Carbon concrete is a new, promising class of materials in the construction industry. This corrosion-resistant reinforcement material leads to a reduction in the concrete cover required for medial shielding. This enables lean construction and the conservation of concrete and energy-intensive cement manufacturing. Bar-type reinforcement is essential for heavily loaded structures. The newly developed helix pultrusion is the first process capable of producing carbon fiber-reinforced polymer (CFRP) reinforcement bars with a topological surface in a single pultrusion process step, with fiber orientation tailored to the specific loads. The manufacturing feasibility and load-bearing capacity were thoroughly tested and compared with other design and process variants. Approaches to increase stiffness and strength while maintaining good concrete anchorage have been presented and fabricated. Tensile testing of the helical rebar variants with a 7.2 mm lead-bearing cross-section was conducted using adapted wedge grips with a 300 mm restraint length. The new helix geometry variants achieved, on average, 40% higher strengths and almost reached the values of the base material. Concrete pull-out tests were carried out to evaluate the bond properties. The helix contour design caused the bar to twist out of the concrete test specimen. Utilizing the Rilem beam test setup, the helical contour bars could also be tested. Compared with the original helix variant, the pull-out forces could be increased from 8.5 kN to up to 22.4 kN, i.e., by a factor of 2.5. It was thus possible to derive a preferred solution that is optimally suited for use in carbon concrete. Its excellent mechanical and physical behavior such as high tensile strength, stiffness, and chemical resistance, make it a good alternative as a reinforcing material for concrete structures [The increasing significance of sustainable materials and technologies in construction, driven by climate change concerns, is evident in the European Union’s goal to achieve a climate-neutral and sustainable European Economic Area by 2050. The target is to reduce COmissions ,7,8,9. Fmissions ,11,12,13missions . Conventructures . Compareructures ,17,18. Dructures ,22.In order to support concrete structures under tensile stress, the installed reinforcement structures must have undercuts in the axial direction. This functionalization helps to transfer the tensile stresses acting on the concrete structures to the inner cross-sectional areas of the reinforcement structures . In prevSelected functionalization variants were exemplarily implemented in production for a large-scale use of novel carbon reinforcement bars. These were subsequently validated experimentally, further developed geometrically, and the technological and economic implementation was investigated ,23. The The analysis of prototypical functionalized rod profiles , in combIn terms of these properties, the variants FormPres, ContMill, and HelixPul showed particular advantages among the investigated options. The ThermPin variant allows direct force introduction into the cross-section of the bar through additional elements as a special case. However, these elements significantly reduce the load-bearing cross-section and thus the load capacity. The connection in the WindForm variant is unfavorable for optimal bonding effectiveness due to the short rib width and a fiber orientation almost perpendicular to the loading direction in the functionalization. In comparison, the ContMill variant with sufficient rib width and axial fiber orientation exhibited good load-bearing behavior even in combination with concrete. The evaluation of tensile tests, damage patterns, and potential economic feasibility of each production technology formed the basis for selecting the optimization variant for load-appropriate rod functionalizations with a thermoplastic matrix for rod-reinforcement structures. The unique design possibility of the helix contour (HelixPul), as the first CFRP rebar design to be based on an economic single-step pultrusion process, showed high potential for further development and effective use in construction and was thus selected as the preferred option for further investigation ,23,24.The application of pultruded fiber-reinforced polymers (FRPs) in the construction sector has a well-established history. Vedernikov et al.  reviewedZhu et al.  experimeYun et al.  characteBenmokrane et al. , inter aTo further investigate the influence of the rebars’ surface geometry on their mechanical properties and the bond strength between the rebars and the concrete matrix, selected parameters of helix-pultruded carbon-reinforced rebars, such as the pitch of the helix contour and their number, have been varied. The structures were then investigated by performing tensile testing and extraction tests.Tape-shaped workpieces (SGL-CF-PA6-Tape) from the manufacturer SGL TECHNOLOGIES GmbH, Meitingen, Germany were used for the production of the helically pultruded CFRP-reinforced rebars. These consist of a thermoplastic polyamide 6 matrix, reinforced with SIGRAFIL R C T50-4.0/240-T140 carbon fibers from the same manufacturer, and have a fiber volume content of 45%. They were characterized by tensile tests, and the determined values were compared with the partially available characteristic values from the manufacturer’s datasheets see . It was n;Number of helix ribs P;Pitch pD;Circumferential diameter eD;Equivalent diameter fA;Projected flank area f/AiA.In order to further develop functionalized HelixPul rod structures, various geometric parameters were defined, which were analyzed and evaluated in terms of production and experimental considerations through parameter investigations. The following parameters were varied for the investigations:iA corresponds to the area of the inner circle with diameter i,D and the projected flank area fA is obtained from:The area iD, eD, P,D and the pitch P are graphically illustrated for the HelixPul variant H81 in The corresponding diameters The necessary geometry of the rod functionalization for the composite effect between the reinforcing structure and the concrete matrix requires a strong undulation, especially of the outer fiber layers, which extends into the center of the cross-section of the reinforcing bars and decreases in intensity from the outside to the inside. This relationship is schematically illustrated in The focus of the further development of the reinforcement bars was therefore on creating a balanced ratio between the outer bar profiling and inner straight layers, in order to achieve a compromise between good bond behavior with the concrete matrix and high stiffness. Carbon reinforcement bars were therefore investigated and further developed not only with respect to their tensile properties but also their bond behavior. Due to manufacturing constraints, the realization of the geometry parameters can only be indirectly achieved through external shaping. Therefore, the control of the inner fiber layers is only possible indirectly through the outer contour design and the pultrusion process.iD of 7.3 mm was defined. Due to the rotational manufacturing technology of the helix-pultrusion process, a circular basic structure of the reinforcement bars is given, around which the functionalization winds spirally in the form of rib-like attachments. The variations of the geometry parameters are summarized in In order to ensure good comparability in terms of assessing the load-bearing capacity of the reinforcement structures, the supporting continuous and undisturbed cross-section of the design variant H81 was adopted as a reference value for all other rebar variants, and an inner diameter P was increased. Insights from preliminary tests showed that the rib height could be lower for a lower fiber undulation and that the undercuts were still sufficiently formed for force transmission in the carbon–concrete composite. In variant H83, the rib height was therefore halved compared to variant H81. This change simultaneously affects the circumferential diameter pD, the projected flank area fA, and the ratio of f/AiA, thus affecting three parameters at the same time. The variation in the number of ribs also leads to a change in rib height, as the ribs must be geometrically narrower. If the rib flank angle is kept constant, a lower rib height automatically results. Therefore, in variant H84, the number of ribs was increased to six, resulting in the most delicate rib structure of all variants. Conversely, when reducing the number of ribs from three to two, as in variant H85, there is also an adjustment possibility with respect to rib width. Compared to a narrow rib, a wider rib can significantly better support itself under load. Due to the greatest potential for improvement and influence, variants H83, H84, and H85 were selected and implemented for manufacturing, material, and composite investigations in addition to variant H81, to test the bar geometries of helix-pultruded carbon reinforcement bars.The variation in the number of ribs over a defined length was influenced by the pitch of the helix, which also affects the orientation of the ribs. The smaller the pitch, the larger the angle of inclination of the ribs. However, since increasing the number of ribs over a defined length reduces the load-bearing potential of the individual ribs in the composite, narrowing the ribs does not necessarily represent an improvement. Therefore, for the design variant H82, the pitch The material characterization using tensile testing was performed on the developed and manufactured HelixPul variants H81, H83, H84, and H85. The test was carried out according to the standards for conventional reinforcement systems: EN ISO 15630-1, DIN EN ISO 7500-1, and DIN EN ISO 6892-1 ,32. Due After conceptualizing and evaluating various reinforcement-bar clamping devices, a clamping mechanism based on a wedge clamping system was developed see . The claThe larger the prism opening angle, the larger the clamping diameter of the bar. According to DIN 2214 for the execution of test prisms, cutting angles of 90° and 108° are suggested. A cutting angle of 108° forms a good compromise for clamping the reference bars and covering different bar diameters and was therefore chosen for the construction of the clamping device.For optimal clamping and load introduction, as well as for the protection of the test specimen from excessively high local clamping forces, prismatic clamping elements are equipped with a compensating layer made of elastomer for testing. The elastomer used is a nitrile rubber from the manufacturer Misumi, designated NB270n-10-300-300. The intermediate layer strips used had the dimensions of 2 mm × 12 mm × 300 mm. Due to the high stress on the compensating material, it cannot be reused multiple times.All tests were carried out at a standard test climate of 23 °C ambient temperature and 50% relative humidity. The testing machine used was a Zwick Z250 with a testing speed of 2 mm/min. Furthermore, the tensile forces occurring during the test were measured and recorded using a 250 kN force transducer, and the resulting deformations were measured using a contact macro extensometer. The specimens used in the study had an overall length of 80 cm, consisting of a free length of 20 cm and an anchoring length of two times 30 cm. The anchored length of the specimens was further secured using screws to fasten the prismatic clamping elements. The screws were tightened progressively from the outside to the inside with stepped torque, beginning at 60 Nm and decreasing by 5 Nm for each subsequent screw to minimize stiffness fluctuations. Afterwards, the rebars were subjected to a load of 20 kN, and subsequently, the screws were retightened. The external screws were tightened up to 80 Nm, while the internal ones were tightened with 2 Nm less each. In case the specimens slipped at higher loads, the test was paused, and the screws were retightened following the specified scheme.maxF, fracture strain maxε, and fracture stress maxσ, which are obtained from:0S is the cross-sectional area of the bar in the direction of tension. Six specimens were tested for each bar configuration to obtain valid results.The testing setup used can be seen in In addition to the evaluation of the pure strength and durability parameters, the reinforcement elements should also exhibit sufficient bond strength to the surrounding clay matrix. This is determined in pull-out tests. A specimen shape was selected that allows measurement results in the direct test procedure. Based on the experience according to , it is bThe selected test specimen allows the adaptation of different bond lengths. In order to map a representative range of composite properties, the bond length had to capture the bar geometry peculiarities  at least one time. The HelixPul test specimens investigated have the same pitch and thus the same bond length. Testing of the bond properties was carried out on a self-compacting concrete with 5 mm crushed granite grit as the largest aggregate. Accompanying compressive strength and flexural tensile tests showed a compressive strength of approx. 100 MPa and a flexural tensile strength of approx. 5 MPa for this formulation. The present HelixPul bar geometry provides a significant input of torsional energy into the concrete body. Due to this high torsional force input, the RILEM RC5 test was applied as an established test method with more massive component measurements. The test is basically a four-point bending test with a fixed pivot point. The latter absorbs the resulting compressive forces, whereas the embedded bar has to absorb the tensile components. In this case, an equal tensile force is generated on both sides of the bar and a pull-out will occur on the side of the weaker composite. The basic setup of the test is shown in Since the carbon rods have a radially lower stiffness than comparable steel rods, a larger-volume Styrofoam insert replaced the recommended plastic sheathing to allow greater bending freedom. For specimen fabrication, the rebar elements were inserted into the Styrofoam elements and positioned in the formwork. In addition, the auxiliary joint was also positioned in the formwork at the intended location to realize a defined pivot point. The formwork was filled with concrete, and the finished specimen was demolded after concrete consolidation. The rebar ends overhanging from the concrete were fitted with an inductive displacement transducer, and the specimens were positioned as a unit in the test fixture.s, and the relative standard deviation ν are also given.The results of the tensile tests conducted on the tested HelixPul geometry variants H81, H83, H84, and H85 are summarized in After testing, the characterized reinforcing bars exhibited a valid failure pattern in the area of the free length. This rules out the possibility that clamping effects, such as transverse forces from clamping, were the cause of damage and thus had a reducing effect on the mechanical properties.All experiments also showed a failure behavior typical for axially reinforced plastics. This is characterized by axial splintering or delamination in combination with uneven failure in the direction of tension. In the manufacturing process, the fibers are pressed into shape slightly under tension, as the matrix material solidifies quickly. This freezes residual stresses, which become visible in the failure shown in Due to the different jacket geometries of the reinforcing bars, the contact macro extensometer occasionally slipped during testing, leading to distortions of the strain measurements, which are evident as jumps in the measurement curves .maxF is related to the respective applied bond length, and thus the characteristic value bond flux maxk (N/mm) is generated. The results of the pull-out test are shown in In the pull-out test, the bar variants of HelixPul H81, H83 and H84 were tested. The measured maximum pull-out force As already described in The RILEM test method described in maxF, deformation max∆l and deflection fmax from the beam tests are summarized in The individual results with the maximum force The test results obtained in The tensile tests conducted to characterize the manufactured tension rod variants H81, H83, H84, and H85 from HelixPul production confirmed the suitability of the restraint principle for profiled rod geometries. The test specimens could be clamped sufficiently firmly and without damage. However, settlement effects of the NBR material occurred in some cases, which could be compensated for by manual re-clamping. For efficient and partially automated testing, it is recommended to use a controllable clamping device, such as one that can be realized through hydraulic force application.To assess the composite tests, morphological analyses were also carried out to examine the failure mechanisms during the pull-out test and the microstructure of the interphase. These optical observations, in addition to the mechanical parameters, offer insights into the prevailing damage mechanisms and potential improvement options. The observation levels can be adjusted as required, ranging from simple visual inspection with the eye/loupe and extending to more advanced techniques such as light microscopy or scanning electron microscopy. During the pull-out tests, the optical findings revealed minimal adhesion of the concrete matrix to the rebar, and no detectable carbon fiber residues were observed in the pull-out channel. Consequently, the investigations focused on utilizing high-resolution photographs and light microscopic images. The pull-out test specimens of HelixPul H81 showed hardly any adhesion to the bar and a precise mapping of the bar surface within the concrete matrix. Thus, positive locking is the dominant force transmission mechanism compare a,b. In cIn summary, the structural morphological investigations reveal that the majority of rebar variants transfer forces from the concrete to the reinforcement element primarily through form closure, rather than material closure. This form closure is primarily attributed to the geometry features, including the helical rib heights of the bars, the strong interlaminar shear strength within the bar, and the high surface quality. However, in cases where the bars exhibit roughness, including micro-roughness, as observed in the manufactured variant HelixPul H83, mineral crystals can grow into the bar structure. This phenomenon leads to an additional increase in bonding forces, further enhancing the effectiveness of force transfer between the concrete and the reinforcing elements.Variation studies for the design of helix-pultruded bar contours as reinforcement elements in concrete were carried out for different geometry parameters. This determination was based on tensile tests and pull-out tests. The results highlight the significant potential for enhancing the bar´s bearing capacity and minimizing the manufacturing impact of the unique helix-pultrusion technique on the bar contours. This was determined by tensile tests and pull-out tests. Upon comparing the individual values obtained from these tests, it becomes evident that the HelixPul H85 variant exhibits the most favorable behavior compared to the other variants, marking it the recommended choice.These findings confirm the initial assumption that novel helix pultrusion, as an efficient manufacturing process for carbon-reinforced bars, can achieve load-adapted and material-appropriate performance. Furthermore, this high potential can be further augmented through additional optimization efforts, such as employing higher-fiber volume contents. Additionally, the use of a thermoplastic matrix system allows for the flexible reshaping of the rebars, enabling adaption to more intricate structural configurations.Reinforcing bar structures made of fiber composites enables thinner construction methods. In particular, carbon reinforcements are corrosion-resistant and reduce the required concrete cover.The newly developed helix-pultrusion process enables efficient manufacturing processes for fiber-composite reinforcement bar structures. It is characterized by few process steps and the effective use of materials. During the work, design parameters were varied, and corresponding variants were manufactured to achieve a more load-oriented material arrangement and utilization.The tensile behavior of the reinforcement bar prototypes was tested using a new, efficient tensioning system developed in-house. Compared to unfunctionalized reference bars, similar bar tensile strength was achieved with the contour adjustments, and the manufacturing influence of helix pultrusion was almost eliminated. The fracture strength of the HelixPul rebars was increased by more than 40% on average.By employing a more complex test arrangement, it was possible to determine torsional moment-free and transverse force-free pull-out characteristics, demonstrating good comparative pull-out behavior. The tensile force transmission in the composite for the HelixPul rebars was increased by more than a factor of 2.5.The determined preferred helix rebar contour, HelixPul H85, proves the suitability of these reinforcing bars from the novel production technology, due to element tension tests and concrete pull-out tests.The novel material—carbon–concrete composite—offers promising possibilities for applications in civil engineering. As part of the project preparation, the authors developed a novel production technology called helical pultrusion. This process enables the efficient and continuous production of surface-topologized rebars in a single process step, which are suitable for use as CFRP rebars in concrete applications. The presented geometric parameter adjustments and the produced rebars aim to enhance the load-bearing properties of the reinforcement elements and to overcome the current disadvantages of carbon–concrete-composite technology on the path towards series production:Through the geometry parameter variation of the HelixPul rebars, the potential and principle of helical pultrusion were successfully demonstrated. It allows the continuous production of reinforcing bars in a single-step forming process using carbon fibers with a thermoplastic matrix through pre-impregnated tapes. The transition to other fiber types and other thermoplastic matrix systems requires only minor adjustments. However, significant challenges in process technology and increasing productivity necessitate the integration of fiber plastic impregnation and in-situ polymerization into the process, along with increasing process speed. Implementing the manufacturing process with a thermoset matrix system requires major adaptations and novel semi-finished products. Therefore, further research is required to enhance the efficiency of the manufacturing process and utilize thermoset matrix systems for improved thermal resistance.


Background: Community-acquired pneumonia (CAP) is a common indication for antibiotic use in hospitalized children and is a key target for pediatric antimicrobial stewardship programs (ASPs). Building upon prior work, we developed and refined an electronic algorithm to identify children hospitalized with CAP and to evaluate the appropriateness of initial antibiotic choice and duration. Methods: We performed a cross-sectional study including children 6 months to 17 years hospitalized for CAP between January 1, 2019, and October 31, 2022, at a tertiary-care children’s hospital. CAP was defined electronically as an International Classification of Disease, Tenth Revision (ICD-10) code for pneumonia, a chest radiograph or chest computed tomography scan (CT) performed within 48 hours of admission, and systemic antibiotics administered within the first 48 hours of hospitalization and continued for at least 2 days. We applied the following exclusion criteria: patients transferred from another healthcare setting, those who died within 48 hours of hospitalization, children with complex chronic conditions, and those with intensive care unit stays >48 hours. Criteria for appropriate antibiotic choice and duration were defined based on established guidelines. Two physicians performed independent medical record reviews of 80 randomly selected patients (10% sample) to evaluate the performance of the electronic algorithm in (1) identifying patients treated for clinician-diagnosed CAP and (2) classifying antibiotic choice and duration as appropriate. A third physician resolved discrepancies. The electronic algorithm was compared to this medical record review, which served as the reference standard. Results: Of 80 children identified by the electronic algorithm, 79 (99%) were diagnosed with CAP based on medical record review. Antibiotic use was classified as the appropriate choice in 75 (94%) of 80 cases, and appropriate duration in 16 (20%) of 80 cases. The sensitivity of the electronic algorithm for identifying appropriate initial antibiotic choice was 94%; specificity could not be calculated because no events of inappropriate antibiotic choice were identified based on chart review. The sensitivity and specificity for determining appropriate duration were 88% and 97%, respectively (Table 1).Conclusions: The electronic algorithm accurately identified children hospitalized with CAP and demonstrated acceptable performance for identifying appropriate antibiotic choice and duration. Use of this electronic algorithm may improve the efficiency of stewardship activities and could facilitate alignment with updated accreditation standards. Future studies validating this algorithm at other centers are needed.Disclosures: None


Characterize antibiotic prescribing behaviors at an Indian palliative care center after the initiation of the Antibiotic Order Form (AOF): an antibiotic stewardship program involving a paper form to track antibiotic use and to provide prescription guidelines.Retrospective chart review.Trivandrum Institute of Palliative Sciences (TIPS) is a palliative care organization in Kerala, India.Antibiotic prescription data and patient data were collected for adult patients treated at TIPS between January 1, 2017, and October 31, 2019. Descriptive statistics and a Zero-Inflated Poisson regression model were used to analyze antibiotic prescriptions. AOF completion and prescription concordance with institutional guidelines were also evaluated.Out of 7,450 unique patients, 675 (9%) were prescribed 1,448 antibiotics. Age was the strongest factor in determining the number of antibiotic courses with each additional year of age decreasing the expected antibiotic prescription count by 2% per year. The most common antibiotics prescribed were topical metronidazole (44%) and penicillins (29%). Among patients who died, 5% were prescribed antibiotics within the final month of life. In total, 32% of antibiotic prescriptions were documented in AOFs, and 18% were concordant with all institutional antibiotic prescribing guidelines.This study is the first to analyze an antibiotic stewardship intervention in a palliative care setting within a low- and middle-income country. This retrospective study provides a benchmark of antibiotic use within Indian palliative care and highlights areas for future stewardship research including topical metronidazole use within palliative care and higher rates of antibiotic use among younger palliative care patients. Antibiotic use among patients receiving palliative and hospice care is a global challenge. When addressing infection among palliative and hospice patients, providers are faced with a dilemma. The challenges include concurrently managing symptoms from possible infection, minimizing the incidence of drug resistance, negotiating an emotionally charged atmosphere, and following the patient’s goals of care. These often conflicting priorities might result in empiric antibiotic use. Yet, there are no established guidelines describing how providers should navigate this clinical setting, and, consequently, antibiotic use varies widely across palliative and hospice care settings. Among patients in the final weeks of life, several studies suggest that at least one-quarter or more of patients receive at least one antibiotic – often in the absence of a documented infection and despite discussions regarding the appropriateness of antibiotic use. The varied use of antibiotics across care settings raises concerns about antibiotic resistance and underscores the need for antibiotic stewardship within palliative and hospice care. Antibiotic stewardship programs (ASPs) are a proven avenue for safely standardizing antibiotic use while improving patient outcomes across multiple clinical settings. However, little has been explored regarding the utility and design of ASPs in palliative and hospice care, and no studies to date have attempted to explore the use of antibiotics or ASPs among palliative care physicians in low- and middle-income countries (LMIC). Given that palliative care research is predominantly focused on Europe and North America, there is a pressing need to explore the LMIC palliative care experience, especially in regard to antibiotic stewardship.A systematic review of palliative and hospice care research across several countries revealed that 4% to 84% of patients in palliative or hospice care receive antibiotics.This study addresses this gap in the literature with a retrospective analysis of records from an Indian palliative care center, Trivandrum Institute of Palliative Sciences (TIPS), following the introduction of an ASP. This study identifies crucial patterns of antibiotic use and suggests potential avenues to support antibiotic stewardship within an LMIC palliative care setting.TIPS is the demonstration site for Pallium India, a charitable organization involved in palliative care education and patient care. Located in Kerala, India, TIPS provides care to inpatients in a 13–15 bed unit, to outpatients in clinics, and in their home-based program to any patient with life-limiting illness. During the time of the study, TIPS cared for approximately 3,300 patients annually  with 6–7 physicians and an annual mortality of 25%–30%. The TIPS patient population includes patients with terminal illnesses such as end-stage cancer as well as patients with chronic care needs such as chronic kidney disease and diabetes. At the time of this study, TIPS used paper clinical charting; however, patient registration, death data, and pharmacy records were recorded in encrypted Microsoft Excel spreadsheets.In December 2016, TIPS leadership launched an internal review to evaluate antibiotic prescribing practices at TIPS. This preliminary review revealed that antibiotic prescriptions were inconsistently documented. Moreover, TIPS physicians did not have access to standardized institutional guidelines when prescribing antibiotics. As a result, TIPS, in partnership with the University of Iowa, instituted an ASP in January 2017 with a goal to establish institutional guidelines for antibiotic use and to document antibiotic use at TIPS.Traditionally, all medications prescribed by TIPS physicians are dispensed by a central pharmacy at TIPS headquarters. Because care teams manage patients across multiple care settings, the TIPS pharmacy dispenses a pre-allotted stock of medications to each care team as well as a Medicine Replacing Book: a standardized paper pad that documents all prescriptions dispensed to patients. When prescribing a medication, TIPS physicians document the medication in both the patient chart as well as a Medicine Replacing Book and dispense the medication directly to the patient. Care teams then return to TIPS each day, submit the Medicine Replacing Book to the pharmacy, and TIPS pharmacists replenish stocks of any dispensed medications.1a and 1b). The AOF contained institutional antibiotic prescribing guidelines on one side and a template for tracking antibiotic prescriptions on the other side. When prescribing antibiotics , TIPS physicians were expected to review the prescribing guidelines, dispense a guideline-concordant antibiotic, and submit the completed AOF alongside their Medication Replacing Book to the TIPS pharmacy at the end of each day. Antibiotic order forms were collated by pharmacy staff and reviewed monthly by the antibiotic stewardship committee. The antibiotic stewardship committee was led by a local champion [AT] who chaired committee discussions regarding antibiotic use and piloted changes to institutional guidelines. All TIPS clinical staff were initially educated regarding AOFs during monthly administrative staff meetings and then reminded annually to continue AOF use. The AOF template and institutional guidelines were updated once by the antibiotic stewardship committee in March 2018 to include multiple changes such as the removal of fluoroquinolones to reflect changes in resistance at local hospitals as well as the addition of topical metronidazole for wound odor management (Supplementary File 2a and 2b).The novel antibiotic stewardship program was initiated with an interprofessional committee of physicians, nurses, and pharmacists, who designed an additional paper form known as the antibiotic order form  and the Pallium India Institutional Ethics Committee (IEC-11/2018). Data included antibiotic prescription data and identifiable patient data for all adult patients (≥18 years of age) prescribed antibiotics by TIPS medical staff between January 1, 2017, and October 31, 2019. The investigator [DT] collated antibiotic prescription data from the available records into an encrypted spreadsheet. Available prescription data did not stratify by care setting , so care setting was not collected. Patient data for antibiotic recipients were abstracted from available TIPS databases including TIPS patient death registry, TIPS patient palliative care registry, and daily clinic logs. All data handling was completed under the supervision of the local TIPS investigator and antibiotic stewardship committee local champion [AT]. This multivariate Poisson analysis was used to model gender, diagnosis, and age with the number of antibiotics prescribed. Statistical analysis was performed in RStudio , and the alpha threshold was set at 0.05.Planned study analysis included descriptive statistics of antibiotic prescription counts, AOF use, and guideline concordance as well as regression analysis of antibiotic prescription counts. However, data such as antibiotic dosage and duration data were available for only a minority of antibiotic prescriptions. Therefore, to evaluate for trends in antibiotic use, patients were categorized into 2 groups: (1) patients who received only 1 antibiotic prescription during the study period; and (2) patients who received 2 or more antibiotic prescriptions during the study period. Patient grouping by number of antibiotic prescriptions was used to compare antibiotic use between patients and to assess if there were target populations for future stewardship intervention. Data was modeled using a Zero-Inflated Poisson regression model. This is a mixture model with a binomial part and a Poisson part that assumes the data contains 2 types of patients: a group receiving one prescription and a group receiving more than 1 prescription where most patients fall into 1 group.3).Of note, patient age refers to the age at the time of registration for palliative care and not the age at the time when receiving antibiotics. Diagnosis refers to the diagnosis that qualifies a patient for palliative care. In order to attain adequately sized diagnosis groupings for the regression model, diagnoses were organized into chronic disease and non-chronic disease categories  that were documented on AOFs. Concordance was assessed according to 4 criteria: antibiotic choice, dose, frequency, and duration in days. Antibiotic dose, frequency, and duration were considered concordant only if the antibiotic choice reflected institutional antibiotic prescribing guidelines. Guidelines were available only for urinary tract infections, wound infections, and respiratory infections. Therefore, analysis of concordance was limited to AOFs indicating 1 of those 3 infection types. Because the AOF template and guidelines were updated in March 2018, analysis of concordance data was completed based on the prescribing guidelines listed on each AOF at the time of use. Prescription data collected from other sources such as Medication Replacement Books do not list indications for antibiotic use and were excluded from concordance analysis.P < 0.001, Table Out of 7,450 patients treated by TIPS staff during the study period, 675 (9%) patients received antibiotics with 1,448 total antibiotic courses prescribed. Patient gender and diagnosis data for antibiotic recipients are categorized in Table Overall, the most common antibiotics prescribed were topical metronidazole  TIPS patients died. Among 675 antibiotic recipients, 282 (42%) died with 133  receiving antibiotics within the final month of life. Since data from deceased patients who did not receive antibiotics were not collected, we were unable to determine which demographic or clinical factors predicted antibiotic use in the final month of life.In total, 458 32%) of all antibiotic prescriptions were documented in AOFs. Topical metronidazole was the least documented antibiotic with only 5% of prescriptions listed on AOFs. Sulfonamides were the most documented class with 89% listed on AOFs  met the criteria for analyzing prescription concordance with institutional prescribing guidelines. Of these, 260 (67%) reflected the guideline-recommended antibiotic choice and 70 (18%) demonstrated concordance across all 4 criteria of antibiotic choice, dose, frequency, and duration. Notably, there was an observed increase in concordance of antibiotic choice from 31% of AOFs in 2017 to 78% and 76% in 2018 and 2019, respectively.To our knowledge, this is the first study to explore how physicians prescribe antibiotics and adhere to antibiotic stewardship guidelines within a low- and middle-income country palliative care setting. The results of this study demonstrate that: (1) younger patients were more likely to receive more than one antibiotic course; (2) nearly half of all deceased antibiotic recipients received antibiotics within the final month of life; and (3) physicians did not consistently complete ASP documentation or follow institutional guidelines when prescribing antibiotics. Moreover, the focus on antibiotic stewardship led to early changes to antibiotic prescribing behaviors at TIPS including fluoroquinolone removal based on regional antibiotic resistance, recognition of variability in prescribing practices, and increased awareness of high topical metronidazole use for the management of wound odor. These results provide a benchmark of antibiotic use and stewardship concordance within an LMIC palliative care setting and highlight avenues for optimizing future antibiotic stewardship interventions. Moreover, younger palliative care patients within LMICs may have a higher incidence of comorbidities such as debilitating traumatic injury, which may increase infection risk and lead to higher antibiotic use. Due to our limited sample size, multivariate analysis of sub-groups such as traumatic injury patients was not possible. Nevertheless, the recognition of higher antibiotic use among younger patients provides a target for future antibiotic stewardship research and intervention.Regarding patient age, younger age appears to be the strongest factor affecting antibiotic prescribing. Age is a known factor affecting access to palliative care therapies with previous studies suggesting that younger age may independently influence palliative care physician perception of patient needs, length of palliative care stay, and access to life-prolonging therapies. Similarly, a survey of antibiotic usage among 16 hospitals across India showed that penicillins with beta-lactamase inhibitors were the most frequently prescribed antibiotics, accounting for 47.6% of antibiotic prescriptions. Yet, penicillin use at TIPS is rising, and increased antibiotic use is a known driver of resistance. Given the heavy reliance of TIPS and other Indian institutions on penicillins, optimizing antibiotic surveillance will be crucial to capturing excessive penicillin use and curbing future resistance.In terms of antibiotic choice, the use of penicillins at TIPS  is comparable to usage rates published by other medical centers in India. A study in Puducherry documented that amoxicillin accounts for ≥30% of antibiotic prescriptions across primary and tertiary care centers in the area. This nursing-led intervention likely complicated antibiotic stewardship documentation and may account for the reduced number of metronidazole prescriptions recorded on AOFs. Considering that the overuse of topical antibiotics may be also linked to increasing antibiotic resistance, tracking and standardizing formulations of topical metronidazole may be another priority for future stewardship initiatives within palliative and hospice care.Likewise, topical metronidazole use at TIPS  may be comparable to other facilities in India and internationally. Metronidazole is exclusively used as a topical agent at TIPS for the management of wound odor. However, due to cost restrictions, topical metronidazole at TIPS is often compounded by nurses on-site by crushing metronidazole tablets into petroleum jelly. This is a widely recommended method for palliative care teams to control wound odor, but, without a standardized formulation, drug concentrations may vary between patients. Existing stewardship guidelines – including AOF guidelines – do not typically reflect the challenges surrounding prescription decisions at the end of life, ultimately limiting their use among terminally ill patients. Therefore, a focus for future stewardship efforts may be to develop guidance to help providers navigate discussions surrounding antibiotic use at the end of life. Instead of strict prescription guidelines, it may be more appropriate for ASPs to provide a model for shared clinical decision-making that integrates patient preferences into existing guidelines to create an individualized care plan.Regarding antibiotic use at the end of life, 47% of antibiotic recipients who died received antibiotics in the final month of life. While this occurred in only 5% of all TIPS patients who died, it is appropriate to review the reasons for antibiotics in this setting, both at the bedside and in the antibiotic stewardship literature. Providers may be attempting to prolong survival, fulfill patient goals of care, or alleviate patient discomfort. A 2020 scoping review suggests that antibiotics do provide a limited survival benefit to some patients nearing the end of life and may also mitigate symptoms associated with certain infections such as urinary tract infections; however, these benefits may come at the cost of prolonging the dying process, compounding adverse effects, and promoting antibiotic resistance.Finally, completion of ASP documentation and concordance of antibiotic prescriptions with institutional antibiotic prescribing guidelines were inconsistent and revealed at least 1 potential area for improvement: leadership engagement. Specifically, engagement of clinical leadership appeared to increase use of AOFs. From January through March 2018, TIPS antibiotic stewardship committee members convened to update existing institutional antibiotic prescribing guidelines due to increasing concerns regarding local fluoroquinolone resistance. There was a marked increase in AOF use during subsequent months and an increase in concordance of antibiotic choice during subsequent years. These results suggest that even inconsistent implementation of an ASP with invested clinical leadership may provide an accessible avenue to introduce antibiotic stewardship into the LMIC palliative care setting with measurable improvements in antibiotic utilization.This study has several limitations. Several of the factors that may affect physician prescribing behavior including patient comorbidities, bacterial culture data, symptom burden before and after antibiotic use, socioeconomic status, location , and length of stay on the inpatient unit were not available for analysis. Additionally, due to severe flooding in 2018, some paper records may have been lost prior to data collection, thus limiting available results. Lastly, the scope of this study was restricted to prescribing behaviors by TIPS medical staff and may not necessarily reflect actual antibiotic consumption by patients.Despite these limitations, this retrospective study is the first to assess the outcomes of a recently launched interprofessional antibiotic stewardship program in an Indian palliative care setting. Results from this study fill a gap in the literature and could potentially help design future prospective studies using the AOF at other healthcare centers. Based on our findings, TIPS officials plan to further evaluate metronidazole use, increase awareness of the need for accurate record keeping by site of care, implement regular review of the AOF, and educate physicians on the consistent use of recommended antibiotic choices, dose, and duration. Further investigation is needed to evaluate perspectives of palliative care physicians, pharmacists, and nurses regarding antibiotic stewardship as well as factors that predict antibiotic use in the final month of life.Thomas et al. supplementary materialThomas et al. supplementary materialThomas et al. supplementary material


The energy-saving design of buildings effectively reduces heating energy consumption, improves indoor environmental problems in traditional houses, and plays an essential role in alleviating the increasingly severe environmental and energy problems. This study reviews the status of research on building energy-saving renovation and environmental improvement in various countries and summarizes their relevant research experiences. It described the theories and methods of survey, experiment, and numerical simulation to provide a theoretical basis for energy-saving renovation and environmental improvement of residential houses. Furthermore, it introduces corncob concrete material to provide new materials for the renovation of an old building. This research is based on the coastal Liyuan houses in Qingdao, and the problems of Liyuan houses are identified through in-field investigation. Then, through experimental simulations, the application and efficiency of corncob materials in the thermal performance improvement of the envelope are explored. The renovation of old residential houses in terms of energy-saving and environmental improvement is proposed from envelope renovation. With the exploration and extension of sustainable strategy attempted in China, the economic and agricultural development level has been improved continuously. However, in the process of fast growth, the balance between human activities and natural system has been broken; thus, the pressure on the ecological environment has gradually increased. In order to fix the balance between economic growth and environmental protection, the guiding policy of "concept of Eco civilization" has gradually gained popularity. Up to now, China has gained certain achievement in the Eco civilization construction. However, it is still confronting high pressure on the ecological environment.Agriculture, as the primary industry in China, is significant to the development of the integral national economy. Since 2017, The Green Production Policy has promoted clean production in agriculture and focused on the management of outstanding agricultural environment problems. In 2018, it was proposed to lead rural revitalization with the concept of green development. In 2019, the policy of high-quality development in agriculture promoted the improvement of rural human settlements comprehensively. In 2020, steadily improve the rural living environment was taken as one of the national development goals.As an opportunity of further optimization, taking advantage scientifically of the agricultural by-products, which has the features of abundant yield and flexible collection, and making use efficiently of biomass resources, can reduce the pollution to the living environment and promote the adaptation process of the living environment significantly. It could eventually achieve coordinated development of life, production and ecology.5.On the other hand, with the high speed in development of China's construction industry, the production of construction waste is also increasing. From preliminary statistics, the total amount of construction waste in China is about 2.1 billion to 2.8 billion tons, and the number is still increasing. The treatment of construction waste has become the major problem, which required reasonable solution urgently. With the continuous reduction of natural resources, it is especially important to improve the recycling efficiency of resources. In the outline of the 12th Five Year Plan for China's national economic and social development, it has been clearly proposed that “to accelerate the construction of a resource-saving production mode, to establish the concept of green development, and to realize improvement in Eco civilization”Construction waste comes from the demolished architecture and it should provide positive benefit to it. The use of construction waste as new building material has gained more attention recently. With the acceleration of urbanization, demolition and reconstruction projects generated a large amount of construction waste, which also accelerate the shortage of new construction resources. How to make full and efficient use of construction waste, especially the trials on making recycling concrete from waste, has become a concerned topic in many countries.The limited natural resources cannot satisfy the fast development in the construction industry gradually. The recycling concrete from construction waste can reduce the consumption of natural resources significantly. The recycling aggregate is an effective utilization of construction waste, which is economical and environmentally friendly as positive solution to the environmental problem. Under the trend of energy saving and environmental protection, the use of recycling aggregate applied in recycling concrete has become the research concerns in architecture.The comprehensive utilization of agricultural and construction waste is one of the most economical, ecological and effective methods. The co-processing of agricultural and construction waste can turn these recycling resources into valuable treasures, effectively alleviate the shortage of resources and energy, which can reduce the pollution of solid waste to the environment, but also achieve collective long-term development in environmental, economic and social benefits.7.In the process of renovation construction in China, the envelope of residential enclosures requires a large amount of energy-saving, environmentally friendly and low-cost insulation materials. Corncob is one renewable resource solution, which is convenient to obtain, energy-saving and environmentally friendly. By using corncob as concrete aggregate, the building cost could be reduced greatly. Corncob has high value as a building material, but it has not been widely used at home and abroad. Whether it can be used as an environmentally friendly building material in the construction industry still needs to be certificated in many aspects. Therefore, the application of corncob as a building material will have great potential and significant economic and social benefits possibilitiesTherefore, this paper makes experiment on fully utilization of the features of corncob, such as light weight, heat preservation and crack resistance, etc. At the same time, it co-disposes all-component recycling sand prepared from construction waste, and jointly prepares corncob-recycled aggregate composite ecological concrete, which develops a new technical solution of green ecological building concrete materials. It provided a solution to expand the high-end utilization of agricultural and recycling construction waste products. The research conclusion of this chapter will promote the coordinated development of rural economy and society, change the mode of rural economic development, and conform to the basic national policy of environmental protection and sustainable development in my country, and conform to the green ecological concept of harmonious coexistence between man and nature.China attaches great importance to the development of circular economy, the core of which is the recycling of resources. The current ecological environment problems have attracted great attention worldwide, and the recycling of resources has important practical significance for protecting and improving the ecological environment, especially in the resource utilization of solid waste in agriculture and construction.6. In addition, corncobs are uniform in texture, light in weight, have certain strength, thermal insulation, and sound absorption performance. They are a valuable secondary resource that urgently needs to be developed and utilized.Agricultural waste is a rich source of renewable energy, but with the development of science and technology, its utilization value is getting lower and lower, and it has gradually become one of the main pollution sources of the rural environment. In order to effectively alleviate this severe situation, it is necessary to make reasonable planning for agricultural waste resources, appropriately increase the comprehensive utilization rate of agricultural waste, and realize the recycling of waste resources, so as to turn agricultural waste from "waste" into "treasure". As an important recyclable resource, corncob has not been used in high value in the process of agricultural development. It is usually treated by means of rural burning for heating, animal husbandry, and returning plow to fields, which not only pollutes the ecological environment, but also Caused a waste of natural resourcesConstruction waste comes from construction and should be returned to construction. The use of construction waste as building materials has attracted much attention. With the acceleration of urbanization, demolition or reconstruction projects will generate a large amount of construction waste, and also bring about a serious shortage of building material resources. How to fully and efficiently utilize construction waste, especially the application of waste concrete, has become a hot topic of joint research in many countries.The limited nature of natural resources gradually does not correspond to the rapid development of the construction industry. The recycling of waste concrete can effectively reduce the consumption of natural resources. Recycled aggregate is an effective use of construction waste resources, which is both economical and environmentally friendly, and has significant benefits. Domestic research on recycled aggregates mainly focuses on the mechanical properties of recycled aggregates and recycled concrete.7 and others strengthened the quality of recycled fine aggregate, the particle size of the recycled fine aggregate after particle shaping was improved, the apparent density was slightly increased, and the water absorption rate was reduced; Guo et al.9 from the physical and the quality of recycled coarse aggregate is improved in chemical aspects. The study found that the 24 h water absorption rate of recycled coarse aggregate after secondary particle shaping treatment is reduced by 2.5%, and the crushing index is reduced by 9%. The performance parameters are close to natural aggregate; After chemical treatment of recycled coarse aggregate, its 24 h water absorption rate decreased by 2.3%, and the crushing index decreased by 5%; It also improves the performance of recycled aggregate, so as to achieve the purpose of improving the performance of recycled aggregate concrete13.Recycled aggregates have poor performance and cannot be directly used in concrete. In order to improve the utilization of recycled aggregates and promote the development of recycled concrete, it is necessary to strengthen the recycled aggregates. LiIn the experimental process of the research on the performance of corncob-recycled aggregate composite ecological concrete, the selected coarse aggregate is corncob in agricultural waste. Similar to corn stover. When selecting raw materials, based on the principle of local materials, the corncob used in the experiment was provided by growers near Chengyang District, Qingdao. Considering that the corncob bone brings more impurities to the corncob during the mechanical threshing process, the corn leaves, corn silk, residual corn kernels and other magazines other than the corncob should be manually sorted and removed before the crushing treatment. The appearance and morphology of corncob before and after selection are shown in Fig. In the test, the agricultural pulverizer Fig.  was usedAfter sieving, corncob particles with different particle sizes were obtained, and their appearance and morphology were shown in Fig. When the service life of some buildings is too long, the performance of recycled coarse aggregate is weakened, and the dismantled construction waste is no longer suitable for use as coarse aggregate for recycling; or the building itself cannot continue to build due to the quality of coarse aggregate. When the construction waste is completely crushed into fine aggregate due to construction requirements, the construction waste can be completely crushed into fine aggregate, which is called full-component recycled sand, which is the fine aggregate used in the test.In the preparation process, the preparation process of full-component reclaimed sand can be subdivided into full-component simple crushing reclaimed sand and full-component particle shaping reclaimed sand. The main feature of the whole-component particle shaping reclaimed sand preparation process is that all the waste concrete blocks are used to prepare the reclaimed sand. That is, during the initial simple crushing, the natural coarse aggregate in the original waste concrete is no longer used to prepare the recycled coarse aggregate, but is all crushed into recycled sand. increase accordingly.However, the performance of full-component particle shaping regenerated sand is better than that of full-component simple crushing regenerated sand, and the economic cost increases less. Therefore, in the experiment, the regenerated sand with full-component particle shaping was finally selected as the fine aggregate for preparing corncob-recycled aggregate ecological concrete.Coarse aggregate corncob is obtained by sorting, crushing and screening of corncob bones, and the waste concrete is crushed and reshaped to obtain full-component regenerated sand, which is used as the fine aggregate for this test; The water consumption, the two together constitute the water consumption of the concrete test, and the mixing ratio plan is designed with the mortar ratio and the volume content of the corncob as the influencing factors.By exploring the relationship between the mass and volume of the corncob, the actual mixing quality of the corncob was determined; the basic performance  and mechanical properties (cube compressive strength under different curing age conditions), and systematically explore the change law of its performance by analyzing the measured test results.Based on the experimental research, using the method of combining data analysis and theoretical analysis, the co-processing of agricultural waste and construction waste is used to prepare corncob-recycled aggregate composite ecological concrete with significant environmental protection effect. The performance provides a certain theoretical basis for the later application and promotion of agricultural engineering.Add reclaimed sand and cement and mix well.Add water and water reducing agent for stirring, and the stirring time is controlled within the range of 10–15 s.Add the mixed corncob particles, and after stirring for 2–3 min, put the concrete mixture into the test mold.Use a vibrating table to vibrate, compact and smooth.In the test, a horizontal mixer was used to prepare a corncob-recycled aggregate composite ecological concrete mixture, as shown in Fig. Because the corncob is light in texture, it will float up during the vibration process, so the vibration time should not be too long, and it is appropriate to control it within 3-5 s in the test. In addition, after the ecological concrete is installed, in order to prevent its moisture from evaporating, it should be immediately covered with plastic wrap to lock the moisture, and marked for easy identification, as shown in Fig. All the authors declared the collection and use of corncob materials applied in this study complied with relevant local institutional, national and international standards and legislation.This section may be divided by subheadings. It should provide a concise and preciseThe specific preparation process of the full-component particle shaping regenerated sand is as follows: firstly, the waste concrete blocks are simply crushed into 6–7 mm particles by a jaw or hammer crusher, and then they are shaped by a particle shaping machine. Sand is called full-component particle shaping regenerated sand. Because the waste concrete contains about 50% of natural coarse aggregate, the full-component particle shaping reclaimed sand contains a large amount of reclaimed sand generated from natural coarse aggregate, which can effectively improve the firmness of the reclaimed sand. According to the relevant requirements in GB/T25176-2010 “Recycled Fine Aggregates for Concrete and Mortar”, the main performance indicators of the regenerated sand with full-component particle shaping are tested. Particle gradationComparing the regenerated sand with full-component particle shaping and natural sand prepared in the test, the particle gradation and fineness modulus are shown in Table Mud content (Micro powder content)Generally, reclaimed sand is generated by crushing natural stones. The particles with a particle size of less than 75 μm in the sand are stone powder, while the particles with a particle size of less than 75 μm in the test concrete regenerated sand are mainly composed of cement stone powder, stone powder and soil. Therefore, the concrete is recycled. The content of particles with a particle size of less than 75 μm in the sand is defined as the content of fine powder. Table Mud contentyThe mud content of the reclaimed sand can be understood as the cement stone chips attached to the surface of the reclaimed sand, washed with water, and scattered by hand. The specific test results are shown in Table SolidityIn this test, the single-stage maximum crushing index is used to test the firmness of the reclaimed concrete. The larger the single-stage maximum crushing index is, the lower the firmness is. The specific test results are shown in Table Density3 lower than those of the natural river sand and are close to the natural sand in terms of density performance.The density of sand includes bulk density, compact density, apparent density and porosity. The specific test results are shown in Table  Water demand ratio and strength ratio of reclaimed rubber sandIn the test, reclaimed sand and standard sand were used to prepare reclaimed sand and standard sand that meet the fluidity requirements, respectively. The water demand ratio of reclaimed sand and the strength ratio of reclaimed sand measured in the experiment are shown in Table Cementitious materialsThe fast-hardening sulfo-aluminate cement SAC42.5 is selected. The aliphatic superplasticizer produced by a building material company in Qingdao is used. Using ordinary tap water, in line with JGJ63-2006. Determination of corncob qualityAdd 50 g, 60 g, 70 g, 80 g, 90 g, and 100 g of mixed corncob particles to the standard mortar test , stir evenly, and put them into a regular container, and then observe the change in the height of the mortar surface.With the change in the number of mixed corncob particles, by actually measuring the height of the freshly mixed mortar in the regular container, the linear relationship between the mass and volume of the corncob is obtained. Based on the linear relationship of corncob mass-volume, the ratio of corncob to the total volume can be calculated, and then the ratio of corncob to cementitious material can be determined according to the different volume contents of corncob set in the experiment (defined as cob glue ratio), and finally, the quality of corncob used for preparing corncob-recycled aggregate composite ecological concrete can be calculated.Determination of test water consumptionThe water consumption of the test should consider the water consumption of the mortar and the water consumption of the corncob, that is, the total water consumption = the water consumption of the mortar + the water consumption of the corncob.Determination of water consumption for mortarIn order to ensure that the concrete mixture has good working performance, the fluidity of the mortar is controlled to be 175–180 mm, and the amount of aliphatic water reducing agent is 2% of the amount of the cementitious material. The results are shown in Table Determination of corncob water consumptionDue to the different harvesting time and storage environment of corncob, the water content of corncob itself is quite different. The water absorption test of corncob used in this experiment should be carried out in advance. During the test, take a certain mass of dry-weight mixed corncob particles and soak them in water, and measure the quality of the corncob at intervals of 10 min until the weight no longer increases, and record the quality of the corncob at this time. The water absorption rate of corncob is calculated according to formula  and 0.0790 W/(m K), and the thermal insulation is good, which provides a feasibility and theoretical basis for biomass materials as thermal insulation building materials. Generally speaking, the thermal insulation properties of materials mainly depend on the thermal resistance. The lower the thermal conductivity of building materials, the better the thermal insulation effect, and the thermal conductivity of different biomass materials variesThe test results showed that the dry density of concrete blocks gradually decreased with the increase of corncob stalks. With the more increase of the amount of corn stalks, the further decrease of concrete compressive strength. There is an obvious linear relationship between the amount of corn stalks added and the compressive strength of concrete Fig. . CompareCorn stalks and corncob are by-products of the same plant. At present, the use of corn stalks as thermal insulation building materials has been studied, but corncob has not been widely used. In engineering, materials with thermal conductivity less than 0.23 W/(m K) are called thermal insulation materials. The thermal conductivity of corncob is less than 0.23 W/(m K), so it can be used in buildings to reduce the thermal conductivity of concrete, thereby improving Thermal insulation properties of concrete. Therefore, in this experiment, the influence law of the thermal conductivity of corncob-recycled aggregate ecological concrete was studied, and the results are shown in Fig. As can be seen from Fig. In addition, when the ratio of mortar to sand is 1:3, the thermal conductivity of eco-concrete is less discrete. When the volume content of corncob is 30%, the thermal insulation of ecological concrete is relatively weak, and the thermal conductivity of concrete is about 0.2479 W/(m·K). When the volume content of corncob increases to 50%, the thermal conductivity of ecological concrete decreases. It is 0.2011 W/(m K), that is, for every 5% increase in the volume content of corncob, the thermal conductivity decreases by about 4.7%, and the decrease is relatively large. It can be seen that the corncob aggregate has a significant effect on the thermal conductivity of ecological concrete.The thermal conductivity of concrete is inseparable from the constituent materials. In addition to recycled coarse aggregates, the influence of cementitious materials and recycled fine aggregates on the thermal conductivity of concrete cannot be ignored. The effect of different cement-sand ratios on the thermal conductivity of ecological concrete is shown in Fig. The recycled fine aggregate produces cracks during the crushing process, which increases the internal porosity of the concrete, increases the water absorption rate, and increases the air content, resulting in a decrease in the thermal conductivity of the corncob-recycled aggregate ecological concrete.As can be seen from Fig. In general, the thermal conductivity of corncob-recycled aggregate composite ecological concrete is between 0.1803 W/(m K) amd 0.2479 W/(m K) under the condition of different mortar ratio and corncob volume content, which is very close to the thermal insulation material, and provides a basis for the research on the preparation of energy-saving thermal insulation wall panels.The corncob itself has high porosity, loose pores, and high-water absorption. With the increase of the volume content of the corncob, the water absorbed by the ecological concrete increases. In the dry state, the dry density of the ecological concrete varies with the ratio of mortar to sand. It gradually decreased with the increase of corncob volume content.3 and the volume content of corncob of 50% was selected, and its thermal conductivity was 0.18.Corncob is a lightweight, loose and porous fibrous material that retains a large amount of air in the pores of the corncob and is a biomass material with a small thermal conductivity. As the volume content of the corncob increases, the independent pores in the ecological concrete structure increase, and the air content inside the concrete increases, the thermal conductivity of the ecological concrete decreases, and the thermal insulation performance is better, which can achieve the effects of heat insulation and heat loss prevention. In the end, the corncob concrete material with the cement-sand ratio of 1:4, the water consumption of 218.3 kg/mThe texture of corncob is uniform, but it is lightweight and porous, and its own compressive strength is low. With the increase of the volume content of corncob, the compressive strength of ecological concrete shows a downward trend; with the continuous reduction of the mortar ratio, the proportion of recycled sand increases, the content of cementitious materials in ecological concrete is reduced, the bonding force between aggregates is weakened, and the porosity is further increased due to insufficient aggregate wrapping, which makes the compressive strength of ecological concrete continue to decrease. Under the conditions of different mortar ratio and volume content, the 28-day compressive strength of ecological concrete is between 2.1 and 5.4 MPa. Although it is lower than the compressive strength of ordinary concrete, it can still meet the requirements of masonry.This section systematically studies the influence of different cement-sand ratios and corncob volume content on the thermal conductivity of corncob-recycled aggregate composite ecological concrete, and draws the following conclusions:


Objective: To investigate adverse events (AEs) associated with denosumab (Dmab) and zoledronic acid (ZA), compare their association strengths, and explore potential applications to provide clinical reference.Methods: We collected data from FAERS from January 2004 to November 2022 and mined AE signals for Dmab and ZA using ROR values. We compared signal intensity for same AEs and investigated off-label use. We also examined their AEs in adjuvant therapy for breast and prostate cancer.Results: 154,735 reports of primary suspect drugs were analyzed in the FAERS database . Dmab and ZA had 333 and 1,379 AE signals, with 189 overlaps. The AEs of Dmab included death (ROR:3.478), osteonecrosis of jaw (ROR:53.025), back pain (ROR:2.432), tooth disorder (ROR:16.18), bone pain (ROR:6.523). For ZA, the AEs included osteonecrosis (ROR:104.866), death (ROR: 3.645), pain (ROR:3.963), osteonecrosis of jaw (ROR: 91.744), tooth extraction (ROR: 142.143). Among overlap signals, Dmab showed higher strength in exostosis of the jaw (ROR: 182.66 vs. 5.769), atypical fractures (ROR: 55.589 vs. 9.123), and atypical femur fractures (ROR:49.824 vs. 4.968). And ZA exhibited stronger associations in abscess jaw (ROR: 84.119 vs. 11.12), gingival ulceration (ROR: 74.125 vs. 4.827), increased bone formation (ROR: 69.344 vs. 3.218). Additionally, we identified 528 off-label uses for Dmab and 206 for ZA, with Dmab mainly used in prostate cancer (1.04%), breast cancer (1.03%), and arthritis (0.42%), while ZA in breast cancer (3.21%), prostate cancer (2.48%), and neoplasm malignant (0.52%). For Dmab in breast cancer treatment, AEs included death (11.6%), disease progression (3.3%), and neutropenia (2.7%), while for ZA included death (19.8%), emotional disorder (12.9%), osteomyelitis (11.7%). For prostate cancer treatment, Dmab`s AEs were death (8.9%), prostate cancer metastatic (1.6%), renal impairment (1.7%), while ZA`s included death (34.4%), general physical health deterioration (19.9%), and hemoglobin decreased (18.9%).Conclusion: Our analysis of FAERS database provided postmarketing surveillance data and revealed different strengths of reported AE signals between Dmab and ZA in some of their common AEs. It’s also worth noting that both drugs have potential off-label applications, which could introduce new AEs. This highlights the necessity for safety monitoring when using Dmab and ZA off-label. Denosumab (Dmab), the first and only one receptor activator of NF-κB ligand (RANKL) inhibitor so far, was approved for marketing by the U.S. Food and Drug Administration (FDA) in 2010 and Zoledronic acid (ZA) approved in 2001. They have similar efficacy such as applying for prevention and treatment of osteoporosis in postmenopausal females, osteoporosis in males, glucocorticoid-induced osteoporosis, hypercalcemia of malignancy, and preventing skeletal-related events (SREs) secondary to solid tumors metastases . HoweverIn the past decade, significant efficacy of both drugs has been extensively documented, whereas, novel AEs not well studied were gradually raised during the clinical application. Furthermore, novel mechanisms as well as application also emerged. We hope this analysis based on FAERS database will provide safety profile in support of future studies in the application of Dmab and ZA. And to provide reference directions for exploring their potential clinical applications.The data for this retrospective pharmacovigilance study were obtained from FAERS, a global spontaneous reporting system that collects safety information on approved drugs and therapeutic biologic products from various sources including manufacturers, healthcare professionals, and consumers. FAERS is the primary source of post-marketing safety monitoring and evaluation for the FDA, and it provides signal detection and quantification of the association between drugs and AEs . The dathttp://openvigil.sourceforge.net/). Individual safety reports (ISRs) for Dmab and ZA were extracted from the FAERS database. ISRs are the count of raw data extracted by OpenVigil 2.1 and an ISR code represents an AE report.To extract adverse event (AE) reports from the FDA Open-FDA program, we utilized the online tool OpenVigil 2.1  included exostosis of jaw (ROR: 182.66 vs. 5.769), atypical fracture (ROR: 55.589 vs. 9.123), and atypical femur fracture (ROR: 49.824 vs. 4.968), while the AE signals of ZA with stronger correlation than Dmab (d > 50) were related to abscess jaw (ROR: 84.119 vs. 11.12), gingival ulceration (ROR: 74.125 vs. 4.827), increased bone formation (ROR: 69.344 vs. 3.218), and bone disorder (ROR: 68.221 vs. 3.189), among others.As there were numerous shared AEs between Dmab and ZA, we conducted a further comparison of the overlapping AE signals. Out of the 189 identical positive AE signals between the two drugs, 29 AEs of Dmab exhibited stronger correlation than ZA, while 160 AEs of Dmab had weaker correlation than ZA, as determined by the ROR value. While analyzing the data, we found off-label use was also a significant signal. Therefore, we further analyzed the data on off-label use. As we observed mixed reports of different specifications for each drug, for example, Dmab had a 60 mg specification for giant cell tumor and hypercalcemia of malignancy, while the 120 mg specification was used for postmenopausal osteoporosis. Similarly, ZA had two different specifications with mixed reports. Therefore, we combined the FDA-approved indications for both specifications of each drug and compared them with the indications in the database to identify off-label uses. We found 528 types of off-label use for Dmab and 206 types for ZA. We conducted a comparison of the AEs associated with Dmab and ZA in off-label use for breast cancer and prostate cancer. After comparing with the FDA-approved instructions and removing similar AEs, we found 451 AEs in Dmab and 848 AEs in ZA for breast cancer treatment. For prostate cancer treatment, we found 341 AEs in Dmab and 583 AEs in ZA that were not mentioned in the drug instructions.In breast cancer treatment, the top AEs associated with Dmab were death (11.6%), disease progression (3.3%), breast cancer metastatic (2.7%), neutropenia (2.7%), emotional disorder (2.3%), and pyrexia (1.7%). For ZA, the most frequent AEs were death 19.8%), emotional disorder (12.9%), osteomyelitis (11.4%), neoplasm progression (10.7%), cardiac disorders (10.6%), and impaired healing (9.5%). Cardiac disorders in patients with breast cancer treated with ZA included tachycardia (3.04%), congestive heart failure (1.35%), arrhythmia (1.01%), and palpitations (1.01%). Both drugs were associated with varying degrees of mental illness such as emotional distress, depression, and personality disorder, particularly in treating breast cancer, even leading to suicidal ideation. %, emotioIn prostate cancer treatment, the top AEs associated with Dmab were death (8.9%), prostate cancer metastatic (1.6%), and renal impairment (1.7%), while for ZA, the most common AEs were death (34.4%), general physical health deterioration (19.9%), and hemoglobin decreased (18.9%). Additionally, ZA was also associated with increased prostatic specific antigen (18.4%) and cardiac disorders (13.6%), while Dmab was associated with emotional disorder (1.6%) and cardiac disorders (1.3%).In this study, we performed a pharmacovigilance analysis using FAERS to investigate suspected AEs and off-label uses associated with Dmab and ZA. The data covers a substantial timeframe from 2004 to 2022, during which these two medications were administered in clinical practice at different time periods. Notably, the reporting rate for AEs can differ not only among various drugs but also for the same drug as time progresses . AdditioNonetheless, the current understanding of these drugs is not yet fully comprehensive, and many AEs still require adequate attention. To better understand the AE profile of these drugs, it is recommended to collect as much clinical data as possible and conduct more in-depth analysis and evaluation.The most frequent AEs of Dmab were death, osteonecrosis of jaw, back pain, tooth disorder, bone pain and hypocalcaemia and those for ZA were osteonecrosis, death, pain, osteonecrosis of jaw, and tooth extraction. The AEs identified in this analysis were generally in line with the known AEs of these drugs, indicating the validity of the study and suggesting that the findings may accurately reflect real-world clinical practices.d > 20), the significant signals were exostosis of jaw, atypical fracture, and atypical femur fracture, suggesting that Dmab may be more prone to these AEs than ZA. Exostosis of jaw may be associated with the widely recognized osteonecrosis of the jaw (ONJ), which is a rare but serious side effect of anti-bone resorption inhibitors. Although a study demonstrated that patients with bone metastases treated with Dmab or ZA had similar incidences of ONJ (d > 50) were mostly related to oral problems, which may also have potential implications for ONJ. Regular dental examinations should be conducted when using Dmab and ZA.It is known that Dmab and ZA share many similar AEs. In our study, we conducted a comparative analysis to assess the signal strength of AEs between these two drugs. Among the signals of Dmab stronger than ZA  for the treatment of bone loss or preventing fracture in non-metastatic prostate cancer, while ZA currently lacks FDA approval. Several small randomized trials have shown that bisphosphonates can increase BMD in patients with non-metastatic prostate cancer . Note thAs for the treatment of osteoarthritis (OA), a study in rabbits with experimental knee osteoarthritis showed that ZA had protective effect on articular cartilage and subchondral bone . An initIn conclusion, mining new indications from the database has the potential to expand drug application range, promote drug research and development, and improve clinical practice. However, it is crucial to conduct further real-world research to validate these new indications and ultimately benefit patients.According to the reports, disease progression was observed more frequently in the treatment of breast cancer and prostate cancer with Dmab or ZA. However, current evidence does not establish a definitive link between tumor progression and drug exposure. Our study also found a high frequency of neutropenia among breast cancer patients treated with Dmab, which is consistent with reports of neutropenia in a phase III study of multiple myeloma patients treated with both Dmab and ZA . Mental It's important to acknowledge several limitations that raise questions about its direct real-world applicability. Looking at the FAERS database, there are several aspects to consider. First, the cases registered in spontaneous reporting systems are only those of drug-induced AEs, not the total number of patients treated with the drugs , making Regarding disproportionality analysis, it solely represents statistical correlation between drugs and AEs and do not permit the establishment of causal associations between reported AEs and specific medications . FurtherAlso, the study’s failure to compare the impact of different specifications on indications may lead to incomplete evaluation of the drugs’ safety and efficacy. Additionally, it solely focuses on potential off-label use, neglecting over-the-label use of different specifications and their safety profiles, potentially overlooking certain safety issues and differences in effectiveness. Furthermore, the article may be affected by selection bias in data or inadequate analysis methods, which may impact the accuracy of drug evaluation and conclusions. The article may also not fully consider other factors affecting drug use, such as individual differences among patients, comorbidities, or the influence of other drugs.However, the FAERS database gathers AE reports associated with drugs and therapeutic biologic products, which is a valuable resource to identify potential safety issues. Despite the aforementioned limitations, disproportionality analysis is now a validated method in the field of drug safety research and surveillance . It has , our findings highlight the importance of safety monitoring when using Dmab and ZA off-label. Moreover, considering the limited research focused on this specific aspect, our study may serve as a reference point for future investigations, contributing to drug safety vigilance efforts. Finally, due to the inherent limitations of spontaneous reporting databases, which inevitably contain potential biases, there is an urgent need for well-designed comparative safety studies to validate these findings.In conclusion, our study found that both Dmab and ZA have similar trends in AE distribution. However, Dmab is statistically associated with a higher risk of jaw exostosis and atypical femur fractures, while ZA has a statistical link to more oral problems. It is important to note that both drugs have potential applications beyond their approved indications, particularly in the treatment of various cancers and osteoarthritis, and some new AEs may come with those off-label use, including mental health disorders, neutropenia and kidney damage, and heart problems. Given the correlation between the analysis results from spontaneous reporting systems databases and clinical safety studies , our fin


Pseudomonas aeruginosa cells are mainly responsible for the recurrence and non-responsiveness to antibiotics of cystic fibrosis (CF) lung infections. The sub-inhibitory antibiotic concentrations found in the CF lung in between successive therapeutic cycles can trigger the entry into the VBNC state, albeit with a strain-specific pattern. Here, we analyzed the VBNC cell induction in the biofilms of two CF P. aeruginosa isolates, exposed to starvation with/without antibiotics, and investigated the putative genetic determinants involved. Total viable bacterial cells were quantified by the validated ecfX-targeting qPCR protocol and the VBNC cells were estimated as the difference between qPCR and cultural counts. The isolates were both subjected to whole genome sequencing, with attention focused on their carriage of aminoglycoside resistance genes and on identifying mutated toxin–antitoxin and quorum sensing systems. The obtained results suggest the variable contribution of different antibiotic resistance mechanisms to VBNC cell abundance, identifying a major contribution from tobramycin efflux, mediated by MexXY efflux pump overexpression. The genome analysis evidenced putative mutation hotspots, which deserve further investigation. Therefore, drug efflux could represent a crucial mechanism through which the VBNC state is entered and a potential target for anti-persistence strategies.Persistent and viable but non-culturable (VBNC)  Pseudomonas aeruginosa lung infections in cystic fibrosis (CF) patients [P. aeruginosa has been detected in sputum samples from hospitalized CF patients undergoing antibiotic treatment [Recurrence and unresponsiveness to antibiotics are two of the main features of patients , most lipatients . Among ppatients , which ipatients , and VBNreatment .P. aeruginosa biofilm model [P. aeruginosa isolates [We previously described the VBNC-inducing role of starvation combined with sublethal concentrations of tobramycin in an in vitro lm model . In an aisolates , one of P. aeruginosa PAO1-N, C30, and AR86 were developed for 48 h in lysogenic broth (LB) and then maintained for 90 days in non-nutrient (NN) broth, , with/without tobramycin. Each strain exhibited a peculiar pattern of adaptation to the stress conditions, with different trends in VBNC subpopulation development (The in vitro biofilms of elopment .P. aeruginosa PAO1-N were detected in the 48-hour biofilms developed in LB (corresponding to time 0 days in p < 0.01) greater abundance.No VBNC cells of P. aeruginosa AR86 exhibited a persistent population entirely made up of culturable cells (between 8 × 106 and 3 × 107 CFU/mL), with no VBNC cells detected after the exposure to starvation  of VBNC cells in 48-hour biofilms grown in the rich medium. However, over the 90 days of stress exposure, arvation D.P. aeruginosa C30 showed a remarkable shift to the VBNC state in the first month of starvation, which was significantly more evident (p < 0.01) in tobramycin-exposed biofilms (92.6% vs. 75% of TVCs). Next, the number of culturable cells gradually increased over time, representing the 100% of the whole P. aeruginosa population after three months  and the minimum biofilm-eradicating concentration (MBEC) of tobramycin of different clones recovered from biofilm subcultures. No increases in MIC (16 and 256 µg/mL) or MBEC (256 and >2048 µg/mL) for P. aeruginosa C30 and AR86, respectively, were observed. In the presence of a subinhibitory concentration of the non-specific efflux-pump inhibitor carbonyl cyanide 3-chlorophenylhydrazone (CCCP), i.e., 100 µM (P. aeruginosa C30 fell to 2 µg/mL (corresponding to susceptibility), while neither P. aeruginosa PAO1-N norAR86 showed significant MIC variations. These results were consistent with previous observations [P. aeruginosa C30.The evolution over time of the tobramycin susceptibility of the , 100 µM , the tobrvations , indicatP. aeruginosa PAO1-N and C30 at both the phenotypic and the gene-expression level. Firstly, the ethidium bromide (EtBr) accumulation in the bacterial cells was monitored in cultures grown in absence/presence of sub-MIC tobramycin  and the greater effect of 100 µM CCCP on C30. Therefore, the inhibition of the efflux pump activity in C30 is responsible for reversion to the tobramycin susceptible phenotype.Compared to the PAO1-N, P. aeruginosa C30 and AR86 after their exposure to tobramycin, the two strains were subjected to whole genome sequencing (WGS) and were compared in terms of their aminoglycoside resistance determinants and sequence type (ST), as well as the presence of mutations in their toxin–antitoxin (TA)/quorum sensing (QS) systems. The two strains belonged to different STs; specifically, P. aeruginosa AR86 belonged to ST260, and P. aeruginosa C30 belonged to ST621. Both these STs are of clinical origin, but they are not among the most difficult-to-eradicate strains in CF patients [To shed some light on the different levels of abundance of VBNC in patients .ant(2”)-Ia gene by P. aeruginosa AR86 [aph(3′)-IIb and aac(6′)-Ib3, known as aminoglycoside modifying enzymes.The sequencing data confirmed the carriage of the adenylyltransferase osa AR86  and highP. aeruginosa PAO1, the two CF isolates showed several nucleotide substitutions in the TA  and AR86  were used [P. aeruginosa PAO1-N, showing a lower decrease in culturable cells and lower amounts of VBNC cells. Potentially, the two CF strains, having already experienced the nutrient-depleted environment of the deepest layers of the lung biofilm, became more prone to growing in unfavorable conditions.We previously pointed out the prominent role of tobramycin subinhibitory concentrations in the shift of biofilms , which iere used . Both isP. aeruginosa C30 exposed to tobramycin produced the greatest amount of VBNC cells (92.6%) after only 30 days of exposure, but these cells were completely lost after two further months. This could have been due either to the metabolic reactivation of the non-culturable cells [P. aeruginosa population in biofilms exposed to starvation and antibiotic pressure for extended periods is currently under consideration for future studies. Interestingly, P. aeruginosa AR86 never developed VBNC cells upon exposure to stress.The most striking results were those obtained in the presence of the 1/4x MIC tobramycin. The laboratory strain PAO1-N shifted to the VBNC state more slowly, but with the continuous induction of VBNC forms over time, confirming previously obtained results . Conversle cells  or to thThe bases of these three strain-specific patterns were investigated at both the phenotypic and the genetic level.We can exclude the evolution of microbial subpopulations less affected by tobramycin than the parental strains, as no increases in MIC or MBEC values were observed in the subcultures recovered throughout the experiment.The genome analyses evidenced relevant features regarding aminoglycoside resistance determinants and mutations in the TA and QS modules, possibly involved in the response to the drug.aph(3′)-IIb gene, mainly related to amikacin resistance, was found in both the CF strains, but also in P. aeruginosa PAO1 (NC_002516.2); (ii) aac(6′)-Ib3 was found in both clinical isolates and, therefore, it was not considered critical in the strain-specific response to the tobramycin sub-MIC; (iii) the presence of the inactivating enzyme ant(2″)-Ia gene was found only in P. aeruginosa AR86 and conferred high-level resistance (MIC = 256 µg/mL); (iv) P. aeruginosa C30 did not harbor further aminoglycoside resistance determinants, and exhibited low-level tobramycin resistance (MIC = 16 µg/mL), which was related to the activity of chromosomal efflux pumps, probably MexXY-OprM, which is the main one involved in aminoglycoside extrusion. This assumption was experimentally verified by the EtBr-efflux and mexY-expression assays, which confirmed a greater activity of MexXY-OprM in this isolate compared to P. aeruginosa PAO1-N. Consistently, P. aeruginosa C30 was the only strain to exhibit increased tobramycin susceptibility when exposed to the efflux pump inhibitor, CCCP, which non-specifically hinders efflux by uncoupling the proton gradient responsible for the pumps’ activity. Considering the tobramycin-induced development of VBNC cells as a hormetic response [P. aeruginosa C30 compared to the laboratory strain, we can assume that the tobramycin binding to the bacterial ribosome triggers the expression of genes (still to be identified) involved in the shift to the VBNC state, similarly to the described development of tolerance to aminoglycosides [P. aeruginosa C30 compared to P. aeruginosa PAO1-N after exposure to tobramycin may thus result in an early shift to the VBNC state of most of the bacterial population, followed by VBNC cell reactivation and multiplication as a consequence of drug extrusion, a behavior not observed in the absence of tobramycin. This is consistent with our previous data, which evidenced the role of the MexXY-OprM pump in the tolerance of P. aeruginosa to tobramycin [P. aeruginosa AR86 might have been due to the modification of tobramycin by Ant(2”)-Ia before reaching the intracellular environment, which prevents the action of tobramycin either as a stress factor or as a gene expression inducer.Concerning acquired aminoglycoside resistance determinants, (i) the  NC_00251.2; (ii) ycosides . The grebramycin  and withbramycin ,15. AccoP. aeruginosa CF isolates and, therefore, they were not considered as critical in affecting the specific shift to the non-culturable state exhibited by each isolate. In particular, no mutations were observed in the DNA-binding domain of the rhlR gene, which is involved in transcription regulation. Nevertheless, the obtained results evidenced the rhlR , P. aeruginosa ATCC 27853, and the previously characterized CF strains P. aeruginosa C30 and AR86, both resistant to tobramycin, through efflux (MIC = 16 µg/mL) and aminoglycoside modification (MIC = 256 µg/mL), respectively, [The reference strains P. aeruginosa ATCC 27853 as control strain. Only discrepancies > 2 folds were considered as significant. The CCCP MIC against each strain was determined in the same way, using the concentration range of 400–12.5 µM. Tobramycin MBEC was determined according to the protocol described by Revest et al. [P. aeruginosa biofilms grown in 96-well plates and in the concentration range of 2048–2 µg/mL.Tobramycin MIC was determined by the broth-microdilution method, according to CLSI guidelines , using tt et al. , using PP. aeruginosa were developed in 35 mm petri dishes in LB broth, and then sub-cultured and maintained in NN broth non-supplemented/supplemented with ¼x MIC tobramycin for 90 days, as previously described [600 = 0.1) were inoculated in the plates and incubated at 37 °C for 48 h, and the medium was refreshed after the first 24 h. Next, the rich LB medium was discarded and, after washing with phosphate-buffered saline, substituted with NN broth. Every 30 days, starting from the shift from the rich to the starving medium, biofilms were assessed for their content of culturable P. aeruginosa or TVCs.The in vitro biofilms of escribed . BrieflyecfX-qPCR protocol and confirmed by live/dead flow-cytometry assays [Biofilms were mechanically detached and lightly sonicated; next, suitable 10-fold biofilms dilutions were used. Abundance of TVCs was determined by the previously validated y assays . CulturaOnly discrepancies ≥0.5 Log were considered as suggestive of the presence of VBNC cells, as previously described ,8. All aP. aeruginosa C30 and AR86 genomes were analyzed by WGS , Birmingham, UK) using the Illumina MiSeq platform and a 2 × 250 bp paired-end approach. The obtained genomes  were compared to the PAO1 genome (accession number NC_002516.2) using ResFinder, MLST , the Comprehensive Antibiotic Resistance Database  and TA (toxin-antitoxin) finder .P. aeruginosa efflux pumps’ activity was measured through the ethidium bromide (EtBr) accumulation assay, performed as previously described [600 = 0.1) of P. aeruginosa PAO1-N and C30 were washed in sterile saline solution and exposed to 2.5 µM EtBr for 1.5 h. Fluorescence emission was measured in relative fluorescence units (RFUs) by a Tecan Spark® multimode microplate reader and recorded before/immediately after exposure to the dye and, subsequently, every 30 min. The signal variation was calculated as:ta is the fluorescence emitted at 0, 30, 60, and 90 min of EtBr exposure and Ft0 is the fluorescence immediately after exposure to the dye. Tests were run in technical triplicates and biological duplicates.escribed . Briefly625 = 0.3) in MHII broth, non-supplemented/supplemented with 1/4x MIC tobramycin, using the RNeasy Mini Kit (Qiagen) according to the manufacturer’s instructions. The RNA was reverse-transcribed (60 ng) using the QuantiTect Rev. transcription kit (Qiagen) according to the manufacturer’s recommendations. The mexY gene was amplified using the primers previously described [P. aeruginosa PAO1-N, grown without tobramycin (100%). The rpoS gene was used for data normalization [Total RNA was extracted from exponential phase cultures .The significance of the differences in the VBNC amounts detected in the different experimental conditions/P. aeruginosa to enter the VBNC state can be influenced by the carriage of different antibiotic resistance mechanisms. Upon exposure to tobramycin, the drug’s inactivation can prevent the formation of non-culturable cells; an active efflux might instead allow a shift to the VBNC state by modulating gene expression while hindering the drug’s ability to reach its intracellular target in a sufficient amount to inhibit protein synthesis. Further studies investigating the role of specific antibiotic resistance mechanisms to the entry into persistent and VBNC states will contribute to the understanding of the onset and behavior of persistent P. aeruginosa infections over time and to the development of more focused antibiotic treatments.In summary, the obtained results suggest that the ability of