Survival and virulence of Acinetobacter baumannii in microbial mixtures.

Acinetobacter species such as A. venetianus and A. guillouiae have been studied for various biotechnology applications, including bioremediation of recalcitrant and harmful environmental contaminants, as well as bioengineering of enzymes and diagnostic materials. Bacteria used in biotechnology are often combined with other microorganisms in mixtures to formulate efficacious commercial products. However, if the mixture contained a closely related Acinetobacter pathogen such as A. baumannii (Ab), it remains unclear whether the survival and virulence of Ab would be masked or augmented. This uncertainty poses a challenge in ensuring the safety of such biotechnology products, since Ab is one of the most significant pathogens for both hospital and community -acquired infections. This research aimed to investigate the growth and virulence of Ab within a mixture of 11 bacterial species formulated as a mock microbial mixture (MM). Growth challenges with environmental stressors (i.e., temperature, pH, sodium, iron, and antibiotics) revealed that Ab could thrive under diverse conditions except in the presence of ciprofloxacin. When cultured alone, Ab exhibited significantly more growth in the presence of almost all the environmental stressors than when it was co-incubated with the MM. During the exposure of A549 lung epithelial cells to the MM, Ab growth was stimulated compared to that in standard mammalian culture media. Cytotoxicity caused by Ab was suppressed in the presence of the MM. Lymphocytes were significantly reduced in mice exposed to Ab with or without MM via intravenous injection. The levels of the splenic cytokines IL-1α, IL-1ß, MCP-1, and MIP-1α were significantly reduced 24 h after exposure to Ab + MM. This study demonstrated that the presence of the MM marginally but significantly reduced the growth and virulence of Ab, which has implications for the safety of mixtures of microorganisms for biotechnological applications. Furthermore, these findings expand our understanding of the virulence of Ab during host-pathogen interactions.

Impact of acidic and alkaline conditions on Staphylococcus aureus and Acinetobacter baumannii interactions and their biofilms.

Bacterial biofilms pose significant challenges due to their association with antibiotic resistance, metabolic adaptation, and survival under harsh conditions. Among notable pathogens forming biofilms, Staphylococcus aureus and Acinetobacter baumannii are concerning pathogens in nosocomial settings. However, their behaviour under acidic (pH 4.5) and alkaline (pH10.5) conditions, especially in co-culture setups, remains insufficiently understood. This study investigates these aspects, by examining growth rates, biofilm formation, pH shifts, phenotypic analysis, and gene expression profiles. The results showed A. baumannii exhibited reduced  growth and biofilm formation at pH 4.5, while S. aureus showed slow growth and low biofilm formation at pH10.5 in mono-cultures. S. aureus leaned towards an acidic pH (6-6.5), whereas A. baumannii shifted towards an alkaline pH (8-9). In co-culture environments, growth rates and biofilm formation increased across all pH conditions, converging towards a neutral pH over time. Phenotypic motility assays indicated that A. baumannii exhibited greater motility in alkaline conditions, while S. aureus showed increased staphyloxanthin production under acidic conditions. Gene expression analyses revealed that the fibronectin-binding protein A (FnbA) and N-acetylglucosaminyl-transferase (icaA) genes, responsible for initial attachment during biofilm formation, were highly expressed in acidic co-culture condition but poorly expressed in alkaline condition. In A. baumannii, the outer membrane protein A (OmpA) gene associated with adhesion and virulence, was upregulated in co-culture. The LuxR gene involved in quorum sensing was upregulated in acidic conditions and poorly expressed at pH 10.5. This study elucidates the metabolic adaptability and biofilm formation tendencies of S. aureus towards acidic conditions and A. baumannii towards alkaline conditions, providing insights for better management of biofilm-related infections.

Unveiling the Self-assembly and Therapeutic Efficacy of Antimicrobial Peptides SA4 Against Multidrug-Resistant A. baumannii.

Infections linked to Acinetobacter baumannii are one of the main risks of modern medicine. Biofilms formed by A. baumannii due to a protective extracellular polysaccharide matrix make them highly tolerant to conventional antibiotics and raise the possibility of antibiotic resistance. Antimicrobial peptides (AMPs) are gaining popularity due to their broad-spectrum actions and key properties of peptide self-assembly, making them a promising alternative to antibiotics. Here, we demonstrate that 12-residue synthetic self-assembled peptide SA4 nanostructures have enough antibacterial action to prevent the growth of mature bacterial biofilms. The SA4 peptide was successfully synthesized by using the solid-phase peptide synthesis method, and its self-assembly was prepared in water. The self-assembled peptide hydrogel formed nanotube structure was observed under a scanning electron microscope and further characterized to confirm their physical and molecular properties. The resulting hydrogel exhibits significant antibacterial activity against MDR A. baumannii strains (MDR-1 and MDR-2), responsible for many nosocomial infections. In addition, at various gel concentrations, this hydrogel has the potential to inhibit about 30-80% of biofilms formed by MDR strains. Furthermore, under a microscope, it has been observed that the rupture of the bacterial cell membrane and cell wall of A. baumannii cells is caused by peptide nanotubes generated by self-assemblies. Thus, peptide-based nanotubes present intriguing avenues for various biomedical applications. This is the first report of bacterial biofilm removal with SA4 peptide nanotubes, and offering a unique treatment for infections linked to biofilms.

Bactericidal effect of bacteria isolated from the marine sponges Hymeniacidon perlevis and Halichondria panicea against carbapenem-resistant Acinetobacter baumannii.

In this study, we evaluated the antimicrobial activity of bacteria isolated from the marine sponges Hymeniacidon perlevis and Halichondria panicea against seven Acinetobacter baumannii strains, the majority of which were clinically relevant carbapenem-resistant A. baumannii strains. We observed the inhibitory activity of 18 (out of 114) sponge-isolated bacterial strains against all A. baumanii strains using medium-throughput solid agar overlay assays. These inhibitory strains belonged to the genera Lactococcus, Pseudomonas, and Vagococcus. In addition, this antimicrobial activity was validated through a liquid co-cultivation challenge using an inhibitory strain of each genus and a green fluorescent protein-tagged A. baumanii strain. Fluorescence measurements indicated that the growth of A. baumanii was inhibited by the sponge isolates. In addition, the inability of A. baumanii to grow after spreading the co-cultures on solid medium allowed us to characterize the activity of the sponge isolates as bactericidal. In conclusion, this study demonstrates that marine sponges are a reservoir of bacteria that deserves to be tapped for antibiotic discovery against A. baumanii.

Investigating the Effectiveness of Ceragenins against Acinetobacter baumannii to Develop New Antimicrobial and Anti-Adhesive Strategies.

A growing body of experimental data indicates that ceragenins (CSAs), which mimic the physicochemical properties of the host's cationic antimicrobial peptide, hold promise for the development of a new group of broad-spectrum antimicrobials. Here, using a set of in vivo experiments, we assessed the potential of ceragenins in the eradication of an important etiological agent of nosocomial infections, Acinetobacter baumannii. Assessment of the bactericidal effect of ceragenins CSA-13, CSA-44, and CSA-131 on clinical isolates of A. baumannii (n = 65) and their effectiveness against bacterial cells embedded in the biofilm matrix after biofilm growth on abiotic surfaces showed a strong bactericidal effect of the tested molecules regardless of bacterial growth pattern. AFM assessment of bacterial cell topography, bacterial cell stiffness, and adhesion showed significant membrane breakdown and rheological changes, indicating the ability of ceragenins to target surface structures of A. baumannii cells. In the cell culture of A549 lung epithelial cells, ceragenin CSA-13 had the ability to inhibit bacterial adhesion to host cells, suggesting that it interferes with the mechanism of bacterial cell invasion. These findings highlight the potential of ceragenins as therapeutic agents in the development of antimicrobial strategies against bacterial infections caused by A. baumannii.

Exploring the antimicrobial potential of crude peptide extracts from Allium sativum and Allium oschaninii against antibiotic-resistant bacterial strains.

CONTEXT: Plant peptides garner attention for their potential antimicrobial properties amid the rising concern over antibiotic-resistant bacteria. OBJECTIVE: This study investigates the antibacterial potential of crude peptide extracts from 27 Thai plants collected locally. MATERIALS AND METHODS: Peptide extracts from 34 plant parts, derived from 27 Thai plants, were tested for their antimicrobial efficacy against four highly resistant bacterial strains: Streptococcus aureus MRSA, Pseudomonas aeruginosa, Acinetobacter baumannii, and Escherichia coli. The stability of these peptide extracts was examined at different temperatures, and the synergistic effects of two selected plant peptide extracts were investigated. Additionally, the time-kill kinetics of the individual extracts and their combination were determined against the tested pathogens. RESULTS: Peptides from Allium sativum L. and Allium oschaninii O. Fedtsch (Amaryllidaceae) were particularly potent, inhibiting bacterial growth with MICs ranging from 1.43 to 86.50 µg/mL. The consistent MICs and MBCs of these extracts across various extraction time points highlight their reliability. Stability tests reveal that these peptides maintain their antimicrobial activity at -20 °C for over a month, emphasizing their durability for future exploration and potential applications in addressing antibiotic resistance. Time-kill assays elucidate the time and concentration-dependent nature of these antimicrobial effects, underscoring their potent initial activity and sustained efficacy over time. DISCUSSION AND CONCLUSIONS: This study highlights the antimicrobial potential of Allium-derived peptides, endorsing them for combating antibiotic resistance and prompting further investigation into their mechanisms.

Bacterial behavior in human blood reveals complement evaders with some persister-like features.

Bacterial bloodstream infections (BSI) are a major health concern and can cause up to 40% mortality. Pseudomonas aeruginosa BSI is often of nosocomial origin and is associated with a particularly poor prognosis. The mechanism of bacterial persistence in blood is still largely unknown. Here, we analyzed the behavior of a cohort of clinical and laboratory Pseudomonas aeruginosa strains in human blood. In this specific environment, complement was the main defensive mechanism, acting either by direct bacterial lysis or by opsonophagocytosis, which required recognition by immune cells. We found highly variable survival rates for different strains in blood, whatever their origin, serotype, or the nature of their secreted toxins (ExoS, ExoU or ExlA) and despite their detection by immune cells. We identified and characterized a complement-tolerant subpopulation of bacterial cells that we named "evaders". Evaders shared some features with bacterial persisters, which tolerate antibiotic treatment. Notably, in bi-phasic killing curves, the evaders represented 0.1-0.001% of the initial bacterial load and displayed transient tolerance. However, the evaders are not dormant and require active metabolism to persist in blood. We detected the evaders for five other major human pathogens: Acinetobacter baumannii, Burkholderia multivorans, enteroaggregative Escherichia coli, Klebsiella pneumoniae, and Yersinia enterocolitica. Thus, the evaders could allow the pathogen to persist within the bloodstream, and may be the cause of fatal bacteremia or dissemination, in particular in the absence of effective antibiotic treatments.

Pilus Production in Acinetobacter baumannii Is Growth Phase Dependent and Essential for Natural Transformation.

Acinetobacter baumannii is a severe threat to human health as a frequently multidrug-resistant hospital-acquired pathogen. Part of the danger from this bacterium comes from its genome plasticity and ability to evolve quickly by taking up and recombining external DNA into its own genome in a process called natural competence for transformation. This mode of horizontal gene transfer is one of the major ways that bacteria can acquire new antimicrobial resistances and toxic traits. Because these processes in A. baumannii are not well studied, we herein characterized new aspects of natural transformability in this species that include the species' competence window. We uncovered a strong correlation with a growth phase-dependent synthesis of a type IV pilus (TFP), which constitutes the central part of competence-induced DNA uptake machinery. We used bacterial genetics and microscopy to demonstrate that the TFP is essential for the natural transformability and surface motility of A. baumannii, whereas pilus-unrelated proteins of the DNA uptake complex do not affect the motility phenotype. Furthermore, TFP biogenesis and assembly is subject to input from two regulatory systems that are homologous to Pseudomonas aeruginosa, namely, the PilSR two-component system and the Pil-Chp chemosensory system. We demonstrated that these systems affect not only the piliation status of cells but also their ability to take up DNA for transformation. Importantly, we report on discrepancies between TFP biogenesis and natural transformability within the same genus by comparing data for our work on A. baumannii to data reported for Acinetobacter baylyi, the latter of which served for decades as a model for natural competence.IMPORTANCE Rapid bacterial evolution has alarming negative impacts on animal and human health which can occur when pathogens acquire antimicrobial resistance traits. As a major cause of antibiotic-resistant opportunistic infections, A. baumannii is a high-priority health threat which has motivated renewed interest in studying how this pathogen acquires new, dangerous traits. In this study, we deciphered a specific time window in which these bacteria can acquire new DNA and correlated that with its ability to produce the external appendages that contribute to the DNA acquisition process. These cell appendages function doubly for motility on surfaces and for DNA uptake. Collectively, we showed that A. baumannii is similar in its TFP production to Pseudomonas aeruginosa, though it differs from the well-studied species A. baylyi.

Enzymatically-crosslinked gelatin hydrogels containing paenipeptin and clarithromycin against carbapenem-resistant pathogen in murine skin wound infection.

BACKGROUND: The recent rise and spread of carbapenem-resistant pathogens pose an urgent threat to public health and has fueled the search for new therapies. Localized delivery of topical antibiotics is an alternative for the treatment of infected wounds caused by drug-resistant pathogens. In this study, we aimed to develop antimicrobial-loaded hydrogels for topical treatment of wound infections in a murine skin wound infection. RESULTS: Paenipeptin analogue 1, a linear lipopeptide, potentiated clarithromycin against multidrug-resistant Acinetobacter baumannii, Enterobacter cloacae, Escherichia coli, and Klebsiella pneumoniae. Enzymatically-crosslinked gelatin hydrogels were developed to encapsulate paenipeptin analogue 1 and clarithromycin. The encapsulated antimicrobials were gradually released from hydrogels during incubation, reaching 75.43 and 53.66% for paenipeptin and clarithromycin, respectively, at 24 h. The antimicrobial-loaded hydrogels containing paenipeptin and clarithromycin synergistically resulted in 5-log reduction in carbapenem-resistant A. baumannii within 6 h in vitro. Moreover, the antimicrobial-loaded hydrogels reduced 3.6- and 2.5-log of carbapenem-resistant A. baumannii when treated at 4 or 20 h post infection, respectively, in a murine skin wound infection. CONCLUSIONS: Enzymatically-crosslinked gelatin hydrogels loaded with paenipeptin analogue 1 and clarithromycin exhibited potent therapeutic efficacy against carbapenem-resistant A. baumannii in murine skin wound infection.

Hydrogen peroxide-producing electrochemical bandage controlled by a wearable potentiostat for treatment of wound infections.

Chronic wound infections caused by biofilm-forming microorganisms represent a major burden to healthcare systems. Treatment of chronic wound infections using conventional antibiotics is often ineffective due to the presence of bacteria with acquired antibiotic resistance and biofilm-associated antibiotic tolerance. We previously developed an electrochemical scaffold that generates hydrogen peroxide (H2 O2 ) at low concentrations in the vicinity of biofilms. The goal of this study was to transition our electrochemical scaffold into an H2 O2 -generating electrochemical bandage (e-bandage) that can be used in vivo. The developed e-bandage uses a xanthan gum-based hydrogel to maintain electrolytic conductivity between e-bandage electrodes and biofilms. The e-bandage is controlled using a lightweight, battery-powered wearable potentiostat suitable for use in animal experiments. We show that e-bandage treatment reduced colony-forming units of Acinetobacter buamannii biofilms (treatment vs. control) in 12 h (7.32 ± 1.70 vs. 9.73 ± 0.09 log10 [CFU/cm2 ]) and 24 h (4.10 ± 12.64 vs. 9.78 ± 0.08 log10 [CFU/cm2 ]) treatments, with 48 h treatment reducing viable cells below the limit of detection of quantitative and broth cultures. The developed H2 O2 -generating e-bandage was effective against in vitro A. baumannii biofilms and should be further evaluated and developed as a potential alternative to topical antibiotic treatment of wound infections.

Performance of VITEK 2, E-test, Kirby-Bauer disk diffusion, and modified Kirby-Bauer disk diffusion compared to reference broth microdilution for testing tigecycline susceptibility of carbapenem-resistant K. pneumoniae and A. baumannii in a multicenter study in China.

Tigecycline is an alternative antibiotic for managing carbapenem-resistant Gram-negative bacterial infections. However, disk diffusion and automated testing often show false-intermediate or false-resistant results in tigecycline susceptibility, misleading clinical antimicrobial therapy. Broth microdilution (BMD) is the reference method for testing tigecycline susceptibility, but it is labor intensive and time consuming to perform in clinical laboratories. Therefore, a simple and accurate method is urgently needed. We evaluated the performance of VITEK 2, E-test, Kirby-Bauer disk diffusion (KB), and modified KB disk diffusion (mKB) versus BMD in testing tigecycline susceptibility of 372 strains of carbapenem-resistant Klebsiella pneumoniae (CRKP) and 346 strains of carbapenem-resistant Acinetobacter baumannii (CRAB). BMD confirmed that 96.8% of CRKP and 91% of CRAB strains were susceptible to tigecycline. E-test, VITEK 2, KB, and mKB yielded categorical agreement of 96.7/59.3%, 69.9/54.3%, 78.5/87.3%, and 96.5%/91% for CRKP/CRAB, respectively. No very major error was found for either CRKP or CRAB by any method. No major error was found for CRKP or CRAB by the mKB method. The mKB method enhanced by R-buffer is simple, accurate, and inexpensive for clinical laboratories to test the susceptibility of CRKP and CRAB isolates to tigecycline.

Octopromycin: Antibacterial and antibiofilm functions of a novel peptide derived from Octopus minor against multidrug-resistant Acinetobacter baumannii.

The emergence of carbapenem-resistant Acinetobacter baumannii has increased the risk of nosocomial infections, which pose a huge health threat. There is an urgent need to develop alternative therapies, including broad-spectrum antimicrobial peptides. In this study, we designed, characterized, and studied the antibacterial, antibiofilm effects and possible mode of actions of a novel synthetic peptide Octopromycin, derived from the proline-rich protein 5 of Octopus minor. Octopromycin consists of 38 amino acids, (+5) net positive charge, high hydrophobic residue ratio (36%), and two α-helix secondary structures. The minimum inhibitory concentration and minimum bactericidal concentration against A. baumannii were 50 and 200 µg/mL, respectively. Time-kill kinetics and bacterial viability assays confirmed the concentration-dependent antibacterial activity of Octopromycin. Field emission scanning electron microscopy images clearly showed ultrastructural alterations in Octopromycin-treated A. baumannii cells. Propidium iodide penetrated into Octopromycin-treated A. baumannii cells, demonstrating the loss of cell membrane integrity. Octopromycin treatment increased the production of reactive oxygen species in a concentration-dependent manner, and it inhibited the biofilm formation and showed biofilm eradication activity against A. baumannii. In vitro and in vivo safety evaluation revealed that Octopromycin was nontoxic to HEK293T and Raw 264.7 cells (<400 µg/mL), as well as mice red blood cells (<300 µg/mL), and zebrafish embryos (<4 µg/mL). An in vivo study results revealed that the A. baumannii-infected fish treated with Octopromycin exhibited a significantly higher relative percent survival (37.5%) than the infected mock-treated fish with PBS (16.6%). Furthermore, a decreased bacterial load and fewer alterations in histological analysis confirmed the successful control of A. baumannii by Octopromycin in vivo. Collectively, the results indicate that the antibacterial peptide Octopromycin may achieve rapid control of A. baumannii through multi-target interactions; it presents a desirable therapeutic option for the prevention and control of the infections.

In vitroassessment of green polyhydroxybutyrate/chitosan blend loaded with kaempferol nanocrystals as a potential dressing for infected wounds.

Despite the major medical advancements in recent decades, treating infected wounds successfully remains a challenge. In this research, a functional blend of Polyhydroxybutyrate (PHB) and Chitosan (Cs) was developed for wound infection mitigation with tailored biological and physicochemical properties. Water insoluble kaempferol (KPF) was pre-formulated to water soluble KPF nanocrystals (KPF-NCs) with fine particle size of 145 ± 11 nm, and high colloidal stability (-31 ± 0.4 mV) to improve its drug transdermal delivery. PHB-Cs-KPF-NCs (1:2 ratio) film owned the best physical properties in terms of high breathability, thermal stability and mechanical strength (33 ± 1 MPa). Besides, XRD and FTIR findings indicated the interaction between Cs, PHB and KPF, reducing the film crystallinity. The scanning electron microscopy of the film displayed a highly interconnected porous morphology. KPF-NCs were integrated in PHB-Cs matrix with a marked encapsulation efficiency of 96.6%. The enhanced drug-loading film showed a sustain release pattern of KPF-NCs over 48 h. Interestingly, the developed blend possessed an impressive blood clotting capacity within 20 min. Furthermore, we presented a new naturally-sourced mixture of Cs+KPF-NCs with powerful antibacterial effects against MDRStaphylococcus aureusandAcentibacter baumanniiat very low concentrations. The membrane evidenced a remarkable antibacterial naturein vitrowith almost 100% cell viability reduction against the study strains after 48 h. By virtue of these advantages, this green blend is highly proposed for optimal wound care.

Antimicrobial activity of Actinobacteria isolated from dry land soil in Yazd, Iran.

Historically, many important secondary metabolites including antibiotics used in clinic are purified from the cultural broths of Actinobacteria, which were inhabited in soil. Yazd is located in the center of Iran, the south of the Dasht-e Kavir and the west of the Dasht-e Lut; accordingly it has a hot, dry climate with long summers. In the present study, 18 strains of Actinobacteria isolated from 60 soil samples from Yazd-Iran. Pure isolates were screened for antibacterial activity against the ATCC strains by using two methods: single line streak method and spot inoculation method. ATCC strains include four antibiotic resistant ATCC strains (Enterococcus faecalis, Staphylococcus aureus, Klebsiella pneumoniae and, Acinetobacter baumannii) and three antibiotic sensitive strains (Pseudomonas aeruginosa, Staphylococcus aureus, Escherichia coli) and, Bacillus subtilis. Seven isolates exhibited antimicrobial activity against the ATCC strains (38.8%). Identification of type I and type II polyketide synthases (pksI, pksII) and nonribosomal peptide synthetase (NRPS) genes were done for these 7 isolates and all of 7 strains, possessed at least one of these genes. The results of this study confirm that soil Actinobacteria bear a great ability to produce antibacterial compounds against resistant and sensitive test organisms.

Acinetobacter spp. in food and drinking water - A review.

Acinetobacter spp. has emerged as a pathogen of major public health concern due to their increased resistance to antibiotics and their association with a wide range of nosocomial infections, community-acquired infections and war and natural disaster-related infections. It is recognized as a ubiquitous organism however, information about the prevalence of different pathogenic species of this genus in food sources and drinking water is scarce. Since the implementation of molecular techniques, the role of foods as a source of several species, including the Acinetobacter baumannii group, has been elucidated. Multidrug resistance was also detected among Acinetobacter spp. isolated from food products. This highlights the importance of foods as potential sources of dissemination of Acinetobacter spp. between the community and clinical environments and reinforces the need for further investigations on the potential health risks of Acinetobacter spp. as foodborne pathogens. The aim of this review was to summarize the published data on the occurrence of Acinetobacter spp. in different food sources and drinking water. This information should be taken into consideration by those responsible for infection control in hospitals and other healthcare facilities.

Profiling of Potential Antibacterial Compounds of Lactic Acid Bacteria against Extremely Drug Resistant (XDR) Acinetobacter baumannii.

A total of 20 of isolates of lactic acid bacteria (LAB) were selected and screened for antagonistic activity against clinical strains of 30 clinical isolates of extremely drug-resistant (XDR) Acinetobacter baumannii using the well diffusion assay method. Results showed that 50% of the highly LAB strains possessed inhibitory activity against (up to 66%) of the XDR A. baumannii strains tested. The supernatant of the twenty LAB strains was subjected to gas chromatography mass spectrometry (GCMS) revealed that the common compound found in the active isolates against XDR A. baumannii was 3-Isobutyl-2,3,6,7,8,8a-hexahydropyrrolo[1,2-a]pyrazine-1,4-dione, a known potential diketopiperazine group. The molecular docking study against potential antibacterial targets with selected ligands was performed to predict the binding mode of interactions, which is responsible for antibacterial activity. The docking analysis of the potent compounds supported the potential antibacterial activity exhibiting high inhibition constant and binding affinity in silico.

Role of PstS in the Pathogenesis of Acinetobacter baumannii Under Microaerobiosis and Normoxia.

Acinetobacter baumannii is a successful pathogen responsible for infections with high mortality rate. During the course of infection it can be found in microaerobic environments, which influences virulence factor expression. From a previous transcriptomic analysis of A. baumannii ATCC 17978 under microaerobiosis, we know the gene pstS is overexpressed under microaerobiosis. Here, we studied its role in A. baumannii virulence. pstS loss significantly decreased bacterial adherence and invasion into A549 cells and increased A549 cell viability. pstS loss also reduced motility and biofilm-forming ability of A. baumannii. In a peritoneal sepsis murine model, the minimum lethal dose required by A. baumannii ATCC 17978 ΔpstS was lower compared to the wild type (4.3 vs 3.2 log colony forming units/mL, respectively), and the bacterial burden in tissues and fluids was lower. Thus, the loss of the phosphate sensor PstS produced a decrease in A. baumannii pathogenesis, supporting its role as a virulence factor.

γ-Glutamyltransferase as a Novel Virulence Factor of Acinetobacter baumannii Inducing Alveolar Wall Destruction and Renal Damage in Systemic Disease.

A thorough understanding of Acinetobacter baumannii pathogenicity is the key to identifying novel drug targets. In the current study, we characterize the γ-glutamyltransferase enzyme (GGT) as a novel virulence factor. A GGT assay showed that the enzyme is secreted via the type II secretion system and results in higher extracellular activity for the hypervirulent AB5075 than the laboratory-adapted strain American Type Culture Collection 17978. Enzyme-linked immunosorbent assay revealed that the former secretes larger amounts of GGT, and a rifampicin messenger RNA stability study showed that one reason for this could be the longer AB5075 ggt transcript half-life. Infection models confirmed that GGT is required for the virulence of A. baumannii. Finally, we show that clinical isolates with significantly higher extracellular GGT activity resulted in more severe infections, and assay of immune response and tissue damage markers confirm this correlation. The current findings establish for the first time the role of the GGT in the pathogenicity of A. baumannii.

Trehalose-6-phosphate-mediated phenotypic change in Acinetobacter baumannii.

The stress protectant trehalose is synthesized in Acinetobacter baumannii from UPD-glucose and glucose-6-phosphase via the OtsA/OtsB pathway. Previous studies proved that deletion of otsB led to a decreased virulence, the inability to grow at 45°C and a slight reduction of growth at high salinities indicating that trehalose is the cause of these phenotypes. We have questioned this conclusion by producing ∆otsA and ∆otsBA mutants and studying their phenotypes. Only deletion of otsB, but not deletion of otsA or otsBA, led to growth impairments at high salt and high temperature. The intracellular concentrations of trehalose and trehalose-6-phosphate were measured by NMR or enzymatic assay. Interestingly, none of the mutants accumulated trehalose any more but the ∆otsB mutant with its defect in trehalose-6-phosphate phosphatase activity accumulated trehalose-6-phosphate. Moreover, expression of otsA in a ∆otsB background under conditions where trehalose synthesis is not induced led to growth inhibition and the accumulation of trehalose-6-phosphate. Our results demonstrate that trehalose-6-phosphate affects multiple physiological activities in A. baumannii ATCC 19606.

Molecular typing of multi-drug resistant Acinetobacter baumannii isolates from clinical and environmental specimens in three Iranian hospitals by pulsed field gel electrophoresis.

BACKGROUND: Multi-drug resistant (MDR) Acinetobacter baumannii is one of the most important causes of nosocomial infections. The purpose of this study was to identify antibiotic resistance patterns, biofilm formation and the clonal relationship of clinical and environmental isolates of A. baumannii by Pulsed Field Gel Electrophoresis method. Forty-three clinical and 26 environmental isolates of the MDR A. baumannii were collected and recognized via API 20NE. Antibiotic resistance of the isolates was assessed by the disk diffusion method, and the biofilm formation test was done by the microtiter plate method. Pulsed Field Gel Electrophoresis (PFGE) was used to assess the genomic features of the bacterial isolates. RESULTS: The resistance rate of clinical and environmental isolates against antibiotics were from 95 to 100%. The difference in antibiotic resistance rates between clinical and environmental isolates was not statistically significant (p > 0.05). Biofilm production capabilities revealed that 31 (44.9%), and 30 (43.5%) isolates had strong and moderate biofilm producer activity, respectively. PFGE typing exhibited eight different clusters (A, B, C, D, E, F, G, and H) with two significant clusters included A and G with 21 (30.4%) and 16 (23.2%) members respectively, which comprises up to 53.6% of all isolates. There was no relationship between biofilm formation and antibiotic resistance patterns with PFGE pulsotypes. CONCLUSIONS: The results show that there is a close relationship between environmental and clinical isolates of A. baumannii. Cross-contamination is also very important that occurs through daily clinical activities between environmental and clinical isolates. Therefore, in order to reduce the clonal contamination of MDR A. baumannii environmental and clinical isolates, it is necessary to use strict infection control strategies.