcub_intor_644
ER  - 

@misc{
author = "Ljubić, Verica and Milošević, Milena and Cvetković, Slobodan and Stojanović, Marijana and Novović, Katarina and Dinić, Miroslav and Popović, Mina",
year = "2022",
abstract = "Using the universal primers (UNI16Sfw and UNI16Srev) [1], the representative isolates were identified by 16S rDNA sequencing. Amplification was carried out in a thermal cycler (Applied Biosystems, ThermoFisher Scientific) and DNA fragments were amplified as follows: initial denaturation at 94 °C for 5 min, followed by 30 cycles consisting of denaturation at 94 °C for 1 min, annealing at 55°C for 1 min, and polymerization at 72°C for 1 min, and a final extension at 72°C for 7 min. The expected length was 1549 bp. Aliquots (5 µl) of the amplified products were subjected to electrophoresis in 1% agarose gel (ThermoFisher Scientific) in TAE buffer (40 mM Tris acetate, 1 mM EDTA, pH 8.2). Gels were stained with ethidium bromide (500 ng/mL) and visualized under UV light (BioDoc Analyze). All amplicons were eluted and purified using GeneJet PCR Purification Kit (ThermoScientific) by following the manufacturer`s protocol. The PCR products that we obtained were sequenced by the Macrogen Sequencing Service (Macrogen, Amsterdam, The Netherlands) and analyzed by using BLAST algorithm (http://www.ncbi.nlm.nih.gov/index.html). Selected isolates were identified as follows: isolate B2 - Lacotbacillus reuteri, isolate H10 - Lactobacillus murinus, and isolate J7 - Klebsiella oxytoca [2]. The most numerous colonies belong to isolate B2, hence it was chosen for further characterization as a potential source for exopolysaccharide (EPS) production.",
publisher = "Springer",
journal = "Biomass Conversion and Biorefinery",
title = "Supplementary information for the article: Ljubic, V.; Milosevic, M.; Cvetkovic, S.; Stojanovic, M.; Novovic, K.; Dinic, M.; Popovic, M. The New Exopolysaccharide Produced by the Probiotic Strain L. Reuteri B2: Extraction, Biological Properties, and Possible Application for Ni2+ Ion Removal from the Contaminated Water. Biomass Conversion and Biorefinery 2022. https://doi.org/10.1007/s13399-022-03292-5.",
url = "https://hdl.handle.net/21.15107/rcub_intor_644"
}

Ljubić, V., Milošević, M., Cvetković, S., Stojanović, M., Novović, K., Dinić, M.,& Popović, M.. (2022). Supplementary information for the article: Ljubic, V.; Milosevic, M.; Cvetkovic, S.; Stojanovic, M.; Novovic, K.; Dinic, M.; Popovic, M. The New Exopolysaccharide Produced by the Probiotic Strain L. Reuteri B2: Extraction, Biological Properties, and Possible Application for Ni2+ Ion Removal from the Contaminated Water. Biomass Conversion and Biorefinery 2022. https://doi.org/10.1007/s13399-022-03292-5.. in Biomass Conversion and Biorefinery
Springer..
https://hdl.handle.net/21.15107/rcub_intor_644

Ljubić V, Milošević M, Cvetković S, Stojanović M, Novović K, Dinić M, Popović M. Supplementary information for the article: Ljubic, V.; Milosevic, M.; Cvetkovic, S.; Stojanovic, M.; Novovic, K.; Dinic, M.; Popovic, M. The New Exopolysaccharide Produced by the Probiotic Strain L. Reuteri B2: Extraction, Biological Properties, and Possible Application for Ni2+ Ion Removal from the Contaminated Water. Biomass Conversion and Biorefinery 2022. https://doi.org/10.1007/s13399-022-03292-5.. in Biomass Conversion and Biorefinery. 2022;.
https://hdl.handle.net/21.15107/rcub_intor_644 .

Ljubić, Verica, Milošević, Milena, Cvetković, Slobodan, Stojanović, Marijana, Novović, Katarina, Dinić, Miroslav, Popović, Mina, "Supplementary information for the article: Ljubic, V.; Milosevic, M.; Cvetkovic, S.; Stojanovic, M.; Novovic, K.; Dinic, M.; Popovic, M. The New Exopolysaccharide Produced by the Probiotic Strain L. Reuteri B2: Extraction, Biological Properties, and Possible Application for Ni2+ Ion Removal from the Contaminated Water. Biomass Conversion and Biorefinery 2022. https://doi.org/10.1007/s13399-022-03292-5." in Biomass Conversion and Biorefinery (2022),
https://hdl.handle.net/21.15107/rcub_intor_644 .

TY  - JOUR
AU  - Miljković, Marija
AU  - Jovanović, Sofija
AU  - O'Connor, Paula M.
AU  - Mirković, Nemanja
AU  - Jovčić, Branko
AU  - Filipić, Brankica
AU  - Dinić, Miroslav
AU  - Studholme, David John
AU  - Fira, Đorđe
AU  - Cotter, Paul D.
AU  - Kojić, Milan
PY  - 2019
UR  - https://imagine.imgge.bg.ac.rs/handle/123456789/1264
UR  - http://intor.torlakinstitut.com/handle/123456789/702
AB  - Bacteria active against multi-drug resistant pathogens, isolated by direct selection of colonies from clover silage samples, produce zones of inhibition against two Gram-negative (Klebsiella pneumoniae Ni9 and Pseudomonas aeruginosa MMA83) and two Gram-positive (Staphylococcus aureus ATCC25923 and Listeria monocytogenes ATCC19111) pathogens. Isolates BGSP7, BGSP9, BGSP11 and BGSP12 produced the largest zones of inhibition against all four pathogens when grown in LB broth with aeration at 37 degrees C. Isolates BGSP7, BGSP9, BGSP11 and BGSP12 were identified as Brevibacillus laterosporus and pulsed field gel electrophoresis and extracellular protein profiles showed that three different strains (BGSP7, BGSP9 and BGSP11) were isolated. A semi-native SDS-PAGE (sodium dodecyl sulphate-polyacrylamide gel electrophoresis) gel overlay assay showed that BGSP7 and BGSP9 produce small antimicrobial molecules of about 1.5 kDa, while BGSP11 produces antimicrobial molecules of 1.5 and 6 kDa active against S. aureus ATCC25923. Amino acid analysis of two antimicrobial molecules (1583.73 Da; from BGSP7 and 1556.31 Da; from BGSP11) revealed that they have a similar composition and differ only by virtue of the presence of a methionine which is present only in BGSP11 molecule. Genome sequencing of the three isolates revealed the presence of gene clusters associated with the production of non-ribosomally synthesized peptides (brevibacillin, bogorol, gramicidin S, plipastatin and tyrocin) and bacteriocins (laterosporulin, a lactococcin 972-like bacteriocin, as well as putative linocin M18, sactipeptide, UviB and lantipeptide-like molecules). Ultimately, the purification of a number of antimicrobial molecules from each isolate suggests that they can be considered as potent biocontrol strains that produce an arsenal of antimicrobial molecules active against Gram-positive and Gram-negative multi-resistant pathogens, fungi and insects.
PB  - Public Library Science, San Francisco
T2  - PLoS One
T1  - Brevibacillus laterosporus strains BGSP7, BGSP9 and BGSP11 isolated from silage produce broad spectrum multi-antimicrobials
IS  - 5
VL  - 14
DO  - 10.1371/journal.pone.0216773
ER  - 

@article{
author = "Miljković, Marija and Jovanović, Sofija and O'Connor, Paula M. and Mirković, Nemanja and Jovčić, Branko and Filipić, Brankica and Dinić, Miroslav and Studholme, David John and Fira, Đorđe and Cotter, Paul D. and Kojić, Milan",
year = "2019",
abstract = "Bacteria active against multi-drug resistant pathogens, isolated by direct selection of colonies from clover silage samples, produce zones of inhibition against two Gram-negative (Klebsiella pneumoniae Ni9 and Pseudomonas aeruginosa MMA83) and two Gram-positive (Staphylococcus aureus ATCC25923 and Listeria monocytogenes ATCC19111) pathogens. Isolates BGSP7, BGSP9, BGSP11 and BGSP12 produced the largest zones of inhibition against all four pathogens when grown in LB broth with aeration at 37 degrees C. Isolates BGSP7, BGSP9, BGSP11 and BGSP12 were identified as Brevibacillus laterosporus and pulsed field gel electrophoresis and extracellular protein profiles showed that three different strains (BGSP7, BGSP9 and BGSP11) were isolated. A semi-native SDS-PAGE (sodium dodecyl sulphate-polyacrylamide gel electrophoresis) gel overlay assay showed that BGSP7 and BGSP9 produce small antimicrobial molecules of about 1.5 kDa, while BGSP11 produces antimicrobial molecules of 1.5 and 6 kDa active against S. aureus ATCC25923. Amino acid analysis of two antimicrobial molecules (1583.73 Da; from BGSP7 and 1556.31 Da; from BGSP11) revealed that they have a similar composition and differ only by virtue of the presence of a methionine which is present only in BGSP11 molecule. Genome sequencing of the three isolates revealed the presence of gene clusters associated with the production of non-ribosomally synthesized peptides (brevibacillin, bogorol, gramicidin S, plipastatin and tyrocin) and bacteriocins (laterosporulin, a lactococcin 972-like bacteriocin, as well as putative linocin M18, sactipeptide, UviB and lantipeptide-like molecules). Ultimately, the purification of a number of antimicrobial molecules from each isolate suggests that they can be considered as potent biocontrol strains that produce an arsenal of antimicrobial molecules active against Gram-positive and Gram-negative multi-resistant pathogens, fungi and insects.",
publisher = "Public Library Science, San Francisco",
journal = "PLoS One",
title = "Brevibacillus laterosporus strains BGSP7, BGSP9 and BGSP11 isolated from silage produce broad spectrum multi-antimicrobials",
number = "5",
volume = "14",
doi = "10.1371/journal.pone.0216773"
}

Miljković, M., Jovanović, S., O'Connor, P. M., Mirković, N., Jovčić, B., Filipić, B., Dinić, M., Studholme, D. J., Fira, Đ., Cotter, P. D.,& Kojić, M.. (2019). Brevibacillus laterosporus strains BGSP7, BGSP9 and BGSP11 isolated from silage produce broad spectrum multi-antimicrobials. in PLoS One
Public Library Science, San Francisco., 14(5).
https://doi.org/10.1371/journal.pone.0216773

Miljković M, Jovanović S, O'Connor PM, Mirković N, Jovčić B, Filipić B, Dinić M, Studholme DJ, Fira Đ, Cotter PD, Kojić M. Brevibacillus laterosporus strains BGSP7, BGSP9 and BGSP11 isolated from silage produce broad spectrum multi-antimicrobials. in PLoS One. 2019;14(5).
doi:10.1371/journal.pone.0216773 .

Miljković, Marija, Jovanović, Sofija, O'Connor, Paula M., Mirković, Nemanja, Jovčić, Branko, Filipić, Brankica, Dinić, Miroslav, Studholme, David John, Fira, Đorđe, Cotter, Paul D., Kojić, Milan, "Brevibacillus laterosporus strains BGSP7, BGSP9 and BGSP11 isolated from silage produce broad spectrum multi-antimicrobials" in PLoS One, 14, no. 5 (2019),
https://doi.org/10.1371/journal.pone.0216773 . .

TY  - JOUR
AU  - Novović, Katarina
AU  - Mihajlović, Sanja
AU  - Dinić, Miroslav
AU  - Malešević, Milka
AU  - Miljković, Marija
AU  - Kojić, Milan
AU  - Jovčić, Branko
PY  - 2018
UR  - https://imagine.imgge.bg.ac.rs/handle/123456789/1119
UR  - http://intor.torlakinstitut.com/handle/123456789/684
AB  - Acinetobacter baumannii has been recognized as one of the most challeging pathogens in clinical settings worldwide. Outer membrane porins play a significant role in Acinetobacter antibiotic resistance and virulence. A. baumannii carbapenem resistance and virulence factor porin Omp33-36 was the subject of this study. We investigated the omp33-36 gene transcriptional response in the growth phase, its response to carbapenems, and the effect of contact with host cells. Additionally, the cytotoxic effect of A. baumannii towards keratinocytes was assessed, as well as correlation between omp33-36 gene transcription and cytotoxicity. Further, Acinetobacter spp. Omp33-36 was classified and its characteristics relevant for vaccine candidature were determined. The level of the omp33-36 gene transcription varied between growth phases, but a common pattern could not be established among different strains. Treatment with subinhibitory concentrations of carbapenems decreased, while contact with keratinocytes increased omp33-36 expression in the analysed A. baumannii strains. Variations in omp33-36 mRNA levels did not correlate with cytotoxicity levels. Decrease of omp33-36 mRNA during treatment with subinhibitory concentrations of carbapenems, indicated the importance of transcriptional changes in reversible resistance to carbapenems due to the absence of Omp33-36. The transcription of omp33-36 increased after contact with keratinocytes, indicating the important role of de novo transcription during the initial phase of A. baumannii infection. Primary structural analysis of Acinetobacter spp. Omp33-36 revealed three distinct groups (among four A. baumannii variants). Although we have shown that Omp33-36 was highly polymorphic, we propose a potential antigen (PLAEAAFL motif) for vaccine development. According to PROVEAN analysis, the highly polymorphic structure of Omp33-36 porin should not influence its function significantly.
PB  - Public Library Science, San Francisco
T2  - PLoS One
T1  - Acinetobacter spp. porin Omp33-36: Classification and transcriptional response to carbapenems and host cells
IS  - 8
VL  - 13
DO  - 10.1371/journal.pone.0201608
ER  - 

@article{
author = "Novović, Katarina and Mihajlović, Sanja and Dinić, Miroslav and Malešević, Milka and Miljković, Marija and Kojić, Milan and Jovčić, Branko",
year = "2018",
abstract = "Acinetobacter baumannii has been recognized as one of the most challeging pathogens in clinical settings worldwide. Outer membrane porins play a significant role in Acinetobacter antibiotic resistance and virulence. A. baumannii carbapenem resistance and virulence factor porin Omp33-36 was the subject of this study. We investigated the omp33-36 gene transcriptional response in the growth phase, its response to carbapenems, and the effect of contact with host cells. Additionally, the cytotoxic effect of A. baumannii towards keratinocytes was assessed, as well as correlation between omp33-36 gene transcription and cytotoxicity. Further, Acinetobacter spp. Omp33-36 was classified and its characteristics relevant for vaccine candidature were determined. The level of the omp33-36 gene transcription varied between growth phases, but a common pattern could not be established among different strains. Treatment with subinhibitory concentrations of carbapenems decreased, while contact with keratinocytes increased omp33-36 expression in the analysed A. baumannii strains. Variations in omp33-36 mRNA levels did not correlate with cytotoxicity levels. Decrease of omp33-36 mRNA during treatment with subinhibitory concentrations of carbapenems, indicated the importance of transcriptional changes in reversible resistance to carbapenems due to the absence of Omp33-36. The transcription of omp33-36 increased after contact with keratinocytes, indicating the important role of de novo transcription during the initial phase of A. baumannii infection. Primary structural analysis of Acinetobacter spp. Omp33-36 revealed three distinct groups (among four A. baumannii variants). Although we have shown that Omp33-36 was highly polymorphic, we propose a potential antigen (PLAEAAFL motif) for vaccine development. According to PROVEAN analysis, the highly polymorphic structure of Omp33-36 porin should not influence its function significantly.",
publisher = "Public Library Science, San Francisco",
journal = "PLoS One",
title = "Acinetobacter spp. porin Omp33-36: Classification and transcriptional response to carbapenems and host cells",
number = "8",
volume = "13",
doi = "10.1371/journal.pone.0201608"
}

Novović, K., Mihajlović, S., Dinić, M., Malešević, M., Miljković, M., Kojić, M.,& Jovčić, B.. (2018). Acinetobacter spp. porin Omp33-36: Classification and transcriptional response to carbapenems and host cells. in PLoS One
Public Library Science, San Francisco., 13(8).
https://doi.org/10.1371/journal.pone.0201608

Novović K, Mihajlović S, Dinić M, Malešević M, Miljković M, Kojić M, Jovčić B. Acinetobacter spp. porin Omp33-36: Classification and transcriptional response to carbapenems and host cells. in PLoS One. 2018;13(8).
doi:10.1371/journal.pone.0201608 .

Novović, Katarina, Mihajlović, Sanja, Dinić, Miroslav, Malešević, Milka, Miljković, Marija, Kojić, Milan, Jovčić, Branko, "Acinetobacter spp. porin Omp33-36: Classification and transcriptional response to carbapenems and host cells" in PLoS One, 13, no. 8 (2018),
https://doi.org/10.1371/journal.pone.0201608 . .