Hetero-antagonism of avibactam and sulbactam with cefiderocol in carbapenem-resistant Acinetobacter spp.

The emergence of Gram-negative bacteria resistant to multiple antibiotics, particularly carbapenem-resistant (CR) Acinetobacter strains, poses a significant threat globally. Despite efforts to develop new antimicrobial therapies, limited progress has been made, with only two drugs-cefiderocol and sulbactam-durlobactam-showing promise for CR-Acinetobacter infections. Cefiderocol, a siderophore cephalosporin, demonstrates promising efficacy in the treatment of Gram-negative infections. However, resistance to cefiderocol has been reported in A. baumannii. Combination therapies, such as cefiderocol with avibactam or sulbactam, show reduced MICs against cefiderocol-non-susceptible strains with in vivo efficacy, although the outcomes can be complex and species-specific. In the present work, the molecular characterization of spontaneous cefiderocol-resistant variants, a CRAB strain displaying antagonism with sulbactam and an A. lwoffii strain showing antagonism with avibactam, were studied. The results reveal intriguing insights into the underlying mechanisms, including mutations affecting efflux pumps, transcriptional regulators, and iron homeostasis genes. Moreover, gene expression analysis reveals significant alterations in outer membrane proteins, iron homeostasis, and ß-lactamases, suggesting adaptive responses to selective pressure. Understanding these mechanisms is crucial for optimizing treatment strategies and preventing adverse clinical outcomes. This study highlights the importance of preemptively assessing drug synergies to navigate the challenges posed by antimicrobial resistance in CR-Acinetobacter infections.

ShlA toxin of Serratia induces P2Y2- and α5ß1-dependent autophagy and bacterial clearance from host cells.

Serratia marcescens is an opportunistic human pathogen involved in antibiotic-resistant hospital acquired infections. Upon contact with the host epithelial cell and prior to internalization, Serratia induces an early autophagic response that is entirely dependent on the ShlA toxin. Once Serratia invades the eukaryotic cell and multiples inside an intracellular vacuole, ShlA expression also promotes an exocytic event that allows bacterial egress from the host cell without compromising its integrity. Several toxins, including ShlA, were shown to induce ATP efflux from eukaryotic cells. Here, we demonstrate that ShlA triggered a nonlytic release of ATP from Chinese hamster ovary (CHO) cells. Enzymatic removal of accumulated extracellular ATP (eATP) or pharmacological blockage of the eATP-P2Y2 purinergic receptor inhibited the ShlA-promoted autophagic response in CHO cells. Despite the intrinsic ecto-ATPase activity of CHO cells, the effective concentration and kinetic profile of eATP was consistent with the established affinity of the P2Y2 receptor and the known kinetics of autophagy induction. Moreover, eATP removal or P2Y2 receptor inhibition also suppressed the ShlA-induced exocytic expulsion of the bacteria from the host cell. Blocking α5ß1 integrin highly inhibited ShlA-dependent autophagy, a result consistent with α5ß1 transactivation by the P2Y2 receptor. In sum, eATP operates as the key signaling molecule that allows the eukaryotic cell to detect the challenge imposed by the contact with the ShlA toxin. Stimulation of P2Y2-dependent pathways evokes the activation of a defensive response to counteract cell damage and promotes the nonlytic clearance of the pathogen from the infected cell.

Antimicrobial activity of the Lacticaseibacillus rhamnosus CRL 2244 and its impact on the phenotypic and transcriptional responses in carbapenem resistant Acinetobacter baumannii.

Carbapenem-resistant Acinetobacter baumannii (CRAB) is a recognized nosocomial pathogen with limited antibiotic treatment options. Lactic acid bacteria (LAB) constitute a promising therapeutic alternative. Here we studied the antibacterial properties of a collection of LAB strains using phenotypic and transcriptomic analysis against A. baumannii clinical strains. One strain, Lacticaseibacillus rhamnosus CRL 2244, demonstrated a potent inhibitory capacity on A. baumannii with a significant killing activity. Scanning electron microscopy images showed changes in the morphology of A. baumannii with an increased formation of outer membrane vesicles. Significant changes in the expression levels of a wide variety of genes were also observed. Interestingly, most of the modified genes were involved in a metabolic pathway known to be associated with the survival of A. baumannii. The paa operon, Hut system, and fatty acid degradation were some of the pathways that were induced. The analysis reveals the impact of Lcb. rhamnosus CRL 2244 on A. baumannii response, resulting in bacterial stress and subsequent cell death. These findings highlight the antibacterial properties of Lcb. rhamnosus CRL 2244 and its potential as an alternative or complementary strategy for treating infections. Further exploration and development of LAB as a treatment option could provide valuable alternatives for combating CRAB infections.

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.

Phenotypic and transcriptional analysis of the antimicrobial effect of lactic acid bacteria on carbapenem-resistant Acinetobacter baumannii: Lacticaseibacillus rhamnosus CRL 2244 an alternative strategy to overcome resistance?"

Carbapenem-resistant Acinetobacter baumannii (CRAB) is a recognized nosocomial pathogen with limited antibiotic treatment options. Lactic acid bacteria (LAB) constitute a promising therapeutic alternative. Here we studied the antibacterial properties of a collection of LAB strains using phenotypic and transcriptomic analysis against A. baumannii clinical strains. One strain, Lacticaseibacillus rhamnosus CRL 2244, demonstrated a potent inhibitory capacity on A. baumannii with a significant killing activity. Scanning electron microscopy images showed changes in the morphology of A. baumannii with an increased formation of outer membrane vesicles. Significant changes in the expression levels of a wide variety of genes were also observed. Interestingly, most of the modified genes were involved in a metabolic pathway known to be associated with the survival of A. baumannii . The paa operon, Hut system, and fatty acid degradation were some of the pathways that were induced. The analysis reveals the impact of Lcb. rhamnosus CRL 2244 on A. baumannii response, resulting in bacterial stress and subsequent cell death. These findings highlight the antibacterial properties of Lcb. rhamnosus CRL 2244 and its potential as an alternative or complementary strategy for treating infections. Further exploration and development of LAB as a treatment option could provide valuable alternatives for combating CRAB infections.

Genomic Comparative Analysis of Two Multi-Drug Resistance (MDR) Acinetobacter baumannii Clinical Strains Assigned to International Clonal Lineage II Recovered Pre- and Post-COVID-19 Pandemic.

BACKGROUND: After the emergence of COVID-19, numerous cases of A. baumannii/SARS-CoV-2 co-infection were reported. Whether the co-infecting A. baumannii strains have distinctive characteristics remains unknown. METHODS AND RESULTS: A. baumannii AMA_NO was isolated in 2021 from a patient with COVID-19. AMA166 was isolated from a mini-BAL used on a patient with pneumonia in 2016. Both genomes were similar, but they possessed 337 (AMA_NO) and 93 (AMA166) unique genes that were associated with biofilm formation, flagellar assembly, antibiotic resistance, secretion systems, and other functions. The antibiotic resistance genes were found within mobile genetic elements. While both strains harbored the carbapenemase-coding gene blaOXA-23, only the strain AMA_NO carried blaNDM-1. Representative functions coded for by virulence genes are the synthesis of the outer core of lipooligosaccharide (OCL5), biosynthesis and export of the capsular polysaccharide (KL2 cluster), high-efficiency iron uptake systems (acinetobactin and baumannoferrin), adherence, and quorum sensing. A comparative phylogenetic analysis including 239 additional sequence type (ST) 2 representative genomes showed high similarity to A. baumannii ABBL141. Since the degree of similarity that was observed between A. baumannii AMA_NO and AMA166 is higher than that found among other ST2 strains, we propose that they derive from a unique background based on core-genome phylogeny and comparative genome analysis. CONCLUSIONS: Acquisition or shedding of specific genes could increase the ability of A. baumannii to infect patients with COVID-19.

The Iron Content of Human Serum Albumin Modulates the Susceptibility of Acinetobacter baumannii to Cefiderocol.

The mortality rates of patients infected with Acinetobacter baumannii who were treated with cefiderocol (CFDC) were not as favorable as those receiving the best available treatment for pulmonary and bloodstream infections. Previous studies showed that the presence of human serum albumin (HSA) or HSA-containing fluids, such as human serum (HS) or human pleural fluid (HPF), in the growth medium is correlated with a decrease in the expression of genes associated with high-affinity siderophore-mediated iron uptake systems. These observations may explain the complexities of the observed clinical performance of CFDC in pulmonary and bloodstream infections, because ferric siderophore transporters enhance the penetration of CFDC into the bacterial cell. The removal of HSA from HS or HPF resulted in a reduction in the minimal inhibitory concentration (MIC) of CFDC. Concomitant with these results, an enhancement in the expression of TonB-dependent transporters known to play a crucial role in transporting iron was observed. In addition to inducing modifications in iron-uptake gene expression, the removal of HSA also decreased the expression of ß-lactamases genes. Taken together, these observations suggest that environmental HSA has a role in the expression levels of select A. baumannii genes. Furthermore, the removal of iron from HSA had the same effect as the removal of HSA upon the expression of genes associated with iron uptake systems, also suggesting that at least one of the mechanisms by which HSA regulates the expression of certain genes is through acting as an iron source.

Human serum albumin (HSA) regulates the expression of histone-like nucleoid structure protein (H-NS) in Acinetobacter baumannii.

According to the Centers for Disease Control and Prevention, Acinetobacter baumannii is listed among the most threatening pathogens. A. baumannii is mainly a nosocomial pathogen with a distinctive ability to survive in multiple environments. These characteristics together with this bacterium's ability to acquire antibiotic resistance determinants make it a notorious pathogen. The presence of human serum albumin (HSA) is associated with modification of expression levels in numerous genes. The presence of HSA in the culture medium is also correlated with a reduction in levels of the global suppressor histone-like nucleoid structure protein, H-NS. Comparative transcriptome analysis of the wild type and isogenic Δhns strains cultured in lysogeny broth (LB) in the presence or absence of HSA revealed that the expression of a subset of eleven genes are modified in the Δhns cultured in LB and the wild-type strain in the presence of HSA, pointing out these genes as candidates to be regulated by the presence of HSA through H-NS. Six and five of these genes were up- or down-regulated, respectively. Three of these genes have functions in quorum sensing (acdA, kar and fadD), one in quorum quenching (aidA), two in stress response (katE, ywrO), three in metabolism (phaC, yedL1, and yedL2), one in biofilm formation (csuAB), and one in ß-oxidation of fatty acids (fadA). The regulation of these genes was assessed by: (i) transcriptional analysis and qPCR at the transcriptional level; and (ii) by determining the phenotypic characteristics of each function. The results of these studies support the hypothesis that HSA-mediated reduction of H-NS levels may be one very important regulatory circuit utilized by A. baumannii to adapt to selected environments, such as those where HSA-containing human fluids are abundant.

Acinetobacter baumannii response to cefiderocol challenge in human urine.

Cefiderocol (CFDC) is a novel chlorocatechol-substituted siderophore antibiotic approved to treat complicated urinary tract infections (cUTI) and hospital-acquired and ventilator-acquired pneumonia (HAP/VAP). Previous work determined that albumin-rich human fluids increase the minimum inhibitory concentration (MICs) of Acinetobacter baumannii against CFDC and reduce the expression of genes related to iron uptake systems. This latter effect may contribute to the need for higher concentrations of CFDC to inhibit growth. The presence of human urine (HU), which contains low albumin concentrations, did not modify MIC values of two carbapenem-resistant A. baumannii. Levels of resistance to CFDC were not modified by HU in strain AMA40 but were reduced in strain AB5075. Expanding the studies to other carbapenem-resistant A. baumannii isolates showed that the presence of HU resulted in unmodified or reduced MIC of CDFC values. The expression of piuA, pirA, bauA, and bfnH determined by qRT-PCR was enhanced in A. baumannii AMA40 and AB5075 by the presence of HU in the culture medium. All four tested genes code for functions related to recognition and transport of ferric-siderophore complexes. The effect of HU on expression of pbp1, pbp3, blaOXA-51-like, blaADC, and blaNDM-1, genes associated with resistance to ß-lactams, as well as genes coding for efflux pumps and porins was variable, showing dependence with the strain analyzed. We conclude that the lack of significant concentrations of albumin and free iron in HU makes this fluid behave differently from others we tested. Unlike other albumin rich fluids, the presence of HU does not impact the antibacterial activity of CFDC when tested against A. baumannii.

Staphylococcus aureus α-Toxin Effect on Acinetobacter baumannii Behavior.

Polymicrobial infections are more challenging to treat and are recognized as responsible for significant morbidity and mortality. It has been demonstrated that multiple Gram-negative organisms take advantage of the effects of Staphylococcus aureus α-toxin on mucosal host defense, resulting in proliferation and dissemination of the co-infecting pathogens. Through phenotypic approaches, we observed a decrease in the motility of A. baumannii A118 after exposure to cell-free conditioned media (CFCM) of S. aureus strains, USA300 and LS1. However, the motility of A. baumannii A118 was increased after exposure to the CFCM of S. aureus strains USA300 Δhla and S. aureus LSI ΔagrA. Hemolytic activity was seen in A118, in the presence of CFCM of S. aureus LS1. Further, A. baumannii A118 showed an increase in biofilm formation and antibiotic resistance to tetracycline, in the presence of CFCM of S. aureus USA300. Transcriptomic analysis of A. baumannii A118, with the addition of CFCM from S. aureus USA300, was carried out to study A. baumannii response to S. aureus' released molecules. The RNA-seq data analysis showed a total of 463 differentially expressed genes, associated with a wide variety of functions, such as biofilm formation, virulence, and antibiotic susceptibility, among others. The present results showed that A. baumannii can sense and respond to molecules secreted by S. aureus. These findings demonstrate that A. baumannii may perceive and respond to changes in its environment; specifically, when in the presence of CFCM from S. aureus.

Human Serum Proteins and Susceptibility of Acinetobacter baumannii to Cefiderocol: Role of Iron Transport.

Cefiderocol, a recently introduced antibiotic, has a chemical structure that includes a cephalosporin that targets cell wall synthesis and a chlorocatechol siderophore moiety that facilitates cell penetration by active iron transporters. Analysis of the effect that human serum, human serum albumin, and human pleural fluid had on growing Acinetobacter baumannii showed that genes related to iron uptake were down-regulated. At the same time, ß-lactamase genes were expressed at higher levels. The minimum inhibitory concentrations of this antimicrobial in A. baumannii cells growing in the presence of human serum, human serum albumin, or human pleural fluid were higher than those measured when these fluids were absent from the culture medium. These results correlate with increased expression levels of ß-lactamase genes and the down-regulation of iron uptake-related genes in cultures containing human serum, human serum albumin, or human pleural fluid. These modifications in gene expression could explain the less-than-ideal clinical response observed in patients with pulmonary or bloodstream A. baumannii infections. The exposure of the infecting cells to the host's fluids could cause reduced cefiderocol transport capabilities and increased resistance to ß-lactams. The regulation of genes that could impact the A. baumannii susceptibility to cefiderocol, or other antibacterials, is an understudied phenomenon that merits further investigation.

Involvement of the Histone-Like Nucleoid Structuring Protein (H-NS) in Acinetobacter baumannii's Natural Transformation.

Most Acinetobacter baumannii strains are naturally competent. Although some information is available about factors that enhance or reduce the frequency of the transformation of this bacterium, the regulatory elements and mechanisms are barely understood. In this article, we describe studies on the role of the histone-like nucleoid structuring protein, H-NS, in the regulation of the expression of genes related to natural competency and the ability to uptake foreign DNA. The expression levels of the natural transformation-related genes pilA, pilT, pilQ, comEA, comEC, comF, and drpA significantly increased in a Δhns derivative of A. baumannii A118. The complementation of the mutant with a recombinant plasmid harboring hns restored the expression levels of six of these genes (pilT remained expressed at high levels) to those of the wild-type strain. The transformation frequency of the A. baumannii A118 Δhns strain was significantly higher than that of the wild-type. Similar, albeit not identical, there were consequences when hns was deleted from the hypervirulent A. baumannii AB5075 strain. In the AB5075 complemented strain, the reduction in gene expression in a few cases was not so pronounced that it reached wild-type levels, and the expression of comEA was enhanced further. In conclusion, the expression of all seven transformation-related genes was enhanced after deleting hns in A. baumannii A118 and AB5075, and these modifications were accompanied by an increase in the cells' transformability. The results highlight a role of H-NS in A. baumannii's natural competence.

Effect of Serum Albumin, a Component of Human Pleural Fluid, on Transcriptional and Phenotypic Changes on Acinetobacter baumannii A118.

Acinetobacter baumannii is a multidrug-resistant pathogen that causes numerous infections associated with high mortality rates. Exposure to human body fluids, such as human pleural fluid (HPF) and human serum, modulates gene expression in A. baumannii, leading to changes in its pathogenic behavior. Diverse degrees of effects at the transcriptional level were observed in susceptible and carbapenem-resistant strains. The transcriptional analysis of AB5075, a hyper-virulent and extensively drug-resistant strain showed changes in genes associated with quorum sensing, quorum quenching, fatty acids metabolism, and high-efficient iron uptake systems. In addition, the distinctive role of human serum albumin (HSA) as a critical component of HPF was evidenced. In the present work, we used model strain to analyze more deeply into the contribution of HSA in triggering A. baumannii's response. By qRT-PCR analysis, changes in the expression level of genes associated with quorum sensing, biofilm formation, and phenylacetic acid pathway were observed. Phenotypic approaches confirmed the transcriptional response. HSA, a predominant component of HPF, can modulate the expression and behavior of genes not only in a hyper-virulent and extensively drug-resistant A. baumannii model, but also in other strains with a different degree of susceptibility and pathogenicity.

Histone-like nucleoid-structuring protein (H-NS) regulatory role in antibiotic resistance in Acinetobacter baumannii.

In the multidrug resistant (MDR) pathogen Acinetobacter baumannii the global repressor H-NS was shown to modulate the expression of genes involved in pathogenesis and stress response. In addition, H-NS inactivation results in an increased resistance to colistin, and in a hypermotile phenotype an altered stress response. To further contribute to the knowledge of this key transcriptional regulator in A. baumannii behavior, we studied the role of H-NS in antimicrobial resistance. Using two well characterized A. baumannii model strains with distinctive resistance profile and pathogenicity traits (AB5075 and A118), complementary transcriptomic and phenotypic approaches were used to study the role of H-NS in antimicrobial resistance, biofilm and quorum sensing gene expression. An increased expression of genes associated with ß-lactam resistance, aminoglycosides, quinolones, chloramphenicol, trimethoprim and sulfonamides resistance in the Δhns mutant background was observed. Genes codifying for efflux pumps were also up-regulated, with the exception of adeFGH. The wild-type transcriptional level was restored in the complemented strain. In addition, the expression of biofilm related genes and biofilm production was lowered when the transcriptional repressor was absent. The quorum network genes aidA, abaI, kar and fadD were up-regulated in Δhns mutant strains. Overall, our results showed the complexity and scope of the regulatory network control by H-NS (genes involved in antibiotic resistance and persistence). These observations brings us one step closer to understanding the regulatory role of hns to combat A. baumannii infections.

Interaction of Acinetobacter baumannii with Human Serum Albumin: Does the Host Determine the Outcome?

Acinetobacter baumannii has become a serious threat to human health due to its extreme antibiotic resistance, environmental persistence, and capacity to survive within the host. Two A. baumannii strains, A118 and AB5075, commonly used as model systems, and three carbapenem-resistant strains, which are becoming ever more dangerous due to the multiple drugs they can resist, were exposed to 3.5% human serum albumin (HSA) and human serum (HS) to evaluate their response with respect to antimicrobial resistance, biofilm formation, and quorum sensing, all features responsible for increasing survival and persistence in the environment and human body. Expression levels of antibiotic resistance genes were modified differently when examined in different strains. The cmlA gene was upregulated or downregulated in conditions of exposure to 3.5% HSA or HS depending on the strain. Expression levels of pbp1 and pbp3 tended to be increased by the presence of HSA and HS, but the effect was not seen in all strains. A. baumannii A118 growing in the presence of HS did not experience increased expression of these genes. Aminoglycoside-modifying enzymes were also expressed at higher or lower levels in the presence of HSA or HS. Still, the response was not uniform; in some cases, expression was enhanced, and in other cases, it was tapered. While A. baumannii AB5075 became more susceptible to rifampicin in the presence of 3.5% HSA or HS, strain A118 did not show any changes. Expression of arr2, a gene involved in resistance to rifampicin present in A. baumannii AMA16, was expressed at higher levels when HS was present in the culture medium. HSA and HS reduced biofilm formation and production of N-Acyl Homoserine Lactone, a compound intimately associated with quorum sensing. In conclusion, HSA, the main component of HS, stimulates a variety of adaptative responses in infecting A. baumannii strains.

Interplay between Meropenem and Human Serum Albumin on Expression of Carbapenem Resistance Genes and Natural Competence in Acinetobacter baumannii.

Acinetobacter baumannii A118, a carbapenem-susceptible strain, and AB5075, carbapenem resistant, were cultured in lysogeny broth (LB) or LB with different supplements, such as 3.5% human serum albumin (HSA), human serum (HS), meropenem, or meropenem plus 3.5% HSA. Natural transformation levels were enhanced in A. baumannii A118 and AB5075 cultured in medium supplemented with 3.5% HSA. Addition of meropenem plus 3.5% HSA caused synergistic enhancement of natural transformation in A. baumannii A118. Medium containing 3.5% HSA or meropenem enhanced the expression levels of the competence and type IV pilus-associated genes. The combination meropenem plus 3.5% HSA produced a synergistic enhancement in the expression levels of many of these genes. The addition of HS, which has a high content of HSA, was also an inducer of these genes. Cultures in medium supplemented with HS or 3.5% HSA also affected resistance genes, which were expressed at higher or lower levels depending on the modification required to enhance resistance. The inducing or repressing activity of these modulators also occurred in three more carbapenem-resistant strains tested. An exception was the A. baumannii AMA16 blaNDM-1 gene, which was repressed in the presence of 3.5% HSA. In conclusion, HSA produces an enhancement of natural transformation and a modification in expression levels of competence genes and antibiotic resistance. Furthermore, when HSA is combined with carbapenems, which may increase the stress response, the expression of genes involved in natural competence is increased in A. baumannii. This process may favor the acquisition of foreign DNA and accelerate evolution.

Human Pleural Fluid and Human Serum Albumin Modulate the Behavior of a Hypervirulent and Multidrug-Resistant (MDR) Acinetobacter baumannii Representative Strain.

Acinetobacter baumannii is a nosocomial pathogen capable of causing serious infections associated with high rates of morbidity and mortality. Due to its antimicrobial drug resistance profile, A. baumannii is categorized as an urgent priority pathogen by the Centers for Disease Control and Prevention in the United States and a priority group 1 critical microorganism by the World Health Organization. Understanding how A. baumannii adapts to different host environments may provide critical insights into strategically targeting this pathogen with novel antimicrobial and biological therapeutics. Exposure to human fluids was previously shown to alter the gene expression profile of a highly drug-susceptible A. baumannii strain A118 leading to persistence and survival of this pathogen. Herein, we explore the impact of human pleural fluid (HPF) and human serum albumin (HSA) on the gene expression profile of a highly multi-drug-resistant strain of A. baumannii AB5075. Differential expression was observed for ~30 genes, whose products are involved in quorum sensing, quorum quenching, iron acquisition, fatty acid metabolism, biofilm formation, secretion systems, and type IV pilus formation. Phenotypic and further transcriptomic analysis using quantitative RT-PCR confirmed RNA-seq data and demonstrated a distinctive role of HSA as the molecule involved in A. baumannii's response.

Cerebrospinal fluid (CSF) augments metabolism and virulence expression factors in Acinetobacter baumannii.

In a recent report by the Centers for Disease Control and Prevention (CDC), multidrug resistant (MDR) Acinetobacter baumannii is a pathogen described as an "urgent threat." Infection with this bacterium manifests as different diseases such as community and nosocomial pneumonia, bloodstream infections, endocarditis, infections of the urinary tract, wound infections, burn infections, skin and soft tissue infections, and meningitis. In particular, nosocomial meningitis, an unwelcome complication of neurosurgery caused by extensively-drug resistant (XDR) A. baumannii, is extremely challenging to manage. Therefore, understanding how A. baumannii adapts to different host environments, such as cerebrospinal fluid (CSF) that may trigger changes in expression of virulence factors that are associated with the successful establishment and progress of this infection is necessary. The present in-vitro work describes, the genetic changes that occur during A. baumannii infiltration into CSF and displays A. baumannii's expansive versatility to persist in a nutrient limited environment while enhancing several virulence factors to survive and persist. While a hypervirulent A. baumannii strain did not show changes in its transcriptome when incubated in the presence of CSF, a low-virulence isolate showed significant differences in gene expression and phenotypic traits. Exposure to 4% CSF caused increased expression of virulence factors such as fimbriae, pilins, and iron chelators, and other virulence determinants that was confirmed in various model systems. Furthermore, although CSF's presence did not enhance bacterial growth, an increase of expression of genes encoding transcription, translation, and the ATP synthesis machinery was observed. This work also explores A. baumannii's response to an essential component, human serum albumin (HSA), within CSF to trigger the differential expression of genes associated with its pathoadaptibility in this environment.

Human pleural fluid triggers global changes in the transcriptional landscape of Acinetobacter baumannii as an adaptive response to stress.

Acinetobacter baumannii is a feared, drug-resistant pathogen, characterized by its ability to resist extreme environmental and nutrient-deprived conditions. Previously, we showed that human serum albumin (HSA) can increase foreign DNA acquisition specifically and alter the expression of genes associated with pathogenicity. Moreover, in a recent genome-wide transcriptomic study, we observed that pleural fluid (PF), an HSA-containing fluid, increases DNA acquisition, can modulate cytotoxicity, and control immune responses by eliciting changes in the A. baumannii metabolic profile. In the present work, using more stringent criteria and focusing on the analysis of genes related to pathogenicity and response to stress, we analyzed our previous RNA-seq data and performed phenotypic assays to further explore the impact of PF on A. baumannii's microbial behavior and the strategies used to overcome environmental stress. We observed that PF triggered differential expression of genes associated with motility, efflux pumps, antimicrobial resistance, biofilm formation, two-component systems (TCSs), capsule synthesis, osmotic stress, and DNA-damage response, among other categories. Phenotypic assays of A. baumannii A118 and two other clinical A. baumannii strains, revealed differences in their responses to PF in motility, biofilm formation, antibiotic susceptibility, osmotic stress, and outer membrane vesicle (OMV) production, suggesting that these changes are strain specific. We conclude that A. baumannii's pathoadaptive responses is induced by HSA-containing fluids and must be part of this bacterium armamentarium to persist in hostile environments.