Colistin-resistant Acinetobacter Baumannii Research Articles

Cinnamic Acid Compounds (p-Coumaric, Ferulic, and p-Methoxycinnamic Acid) as Effective Antibacterial Agents Against Colistin-Resistant Acinetobacter baumannii.

Colistin-resistant Acinetobacter baumannii (COLR-Ab) is an opportunistic pathogen commonly associated with nosocomial infections, and it is difficult to treat with current antibiotics. Therefore, new antimicrobial agents need to be developed for treatment. Based on this information, we investigated the antimicrobial, antibiofilm, and combination activities of p-coumaric acid (p-CA), ferulic acid (FA), and p-methoxycinnamic acid (p-MCA) against five COLR-Ab isolates. p-CA, FA, and p-MCA exhibited antimicrobial activity against COLR-Ab isolates, with minimum inhibitory concentration (MIC) values in the range of 256-128 µg/mL, 1024-512 µg/mL, and 512-128 µg/mL, respectively. The combination effects of the compounds with colistin (COL) were evaluated using a checkerboard synergy test. The combinations exhibited a synergistic effect and caused a 128- to 16-fold decrease in COL MIC values. In addition, the biofilm production capacities of the COLR-Ab isolates and the antibiofilm activities of the compounds were determined using the microtitre plate-based crystal violet (CV) technique. The compounds showed effective antibiofilm activity against strong and moderate biofilm-producing isolates, inhibiting biofilm formation by 77.5% and 19.7%. Spectrometric measurements were used to examine the effect of compounds on membrane permeability; 1.9-, 1.66-, and 1.34-fold increases in absorbance values were observed at MIC concentrations of p-CA, FA, and p-MCA, respectively. Furthermore, morphological changes caused by the compounds in the isolate were observed using scanning electron microscopy (SEM) micrographs. According to the WST assay, the compounds did not show any statistically significant cytotoxic effect on the cells (p > 0.05). These findings indicate that p-CA, FA, and p-MCA may be potential new alternative candidates against resistant A. baumannii.

The Prevalence of Carbapenem and Colistin-Resistant Gram-Negative Bacteria Isolated from Hospital-Admitted Patients with Bacteremia

Carbapenem-resistance, a global health concern augmented with resistance against last-resort antibiotic colistin has become a great challenge. Bacteremia caused by Carbapenem-resistant (CR) Gram-negative bacteria (GNB) has been linked to prolonged hospitalization and high mortality. Colistin-resistant Acinetobacter baumannii (CoRAB) represent a serious threat to patients admitted to Intensive Care Unit (ICU) with mortality estimates up to 100%. This study aimed to estimate the current burden of CR and CoR GNB in admitted patients. In this prospective study, a total of 21,600 blood cultures were processed in automated BACTEC system. Conventional methods and Automated Profile Index (API) 20E and 20NE were used for GNB identification. Carbapenem and colistin Antimicrobial Susceptibility Testing (AST) was determined using disc-diffusion and broth microdilution (BMD) methods respectively. The pooled CR was 25.59%. CR was highest in Klebsiella spp. (44.2%) and Acinetobacter baumannii (34.66%). The overall CoR among 2903 GNB was 0.96% while among 743 CR-GNB was 3.76%. CoR rates are lower in Sindh compared to Punjab. CoR has reached up to 5.20% in Klebsiella spp. and up to 3.8% in A. baumannii. All Pseudomonas aeruginosa isolates were sensitive to colistin. Significant proportion of CoRKP and CoRAB in ICU alarms the situation and calls for to seek ways to minimize the emergence of CoR. Klebsiella spp. and A. baumannii remain the predominant CR and CoR GNB in Bloodstream infections (BSI). Presence of CoR-E. coli in pediatric wards highlight the poor hygienic practices and fecal transmission.

Outer membrane permeability of mcr-positive bacteria reveals potent synergy of colistin and macromolecular antibiotics against colistin-resistant Acinetobacter baumannii.

Colistin (CT) is the last-resort of antibiotic against multidrug-resistance (MDR) Acinetobacter baumannii (A. baumannii) infection. However, colistin resistance is increasingly reported in A. baumannii isolates partially due to the global emergence and dissemination of plasmid-borne mobile colistin resistance (mcr) gene and is a threat to human health. Thus, available treatment strategies urgently required in the fight against colistin-resistant A. baumannii. Here, we showed that mcr confers damaged outer membrane (OM) permeability in A. baumannii, which could compromise the viability of A. baumannii. Consistently, A. baumannii with colistin resistance exhibits increased susceptibility to macromolecular antibiotics such as rifampicin (RIF) and erythromycin (ERY). Moreover, the combination therapy of colistin and rifampicin demonstrates efficacy against colistin-resistant A. baumannii, regardless of the presence of mcr. Altogether, our data suggest that the synergy of colistin in combination with macromolecular hydrophobic antibiotics poses a promising therapeutic alternative for colistin-resistant A. baumannii.

(E, E)-farnesol and myristic acid-loaded lipid nanoparticles overcome colistin resistance in Acinetobacter baumannii

The rise of colistin-resistant Acinetobacter baumannii has severely limited treatment options for infections caused by this pathogen. While terpene alcohols and fatty acids have shown potential to enhance colistin’s efficacy, but their high lipophilicity limits their clinical application. To address this, we developed water-dispersible lipid nanoparticles (LNPs) in two sizes (40 nm and 130 nm), loaded with these compounds to act as colistin adjuvants. Among eleven LNP formulations, six significantly reduced colistin’s minimum inhibitory concentration (MIC) by 16- to 64-fold. The most effective, featuring (E,E)-farnesol and myristic acid, were further examined for bactericidal activity, membrane disruption, cytotoxicity, and in vivo efficacy in Galleria mellonella larvae. Time-kill studies demonstrated that at an adjuvant concentration of 60 mg/L, these LNPs eradicated bacteria when combined with 4 mg/L free colistin for resistant isolates (MIC = 128 mg/L) and 0.06 mg/L for susceptible isolates (MIC = 0.5 mg/L), without regrowth. Myristic acid-loaded LNPs combined with free colistin at 1/8 MIC resulted in a 4.2-fold higher mortality rate than the combination with (E,E)-farnesol-loaded LNPs in resistant strains. This result was correlated with a 45-fold faster increase in inner membrane permeability, measured by propidium iodide (PI) uptake, in the presence of myristic acid-loaded LNPs compared with a 13-fold faster increase with (E,E)-farnesol-loaded LNPs. DiSC3(5) assays revealed that LNPs alone depolarised the bacterial inner membrane, with enhanced effects when combined with colistin at 1/8 MIC, a result not observed with colistin alone at this concentration. As with PI uptake, this inner membrane depolarising effect was more pronounced with myristic acid-loaded LNPs than with (E,E)-farnesol-loaded LNPs in resistant strains, suggesting that the colistin adjuvant effect of these lipophilic compounds is due to their ability to help colistin destabilise the bacterial inner membrane. Cytotoxicity assays demonstrated no adverse effects on bone marrow macrophages after 6 h of exposure, although some toxicity was observed after 24 h. No mortality was observed in Galleria mellonella larvae over 7 days following three consecutive days of treatment with colistin and LNPs. Notably, the combination of (E,E)-farnesol-loaded LNPs and colistin significantly improved the survival of Galleria infected with A.baumannii. These results suggest that lipophilic-adjuvant-loaded LNPs may offer a promising strategy to enhance colistin efficacy and combat antibiotic-resistant A. baumannii infections.

Discovery of new antimicrobial thiophene derivatives with activity against drug-resistant Gram negative-bacteria.

Our aim is to identify new small molecules with antimicrobial potential, especially against colistin-resistant (Col-R) Acinetobacter baumannii and Escherichia coli. After initial hits identification by fingerprint similarity, MIC of 24 heterocyclic derivatives for A. baumannii and E. coli reference strains, and bactericidal activity of selected thiophenes against Col-R strains were determined. We analyzed changes in bacterial membrane permeability and the OMPs profile. Additionally, we determined bacterial adherence to host cells and performed molecular docking studies to assess their binding to bacterial targets. The compounds' MICs ranged from 4 to >64mg/L. Thiophene derivatives 4, 5 and 8 exhibited MIC50 values between 16 and 32mg/L for Col-R A. baumannii and 8 and 32mg/L for Col-R E. coli. The time-kill curve assay demonstrated that thiophenes 4 and 8 had bactericidal effects against Col-R A. baumannii and E. coli. Furthermore, treatment with them resulted in increased membrane permeabilization and reduced adherence of these isolates to host cells. Finally, the docking studies showed a stronger binding affinity to CarO1 and Omp33 of A. baumannii and OmpW and OmpC of E. coli. These findings indicate that thiophene derivatives possess antibacterial activity against Col-R A. baumannii and E. coli, suggesting that they may enhance the repertoire of drug treatments against bacteria.

Isolation, Identification, and Antibiogram of Colistin-resistant Acinetobacter baumannii from Rivers in and around Kathmandu Valley

Background: Acinetobacter baumannii, an opportunistic Gram-negative pathogen, poses an escalating threat in clinical settings due to the rise of multidrug-resistant infections. Despite its clinical significance, there exists a considerable gap in understanding its environmental dissemination.
 Aims and Objectives: The primary objective is to examine the distribution of A. baumannii and its antibiotic resistance in river ecosystems. Specifically, we aim to identify strains resistant to Colistin, a last-resort antibiotic, and elucidate the susceptibility patterns to other antibiotics.
 Materials and Methods: Water samples from 10 rivers were collected and subjected to analysis using Leeds Acinetobacter Agar Base and a series of biochemical tests. Antibiotic susceptibility testing, focusing on Colistin resistance, was performed using standard procedures.
 Results: Out of the 284 isolated strains, 14 (4.9%) exhibited resistance to Colistin, while demonstrating varying susceptibility patterns to other antibiotics. Notably, Gentamycin showed effectiveness against resistant strains (14.28%), while Ceftazidime resistance was complete. Colistin-sensitive strains displayed high susceptibility to Ciprofloxacin (84.44%) and lower susceptibility to Chloramphenicol (53.33%). Carbapenem susceptibility was observed across all isolates.
 Conclusion: The study underscores a concerning environmental presence of multidrug-resistant A. baumannii in rivers around Kathmandu Valley, with Sundarijal being the exception. The findings emphasize the necessity of scrutinizing environmental reservoirs for pathogen spread, advocating for heightened awareness of potential health implications beyond clinical settings. Urgent attention is needed to comprehend and counteract the emergence and dissemination of antibiotic resistance, necessitating comprehensive strategies and continued surveillance

Molecular mechanisms of colistin resistance mediated by pmrCAB genes in Acinetobacter baumannii isolated from patients hospitalized in Isfahan medical centers

Abstract Introduction In recent years, colistin-resistant Acinetobacter baumannii (A. baumannii) has been found all over the world. In this current study, the main purpose was to examine the occurrence of extensively drug resistant (XDR), resistance to colistin and characterization and mutations in pmrCAB genes among A. baumannii obtained from inpatients. Materials and Methods A total of 108 clinical isolates of A. baumannii were collected from several hospitals located in Isfahan, Iran. The Kirby-Bauer assay was performed to assess the antimicrobial resistance. The Phoenix automated system was utilized to determine the minimum inhibitory concentration (MIC) of colistin for each of the bacterial isolates. Polymerase chain reaction was used to screen for pmrCAB genes that mediate colistin resistance, and sequencing was used to determine the amplicon’s nucleotide sequence. Results The results revealed that all A. baumannii isolates (100%) were resistant to piperacillin tazobactam, meropenem and ciprofloxacin. All isolates were classified as XDR, with seven isolates being pan-drug resistant (PDR). Colistin resistance (CoR) was found in 6.48% (7/108) of studied isolates, all of which were positive for pmrCAB genes. The sequencing results showed a substitution in pmrB and two isolates showed a substitution in pmrC. Conclusions In conclusion, this study is the initial report of the existence and mutations of pmrB and C genes in the clinical isolate of A. baumannii in our region. This outcome highlights the necessity to explore additional mutations in the PMR operon of A. baumannii in forthcoming studies. Moreover, our results highlight the high occurrence of XDR-A. baumannii strain in Isfahan, Iran.

The efficacy of mesenchymal stem cell treatment and colistin-fosfomycin combination on colistin-resistant Acinetobacter baumannii sepsis model.

This study examines the role of mesenchymal stem cells (MSCs) in an experimental sepsis model developed with colistin-resistant Acinetobacter baumannii (CRAB). BALB-c mice were divided into treatment groups (MSC, MSC + colistin (C)-fosfomycin (F), and C-F and control groups (positive and negative)). CRAB was administered to mice through intraperitoneal injection. Three hours later, C, F, and MSC were given intraperitoneally to the treatment groups. Colistin administration was repeated every 12 h, F administration was done every 4 h, and the second dose of MSC was administered after 48 h. Mice were sacrificed at 24 and 72 h. The bacterial load was determined as colony-forming units per gram (cfu/g). Histopathological examination was conducted on the left lung, liver, and both kidneys. IL-6 and C-reactive protein (CRP) levels in mouse sera were determined by enzyme-linked immunosorbent assay. Among the treatment groups, the C-F group had the lowest colony count in the lung (1.24 ± 1.66 cfu/g) and liver (1.03 ± 1.08 cfu/g). The highest bacterial clearance was observed at 72 h compared to 24 h in the MSC-treated groups (p = 0.008). The MSC + C-F group showed the lowest histopathological score in the liver and kidney (p = 0.009). In the negative control group, the IL-6 level at the 24th hour was the lowest (p < 0.001). Among the treatment groups, the CRP level was the lowest in the MSC + C-F group at 24 and 72 h. In a CRAB sepsis model, adding MSCs to a colistin-fosfomycin treatment may be beneficial in terms of reducing bacterial loads and preventing histopathological damage.

Emergence of colistin resistant Acinetobacter baumannii clonal complex 2 (CC2) among hospitalized patients in Iran.

Acinetobacter baumannii is a major causative agent of serious nosocomial infections. This study was carried out to investigate the molecular characterization of colistin resistant isolates of A. baumannii from hospitalized patients, based on multilocus sequence typing (MLST). A cross-sectional study was conducted to collect A. baumannii from clinical samples in Isfahan from 2021 to 2022. Isolates were identified as A. baumannii using biochemical tests and PCR of blaOXA-51. Antibiotic susceptibility testing was carried out using the Kirby-Bauer method and minimum inhibitory concentration (MIC) values were determined for colistin. Additionally, MLST was performed according to the Pasteur scheme to assess the relationship between colistin resistant A. baumannii. A total of 70 non-repetitive A.baumannii isolates were obtained from different clinical samples. MIC results showed that seven A.baumannii isolates were resistant to colistin. The antibiotic susceptibility pattern revealed that all seven colistin resistant strains were resistant to all tested antibiotics. Based on MLST analysis, the colistin resistant isolates were assigned to five unique STs namely, ST2 (3; 42.9%) followed by ST78 (1;14.3%), ST1077 (1; 14.3%), ST415 (1; 14.3%) and ST391 (1; 14.3%). Among them ST2, ST391 and ST415 belong to clonal complex 2. Colistin resistant A. baumannii ST2 is the main circulating clone in clinical settings in Iran, but additionally ST415, ST391, and ST1077 are found for the first time in our country. Intensive control procedures and strict adherence to surveillance programs are recommended to decrease the spread of carbapenem and colistin resistant A. baumannii strain.

Evaluation of potential synergistic activity of antimicrobial combinations against Colistin resistant Acinetobacter baumannii

Evaluation of potential synergistic activity of antimicrobial combinations against Colistin resistant Acinetobacter baumannii

Risk factors associated with colistin-resistant Acinetobacter baumannii infection

Acinetobacter baumannii is an important cause of healthcare-associated infections and is resistant to almost all antimicrobial agents, with strains recently reported to be resistant to colsitin. In this study, we aimed to identify the risk factors associated with colistin-resistant A. baumannii infections by comparing colistin-resistant and -susceptible A. baumannii isolates. We retrospectively reviewed the medical records of 51 and 100 cases in which colistinresistant and -susceptible A. baumannii were isolated, respectively. Univariate analysis showed that compared with patients with colistin-sensitive infections, patients with colistinresistant A. baumanni infections had a combined pulmonary disease (P = 0.017), were admitted to intensive care unit (P = 0.020), and had prior mechanical ventilation (P = 0.003), tracheostomy (P = 0.043), percutaneous drainage (P = 0.070), hemodialysis (P = 0.002); use of colistin (P = 0.000), carbapenem (P = 0.000), and teicoplanin (P = 0.004); and co-infection (P = 0.035). Multivariate analysis indicated that eight variables were related to the likelihood of colistin-resistant A. baumanni infections: use of teicoplanin (Odds ratio [OR]: 3.140, 95% confidence interval [CI]: 0.529–18.650), prior hemodialysis (OR: 2.722, 95% CI: 0.851–8.709), combined pulmonary disease (OR: 2.286, 95% CI: 0.998–5.283), prior use of carbapenem (OR: 0.199, 95% CI: 0.863–5.603), co-infection (OR: 1.706, 95% CI: 0.746–3.898), prior mechanical ventilation (OR: 1.614, 95% CI, 0.684–3.809), intensive care unit admission (OR: 1.387, 95% CI: 0.560–3.435), and prior tracheostomy (OR: 1.102, 95% CI: 0.344–3.527); however, no statistical differences were observed. Although colistin use could not be proven in multivariate analysis, the possibility of being a risk factor cannot be ruled out.

Quercetin: Synergistic Interaction with Antibiotics against Colistin-Resistant Acinetobacter baumannii.

Infections caused by resistant strains of Acinetobacter baumannii are now a global problem that requires the immediate development of new antimicrobial drugs. Combination therapy is one of the strategies used to solve this problem. Based on this information, the purpose of this study was to determine whether quercetin (QUE), in combination with three antibiotics, is effective against colistin-resistant A. baumannii strains (ColR-Ab). The effects of the combination of QUE with colistin (COL), amikacin (AMK), and meropenem (MEM) were evaluated according to the checkerboard synergy test. The combinations of QUE + COL and QUE + AMK showed synergistic activity on ColR-Ab strains with FICI values in the range of 0.1875-0.5 and 0.1875-0.2825, respectively. A 4- to 16-fold decrease in COL MIC and a 16- to 64-fold decrease in AMK MIC values were detected. Synergistic activity was confirmed by the time-kill test, and these combinations were found to be bactericidal at the end of 24 h. According to spectrophotometric measurements, the combinations of QUE + COL and QUE + AMK induced membrane damage, leading to the leakage of nucleic acids. Cell lysis and cell death were confirmed with SEM observations. The detected synergy offers an opportunity for the future development of treatment strategies for potential infections caused by ColR-Ab strains.

In Vitro Synergistic Activity of Antimicrobial Combinations against Carbapenem- and Colistin-Resistant Acinetobacter baumannii and Klebsiella pneumoniae.

Polymyxins are commonly used as the last resort for the treatment of MDR Acinetobacter baumannii and Klebsiella pneumoniae nosocomial infections; however, apart from the already known toxicity issues, resistance to these agents is emerging. In the present study, we assessed the in vitro synergistic activity of antimicrobial combinations against carbapenem-resistant and colistin-resistant A. baumannii and K. pneumoniae in an effort to provide more options for their treatment. Two hundred A. baumannii and one hundred and six K. pneumoniae single clinical isolates with resistance to carbapenems and colistin, recovered between 1 January 2021 and 31 July 2022,were included. A. baumannii were tested by the MIC test strip fixed-ratio method for combinations of colistin with either meropenem or rifampicin or daptomycin. K. pneumoniae were tested for the combinations of colistin with meropenem and ceftazidime/avibactam with aztreonam. Synergy was observed at: 98.99% for colistin and meropenem against A. baumannii; 91.52% for colistin and rifampicin; and 100% for colistin and daptomycin. Synergy was also observed at: 73.56% for colistin and meropenem against K. pneumoniae and; and 93% for ceftazidime/avibactam with aztreonam. The tested antimicrobial combinations presented high synergy rates, rendering them valuable options against A. baumannii and K. pneumoniae infections.

Comparative genomics and molecular epidemiology of colistin-resistant Acinetobacter baumannii

This study aimed to investigate the prevalence and resistance mechanisms of colistin-resistant Acinetobacter baumannii (ColRAB) isolates in Serbia, assess their genetic relatedness to other circulating A. baumannii isolates in the neighbouring European countries, and analyse the global genomic epidemiology of ColRAB isolates. A total of 784 isolates of A. baumannii were recovered from hospitalised patients in Serbia between 2018 and 2021. The antimicrobial susceptibility testing was performed using disk diffusion and broth microdilution. All ColRAB isolates were subjected to DNA isolation and whole-genome sequencing (WGS). Overall, 3.94 % (n = 30) isolates were confirmed as ColRAB. Results of mutational and transcriptional analysis of genes associated with colistin resistance indicate the central role of the two-component regulating system, PmrAB, and increased expression of the pmrC gene in ColRAB. Most of the isolates (n = 29, 96.6 %) belonged to international clone II, with the most common sequence type being STPas2 (n = 23, 76.6 %). Based on the WGS analysis, ColRAB isolates belonging to the same ST isolated in various countries were grouped into the same clusters, indicating the global dissemination of several high-risk clonal lineages. Phylogenomic analysis of ColRAB isolates, together with all previously published A. baumannii genomes from South-Eastern European countries, showed that colistin resistance arose independently in several clonal lineages. Comparative genomic analysis revealed multiple genes with various roles (transcriptional regulation, transmembrane transport, outer membrane assembly, etc.), which might be associated with colistin resistance in A. baumannii. The obtained findings serve as the basis for further studies, contributing to a better understanding of colistin resistance mechanisms in A. baumannii.

C-Locked Analogs of the Antimicrobial Peptide BP214.

BP214 is an all-D antimicrobial peptide amide, kklfkkilryl, which shows an excellent activity against colistin-resistant Acinetobacter baumannii and a low hemolytic activity. The aim of the present work was to investigate how C-terminus-to-side chain macrocyclization and fatty acid modification affect the antimicrobial and hemolytic activity of this peptide. In total, 18 analogs of BP214 were synthesized using a combination of Fmoc-based solid-phase peptide synthesis and the submonomer approach. Cyclization was achieved by reacting the ε-amino group of a C-terminal lysine residue with a bromoacetylgroup attached to the Nα amino group of the N-terminal amino acid, generating a secondary amine at which the exocyclic lipopeptide tail was assembled. Three different ring sizes (i.e., 3–5 amino acid residues) of C-locked analogs combined with fatty acids of different lengths (i.e., C10–C14) were investigated. The antimicrobial activity of the analogs was tested against Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae, Acinetobacter baumannii, and Pseudomonas aeruginosa. The most promising compound was analog 13 (MIC = 4 µg/mL (2.4 µM) against E. coli and 36% hemolysis of red blood cells at 150 µM). In a time-kill assay, this peptide showed a significant, concentration-dependent reduction in viable E. coli cells comparable to that seen for colistin.

Synergistic activities of colistin combined with other antimicrobial agents against colistin-resistant Acinetobacter baumannii clinical isolates.

Emerging resistance to colistin in Acinetobacter baumannii clinical strains is concerning because of the limited therapeutic choices for these important clinical pathogens. We studied the in vitro activities of different colistin-based antimicrobial agent combinations against colistin-resistant Acinetobacter baumannii. Fourteen clinical isolates of colistin-resistant Acinetobacter baumannii were obtained between 2015 and 2016. To identify colistin-based combinations with synergistic activities, multiple two antimicrobial combinations based on 8 commercially available drugs were evaluated by the checkerboard method. The most effective colistin-based combinations were vancomycin, aztreonam, ceftazidime and imipenem which showed synergistic activities against all examined strains. Colistin-rifampin showed synergy against four strains. Colistin-tigecycline and colistin-amikacin mostly showed indifferent results. By using the checkerboard tests, we were able to find the most promising colistin-based combinations that may provide more therapeutic options against colistin-resistant Acinetobacter baumannii.

Overcoming addition of phosphoethanolamine to lipid A mediated colistin resistance in Acinetobacter baumannii clinical isolates with colistin–sulbactam combination therapy

Overcoming colistin-resistant Acinetobacter baumannii (CoR-AB) has become a major concern due to the lack of effective antibiotics. This study aimed to explore the prevalence of CoR-AB clinical isolates in Thailand, their mechanisms of resistance, and test the efficacy of colistin plus sulbactam against CoR-AB isolates. The colistin resistance rate among carbapenem-resistant A. baumannii was 15.14%. The mcr gene or its variants were not detected in CoR-AB isolates by PCR screening. The lipid A mass spectra of CoR-AB isolates showed the additional [M–H]− ion peak at m/z = 2034 that correlated to the phosphoethanolamine (pEtN) addition to lipid A (N = 27/30). The important amino acid substitutions were found at position S14P, A138T, A227V in PmrB that are associated with overexpression of the pEtN transferase (PmrC) and contributed the pEtN addition. The lipopolysacccharide production genes (lpxACD) were not related to lipid A mass spectra. A colistin plus sulbactam combination exhibited the synergy rate at 86.7% against CoR-AB isolates compare to sulbactam (85.89% resistance) or colistin (15.14% resistance) alone. The excellent synergistic activity of colistin plus sulbactam combination has the potential for the treatment of CoR-AB infections.

MICROBIOLOGICAL ISOLATES AND IT’S RESISTOTYPE FROM CLINIC OF VASCULAR SURGERY FOR THE FIRST QUATER OF 2022

One of the most common complications of surgical exposures is the surgical site infection (SSI). Although it varies between different surgical profiles, it can reach up to one third of all complications. In vascular surgery patients ischemic ulcers are very common, as well as factors, compromising the immune system such as diabetes, chronic kidney disease etc. One of the main surgical exposures in vascular surgery is inguinal, providing access to the femoral artery and its bifurcation. Although it allows a wide range of reconstructions, implanting different types of prosthetic materials, stents and providing anastomosis site, it contains lymph nodes, which can contaminate the reconstruction and cause SSI with severe consequences. Patients, prone to SSI due to concomitant diseases, are threatened by sepsis, limb loss and even death, which makes prevention of those type of complications essential. Aim: To investigate etiological spectrum of microbiological isolates and their resistance against most common antimicrobials among vascular surgery patients. Materials and methods: The study is retrospective, conducted in the period 01 January 2022 – 31 March 2022. All of the samples were obtained from patients of Clinic of Vascular Surgery. After isolation of pure culture from the samples, the strains were identified by MALDI TOF MS and Vitec – 2 Compact. Antibiotic resistance was determined with Bauer-Kirby disk diffusion method. Results: From all 419 of the patients, hospitalized in the Clinic of Vascular surgery for this period, 28 isolates from 26 (6,21%) patients were obtained, of which Gram-negative were 19 (67,86%) and Gram-positive - 9 (32,14%). From Gram-negative - enterobacteria – 14 (73,68%), and non-fermenting gram-negative bacteria (NFGNB) were 5 (26,32%). Only 3 (21,43%) from all enterobacteria were extended spectrum beta-lactamases producing strains (ESBLs). No strains, resistant to carbapenems (RCP) were isolated. Five (55,55%) of the Gram-positive isolates were Staphylococcus aureus, 4 (80%) of which were methicillin resistant Staphylococcus aureus (MRSA). Two of the Gram-positive species isolated were Enterococcus faecalis, of which 1 with a high-level aminoglycoside resistance (HLAR). No Vancomycin resistant enterococci (VRE) were discovered. There were no colistin-resistant Acinetobacter baumannii and Pseudomonas aeruginosa strains. Conclusion: From all 28 isolates 8 (28,57%) were with acquired types of antimicrobial resistance. With almost one third of the isolates that are problematic in terms of antibiotic susceptibility, treatment of those patients can be challenging. Prevention of in hospital contamination with polyresistant strains, associated with medical care it is crucial for reducing the number of severe complications, decreasing of hospital stay and cost for treatment.

Whole-Genome Sequencing of a Colistin-Resistant Acinetobacter baumannii Strain Isolated at a Tertiary Health Facility in Pretoria, South Africa.

Background: Acinetobacter baumannii’s (A. baumannii) growing resistance to all available antibiotics is of concern. The study describes a colistin-resistant A. baumannii isolated at a clinical facility from a tracheal aspirate sample. Furthermore, it determines the isolates’ niche establishment ability within the tertiary health facility. Methods: An antimicrobial susceptibility test, conventional PCR, quantitative real-time PCR, phenotypic evaluation of the efflux pump, and whole-genome sequencing and analysis were performed on the isolate. Results: The antimicrobial susceptibility pattern revealed a resistance to piperacillin/tazobactam, ceftazidime, cefepime, cefotaxime/ceftriaxone, imipenem, meropenem, gentamycin, ciprofloxacin, trimethoprim/sulfamethoxazole, tigecycline, and colistin. A broth microdilution test confirmed the colistin resistance. Conventional PCR and quantitative real-time PCR investigations revealed the presence of adeB, adeR, and adeS, while mcr-1 was not detected. A MIC of 0.38 µg/mL and 0.25 µg/mL was recorded before and after exposure to an AdeABC efflux pump inhibitor. The whole-genome sequence analysis of antimicrobial resistance-associated genes detected beta-lactam: blaOXA-66; blaOXA-23; blaADC-25; blaADC-73; blaA1; blaA2, and blaMBL; aminoglycoside: aph(6)-Id; aph(3″)-Ib; ant(3″)-IIa and armA) and a colistin resistance-associated gene lpsB. The whole-genome sequence virulence analysis revealed a biofilm formation system and cell–cell adhesion-associated genes: bap, bfmR, bfmS, csuA, csuA/B, csuB, csuC, csuD, csuE, pgaA, pgaB, pgaC, and pgaD; and quorum sensing-associated genes: abaI and abaR and iron acquisition system associated genes: barA, barB, basA, basB, basC, basD, basF, basG, basH, basI, basJ, bauA, bauB, bauC, bauD, bauE, bauF, and entE. A sequence type classification based on the Pasteur scheme revealed that the isolate belongs to sequence type ST2. Conclusions: The mosaic of the virulence factors coupled with the resistance-associated genes and the phenotypic resistance profile highlights the risk that this strain is at this South African tertiary health facility.

Whole-genome sequencing for the characterization of resistance mechanisms and epidemiology of colistin-resistant Acinetobacter baumannii.

BackgroundMultidrug-resistant Acinetobacter baumannii is an important causal pathogen of healthcare-associated infections, and colistin-resistant strains have recently emerged owing to the increased use of colistin. Using next-generation sequencing (NGS), a single whole-genome sequencing (WGS) protocol can identify and type pathogens, analyze genetic relationships among different pathogens, predict pathogenic transmissions, and detect antibiotic resistance genes. However, only a few studies have applied NGS in studying the resistance mechanism and epidemiology of colistin-resistant A. baumannii. This study aimed to elucidate the resistance mechanism of colistin-resistant A. baumannii and analyze its molecular epidemiology through WGS.Materials and methodsThe subjects in this study were patients who visited a university hospital between 2014 and 2018. Thirty colistin-resistant strains with high minimum inhibitory concentrations were selected from various patient samples, and WGS was performed. Comparative genomic analysis was performed for the 27 colistin-resistant A. baumannii strains using a colistin-susceptible strain as the reference genome.ResultsThe WGS analysis found no mutation for lpxA, lpxC, lpx D, pmrA, pmrB, and mcr1, the genes known to be associated with colistin resistance. Fifty-seven coding sequences (CDS) showed differences; they included 13 CDS with known names and functions that contained 21 genes. From the whole-genome multi-locus sequence typing (wgMLST) and single nucleotide polymorphism (SNP) analyses, two major clusters were found for the colistin-resistant A. baumannii strains. However, no differences were observed by the time of detection for each cluster, the samples, the pattern of antibiotic resistance, or the patient characteristics. In the conventional MLST following the Oxford scheme, the typing result showed ST1809, ST451, ST191, ST1837, and ST369 in the global clone 2 (GC2), without any relation with the results of wgMLST and SNP analyses.ConclusionBased on the findings of the resistance gene analysis through WGS and comparative genomic analysis, the potential genes associated with colistin-resistance or CDS were examined. Furthermore, the analysis of molecular epidemiology through WGS regarding colistin-resistant A. baumannii may prove helpful in preventing infection by multidrug-resistant bacteria and controlling healthcare-associated infections.