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1.
PeerJ ; 12: e17776, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39224820

RESUMO

Background: The tcdA gene codes for an important toxin produced by Clostridioides difficile (C. difficile), but there is currently no simple and cost-effective method of detecting it. This article establishes and validates a rapid and visual loop-mediated isothermal amplification (LAMP) assay for the detection of the tcdA gene. Methods: Three sets of primers were designed and optimized to amplify the tcdA gene in C. difficile using a LAMP assay. To evaluate the specificity of the LAMP assay, C. difficile VPI10463 was used as a positive control, while 26 pathogenic bacterial strains lacking the tcdA gene and distilled water were utilized as negative controls. For sensitivity analysis, the LAMP assay was compared to PCR using ten-fold serial dilutions of DNA from C. difficile VPI10463, ranging from 207 ng/µl to 0.000207 pg/µl. The tcdA gene of C.difficile was detected in 164 stool specimens using both LAMP and polymerase chain reaction (PCR). Positive and negative results were distinguished using real-time monitoring of turbidity and chromogenic reaction. Results: At a temperature of 66 °C, the target DNA was successfully amplified with a set of primers designated, and visualized within 60 min. Under the same conditions, the target DNA was not amplified with the tcdA12 primers for 26 pathogenic bacterial strains that do not carry the tcdA gene. The detection limit of LAMP was 20.700 pg/µl, which was 10 times more sensitive than that of conventional PCR. The detection rate of tcdA in 164 stool specimens using the LAMP method was 17% (28/164), significantly higher than the 10% (16/164) detection rate of the PCR method (X2 = 47, p < 0.01). Conclusion: LAMP method is an effective technique for the rapid and visual detection of the tcdA gene of C. difficile, and shows potential advantages over PCR in terms of speed, simplicity, and sensitivity. The tcdA-LAMP assay is particularly suitable for medical diagnostic environments with limited resources and is a promising diagnostic strategy for the screening and detection of C. difficile infection in populations at high risk.


Assuntos
Toxinas Bacterianas , Clostridioides difficile , Infecções por Clostridium , Enterotoxinas , Fezes , Técnicas de Amplificação de Ácido Nucleico , Sensibilidade e Especificidade , Clostridioides difficile/genética , Clostridioides difficile/isolamento & purificação , Técnicas de Amplificação de Ácido Nucleico/métodos , Humanos , Toxinas Bacterianas/genética , Infecções por Clostridium/diagnóstico , Infecções por Clostridium/microbiologia , Fezes/microbiologia , Fezes/química , Enterotoxinas/genética , Primers do DNA/genética , Técnicas de Diagnóstico Molecular/métodos , Reação em Cadeia da Polimerase/métodos , Adulto , Pessoa de Meia-Idade
2.
Gut Microbes ; 16(1): 2399215, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39284098

RESUMO

Enterotoxigenic Escherichia coli (ETEC) are a major cause of diarrheal illness in humans and animals, induced by enterotoxins produced by these pathogens. Despite the crucial role of neutrophils in combatting bacterial infections, our understanding of how enterotoxins impact neutrophil function is limited. To address this knowledge gap, we used heat-labile enterotoxin (LT) and heat-stable enterotoxin a (STa) to investigate their impact on the effector functions of neutrophils. Our study reveals that pSTa does not exert any discernible effect on the function of neutrophils. In contrast, LT altered the migration and phagocytosis of neutrophils and induced the production of inflammatory factors via activation of cAMP/PKA and ERK1/2 signaling. LT also attenuated the release of neutrophil extracellular traps by neutrophils via the PKA signaling pathway. Our findings provide novel insights into the impact of LT on neutrophil function, shedding light on the underlying mechanisms that govern its immunoregulatory effects. This might help ETEC in subverting the immune system and establishing infection.


Assuntos
Toxinas Bacterianas , Proteínas Quinases Dependentes de AMP Cíclico , AMP Cíclico , Escherichia coli Enterotoxigênica , Enterotoxinas , Infecções por Escherichia coli , Proteínas de Escherichia coli , Neutrófilos , Fagocitose , Enterotoxinas/metabolismo , Neutrófilos/imunologia , Neutrófilos/metabolismo , Humanos , AMP Cíclico/metabolismo , Proteínas de Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Toxinas Bacterianas/metabolismo , Toxinas Bacterianas/imunologia , Infecções por Escherichia coli/imunologia , Infecções por Escherichia coli/microbiologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Armadilhas Extracelulares/metabolismo , Armadilhas Extracelulares/imunologia , Transdução de Sinais
3.
PLoS Biol ; 22(9): e3002734, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-39226241

RESUMO

Vibrio coralliilyticus is a pathogen of coral and shellfish, leading to devastating economic and ecological consequences worldwide. Although rising ocean temperatures correlate with increased V. coralliilyticus pathogenicity, the specific molecular mechanisms and determinants contributing to virulence remain poorly understood. Here, we systematically analyzed the type VI secretion system (T6SS), a contact-dependent toxin delivery apparatus, in V. coralliilyticus. We identified 2 omnipresent T6SSs that are activated at temperatures in which V. coralliilyticus becomes virulent; T6SS1 is an antibacterial system mediating interbacterial competition, whereas T6SS2 mediates anti-eukaryotic toxicity and contributes to mortality during infection of an aquatic model organism, Artemia salina. Using comparative proteomics, we identified the T6SS1 and T6SS2 toxin arsenals of 3 V. coralliilyticus strains with distinct disease etiologies. Remarkably, T6SS2 secretes at least 9 novel anti-eukaryotic toxins comprising core and accessory repertoires. We propose that T6SSs differently contribute to V. coralliilyticus's virulence: T6SS2 plays a direct role by targeting the host, while T6SS1 plays an indirect role by eliminating competitors.


Assuntos
Antozoários , Sistemas de Secreção Tipo VI , Vibrio , Animais , Vibrio/patogenicidade , Vibrio/genética , Vibrio/metabolismo , Sistemas de Secreção Tipo VI/metabolismo , Sistemas de Secreção Tipo VI/genética , Virulência , Antozoários/microbiologia , Artemia/microbiologia , Toxinas Bacterianas/metabolismo , Toxinas Bacterianas/genética , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Vibrioses/microbiologia , Proteômica/métodos , Fatores de Virulência/metabolismo
4.
Nat Commun ; 15(1): 7719, 2024 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-39231966

RESUMO

Nucleotidyltransferases (NTases) control diverse physiological processes, including RNA modification, DNA replication and repair, and antibiotic resistance. The Mycobacterium tuberculosis NTase toxin family, MenT, modifies tRNAs to block translation. MenT toxin activity can be stringently regulated by diverse MenA antitoxins. There has been no unifying mechanism linking antitoxicity across MenT homologues. Here we demonstrate through structural, biochemical, biophysical and computational studies that despite lacking kinase motifs, antitoxin MenA1 induces auto-phosphorylation of MenT1 by repositioning the MenT1 phosphoacceptor T39 active site residue towards bound nucleotide. Finally, we expand this predictive model to explain how unrelated antitoxin MenA3 is similarly able to induce auto-phosphorylation of cognate toxin MenT3. Our study reveals a conserved mechanism for the control of tuberculosis toxins, and demonstrates how active site auto-phosphorylation can regulate the activity of widespread NTases.


Assuntos
Domínio Catalítico , Mycobacterium tuberculosis , Nucleotidiltransferases , Fosforilação , Mycobacterium tuberculosis/metabolismo , Mycobacterium tuberculosis/genética , Nucleotidiltransferases/metabolismo , Nucleotidiltransferases/genética , Toxinas Bacterianas/metabolismo , Toxinas Bacterianas/genética , Toxinas Bacterianas/química , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/química , Modelos Moleculares , RNA de Transferência/metabolismo , RNA de Transferência/genética , Cristalografia por Raios X
5.
Virulence ; 15(1): 2399798, 2024 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-39229975

RESUMO

Staphylococcus aureus is the most common cause of skin and soft tissue infections (SSTIs) with Methicillin-Resistant S. aureus (MRSA) strains being a major contributor in both community and hospital settings. S. aureus relies on metabolic diversity and a large repertoire of virulence factors to cause disease. This includes α-hemolysin (Hla), an integral player in tissue damage found in various models, including SSTIs. Previously, we identified a role for the Spx adapter protein, YjbH, in the regulation of several virulence factors and as an inhibitor of pathogenesis in a sepsis model. In this study, we found that YjbH is critical for tissue damage during SSTI, and its absence leads to decreased proinflammatory chemokines and cytokines in the skin. We identified no contribution of YjbI, encoded on the same transcript as YjbH. Using a combination of reporters and quantitative hemolysis assays, we demonstrated that YjbH impacts Hla expression and activity both in vitro and in vivo. Additionally, expression of Hla from a non-native promoter reversed the tissue damage phenotype of the ΔyjbIH mutant. Lastly, we identified reduced Agr activity as the likely cause for reduced Hla production in the ΔyjbH mutant. This work continues to define the importance of YjbH in the pathogenesis of S. aureus infection as well as identify a new pathway important for Hla production.


Assuntos
Proteínas de Bactérias , Toxinas Bacterianas , Regulação Bacteriana da Expressão Gênica , Proteínas Hemolisinas , Staphylococcus aureus , Transativadores , Proteínas Hemolisinas/metabolismo , Proteínas Hemolisinas/genética , Toxinas Bacterianas/metabolismo , Toxinas Bacterianas/imunologia , Toxinas Bacterianas/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Staphylococcus aureus/patogenicidade , Staphylococcus aureus/imunologia , Staphylococcus aureus/genética , Camundongos , Animais , Transativadores/genética , Transativadores/metabolismo , Infecções Cutâneas Estafilocócicas/microbiologia , Infecções Cutâneas Estafilocócicas/imunologia , Infecções Cutâneas Estafilocócicas/patologia , Staphylococcus aureus Resistente à Meticilina/patogenicidade , Staphylococcus aureus Resistente à Meticilina/genética , Staphylococcus aureus Resistente à Meticilina/imunologia , Pele/microbiologia , Pele/patologia , Pele/imunologia , Fatores de Virulência/genética , Humanos , Infecções dos Tecidos Moles/microbiologia , Infecções dos Tecidos Moles/imunologia , Infecções Estafilocócicas/imunologia , Infecções Estafilocócicas/microbiologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Citocinas/metabolismo , Citocinas/imunologia , Citocinas/genética
6.
Sci Rep ; 14(1): 21257, 2024 09 11.
Artigo em Inglês | MEDLINE | ID: mdl-39261531

RESUMO

The bacterium Clostridium botulinum, well-known for producing botulinum neurotoxins, which cause the severe paralytic illness known as botulism, produces C2 toxin, a binary AB-toxin with ADP-ribosyltranferase activity. C2 toxin possesses two separate protein components, an enzymatically active A-component C2I and the binding and translocation B-component C2II. After proteolytic activation of C2II to C2IIa, the heptameric structure binds C2I and is taken up via receptor-mediated endocytosis into the target cells. Due to acidification of endosomes, the C2IIa/C2I complex undergoes conformational changes and consequently C2IIa forms a pore into the endosomal membrane and C2I can translocate into the cytoplasm, where it ADP-ribosylates G-actin, a key component of the cytoskeleton. This modification disrupts the actin cytoskeleton, resulting in the collapse of cytoskeleton and ultimately cell death. Here, we show that the serine-protease inhibitor α1-antitrypsin (α1AT) which we identified previously from a hemofiltrate library screen for PT from Bordetella pertussis is a multitoxin inhibitor. α1AT inhibits intoxication of cells with C2 toxin via inhibition of binding to cells and inhibition of enzyme activity of C2I. Moreover, diphtheria toxin and an anthrax fusion toxin are inhibited by α1AT. Since α1AT is commercially available as a drug for treatment of the α1AT deficiency, it could be repurposed for treatment of toxin-mediated diseases.


Assuntos
Toxinas Bacterianas , Toxinas Botulínicas , alfa 1-Antitripsina , Toxinas Botulínicas/metabolismo , Toxinas Botulínicas/antagonistas & inibidores , Toxinas Botulínicas/química , Humanos , alfa 1-Antitripsina/metabolismo , alfa 1-Antitripsina/química , Toxinas Bacterianas/metabolismo , Toxina Diftérica/metabolismo , Corynebacterium diphtheriae/metabolismo , Corynebacterium diphtheriae/efeitos dos fármacos , Antígenos de Bactérias/metabolismo , Animais , Clostridium botulinum/metabolismo , Bacillus anthracis/metabolismo , Bacillus anthracis/efeitos dos fármacos
7.
Int J Mol Sci ; 25(17)2024 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-39273577

RESUMO

Tuberculosis is a worldwide plague caused by the pathogen Mycobacterium tuberculosis (M. tb). Toxin-antitoxin (TA) systems are genetic elements abundantly present in prokaryotic organisms and regulate important cellular processes. MazEF is a TA system implicated in the formation of "persisters cells" of M. tb, which contain more than 10 such members. However, the exact function and inhibition mode of each MazF are not fully understood. Here we report crystal structures of MazF-mt3 in its apo form and in complex with the C-terminal half of MazE-mt3. Structural analysis suggested that two long but disordered ß1-ß2 loops would interfere with the binding of the cognate MazE-mt3 antitoxin. Similar loops are also present in the MazF-mt1 and -mt9 but are sustainably shortened in other M. tb MazF members, and these TA pairs behave distinctly in terms of their binding modes and their RNase activities. Systematic crystallographic and biochemical studies further revealed that the biochemical activities of M. tb toxins were combined results between the interferences from the characteristic loops and the electrostatic interactions between the cognate TA pairs. This study provides structural insight into the binding mode and the inhibition mechanism of the MazE/F TA pairs, which facilitate the structure-based peptide designs.


Assuntos
Proteínas de Bactérias , Endorribonucleases , Mycobacterium tuberculosis , Sistemas Toxina-Antitoxina , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Mycobacterium tuberculosis/metabolismo , Mycobacterium tuberculosis/genética , Sistemas Toxina-Antitoxina/genética , Endorribonucleases/química , Endorribonucleases/metabolismo , Endorribonucleases/genética , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Toxinas Bacterianas/química , Toxinas Bacterianas/metabolismo , Toxinas Bacterianas/genética , Ligação Proteica , Cristalografia por Raios X , Modelos Moleculares , Antitoxinas/química , Antitoxinas/metabolismo , Antitoxinas/genética , Sequência de Aminoácidos
8.
Immunohorizons ; 8(9): 606-621, 2024 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-39240270

RESUMO

The human body harbors a substantial population of bacteria, which may outnumber host cells. Thus, there are multiple interactions between both cell types. Given the common presence of Staphylococcus aureus in the human body and the role of Th17 cells in controlling this pathogen on mucous membranes, we sought to investigate the effect of α-hemolysin, which is produced by this bacterium, on differentiating Th17 cells. RNA sequencing analysis revealed that α-hemolysin influences the expression of signature genes for Th17 cells as well as genes involved in epigenetic regulation. We observed alterations in various histone marks and genome methylation levels via whole-genome bisulfite sequencing. Our findings underscore how bacterial proteins can significantly influence the transcriptome, epigenome, and phenotype of human Th17 cells, highlighting the intricate and complex nature of the interaction between immune cells and the microbiota.


Assuntos
Toxinas Bacterianas , Epigênese Genética , Proteínas Hemolisinas , Staphylococcus aureus , Células Th17 , Staphylococcus aureus/imunologia , Staphylococcus aureus/genética , Proteínas Hemolisinas/genética , Proteínas Hemolisinas/imunologia , Proteínas Hemolisinas/metabolismo , Humanos , Células Th17/imunologia , Células Th17/metabolismo , Toxinas Bacterianas/imunologia , Toxinas Bacterianas/genética , Infecções Estafilocócicas/imunologia , Infecções Estafilocócicas/microbiologia , Metilação de DNA , Diferenciação Celular , Transcriptoma
9.
BMC Microbiol ; 24(1): 334, 2024 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-39251908

RESUMO

BACKGROUND: Characteristics of non-clinical strains of methicillin-resistant Staphylococcus aureus (MRSA) especially from fishery environment are poorly understood. This research, in addition to comprehensive characterisation, sought to delineate the genetic relatedness between the MRSA strains originating from clinical as well as non-clinical settings. Out of 39 methicillin-resistant staphylococcal isolates from 197 fish samples, 6 (Three each of methicillin-resistant S. haemolyticus (MRSH) and MRSA) with distinct resistance profiles were selected for whole-genome sequencing. Using respective bioinformatics tools, MRSA genomes were comprehensively characterized for resistome, virulomes, molecular epidemiology and phylogenetic analysis. Simultaneously, MRSH genomes were specifically examined to characterize antimicrobial resistance genes (ARGs), owing to the fact that MRSH is often recognized as a reservoir for resistance determinants. RESULTS: Three MRSA clones identified in this study include ST672-IVd/t13599 (sequence type-SCCmec type/spa type), ST88-V/t2526, and ST672-IVa/t1309. Though, the isolates were phenotypically vancomycin-sensitive, five of the six genomes carried vancomycin resistance genes including the VanT (VanG cluster) or VanY (VanM cluster). Among the three MRSA, only one harbored the gene encoding Panton-Valentine Leukocidin (PVL) toxin, while staphylococcal enterotoxin (SEs) genes such as sea and seb, associated with staphylococcal food poisoning were identified in two other MRSA. Genomes of MRSH carried a composite of type V staphylococcal cassette chromosome mec (SCCmec) elements (5C2 & 5). This finding may be explained by the inversion and recombination events that may facilitate the integration of type V elements to the SCC elements of S. aureus with a methicillin-susceptible phenotype. Phylogenetically, MRSA from a non-clinical setting displayed a considerable relatedness to that from clinical settings. CONCLUSION: This study highlights the genetic diversity and resistance profiles of MRSA and MRSH, with non-clinical MRSA showing notable relatedness to clinical strains. Future research should explore resistance gene transfer mechanisms and environmental reservoirs to better manage MRSA spread.


Assuntos
Peixes , Genoma Bacteriano , Staphylococcus aureus Resistente à Meticilina , Filogenia , Intoxicação Alimentar Estafilocócica , Staphylococcus aureus Resistente à Meticilina/genética , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Staphylococcus aureus Resistente à Meticilina/patogenicidade , Animais , Peixes/microbiologia , Intoxicação Alimentar Estafilocócica/microbiologia , Genoma Bacteriano/genética , Antibacterianos/farmacologia , Sequenciamento Completo do Genoma , Virulência/genética , Testes de Sensibilidade Microbiana , Humanos , Fatores de Virulência/genética , Alimentos Marinhos/microbiologia , Microbiologia de Alimentos , Toxinas Bacterianas/genética , Epidemiologia Molecular , Staphylococcus haemolyticus/genética , Staphylococcus haemolyticus/efeitos dos fármacos , Staphylococcus haemolyticus/isolamento & purificação , Staphylococcus haemolyticus/patogenicidade
10.
Arch Microbiol ; 206(10): 401, 2024 Sep 11.
Artigo em Inglês | MEDLINE | ID: mdl-39261350

RESUMO

Staphylococcus aureus is a notorious pathogen predominantly involved in skin and soft tissue infections, exhibiting a distinct innate sex bias. This study explores the influence of testosterone on the virulence of S. aureus and elucidates its underlying mechanisms. Utilizing a skin abscess model in intact and castrated male mice, we assessed the effects of testosterone on S. aureus pathogenicity. Compared to controls, castrated mice showed significantly reduced abscess sizes and decreased bacterial loads, highlighting the role of testosterone in modulating the severity of S. aureus infections. In vitro experiments revealed that testosterone enhances the hemolytic activity, cytotoxicity, and oxidative stress resistance of S. aureus. Real-time quantitative PCR analysis showed a significant upregulation of the genes encoding α-hemolysin (hla) and phenol-soluble modulin (psmα). Importantly, testosterone treatment significantly enhanced the expression of the accessory gene regulator (Agr) quorum-sensing system components (agrC, agrA, agrB, agrD), while the SaeRS system (saeR, saeS, and sbi) exhibited only slight changes. Gene knockout experiments revealed that deletion of agrC, rather than saeRS and agrBD, abolishes the testosterone-induced enhancement of hemolysis and gene expression, underscoring the key role of AgrC. Molecular docking simulations indicated a direct interaction between testosterone and AgrC protein, with a strong binding affinity at the active site residue SER201. This study provides new insights into the mechanistic basis of how testosterone enhances the pathogenicity of S. aureus, potentially contributing to increased male susceptibility to S. aureus infections and offering a targeted approach for therapeutic interventions.


Assuntos
Proteínas de Bactérias , Infecções Estafilocócicas , Staphylococcus aureus , Testosterona , Masculino , Testosterona/farmacologia , Testosterona/metabolismo , Animais , Staphylococcus aureus/genética , Staphylococcus aureus/patogenicidade , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/metabolismo , Camundongos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Virulência , Infecções Estafilocócicas/microbiologia , Transativadores/genética , Transativadores/metabolismo , Regulação Bacteriana da Expressão Gênica , Percepção de Quorum , Simulação de Acoplamento Molecular , Toxinas Bacterianas/metabolismo , Toxinas Bacterianas/genética , Abscesso/microbiologia , Hemólise , Proteínas Hemolisinas/metabolismo , Proteínas Hemolisinas/genética
11.
Proc Natl Acad Sci U S A ; 121(38): e2410679121, 2024 Sep 17.
Artigo em Inglês | MEDLINE | ID: mdl-39264739

RESUMO

Enterotoxigenic Escherichia coli (ETEC) cause hundreds of millions of diarrheal illnesses annually ranging from mildly symptomatic cases to severe, life-threatening cholera-like diarrhea. Although ETEC are associated with long-term sequelae including malnutrition, the acute diarrheal illness is largely self-limited. Recent studies indicate that in addition to causing diarrhea, the ETEC heat-labile toxin (LT) modulates the expression of many genes in intestinal epithelia, including carcinoembryonic cell adhesion molecules (CEACAMs) which ETEC exploit as receptors, enabling toxin delivery. Here, however, we demonstrate that LT also enhances the expression of CEACAMs on extracellular vesicles (EV) shed by intestinal epithelia and that CEACAM-laden EV increase in abundance during human infections, mitigate pathogen-host interactions, scavenge free ETEC toxins, and accelerate ETEC clearance from the gastrointestinal tract. Collectively, these findings indicate that CEACAMs play a multifaceted role in ETEC pathogen-host interactions, transiently favoring the pathogen, but ultimately contributing to innate responses that extinguish these common infections.


Assuntos
Toxinas Bacterianas , Escherichia coli Enterotoxigênica , Enterotoxinas , Infecções por Escherichia coli , Proteínas de Escherichia coli , Interações Hospedeiro-Patógeno , Escherichia coli Enterotoxigênica/metabolismo , Humanos , Infecções por Escherichia coli/microbiologia , Infecções por Escherichia coli/imunologia , Infecções por Escherichia coli/metabolismo , Proteínas de Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Enterotoxinas/metabolismo , Toxinas Bacterianas/metabolismo , Vesículas Extracelulares/metabolismo , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiologia , Animais , Camundongos , Antígenos CD/metabolismo , Antígenos CD/genética , Antígeno Carcinoembrionário/metabolismo , Antígeno Carcinoembrionário/genética , Moléculas de Adesão Celular/metabolismo , Moléculas de Adesão Celular/genética , Diarreia/microbiologia , Diarreia/metabolismo
12.
Arch Microbiol ; 206(9): 381, 2024 Aug 17.
Artigo em Inglês | MEDLINE | ID: mdl-39153128

RESUMO

The bacterial type II toxin-antitoxin (TA) system is a rich genetic element that participates in various physiological processes. Aeromonas veronii is the main bacterial pathogen threatening the freshwater aquaculture industry. However, the distribution of type II TA system in A. veronii was seldom documented and its roles in the life activities of A. veronii were still unexplored. In this study, a novel type II TA system AvtA-AvtT was predicted in a fish pathogen Aeromonas veronii biovar sobria with multi-drug resistance using TADB 2.0. Through an Escherichia coli host killing and rescue assay, we demonstrated that AvtA and AvtT worked as a genuine TA system, and the predicted toxin AvtT actually functioned as an antitoxin while the predicted antitoxin AvtA actually functioned as a toxin. The binding ability of AvtA with AvtT proteins were confirmed by dot blotting analysis and co-immunoprecipitation assay. Furthermore, we found that the toxin and antitoxin labelled with fluorescent proteins were co-localized. In addition, it was found that the transcription of AvtAT bicistronic operon was repressed by the AvtAT protein complex. Deletion of avtA gene and avtT gene had no obvious effect on the drug susceptibility. This study provides first characterization of type II TA system AvtA-AvtT in aquatic pathogen A. veronii.


Assuntos
Aeromonas veronii , Proteínas de Bactérias , Sistemas Toxina-Antitoxina , Aeromonas veronii/genética , Aeromonas veronii/metabolismo , Sistemas Toxina-Antitoxina/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Toxinas Bacterianas/metabolismo , Toxinas Bacterianas/genética , Óperon , Escherichia coli/genética , Escherichia coli/metabolismo , Escherichia coli/efeitos dos fármacos , Antitoxinas/genética , Antitoxinas/metabolismo , Regulação Bacteriana da Expressão Gênica
13.
Nat Commun ; 15(1): 7416, 2024 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-39198411

RESUMO

The human gut pathogen Clostridioides difficile displays substantial inter-strain genetic variability and confronts a changeable nutrient landscape in the gut. We examined how human gut microbiota inter-species interactions influence the growth and toxin production of various C. difficile strains across different nutrient environments. Negative interactions influencing C. difficile growth are prevalent in an environment containing a single highly accessible resource and sparse in an environment containing C. difficile-preferred carbohydrates. C. difficile toxin production displays significant community-context dependent variation and does not trend with growth-mediated inter-species interactions. C. difficile strains exhibit differences in interactions with Clostridium scindens and the ability to compete for proline. Further, C. difficile shows substantial differences in transcriptional profiles in co-culture with C. scindens or Clostridium hiranonis. C. difficile exhibits massive alterations in metabolism and other cellular processes in co-culture with C. hiranonis, reflecting their similar metabolic niches. C. hiranonis uniquely inhibits the growth and toxin production of diverse C. difficile strains across different nutrient environments and robustly ameliorates disease severity in mice. In sum, understanding the impact of C. difficile strain variability and nutrient environments on inter-species interactions could help improve the effectiveness of anti-C. difficile strategies.


Assuntos
Clostridioides difficile , Infecções por Clostridium , Técnicas de Cocultura , Microbioma Gastrointestinal , Clostridioides difficile/genética , Clostridioides difficile/metabolismo , Clostridioides difficile/fisiologia , Humanos , Animais , Camundongos , Infecções por Clostridium/microbiologia , Nutrientes/metabolismo , Toxinas Bacterianas/metabolismo , Toxinas Bacterianas/genética , Interações Microbianas , Clostridium/metabolismo , Clostridium/genética , Feminino , Antibiose , Camundongos Endogâmicos C57BL
14.
Mar Drugs ; 22(8)2024 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-39195450

RESUMO

Cyanobacterial biodiversity and potential toxicity in coastal lagoons have barely been studied despite these transitional water systems being very important in conservation and for the preservation of economic resources. Most of these transitional systems have been affected by eutrophication, and climate change will severely affect them by promoting cyanobacteria growth, especially in Mediterranean areas. This study aims to characterize the diversity of epipelic and epiphytic cyanobacteria species in a Mediterranean coastal lagoon and their potential for toxins production (microcystins and saxitoxins). Strains were isolated and genetically identified. Toxins were extracted and quantified by LC/MS-MS. All the taxa belong to the former Oscillatoriales. The presence of Nodosilinea and Toxifilum is reported for the first time for Spanish waters, but Pseudanabaena, Phormidium, Geitlerinema and Synechococcus also formed part of benthic mats. All the strains contained Microcystin-YR (MC-YR), but saxitoxin (STX) was present only in the extracts of Nodosilinea and Pseudanabena. MC-LY, MC-LW and [D-Asp3] MC-LR were detected in the extracts of Synechococcus and MC-LF in Toxifilum, but at concentrations that did not permit quantification. Toxins production by epipelic and epiphytic strains in coastal lagoons may represent a hazard, but also an opportunity to obtain potentially interesting compounds that should be further studied.


Assuntos
Mudança Climática , Cianobactérias , Microcistinas , Cianobactérias/metabolismo , Microcistinas/análise , Saxitoxina/toxicidade , Saxitoxina/análise , Toxinas Marinhas/análise , Toxinas Marinhas/toxicidade , Toxinas Bacterianas , Espectrometria de Massas em Tandem , Eutrofização , Água do Mar/microbiologia , Salinidade , Espanha
15.
PLoS One ; 19(8): e0308615, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39146363

RESUMO

Staphylococcus aureus infection and colonization in patients may be transmitted to healthcare providers and the environment and subsequently cause healthcare-associated infections in other patients. Pathogenic S. aureus strains produce virulence factors, such as Panton-Valentine Leukocidin (PVL), that contribute to the severity of infections and aid in their spread. The emergence of antimicrobial resistance (AMR) is additional concern with respect to S. aureus infection. In this study, the virulence genes and antibiotic resistance profiles of S. aureus were characterized from patients' clinical isolates, healthcare workers' (HCWs') nasal colonization screenings, and the environment at a tertiary healthcare hospital in Addis Ababa, Ethiopia. A total of 365 samples were collected from September 2021 to September 2022: 73 patients' clinical specimens, 202 colonization screenings from HCWs, and 90 hospital environment's swabs. Fifty-one (25.2%) HCW and 10/90 (11.1%) environment S. aureus isolates were identified. Among the 134 isolates, 10 (7.5%) were methicillin-resistant S. aureus (MRSA). Three (4.1%), five (9.8%), and two (20.0%) of the MRSA isolates were identified from the patients, HCWs, and the environment, respectively. Overall, 118 (88.1%) were ampicillin and penicillin resistant; 70 (52.2%) were trimethoprim sulfamethoxazole resistant; and 28 (20.9%) were erythromycin resistant. S. aureus isolates from patients were more resistant to antibiotics than isolates from HCWs or the hospital environment (p<0.05). A total of 92/134 (68.6%) isolates possessed the lukfF-PV gene, which was identified in 62 (85.0%), 26 (51.0%), and 4 (40.0%) of the patient, HCWs, and the environment, respectively. The proportion of lukfF-PV gene containing S. aureus isolated from patient samples was statistically significant. Four (40.0%) of the MRSA isolates also had the lukfF-PV gene. The identification of highly AMR and virulence factors from patients, HCWs and the environment is concerning. Further studies are needed to identify potential transmission links and improve infection prevention and control.


Assuntos
Antibacterianos , Pessoal de Saúde , Infecções Estafilocócicas , Staphylococcus aureus , Centros de Atenção Terciária , Humanos , Etiópia/epidemiologia , Feminino , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/epidemiologia , Infecções Estafilocócicas/tratamento farmacológico , Adulto , Masculino , Staphylococcus aureus/genética , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/isolamento & purificação , Antibacterianos/farmacologia , Pessoa de Meia-Idade , Testes de Sensibilidade Microbiana , Adolescente , Staphylococcus aureus Resistente à Meticilina/genética , Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Adulto Jovem , Fatores de Virulência/genética , Leucocidinas/genética , Criança , Exotoxinas/genética , Pré-Escolar , Infecção Hospitalar/microbiologia , Infecção Hospitalar/epidemiologia , Farmacorresistência Bacteriana/genética , Lactente , Idoso , Toxinas Bacterianas
16.
Gut Microbes ; 16(1): 2390133, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39132815

RESUMO

Vancomycin (VAN) treatment in Clostridioides difficile infection (CDI) suffers from a relatively high rate of recurrence, with a variety of reasons behind this, including biofilm-induced recurrent infections. C. difficile can form monophyletic or symbiotic biofilms with other microbes in the gut, and these biofilms protect C. difficile from being killed by antibiotics. In this study, we analyzed the ecological relationship between Bacteroides thetaiotaomicron and C. difficile and their formation of symbiotic biofilm in the VAN environment. The production of symbiotic biofilm formed by C. difficile and B. thetaiotaomicron was higher than that of C. difficile and B. thetaiotaomicron alone in the VAN environment. In symbiotic biofilms, C. difficile was characterized by increased production of the toxin protein TcdA and TcdB, up-regulation of the expression levels of the virulence genes tcdA and tcdB, enhanced bacterial cell swimming motility and c-di-GMP content, and increased adhesion to Caco-2 cells. The scanning electron microscope (SEM) combined with confocal laser scanning microscopy (CLSM) results indicated that the symbiotic biofilm was elevated in thickness, dense, and had an increased amount of mixed bacteria, while the fluorescence in situ hybridization (FISH) probe and plate colony counting results further indicated that the symbiotic biofilm had a significant increase in the amount of C. difficile cells, and was able to better tolerate the killing of the simulated intestinal fluid. Taken together, C. difficile and B. thetaiotaomicron become collaborative in the VAN environment, and targeted deletion or attenuation of host gut B. thetaiotaomicron content may improve the actual efficacy of VAN in CDI treatment.


Assuntos
Antibacterianos , Proteínas de Bactérias , Bacteroides thetaiotaomicron , Biofilmes , Clostridioides difficile , Simbiose , Vancomicina , Biofilmes/efeitos dos fármacos , Biofilmes/crescimento & desenvolvimento , Clostridioides difficile/efeitos dos fármacos , Clostridioides difficile/fisiologia , Clostridioides difficile/genética , Humanos , Vancomicina/farmacologia , Antibacterianos/farmacologia , Células CACO-2 , Bacteroides thetaiotaomicron/efeitos dos fármacos , Bacteroides thetaiotaomicron/metabolismo , Bacteroides thetaiotaomicron/fisiologia , Bacteroides thetaiotaomicron/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Toxinas Bacterianas/metabolismo , Toxinas Bacterianas/genética , Enterotoxinas/metabolismo , Enterotoxinas/genética , Aderência Bacteriana/efeitos dos fármacos
17.
Proc Natl Acad Sci U S A ; 121(33): e2406775121, 2024 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-39116134

RESUMO

Biofilm-protected pathogenic Staphylococcus aureus causes chronic infections that are difficult to treat. An essential building block of these biofilms are functional amyloid fibrils that assemble from phenol-soluble modulins (PSMs). PSMα1 cross-seeds other PSMs into cross-ß amyloid folds and is therefore a key element in initiating biofilm formation. However, the paucity of high-resolution structures hinders efforts to prevent amyloid assembly and biofilm formation. Here, we present a 3.5 Å resolution density map of the major PSMα1 fibril form revealing a left-handed cross-ß fibril composed of two C2-symmetric U-shaped protofilaments whose subunits are unusually tilted out-of-plane. Monomeric α-helical PSMα1 is extremely cytotoxic to cells, despite the moderate toxicity of the cross-ß fibril. We suggest mechanistic insights into the PSM functional amyloid formation and conformation transformation on the path from monomer-to-fibril formation. Details of PSMα1 assembly and fibril polymorphism suggest how S. aureus utilizes functional amyloids to form biofilms and establish a framework for developing therapeutics against infection and antimicrobial resistance.


Assuntos
Amiloide , Biofilmes , Staphylococcus aureus , Staphylococcus aureus/metabolismo , Staphylococcus aureus/fisiologia , Biofilmes/crescimento & desenvolvimento , Amiloide/metabolismo , Amiloide/química , Toxinas Bacterianas/metabolismo , Toxinas Bacterianas/química , Conformação Proteica , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/química , Modelos Moleculares
18.
J Hazard Mater ; 478: 135452, 2024 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-39121740

RESUMO

Thermostable direct hemolysin (TDH) is a key virulence factor of Vibrio parahaemolyticus, capable of causing seafood-mediated outbreaks of gastroenteritis, posing a threat to the aquatic environment and global public health. In the present study, we explored a multivalent aptamer-mediated inhibition strategy to mitigate TDH toxicity. Based on the characteristic structure of TDH, a stable multivalent aptamer, Ap3-5, was rationally designed by truncation, key fragment evolution, and end fixation. Ap3-5 exhibited strong affinity (Kd=39.24 nM), and thermal (Tm=57.6 °C) and enzymatic stability. In silico studies also revealed that Ap3-5 occupied more active sites of TDH and covered its central pore, indicating its potential as a blocking agent for inhibiting TDH toxicity. In the hemolysis assay, Ap3-5 significantly suppressed the hemolytic effect of TDH. A cellular study revealed a substantial (∼80 %) reduction in TDH cytotoxicity. Supporting these findings, in vivo trials confirmed the inhibitory action of Ap3-5 on both the acute and intestinal toxicity of TDH. Overall, benefiting from the strong binding affinity, high stability, and multisite occupation of the multivalent aptamer with TDH, Ap3-5 displayed robust potential against TDH toxicity by inhibiting membrane pore formation, providing a new approach for alleviating bacterial infections.


Assuntos
Aptâmeros de Nucleotídeos , Toxinas Bacterianas , Proteínas Hemolisinas , Hemólise , Vibrio parahaemolyticus , Vibrio parahaemolyticus/efeitos dos fármacos , Vibrio parahaemolyticus/genética , Proteínas Hemolisinas/toxicidade , Proteínas Hemolisinas/química , Aptâmeros de Nucleotídeos/química , Toxinas Bacterianas/toxicidade , Toxinas Bacterianas/química , Hemólise/efeitos dos fármacos , Animais , Humanos , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/toxicidade , Proteínas de Bactérias/genética
19.
Toxicon ; 249: 108059, 2024 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-39117156

RESUMO

The aim of our study was to investigate the effects of cyanobacterial metabolites: microcystin-LR (MC-LR) anabaenopeptin-A (ANA-A), cylindrospermopsin (CYL), their binary and ternary mixtures on rainbow trout (Oncorhynchus mykiss) gill (RTgill-W1) cell line. We determined the following cell parameters: Hoechst and propidium iodide (PI) double staining, intracellular ATP level with luminometric assay, glutathione level with ThiolTracker Violet®- glutathione detection reagent and cytoskeletal F-actin fluorescence. The results showed that although reduction of Hoechst fluorescence was observed in both binary and ternary combinations of cyanobacterial metabolites, the mixture of MC-LR + ANA-A + CYL was the most potent inhibitor (EC50 = 148 nM). PI fluorescence and ATP levels were more increased in the cells exposed to the mixtures than those exposed to the individual metabolites with synergistic toxic changes suggesting apoptosis as the mechanism of cell death. Reduced glutathione level was also decreased in cells exposed both to single metabolites and their mixtures with the highest decrease and synergistic effects at 334 nM MC-LR+334 nM ANA-A+ 334 nM CYL suggesting induction oxidative stress by the tested compounds. Reduction of F-actin fluorescence was found in the cells from all of the groups exposed to individual metabolites and their mixtures, however the highest level of inhibition showed the binary MC-LR + CYL and the ternary MC-LR + ANA-A + CYL with synergistic interactions. The study suggests that in natural conditions fish gill cells may be very sensitive to individual cyanobacterial metabolites and more prone to their binary and ternary mixtures.


Assuntos
Alcaloides , Toxinas de Cianobactérias , Cianobactérias , Toxinas Marinhas , Microcistinas , Oncorhynchus mykiss , Uracila , Microcistinas/metabolismo , Animais , Alcaloides/farmacologia , Uracila/análogos & derivados , Linhagem Celular , Cianobactérias/metabolismo , Oncorhynchus mykiss/metabolismo , Brânquias/metabolismo , Brânquias/efeitos dos fármacos , Glutationa/metabolismo , Toxinas Bacterianas , Trifosfato de Adenosina/metabolismo , Apoptose/efeitos dos fármacos , Peptídeos Cíclicos/farmacologia
20.
Microb Pathog ; 195: 106865, 2024 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-39153578

RESUMO

Community-Acquired Respiratory Distress Syndrome Toxin (CARDS TX) is a unique exotoxin produced by Mycoplasma pneumoniae (MP) and has been confirmed to possess ADP-ribosyltransferase (ART) and vacuolating activities. CARDS TX binds to receptors on the surfaces of mammalian cells followed by entry into the cells through clathrin-mediated endocytosis, and exerts cytotoxic effects by undergoing retrograde transport and finally cleavage on endosomes and cellular organelles. In addition, CARDS TX can trigger severe inflammatory reactions resulting in airway dysfunction, producing allergic inflammation and asthma-like conditions. As a newly discovered virulence factor of MP, CARDS TX has been extensively studied in recent years. As resistance to macrolide drugs has increased significantly in recent years and there is no vaccine against MP, the development of a vaccine targeting CARDS TX is considered a potential preventive measure. This review focuses on recent studies and insights into this toxin, providing directions for a better understanding of MP pathogenesis and treatment. IMPORTANCE: A serious hazard to worldwide public health in recent years, Mycoplasma pneumoniae (MP) is a prominent bacterium that causes community-acquired pneumonia (CAP) in hospitalized children. Due to their high prevalence and fatality rates, MP infections often cause both respiratory illnesses and extensive extrapulmonary symptoms. It has recently been shown that MP produces a distinct exotoxin known as Community-Acquired Respiratory Distress Syndrome Toxin (CARDS TX). Mycoplasma pneumoniae pneumonia (MPP)-like tissue injury is caused by this toxin because it has both ADP-ribosyltransferase and vacuolating properties. A better knowledge of MP etiology and therapy is provided by this review, which focuses on latest research and insights into this toxin.


Assuntos
ADP Ribose Transferases , Proteínas de Bactérias , Toxinas Bacterianas , Infecções Comunitárias Adquiridas , Mycoplasma pneumoniae , Pneumonia por Mycoplasma , Fatores de Virulência , Mycoplasma pneumoniae/patogenicidade , Humanos , Infecções Comunitárias Adquiridas/microbiologia , Toxinas Bacterianas/metabolismo , Pneumonia por Mycoplasma/microbiologia , Fatores de Virulência/metabolismo , ADP Ribose Transferases/metabolismo , Proteínas de Bactérias/metabolismo , Animais , Virulência , Exotoxinas/metabolismo , Síndrome do Desconforto Respiratório/microbiologia , Síndrome do Desconforto Respiratório/etiologia
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