Colistin resistance in acinetobacter baumannii: Molecular mechanisms and epidemiology

: Acinetobacter baumannii is recognized as a clinically significant pathogen causing a wide spectrum of nosocomial infections. Colistin was considered a last-resort antibiotic for the treatment of infections caused by multidrug-resistant A. baumannii. Since the reintroduction of colistin, a number of mechanisms of colistin resistance in A. baumannii have been reported, including complete loss of LPS by inactivation of the biosynthetic pathway, modifications of target LPS driven by the addition of phosphoethanolamine (PEtN) moieties to lipid A mediated by the chromosomal pmrCAB operon and eptA gene-encoded enzymes or plasmid-encoded mcr genes and efflux of colistin from the cell. In addition to resistance to colistin, widespread heteroresistance is another feature of A. baumannii that leads to colistin treatment failure. This review aims to present a critical assessment of relevant published (>50 experimental papers) up-to-date knowledge on the molecular mechanisms of colistin resistance in A. baumannii with a detailed review of implicated mutations and the global distribution of colistin-resistant strains

Biogenic silencers of Pseudomonas aeruginosa virulence

Pseudomonas aeruginosa jedan je od najznačajnijih uzročnika unutarbolničkih infekcija čiji je terapijski tretman konvencionalnim antibioticima sve češće neefikasan usled rezistencije na antibiotike. Inovativni vidovi kontrole infekcija, poput utišavanja međućelijske komunikacije bakterija, a time i onemogućavanja virulencije i inhibicije patogenog fenotipa su stoga od izuzetnog značaja. U ovom radu biće predstavljena istraživanja koja su bazirana na prirodnom svojstvu bakterija koje dele ekološke niše da sarađuju, ali i kompetiraju, na osnovu čega su analizirane Delftia tsuruhatensis i Burkholderia cepacia koje tokom infekcija kolokalizuju sa P. aeruginosa. Pokazano je da D. tsuruhatensis 11304 produkuje C18-HSL koji inhibira virulenciju P. aeruginosa i rekonstituiše osetljivost na antibiotike, a takođe je po prvi put u literaturi opisano prisustvo dihidroksi- C18-HSL u biološkim uzorcima. Opisane su i laktonaze vrste B. cepacia BCC4135 koje degraduju autoinducere komunikacije P. aeruginosa i inhibiraju ekspresiju faktora virulencije. Utvrđena je njihova supstratna specifičnost i ukazano na različitu biološku funkciju u zavisnosti od lokalizacije.Pseudomonas aeruginosa is a leading cause of nosocomial infections, whose therapeutic treatment with conventional antibiotics is increasingly ineffective due to antibiotic resistance. Inovative approaches of infection control, such as silencing the bacterial quorum sensing system and thus virulence and pathogenic phenotype inhibition are of great importance. In this study, there will be presented research based on natural feature of bacteria that share the same ecological niche to coordinate, but also to compete, based on which Delftia tsuruhatensis and Burkholderia cepacia that colocalize with P. aeruginosa during infections were analysed. D. tsuruhatensis 11304 has been shown to produce C18-HSL which inhibits P. aeruginosa virulence and reconstitutes antibiotic susceptibility, and the presence of dihydroxy-C18-HSL in biological samples has also been described for the first time in the literature. B. cepacia BCC4135 lactonases that degrade autoinducers of P. aeruginosa quorum sensing system and inhibit virulence factor expression have also been reported. Their substrate specificity was determined and different biological function depending on their localization was indicated.Jedan deo ovog rada realizovan je na Dipartimento di Scienze Chimiche, Università di Napoli Federico II, Napulj, Italija pod rukovodstvom prof dr Antonio Molinaro i dr Flaviana Di Lorenzo, kojima se ovom prilikom zahvaljujem. Hvala dr sci med Zorici Vasiljević sa Instituta za zdravstvenu zaštitu majke i deteta Srbije ‘’Dr Vukan Čupić‘’ koja je obezbedila kliničke izolate korišćene u ovom radu, kao i dr Milanu Kojiću i drugim saradnicima Laboratorije za molekularnu mikrobiologiju Instituta za molekularnu genetiku i genetičko inženjerstvo za izuzetnu pomoć tokom izrade eksperimenata

Molecular characterization of semi-hard homemade cheese microflora

U poslednjoj deceniji zabeležen je nagli razvoj molekularnih tehnika baziranih na 16S i 23S rRNK, koje se koriste u izučavanju biodiverziteta mikroorganizama. U ovom radu ispitivana je mikroflora polutvrdog sira pripremljenog u domaćinstvu. Zbog visokog sadržaja masti u ovom siru razvili smo novu tehniku za izolaciju totalne DNK iz sira (metod je baziran na bead beating-u). Brza izolacija DNK iz mikroflore sira omogućila nam je molekularnu identifikaciju BMK (BAKTERIJE MLEČNE KISELINE) na osnovu umnožavanja gena za 16S rRNK PCR metodom. Za umnožavanje su korišćeni prajmeri specifični za gene 16S rRNK lakto-bacila, ali su uslovi PCR reakcije bili takvi da su omogućavali i umnožavanje gena 16S rRNK laktokoka. Rezultati RFLP analize pokazali su da mikrofloru sira pripremljenog u domaćinstvu čine predominantno laktokoke.This decade has shown an impressive development in the application of molecular techniques based on 16S and 23S rRNA genes to study the microbial diversity in various ecosystems. Microflora of semi-hard homemade cheese was examined in this work. We developed a novel technique for DNA extraction (a bead beating based method) due to high fat content of this cheese. Rapid extraction of DNA from cheese microflora enabled a molecular identification of the LAB (Lactic Acid Bacteria) strains based on PCR amplification of 16S RNA coding sequences. The specific primers for 76S RNA gene of lactobacilli were used for amplification. The PCR reaction was performed at lower temperature, where the specificity of the annealing reaction was reduced and lactococcal sequences of 16S RNA genes were also amplified. The results of RFLP analysis revealed that the microflora of Doboj homemade cheese encompases mostly lactococci

ACINETOBACTER BAUMANNII RESISTANT TO LAST-LINE ANTIBIOTICS: AN EMERGING THREAT IN THE WESTERN BALKANS

Acinetobacter baumannii is considered one of the greatest threats to public health on a global scale. This Gram-negative pathogen causes severe infections, mostly of nosocomial origin, with a high mortality rate. In recent years, the rapid increase in the emergence and spread of antibiotic resistance in A. baumannii has significantly limited the effective therapeutic options against infections caused by this bacterium. The last-line antibiotics used in the treatment of multidrug-resistant (MDR) A. baumannii are carbapenems, tigecycline and polymyxins. However, resistance to these antibiotics is steadily increasing, especially to carbapenems, leading to an extensively drug-resistant (XDR) and even pandrug-resistant (PDR) phenotype of A. baumannii. In 2021, the European Centre for Disease Prevention and Control (ECDC) reported that resistance of Acinetobacter spp. to carbapenems reached 50% or more, mostly in Southern and Eastern European countries. Although the Western Balkans is a part of this region, detailed studies on the epidemiology and antibiotic resistance of A. baumannii are mainly limited to Serbia and Croatia. In most cases, carbapenem resistance in A. baumannii is due to the production of carbapenemases, in particular b-lactamases belonging to the class D known as oxacillinases. The studies from the Western Balkan countries revealed that besides the intrinsic blaOXA-51-like gene, the most prevalent acquired oxacillinase gene was the blaOXA- 24-like followed by the blaOXA-23-like, while the blaOXA-58-like and metallo- b-lactamase blaNDM-1 genes were less common. Although significantly lower compared to carbapenem-resistant, the number of A. baumannii isolates resistant to tigecycline and colistin is on a continual rise in the Western Balkans. As worldwide, the main mechanism conferring tigecycline resistance to A. baumannii from the Western Balkans was overexpression of efflux pumps. Also, the majority of reported alternations leading to colistin resistance in A. baumannii were found in the pmrCAB operon, which is responsible for the modification of the colistin target, LPS.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

Supplementary data for the article: Vatić, S.; Mirković, N.; Milošević, J. R.; Jovčić, B.; Polović, N. Đ. Broad Range of Substrate Specificities in Papain and Fig Latex Enzymes Preparations Improve Enumeration of Listeria Monocytogenes. International Journal of Food Microbiology 2020, 334, 108851. https://doi.org/10.1016/j.ijfoodmicro.2020.108851

Supplementary material for: [https://doi.org/10.1016/j.ijfoodmicro.2020.108851]Related to published version: [https://cherry.chem.bg.ac.rs/handle/123456789/4090

Effect of methionine and cysteine deprivation on growth of different natural isolates of Lactobacillus spp. in chemically defined media

Cilj ovog rada je bio da se utvrdi sposobnost prirodnih izolata laktobacila,izolovanih iz različitih ekoloških niša da rastu u hemijski definisanom medijumu sa ili bez prisustva aminokiselina koje sadrže sumpor, metionin i/ili cistein. Dobijeni rezultati su pokazali da je esencijalna aminokiselina za rast izolata L. paracasei subsp. paracasei BGHN14 i BGSJ2-8 cistein, dok je za izolate BGHN40, BGCG31, BGHV54T, koji pripadaju vrsti L. plantarum-metionin. Metionin je esencijalna aminokiselina za rast soja L. rhamnosus BGHV58T. Ostali analizirani sojevi,kao što su L. plantarum BGSJ3-18,BGZB19,BGHV52Ta i BGHV43T za svoj rast zahtevaju prisustvo obe aminokiseline u medijumu.The purpose of this study was to determine the ability of natural isolates of lactobacilli from different ecological niches to grow in a chemically defined medium in the presence or absence of sulphur-containing amino acids, methionine and/or cysteine. The obtained results indicate that cysteine is essential for growth of L. paracasei subsp. paracasei BGHN14 and BGSJ2-8, while methionine is essential for isolates BGHN40, BGCG31, and BGHV54T of the species L. plantarum. Methionine is also essential for growth of L. rhamnosus BGHV58T. Other analyzed strains, such as L. plantarum BGSJ3-18, BGZB19, BGHV52Ta, and BGHV43T, require the presence of both amino acids for their growth

Supplementary data for the article: Milošević, J.; Petrić, J.; Jovčić, B.; Janković, B.; Polović, N. Exploring the Potential of Infrared Spectroscopy in Qualitative and Quantitative Monitoring of Ovalbumin Amyloid Fibrillation. Spectrochimica Acta - Part A: Molecular and Biomolecular Spectroscopy 2020, 229. https://doi.org/10.1016/j.saa.2019.117882

Supplementary material for: [https://doi.org/10.1016/j.saa.2019.117882]Related to published version: [http://cherry.chem.bg.ac.rs/handle/123456789/3884]Related to accepted version: [http://cherry.chem.bg.ac.rs/handle/123456789/3895

Post-translational regulation of the RpoS and PsrA genes in pseudomonas putida WCS358: The role of ClpXP protease

RpoS i PsrA proteini su ključni transkripcioni regulatori koji se kod pseudomonada aktiviraju u odgovoru na stacionarnu fazu rasta. Cilj ove studije bio je utvrđivanje uloge ClpXP (ATP zavisna serin proteaza) u stabilnosti RpoS i PsrA proteina tokom različitih faza rasta u Pseudomonas putida WCS358. "Western blot" analiza proteinskih ekstrakata P. putida WCS358 i P. putida WCS358 clpX::Km iz rane eksponencijalne, Kasne eksponencijalne i stacionarne faze rasta, sa antitelima na RpoS i PsrA, pokazala je da ClpXP degraduje RpoS i PsrA u ranoj eksponencijalnoj fazi rasta.The RpoS and PsrA proteins are key transcriptional regulators that are activated in response to the stationary phase of growth in pseudomonads. This study was designed to establish whether ClpXP (ATP-dependent serine protease) regulates levels of RpoS and PsrA in Pseudomonas putida WCS358. Western blot analysis of P. putida WCS358 protein extracts from the early exponentianl, late exponential, and stationary phases of growth with antibodies against RpoS and PsrA revealed that these proteins are degraded by ClpXP in the early exponential phase of growth. The obtained results demonstrate a role for ClpXP protease in post-translational regulation of proteins encoded by the rpoS and psrA genes in Pseudomonas spp

Emergence of VIM-2 metallo-beta-lactamase-producing Pseudomonas aeruginosa isolates in a paediatric hospital in Serbia

Molecular detection and surveillance of the resistance genes harboured by Pseudomonas aeruginosa are becoming increasingly important in assessing and controlling spread and colonization in hospitals, and in guiding the antibiotic treatment of infections. Metallo-blactamase (MBL)-producing P. aeruginosa strains are slowly but steadily increasing within hospitals, causing outbreaks and/or hyperendemic situations in some centres, mostly in the Far East and the south of Europe (Queenan & Bush, 2007). The global dissemination of MBL-producing P. aeruginosa strains has also reached the Balkan region (Lepsanovic et al., 2008; Sardelic et al., 2003). The objective of our study was to detect and characterize P. aeruginosa isolates producing MBLs from the 400-bed paediatric tertiary care hospital Mother and Child Health Institute of Serbia ‘Dr Vukan Cupic