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Fault zones exhibit 3D variable thickness, a feature that remains inadequately explored, particularly with regard to the impact on fluid flow. Upon analyzing an analytic solution, we examine 3D thermal‐hydraulic (TH) dynamical models through a benchmark experiment, which incorporates a fault zone with thickness variations corresponding to realistic orders of magnitude. The findings emphasize an area of interest where vigorous convection drives fluid flow, resulting in a temperature increase to 150°C at a shallow depth of 2.7 km in the thickest sections of the fault zone. Moreover, by considering various tectonic regimes (compressional, extensional, and strike‐slip) within 3D thermal‐hydraulic‐mechanical (THM) models and comparing them to the benchmark experiment, we observe variations in fluid pressure induced by poroelastic forces acting on fluid flow within the area of interest. These tectonic‐induced pressure changes influence the thermal distribution of the region and the intensity of temperature anomalies. Outcomes of this study emphasize the impact of poroelasticity‐driven forces on transfer processes and highlight the importance of addressing fault geometry as a crucial parameter in future investigations of fluid flow in fractured systems. Such research has relevant applications in geothermal energy, CO2 storage, and mineral deposits. Plain Language Summary: Exploring critical parameters affecting the fluid flow within fault zones in the Earth's crust is of fundamental scientific and economic interest. Among them, fault zone thickness and tectonic regimes are two parameters whose role remains unexplored. The results of this study show that fluid flow by convection is vigorous in the zone where fault thickness is largest. In this area of interest, the tectonic regimes impact convective dynamics and modify the thermal distribution of the system. These generic outcomes are discussed in relation to real‐cases scenarios and emphasize the fundamental role of considering a Geometric parameter (G), the fault zone thickness variation, as well as tectonic regimes, during the exploratory phase for geothermal energy, CO2 storage, and mineral exploration. Key Points: Based on a critical Rayleigh number analysis and 3D numerical modeling, we explore the effects of fault zone thickness variations on fluid flowIn line with the critical Rayleigh number analysis, our numerical results reveal that thermal convection is more efficient in areas where the fault zone thickness is largestThe poroelasticity‐driven force alters the convective dynamics within the zone of interest in the fault zone (i.e., the thickest part) [ABSTRACT FROM AUTHOR]

Antibiotic resistance is frequently observed shortly after the clinical introduction of an antibiotic. Whether and how frequently that resistance occurred before the introduction is harder to determine, as isolates could not have been tested for resistance before an antibiotic was discovered. Historical collections, like the British National Collection of Type Cultures (NCTC), stretching back to 1885, provide a window into this history. Here we match 1,817 sequenced high-quality genomes from the NCTC collection to their respective year of isolation to study resistance genes before and concurrent with the age of antibiotics. Concordant with previous work, we find resistance genes in both pathogens and environmental samples before the age of antibiotics. While generally rare before the introduction of an antibiotic, we find an associated increase in frequency with antibiotic introduction. Finally, we observe a trend of resistance elements becoming both increasingly mobile and nested within multiple mobile elements as time goes on. More broadly, our findings suggest that likely-functional antibiotic resistance genes were circulating in clinically relevant isolates before the age of antibiotics, but human usage is associated with increasing both their overall prevalence and mobility., Data Summary: Genome assemblies downloaded and analyzed are in Supplementary Table 1, and computational tools used are found in the Methods. The authors confirm all supporting data, code and protocols have been provided within the article or through supplementary data files., Impact Statement: Historical collections of microbial isolates enable researchers to both investigate the past and identify interesting trends over time. In this study, we queried over 1,800 isolate genomes in one such collection for genomic variation linked to antibiotic resistance. We show that numerous isolates cultured before the introduction of a given antibiotic contain genomic variation linked to antibiotic resistance; however, this phenomenon remained relatively rare. We demonstrate a strong association between the year a given antibiotic was clinically introduced and a rise in prevalence of genomic resistance to that antibiotic. Finally, we show that while mobile elements are common throughout the isolates and timeframe analyzed, genomic resistance has become increasingly mobile as time has gone on. This study shows that as expected, the clinical introduction of a given antibiotic is correlated with an increase in resistance to that antibiotic but also was linked with increased mobility of genes and alleles conferring resistance. However, we note that the effect of deposition bias in the collection cannot be excluded. Our work also indicates that numerous microbial pangenomes of pathogens naturally contained genomic resistance to a given antibiotic even before anthropogenic use of that antibiotic. Taken together, we demonstrate that although human use may affect the prevalence and mobility of genomic resistance in clinical isolates; for most antibiotics, genomic resistance existed within the pangenomes of sampled pathogens prior to clinical introduction. Quantifying and understanding the impact of antibiotic introduction in the past helps us understand how the introduction of novel antibiotics can impact bacteria; allowing better reaction to novel resistant infections as they arise.

Tackling the challenge created by antibiotic resistance requires understanding the mechanisms behind its evolution. Like any evolutionary process, the evolution of antimicrobial resistance (AMR) is driven by the underlying variation in a bacterial population and the selective pressures acting upon it. Importantly, both selection and variation will depend on the scale at which resistance evolution is considered (from evolution within a single patient to the host population level). While laboratory experiments have generated fundamental insights into the mechanisms underlying antibiotic resistance evolution, the technological advances in whole genome sequencing now allow us to probe antibiotic resistance evolution beyond the lab and directly record it in individual patients and host populations. Here we review the evolutionary forces driving antibiotic resistance at each of these scales, highlight gaps in our current understanding of AMR evolution, and discuss future steps toward evolution-guided interventions.

Background: Antimicrobial resistance (AMR) poses a serious threat to global healthcare, and inadequate education has been identified as a major challenge by the WHO. The human , animal and agricultural sectors contribute to the emergence of AMR. Gamification has emerged as an innovative tool to improve knowledge and change behaviours. Our study provides an overview of the literature on existing games in prescribers' education across the One Health sectors, with a particular focus on the impact of gamification on learning., Methods: Using the PRISMA guidelines, we searched Cochrane, PubMed, Scopus and Google Scholar for articles related to gamification for future prescribers of antimicrobials from inception until 28 March 2023. Retrieval and screening of articles was done using a structured search protocol with strict inclusion/exclusion criteria., Results: A total of 120 articles were retrieved, of which 6 articles met the inclusion criteria for final analysis. High-income countries had the most studies, with one global study incorporating low- to middle-income countries. All games were evaluated in the human sector. Board and card games, featuring scoring and point systems, were the most prevalent game types. Most games focused on improving knowledge and prescribing behaviours of medical students, with bacteria or antibiotics as the only content. All studies highlighted the significant potential of gamification in mitigating AMR, promoting antimicrobial stewardship, and improving retention of information compared with conventional lectures., Conclusions: Our review found an absence of studies in the animal and environmental sectors, disproportionately focused on medical students with questionable sample size, inadequate assessment of game content and effectiveness, and opportunities for game developers., (© The Author(s) 2023. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy.)

Integrons are mobile genetic elements that have played an important role in the dissemination of antibiotic resistance. Under stress, the integron can generate combinatorial variation in resistance cassette expression by cassette reshuffling, accelerating the evolution of resistance. However, the flexibility of the integron integrase site recognition motif hints at potential off-target effects of the integrase on the rest of the genome that may have important evolutionary consequences. Here, we test this hypothesis by selecting for increased-piperacillin-resistance populations of Pseudomonas aeruginosa with a mobile integron containing a difficult-to-mobilize β-lactamase cassette to minimize the potential for adaptive cassette reshuffling. We found that integron activity can decrease the overall survival rate but also improve the fitness of the surviving populations. Off-target inversions mediated by the integron accelerated plasmid adaptation by disrupting costly conjugative genes otherwise mutated in control populations lacking a functional integrase. Plasmids containing integron-mediated inversions were associated with lower plasmid costs and higher stability than plasmids carrying mutations albeit at the cost of a reduced conjugative ability. These findings highlight the potential for integrons to create structural variation that can drive bacterial evolution, and they provide an interesting example showing how antibiotic pressure can drive the loss of conjugative genes. IMPORTANCE Tackling the public health challenge created by antibiotic resistance requires understanding the mechanisms driving its evolution. Mobile integrons are widespread genetic platforms heavily involved in the spread of antibiotic resistance. Through the action of the integrase enzyme, integrons allow bacteria to capture, excise, and shuffle antibiotic resistance gene cassettes. This integrase enzyme is characterized by its ability to recognize a wide range of recombination sites, which allows it to easily capture diverse resistance cassettes but which may also lead to off-target reactions with the rest of the genome. Using experimental evolution, we tested the off-target impact of integron activity. We found that integrons increased the fitness of the surviving bacteria through extensive genomic rearrangements of the plasmids carrying the integrons, reducing their ability to spread horizontally. These results show that integrons not only accelerate resistance evolution but also can generate extensive structural variation, driving bacterial evolution beyond antibiotic resistance.

Accurate surveillance of the COVID-19 pandemic can be weakened by under-reporting of cases, particularly due to asymptomatic or pre-symptomatic infections, resulting in bias. Quantification of SARS-CoV-2 RNA in wastewater can be used to infer infection prevalence, but uncertainty in sensitivity and considerable variability has meant that accurate measurement remains elusive. Here, we use data from 45 sewage sites in England, covering 31% of the population, and estimate SARS-CoV-2 prevalence to within 1.1% of estimates from representative prevalence surveys (with 95% confidence). Using machine learning and phenomenological models, we show that differences between sampled sites, particularly the wastewater flow rate, influence prevalence estimation and require careful interpretation. We find that SARS-CoV-2 signals in wastewater appear 4-5 days earlier in comparison to clinical testing data but are coincident with prevalence surveys suggesting that wastewater surveillance can be a leading indicator for symptomatic viral infections. Surveillance for viruses in wastewater complements and strengthens clinical surveillance, with significant implications for public health., (© 2022. Crown.)

The COVID-19 pandemic has put unprecedented pressure on public health resources around the world. From adversity, opportunities have arisen to measure the state and dynamics of human disease at a scale not seen before. In the United Kingdom, the evidence that wastewater could be used to monitor the SARS-CoV-2 virus prompted the development of National wastewater surveillance programmes. The scale and pace of this work has proven to be unique in monitoring of virus dynamics at a national level, demonstrating the importance of wastewater-based epidemiology (WBE) for public health protection. Beyond COVID-19, it can provide additional value for monitoring and informing on a range of biological and chemical markers of human health. A discussion of measurement uncertainty associated with surveillance of wastewater, focusing on lessons-learned from the UK programmes monitoring COVID-19 is presented, showing that sources of uncertainty impacting measurement quality and interpretation of data for public health decision-making, are varied and complex. While some factors remain poorly understood, we present approaches taken by the UK programmes to manage and mitigate the more tractable sources of uncertainty. This work provides a platform to integrate uncertainty management into WBE activities as part of global One Health initiatives beyond the pandemic., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

Mobile integrons are widespread genetic platforms that allow bacteria to modulate the expression of antibiotic resistance cassettes by shuffling their position from a common promoter. Antibiotic stress induces the expression of an integrase that excises and integrates cassettes, and this unique recombination and expression system is thought to allow bacteria to 'evolve on demand' in response to antibiotic pressure. To test this hypothesis, we inserted a custom three-cassette integron into Pseudomonas aeruginosa and used experimental evolution to measure the impact of integrase activity on adaptation to gentamicin. Crucially, integrase activity accelerated evolution by increasing the expression of a gentamicin resistance cassette through duplications and by eliminating redundant cassettes. Importantly, we found no evidence of deleterious off-target effects of integrase activity. In summary, integrons accelerate resistance evolution by rapidly generating combinatorial variation in cassette composition while maintaining genomic integrity., Competing Interests: CS, JE, RM No competing interests declared, (© 2021, Souque et al.)

Intracellular pathogens include all viruses, many bacteria and parasites capable of invading and surviving within host cells. Key to survival is the subversion of host cell pathways by the pathogen for the purpose of propagation and evading the immune system. The intracellular bacterium Shigella flexneri, the causative agent of bacillary dysentery, invades host cells in a vacuole that is subsequently ruptured to allow growth of the pathogen within the host cytoplasm. S. flexneri invasion has been classically described as a macropinocytosis-like process, however the underlying details and the role of macropinosomes in the intracellular bacterial lifestyle have remained elusive. We applied dynamic imaging and advanced large volume correlative light electron microscopy (CLEM) to study the highly transient events of S. flexneri's early invasion into host epithelial cells and elucidate some of its fundamental features. First, we demonstrate a clear distinction between two compartments formed during the first step of invasion: the bacterial containing vacuole and surrounding macropinosomes, often considered identical. Next, we report a functional link between macropinosomes and the process of vacuolar rupture, demonstrating that rupture timing is dependent on the availability of macropinosomes as well as the activity of the small GTPase Rab11 recruited directly to macropinosomes. We go on to reveal that the bacterial containing vacuole and macropinosomes come into direct contact at the onset of vacuolar rupture. Finally, we demonstrate that S. flexneri does not subvert pre-existing host endocytic vesicles during the invasion steps leading to vacuolar rupture, and propose that macropinosomes are the major compartment involved in these events. These results provide the basis for a new model of the early steps of S. flexneri epithelial cell invasion, establishing a different view of the enigmatic process of cytoplasmic access by invasive bacterial pathogens.