Acinetobacter baumannii as an oro-dental pathogen: a red alert!!

OBJECTIVES: This review highlights the existence and association of Acinetobacter baumannii with the oro-dental diseases, transforming this systemic pathogen into an oral pathogen. The review also hypothesizes possible reasons for the categorization of this pathogen as code blue due to its stealthy entry into the oral cavity. METHODOLOGY: Study data were retrieved from various search engines reporting specifically on the association of A. baumannii in dental diseases and tray set-ups. Articles were also examined regarding obtained outcomes on A. baumannii biofilm formation, iron acquisitions, magnitude of antimicrobial resistance, and its role in the oral cancers. RESULTS: A. baumannii is associated with the oro-dental diseases and various virulence factors attribute for the establishment and progression of oro-mucosal infections. Its presence in the oral cavity is frequent in oral microbiomes, conditions of impaired host immunity, age related illnesses, and hospitalized individuals. Many sources also contribute for its prevalence in the dental health care environment and the presence of drug resistant traits is also observed. Its association with oral cancers and oral squamous cell carcinoma is also evident. CONCLUSIONS: The review calls for awareness on the emergence of A. baumannii in dental clinics and for the need for educational programs to monitor and control the sudden outbreaks of such virulent and resistant traits in the dental health care settings.

Detection of multidrug-resistant bacteria in the nasal cavities and evaluation of sinus disorders in patients undergoing Le Fort I osteotomy.

INTRODUCTION: Orthognathic surgery can lead to sinus alterations, including sinusitis, attributed to the exposure of maxillary sinuses during Le Fort I osteotomy. Furthermore, being a hospital-based procedure, there is potential risk of complications arising from bacteria prevalent in such environments. This study evaluated maxillary sinusitis occurrence and the presence of multidrug-resistant bacteria in the nasal cavity before and after orthognathic surgery. METHODS: Ten patients with dentofacial deformities underwent Le Fort I osteotomy. Clinical evaluations using SNOT-22 questionnaire were performed, and nasal cavity samples were collected pre-surgery and 3-6 months post-surgery to quantify total mesophilic bacteria and detect Staphylococcus aureus, Acinetobacter baumannii, and Klebsiella pneumoniae. Cone Beam Computed Tomography (CBCT) was performed pre- and post-operatively, and the results were evaluated using the Lund-Mackay system. This study was registered and approved by the Research Ethics Committee of PUCRS (No. 4.683.066). RESULTS: The evaluation of SNOT-22 revealed that five patients showed an improvement in symptoms, while two remained in the same range of interpretation. One patient developed post-operative maxillary sinusitis, which was not detected at the time of evaluation by SNOT-22 or CBCT. CBCT showed a worsening sinus condition in three patients, two of whom had a significant increase in total bacteria count in their nasal cavities. The Brodsky scale was used to assess hypertrophy in palatine tonsils, where 60% of the subjects had grade 1 tonsils, 20% had grade 2 and 20% had grade 3. None of the patients had grade 4 tonsils, which would indicate more than 75% obstruction. Two patients harboured S. aureus and K. pneumoniae in their nasal cavities. Notably, K. pneumoniae, which was multidrug-resistant, was present in the nasal cavity of patients even before surgery, but this did not result in maxillary sinusitis, likely due to the patients' young and healthy condition. CONCLUSION: There was an improvement in signs and symptoms of maxillary sinusitis and quality of life in most patients after orthognathic surgery. However, some patients may still harbour multidrug-resistant bacteria, even if they are asymptomatic. Therefore, a thorough pre-operative assessment is essential to avoid difficult-to-treat post-operative complications.

Risk factors for bacteremic pneumonia and mortality (28-day mortality) in patients with Acinetobacter baumannii bacteremia.

BACKGROUND: Patients infected with Acinetobacter baumannii (AB) bacteremia in hospital have high morbidity and mortality. We analyzed the clinical characteristics of pneumonia and nonpneumonia-related AB bloodstream infections (AB BSIs) and explored the possible independent risk factors for the incidence and prognosis of pneumonia-related AB BSIs. METHODS: A retrospective monocentric observational study was performed. All 117 episodes of hospital-acquired AB bacteremia sorted into groups of pneumonia-related AB BSIs (n = 45) and nonpneumonia-related AB BSIs (n = 72) were eligible. Univariate/multivariate logistic regression analysis was used to explore the independent risk factors. The primary outcome was the antibiotic susceptibility in vitro of pneumonia-related AB BSIs group. The secondary outcome was the independent risk factor for the pneumonia-related AB BSIs group. RESULTS: Among 117 patients with AB BSIs, the pneumonia-related group had a greater risk of multidrug resistant A. baumannii (MDRAB) infection (84.44%) and carbapenem-resistant A. baumannii (CRAB) infection (80%). Polymyxin, minocycline and amikacin had relatively high susceptibility rates (> 80%) in the nonpneumonia-related group. However, in the pneumonia-related group, only polymyxin had a drug susceptibility rate of over 80%. Univariate analysis showed that survival time (day), CRAB, MDRAB, length of hospital stay prior to culture, length of ICU stay prior to culture, immunocompromised status, antibiotics used prior to culture (n > = 3 types), endotracheal tube, fiberoptic bronchoscopy, PITT, SOFA and invasive interventions (n > = 3 types) were associated with pneumonia-related AB bacteremia. The multivariate logistic regression analysis revealed that recent surgery (within 1 mo) [P = 0.043; 0.306 (0.098-0.962)] and invasive interventions (n > = 3 types) [P = 0.021; 0.072 (0.008-0.671)] were independent risk factors related to pneumonia-related AB bacteremia. Multivariate logistic regression analysis revealed that length of ICU stay prior to culture [P = 0.009; 0.959 (0.930-0.990)] and recent surgery (within 1 mo) [P = 0.004; 0.260 (0.105-0.646)] were independent risk factors for mortality in patients with pneumonia-related AB bacteremia. The Kaplan‒Meier curve and the timing test showed that patients with pneumonia-related AB bacteremia had shorter survival time compared to those with nonpneumonia-related AB bacteremia. CONCLUSIONS: Our study found that A. baumannii had a high rate of antibiotic resistance in vitro in the pneumonia-related bacteremia group, and was only sensitive to polymyxin. Recent surgery was a significantly independent predictor in patients with pneumonia-related AB bacteremia.

Characterization of Acinetobacter baumannii-calcoaceticus complex isolates and microbiological outcome for patients treated with sulbactam-durlobactam in a phase 3 trial (ATTACK).

Acinetobacter baumannii-calcoaceticus complex (ABC) causes severe, difficult-to-treat infections that are frequently antibiotic resistant. Sulbactam-durlobactam (SUL-DUR) is a targeted ß-lactam/ß-lactamase inhibitor combination antibiotic designed to treat ABC infections, including those caused by multidrug-resistant strains. In a global, pathogen-specific, randomized, controlled phase 3 trial (ATTACK), the efficacy and safety of SUL-DUR were compared to colistin, both dosed with imipenem-cilastatin as background therapy, in patients with serious infections caused by carbapenem-resistant ABC. Results from ATTACK showed that SUL-DUR met the criteria for non-inferiority to colistin for the primary efficacy endpoint of 28-day all-cause mortality with improved clinical and microbiological outcomes compared to colistin. This report describes the characterization of the baseline ABC isolates from patients enrolled in ATTACK, including an analysis of the correlation of microbiological outcomes with SUL-DUR MIC values and the molecular drivers of SUL-DUR resistance.

Emergence of eravacycline heteroresistance in carbapenem-resistant Acinetobacter baumannii isolates in China.

Carbapenem-resistant Acinetobacter baumannii (CRAB) is resistant to almost all antibiotics. Eravacycline, a newer treatment option, has the potential to treat CRAB infections, however, the mechanism by which CRAB isolates develop resistance to eravacycline has yet to be clarified. This study sought to investigate the features and mechanisms of eravacycline heteroresistance among CRAB clinical isolates. A total of 287 isolates were collected in China from 2020 to 2022. The minimum inhibitory concentration (MIC) of eravacycline and other clinically available agents against A. baumannii were determined using broth microdilution. The frequency of eravacycline heteroresistance was determined by population analysis profiling (PAP). Mutations and expression levels of resistance genes in heteroresistant isolates were determined by polymerase chain reaction (PCR) and quantitative real-time PCR (qRT-PCR), respectively. Antisense RNA silencing was used to validate the function of eravacycline heteroresistant candidate genes. Twenty-five eravacycline heteroresistant isolates (17.36%) were detected among 144 CRAB isolates with eravacycline MIC values ≤4 mg/L while no eravacycline heteroresistant strains were detected in carbapenem-susceptible A. baumannii (CSAB) isolates. All eravacycline heteroresistant strains contained OXA-23 carbapenemase and the predominant multilocus sequence typing (MLST) was ST208 (72%). Cross-resistance was observed between eravacycline, tigecycline, and levofloxacin in the resistant subpopulations. The addition of efflux pump inhibitors significantly reduced the eravacycline MIC in resistant subpopulations and weakened the formation of eravacycline heteroresistance in CRAB isolates. The expression levels of adeABC and adeRS were significantly higher in resistant subpopulations than in eravacycline heteroresistant parental strains (P < 0.05). An ISAba1 insertion in the adeS gene was identified in 40% (10/25) of the resistant subpopulations. Decreasing the expression of adeABC or adeRS by antisense RNA silencing significantly inhibited eravacycline heteroresistance. In conclusion, this study identified the emergence of eravacycline heteroresistance in CRAB isolates in China, which is associated with high expression of AdeABC and AdeRS.

Post Trauma Acinetobacter Baumanii Meningitis Treatment Approach.

A. baumannii, which is said to be associated with nosocomial infections is considered a significant mortality risk if not adequately addressed. A. baumannii infections typically occur in the aftermath of surgery or trauma. Our patient developed complicated A. baumannii meningitis with lateral ventriculitis and a lumbar abscess post surgery after suffering from a fall. The patient was treated with a 21-day regimen of intrathecally administered colistin and polymyxin B. Following this therapeutic period, the patient's condition improved, ultimately leading to successful recovery and subsequent discharge. This case report serves to highlight the ability of intrathecal administration of antibiotics, that normally have limited potential of crossing the blood-brain-barrier, to lead to improved survival outcomes in multi-drug resistant nosocomial meningitis.

Efflux Pump Inhibitor Potentiates the Antimicrobial Photodynamic Inactivation of Multidrug-Resistant Acinetobacter baumannii.

Background: Acinetobacter baumannii, a nosocomial pathogen, poses a major public health problem due to generating resistance to several antimicrobial agents. Antimicrobial photodynamic inactivation (APDI) employs a nontoxic dye as a photosensitizer (PS) and light to produce reactive oxygen species that destroy bacterial cells. The intracellular concentration of PS could be affected by factors such as the function of efflux pumps to emit PS from the cytosol. Objective: To evaluate the augmentation effect of an efflux pump inhibitor, verapamil, three multidrug-resistant A. baumannii were subjected to APDI by erythrosine B (EB). Methods and results: The combination of EB and verapamil along with irradiation at 530 nm induced a lethal effect and more than 3 log colony-forming unit reduction to all A. baumannii strains in planktonic state. In contrast, EB and irradiation alone could produce only a sublethal effect on two of the strains. Conclusions: These data suggest that verapamil increases the intracellular concentration of EB, which potentiates the lethal efficacy of APDI. Verapamil could be applied with EB and green light to improve their antimicrobial efficacy against A. baumannii-localized infections.

Coupling of zinc and GTP binding drives G-domain folding in Acinetobacter baumannii ZigA.

COG0523 proteins, also known as nucleotide-dependent metallochaperones, are a poorly understood class of small P-loop G3E GTPases. Multiple family members play critical roles in bacterial pathogen survival during an infection as part of the adaptive response to host-mediated "nutritional immunity." Our understanding of the structure, dynamics, and molecular-level function of COG0523 proteins, apart from the eukaryotic homolog, Zng1, remains in its infancy. Here, we use X-ray absorption spectroscopy to establish that Acinetobacter baumannii (Ab) ZigA coordinates ZnII using all three cysteines derived from the invariant CXCC motif to form an S3(N/O) coordination complex, a feature inconsistent with the ZnII-bound crystal structure of a distantly related COG0523 protein of unknown function from Escherichia coli, EcYjiA. The binding of ZnII and guanine nucleotides is thermodynamically linked in AbZigA, and this linkage is more favorable for the substrate GTP relative to the product GDP. Part of this coupling originates with nucleotide-induced stabilization of the G-domain tertiary structure as revealed by global thermodynamics measurements and hydrogen-deuterium exchange mass spectrometry (HDX-MS). HDX-MS also reveals that the HDX behavior of the G2 (switch 1) loop is highly sensitive to nucleotide status and becomes more exchange labile in the GDP (product)-bound state. Significant long-range perturbation of local stability in both the G-domain and the C-terminal domain define a candidate binding pocket for a client protein that appears sensitive to nucleotide status (GDP versus GTP). We place these new insights into the structure, dynamics, and energetics of intermolecular metal transfer into the context of a model for AbZigA metallochaperone function.

High-Throughput Host-Microbe Single-Cell RNA Sequencing Reveals Ferroptosis-Associated Heterogeneity during Acinetobacter baumannii Infection.

Interactions between host and bacterial cells are integral to human physiology. The complexity of host-microbe interactions extends to different cell types, spatial aspects, and phenotypic heterogeneity, requiring high-resolution approaches to capture their full complexity. The latest breakthroughs in single-cell RNA sequencing (scRNA-seq) have opened up a new era of studies in host-pathogen interactions. Here, we first report a high-throughput cross-species dual scRNA-seq technology by using random primers to simultaneously capture both eukaryotic and bacterial RNAs (scRandom-seq). Using reference cells, scRandom-seq can detect individual eukaryotic and bacterial cells with high throughput and high specificity. Acinetobacter baumannii (A.b) is a highly opportunistic and nosocomial pathogen that displays resistance to many antibiotics, posing a significant threat to human health, calling for discoveries and treatment. In the A.b infection model, scRandom-seq witnessed polarization of THP-1 derived-macrophages and the intracellular A.b-induced ferroptosis-stress in host cells. The inhibition of ferroptosis by Ferrostatin-1 (Fer-1) resulted in the improvement of cell vitality and resistance to A.b infection, indicating the potential to resist related infections. scRandom-seq provides a high-throughput cross-species dual single-cell RNA profiling tool that will facilitate future discoveries in unraveling the complex interactions of host-microbe interactions in infection systems and tumor micro-environments.

Activity of zinc oxide and zinc borate nanoparticles against resistant bacteria in an experimental lung cancer model.

BACKGROUND: Recent research indicates a prevalence of typical lung infections, such as pneumonia, in lung cancer patients. Klebsiella pneumoniae, Pseudomonas aeruginosa, and Acinetobacter baumannii stand out as antibiotic-resistant pathogens. Given this, there is a growing interest in alternative therapeutic avenues. Boron and zinc derivatives exhibit antimicrobial, antiviral, and antifungal properties. OBJECTIVES: This research aimed to establish the effectiveness of ZnO and ZB NPs in combating bacterial infections in lung cancer cell lines. METHODS: Initially, this study determined the minimal inhibitory concentration (MIC) and fractional inhibitory concentration (FIC) of zinc oxide nanoparticles (ZnO NPs) and zinc borate (ZB) on chosen benchmark strains. Subsequent steps involved gauging treatment success through a lung cancer-bacteria combined culture and immunohistochemical analysis. RESULTS: The inhibitory impact of ZnO NPs on bacteria was charted as follows: 0.97 µg/mL for K. pneumoniae 700603, 1.95 µg/mL for P. aeruginosa 27853, and 7.81 µg/mL for Acinetobacter baumannii 19,606. In comparison, the antibacterial influence of zinc borate was measured as 7.81 µg/mL for Klebsiella pneumoniae 700603 and 500 µg/mL for both P. aeruginosa 27853 and A.baumannii 19606. After 24 h, the cytotoxicity of ZnO NPs and ZB was analyzed using the MTT technique. The lowest cell viability was marked in the 500 µg/mL ZB NPs group, with a viability rate of 48.83% (P < 0.001). However, marked deviations appeared at ZB concentrations of 61.5 µg/mL (P < 0.05) and ZnO NPs at 125 µg/mL. CONCLUSION: A synergistic microbial inhibitory effect was observed when ZnO NP and ZB were combined against the bacteria under investigation.

Phage-mediated colistin resistance in Acinetobacter baumannii.

AIMS: Antimicrobial resistance is a global threat to human health, and Acinetobacter baumannii is a paradigmatic example of how rapidly bacteria become resistant to clinically relevant antimicrobials. The emergence of multidrug-resistant A. baumannii strains has forced the revival of colistin as a last-resort drug, suddenly leading to the emergence of colistin resistance. We investigated the genetic and molecular basis of colistin resistance in A. baumannii, and the mechanisms implicated in its regulation and dissemination. METHODS: Comparative genomic analysis was combined with genetic, biochemical, and phenotypic assays to characterize Φ19606, an A. baumannii temperate bacteriophage that carries a colistin resistance gene. RESULTS: Ф19606 was detected in 41% of 523 A. baumannii complete genomes and demonstrated to act as a mobile vehicle of the colistin resistance gene eptA1, encoding a functional lipid A phosphoethanolamine transferase. The eptA1 gene is coregulated with its chromosomal homolog pmrC via the PmrAB two-component system and confers colistin resistance when induced by low calcium and magnesium levels. Resistance selection assays showed that the eptA1-harbouring phage Ф19606 promotes the emergence of spontaneous colistin-resistant mutants. CONCLUSIONS: Φ19606 is an unprecedented example of a self-transmissible phage vector implicated in the dissemination of colistin resistance.

Regulation of overexpressed efflux pump encoding genes by cinnamon oil and trimethoprim to abolish carbapenem-resistant Acinetobacter baumannii clinical strains.

Resistance mechanisms are a shelter for Acinetobacter baumannii to adapt to our environment which causes difficulty for the infections to be treated and WHO declares this organism on the top of pathogens priority for new drug development. The most common mechanism that develops drug resistance is the overexpression of the efflux pump, especially Resistance-nodulation-cell division (RND) family, to almost most antibiotics. The study is designed to detect RND efflux pump genes in A. baumannii, and its correlation to multidrug resistance, in particular, the carbapenems resistance Acinetobacter baumannii (CRAB), and using different inhibitors that restore the antibiotic susceptibility of imipenem. Clinical A. baumannii isolates were recovered from different Egyptian hospitals in Intensive care unit (ICU). The expression of genes in two strains was analyzed using RT-PCR before and after inhibitor treatment. About 100 clinical A. baumannii isolates were recovered and identified and recorded as MDR strains with 75% strains resistant to imipenem. adeB, adeC, adeK, and adeJ were detected in thirty- seven the carbapenems resistance Acinetobacter baumannii (CRAB) strains. Cinnamomum verum oil, Trimethoprim, and Omeprazole was promising inhibitor against 90% of the carbapenems resistance Acinetobacter baumannii (CRAB) strains with a 2-6-fold decrease in imipenem MIC. Downregulation of four genes was associated with the addition of those inhibitors to imipenem for two the carbapenems resistance Acinetobacter baumannii (CRAB) (ACN15 and ACN99) strains, and the effect was confirmed in 24 h killing kinetics. Our investigation points to the carbapenems resistance Acinetobacter baumannii (CRAB) strain's prevalence in Egyptian hospitals with the idea to revive the imipenem activity using natural and chemical drugs as inhibitors that possessed high synergistic activity.

Multi-drug Resistant Acinetobacter baumannii: Phenotypic and Genotypic Resistance Profiles and the Associated Risk Factors in Teaching Hospital in Jordan.

BACKGROUND: This study aimed to determine the prevalence of Antimicrobial Resistance Genes (ARGs), with a focus on colistin resistance in clinical A. baumannii, as well as the risk factors associated with A. baumannii infection in Jordanian patients. METHODS: In total, 150 A. baumannii isolates were obtained from patients at a teaching hospital. The isolates were tested for antimicrobial susceptibility using disc diffusion and microdilution methods. PCR amplification was used to detect ARGs, and statistical analysis was conducted to evaluate the influence of identified risk factors on the ARGs acquisition. RESULTS: More than 90% of A. baumannii isolates were resistant to monobactam, carbapenem, cephalosporins, Fluoroquinolones, penicillin, and ß-lactam agents. Moreover, 20.6% of the isolates (n = 31) were colistin-resistant. Several ARGs were also detected in A. baumannii isolates. Univariate analysis indicated that risk factors and the carriage of ARGs were significantly associated P ≤ (0.05) with gender, invasive devices, immunodeficiency, systemic diseases, tumors, and covid-19. Logistic regression analysis indicated seven risk factors, and three protective factors were associated with the ARGs (armA, strA, and strB) P ≤ (0.05). In contrast, tetB and TEM were associated with 2 risk factors each P ≤ (0.05). CONCLUSION: Our study indicates a high prevalence of MDR A. baumannii infections in ICU patients, as well as describing the case of colistin-resistant A. baumannii for the first time in Jordan. Additionally, the risk factors associated with ARGs-producing A. baumannii infections among ICU patients suggest a rapid emergence and spread of MDR A. baumannii without adequate surveillance and control measures.

Recovery and genetic characterization of clinically-relevant ST2 carbapenem-resistant Acinetobacter baumannii isolates from untreated hospital sewage in Zhejiang Province, China.

The global transmission of carbapenem-resistant Acinetobacter baumannii (CRAB) poses a significant and grave threat to human health. To investigate the potential relationship between hospital sewage and the transmission of CRAB within healthcare facilities, isolates of Acinetobacter spp. obtained from untreated hospital sewage samples were subjected to antimicrobial susceptibility tests, genome sequencing, and bioinformatic and phylogenetic tree analysis, and that data were matched with those of the clinical isolates. Among the 70 Acinetobacter spp. sewage isolates tested, A. baumannii was the most prevalent and detectable in 5 hospitals, followed by A. nosocomialis and A. gerneri. Worryingly, 57.14 % (40/70) of the isolates were MDR, with 25.71 % (18/70) being resistant to carbapenem. When utilizing the Pasteur scheme, ST2 was the predominant type among these CRAB isolates, with Tn2006 (ΔISAba1-blaOXA-23-ATPase-yeeB-yeeA-ΔISAba1) and Tn2009 (ΔISAba1-blaOXA-23-ATPase-hp-parA-yeeC-hp-yeeB-ΔISAba1) being the key mobile genetic elements that encode carbapenem resistance. Seven A. gerneri isolates which harbored Tn2008 (ISAba1-blaOXA-23 -ATPase) and the blaPER-1 gene were also identified. Besides, an A. soil isolate was found to exhibit high-level of meropenem resistance (MIC ≥128 mg/L) and harbor a blaNDM-1 gene located in a core genetic structure of ISAba125-blaNDM-1-ble-trpF-dsbC-cutA. To investigate the genetic relatedness between isolates recovered from hospital sewage and those collected from ICUs, a phylogenetic tree was constructed for 242 clinical isolates and 9 sewage isolates. The results revealed the presence of two evolutionary clades, each containing isolates from both ICU and sewage water, suggesting that CRAB isolates in untreated sewage water were also the transmission clones or closely related evolutionary isolates recoverable in hospital settings. Findings in this work confirm that hospital sewage is a potential reservoir of CRAB.

Sub-minimum inhibitory concentrations (sub-MICs) of colistin on Acinetobacter baumannii biofilm formation potency, adherence, and invasion to epithelial host cells: an experimental study in an Iranian children's referral hospital.

Here, we described the efficacy of colistin sub-minimum inhibitory concentrations (sub-MICs) on biofilm-forming activity, host epithelial cell adherence, and invasion capacity of Acinetobacter baumannii strains collected from children admitted to the Children's Medical Center Hospital. Biofilm formation potency of A. baumannii clinical isolates was measured using a 96-well microtiter plate assay. Distribution of biofilm-related genes, including bap, abaI, ompA, csuE, and blaPER-1, was detected by PCR. The mRNA expression level of ompA and csuE was measured by qPCR in the presence of » and ½ MICs of colistin. A. baumannii adhesion and invasion to eukaryotic host cells were phenotypically assayed at sub-MICs of colistin. Eighty percent (56/70) and 35.7% (25/70) of A. baumannii isolates were multidrug-resistant (MDR) and extensively drug-resistant (XDR) phenotypes, respectively. The strong, moderate, and weak biofilm producers of A. baumannii were 37.1% (26/70), 32.8%, (23/70), and 22.8% (16/70), respectively. The frequencies of biofilm-associated genes were 100% for abaI, ompA, and csuE, followed by 22.8% (16/70) and 24.3% (17/70) for bap and blaPER-1, respectively. The downregulation of csuE and ompA expression levels was observed in the sub-MIC of colistin. In vitro cell culture study showed a decreased capability of A. baumannii to adhere to the human epithelial cells at sub-inhibitory doses of colistin; however, none of the isolates could invade HEp-2 cells. Our study showed that the genes encoding biofilm-associated proteins undergo downregulation in expression levels after exposure to sub-MICs of colistin in A. baumannii. Longitudinal in vivo studies are needed to fully understand the clinical aspects of pathogenicity mechanisms and evolutionary dynamics of drug resistance.IMPORTANCESince the toxicity of colistin is dose dependent, there is a focus on strategies that reduce the dose while maintaining the therapeutic effect of the drug. Our findings about sub-inhibitory doses of colistin provide a novel insight into the logical use of colistin to treat and control Acinetobacter baumannii-related infections in clinical practice.

Novel Antimicrobial Peptide SAAP Mutant as a Better Adjuvant to Sulbactam-Based Treatments Against Clinical Strains of XDR Acinetobacter baumannii.

The production of extended spectrum ß-lactamases (ESBLs) in extensively drug-resistant (XDR) strains of Acinetobacter baumannii has created havoc amongst clinicians making the treatment procedure challenging. Carbapenem-resistant strains have displayed total ineffectiveness towards newer combinations of ß-lactam-ß-lactamase inhibitors (ßL-ßLI) in tertiary healthcare settings. Therefore, the present study was aimed to design potential ß-lactamase antimicrobial peptide (AMP) inhibitors against ESBLs produced by the strains. We have constructed an AMP mutant library with higher antimicrobial efficacy (range: ~ 15 to 27%) than their parent peptides. The mutants were thoroughly screened based on different physicochemical and immunogenic properties revealing three peptides, namely SAAP-148, HFIAP-1, myticalin-C6 and their mutants with safe pharmacokinetics profile. Molecular docking highlighted SAAP-148_M15 displaying maximum inhibitory potential with lowest binding energies against NDM1 (- 1148.7 kcal/mol), followed by OXA23 (- 1032.5 kcal/mol) and OXA58 (- 925.3 kcal/mol). The intermolecular interaction profiles displayed SAAP-148_M15 exhibiting hydrogen bonds and van der Waals hydrophobic interactions with the crucial residues of metallo ß-lactamase [IPR001279] and penicillin-binding transpeptidase [IPR001460] domains. Coarse-grained clustering and molecular dynamics simulations (MDS) further validated the stable backbone profile and minimal residue-level fluctuations of the protein-peptide complex that were maintained throughout the simulation timeframe. The present study hypothesised that the combination of sulbactam (ßL) with SAAP-148_M15 (ßLI) holds immense potential in inhibiting the ESBLs alongside restoration of sulbactam activity. The current in silico findings upon further experimental validations can pave path towards designing of successful therapeutic strategy against XDR strains of A. baumannii.

Sulbactam-durlobactam susceptibility test method development and quality control ranges for MIC and disk diffusion tests.

Sulbactam-durlobactam is a ß-lactam/ß-lactamase inhibitor combination developed to treat hospital-acquired and ventilator-associated bacterial pneumonia caused by Acinetobacter baumannii-calcoaceticus complex (ABC). Durlobactam is a diazabicyclooctane ß-lactamase inhibitor with potent activity against Ambler classes A, C, and D serine ß-lactamases and restores sulbactam activity against multidrug-resistant ABC. Studies were conducted to establish sulbactam-durlobactam antimicrobial susceptibility testing methods for both broth microdilution minimal inhibitory concentration (MIC) and disk diffusion tests as well as quality control (QC) ranges. To establish the MIC test method, combinations of sulbactam and durlobactam were evaluated using a panel of genetically characterized A. baumannii isolates which were categorized as predicted to be susceptible or resistant based on the spectrum of ß-lactamase inhibition by durlobactam. MIC testing with doubling dilutions of sulbactam with a fixed concentration of 4 µg/mL of durlobactam resulted in the greatest discrimination of the pre-defined susceptible and resistant strains. Similarly, the sulbactam/durlobactam 10/10 µg disk concentration showed the best discrimination as well as correlation with the MIC test. A. baumannii NCTC 13304 was selected for QC purposes because it assesses the activity of both sulbactam and durlobactam with clear endpoints. Multi-laboratory QC studies were conducted according to CLSI M23 Tier 2 criteria. A sulbactam-durlobactam broth MIC QC range of 0.5/4-2/4 µg/mL and a zone diameter QC range of 24-30 mm were determined for A. baumannii NCTC 13304 and have been approved by CLSI. These studies will enable clinical laboratories to perform susceptibility tests with accurate and reproducible methods.

In vivo dose response and efficacy of the ß-lactamase inhibitor, durlobactam, in combination with sulbactam against the Acinetobacter baumannii-calcoaceticus complex.

Multi-drug resistant (MDR) Acinetobacter baumannii is emerging as a pathogen of increasing prevalence and concern. Infections associated with this Gram-negative pathogen are often associated with increased morbidity and mortality and few therapeutic options. The ß-lactamase inhibitor sulbactam used commonly in combination with ampicillin demonstrates intrinsic antibacterial activity against A. baumannii acting as an inhibitor of PBP1 and PBP3, which participate in cell wall biosynthesis. The production of ß-lactamases, particularly class D oxacillinases, however, has limited the utility of sulbactam resorting to increased doses and the need for alternate therapies. Durlobactam is a non-ß-lactam ß-lactamase inhibitor that demonstrates broad ß-lactamase inhibition including class D enzymes produced by A. baumannii and has shown potent in vitro activity against MDR A. baumannii, particularly carbapenem-resistant isolates in susceptibility and pharmacodynamic model systems. The objective of this study is to evaluate the exposure-response relationship of sulbactam and durlobactam in combination using in vivo neutropenic thigh and lung models to establish PK/PD exposure magnitudes to project clinically effective doses. Utilizing established PK/PD determinants of %T>MIC and AUC/MIC for sulbactam and durlobactam, respectively, non-linear regressional analysis of drug exposure was evaluated relative to the 24-hour change in bacterial burden (log10 CFU/g). Co-modeling of the data across multiple strains exhibiting a broad range of MIC susceptibility suggested net 1-log10 CFU/g0 reduction can be achieved when sulbactam T>MIC exceeds 50% of the dosing interval and durlobactam AUC/MIC is 10. These data were ultimately used to support sulbactam-durlobactam dose selection for Phase 3 clinical trials.

I n vitro pharmacokinetics/pharmacodynamics of the ß-lactamase inhibitor, durlobactam, in combination with sulbactam against Acinetobacter baumannii-calcoaceticus complex.

Infections caused by Acinetobacter baumannii are increasingly multidrug resistant and associated with high rates of morbidity and mortality. Sulbactam is a ß-lactamase inhibitor with intrinsic antibacterial activity against A. baumannii. Durlobactam is a non-ß-lactam ß-lactamase inhibitor with an extended spectrum of activity compared to other inhibitors of its class. In vitro pharmacodynamic infection models were undertaken to establish the pharmacokinetic/pharmacodynamic (PK/PD) index and magnitudes associated with sulbactam and durlobactam efficacy and to simulate epithelial lining fluid (ELF) exposures at clinical doses to understand sulbactam-durlobactam activity with and without co-administration of a carbapenem. Hollow fiber infection models (HFIMs) and one-compartment systems were used to identify the PK/PD indices and exposure magnitudes associated of 1-log10 and 2-log10 colony-forming unit (CFU)/mL reductions. Sulbactam and durlobactam demonstrated PK/PD drivers of % time above the minimum inhibition concentration (%T > MIC) and area under the plasma concentration-time curve from time 0 to 24 h (AUC0-24)/MIC, respectively. Against a sulbactam-susceptible strain, sulbactam %T > MIC of 71.5 and 82.0 were associated with 1-log10 and 2-log10 CFU/mL reductions, respectively, in the HFIM. Against a non-susceptible strain, durlobactam restored the activity of sulbactam with an AUC0-24/MICs of 34.0 and 46.8 using a polysulfone cartridge to achieve a 1-log10 and 2-log10 CFU/mL reduction. These magnitudes were reduced to 13.8 and 24.2, respectively, using a polyvinylidene fluoride cartridge with a membrane pore size of 0.1 µm. In the one-compartment model, durlobactam AUC0-24/MIC to achieve 1-log10 and 2-log10 CFU/mL reduction were 7.6 and 33.4, respectively. Simulations of clinical ELF exposures in the HFIM showed cidal activity at MICs ≤4 µg/mL. Penicillin binding protein 3 mutant strains with MICs of 8 µg/mL may benefit from the addition of a carbapenem at clinical exposures.

Inhibitory Effects of Carvacrol on Biofilm Formation in Colistin Heteroresistant Acinetobacter baumannii Clinical Isolates.

BACKGROUND: The ability of bacteria to form biofilm is an essential strategy for creating stable infections. This issue is more critical in Acinetobacter bauamannii as a hospital pathogen. Today, the control of biofilm formation and solutions to prevent or remove biofilm is being developed. Carvacrol has been considered an anti-biofilm compound in significant bacteria. This study investigated the anti-biofilm effect of Carvacrol on biofilm formation in clinical colistin heteroresistant isolates of A. baumannii. METHODS: 22 clinical strains of A. baumannii were collected from Motahari Hospital in Tehran, Iran, in 2019. Biochemical and genotypic methods confirmed these isolates. Colistin heteroresistance was determined by the Standard PAP method. Carvacrol's antibacterial and anti-biofilm activity was determined according to the standard protocol. RESULTS: About 12 isolates were considered strong biofilm producers and were used for analysis. Six isolates had hetero-resistance to colistin. Carvacrol at a 512 g/ml concentration showed the best antibacterial activity against all isolates. The sub-MIC of Carvacrol (256 g/ml) reduced the biofilm formation capacity, which was statistically significant (p < 0.05). CONCLUSION: The results of this study showed that sub-MIC of Carvacrol has anti-biofilm effects in clinical A.baumannii colistin hetero-resistance isolates.