ACS Infectious Diseases最新文献

The F₁F_O-ATP synthase is essential to the aerobe Acinetobacter baumannii. Its F_O-domain utilizes the proton motive force to rotate the turbine (c₁₀-ring) inside the stator (a subunit), which generates a torque that is translated to the catalytic F₁-domain for adenosine 5′-triphosphate (ATP) synthesis. Here, we investigated key features of the F_O-domain, including the proton intake channel, proton donor and acceptor residues, an A. baumannii unique subunit a helix, and the proton exit pathway. By employing a heterologous system, we generated mutants and studied their growth kinetic properties in minimal media, as well as the ATP synthesis activity of their inverted-membrane vesicles. The findings highlight the front entry as the main proton uptake pathway and the key residues involved in proton translocation. Molecular dynamics (MD) simulations confirm the role of these charged residues, which interact with water molecules to facilitate a water-mediated proton transfer in a Grotthuss-like mechanism. Similarly, the exit channel with R224 of subunit a playing a central role is described. Importantly, the sequential flow of proton intake, turbine rotation, and proton release are modulated by the unique a subunit helix, which functions like a molecular ratchet to facilitate effective proton transfer for the final formation of ATP. The importance in function, difference in amino acid content, and uniqueness in regulation by its specific molecular ratchet make the A. baumannii proton pathway an attractive inhibitor target, where a cork-like molecule could prevent proton intake and/or release with the consequence of ATP synthesis and cell growth inhibition.

Polymyxins are considered last-resort antibiotics for multidrug-resistant Gram-negative bacteria. However, their clinical utility is limited by toxicities, particularly nephrotoxicity and neurotoxicity, as well as the emergence of resistance. This review addresses these challenges and evaluates strategic interventions aimed at enhancing the efficacy of polymyxins while mitigating their adverse effects. Combination therapies have emerged as a cornerstone strategy. These therapies can be categorized into five frameworks: structural barrier disruption, bioenergetic flux modulation, metabolic homeostasis disruption, resistance neutralization, and virulence disarming. In addition to synergistic agents, complementary strategies such as detoxifying adjuvants and advanced delivery systems have been systematically integrated to overcome the intrinsic limitations of polymyxins. Collectively, these multifaceted strategies enhance the antibacterial activity of polymyxins against Gram-negative bacteria, while simultaneously reducing effective doses, minimizing toxicity, and mitigating the development of resistance. These innovations represent a pivotal advance in revitalizing polymyxin therapy in the era of multidrug resistance.

Bacterial glycans are complex and often presented on the surface of the cell as a level of protection. These glycans offer an opportunity to screen for new antibiotic targets and immunological markers. Here recent developments in the field of glycan arrays are presented as opportunities to advance therapies for human health.

Bacterial strains are distinguished by surface glycans composed of defined sugar sequences that include “rare” monosaccharides, which are absent in human glycans and help to mediate host–microbe interactions. One of the most prevalent rare sugars is l-Rhamnose (l-Rha), and human sera are generally enriched in anti-l-Rha antibodies; however, the source of l-Rha antigens is unknown. Here, we synthesize a surface glycan l-Rha-N-acetyl glucosamine disaccharide sequence, which is found across many bacterial species, to evaluate binding motifs of human anti-glycan antibodies in clinical and commercial human sera. We find that sera are enriched in IgG antibodies that react with this disaccharide probe. Through capture of bound antibodies and analysis with surface glycan sequences from different strains, we observe that bound human antibodies appear to recognize free or branched, but not internal, l-Rha motifs. Overall, this work details the isolation of naturally occurring anti-l-Rha human antibodies and promotes an understanding of their carbohydrate recognition epitopes.

Mycobacterium tuberculosis (Mtb), the causative agent of human tuberculosis (TB), employs its de novo histidine (His) biosynthesis to escape host-inflicted His starvation. This makes the enzymes involved in this biosynthetic pathway promising anti-TB drug targets. In this study, employing the high-resolution crystal structure of imidazole glycerol phosphate dehydratase (IGPD) of the Mtb His pathway, three triazole scaffold molecules were identified as potential inhibitors of this enzyme. These high-resolution crystal structures of the enzyme–inhibitor complexes elucidated the key interactions responsible for their binding specificity and affinity. We also studied the interactions of these inhibitors with the enzyme at the atomic level and tested their cytotoxicity and efficacy in in vitro and in vivo models. Our findings revealed that the most prominent inhibitor, SF2, was safe in mice and effectively inhibited the in vitro growth of both free as well as in macrophage-internalized wild-type and drug-resistant Mtb clinical isolates. Notably, SF2 also showed a marginal reduction in the bacterial load in organs of mice infected with Mtb. Collectively, these findings advocate the chemical inhibition of IGPD of the His pathway as a novel anti-Mtb therapeutic strategy.

Methicillin-resistant Staphylococcus aureus (MRSA) ST398 carries two hyaluronidase genes, hysA and its homologue hysA^νSaβ, the latter located on the genomic island νSaβ. However, the prevalence of hysA^νSaβ and its contribution to virulence remain unclear. Here, we report that the hysA^νSaβ gene is present in 18.3% (4707/25,752) of S. aureus in the NCBI database, with ST398 being the most prevalent sequence type (30.9%, 1457/4707). In ST398, the hysA^νSaβ gene is flanked by IS21 and IS3, with >99.0% nucleotide identity across strains, suggesting horizontal acquisition. In a mouse skin infection model, a wild-type ST398 MRSA strain carrying both hysA and hysA^νSaβ formed significantly larger abscesses than isogenic mutants lacking one or both hyaluronidase genes. Wild-type infection led to a higher bacterial load and sustained induction of chemokines (CCL5, CXCL1, CCL4) and pro-inflammatory cytokines (IL-1β, IL-6, IL-33), resulting in prolonged neutrophil recruitment and severe inflammation. Consistently, hysA and hysA^νSaβ enhanced the survival of MRSA ST398 inside RAW 264.7 macrophages and neutrophils. In vitro, a double knockout strain (ΔhysA-ΔhysA^νSaβ) grew more slowly with hyaluronic acid (HA) as the sole carbon source, accompanied by intracellular accumulation of specific amino acids (proline, valine, threonine, and phenylalanine) and downregulation of amino acid biosynthesis pathways. Moreover, RAW 264.7 macrophages infected with ΔhysA-ΔhysA^νSaβ showed a marked upregulation of the oxidative phosphorylation (OXPHOS) pathway compared to uninfected controls, suggesting an enhanced cellular metabolic and inflammatory response that could improve bacterial clearance. Our findings highlight the functionally redundant roles of hysA and hysA^νSaβ in MRSA ST398 pathogenesis, suggesting that these hyaluronidases are potential targets for antistaphylococcal therapy.

Leishmania major, an intracellular protozoan parasite, resides within parasitophorous vacuoles in host macrophages and relies on host-pathway manipulation for survival. Here, we uncover a novel role of the Leishmania surface metalloprotease GP63 in stabilizing the parasitophorous vacuoles through targeted subversion of host vesicular trafficking and apoptosis. We demonstrate that GP63 is essential for the selective recruitment of the Golgi-associated adaptor protein PIST to the parasitophorous vacuoles, a process that is impaired in GP63-deficient (LmGP63^–/–) parasites. GP63 facilitates PIST-Golgin160 complex formation by suppressing caspase-3 activation, preventing Golgin160 cleavage. Caspase inhibition via Z-VAD-FMK further enhances this complex’s recruitment. Moreover, GP63 selectively modulates autophagy by promoting PIST-Beclin1 colocalization while excluding LC3 from the parasitophorous vacuoles. These findings identify GP63 as a central effector that orchestrates host vesicular and apoptotic pathways to maintain parasitophorous vacuoles integrity and promote chronic infection, offering insights into potential therapeutic targets against Leishmaniasis.

Chagas disease (CD), caused by Trypanosoma cruzi, has been one of the leading causes of cardiac death in Latin America. Its pathogenesis and progression are still poorly understood. Thus, we performed an untargeted metabolomics analysis to understand the metabolic changes involved in the final acute phase of CD. Male mice’s chagasic hearts (60 days postinfection) were compared to healthy tissues. Two hundred and fifty-one significant metabolites or chemical classes were annotated. Disturbances in energy metabolism and dysregulation of amino acids were observed. Pathway analyses indicated increased inflammatory activity in infected individuals, as observed by eicosanoid (prostaglandin and thromboxane) changes. The accumulation of some sphingomyelins, correlated with myocarditis, suggests heart tissue damage from the infection. The metabolic changes observed contribute to understanding disease progression and the cardiac effects caused by the parasite, bringing new insights into the discovery and development of new therapies.

Streptococcus gordonii sp. firmicutes is an early colonizer of the oral microbiome and contributes positively to oral health. While this species has been found to produce hydrogen peroxide by spxB expression, the relationship of this expression to the competence regulon has not yet been explored. To this end, this study sought to investigate the connection of the S. gordonii competence regulon quorum sensing (QS) circuitry with downstream proliferative phenotypic expression resulting from competence-stimulating peptide (CSP) exposure, with specific attention to peroxide formation. Following confirmation of the native CSP, RNA-seq was completed to gain insights into transcriptomic variations resulting from CSP incubation. Later, structure–activity relationship (SAR) analyses of the native CSP were completed. The results revealed residues integral to CSP:ComD binding and activation, while indicating which residues were considered dispensable to this process. Phenotypic assessment revealed that peroxide formation was modulated via the competence regulon. Finally, interspecies competition assays were carried out to understand the interactions between S. gordonii and S. mutans, with S. gordonii demonstrating a profound capability of antagonizing S. mutans growth and proliferation. Our results support that this antagonism is mainly attributed to hydrogen peroxide production by S. gordonii. This finding suggests that S. gordonii may be exploited for its beneficial proliferative phenotypes downstream of the competence regulon.

Crassostrea gigas big defensins (Cg-BigDefs) are a family of two-domain antimicrobial peptides with broad antibacterial activity. The C-terminal domain of Cg-BigDefs harbors a β-defensin-like structure, whereas the ancestral N-terminal domain adopts a globular structure. Here, we developed molecular tools to track the fine interactions of these two domains with Staphylococcus aureus and to gain insight into Cg-BigDef1 mechanisms of action. By using super-resolution microscopy and S. aureus mutants with specific deletions of cell wall components, we found that teichoic acids (TAs) play a key role in the Cg-BigDef1 interaction with S. aureus. A ΔtagO S. aureus mutant lacking cell wall teichoic acids (WTAs) exhibited increased resistance to Cg-BigDef1. Consistently, the binding of Cg-BigDef1 to S. aureus cell wall was significantly reduced in the ΔtagO mutant. In contrast, a ΔdltA S. aureus mutant unable to transfer d-alanine onto lipoteichoic acid (LTA) showed increased susceptibility to Cg-BigDef1 and lysed rapidly in contact with the peptide. Cg-BigDef1 bound to S. aureus cell wall. In addition, competitive binding with exogenously added LTA was sufficient to impair Cg-BigDef1 antimicrobial activity against S. aureus. These data suggest that TAs are conserved molecular motifs recognized by Cg-BigDef1. Finally, we found that Cg-BigDef1 interaction with S. aureus was mediated by its N-terminal domain, which enables the C-terminal β-defensin-like domain to interact with the bacterial cell wall. Altogether, our results identify TAs as important targets for Cg-BigDef1. This interaction appears to be mediated by the ancestral N-terminal domain characteristic of this peptide family.