Yet, 80%–90% of antibiotic use in humans occurs in outpatient settings, and the Centers for Disease Control and Prevention (CDC) estimates that nearly 50% of this prescribing is inappropriate. The Joint Commission recently introduced ASP requirements for accredited ambulatory care organizations.Although antibiotic stewardship programs (ASPs) have an established role in the acute-care setting, ASPs remain nascent in outpatient settings. Several institutions have investigated the accuracy of prescriber-selected indications for inpatient antibiotic orders, but none have assessed the accuracy of outpatient antibiotic indication fields. The objective of this study was to validate the accuracy of the indication field upon antibiotic prescribing in the ambulatory care setting so this field may be used to for benchmarking to facilitate future stewardship interventions.It is vital for institutions to assess their general prescribing practices to set a foundation for future stewardship efforts. The use of antibiotic indication fields within computerized prescriber order entry (CPOE) is advocated for by the CDC to facilitate benchmarking and communication.This retrospective quality assurance project was conducted within an existing ASP to validate the accuracy of a mandatory antibiotic indication field when prescribing outpatient antibiotics. This study was conducted at an academic medical center with multiple ambulatory care clinics, urgent care clinics and an emergency department (ED). The primary end point was the proportion of indication fields that accurately reflected the reason the antibiotic was prescribed. We utilized an electronic health record (EHR)–specific reporting tool, Epic Reporting Workbench , to identify patients aged ≥18 years who received an outpatient oral antibiotic prescription between September 1, 2021, and October 22, 2022. Investigators extracted the antibiotic order date, prescriber name, drug name, dose, directions, duration, and selected indication from the medical record. The mandatory indication field includes checkboxes for common outpatient infectious syndromes listed in Table Corresponding chart notes were reviewed by an investigator (J.P.) for the outpatient encounter associated with the prescription. If the infection documented in the chart note from the ambulatory visit was consistent with the indication selected upon ordering the antibiotic, then the use of the indication field documentation was considered accurate. In addition, suboptimal use was characterized and defined as the selection of the “other” indication field when there was a more appropriate default option available but entered free text matched the chart documentation. Descriptive statistics were used. This study was approved by the University of Illinois at Chicago Institutional Review Board.Among 4,323 outpatient antibiotic prescriptions that met the inclusion criteria, we selected 350 for validation through random sampling. All prescriptions had a corresponding chart note that clearly identified the antibiotic indication. Table Our study is the first to evaluate the accuracy of a required indication field when prescribing antibiotics in the outpatient setting. Overall, 92.5% of selected indications were accurate and matched the infectious syndrome on chart documentation. Accuracy was similarly high among the ED and other outpatient settings, and these indication data can serve as a foundation for benchmarking and systematic stewardship interventions.Through the course of this review, we identified ways to optimize the indication field listing. Half of the indication inaccuracies were due to prescribers selecting “genital tract infection” instead of “urinary tract infection” for patients with UTIs. Therefore, we have eliminated “genital tract infection” and replaced it with “sexually transmitted infection” to further direct prescribers to the appropriate choice. To provide additional actionable data, we also changed “urinary tract infection” to “UTI, pyelonephritis” and “UTI, cystitis,” and we split “otitis/sinusitis” into 2 unique headings. Furthermore, because the “surgical site infection” indication was selected for only 1 patient and because these infections can be captured by the “skin and soft-tissue infection” indication, we eliminated the former indication. These researchers reported that 43 (86%) of the 50 randomized CPOE indications were accurate, with most inaccurate designations being “other.” With the addition of an indication to account for the selection of “other,” it was estimated that accuracy would have improved to 92%. This validation study allowed for additional benchmarking and assessment institutional guideline adherence.Prior studies have evaluated the accuracy of the indication field for inpatient antibiotic orders. One of the earliest was conducted at a large academic medical center and evaluated indications entered via CPOE. assessed the accuracy of prescriber-selected indications in a pediatric ICU and adult medicine step-down unit. This analysis evaluated the frequency at which antibiotic indications chosen by the prescriber matched the true indications 48–72 hours after the initial order. The study reported a 90% concordance between the provider selected indication and independent review by infectious disease physicians. Similar to our study, one of the highest rates of discordance was seen with genitourinary and urinary tract infections, but this finding was not further elaborated.Heil et al reported a method for monitoring prescribing patterns by generating indication lists for selected broad-spectrum antibiotics. Indication matching was determined by ensuring the chart-documented diagnosis mirrored the indication in the order. The matching rate was 74%–80%. Notably, prescribers chose “other” for 41% of orders, but in most cases the true indication was not available on the prepopulated indication list. Timmons et al attempted to characterize clinical appropriateness of prescribing as well, and they reported that 13.6% of antibiotic orders were inappropriate. This study sheds light on the scope of potential opportunities to assess clinical appropriateness with validated indication fields.In another inpatient medical center, Timmons et al focused on respiratory tract infection and UTIs in 3 hospitals in the Netherlands. Similar to our study, prescription indications and chart notes were manually screened, and accuracy ranged from 78.2% to 96.7%.A recent study by van den Broek et alThrough our EHR, we can query specific indication fields used and key elements of the prescription including dose, duration, prescriber, clinic, and other information. These data, when combined with our now-validated indication fields can facilitate benchmarking of outpatient antibiotic use and can serve as a foundation from which to investigate opportunities to improve care. Stratifying these data by prescriber and/or department can shed light on prescribing practices amenable to systematic intervention or provider comparisons. Some system-oriented solutions may include generating indication-based order sets, review and modification of specialty-specific preference lists, modification of order defaults, and implementation of clinical decision support. More targeted antibiotic stewardship interventions may include educational efforts to address inappropriate prescribing patterns for specific prescribers or indications.Generalizability across institutions is a limitation of this single-center study; additional data are needed from other outpatient centers. Additionally, appropriate use of the indication fields may change over time, which may warrant routine monitoring of this tool. Notably, not all outpatient antibiotic orders have an associated note. Although we did not encounter this within our study, scenarios in which providers enter orders without EHR documentation may occur.In this retrospective quality assurance project, outpatient prescribers appropriately utilized the mandatory indication function in the CPOE. This project, along with the aforementioned studies, support the importance of a required indication field because it can inform quality care. The indication field offers a method to standardize and support reliable and efficient assessments of antibiotic use. This method provides institutions the opportunity to identify inappropriate antibiotic use and to prompt changes in prescribing practice. Other outpatient centers are encouraged to implement mandatory indication fields for antibiotic orders.


Studies on intracranial aneurysms (IAs) using animal models have evolved for decades. This study aimed to analyze major contributors and trends in IA-related animal research using bibliometric analysis.IA-related animal studies were retrieved from the Web of Science database. Microsoft Excel 2010, GraphPad Prism 6, VOSviewer, and CiteSpace were used to collect and analyze the characteristics of this field.Stroke was the most prolific journal that accepted publications related to IA research using animal models. Circulation has the highest impact factor with IA-related animal studies. Hashimoto N contributed the largest number of articles. Meng hui journal published the first and second highest cited publications. The keywords “subarachnoid hemorrhage,” “macrophage,” “rupture,” “mice,” “elastase,” “gene,” “protein,” “proliferation,” and “risk factors” might be a new trend for studying IA-related animal research.A total of 273 publications were retrieved. All publications were published between 1976 and 2021, and the peak publication year is 2019. Rat model were used in most of the publications, followed by mice and rabbits. Japan (35.5%), the United States (30.0%), and China (20.1%) were the top three most prolific countries. Although China ranks third in the number of publications, it still lacks high-quality articles and influential institutions. Stroke, Journal of Neurosurgery, Neurosurgery, and Circulation for their high-quality IA-related animal studies. Using animal IA models, especially mice, to investigate the molecular mechanisms of IA may be the frontier topic now and in future.Japan and the Unites States contributed the most to IA–related animal studies, in terms of both researchers and institutions. Although China ranks third in terms of the number of publications, it should strengthen the quality of its publications. Researchers should pay attention to the latest progress of  It is asm of IA . Currentsm of IA . Howeversm of IA –10. Prevsm of IA  and scolsm of IA . In addism of IA . TherefoRQ1. The publication trend for IA research using different animals.RQ2. The most contributing authors, institutions, countries, journals, and their cooperative networks.RQ3. The most influential publications in this field.RQ4. The current and frontier topics for IA research using different animals.To avoid bias introduced by database retrieval, we performed an electronic search using the Web of Science (WOS). Expanded database  from its date of inception (1945) to 1 July 2021. The Web of Science  is one of the most professional and authoritative citation-based databases with a powerful indexing function, which contains not only basic bibliometric parameters, including title, author, institution, country/region, and author keywords, but also reference information. Thus, WOS has been widely used in bibliometric studies , 15.*” or “intracranial aneurysm*” or “brain aneurysm*”) and TS= . First, two investigators (Qian Zhang and Jing Liu) independently searched and screened the database. Second, Shifu Li and Jia Chen re-evaluated the obtained publications by reading the title and abstract. Any discrepancies were resolved by discussion until a consensus was reached.To obtain publications primarily focused on animal IA models, an advanced search was performed using the following items: TI =  and clinical outcomes of patients were excluded. Our search was limited to articles published in English.The following bibliometric parameters were extracted: title, keywords, authors, journal, year, citation counts, country, institution, and species of animal . The data were then imported into Microsoft Excel 2010 , GraphPad Prism 6 , VOSviewer , and CiteSpace (version 5.8 R3); then, the major contributors were analyzed, and the knowledge network was visualized , 17.The initial search of the WOS identified 375 publications. After limiting the search to English language articles and excluding irrelevant publications that focused on SAH and that lacked keywords and cited references , 273 publications were obtained. Of the 273 articles, 256 publications were original research and 17 publications were review articles .Concurrence of iris aneurysms and cerebral hemorrhage in hypertensive rabbits.” The authors found that a striking rate (42.5%) of hemorrhagic stroke occurred during the formation of iris aneurysms in hypertensive rabbits , followed by mice (n = 81) and rabbits (n = 41). Approximately 80% of the studies used these three animals. The first publication year of IA research involving rats, mice, rabbits, swine, canines, zebrafish, primates was in 1978  was the study by Meng et al.  could crosstalk with different cells, such as inflammable cells or vascular mural cells, to mediate the initiation, progression, and rupture of IA , and the number of citations per publication was 0.7 . The 10 most cited publications are listed in g et al. , entitleormation . InteresA total of 1022 scholars were involved in the selected IA-related animal studies. A total of 301 institutions participated in IA-related animal studies. The Kyoto University Graduate School of Medicine (Japan) ranked first, with 58 publications , followed by the University of California (USA), with 23 publications, and the National Cerebral Cardiovascular Center (Japan), with 21 publications. Stroke , with 36 publications , followed by the Journal of Neurosurgery , with 22 publications (8.0%), and Neurosurgery , with 11 publications (4.0%). Of the ten most cited publications, four articles were published in Stroke. Circulation has the highest impact factor  in this field.A total of 104 journals accepted IA-related animal studies. https://graphica.app/). Countries in one cluster with the same color were closely cooperated , where, by using a Sprague–Dawley (SD) rat IA model, the authors found that macrophage-derived MMP-2 and−9 could promote the progression of IA , two publications in the 10 most cited articles indicate that Circulation poses a high standard for IA-related animal studies.In terms of the journal, the three most prolific journals were n = 97, 35.5%), the United States , and China . These countries have larger aging populations and higher levels of economic development and may pay special attention to basic research of IA. We believe these countries will continue to do so in future because scientific research can effectively guide national healthcare policies  by authors from Japan and the United States was much higher than that from China .The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


The article presents selected types of phase change materials (PCM) and their properties in terms of applications in various fields of science such as construction and concrete technology. The aim of the article is to present a comparative analysis between the results of the laboratory tests and numerical simulations. The analysis contains two types of PCM (powder and in liquid), which were dosed in a hybrid system to the concrete mix. The purpose of using PCM is to allow the technological barrier to be exceeded in hot and dry climate conditions, enabling the construction of non-cracking concrete structures. The paper presents a parametric analysis of the influence of various modeling elements on the obtained results. The procedure of generating and absorbing heat caused by the applied PCM was also implemented using user subroutine into finite element code (Abaqus/Standard). The numerically obtained results are consistent with the experimental results. The presented results demonstrate that the use of PCM improves the conditions of concrete maturation by reducing the average temperature of the mixture in its entire volume. The process of concreting in dry and hot conditions differs significantly from the performance of these works in a temperate climate. At a concrete temperature above 32 °C, blowing wind and insufficient air humidity intensify the phenomena of accelerated evaporation of water from the concrete mix. The process is exponential, and even small changes in temperature cause a significant loss of water from the maturing young concrete, which reduces its compressive strength and causes the appearance of shrinkage cracks ,2,3,4. TA necessary condition for the continuation of the correct mixture maturation process is to maintain the relative humidity inside the concrete higher than 80%. The essence of maintenance is to keep the concrete saturated with water or as saturated as possible, until the spaces originally filled with water in fresh cement slurry are filled to the desired degree by cement hydration products .If the concrete element is constructed without external barriers, it can freely expand during heating and contract during cooling, without causing stresses . In pracPractical measurements to mitigate these effects include actions such as changes in the formulation of the concrete mix , modifications to the structure design , or component cooling mixes or installation of cooling pipes or replacement of part of the mixing water with crushed ice. Additionally, the use of PCM with the appropriate phase change temperature may be a good solution in reducing the risk of thermal cracking in young concrete ,10,11,12The current article presents laboratory tests showing the effect of PCM on the maturation process of concrete in slabs with dimensions of 52 × 52 × 20 cm. Temperature changes at different points of the slabs were compared for the case without PCM and with different PCM contents. The results of computer simulations are also presented, taking into account the complexity of the conditions in the thermal chamber during the tests, with particular emphasis on the heat that PCM return and absorb during concrete maturation. The results of laboratory and numerical tests were compared.use of PCM pipes in concrete,use of lightweight aggregates impregnated with PCM,adding powder or in liquid PCM microcapsules to the concrete,impregnation of concrete with PCM .In the current research the applicability of PCM is discussed in order to reduce the negative thermal effects in concrete in the initial stage of maturation. PCM are to ensure internal care, preventing the formation of negative thermal gradients inside the manufactured element. Various techniques may be used to add PCM to the concrete mix in order to control temperature changes as it matures. The main methods are the following see :use of PThere are many possibilities of PCM application in individual fields of science and practice: in construction, transport, agri-food industry, automotive, chemical, medicine, and the textile industry. PCM undergo a phase change within a certain temperature range. When the temperature of the mixture reaches the phase transition temperature, the PCM absorb excess heat, they switch into the liquid, and prevent the temperature from exceeding unfavorable levels inside the concreted element. When the temperature drops below the phase transition temperature, they solidify and give up the excess of stored heat, thanks to which they stabilize the temperature inside the concreted element. The described mechanism of action of PCM in the maturing concrete is favored by typical climatic conditions of the Middle East, characterized by a significant diurnal temperature amplitude .When introducing PCM into concrete, the following requirements should be taken into account. First, the introduced PCM should not interact with the mixture components during the cement hydration process. This can negatively affect the properties of mature concrete. Second, the phase transition period of the PCM should fall within the preferred range for the hydration process. This temperature range is from 19 °C to 34 °C. The last requirement is that the PCM has a high heat capacity. The most frequently used PCM to control the temperature of young concrete is microencapsulated Micronal powder ,19,20,21The issues of the use of PCM in construction, with particular use of the temperature of the concrete mix to regulate  in dry and hot climate conditions, has been described in detail in the doctoral dissertation  of the cIn the below described laboratory experiment, two types of PCM were selected for the research. The first is Micronal, an encapsulated additive, which is in the form of a dry powder from BASF . The secwithout  and maintained humidity from 0% RH to 100%. Temperature and relative humidity control system in the climatic chamber shown in It is expected that the excess heat generated in the concrete mix as a result of hydration and the heat from the surroundings during the initial maturation period of young concrete will be absorbed during the melting process of the PCM. Conversely, excessively absorbed heat will be released when the ambient temperature drops below the phase transition temperature of the PCM, i.e., during solidification of the PCM. In this way, the microcapsules are supposed to regulate the temperature of the concrete mix, making the temperature of the maturing concrete uniform and reducing the thermal peak and shifting it in time, as well as enabling the technological barrier associated with quick cement hydration during concreting in dry and hot climates to be exceeded. For the analysis of this phenomenon, cyclic daily cycles of temperature fluctuations (range from 13.6 °C—night to + 43.6 °C—day) were adopted and the response temperatures of two types of samples were recorded.It should be clearly stated that the conducted laboratory experiment strictly relates to the practice of concreting in dry and hot conditions. The production and transport of concrete in a dry climate takes place in the evening and night conditions, so that the laying of concrete takes place at the lowest possible ambient temperatures, to delay the evaporation of water from the concrete mix. With increasing ambient temperature, with sunrise, we are already beyond the hydration thermal peak of young concrete and the addition of phase change materials additionally lowers the hardening temperature and reduces the shrinkage effect in the structure.In the laboratory tests performed, this rule was followed. The concrete mix was prepared at a temperature of about 25 °C and, after shaping three slabs, they were placed in the chamber, which cooled the concrete in the coming hours to simulate the effect of the night chill. The later diurnal cycles simulate—hourly intervals—an increase or decrease in ambient temperature, as occurs in the summer, for example in Damascus.without (P1) and the chamber were at a comparable level (see PCM (P2) and No. 3PCM (P3), the initial temperature in the chamber was at the same level as for No. 1without (P1), but the initial temperature of both the PCM concrete slabs was lower, because when the phase transition temperature is exceeded, the thermal peak in the concrete is reduced and heat is accumulated by the phase change capsules . The initial temperature of the unmodified concrete slab—No. 1evel see . For slaules see .The temperature curve (second slab) quickly lowers after reaching the maximum, which proves that PCM quickly release the absorbed heat to the environment. This feature is very important for the durability of the structure, not only for the structure made in hot climates, but also for the structure erected in Polish climate, e.g., a massive structure. The experimental studies using the previously described test stand , which wThe energy balance of the system according to the first law of thermodynamics can be represented by the following equation ,34,35:, In the finite element method , the following interpolations of the temperature and its rate inside the finite elements are used:In this equation In Equation (8), In the discussed case of transient heat flow, the following backward differential algorithm was used:We write the heat flow equation for the moment Assuming that When solving the above equation using the modified Newton’s method, it should be decomposed into the Taylor series around the approximate solution In order to determine the extent to which all the parameters of the simulation  affect the change and temperature distribution in the concrete slab, computer simulations of the process were carried out in the Abaqus/Standard program. In this work, the focus was only on the simulation of thermal problems and the part related to the strength (displacement) analysis in the potential sequential temperature-displacement analysis was omitted .The regular geometry of the slab (dimensions 0.52 m × 0.52 m × 0.2 m) allows the use of rectangular finite elements (diffusive heat transfer elements). The use of linear elements with the designation: DC3D8 (8-node linear heat transfer brick) and parabolic ones with the designation: DC3D20 (A 20-node quadratic heat transfer brick) was considered. In all kinds of computer simulations, the first point should be to determine the effect of discretization on the results obtained. The influence of the element type along with the mesh density was also analyzed in the current study. The ambient temperature was assumed to be the temperature set by the programmable controller and then measured in the thermal chamber, which is assumed during the experiment to determine the rhythm of daily temperature changes in Middle East (Syria), termined . ConvectThe second analyzed parameter is the coefficient of thermal conductivity For discretization purposes, material constants The temperature distributions are presented in The conducted analyses showed that in this case the size of the finite elements and their type are not of great importance . In furtThe influence of thermal conductivity was analyzed in the study because it determines the rate of temperature changes inside the concrete slab. The coefficient For the smaller The convection phenomenon in which the heat flux The influence of its value on the results was determined, i.e., temperature changes at measurement points for two values of the coefficient The influence of specific heat The presented results show that for a lower specific heat In maturing concrete, heat is generated by an exothermic hydration reaction. As a result, the temperature of the concrete rises. In addition, thermal energy is absorbed and released by the PCM, which reduces the temperature inside the concrete slab. The general model was assumed in the following additive form:The history of temperature changes corresponding to the nce with , that th with [℃ .It should be noted that the experimental curves ,38 deal ux rH(t) . HoweverAnother analyzed issue is the absorption and emission of heat by the phase change material. The two different PCM have different latent heat and a different transformation temperature. The heat flux associated with the change of the physical state i = 1, heat is released but for i = 2 and 3 it is absorbed).In this equation, the parameters HETVAL user subroutine has been programmed to account for the heat generated by the phase change. The procedure interface is as follows  .*Depvar function and later written into Abaqus output files as SDV variables. CMNAME is the material name, TEMP(1) is the current temperature, and The following variables are defined in the subroutine: FLUX(1) is heat flux, The parameters are presented in The above analysis of the influence of the model parameters, those that determine the behavior of the material and those that determine the boundary conditions and the interaction of the concrete slab with the environment , led to The following figures compare the temperatures in the slabs with and without PCM during concrete maturation see  at diffeThe important aspect is also the fact that PCM start to work at the beginning before we place the specimen in the thermal chamber. Thanks to this the initial temperature of the concrete mix is about 8 °C lower in case of the slab with PCM.In hot and dry climate conditions, it is justified to use technological flexibility in the form of several maintenance options depending on the conditions of concreting and maturing, technological possibilities of the plant, and the type of concrete element. PCM used for the modification of cement concrete make it possible to exceed typical technological barriers in the case of concreting in unfavorable conditions (hot and dry climate). The use of PCM as a cement concrete modifier makes it possible to limit the intensity of the thermal peak and to standardize the temperature in the cross-section of the concrete element, which reduces the risk of element cracking associated with it . The addition of PCM to concrete also reduces the magnitude of temperature changes during the operation of the concrete element, which leads to increased durability and an extension of the life cycle of such element. The addition of two PCM directly to the maturing concrete delayed the appearance of the thermal peak , which allowed the construction of a correct concrete structure resistant to cracks and scratches. Thanks to PCM, the initial (starting) temperature of the concrete mix is 8 °C lower than in the slab without the addition of PCM, which helps to build the correct structure of the young concrete. The use of PCM saves a huge portion of energy without any significant reduction in mechanical strength .The research work considered the possibility of adding PCM to concretes with both high-caloric and low-caloric cements. Of course, only adding PCM to concretes with high-calorific cements is effective, because then the thermal peak in the concrete is lowered by a few degrees °C, and, furthermore, it is shifted in time by several hours , which has a direct impact on contraction and bleeding phenomena. In practice, for the construction of certain structures  mainly high-calorific cements are used due to the guarantee of maintaining the subsequent mechanical properties. PCM is preferred as a concrete curing method, because other curing methods associated with lowering the curing temperature of concrete are simply more expensive.The tests carried out so far confirm that the applied powdered and liquid PCM are characterized by the fact that when the PCM reaches the phase change temperature, by absorbing the excess heat, they do not allow the unfavorable temperature level inside the concrete element to be exceeded. When the temperature drops below the phase transition temperature, the PCM solidify and give up the excess stored heat, stabilizing the temperature inside the concrete element. The described mechanism of action of PCM in the maturing concrete is favored by the climatic conditions of the Middle East, characterized by a significant diurnal temperature amplitude. The heat accumulated during a hot day in concrete walls of residential buildings with the addition of PCM is transferred during a cold night, thus adjusting the thermal comfort to the needs of residents . In ordeThe presented calculations have shown that supporting experimental research with calculations can help in understanding the processes taking place, including in the case of the use of PCM. The results obtained in the simulations reflect the qualitative and quantitative results of the experimental research surprisingly well. This aspect was the primary goal of the research.


Antibiotic resistance is a global health crisis. New classes of antibiotics that can treat drug-resistant infections are urgently needed. To communicate this message, researchers have used antibiotic development timelines, but these are often contradictory or imprecise. We conducted a systematic literature review to produce an antibiotic timeline that incorporates the dates of discovery, first use, and initial reports of the emergence of resistance for the 38 classes of clinically used antibiotics. From our timeline, we derive lessons for identifying new antibiotics that are less prone to resistance. These include a required focus on molecules that exhibit multiple modes of action, possess unusually long ‘resistance windows’, or those that engage cellular targets whose molecular architectures are at least in part decoupled from evolutionary pressures. Our analysis also further highlights the importance of safeguarding antibiotics as a mechanism for mitigating the development of resistance. We have made our data and sources freely available so that the research community can adapt them to their own needs. Antibiotic resistance—bacterial infections that no longer respond to the drugs used to treat them—is a global health crisis of growing concern ,2. The dIn previous reviews, the definitions used for the date of the discovery of an antibiotic, its first clinical use, and the emergence of resistance to it, either are not disclosed or lack internal consistency. Examples include the non-disclosure of methods used to generate a timeline for antibiotic discovery ; inconsiDiscovery: when a compound or extract—not merely an organism—was first reported to have antibiotic activity. In some cases, this was decades after the compound was first discovered or synthesized.First clinical use: the first use of the antibiotic to treat a bacterial infection: in modern terms, a phase II trial. Clinical studies of tolerance and toxicity are not included. The approval of an antibiotic for human use is not used because several of these drugs predate regulatory bodies.Resistance: the first report of clinically isolated bacteria resistant to the antibiotic.We conducted a systematic literature review to produce a more precise timeline for antibiotic discovery, introduction, and resistance. Antibiotics can be classified by their origins, structures, and mechanisms of action. We took the 38 classes of antibiotics in clinical use , as defiWe hope that our timeline will be a useful resource to the antibiotic research and public health communities. We have included our data and sources so that the timeline can be verified, updated, and modified.Clinical Microbiology Reviews defined the “discovery void”, during which no new antibiotic classes have been discovered, as starting from 1987, and several sources have repeated this claim [is claim ,13,14,15is claim .The development window: how long after its discovery the antibiotic was first used in the clinic.The resistance window: how long after its first use clinical resistance was reported.There are some obvious outliers in this analysis. The antibiotics with long development windows were either technically challenging to optimize or shelved because they were not considered to be promising drugs until the antibiotic resistance crisis worsened ,18,19,20More promisingly, there are a few examples of antibiotics with unusually long resistance windows, from which we can derive some lessons for designing or identifying “resistance-proof” antibiotics ,24.The polypeptide antibiotic tyrothricin has only been used topically, which is likely part of the reason for its long resistance window . HoweverGlycopeptides such as vancomycin bind D-Ala-D-Ala residues at the ends of glycan chains, preventing the binding of peptidoglycan biosynthetic enzymes . This moMycobacterium leprae, which made resistance to these antibiotics more difficult to definitively prove [The long resistance window for sulfones is probably due to their main indication as drugs for leprosy . Leprosyly prove . Enniatily prove . In vivoly prove . Presumaly prove ,41. The This work represents the first comprehensive and consistent timeline for antibiotic discovery, development, and resistance. It should prove useful for communicating the alarmingly low number of new antibiotic classes that are reaching the clinic, and we have also shown how the data can be used to identify antibiotic classes for which resistance is more difficult to evolve. Our findings reveal a correlation between pharmacophore novelty and a reduced ‘development window’, and also serve to highlight the importance of prioritizing molecules with expanded ‘resistance windows’ to ensure the long-term safeguarding of antibiotics. By making our data fully available, and our methods transparent, we hope that future researchers can use and adapt our timeline for their own science communication.We conducted a systematic literature review by searching the Web of Science Core Collection and PubMed databases for the names of antibiotic classes or their first members, and found the earliest dates of discovery, use, and resistance, defined in


Salmonella Typhimurium under continuous sublethal selective pressure. Salmonella Typhimurium ATCC 19585 (STATCC) and S. Typhimurium CCARM 8009 (STCCARM) were sequentially cultured for 3 days at 37 °C in trypticase soy broth containing 1/2 × MICs of cefotaxime (CEF1/2), chloramphenicol (CHL1/2), gentamicin (GEN1/2), and polymyxin B (POL1/2). The STATCC and STCCARM exposed to CEF1/2, CHL1/2, GEN1/2, and POL1/2 were evaluated using antibiotic susceptibility, cross-resistance, and relative fitness. The susceptibilities of STATCC exposed to GEN1/2 and POL1/2 were increased by a 2-fold (gentamicin) and 8-fold (polymyxin B) increase in minimum inhibitory concentration (MIC) values, respectively. The MIC values of STCCARM exposed to CEF1/2, CHL1/2, GEN1/2, and POL1/2 were increased by 4-fold (cefotaxime), 2-fold (chloramphenicol), 2-fold (gentamicin), and 8-fold (polymyxin B). The highest heterogeneous fractions were observed for the STATCC exposed to CEF1/2 (38%) and POL1/2 (82%). The STCCARM exposed to GEN1/2 was cross-resistant to cefotaxime (p < 0.05), chloramphenicol (p < 0.01), and polymyxin B (p < 0.05). The highest relative fitness levels were 0.92 and 0.96, respectively, in STATCC exposed to CEF1/2 and STCCARM exposed to POL1/2. This study provides new insight into the fate of persistent cells and also guidance for antibiotic use.This study was designed to evaluate the adaptive resistance of  Over the last few decades, the emergence and spread of antibiotic resistance have become some of the top threats to global health ,2. BacteSalmonella Typhimurium wild-type strain (STATCC), which serves as the original parent strain, and the antibiotic-resistant strain (STCCARM), which encodes pre-resistome genes that can respond to several stress conditions [Antibiotic tolerance in persister cells is due to dormancy, leading to fewer active target sites for antibiotics . Persistnditions , by expoIn the early stage of antibiotic discovery, antibiotics became a successful treatment for bacterial infections. However, the overuse and misuse of antibiotics have become major causes of the development of antibiotic resistance in bacteria . BacteriATCC and STCCARM exposed to a half MICs of antibiotics for 3 days at 37 °C   . For STAd CHL1/2 A,B, corrd CHL1/2 A. STATCCce genes . Thus, Sux pumps . On the ux pumps . Thus, tectively C,D. For  4-fold; E, chlora 4-fold; F, gentam 4-fold; G, and po 4-fold; H. BacterATCC and STCCARM exposed to CEF1/2, CHL1/2, GEN1/2, and POL1/2 was evaluated with the formation of persister and resistant cells (ATCC exposed to CEF1/2 (38%), followed by CHL1/2 (23%). This is in good agreement with a previous report which stated that the continuous exposure of bacteria to antibiotics can promote the formation of persister cells [ATCC exposed to POL1/2 showed the highest resistance percentage (83%), followed by GEN1/2 (30%) .The bacterial heterogeneity of STnt cells . Compareer cells . The for CEF1/2 3%, follower cells ,25 and ier cells ,27. For er cells ,29. In aer cells . These rer cells . Furthery CHL1/2 %. This ier cells . STATCC /2 (30%) A. Resister cells B. SimilaSalmonella Typhimurium has several resistance mechanisms to antimicrobial peptides, such as polymyxin B, which regulate the activation of the two-component regulatory PmrAB system and the increased secretion of outer membrane vesicles containing antimicrobial agents from the bacterial envelope [ATCC and STCCARM strains used in this study are able to activate the resistance mechanisms that can lead to easy adaptation to polymyxin B and a large proportion of polymyxin-B-resistant cells. This observation suggests that the PmrAB operon and outer membrane vesicles are more likely to be associated with resistance development than persistence formation [envelope . This imormation . Persistormation . Therefoormation . The surormation ,33.ATCC and STCCARM exposed to CEF1/2, CHL1/2, GEN1/2, and POL1/2 was evaluated using different classes of antibiotic disks  were cross-resistant to cefotaxime (1/2 (p < 0.05) to chloramphenicol (1/2 (p < 0.05), GEN1/2 (p < 0.001), and POL1/2 (p < 0.001) to polymyxin B , while those exposed to CHL1/2 showed the lowest relative fitness level (0.47), indicating the highest fitness cost. The highest fitness level achieved was that of STCCARM exposed to POL1/2 (0.96), followed by that achieved by GEN1/2 (0.82) and CHL1/2 (0.70)  . Antibioibiotics . As a re level 0., indicatSalmonella enterica subsp. enterica serovar Typhimurium ATCC 19585 (STATCC) and S. Typhimurium CCARM 8009 (STCCARM) were obtained from the American Type Culture Collection  and the Culture Collection of Antibiotic Resistant Microbes , respectively. STATCC and STCCARM were used as representative of wild-type, antibiotic-sensitive and antibiotic-resistant strains, respectively [g for 15 min at 4 °C. The collected cells were diluted with PBS to 108 CFU/mL for further assays.Strains of ectively . All strATCC and STCCARM were determined according to the microbroth dilution assay [5 CFU/mL of STATCC or STCCARM was inoculated in the microtiter plates. After inoculation, the plates were incubated at 37 °C for 18 h to measure optical density (OD) at 600 nm using a microplate reader  and to determine minimum inhibitory concentration (MIC) where no visible growth was observed.The antibiotic susceptibilities of STon assay . The antATCC and STCCARM were successively cultured in TSB containing 1/2 × MICs of cefotaxime , chloramphenicol , gentamicin , and polymyxin B  for 3 days at 37 °C. The antibiotic-treated STATCC and STCCARM cells  were daily resuspended in 1 mL of TSB containing a half MIC of each antibiotic. After exposure to antibiotics, the growths of untreated control and STATCC and STCCARM exposed to CEF1/2, CHL1/2, GEN1/2, and POL1/2 as a function of antibiotic concentration were used to estimate the fractions of heterogeneous cells. The area under curve was calculated using sub-MIC and resistance-selective windows [ATCC and STCCARM exposed to CEF1/2, CHL1/2, GEN1/2, and POL1/2 were compared with those of the untreated control to measure heterogeneity.ST windows . The selATCC and STCCARM exposed to CEF1/2, CHL1/2, GEN1/2, and POL1/2 was evaluated using antibiotic disk diffusion assay. Antibiotic disks  used in this study were cefotaxime (30 μg), chloramphenicol (30 μg), gentamicin (10 μg), and polymyxin B (300 μg). The antibiotic-exposed STATCC and STCCARM cells were spread on Mueller–Hinton agar plates. After the plates were dried for 10 min, antibiotic disks were placed on the surface of agar plates, and the plates were incubated at 37 °C for 24 h. The diameter of the inhibition zone was measured using a digital vernier caliper .The cross-resistance of STATCC and STCCARM exposed to CEF1/2, CHL1/2, GEN1/2, and POL1/2 was determined by culturing in antibiotic-free TSB at 37 °C for 24 h. The relative fitness was calculated to evaluate the cost of resistance of STATCC and STCCARM exposed to CEF1/2, CHL1/2, GEN1/2, and POL1/2, which were expressed as the ratio of the growth of antibiotic-treated cells to antibiotic-untreated control cells cultured in antibiotic-free TSB.The relative fitness of STData were analyzed using the Statistical Analysis System (SAS) software. All analyses were performed with three biological replicates. The general linear model (GLM) and Fisher’s least significant difference (LSD) procedures were used to determine mean differences at 5%, 1%, and 0.1% levels of significance.In conclusion, the most significant finding was that bacterial persistence can cause a significant impact on the development of antibiotic resistance. Bacteria exposed to antibiotic selection pressure promote the emergence and spread of antibiotic resistance. Antibiotic resistance and persistence are characterized by transferable genetic variation and transient phenotypic variations, respectively, in bacteria. Although bacterial persistence can contribute to the evolution of antibiotic resistance, antibiotic-tolerant bacteria show growth defects in the absence of antibiotics. The population-based study of persistence is not sufficient to specifically identify and characterize persister cells in mixed populations. Thus, single-cell-based approaches need to accurately identify specific markers and distinguish persister cells. The evolution of antibiotic persistence cannot go unnoticed in clinical practice. It is essential to understand the mechanisms of persister formation when designing effective antibiotic strategies.


Bloodstream infection is a significant cause of morbidity and mortality. Early diagnosis and appropriate antibiotic treatment contribute to a favorable prognosis. We demonstrate a reduction of time to proper antibiotics and reduced mortality utilizing prompt diagnosis and antibiotic stewardship by infectious diseases physicians at a general hospital in Thailand. Furthermore, previous studies have reported survival benefits of earlier antibiotic administration. Therefore, we assessed time to appropriate antibiotic therapy, clinical outcome, and microbiological outcome after rapid diagnosis was implemented in combination with antibiotic stewardship.Early administration of optimal antibiotics is considered the cornerstone therapy for managing severe sepsis and septic shock.We obtained data from the internal medicine ward at Lampang Hospital. The medical records of 403 patients were included from June 1, 2019, to December 31, 2019  were retrospectively reviewed with informed consent waived. For the postintervention period , the medical records of 309 patients were reviewed with informed consent. All patients were aged ≥18 years and had clinically significant bloodstream infections (BSIs). We applied the following exclusion criteria : contaminated positive blood cultures, polymicrobial bacteremia> 2 species, and/or palliative care. The study protocol was approved by Lampang Hospital Ethics Committees (code no. 119/62). The medical technician enrolled postintervention candidates for 8 hours daily (8:00 a.m.–4:00 p.m.) and notified the ID physician when needed. Empirical antibiotics were adjusted based on clinical syndrome and antibiotic adjustments according to AST results were considered by the ID physician.Laboratory techniques included MicroScan Walk Away DxM 1096 before rapid diagnosis was implemented in combination with antibiotic stewardship and Phoenix M50 thereafter. Antimicrobial susceptibility testing (AST) was performed using CLSI 2019–2020.P < .05. Statistical analysis was carried out using Stata version 14.2 software .We compared the median time to appropriate antibiotics before and after the intervention. Overall survival was assessed using Kaplan-Meier survival analysis. Statistical significance was defined as a P = .03). Frequent comorbidities were diabetes mellitus, chronic renal failure, and malignancy. Secondary BSIs were present in 73% of patients before the intervention and 80% of the intervention. Common sites of infection were genitourinary tract, intra-abdominal tract, and respiratory tract. Septic shock (P = .03), Pitt bacteremia score (P = .007), and SOFA score (P = .006) were significantly higher in the preintervention group (Table P = .04).We included 403 patients before the intervention and 309 patients after the intervention. Community-acquired infection was the most common infection type (up Table . Gram-neP < .001), and de-escalation increased after the intervention . Time to appropriate antibiotic decreased from 106.6 hours before the intervention to 61.8 hours after the intervention (P < .001) , and 14-day mortality decreased after the intervention . However, recurrent bacteremia in 1 month remained the same rate (11% vs 13%). Before and after the intervention, respectively, the most common appropriate antibiotics were ceftriaxone (33% vs 43%), followed by ceftazidime (4% vs 6%) and cloxacillin (2% vs 7%).Overall appropriate antibiotic increased after the intervention (49% vs 80%;  We documented earlier times to appropriate antibiotic therapy with higher antibiotic adjustment, especially de-escalation in the postintervention group. Rapid diagnosis and administration of appropriate antibiotic therapy and earlier AST-based de-escalation increased the survival rate. Our findings strongly support using β-lactam monotherapy due to mainly community-acquired BSI with antibiotic-susceptible pathogens because there is no survival benefit from empirical combination therapy over monotherapy in patients with community-acquired, gram-negative BSIs. Although microbiological cure significantly increased, there was no difference in the proportion of recurrent bacteremia cases within 30 days between the pre- and postintervention groups. Therefore, repeated blood culture may have little utility in those who demonstrate early favorable clinical response. Finally, our clinical outcomes in BSIs were improved by emphasizing the importance of combining rapid diagnosis with antibiotic stewardship.Our results highlight the importance of implementing a rapid diagnosis combination with antibiotic stewardship to improve outcomes for patients with BSI. We demonstrated that community-acquired BSI and gram-negative bacteria, especially Enterobacterales, were the most common causes of BSI. Patients in the preintervention group had a higher rate of septic shock, higher Pitt bacteremia scores, and higher SOFA scores, so they had a greater likelihood of an unfavorable clinical response.This study had several limitations. The quasi-experimental design limited data accessibility. Second, the variation of time to enrollment affects time to antibiotic adjustment. Third, it is difficult to determine appropriate duration of antibiotic, conversion to an oral antibiotic, and source control measures. Lastly, cultures from nonsterile sites are challenging to interpret.In conclusion, antibiotic stewardship combined with rapid diagnosis contributes to appropriate early antibiotic therapy and enhances recovery.


To explore the associations between higher antibiotic use rates (AURs) and adverse outcomes in very-low-birth-weight (VLBW) infants without culture-proven sepsis or necrotizing enterocolitis (NEC) in a multicenter of China.A prospective cohort study was performed on VLBW infants admitted to 24 neonatal intensive care units from January 1, 2018, to December 31, 2018. AUR was calculated as calendar days of antibiotic therapy divided by total hospital days. The composite primary outcome was defined as mortality or severe morbidity, including any of the following: severe neurologic injury, bronchopulmonary dysplasia (BPD), and stage 3 or higher retinopathy of prematurity.A total of 1,034 VLBW infants who received antibiotics without culture-proven sepsis or NEC were included in this study. The overall AUR of eligible VLBW infants was 55%, and the AUR of each eligible VLBW infant ranged from 3 to 100%, with a median of 56% . After generalized propensity score and logistic regression analysis of 4 groups of VLBW infants with different AUR range, infants in the higher quartile AUR,  and , had higher odds of composite primary outcome  and BPD  than those in the lowest AUR (Q1).Antibiotic overexposure in VLBW infants without culture-proven sepsis or NEC was associated with increased risk of composite primary outcome and BPD. Rational empirical antibiotic use in VLBW infants is urgently needed in China. As the world’s most populous country, China has the second largest number of premature babies in the world, with an increasing number of very-low-birth-weight (VLBW) infants in recent years . Due to This multicenter, prospective cohort study collected data for all VLBW infants admitted to 24 participating neonatal intensive care units (NICUs) between January 1, 2018, and December 31, 2018, from the Sina-northern Neonatal Network (SNN). SNN is a regional multicenter neonatal clinical research database in China that includes preterm inpatients with birth weights (BWs) < 1500 g or gestational ages (GAs) < 32 wk. Due to more than 50% missing antibiotic data from 4 hospitals, 24 units with relatively complete data were included in this study. This study was approved by the Ethics Committee of Shandong Provincial Hospital affiliated to Shandong First Medical University and Shandong University (LCYJ: NO.2019–132). The parents of all the subjects agreed to participate in this study and signed the informed consent form, and all data were deidentified.Infants with BW < 1500 g who were admitted to the 24 participating hospitals between January 1, 2018, and December 31, 2018, were included in the study. The exclusion criteria were as follows: infants with major congenital anomalies, infants with missing antibiotic data, infants with redirection of intensive care, infants diagnosed with culture-proven sepsis or NEC, and infants without antibiotic use.Data on demographic information, Neonatal Acute Physiology version II (SNAP-II) scores, diagnoses, laboratory, and drug data, including drug names, classes, and dates, were collected prospectively in the database.AUR was calculated as calendar days of antibiotic therapy divided by total hospital days . The terThe SNAP-II score  is a valp values at the 5% testing level.All eligible VLBW infants were divided into 4 groups based on their AUR quartiles (Q1−Q4). Demographic data are expressed as the mean [SD] or percentages. In the univariate analysis, the Kruskal−Wallis test or chi-square test for continuous and categorical variables were used, respectively. Potential confounders and other covariates on the basis of findings in the univariate analysis were then adjusted in propensity score matching methods with multilevel treatments to control for severity of illness of patients. Balance was assessed by the generalized propensity score (GPS) . Propensn = 8), missing antibiotic data (n = 91), infants with redirection of intensive care (n = 150), infants diagnosed with culture-proven sepsis or NEC (n = 81), and infants without antibiotic use (n = 41), the remaining 1034 VLBW infants were included in the present study . The comparisons of infant demographic characteristics and clinical outcomes are shown in Table GA, sex, Apgar score at 5 min, SNAP-II score, cesarean delivery, mechanical ventilation, and vasoactive drugs were adjusted for propensity score matching methods with multilevel treatments. For each group, the normalized difference for GPS were calculated for that group and plotted a box-whisker plot of GPS for infants in each group and a box-whisker plot of GPS for the remaining infants in the same figure Fig.  a−d. GroIn this multicenter prospective cohort study of China, it was found that the overall AUR of VLBW infants who received antibiotics without culture-proven sepsis or NEC was 55%, and the median AUR of each VLBW infant was 56% (IQR 33%/86%), which were both much higher than those in developed countries , 14. MorOver half of the VLBW infants in the present study were treated with antibiotics for more than half of the patient's length of stay (AUR > 56%). However, the median AUR of VLBW infants was only 17% in a multicenter study of the Canadian Newborn Network (CNN) . Data inIn the analysis of the associations between different AURs and adverse outcomes in VLBW infants, severe neurologic injury, BPD, and ROP were included in the primary outcome since their chronological sequence usually occurs after the median age of infection in preterm neonates. The authors found increased adjusted ORs of the composite primary outcome and BPD in VLBW infants in the higher AUR quartile and significantly increased ORs of mortality in VLBW infants in the highest AUR quartile compared to the lowest quartile. The findings of the present study were similar to those reported in the literature. Cantey et al.  reportedAs far as the authors' know, this study is the first multicenter, observational cohort study with a large sample to investigate the real-life situation of antibiotic overexposure and its adverse outcomes in VLBW infants in China. Antibiotic overuse has increasingly become a serious problem worldwide, with potential for the emergence of multidrug-resistant bacteria and even ‘superbugs’. However, this study presents only an overview of antibiotic use in VLBW infants and does not further represent data about the reasons for antibiotic overuse. The authors are planning to develop multicenter ASP and lead a deep survey about reasons for inappropriate antibiotic use and differences in antibiotic use strategies among NICUs. Another limitation of this study is confounding by indication as an observational study, which could lead to a false association between adverse outcomes and treatment. In this study, GPS was used to address the limitations of confounding bias of multilevel AUR. The idea of GPS for multilevel treatments is relatively new, but it is more advantageous than simple multivariable logistic regression due to the weak nonconfoundedness assumption that assignment to a certain treatment level is independent of the potential outcome given the GPS . In addiAntibiotic overexposure in VLBW infants without culture-proven sepsis or NEC was common and was associated with an increased risk of composite primary outcomes and BPD, which is a serious health issue in China. More efforts should be focused on the development and implementation of antimicrobial stewardship programs that can help reduce the burden of antibiotic use in countries where bacterial resistance is rising, such as China.


Principal Component Analyses (PCA) did not reveal separation of samples into distinct microbiota-based clusters by antibiotics use, suggesting the microbiota composition was not affected by the antibiotics commonly used in this population. Additionally, the fecal microbial diversity of the subjects in our study cohort was lower when compared to that of healthy Dutch adults (median 3.95 (IQR 3.72–4.13) vs median 3.69 (IQR3.31–4.11), p = 0.028, despite the higher dietary fiber content in the Vietnamese as compared to western diet. Our findings support the hypothesis that frequent antibiotic exposure may push the microbiota to a different steady state that is less diverse but more resilient to disruption by subsequent antibiotic use.The human gut microbiota has been shown to be significantly perturbed by antibiotic use, while recovering to the pre-treatment state several weeks after short antibiotic exposure. The effects of antibiotics on the gut microbiota have however been mainly documented in high-income settings with lower levels of antibiotic resistance as compared to lower and middle income countries (LMIC). This study aimed to examine the long-term consequences of repeated exposure to commonly use antibiotics on the fecal microbiota of residents living in a low income setting with high prevalence of antibiotic resistance. Fecal samples from household individuals (n = 63) participating in a rural cohort in northern Vietnam were collected monthly for a period of 6 months. Using 16S V4 rRNA gene region amplicon sequencing and linear mixed-effects models analysis, we observed only a minor and transient effect of antibiotics on the microbial richness (ß = − 31.3, 95%CI = − 55.3, − 7.3,  Transient or long term alterations can however still occur in response to various external factors such as diet, living conditions and medical drug use5. In general, the human gut microbiota is rather resilient and recovers after drastic pulse perturbations, such as courses of antibiotics. Maintaining a resilient and robust microbiome is pivotal to minimize the period of microbial disruption upon antibiotic use which may put the host at increased risk of opportunistic infections6.After the initial maturation of the gut microbiota during the first years of life, it exists as a highly diverse and relatively stable ecosystem in healthy individuals8. Regarding the latter, four main factors have been recognized to play an essential role in the resilience and recovery of the human gut microbiota: initial composition, capability to maintain its functional redundancy, abundance of (opportunistic) pathogens and the antibiotic resistance genes (ARGs) reservoir10. Indeed, when a simulated human gut microbiota was exposed to amoxicillin, it was shown that its biodiversity was extended by increasing abundance of pathogens carrying resistance genes11. A study on 12 healthy adults found that bacterial species harboring β-lactam (ARGs) modulated the resilience/recovery patterns by increasing the chances of survival and novel colonization after a broad-spectrum antibiotic treatment12.Microbiota resilience after antibiotic exposure has been shown to be highly dependent on diet, environmental context, past perturbations, and specific properties of the used antibiotics, as well as the individual-specific microbiome13. For instance, previous studies demonstrated that the abundance of Enterobacteriaceae was decreased after treatment with penicillin and cephalosporin antibiotics while the abundance of enterococci and Bacteroides spp was increased14. Moreover, the ensuing perturbations related to the use of β-lactam antibiotics is conductive for the selective outgrowth of opportunistic pathogens as illustrated by the increased risk of Clostridioides difficile infection16. A recent study illustrated that a course of penicillin or amoxicillin does not alter human gut microbiota of western obese individual17, whereas other studies found more profound yet temporary microbiota alterations upon amoxicillin treatment18. Previous culture-based studies reported that bacterial abundance returned to pre-antibiotics state 4 to 14 days after discontinuation of cephalosporin use21, but some studies observed an altered state that lasted longer than 42 days23.β-lactam antibiotic therapy has been associated with profound alterations in the gut microbiota of both healthy individuals and patients, including a significant decrease in species richness, changes in abundance of specific bacterial taxa and a relative increase in antimicrobial resistance24. The effects of antibiotics on the microbiota composition in these settings may differ given the different initial microbiota composition resulting from different lifestyles, dietary habits and history of antibiotic exposures. The fecal microbiota composition of rural populations in LMICs like Vietnam, is expected to be more diverse than in high income countries25 at least in part due to the lower fat and higher fibre-content of the diet27 as compared to the Western diets which are typically enriched in saturated fats and simple carbohydrates29. We therefore conducted this study (i) to explore the long-term impact of antibiotics on diversity of the human gut microbiota in a resource limited setting and (ii) to evaluate whether repeated exposure to commonly used antibiotics induces the human gut microbiota with high prevalence of antibiotic resistance to be resilient to further antibiotic effects.The majority of studies were conducted in high-income settings or settings with a low prevalence of antibiotic resistance. Studies on impact of antibiotics on microbiota conducted in low- and middle-income countries (LMICs) with a high consumption of antibiotics and high prevalence of antibiotic resistance are underrepresented30.This study was conducted within the context of an existing prospective household-based community cohort in Ha Nam in northern Vietnam, which was established to quantify the burden of influenza and to gain insights into the influenza virus transmission in a tropical setting. A full description of the demographics and lifestyle of the Ha Nam cohort has been published previouslyTo examine the impact of antibiotic use on shifts in the gut microbiota of residents of this community, we conducted a longitudinal study over a period of six months from November 2014 to June 2015. The study design encompassed weekly interviews to collect information on health issues and consumption of antibiotic use in the preceding week and monthly fecal samples for microbial profiling. The interviews included questions to indicate the indications for which antibiotics were used in the preceding week ).From 80 households (HHs), we selected 11 HHs in which at least one household member used antibiotics during the six months follow-up. In total this resulted in 41 participants of whom 32 used at least one course of antibiotics.www.randomizer.org) for the selection.In addition, we randomly selected a control group from 11 HHs (22 participants) where none of the household members used antibiotics during the study period. We used randomizer (2), samples collected within two to four weeks after last antibiotic exposure (E4), and samples collected without any antibiotic exposure in the preceding four weeks  of 106 subjects with no history of antibiotic use during the past year. Differences in Shannon diversity and observed species richness in fecal samples of Dutch and Vietnamese individuals were examined by linear regression analyses with adjustment for age and sex as potential confounding factors.To compare the microbial diversity between our cohort and a cohort with less antibiotic use, we included a random selection of healthy Dutch adults (≥ 18 years) who participated in the Carriage Of Multiresistant Bacteria After Travel (COMBAT) Study. This study was conducted to evaluate the travel-associated risk factors on the acquisition, persistence and transmission of antimicrobial resistance (AMR) in gut microbiota of healthy people. The detailed information of demographic characterization of the cohort has been reported previouslyWe used the term “microbial resistance to pulse perturbations” to indicate the resistance of the microbiota to change upon exposure to antibiotic use. Microbial resilience, on the other hand, reflects the recovery speed upon a shift in the microbiota. In this study, to examine the microbial resistance to pulse perturbations, we stratified Vietnamese individuals who used antibiotics according to their microbial resistance. For this purpose, we calculated the Aitchison distance between the samples of each subject collected before and immediately after antibiotic use. Subjects were classified as having a low microbial resistance when the Aitchison distance was above the median, whereas subjects with an Aitchison distance below the median were classified as having a high microbial resistance to pulse perturbations changes.We also examined if the number of different antimicrobial resistance genes (ARGs) encoding extended-spectrum beta-lactamases, carbapenemases and plasmid-mediated colistin resistance in fecal samples was associated with the microbial community structure.32. Briefly, fecal samples were firstly subjected to mechanical lysis by repeated Bead-Beating (RBB) followed by column-based purification.Fecal samples were transported within 4 h after defecation to the National Institute of Hygiene and Epidemiology (NIHE) in cold conditions (4–8 °C). Within 3 h after transport, samples were stored at − 80 °C until DNA extraction. Microbial DNA was manually extracted from frozen feces according to protocol Q of the International Human Microbiome Standards consortium33. In brief, we amplified the V4 region of the 16S rRNA gene from each DNA sample in triplicate using the 515 f./806r primer pair34. PCR amplicons of the triplicate reactions were pooled; next products were purified using AMPure XP purification  according to the manufacturer’s instructions and eluted in 25 µl 1 × low TE . Subsequently, we quantified DNA concentration by Quant-iT PicoGreen dsDNA reagent kit  using a Victor3 Multilabel Counter . The purified amplicons were mixed in equimolar concentrations to ensure equal representation of each sample and, were sequenced on an Illumina MiSeq instrument using the V3 reagent kit (2 × 250 cycles).We performed amplicon libraries preparation and sequencing as described previously35. We set maximum number of expected errors (E_max) at 0.5 to remove low-quality reads (with expected errors (E) > E_max) from demultiplexed reads. We then remained good-quality regions of reads for further steps by truncation at positions 200 of forward and 140 of reverse reads. Samples with total read counts below 12,000 were not further analyzed. Merged reads were used to generate amplicon sequence variants (ASV) using DADA2 (V1.16.0)35. Identified ASVs were aligned to construct a phylogeny with FastTree236. Taxonomy was assigned using the RDP v16  with the set formatted for DADA2 package37.After demultiplexing raw reads, we visualized the quality profiles of forward and revered reads by uploading the Fastq files into R using package DADA2 (v1.16.0)ASVs that were present in less than 5% of all samples as well as reads with a total relative abundance < 0.01% were removed from downstream analysis. At ASV level of taxonomic aggregation, we calculated the alpha-diversity (observed richness and Shannon index) and beta-diversity (Aitchison and Bray–Curtis distances) as measures of within- and between-sample microbial diversity, respectively. To examine the change in alpha-diversity from baseline (M1) to the subsequent time points , we next calculated the delta of the alpha diversity , ggthemes (v.4.2.0), circlize (v.0.4.10), ComplexHeatmap (v.2.0.0)39, ggplot2 (v.3.3.2)38, lme4 (v.1.1–27.1), glmmTMB (v.1.1.2.3), ggstatsplot (v.0.9.0)40,dplyr (v.1.0.2)38, phyloseq (v.1.28.0), microViz (0.7.10.9004)41.All statistical and computational analysis and visualizations were performed in R v4.1.1 (2021–08-10) using the following packages: reshape2 (v.1.4.4), purrr (v.0.3.4), readr (v.1.3.1), tidyr (v.1.1.2), tibble (v.3.0.4), tidyverse (v.1.3.0)Genus (Counts) ~ Exp_ABx_3codes + Age + Time_point + Age *Exp_ABx_3codes + offset ) + (1|ParticipantID). In the model, the variance ‘Exp_ABx_3codes’ is a logical values including three categories that was grouped by number of days since the last antibiotic use prior to sampling as described in the study design.Linear mixed effects models were used to test for the effects of antibiotic use, age, and time points on alpha diversity , USA). The model included exposure to antibiotics, age and time point of sample collection as fixed effects and participant ID as a random effect to control for repeated measures. The variance inflation factors (VIF) were calculated prior to model fitting to assess correlated predictor parameters, none were excluded on the basis of collinearity. In addition, the residuals were plotted and checked for homoscedasticity. We next used generalized linear mixed effects models using a negative binomial distribution to test the effect of antibiotic use on the relative abundance of individual bacterial genera . For each genus, we fitted the model p < 0.05 upon Bonferroni correction) are presented42.To examine differentially abundant genus-level taxa between Dutch and Vietnamese individuals, we conducted Analysis of Compositions of Microbiomes with Bias Correction (ANCOM-BC). Differentially abundant genera , the National Institute of Hygiene and Epidemiology, Vietnam (NIHE) institutional review board and National Hospital for Tropical Diseases, Vietnam.All methods were performed in accordance with the relevant guidelines and regulations.We confirm that informed consent was obtained from all participants and/or their legal guardians.To analysis the impact of antibiotic use on the microbiota in the Ha Nam cohort. This observational study investigated gut microbiota responses among 63 individuals belonging to 11 HHs that used at least one course of antibiotics (n = 32) or 11 HHs that did not use antibiotics (n = 31) during the study period. The median age was 28 years (IQR = 8.5–45.2) for those that consumed antibiotics (including 6 subjects aged < 6 years) versus 41 years (IQR = 17.5–52.5) for those who did not (no subjects aged < 6 years).Among the 32 individuals who used antibiotics during the study period, 20 individuals consumed two or more courses, which resulted in a total consumption of 69 courses of oral antibiotics . First and second generation cephalosporins (n = 52) were most frequently consumed, followed by penicillin (n = 15), trimethoprim (n = 1) and lincosamide antibiotic (n = 1).The symptoms for which participants reported to have used antibiotics were symptoms of the upper respiratory tract (fever accompanied by cough) for 50 courses, urinary tract combined with upper respiratory tract symptoms for 15 courses and urinary tract combined with stomach complaints for 4 courses. In none of these cases individuals reported to have symptoms of diarrhoea or other intestinal tract symptoms. Therefore, it is unlikely that confounding by indication  affected our findings.A total of 373 samples were collected from 63 individuals at month 1 (n = 60), month 2 (n = 62), month 3 (n = 63), month 4 (n = 63), month 5 (n = 63) and month 6 (n = 62) of follow-up.After filtering of low-quality reads the remaining reads with a median sequencing depth of 81,488 reads per sample  were clustered into 5,367 amplicon sequence variants (ASVs). Upon removal of rare ASVs, defined as ASVs present in less than 5% (n = 19) of the samples and ASVs with an overall abundance of less than 0.01%, a total of 460 ASVs were retained.We first investigated the richness and diversity of the microbiota. Overall, significant differences during the study period were not observed for both richness and biodiversity Fig. . Althougp = 0.011) and had already disappeared when antibiotics were consumed 2–4 weeks prior to fecal sampling . Also, the microbial diversity, as measured by the Shannon index, was reduced in samples collected within two weeks after antibiotic use, although this was not statistically significant . These findings suggest that in the current population the use of antibiotics has only an effect on the microbial richness for a short period, while the microbial diversity is even less affected. In contrast the reduction of microbial diversity remained significantly lower than that in the initial composition after 42 days of treatment with beta–lactam antibiotics, as previously described12.To account for the longitudinal design of our study and for potential confounding by age and carriage of extended-spectrum beta lactamases (ESBL) and carbapenemases, we next assessed the effects of antibiotics on the alpha diversity by linear mixed-effects models. Neither the follow-up time-point at which the fecal samples were collected, nor age, significantly affected the observed richness and Shannon index  Table . Also, t44, we hypothesized that the microbiota diversity of individuals in this cohort might have already been reduced by extensive past antibiotic use to an extent that the effect of further perturbations would be limited. To test this hypothesis, we compared the microbial richness and diversity of the adults in the Ha Nam cohort (n = 63) to that of a cohort of healthy Dutch adults (n = 106). Several previous studies have shown that rural communities in LMICs have a higher microbial richness and diversity compared to individuals in Western countries, mainly as a result of a diet that is rich in fibers and low in animal fat and protein. Indeed, most Vietnamese adults take grains (96.4%) and vegetables (93.9%) every day45. Meanwhile, less than 10% of the Dutch take the recommended amount of food originate from plant46. Moreover, on average, each Vietnamese person consumes 134 g meat per day47, much lower than the Dutch’s 46. However, the microbiota of the Ha Nam cohort participants showed a statistically significantly lower microbial diversity at baseline with Shannon median 3.69 (IQR 3.31–4.11) when compared to the Dutch individuals with median 3.95 (IQR 3.72–4.13), (p = 0.028). This supports the hypothesis of a reduction in microbial diversity as a consequence of frequent antibiotic exposure similar to that of Dutch individuals 48, were depleted in Dutch as compared to Vietnamese individuals. Next to these vanishing microbes, also members of the Enterobacteriaceae family including, Klebsiella and Escherichia-Shigella were enriched in the Vietnamese individuals. On the other hand, Akkermansia and Alistipes, genera of which members have been classified as BloSSUM (Bloom or Selected in Societies with Urbanization/Modernization) microbial taxa, were enriched in the Dutch individuals .Analysis of the differentially abundant taxa at genus level showed that many bacterial genera, including p < 0.001, Wilcoxon test) . However, Principal Component Analyses (PCA) did neither reveal separation of samples into distinct microbiota-based clusters by time points nor by antibiotics use  49. Possibly, individuals in the control group have used antibiotics without being aware or recording, leading to an alternative microbiota composition with a similar richness and diversity to their prior composition. Consequently, it is difficult to observe the difference in richness and biodiversity between them and antibiotics users. In addition, people of rural communities of LMICs like Vietnam, normally have a diet with higher content than western diets, while studies having shown fiber rich diets were associated with higher diversity of microbiota54. When comparing the microbiota of participants of the Ha Nam cohort to that the microbiota of Dutch individuals, we observed differences in the abundance of specific microbial genera in line with previous studies comparing traditional and industrialized populations48. However, in contrast, our results showed a significantly lower diversity in the microbiota of the participants of the Ha Nam cohort as compared to those from a Dutch cohort with less antibiotic use, supporting our hypothesis that Ha Nam cohort participants have chronically disrupted microbiota, with a lower diversity, due to frequent antibiotic exposure.Low vaccination coverage, coupled with poor water, sanitation, and hygiene (WASH) infrastructure, residents vulnerable to infection and dependent on antibiotics for treatment55. This contrasts with another recent study, where the use of both access- (59.0%) and watch (39.3%) group antibiotics was commonly observed in rural Vietnam56.This suggest that it needs an investigation the response of the human gut microbiota to the new patterns of antibiotic use. Nonetheless, the results presented here support the hypothesis that frequent or chronic antibiotic exposure may push the microbiota to a different steady state that is less diverse but more resilient to disruption by antibiotic use. Further research including comparisons between populations with different exposures is needed to provide more evidence for this hypothesis. Furthermore, the potential health consequences of this altered microbiota composition should be considered.The study has several limitations. The objective, to evaluate the effect of antibiotic use on microbiota in a community, may require a longer follow-up study, and the sample size was constrained by monthly sample collection. Currently, the definition of a healthy microbiome has not been established. Therefore, we could not conclude with certainty that the gut microbiota of this cohort is in an unhealthy steady state. Moreover, recorded antibiotic use in our community was predominantly with earlier generation cephalosporins (access group), which are less potent compared to later generation (watch group) or other restricted antibiotic (reverse group) classesThis study has added a line to the long-term impacts of antibiotic use that, with frequent exposure to antibiotic, the gut microbiota adapts to a state that is little influenced by antibiotics of longitudinal study targeting long term influence of antibiotics on microbiota in similar settings.Supplementary Information 1.Supplementary Information 2.