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An understanding of why certain Acinetobacter species are more successful in causing nosocomial infections, transmission and epidemic spread in healthcare institutions compared with other species is lacking. We used genomic, phenotypic and virulence studies to identify differences between Acinetobacter species. Fourteen strains representing nine species were examined. Genomic analysis of six strains showed that the A. baumannii core genome contains many genes important for diverse metabolism and survival in the host. Most of the A. baumannii core genes were also present in one or more of the less clinically successful species. In contrast, when the accessory genome of an individual A. baumannii strain was compared to a strain of a less successful species (A. calcoaceticus RUH2202), many operons with putative virulence function were found to be present only in the A. baumannii strain, including the csu operon, the acinetobactin chromosomal cluster, and bacterial defence mechanisms. Phenotype microarray analysis showed that compared to A. calcoaceticus (RUH2202), A. baumannii ATCC 19606(T) was able to utilise nitrogen sources more effectively and was more tolerant to pH, osmotic and antimicrobial stress. Virulence differences were also observed, with A. baumannii ATCC 19606(T), A. pittii SH024, and A. nosocomialis RUH2624 persisting and forming larger biofilms on human skin than A. calcoaceticus. A. baumannii ATCC 19606(T) and A. pittii SH024 were also able to survive in a murine thigh infection model, whereas the other two species were eradicated. The current study provides important insights into the elucidation of differences in clinical relevance among Acinetobacter species.

The Acinetobacter calcoaceticus-baumannii complex is a significant cause of nosocomial infections. While A. baumannii is the most common and clinically significant member globally, recent studies suggest that A. pittii is an emerging pathogen within the complex. Clinical A. pittii isolates have not been well characterized. In this study, we investigated the genomic and phenotypic characteristics of six clinical A. pittii isolates from five healthcare facilities. All isolates were resistant to ceftazidime and ertapenem, and one was resistant to imipenem and meropenem. Using whole genome sequencing and phylogenetic analysis, we found that isolates were not linked genetically, harbored OXA-type β-lactamase genes, and exhibited a wide array of virulence factors. Plasmids were identified in two isolates. Transfer of a plasmid carrying bla OXA−72 from AP290R to a carbapenem-susceptible A. bauma nnii recipient strain conferred carbapenem resistance. A. pittii isolates varied in motility and biofilm production. When tested in a Zophobas morio survival model, the carbapenem-resistant strain AP290R showed high virulence and led to increased mortality, reaching 70% within 24 h post-infection. This study highlights the potential of A. pittii to serve as a reservoir of carbapenem resistance and cause severe infections. Infection control efforts to limit the spread of Acinetobacter calcoaceticus-baumannii complex should include A. pittii.

Acinetobacter baumannii is an emergent and global nosocomial pathogen. In addition to A. baumannii, other Acinetobacter species, especially those in the Acinetobacter calcoaceticus-baumannii (Acb) complex, have also been associated with serious human infection. Although mechanisms of attachment, persistence on abiotic surfaces, and pathogenesis in A. baumannii have been identified, the genetic mechanisms that explain the emergence of A. baumannii as the most widespread and virulent Acinetobacter species are not fully understood. Recent whole genome sequencing has provided insight into the phylogenetic structure of the genus Acinetobacter. However, a global comparison of genomic features between Acinetobacter spp. has not been described in the literature. In this study, 136 Acinetobacter genomes, including 67 sequenced in this study, were compared to identify the acquisition and loss of genes in the expansion of the Acinetobacter genus. A whole genome phylogeny confirmed that A. baumannii is a monophyletic clade and that the larger Acb complex is also a well-supported monophyletic group. The whole genome phylogeny provided the framework for a global genomic comparison based on a blast score ratio (BSR) analysis. The BSR analysis demonstrated that specific genes have been both lost and acquired in the evolution of A. baumannii. In addition, several genes associated with A. baumannii pathogenesis were found to be more conserved in the Acb complex, and especially in A. baumannii, than in other Acinetobacter genomes; until recently, a global analysis of the distribution and conservation of virulence factors across the genus was not possible. The results demonstrate that the acquisition of specific virulence factors has likely contributed to the widespread persistence and virulence of A. baumannii. The identification of novel features associated with transcriptional regulation and acquired by clades in the Acb complex presents targets for better understanding the evolution of pathogenesis and virulence in the expansion of the genus.

Acinetobacter baumannii and Acinetobacter nosocomialis are the imperious pathogens in the intensive care units. We aimed to explore the genomic features of these pathogens to understand the factors influencing their plasticity. Using next-generation sequencing, two carbapenem-resistant A. baumannii (AbaBS-3, AbaETR-4) isolates and a pan-susceptible A. nosocomialis (AbaAS-5) isolate were characterised. All genomes exhibited 94% similarity with a degree of heterogeneity. AbaBS-3 and AbaETR-4 harboured antibiotic resistance gene (ARG) repertoire to most antibiotic classes. Carbapenem resistance was due to blaOXA-23 and blaOXA-66 besides the antibiotic efflux pumps. Diverse mobile genetic elements (MGE), insertion sequences (IS), prophages and virulence determinants with a plethora of stress response genes were identified in all three genomes. Class-1 integron in AbaETR-4, encoded genes that confer resistance to aminoglycosides, phenicol, sulfonamides and disinfectants. Substitutions in LpxACD and PmrCAB of AbaETR-4 confirmed the colistin resistance in vitro. Novel mutations in piuA, responsible for transporting cefiderocol, were found in AbaBS-3 and AbaETR-4. Plasmids carrying toxin-antitoxin systems, ARGs and ISs were present in these genomes. All three genomes harboured diverse protein secretion systems, virulence determinants related to immune evasion, adherence, biofilm formation and iron acquisition systems. AbaAS-5 exclusively harboured serine protease pkf, and CpaA substrate of type-II secretion system but lacked the acinetobactin-iron acquisition system. Our work delivers a holistic genome characterization of A. baumannii, coupled with a trailblazing attempt to study A. nosocomialis from India. The presence of ARGs and potential virulence factors interspersed with MGE is a cause for concern, depicting the dynamic adaptability mediated by genetic recombination.

The genus Acinetobacter comprises both environmental and clinically relevant species associated with hospital-acquired infections. Among them, Acinetobacter baumannii is a critical priority bacterial pathogen, for which the research and development of new strategies for antimicrobial treatment are urgently needed. Acinetobacter spp. produce a variety of structurally diverse capsular polysaccharides (CPSs), which surround the bacterial cells with a thick protective layer. These surface structures are primary receptors for capsule-specific bacteriophages, that is, phages carrying tailspikes with CPS-depolymerizing/modifying activities. Phage tailspike proteins (TSPs) exhibit hydrolase, lyase, or esterase activities toward the corresponding CPSs of a certain structure. In this study, the data on all lytic capsule-specific phages infecting Acinetobacter spp. with genomes deposited in the NCBI GenBank database by January 2024 were summarized. Among the 149 identified TSPs encoded in the genomes of 143 phages, the capsular specificity (K specificity) of 46 proteins has been experimentally determined or predicted previously. The specificity of 63 TSPs toward CPSs, produced by various Acinetobacter K types, was predicted in this study using a bioinformatic analysis. A comprehensive phylogenetic analysis confirmed the prediction and revealed the possibility of the genetic exchange of gene regions corresponding to the CPS-recognizing/degrading parts of different TSPs between morphologically and taxonomically distant groups of capsule-specific Acinetobacter phages.

The genus Acinetobacter is comprised of a diverse group of species, several of which have raised interest due to potential applications in bioremediation and agricultural purposes. In this work, we show that many species within the genus Acinetobacter possess the genetic requirements to assemble a functional type VI secretion system (T6SS). This secretion system is widespread among Gram negative bacteria, and can be used for toxicity against other bacteria and eukaryotic cells. The most studied species within this genus is A. baumannii, an emerging nosocomial pathogen that has become a significant threat to healthcare systems worldwide. The ability of A. baumannii to develop multidrug resistance has severely reduced treatment options, and strains resistant to most clinically useful antibiotics are frequently being isolated. Despite the widespread dissemination of A. baumannii, little is known about the virulence factors this bacterium utilizes to cause infection. We determined that the T6SS is conserved and syntenic among A. baumannii strains, although expression and secretion of the hallmark protein Hcp varies between strains, and is dependent on TssM, a known structural protein required for T6SS function. Unlike other bacteria, A. baumannii ATCC 17978 does not appear to use its T6SS to kill Escherichia coli or other Acinetobacter species. Deletion of tssM does not affect virulence in several infection models, including mice, and did not alter biofilm formation. These results suggest that the T6SS fulfils an important but as-yet-unidentified role in the various lifestyles of the Acinetobacter spp.

Detection of Acinetobacter baumannii has been relying primarily on bacterial culture that often fails to return useful results in time. Although DNA-based assays are more sensitive than bacterial culture in detecting the pathogen, the molecular results are often inconsistent and challenged by doubts on false positives, such as those due to system- and environment-derived contaminations. In addition, these molecular tools require expensive laboratory instruments. Therefore, establishing molecular tools for field use require simpler molecular platforms. The loop-mediated isothermal amplification method is relatively simple and can be improved for better use in a routine clinical bacteriology laboratory. A simple and portable device capable of performing both the amplification and detection (by fluorescence) of LAMP in the same platform has been developed in recent years. This method is referred to as real-time loop-mediated isothermal amplification. In this study, we attempted to utilize this method for rapid detection of A. baumannii. Species-specific primers were designed to test the utility of this method. Clinical samples of A. baumannii were used to determine the sensitivity and specificity of this system compared to bacterial culture and a polymerase chain reaction method. All positive samples isolated from sputum were confirmed to be the species of Acinetobacter by 16S rRNA gene sequencing. The RealAmp method was found to be simpler and allowed real-time detection of DNA amplification, and could distinguish A. baumannii from Acinetobacter calcoaceticus and Acinetobacter genomic species 3. DNA was extracted by simple boiling method. Compared to bacterial culture, the sensitivity and specificity of RealAmp in detecting A. baumannii was 98.9% and 75.0%, respectively. The RealAmp assay only requires a single unit, and the assay positivity can be verified by visual inspection. Therefore, this assay has great potential of field use as a molecular tool for detection of A. baumannii.

β-lactamases represent the main mechanism of antimicrobial resistance in Gram-negative pathogens. Their catalytic function (cleaving β-lactam antibiotics) occurs in the bacterial periplasm, where they are commonly reported as soluble proteins. A bioinformatic analysis reveals a significant number of putative lipidated β-lactamases, expected to be attached to the outer bacterial membrane. Notably, 60% of Class D OXA β-lactamases (all from Acinetobacter spp.) are predicted as membrane-anchored proteins. We demonstrate that two clinically relevant carbapenemases, OXA-23 and OXA-24/40, are membrane-bound proteins in A. baumannii . This cellular localization favors the secretion of these enzymes into outer membrane vesicles that transport them outside the boundaries of the cell. β-lactamase-loaded vesicles can protect populations of antibiotic-susceptible bacteria, enabling them to thrive in the presence of β-lactam antibiotics. The ubiquity of this phenomenon suggests that it may have influenced the dissemination of resistance mediated by Acinetobacter spp., particularly in polymicrobial infections, being a potent evolutionary advantage.

poses a profound global health threat because of multidrug resistance and its association with nosocomial infections. However, standard clinical diagnostics often report it together with other species as complex (ABC), which unavoidably conceals the attribution of non- species. This study reported orthopedic infection cases associated with different species and characterized the genomes of the culture isolates to evaluate their potential impact on the clinical treatment. Nine in-patients with complex identified by culture during hospitalization were enrolled by the Orthopedics Department from a local hospital in Qingdao, China. Their clinical data were reviewed. One ABC isolate from each patient was tested for drug susceptibility and subjected for whole-genome sequencing, followed by bioinformatic analyses. Through whole-genome analysis, nine ABC isolates were identified as six , two , and one with distinct antibiotic resistance profiles and phylogenetic characteristics, indicating progressing pathogen transmission across broad geographic regions in One Health perspective. All and strains carried multidrug resistance genes, while bore only and . Phenotypically, eight isolates were susceptible to almost all the antibiotics tested, with only one being multidrug resistant. Despite this, eight patients received cephalosporins following positive reports of complex. Our study highlighted the limitation of current clinical diagnostic approaches for non- cases, which tended to be overtreated, and suggested that etiology landscape should be explored further beyond to avoid antibiotic misuse.

Acinetobacter baumannii is considered one of the most persistent pathogens responsible for nosocomial infections. Due to the emergence of multidrug resistant strains, as well as high morbidity and mortality caused by this pathogen, A. baumannii was placed on the World Health Organization (WHO) drug-resistant bacteria and antimicrobial resistance research priority list. This review summarizes current studies on mechanisms that protect A. baumannii against multiple stresses caused by the host immune response, outside host environment, and antibiotic treatment. We particularly focus on the ability of A. baumannii to survive long-term desiccation on abiotic surfaces and the population heterogeneity in A. baumannii biofilms. Insight into these protective mechanisms may provide clues for the development of new strategies to fight multidrug resistant strains of A. baumannii.