TY  - CONF
AU  - Vukotić, Goran
AU  - Obradović, Mina
AU  - Plačkić, Nikola
AU  - Kljajević, Nemanja
AU  - Pavić, Aleksandar
AU  - Kekić, Dušan
AU  - Gajić, Ina
AU  - Kojić, Milan
AU  - Stanisavljević, Nemanja
PY  - 2024
UR  - http://intor.torlakinstitut.com/handle/123456789/885
AB  - Antimicrobial resistance (AMR) arises whenbacteria and other microbes stop respondingto medications. AMR is now recognized as oneof serious global health threats, repeatedlyappearing in the World Health Organization’s(WHO) lists of urgent global health challenges,including the 2024 list. It is taking a fatal toll– nearly 5 million deaths globally per year areassociated with AMR, encompassing 1.27 milliondirectly attributed to AMR. The COVID-19pandemic paved the way for aggravation ofbacterial AMR – primarily due to enhancementin unspecific and unjustified prescription anduse of broad-spectrum antibiotics, resulting inwhat is now recognized as „silent pandemic ofAMR“. Bacteriophages (phages) are natural andspecific predators of bacteria - viruses that caninfect, replicate inside and lyse arguably anybacteria. Their therapeutic potential is beinghastily evaluated through different approaches:in silico, in vitro, ex vivo and in vivo – in laboratoryanimals as well as in human case and clinicalstudies. Although the results are promising,bacteria rapidly develop resistance againstphages, which why the isolation and researchof new phages is needed. Our work is concentratedon three bacterial species for which criticalpriority by WHO has been declared – carbapenem-resistant Acinetobacter baumannii,Pseudomonas aeruginosa and Klebsiella pneumoniae.Twenty distinct pathogenic strains ofA. baumannii, 6 K. pneumoniae and 6 P. aeruginosawere used as targets for bacteriophageisolation, and total of 14, 22 and 8 potentiallydistinct phages were collected, respectively. Allstrains were nosocomial isolates obtained fromvarious tissues, including from terminally ill patients.Six phages were characterized in detail.In particular, phage vB_AbaM_ISTD was appliedagainst A. baumannii in zebrafish embryomodel of systemic infection, and demonstratedpowerful therapeutic potential, eradicating theinfection. Interestingly, its DNA was characterizedwith highly modified thymidine (amassing1228 Da), making it the largest non-canonicaldeoxynucleoside reported so far.
PB  - Serbian Society for Microbiology
C3  - XIII Congress of microbiologists of Serbia: From biotechnology to human and planetary health
T1  - Bacteriophages of multidrug-resistant nosocomial pathogens – Belgrade experience
EP  - 121
SP  - 121
ER  - 

@conference{
author = "Vukotić, Goran and Obradović, Mina and Plačkić, Nikola and Kljajević, Nemanja and Pavić, Aleksandar and Kekić, Dušan and Gajić, Ina and Kojić, Milan and Stanisavljević, Nemanja",
year = "2024",
abstract = "Antimicrobial resistance (AMR) arises whenbacteria and other microbes stop respondingto medications. AMR is now recognized as oneof serious global health threats, repeatedlyappearing in the World Health Organization’s(WHO) lists of urgent global health challenges,including the 2024 list. It is taking a fatal toll– nearly 5 million deaths globally per year areassociated with AMR, encompassing 1.27 milliondirectly attributed to AMR. The COVID-19pandemic paved the way for aggravation ofbacterial AMR – primarily due to enhancementin unspecific and unjustified prescription anduse of broad-spectrum antibiotics, resulting inwhat is now recognized as „silent pandemic ofAMR“. Bacteriophages (phages) are natural andspecific predators of bacteria - viruses that caninfect, replicate inside and lyse arguably anybacteria. Their therapeutic potential is beinghastily evaluated through different approaches:in silico, in vitro, ex vivo and in vivo – in laboratoryanimals as well as in human case and clinicalstudies. Although the results are promising,bacteria rapidly develop resistance againstphages, which why the isolation and researchof new phages is needed. Our work is concentratedon three bacterial species for which criticalpriority by WHO has been declared – carbapenem-resistant Acinetobacter baumannii,Pseudomonas aeruginosa and Klebsiella pneumoniae.Twenty distinct pathogenic strains ofA. baumannii, 6 K. pneumoniae and 6 P. aeruginosawere used as targets for bacteriophageisolation, and total of 14, 22 and 8 potentiallydistinct phages were collected, respectively. Allstrains were nosocomial isolates obtained fromvarious tissues, including from terminally ill patients.Six phages were characterized in detail.In particular, phage vB_AbaM_ISTD was appliedagainst A. baumannii in zebrafish embryomodel of systemic infection, and demonstratedpowerful therapeutic potential, eradicating theinfection. Interestingly, its DNA was characterizedwith highly modified thymidine (amassing1228 Da), making it the largest non-canonicaldeoxynucleoside reported so far.",
publisher = "Serbian Society for Microbiology",
journal = "XIII Congress of microbiologists of Serbia: From biotechnology to human and planetary health",
title = "Bacteriophages of multidrug-resistant nosocomial pathogens – Belgrade experience",
pages = "121-121"
}

Vukotić, G., Obradović, M., Plačkić, N., Kljajević, N., Pavić, A., Kekić, D., Gajić, I., Kojić, M.,& Stanisavljević, N.. (2024). Bacteriophages of multidrug-resistant nosocomial pathogens – Belgrade experience. in XIII Congress of microbiologists of Serbia: From biotechnology to human and planetary health
Serbian Society for Microbiology., 121-121.

Vukotić G, Obradović M, Plačkić N, Kljajević N, Pavić A, Kekić D, Gajić I, Kojić M, Stanisavljević N. Bacteriophages of multidrug-resistant nosocomial pathogens – Belgrade experience. in XIII Congress of microbiologists of Serbia: From biotechnology to human and planetary health. 2024;:121-121..

Vukotić, Goran, Obradović, Mina, Plačkić, Nikola, Kljajević, Nemanja, Pavić, Aleksandar, Kekić, Dušan, Gajić, Ina, Kojić, Milan, Stanisavljević, Nemanja, "Bacteriophages of multidrug-resistant nosocomial pathogens – Belgrade experience" in XIII Congress of microbiologists of Serbia: From biotechnology to human and planetary health (2024):121-121.

TY  - CONF
AU  - Malešević, Milka
AU  - Stanisavljević, Nemanja
AU  - Rašić, Slađan
AU  - Vukotić, Goran
AU  - Gardijan, Lazar
AU  - Obradović, Mina
AU  - Kojić, Milan
PY  - 2023
UR  - http://intor.torlakinstitut.com/handle/123456789/823
AB  - Introduction: Brevibacillus laterosporus is a promising microbiological agent that can be used to prevent and control destructive diseases affecting honey bee colonies. In the presentstudy, the short-termeffect of the B. laterosporus BGSP11 bee diet on microbiota and mycobiota was investigated.Methods: The honey bee diet was supplemented with spores of B. laterosporus BGSP11 at a concentration of 1×108 CFU/mL in sucrose solution. Metabarcoding analysis of the bee microbial community profile was performed based on 16S RNA (bacteriobiota) and Internally Transcribes Spacer (ITS) region(mycobiota) obtained using MiSeq Illumina sequencing. The QIIME2 v2021.4 pipeline was used to analyze the obtained amplicon data library.Results: The results show that the BGSP11 bee diet slightly altered the bee microbiota and did not leadto potentially harmful changes in the bacterial microbiota. Moreover, it can potentially induce positivechanges, mainly reflected in the reduction of opportunistic bacteria. On the other hand, the treatmenthad a greater effect on mycobiota. However, the changesin the bee mycobiome caused by the treatmentcannot be considered a priori as beneficial or harmful,since the interaction between the bee and its mycobiome is not sufficiently studied. The observed positive changes in the bee mycobiome are mainlyreflected in the reduction of phytopathogenic fungi that may affect the organoleptic and techno-functional properties of honey.Conclusion: This pilot study suggests that the introduction of BGSP11 in beekeeping practice as a biological agent could be considered due to no harmful effects observed on the microbiota of bees.
PB  - Institute of Molecular Genetics and Genetic Engineering (IMGGE), University of Belgrade
C3  - CoMBoS2 – the Second Congress of Molecular Biologists of Serbia, Abstract Book – Trends in Molecular Biology, Special issue 06-08 October 2023, Belgrade, Serbia
T1  - Short-term effect of Brevibacillus laterosporus supplemented diet on worker honey bee microbiome
EP  - 112
SP  - 112
UR  - https://hdl.handle.net/21.15107/rcub_intor_823
ER  - 

@conference{
author = "Malešević, Milka and Stanisavljević, Nemanja and Rašić, Slađan and Vukotić, Goran and Gardijan, Lazar and Obradović, Mina and Kojić, Milan",
year = "2023",
abstract = "Introduction: Brevibacillus laterosporus is a promising microbiological agent that can be used to prevent and control destructive diseases affecting honey bee colonies. In the presentstudy, the short-termeffect of the B. laterosporus BGSP11 bee diet on microbiota and mycobiota was investigated.Methods: The honey bee diet was supplemented with spores of B. laterosporus BGSP11 at a concentration of 1×108 CFU/mL in sucrose solution. Metabarcoding analysis of the bee microbial community profile was performed based on 16S RNA (bacteriobiota) and Internally Transcribes Spacer (ITS) region(mycobiota) obtained using MiSeq Illumina sequencing. The QIIME2 v2021.4 pipeline was used to analyze the obtained amplicon data library.Results: The results show that the BGSP11 bee diet slightly altered the bee microbiota and did not leadto potentially harmful changes in the bacterial microbiota. Moreover, it can potentially induce positivechanges, mainly reflected in the reduction of opportunistic bacteria. On the other hand, the treatmenthad a greater effect on mycobiota. However, the changesin the bee mycobiome caused by the treatmentcannot be considered a priori as beneficial or harmful,since the interaction between the bee and its mycobiome is not sufficiently studied. The observed positive changes in the bee mycobiome are mainlyreflected in the reduction of phytopathogenic fungi that may affect the organoleptic and techno-functional properties of honey.Conclusion: This pilot study suggests that the introduction of BGSP11 in beekeeping practice as a biological agent could be considered due to no harmful effects observed on the microbiota of bees.",
publisher = "Institute of Molecular Genetics and Genetic Engineering (IMGGE), University of Belgrade",
journal = "CoMBoS2 – the Second Congress of Molecular Biologists of Serbia, Abstract Book – Trends in Molecular Biology, Special issue 06-08 October 2023, Belgrade, Serbia",
title = "Short-term effect of Brevibacillus laterosporus supplemented diet on worker honey bee microbiome",
pages = "112-112",
url = "https://hdl.handle.net/21.15107/rcub_intor_823"
}

Malešević, M., Stanisavljević, N., Rašić, S., Vukotić, G., Gardijan, L., Obradović, M.,& Kojić, M.. (2023). Short-term effect of Brevibacillus laterosporus supplemented diet on worker honey bee microbiome. in CoMBoS2 – the Second Congress of Molecular Biologists of Serbia, Abstract Book – Trends in Molecular Biology, Special issue 06-08 October 2023, Belgrade, Serbia
Institute of Molecular Genetics and Genetic Engineering (IMGGE), University of Belgrade., 112-112.
https://hdl.handle.net/21.15107/rcub_intor_823

Malešević M, Stanisavljević N, Rašić S, Vukotić G, Gardijan L, Obradović M, Kojić M. Short-term effect of Brevibacillus laterosporus supplemented diet on worker honey bee microbiome. in CoMBoS2 – the Second Congress of Molecular Biologists of Serbia, Abstract Book – Trends in Molecular Biology, Special issue 06-08 October 2023, Belgrade, Serbia. 2023;:112-112.
https://hdl.handle.net/21.15107/rcub_intor_823 .

Malešević, Milka, Stanisavljević, Nemanja, Rašić, Slađan, Vukotić, Goran, Gardijan, Lazar, Obradović, Mina, Kojić, Milan, "Short-term effect of Brevibacillus laterosporus supplemented diet on worker honey bee microbiome" in CoMBoS2 – the Second Congress of Molecular Biologists of Serbia, Abstract Book – Trends in Molecular Biology, Special issue 06-08 October 2023, Belgrade, Serbia (2023):112-112,
https://hdl.handle.net/21.15107/rcub_intor_823 .

TY  - JOUR
AU  - Mirković, Nemanja
AU  - Obradović, Mina
AU  - O'Connor, Paula M.
AU  - Filipić, Brankica
AU  - Jovčić, Branko
AU  - Cotter, Paul D.
AU  - Kojić, Milan
PY  - 2021
UR  - https://imagine.imgge.bg.ac.rs/handle/123456789/1429
UR  - http://intor.torlakinstitut.com/handle/123456789/706
AB  - Screening for producers of potent antimicrobial peptides, resulted in the isolation of Bacillus cereus BGNM1 with strong antimicrobial activity against Listeria monocytogenes. Genome sequence analysis revealed that BGNM1 contains the gene cluster associated with the production of the lantibiotic, thusin, previously identified in B. thuringiensis. Purification of the antimicrobial activity confirmed that strain BGMN1 produces thusin. Both thusin sensitive and resistant strains were detected among clinical isolates of Streptococcus agalactiae. Random mutagenesis of a thusin sensitive strain, S. agalactiae B782, was performed in an attempt to identify the receptor protein for thusin. Three independent thusin resistant mutants were selected and their complete genomes sequenced. Comparative sequence analysis of these mutants with the WT strain revealed that duplication of a region encoding a 79 amino acids repeat in a C-protein alpha-antigen was a common difference, suggesting it to be responsible for increased resistance to thusin. Since induced thusin resistant mutants showed higher level of resistance than the naturally resistant B761 strain, complete genome sequencing of strain B761 was performed to check the integrity of the C-protein alpha-antigen-encoding gene. This analysis revealed that this gene is deleted in B761, providing further evidence that this protein promotes interaction of the thusin with receptor.
PB  - Springer, Dordrecht
T2  - Antonie Van Leeuwenhoek International Journal of General and Molecular Microbiology
T1  - C-protein alpha-antigen modulates the lantibiotic thusin resistance in Streptococcus agalactiae
EP  - 1607
IS  - 10
SP  - 1595
VL  - 114
DO  - 10.1007/s10482-021-01626-3
ER  - 

@article{
author = "Mirković, Nemanja and Obradović, Mina and O'Connor, Paula M. and Filipić, Brankica and Jovčić, Branko and Cotter, Paul D. and Kojić, Milan",
year = "2021",
abstract = "Screening for producers of potent antimicrobial peptides, resulted in the isolation of Bacillus cereus BGNM1 with strong antimicrobial activity against Listeria monocytogenes. Genome sequence analysis revealed that BGNM1 contains the gene cluster associated with the production of the lantibiotic, thusin, previously identified in B. thuringiensis. Purification of the antimicrobial activity confirmed that strain BGMN1 produces thusin. Both thusin sensitive and resistant strains were detected among clinical isolates of Streptococcus agalactiae. Random mutagenesis of a thusin sensitive strain, S. agalactiae B782, was performed in an attempt to identify the receptor protein for thusin. Three independent thusin resistant mutants were selected and their complete genomes sequenced. Comparative sequence analysis of these mutants with the WT strain revealed that duplication of a region encoding a 79 amino acids repeat in a C-protein alpha-antigen was a common difference, suggesting it to be responsible for increased resistance to thusin. Since induced thusin resistant mutants showed higher level of resistance than the naturally resistant B761 strain, complete genome sequencing of strain B761 was performed to check the integrity of the C-protein alpha-antigen-encoding gene. This analysis revealed that this gene is deleted in B761, providing further evidence that this protein promotes interaction of the thusin with receptor.",
publisher = "Springer, Dordrecht",
journal = "Antonie Van Leeuwenhoek International Journal of General and Molecular Microbiology",
title = "C-protein alpha-antigen modulates the lantibiotic thusin resistance in Streptococcus agalactiae",
pages = "1607-1595",
number = "10",
volume = "114",
doi = "10.1007/s10482-021-01626-3"
}

Mirković, N., Obradović, M., O'Connor, P. M., Filipić, B., Jovčić, B., Cotter, P. D.,& Kojić, M.. (2021). C-protein alpha-antigen modulates the lantibiotic thusin resistance in Streptococcus agalactiae. in Antonie Van Leeuwenhoek International Journal of General and Molecular Microbiology
Springer, Dordrecht., 114(10), 1595-1607.
https://doi.org/10.1007/s10482-021-01626-3

Mirković N, Obradović M, O'Connor PM, Filipić B, Jovčić B, Cotter PD, Kojić M. C-protein alpha-antigen modulates the lantibiotic thusin resistance in Streptococcus agalactiae. in Antonie Van Leeuwenhoek International Journal of General and Molecular Microbiology. 2021;114(10):1595-1607.
doi:10.1007/s10482-021-01626-3 .

Mirković, Nemanja, Obradović, Mina, O'Connor, Paula M., Filipić, Brankica, Jovčić, Branko, Cotter, Paul D., Kojić, Milan, "C-protein alpha-antigen modulates the lantibiotic thusin resistance in Streptococcus agalactiae" in Antonie Van Leeuwenhoek International Journal of General and Molecular Microbiology, 114, no. 10 (2021):1595-1607,
https://doi.org/10.1007/s10482-021-01626-3 . .

TY  - JOUR
AU  - Vukotić, Goran
AU  - Obradović, Mina
AU  - Novović, Katarina
AU  - Di Luca, Mariagrazia
AU  - Jovčić, Branko
AU  - Fira, Đorđe
AU  - Neve, Horst
AU  - Kojić, Milan
AU  - McAuliffe, Olivia
PY  - 2020
UR  - http://intor.torlakinstitut.com/handle/123456789/721
AB  - Acinetobacter baumanniiis a leading cause of healthcare-associated infections worldwide. Its various intrinsic and acquired mechanisms of antibiotic resistance make the therapeutic challenge even more serious. One of the promising alternative treatments that is increasingly highlighted is phage therapy, the therapeutic use of bacteriophages to treat bacterial infections. Two phages active against nosocomial carbapenem-resistantA. baumanniistrain 6077/12, vB_AbaM_ISTD, and vB_AbaM_NOVI, were isolated from Belgrade wastewaters, purified, and concentrated using CsCl gradient ultracentrifugation. The phages were screened against 103 clinical isolates ofA. baumanniifrom a laboratory collection and characterized based on plaque and virion morphology, host range, adsorption rate, and one-step growth curve. Given that phage ISTD showed a broader host range, better adsorption rate, shorter latent period, and larger burst size, its ability to lyse planktonic and biofilm-embedded cells was tested in detail. Phage ISTD yielded a 3.5- and 2-log reduction in planktonic and biofilm-associated viable bacterial cell count, respectively, but the effect was time-dependent. Both phages produced growing turbid halos around plaques indicating the synthesis of depolymerases, enzymes capable of degrading bacterial exopolysaccharides. Halos tested positive for presence of phages in the proximity of the plaque, but not further from the plaque, which indicates that the observed halo enlargement is a consequence of enzyme diffusion through the agar, independently of the phages. This notion was also supported by the growing halos induced by phage preparations applied on pregrown bacterial lawns, indicating that depolymerizing effect was achieved also on non-dividing sensitive cells. Overall, good rates of growth, fast adsorption rate, broad host range, and high depolymerizing activity, as well as antibacterial effectiveness against planktonic and biofilm-associated bacteria, make these phages good candidates for potential application in combatingA. baumanniiinfections.
PB  - Frontiers Media Sa, Lausanne
T2  - Frontiers in Medicine
T1  - Characterization, Antibiofilm, and Depolymerizing Activity of Two Phages Active on Carbapenem-ResistantAcinetobacter baumannii
VL  - 7
DO  - 10.3389/fmed.2020.00426
ER  - 

@article{
author = "Vukotić, Goran and Obradović, Mina and Novović, Katarina and Di Luca, Mariagrazia and Jovčić, Branko and Fira, Đorđe and Neve, Horst and Kojić, Milan and McAuliffe, Olivia",
year = "2020",
abstract = "Acinetobacter baumanniiis a leading cause of healthcare-associated infections worldwide. Its various intrinsic and acquired mechanisms of antibiotic resistance make the therapeutic challenge even more serious. One of the promising alternative treatments that is increasingly highlighted is phage therapy, the therapeutic use of bacteriophages to treat bacterial infections. Two phages active against nosocomial carbapenem-resistantA. baumanniistrain 6077/12, vB_AbaM_ISTD, and vB_AbaM_NOVI, were isolated from Belgrade wastewaters, purified, and concentrated using CsCl gradient ultracentrifugation. The phages were screened against 103 clinical isolates ofA. baumanniifrom a laboratory collection and characterized based on plaque and virion morphology, host range, adsorption rate, and one-step growth curve. Given that phage ISTD showed a broader host range, better adsorption rate, shorter latent period, and larger burst size, its ability to lyse planktonic and biofilm-embedded cells was tested in detail. Phage ISTD yielded a 3.5- and 2-log reduction in planktonic and biofilm-associated viable bacterial cell count, respectively, but the effect was time-dependent. Both phages produced growing turbid halos around plaques indicating the synthesis of depolymerases, enzymes capable of degrading bacterial exopolysaccharides. Halos tested positive for presence of phages in the proximity of the plaque, but not further from the plaque, which indicates that the observed halo enlargement is a consequence of enzyme diffusion through the agar, independently of the phages. This notion was also supported by the growing halos induced by phage preparations applied on pregrown bacterial lawns, indicating that depolymerizing effect was achieved also on non-dividing sensitive cells. Overall, good rates of growth, fast adsorption rate, broad host range, and high depolymerizing activity, as well as antibacterial effectiveness against planktonic and biofilm-associated bacteria, make these phages good candidates for potential application in combatingA. baumanniiinfections.",
publisher = "Frontiers Media Sa, Lausanne",
journal = "Frontiers in Medicine",
title = "Characterization, Antibiofilm, and Depolymerizing Activity of Two Phages Active on Carbapenem-ResistantAcinetobacter baumannii",
volume = "7",
doi = "10.3389/fmed.2020.00426"
}

Vukotić, G., Obradović, M., Novović, K., Di Luca, M., Jovčić, B., Fira, Đ., Neve, H., Kojić, M.,& McAuliffe, O.. (2020). Characterization, Antibiofilm, and Depolymerizing Activity of Two Phages Active on Carbapenem-ResistantAcinetobacter baumannii. in Frontiers in Medicine
Frontiers Media Sa, Lausanne., 7.
https://doi.org/10.3389/fmed.2020.00426

Vukotić G, Obradović M, Novović K, Di Luca M, Jovčić B, Fira Đ, Neve H, Kojić M, McAuliffe O. Characterization, Antibiofilm, and Depolymerizing Activity of Two Phages Active on Carbapenem-ResistantAcinetobacter baumannii. in Frontiers in Medicine. 2020;7.
doi:10.3389/fmed.2020.00426 .

Vukotić, Goran, Obradović, Mina, Novović, Katarina, Di Luca, Mariagrazia, Jovčić, Branko, Fira, Đorđe, Neve, Horst, Kojić, Milan, McAuliffe, Olivia, "Characterization, Antibiofilm, and Depolymerizing Activity of Two Phages Active on Carbapenem-ResistantAcinetobacter baumannii" in Frontiers in Medicine, 7 (2020),
https://doi.org/10.3389/fmed.2020.00426 . .