This study aimed to assess the knowledge on antibiotics and antimicrobial resistance (AMR) and the antibiotic use among the general public in Southern Italy and to analyze whether sociodemographic characteristics could be associated with poor knowledge and improper practices.From March to November 2019, a face-to-face interview was conducted with adult subjects attending the waiting room of 27 randomly selected general practitioners (GPs) in Southern Italy. The questionnaire covered sociodemographic characteristics, knowledge on antibiotics and AMR and practices regarding the consumption of and self-medication with antibiotics.The response rate was 89.7%. In the sample, 29.2% thought that antibiotics are effective for viral infections, and 49.5% correctly recognized the definition of AMR. Predictors of good knowledge about antibiotics and AMR were female gender and a higher education level. Almost half of the respondents had used antibiotics in the previous year and 23.6% took antibiotics to treat a common cold and/or fever. Among participants, 25.5% reported to have bought antibiotics without a prescription, and 30.6% were classified as antibiotic self-medication users. Use of antibiotics in the previous 12 months and having taken an antibiotic after a phone consultation with the GP were positively associated with both antibiotic use for a common cold and/or fever and self-medication with antibiotics.The findings of this study highlighted a considerable antibiotic consumption in the adult population of Southern Italy together with misconceptions regarding the correct indication for antibiotic use that could foster indiscriminate antibiotic use.The findings of this study highlighted a considerable antibiotic consumption in the adult Italian population together with misconceptions regarding the correct indication for antibiotic use that could foster indiscriminate antibiotic use. Almost a quarter of the respondents took antibiotics to treat a common cold and/or fever and reported to have bought antibiotics without a prescription. Since their discovery, antibacterial drugs have become an essential part of the modern healthcare landscape, allowing treatment of previously life-threatening bacterial infections. However, the enormous and irresponsible use of the antibiotics in human, animal and environmental sectors has significantly contributed to the advent of resistant strains. The rapid emergence of antimicrobial resistance (AMR) is currently a worldwide public health crisis, and it has been cited by the World Health Organization as one of the top global public health problems , 2. The Preserving the continued effectiveness of existing antimicrobials includes a combination of interventions, such as strengthening health systems and surveillance, improving hygiene and infection control and appropriate use of antimicrobials in hospitals and in the community . SpecifiDespite available data on public perceptions of AMR and practices regarding antibiotic use in Southern Italy on subgroups of population  and 10 community-based pediatrician (CBP) from a list of geographical units provided by the health districts as being responsible for coordinating and providing primary care. All consecutive subjects were recruited at randomly selected days from those attending GP and CBP clinics, with 20–30 individuals selected from each physician’s clinic. Two trained physicians, not involved in patient care, conducted a structured face-to-face interview with the subjects attending the waiting room of the selected physicians. Data collectors assured the participants of confidentiality and informed them of the purpose and methods of the investigation, and a signed informed consent form was obtained from all participants who agreed to participate in the study.All the information was self-reported by the participants, no medical records or interviews by any GP or CBP were used as sources of data. No incentives were offered for participation.Sample size was determined in order to warrant estimation of proportions with an expected margin of error of 5%, assuming an intended confidence interval level (CI) of 95%. We used the prevalence of knowledge of antibiotics and AMR and practices regarding antibiotics use and self-medication obtained from similar studies , 16. BasThe questionnaire was developed based on the literature review of comparable studies , 16–21. The questionnaire covered a broad range of issues related to sociodemographic characteristics of the participants , knowledge on antibiotics and AMR and practices regarding the consumption of antibiotics.Each question elicited responses in a variety of formats: yes/no/do not know, closed-ended with multiple answers possible and open options.To assess knowledge, the participants were asked to respond to 10 different statements : two statements were related to the identification of antibiotics (Amoxicillin is an antibiotic/Paracetamol is an antibiotic); four related to antibiotic use with or without an indication ; two were about the side effects of antibiotics ; and two were regarding the awareness of AMR (AMR is a phenomenon that takes place when a bacterium loses its sensitivity to an antibiotic/Infection prevention measures limit the development of AMR). An overall knowledge score was calculated by assigning one point for each correct response and summing the scores to each statement. The total score ranged from 0 to 10. The overall median knowledge score of the respondents was then estimated and a ≤50th percentile score was interpreted as poor knowledge, whereas a >50th percentile score as good knowledge.The respondents’ antibiotic use and self-medication practices were measured with questions asking if they and/or their minor son/daughter (i.e. younger than 18 years old) had been prescribed antibiotics within the previous year; frequency of prescribing in the same period; having ever used an antibiotic without indication (Have you ever taken an antibiotic for common cold?/Have you ever taken an antibiotic for a fever?); whether they had adhered to the GP’s prescription; whether they had ever stored and used leftover antibiotics; whether they had ever purchased an antibiotic without a prescription of the GP and whether they had ever begun an antibiotic therapy after a phone consultation with the GP, without a clinical examination. The responders were classified as antibiotic self-medication users if they reported that they had ever bought an antibiotic without a prescription from the GP and/or had ever used leftover antibiotics.Ethical approval was granted by the Regional Human Research Ethics Committee (ID No. 122/2019/04/18).t-test and χ2 test) were conducted to examine the unadjusted relationships between the outcomes of interest and the independent variables. Furthermore, multivariate analyses were conducted using stepwise multivariate logistic regression modeling techniques. All significant variables identified in the bivariate analysis, and other variables considered as potential determinants of outcomes of interest were included in the analysis. A significance level of 5% was used for hypothesis testing. The following outcomes of interest were investigated: knowledge about antibiotics and AMR (Model 1), antibiotic use in the previous 12 months (Model 2), antibiotic use for common cold and/or fever (Model 3) and self-medication with antibiotics (Model 4). The following variables were included in all models: gender , age , nationality , education level , employment status , marital status  and having at least one minor son/daughter who had used antibiotics in the previous 12 months . In Models 3 and 4, the following variables were included: knowledge about antibiotics and AMR , use of antibiotics in the previous 12 months , having interrupted an antibiotic course , having taken an antibiotic after a phone consultation with the GP . In Model 3, the independent variables having purchased an antibiotic without a prescription  and storing leftover antibiotics  were also included. We applied a procedure to obtain the final models with P values of >0.2 and <0.4 for the inclusion and exclusion of the variables in the models, respectively. Adjusted odds ratio (OR) and 95% CI were calculated.Statistical analysis was performed using STATA software program, version 16 . Data weThe data set was deposited in Mendeley Data repository (doi:10.17632/6rkz746j35.1).Of the original sample of 633 subjects, 568 agreed to participate for a response rate of 89.7%.More than half were female (58.8%), and the mean age observed was 47.8 years (±16.7). The majority held high school qualification (47.5%), was employed (53.7%) and had at least one minor son/daughter (58.1%).The overall median knowledge score was 8 (interquartile range 6–9), and only 17.1% of all the respondents gave the correct answer to all 10 statements.The model predicting good knowledge about antibiotics and AMR showed multiple significant independent sociodemographic associations. Every level of education from secondary school upward was positively associated with good knowledge. The effect size for high school  was lower than bachelor/university/doctoral degree . Female gender  was positively associated and foreign nationality  negatively associated with correct knowledge on antibiotics and AMR  with antibiotic use in previous 12 months, whereas an independent negative association with employment status  was shown.When asked about the adherence to the prescribed antibiotic regimen, 30.5% of the study population reported that they had interrupted antibiotic use before they completed the course. The main reasons for stopping taking antibiotics were symptom relief (42.8%), having forgotten to complete the full course of antimicrobial therapy (15.6%) and onset of side effects (34.1%). Furthermore, almost half (43.1%) of the participants stated that they started antibiotic therapy at least once just after a phone consultation with the GP, without a clinical examination .In the sample, 23.6% took antibiotics to treat a common cold and/or fever. The model predicting this practice, with results presented in A quarter of the sample (25.5%) self-reported to have bought antibiotics without a prescription and 68% to have kept leftover antibiotics at home, and almost a quarter (23.8%) of them reported using antibiotics leftover from a previous course without consulting a GP. Overall, 30.6% of the sample was classified as an antibiotic self-medication user, namely they had taken leftover antibiotics and/or had bought antibiotics without a prescription. Model 4 showed an independent positive association of the interruption of an antibiotic course , of the use of antibiotics in the previous 12 months  and of the consumption of an antibiotic after a phone consultation with the GP without medical examination  with self-medication with antibiotics .This study has tried to investigate the knowledge and practices of the general population regarding antibiotic use and AMR. The findings of the present survey provide an up-to-date insight that will aid in the design of the community educational campaigns to promote prudent antibiotic use.Our results showed fairly poor knowledge of antibiotic use and AMR among the Italian population with less than 2-in-10 that correctly responded to all statements about the investigated knowledge. Many subjects failed to identify that antibiotics have no significant therapeutic effects on viruses. Similarly, European as well as international studies on the public knowledge of antibiotic use indicated a widespread ignorance regarding the ineffectiveness of antibiotic treatment for viral infections , 23–25. Confusion concerning the phenomenon of AMR was also detected. Among knowledge statements, those regarding AMR showed the highest percentage of failure, and these findings pointed out that the recognition of AMR and the measures for its prevention remain partially unknown among the public in Italy. The results of multivariate analysis underlined the heterogeneity of public knowledge regarding antibiotics and AMR on the basis of sociodemographic factors. These figures could be helpful as a considerable inter- and intraregional variability in antibiotic consumption and prescribing pattern has been already described in Italy  as well It is worthwhile to underline that the proportion of individuals who reported having taken at least one antibiotic for himself/herself (47.9%) or for their minor son/daughter (55.4%) in the previous 12 months is among the highest reported in European Member States, emphasizing that antibiotic consumption is considerable in the study area as well as in the country as a whole . A similWe investigated two types of practices that are known to contribute to self-medication: obtaining antibiotics from a pharmacy without a prescription and using leftover antibiotics from a previous course. In Italy, availability of over-the-counter antibiotics without prescription has already been described , althougThe findings of multivariate analysis identified groups of participants who were more prone to use self-medication antibiotics as well as to take antibiotics without an indication (a common cold and/or fever). As expected, respondents’ adequate knowledge of antibiotics was identified to positively correlate with correct antibiotic use, which is consistent with previous studies , 23, 32.The current finding that almost 3-in-10 of those being prescribed an antibiotic did not follow the established regimen and discontinue treatment prematurely mainly due to relief of symptoms is of concern. Similar results were obtained in studies conducted in other countries, including the UK (11.3%) , PortugaIn our study, the mass media, including the Internet, were indicated as the main sources of AMR information. The role of the Internet as a common source of health- and antibiotic-related information was reported by previous research conducted in the Italian population , 39. HeaThe strengths of this study include a high response rate, an even distribution of men and women, all the desired age groups and subjects having high as well as low levels of education. The high response rate of a survey is a key data quality indicator since reducing nonresponse bias coupled with random sampling ensures generalizability of findings to the population of interest. Another strength is that the respondents did not have the opportunity to take the questionnaires home. Consequently, they could not look up the correct answers online or confer with someone else before responding, that could lead to some overestimation of knowledge and of correct practices regarding antibiotic use. Limitations of this study attain to the cross-sectional design, not allowing to draw conclusions on causality about the observed associations, and to the self-reporting of practices, only a proxy of real practices. Another of the study’s limitation is the potential overestimation of positive outcomes. It is possible that people who were more informed or interested in the topic were more willing to participate, but the high response rate reduces this limitation to a minimum. To overcome the difficulty of identifying a sampling frame of all primary care patients, we first selected a random sample of clusters (27 clinics), and then all consecutive subjects attending GP and CBP clinics during randomly selected days were invited to participate. Since enrollment of patients within practices was not random, issues of representativeness may arise and indeed selected patients may be representative of those who attend GP and CBP clinics. However, in an attempt to overcome this limit, we compared the sociodemographic characteristics of the study sample in some clinics with those of all the patients referring to that specific clinic, and no substantial differences were found. Furthermore, our study involved two Italian Regions, which might not represent all the adult population in Italy. However, we are confident that the findings of the study may be representative at least for the Regions of Southern Italy.Even with these potential limitations, the findings of the present study highlighted a considerable antibiotic consumption in the adult population of Southern Italy together with misconceptions regarding the correct indication for antibiotic use that could foster indiscriminate antibiotic use. Policy makers should consider multifaceted interventions to tackle the determinants of antibiotic misuse at the healthcare system as well as at the patient level.No external funding for this manuscript.Conflicts of interest: None declared.


The distribution of pores, wet and dry density and compressive strengths were evaluated. In addition, the creep deformations of foamed concrete with different densities were measured. The difference in density of up to 170 kg/m3 for the highest densities was obtained. Foamed concrete with higher densities (700 and 800 kg/m3) showed similar characteristics of pores, which were different from those of samples with a density of 500 kg/m3. Compressive strength equal to 5.9 ± 0.2, 5.1 ± 0.2, 3.8 ± 0.3 and 1.4 ± 0.2 MPa was obtained for foamed concrete with a density of 500, 700, 800 and 1000 kg/m3, respectively. The obtained compressive strengths were higher than those found in the literature for the foamed concrete with the same densities. With increasing density, smaller creep deformations were obtained. Creep deformations were 509, 495 and 455 με for samples with densities of around 500, 700 and 1000 kg/m3 respectively. Deformation under long-term loading took place up to 90 days, regardless of the density of the foamed concrete.The components of foamed concrete have a significant effect on its properties. Protein-based foamed concrete is used much more often. This study aims to assess the properties of foamed concrete with a density of around 500, 700, 800 and 1000 kg/m Foamession [3 ,10,11) assion [3 ,13. Foamssion [3 ,15,16. Fssion [3 ,18,19,20on [3 [3 ,22,23, won [3 [3 ,25,26. Ion [3 [3 , Fadila on [3 [3 , Kang [2on [3 [3 , Fernandon [3 [3  and Porton [3 [3  producedon [3 [3  used foaon [3 [3 ,34,35, Don [3 [3 ,37,38, Ton [3 [3  and Lee on [3 [3  used foaon [3 [3 . In addion [3 [3 ,43,44.3 for a protein-based foaming agent than for a synthetic-based foaming agent. This impact was far more pronounced than that observed by Panesar [3 formed by using a synthetic polymer-based foaming agent. Evaluation of the foamed concrete was carried out to examine the distribution of pores and properties such as wet and dry density and compressive strength. In addition, unlike other scientists, we measured the creep deformations of foamed concrete with different densities. Assessment of behavior under long-term load is very important for structures such as roads or car park pavements, floor structures or building foundations, where foamed concrete is currently used, as mentioned above.The components of foamed concrete , tap water and foaming agent. The industrial Portland cement was CEM I 42.5R, according to PN-EN 197-1:2012. Its chemical composition measured as per PN-EN 196-2:2013-11  and physical properties measured according to PN-EN 196-6:2011 are given in 3 was used as foaming agent. The FTIR spectra of the foaming agent are shown in A commercial liquid polymer admixture with specific gravity of 1.02 g/cm3 with a tolerance of ±50 kg/m3, marked as FC500, FC700, FC800 and FC1000, respectively. The synthetic foaming agent  contents were 8.0, 6.0, 5.0 and 4.0 dm3 per 100 kg of cement. As reported in effw/c ratio was fixed equal to 0.44 for all specimens, where effw is water content, which includes tap water and liquid foaming agent; c is the cement content. This was based on the results of Jones and McCarthy [The specimens of foamed concrete with four different densities were produced to investigate the effect of synthetic polymer foaming agent on the properties of foamed cementitious mix . The denMcCarthy , XianjunMcCarthy  and the McCarthy ,78,79.3, the appropriate amount of liquid polymer agent was pressurized with air at approximately 5 bars using a foam generator. The foam content was established on the basis of the target dry density (considering earlier experiments); see 3 was allowed on the achieved dry density. The entire manufacturing process of foamed concrete must carefully consider the densities of the mix, the foaming production rate and other factors in order to prepare high-quality foamed concrete. The key factors in producing stable foamed concrete included the pressurizing of the foaming agent at stable pressure and mixing the components in constant rotational speed.The foamed concrete specimens were prepared with the pre-forming method. First, water and cement were mixed, and in the next step, the foam was added and all components were mixed a. In ordAll specimens were poured into steel molds b,c and wThe tests on hardened concrete were carried out after 28 days of curing. The material (density and distribution of pores) and mechanical properties of the concrete (compressive strength and creep deformation) were tested. Because foamed concrete is a relatively new material, there are no test standards for it in Poland or any other country .The density of the mixture was measured as per PN-EN 12350-6:2011 after pouring the mixture into the mold. The six specimens for each mix were measured.The density of hardened foamed concrete was measured with 150 mm × 150 mm × 150 mm standard cubes as per PN-EN 12390-7:2011. The as-received specimen, i.e., naturally dried samples, was assessed. Ten measured specimens for each mix were tested.The distribution of air bubbles in the foamed concrete was analyzed using a LM microscope .Compressive strength was measured on samples 150 mm × 150 mm × 150 mm, according to PN-EN 12390-3:2011 + AC:2012, using a compression machine MEGA 6-3000-100  having 3000 kN as max. load capacity. The loading rate was assumed as per PN-EN 772-1 + A1:2015-10 as for cellular concrete masonry units. Three samples per mix were used.cε(t)—creep deformation in time t;totε(t)—total measured creep deformation of foamed concrete sample long-term loading in time t;aε—immediate strain determined after 5 min of obtaining 1/3 of compressive strength;sε(t)—shrinkage in time t.The creep test was carried out on beams 150 mm × 150 mm × 450 mm using a concrete creep testing machine  in a room at 20 ± 1 °C. The load was applied continuously to the level of 1/3 of the compressive strength. For this purpose, the average compressive strength was determined on three vertically oriented base samples; see Therefore, at the same time as the start of the long-term loading of samples, the next three samples of 150 mm × 150 mm × 450 mm were tested in the shrinkage test using Amsler’s apparatus . The initial distance between the measuring points on the sample was determined. Subsequently, the samples were stored in a curing room at 20 ± 2 °C and 50 ± 3% humidity, and the distance between the measuring points on the sample was measured.s—εshrinkage in i-period;l0, t—linitial distance between measuring points and distance after a specified curing time.The shrinkage was determined by Equation (2). Three specimens for each density were tested.3 for foaming agent content of 4.0, 5.0, 6.0 and 8.0 dm3 per 100 kg of cement, respectively. The volume of foam commonly created air-voids and resulted in lower density [3. On the basis of the achieved results, it was observed that the densities of the hardened sample from one batch of the mix were obtained with a standard deviation of ±30 kg/m3 and with tolerance at ±50 kg/m3 of the target value. This is in line with the practical method used in industrial concrete production [3, which might reach a difference of up to ±100 kg/m3 for high density [oduction ,72. The 3 depending on the density of foamed concrete. Larger differences were demonstrated for higher densities; see 3 for synthetic-based foamed concrete, with density in the range of 400 to 800 kg/m3 and 40 to 100 kg/m3 for protein-based foamed concrete. The densities of the mixture and hardened samples decreased with the increase in foaming agent content . These rThe decrease in density with increasing of the foaming agent content is associated with the change in the air-void structure of foamed concretes; see 3 density  had similar characteristics of pores, such as number, area, perimeter and shape descriptors . The size of the pores ranged from 0.26 to even 3.45 mm for foamed concrete with a density of 700 kg/m3. This was not the case in samples with a density of 500 kg/m3; see 3 were more circular and round. The average size of pores ranged from 0.43 to 0.65 mm. The same size of pores was determined by Zhu et al. [3 had smaller air-voids. This is consistent with the observation that smaller size of pores corresponds to higher density [3; Hilal et al. [3; and Sun et al. [3. At the same time, more large air bubbles of up to 3.5 mm in diameter were observed in higher densities; see Based on u et al.  for anio density . For exa density  obtainedl et al.  achievedn et al.  obtained3 per 100 kg of cement, the 28-day compressive strength was equal to 5.9 ± 0.2, 5.1 ± 0.2, 3.8 ± 0.3 and 1.4 ± 0.2 MPa, respectively. The addition of foaming agent decreased compressive strength because the volume of foam created pores and resulted in lower density.The results of compressive strength for samples with different densities are shown in The correlation between density of hardened foamed concrete and its compressive strength is exponential, which is in line with observations made by other scientists ,59.3. The compressive strength obtained for higher densities influences the shape of the approximation curve. Moreover, despite the fact that the values were higher than in this study, the dispersion of the obtained results was greater. The presented diagram confirms that, depending on many factors , the obtained results may be different. This is also confirmed by the results of other scientists; e.g., Castillo-Lara et al. [3. Meanwhile, She et al. [3. This is due to the good structure of the foam (homogeneity of the bubbles) and its properties [3 was determined by using an organic stabilizer or 81% when nano-silica was used additionally [3 was obtained [The results obtained in this study showed that the compressive strengths were higher than the results achieved by other scientists for the same densities, but lower than in the authors’ previous research , marked a et al.  obtainede et al.  achievedoperties . In addioperties  to preveoperties  and thustionally . Moreoveobtained . In addi3 density. Brittle behavior of foamed concrete specimens with higher density can be observed.Images showing the failure of foamed concrete samples with different densities are presented in 3, respectively. Deformation under long-term loading took place up to 90 days, regardless of the density of foamed concrete. After 90 days, the differences in the values in two consecutive measurements were less than 0.2%, and therefore the creep process was considered complete. For foamed concrete with a density of around 700 and 1000 kg/m3, around 2.6% and 10.6% decrease compared to samples with density of 500 kg/m3 were obtained.The results of long-term loading of foamed concrete samples with different densities are shown in 3; see 3. Drying shrinkage of foamed concrete was relatively high. It was found to be 4 to 10 times higher than normal concrete due to the aggregate type in the mix design, higher cement and water content and mineral admixture in foamed concrete [3, which was made of Portland cement (type I), water (effw/c = 0.5) and sand [effw/c = 0.43) and ground granulated blast-furnace slag, Wan et al. [effw/c ratio of 0.5) with a density of around 600 kg/m3 on day 90 was from 2500 to 3020 με depending on the type of foaming agent used. Meanwhile, drying shrinkages of normal concrete are reported to be between 200 and 800 με [The simultaneous measurement of shrinkage strains took place in concrete for 30 to 50 days of measurement for foamed concrete with a density of around 500–1000 kg/mconcrete . It is rand sand . For then et al.  obtainedn et al.  determind 800 με , and ford 800 με .Drying shrinkage can be modified by selecting a suitable foaming agent type at an appropriate volume ,87, alth3 per 100 kg of cement), and the results obtained foamed concrete with a target density of 500, 700, 800 and 1000 kg/m3 with a tolerance ±50 kg/m3. Based on the results of this experimental investigation, the following key conclusions can be drawn:3 for foaming agent content of 4.0, 5.0, 6.0 and 8.0 dm3 per 100 kg of cement, respectively. The volume of foam commonly created air-voids and resulted in lower density.The density of hardened foamed concrete was 970 ± 30, 800 ± 30, 720 ± 20 and 550 ± 20 kg/m3 depending on the density of foamed concrete samples; larger differences were demonstrated for higher densities. In this way, the difference between the density of the mixture and the hardened foamed concrete was obtained in order to design the mixture with the target parameters.The differences between the density of the mixture and the density of the hardened sample were up to 170 kg/m3 had similar characteristics of pores, such as number, area, perimeter and shape descriptors . It can be observed that the size of pores was from 0.26 to even 3.45 mm for foamed concrete with a density of 700 kg/m3, while the average size of pores for foamed concrete with a density of 500 kg/m3 was from 0.43 to 0.65 mm.Foamed concrete with a density of 700 and 800 kg/m3 was greater and the average thickness of air bubbles wall was thinner than for the other densities.The number of air-voids for foamed concrete with a density of 500 kg/m3 had smaller air-voids, but more large air bubbles up to 3.5 mm in diameter were observed by higher density.Specimens with a density of around 700 and 800 kg/m3 per 100 kg of cement, the compressive strength was equal to 5.9 ± 0.2, 5.1 ± 0.2, 3.8 ± 0.3 and 1.4 ± 0.2 MPa, respectively. The obtained compressive strengths were higher than those found in the literature for the same densities. This was most likely related to the fact that the synthetic foaming agent contained a stabilizer foam.With the addition of foaming agent of 8.0, 6.0, 5.0 and 4.0 dm3. Deformation under long-term loading took place up to 90 days, regardless of the density of the foamed concrete.With increasing density of foamed concrete, smaller creep deformations were obtained. Creep deformations were 509, 495 and 455 με for samples with density of around 500, 700 and 1000 kg/mThe aim of the study was to assess the properties of foamed concrete made of a synthetic polymer foaming agent. This experimental work has included four different foaming agent contents (4.0, 5.0, 6.0 and 8.0 dm


The human gut microbiome is a dynamic collection of bacteria, archaea, fungi, and viruses that performs essential functions for immune development, pathogen colonization resistance, and food metabolism. Perturbation of the gut microbiome’s ecological balance, commonly by antibiotics, can cause and exacerbate diseases. To predict and successfully rescue such perturbations, first, we must understand the underlying taxonomic and functional dynamics of the microbiome as it changes throughout infancy, childhood, and adulthood. We offer an overview of the healthy gut bacterial architecture over these life stages and comment on vulnerability to short and long courses of antibiotics. Second, the resilience of the microbiome after antibiotic perturbation depends on key characteristics, such as the nature, timing, duration, and spectrum of a course of antibiotics, as well as microbiome modulatory factors such as age, travel, underlying illness, antibiotic resistance pattern, and diet. In this review, we discuss acute and chronic antibiotic perturbations to the microbiome and resistome in the context of microbiome stability and dynamics. We specifically discuss key taxonomic and resistance gene changes that accompany antibiotic treatment of neonates, children, and adults. Restoration of a healthy gut microbial ecosystem after routine antibiotics will require rationally managed exposure to specific antibiotics and microbes. To that end, we review the use of fecal microbiota transplantation and probiotics to direct recolonization of the gut ecosystem. We conclude with our perspectives on how best to assess, predict, and aid recovery of the microbiome after antibiotic perturbation. The human gut microbiome consists of bacteria, viruses, and fungi ideally living symbiotically with their human host, though this review will focus exclusively on bacterial residents within the gut microbiome . Individα) and between (β) samples can be used to compare microbial communities over time and between disease states . Our abiewed in , 23. Howiewed in , 24, 25.The most common external perturbations to the microbiome are diet, medications , and the environment –31. In tImmediately after birth, bacteria, viruses, and fungi colonize the sterile human gut with a subsequent rapid accumulation of species and diversity from the mother and the environment –34. Gut Clostridioides difficile was universally lower and Fusobacterium nucleatum higher than the actual abundance of the mock community [The overall diversity and community architecture of healthy adult microbiomes do not change dramatically in the absence of significant perturbation , 40, 41.ommunity . Conversommunity . TherefoAlthough the dichotomy of stable  versus dynamic  microbiomes is likely oversimplified, it is important to understand that the impact of an intervention/perturbation depends on the context Fig. . That isβ diversity between consecutive samples [Large-scale studies of the microbiome have demonstrated that the dynamic first 2 years of life respond most dramatically to antibiotic perturbation , 50. Ant samples . Among t samples , 28. Pri samples , 53  as well as antibiotics not used in the NICU (tetracyclines and chloramphenicol) [E. coli. Accordingly, these novel ARGs are likely still an underestimate of the total resistome. To date, similar methods have been unsuccessful in gram-positive bacteria. Resistance to antimicrobials that exclusively or preferentially target gram positives such as vancomycin, linezolid, and clindamycin would require other methods to identify. Therefore, frequent antibiotic use during times of microbial change acutely disturbs the microbiome and enriches for potential pathogens and ARGs.Antibiotic treatment in infancy enriches for AR organisms in the stool as determined by selective culturing and DNA sequencing , 54. To ve media , 54, 67.ve media . GreaterBifidobacterium spp. and, thereafter, several microbiome states, so-called enterotypes, account for the bulk of gut community structure [Childhood is a time of immense microbiome dynamics and environmental changes including dietary shifts and introductions , 26, 69.tructure , 37. Tratructure , 69.Ruminococcus even though the overall number of species and diversity was similar between exposed and unexposed groups after 1 year of life  acquired in the NICU persisted in the gut microbiome up to 1 year after NICU discharge with greater than 99.997% identity between them. Machine learning algorithms classified fecal samples post-NICU discharge deriving from an antibiotic-naive or frequently exposed infant with 96% accuracy [Frequent antibiotic use in the NICU delays preterm neonatal microbiome maturation initially, but the microbiota recovers to a similar taxonomic composition to antibiotic-naive term controls by 15 months of life . Similaraccuracy . Among tBacteroides fragilis, a key commensal important for immune education and bacterial tolerance [Eubacterium rectale) in lieu of initial colonization and in vivo evolution. That is, because of frequent antibiotic administration, beneficial microbes are not surviving and undergoing in vivo adaptation, potentially inhibiting the microbiome plasticity expected of a healthy, interconnected, adult microbiome [In addition to allowing MDRO potential pathogens to maintain a foothold within the GI tract, frequent antibiotic exposure in this critical microbiome developmental period diminished strain diversity, enriched for ARGs, and conferred a less stable composition . Researcolerance , 89. Harolerance , 90, 91.crobiome , 92. Concrobiome . FrequenDorea spp. and Faecalibacterium prausnitzii relative to healthy children living in the same geographical area [Enterobacteriaceae dramatically increased in children with SAM during treatment with ampicillin/amoxicillin and gentamicin [The main arguments against the use of antibiotics in vulnerable populations are the loss of diversity and selection for AR pathogens, which is especially worrisome when patients are immunocompromised. Treating the gut microbiota of children with severe acute malnutrition (SAM) therefore constitutes a special case combining all of the factors. In healthy children older than 2 years, some adult-like stability and resilience to antibiotics should be characteristic. Malnourished children however exhibit an age-regressed microbiome which is often lacking taxa of a healthy child or adult . Specifical area . Broad-sntamicin . The incCampylobacter spp., which can cause diarrheal illnesses worldwide [The data to address the long-term concerns of antimicrobial treatment of SAM is accumulating in placebo-controlled randomized trials of mass drug administration. Azithromycin and amoxicillin have been trialed to improve all-cause mortality and SAM in sub-Saharan Africa –95. Reseorldwide . As a trorldwide . Similarorldwide , 58 . Additionally, untreated siblings of amoxicillin-treated children were more likely than placebo-controlled siblings to acquire an ESBL Enterobacteriaceae. Therefore, the administration of antibiotics can potentially affect the long-term AR of specific genera and can lead to environmental dissemination. However, given the repeated demonstration of their positive effect on childhood health, we support this practice. Further research demonstrating the environmental impact of increased AR in this population needs to be performed.Amoxicillin is also routinely prescribed for uncomplicated SAM in sub-Saharan Africa; however, similar concerns about the long-term consequences of this practice abound . To addr threats , 98, 99.ulturing . This reThe relatively stable adult microbiome requires more sensitive measurement techniques to assess long-term perturbations. Once the microbiome has stabilized around age 2–3 , 35, minClostridiales, among the most abundant pre-ciprofloxacin taxa, were absent in all samples after treatment [The response to the same antibiotic appears individualized at the taxa level likely reflective of underlying co-morbidities, local environment, or prior antibiotic history commensurate with ARG abundance Fig.  84, 101101. AddiClostridioides difficile infection (CDI) and steroid non-responsive colitis [Klebsiella spp., Pseudomonas, and vancomycin-resistant enterococci with allo-FMT , thoughiewed in , 112. Siiewed in –115. Reciewed in ) delayedrichness . This prE. coli derived from donor FMT, one of whom succumbed to the infection [Despite the benefits of FMT detailed above, the introduction of fecal microbes into another individual is not without risk. Recently, two immunocompromised patients became bacteremic with ESBL producing nfection . Accordinfection . One othnfection , 105. ThThe state of the microbiome, the duration, route, and spectrum of antibiotic activity, other co-morbidities, diet, and post-antibiotic environment all factor into the expected acute and chronic disruption and resilience from perturbation Fig.  7, 28, , 59, 74.


IgG4-related disease (IgG4-RD) is a slowly progressing inflammatory disease that can involve multiple organ systems. There is considerable overlap between IgG4-RDs and anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). Herein, we present an unusual case of IgG4-associated tubulointerstitial nephritis (IgG4-TIN) and ANCA-associated glomerulonephritis (ANCA-GN) co-occurring with C3 glomerulonephritis (C3GN).A 72-year-old male was admitted to hospital because of fever and fatigue. He was diagnosed with elevated serum creatinine and IgG4 levels, and was positive for ANCA.Initially, the pathology supported a diagnosis of IgG4-TIN and ANCA-GN; however, further examination revealed he also had C3GN.The patient was treated with methylprednisolone and cyclophosphamide and received regular follow-up care.After treatment, the patient no longer exhibited fever or fatigue and had no complications. The seven-month follow-up showed downward trends in IgG4 and MPO-ANCA levels and stable 24-hour urine protein, serum creatinine levels.Anti-neutrophil cytoplasmic antibody-associated glomerulonephritis and IgG4-associated tubulointerstitial nephritis with C3glomerulonephritis rarely occur simultaneously. Laboratory analysis and pathology are both needed to ensure diagnostic accuracy. However, in this case, the three diseases overlapped to such a large extent that achieving a definitive diagnosis was particularly challenging. Timely and accurate diagnosis is crucial for selecting the best treatment course and optimizing patient outcome. There is substantial overlap in the types of organs involved and the histopathology of IgG4-RD and anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). C3 glomerulonephritis (C3GN) was first characterized by Verroust in 1974 as a type of glomerulopathy, named by Fakhoufi et al, that is characterized by immunoreactive C3 complement protein deposited along the capillary loop or mesangial area of glomeruli, without immunoglobulin deposition. There are no reports of co-occurrence of these three kidney diseases in a single patient. Here, we describe a case of ANCA-GN and IgG4-TIN with C3GN simultaneously affecting 1 patient. We analyzed the clinical characteristics, pathology, common pathophysiologies, and histopathological characteristics of ANCA-GN, IgG4-TIN, and C3GN to provide clinical insight and facilitate more accurate clinical diagnoses.IgG4-related disease (IgG4-RD) is a fibro-inflammatory condition that can affect every organ system in the body.29/L; hemoglobin, 72 g/L, and platelets, 400 x 109/L. Abdominal CT results were normal. A bone marrow smear showed signs of hyperplastic anemia. Granulocyte-poisoning particles in peripheral blood, vacuole degeneration, and, occasionally, late granulocytes were also present.A 72-year-old male was admitted to our hospital after experiencing 2 months of intermittent fever, fatigue, and full-body discomfort. He was treated with antibiotics at a local hospital, but there was no improvement in his symptoms. The fever was apparently unprovoked. He had no known personal or family history of renal dysfunction. Physical examination indicated his blood pressure was 137/74 mmHg, heart rate was 100 beats/min, and cardiopulmonary and neurological functions were normal, with no costovertebral knocking tenderness. A urinalysis performed upon admission showed 3+ occult blood and his 24-hour urine protein level was 1.24 g. Serum biochemistry revealed the following: creatinine, 233.9 μmol/L; MPO-ANCA, 203.45 AU/mL, p-ANCA, 1:32, IgG, 22.90 g/L; IgG4, 7.230 g/L; complement C3, 0.82 g/L, erythrocyte sedimentation rate (ESR), 120 mm/h; and C-reactive protein (CRP), 141 mg/L. Routine blood analysis revealed the following: white blood cells, 11.45 x 10Among sixteen glomeruli examined in a renal biopsy, one had glomerular sclerosis, and eight had cellular crescent formations, we observed 2 cellular fibrinous crescents, and one renal bulb with segmental celluloid necrosis. Furthermore, there was evidence of local mild edema and diffuse inflammatory cell infiltration into the renal stroma, predominantly lymphocytes and plasma cells, macrophages, a small number of neutrophils and eosinophils, in addition to stromal foci with mild fibrosis. Immunofluorescent staining of three glomeruli indicated that these were highly C3-immunoreactive yet negative for IgA, IgM, IgG, C4, and complement F. Immunohistochemical analysis demonstrated that most of the IgG-positive infiltrating inflammatory cells were IgG4 immunoreactive, accounting for over 40% of the IgG-positive cells.Our patient was admitted due to intermittent fever and fatigue. Upon admission, further testing revealed albuminuria, hematuria, elevated serum creatinine levels, serum IgG4 antibody levels three-fold higher than typical healthy values, and MPO-ANCA-positive serum. These serological indicators are characteristic of IgG4-TIN and ANCA-GN. The pathological results from a renal puncture indicated that 67% of the glomeruli were marked by crescent-shaped scars with segmental cellulose-like necrosis, with no obvious immune deposits, which is in line with the pathological characteristics of ANCA-GN Fig. A. The reWe prescribed methylprednisolone and cyclophosphamide Fig. . The patHe continued with these treatments and received regular follow-up care at our hospital. The monthly follow-up showed downward trends in MPO-ANCA, 24-hour urine protein, IgG4, CRP, and ESR, but creatinine remained around 170 μmol/L Fig. . Due to 3 The histological hallmarks characteristic of IgG4-RD include storiform fibrosis, obliterative phlebitis, tissue eosinophilia, and lymphoplasmacytic infiltration in which the ratio of IgG4- to IgG-positive plasma cells usually exceeds 0.40; interestingly, these features also affect seemingly unrelated organs. To date, diagnostic criteria for IgG4-RD have been proposed by two groups.,8 IgG4-RD generally responds to glucocorticoids during its inflammatory stage, but recurrent or refractory cases are common. The extent of the fibrosis is a useful predictor of responsiveness to immunosuppressive therapies.,10Given the presence of autoantibodies, IgG4-RD is categorized as an autoimmune disease; however, there is no evidence that these autoantibodies contribute directly to the pathogenesis or progression of the disease. IgG4-RD lesions are infiltrated by T helper cells, which likely cause progressive fibrosis and organ damage. The pathogenesis of the disease is not clear. Patients with IgG4-TIN generally present with proteinuria, hematuria, decreased kidney function, hypocomplementemia, and radiologic abnormalities. IgG4-TIN has the same histopathology as in other organs, however, it can be distinguished from many other forms of organ involvement by the presence of extremely low concentrations of complement protein. One plausible explanation is that hypocomplementaemia in IgG4-RD results from the formation of immune complexes that contain IgG1 or IgG3, which bind complement more effectively, which supports the notion that IgG4 does not directly contribute to the pathogenesis of IgG4-TIN.‘IgG4-related tubulointerstitial nephritis (IgG4-TIN)’ was first proposed by Saeki T. Kidney involvement seems to be more frequent in MPA than GPA but some groups have not been able to confirm this result. By contrast, ear nose and throat involvement is more frequent in GPA than MPA. In ANCA-GN, immunoglobulin deposition in glomeruli is considered to be infrequent and minimal and, therefore, it is classified as pauci-immune glomerulonephritis. However, studies have shown granular IgG and/or IgM deposition in ANCA-GN. MPO-ANCA-GN is characterized by pauci-immune necrotizing glomerulonephritis (NGN) with crescent formation. MPO-ANCAs bind to MPO molecules on the surface of primed neutrophils/monocytes and induce the release of reactive oxygen species and proteases near the endothelium, thereby causing glomerular and systemic capillary inflammation. Recent studies have also implicated other factors, including immunoglobulins, that are associated with glomeruli. AAV can eventually lead to acute renal failure, life-threatening pulmonary hemorrhage, and gastrointestinal dysfunction and, as such, are potentially fatal.AAV is a heterogeneous group of diseases that show necrotizing inflammation of small vessels with a wide range of clinical presentations and includes: granulomatosis with polyangiitis (GPA), eosinophilic GPA (EGPA), and microscopic polyangiitis (MPA).The similarities between the organs affected, biochemistry, and pathogenesis of IgG4-RD and AAV warrants elaboration.1)IgG4-RD in patients who are ANCA-positive is limited to the head and neck and does not involve other commonly affected organs, such as the lungs, pancreas, biliary tree, and retroperitoneum. By contrast, AAV in patients with established IgG4-RD presents with systemic manifestations beyond the ear/nose/throat region. One case study reported a patient with histological features of the left lacrimal gland that were diagnostic for IgG4-RD while the patient's lung biopsy showed typical features of GPA.2) IgG4 infiltration is not unique to IgG4-RD. Tao Su et al reported a case of typical IgG4-TIN concurrent with IgG4 MPO-ANCA-positive necrotizing crescentic GN. Raissian et al reported that moderate to marked IgG4-positive plasma cell infiltration is observed in 31.7% of patients with pauci-immune vasculitis, especially in MPO-ANCA-associated EGPA or Churg–Strauss syndrome. Moreover, ANCA is sometimes absent in AAV cases. Toru Sakairi et al reported an ANCA-negative case with histological and clinical findings similar to those seen in IgG4-TIN; however, the presence of cellular crescent formation in a few glomeruli and fibrinoid necrosis in an interlobular artery led to a diagnosis of small-vessel vasculitis, which is considered to be a renal manifestation of AAV. Serological indicators are not sufficient to define a disease; rather, to achieve a definitive diagnosis, these must be combined with pathological features.Although serum IgG4 is regarded as a sensitive marker of IgG4-RD, it lacks diagnostic specificity.3) Whether IgG4 can induce an autoantigen-mediated secondary inflammatory response requires further study.IgG is a potent neutrophil stimulus, particularly when presented as ANCAs in AAV. Normal IgG binds flowing neutrophils efficiently with an affinity in the order of IgG3 > IgG1 > IgG2 > IgG4. Patients with IgG4-RD exhibit diffuse infiltration of IgG4-producing plasma cells into the interstitial renal space. One study reported that IgG1 and IgG4 were dominant in both MPO-ANCA-positive IgG preparations and PR3-ANCA. C3GN affects all ages, and does not have a sex predilection. Most patients present with hypertension, proteinuria, and hematuria. In both the short- and long-term, renal function remains stable in the majority of patients. Alternative pathway abnormalities are heterogeneous and can have both acquired and genetic origins. The most common acquired abnormality is the presence of C3 nephritic factors (C3Nef), while the most common genetic contribution arises from the H402 and V62 alleles of factor H. In addition, other risk factors include factor H autoantibodies and mutations in CFH, CFI, and CFHR genes. These factors result in the dysregulation of the alternative pathway of complement activation, causing aberrant complement protein deposition on glomeruli and, consequently, glomerular injury. Light microscopy-based studies of C3 glomerulonephritis have demonstrated its proliferative pattern. Immunohistochemistry studies have revealed a dominant C3 staining pattern with minimal or no Ig immunoreactivity. Electron microscopy studies have shown that C3 deposits are located in the mesangium and along the capillary wall.,26 Although immunosuppressive agents and terminal complement pathway blockers are beneficial for some patients, there is no treatment for this disease. Limited data from patients with renal transplants indicate that the recurrence rate of C3GN is high.,28C3GN is a rare disorder characterized by excess alternative complement pathway activation. In C3GN, C3Nefs can stabilize C3 inverters by prolonging their half-life, increasing C3 fragmentation, and thereby decreasing serum complement C3 levels; moreover, abnormal complement regulatory proteins can also lead to excessive activation of complement alternative pathways. Aadel Chaudhuri et al reported a case of C3GN co-occurring with GPA, but this diagnosis remains unclear as the incidence of GPA is high and other reports have described cases of GPA with glomerular immune deposits. Therefore, it is difficult to determine whether this patient truly harbored both diseases or a variant of only one of the diseases. This case nicely demonstrates the pathogenic similarities between AAV and C3GN; however, the specificities of the underlying mechanisms require further investigation.,32Studies have shown that C3GN and AAV overlap in their pathogenesis. Hillhorst et al retrospectively analyzed 187 renal biopsies from patients with ANCA-positive glomerulonephritis. Among these, 78 stained positive for C3 complement protein using immunohistochemistry. They found that complement activation in AAV occurs predominantly via the alternative pathway.To our knowledge, this the first reported case of a patient with concurrent IgG4-TIN, ANCA-GN, and C3GN. This rare diagnosis was substantiated by several factors. The patient's IgG4 level was above normal and he was positive for MPO-ANCA. Pathology findings included IgG4-positive plasma cell infiltration, crescent-shaped glomeruli, deposition of immunoreactive C3, and the presence of electron-dense sediment in the superior subcutaneous region, basement membrane, and mesangial area. Taken together, these clinical features led us to diagnose concurrent IgG4-TIN and MPO-ANCA-associated necrotizing crescentic GN with C3GN. The patient responded well to methylprednisolone and cyclophosphamide treatment with marked improvement in renal function.4,29,31,32In conclusion, here we describe a rare case of concurrent IgG4-TIN and MPO-ANCA-associated necrotizing crescentic GN with C3GN. This case exemplifies the difficulty of differentially diagnosing three separate diseases and also provides insight into the relationship between IgG4-RD and AAV and how to accurately identify them.Conceptualization: Jianan Feng, Weixia SunData curation: Jinyu Yu, Xueyao WangFunding acquisition: Weixia SunInvestigation: Yue Wang, Yang LiuProject administration: Zhonggao XuResources: Weixia SunSoftware: Jinyu Yu, Xueyao WangValidation: Weixia SunVisualization: Jianan FengWriting – original draft: Jianan FengWriting – review & editing: Weixia Sun


In 2011, the IgG4-related kidney disease (IgG4-RKD) working group of the Japanese Society of Nephrology proposed diagnostic criteria for IgG4-RKD. The aim of the present study was to validate those criteria and develop a revised version.Between April 2012 and May 2019, we retrospectively collected Japanese patients with kidney disease, for whom data on serum IgG4 values and/or immunohistological staining for IgG4 in renal biopsy samples were available. These patients were classified as IgG4-RKD or non-IgG4-RKD based on the diagnostic criteria for IgG4-RKD 2011, and the results were evaluated by expert opinion. Accordingly, we developed some revised versions of the criteria, and the version showing the best performance in the present cohort was proposed as the IgG4-RKD criteria for 2020.Of 105 included patients, the expert panel diagnosed 55 as having true IgG4-RKD and 50 as mimickers. The diagnostic criteria for IgG4-RKD 2011 had a sensitivity of 72.7% and a specificity of 90.0% in this cohort. Of the 15 patients with true IgG4-RKD who were classified as non-IgG4-RKD, all lacked biopsy-proven extra-renal lesions, although many had clinical findings highly suggestive of IgG4-RD. The revised version to which “bilateral lacrimal, submandibular or parotid swelling, imaging findings compatible with type 1 autoimmune pancreatitis or retroperitoneal fibrosis” was added as an item pertaining to extra-renal organ(s) improved the sensitivity to 90.9% while the specificity remained at 90.0%.The revised version has considerably improved test performance after addition of the new extra-renal organ item . IgG4-related disease (IgG4-RD) is a fibroinflammatory condition characterized by infiltration of abundant IgG4-positive plasma cells with fibrosis, and usually an elevated serum IgG4 concentration , 2. In 2The aim of the present study was to determine the validity and potential problems of IgG4-RKD 2011 and devise modifications to finally develop a revised version for 2020.Among patients diagnosed as having various forms of renal injury between April 2012 and May 2019 at the institutions affiliated to the IgG4-RKD working group, we retrospectively selected those for whom serum IgG4 values and/or data for immunohistological staining of IgG4 in renal biopsy samples were known, and then for study enrollment further selected those with sufficient clinical information, such as serological and radiological data and response to treatment. The IgG4-RKD working group consists of nephrologists, rheumatologists, and pathologists, all highly experienced in the diagnosis of IgG4-RKD. For each case, the physicians reported their degree of diagnostic confidence as “uncertain”, “slightly confident”, “confident” or “very confident” for IgG4-RKD or a mimicker, along with the clinical, serological, radiological, and histological data. Among those initially selected, only patients for whom investigators were “confident” or “very confident” about the diagnosis were included in this study, i.e., diagnoses made by physicians associated with the IgG4-RKD working group (experts) were regarded as the diagnostic gold standard for this study.Thereafter, the included patients were classified as having “definite”, “probable”, “possible” or “unlikely” IgG4-RKD using the IgG4-RKD 2011 diagnostic criteria, and the results were compared with the expert opinions.t test, Mann–Whitney U test or Wilcoxon signed-rank test, and the significance of differences in frequencies was analyzed using Fisher’s exact probability test. Data are presented as means ± SD for continuous variables and as frequencies and percentages for categorical variables. A probability of P < 0.05 was considered to indicate statistical significance. Statistical analysis was performed using SPSS version 19 software .The significance of differences between groups was determined using the paired Student’s n = 8; microscopic polyangiitis (MPA) 5 patients, eosinophilic granulomatosis with polyangiitis (EGPA) 3 patients], idiopathic tubulointerstitial nephritis (TIN) (n = 5), drug-induced TIN (n = 5), nephrosclerosis (n = 4), Sjögren’s syndrome (n = 4), sarcoidosis (n = 3), multicentric Castleman’s disease (MCD) (n = 3), necrotizing glomerulonephritis without ANCA (n = 3), membranous nephropathy (n = 3), tubulointerstitial nephritis and uveitis (TINU) syndrome (n = 2), TIN associated with inflammatory bowel disease (n = 2), TIN with IgM-positive plasma cells (n = 2), TIN associated with infection (n = 2), IgA nephropathy (n = 1), diabetic nephropathy (n = 1), malignant lymphoma (n = 1) and antibody-mediated rejection after renal transplantation (n = 1).Among the 116 collected patients, 105 for whom investigators were diagnostically “confident” or “very confident” were assessed. On the basis of expert opinion, 55 patients were diagnosed as having true IgG4-RKD and 50 as mimickers. Descriptions of patients in both groups are given in Table Patients with true IgG4-RKD were significantly older with a male predominance and showed higher serum levels of IgG4 than mimickers. The frequency of allergy did not differ between the two groups. Most patients with true IgG4-RKD had an elevated serum IgG4 level and dense infiltration of IgG4-positive plasma cells in the renal tissue . On the other hand, 18 of the 50 mimickers (36.0%) had an elevated serum IgG4 level . Dense infiltration of IgG4-positive plasma cells in the kidney was evident in 13 of 40 mimickers . Storiform fibrosis was evident in 28 of the 51 patients with true IgG4-RKD (54.9%) and 3 of the 50 mimickers (6.0%)  . Conversely, no case was classified as “probable” among the mimickers. Therefore, we defined “definite” and “probable” cases based on the IgG4-RKD diagnostic criteria as “IgG4-RKD”, and “possible” and “unlikely” cases as “non-IgG4-RKD” in this study.As a result, 40 of the 55 patients with true IgG4-RKD were classified as IgG4-RKD (39 definite and 1 probable): i.e., the sensitivity was 72.7%  and 45 of the 50 mimickers were classified as non-IgG4-RKD (15 possible and 30 unlikely): i.e., the specificity was 90.0% .n = 3), MCD (n = 1) and malignant lymphoma (n = 1).Table The results of the validation study and the comparison with CD for IgG4-RD highlighted the points below as potentially problematic. We developed some revised versions of the criteria and examined the sensitivity, specificity, positive likelihood ratio, and negative likelihood ratio for each revised version in the present cohort. On this basis, we selected the version showing the best performance  as the IgG4-RKD criteria for 2020.In the CD criteria for IgG4-RD 2011 , the patIn extra-renal organ(s), we changed the term “(a) and/or (b)” in the previous version to “(a) and (b)”, according to the other organ-specific classification criteria .In the present cohort, “storiform fibrosis” in the renal pathology was evident in 54.9% of patients with true IgG4-RKD, which was a lower frequency than expected Table . FourteeIn the IgG4-RKD 2011 diagnostic criteria, histological evidence in the affected organ(s) was mandatory for the diagnosis of the extra-renal lesion(s). However, some characteristic clinical or radiological findings have been shown to be quite useful for the diagnosis of IgG4-RD, as is the case for histological features. For example, the clinical finding “two or more sets of the bilateral lacrimal, parotid, sublingual or submandibular glands swellings” or the radiological finding “diffuse pancreas enlargement and capsule-like rim with decreased enhancement” are considered to be highly suggestive for IgG4-RD, even in the absence of histology . Table 5C-1; (1) bilateral lacrimal gland swelling or (2) bilateral submandibular or parotid gland swelling or (3) imaging findings compatible with type 1 autoimmune pancreatitis, or 4) imaging features of retroperitoneal fibrosis.C-2; (1) and (2), or (3), or (4).C-3; (1) or (2), or (3).C-4; (1) and (2), or (3).The sensitivity and specificity of the modified classification criteria for IgG4-RKD using each pattern were: (C-1) 90.9% and 90.0%, (C-2) 87.3% and 90.0%, (C-3) 85.5% and 90.0% and (C-4) 80.0% and 90.0%, respectively.Table An elevated serum IgG4 level and infiltration of abundant IgG4-positive plasma cells have been considered features of ANCA-associated vasculitis, including MPA, MCD, and malignant lymphoma –9, and iThe revised version of the diagnostic criteria for IgG4-RKD 2011 (i.e. the diagnostic criteria for IgG4-RKD 2020) are shown in Table Using the revised criteria, 48 of 55 true IgG4-RKD cases were classified as “definite”, 2 as “probable”, and 5 as “possible”, i.e. the sensitivity improved to 90.9%. The performance among the mimickers was unchanged from the previous version , i.e. the specificity remained at 90.0%.The revised version of the diagnostic algorithm for IgG4-RKD  is shown in Fig. The kidney is one of the organs most frequently affected in IgG4-RD . AlthougTo test some revised versions, we prepared several patterns  and examined the sensitivity and specificity of each. With regard to infiltrating IgG4-positive cells in the renal histology, we confirmed that “(a) IgG4-positive plasma cells > 10/hpf and/or (b) IgG4/IgG-positive plasma cells > 40%” was more appropriate than “(a) and (b)” for diagnosis of IgG4-TIN, as had been previously suggested by Kawano, et al. . In addiWith regard to “imaging or clinical findings compatible with extra-renal organ(s)”, we selected plan C-1  based on the results for the positive likelihood ratio and negative likelihood ratio. However, it should always be borne in mind that such imaging or clinical features might be evident in mimickers. In particular, retroperitoneal fibrosis as an imaging feature has been demonstrated in ANCA-associated vasculitis or malignant lymphoma , 15, 16.In summary, we have validated the diagnostic criteria for IgG4-RKD 2011 and proposed a revised version (the diagnostic criteria for IgG4-RKD 2020). Use of the revised criteria improved the sensitivity to 90.9%, while the specificity remained at 90.0%, being unchanged from the previous version. Prospective studies to validate the revised version in a separate cohort will be necessary.


Respiratory tract infections (RTIs) account for a large part of antibiotic prescriptions in primary care. However, guidelines advise restrictive antibiotic prescribing for RTIs. Only in certain circumstances, depending on, e.g., comorbidity, are antibiotics indicated. Most studies on guideline adherence do not account for this. We aimed to assess guideline adherence for antibiotic prescribing for RTIs as well as its variation between general practices (GPs), accounting for patient characteristics. We used data from electronic health records of GPs in the Netherlands. We selected patients who consulted their GP for acute cough, rhinitis, rhinosinusitis or sore throat in 2014. For each disease episode we assessed whether, according to the GP guideline, there was an indication for antibiotics, using the patient’s sociodemographic characteristics, comorbidity and co-medication. We assessed antibiotic prescribing for episodes with no or an unsure indication according to the guidelines. We analysed 248,896 episodes. Diagnoses with high rates of antibiotic prescribing when there was no indication include acute tonsillitis (57%), strep throat (56%), acute bronchitis (51%) and acute sinusitis (48%). Prescribing rates vary greatly between diagnoses and practices. Reduction of inappropriate antibiotic prescribing remains a key target to tackle antimicrobial resistance. Insight into reasons for guideline non-adherence may guide successful implementation of the variety of interventions already available for GPs and patients. Increasing antimicrobial resistance is globally recognized as a major threat to human health . AntimicThe evidence of effectiveness is collected and summarized in recommendations for physicians in clinical practice guidelines. These guidelines support physicians in their decision on whether or not to prescribe an antibiotic. Guidelines also provide advice for specific patient groups, such as for children, elderly patients or patients with a poor immune response. Most studies assessing antibiotic prescribing do not take into account these patient characteristics which may justify prescription of antibiotics for certain indications ,5. An exThe Netherlands, the setting of this study, has maintained a relatively low antibiotic prescribing in primary care compared with other countries, with a corresponding low antimicrobial resistance rate ,8. The DThe aim of the present study was to assess: (i) Guideline adherence to antibiotic prescribing for RTIs in the Netherlands, taking into account patient characteristics, comorbidity and co-medications that may justify prescribing of antibiotics according to guidelines; (ii) practice variation, while accounting for patient characteristics, comorbidity and co-medications.Data from the Nivel Primary Care Database  ;allergic and non-allergic rhinitis ,acute rhinosinusitis acute sore throat .The study sample consisted of the 206,473 registered patients from the 307 selected general practices (136 to 3247 per practice), who, according to Nivel-PCD, contacted their GP in 2014 for: We excluded episodes with an ICPC-code for which an antibiotic is indicated according to the guideline . For the included diagnoses, the indication for antibiotics depends on, for example, the patient’s age and comorbidity . For each selected episode of illness, we assessed whether or not an antibiotic (ATC subgroup J01) had been prescribed using Nivel-PCD prescription data, linking prescriptions to episodes based on dates and ICPC codes. This outcome (antibiotics: Yes/no) was used as the dependent variable. First, descriptive statistical analyses were performed in order to gain insight into numbers of relevant illness episodes, the indication for antibiotics and the GP’s prescription of antibiotics. Next, logistic multilevel regression analyses were performed to estimate practice variation. For each ICPC code, and separately for episodes with no or an unsure antibiotic indication, we assessed the practice variation in antibiotic prescribing. The models contained two levels, as the data are hierarchically structured with patients nested in general practices. A multilevel analysis takes into account the nested structure of the data as well as the differences in the number of patients per practice. As the majority of patients (87%) had one episode per ICPC, we only analysed the first episode per patient per ICPC. We included the GP’s prescription of antibiotics as outcome variable (an antibiotic prescribed yes or no), and patient’s age and sex as control variables. The level of significance was set at 0.05. All analyses were performed using STATA, version 14.0.We included 248,896 RTI episodes with no or an unsure indication for antibiotics according to the guidelines of 206,473 unique patients. Acute upper respiratory tract infections and cough had the highest prevalence . This is in line with other European studies ,17,18,19Moreover the wide 95% practice range suggests that there is room for improvement, although antibiotic prescribing to RTIs in the Netherlands is generally lower than in other countries. Over the past years many interventions to reduce inappropriate prescribing of antibiotics for RTIs have been developed, tested and evaluated ,21. ReceThe study of practice variation helps to pinpoint practices with high antibiotic prescribing to target interventions as well as practices with low antibiotic prescribing. This allows to find out which modifiable factors are associated with low and high antibiotic prescribing, such as the level of shared decision making that GPs use  or the rNotably, we found that antibiotic prescribing was nearly as high in situations where antibiotics could be considered according to the guideline as in situations with no indication for antibiotics. This may indicate that additional patient related factors, that were beyond the scope of this study, play an important role in the decision to prescribe an antibiotic. An example of such factor may be patient preference, or the GP’s perception of the patient’s preference ,27,28,29The main strength of the current study is that we used data from a large nationally representative database from which antibiotic prescribing as well as relevant patient characteristics such as comorbidities, co-medication and CRP measurements could be retrieved. Using a registration database has the downside that the results depend on the accuracy and completeness of coding by the participating GPs. Differences in coding between GPs may partly explain the large practice variation between GPs we observed. Some GPs may for example code bronchitis (diagnosis) as acute cough (symptom). This may affect antibiotic prescribing rates in these practices. However, nationally there has been a lot of attention for completeness and quality of coding by GPs, and this resulted in improved diagnostic coding in the years before this study, so we do not expect this to have a large impact on our results .While we were able to assess many patient characteristics, we had no information on disease severity and prolonged or recurring fever. According to guidelines, these factors may justify the prescribing of antibiotics, even in patients and for diagnoses for which antibiotics are usually not recommended. As a result disease severity and fever may greatly influence the decision to treat an RTI with an antibiotic. This implies that we overestimated the percentage of antibiotic treatment in RTIs without an antibiotic indication. However, we do not expect the percentage of patients with severe disease or fever to differ greatly between practices, so this does not explain the wide 95% practice range. Indeed, a study by Stuart et al. showed that the difference in patient’s illness severity scores account for only a minority of RTI antibiotic prescribing variation between general practices .The current study focused on practice variation in antibiotic treatment for RTIs in which antibiotics are not indicated or possibly indicated. We did not assess antibiotic prescribing for RTIs for which antibiotics are indicated, such as pneumonia. A study on guideline adherence in RTIs for which antibiotics are indicated may reveal cases of undertreatment with antibiotics. Furthermore we did not assess symptom relief or other outcomes of patients who did or did not receive an antibiotic. Therefore we cannot conclude whether the great observed practice variation led to different patient outcomes.The current study suggests that, although antibiotic prescribing for RTIs in the Netherlands is low compared to many other countries, improvement is possible and warranted particularly for bronchitis, sinusitis, strep throat and acute tonsillitis. Reduction of inappropriate antibiotic prescribing remains a key target to tackle the global threat of antimicrobial resistance. To further improve guideline adherence, insight into reasons for non-adherence is required to guide improvement and successful implementation of the variety of interventions that are already available for GPs and patients. We can benefit from practice variation by studying differences between high and low antibiotic prescribers.


Methionine is a proteogenic sulfur amino acid with a vital role in intermediary metabolism. However, excess Methionine (Met) intake is toxic, leading to hyperhomocysteinemia. Betaine supplementation effectively ameliorates biochemical abnormalities. However, a lack of genetic information hinders the understanding of the mechanisms underlying methionine excess-mediated effects and whether Bet can effectively suppress these effects in geese. This study was performed to evaluate the effects of excess methionine on growth performance, serum homocysteine levels, apoptotic rates, and Bax and Bcl-2 protein levels in geese and to study the role of betaine in relieving excess Met-induced hyperhomocysteinemia. It was found that excess methionine reduces body weight induced by myocardial apoptosis, and betaine can be used to effectively lower plasma homocysteine levels.p < 0.05). The serum homocysteine levels in geese at 70 d of age in the detoxification group were significantly lower than those in the Met toxicity group (p < 0.05). Compared with the control, Met significantly increased cardiomyocyte apoptosis rates, while Bet reduced them. In conclusion, our results suggest that excess methionine reduces body weight induced by myocardial apoptosis, and Bet can be used to effectively lower plasma homocysteine levels.The objective of our study was to investigate the effects of excess Methionine (Met) on the growth performance, serum homocysteine levels, apoptotic rates, and Bax and Bcl-2 protein levels in geese and to study the role of Bet (betaine) in relieving excess Met-induced hyperhomocysteinemia (HHcy). In this study, 150 healthy male 14-day-old Yangzhou geese of similar body weight were randomly distributed into three groups with five replicates per treatment and 10 geese per replicate: the control group (fed a control diet), the Met toxicity group (fed the control diet +1% Met), and the Bet detoxification group (fed the control diet +1% Met +0.2% Bet). At 28, 49, and 70 d of age, the geese in the Met toxicity group had significantly lower body weights than those in the control group ( Methionine (Met) is a sulfur amino acid (SAA) with a vital role in intermediary metabolism ,2. ObserHomocysteine (Hcy) is a nonproteogenic SAA generated by the intrahepatic transmethylation of dietary Met . Hcy funThe feature of atherosclerotic lesions is apoptotic cell death in both animals and humans ,13. It hExcess Met causes marked growth depression in birds. In general, considerable evidence has revealed that excess Met is toxic to many vertebrates, including humans , mice 1, and pouThe animal study was approved by the Institutional Animal Care and Use Committee (IACUC) of the Yangzhou University Animal Experiments Ethics Committee under permit number SYXK(Su) IACUC 2019-0029. All geese experimental procedures were performed according to the Regulations for the Administration of Affairs Concerning Experimental Animals approved by the State Council of the People’s Republic of China.DL-Met. The geese were fed in separate plastic-floored pens with 2 cm2 square holes that were laid 70 cm above the ground. All manure was cleaned at the end of the trial. All geese were fed and watered ad libitum for 56 d. Water was provided in a half-open plastic cylindrical water tank, and the feed was provided in feeders on one side of each pen. The geese were reared indoors under similar environmental conditions : 65.5 ± 5.0%; lighting period: 16 h; space allocation: 0.5 m2/gander).This study was conducted with 150 healthy male Yangzhou geese at 14 d of age from a commercial hatchery . All of the geese had similar body weights (BWs) 473 ± 0.36 g) and were randomized into three groups that included five replicates per treatment and ten geese per replicate. A basal corn/soybean meal diet with two stages (14–28 d and 29–70 d) was formulated to meet the nutritional needs of the geese National Research Council (NRC), 1994 . The con g and weFeed intake (FI) by pen was measured on a daily basis, and BW was recorded at 14, 28, 49, and 70 d of age. The average daily feed intake (ADFI), average daily gain (ADG), and feed-to-gain ratio (F/G) were calculated from 14 to 28 d and from 29 to 70 d of age, and mortality was recorded as it occurred.Calculations:ADFI = feed consumption/feeding days in the whole periodADG = total weight gain/feeding days in the whole periodF/G = ADFI/ADGWhen the geese reached 70 d of age, 2 geese from each treatment replicate  were randomly selected for blood collection from their wing veins.Blood drawn from wing veins was cooled in ice water and centrifuged for 10 min at 4500 rpm to obtain plasma for the measurement of biochemical indices. The plasma was stored at −20 °C until analysis. The plasma Hcy concentrations in peripheral blood were determined according to the previously described methods of Feussner et al. . The thi6 cells/mL. Then, aliquots of 100 μL of cell suspension (1 × 105 cells) were incubated with fluorescein isothiocyanate (FITC)-annexin V and propidium iodide for 15 min at room temperature in the dark. The apoptotic rate was analyzed using flow cytometry(Thermo Fisher Technology (China) Co., Ltd, Shanghai, China) within 1 h.An annexin V apoptosis detection kit  was utilized to measure apoptosis in heart tissue following the manufacturer’s instructions. After treatments, cardiomyocytes were washed twice with cold phosphate-buffered solution (PBS), trypsinized, and then resuspended in binding buffer at a concentration of 1 × 10Total protein lysates were collected for standard immunoblot analysis. The protein concentrations were determined by bicinchoninic acid (BCA) protein assay(Thermo Fisher Technology (China) Co., Ltd, Shanghai, China). Aliquots of protein lysates (30 mg/lane) were loaded into gels for sodium dodecyl sulfide-polyacrylamide gel electrophoresis (SDS-PAGE), and the separated proteins were transferred to a polyvinylidene fluoride (PVDF) membrane(Thermo Fisher Technology (China) Co., Ltd, Shanghai, China). The membrane was blocked and incubated with Bcl-2 and Bax antibodies overnight at 4 °C. The Bcl-2 and Bax antibodies were purchased from AmyJet Scientific  . The blots were washed with TBST  and incubated with corresponding horseradish peroxidase-conjugated secondary antibodies . Finally, the blots were visualized with enhanced chemiluminescence and quantified by densitometry.p < 0.05 was considered to indicate statistical significance.The data are expressed as the means ± standard deviations (SDs) and were subjected to Kolmogorov-Smirnov (KS) testing to confirm normality. Different groups were compared using one-way analysis of variance (ANOVA) followed by the Student-Newman-Keuls post hoc test with SPSS 17.0 . p > 0.05), and there was no treatment effect between groups. The effects of Bet on the growth performance of geese with HHcy are shown in p < 0.05). There was no significant difference in BW at 70 d between the Met toxicity group and the Bet detoxification group (p > 0.05). The ADFI and ADG for geese at 14–28 d of age were significantly higher in the Met toxicity group and the Bet detoxification group than in the control group (p < 0.05).The data for mortality at 70 d were transformed before analysis. Mortality was greater in the Met toxicity group than in the control group and the Bet detoxification group (p < 0.05). The Hcy levels in geese at 70 d of age in the detoxification group were significantly lower than those in the Met toxicity group (p < 0.05) but were not significantly different from those in the control group (p > 0.05). The serum Hcy level was a sensitive index for the identification of geese with HHcy. The Hcy values for the geese in the control group were all between 16 and 21 μmol/L, while the Hcy values for the geese in the Met toxicity group were all higher than 30 μmol/L.The effects of Bet on serum biochemical indices of HHcy in geese are shown in p < 0.05), but the Bet detoxification diet reduced the apoptosis rates.The effects of Bet on cardiomyocyte apoptosis in geese with HHcy are shown in p < 0.05) (The effects of Bet on the expression of apoptosis-related proteins in geese with HHcy are shown in  < 0.05) . There wExcess Met causes marked growth depression in birds, and very high levels of Met can lead to HHcy. However, several studies have suggested that dietary Bet might alleviate HHcy. In the present study, we investigated the effect of Bet on Met loading-induced elevations in plasma Hcy levels. We first established a model of HHcy in geese via Met loading. In previous Met loading studies, Met has been administered orally to humans ,23 and rDL-Met and excess L-Met are toxic to starter Pekin ducks and that both Met sources exert equivalent growth-depressing effects. Our results are in agreement with the findings of many previous studies on the toxicity of Met and its hydroxy analog in broilers [In this study, the BWs of geese at 28, 49, and 70 d of age were significantly lower in the Met toxicity group than in the control group, and Bet did not improve growth performance in geese at 28, 49, and 70 d of age. In previous studies, excess levels of Met or a Met hydroxy analog have been found to be toxic and to cause growth depression in ducks  and broibroilers . At presbroilers  and ratsbroilers . It is ubroilers .DL-Met supplementation can lead to HHcy in geese. Hyperhomocysteinemia (HHcy) was defined as mild HHcy (Hcy: 15–30 μmol/L), moderate HHcy (Hcy: 30–100 μmol/L), and severe HHcy (Hcy > 100 μmol/L) by measuring the level of Hcy in human blood [Hcy is a nonessential, thiol-containing and potentially cytotoxic 4-carbon α-amino acid formed through the demethylation of Met during Met metabolism . Traditian blood ,30. The Several studies have suggested that dietary Bet might be a determinant of plasma Hcy concentrations ,32,33. SThe Hcy-lowering effect of Bet can most likely be ascribed to an increase in Bet-dependent remethylation owing to increased Bet availability and enhanced activity of the enzyme Bet methyltransferase in the liver and kidneys. This possibility is supported by the results of studies by Barak et al.  and SaarApoptosis plays an essential role in maintaining cellular homeostasis during development, pathophysiological, and differentiation processes in multicellular organisms . HHcy isPrevious studies have demonstrated that the process of apoptosis is mediated by proteins in the Bcl-2 family and is accomplished through the Fas pathway or the Caspase-dependent apoptotic pathway, which relies on active mitochondrial control . Once thp < 0.05), but the presence of Bet reduced the apoptosis rates of these cells. The hearts of Bet-supplemented geese had reduced absolute numbers of apoptotic cells, showing that Bet administration appears to reverse the biochemical alterations associated with heart apoptosis.Elucidation of the metabolic events linking excess Met intake to pathology and their dependence on other underlying metabolic processes and pathologic conditions will provide crucial insights into the roles of these important diet-dependent pathways in cardiovascular health and disease. Overall, our results indicated that the supplementation of feed with 1% Met decreased growth performance and increased plasma Hcy levels, indicating that excess Met can lead to HHcy. Bet was able to effectively lower fasting plasma Hcy levels and to prevent plasma Hcy levels from increasing after Met intake. Met toxicity significantly increased the apoptosis rates of cardiomyocytes (


Uncomplicated urinary tract infection (uUTI) is very common among women in primary care. The risk of developing pyelonephritis remains low after uUTI, nonetheless, empiric antibiotic therapy is frequently prescribed for symptomatic purposes. This may lead to adverse effects and antibiotic resistance. Furthermore, patients may express the will to limit the use of antibiotics. Some European countries recommend discussing a delayed prescription with the patient and developing a shared decision. The aim of this study is to create a patient decision aid (PtDA) used in primary care settings to make a shared decision between practitioners and women about whether or not to treat uUTI with antibiotics.We followed the steps recommended by the International Patient Decision Aids Standards, with a scoping phase, a design phase (including focus groups and literature review), and an alpha-testing phase. A steering group, made of patients and physicians, met throughout the study to develop a prototype PtDA.The information included in the PtDA is the definition of uUTI, information on the options, their benefits, risks, and consequences, based on a review of the literature. The results of the focus group made possible to determine the patient’s values and preferences to consider in decision-making, including: the discomfort felt, the impact on daily life, patients’ perceptions of antibiotics, and the position relative to the risk of adverse effect. The choices in presentation, organisation and design are the result of the work of the steering group, improved by feedback from alpha testing. We confirmed the need for shared decision-making and the equipoise in this situation.We developed a PtDA to be used in primary care for sharing decision on the use of antibiotic in uUTI. It needs to be validated in a beta-testing phase, with complementary advice from peers, and then tested in a clinical study comparing its use with the systematic prescription approach. Uncomplicated urinary tract infection (uUTI), or cystitis, is among the leading reasons for infectious disease consultation in primary care . Nearly There is no standard definition of uUTI internationally accepted. In France, uUTI in women is defined as an acute urinary tract infection in adult under 75 years-old, without signs of pyelonephritis or risk factor for more severe disease .Acute cystitis affects several facets of women’s life: social and intimate relationships, self-esteem, and ability to work. Depending on the situation, the impact can vary from a woman to another: from small discomfort to severe disability impacting personal and/or professional life , 4. NoneCurrently, the systematic and immediate prescription of empiric antibiotic treatment is often recommended to reduce symptoms , even thSociété de Pathologie Infectieuse de Langue Française) recommends fosfomycin  [40 [4010] .“It restricts social life, because you always have to be near a bathroom (laughter)” (P3.2), “You don’t dare to have sex anymore” (P4.4).Participants spoke of their personal or reported experiences with cystitis and their impact on their social and sexual lives: “It’s a feeling of guilt, actually. Well, in a way it is, because we feel that our intimacy as women is being attacked” (P3.4).This experience touched on intimacy and could be perceived as taboo: “If there’s blood in my urine it means that the kidneys must be affected” (P3.2), “It’s going to make an infection maybe a bit generalized in that area, maybe causing problems to have children...” (P1.2).They feared that cystitis could be complicated by renal, gynaecological, or fertility problems: “I told my mother about it and she said: don’t worry, drink lots of water, it will pass, this antibiotic works well; because she often had it” (P2.2).Their knowledge about the risk factors and treatment of cystitis was part of a lay knowledge shared amongst women: “The doctor or the ones I saw, made me feel like it was my fault because I didn’t wash (myself) well. Afterwards, we are told once we hold it back! We don’t have to be told every time” (P3.2).Some participants described a feeling of infantilization and guilt during the consultation with the physician: “What is important with cystitis, I learned from the doctor who took the time to explain it to me. [...] We are not in a normal state when we are sick. So he really needs to listen to us” (P2.5).They wanted a personalized exchange, where they could express their experiences: “And in rather simple terms, so that everyone can understand it... Not in doctor’s language” (P1.3). They suggested a pictorial presentation: “The more graphic, the more people are affected” (P1.3), accompanied by the doctor: “The diagram is nice, but if the doctor doesn’t explain it to you, [...] she won’t understand anything” (P3.3).The participants wished to clearly define cystitis and its risk factors, with a vocabulary accessible to all, without medical jargon: “Isn’t there something more effective and long-lasting, [...] rather than just immediately stopping the pain?” (P1.2).Their expectations of treatment could be the rapid relief of symptoms, or the prevention of recurrence in the longer term: “Do you have something to offer me that is not antibiotics? I have time now, I can stay at home, if it’s really not going well we’ll switch to antibiotics, but why don’t we test something else? Maybe there should be a second option” (P4.4).Some patients have expressed an interest in being involved in the decision related to the antibiotic: “This antibiotic was really a miracle” (P6.1), “Every antibiotic [...] that we swallow, we know that there are side-effects” (P1.1).The action of the antibiotic was seen as magical, but could lead to side effects and resistance: “Having the choice between a chemical molecule and something a bit more natural, something less harsh, I’ll take what is less harsh” (P2.3).The participants considered alternative treatments, described as natural, such as cranberry or hydration: The draft version of the prototype Fig.  includedThe title explicitly described the decision of whether to take an antibiotic or not. The elements of the PtDA were then arranged according to these two choices.The common symptoms and etiology of uUTI were briefly described and illustrated with a diagram of the bladder.The treatment options that were presented included antibiotic treatment, hydration, and cranberry. The practical modality of a single-dose antibiotic therapy was specified.The average duration of symptoms, the risks of recurrence, resistance and complications were detailed according to whether the antibiotic was taken or not.Coloured pictograms numerically represented the evolution of symptoms after three days according to the choice of treatment and the incidence of adverse effects.The patient values to report were physical discomfort, the impact on their daily life , their general opinion on antibiotics and their adverse effects. A free space allowed for the collection of additional value.Deliberation was facilitated by sliders polarized according to the two options, for each expressed value. A final slider helped in the decision-making process.The chosen format of the PtDA was a double-sided A4 sheet of paper. It was intended to be used during a discussion with the physician during the consultation and not by the patient alone.The results of the alpha-testing phase and the second meeting of the steering group are presented in Table The literature review showed a need to increase patient’s involvement in the decision-making process for uUTI treatment. Patient’s feedbacks during focus groups confirmed this need  7]. The. The7]. Hence, we developed a prototype of PtDA to help physicians and patients to make a shared decision regarding antibiotic treatment for uUTI. This prototype aims to provide support during consultations, in addition to the information delivered orally by the physician .Of note, we recruited more patients face-to-face during medical consultations than through patient support and community groups. The ease of face-to-face recruitment was previously described in the literature . MoreoveThe medium-sized focus groups in our study allowed rich and varied exchanges, including those involving intimate topics, as already described in the literature, where a focus group of four to six participants has previously been documented to facilitates communication . The ageSome elements collected from the focus groups corresponded with the existing literature, such as representations of uUTI and its risk factors , represeSome of these representations correspond to known risk factors , however, the steering group decided not to mention risk factors because they were not directly involved in the decision-making process. Other representations were beliefs (risk of infertility) that were absent in the literature. Some patients wanted to be able to discuss them. This information could be added in a leaflet handed over to the patient.Most of the patients’ values expressed in the focus groups could be integrated into the PtDA, particularly in the slider. An empty slider allows the patient to express additional values, like her expectations regarding treatment .One concern expressed among the interviewed patients and doctors was the risk of pyelonephritis. There is little data on the natural course of uUTI without antibiotics, and the meta-analysis comparing antibiotics to placebo did not show a significant increase in the risk of pyelonephritis between the two . HoweverSome of the interviewed physicians were concerned that the use of the PtDA would lengthen the consultation. There is little evidence on the impact of shared-decision making on the length of consultations , but a sA noticeable limitation of this PtDA is the lack of medical perspectives exterior to the study. The number of interviews with physicians during the alpha-testing phase was low. As already reported in a Cochrane review, during PtDAs development, patients’ views were more often collected than those of physicians .The steering group chose to present the probability of symptoms after three days. This time frame made possible to present data in the PtDAs regarding the first-line antibiotic in France (fosfomycin) and placebo. This short delay is in line with the French guidelines which recommend another consultation after three days in case of treatment failure, but also with the British guidelines proposing a 48-h delay for the delayed prescription of antibiotics. Indeed, delayed prescribing could reduce antibiotic use . This opOur PtDA was created in line with international standards and will soon be beta-tested in a larger study . Helped The possibility to implement this approach in France will also be studied during the beta-test. Indeed, there may be some difficulties to implement the PtDA in clinical practice. The PtDA leads to a discussion with the patient about the choice of using antiobiotic or not, whereas actual French guidelines advise a systematic antibiotic prescription. Furthermore, systematic antibiotic prescription requires less investment from the practitioner. Nevertheless, practitioner investment can lead to better outcomes: if the practitioner is optimistic about diagnosis and prognosis, symptoms could resolve faster . Thus, iuUTI is no longer considered as a pathology for which the symptomatology is experienced similarly for all patients and for which a single treatment should be offered. The literature review suggests that uUTI is an equipoise situation, hence, shared-decision making seems to be the best way to broach this subject. Developing a tool to help for shared-decision making in primary care consultation is necessary to move forward and to assess its efficiency compared to a systematic antibiotic treatment approach. To do so, we developed one of the first PtDA for uUTI treatment in primary care, in line with international standards.The impact of our PtDA on patients’ satisfaction and antibiotic prescription remains to be evaluated, but its conception has already brought a lot of information on patients’ and practitioners’ perceptions of uUTI.