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Rheumatoid arthritis-associated interstitial lung disease (RA-ILD) is a serious and common extra-articular disease manifestation. Patients with RA-ILD experience reduced bacterial diversity and gut bacteriome alterations. However, the gut mycobiome and virome in these patients have been largely neglected. In this study, we performed whole-metagenome shotgun sequencing on fecal samples from 30 patients with RA-ILD, and 30 with RA-non-ILD, and 40 matched healthy controls. The gut bacteriome and mycobiome were explored using a reference-based approach, while the gut virome was profiled based on a nonredundant viral operational taxonomic unit (vOTU) catalog. The results revealed significant alterations in the gut microbiomes of both RA-ILD and RA-non-ILD groups compared with healthy controls. These alterations encompassed changes in the relative abundances of 351 bacterial species, 65 fungal species, and 4,367 vOTUs. Bacteria such as Bifidobacterium longum, Dorea formicigenerans, and Collinsella aerofaciens were enriched in both patient groups. Ruminococcus gnavus (RA-ILD), Gemmiger formicilis, and Ruminococcus bromii (RA-non-ILD) were uniquely enriched. Conversely, Faecalibacterium prausnitzii, Bacteroides spp., and Roseburia inulinivorans showed depletion in both patient groups. Mycobiome analysis revealed depletion of certain fungi, including Saccharomyces cerevisiae and Candida albicans, in patients with RA compared with healthy subjects. Notably, gut virome alterations were characterized by an increase in Siphoviridae and a decrease in Myoviridae, Microviridae, and Autographiviridae in both patient groups. Hence, multikingdom gut microbial signatures showed promise as diagnostic indicators for both RA-ILD and RA-non-ILD. Overall, this study provides comprehensive insights into the fecal virome, bacteriome, and mycobiome landscapes of RA-ILD and RA-non-ILD gut microbiota, thereby offering potential biomarkers for further mechanistic and clinical research.
Assuntos
Artrite Reumatoide , Bactérias , Fezes , Microbioma Gastrointestinal , Doenças Pulmonares Intersticiais , Humanos , Doenças Pulmonares Intersticiais/microbiologia , Doenças Pulmonares Intersticiais/virologia , Artrite Reumatoide/complicações , Artrite Reumatoide/microbiologia , Fezes/microbiologia , Fezes/virologia , Feminino , Masculino , Pessoa de Meia-Idade , Bactérias/classificação , Bactérias/isolamento & purificação , Bactérias/genética , Idoso , Viroma , Micobioma , Adulto , Vírus/classificação , Vírus/isolamento & purificação , Vírus/genética , Fungos/isolamento & purificação , Fungos/classificaçãoRESUMO
Introduction: The type VI secretion system (T6SS) is a crucial virulence factor in the nosocomial pathogen Acinetobacter baumannii. However, its association with drug resistance is less well known. Notably, the roles that different T6SS components play in the process of antimicrobial resistance, as well as in virulence, have not been systematically revealed. Methods: The importance of three representative T6SS core genes involved in the drug resistance and virulence of A. baumannii, namely, tssB, tssD (hcp), and tssM was elucidated. Results: A higher ratio of the three core genes was detected in drug-resistant strains than in susceptible strains among our 114 A. baumannii clinical isolates. Upon deletion of tssB in AB795639, increased antimicrobial resistance to cefuroxime and ceftriaxone was observed, alongside reduced resistance to gentamicin. The ΔtssD mutant showed decreased resistance to ciprofloxacin, norfloxacin, ofloxacin, tetracycline, and doxycycline, but increased resistance to tobramycin and streptomycin. The tssM-lacking mutant showed an increased sensitivity to ofloxacin, polymyxin B, and furazolidone. In addition, a significant reduction in biofilm formation was observed only with the ΔtssM mutant. Moreover, the ΔtssM strain, followed by the ΔtssD mutant, showed decreased survival in human serum, with attenuated competition with Escherichia coli and impaired lethality in Galleria mellonella. Discussion: The above results suggest that T6SS plays an important role, participating in the antibiotic resistance of A. baumannii, especially in terms of intrinsic resistance. Meanwhile, tssM and tssD contribute to bacterial virulence to a greater degree, with tssM being associated with greater importance.
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Acinetobacter baumannii , Sistemas de Secreção Tipo VI , Humanos , Virulência/genética , Sistemas de Secreção Tipo VI/genética , Resistência Microbiana a Medicamentos , Antibacterianos/farmacologia , OfloxacinoRESUMO
Infections led by Acinetobacter baumannii strains are of great concern in healthcare environments due to the strong ability of the bacteria to spread through different apparatuses and develop drug resistance. Severe diseases can be caused by A. baumannii in critically ill patients, but its biological process and mechanism are not well understood. Secretion systems have recently been demonstrated to be involved in the pathogenic process, and five types of secretion systems out of the currently known six from Gram-negative bacteria have been found in A. baumannii. They can promote the fitness and pathogenesis of the bacteria by releasing a variety of effectors. Additionally, antibiotic resistance is found to be related to some types of secretion systems. In this review, we describe the genetic and structural compositions of the five secretion systems that exist in Acinetobacter. In addition, the function and molecular mechanism of each secretion system are summarized to explain how they enable these critical pathogens to overcome eukaryotic hosts and prokaryotic competitors to cause diseases.
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Acinetobacter baumannii is a critical biofilm-forming pathogen that has presented great challenges in the clinic due to multidrug resistance. Thus, new methods of intervention are needed to control biofilm-associated infections. In this study, among three tested Lactobacillus species, Lactobacillus rhamnosus showed significant antimaturation and antiadherence effects against A. baumannii biofilm. Lactic acid (LA) and acetic acid (AA) were the most effective antibiofilm biosurfactants (BSs) produced by L. rhamnosus. This antibiofilm phenomenon produced by LA and AA was due to the strong bactericidal effect, which worked from very early time points, as determined by colony enumeration and confocal laser scanning microscope. The cell destruction of A. baumannii appeared in both the cell envelope and cytoplasm. A discontinuous cell envelope, the leakage of cell contents, and the increased extracellular activity of ATPase demonstrated the disruption of the cell membrane by LA and AA. These effects also demonstrated the occurrence of protein lysis. In addition, bacterial DNA interacted with and was damaged by LA and AA, resulting in significantly reduced expression of biofilm and DNA repair genes. The results highlight the possibility and importance of using probiotics in clinical prevention. Probiotics can be utilized as novel biocides to block and decrease biofilm formation and microbial contamination in medical equipment and during the treatment of infections. IMPORTANCE A. baumannii biofilm is a significant virulence factor that causes the biofilm colonization of invasive illnesses. Rising bacterial resistance to synthetic antimicrobials has prompted researchers to look at natural alternatives, such as probiotics and their derivatives. In this study, L. rhamnosus and its BSs (LA and AA) demonstrated remarkable antibiofilm and antimicrobial characteristics, with a significant inhibitory effect on A. baumannii. These effects were achieved by several mechanisms, including the disruption of the cell envelope membrane, protein lysis, reduced expression of biofilm-related genes, and destruction of bacterial DNA. The results provide support for the possibility of using probiotics and their derivatives in the clinical prevention and therapy of A. baumannii infections.
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Introduction: An extended-spectrum beta-lactamase (ESBL)-hypervirulent Klebsiella pneumoniae (HvKP) strain HKE9 was isolated from the blood in an outpatient. Methods: The effect of the global regulatory factor RpoS on antimicrobial resistance, pathogenicity, and environmental adaptability was elucidated. Results: HKE9 is a novel ST3355 (K20/O2a) hypervirulent strain with a positive string test and resistant to cephems except cefotetan. It has a genome size of 5.6M, including two plasmids. CTX-M-15 was found in plasmid 2, and only ompk37 was found in the chromosome. HKE9 could produce bacterial siderophores, and genes of enterobactin, yersiniabactin, aerobactin, and salmochelin have been retrieved in the genome. As a global regulatory factor, knockout of rpoS did not change antimicrobial resistance or hemolytic phenotype while increasing the virulence to Galleria mellonella larvae and showing higher viscosity. Moreover, rpoS knockout can increase bacterial competitiveness and cell adhesion ability. Interestingly, HKE9-M-rpoS decreased resistance to acidic pH, high osmotic pressure, heat shock, and ultraviolet and became sensitive to disinfectants (H2O2, alcohol, and sodium hypochlorite). Although there were 13 Type 6 secretion system (T6SS) core genes divided into two segments with tle1 between segments in the chromosome, transcriptomic analysis showed that rpoS negatively regulated T4SS located on plasmid 2, type 1, and type 3 fimbriae and positively regulate genes responsible for acidic response, hyperosmotic pressure, heat shock, oxidative stress, alcohol and hypochlorous acid metabolism, and quorum sensing. Discussion: Here, this novel ST3355 ESBL-HvKP strain HKE9 may spread via various clonal types. The important regulation effect of rpoS is the enhanced tolerance and resistance to environmental stress and disinfectants, which may be at the cost of reducing virulence and regulated by T4SS.
Assuntos
Anti-Infecciosos , Desinfetantes , Animais , Virulência/genética , Klebsiella pneumoniae , Fatores de Virulência/genética , Fatores de Virulência/farmacologia , Transcriptoma , Peróxido de Hidrogênio/farmacologia , beta-Lactamases/genética , beta-Lactamases/metabolismo , Fenótipo , Desinfetantes/farmacologia , Anti-Infecciosos/farmacologiaRESUMO
The modified carbapenem inactivation method (mCIM) recommended by the Clinical and Laboratory Standards Institute is not applicable for detecting carbapenemases in Acinetobacter baumannii. Four currently reported phenotypic detection methods, namely, the modified Hodge test, the mCIM, the adjusted mCIM, and the simplified carbapenem inactivation method (sCIM), did not perform well in our 90 clinical A. baumannii isolates. Thus, the minimal inhibitory concentrations (MICs) of carbapenems and the existence and expression of carbapenemase-encoding genes were detected to explain the results. According to the E-test, which was more accurate than the VITEK 2 system, 80.0 and 41.1% were resistant to imipenem (IPM) and meropenem (MEM), respectively, and 14.4 and 53.3% exhibited intermediate resistance, respectively. Five ß-lactamase genes were found, of which blaOXA-51-like, blaTEM, and blaOXA-23-like were detected more frequently in 85 non-susceptible strains. The expression of blaOXA-23-like was positively correlated with the MIC values of IPM and MEM. Therefore, an improved approach based on the mCIM, designated the optimized CIM (oCIM), was developed in this study to detect carbapenemases more accurately and reproducibly. The condition was improved by evaluating the factors of A. baumannii inoculum, incubation broth volume, and MEM disk incubation time. Obvious high sensitivity (92.94%) and specificity (100.00%) were obtained using the oCIM, which was cost-effective and reproducible in routine laboratory work.
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Acinetobacter baumannii is an important nosocomial pathogen that can develop various resistance mechanisms to many antibiotics. However, little is known about how it evolves from an antibiotic sensitive to a resistant phenotype. In this study, we investigated the transition of outer membrane proteins (OMPs) under antibiotic stress and identified YiaD as an OMP marker involved in the development of adaptive resistance to meropenem (MEM) in A. baumannii. Following stimulation of a carbapenem-sensitive strain AB5116 with sub-MIC of MEM, yiaD showed significantly decreased expression, and this decrease continued with prolonged stimulation for 8 h. The downregulation of yiaD was not only observed in clinically sensitive strains but also in 45 carbapenem-resistant isolates that produced the ß-lactamases TEM and OXA-23. However, the extent of the reduction of yiaD expression in resistant strains was less than that in sensitive strains. Lack of yiaD resulted in a 4-fold increase in the MIC of AB5116 to MEM. The same level of depressed susceptibility induced by yiaD deletion was observed in both a growth curve test and a survival rate assay. Moreover, the colony shape became enlarged and irregular after loss of yiaD, and the biofilm formation ability of A. baumannii was influenced by YiaD. These results suggest that YiaD could respond to the stimulus of MEM in A. baumannii with a downregulation trend that kept pace with the prolonged stimulation time, indicating that it participates in various routes to benefit MEM resistance evolution in both carbapenem-sensitive and -resistant A. baumannii strains. IMPORTANCE Acinetobacter baumannii can develop various resistance mechanisms to carbapenems. However, the factors involved in the evolutionary process that leads from transition to the sensitive to resistant phenotype are not clear. The outer membrane protein YiaD of A. baumannii was downregulated under the stress of meropenem (MEM), and its expression level was continuously reduced with prolonged stimulation time. The downregulation of yiaD was not only observed in sensitive strains but also in carbapenem-resistant isolates producing the ß-lactamases TEM and OXA-23. However, the extent of yiaD reduction was less in resistant strains than in sensitive strains. Lack of yiaD resulted in an increased MEM MIC, enlarged and irregular colonies, and decreased biofilm formation ability. These results suggest that YiaD responds to MEM stimulus in A. baumannii and participates in the adaptive resistance of MEM in both carbapenem-sensitive and -resistant strains.