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This Correspondence summarizes the session "From Petri Dish to Planet Earth: Microbiology by All" at FEMS MICRO Milan 2025 highlighting the urgent need for microbial literacy and societal engagement with microbiology. Chaired by Rup Lal and Juan-Luis Ramos, the session convened scientists, educators, and communicators to discuss strategies for promoting microbiology among communities, policymakers, and students. Central contributions focused on the International Microbiology Literacy Initiative (IMiLI), emphasizing global collaboration, intergenerational dialogue, and community-centered educational tools. Discussions showcased practical examples of microbial applications for sustainability and public engagement. The session conclusions, presented by Princy Hira, reaffirmed IMiLI's mission to expand microbial literacy across borders and generations, recognizing educators as key drivers and science communicators as essential outreach ambassadors.
Antiracist pedagogy is a teaching philosophy aimed at critically examining the interconnection between race, power, education, and one's field of study. After participating in the Seminar for Transformation Around Antiracist Teaching (START) program at Brown University, an intergenerational biology team composed of a professor, doctoral student, and an undergraduate student developed new assignments covering broad topics in bioethics for an Introductory Microbiology course. Here, we share our process and perspectives on engaging students in the exploration of how the field of microbiology is deeply connected to history, society, and culture.
The Real-Lab-Day active learning methodology is an innovative approach aimed at deepening undergraduate students' understanding of the scientific process by integrating theory with practical laboratory experience in microbiology. This study aimed to reassess the educational impact of Real-Lab-Day ten years after its initial implementation and to explore its potential for optimizing experimental microbiology protocols. Undergraduate students participated in authentic laboratory experiments under the supervision of graduate mentors. To assess the impact of the activity, formative questionnaires were administered to evaluate students' perceptions of learning, engagement, and motivation. In parallel, the methodology was employed to optimize a bacterial pathogenicity protocol using enteroinvasive Escherichia coli (EIEC) as a model organism. Our data revealed sustained student approval and confirmed the effectiveness of the methodology in consolidating theoretical knowledge, developing practical and cognitive skills, and fostering interest in microbiological research. Furthermore, Real-Lab-Day enabled the successful optimization of bacterial invasion and dissemination protocol. Overall, Real-Lab-Day emerges as a multifunctional pedagogical strategy that effectively connects teaching and research. It enhances student learning experiences while also contributing to the optimization and validation of experimental protocols for use in practical classes. The methodology aligns with contemporary higher education practices and holds strong potential for future educational research.
This work describes a laboratory activity designed to illustrate the phenomenon of bacterial quorum sensing (QS), a communication mechanism in bacterial communities. The activity focuses on the bioluminescence production regulated by QS of bacteria that live in symbiosis with cephalopods. This activity targets undergraduate students in biology, biochemistry, or other sciences and aims to promote their interest in microbiology and to help students to understand the role and mechanism of QS in microorganisms by means of a visual example of symbiotic interactions between bacteria and animals. At the same time, students are expected to develop lab skills in bacterial isolation, pure culture obtention, and interpretation of microbiological results. The work also provides references and resources to help students understand the subject and teachers assess student learning.
Lichen symbioses host complex microbial communities whose functional organization remains poorly understood. In Peltigera lichens, bacterial partners mediate key nutrient transformations, but it is unclear whether the spatial distribution of phosphorus-cycling functions follows the hierarchical control previously observed for community composition. We hypothesized that Peltigera microbiomes follow a thallus-to-soil gradient of control, in which host-driven specialization within thalli transitions toward environmentally driven reconfiguration in the substrate and soil. To test this, we quantified five bacterial genes involved in phosphorus turnover (gcd, phoD, phoN, phnX, and appA) across thalli, underlying substrates, and adjacent soils of several Peltigera species collected along contrasting Patagonian bioclimates. Absolute and relative gene abundances, together with diversity and variance partitioning analyses, were used to evaluate the influence of host identity, edaphic properties, and climate. Gene profiles revealed a shift from host-associated specialization to environmentally filtered assemblages, indicating that symbiotic and abiotic factors jointly structure phosphorus-cycling potential. The coexistence of functional specialization and redundancy provides a plausible mechanism for sustaining phosphorus turnover under changing environmental conditions.
Photobacterium damselae subsp. piscicida (Pdp) is a host-adapted primary pathogen impacting finfish aquaculture worldwide, whose virulence evolution is driven by the mobilome. The pPHDPT3 plasmid encoding a type III secretion system is critical for Pdp virulence but unstable in vitro in European and Japanese isolates. Here we show that a stable ancestral variant is conserved in Australian isolates. The elusive pPHDPT3 variant has undergone gene loss and accumulated a ∼5.5 kb quadruple direct repeat, which could explain plasmid loss via the classic dimer catastrophe scenario. In addition, we hypothesise that a 189-aa serine recombinase encoded within this repeat, and also on pPHDP10 plasmid, may act as a plasmid resolvase, with its frequent loss further exacerbating the dimer catastrophe.
Shiga toxin (Stx) is the primary virulence factor of Shiga toxin-producing Escherichia coli (STEC). Stx is categorized into Stx1 and Stx2 and further classified into several subtypes based on amino acid sequence variations. During routine surveillance of STEC isolates in Japan, we identified strain 2021H102 which harbored an uncommon stx subtype. Whole genome sequencing combined with matrix-assisted laser desorption/ionization-time of flight mass spectrometry analysis revealed that 2021H102 was a Stx2j-producing E. coli. Despite similarity among the sequences of Stx2j encoding prophages, phylogenetic analysis revealed that stx2j-positive E. coli genomes are diverse. 2021H102 clustered with genomes identified in the United States by core-genome single nucleotide variant-based phylogenetic analysis, implying that 2021H102 may have been an imported case. Several detection PCR primers failed to amplify stx2j, implying that stx2j-positive STEC might not be detected in some clinical laboratories. To our knowledge, this is the first report of Stx2j-producing E. coli isolated from outside of North American continent.
It is increasingly important to understand the response of freshwater communities and ecosystems to fertilizers given their widespread usage and the propensity for these fertilizers to runoff into rivers and lakes. Dispersal, an important ecological factor mediated by landscape connectivity, could potentially counteract the impacts of anthropogenic stressors through the reintroduction of communities unperturbed by local stressors. However, this potential has not yet been studied in the context of nutrient stressed natural communities. Here, we investigate the impacts of nutrient enrichment and connectivity on freshwater bacterioplankton communities. We subjected mesocosms stocked with native bacterioplankton communities to different combinations of nutrient enrichment and connectivity (volumes of water transferred between mesocosms). We show that nutrient enrichment strongly structures the bacterioplankton community, favoring nutrient tolerant taxa and depressing taxonomic diversity. Connectivity, however, interacts with nutrient enrichment to restore functional diversity in communities subjected to the highest levels of nutrient stress. Despite the ameliorating effects of dispersal, nutrient enrichment leaves a consistent signature in communities, driving a shift from more heterotrophic to more phototrophic communities. Taken together, our results demonstrate that while nutrient enrichment significantly impacts freshwater bacterioplankton communities, connectivity can help restore functional diversity to a certain extent.
Ceftriaxone tolerance in Neisseria gonorrhoeae is increasingly recognized as a phenotype that allows prolonged bacterial survival at inhibitory drug concentrations without altering minimum inhibitory concentrations (MICs). Host environmental factors are likely to influence such tolerance phenotypes, but experimental evidence remains limited. Zinc is a host-relevant transition metal that shapes bacterial stress responses and ribosomal function. Transcriptomic analyses have previously linked ceftriaxone tolerance in N. gonorrhoeae to a zinc-sparing ribosomal programme. In this study, we examined whether zinc availability modifies ceftriaxone killing kinetics in a paired tolerant and non-tolerant clinical isolate of N. gonorrhoeae. Both isolates had identical ceftriaxone MICs of 0.008 μg/mL. Using time-kill assays, we show that zinc supplementation attenuates early ceftriaxone-mediated killing in both phenotypes, resulting in shallower killing slopes and increased areas under the killing curve between 2 and 8 h after drug exposure. Zinc was associated with transiently higher viable counts during the mid-phase of killing, with a numerically greater effect observed in the tolerant isolate. Despite these differences in early and mid-phase dynamics, all ceftriaxone-exposed cultures were at or below the limit of detection by 24 h, irrespective of zinc availability. Within the limitations of this in vitro study using a single paired tolerant and non-tolerant isolate, these findings provide functional evidence that zinc availability can act as a conditional modifier of ceftriaxone killing dynamics in N. gonorrhoeae, influencing short-term survival without altering MICs or final outcomes.
In response to the loss of microbial efficiency caused by environmental stress in biomanufacturing, CRISPR-Cas gene editing technology has become a core tool for enhancing stress tolerance by accurately targeting genomic loci. This article systematically reviews the progress of its application. By optimizing engineered nucleases, gRNA design, and innovative delivery strategies, this technology successfully regulates key pathways in oxidative stress responses. It integrates functional genome screening with dynamic regulation to examine the networks of multi-gene collaborative tolerance. In the construction of high-stress-tolerant industrial chassis cells, the stress survival rate (>90% in Bacillus subtilis under thermal stress) and product synthesis ability (such as cellulose producing ethanol up to 4.5 g/l) of strains such as Escherichia coli and Corynebacterium glutamicum were significantly improved. Current challenges focus on delivery efficiency, off-target risks, and complex regulatory bottlenecks. In the future, the development of new editing tools and intelligent circuits will promote their industrial application in sustainable biomanufacturing.
A 3 × 3 factorial design was used to systematically evaluate the impact of three widely used emulsification methods [ultrasonication (UT), high-shear homogenization (HSH), or vortex mixing (VM)] at three treatment times (30, 60, or 120 s) on the antimicrobial efficacy of sage (SEO) and garlic (GEO) essential oil emulsions. Independent of essential oil used, antimicrobial activity was significantly (P < 0.05) influenced by both emulsification method and treatment time, with Gram-negative bacteria showing greater resistance. From the tested methods, emulsions prepared by UT or HSH for at least 60 s exhibited significantly (P < 0.05) higher antimicrobial activity, which might be associated with their significantly (P < 0.05) smaller particle size and polydispersity index. Additionally, UT and HSH produced more stable SEO emulsions, while GEO required ≥60 s treatment for stability and VM led to rapid destabilization regardless of oil or time. This study highlights the importance of selecting an appropriate emulsification method for producing essential oil emulsions with enhanced antimicrobial activity, which is important not only for antimicrobial testing but also for the overall functional performance of the emulsion.
Nutritional exchanges fuel the evolutionary and ecological dominance of multi-partner symbioses among reef-building corals and microbial associates in oligotrophic tropical marine ecosystems. Mutualistic relationships with endosymbiotic dinoflagellates (Family Symbiodiniaceae) are central to coral holobiont metabolism, yet their metabolic contributions are sensitive to nutrient availability. Symbiodiniaceae may compensate for oligotrophic environments via metabolic exchanges with prokaryotic partners. Bacterial production of ligands with affinities for otherwise insoluble elements promotes uptake and exchanges. Bacterial secretion of small molecules with high Ferric (Fe3+) iron affinities, herein referred to as siderophore production, presents one example of microbial metabolic cooperation. We isolated 78 pure bacterial culture lines from 14 Symbiodiniaceae cultures to screen for siderophore production using a Chrome Azurol S (CAS) overlay assay. Colorimetric changes observed on CAS overlays indicated ubiquitous siderophore production across 22 bacterial genera. Many of the isolated bacterial cultures corresponded to known 'core' Symbiodiniaceae microbiome. These results suggest an avenue of bacterial metabolism may facilitate biotic iron exchange among coral holobiont partners. Future characterization of the identity siderophores secreted will inform predictions on their impacts on iron exchange within the coral holobiont. Ultimately, a greater ability to acquire iron via siderophore production may improve the coral holobiont's tolerance to environmental stressors.
In vitro multispecies biofilm models are widely used to study oral diseases and to evaluate treatment strategies for conditions such as peri-implantitis, often relying on accurate species quantification to assess treatment efficiency. However, the influence of DNA extraction methodology on downstream quantitative analysis has not yet been addressed for such model systems. Here, we evaluated three mechanistically distinct protocols, a custom phenol-chloroform approach, and two commercial kits employing different lysis strategies. These were applied to planktonic cultures of six peri-implantitis-associated species individually, as well as to defined multispecies biofilms grown on implant surfaces. Pure culture DNA yields differed substantially between methods, revealing pronounced species-dependent variation. Species-specific quantification of biofilm replicates by quantitative PCR resulted in community profiles that appeared dominated by either commensal early- or pathogenic late-colonizers, depending solely on the extraction approach employed. These findings demonstrate that DNA extraction is a critical yet often overlooked variable, capable of fundamentally altering the apparent community composition of in vitro biofilm models. Our work is intended to serve as a warning, emphasizing the need for method validation and standardization when applying DNA-based community profiling to biofilm models before drawing conclusions on relative species abundances.
Antimicrobial resistance represents a major global health concern, particularly in countries where multidrug-resistant (MDR) pathogens are widespread. Biofilm formation further complicates therapeutic strategies. This study investigated the synergistic effects of combining norfloxacin (NOR) and sulfadiazine (SDZ) against three Escherichia coli strains: a reference, a quinolone-resistant clinical isolate, and a highly resistant extended-spectrum β-lactamase (ESBL)-producing strain. Checkerboard assays and isobolograms revealed synergistic or partially synergistic effects across all strains, with Fractional Inhibitory Concentration Index (FICI) values ranging from 0.37 to 1.0. Notably, the ESBL strain displayed enhanced synergy (FICI = 0.75) under white LED light irradiation. Reactive oxygen species (ROS) analysis showed that SDZ generated higher levels than NOR, particularly in the quinolone-resistant clinical isolate, while the NOR-SDZ combination yielded lower levels. Scanning electron microscopy of biofilms confirmed that the drug combination caused greater structural disruption than either monotherapy, especially at NOR (FIC × 100) and SDZ (FIC × 10). Subinhibitory monotherapies modulated the biofilm phenotype, underscoring the benefits of combined treatments. Overall, these findings highlight the NOR-SDZ combination as a promising therapeutic approach against MDR E. coli, where drug synergy and biofilm disruption emerge as key strategies to combat antimicrobial resistance.
Pseudomonas plecoglossicida is the etiological agent of visceral white spot disease, which induces significant mortality in economically important fish such as the large yellow croaker. In this study, we integrated recombinase polymerase amplification (RPA) with CRISPR/Cas12a-mediated detection to establish a fluorescence-based assay for rapid identification of P. plecoglossicida. The complete single-tube, two-stage RPA-CRISPR/Cas12a workflow can be performed within ~45 min. Using purified genomic DNA, the assay achieved an analytical detection limit of 1.65 copies μl-1 and showed no cross-reactivity with several other common fish pathogens. Its applicability was further evaluated using crude DNA extracts from spleen, liver, and kidney tissues of experimentally infected large yellow croakers. Overall, with its rapid turnaround, minimal equipment requirement, and high sensitivity, the RPA-CRISPR/Cas12a assay represents a promising diagnostic tool for rapid detection of P. plecoglossicida, thereby helping to control the spread of infection.
Ankylosing spondylitis (AS) is a chronic inflammatory disease with a global prevalence, primarily affecting the axial skeleton. While traditional therapies are often employed before surgical interventions, pharmacotherapy is typically reserved for cases of unstable AS condition. The gut microbiota has been implicated in numerous pathological conditions. However, its specific role in AS remains poorly understood. To address this knowledge gap, we applied Mendelian randomization (MR) to explore potential causal relationships between gut microbiota and AS. For this analysis, we utilized two large-scale genome-wide association study datasets: one comprising 18 340 individuals across 24 cohorts for human gut microbiota, and another consisting of 9069 AS cases and 1550 controls for AS susceptibility. Following heterogeneity testing, assessment of horizontal pleiotropy and application of multiple MR methods and 'leave-one-out' analysis, we identified 10 bacterial traits associated with AS risk. These include four risk-increasing factors: Actinobacteria (class), Streptococcaceae (family), Enterorhabdus (genus), and the Ruminococcaceae NK4A214 group (genus); and six risk-decreasing factors: Lactobacillaceae (family), Rikenellaceae (family), Anaerotruncus (genus), Eubacterium oxidoreducens group (genus), Howardella (genus), and Oscillibacter (genus). This study provides novel insights into the gut-spine axis and suggests potential avenues for AS management through microbiota-targeted interventions.
The lizard microbiome is a dynamic community that plays a crucial role in the health and survival of these animals. As global change poses significant threats to lizard populations around the world, understanding the interactions between lizards and their microbial communities is increasingly important. Here, we synthesize a rapidly growing body of research on the composition, diversity, transmission, and functional roles of lizard microbiomes. We discuss the implications of microbiome variation for lizard physiology, as well as the potential for microbiomes to inform conservation strategies for threatened species. Finally, we highlight priorities for future research, which include the need to quantify microbiome diversity and function across additional taxa, as lizards remain under-represented in the microbiome literature. We also stress the importance of experimental and field research that can reveal the adaptive significance of lizard microbiomes in the face of environmental change. Our synthesis highlights the contributions of lizard microbiome science to the fields of ecology, evolution, and conservation biology and demonstrates how the microbial communities that live in and on lizards enhance our understanding of their biodiversity and inform efforts to protect vulnerable populations.
Salmonella is a major zoonotic bacterial pathogen with significant impacts on public health, food safety, and veterinary medicine. The genus Salmonella includes exactly two recognized species, S. enterica and S. bongori. Most routine molecular surveillance assays target conserved genus-level genes, such as invA, which do not distinguish Salmonella enterica and Salmonella bongori, potentially obscuring species-specific ecology and source attribution. Here, we present the first introduction of multiple accessory-genes sequence typing (MAST), a pan-genome-guided framework for binary (two species) bacterial PCR target screening, primer design, and experimental validation, demonstrated here in Salmonella. Applying MAST to 2236 high-quality Salmonella genomes, we identified sopD2 as a species-discriminative locus. sopD2-targeted primers with the highest MAST Diff score (= 0.999) demonstrated high analytical specificity for Salmonella, with no cross-amplification observed against Escherichia coli, Streptococcus suis, or Pasteurella multocida, and enabled clear PCR differentiation of S. enterica from S. bongori across laboratory isolates. Taken together, our results position MAST as a broadly applicable and practical pipeline for species-level PCR marker discovery across diverse bacterial taxa and validate sopD2 as a robust target for discriminating Salmonella species, enhancing the accuracy of molecular surveillance and the clarity of source attribution.
Given the ability of microorganisms to develop drug resistance at a faster pace than the development of new drugs, it becomes urgent to seek alternatives capable of restoring the efficacy of antibiotics, especially against pathogens such as Staphylococcus aureus, known for its versatility in expressing resistance mechanisms. Synthetic chalcones have been extensively investigated as efflux pump inhibitors due to their structural flexibility and ease of chemical modification. Can cite studies that show chalcones derived from Croton anisodontus for their ability to inhibit the NorA and MepA efflux pumps in S. aureus strains (1199B and K2068) and studies chalcones act as inhibitors of the NorA efflux pump in whole cells and everted membrane vesicles of S. aureus. Among these mechanisms, the NorA and MepA efflux pumps stand out, as they are responsible for expelling antibiotics from the bacterial cell interior. In this context, the present study investigated the potential of the chalcone (E)-1-(3-aminophenyl)-3-(4-chlorophenyl)prop-2-en-1-one as a possible inhibitor of these efflux systems. Initial screening was conducted through in silico studies, followed by in vitro assays, determination of minimum inhibitory concentration (MIC), and fluorescence tests using ethidium bromide EtBr and SYTOX Green. Molecular docking results revealed that this chalcone interacts with amino acid residues described in the literature as essential for efflux pump activity (Glu222 and Phe303 in NorA; Tyr35 and Met172 in MepA). The binding distances and energies obtained were consistent with potential inhibitory activity, suggesting that chalcone may interfere with the functionality of these proteins. In vitro assays showed a significant reduction in the MIC of antibiotics such as norfloxacin and ciprofloxacin in resistant strains, indicating a potentiating effect. Furthermore, fluorescence tests demonstrated increased retention of ethidium bromide by up to 736.3% for NorA and 234.8% for MepA, supporting the inhibition of efflux pumps. Increased bacterial membrane permeability was also observed, evidenced by an increase in SYTOX Green fluorescence by 70-855%. Although the results demonstrate the potential of chalcone as a therapeutic adjuvant, the ADMET analyses revealed toxicological alerts that require further investigation. These findings suggest that rational structural modifications may be necessary to ensure its safety and efficacy in combating resistant bacterial infections.
Barley (Hordeum vulgare) is susceptible to Puccinia hordei (leaf rust), a biotrophic foliar pathogen contributing to global yield losses. With rising food demand and increasing disease pressure, sustainable crop protection strategies are urgently needed to support UN Sustainable Development Goal 2: Zero Hunger. Arbuscular mycorrhizal (AM) fungi, including Rhizophagus irregularis, form symbioses with barley roots and can modulate host immunity through mycorrhiza-induced resistance (MIR). Here, we tested whether R. irregularis colonization alters barley growth and defence responses during P. hordei infection. AM fungal colonization did not significantly reduce disease severity or mitigate pathogen-associated biomass loss at a single post-infection time point. However, co-infected plants showed enhanced expression of defence genes (PR1, PR2, PR3, and WRKY28), which remained low in plants colonized by AM fungi alone, consistent with immune priming. RNA sequencing revealed AM fungal-associated reprogramming of the leaf transcriptome, including enrichment of defence, metabolism and ubiquitination-related processes. These results indicate that R. irregularis reshapes barley immune regulatory networks at transcriptional and post-translational levels. Although these molecular changes did not translate into measurable phenotypic protection within the short experimental timeframe, they highlight the complexity and context dependence of MIR in cereal-rust interactions.