Escherichia coli (UPEC). However, previous antibiotic therapy may impart a selective pressure that influences the population structure and pathogenic potential of infecting UPEC strains. Here, we conducted a 3-year study using whole-genome-sequencing analysis and retrospective medical record review to characterize how antibiotic exposure influenced the phenotypic antibiotic resistance, acquired resistome, virulome, and population structure of 88 UTI-causing E. coli strains from dogs. A majority of UTI-associated E. coli strains were from phylogroup B2 and clustered within sequence type 372. Previous antibiotic exposure was associated with a population shift toward UPEC from phylogroups other than the typical urovirulent phylogroup B2. The specific virulence profiles within the accessory virulome that were associated with antibiotic use were elicited by the effect of antibiotics on UPEC phylogenetic structure. Among phylogroup B2, antibiotic exposure increased the quantity of genes within the resistome and the odds of developing reduced susceptibility to at least one antibiotic. Non-B2 UPEC strains harbored a more diverse and greater resistome that conferred reduced susceptibility to multiple antibiotic classes following antibiotic exposure. Collectively, these data suggest that previous antibiotic exposure establishes an environment that provides a selective edge to non-B2 UPEC strains through their diverse and abundant antibiotic resistance genes, despite their lack of urovirulence genes. Our findings highlight the necessity for judicious use of antibiotics as we uncover another mechanism by which antibiotic exposure and resistance can influence the dynamics of bacterial infectious disease.Antibiotic therapy is the standard of care for urinary tract infections (UTIs) caused by uropathogenic IMPORTANCE Urinary tract infections (UTIs) are one of the most common infections of dogs and humans. While antibiotic therapy is the standard of care for UTIs and other infections, antibiotic exposure may influence the pathogenic profile of subsequent infections. We used whole-genome sequencing and retrospective medical record review to characterize the effect of systemic antibiotic therapy on the resistance, virulence, and population structure of 88 UTI-causing UPEC strains isolated from dogs. Our results indicate that antibiotic exposure alters the population structure of infecting UPEC strains, providing a selective edge for non-B2 phylogroups that harbor diverse and abundant resistance gene catalogues but fewer urovirulence genes. These findings highlight how antibiotic resistance can influence pathogen infection dynamics and have clinical implications for the judicious use of antibiotics for bacterial infections. Escherichia coli (UPEC), strains of E. coli that are well-adapted to colonize the urinary tract, are the most common cause of UTIs  are one of the most common bacterial infections affecting humans and companion animals , 2. Urop of UTIs . The intotential . Among tfections , 5, 6. Wfections . More renfection , 9. CollE. coli  and a history of previous UTI  (P = 0.035) as the only significant difference between the antibiotic-exposed and -unexposed groups .Of the 124 isolates from 112 different dogs, 88 (71%) unique isolates from 79 different dogs had a complete history and medical record that allowed us to retrospectively characterize their antibiotic exposure over the previous 30 days. Eight dogs submitted repeat samples, and one dog submitted three samples over the study period. All dogs had a diagnostic sample collected by cystocentesis. Forty-three isolates (48.9%) came from patients that had received at least one antibiotic in the previous 30 days. Univariate analysis showed that the dogs exposed and unexposed to antibiotics were largely the same except for differences in the presence of comorbidities  . MultivaP = 0.031) or antipsychotic/sedative (P = 0.011) compared to dogs not taking those medications. However, upon further investigation, we found that these factors confounded one another. After controlling for dogs taking analgesics at the time of diagnosis, antipsychotic or sedative use did not influence the number of virulence genes harbored by the infecting UPEC strain . Similarly, after adjusting for antipsychotic or sedative use, UPEC strains from dogs that took analgesics did not carry significantly more virulence genes . Since patient demographics and clinical factors did not alter the phylogenetics and pathogenomics of the UPEC strains, these variables were excluded from all subsequent multivariable models investigating phylogroup, virulome, resistome, and reduced-antibiotic-susceptibility outcomes.A majority of the epidemiological variables collected for dogs in this study were not associated with UPEC strain phylogroup, virulence, or resistance  to E. InCore-genome single-nucleotide polymorphism (SNP) phylogeny revealed a UPEC population structure among the dogs that was segregated into six phylogroups, with 1.1% (1/88) for phylogroup A, 14.8% (13/88) for B1, 72.7% (64/88) for phylogroup B2, 10.2% (9/88) for phylogroup D, and 1.1% (1/88) for phylogroup F . FurtherP = 0.004) of having an E. coli strain from a phylogroup other than B2 compared to dogs that did not receive antibiotics. Population structure was not influenced by sex, breed, age, comorbidity status, the use of analgesics, immunosuppressants, or sedatives/antipsychotics, or the presence or history of recurrent UTIs.Among dogs receiving antibiotic therapy in the previous 30 days, 25.6% (11/43) of UPEC strains were in phylogroup B1, 58.1% (25/43) were in B2, and 16.3% (7/43) were in D . Dogs thE. coli pathogenesis. The core virulome consisted of 91 (20.0%) virulence genes, most of which contributed to iron acquisition (ent and fep), amino acid metabolism and transport (artJ and cadA), motility , and fimbrial assembly   . The accP value of <0.1) (yfcV and ygiL), autotransporter (upaB and usp), porin (ompA and ompT), and fitness  genes associated with extraintestinal infections and UTIs. Those upregulated in the unexposed group included genes involved in secretion systems  and fimbriae  and other genes (hlyE and ehaA) that are associated with intestinal colonization and pathogenicity. Because the B2 phylogroup is associated with UTIs and other extraintestinal infections and other phylogroups are associated with intestinal colonization, we tested to see if there was a phylogroup effect among the 58 differentially abundant genes. Among those genes, 56 were significantly associated with either the B2 or the non-B2 phylogroups  . Abundanlogroups .P < 0.001) than non-B2 UPEC strains after controlling for antibiotic exposure. Contrastingly, antibiotic exposure within the previous 30 days did not significantly alter virulence gene carriage after controlling for phylogroup status   . Using t= 0.233) .z = −3.1; P = 0.0019) . Both antibiotic exposure  and being in phylogroups other than B2  increased the odds of harboring resistance genes. The resistome was greatest among phylogroup B1, with an average of 3.9 resistance genes per isolate and a range of 0 to 14.The quantity of acquired resistance genes among UPEC strains was significantly greater among dogs that were treated with antibiotics within 30 days of their UTI ( 0.0019) . UPEC isP = 0.036) and carried more resistance genes (P = 0.0359) than B2 UPEC strains not exposed to antibiotics. Contrastingly, there was no effect of antibiotic exposure on the likelihood of carrying resistance genes or the quantity of resistance genes among UPEC strains in phylogroups other than B2 (P > 0.05).The effect of previous antibiotic use on the odds of harboring a resistance gene and the number of resistance genes in the resistome were different between phylogroups. Those UPEC strains in phylogroup B2 that were exposed to antibiotics were at greater odds of carrying a resistance gene  had reduced susceptibility to at least one drug tested. The rates of reduced susceptibility were highest for ampicillin  and amoxicillin-clavulanic acid  .P = 0.002). While the phylogroup classification was not associated with reduced susceptibility among the strains tested (P = 0.249), the effect of antibiotic exposure varied within the phylogroup groupings. Antibiotic exposure was significantly associated with reduced susceptibility in phylogroup B2 (P = 0.013), while it was only marginally associated with reduced susceptibility in other phylogroups (P = 0.06)  .P = 0.020) (P = −0.011). In addition, the association between antibiotic exposure and MRS was more pronounced in UPEC strains from phylogroups other than B2 (P = 0.016).Eighteen (20.7%) of the isolates were resistant to drugs from three or more categories. Similar to reduced susceptibility, multidrug reduced susceptibility (MRS) was associated with previous antibiotic exposure  . HoweverAmong the nine dogs that submitted more than one isolate during the study, we found that most dogs were reinfected by a new UPEC strain (7/9), but two dogs were reinfected with a clonal strain . It was Among our subset of dogs with repeat isolates, we observed trends similar to those noted in our larger sample: antibiotic exposure within the previous 30 days was associated with a propensity away from phylogroup B2, increased resistance, and decreased virulence. UPEC strains isolated from repeat infections did not differ in their quantities of virulence or resistance genes or their antimicrobial susceptibilities. We did observe an inverse trend in the virulomes and resistomes among nonclonal repeat isolates, but this trend was not statistically significant.E. coli isolates from animals and explore how antibiotic exposure influences the population structure and pathogenic profiles of UTI-causing strains. Our data support our hypothesis that exposure to systemic antibiotics can alter the population structure of infecting UPEC strains away from the typical urovirulent phylogroup B2 toward other, less-well-adapted phylogroups. This observation is circumstantially supported by others who found that dogs with UTI-associated E. coli strains with common urovirulence genes were less likely to be previously exposed to antibiotics   . Our datThe UPEC community virulome was quite diverse, as only 20% of the 456 genes were present among all strains. Collectively, our data indicate that virulence gene prevalence was dictated by phylogroup association. Not surprisingly, B2 UPEC strains were enriched with typical urovirulence genes necessary for colonization of the bladder, while non-B2 UPEC strains harbored genes associated with colonization of the intestines. The types of virulence factors that cluster by phylogroup also correlate well with the evolutionary emergence of these strains, as a majority of the virulence factors required to colonize extraintestinal niches clustered within phylogroup B2, while those necessary for colonization of the intestine as pathogens or commensals clustered in later derived phylogroups , 29. At E. coli that were less virulent . A total of 124 unique isolates from 112 different dogs were collected for this study. Isolates were frozen in a 1:1 mixture of tryptic soy broth and 60% glycerol at −80° C until phenotypic and molecular characterization. This project was determined to be exempt from IACUC review because the samples were post-diagnostic specimens originally collected for another purpose.Canine The medical record for each of the 112 different dogs included in the study was reviewed for patient information including age, breed, sex/neutering status, comorbidities known to increase the risk of UTI, history of a recurrent UTI, current recurrent UTI state, which was defined using guidelines previously established in veterinary medicine, current use of antipsychotics/sedatives, analgesics, and/or immunosuppressive medication, diagnosis or suspicion of pyelonephritis, and antibiotic use within the previous 30 days . ComorbiE. coli strains ATCC 25922 and ATCC 35218 were used as quality controls. Multidrug reduced susceptibility was defined as reduced susceptibility to at least one antibiotic from three or more drug categories  broth microdilution system following Clinical and Laboratory Standards Institute (CLSI) guidelines . Isolatetegories .N50 of >99.5%, and percent protein-encoding-feature coverage of >95% were included in this study (https://cge.food.dtu.dk/services/ResFinder/) online database with default parameters from the Center for Genomic Epidemiology  (in silico Clermont phylotyping tool (http://clermontyping.iame-research.center)  with default settings (E. coli K-12 strain MG1655 (GenBank accession number LR881938) was used as the reference strain. Phylogenetic trees were visualized using the Interactive Tree of Life (iTOL) interface (https://itol.embl.de) (Isolates were further characterized into sequence types and sequence type complexes using the Achtman multilocus sequence typing (MLST) criteria via the PubMLST online database (lst.org) . Isolate.center) . A de nosettings . E. coliembl.de) .P < 0.05) in the univariate analysis, we employed a forward-selection model building approach to construct multivariate models to calculate adjusted associations and their P values   .PRJNA773999.Genome sequences were submitted to GenBank and are available under BioProject accession number


According to the test results, the incorporation of graphene oxide in appropriate admixtures could improve the mechanical properties of concrete in these two working environments. It is worth noting that this effect is most pronounced when 0.05 wt% graphene oxide is incorporated. In the sulfate attack environment, the MOEdy and UTS of the GOC0.05% specimen at 120 cycles decreased by 22.28% and 24.23%, respectively, compared with the normal concrete specimens. In the freeze–thaw environment, the MOEdy and UTS of the GOC0.05% specimen at 90 cycles decreased by 13.96% and 7.58%, respectively, compared with the normal concrete specimens. The scanning electron microscope (SEM) analysis showed that graphene oxide could adjust the aggregation state of cement hydration products and its own reaction with some cement hydration crystals to form strong covalent bonds, thereby improving and enhancing the microstructure density.In this paper, firstly, the effects of graphene oxide on the mechanical properties of concrete were investigated. Secondly, the degradation and mechanism of the mechanical properties of graphene oxide concrete (GOC) under sulfate attack and a freeze–thaw environment were investigated. In addition, the dynamic modulus of elasticity (MOE Concrete is a versatile building material used for construction over the last few decades. It is made by mixing cement, water, and other materials. On the one hand, to achieve recycling and reuse, some academics have mixed waste materials  as well as the loss of the internal alkaline environment [Cement is widely used in construction projects as the primary constituent of concrete and mortar. Nevertheless, the presence of brittle cracking defects in concrete and mortar results in a reduction in their ultimate compressive strength (UCS) and dynamic modulus of elasticity  within the internal structure of the specimens. Similarly, Cheng et al. [Sulfate attack has been identified as a prominent factor contributing to the reduced durability of concrete ,34, withg et al.  investigg et al. ; they alg et al. .There is little research on the durability of GOC with respect to sulfate attack and freeze–thaw cycles. Because graphene oxide nanomaterials have a significant impact on the long-term mechanical properties and durability of structures in construction projects, it is imperative to investigate the degradation patterns and mechanisms that affect the mechanical properties of GOC in various corrosive environments. The adoption of such an approach is of immense importance in order to obtain a thorough understanding of the performance of this novel material and its use in the construction of structures. The primary objective of this work is to investigate the effect of graphene oxide nanomaterials on the mechanical properties of concrete, with a particular focus on the degradation of the mechanical properties of GOC in two corrosive environments (sulfate attack and freeze–thaw cycle) during its service life. The influence of different levels of graphene oxide incorporation on the mechanical properties of the specimens exposed to the environment was studied using SEM analysis of the microstructure of the concrete.The specimens were meticulously prepared according to the guidelines of GB/T 50081-2019 . The dim(1)Complex Portland cementAll the indexes  of P.C. 42.5  complex Portland cement meet the requirements of GB175-2020 [(2)Multilayer graphene oxide dispersionHummers method [The multilayer graphene oxide was subjected to freezing, drying, and dispersing using the modified s method . The resThe microstructures of the graphene oxide sheet were investigated using scanning electron microscopy (SEM)  and transmission electron microscopy (TEM) , as shown in (3)Concrete aggregatesThe GOC specimens use a coarse aggregate consisting of crushed stone graded 5–20 mm, while the fine aggregate consists of mechanism sand with the specifications shown in It should be noted that the specimens were cured at a pool with a constant temperature around 20 ± 1 °C for 28 days, and were taken out 15 min before the test. The main procedure of the test in this paper is shown in The GOC specimens used in the test were prepared with a water–cement ratio of 0.50 and a sand content of 35%. Before the test, graphene oxide dispersions with different mass ratios  were first mixed with water. The concrete mixture was prepared according to the proportions shown in (1)Basic test0.02% indicates that the specimen contains 0.02 wt% graphene oxide.For the basic test, the parameter of interest was the incorporation of graphene oxide at levels of 0, 0.02, 0.05, 0.08, 0.11, 0.14, and 0.17 wt%. The curing time of the specimens was set at 28 days. The GOC(2)Sulfate attack test2SO4 solution with 10 wt% concentration. The specimens were subjected to a series of dry–wet cycles, specifically 0, 30, 60, 90, and 120 cycles, as shown in 30-GOC0.02% indicates a GOC specimen with 30 wet-dry cycles and 0.02 wt% graphene oxide.The sulfate attack test used a Na(3)Freeze–thaw cycle test30-GOC0.02% indicates a GOC specimen with 30 freeze–thaw cycles and 0.02 wt% graphene oxide.The specimens were subjected to a series of freeze–thaw cycles, specifically 0, 30, 60, and 90 cycles, as shown in dy and UCS and SEM of the specimens. The detailed steps are as follows:Once the number of cycles reached a predetermined test design, the surface moisture of the removed specimen was dried to assess surface damage.dy of the specimen. A layer of petroleum jelly was applied as a coupling medium on the test surface of the specimen, which was positioned in the center of the polystyrene plate. Then, the excitation transducer rod of the DT-20 device  was gently pressed at 1/2 of the center line of the test surface, while the receiving transducer rod was gently pressed at a distance of 5 mm from the end of the center line of the test surface. The test results obtained were calculated and processed according to Equation (1),aE is the MOEdy of the specimen (MPa); a is the cross-sectional length (mm); L is the length (mm); W is the mass (kg); and f is the vibration frequency of the fundamental frequency (Hz).The resonance method was used to measure the MOEf100 is the UCS of the test specimen of 100 mm ×100 mm × 100 mm and f150 is the UCS of the standard specimen of 150 mm × 150 mm × 150 mm.The HUT-1000 device  was used to measure the UCS of specimens. The specimen was placed in the center of the lower platen of this device and pressing was started. The loading speed was 1.2 ± 0.2 kN/s. When the load became sharply smaller and the specimen deformation sharply increased, the setup automatically controlled to stop the loading and the data were recorded when the specimen was destroyed. The test results were calculated and processed according to the following Equation (2),New fracture surfaces of the specimens were observed using the Quattro setup to investigate the effect of graphene oxide incorporation on the concrete microstructure at different cycle counts.The test was performed according to the guidelines of GB/T 50081-2019  and incldy and UCS of the GOC basically reached a peak at 0.08 wt% graphene oxide incorporation. After that, further increases in graphene oxide incorporation did not significantly improve the mechanical properties of the GOC. Then, the results of the mechanical properties of GOC with different levels of graphene oxide incorporation  were systematically investigated through sulfate attack and freeze–thaw cycle tests.The mechanical properties of GOC with different levels of graphene oxide incorporation  were investigated. It was observed that the MOE0%). Thereafter, the UCS did not show a significant further increase with the increasing incorporation of graphene oxide, with all specimens showing an increase of approximately 15% compared with the normal concrete specimen.The UCS of GOC with a curing age of 28 days showed a rapid and then slow increase with the incorporation of graphene oxide, as shown in 2), C-S-H, ettringite (AFt), monosulfate (AFm), hydrated cement particles, micropores, and water or aqueous solution filled in the micropores. To explain the obtained results, four representative groups of specimens were selected for microstructural analysis. The specimens were designated as GOC0%, GOC0.08%, GOC0.14%, and GOC0.17%, and the SEM images are shown in Hardened cement paste is a solid-liquid-gas multiphase system consisting of CH (Ca(OH)0% specimen, and it can also be observed that some cracks are distributed around the C-S-H. As shown in 0.08% specimen tends to be smaller and the number tends to be reduced, and its graphene oxide sheet and clusters inhibit the growth and diffusion of surrounding cracks to some extent [0% specimen, the cement hydration products of the GOC0.08% specimen are more regular in morphology and arrangement. With a further increase in graphene oxide incorporation (GOC0.14%), as shown in From e extent . CompareAt present, some scholars  have shoIn summary, at graphene oxide incorporation levels of 0–0.08 wt%, the graphene oxide can be well dispersed, participate in and regulate the cement hydration process, make the formation and aggregation of the cement hydration products more regular and effective, and reduce the cracks and pores inside the concrete ,48. WhenThe sulfate attack on concrete can be divided into two main categories: physical and chemical erosion . During This damage usually causes spalling, and many microcracks appear on the surface of specimens. The specimens show spalling at the edges and corners as the attack becomes more severe. As shown in dy and UCS of the GOC specimens under the sulfate attack test were both improved compared with the normal concrete specimens. With an increase in cycle number, the compressive indexes of the specimens showed a trend of “increases at first, and then decreases”, and the MOEdy and UCS of the specimens reached the maximum at 60 cycles. The MOEdy loss rate of the GOC specimens ranged from 22.282% to 28.252% , and the UCS loss rate ranged from 24.229% to 25.597%  after 120 cycles. With the increase in graphene oxide incorporation, the compressive indexes of the GOC specimens were all improved relative to normal concrete specimens, with the most significant improvement at 0.05 wt% graphene oxide incorporation, indicating that graphene oxide improved the concrete resistance to sulfate attack.As shown in To explain the results obtained in the previous section, the microstructure of specimens with different graphene oxide incorporation levels was compared and analyzed under 120 sulfate dry–wet cycles. The SEM images are shown in The results show that graphene oxide can reduce the number of products generated by sulfate attack so that the microstructure of the concrete remains relatively dense when subjected to sulfate attack. On the other hand, it improves the erosion resistance of concrete by promoting the formation of regular cement hydration products and optimizing the crystal arrangement to reduce the size and number of microcracks and pores.Freeze–thaw damage in concrete is primarily caused by the freezing of water filling the capillary pores, which causes the concrete to expand. In turn, this forces the unfrozen water from the freezing area to flow around the capillary pores, creating a large migration pressure within the concrete. If the water content in the concrete capillary pore is too high, the expansion pressure on the pore wall will be significant, creating a large tensile stress around the pore. If this tensile stress exceeds the ultimate tensile strength of the concrete, internal microcracks will be generated, causing damage to the concrete structure.0.05% is taken as an example) showed less damage compared with them.The freeze–thaw damage in concrete is usually a combination of internal erosion and surface stripping, and the change in surface appearance is the most intuitive. Concrete surface damage includes water damage, large holes, pockmarks, cracks, slagging, and spalling. As shown in dy and UCS) showed a trend of “first increase, and then decrease” as the number of cycles gradually increased, reaching a maximum at 30 freeze–thaw cycles. The compressive indexes of the GOC specimens were improved compared with the normal concrete specimens, and the compressive indexes improved more obviously with the increase in graphene oxide incorporation. After 90 freeze–thaw cycles, the MOEdy loss rate of the GOC specimens ranged from 13.960% to 19.140% , and the UCS loss rate ranged from 7.590% to 11.350% . The results showed that graphene oxide improved the frost-resisting durability of concrete.As shown in 90-GOC0% specimens under repeated freeze–thaw cycles can be observed in 90-GOC0.02% specimens, but some cracks caused by freeze–thaw damage still appeared distributed on the surface of the cement matrix. With a further increase in graphene oxide incorporation, small-scale agglomeration or clustering occurred in the FtC90-GOC0.05% specimens, as shown in 90-GOC0.08% specimen, as shown in To explain the results obtained in the previous section, the microstructure of specimens with different graphene oxide incorporation levels was compared and analyzed under 90 freeze–thaw cycles. The SEM images are shown in The results show that the number of defects in the concrete can be reduced. The microstructure is relatively dense, and strong covalent bonds with cement hydration products are formed by incorporating appropriate amounts of graphene oxide. This practice improves the performance of concrete against freeze–thaw damage and increases its freeze–thaw durability.(1)The mechanical properties of GOC specimens are higher than those of normal concrete specimens, showing a trend of “rapid and then slow increase,” in which the UCS of 0.08 wt% GOC reaches the maximum (51.86 MPa) for the first time, which increases by 14.79% compared with normal concrete specimens. SEM shows that this increase is related to the formation of strong covalent bonds.(2)dy and UTS of the SAGOC0.05% specimen under 120 sulfate attack cycles decrease by 22.28% and 24.23%, respectively, compared with the normal concrete specimen . SEM shows that graphene oxide can improve the microstructure of concrete in a sulfate attack environment, which is related to the cement matrix’s co-resistance to the degradation of the concrete’s mechanical properties due to swelling damage from corrosion products.For the sulfate attack environment, the performance of GOC shows a trend of “first increase, and then decrease” with the increase in the number of cycles. The addition of graphene oxide significantly improves the mechanical properties of concrete against sulfate attack, with the most significant improvement being achieved when 0.05 wt% graphene oxide is added. The MOE(3)dy and UTS of the FtGOC0.05% specimen under 90 freeze–thaw cycles decrease by 13.96% and 7.58%, respectively, compared with the normal concrete specimen . SEM shows that this is related to the improvement of the concrete microstructure by graphene oxide and the inhibition of crack propagation and penetration under the freeze–thaw cycle.For the freeze–thaw cycle environment, the performance of GOC shows a trend of “first increase, and then decrease” with increasing cycle times. The resistance of concrete to mechanical degradation is significantly improved by the incorporation of graphene oxide, and this improvement is most significant at 0.05 wt% graphene oxide incorporation. The MOEThis paper investigates the degradation of the mechanical properties of GOC under sulfate attack and freeze–thaw cycles. Based on the mechanical behavior of graphene oxide cement slurry and the microstructure of graphene oxide concrete slurry observed using SEM, the main conclusions are as follows:(1)The service environment of building structures is complex and variable. The experimental corrosion environment set up in this paper is only sulfate erosion or freeze–thaw cycle alone. Therefore, it is necessary to carry out a variety of environmental effects coupled erosion tests in subsequent research, such as the coupling erosion of chlorides and sulfates, the coupling of freeze–thaw and sulfate erosion, the coupling of chloride salt and freeze–thaw erosion, etc.(2)The erosion of building structures in the actual service environment is a long-term process. In this paper, only the degradation of the mechanical properties of GOC under a fixed number of corrosion times set in the laboratory is investigated. Therefore, long-term erosion tests need to be conducted in subsequent studies.(3)In this paper, only standard concrete specimens (100 mm × 100 mm × 100 mm) are used, and the mechanical properties of GOC members  need to be examined in subsequent studies.In this paper, the degradation study of the mechanical properties of GOC under a corrosive environment is still insufficient, and some problems are still to be solved, mainly in the following aspects:


The combination of these two components can be an effective strategy for controlling the perovskite crystallization process. In addition, passivation of surface defects/bulk and the engineering of energy and optoelectronic properties of perovskite-based materials can be achieved following this method. In this minireview, we summarized the existing literature reports on the structural specificity and properties of hybrid POSS perovskites.Polyhedral oligomeric silsesquioxanes (POSS) and hybrid organo-halide perovskites are two important types of hybrid nanoscale frameworks with great potential in materials chemistry. Both are currently under intensive investigation for a wide range of possible applications. Recent results suggest that POSS can be attractive passivating and structure-controlling agents for perovskite materials. In this review, we present the importance of POSS in engineering the structures of inorganic cesium-halide perovskites CsPbX While the inorganic cage of the best-known POSS is usually octahedral, the chemical structure of organic functional groups R on the silicon atoms may vary  of general the chemical composition (RSiOmay vary  1]. The. The3/2)3  are typically obtained when small organic amine molecules are used as the precursors of cationic species, whereas A2BX4 two-dimensional perovskite structures are produced when larger amine molecules are employed.Interesting results obtained in the field of materials engineering have inspired the use of polyhedral silsesquioxanes to improve the structure and properties of hybrid organo-halide perovskites. Materials based on this type of perovskite material are of great interest in the field of organic electronics, both in basic research and in a wide range of applications ,23,24,253  perovskite nanocrystals (NCs) are dominant in this field, owing to their high photoluminescence quantum yield  and narrow emission peaks, as well as their easy solution processing . Al. Al44]. 2 sorption studies corroborated the PXRD data. All the hybrid materials containing A-POSS  contained micropores smaller than 2 nm and sorption/desorption isotherms of type I were obtained. The amount of adsorbed N2 was greater for Cu-A-POSS/EA than Cu-A-POSS, and the micropore volume and BET surface increased linearly with the replacement ratio (x) of POSS with EA(44% increment if x = 0.202). When x was >20%, the micropore volume and BET surface area decreased and an increase in the fraction of nonporous layered perovskites was observed with PXRD (EA-derived layered perovskites are nonporous). On the contrary, the shrinkage of the interlayer pores was observed in the case of the layered perovskites separated by mixtures of the cubic silsesquioxanes and PEA (comparing to Cu-PEA). It implies the influence of π–π stacking of the aromatic amine molecules on the porosity of the layered perovskites. The reduction in the interlayer distance suggests that the ordering of PEA was altered in the presence of A-POSS. The effect allows for the precise structural engineering of the interlayer distance and, correspondingly, the properties of such complex systems.The observed phenomena were explained by detailed structural analysis. The PXRD patterns showed differences between the interlayer thicknesses of the perovskites obtained from EA (1.06 nm) or PEA (1.86 nm) alone and those containing a mixture of one of the amines and A-POSS (1.71 nm regardless of the type of organic amine). Increasing the amount of EA resulted in a product of bimodal structure with the layer distances of 1.06 nm (EA) and 1.71 nm (A-POSS or a mixture of A-POSS and EA), as indicated by the characteristic diffraction patterns. The results of the N2 adsorption isotherm was not altered, but the amount of N2 adsorbed (as well as the micropore volume and the BET surface area) was smaller and it even slightly decreased as the PEA fraction increased. This general trend was supported by the results obtained with the other amines. The micropore volume increased when A-POSS was partly replaced with the small organic amines  and decreased when the bulky aromatic amines (PEA or 4-phenylbutylamine) were used. Despite the changes in the structure of the silsesquioxane interlayers, the magnetic moment measurements proved that the hybrid layered perovskites retain the ferromagnetic ordering at T < 20 K. Thus, the addition of the organic amines did not cause defects in the perovskite layers, but by adjusting their type and amount, desired and interesting structural changes can be achieved.The diffraction patterns of samples containing A-POSS and PEA (<8.8%) resembled those obtained with A-POSS only . In this case, the type of N3 structure are promising and valuable emissive materials in electroluminescent devices. However, the quality of an as-prepared film of NCs may be relatively poor due to the presence of the long-chain surface ligands used during the coating process, which hampers the efficiency of the charge injection. The processing problems may lead to uneven, patchy film coverage over the device sublayers. Perovskite nanocrystals can be protected from moisture by encapsulation in a hydrophobic polymer matrix, which provides a physical barrier  were used as a surface protecting additive to the CsPbX3 (X = Br or I) nanocrystals 73 perovskite, and high output performance. Moreover, the silsesquioxane coating prevented anion exchange between the perovskite nanocrystals in the solid state, thereby increasing the stability of the mixtures of the perovskite NC powders with the different halide compositions. The distinct emission spectra of the different POSS-passivated CsPbX3 were preserved, while the uncoated NCs underwent ion exchange, resulting in a broadening of their characteristic PL signals in the solid state (The treatment provided the moisture resistant hybrid perovskite nanopowders-quantum dots (PQD) that can be used as solid state luminophores in all-perovskite white light-emitting devices. The hybrid silsesquioxane molecules acted as a hole-blocking layer between the thin coating made of the perovskite NCs and the 1,3,5-tris(N-phenylbenzimidazol-2-yl) benzene (TPBi) film that operated as the electron-transporting layer. The POSS-PQD exhibited 3. The POSS-passivated perovskites were thus used as solid state luminophores for the fabrication of all-perovskite (a single down-conversion layer) white LEDs with a CIE chromaticity coordinate of , CRI = 81, and luminous efficiency of 14.1 lm W−1. The characteristic electroluminescence spectrum 7[HS(CH2)3SiO3/2] attached onto the surface of the NCs and, at larger concentrations of CsPbX3, separated the perovskite crystals in the suspension, prohibiting the formation of larger clusters by breaking the inter-NC attachments. The presence of POSS did not influence the absorption and PL spectra of the NCs, nor altered the crystal structure and shape of the monodisperse cube-shaped CsPbBr3 nanoparticles, as evidenced by the respective XRD patterns and the TEM images  value was not high and the average PL lifetimes were reduced . Nevertheless, with the optimized POSS concentrations, higher loading of the separated NCs resulted in an increase in the overall LED brightness.The improvement in the morphology and coverage of the CsPbBr2 at 11.5 V; LE = 1.20 cd/A; EQE = 0.35%) than that obtained without the hole-blocking layer. In addition, POSS enhanced the stability of the LED devices and their operation lifetime was five times longer.It was more beneficial to use POSS as a separate layer on top of the active perovskite NC layer. In this case, the average PL decay time only decreased from 434 to 342 ns, and the average recombination rate increased by 21%. The effect of the upper POSS layer was attributed to the more efficient electron and hole recombination in the NCs zone and the blocking of hole transport between the perovskite NCs and the TPBi layers. In this case, the peak LED luminance was almost eight times higher (2983 cd/m3 NCs can be passivated through the coordination of atoms in the top layer with the carboxylate groups of (iBuSiO3/2)7[(H3C)H2C=CC(O)O(CH2)3SiO3/2] bearing methacrylic acid ligands (POSS-ME)   51]. Th. Th3 NCs8–11 nm, ), wherea, [3 NCs .3/2)7[(H3C)H2C=CC(O)O(CH2)3SiO3/2] yielded composite materials . Their diffraction patterns corresponded to that of the orthorhombic perovskite NCs crystals. The thin films cast from the dispersions of PMPOPNC in the organic solvents contained well-dispersed crystalline nanoparticles with a size of ∼12 nm, embedded within the amorphous PMMA-co-P(ME-POSS) (PMPO) matrix. No phase separation was observed. The nanoparticles of PMPOPNC were larger (average size 68 nm) than the free ME-NCs and their size distribution was wider. The PL spectra of PMPNC and PMPOPNC corroborated those of the free ME-NCs and the ME-NC/PMMA composites (λe at 516 nm). Their PLQY  was similar to that of the free ME-NCs (PLQY above 80%) and larger than the PLQY of the MENCs/PMMA blend (ca. 54%). The emission wavelength and PLQY of the PMPOPNC can be tuned by adjusting the ratio of halides in the perovskite structure CsPbX3 by using different lead halide precursors.MA-NCs can further participate in radical polymerization reactions. The copolymerization with methyl methacrylate and/or . The effect was assigned to the micro-nanorough features revealed in the SEM micrographs, caused by the migration of the POSS-containing fraction to the air-solid interface (confirmed by XPS gradient concentration data). In addition, the PMPOPNC films were not prone to anions exchange. Remarkably, PMPOPNC retained its luminescence over the three cycles of heating to 120 °C (followed cooling to room temperature) much better  than the ME-NCs, which degraded significantly (PLQY < 40%) upon the thermal treatment. The photostability of PMPOPNC under blue light was also improved  compared to the native NCs. The enhanced hydrolytic resistance, thermal stability, and good solubility in organic solvents of PMPOPNC makes it potentially suitable for use as a solution-processable luminescent ink and as a green fluorophore for the fabrication of white LEDs.3/2)7(R′SiO3/2), where R = iBu and R′ is a reactive organic residue that enables the adsorption of POSS on the surface of perovskite crystals or thin films. The passivated perovskites can be applied as water resistant light-emitting materials.As was mentioned in the previous section, apart from the employment of the POSS coated perovskite NCs as solid state luminophores, the large size of the POSS macromolecules and the presence of hydrophobic organic groups grafted to Si atoms can be an advantage with regard to their barrier action against moisture. The significant potential in this area is offered by monofunctional polyhedral silsesquioxanes of type (RSiO3 (X = Br or I) perovskite NCs by (iBuSiO3/2)7[HS(CH2)3SiO3/2] (POSS-SH) containing a thiol group resulted in an increase in the water resistance and anion exchange prevention, both in water and in the solid state  (POSS-NH2) can be applied as a capping ligand for (CH3NH3)PbBr3 (MAPbBr3) (MA—methylammonium) propyl-heptaisobutyl substituted POSS (iBuSiOmmonium) . POSS-NH2 made the surface of a thin layer of the perovskite NCs waterproof, influenced the structure of the film, and helped to tune its optoelectronic behaviour  was spin-coated as an ultra-thin passivation layer over the hole transporting layer of nickel-oxide (NOx) in a fluorine-doped tin oxide (FTO)/NOX/POSS/MAPbI3/PC61BM/Bathocuproine (BCP)/Ag perovskite solar cell of inverted structure , including amine (-NH2), phosphine (-PH2), hydroxyl (-OH), and thiol (-SH) residues 0.85(MA)0.15Pb(I3)0.85(Br3)0.15 perovskite thin films in terms of both their power conversion efficiency and their long-term stability. The conclusions drawn from the measurements of wetting angle changes and X-ray diffraction data, as well as the stability of the solar cells in the ambient atmosphere, supported the results of the DFT calculations. Three different passivation mechanisms were considered to explain the observed phenomena (2) and the iodide ions on the surface of the model perovskites. It was noted that the coordination involved electrostatic interactions between the central Cs+ ions in the perovskite structure and the partially negatively charged atoms of nitrogen and sulfur. Moreover, the secondary interactions of the agostic C-H⋯surface type also contributed to the overall energy. The second hypothesis involved the contacts between the perovskite film surface and the anionic POSS species, resulting from the chemical reactions on the MAPbI3 surface with the formation of direct Pb-N  bonds by the release of hydrogen iodide (HI). It was considered the least likely. The third (acid) hypothesis postulated the formation of a protonated, cationic POSS (POSS-NH3+/POSS-SH2+) that could replace the MA+ cations in the top layer of the MAPbI3 film, resulting in a passivated hydrophobic surface.POSS-NHhenomena . The fir−1) than the POSS-SH to the surface of the perovskite film.It was concluded that, in general, at this level of theory calculations, adsorption is significant and corresponds to the formation of a weak covalent bond, although secondary interactions may also play a role. Although all the POSS systems showed a consistent preference for the third hypothesis, the differences in the adsorption energies were similar for the 1st and 3rd approach. Both models of adsorption can possibly take place in parallel. The POSS with the amine function adsorbed stronger (with 86 (56) kJ mol3 surface that was hydrophilic (contact angle 47°) and polycrystalline  before the passivation became more hydrophobic after both the coating with POSS-NH2 (contact angle 107°) and with POSS-SH (contact angle 109°) (2 and POSS-SH (PL peaks intensities increased and were blue-shifted). The wetting angle of the perovskite film after the POSS-NH2 deposition and the subsequent washing with chlorobenzene decreased to 76° (67° in the case of the POSS-SH passivated film), but was still significantly higher than that of the pristine material. The perovskite films passivated by POSS-NH2 showed strong repulsion of the water droplet within 40 s (6 s for POSS-SH passivated sample). This shows that POSS-NH2 adsorbs more strongly on the MAPbI3 surface than POSS-SH.The contact angle investigations revealed that the MAPbIle 109°) . The sim3 and the passivated films (2 compared to the 2θ = 14.49° that corresponds to (110) crystal plane in MAPbI3 indicated the degree of moisture-induced degradation of the latter (resulting in the formation of PBI2). The increase in the intensity of the (001) peak was significantly retarded in the passivated samples. POSS-NH2 protected the perovskite film more efficiently than POSS-SH.X-ray diffraction was employed to follow the changes in the structure of the pristine MAPbIed films . The rel3-based devices. The efficiency of 16.25% was recorded for the MAPbI3-only specimen, while the passivated samples had efficiencies of around 16.01% and 15.89% for POSS-NH2 and POSS-SH, respectively. After passivation with POSS, a slight decrease in the current density (JSC) and open circuit voltage was observed. This effect was explained by the partial blocking of charge carriers at the interface by the bulky POSS molecules. Correspondingly, an increase in the transport resistance was found by the electrochemical impedance spectroscopy (EIS) measurements in the POSS-passivated devices. The increase in the recombination resistance and the reduction in the recombination loss at the perovskite-HTM interface was linked to the adsorption of POSS-NH2 on the surface of the perovskite film.Passivation with POSS had a limited effect on the photovoltaic performance, as shown by the J-V characteristics of the MAPbI3-based solar cells, based on the passivated films under ambient environment, was shown by the stable VOC and JSC values during 90 days. After that time, the solar cells made of the perovskites passivated by POSS-NH2 and POSS-SH retained, respectively, 100% and 94% of their initial JSC . The better protection offered by POSS-NH2 corroborated the stronger interactions of this compound with the perovskite film. The protective effect provided by POSS was less pronounced in the case of the solar cells based on (FA)0.85(MA)0.15Pb(I3)0.85(Br3)0.15 with formamidinium ions embedded in the perovskite structure. Passivation with POSS played a minor role in such stable systems, which already had good moisture resistance.The stability of the MAPbIap-POSS-PMMA-b-PDMAEMA, methylmethacrylate, MMA, and 2-(dimethylamino)ethylmethacrylate, DMAEMA) further applied to prepare the stable core-shell colloidal perovskite nanocrystal-polymer micelle composites (ap-POSS-PMMA-b-PDMAEMA@CsPbBr3) propyl-heptaisobutyl-POSS was used for the synthesis of an amphiphilic copolymer (CsPbBr3) . The preap-POSS-PMMA-b-PDMAEMA@CsPbBr3 was about 75% of the initial PLQY, while the CsPbBr3 NCs were fully degraded within 0.5 h under the same conditions.The NCs growth inside the inverted micelles proved to be an effective approach to engineering the morphology of the perovskite composites. As a result, multi-nanocubes of perovskite nanocrystal-polymer composites (6–8 nm size and PLQY > 60%) were obtained. It was demonstrated that the observed excited carrier dynamics was related to the decreased number of internal band-gap trap states. The hybrid composites were much more water-resistant than the bare NCs or the previously reported systems. After 5 h in water, the relative photoluminescence quantum yield of 3 NCs  make them good candidates for fluorescence resonance energy transfer (FRET) aptasensor probes, on the condition of adequate hydrolytic stability. Passivation of the perovskite NCs with functionalized POSS provides perovskite quantum dots with a hydrophobic shell, while not shielding the PL emission. For example, octaaminophenyl-POSS  MXene particles as fluorescence quenchers. The device was used for screening pathogenic bacteria (Vibrio parahaemolyticus) in seawater.The encapsulation of PQD with POSS did not change the position (519 nm) and intensity of the PL peak. The hybrid particles were hydrolytically stable and only a 10% decrease in the PL intensity was observed after the immersion of POSS-PQD into water for 24 h. POSS-PQDs-Apt were used as a signal probe to prepare a composite turn-on aptasensor containing titanium carbide (Ti3C2 MXene) was developed for the on-site detection of live Salmonella typhimurium and Vibrio parahaemolyticus in water [Analogously, a fluorescent aptasensor based on DNA aptamer-functionalized multicolor polyhedral oligomeric silsesquioxane-perovskite quantum dots (cDNA-POSS-PQDs) , both nanocrystals and thin films. Depending on the chemical structure of POSS, they can act as an interlayer component of perovskite structures  or as a passivating and structure-controlling agent of perovskite nanocrystals 7(R′SiO3/2)1 type). The literature reports show that the crystallization kinetics of perovskites can be modified by the presence of structure-directing POSS. This approach results in a reduction in the number of crystallographic defects in the perovskite-based materials and, consequently, an improvement/modification in their optoelectronic properties. In addition, POSS can enhance the hydrolytic and thermal stability of perovskite structures.Polyhedral oligomeric silsesquioxanes can be effectively used for the modification of inorganic cesium-halide perovskites CsPbX3 counterparts, seem to be a slightly more complicated solution. However, because POSS are widely sought after in materials chemistry, high-throughput, low-cost synthesis protocols for functionalized silsesquioxane cubes are constantly being developed and improved. It should also be emphasized that, usually, only very small amounts of POSS are needed to achieve an outstanding effect of changing the structure and properties of perovskites. Moreover, the use of higher concentrations is not recommended due to the insulating effect of silsesquioxane cages. These reasons allow for us to hope for a renewed interest in the use of POSS for thin-film solar cells, either in the form of nanocrystal components or passivating layers.The reason why POSS-perovskites are still somewhat underdeveloped and even slightly abandoned in this branch of materials chemistry is the fact that quite satisfactory results can be obtained by using simple trialkoxysilanes for the hydrophobization of perovskite coatings. In this context, octahedral POSS ligands, which are not as readily available as their R′Si(OR)Hybrid materials formed from the combination of POSS and perovskites appear to have an even larger potential in materials chemistry. These nanoscale structures are currently being intensively studied for a wide range of possible applications. Efficient light-emitting diodes, luminescent inks, and light absorbers in solar cells with increased lifetime are the immediate answer. However, POSS/perovskite hybrids have also been used as fluorescent probes and aptasensors that operate in situ in aqueous environments. We hope that inciting interest in the hybrid combinations of polyhedral silsesquioxanes and perovskite structures will result in embracing challenges, which will lead to new, interesting solutions in this field, going beyond the already beaten paths.


A series of fibrous meshes based on liquid crystalline polyurethane/POSS composites were prepared. Two types of polyhedral oligomeric silsesquioxanes (POSSs) of different structures were chosen to show their influence on electrospun fibers: aromatic-substituted Trisilanolphenyl POSS (TSP-POSS) and isobutyl-substituted Trisilanolisobutyl POSS (TSI-POSS) in amounts of 2 and 6 wt%. The process parameters were selected so that the obtained materials showed the highest possible fiber integrity. Moreover, 20 wt% solutions of LCPU/POSS composites in hexafluoroisopropanol (HFIP) were found to give the best processability. The morphology of the obtained meshes showed significant dependencies between the type and amount of silsesquioxane nanoparticles and fiber morphology, as well as thermal and mechanical properties. In total, 2 wt%. POSS was found to enhance the mechanical properties of produced mesh without disrupting the fiber morphology. Higher concentrations of silsesquioxanes significantly increased the fibers’ diameters and their inhomogeneity, resulting in a lower mechanical response. A calorimetric study confirmed the existence of liquid crystalline phase formation. Among all techniques used in fiber preparation, electrospinning has attracted broad interest due to its ability to form fibers with diameters down to the nanoscale. It can be applied to various types of polymers and composites by dissolving them in a suitable solvent or being applied directly from their melt . VariousDespite the fact that nano-structured fibers have been applied in many fields of research, there is still room for improvement in their processability or the creation of enhanced mechanical and thermal properties or new unique functions ,19,20. ACombining crystalline properties with anisotropic orientation makes liquid crystalline polymers a unique class of materials. Over the past few decades, such polymers have become one of the most interesting matrices for the fiber industry ,22,23. TPOSS molecules are one of the most studied nanostructures in the field of polymer composites ,28,29,30Our research is focused on the preparation of liquid crystalline polyurethane (LCPU)/POSS nanofibers using composite elastomer as a raw material. The electrospinning of polyurethane/POSS composites introduced as physical blends  and chemThe synthesis of liquid crystalline polyurethanes with POSS has already been described in ,53, alonFor the preparation of nanofibers, LCPU/POSS composites were dissolved in hexafluoroisopropanol (HFIP) to obtain 10 mL of 20 wt% solutions because, in test procedures, lower concentrations caused the formation of beads. The solutions were placed in 20 mL syringes equipped with steel needles of 1 mm in diameter before being connected to a charge Master SIMCO ION voltage source. The electrospinning process was carried out at 30 °C, with an applied voltage of 10 kV and a flow rate of 3 mL/h. The obtained fibers were collected on a rotating disc collector covered with aluminum foil with a needle-to-collector distance of 10 cm. The humidity level during the process was 55%. The process was carried out for 3 h and led to the formation of fibrous meshes, which were then dried for 48 h at 50 °C under vacuum.The morphology of the obtained fibers was studied using a JEOL InTouchScope JSM-6010LV scanning electron microscope  scanning electron microscope. In order to assess the distribution of POSS nanoparticles, an analysis of silicon distribution (mapping) was performed in selected areas of the samples using energy-dispersive X-ray spectroscopy (EDS). For the determination of the average fiber diameter, a minimum of 10 microphotographs were taken and analyzed using ImageJ software v. 1.51w .In order to determine the liquid crystalline effects, a heating and a cooling run was performed in the range of −65 to 180 °C with a rate of 10 K/min. The samples were prepared in standard aluminum pans. A single layer of the electrospun mat of approximately 0.1 mm was used for the measurement in order to minimize heat transfer lags, therefore the mass was quite low, in the order of 1 mg. The analysis was performed on a Mettler Toledo 823 Differential scanning calorimeter .The crystalline structure was evaluated with a Bruker 2D Phaser diffractometer . The measurement of the 2θ range was −0.005° to 2° at a step of 0.002°. An empty holder measurement was taken in order to record background data, which was then subtracted from the sample intensity, taking into account the transmission of the sample:θ was converted to momentum transfer The SAXS measurements were performed with a SmartLab SE instrument , an X-ray powder diffractometer equipped with a semiconductor (2D) X-ray detector Hypix 400 , and a 2.2 kW Cu LFF (Long Fine Focus)  anode lamp. The nominal 2Measurements were carried out in dynamic conditions in the temperature range of −80–80 °C at a heating rate of 2 K/min and in an inert gas atmosphere. Measurements were made for the frequencies 1; 2.5; 5; 10 Hz with target amplitude Fibrous mat samples in the form of paddles were tested according to the ISO 527-2 5A  and ISO The type and concentration of POSS in the neat composite plays an important role in fiber formation . Fibers Generally, the addition of POSS nanoparticles to the electrospun system is known to cause a decrease in fiber diameter ,57. InteTo quantify the observations described in the previous paragraph, the distribution of fiber diameters was evaluated and shown in the form of histograms in The above histograms present the fiber diameter distribution in each of the composite meshes. We can observe that the addition of the phenyl-substituted TSPPOSS does not significantly influence the dispersity in the diameter of the obtained fibers. In contrast to that, electrospinning the composite with isobutyl-substituted POSS leads to disturbance in homogeneity, particularly in the case of 6 wt% inclusion. The difference between both silsesquioxane molecules lies in the surrounding cage, which may lead to the conclusion that particles bearing aromatic rings are favorable in electrospinning processes.The scattering curve of the matrix shows a weak and broad peak of around 0.08 Å superimposed on an upturn on the low q side . The peaIn lieu of a reliable model for the study of such systems, which tend to show high dispersity in sizes of inhomogeneities, as well as a lack of well-defined shape of the scattering centers, we used the so-called broad peak function to quantify the curves at hand :(2)IbroaIn this model, A Porod term was used for the low This term is related to scattering from larger objects.  objects .The fit is excellent for all the materials under investigation . The cald0 increases. In addition, modification via isobutyl-substituted POSS (TSI POSS) increases the exponent m. This is a result that is not easy to interpret. Ballard et al. [The Porod model describes scattering from larger objects. Here, it is most likely the fiber itself. Essentially, what we see is the surface of the structure. For smooth surfaces ry rough . TSIPOSSd et al.  associatTo investigate the crystalline structure and formation of possible POSS-rich regions, XRD studies were performed. The diffractograms of the samples and neat silsesquioxanes are presented in θ of 25°, a sharper peak can be observed, which corresponds to the crystalline structure form of a mesogenic unit, which was discussed in our previous works [The pattern of the matrix is mainly amorphous, with the main halo at 22°. Around 2us works ,53. The The formation of liquid crystalline phases was investigated via differential scanning calorimetry. The DSC curves are presented in The calorimetric study shows that the obtained fibrous composites have a complex behavior. In the heating run A, the fiIt can be hypothesized that when a liquid crystalline elastomer is electrospun, crystalline to liquid crystalline phase transition requires less heat to pass through LC phases, possibly due to smaller domains. At higher temperatures, above 120 °C, weaker and broader melting effects can be observed. Interestingly, for a starting material in the heating mode, only a single melting was observed, which can lead to the conclusion that when an LC elastomer is electrospun, more than one type of crystalline or liquid-crystalline domain is formed. Perhaps fast solvent evaporation during electrospinning causes the formation of smaller domains because they do not have as much time to organize themselves as they do during slow evaporation in the elastomer synthesis process .In the cooling run B, a fainInterestingly, there is no correlation of peak temperature with long period as is known for lamellar crystals or in polyurethanes . It is pAll composite meshes show a modulus higher than that for neat material . The glaReinforcement of the modulus after the incorporation of POSS is a well-known phenomenon on the nanoscale in polyurethanes ,69,70. TgT in the vicinity of 0 °C [gT upon the addition of phenyl-substituted POSS, if we assume that the dominant mechanism is the third one, i.e., the participation of diluted POSS in the glass transition.The glass transition temperature, as quantified by the peak temperature of tanδ curves, increases by several degrees with increasing POSS content. The effect is more pronounced for the phenyl-substituted moiety. Such an increase in the glass transition temperature is associated with a slowing of dynamics. In polyurethane POSS systems, several mechanisms may give rise to this effect: (i) the restriction of dynamics due to crosslinking; (ii) morphological changes, that is, decrease in microphase separation; (iii) restriction of dynamics due to the participation of POSS in the glass transition, in the sense of mixing models, such as Fox; and (iv) anchoring of polymer chains on nanoaggregates . Accordi of 0 °C . This obThe measurement of tensile strength shows the reinforcement effect of both types of POSS on the LCPU matrix .We found that 2 wt% POSS loading in composites leads to a significant increase in tensile strength at break by more than 500%, along with increased elongation at break. Interestingly, when 6 wt% of POSS materials are electrospun, the resulting meshes exhibit much lower tensile strength—lower than that for neat samples. This should be correlated with the higher degree of ordering in the 2 wt% materials, as observed via DSC. Moreover, the mesoscopic morphology of the fibrous mat may play a role . It is hFibrous mats were prepared via electrospinning hybrid organic–inorganic liquid crystalline polyurethanes (LCPUs) modified using POSS as crosslinking agents. It was possible to obtain fibers with diameters in the sub-micrometer range. The spatial distribution of POSS moieties within the fibers was high, despite the occurrence of rare aggregates with sizes in the order of micrometers. Crosslinking with POSS resulted in somewhat thicker and more disperse fibers. The type of organic vertex groups of POSS moieties played an important role in this. That is, the isobutyl-substituted moieties had only a small effect, while the phenyl-substituted POSS caused a much more pronounced one, resulting in a content of 6 wt% in a very broad distribution of diameters in the order of μm.This should be correlated with the internal micromorphology of the fibers. Although SAXS detected no specific POSS-related inhomogeneities at the nanoscale, the microphase separation changed significantly. Isobutyl-substituted POSS caused larger and less distanced mesogen-related inhomogeneities than the matrix, and phenyl-substituted ones resulted in inhomogeneities of similar size as those of the matrix but significantly more distanced from each other. The surface of the fibers was found to be rather rough, but POSS had no effect on that. On the atomic scale, there was only very minor indication of POSS crystallinity, despite the presence of some weak and broad peaks, possibly related to the micrometric aggregates observed via scanning electron microscopy.The glass transition temperature increased with crosslinking, with the benzene-substituted POSS having a more pronounced effect. This is due to the combined effect of deceleration of dynamics due to crosslinking, which is presumably the same in both types of composites, and changes in microphase separation, which are more pronounced in the case of the phenyl-substituted POSS. The slower mobility of the POSSs themselves possibly plays a role too.The mechanical modulus and elongation at break also increased with POSS incorporation, but interestingly, the effect is more pronounced when a smaller amount of POSS is introduced into the system.


IgG4-related disease (IgG4-RD) is a newly recognised fibro-inflammatory clinical entity with multiorgan involvement. Although most patients with IgG4-RD respond well to glucocorticoids, they have limited treatment options for disease control and high relapse rates were reported during long-term treatment.10.1136/ard-2022-223529.supp1Supplementary dataA 54-year-old man, who was diagnosed with IgG4-RD in 2017, presented in our department with re-enlarged bilateral submandibular glands, parotid glands and 12 lymph nodes after withdrawal of glucocorticoids . His 2019 American College of Rheumatology/European League Against Rheumatism classification criteria (ACR/EULAR)+CD24-CD38hi plasmablast levels during the 24 weeks of follow-up (+ T, CD8+ T and B cell counts contributed to the status of ‘non-remission’ (Besides the aforementioned case, based on the criteria listed in ollow-up , while nollow-up . Howeverollow-up . Analysemission’ . This reTo the best of our knowledge, this is the first study reporting the therapeutic potential of BLyS/APRIL-targeting biologics in IgG4-RD. Patients with IgG4-RD patients have limited treatment options, especially for those who were unable or unwilling to use long-term glucocorticoid therapy due to various conditions. Here, we reported a 60% partial remission rate at week 24, a duration widely adopted for the observation of IgG4-RD remission


Colorectal cancer (CRC) is the most common malignancy in the world, and novel molecular targeted therapies for CRC have been vigorously pursued. We searched for novel combination therapies based on the expression patterns of membrane proteins in CRC cell lines.A positive correlation was observed between the expression of human pidermal growth factor receptor (HER) 3 and mesenchymal‐to‐epithelial transition factor (MET) on the cell surface of CRC cell lines. The brief stimulation of HER3/MET‐high SW1116 CRC cells with both neuregulin‐1 (NRG1) and hepatocyte growth factor enhanced ERK phosphorylation and cell proliferation more than each stimulation alone. In addition, a prolonged NRG1 stimulation resulted in the tyrosine phosphorylation of MET. In this context, the Forkhead Box protein M1 (FOXM1)‐regulated tyrosine phosphorylation of MET by NRG1 was demonstrated, suggesting the existence of a signaling pathway mediated by FOXM1 upon the NRG1 stimulation. Since the co‐expression of HER3 and MET was also demonstrated in in vivo CRC tissues by immunohistochemistry, we investigated whether the co‐inhibition of HER3 and MET could be an effective therapy for CRC. We established HER3‐and/or MET‐KO SW1116 cell lines, and HER3/MET‐double KO resulted in the inhibition of in vitro cell proliferation and in vivo tumor growth in nude mice by SW1116 cells. Furthermore, the combination of patritumab, an anti‐HER3 fully human mAb, and PHA665752, a MET inhibitor, markedly inhibited in vitro cell proliferation, 3D‐colony formation, and in vivo tumor growth in nude mice by SW1116 cellsThe dual targeting of HER3/MET has potential as CRC therapy. The present study demonstrated that HER3 and MET were not directly associated with each other in CRC but cooperated for the cellular and tumor growth of CRC. In this context, the combination of anti‐HER3 (patritumab) and anti‐MET (PHA665752) drugs was definitely effective against HER3/MET‐high SW1116 CRC cells, indicating the potential of the dual inhibition of HER3 and MET as targeting therapy in human CRC. The activation of HER family proteins stimulates intracellular events, such as the MAPK and PI3K‐AKT pathways,Colorectal cancer (CRC) is one of the leading causes of human cancer death worldwide.The phosphor‐transfer reaction of HER3 is catalytically impaired, resulting in very weak kinase activity; however, HER3 may be activated by forming heterodimers with other HER family receptors. By the binding to HER3 ligands, like neuregulin 1 (NRG1), HER3 plays important roles, such as in the differentiation and proliferation of normal and cancer cells.Mesenchymal‐to‐epithelial transition factor (MET or c‐MET) is a receptor‐type tyrosine kinase that is activated by the binding of hepatocyte growth factor (HGF). The HGF/MET pathway is essential in cell proliferation, motility, and the development of resistance to cancer therapy.Growing evidence has suggested that the presence of different cell populations within a single tumor may limit the effectiveness of therapies targeting a single molecule.In the present study, we demonstrate that the cell surface expression of HER3 and MET proteins positively correlates in human CRC, the NRG1‐stimulated tyrosine phosphorylation of MET is regulated by the transcription regulator, Forkhead Box protein M1 (FOXM1), and the co‐inhibition of HER3 and MET represents an effective therapy for human CRC.22.1Female F344 rats and Male KSN nude mice were obtained from Shimizu Animal Farm . Animals were individually raised in plastic cages and maintained under specific pathogen‐free conditions and housed in under a standard light/dark cycle at a constant temperature of 23 ± 1°C. Experiments were approved by the Committee for the Care and Use of Laboratory Animals at Kindai University (KAPS‐23‐004) and performed following the institutional guidelines and the United States National Institutes of Health Guide for the Care and Use of Laboratory Animals.2.22) at 37°C. Aseptic processing was strictly controlled by air purifiers .The following 23 human CRC cells , and P3X63Ag8.653 mouse myeloma cells were obtained from the American Type Cell Collection . CCD841 non‐neoplastic colon cell line was obtained from the Japanese Collection of Research Bioresources Cell Bank. The RH7777 rat hepatoma cell line was donated by Dr. Chiba K (Mitsubishi Tanabe Pharma). These cells and human HEK293F (Invitrogen) were cultured in RD medium2.3−c transporter/xCT (Ab 2–31 and Ab2‐25), epithelial cell adhesion molecule/EpCAM (1D12), standard CD44/CD44s (RV7), variant CD44/CD44v8 (RV14), CD44v9 (RV3), and CD98 heavy chain  of human cells were used. Regarding HER3,All rat mAb used in the present study were produced by Masuko T, et al.2.45) were mixed with the primary rat mAb (10 μg/mL) on ice for 1 h and then incubated with 50 μL of the phycoerithrin (PE)‐labeled donkey anti‐rat IgG (H+L) secondary antibody  at 4°C for 1 h. Cells were washed and suspended in 0.2% BSA‐PBS and filtered through a nylon mesh . BD LSR Fortessa (Becton Dickinson) were used to perform FCM. The subtracted mean fluorescence intensity (ΔMFI) or signal‐to‐noise ratio (S/N) was calculated to quantify cell‐surface protein expression.Cells  at room temperature for 2 h. They were then incubated with Ab4 anti‐HER3 (10 μg/mL) or Ab57 anti‐MET mAb (10 μg/mL) primary rat mAb diluted in 1% BSA‐PBS at 4°C overnight. After washing, sections were incubated with biotinylated goat anti‐rat IgG (Vector Laboratories) diluted 1:200 in 2% BSA‐PBS at room temperature for 30 min. After washing, samples were treated with avidin‐biotin‐peroxidase complex (ABC) reagent (Vector Laboratories) diluted 1:50 in 2% BSA‐PBS at room temperature for 30 min. After washing, sections were incubated with 0.05% 3,3′‐diaminobenzidine  and 0.01% H2O2 in 0.1 M Tris–HCl (pH 7.4), and then counterstained with hematoxylin. Sections were dehydrated, cleared in xylene, and mounted in Permount (Fisher Scientific). IHC scores for HER3 or MET expression were set based on the standard HER2 test. IHC gave a score of 0–3, indicating the amount of HER3/MET in tissue samples. HER3/MET expression in normal (adjacent to or distant from CRC) and cancer tissues was estimated as score 0 (negative), score 1 (weak or borderline), score 2 (intermediate), or score 3 (strong) by two pathologists.Colon adenocarcinoma with a matched adjacent colon tissue array  was used. Following deparaffinization and rehydration, tissue sections were treated with solution  at 98°C for 5 min 3 times by a microwave to retrieve the antigen. After three washes with PBS (“washing” hereafter), these sections were incubated with 3% H2.6g for 1 min. Beads were washed 3 times with lysis buffer and once with IP wash buffer (50 mM Tris (pH 8.0)). SDS sample buffer  was added to the beads and heat treated at 95°C for 4 min. The supernatant, separated by centrifugation at 10000 × g for 1 min was used for Western blot (WB).Cells were solubilized by lysis buffer , 1% Nonidet P‐40), and protease inhibitor cocktail  at 4°C for 20 min. Cell lysates were reacted with anti‐MET or anti‐HER2 mAb (10 μg/mL) at 4°C overnight. Antibody‐bound proteins were mixed with Protein G Sepharose 4 Fast Flow  for 2 h, and beads were collected by centrifugation at 9000 × 2.7Whole cell extracts were prepared using lysate buffer (50 mM Tris–HCl (pH 7.4), 150 mM NaCl, 1% NP40, and 0.1% SDS) with a Protease and Phosphatase Inhibitor Cocktail . Protein concentrations were measured using the BCA assay . Samples were diluted in 5 × SDS buffer  and treated at 95°C for 5 min. Proteins (20 μg/lane) were separated by 8 ~ 10% SDS‐PAGE and transferred onto polyvinylidene fluoride (PVDF) membranes. Non‐specific binding sites on PVDF membranes were blocked with 5% Skim milk (Yukijirushi) for 1 h. Membranes were incubated with antibodies against HER3, MET, phospho‐HER3, phospho‐MET, phospho‐ERK, ERK, FOXM1, and GAPDH (CST) for 16 h, and then with horseradish peroxidase‐labeled goat anti‐rabbit or mouse IgG (H+L) pAb . Immunoreactive proteins were visualized by an ECL system  using Chemi‐Lumi One Super  in Kindai University, and by exposing X‐ray film  using a detection solution  or by an ECL system  in Tohoku University.2.8https://crispr.dbcls.jp/). SW1116 cells were used to generate HER3 and/or MET and the FOXM1‐KO cell line using the pX330 (Addgene) and pCAG‐EG × ×FP (Addgene) CRISPR/Cas9 vectors. The gene‐specific region of gRNA sequences was designed by the CRISPR design tool from CRISPR direct (https:/crispr.dbcls.jp/). Single clones were picked up and the KO efficiency was assessed by WB and FCM. Cells were seeded onto 35‐mm dishes  in 1 mL of RD medium, and plasmid DNA (5 μg) was introduced into cells of approximately 80% confluency using Xfect transfection reagent (Takara Bio Inc.). The co‐transfection of pX330 and pUC19  containing the puromycin‐resistant gene was also performed, and cells were cultured with puromycin  for 10 days.KO was performed as recently described2.9The cell cycle distribution was analyzed by FCM. Briefly, cells were incubated for 96 h, washed with cold PBS, fixed in 70% ethanol overnight at −20°C, and stained with DAPI  reagent at 4°C for 15 min. The fluorescence intensity of the final mixture was analyzed by BD LSR Fortessa.2.10Cell proliferation was assessed by WST‐8 . After cells were cultured in 96‐well plates, WST‐8 (10 μL /well) was added to each well at different time points and incubated at 37°C for 2 h. Absorbance (measurement: 450 nm/reference: 600 nm) was measured using a fluorescence spectrophotometer .2.116 cells) in 0.2 mL of PBS were s.c. injected into the flank of KSN mice, and visible tumors were confirmed in all mice. At this point (Day 1), patritumab, PHA665752, the combination of patritumab and PHA665752, or PBS as a control was i.p. injected, followed by three additional injections on Days 7, 13, and 19. WT, HER3‐KO, MET‐KO, and HER3/MET‐double KO (dKO) SW1116 (5 × 106 cells/mouse) in 0.2 mL of PBS were s.c. injected into the flank and visible tumors were confirmed in all mice. Tumor volumes were measured with a digital caliper every 3 days and quantified using the following formula: volume [mm3] = (length [mm]) × (width [mm])2 × 0.5.SW1116 (5 × 102.12Matrigel® (CORNING®) was added at a volume of 150 μL to 8‐well chamber slides (WATSON Bio Lab) and incubated at 37°C for 15 min, after which 5000 cells were seeded. Patritumab and the MET inhibitor were added the next day, and after a 4‐day incubation, 50 cells were measured as one colony after imaging with BIOZERO (KEYENCE) in nine fields of view/one well.2.132 values and subtracted from the median. A clustered heat map analysis and PCA were conducted with the R Project for Statistical Computing (https://www.r‐project.org/). Data for correlations were analyzed using Pearson's correlation coefficient. Experimental data were analyzed using Prism 5 for Win (GraphPad Software). The criterion for significance was at least.Regarding the expression levels of cell surface proteins, S/N or ΔMFI was calculated. The results obtained were transformed into logp < 0 0.05. Combined effects by mAb or inhibitors were considered to be additive (#) or synergistic (##), as described elsewhere.33.1To identify targets for simultaneous inhibition, we initially examined the expression of cancer‐associated cell‐surface proteins in 24 human colon cell lines and analyzed the expression levels of 14 cell‐surface proteins with specific mAb by FCM. Protein /cell  types were classified by the clustered heatmap Figure . Cell li3.2We examined the effects of NRG1 and HGF on the phosphorylation of HER3 and MET and on the proliferation of SW1116 cells. A brief ligand stimulation resulted in the phosphorylation of the receptor for each ligand Figure . After a3.3To analyze potential crosstalk between HER3 and MET, we initially examined MET activation by NRG1 in SW1116 cells. NRG1 induces the formation of the HER2/HER3 heterodimer and the activation of HER3. We confirmed the formation of HER2/HER3 heterodimers and demonstrated MET phosphorylation by the NRG1 stimulation for 12 h Figure . MET pho3.4To analyze the effects of HER3‐ and/or MET‐KO on cell growth, we established HER3 and/or MET‐KO SW1116 cells. Using FCM and WB analyses, we confirmed that HER3 proteins disappeared in both HER3‐KO and HER3/MET‐double KO (dKO) cells, whereas MET proteins disappeared in both MET‐KO and HER3/MET‐dKO cells Figure . ERK pho3.5To examine the efficacy of the combined treatment of anti‐HER3 and anti‐MET drugs against human CRC cells, we evaluated the effects of patritumab and PHA665752 against SW1116 cells using in vitro colony formation in 3D‐culture and in vivo tumor growth in nude mice. The combined treatment of patritumab and PHA665752 resulted in the significantly stronger inhibition of colony formation Figure  and tumo4Due to the heterogeneity of cancer cells or cancer tissues, single‐agent therapy often resulted in limited efficacyr) of HER2 versus HER3 = 0.711 (data not shown). A positive correlation between the expression of HER3 and MET , but not in 25 breast and 22 lung cancer cell lines  (data not shown).In the present study, we analyzed the expression of 14 cancer‐associated proteins in 24 CRC cell lines by FCM. A positive correlation was reported between the expression of HER2 and HER3 in primary colorectal cancer.T Figure  was noteRegarding the effects of HER3 and MET on cancer cell proliferation, the combined stimulation of both ligands enhanced cell proliferation more than either ligand alone Figure , suggestEffects of NRG1 and HGF on colon cancer cell proliferation shown in Figures Contrary to expectations, MET‐KO promoted in vitro cell proliferation and in vivo tumor growth of SW1116. Although a marked increase in the percentage of cells in the S phase may be involved in these phenomena, the underlying mechanisms remain unclear. In HER3/MET‐dKO SW1116 cells, in vitro cell proliferation and in vivo tumor growth were significantly inhibited and G0/G1 arrest was induced, indicating that simultaneous HER3/MET targeting was effective Figure . Since FWe showed that HER3 and MET indirectly affected each other in the presence of each ligand; however, essential or ultimate effectors transcriptionally regulated by FOXM1 that are responsible for the NRG1‐induced phosphorylation of MET and anti‐tumor effects by the dual targeting of HER3/MET remains to be solved. As to possible mechanism of “NRG1‐stimulated and FOXM1‐mediated MET phosphorylation,” we remark cell‐surface expression of MET. Total MET expression seems not altered in FOXM1 induction in Figure In recent years, the combination therapy has been attracting attention, because of the possible additive or synergistic potential for effective therapeutic effects through independent mechanisms of action.The present results demonstrated that HER3 and MET were not directly associated with each other in CRC but cooperated for the cellular and tumor growth of CRC Figure . The comAkitaka Yamasaki: Investigation (lead); methodology (lead); writing – original draft . Rikuto Miyake: Investigation (supporting); methodology (supporting). Yuta Hara: Formal analysis (supporting); investigation (supporting); methodology (supporting). Hideki Okuno: Investigation (supporting); methodology (supporting). Takuya Imaida: Investigation (supporting); methodology (supporting). Kouki Okita: Investigation (supporting); methodology (supporting). Shogo Okazaki: Formal analysis (supporting); investigation (supporting). Yasutoshi Akiyama: Investigation (supporting); validation (supporting). Kenji Hirotani: Methodology (supporting). Yuichi Endo: Investigation (supporting); validation (supporting). Kazue Masuko: Investigation (supporting); methodology (supporting); validation (supporting). Takashi Masuko: Conceptualization (lead); funding acquisition (lead); investigation (supporting); methodology (supporting); writing – original draft ; writing – review and editing (lead). Yoshihisa Tomioka: Conceptualization ; investigation (supporting); supervision (supporting).This work was supported by MEXT/JSPS KAKENHI Grant Number 18H05463 .Masuko T and Endo Y were supported by commissioned research fund to Kindai University  from HEALIOS K.K. (Japan). The other authors declare that they have no conflicts of interest.This study was approved by the Institutional Review Board of the Pharmaceutical department , Kindai University.Figure S1:Click here for additional data file.Figure S2:Click here for additional data file.Figure S3:Click here for additional data file.Figure S4:Click here for additional data file.Figure S5:Click here for additional data file.


ObjectiveThe purpose of this study is to evaluate the antibiotic prescription patterns of endodontists, general dentists, and other dental specialists for endodontic procedures in India by using the knowledge, attitude, and practices (KAP) survey method.MethodsThis cross-sectional study was carried out from February 2022 to May 2022 and involved dentists across India. A self-made questionnaire survey was created to judge the knowledge of various dental practitioners, which includes general dentists, endodontists, other dental specialists, and post-graduates with regard to antibiotic usage guidelines for endodontic purposes. A total of about 310 dental practitioners were surveyed across India. The questionnaire was circulated via social platforms such as WhatsApp, Instagram, and Facebook Messenger.Statistical analysisData for KAP regarding antibiotic prescription patterns among general dentists, endodontists, other dental specialists, and postgraduates were entered into Microsoft Excel and analyzed using IBM SPSS Statistics for Windows, Version 20 . Descriptive statistics of the study population were examined. The level of statistical significance was determined at p<0.05.ResultsFor the percentage of patients who were prescribed systemic antibiotics every day for endodontic reasons, about 38.6% (119) responded 0-10% and 27.3% (84) responded 10-30%. For the order of antibiotics that they prefer from most to least, about 85.4% (263) responded amoxicillin > metronidazole > doxycycline > azithromycin > clindamycin > ciprofloxacin. For the question of whether they use local antibiotics, about 35% answered yes of which 25% were endodontists, 2% were general dentists, 5% were other dental specialists, and 3% were post-graduates. About 77.3% of the total participants were unaware of the antimicrobial stewardship concept and AwaRe classification from WHO. About 53.2% (164) attended CDE programs with regard to antibiotic usage.ConclusionIt is evident from the results of the present study that there is over-prescription of antibiotics by practitioners especially by general dentists without following proper guidelines for endodontic treatments. More emphasis should be made on the proper prescription pattern of antibiotics, proper understanding of endodontic diagnosis, and the need for antibiotics at the undergraduate level. In addition, proper awareness, as well as proper prescription of antibiotics, should be made for existing dental professionals. Antibiotics were introduced in the late 1920s. These life-saving medicines should be used appropriately, or they would result in the development of resistance. Unfortunately, antibiotic over-prescription has resulted in the development of newer strains of bacteria that are resistant to these antibiotics. A large number of antibiotics that were effective in the past are not prescribed nowadays because of the development of resistance [Agnihotry et al. 2019) stated that the prescription of systemic antibiotics has become a common part of dental practice and has increased to a greater extent in the last two decades. Dental antibiotic prescription accounts for about 7-10% of the total antibiotic that has been prescribed worldwide for other medical reasons 019 state. The oveClinical situations that require antibiotic therapy include oral infections in medically compromised patients and those with systemic involvement such as malaise, elevated body temperature, trismus, lymphadenopathy, and progressive and persistent infections. Endodontic conditions, such as symptomatic reversible and irreversible pulpitis, pulpal necrosis, acute apical periodontitis, acute apical abscess with no systemic involvement, and chronic apical abscess, do not require antibiotic coverage .The primary cause of pulpal and periapical pathosis is micro-organisms . SystemiAbuse and irrational prescription are the main contributing factors to the emergence of antimicrobial resistance (AMR). Hence, the primary goal of this knowledge, attitude, and practices (KAP) survey is to assess the antibiotic prescription pattern among general dentists, endodontists, other dental specialty persons, and post-graduates in India for endodontic procedures.The null hypothesis is that there is no difference in the KAP regarding antibiotic prescription patterns among endodontists and other dentists.The research hypothesis is that there is a difference in the KAP regarding antibiotic prescription patterns among endodontists and other dentists.This cross-sectional study was carried out from February 2022 to May 2022 and involved dentists across India. Ethical approval was obtained from the Institutional Human Ethics Committee registered with the Central Drugs Standard Control Organisation (reg. no.: ECR/212/Inst/TN/2013/RR-19) of Chettinad Academy of Research and Education . The purpose of this study is to evaluate dental professionals' KAP about the usage of antibiotics for endodontic reasons. The current study's inclusion criteria include general dentists, endodontists, other dental specialists, and post-graduates of all dental specialties in India. In addition, dental professionals who perform endodontic procedures in their day-to-day practice were only included in the survey. Practitioners who refused to take part in the study and those who do not perform endodontic procedures were excluded.A self-made questionnaire survey was created to judge the knowledge of various dental practitioners with regard to antibiotic usage guidelines for endodontic purposes. The questionnaire consisted of two parts: Part A contained demographic details and Part B contained the questionnaire proper with 16 questions regarding endodontic practice. They were presented as follows: six multiple-choice questions, one open-ended question, and nine yes/ no questions.The sample size of the current study was estimated using statistical power analysis G*Power software (version 3.1.9.2) and considering χ² tests, goodness-of-fit tests, contingency tables, and other input parameters:1. α error as 0.05 at 95% CI2. β error as 0.203. Power of the test (1-β error) as 80%4. Degrees of freedom as 45. Effect size (Cohen’s w statistic) as 0.1880 )The required total sample size for the current study was 338 participants with an output actual power of 80.04%. The questionnaire was circulated based on 338 participants, but only 310 participants responded to the questionnaire, thereby having a non-response rate of 8.87%. The questionnaire was circulated via social platforms like WhatsApp, Instagram, and Facebook Messenger.Data regarding KAP regarding antibiotic prescription patterns among general dentists, endodontists, other dental specialists, and postgraduates were entered into Microsoft Excel and analyzed using IBM SPSS Statistics for Windows, Version 20. The descriptive statistics of the study population were examined. The relationship between the designation and years of experience and KAP regarding antibiotic usage was analyzed using the Chi-square test. The level of statistical significance was determined at p<0.05.In this study, among 308 dentists who answered the questionnaire survey, 32.5% 100) were general dentists, 24.4% were from other specialty PGs, 20.8% were endodontic PGs, 13% were endodontists, and only 9.4% were other dental specialists of which 2% were pedodontists, 2% were prosthodontists, 1.2% were orthodontists, 1.2% were oral surgeons, 1% were oral pathologists, 1% were periodontists, and the remaining 1% were public health dentists. In addition, 77.6% (239) had less than five years of clinical experience, 14.6% had 5-15 years of clinical experience, and only 7.8% had more than 15 years of clinical experience  responded 0-5, and 35.7% (110) responded 6-10. For the percentage of patients who were prescribed systemic antibiotics every day for endodontic reasons, about 38.6% (119) responded 0-10%, and 27.3% (84) responded 10-30%. For the question conditions for which antibiotics were being prescribed, about 15.9% (49) responded dentoalveolar abscess, replantation after avulsion, and 84.1% (259) responded pain relief, reversible pulpitis, irreversible pulpitis, and endodontic flareups. For the most commonly prescribed antibiotics, about 87%(268) responded amoxicillin, and 11% (34) responded metronidazole. In the clinical situation for which they prescribe antibiotics, about 41.9% (129) responded based on radiological findings like periapical lesions and 25%(77) responded that they would prescribe in the presence of systemic disorders like diabetes mellitus, hypertension, and cardiac conditions. For the order of antibiotics they would prefer from most to least, about 85.4% (263) responded amoxicillin > metronidazole > doxycycline > azithromycin > clindamycin > ciprofloxacin. For the question of whether they use local antibiotics, about 35% answered yes of which 25% were endodontists, 2% were general dentists, 5% were other dental specialists, and 3% were post-graduates.Regarding the prophylactic prescription of antibiotics prior to apical surgery, 76.6% (236) responded "yes," and 84.7% (261) responded "no." For the question of whether they advise antibiotic culture tests for their patients, 61.4% (189) responded that their patients had self-prescribed antibiotics, and 50% responded that their patients haven’t responded to the prescribed antibiotics. With regard to the use of a combination of antibiotics for synergistic effects, 70.1% (216) responded "yes," 67.5% (208) prescribed drugs based on drug dosage formula, half-life, and weight of the patient, and 71.8% (221) upgraded themselves with the new guidelines and updates regarding antibiotics prescription patterns. About 77.3% of the total participants were unaware of the AMS concept and AwaRe classification from WHO. About 53.2% (164) have attended CDE programs with regard to antibiotic usage.A recent study by the American Association of Endodontists (AAE) states that general dentists perform the majority of root canal treatments when compared with endodontists. Studies have reported that about 10 million deaths would occur per year globally by 2050, which makes drug resistance a curse of the medical profession unless sustained actions are not taken .Various contributing factors for AMR include inappropriate antibiotic prescription by medical or dental practitioners . Once reThe British Society for Antimicrobial Chemotherapy states that improper antibacterial drug prescription by dental practitioners is a major contributing factor in the development of drug-resistant strains. Factors such as inappropriate dosing, duration, and prophylaxis may contribute to the development of resistant strains. Therefore, it is essential to evaluate the prescription pattern in India to know whether there is a need for continuous education on antibiotic prescription patterns for dentists .AMR has been acknowledged as a global danger to public health by the World Health Assembly's acceptance of the global action plan on AMR in May 2015 and the political statement of the high-level meeting of the general assembly on AMR in September 2017 .This study shows that though a large number of respondents 38.6% (119) found the requirement to prescribe antibiotics was only 0-10% of the endodontic patients who have been examined every day, there were also 27.3% (84) who prescribed antibiotics in the range of 10-30% and 19.5% (60) who prescribed antibiotics between 30% and 50%, which may contribute to over-prescription of antibiotic to some extent. In addition, the majority of general dentists prescribe antibiotics for 10-30% of endodontic cases they handle in their day-to-day practice. This may be due to anxiety about pain development, preventing endodontic flareups, improving patient comfort, and the lack of awareness of proper antibiotic prescription patterns.According to a systematic review by James et al., antibiotics were not essential for irreversible pulpitis and pain relief ,14. In lAbout 87% of participants chose Amoxicillin as the first line of drug, followed by metronidazole (11%). This is consistent with the study done by Maslamani et al. . AmoxiciThe 2017 AAE guidelines regarding antibiotic prophylaxis given recommend prescribing antibiotics for diabetic patients with poor glycemic control . The ESEStudies conducted by Amisha et al. 2020 and Singh et al. found that <40% of dentists have advised antibiotic culture test, and in this study, only 15.3% of dentists have advised it ,22. FurtA combination of antibiotics is prescribed by more than 70% of dentists. Amoxicillin + clavulanic acid is one such combination that may be used for serious oral infections and in cases where resistant species might be suspected, which is refractory to normal endodontic procedures .The antimicrobial stewardship (AMS) concept was introduced by WHO to take care of the health and well-being of the people and provide guidelines for health systems both nationally and globally. The three pillars that strengthen the health systems are AMS, medicine and patient safety, and infection prevention and control. AMR shall be controlled with the help of AMS, antibiotic surveillance, the AWaRe classification, and the essential medicine list (EMS) from WHO . In AWaRIn this study, about 77% of dentists were unaware of the AMS concept and AWaRe classification from WHO, and this clearly states the reason for the overuse or misuse of drugs. In addition, 53.2% of dentists have not regularly attended CDE programs with regard to antibiotic resistance.The endodontic flareups shall be prevented by following the proper working length of the tooth being prepared, thereby preventing apical extrusion of debris. NiTi rotary instruments showed less debris extrusion when compared with the step-back technique using manual instruments . FurtherIt is evident from the results of the present study that there is an over-prescription of antibiotics by practitioners, especially by general dentists, without following proper guidelines for endodontic treatments. More emphasis should be made on the proper prescription pattern of antibiotics at the undergraduate level. In addition, dental practitioners must be updated on recent guidelines for antibiotic prescription, the WHO’s EML, and the AWaRe classification with the help of CDE programs and by following proper endodontic diagnosis and treatment protocols to prevent endodontic flareups, thereby reducing the need for antibiotics. Patients should be warned about the adverse effects of self-prescribing antibiotics.