27 research outputs found
Influence of newly-synthesized chalcone derivatives on growth, biofilm production, and virulence factors expression of multiresistant Acinetobacter baumannii strains
Acinetobacter baumannii je nozokomijalni, multirezistentni patogen, koga karakterišesposobnost perzistencije na neživim površinama i mogućnost veoma brzog sticanja rezistencije naantibiotike. Danas su u svetu rasprostranjeni izrazito rezistentni sojevi protiv kojih u mnogimzdravstvenim ustanovama ne postoji efikasna terapija, a pronalazak alternativnih terapijskihpristupa je od izuzetne važnosti. Halkoni su jedinjenja sa potvrđenim antimikrobnim svojstvima ipokazanim različitim antivirulentnim aktivnostima. Ciljevi istraživanja ovog rada bili suodređivanje profila rezistencije, ispitivanje mogućnosti kontaminacije antiseptika i ispitivanjeprodukcije biofilma identifikovanih kliničkih izolata A. baumannii, kao i sinteza derivatahidroksihalkona i ispitivanje njihovih antimikrobnih i antivirulentnih aktivnosti protiv ovih izolata.Osetljivost izolata na antibiotike ispitana je kombinacijom difuzionih, dilucionih iautomatizovanih metoda, a identifikovani kolistin-rezistentni izolati dodatno su podvrgnutisekvenciranju celog genoma (WGS) i genetički su okarakterisani. Takođe, mehanizmi rezistencijena kolistin ispitani su primenom komparativne analize genoma i Real-Time kvantitativne lančanereakcije polimeraze (RT-qPCR). Time-kill test je primenjen za ispitivanje perzistencije uantisepticima, a nivo produkcije biofilma ispitan je pod različitim uslovima kultivacije in vitrostatičkom metodom uz bojenje safraninom. Derivati hidroksihalkona sintetisani su pomoću Claisen-Schmidt kondenzacije i njihove antimikrobne aktivnosti, samih i u kombinaciji sa antibioticima,ispitane su bujon-mikrodilucionom, Time-kill i Checkerboard analizom. Antivirulentne aktivnostiodabranih halkona procenjene su posredstvom uticaja na produkciju biofilma (monomikrobnog ipolimikrobnog), vijabilnost biofilmskih ćelija, ekspresiju motiliteta, gensku ekspresiju faktoravirulencije (OmpA, Bap i AbaI), adheziju A. baumannii na komponente ekstracelularnog matriksa(ECM), kao što su fibronektin i kolagen, i aktivnost sistema međućelijske komunikacije (Quorum-Sensing, QS).Klinički izolati A. baumannii gotovo uniformno bili su rezistentni na karbapeneme, a čakskoro 19% izolata bilo je rezistentno na kolistin, pripadajući tako ekstenzivno rezistentnom ilipanrezistentnom fenotipu. Izolati su pokazali sposobnost kontaminacije antiseptika i produkcijevelikih količina biofilma. Nutritivni sastav hranljivih medijuma značajno je uticao na nivoprodukcije biofilma, dok se visok nivo produkcije održao pri širokom opsegu različitih temperaturainkubacije i u prisustvu subinhibitornih koncentracija antibiotika. Sintetisani halkoni ispoljili suumerenu antimikrobnu aktivnost, pri čemu su metoksi-supstituisani derivati u proseku najjačeinhibirali rast. Takođe, zabeleženo je nekoliko sinergističkih interakcija halkona sa meropenemom,a inhibicija efluksnih pumpi predložena je kao potencijalni mehanizam. Halkoni su pokazalisposobnost značajne inhibicije motiliteta i produkcije biofilma, a metoksi-supstituisani derivat (o-OCH3) ispoljio je značajnu antivirulentnu aktivnost posredstvom nishodne regulacije ekspresijeompA, bap i abaI gena i inhibicije adhezije na komponente ECM. Na osnovu ovih rezultata, o-OCH3 halkon je identifikovan kao potentni antivirulentni agens protiv A. baumannii.Acinetobacter baumannii is a nosocomial, multiresistant pathogen, able to persist on abioticsurfaces and to rapidly acquire antibiotic resistance. Nowadays, highly resistant strains are widelydisseminated throughout the world, and the discovery of alternative therapeutic strategies is of utterimportance. Chalcones are compounds whose antimicrobial properties are well-known and forwhich different antivirulence activities have been demonstrated. The aims of this research were todetermine resistance profiles, to evaluate the possibility of antiseptic contamination, and to analyzethe biofilm production of identified A. baumannii clinical isolates, as well as to synthesizehydroxychalcone derivatives and to investigate their antimicrobial and antivirulence activitiesagainst these isolates.Antibiotic susceptibility of the isolates was tested by combination of diffusion, dilution, andautomated methods, and additionally, identified colistin-resistant isolates were subjected to wholegenome sequencing (WGS) and were genetically characterized. Also, colistin resistancemechanisms were explored by using comparative genome analysis and Real-Time quantitativepolymerase chain reaction (RT-qPCR). Time-kill test was used for the measurement of bacterialsurvival in antiseptics, whereas the level of biofilm production under different cultivationconditions was quantified by in vitro static method using safranin stain. Hydroxychalconederivatives were synthesized by Claisen-Schmidt condensation, and their antimicrobial activities,alone and in combination with antibiotics, were investigated using broth-microdilution, Time-kill,and Checkerboard analyses. Antivirulence activities of selected chalcones were evaluated based onthe impact on biofilm production (monomicrobial and polymicrobial), biofilm cell viability,motility, virulence factors (OmpA, Bap, and AbaI) gene expression, fibronectin- and collagen-mediated adhesion, and quorum-sensing (QS) activity.A. baumannii clinical isolates expressed extensive drug-resistant or pan-drug resistantphenotypes, being nearly uniformly resistant to carbapenems. Almost 19% of isolates were resistantto colistin as well. The isolates proved to be able to contaminate the antiseptic solutions and toproduce large quantities of biofilms. Nutritional composition of growth media significantly affectedthe level of biofilm production. In contrast, wide range of different incubation temperatures and thepresence of antibiotics at subinhibitory concentrations had little effect, and the bacteria managed tomaintain high level of biofilm production. Moderate antimicrobial activity was displayed bysynthesized chalcones, among which methoxy-substituted derivatives achieved greatest growthinhibition in average. Also, synergistic activity of chalcones and meropenem was present in severalcases, for which efflux pump inhibition was proposed as the potential mechanism. The chalconessignificantly inhibited motility and biofilm production, whereas methoxy-substituted derivative (o-OCH3) also displayed significant antivirulence activity, by downregulating the ompA, bap, and abaIgene expression and by inhibiting fibronectin- and collagen-mediated adhesion. It can be concludedthat o-OCH3 has been identified as a potent antivirulence agent against A. baumannii
Influence of newly-synthesized chalcone derivatives on growth, biofilm production, and virulence factors expression of multiresistant Acinetobacter baumannii strains
Acinetobacter baumannii je nozokomijalni, multirezistentni patogen, koga karakteriše
sposobnost perzistencije na neživim površinama i mogućnost veoma brzog sticanja rezistencije na
antibiotike. Danas su u svetu rasprostranjeni izrazito rezistentni sojevi protiv kojih u mnogim
zdravstvenim ustanovama ne postoji efikasna terapija, a pronalazak alternativnih terapijskih
pristupa je od izuzetne važnosti. Halkoni su jedinjenja sa potvrđenim antimikrobnim svojstvima i
pokazanim različitim antivirulentnim aktivnostima. Ciljevi istraživanja ovog rada bili su
određivanje profila rezistencije, ispitivanje mogućnosti kontaminacije antiseptika i ispitivanje
produkcije biofilma identifikovanih kliničkih izolata A. baumannii, kao i sinteza derivata
hidroksihalkona i ispitivanje njihovih antimikrobnih i antivirulentnih aktivnosti protiv ovih izolata.
Osetljivost izolata na antibiotike ispitana je kombinacijom difuzionih, dilucionih i
automatizovanih metoda, a identifikovani kolistin-rezistentni izolati dodatno su podvrgnuti
sekvenciranju celog genoma (WGS) i genetički su okarakterisani. Takođe, mehanizmi rezistencije
na kolistin ispitani su primenom komparativne analize genoma i Real-Time kvantitativne lančane
reakcije polimeraze (RT-qPCR). Time-kill test je primenjen za ispitivanje perzistencije u
antisepticima, a nivo produkcije biofilma ispitan je pod različitim uslovima kultivacije in vitro
statičkom metodom uz bojenje safraninom. Derivati hidroksihalkona sintetisani su pomoću Claisen-
Schmidt kondenzacije i njihove antimikrobne aktivnosti, samih i u kombinaciji sa antibioticima,
ispitane su bujon-mikrodilucionom, Time-kill i Checkerboard analizom. Antivirulentne aktivnosti
odabranih halkona procenjene su posredstvom uticaja na produkciju biofilma (monomikrobnog i
polimikrobnog), vijabilnost biofilmskih ćelija, ekspresiju motiliteta, gensku ekspresiju faktora
virulencije (OmpA, Bap i AbaI), adheziju A. baumannii na komponente ekstracelularnog matriksa
(ECM), kao što su fibronektin i kolagen, i aktivnost sistema međućelijske komunikacije (Quorum-
Sensing, QS).
Klinički izolati A. baumannii gotovo uniformno bili su rezistentni na karbapeneme, a čak
skoro 19% izolata bilo je rezistentno na kolistin, pripadajući tako ekstenzivno rezistentnom ili
panrezistentnom fenotipu. Izolati su pokazali sposobnost kontaminacije antiseptika i produkcije
velikih količina biofilma. Nutritivni sastav hranljivih medijuma značajno je uticao na nivo
produkcije biofilma, dok se visok nivo produkcije održao pri širokom opsegu različitih temperatura
inkubacije i u prisustvu subinhibitornih koncentracija antibiotika. Sintetisani halkoni ispoljili su
umerenu antimikrobnu aktivnost, pri čemu su metoksi-supstituisani derivati u proseku najjače
inhibirali rast. Takođe, zabeleženo je nekoliko sinergističkih interakcija halkona sa meropenemom,
a inhibicija efluksnih pumpi predložena je kao potencijalni mehanizam. Halkoni su pokazali
sposobnost značajne inhibicije motiliteta i produkcije biofilma, a metoksi-supstituisani derivat (o-
OCH3) ispoljio je značajnu antivirulentnu aktivnost posredstvom nishodne regulacije ekspresije
ompA, bap i abaI gena i inhibicije adhezije na komponente ECM. Na osnovu ovih rezultata, o-
OCH3 halkon je identifikovan kao potentni antivirulentni agens protiv A. baumannii.Acinetobacter baumannii is a nosocomial, multiresistant pathogen, able to persist on abiotic
surfaces and to rapidly acquire antibiotic resistance. Nowadays, highly resistant strains are widely
disseminated throughout the world, and the discovery of alternative therapeutic strategies is of utter
importance. Chalcones are compounds whose antimicrobial properties are well-known and for
which different antivirulence activities have been demonstrated. The aims of this research were to
determine resistance profiles, to evaluate the possibility of antiseptic contamination, and to analyze
the biofilm production of identified A. baumannii clinical isolates, as well as to synthesize
hydroxychalcone derivatives and to investigate their antimicrobial and antivirulence activities
against these isolates.
Antibiotic susceptibility of the isolates was tested by combination of diffusion, dilution, and
automated methods, and additionally, identified colistin-resistant isolates were subjected to whole
genome sequencing (WGS) and were genetically characterized. Also, colistin resistance
mechanisms were explored by using comparative genome analysis and Real-Time quantitative
polymerase chain reaction (RT-qPCR). Time-kill test was used for the measurement of bacterial
survival in antiseptics, whereas the level of biofilm production under different cultivation
conditions was quantified by in vitro static method using safranin stain. Hydroxychalcone
derivatives were synthesized by Claisen-Schmidt condensation, and their antimicrobial activities,
alone and in combination with antibiotics, were investigated using broth-microdilution, Time-kill,
and Checkerboard analyses. Antivirulence activities of selected chalcones were evaluated based on
the impact on biofilm production (monomicrobial and polymicrobial), biofilm cell viability,
motility, virulence factors (OmpA, Bap, and AbaI) gene expression, fibronectin- and collagen-
mediated adhesion, and quorum-sensing (QS) activity.
A. baumannii clinical isolates expressed extensive drug-resistant or pan-drug resistant
phenotypes, being nearly uniformly resistant to carbapenems. Almost 19% of isolates were resistant
to colistin as well. The isolates proved to be able to contaminate the antiseptic solutions and to
produce large quantities of biofilms. Nutritional composition of growth media significantly affected
the level of biofilm production. In contrast, wide range of different incubation temperatures and the
presence of antibiotics at subinhibitory concentrations had little effect, and the bacteria managed to
maintain high level of biofilm production. Moderate antimicrobial activity was displayed by
synthesized chalcones, among which methoxy-substituted derivatives achieved greatest growth
inhibition in average. Also, synergistic activity of chalcones and meropenem was present in several
cases, for which efflux pump inhibition was proposed as the potential mechanism. The chalcones
significantly inhibited motility and biofilm production, whereas methoxy-substituted derivative (o-
OCH3) also displayed significant antivirulence activity, by downregulating the ompA, bap, and abaI
gene expression and by inhibiting fibronectin- and collagen-mediated adhesion. It can be concluded
that o-OCH3 has been identified as a potent antivirulence agent against A. baumannii
Influence of newly-synthesized chalcone derivatives on growth, biofilm production, and virulence factors expression of multiresistant Acinetobacter baumannii strains
Acinetobacter baumannii je nozokomijalni, multirezistentni patogen, koga karakteriše sposobnost perzistencije na neživim površinama i mogućnost veoma brzog sticanja rezistencije na antibiotike. Danas su u svetu rasprostranjeni izrazito rezistentni sojevi protiv kojih u mnogim zdravstvenim ustanovama ne postoji efikasna terapija, a pronalazak alternativnih terapijskih pristupa je od izuzetne važnosti. Halkoni su jedinjenja sa potvrđenim antimikrobnim svojstvima i pokazanim različitim antivirulentnim aktivnostima. Ciljevi istraživanja ovog rada bili su određivanje profila rezistencije, ispitivanje mogućnosti kontaminacije antiseptika i ispitivanje produkcije biofilma identifikovanih kliničkih izolata A. baumannii, kao i sinteza derivata hidroksihalkona i ispitivanje njihovih antimikrobnih i antivirulentnih aktivnosti protiv ovih izolata. Osetljivost izolata na antibiotike ispitana je kombinacijom difuzionih, dilucionih i automatizovanih metoda, a identifikovani kolistin-rezistentni izolati dodatno su podvrgnuti sekvenciranju celog genoma (WGS) i genetički su okarakterisani. Takođe, mehanizmi rezistencije na kolistin ispitani su primenom komparativne analize genoma i Real-Time kvantitativne lančane reakcije polimeraze (RT-qPCR). Time-kill test je primenjen za ispitivanje perzistencije u antisepticima, a nivo produkcije biofilma ispitan je pod različitim uslovima kultivacije in vitro statičkom metodom uz bojenje safraninom. Derivati hidroksihalkona sintetisani su pomoću Claisen- Schmidt kondenzacije i njihove antimikrobne aktivnosti, samih i u kombinaciji sa antibioticima, ispitane su bujon-mikrodilucionom, Time-kill i Checkerboard analizom. Antivirulentne aktivnosti odabranih halkona procenjene su posredstvom uticaja na produkciju biofilma (monomikrobnog i polimikrobnog), vijabilnost biofilmskih ćelija, ekspresiju motiliteta, gensku ekspresiju faktora virulencije (OmpA, Bap i AbaI), adheziju A. baumannii na komponente ekstracelularnog matriksa (ECM), kao što su fibronektin i kolagen, i aktivnost sistema međućelijske komunikacije (Quorum- Sensing, QS). Klinički izolati A. baumannii gotovo uniformno bili su rezistentni na karbapeneme, a čak skoro 19% izolata bilo je rezistentno na kolistin, pripadajući tako ekstenzivno rezistentnom ili panrezistentnom fenotipu. Izolati su pokazali sposobnost kontaminacije antiseptika i produkcije velikih količina biofilma. Nutritivni sastav hranljivih medijuma značajno je uticao na nivo produkcije biofilma, dok se visok nivo produkcije održao pri širokom opsegu različitih temperatura inkubacije i u prisustvu subinhibitornih koncentracija antibiotika. Sintetisani halkoni ispoljili su umerenu antimikrobnu aktivnost, pri čemu su metoksi-supstituisani derivati u proseku najjače inhibirali rast. Takođe, zabeleženo je nekoliko sinergističkih interakcija halkona sa meropenemom, a inhibicija efluksnih pumpi predložena je kao potencijalni mehanizam. Halkoni su pokazali sposobnost značajne inhibicije motiliteta i produkcije biofilma, a metoksi-supstituisani derivat (o- OCH3) ispoljio je značajnu antivirulentnu aktivnost posredstvom nishodne regulacije ekspresije ompA, bap i abaI gena i inhibicije adhezije na komponente ECM. Na osnovu ovih rezultata, o- OCH3 halkon je identifikovan kao potentni antivirulentni agens protiv A. baumannii.Acinetobacter baumannii is a nosocomial, multiresistant pathogen, able to persist on abiotic surfaces and to rapidly acquire antibiotic resistance. Nowadays, highly resistant strains are widely disseminated throughout the world, and the discovery of alternative therapeutic strategies is of utter importance. Chalcones are compounds whose antimicrobial properties are well-known and for which different antivirulence activities have been demonstrated. The aims of this research were to determine resistance profiles, to evaluate the possibility of antiseptic contamination, and to analyze the biofilm production of identified A. baumannii clinical isolates, as well as to synthesize hydroxychalcone derivatives and to investigate their antimicrobial and antivirulence activities against these isolates. Antibiotic susceptibility of the isolates was tested by combination of diffusion, dilution, and automated methods, and additionally, identified colistin-resistant isolates were subjected to whole genome sequencing (WGS) and were genetically characterized. Also, colistin resistance mechanisms were explored by using comparative genome analysis and Real-Time quantitative polymerase chain reaction (RT-qPCR). Time-kill test was used for the measurement of bacterial survival in antiseptics, whereas the level of biofilm production under different cultivation conditions was quantified by in vitro static method using safranin stain. Hydroxychalcone derivatives were synthesized by Claisen-Schmidt condensation, and their antimicrobial activities, alone and in combination with antibiotics, were investigated using broth-microdilution, Time-kill, and Checkerboard analyses. Antivirulence activities of selected chalcones were evaluated based on the impact on biofilm production (monomicrobial and polymicrobial), biofilm cell viability, motility, virulence factors (OmpA, Bap, and AbaI) gene expression, fibronectin- and collagen- mediated adhesion, and quorum-sensing (QS) activity. A. baumannii clinical isolates expressed extensive drug-resistant or pan-drug resistant phenotypes, being nearly uniformly resistant to carbapenems. Almost 19% of isolates were resistant to colistin as well. The isolates proved to be able to contaminate the antiseptic solutions and to produce large quantities of biofilms. Nutritional composition of growth media significantly affected the level of biofilm production. In contrast, wide range of different incubation temperatures and the presence of antibiotics at subinhibitory concentrations had little effect, and the bacteria managed to maintain high level of biofilm production. Moderate antimicrobial activity was displayed by synthesized chalcones, among which methoxy-substituted derivatives achieved greatest growth inhibition in average. Also, synergistic activity of chalcones and meropenem was present in several cases, for which efflux pump inhibition was proposed as the potential mechanism. The chalcones significantly inhibited motility and biofilm production, whereas methoxy-substituted derivative (o- OCH3) also displayed significant antivirulence activity, by downregulating the ompA, bap, and abaI gene expression and by inhibiting fibronectin- and collagen-mediated adhesion. It can be concluded that o-OCH3 has been identified as a potent antivirulence agent against A. baumannii
Sastav i antimikrobna aktivnost etarskih ulja Salvia fruticosa i Salvia ringens (Lamiaceae)
Background/Aim. Plant essential oils (EOs) can have a significant antibacterial effect especially through additive or synergistic action as antibiotic adjuvants. We investi-gated the composition and activity of EOs of two species of genus Salvia (S) from Greece with the aim to deter-mine their antimicrobial activity as well as the activity in combination with selected antibiotics. Methods. The aerial parts of wild-growing S. fruticosa and S. ringens were subjected to a steam distillation and the obtained EOs were analyzed by gas chromatography and gas chroma-tography/mass spectrometry. The broth-microdilution method was used in order to determine the minimum in-hibitory concentrations (MICs) of EOs on seven strains of bacteria and one yeast. Antimicrobial activity of the combination of EO and antibiotics was investigated by checkerboard method and estimated by calculating frac-tional inhibitory concentration (FIC) of each component and fractional inhibitory concentration index (FICI). Re-sults. Dominant component of S. fruticosa EO was trans-thujone (54.2%) and for S. ringens EO it was α-pinene (28.1%). The MICs of EOs of both species were in the range from 200 μg/mL to > 500 μg/mL. The strongest antimicrobial effect was achieved against Bacillus subtilis and Candida albicans. According to FICI values, EO of S. fruticosa had additive effect with ciprofloxacin against most of bacterial strains but not with amikacin. Conclu-sion. The essential oils of S. ringens and S. fruticosa showed modest antimicrobial activity. However, the es-sential oil of S. fruticosa showed a promising additive ef-fect in combination with ciprofloxacin.Uvod/Cilj. Etarska ulja različitih biljaka mogu imati značajna antibakterijska svojstva, posebno kao adjuvanti antibiotika sa kojima ostvaruju aditivno ili sinergistično dejstvo. Ispitivali smo sastav i aktivnost etarskih ulja dve vrste roda Salvia (S) iz Grčke sa ciljem da odredimo njihovu antimkrobnu aktivnost, kao i dejstvo u kombi-naciji sa odabranim antibioticima. Metode. Nadzemni delovi samoniklih S. fruticosa i S. ringens su destilovani vodenom parom i dobijena etarska ulja su analizirana gasnom hromatografijom i gasnom hromatografijom sa masenom spektrometrijom. Radi određivanja minimalnih inhibitornih koncentracija (MICs) etarskog ulja na sedam sojeva bakterija i na jednoj patogenoj gljivici korišćena je mikrodiluciona metoda. Antimikrobna aktivnost kombi-nacije etarskog ulja i antibiotika ispitana je checkerboard metodom i procenjena je na osnovu frakcione inhibi-torne koncentracije (FIC) svake komponente i indeksa frakcione inhibitorne koncentracije (FICI). Resultati. Dominantna komponenta etarskog ulja S. fruticosa je bio trans-tujon (54,2%), a etarskog ulja S. ringens α-pinen (28,1%). MICs etarskog ulja obe vrste su bile u opsegu od 200 μg/mL do > 500 μg/mL. Najsnažnija anti-mikrobna aktivnost ostvarena je protiv Bacillus subtilis i Candida albicans. Na osnovu FICI vrednosti, etarsko ulje S. fruticosa je sa ciprofloksacinom, ali ne i sa amikacinom imalo aditivni efekat protiv većine bakterijskih sojeva. Zaključak. Etarska ulja S. ringens i S. fruticosa su pokazala skromnu antimikrobnu aktivnost, ali je etarsko ulje S. fru-ticosa u kombinaciji sa ciprofloksacinom ispoljilo značajan aditivni efekat
HERBAL PRODUCTS AS AN ALTERNATIVE TO ANTIBIOTICS: APPLICATION POSSIBILITIES AND LIMITATIONS
Antimicrobial resistance (AMR) has developed as
one of the top 10 global public health threats
facing humanity. As the nosocomial bacterial
strains are being increasingly resistant to most
clinically available antibiotics, there is a constant
need for exploration of new substances
that could kill them or inhibit their growth, or
alternatively inhibit some of their essential virulence
factors to counteract the lack of new antibacterials
and the rise of antibiotic resistance,
plants could represent a potential solution.
Plants produce a variety of bioactive secondary
metabolites that could be used to fuel the future
discovery pipeline. Aim of the present study was
to examine inhibitory activity of the supercritical
extract of J. communis L. green pseudofructus
(7SCO2) against the growth, biofilm production
and several virulence factors of significant nosocomial
bacterial pathogens. The extract was
obtained by fractional extraction with supercritical
CO2, and the qualitative and quantitative
analysis was performed using the GC-FID/MS
method. Clinical isolates of Pseudomonas aeruginosa,Acinetobacter baumannii, Staphylococcus
aureus (methicillin-sensitive-MSSA and methicillin-
resistant - MRSA), Enterococcus faecalis, and
Klebsiella pneumoniae, as well as their antibiotic
resistance profiles, were obtained from the Clinical
Hospital Centre “Dr Dragiša Mišović Dedinje”.
Minimum inhibitory concentrations (MICs) of
the 7SCO2 were determined by broth-microdilution
method. Examination of the anti-adhesive
effect of the extract was carried out using the
spectrophotometric method. The pyocyanin
production of Pseudomonas aeruginosa was determined
by the method described by Rampioni
et al. Most significant findings of this study
are potent antivirulence activity of the 7SCO2
against P. aeruginosa through the inhibition of
pyocyanin production. In addition, the biofilm
production of A. baumannii was inhibited by the
7SCO2 in concentration 50 μg/mL. Finally, notable
antivirulence activity of the 7SCO2 against
E. faecalis and S. aureus was detected, since it
significantly inhibited collagen and laminin adhesion
of these pathogens.Book of abstract: From biotechnology to human and planetary health XIII congress of microbiologists of Serbia with international participation Mikromed regio 5, ums series 24: 4th – 6th april 2024, Mona Plaza hotel, Belgrade, Serbi
Evaluation of novel compounds as anti-bacterial or anti-virulence agents
Antimicrobial resistance is a global threat, leading to an alarming increase in the prevalence of bacterial infections that can no longer be treated with available antibiotics. The World Health Organization estimates that by 2050 up to 10 million deaths per year could be associated with antimicrobial resistance, which would equal the annual number of cancer deaths worldwide. To overcome this emerging crisis, novel anti-bacterial compounds are urgently needed. There are two possible approaches in the fight against bacterial infections: a) targeting structures within bacterial cells, similar to existing antibiotics; and/or b) targeting virulence factors rather than bacterial growth. Here, for the first time, we provide a comprehensive overview of the key steps in the evaluation of potential new anti-bacterial and/or anti-virulence compounds. The methods described in this review include: a) in silico methods for the evaluation of novel compounds; b) anti-bacterial assays (MIC, MBC, Time-kill); b) anti-virulence assays (anti-biofilm, anti-quorum sensing, anti-adhesion); and c) evaluation of safety aspects (cytotoxicity assay and Ames test). Overall, we provide a detailed description of the methods that are an essential tool for chemists, computational chemists, microbiologists, and toxicologists in the evaluation of potential novel antimicrobial compounds. These methods are cost-effective and have high predictive value. They are widely used in preclinical studies to identify new molecular candidates, for further investigation in animal and human trials
АНТИОКСИДАТИВЕН КАПАЦИТЕТ И АНТИМИКРОБНИ ЕФЕКТИ НА КОМПЛЕКСИ НА ЦИНК СО ФЛАВОНОИДИ – ДАЛИ ПОСТОИ СИНЕРГИЗАМ?
The presence of residual cardiovascular disease (CVD) risk is a current dilemma in clinical practice; indeed, despite optimal management and treatment, a considerable proportion of patients still undergo major CV events. Novel lipoprotein biomarkers are suggested as possible targets for improving the outcomes of patients at higher risk for CVD, and their impact on major CV events and mortality have previously been investigated. Innovative antidiabetic therapies have recently shown a significant reduction in atherogenic lipoproteins, beyond their effects on glucose parameters; it has also been suggested that such anti-atherogenic effect may represent a valuable mechanistic explanation for the cardiovascular benefit of, at least, some of the novel antidiabetic agents, such as glucagon-like peptide-1 receptor agonists. This emphasizes the need for further research in the field in order to clearly assess the effects of innovative treatments on different novel biomarkers, including atherogenic lipoproteins, such as small dense low-density lipoprotein (LDL), lipoprotein(a) (Lp(a)) and dysfunctional high-density lipoprotein (HDL). The current article discusses the clinical importance of novel lipid biomarkers for better management of patients in order to overcome residual cardiovascular risk.Постои постојана потреба од ефикасни лекови, комбинации на лекови и методи за спречување на бактериски и вирусни инфекции, вклучувајќи го и коронавирусот. Познато е дека улогата на елементи во траги во зајакнување на човековиот имун систем е значајна. Витамините, микроелементите, вклучувајќи цинк, железо, селен, магнезиум и бакар, масните киселини омега-3 играат значајна физиолошка улога во унапредување на имуниот систем. Цинкот е неопходен микроелемент за основните ензимски физиолошки процеси. Тој игра важна улога во делбата на клетките и е вклучен во развојот на клетки одговорни за неспецифичен имунитет. Познато е дека недостигот на цинк ги предиспонира пациентите на вирусни инфекции поради намалениот антивирусен имунитет. Од друга страна, флавоноидите како метаболити на растенијата играат важна улога во спречување на оксидативен стрес. Овој труд има за цел да ја дискутира in vitro улогата на цинкот, флавонидите и нивните комплекси, како и нивните антиоксидативни и антимикорбни активности. Разгледана е и оправданоста за истовремена употреба на цинк и флавоноиди
Inhibicija Α‐amilaze in vitro i in vivo hipoglikemijski efekat metanolnog ekstrakta herbe Alchemilla viridiflora Rothm
Dry methanol extract (DER 3.95:1) of aerial flowering parts of Alchemilla viridiflora
Rothm., Rosaceae, obtained after successive extraction with cyclohexane and
dichloromethane, predominantly contains polyphenols, tannins (3.74%), mostly ellagic, and
flavonoids (0.30%), hexosides of quercetin and kaempferol. It shows ACE inhibitory and
anti-Helicobacter pylori activity in vitro (1). Based on the traditional use of the common A.
vulgaris in the treatment of diabetes (2), the effect of this A. viridiflora extract on α-amylase
activity in vitro and lowering blood glucose in rats were investigated. α-Amylase inhibitory
activity was assessed using the 3,5-dinitrosalicylic acid method. Extract in doses of 50, 100
and 200 mg/kg, p.o., was administered to streptozotocin-induced diabetic rats for 20 days,
and blood glucose level and weight of rats were monitored during treatment. Extract
inhibited α-amylase activity at IC 50 5.47 ± 0.30 mg/mL. A dose of 200 mg/kg significantly
decreased blood glucose level after 10 (32.2%) and 20 days of treatment (38.3%). The effect
of this dose was similar to the effect of the reference drug, glibenclamide. A dose of 100
mg/kg significantly increased glucose level by day 10 (50.5%) and a reduction in blood sugar
level of 51.2% was observed only after 10 days. Body weight was correlated with changes in
blood sugar levels. A dose of 50 mg/kg had no statistically significant effect. Results indicate
a significant potential of A. viridiflora aerial parts in the treatment of diabetes, which must be
confirmed by additional studies, and active ingredients need to be identified.Suvi metanolni ekstrakt (DER 3,95:1) nadzemnih delova u cvetu Alchemilla viridiflora
Rothm., Rosaceae, dobijen nakon sukcesivne ekstrakcije cikloheksanom i dihlormetanom,
sadrži visok nivo polifenola, tanina (3,74%), uglavnom elagnih, i flavonoida (0,30%),
heksozida kvercetina i kemferola. Pokazuje ACE inhibitornu i anti-Helicobacter pylori
aktivnost in vitro (1). Na osnovu podataka o tradicionalnoj upotrebi herbe A. vulgaris,
najčešće korišćene Alchemilla vrste, u lečenju dijabetesa (2), ispitivan je uticaj navedenog
ekstrakta A. viridiflora na aktivnost α-amilaze in vitro i na snižavanje nivoa glukoze u krvi
kod pacova. Uticaj na aktivnost α-amilaze procenjen je primenom metode sa 3,5-
dinitrosalicilnom kiselinom. Ekstrakt A. viridiflora u dozama od 50, 100 i 200 mg/kg, p.o.,
aplikovan je pacovima sa dijabetesom izazvanim streptozotocinom tokom 20 dana, a nivoi
glukoze u krvi i težina pacova su praćeni u određenim vremenskim intervalima tokom
trajanja tretmana. Ekstrakt je inhibirao aktivnost α-amilaze, a vrednost IC 50 iznosila je
5,47±0,30 mg/mL. Doza od 200 mg/kg značajno je smanjila nivo glukoze u krvi nakon 10
(32,2%) i 20 dana tretmana (38,3%). Efekat ove doze bio je sličan efektu referentnog leka,
glibenklamida. S druge strane, doza od 100 mg/kg značajno je povećala nivo glukoze do 10.
dana (50,5%) a do značajnog poboljšanja, tj. sniženja nivoa šećera od 51,2% dovela je tek
nakon 10 dana. Telesna težina bila je u korelaciji sa promenama nivoa šećera u krvi. Doza od
50 mg/kg nije imala statistički značajan efekat. Rezultati ukazuju na značajan potencijal
herbe A. viridiflora u lečenju dijabetesa, što mora biti potvrđeno dodatnim studijama, a
aktivni sastojci identifikovani.VIII Kongres farmaceuta Srbije sa međunarodnim učešćem, 12-15.10.2022. Beogra
Antimicrobial activity of Hyssopus officinalis L. subsp. aristatus (Godr.) Nyman (Lamiaceae) essential oils from Montenegro and Serbia
In this study, antimicrobial activity of essential oils extracted from the aerial flowering parts (herbs) of Hyssopus officinalis subsp. aristatus (Godr.) Nyman (Lamiaceae) collected from five different locations in Montenegro, or purchased in Serbia, were investigated. In addition, their antibacterial activity in combination with antibiotics was studied. The antimicrobial activity against selected standard bacterial and yeast strains was investigated using the broth microdilution method. Two standard antibiotics were used for comparison: the aminoglycoside antibiotic amikacin and the cephalosporin antibiotic ceftriaxone. The antimicrobial activity of the essential oil-amikacin combination was investigated using the checkerboard assay. The main components of the essential oils were 1,8-cineole, cis-pinocamphone, β-pinene and limonene in varying quantities. Most of the tested essential oils showed no significant antimicrobial activity. However, an essential oil rich in cis-pinocamphone showed moderate activity against both Staphylococcus aureus and Escherichia coli (MIC = 400 µg/mL). The overall effect of the essential oils and antibiotic combinations against E. coli or S. aureus ranged from additive (FICI = 0.625) to indifferent (FICI = 1.5), depending on the source of the essential oil
Comparative Study of the Antimicrobial Activity of Selenium Nanoparticles With Different Surface Chemistry and Structure
Although selenium nanoparticles (SeNPs) have gained attention in the scientific community mostly through investigation of their anticancer activity, a great potential of this nanomaterial was recognized recently regarding its antimicrobial activity. The particle form, size, and surface chemistry have been recognized as crucial parameters determining the interaction of nanomaterials with biological entities. Furthermore, considering a narrow boundary between beneficial and toxic effects for selenium per se, it is clear that investigations of biomedical applications of SeNPs are very demanding and must be done with great precautions. The goal of this work is to evaluate the effects of SeNPs surface chemistry and structure on antimicrobial activity against several common bacterial strains, including Staphylococcus aureus (ATCC 6538), Enterococcus faecalis (ATCC 29212), Bacillus subtilis (ATCC 6633), and Kocuria rhizophila (ATCC 9341), as well as Escherichia coli (ATCC 8739), Salmonella Abony (NCTC 6017), Klebsiella pneumoniae (NCIMB 9111) and Pseudomonas aeruginosa (ATCC 9027), and the standard yeast strain Candida albicans (ATCC 10231). Three types of SeNPs were synthesized by chemical reduction approach using different stabilizers and reducing agents: (i) bovine serum albumin (BSA) + ascorbic acid, (ii) chitosan + ascorbic acid, and (iii) with glucose. A thorough physicochemical characterization of the obtained SeNPs was performed to determine the effects of varying synthesis parameters on their morphology, size, structure, and surface chemistry. All SeNPs were amorphous, with spherical morphology and size in the range 70–300 nm. However, the SeNPs obtained under different synthesis conditions, i.e. by using different stabilizers as well as reducing agents, exhibited different antimicrobial activity as well as cytotoxicity which are crucial for their applications. In this paper, the antimicrobial screening of the selected systems is presented, which was determined by the broth microdilution method, and inhibitory influence on the production of monomicrobial and dual-species biofilm was evaluated. The potential mechanism of action of different systems is proposed. Additionally, the cytotoxicity of SeNPs was examined on the MRC-5 cell line, in the same concentration interval as for antimicrobial testing. It was shown that formulation SeNPs-BSA expressed a significantly lower cytotoxic effect than the other two formulations