This work experimentally and numerically explored how varied steel-polypropylene fibre mixtures affected simply supported reinforced concrete deep beams. Due to their better mechanical qualities and durability, fibre-reinforced polymer composites are becoming more popular in construction, with hybrid polymer-reinforced concrete (HPRC) promising to increase the strength and ductility of reinforced concrete structures. The study evaluated how different combinations of steel fibres (SF) and polypropylene fibres (PPF) affected beam behaviour experimentally and numerically. The study’s focus on deep beams, research of fibre combinations and percentages, and integration of experimental and numerical analysis provide unique insights. The two experimental deep beams were the same size and were composed of hybrid polymer concrete or normal concrete without fibres. Fibres increased deep beam strength and ductility in experiments. The calibrated concrete damage plasticity model in ABAQUS was used to numerically calibrate HPRC deep beams with different fibre combinations at varied percentages. Based on six experimental concrete mixtures, calibrated numerical models of deep beams with different material combinations were investigated. The numerical analysis confirmed that fibres increased deep beam strength and ductility. HPRC deep beams with fibre performed better than those without fibres in numerical analysis. The study also determined the best fibre percentage to improve deep beam behaviour where a combination of 0.75% SF and 0.25% PPF was recommended to enhance load-bearing capacity and crack distribution, while a higher content of PPF was suggested for reducing deflection. The deep reinforced concrete beams are commonly used, which are structural components with large depths relative to their span. They can withstand bending moments, shear stresses, and deflection because of their thick sections. Shear, flexure, and compression all interact, making for a challenging design environment when working with deep reinforced concrete beams. Strut-and-tie models or finite element analysis are commonly used when constructing deep beams out of reinforced concrete, with consideration given to factors such as the location and size of the supports, the type and amount of reinforcing steel, and the shear and flexural strengths of the concrete. Several models for estimating the shear strength of deep beams made of reinforced concrete (RC) with and without web reinforcement were analysed by Shahnewaz et al. . Each moHybrid concrete made from steel and polypropylene (PP) fibres combines the advantages of both types of fibre-reinforced concrete. Steel fibres improve the concrete’s strength and ductility, while polypropylene fibres make it more durable and crack-resistant. In hostile environments where corrosion is a key problem, steel-PP hybrid concrete showed encouraging results in increasing the mechanical characteristics and performance of concrete structures. Bridge decks, pavements, and earthquake-resistant structures are just some of the places one might see this tough stuff put to use. Steel-PP hybrid concrete is a ground-breaking innovation in the industry, with the potential to greatly enhance the functionality and durability of concrete buildings. Crack arresting in reinforced concrete was a topic investigated by Qian and Stroeven . The totIn structural engineering, the concrete damage plasticity (CDP) model is a popular numerical model for evaluating the response of concrete structures to different loads. Concrete’s complicated behaviour, including cracking, crushing, and plastic deformation, are all accounted for in the CDP model, which is a continuum-based model. The post-peak behaviour of concrete may be captured by this non-linear model, making it a vital tool for the design and analysis of structures subjected to severe stresses. The CDP model was effectively implemented in several contexts, such as earthquake engineering, blast-resistant building design, and reinforced concrete construction. particularly in earthquake-prone places where ductility is of crucial concern, this finding may have significant consequences for the design and construction of reinforced concrete structures. The concrete damage plasticity (CDP) model was first proposed by Lubliner et al. , and it The use of the CDP (concrete damaged plasticity) model in modelling fibre hybrid concrete is a suitable approach to capture the behaviour of such materials. The CDP model is widely used in simulating the response of concrete structures, including fibre-reinforced concrete. Several studies took into consideration modelling fibres inside concrete where having an experimental study will lead to high cost and time consumption. The behaviour of hybrid concrete is reflected by presenting CDP parameters that were changed to fit the required compression and tension behaviour ,17,18,19Ibrahim and Rad  used a pThis study aims to investigate the effect of using different mixes of steel fibres (SF) and polypropylene fibres (PPF) on the behaviour of simply supported reinforced concrete deep beams, both experimentally and numerically. The study aims to determine the most effective percentage of fibres for improving the strength and ductility of the deep beams. The novelty of the study was highlighted in the investigation of hybrid polymer reinforced concrete (HPRC) and the use of a numerical calibration process. The numerical process was held using the concrete damage plasticity (CDP) model in ABAQUS to investigate the behaviour of HPRC deep beams with different material combinations based on the experimental results. Firstly, two reinforced deep beams were tested experimentally, one with normal concrete (0%SF–0%PPF) and the other with (75%SF–25%PPF). The test results were then compared to study the effect of fibre existence. Additionally, six concrete mixes were prepared and tested to obtain the concrete parameters for each, with different percentages of SF and PPF, including (0%SF–0%PPF), (1%SF–0%PPF), (0.75%SF–0.25%PPF), (0.5%SF–0.5%PPF), (0.25%SF–0.75%PPF), and (0%SF–1%PPF). Numerical models were then calibrated using ABAQUS based on the experimental results, and calibrated CDM parameters were used to match the experimental concrete behaviour. The calibrated models were then compared, taking into account ultimate load, deflection, cracking patterns, and concrete damage in tension and compression, to investigate the most effective percentage of fibres for improving the strength and ductility of the deep beams. The study aimed to explore the potential of hybrid polymer-reinforced concrete (HPRC) to enhance the strength and ductility of reinforced concrete structures using a combination of steel and polypropylene fibres. The recent studies did not cover the behaviour of deep beams with these ranges of fibres percentages; also, the presented comparisons showed the compression and tension behaviour of the tested mixes in details by presenting stress–strain relationships. Moreover, this study showed the significance of having hybrid concrete on the concrete and steel damages using ABAQUS.This study’s significance lies in the insights it provides for improving fibre-reinforced polymer composites for building applications. The research shed light on the performance of HPRC in reinforced concrete deep beams and how it might be improved. Engineers and researchers can use the results as a guide when deciding which fibre combinations and percentages to use to achieve specific structural goals. There are several highlighting features of this study that make it a novel study. It begins by investigating deep beams experimentally, a specific structural feature that plays a significant role in many different building projects and where hybrid polymer concrete can be useful. To further our comprehension of how various fibre combinations affect the behaviour of deep beams, this study also examined the combined impacts of SF and PPF, taking varying percentages into account; these percentages were derived by testing several specimens and then, their tension and compression behaviours were investigated. Finally, experimental testing and numerical analysis were utilised in this study for a more thorough evaluation and validation of the findings, which increases the study’s credibility and practicality.After this introduction, The concrete damage plasticity (CDP) model is a constitutive model used to simulate the nonlinear behaviour of concrete subjected to loading, such as compression, tension, and bending. It is based on the concept of continuum damage mechanics, which considers that the material undergoes damage and degrades as it is subjected to loading. It considers two different types of damage: tensile and compressive. In the tensile damage, the model assumes that the material undergoes microcracking, which leads to a reduction in the material’s stiffness and strength. In compressive damage, the model assumes that the material undergoes crushing, which leads to a reduction in the material’s stiffness and strength. It also considers the effects of plasticity, which means that the material undergoes permanent deformation even after the load is removed. The model uses an isotropic hardening rule to capture the plastic behaviour of the material. To implement the CDP model, a set of parameters is required to describe the material’s behaviour. These parameters include tensile and compressive strengths, Young’s modulus, and others which are typically obtained through experimental testing.Detailed explanations of the procedure can be found in the relevant publications and monographs. In this article, a quick overview of the concrete constitutive model is provided and then extended in a few key ways. The entire strain tensor value il below ,21,22,23The scalar damage elasticity equation also gives us insight into the interactions between forces and strains within a material:The scalar reduction relation relates the internal force The effective internal force This internal force–strain relationship forms the constitutive model of damage plasticity:Concrete’s uniaxial compressive and tensile reaction to compression and tension stress, as described by the concrete damage plasticity model, is shown by:Using the expressions: Dilatation angle: The dilatation angle is a parameter that characterizes the volumetric change in a material during plastic deformation. It represents the angle between the direction of the maximum principal stress and the direction of maximum volume change. In Abaqus CDP, the dilatation angle is used to control the volumetric response of the material, particularly in compression.Eccentricity: The eccentricity parameter is used to describe the anisotropic behaviour of a material during plastic deformation. It represents the deviation of the plastic strain from the direction of maximum shear stress and can be used to control the material’s yield surface shape in the stress space.fb0/fc0: fb0 and fc0 are parameters used to describe the initial compressive and tensile strength of concrete in CDP models. They are used to control the initial stiffness and strength of the material and are typically determined experimentally.K: The isotropic hardening parameter “k” is used to describe the hardening behaviour of a material during plastic deformation. It determines the rate at which the yield stress increases with plastic strain or plastic strain rate and is an important parameter in accurately simulating the material’s plastic behaviour under various loading conditions.Viscosity parameter: The viscosity parameter is used to describe the material’s viscous behaviour, which is the ability of a material to resist deformation over time when subjected to a constant load. In CDP models, the viscosity parameter can be used to control the rate at which the material’s yield stress increases with increasing strain rate, or to simulate the material’s creep behaviour.In conclusion, the calibrated model represents tension and compression concrete behaviour using the CDP constitutive model, whereas these two behaviours are shown utilising the concrete’s properties in tension and compression according to The behaviour of a deep beam reinforced with steel and polypropylene fibres was studied through experiments conducted at the University of Technology’s lab. The experiments involved the construction and testing of a pair of deep, simply supported beams subjected to symmetric two-point loading. The overall height (h) of the two beams was 400 mm, the width (b) was 150 mm, the total length (L) was 1400 mm, and the clear span (Ln) was 1070 mm. The Ln/h ratio was equal to 2.675, which was less than the 4 that is recommended by the provision of the ACI 318M-14 code . The pri® Company’s hooked end SF class C, type A was used throughout the experimental procedure. In addition, the manufacturer provided data in As can be seen in Furthermore, deformed longitudinal steel bars with a nominal diameter of 16 mm were used in this investigation, and deformed steel with a diameter of 4 mm was used for both vertical and horizontal shear reinforcement. The characteristics of the steel bars used in this experiment are shown in Steel-polypropylene fibres added to concrete might have both positive and negative results depending on the specific circumstances. Adding steel fibres to concrete is a common practice for increasing the material’s strength and durability. Steel fibres enhance concrete’s tensile strength and toughness, which helps prevent cracking and boosts the material’s overall performance. On the other side, polypropylene fibres are frequently utilised to limit concrete’s potential to crack. They do not do much to boost the concrete’s strength, but they can keep it from shrinking and splitting as much. Polypropylene fibres are sometimes added to concrete mixes to improve durability and reduce the potential for cracking, both of which can help to maintain the compressive strength of the concrete over time, despite the fact that they can reduce the stiffness of the concrete and, thus, lead to a lower modulus of elasticity. However, the concrete’s compressive strength can be reduced if an excessive number of polypropylene fibres is added during the mixing process. This is due to the fact that the fibres may form air pockets in the concrete, thus weakening the material.Adding steel fibres to concrete increases its tensile strength and makes it less likely to crack in tension. As can be seen in The beams that were tested experimentally in this study had the properties shown in Generally, it can be seen that the type of the crack spread was different for the two tested beams, the crack that caused the failure in beam M1B seemed to be wide and straight, accompanied by compression crushing of concrete near the loading area; on the other hand, the beam with fibres seemed to have a different crack initiating method, where the crack that caused the failure consisted of small tight cracks and no compression cracks are presented. In this, the effect of the fibres occurred in tightening the cracks and increasing the ultimate load, as presented in In order to further investigate the effect of having different fibre percentages, this section considered validating the two tested deep beams using ABAQUS. The hybrid concrete beam M3B was validated using the concrete damage plasticity (CDP) model by having a calibration process in order to obtain the parameters that reflect the existence of the effect of the fibres on the beam behaviour. Starting with defining the material properties of the hybrid concrete, such as elastic modulus, Poisson’s ratio and material strength and damage parameters. Then, calibrating the CDP model for the hybrid concrete using experimental data, such as the stress–strain curves or the test results. Finally, fitting the damage parameters to the experimental data to calibrate the model to the actual behaviour of the hybrid concrete beams. Moreover, the six mixes presented in The 3D finite element models were developed using data from laboratory experiments. Using the CDP model to represent concrete within ABAQUS , six simThe finite element model was constructed from concrete and reinforcement materials, with the concrete represented by a solid element with eight nodes  and the reinforcement bars represented by beam elements with a 2-node linear beam in space (B31: Timoshenko beam). An embedded zone was used to mimic the link between the longitudinal and transverse reinforcements and the concrete, while the boundaries were set so that they would be equivalent to those observed in a laboratory setting. In addition, identical experimental circumstances were achieved by applying a vertical concentrated load at each point load of the beam and then distributing the loads via the coupling effect. Both numerical accuracy and computation time can be affected by the mesh size, and so, a size study was performed to see how different mesh sizes performed and to choose the most suitable size. The optimal mesh size was then applied to provide a precise result, with the total number of elements equal to 25,110 for the beams.The numerical models calibrated with the CDP model in ABAQUS showed good agreement with the experimental results, validating the use of the model for predicting the behaviour of HPRC deep beams as shown in M1B (0%SF–0%PPF): This is the fibre-free control beam. It has the lowest loading capacity where the absence of fibre reinforcement makes the beam more subject to fracture under stress. In comparison to fibre-reinforced beams, the damaged area seems more numerous (more red areas). Moreover, in this situation, the beam is more subject to deflection under load, increasing sagging and deformation. Therefore, beam M1B has the lowest load capacity and the highest deflection values when compared to the other mixtures even though the M1 mix has a better concrete compression strength than M3, M4, M5, and M6, but the absence of fibres was reflected by the general behaviour of the beam. Moreover, reinforcement steel bars undergo high stresses especially in the flexural area and along the major shear-damaged area.M2B (1%SF–0%PPF): Adding 1% SF to concrete improves its tensile and flexural strength and enhances its resistance to cracking. However, it may also lead to an increase in brittleness. It leads to smaller and less frequent cracks compared to the control beam without fibres. The flexural damage that appeared may be narrower and more tightly spaced. However, it does not cover the whole part, as was observed in the M1B model. The addition of SF helped to reduce deflection by providing additional stiffness and strength to the beam. The addition of 1% SF increased the load capacity of the beam remarkably, due to the additional tensile strength provided by the fibres. This mix has enhancement in the general behaviour of the beam if compared with the M1B case, especially in the flexural part; however, it was not the best tested case. In addition, it can be seen that the longitudinal reinforcement bars in the mid span flexural area are not damaged even after the failure, which reflects the steel fibres’ effectivity in tension strength.M3B (0.75%SF–0.25%PPF): Adding 0.25% PPF to the mix in addition to 0.75% SF can further improve the toughness and ductility of the material compared to the mix with 1% SF only. The addition of 0.25% PPF can affect the brittleness of the concrete. The damage that appeared in the middle of the beam seems a bit higher than in the previous case, however, it is still better if compared with the M1B case. The combined effect of SF and PPF improved the stiffness and resistance to deflection of the beam, as the PPF helps to absorb energy and reduce the risk of brittle failure. That is why M3B gave the best results in terms of load capacity between all the tested models. It can be seen also that the reinforcement steel damage was not too severe if compared with the M1B model.M4B (0.5%SF–0.5%PPF): Using equal percentages of SF and PPF can balance the properties of the concrete and provide improved flexural strength, ductility, and resistance to cracking and fatigue. The combination of SF and PPF can lead to a more balanced distribution of cracks across the beam, with smaller and less frequent cracks compared to the control beam (without fibres). The balanced combination of SF and PPF can provide a further improvement in load capacity, as the fibres work together to resist the applied loads if compared with the M1 mix. However, this improvement in the load capacity is not the best if compared with the M3 mix, where it can be seen that the results curves from now on for the next beams will have less loading capacity but less deflection values also.M5B (0.25%SF–0.75%PPF): Using a higher percentage of PPF (0.75%) with a lower percentage of SF (0.25%) can increase the toughness and energy absorption of the material. The higher percentage of PPF produced smaller deflection values, with a more occurrence of the damaged area in the mid-span of the beam. However, reducing the SF and using higher PPF weakened the general behaviour of the beam, where it can be seen that the longitudinal reinforcement started to present some damaged areas.M6B (0%SF–1%PPF): Adding 1% PPF to the mix can enhance the durability of the material and improve its resistance to shrinkage cracking, as well as provide some enhancement to its flexural strength. Beam M6B has a better load capacity than M1B, but it comes last if compared with the other hybrid beams even though it has the best enhanced and reduced deflection value among all the six mixes. Additionally, in this model, the damaged areas in the shear part happen to be less intensive if compared with all other cases, but it has the highest damaged flexural area among the hybrid models. On the other hand, because of the absence of steel fibres, the longitudinal reinforcement in this case is almost totally damaged.The addition of different percentages of steel fibres (SF) and polypropylene fibres (PPF) to concrete had various effects on the crack pattern, load capacity, deflection, and damage of reinforced concrete beams. After the validation process, the six beams were modelled and tested numerically, revealing interesting results. Load deflection curves are presented in The effect of different percentages of steel fibres (SF) and polypropylene fibres (PPF) on the behaviour of concrete beams was evaluated in terms of their impact on crack pattern, load-carrying capacity, deflection, and steel and concrete damage. In summary, the addition of fibres to concrete can significantly affect the crack pattern and the damage behaviour of concrete beams. It also improves the load-carrying capacity of the beam by enhancing its strength and reduces the deflection, where fibres existence affect the deflection of the beams by changing its stiffness and ductility. In general, the addition of fibres can increase the beam’s stiffness, which can reduce the amount of deflection under loading. However, the effect of the fibre content on the load-carrying capacity can be complex, as an increase in fibre content can also lead to an increase in brittleness, which can reduce the overall strength of the material; this occurred in the numerical models where increasing PPF percentage weakened the beams but not using any PPF percentages did not give the best behaviour among the tested models. Regarding steel and concrete damage, the use of fibres can also affect the damage behaviour of the steel and concrete in the beam. SF can improve the behaviour of steel bars and concrete in flexural areas. On the other hand, PPF can reduce the damage a bit in the shear area but it has almost lighter effect on the flexural parts.By investigating the effect of different mixes of SF and PPF, this study aimed to contribute to the knowledge base on HPRC and its potential to improve the performance of reinforced concrete deep beams.The experimental results showed that the addition of SF and PPF could significantly enhance the load-bearing capacity of reinforced concrete deep beams. The presence of fibres also affected the ductility of the beams, as evidenced by the decrease in deflection before failure. The cracking patterns of the beams with fibres were also more cohesive, uniform and narrow compared to the beams without fibres.The numerical models calibrated with the CDP model in ABAQUS showed good agreement with the experimental results. The results showed that the optimal percentage of fibres depends on the desired performance. For instance, if the goal is to improve the load-bearing capacity and crack distribution, then a 0.75%SF–0.25%PPF combination is preferred. Conversely, if the aim is to improve the deflection, i.e., producing lower deflection values, then PPF higher content is preferable.In conclusion, the addition of steel fibres (SF) and polypropylene fibres (PPF) to concrete has varying effects on the crack pattern, load capacity, deflection, and damage of reinforced concrete beams. The control beam without fibres (M1B) has the lowest load capacity and highest deflection values compared to the other mixes. The addition of 1% SF (M2B) improved the tensile and flexural strength of the beam, reduced deflection, and increased load capacity. The addition of 0.25% PPF with 0.75% SF (M3B) improved toughness and ductility, reduced brittleness, and provided the best results in terms of load capacity. Using equal percentages of SF and PPF (M4B) balanced the properties of the concrete and provided improved flexural strength. Using a higher percentage of PPF with a lower percentage of SF (M5B) weakened the general behaviour of the beam. Adding 1% PPF (M6B) provided some enhancement to flexural strength if compared with the normal concrete but resulted in almost total damage to the longitudinal reinforcement due to the absence of steel fibres. Overall, the addition of SF and PPF can improve the properties of reinforced concrete beams, but the combination and percentage of fibres used should be carefully considered to achieve optimal results.Finding the optimized ratio of steel fibres (SF) to polypropylene fibres (PPF) for various performance criteria is a potential area for future study;Although SF and PPF were the primary focus of this study, other fibre types such as carbon fibres and glass fibres exist and can be considered;Long-term impacts of fibre addition on the behaviour of reinforced concrete beams were not the primary focus of the present study. The long-term effects of these fibres, such as their resistance to moisture, temperature changes, and chemical exposure, can be studied in future studies;Although deep beams were the focus of this study, columns, slabs, and walls are all potential targets for future research into the effects of SF and PPF.The following are some ideas for further research based on the results of this study:


Escherichia coli (E. coli) has been developed. In contrast to the traditional methods, the designed device has the rapid sensing ability to screen different antibiotics simultaneously, reducing the overall time of diagnosis. Screen-printed electrodes with integrated miniaturized reservoirs with a thermostat were developed. The designed device proffers simultaneous incubator-free culturing and detects antibiotic susceptibility within 6 h, seven times faster than the conventional method. Four antibiotics, namely amoxicillin–clavulanic acid, ciprofloxacin, ofloxacin, and cefpodoxime, were tested against E. coli. Tap water and synthetic urine samples were also tested for antibiotic susceptibility. The results show that the device could be used for antibiotic resistance susceptibility testing against E. coli with four antibiotics within six hours. The developed rapid, low-cost, user-friendly device will aid in antibiotic screening applications, enable the patient to receive the appropriate treatment, and help to lower the risk of anti-microbial resistance.Antimicrobial resistance (AMR) is a global health threat, progressively emerging as a significant public health issue. Therefore, an antibiotic susceptibility study is a powerful method for combating antimicrobial resistance. Antibiotic susceptibility study collectively helps in evaluating both genotypic and phenotypic resistance. However, current traditional antibiotic susceptibility study methods are time-consuming, laborious, and expensive. Hence, there is a pressing need to develop simple, rapid, miniature, and affordable devices to prevent antimicrobial resistance. Herein, a miniaturized, user-friendly device for the electrochemical antibiotic susceptibility study of  The accurate and early detection of microbial infection, followed by appropriate treatment via antibiotic administration, is pivotal in reducing the fatality and severity of the disease in a patient ,2. AntibEscherichia coli [E. coli is the most known bacterium that causes multidrug resistance [E. coli multidrug resistance [Various pathogens, like bacteria, fungi, viruses, and parasites, cause infection and form resistance . Among thia coli . E. colisistance . Impropesistance . Therefosistance ,13. AST sistance , which isistance . Often, sistance . Hence, sistance . In thissistance ,19. Micrsistance ,21. Howesistance . Hence, sistance ,24.E. coli was used as a model microbe for testing the device. A screen-printed electrode system modified with graphitized mesoporous carbon (GMC) was used for testing [E. coli [E. coli was validated using Streptococcus pneumoniae, Pseudomonas aeruginosa, and Shewanella putrefaciens bacteria. The real sample analysis was done using artificial urine and water samples. The obtained results were further validated with the conventional broth dilution method. To the extent feasible, this is a benchmarking prototype study that has yet to be explored further. The strategy with further optimizations can also be used for other microorganisms in real-time. The present study is an extension of our previous work . Herein, testing . GMC is [E. coli ,30. The E. coli culture was acquired from the Biological Science Department, BITS Pilani Hyderabad campus. Clavam 625 (amoxicillin and clavulanic acid), Zenflox 200 (ofloxacin), Monocef-O 200 (cefpodoxime), Cifran 500 (ciprofloxacin), and Azee-500 (azithromycin) were purchased from a local medical store. Polydimethylsiloxane (PDMS) was purchased from Delta Silicon, Mumbai, India. A CO2 laser (VLS 3.20) was procured from Universal Laser Systems, Scottsdale, AZ, USA.Luria broth and Luria agar were procured from Thermo Fisher Scientific, Delphi, India. Potassium chloride, carbon ink, glass slides (75 × 50 mm), ammonium phosphate, sodium sulfate, ammonium diphosphate, magnesium chloride, calcium chloride, creatinine, and urea were purchased from Sigma Aldrich, Ltd. . A three-electrode system was fabricated using the screen-printing technique. The design of the requisite dimension was first drawn on SolidWorks software. A polyvinyl chloride (PVC) sheet was attached to a glass side (75 × 50 mm), and the laser was scribed over the PVC sheet to prepare the mask. Carbon ink was laid down over the obtained mask with the help of a squeeze and kept in the oven for 30 min at 60 °C. The PVC sheet was removed after drying, and the screen-printed electrodes were obtained . Figure 3) was prepared on an acrylic sheet to develop the microfluidic device. To create a PDMS mixture, epoxy and curing agent were mixed in a 10:1 ratio and degassed for 30 min to remove oxygen bubbles. Following this, PDMS was run over the mold and baked for an hour at 60 °C. Post-curing, the reservoir was cut from the mold and bonded over the three-electrode system using the plasma bonding method. The developed microfluidic device integrated with the laser-induced graphene (LIG) heater is shown in A mold (2 × 1.4 × 17 mm2 laser (VLS 3.60) was exposed on the polyimide sheet with power and speed of 6.5% and 4.5% to obtain laser-induced graphene [For the fabrication of the LIG heater, a polyamide sheet of the required dimension of 25 × 25 mm was initially pasted onto a glass slide surface using double-sided tape. The COgraphene . After eElectrochemical analysis was used to determine the effect of antibiotics on bacterial growth. A Potentiostat (CHI 1030E) was utilized to record the electrochemical response. A three-electrode setup was employed, with a reference electrode of Ag/AgCl, a working electrode of GMC, and a counter electrode of plan carbon ink.The real samples used for analysis were synthetic urine and tap water . The synE. coli (DH5α strain) for MIC calculation. An agar plate containing Luria–Bertani (LB) broth was used to sustain E. coli cells. A colony of E. coli cells was removed from the agar plate and suspended in 5 mL of LB liquid media. Overnight at 37 °C, the cells were cultured in the medium on a shaker at 200 rpm. After that, fresh LB medium was used to dilute the cell suspension until it reached an optical density of 0.01 at 600 nm.The MIC describes the resistance or susceptibility of the particular bacteria toward valuable antibiotics. The model microorganism used was The antibiotic stock solution (1 mg/mL) of cefpodoxime, ofloxacin, amoxicillin, clavulanic acid, and ciprofloxacin was prepared in sterile water. Several concentrations, ranging from 100 to 500 g/mL, were prepared from the stock solution, and the MIC was measured using a disk diffusion approach. The LB agar media was prepared and poured into a Petri plate. After solidifying the agar gel, the bacterial culture was spread over the plate. Using sterile forceps, an antibiotic disk of cefpodoxime, ofloxacin, amoxicillin, clavulanic acid, and ciprofloxacin was applied to the plate and incubated for 12 to 24 h at 37 °C. The minimum inhibition zone formed at the edge of the antibiotic disk was calculated. E. coli bacterial inhibition. The bacterial concentration would decline in the device with increased time and a constant temperature. In To carry out the electrochemical investigation of bacterial growth inhibition, 100 µg/mL antibiotic concentration of cefpodoxime, amoxicillin and clavulanic acid, ciprofloxacin, and ofloxacin with bacterial culture media was injected into four miniaturized reservoirs through the inlet. Before the analysis, the reservoir was washed with 0.1 M PBS to prevent cross-contamination. The LIG heater was used to incubate the bacterial culture throughout the experiment, which is necessary for bacterial growth. The electrochemical detection was carried out using CV for 6 h, and after every hour, the response was recorded. E. coli was checked in the developed microfluidic device. Four variants, namely Streptococcus, Shewanella, Pseudomonas, and E. coli, were tested with Clavam antibiotic. The first E. coli with Clavam antibiotic was injected into the device, and CV response was recorded, Pseudomonas, Streptococcus, and Shewanella were then added into the same device, and the CV response was measured. A similar current histogram for E. coli and an additional variant mixed with E. coli is shown in E. coli and other bacterial species confirms that ciprofloxacin did not affect Shewanella putrefaciens, Pseudomonas aeruginosa, and Streptococcus pneumoniae [E. coli species or that the developed sensor is specific toward E. coli species.The specificity of the Clavam antibiotic toward eumoniae . The effA urine sample is mainly used for examining a medical condition. However, obtaining the same quality urine for many illness detections takes time and effort. Hence, synthetic urine is used for experimentation purposes . Along wE. coli culture was inoculated into the synthetic urine and tap water before being injected into the device. The electrochemical response was checked for 7 h at hourly intervals. The CV response of a water sample and synthetic urine is shown in E. coli and detected the antibiotic susceptibility within a concise 6-h timeframe. As part of our methodology, a three-electrode system was added to our miniaturized platform, with the working electrode modified with GMC, the reference electrode using Ag/AgCl, and the counter electrode composed of bare carbon ink. Electrochemical detection was performed using cyclic voltammetry, which enabled accurate measurements. The working electrode was modified with GMC to improve sensor performance and increase precision. Four antibiotics of the same concentration were tested in the susceptibility study, and the interference of the antibiotic toward E. coli was detected. Further antibiotic susceptibility study was performed in both artificial urine and water samples, ensuring the versatility and robustness of our device’s performance. The obtained outcomes were validated with the conventional approach, confirming the reliability of our findings. To address the pressing demand for rapid and accurate antibiotic susceptibility testing, integration with Cyber-Physical System (CPS) augmented techniques, like data mining and machine learning, with essential automation will aid in diagnosing antibiotic susceptibility rapidly. These technologies will advance the diagnostic process and enhance its accuracy. Moreover, as bacterial resistance to conventional treatments continues to rise, there is a critical need for technologies that accurately distinguish between resistant, susceptible, and persistent bacterial strains. These advancements pave the way for precision medicine, yielding optimal diagnostic results in the ever-evolving landscape of bacterial infections.This study developed a label-free, simple, cost-effective, miniaturized electrochemical microfluidic device to diagnose microbial resistance and bacterial infection rapidly. The designed device demonstrated remarkable sensitivity toward


Pfn1 gene selectively in EC (Pfn1EC-KO). Widespread deletion of EC Pfn1 in adult mice leads to severe health complications presenting overt pathologies  in major organ systems and evidence for inflammatory infiltrates, ultimately compromising the survival of animals within 3 weeks of gene ablation. Mice deficient in endothelial Pfn1 exhibit selective bias toward the proinflammatory myeloid-derived population of immune cells, a finding further supported by systemic elevation of proinflammatory cytokines. We further show that triggering Pfn1 depletion not only directly upregulates proinflammatory cytokine/chemokine gene expression in EC but also potentiates the paracrine effect of EC on proinflammatory gene expression in macrophages. Consistent with these findings, we provide further evidence for increased activation of Interferon Regulatory Factor 7 (IRF7) and STAT1 in EC when depleted of Pfn1. Collectively, these findings for the first time demonstrate a prominent immunological consequence of loss of endothelial Pfn1 and an indispensable role of endothelial Pfn1 in mammalian survival unlike tolerable phenotypes of Pfn1 loss in other differentiated cell types.Actin-binding protein Profilin1 is an important regulator of actin cytoskeletal dynamics in cells and critical for embryonic development in higher eukaryotes. The objective of the present study was to examine the consequence of loss-of-function of Pfn1 in vascular endothelial cells (ECs) in vivo. We utilized a mouse model engineered for tamoxifen-inducible biallelic inactivation of the  Actin cytoskeleton plays a key role in virtually all aspects of cell biology. In this study, we show that selective disruption of actin cytoskeleton regulatory gene Pfn1 in vascular endothelial cells leads to multiorgan pathology and death in mammals demonstrating an indispensable role of vascular endothelial Pfn1 for life. Loss of Pfn1 function in endothelial cells has a major proinflammatory consequence in vivo. We further propose that Pfn1 plays an important role in vascular endothelial and immune cell crosstalk.Profilins (Pfn) belong to a family of small actin monomer-binding proteins that are found in all eukaryotes. Besides actin, Pfn also interacts with a host of other cellular proteins harboring poly-L-proline (PLP) domains as well as poly-phosphoinositides (PPIs). Among the various isoforms of Pfn , Pfn1 is the only variant that is expressed in almost all mammalian cell types and much more abundantly than other variants . Pfn1 plSince the global absence of Pfn1 expression leads to early embryonic lethality during mammalian development , a numbeflox/flox:Cdh5-Cre-ERT2 mice that displayed defect in developmental retinal angiogenesis upon TMX-induced excision of the Pfn1 gene (flox/flox:Cdh5-Cre-ERT2 mice (referred to as Pfn1EC-KO from hereon) utilizing a 5-day time course injection of TMX. Littermate control Cre-negative mice (referred to as Pfn1WT from hereon) were also subjected to TMX administration following the same protocol. Genotyping performed on ear-clipped tissues confirmed the expected 700-bp amplicon upon deletion of Exon 1 of the Pfn1 gene in Cre+ mice  and hemorrhage in the thorax and abdominal cavities  showed a decline in Pfn1EC-KO animals  or the late-stage (18 days after last TMX injection) of Pfn1-depleted condition. Mid-stage immune profiling showed evidence for a dramatically (as high as ∼10-fold) elevated total leukocytes, various lymphocytes, and monocyte population in the peripheral blood (but not in the spleen) of Pfn11WT mice . End-staer cells , a findi animals . Howeverner Fig. . A greatWT and Pfn1EC-KO animals for a panel of 32 immunomodulatory cytokines and chemokines using a Luminex-based assay at either mid-stage or end-stage of Pfn1 depletion. Based on these analyses, mid-stage Pfn1EC-KO mice showed a statistically significant 2.45-fold increase in the circulating level of proinflammatory cytokine IL6 (P = 0.015) relative to Pfn1WT mice , circulating levels of several other factors including VEGF, G-CSF, IL1β, IL9, IL10, IP10/CXCL10, and MCP-1/CCL2 increased in statistically significant manners in end-stage Pfn1EC-KO relative to Pfn1WTanimals –immortalized EC isolated from Pfn1flox/flox FVB mice  did not reach statistical significance , monocytes/macrophages from Pfn1EC-KO mice indeed showed a trend of higher IL6 expression relative to those harvested from Pfn1WT mice  analyses to transcriptionally profile Ad-GFP- vs Ad-Cre-transduced EC in an acute setting ensuring that cell viability was not compromised. With the absolute fold change in expression and the false-discovery rate (FDR) set to >1.5 and <0.05, respectively, we found a total of 1,035 differentially expressed genes (DEGs—789 upregulated and 246 downregulated) in Pfn1 KO relative to control EC Fig. , with STFirst, the induction of loss of EC Pfn1 in adult mammals leads to severe multiorgan pathology ultimately compromising survival, demonstrating that endothelial Pfn1 function is indispensable for life. These results contrast the characteristics of all other previously reported conditional Pfn1 KO mouse models where complete loss of Pfn1 function in various other types of differentiated cells resulted in discernible phenotypes but was tolerable. These results should be informative for the scientific community when choosing an appropriate mouse model for studies that pertain to the investigation into the role of endothelial Pfn1 in cardiovascular diseases and other chronic pathologies. Second, loss of endothelial Pfn1 leads to an inflammatory response marked by pronounced elevation of a number of major proinflammatory cytokines causing enrichment of myeloid-derived immune cells with proinflammatory characteristics. These data demonstrate for the first time that perturbation of Pfn1 expression in EC has dramatic immunological consequences. Third, loss of Pfn1 not only stimulates endothelial–intrinsic transcription/release of proinflammatory factors (consistent with the signature of increased IRF7 and STAT1 activation) but also potentiates endothelial-directed proinflammatory gene induction in macrophages in a paracrine manner thereby amplifying the inflammatory milieu, as schematized in Fig. In this study, we report three novel findings. EC-KO mice at the organ level found in this study are disrupted vasculature, stunted growth, tissue infarct, fibrosis, and immune cell infiltration. Disrupted vasculature in organs (kidney and liver) in Pfn1EC-KO mice is consistent with endothelial Pfn1 dependency for angiogenesis previously shown by our group in early postnatal developmental setting  in Pfn1EC-KO mice. Therefore, VEGF upregulation (compounded by apoptotic cell death) could be another major driving force for inducing vascular leakiness in Pfn1EC-KO mice.The salient pathological features of Pfn1 setting  and, mor setting . In the  in mice , similarEna/VASP . Therefosruption . Anothersruption . We attrA key question that emerges from our study is whether the proinflammatory consequence of loss of endothelial Pfn1 is specific to some unique functional aspects of Pfn1 or a generic feature expected to result from the loss of any other key actin polymerization regulatory proteins. Disruption of endothelial barrier function contributing to local inflammation and cell death could certainly be a common phenotypic consequence of disruption of key regulators of actin polymerization. However, our cell culture–based studies showed that loss of endothelial Pfn1 leads to upregulation of proinflammatory factors not only in an EC-intrinsic manner but also in macrophages through a paracrine mechanism. Furthermore, our RNAseq studies revealed that many of the biological pathways that are predicted to be activated in EC upon Pfn1 depletion are related to immunological functions. Therefore, we reason that there are additional cellular consequences capable of impacting immune response that are unique to the loss of Pfn1.EC-KO mice. In the context of these data, it is interesting to note that in atherosclerosis setting, global heterozygous deletion of the Pfn1 gene reduces inflammation and macrophage activity conferring partial protection from the severity of atherosclerosis  that are elevated either in vitro or in vivo upon loss of endothelial Pfn1  for 24 h.Generation and culture conditions of hTERT-immortalized mouse EC bearing floxed-Pfn1 alleles have been described recently . For Pfnflox/flox mice, as described in our previous study  and mouse EC CM were probed for cytokine/chemokine expression levels by Luminex analysis by service provided by Eve Technologies  using the MD32  panel. For mouse serum analyses, analyte values were normalized to the average of wild-type (WT) animals within litters. For CM analyses, analyte values were normalized to the corresponding average WT CM values per analyte basis.Whole blood or spleen was passed through a 70-μm cell strainer, treated with 1× RBC lysis buffer to reduce red blood cell contamination, and incubated with Brilliant Stain Buffer  to prevent signal overlap between blue and ultraviolet markers. Samples were stained with either a full panel of immune cell–specific antibodies or buffer only for 30 min at 4°C, spun, and fixed in fixation/permeabilization buffer overnight, followed by next-day intracellular factor staining for 30 min at room temperature  Detection IHC kit , and slides were incubated overnight at 4°C with primary anti-CD31 antibody . This was followed by incubation with a biotin-labeled anti-rabbit secondary antibody for 10 min, washing with PBS-Tween, and incubation with streptavidin–peroxidase for 10 min. The staining was then detected with DAB substrate by incubation for 5–6 min. Slides were counter-stained with hematoxylin for 2 min before dehydration, mounting, and imaging.For H&E staining, the same initial steps were adopted before incubation of histosections with Mayer's hematoxylin (Lillie's modification) for 2 to 3 min. Slides were washed in deionized water, color adjusted with 5% glacial acetic acid, incubated with bluing reagent for 10–15 s, washed in DI, and stained with eosin for 2–3 min before dehydration and mounting. For apoptotic cell analysis, Click-IT TUNEL colorimetric kit  staining was performed according to the manufacturer-supplied protocol. Masson's trichrome staining was performed by the McGowan Institute of Regenerative Medicine Histology Core at the University of Pittsburgh.4 and 1% K3Fe(CN)6 for 1 h. Following three PBS washes, the tissue was dehydrated through a graded series of 30–100% ethanol and 100% propylene oxide and then infiltrated in 1:1 mixture of propylene oxide:Polybed 812 epoxy resin  for 1 h. After several changes of 100% resin over 24 h, tissue was embedded in molds and cured at 37°C overnight, followed by additional hardening at 65°C for 2 more days. Ultrathin (60 nm) cross-sections of the liver or kidney were collected on copper grids and stained with 1% uranyl acetate for 10 min, followed by 1% lead citrate for 7 min. Sections were imaged using a JEOL JEM 1400 Flash transmission electron microscope  at 80 kV with a bottom-mount AMT 2k digital camera .For TEM, tissues were harvested and immersion fixed in 2.5% glutaraldehyde and 2% paraformaldehyde in PBS overnight at 4°C. Following fixation, tissues were washed 3× in PBS and then postfixed in aqueous 1% OsOTotal cell lysate (TCL) was collected using a modified RIPA buffer , 1 mM EDTA, 50 mM NaF, 1 mM sodium pervanadate, along with 6× sample buffer with SDS diluted to 2% in final buffer. TCL was run on an SDS–PAGE and immunoblotted using antibodies specific for Pfn1 , IRF7 , p-IRF7 , STAT1 , p-STAT1 , and GAPDH .For assessing Pfn1 expression in ECs in vivo, single cell isolate was collected by collagenase-mediated kidney digestion and labeled with MACS mouse CD31 magnetic beads  prior to isolation using magnetic LS columns according to manufacturer's recommended protocol. Unlabeled effluent was collected as a negative control. To assess gene expression in circulating myeloid populations, whole blood was labeled with MACS mouse F4/80 beads  and isolated using LS columns. Cellular RNA was extracted using the Qiagen RNeasy Mini Kit  as per the manufacturer's protocol. Complementary DNA (cDNA) synthesis from the extracted RNA was then performed using the Qiagen QuantiTect Reverse Transcription kit  using the Applied Biosystems Veriti Thermal Cycler  following the manufacturer's instructions. Collected cDNA was amplified using SYBR Green Gene Expression Master Mix on the StepOnePlus real-time PCR system (Applied Biosystems) using primers for specific genes of interest. For product amplification, the plate was ramped to 50°C for 4 min, followed by 2 min at 95°C preamplification cycling. Cycling conditions of 95°C for 20 s, 49.7°C for 25 s, and 95°C for 50 s were used for 60 cycles  for quality control analyses, preparation of multiplexed paired-end libraries, polyA selection, and sequencing on an Illumina platform with 20 to 30 million reads per sample. FASTQ data processing and downstream analyses are described in detail in our recent study .U test was used for statistical comparison of data between the groups. For experiments involving a larger sample size and normal sample distribution, an unpaired t-test was used. A P-value less than 0.05 was considered to be statistically significant.All statistical tests were performed using GraphPad Prism 9 software. For experiments with a small sample size and nonnormal distribution of data, nonparametric Mann–Whitney pgad305_Supplementary_DataClick here for additional data file.


Basalt fibers are a type of reinforcing fiber that can be added to concrete to improve its strength, durability, resistance to cracking, and overall performance. The addition of basalt fibers with high tensile strength has a particularly favorable impact on the splitting tensile strength of concrete. The current study presents a data set of experimental results of splitting tests curated from the literature. Some of the best-performing ensemble learning techniques such as Extreme Gradient Boosting (XGBoost), Light Gradient Boosting Machine (LightGBM), Random Forest, and Categorical Boosting (CatBoost) have been applied to the prediction of the splitting tensile strength of concrete reinforced with basalt fibers. State-of-the-art performance metrics such as the root mean squared error, mean absolute error and the coefficient of determination have been used for measuring the accuracy of the prediction. The impact of each input feature on the model prediction has been visualized using the Shapley Additive Explanations (SHAP) algorithm and individual conditional expectation (ICE) plots. A coefficient of determination greater than 0.9 could be achieved by the XGBoost algorithm in the prediction of the splitting tensile strength. Concrete is a commonly utilized construction material globally, produced by blending cement with sand, gravel, and water. Additional chemical and mineral components can be incorporated into concrete as needed ,2. FurthOne of the new developments in the field of fiber-reinforced concrete is the usage of basalt fiber (BF), which gives excellent results in improving the compressive and flexural strength of concrete composites. Basalt fiber has high tensile strength and ductility, and its resistance to corrosion under alkali, acidic, and salt effects is better than fiber types such as steel, carbon, and aramid, making it a suitable fiber for bridge, shoreline, and road structures ,17. TablPrevious studies have demonstrated the effectiveness of basalt fiber reinforcement in a structural engineering context. Kabay  studied With the evolution of artificial intelligence (AI), machine learning (ML) algorithms are commonly used because ML can provide fast and economical solutions for civil engineering problems ,40,41,422 = 0.99) and a more reliable result than the LR and SVR algorithms. Li et al. [2 = 0.938) outperformed SVM and ANN, especially in FD prediction. Sensitivity analysis to determine the effect of inputs on MDS and FD response variables showed that fiber length and cell pressure have a significant effect on MDS predictions. Najjar et al. [Hasanzadeh et al.  presentei et al.  predictei et al.  used stai et al.  proposedi et al.  proposedi et al.  investigr et al.  developer et al.  used RanIn this study, ensemble learning techniques such as Extreme Gradient Boosting (XGBoost), Light Gradient Boosting Machine (LightGBM), Random Forest, and Categorical Boosting (CatBoost) have been applied to the prediction of the splitting tensile strength of concrete reinforced with basalt fibers. A dataset of 135 splitting tensile strength experiments has been curated from the literature. Based on this dataset, the splitting tensile strength has been predicted as a function of 12 different independent input features. Furthermore, the impact of each input feature on the model predictions has been quantified using SHAP analysis and individual conditional expectation (ICE) plots. The features have been ranked according to their importance, and the most impactful features have been determined. The introduction of basalt fibers into concrete is a relatively recent development in structural engineering, and there is a lack of reliable techniques for the prediction of basalt fiber-reinforced-concrete strength. The current study aims at developing novel data-driven methodologies for the prediction of the splitting tensile strength of concrete reinforced with basalt fibers. Due to the well-known weakness of concrete under tensile forces, and the increasing application of fiber-reinforced concrete, the availability of sound methodologies for the assessment of basalt fiber-reinforced concrete tensile strength bears great significance. This section presents the statistical properties of the dataset used in training the machine learning models in detail. The original dataset collected from the literature ,73,74,75The stars in the upper right of the correlation values in The range of each feature in the model is shown in The relationship between the fiber content (d Sawant  are givex.The ensemble learning approach combines the predictions of single learning models such as single decision trees to obtain strong learner models. Ensemble learning methods such as eXtreme Gradient Boosting (XGBoost), Categorical Boosting (CatBoost), Light Gradient Boosting Machine (LightGBM), and Random Forest have been successfully applied in the area of engineering systems response prediction. The output of ensemble learning algorithms can be summarized as in Equation (4) where The ensemble learning models aim to minimize the error between the predictions and the actual values which can be represented as in Equation (5), where L is the loss function, n is the total number of samples in the training set, inimized ,79,80.The predictions of the equations from the literature as well as the ensemble learning techniques are presented in the following section. The predictions are compared with the experimental measurements, and the accuracies of different modeling techniques are quantified using coefficient of determination, root mean squared error and mean absolute error. The calculations of these error metrics are given in The predictions of Equations (2) and (3) have been presented in The predictions of the ensemble learning models have been plotted against the actual measurements in The ensemble models are combinations of iteratively generated decision trees. The accuracies of the ensemble learning models are visualized in radar chart form in F is the set of all input features and S is a subset of F which does not contain the feature with index i. According to Equation (6), the impact of a feature is determined by the differences in the model outputs when a feature is included and withheld from the set of input features [The SHAP methodology effectively visualizes the impact of each input feature on the predictions of an ML model. This technique has its origin in cooperative game theory, where the Shapley values are used to measure the contribution of each player to a coalition. The Shapley values features ,82,83.(The SHAP summary plot in The individual conditional expectation plots in Basalt fibers are finding applications in structural engineering in recent years for reinforcing concrete. The splitting tensile strength is one of the decisive parameters that define the strength of concrete. However, the currently available equations for the prediction of this quantity are not sufficiently developed to include the effects of a large number of variables. The current study presents an effective way of predicting the splitting tensile strength of basalt fiber-reinforced concrete with reasonable accuracy based on data obtained from experiments. A total of 135 experimental results were used in training and testing four different ensemble learning algorithms. Among these algorithms, XGBoost delivered the highest accuracy after hyperparameter tuning with an A limitation of the current study is the size of the dataset used in training and testing the ML models. In order to develop predictive models that could be reliably generalized, large datasets are necessary. Moreover, it should be noted that the database used in the study consists of experimental results with small-scale specimens, and size–scale effects are built into the results. Future research in this area may focus on obtaining larger data sets. These larger datasets could also be used for developing reliable closed-form predictive equations for the splitting tensile strength. Furthermore, additional mechanical properties of basalt fiber-reinforced concrete, such as flexural strength, could be investigated.


Mud powders in aggregates are often found to cause deterioration of concrete properties. Based on a study of the mechanical properties of bridge deck leveling concrete (BDLC) containing different mud powders at various ages, the effects of mud powders on concrete durability were evaluated through rapid chloride permeability testing, freeze–thaw testing, and the coupling of salt solution and a freeze–thaw test. The properties of the interfacial transition zone (ITZ) were also investigated via microhardness testing. The test results showed that mud powder reduced the compressive strength, static compressive elastic modulus, and bond strength at early stages of curing. Moreover, mud powder was found to reduce the tensile properties and durability of concrete, with clay powder causing a greater reduction than mud powder in river sands and coarse aggregate. In addition, the width of the ITZ of concrete containing mud powder was found to increase by 23.1–48.3%. A significant correlation between the ITZ and the tensile properties, as well as the durability of concrete, was also observed. Therefore, in order to improve the tensile properties and durability of BDLC in seasonally frozen regions, the content of mud powder in the aggregates should be minimized according to the different compositions of mud powders. The coupling effect of salt solution and a freeze–thaw cycle should also be taken into consideration. In recent years, increasingly heavy axle loads and surging traffic volumes have led to the occurrence of cracks, pumping distress, interlayer bond failures, and other issues in local areas of bridge decks. This is mainly attributed to the early deterioration of the bridge deck pavement layer, allowing water ingress into the bridge structure ,2,3,4. SThe strength of concrete stands as the foremost factor in structural design; however, in numerous practical scenarios, durability assumes primacy ,9. WithiChina defines the content of fine particles in aggregates as the proportion of particles with a particle size less than 75 μm ,24. In pAs a portion of the fine mud powder resides on the surface of aggregates in the form of a surface cover layer, it obstructs the bonding between aggregates and the cementitious paste, potentially leading to a reduction in both the strength and durability of the concrete . StudiesIn this study, taking into account the operational requirements of BDLC in seasonally frozen regions, we conducted a comprehensive comparative analysis to assess the influence of various compositions of mud powder on the mechanical properties of BDLC. Our evaluation encompassed the relative permeability of different concrete mixtures, employing the electrical flux method. Additionally, we designed freeze–thaw tests, as well as the coupling of salt solution and freeze–thaw tests, to gauge the frost resistance of concrete. Parameters such as mass loss rate, relative dynamic elastic modulus, and relative durability index were considered. The characteristics of the ITZ were examined at a microscopic scale through the incorporation of microhardness testing. We also conducted a comprehensive assessment of how various types of mud powder influence the performance of BDLC. The primary objective of this research is to conduct a comparative and analytical study of the impacts of commonly encountered mud powder varieties on the mechanical properties and durability of concrete in practical engineering applications. The ultimate aim is to establish a foundational understanding for the construction of BDLC in regions characterized by seasonal freezing conditions.Based on the construction technical plan for the Jiamusi Transit Section PPP Project of the HeDa Expressway, specifically for the Songhua River Bridge, the primary cementitious materials employed were ordinary Portland cement with a strength grade of 52.5  and fly The river sands and coarse aggregates underwent a cleaning process using water. After this cleaning, the resulting mud liquid, which included materials from both the river sands and coarse aggregates, was passed through a square-hole sieve with a mesh size of 75 μm. This step ensured that particles with a size exceeding 75 μm within the aggregates were retained and not washed away. To facilitate a meaningful comparison of the effects of various mud powder compositions on concrete performance, the mud content of the aggregates utilized in the real-world project was adopted as the mud content benchmark for the test mixture. Furthermore, to maintain control over the variables, different mud powders were introduced into the cleaned aggregates, according to the specifications outlined in The concrete mixtures were prepared using a double horizontal shaft laboratory mixer. Once mixed, the specimens were compacted into molds and left to demold for 1 d at a 20 ± 5 °C and relative humidity of more than 50%. Following the demolding stage, the specimens were subjected to standard curing conditions, with humidity levels nominally at 95% and a temperature of approximately 23 °C, until they were ready for testing.In accordance with the guidelines outlined in JTG3420-2020 , a serieH represents the roughness Mass loss rate exceeding 5%(b)Relative dynamic modulus of elasticity declining to less than 60%The freeze–thaw test was conducted using the quick-freeze method in accordance with the guidelines outlined in JTG3420-2020 . SpecimeThe specimens were sectioned into square slices, each with a thickness of 7 ± 2 mm, utilizing a cutting machine, as illustrated in Vickers hardness testing was employed to assess the properties of the ITZ; images from the eyepiece of the microhardness tester are shown in To enhance the precision of measurements, the diagonal length of the indents was determined using a field emission scanning electron microscope  Co., Ltd., Shanghai, China). Subsequently, Vickers hardness values in proximity to the ITZ were computed using Equation (2). The indentations made in the paste, aggregate, and ITZ can be observed in HV is the Vickers hardness (MPa); P is the applied load (N); D is the average diagonal length of the indents (mm).In which, The test results for compressive strength at various ages are depicted in 2 adjusted) values for C-0 and C-K exceed 0.99, which could be attributed to the uniformity of the mixtures.The relationship between the square root of compressive strength and static compressive elastic modulus is illustrated in The splitting tensile strength of C-0 consistently surpasses that of C-SC, C-C, C-S, and C-K from 7 days to 28 days, as depicted in In summary, C-0 exhibits superior tensile properties, including splitting tensile strength and flexural-tensile strength, when compared to C-SC, C-C, C-S, and C-K. This difference can be attributed to the presence of mud powder in C-SC, C-C, C-S, and C-K, which enhances the overall density of the concrete. However, in these mixtures, the absence of pozzolanic reactions results in poor bonding around the mud powder particles, ultimately leading to reduced tensile properties in the concrete. Furthermore, when compared to the mud powder found in the river sands and coarse aggregate in the actual project, the clay powder has an additional detrimental impact, further diminishing the flexural-tensile strength of the concrete.The relationship between splitting tensile strength and flexural-tensile strength is illustrated in The splitting tensile bond strength at 7 days, 14 days, and 28 days is depicted in In actual construction projects, the compressive strength of BDLC typically meets its performance requirements and has minimal impact on its service life. However, when considering the structure and load-bearing characteristics of BDLC, it becomes evident that the tensile properties hold significantly more importance. While mud powder can enhance the compressive strength of concrete in later stages, it tends to diminish the tensile properties of BDLC, consequently potentially reducing its overall service life.Electrical flux serves as a valuable comparative index for assessing the chloride ion penetration resistance of concretes with different mud powders. The results of the rapid chloride permeability test conducted at 7 days and 28 days are presented in The results of the freeze–thaw cycle in water are presented in The relative dynamic elastic modulus of the specimens exhibits a gradual decline as the freeze–thaw cycles progress. The relative dynamic elastic modulus of C-K significantly lags behind that of C-0, C-SC, C-C, and C-S after 325 freeze–thaw cycles, as illustrated in The mass loss rate, relative dynamic elastic modulus, and relative durability index at the conclusion of the test are compiled in The test results for frost resistance indexes under the combined effect of freeze–thaw and salt solution are presented in Regarding the relative dynamic elastic modulus, there is minimal distinction between the specimens during the initial 100 cycles, as illustrated in Furthermore, N is the number of freeze–thaw cycles at which the relative dynamic elastic modulus of the specimen drops below 60% or the mass loss rate of the specimen reaches 5%; P is the average value of relative dynamic elastic modulus of three specimens after n freeze–thaw cycles.At present, there is no established standard test for evaluating the resistance of concrete to salt freeze–thaw. To provide a more logical assessment of the combined effect of freeze–thaw and salt solution on concrete, the formula for calculating the relative durability index is adjusted from Equation (3) to (4), as per the guidelines outlined in JTG3420-2020 . The resIn summary, concrete without mud powder exhibits superior frost resistance, while the inclusion of clay powder has a more detrimental effect on frost resistance compared to mud powder present in river sands and coarse aggregate. Moreover, the coupling effect of salt solution and freeze–thaw cycle significantly diminishes the maximum number of freeze–thaw cycles a concrete structure can endure and exacerbates the extent of freeze–thaw damage. To enhance the durability of the BDLC in regions with seasonal freezing conditions, it is advisable to minimize the mud powder content in the aggregates by considering various mud powder compositions and to give strong consideration to the effects of salt solution and freeze–thaw cycles.n, The microhardness data for ITZ at 7 days and 28 days are shown in At the 7-day mark, C-0 exhibits the thinnest ITZ thickness, with C-SC, C-C, C-S, and C-K showing, respectively, ITZ thicknesses that are 27.7%, 24.3%, 28.1%, and 36.2% greater than C-0. Between 7 and 28 days, the ITZ thickness in C-0 and the other concrete types decreases by 31.4%, 33.0%, 32.1%, 31.4%, and 25.3%, respectively. Even after this reduction, the ITZ thicknesses in C-SC, C-C, C-S, and C-K remain notably higher, with differences of 24.7%, 23.1%, 28.2%, and 48.3%, respectively, in comparison to C-0. This observation underscores the fact that the presence of mud powder reduces the ITZ properties, with clay powder exhibiting the most significant impact in diminishing ITZ thickness.When considering the results presented in Simultaneously, when we take into account the cumulative impacts of various mud powders on concrete’s chloride ion penetration resistance and frost resistance, as detailed in (1)In the initial stages of curing, concrete without mud powder in aggregates demonstrates superior performance in terms of compressive strength, static compressive elastic modulus, and splitting tensile bond strength. However, the compressive strength and static compressive elastic modulus of the concrete with mud powder surpass those of the concrete without mud powder at the 14 days and 28 days.(2)During the curing period spanning from 7 days to 28 days, the inclusion of mud powder in the aggregate resulted in a reduction of both the splitting tensile strength and flexural-tensile strength of the concrete. Interestingly, there was a notable positive correlation between the splitting tensile strength and bending tensile strength of the concrete.(3)Concrete without mud powder demonstrates better resistance to chloride ion penetration, whereas the chloride ion penetration resistance of concrete containing clay powder is lower compared to that of concrete containing mud powder in river sands and coarse aggregate.(4)The presence of mud powder in aggregates diminishes the early-stage frost resistance of concrete, with clay powder leading to a more substantial reduction in frost resistance compared to mud powder derived from river sands and coarse aggregates. Furthermore, the coupling of salt solution and freeze–thaw cycles significantly intensifies the extent of freeze–thaw damage in comparison to freeze–thaw cycles in water only.(5)The compressive strength of bridge deck leveling concrete (BDLC) typically fulfills the structural demands for bridge construction. However, the tensile characteristics and durability predominantly influence the service life of BDLC. While mud powder does contribute to the increased compressive strength of concrete in the later stages, its presence in the aggregates results in reduced tensile properties and diminished durability of BDLC. As a result, it is crucial to minimize the mud powder content in the aggregates, considering the various compositions of mud powder and the specific operational conditions of BDLC in regions subject to seasonal freezing.(6)No significant correlation was observed between the ITZ and compressive strength or static compressive elastic modulus. However, a clear relationship was established between the properties of the ITZ and tensile characteristics and durability. Further research is required to fully understand the nature and characteristics of the ITZ.The study focused on typical mud powders found in the aggregates employed in a real-world construction project. It conducted a comprehensive comparative analysis of the influence of varying mud powder compositions on concrete’s mechanical attributes and durability. Moreover, the research utilized microhardness measurements to assess the characteristics of the interfacial transition zone (ITZ). The primary findings of this investigation can be summarized as follows:


Here, we report the discovery of nitrofurantoin heteroresistance in E. coli clinical isolates and a novel genetic mechanism associated with this phenomenon. Subpopulations with lower nitrofurantoin susceptibility than major populations  in two E. coli blood isolates (previously whole-genome sequenced) were identified using population analysis profiling. Each isolate was known to have a loss-of-function mutation in nfsA. From each isolate, four nitrofurantoin-resistant isolates were derived at a nitrofurantoin concentration of 32 mg l−1, and a comparator isolate was obtained without any nitrofurantoin exposure. Genomes of derived isolates were sequenced on Illumina and Nanopore MinION systems. Genetic variation between isolates was determined based on genome assemblies and read mapping. Nitrofurantoin minimum inhibitory concentrations (MICs) of both blood isolates were 64 mg l−1, with MICs of major nitrofurantoin-susceptible populations varying from 4 to 8 mg l−1. Two to 99 c.f.u. per million demonstrated growth at the nitrofurantoin concentration of 32 mg l−1, which is distinct from that of a homogeneously susceptible or resistant isolate. Derived nitrofurantoin-resistant isolates had 11–66 kb deletions in chromosomal regions harbouring nfsB, and all deletions were immediately adjacent to IS1-family insertion sequences. Our findings demonstrate that the IS1-associated large-scale genetic deletion is a hitherto unrecognized mechanism of nitrofurantoin heteroresistance and could compromise UTI management. Further, frequencies of resistant subpopulations from nitrofurantoin-heteroresistant isolates may challenge conventional nitrofurantoin susceptibility testing in clinical settings.Nitrofurantoin is a broad-spectrum first-line antimicrobial used for managing uncomplicated urinary tract infection (UTI). Loss-of-function mutations in chromosomal genes  Whole-genome sequencing reads and genome assemblies generated in this study have been deposited under BioProject PRJEB58678 in the European Nucleotide Archive (ENA). Accession numbers are listed in Table S1, available in the online version of this article. Previously generated Illumina whole-genome sequencing reads of progenitor isolates EC0026B and EC0880B are available under ENA accessions ERR3142418 and ERR3142524, respectively.Escherichia coli. Our work discovered nitrofurantoin heteroresistance in distinct E. coli clinical isolates and attributed this phenotype to IS1-associated deletion of chromosomal regions harbouring the oxygen-insensitive nitroreductase gene nfsB in a genetic background where nfsA was inactivated. Our findings demonstrate a novel genetic mechanism of nitrofurantoin heteroresistance and suggest surveillance for this phenotype in E. coli urinary isolates for improving UTI management.Nitrofurantoin is widely used for treating and preventing urinary tract infection (UTI). The prevalence of nitrofurantoin resistance is generally low in Escherichia coli has been associated with inactivating mutations in chromosomal genes nfsA, nfsB and ribE [E. coli clinical isolates (<7 % in Europe) [Nitrofurantoin is a widely used first-line antimicrobial for treatment and prophylaxis of urinary tract infection (UTI) . Reducedand ribE , which eand ribE . Despite Europe) , concern Europe) .E. coli clinical isolates may show heteroresistance against polymyxins and carbapenems [E. coli or other species had not been reported until our recent observation of potential examples in two E. coli blood isolates [Studies have reported that bapenems , a phenobapenems . Nitrofuisolates . We therE. coli isolates, EC0026B (SAMEA104039660) and EC0880B (SAMEA104040147), of Achtman multi-locus sequence types ST484 and ST58, respectively, were previously collected from independent bloodstream infections [nfsA gene [Two fections  and were26B SAMEA4039660 afections  were incfections  and are 26B SAMEA4039660 aN,N-dimethylformamide  to produce a stock solution of 50 mg ml-1 nitrofurantoin, which was then infused into cation-adjusted Mueller-Hinton  agar to form a series of two-fold increments in nitrofurantoin concentrations (4–256 mg l−1). One colony of each isolate was inoculated into nitrofurantoin-free MH2 broth and grown aerobically overnight at 37 °C with shaking (180 r.p.m.). Seven serial 10-fold dilutions of each broth culture were created with PBS . Aliquots (10 µl) of the original broth culture and serial dilutions were spread in octants of MH2 agar plates following the nitrofurantoin-concentration gradient and incubated aerobically overnight at 35 °C. For each agar plate, colony-forming units were counted in the octant where colonies were well separated, and thereby the nitrofurantoin minimum inhibitory concentration (MIC) of each isolate was determined. A maximum non-inhibitory nitrofurantoin concentration of each isolate was defined as the greatest concentration at which ≥70 % cells grew when compared to the growth without nitrofurantoin.Nitrofurantoin-resistant subpopulations of each isolate  were sought in three independent PAP experiments to confirm reproducibility . Specifie.g. 32 mg l−1, if the isolate’s MIC was 64 mg l−1) were randomly chosen as resistant isolates . To extract DNA and compare isolates with or without an extended time of nitrofurantoin exposure, these resistant isolates were divided into two groups (two isolates per group) for inoculating MH2 broth with (0.5× MIC) or without nitrofurantoin and aerobically incubated overnight at 35 °C (e.g. EC0026BC) and was grown in nitrofurantoin-free MH2 broth before DNA extraction , and purified using a GeneJET Genomic DNA Purification Kit (ThermoFisher Scientific). The mean concentration of DNA in each extract was estimated from three reads obtained with a Qubit dsDNA BR Assay Kit (ThermoFisher Scientific).Extracted DNA of each isolate was aliquoted for WGS. Short-read sequencing  was conducted on an Illumina HiSeq 2500 system (Illumina), and long-read sequencing was conducted on a MinION R9.4.1 flow cell (Oxford Nanopore Technologies) for isolates having adequate yields of DNA. For MinION sequencing, DNA libraries were prepared using a Rapid Barcoding Kit SQK-RBK004 (Oxford Nanopore Technologies), and base-calling, demultiplexing and barcode trimming were conducted with Guppy v5.0.16 (community.nanoporetech.com/downloads) and its built-in high-accuracy model. For quality control, Illumina reads were trimmed for a minimal base quality of Phred Q20 (10 bp sliding window) and filtered for a minimal length of 50 bp using Trimmomatic ; MinION de novo using Raven v1.7.0 [dnaA and rep genes, respectively. Scripts used for aforementioned steps are available at github.com/wanyuac/Assembly_toolkit. Genome annotation was conducted with Prokka v1.14.6 [For isolates having MinION and Illumina reads, MinION reads were assembled n v1.7.0  and polin v1.7.0 . All assn v1.7.0 . Complet v1.14.6 . Inserti v1.14.6  for confFor isolates derived from the same progenitor, the complete chromosome sequence of the comparator isolate was used as a reference to identify chromosomal genetic variation in resistant isolates. Since chromosomes of isolates having MinION reads were fully assembled, isolate-specific alignments of these chromosomes against references were performed using Minimap2 v2.24 . ChromosFor each isolate, mutations were filtered to exclude those in repetitive chromosomal regions   for accuE. coli blood isolates EC0026B and EC0880B. Each heteroresistant isolate had a nitrofurantoin MIC of 64 mg l−1, which was 8–16 times its maximum non-inhibitory concentration  was four times its maximum non-inhibitory concentration, and positive control IN09 (MIC=128 mg l−1) showed a drastic transition from full growth to complete inhibition when the nitrofurantoin concentration doubled from 64 mg l−1.PAP assays confirmed nitrofurantoin heteroresistance in both R1, since its DNA yield was not adequate for MinION sequencing (Table S2).Four resistant isolates and one comparator isolate were derived from each of isolates EC0026B and EC0880B , providiR1–R4 showed different deletions of 11–20 kb chromosomal regions harbouring nfsB  of another IS1A, which interrupted the gene vgrG. These two IS1A elements and the relevant reference sequence in the ISFinder database mutually differed by two nucleotide substitutions in the second open reading frame (insB) of the IS1A transposase gene [Resistant isolates EC0026Bing nfsB , and eacce IS150 . The othase gene .1A were found in chromosomes of EC0026BC, EC0026BR1 and EC0026BR4, and 23 copies were found in EC0026BR2 and EC0026BR3, which is consistent with deletions illustrated in 1A elements demonstrated 99–100% nucleotide identities and a 100 % coverage to the reference IS1A sequence in the ISFinder database. The explicit copy number of IS1A could not be determined in the Unicycler short-read-only assembly of EC0026B. Nevertheless, in the assembly graph, the mean and median depths of assembled segments from which the complete IS1A was recovered suggest 20–22 copies of this element with >98 % nucleotide identities to the reference sequence.Twenty-four copies of ISR1, whereas no point mutation was found in EC0026BR2, EC0026BR3, or EC0026BR4  of chromosomal regions harbouring nfsB  and a 142 bp truncation of the 5′ end of wzc in the conserved capsule locus wzi-wza-wzb-wzc [Unexpectedly, an additional chromosomal 4289 bp deletion was identified in EC0880B80BR1–R4  and imme of nfsB . This de-wzb-wzc .1X2 were found in chromosomes of EC0880BC, EC0880BR2, EC0880BR3 and EC0880BR4, with 96–98% nucleotide identities and a 100 % coverage to the reference IS1X2 sequence. The precise copy number of IS1X2 could not be determined in the Unicycler short-read-only assembly of EC0880BR1, although 100 % of this insertion sequence was recovered from five connected nodes (with 96–100% nucleotide identities to the reference sequence) in the assembly graph of this genome. These five nodes had a median read depth of 11-fold, suggesting 11 copies of IS1X2, which is consistent with the identified deletion in this isolate’s chromosome  that could grow at 32 mg l−1 (0.5× MIC) nitrofurantoin. The resistant subpopulation in each nitrofurantoin-heteroresistant isolate was more frequent than a minimum of 10−7 proposed to define heteroresistance [nfsA and ribE were identical between each progenitor isolate (EC0026B or EC0880B) and its derived resistant and comparator isolates, and neither progenitor isolate produces the multidrug-efflux pump OqxAB [1-associated deletion of nfsB regions. This kind of deletion probably occurred in a UTI-associated E. coli ST131 strain, whose nitrofurantoin MIC increased from 8 to 128 mg l−1 upon intermittent in vivo exposures to a therapeutic dose of nitrofurantoin in six months [We confirmed nitrofurantoin heteroresistance in sistance . Since nx months .1 element, IS1X2, was associated with the truncation of capsule locus wzi-wza-wzb-wzc in EC0880BC. All deletion events observed in our study are consistent with the abortive transposition model of IS1-mediated deletion, where a DNA duplex nicks at an end of IS1 and ligates to a target site on the same molecule, causing a loss of DNA between these two breakpoints [1X2 had interrupted nfsA in EC0880B, and we have previously elucidated that IS1R interrupted nfsA in two nitrofurantoin-resistant isolates IN01 and IN02 [1-associated deletion and interruption of nfsA or nfsB reduces nitrofurantoin susceptibility of E. coli and such genetic variation may occur at other loci, potentially altering phenotypes. As such, we emphasise the importance of monitoring the prevalence and genomic locations of IS1 using long-read sequencing.We also show that an ISakpoints . Moreoveand IN02 . Taken tE. coli strain K-12 MG1655 was reported previously, with a small subpopulation growing at a maximum nitrofurantoin concentration of 8 mg l−1 while the majority of cells had an MIC of 4 mg l−1 [E. coli strain ATCC 25922, which showed a maximum non-inhibitory nitrofurantoin concentration of 4 mg l−1 and a subpopulation growing at 8 mg l−1 with an average frequency of 9 % (nfsA may have been prerequisites for the level of resistance in both heteroresistant isolates (nitrofurantoin MIC=64 mg l−1), since loss of nfsA or nfsB function alone only has limited (MIC ≤32 mg l−1) or no impact on the nitrofurantoin susceptibility of E. coli [Nitrofurantoin heteroresistance in 4 mg l−1 . Our PAPy of 9 % . Nonethe. coli .−1) determined by the European Committee on Antimicrobial Susceptibility Testing [nfsB in these isolates, which already had inactivating mutations in nfsA, would cause an irreversible reduction in their nitrofurantoin susceptibility, paving the way to the emergence of nitrofurantoin-resistant cells if there were to be subsequent resistance mutations, such as those in ribE and the marA-marB intergenic region [nfsB reduces the reproduction rate of E. coli [ΔnfsB mutants may only occur under selective pressure of nitrofurantoin.Since nitrofurantoin MICs of both EC0026B and EC0880B were at the clinical breakpoint for nitrofurantoin resistance  was 10–100 times the reported frequencies of spontaneous resistance point mutations in E. coli [1 is present [1 provides E. coli with adaptive fitness. Specifically, in the absence of nitrofurantoin, the susceptible major population in an IS1-related nitrofurantoin-heteroresistant isolate can transmit and establish colonisation or infection as homogeneously nitrofurantoin-susceptible E. coli does and probably outcompete nitrofurantoin-resistant E. coli that has lost functions of nfsA and/or nfsB; and in the presence of nitrofurantoin, the infection/colonisation may persist with the resistant subpopulation. Third, nitrofurantoin-heteroresistant isolates may not be detected by routine nitrofurantoin-susceptibility tests in clinical settings if the resistant subpopulation has a frequency below the test’s detection limit.We consider IS. coli . An earl present . Second,E. coli, our study took advantage of hybrid de novo assemblies in accurately resolving complete bacterial genomes and identifying structural variation. Further work is needed to elucidate how IS1 is associated with genetic deletions and to compare fitness between resistant mutants and heteroresistant progenitor isolates. Moreover, since this work focused on two E. coli blood isolates in which gene-deletion events were associated with different IS1 variants, the prevalence of IS1-associated nitrofurantoin heteroresistance and genomic rearrangements in E. coli have yet to be fully assessed within a wider strain collection including urinary isolates. This future work would determine whether gene deletions require specific combinations of isolates and insertion sequences, as might be inferred from the isolate-specific nucleotide substitutions and indels in resistant isolates derived from EC0026B and EC0880B (1 in clinical E. coli isolates will help us to understand how this insertion sequence contributes to bacterial evolution.To determine the mechanism of nitrofurantoin heteroresistance in  EC0880B . FinallyE. coli can be stably heteroresistant to nitrofurantoin, with potential clinical significance, and such heteroresistance may be a precursor to nitrofurantoin resistance. IS1-family elements are associated with large-scale deletion of genomic regions containing nfsB and are potentially important agents of genomic and phenotypic changes in E. coli.Click here for additional data file.


Nitrofurantoin has been utilized for the prevention and treatment of urinary tract infections (UTIs) since the 1950s, and it has been prescribed with increasing frequency since being recommended as a first-line therapy. The adverse neurological and psychiatric effects of antibiotic medications have been well-established. There is evidence to suggest a direct association between acute psychosis and antibiotic exposures. Nitrofurantoin-induced adverse effects have been reported recurrently; however, to the best of our knowledge, a combination of auditory and visual hallucinations with normal baseline mentation and cognition in an immunocompetent geriatric patient, without previously reported hallucinations, have not been reported in the literature so far. We present a case of an 86-year-old Caucasian female who was admitted with audio and visual hallucinations on the fifth day of starting nitrofurantoin therapy for UTI. During her stay, after ruling out all other probable etiologies, it was determined that the likely pathogenesis of the patient’s neuropsychiatric effects was the use of nitrofurantoin. Urinary tract infections (UTIs), the most common bacterial infections worldwide, are primarily seen in women, with about 50% of women experiencing at least one incidence of UTI in their lifetime ,2. Its sNitrofurantoin has been available as an antibiotic since the 1950s, and it obtained FDA approval in 1989. Its predominant use is in both the treatment and prophylaxis of uncomplicated UTIs. Even though it has been prescribed and used for decades, its mechanism of action is not fully understood. It is thought to inhibit the bacterial enzymatic synthesis of nucleic acid and cellular structures -11.An 86-year-old Caucasian female with a pertinent past medical history of paroxysmal atrial fibrillation, insulin-dependent type II DM, diastolic congestive heart failure, recurrent UTIs, hypertension, hypothyroidism, and iron deficiency anemia was admitted with concerns of auditory and visual hallucinations that she had experienced on the fourth and fifth days of starting nitrofurantoin therapy. The patient reported seeing various people whom she did not know in her house interacting among themselves but not with her. The next day, she experienced a vision in which she saw a person with a knife coming toward her. She had also heard music playing in her house a few days prior to admission. Per the patient’s daughter, apart from her hallucinations, the patient had no abnormalities in her baseline cognitive function and exhibited appropriate mentation for age and person on admittance.Upon being admitted, nitrofurantoin was discontinued due to concerns about it causing encephalopathy. The patient’s vitals were within normal limits, and her only lab abnormalities included hyperglycemia (142 mg/dL) and mild anemia (10.4 g/dL). The patient's home medications included apixaban, duloxetine, lisinopril, levothyroxine, and a short course of nitrofurantoin. The patient had a bedside neurological exam, including cranial nerves, mental status, as well as alertness and orientation. The exam was unremarkable. The patient was alert and oriented to person, time, and place with no other signs of delirium sans the visual hallucinations. In general, the patient was well-kempt and in no general distress with a normocephalic and atraumatic head. The patient wore lipstick, had a full cognitive, pleasant conversation, and had positive hyperacuity of her environment. The remainder of her exam did not reveal any acute findings. During the examination, she reported having hallucinations in the room, and she described these as vivid and detailed. She described a door, visible only to her, next to the clothing chest, through which various people entered from the roof. She indicated having little interaction with the visitors. She described seeing children visiting as well. The accuracy of her descriptions was consistent throughout the duration of her hospital stay. The patient indicated that she understood that the hallucinations were not real and she was able to differentiate her hallucinations from reality. Her awareness of this persisted throughout her hospital stay without change or medical staff reassurance.An MRI would have been the best imaging modality for identifying structural brain abnormalities; however, it was contraindicated due to a noncompatible pacemaker. A head CT without contrast indicated advanced chronic small vessel ischemia and diffuse cortical atrophy; however, no acute findings or exacerbations that would explain the patient's symptoms were found Figure . The smaNitrofurantoin is a commonly prescribed antibiotic for treating UTIs. Though the mechanism of nitrofurantoin is incompletely understood, it is commonly accepted that the medication works through the action of inhibiting various enzymes involved in the synthesis of DNA, RNA, and proteins . It is wThis report described a unique case of nitrofurantoin-induced audiovisual hallucinations. Our case illustrates that while it remains a diagnosis of exclusion, older patients with comorbidities such as diabetes and frequent UTIs can present with the symptoms described here as an adverse effect of antibiotic use. Various studies have associated hallucinations with antibiotic use. With the increasing use of nitrofurantoin for uncomplicated UTIs and RUTIs, it is pertinent to assess its potential complications in various patient populations.