Sulbactam-durlobactam is the first-line agent against carbapenem-resistant Acinetobacter baumannii. Resistance can be conferred by metallo-β-lactamases alone or penicillin-binding protein 3 alterations plus β-lactamase overproduction or enhanced efflux. Other reported mechanisms remain associative and resistance in guideline-recommended carbapenem addition is unclear, underscoring the need for mechanistic validation and further studies.
Methanogens are an ancestral group of archaea that occupy a unique niche within the human gut microbiome by virtue of their methane production. In this process, they serve as hydrogen sinks, allowing continued bacterial fermentation and influencing short-chain fatty acid production. Available evidence suggests that methanogen abundance may be declining in parallel with the broader reduction in gut microbial diversity accompanying industrialization. We describe the evolution of methanogens, their ecological roles in the human microbiome, and evidence for their apparent decline. If confirmed, reductions in methanogen prevalence and abundance may have substantial metabolic consequences, reframing these archaea as keystone species in need of scientific attention and conservation efforts.
Bacteria live in dense communities where competition influences the composition and, therefore, the function of these communities. Beyond competing for resources, bacteria engage in antagonism by deploying a range of molecular weapon systems to inhibit and kill other bacteria. Investing in antagonism is expected to incur a fitness trade-off, but the nature of this trade-off at the level of molecular physiology remains underexplained. Applying recent advances about the physiological constraints faced by bacterial cells may help us better understand existing studies and design new investigations into interbacterial antagonism. Bacterial cells face two important constraints: a finite amount of protein and a maximum translation speed for ribosomes. As a result, the only way for a cell to grow faster is to allocate more of its finite proteome to synthesizing ribosomes. A cell choosing to attack competitors must therefore allocate some of its limited proteome budget to antagonistic proteins instead of other functions. Conversely, being attacked and resisting the effects of such attacks also require an investment of proteomic resources. The extent to which proteome allocation constraints influence bacterial physiology is not fully understood; consequently, how these constraints influence interbacterial antagonism has not been investigated. Here, I will discuss how proteome allocation constraints can re-contextualize our existing understanding of the costs of both deploying and resisting attacks and how investigation of these constraints may further our understanding of interbacterial antagonism.
Increased intestinal colonization by adherent-invasive Escherichia coli (AIEC) has been implicated in Crohn's disease, an inflammatory bowel disease characterized by abnormal immune responses and chronic inflammation in the gastrointestinal tract. LPS-Toll-like receptor 4 signalling may contribute to AIEC-associated intestinal inflammation. Here, we show that disruption of rfaG-dependent cell envelope biogenesis is associated with reduced type 1 fimbrial expression, epithelial invasion and intestinal colonization in the AIEC strain LF82. Deletion of rfaG, a gene involved in cell envelope biogenesis, resulted in reduced type 1 fimbrial expression and activity, accompanied by reduced transcription of fim genes. The rfaG mutant also exhibited impaired invasion of epithelial cells compared with the parental LF82 strain. Finally, the rfaG mutant showed reduced competitive colonization in the murine intestinal tract, whereas chromosomal complementation with rfaG restored this colonization defect. Inducible expression of type 1 fimbriae also partially reversed the reduced colonization at the early stage of infection, raising the possibility that additional rfaG-dependent mechanisms, independently of type 1 fimbrial expression, contribute to intestinal colonization. These findings suggest that rfaG-dependent cell envelope biogenesis contributes to the coordinated expression of surface-associated virulence factors associated with intestinal colonization in AIEC.
Group B Streptococcus (GBS) is a leading cause of neonatal sepsis and meningitis worldwide. Capsular polysaccharide (CPS) is a major virulence factor that aids GBS in colonizing the gastrointestinal (GI) and vaginal tracts, surviving in whole blood and evading the immune system. CPS is also the basis for a GBS CPS-protein conjugate vaccine candidate. In addition to its function on the surface of the bacteria, CPS can also be released and shed into the environment. In Streptococcus pneumoniae , shed CPS has been shown to absorb antimicrobial peptides and CPS-specific antibodies, allowing the bacteria to evade the innate immune system and potentially limiting vaccine protection. In this work, we show that CPS shedding occurs in ~75% of GBS isolates that we screened from a large library of clinical strains. We then identified ypmS as a genetic determinant of CPS shedding using an indexed transposon library. We used a clean in-frame deletion mutant of ypmS to assess the importance of shedding in GBS colonization and immune evasion. We explored the role of ypmS in colonization using murine models of GI and vaginal co-colonization and showed that the wild-type strain outcompeted the Δ ypmS mutant in both scenarios. We also assessed the effect of shed capsule on opsonophagocytosis. Free CPS was protective against bacterial killing in an opsonophagocytic killing assay using CPS-specific antibodies. With this work, we have identified a novel genetic cause of CPS shedding and shown its contribution to GBS colonization fitness and immune evasion.
Cyanobacteria are a globally abundant bacterial phylum that make key contributions to primary production and nutrient cycling in aquatic ecosystems. However, some cyanobacteria also cause substantial adverse environmental impacts through harmful blooms and the production of potent toxins. Given their global significance, understanding the processes governing cyanobacterial ecology is critical for assessing aquatic ecosystem function. While abiotic environmental factors have traditionally been considered most important, a growing body of literature indicates that interactions with other microorganisms, in particular heterotrophic bacteria, shape cyanobacterial growth, population dynamics, and ecological success. In this review, we provide a synthesis of the importance of interbacterial interactions in controlling cyanobacterial populations and their ecological impacts on natural aquatic environments and argue that these relationships should be incorporated into conceptual and predictive frameworks of cyanobacterial ecology.
Atomic force microscopy (AFM) has become a powerful tool to provide nanoscale insights into the spatial organization, biomechanics, and intermolecular interactions of the cell envelopes of living bacteria in real time and under near-native conditions. Focusing on recent literature, we discuss how AFM has advanced our understanding of Gram-positive, Gram-negative, and mycobacterial cell envelope architectures, as well as the mechanisms of action of established and novel antimicrobials. We also summarize how AFM can be used to probe the adhesion, virulence, and immune evasion of pathogens through single-molecule and single-cell force spectroscopy.
Prokaryotic toxin-antitoxin (TA) systems are genetic modules involved in stress responses, including antiphage defense and the maintenance of mobile genetic elements. Although canonical TA systems consist of two neighboring genes encoding one toxin and one antitoxin, many have evolved into tripartite systems through the incorporation of a third component that serves as a chaperone, a regulator, or an additional antitoxin or toxin to increase functional diversity and regulatory potential. This review compares tripartite TA systems [TA-chaperone (TAC), TA-regulator (TAR), dual-antitoxin, and dual-toxin systems] with bipartite systems, summarizing their neutralizing mechanisms and phage-triggered activation pathways. The regulatory capacity and modularity of tripartite TAs illuminate the evolutionary trajectory of these 'selfish' genetic elements, reflecting complex coevolutionary dynamics among TA systems, host bacteria, and phages.
Positive species interactions underpin ecological resilience, which is critical for counteracting anthropogenic pressures on ecosystems. Current research focuses on intra-holobiont interactions between hosts and their microbiomes, reflecting their recognized importance for host well-being, but overlooks interactions among holobionts. We introduce the meta-holobiont framework for the marine environment, which conceptualizes ecological communities as networks of interacting holobionts connected through microbial transmission and metabolite production. Marine connectivity can facilitate horizontal transmission of microorganisms, potentially enhancing host fitness and stress tolerance across all life stages. Conceptual and empirical findings from sessile benthic species support this view. We argue that investigating meta-holobiont interactions is crucial for advancing knowledge of ecological dynamics and may have important practical implications, particularly for marine restoration strategies.
For the clinical treatment of infections with Enterobacteriaceae , the development of resistance to last-resort antimicrobials like colistin is of concern, as it limits treatment options. Since 2015, ten families of mobile colistin resistance ( mcr ) genes have been discovered; however, our understanding of MCR remains limited because (i) mcr-1 is the most studied variant, (ii) mcr variants are rarely compared and (iii) mcr variants are primarily studied in Escherichia coli , even though studies have shown that mcr variants can confer significantly different phenotypes and that lipid A (i.e. the target of colistin and mcr- modification) structurally differs between bacterial species. To fill this gap, we examined how mcr-3 and mcr-9 , as two less frequently studied mcr variants, impact the colistin resistance of laboratory-adapted and real-world (i.e. clinical or food) isolates of E. coli , Salmonella enterica and Klebsiella pneumoniae . We found that while mcr-3 consistently conferred colistin resistance to all tested strains, mcr-9 only conferred resistance to two strains. The fold-change in colistin minimum inhibitory concentrations was significantly impacted by both the bacterial species and the mcr variant but not by whether a strain was a laboratory-adapted or real-world isolate. Overall, our results suggest that while laboratory-adapted strains may provide a good estimate of mcr -mediated colistin resistance of real-world isolates, findings of mcr -mediated phenotypes in one bacterial species should not be extrapolated to another.
Aerated subterranean ecosystems are emerging as globally widespread yet understudied sinks for atmospheric greenhouse gases (GHGs), especially methane. Subterranean GHG cycling is driven by atmospheric exchange between the surface and subsurface, and by uptake by aerotrophic bacteria inhabiting cave surfaces. Despite growing evidence for these novel biogeochemical processes, aerated subterranean ecosystems remain largely omitted from Earth system models, GHG budgets, ecosystem service valuations, and climate change mitigation efforts. Here, we synthesise the current understanding of the subterranean microbial methane sink and provide a framework for upscaling uptake based on ventilation and biological determinants. We propose that enhanced subterranean aerotrophy could provide a novel pathway for atmospheric methane removal by leveraging caves and cave proxies as passive methane biofilters. We speculate that decommissioned mines might provide a similar opportunity. Based on this mechanistic understanding, we discuss key uncertainties and outline a research agenda to direct future research on subterranean ecosystems as a new potential lever for climate change mitigation.
Forests are not a uniform methane (CH4) sink; soils, stems, and canopies act as sources or sinks depending on local conditions. We synthesize emerging evidence, expose the limitations of component-based approaches, and advocate integrated observational-modeling frameworks to accurately quantify net forest CH4 exchange and climate feedbacks.
For most organisms, we rely on the ability to observe them from birth to death in order to fully understand their ecology, life cycles, adaptations, and interactions with other organisms. For fungi, this is a challenge, and little is known about how old they can become and in what form they persist in the environment. In this opinion article, we highlight the complexity of determining the longevity of fungi. We provide an overview of how the age of fungal individuals and the persistence of mycelial structures have been measured in different groups of fungi. Finally, we propose ideas for advancing the study of fungal longevities and discuss how differences in fungal longevity and mycelial persistence can impact the ways we study and manage fungi in ecosystems.
Acidic, metal-contaminated environments harbour specialized microbial consortia adapted to extreme stress. We examined an environmental Euglena mutabilis culture naturally associated with Talaromyces and Acidiphilium acidophilum and exposed it to cadmium (Cd). Integrated transcriptomic, hormonal, structural and taxonomic analyses revealed a coordinated Cd-tolerance strategy. RNA sequencing showed differential regulation of metal transporters consistent with a shift from Cd uptake to intracellular sequestration. Transmission electron microscopy confirmed Cd compartmentalization within chloroplasts and increased paramylon granules. Cd exposure suppressed light-harvesting complex genes and formate/nitrite transporters while maintaining core photosynthetic function. Hormone profiling indicated strong repression of bioactive auxin and cytokinin (CK) free bases, alongside accumulation of CK nucleotides and downregulation of CK biosynthetic and activation genes. Metagenomics revealed Cd-driven enrichment of Talaromyces and Acidiphilium, implicating them in detoxification and stress support. Together, these responses highlight early Cd uptake followed by chloroplast-based detoxification, metabolic buffering via paramylon, hormonal downregulation of growth and community-mediated resilience.
Gut microbial metabolites, particularly short-chain fatty acids (SCFAs), bile acid (BAs) derivatives, and tryptophan-derived metabolites (TDMs), act as signaling molecules, receptor ligands, and epigenetic modulators in regulating host metabolic and immune homeostasis. They exert local effects in the gut and reach distant tissues via hepatic, lymphatic, neural, and immune-hormonal routes, thereby regulating immunity, metabolism, and neural function. Disruptions in these pathways are associated with inflammatory diseases, neurodegeneration, and cancer. This review discusses the importance of SCFAs, BAs, and TDMs in different host metabolic and immunological contexts, dissects their molecular mechanisms of action on target cells, evaluates the therapeutic implications of targeting these microbial pathways, and outlines a roadmap for possible future clinical translations.
The type VI secretion system (T6SS) is a molecular nanomachine that provides Gram-negative bacteria with competitive advantages during host colonization and interbacterial interactions. Citrobacter rodentium, a mouse-specific attaching and effacing pathogen, employs a T6SS encoded by the cts1 locus to outcompete the gut microbiota and establish infection. However, under standard laboratory growth conditions, cts1 expression remains silent and the regulatory mechanisms governing its activation are unknown. Here, we investigated the regulation of cts1, which is organized into two divergent gene clusters, cts1L and cts1R. Transcriptional fusions of the cts1L-cts1R intergenic region in both orientations were inactive in wild-type C. rodentium, but became active in the absence of the global regulator H-NS. Systematic deletion and site-directed mutagenesis of this region identified four promoter regions, one driving cts1L transcription and three directing cts1R expression (P1, P2 and P3). Additional promoters were identified within the cts1R cluster. All cts1 promoters were differentially repressed by H-NS. Moreover, expression of the response regulator CpxR from a low-copy plasmid enhanced promoter activity in the Δhns background. Electrophoretic mobility shift assays showed that H-NS binds the cts1 regulatory region directly, whereas phosphorylated CpxR bound the internal cts1R promoters but not the intergenic P1-P3 regions, suggesting that CpxR activates the latter indirectly. Together, these findings demonstrate that cts1 expression is controlled by a complex hierarchical regulatory network in which H-NS acts as the primary repressor and CpxR contributes to activation only once H-NS-mediated repression is relieved, in a context-dependent manner. This multifactorial regulation likely involves additional, yet-unidentified factors that fine-tune cts1 expression in response to environmental cues.
Industrial adoption of microbial cell-free protein synthesis (CFPS), an in vitro protein production platform that uses transcription and translation machinery from lysed cells, remains limited despite sustained technical progress across the field. This perspective positions microbial CFPS within a manufacturing context, focusing on its current capabilities, the limits of those capabilities and the concrete steps required for industrial adoption. In CFPS, the absence of cell growth constraints supports rapid protein production, simplified upstream workflows and direct control over reaction conditions, while also lending itself to formats such as lyophilized reagents for storage and transport. Beyond basic protein expression, microbial CFPS is increasingly used as an adaptable production environment in which reaction composition can be tuned, accessory enzymatic functions can be introduced and post-translational modification strategies can be engineered to better align with product requirements. The expanding range of microbial chassis further broadens the design space, allowing platform choice to be guided by functional, regulatory and deployment considerations rather than yield alone. However, increased capability has not yet translated into routine manufacturing use, and persistent barriers include reproducibility, vulnerable supply chains, limited scale-up practice, downstream purification suitable for therapeutic products and the absence of CFPS-specific quality and regulatory expectations. Overall, microbial CFPS is best evaluated not as a replacement for existing biomanufacturing but as a complementary manufacturing paradigm whose industrial relevance will depend on establishing standardization, economics and regulatory readiness.
The type III secretion system (T3SS) is a virulence mechanism commonly used by Gram-negative bacterial pathogens to deliver virulence proteins, known as effectors, into infected cells. The T3SS secretes a range of different substrates: first the needle subunits, then the translocon pore components and finally a pathogen-specific range of effector proteins. Each of these classes of substrates interacts with a corresponding class of bacterial chaperones, which are required for their efficient secretion. The requirement for these chaperones has been attributed to multiple functions, including preventing premature substrate activity, maintaining substrate stability in the bacterial cytoplasm and mediating substrate targeting and secretion hierarchy. Here, we bring together what is known about the function of T3SS chaperones in a range of different bacterial pathogens. Through analysis of the conservation of chaperone sequence and structure, we discuss how these proteins interact with and support the secretion of diverse substrates. Finally, we evaluate the extent to which chaperones are universally required for effector secretion.
