Successful Treatment of Carbapenem-Resistant Acinetobacter baumannii Meningitis with Sulbactam-Durlobactam.

BACKGROUND: The treatment of carbapenem-resistant Acinetobacter baumannii/calcoaceticus complex (CRAB) presents significant treatment challenges. METHODS: We report the case of a 42-year-old woman with CRAB meningitis who experienced persistently positive cerebrospinal fluid (CSF) cultures for 13 days despite treatment with high-dose ampicillin-sulbactam and cefiderocol. On day 13, she was transitioned to sulbactam-durlobactam and meropenem; four subsequent CSF cultures remained negative. After 14 days of sulbactam-durlobactam, she was cured of infection. Whole genome sequencing investigations identified putative mechanisms that contributed to reduced cefiderocol susceptibility observed during cefiderocol therapy. Blood and CSF samples were collected pre-dose and 3-hours post initiation of a sulbactam-durlobactam infusion. RESULTS: The CRAB isolate belonged to sequence type 2. An acquired blaOXA-23 and an intrinsic blaOXA-51-like (i.e., blaOXA-66) carbapenemase gene were identified. The paradoxical effect (i.e., no growth at lower cefiderocol dilutions but growth at higher dilutions) was observed by broth microdilution after 8 days of cefiderocol exposure but not by disk diffusion. Potential markers of resistance to cefiderocol included mutations in the start codon of piuA and piuC iron transport genes and a A515V substitution in PBP3, the primary target of cefiderocol. Sulbactam and durlobactam were detected in CSF at both timepoints, indicating CSF penetration. CONCLUSIONS: This case describes successful treatment of refractory CRAB meningitis with the administration of sulbactam-durlobactam and meropenem and highlights the need to be cognizant of the paradoxical effect that can be observed with broth microdilution testing of CRAB isolates with cefiderocol.

Epidemiological insights into seasonal, sex­specific and age­related distribution of bacterial pathogens in urinary tract infections.

Urinary tract infections (UTIs) are prevalent and recurrent bacterial infections that affect individuals worldwide, posing a significant burden on healthcare systems. The present study aimed to explore the epidemiology of UTIs, investigating the seasonal, gender-specific and age-related bacterial pathogen distribution to guide clinical diagnosis. Data were retrospectively collected from electronic medical records and laboratory reports of 926 UTIs diagnosed in Fuding Hospital (Fujian University of Traditional Chinese Medicine, Fuding, China). Bacterial isolates were identified using standard microbiological techniques. χ2 tests were performed to assess associations between pathogens and the seasons, sex and age groups. Significant associations were found between bacterial species and seasons. Enterococcus faecium exhibited a substantial prevalence in spring (χ2, 12.824; P=0.005), while Acinetobacter baumannii demonstrated increased prevalence in autumn (χ2, 16.404; P=0.001). Female patients showed a higher incidence of UTIs. Gram-positive bacteria were more prevalent in males, with Staphylococcus aureus showing significant male predominance (χ2, 14.607; P<0.001). E. faecium displayed an age-related increase in prevalence (χ2, 17.775; P<0.001), whereas Escherichia coli tended to be more prevalent in younger patients (χ2, 12.813; P=0.005). These findings highlight the complex nature of UTIs and offer insights for tailored diagnostic and preventive strategies, potentially enhancing healthcare outcomes.

Critical role of growth medium for detecting drug interactions in Gram-negative bacteria that model in vivo responses.

The rise in infections caused by multidrug-resistant (MDR) bacteria has necessitated a variety of clinical approaches, including the use of antibiotic combinations. Here, we tested the hypothesis that drug-drug interactions vary in different media, and determined which in vitro models best predict drug interactions in the lungs. We systematically studied pair-wise antibiotic interactions in three different media, CAMHB, (a rich lab medium standard for antibiotic susceptibility testing), a urine mimetic medium (UMM), and a minimal medium of M9 salts supplemented with glucose and iron (M9Glu) with three Gram-negative ESKAPE pathogens, Acinetobacter baumannii (Ab), Klebsiella pneumoniae (Kp), and Pseudomonas aeruginosa (Pa). There were pronounced differences in responses to antibiotic combinations between the three bacterial species grown in the same medium. However, within species, PaO1 responded to drug combinations similarly when grown in all three different media, whereas Ab17978 and other Ab clinical isolates responded similarly when grown in CAMHB and M9Glu medium. By contrast, drug interactions in Kp43816, and other Kp clinical isolates poorly correlated across different media. To assess whether any of these media were predictive of antibiotic interactions against Kp in the lungs of mice, we tested three antibiotic combination pairs. In vitro measurements in M9Glu, but not rich medium or UMM, predicted in vivo outcomes. This work demonstrates that antibiotic interactions are highly variable across three Gram-negative pathogens and highlights the importance of growth medium by showing a superior correlation between in vitro interactions in a minimal growth medium and in vivo outcomes. IMPORTANCE: Drug-resistant bacterial infections are a growing concern and have only continued to increase during the SARS-CoV-2 pandemic. Though not routinely used for Gram-negative bacteria, drug combinations are sometimes used for serious infections and may become more widely used as the prevalence of extremely drug-resistant organisms increases. To date, reliable methods are not available for identifying beneficial drug combinations for a particular infection. Our study shows variability across strains in how drug interactions are impacted by growth conditions. It also demonstrates that testing drug combinations in tissue-relevant growth conditions for some strains better models what happens during infection and may better inform combination therapy selection.

Anti-Acinetobacter Baumannii single-chain variable fragments provide therapeutic efficacy in an immunocompromised mouse pneumonia model.

BACKGROUND: The emergence of carbapenem-resistant and extensively drug-resistant (XDR) Acinetobacter baumannii as well as inadequate effective antibiotics calls for an urgent effort to find new antibacterial agents. The therapeutic efficacy of two human scFvs, EB211 and EB279, showing growth inhibitory activity against A. baumannii in vitro, was investigated in immunocompromised mice with A. baumannii pneumonia. RESULTS: The data revealed that infected mice treated with EB211, EB279, and a combination of the two scFvs showed better survival, reduced bacterial load in the lungs, and no marked pathological abnormalities in the kidneys, liver, and lungs when compared to the control groups receiving normal saline or an irrelevant scFv. CONCLUSIONS: The results from this study suggest that the scFvs with direct growth inhibitory activity could offer promising results in the treatment of pneumonia caused by XDR A. baumannii.

Antimicrobial activity of spiculisporic acid isolated from endophytic fungus Aspergillus cejpii of Hedera helix against MRSA.

The surge in multidrug-resistant pathogens worldwide has jeopardized the clinical efficiency of many current antibiotics. This problem steered many researchers in their quest to discover new effective antimicrobial agents from natural origins including plants or their residing endophytes. In this work, we aimed to identify the endophytic fungi derived from Hedera helix L. and investigate their potential antimicrobial activity. Bioguided fractionation approach was conducted to isolate the pure compounds from the most active fungal fraction. Out of a total of six different isolated endophytic fungal strains, only Aspergillus cejpii showed the highest activity against all tested microbial strains. The most active fraction was the dichloromethane/methanol fraction (DCM:MeOH), where it showed significant activity against Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Serratia marcescens, Acinetobacter baumannii, Salmonella typhi, and three drug-resistant clinical isolate strains including Methicillin-resistant Staphylococcus aureus (MRSA, H1), Pseudomonas aeruginosa (PS 16), and Acinetobacter baumannii (ACT 322) using tetracyline and kanamycin as the control antibiotics. Bioguided fractionation of the active fraction led to the isolation of the γ-butenolide, spiculisporic acid. Structure elucidation was carried out using 1H and 13C-NMR spectroscopic analysis. The compound showed good antimicrobial activities with minimum inhibitory concentration (MIC) values ranging from 3.9 to 31.25 µg/mL against all tested strains. Gas chromatography coupled to mass spectrometry (GC-MS) profiling was also carried out to identify the metabolites in the microbial crude extract. In conclusion, endophytic fungi, Aspergillus cejpii, isolated from Hedera helix L. roots showed promising antimicrobial activity which merits further in-depth investigations for potential utilization as a source of new antibiotics in the future. It can also be considered as a novel source for spiculisporic acid.

Revisiting the role of Acinetobacter spp. in side-stream enhanced biological phosphorus removal (S2EBPR) systems.

We piloted the incorporation of side-stream enhanced biological phosphorus removal (S2EBPR) with A/B stage short-cut nitrogen removal processes to enable simultaneous carbon-energy-efficient nutrients removal. This unique configuration and system conditions exerted selective force on microbial populations distinct from those in conventional EBPR. Interestingly, effective P removal was achieved with the predominance of Acinetobacter (21.5 ± 0.1 %) with nearly negligible level of known conical PAOs (Ca. Accumulibacter and Tetrasphaera were 0.04 ± 0.10 % and 0.47 ± 0.32 %, respectively). Using a combination of techniques, such as fluorescence in situ hybridization (FISH) coupled with single cell Raman spectroscopy (SCRS), the metabolic tracing of Acinetobacter-like cells exerted PAO-like phenotypic profiling. In addition, comparative metagenomics analysis of the closely related Acinetobacter spp. revealed the EBPR relevant metabolic pathways. Further oligotyping analysis of 16s rRNA V4 region revealed sub-clusters (microdiversity) of the Acinetobacter and revealed that the sub-group (oligo type 1, identical (100 % alignment identity) hits from Acinetobacter_midas_s_49494, and Acinetobacter_midas_s_55652) correlated with EBPR activities parameters, provided strong evidence that the identified Acinetobacter most likely contributed to the overall P removal in our A/B-shortcut N-S2EBPR system. To the best of our knowledge, this is the first study to confirm the in situ EBPR activity of Acinetobacter using combined genomics and SCRS Raman techniques. Further research is needed to identify the specific taxon, and phenotype of the Acinetobacter that are responsible for the P-removal.

Molecular detection of Bartonella quintana, Acinetobacter baumannii and Acinetobacter haemolyticus in Pediculus humanus lice in Nigeria, West Africa.

The human lice Pediculus humanus is distributed worldwide but, it thrives and flourishes under conflict situations where people are forced to live in crowded unhygienic conditions. Molecular methods were used to identify and screen human lice for the DNA of pathogens of public health importance in an area that has been under insurgency related to religious and political conflicts with tens of thousands of internally displaced people (IDP). DNA of Bartonella quintana, Acinetobacter baumannii and Acinetobacter haemolyticus was detected in 18.3%, 40.0% and 1.7%, respectively, of human lice collected from children in Maiduguri, Nigeria. More body lice than head lice were positive for pathogen's DNA (64.3% vs. 44.4%; χ2 = 1.3, p = 0.33), but the difference was not significant. Two lice samples were found to harbour mixed DNA of B. quintana and A. baumannii. Phylogenetic analysis of the cytochrome b (cytb) gene sequences of the positive lice specimens placed them into clades A and E. This is the first report on the molecular identification of human lice and the detection of the DNA of pathogens of public health importance in lice in Nigeria, West Africa. The findings of this study will assist policy makers and medical practitioners in formulating a holistic healthcare delivery to IDPs.

Physical compatibility of sulbactam/durlobactam with select intravenous drugs during simulated Y-site administration.

PURPOSE: Sulbactam/durlobactam is a combination antibiotic designed to target Acinetobacter baumannii, including carbapenem-resistant and multidrug-resistant strains. The objective of this study was to determine the physical compatibility of sulbactam/durlobactam solution during simulated Y-site administration with 95 intravenous (IV) drugs. METHODS: Vials of sulbactam/durlobactam solution were diluted in 0.9% sodium chloride injection to a volume of 100 mL (the final concentration of both drugs was 15 mg/mL). All other IV drugs were reconstituted according to the manufacturer's recommendations and diluted with 0.9% sodium chloride injection to the upper range of concentrations used clinically or tested undiluted as intended for administration. Y-site conditions were simulated by mixing 5 mL of sulbactam/durlobactam with 5 mL of the tested drug solutions in a 1:1 ratio. Solutions were inspected for physical characteristics (clarity, color, and Tyndall effect), turbidity, and pH changes before admixture, immediately post admixture, and over 4 hours. Incompatibility was defined as any observed precipitation, significant color change, positive Tyndall test, or turbidity change of ≥0.5 nephelometric turbidity unit during the observation period. RESULTS: Sulbactam/durlobactam was physically compatible with 38 out of 42 antimicrobials tested (90.5%) and compatible overall with 86 of 95 drugs tested (90.5%). Incompatibility was observed with albumin, amiodarone hydrochloride, ceftaroline fosamil, ciprofloxacin, daptomycin, levofloxacin, phenytoin sodium, vecuronium, and propofol. CONCLUSION: The Y-site compatibility of sulbactam/durlobactam with 95 IV drugs was described. These compatibility data will assist pharmacists and nurses to safely coordinate administration of IV medications with sulbactam/durlobactam.

A Retrospective Case-Control Study of Eravacycline for the Treatment of Carbapenem-Resistant Acinetobacter Infections in Patients With Burn Injuries.

Thermal injuries lead to a deficiency in one's natural, protective barrier, resulting in increased susceptibility to pathogens, and often require multiple courses of broad-spectrum antibiotics. Eravacycline (ERA) has shown adequate in vitro activity against multiple multi-drug resistant (MDR) pathogens including Acinetobacter sp. Due to the increasing prevalence of MDR bacteria and the heightened susceptibility of patients with burns to infection, studies are needed to examine the clinical effect of eravacycline in this population. The objective of this retrospective, case-control study was to compare the outcomes of patients with thermal injuries treated with eravacycline versus a matched control for carbapenem-resistant Acinetobacter baumannii (CRAB) infections. Patients with thermal injury admitted to an American Burn Associated-verified burn center from May 1, 2019 to July 31, 2022, who received eravacycline, were randomly matched 4:1 to a historical cohort using a previously established, de-identified dataset of patients treated with colistimethate sodium (CMS) in the same burn center (March 1, 2009 to March 31, 2014), based on % total body surface area (%TBSA), age, and CRAB. A composite favorable outcome was defined as 30-day survival, completion of the course, lack of 14-day recurrence, and lack of acute kidney injury (AKI). Treatment with eravacycline over CMS was not more or less likely to be associated with a favorable response [odds ratio (95% confidence interval), 2.066 (0.456-9.361), P = .347]. Patients treated with CMS had nearly 9-fold higher odds of new-onset AKI versus ERA [8.816 (0.911-85.308), P = .06]. Adverse events were uncommon in the ERA group. There was no difference in mortality.

Perillaldehyde: A promising antibacterial agent for the treatment of pneumonia caused by Acinetobacter baumannii infection.

Perillaldehyde is a monoterpene compound mainly from the medicinal plant Perilla frutescens (L.) Britt., which has hypolipidemic, antioxidant, antibacterial and anti-inflammatory functions. In this investigation, we discovered that Perillaldehyde had powerful antimicrobial activity against Acinetobacter baumannii 5F1, and its minimum inhibitory concentration was 287.08 µg/mL. A. baumannii is a conditionally pathogenic bacterium with a high clinical resistance rate and is a major source of hospital infections, especially in intensive care units, which is one of the main causes of pneumonia. Inflammatory immune response is characteristic of pneumonia caused by A. baumannii infection. The results of our in vitro experiments indicate that Perillaldehyde disrupts the cell membrane of A. baumannii 5F1 and inhibits its quorum sensing to inhibit biofilm formation, among other effects. With an experimental model of murine pneumonia, we investigated that Perillaldehyde decreased NLRP3 inflammasome activation and TNF-α expression in lung tissues by inhibiting the NF-κB pathway, and also impacted MAPKs protein signaling pathway through the activation of TLR4. Notably, the use of high doses of Perillaldehyde for the treatment of pneumonia caused by A. baumannii 5F1 infection resulted in a survival rate of up to 80 % in mice. In summary, we demonstrated that Perillaldehyde is promising as a new drug for the treatment of pneumonia caused by A. baumannii 5F1 infection.

Clinical efficacy of cefiderocol-based regimens in patients with carbapenem-resistant Acinetobacter baumannii infections: A systematic review with meta-analysis.

OBJECTIVES: To perform a systematic review with meta-analysis to assess the clinical efficacy of cefiderocol-based regimens for the treatment of carbapenem-resistant Acinetobacter baumannii (CRAB) infections. METHODS: Two authors independently searched PubMed-MEDLINE, Scopus, and Cochrane databases, from inception to 02 July 2023, for randomised controlled trials (RCTs) or observational studies comparing clinical efficacy of cefiderocol-based vs. non-cefiderocol-based regimens in patients with CRAB infections. Data were extracted by the two authors independently, and the quality of included studies was independently assessed using ROB 2.0 or ROBINS-I tools. Primary outcome was mortality rate. Meta-analysis was performed by pooling odds ratios (ORs) retrieved from studies providing adjustment for confounders using a random-effects model with the inverse variance method. Multiple subgroups and sensitivity analyses were conducted to investigate the source of heterogeneity. RESULTS: A total of 530 articles were screened, and 6 studies (1 RCT and 5 observational; N=561; 247 cefiderocol-based vs. 314 non-cefiderocol-based regimens) were included. Cefiderocol did not significantly reduce in-hospital mortality compared to alternative therapies (predominantly colistin-based), but the confidence intervals around the effect estimate included clinically important benefit (N=5; OR 0.64; 95%CI 0.40-1.04; I2=57.5%). When only observational studies providing adjustment for confounders were considered, a lower risk of mortality was found in patients treated with cefiderocol-based regimens (N=4; OR 0.53; 95%CI 0.39-0.71; I2=0.0%). CONCLUSIONS: Cefiderocol-based regimens were associated with a significantly lower risk of mortality in patients with CRAB infections in observational studies providing proper adjustment for confounders.

Genotypic Evolution of Klebsiella pneumoniae Sequence Type 512 during Ceftazidime/Avibactam, Meropenem/Vaborbactam, and Cefiderocol Treatment, Italy.

In February 2022, a critically ill patient colonized with a carbapenem-resistant K. pneumoniae producing KPC-3 and VIM-1 carbapenemases was hospitalized for SARS-CoV-2 in the intensive care unit of Policlinico Umberto I hospital in Rome, Italy. During 95 days of hospitalization, ceftazidime/avibactam, meropenem/vaborbactam, and cefiderocol were administered consecutively to treat 3 respiratory tract infections sustained by different bacterial agents. Those therapies altered the resistome of K. pneumoniae sequence type 512 colonizing or infecting the patient during the hospitalization period. In vivo evolution of the K. pneumoniae sequence type 512 resistome occurred through plasmid loss, outer membrane porin alteration, and a nonsense mutation in the cirA siderophore gene, resulting in high levels of cefiderocol resistance. Cross-selection can occur between K. pneumoniae and treatments prescribed for other infective agents. K. pneumoniae can stably colonize a patient, and antimicrobial-selective pressure can promote progressive K. pneumoniae resistome evolution, indicating a substantial public health threat.

Effects of the combination of chitosan and Acinetobacter KU011TH on the growth and health performances and disease resistance of juvenile hybrid catfish (Clarias gariepinus × C. macrocephalus).

Aquatic animal health management has become a crucial component in the goal of increasing catfish aquaculture productivity. Additionally, hybrid catfish (Clarias gariepinus × C. macrocephalus) has been promoted as a highly profitable freshwater fish in Asia. Interestingly, the crucial diseases induced by Aeromonas hydrophila have been reported to greatly impede catfish production. To overcome this challenge, the aim was to investigate the effects of the oral administration of potentially synbiotic chitosan (CH) and Acinetobacter KU011TH (AK) on the growth performance, immunological responses, and disease resistance of hybrid catfish against A. hydrophila. The control group was fed a basal diet (A), the diet fed to treatment group B was supplemented with 20 mL of CH/kg diet (B), and the experimental feed fed to groups C-D was mixed with 1 × 108, 1 × 109 and 1 × 1010 CFU/mL AK coated with 20 mL of CH/kg diet. Five different groups of juvenile hybrid catfish were continuously fed the 5 formulated feeds for 4 weeks. The results revealed that all tested feeds did not significantly enhance the hybrid catfish's average daily gain, specific growth rate, feed conversion ratio, hematocrit and erythrocyte counts. Interestingly, the application of CH and AK significantly increased the leukocyte counts, respiratory burst, lysozyme activity, alternative complement pathway hemolytic activity, and bactericidal activity (P < 0.05). The expression levels of the immune-related genes in the whole blood, head kidney, and spleen were significantly increased after CH-AK application (P < 0.05), but this finding was not observed in the liver (P > 0.05). Additionally, after 14 days of A. hydrophila peritoneal injection, the fish in group C showed significantly higher survival rates of approximately 70.0 % compared with the control fish in groups B, D, and E (52.5 %, 40.0 %, 45.0 %, and 45.0 %, respectively) (P < 0.05). These results collectively suggest that short-term application of the diet fed to group C effectively boosted the immune responses and disease resistance of hybrid catfish against A. hydrophila.

Synergistic Antibiofilm Action of Cinnamomum verum and Brazilian Green Propolis Hydroethanolic Extracts against Multidrug-Resistant Strains of Acinetobacter baumannii and Pseudomonas aeruginosa and Their Biocompatibility on Human Keratinocytes.

The accumulated dental biofilm can be a source of oral bacteria that are aspirated into the lower respiratory tract causing ventilator-associated pneumonia in hospitalized patients. The aim of this study was to evaluate the synergistic antibiofilm action of the produced and phytochemically characterized extracts of Cinnamomum verum and Brazilian green propolis (BGP) hydroethanolic extracts against multidrug-resistant clinical strains of Acinetobacter baumannii and Pseudomonas aeruginosa, in addition to their biocompatibility on human keratinocyte cell lines (HaCaT). For this, High-performance liquid chromatography analysis of the plant extracts was performed; then the minimum inhibitory and minimum bactericidal concentrations of the extracts were determined; and antibiofilm activity was evaluated with MTT assay to prevent biofilm formation and to reduce the mature biofilms. The cytotoxicity of the extracts was verified using the MTT colorimetric test, evaluating the cellular enzymatic activity. The data were analyzed with one-way ANOVA and Tukey's tests as well as Kruskal-Wallis and Dunn's tests, considering a significance level of 5%. It was possible to identify the cinnamic aldehyde in C. verum and p-coumaric, caffeic, and caffeoylquinic acids as well as flavonoids such as kaempferol and kaempferide and Artepillin-C in BGP. The combined extracts were effective in preventing biofilm formation and reducing the mature biofilms of A. baumannii and P. aeruginosa. Moreover, both extracts were biocompatible in different concentrations. Therefore, C. verum and BGP hydroethanolic extracts have bactericidal and antibiofilm action against multidrug resistant strains of A. baumannii and P. aeruginosa. In addition, the combined extracts were capable of expressively inhibiting the formation of A. baumannii and P. aeruginosa biofilms (prophylactic effect) acting similarly to 0.12% chlorhexidine gluconate.

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

INTRODUCTION: This study examines the role of mesenchymal stem cells (MSCs) in an experimental sepsis model developed with colistin-resistant Acinetobacter baumannii (CRAB). MATERIALS AND METHODS: 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. RESULTS: 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. CONCLUSION: 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.

Co-culture of Acinetobacter sp. and Scedosporium sp. immobilized beads for optimized biosurfactant production and degradation of crude oil.

The widespread exploration and exploitation of crude oil has increased the prevalence of petroleum hydrocarbon pollution in the marine and coastal environment. Bioremediation of petroleum hydrocarbons using cell immobilization techniques is gaining increasing attention. In this study, the crude oil degradation performance of bacterial and fungal co-culture was optimized by entrapping both cells in sodium-alginate and polyvinyl alcohol composite beads. Results indicate that fungal cells remained active after entrapment and throughout the experiment, while bacterial cells were non-viable at the end of the experimental period in treatments with the bacterial-fungal ratio of 1:2. A remarkable decrease in surface tension from 72 mN/m to 36.51 mN/m was achieved in treatments with the bacterial-fungal ratio of 3:1. This resulted in a significant (P < 0.05) total petroleum hydrocarbon (TPH) removal rate of 89.4%, and the highest degradation of n-alkanes fractions (from 2129.01 mg/L to 118.53 mg/L), compared to the other treatments. Whereas PAHs removal was highest in treatments with the most fungal abundance (from 980.96 µg/L to 177.3 µg/L). Furthermore, enzymes analysis test revealed that catalase had the most effect on microbial degradation of the target substrate, while protease had no significant impact on the degradation process. High expression of almA and PAH-RHDa genes was achieved in the co-culture treatments, which correlated significantly (P < 0.05) with n-alkanes and PAHs removal, respectively. These results indicate that the application of immobilized bacterial and fungal cells in defined co-culture systems is an effective strategy for enhanced biodegradation of petroleum hydrocarbons in aqueous systems.

Comparative transcriptomics analysis of multidrug-resistant Acinetobacter baumannii in response to treatment with the terpenic compounds thymol and carvacrol.

Acinetobacter baumannii is a gram-negative opportunistic bacterium that has become a major public health concern and a substantial medical challenge due to its ability to acquire multidrug resistance (MDR), extended-drug resistance, or pan-drug resistance. In this study, we evaluated the antibacterial activity of thymol and carvacrol alone or in combination against clinical isolates of MDR A. baumannii. Additionally, we used RNA-sequency to perform a comparative transcriptomic analysis of the effects of carvacrol and thymol on the Acb35 strain under different treatment conditions. Our results demonstrated that thymol and carvacrol alone, effectively inhibited the bacterial growth of MDR A. baumannii isolates, with a minimum inhibitory concentration (MIC) lower than 500 µg/mL. Furthermore, the combination of thymol and carvacrol exhibited either synergistic (FICI ≤ 0.5) or additive effects (0.5 < FICI ≤ 4), enhancing their antibacterial activity. Importantly, these compounds were found to be non-cytotoxic to Vero cells and did not cause hemolysis in erythrocytes at concentrations that effectively inhibited bacterial growth. Transcriptomic analysis revealed the down-regulation of mRNA associated with ribosomal subunit assemblies under all experimental conditions tested. However, the up-regulation of specific genes encoding stress response proteins and transcriptional regulators varied depending on the experimental condition, particularly in response to the treatment with carvacrol and thymol in combination. Based on our findings, thymol and carvacrol demonstrate promising potential as chemotherapeutic agents for controlling MDR A. baumannii infections. These compounds exhibit strong antibacterial activity, particularly in combination and lower cytotoxicity towards mammalian cells. The observed effects on gene expression provide insights into the underlying mechanisms of action, highlighting the regulation of stress response pathways.

Induced Heteroresistance in Carbapenem-Resistant Acinetobacter baumannii (CRAB) via Exposure to Human Pleural Fluid (HPF) and Its Impact on Cefiderocol Susceptibility.

Infections caused by Carbapenem-resistant Acinetobacter baumannii (CRAB) isolates, such as hospital-acquired pneumonia (HAP), bacteremia, and skin and soft tissue infections, among others, are particularly challenging to treat. Cefiderocol, a chlorocatechol-substituted siderophore antibiotic, was approved by the U.S. Food and Drug Administration (FDA) in 2019 and prescribed for the treatment of CRAB infections. Despite the initial positive treatment outcomes with this antimicrobial, recent studies reported a higher-than-average all-cause mortality rate in patients treated with cefiderocol compared to the best available therapy. The cause(s) behind these outcomes remains unconfirmed. A plausible hypothesis is heteroresistance, a phenotype characterized by the survival of a small proportion of cells in a population that is seemingly isogenic. Recent results have demonstrated that the addition of human fluids to CRAB cultures leads to cefiderocol heteroresistance. Here, we describe the molecular and phenotypic analyses of CRAB heteroresistant bacterial subpopulations to better understand the nature of the less-than-expected successful outcomes after cefiderocol treatment. Isolation of heteroresistant variants of the CRAB strain AMA40 was carried out in cultures supplemented with cefiderocol and human pleural fluid (HPF). Two AMA40 variants, AMA40 IHC1 and IHC2, were resistant to cefiderocol. To identify mutations and gene expression changes associated with cefiderocol heteroresistance, we subjected these variants to whole genome sequencing and global transcriptional analysis. We then assessed the impact of these mutations on the pharmacodynamic activity of cefiderocol via susceptibility testing, EDTA and boronic acid inhibition analysis, biofilm formation, and static time-kill assays. Heteroresistant variants AMA40 IHC1 and AMA40 IHC2 have 53 chromosomal mutations, of which 40 are common to both strains. None of the mutations occurred in genes associated with high affinity iron-uptake systems or ß-lactam resistance. However, transcriptional analyses demonstrated significant modifications in levels of expression of genes associated with iron-uptake systems or ß-lactam resistance. The blaNDM-1 and blaADC-2, as well as various iron-uptake system genes, were expressed at higher levels than the parental strain. On the other hand, the carO and ompA genes' expression was reduced. One of the mutations common to both heteroresistant strains was mapped within ppiA, a gene associated with iron homeostasis in other species. Static time-kill assays demonstrated that supplementing cation-adjusted Mueller-Hinton broth with human serum albumin (HAS), the main protein component of HPF, considerably reduced cefiderocol killing activity for all three strains tested. Notably, collateral resistance to amikacin was observed in both variants. We conclude that exposing CRAB to fluids with high HSA concentrations facilitates the rise of heteroresistance associated with point mutations and transcriptional upregulation of genes coding for ß-lactamases and biofilm formation. The findings from this study hold significant implications for understanding the emergence of CRAB resistance mechanisms against cefiderocol treatment. This understanding is vital for the development of treatment guidelines that can effectively address the challenges posed by CRAB infections.

Synergistic combination of carvedilol, amlodipine, amitriptyline, and antibiotics as an alternative treatment approach for the susceptible and multidrug-resistant A. baumannii infections via drug repurposing.

We evaluated in vitro activity of 13 drugs used in the treatment of some non-communicable diseases via repurposing to determine their potential use in the treatment of Acinetobacter baumannii infections caused by susceptible and multidrug-resistant strains. A. baumannii is a multidrug-resistant Gram-negative bacteria causing nosocomial infections, especially in intensive care units. It has been identified in the WHO critical pathogen list and this emphasises urgent need for new treatment options. As the development of new therapeutics is expensive and time consuming, finding new uses of existing drugs via drug repositioning has been favoured. Antimicrobial susceptibility tests were conducted on all 13 drugs according to CLSI. Drugs with MIC values below 128 µg/mL and control antibiotics were further subjected to synergetic effect and bacterial time-kill analysis. Carvedilol-gentamicin (FICI 0.2813) and carvedilol-amlodipine (FICI 0.5625) were determined to have synergetic and additive effect, respectively, on the susceptible A. baumannii strain, and amlodipine-tetracycline (FICI 0.75) and amitriptyline-tetracycline (FICI 0.75) to have additive effect on the multidrug-resistant A. baumannii strain. Most remarkably, both amlodipine and amitriptyline reduced the MIC of multidrug-resistant, including some carbapenems, A. baumannii reference antibiotic tetracycline from 2 to 0.5 µg/mL, for 4-folds. All these results were further supported by bacterial time-kill assay and all combinations showed bactericidal activity, at certain hours, at 4XMIC. Combinations proposed in this study may provide treatment options for both susceptible and multidrug-resistant A. baumannii infections but requires further pharmacokinetics and pharmacodynamics analyses and in vivo re-evaluations using appropriate models.