The objective of this study was to evaluate the knowledge, attitudes, and practices (KAP) of broiler poultry farmers (BPF) in Nepal concerning the use of antimicrobials (AMU) and the issue of antimicrobial resistance (AMR). A total of 500 farmers from all seven provinces of Nepal, spanning 40 districts, participated in this study. The collected data, encompassing AMU, demographic distribution, and KAP regarding AMU and AMR among BPF, underwent descriptive statistic and logistic regression analysis. This study’s results revealed that BPF possessed limited understanding and engagement in practices related to AMU and AMR, whereas they exhibited positive attitudes. This highlights the importance of implementing effective regulatory measures, improving veterinary services, and providing training and awareness programs to address the issue of AMR in the poultry farming sector.p = 0.02), and those using antimicrobials for preventive purposes had a more favorable attitude compared to those using them for other purposes . In light of these findings, this study recommends the implementation of effective regulatory measures for drug usage, along with awareness programs addressing AMU and AMR to address the issue of AMR in poultry production.The purpose of this study was twofold: first, to conduct a situational analysis, and second, to assess the knowledge, attitudes, and practices (KAP) of broiler poultry farmers (BPF) regarding antimicrobial use (AMU) and antimicrobial resistance (AMR). Data were collected from 500 BPF across 40 districts, representing 88.1% of Nepal’s broiler poultry population. Among these farmers, 81.0% were male, 59.6% had up to 4 years of experience in poultry farming, and 50.8% had completed at least a high school education. The most used antimicrobials on the farms were doxycycline (23.5%), neomycin (17.1%), and colistin sulfate (9.6%) out of 27 reported antimicrobials. While the BPF exhibited limited knowledge (62.6%) and practice (55.5%) related to AMU and AMR, their overall attitude toward these issues was positive (91.6%). This study also identified significant factors influencing farmers’ attitudes toward AMU and AMR. Farmers aged 31–40 showed a stronger inclination compared to other age groups (OR = 4.2,  The global concern regarding antimicrobial resistance (AMR) has gained significant importance, particularly within the context of the One Health approach. The improper use of antimicrobials in both human and veterinary medicine is leading to the emergence of resistant bacteria . The emeAntimicrobials play a crucial role in managing and controlling bacterial infections and promoting growth in food-producing animals, such as poultry . HoweverEscherichia coli and Enterococcus faecalis [In Nepal, commonly used antimicrobials for poultry production encompassed amikacin, gentamicin, streptomycin, cefotaxime, ceftazidime, ciprofloxacin, ampicillin, colistin, sulfamethoxazole, and tetracycline . Howeverfaecalis ,13. The faecalis .Poultry meat and eggs play a significant role as a primary source of animal protein in Nepal, contributing to the country’s economic growth. In 2021, the Nepalese government achieved self-sufficiency in poultry meat and egg production. While Nepal’s Ministry of Health and Population approved the National Antibiotic Containment Action Plan in 2016 and the National Antibiotic Treatment Guideline in 2014, both aimed at human health [The knowledge, attitude, and practice (KAP) study is a widely employed quantitative research method that helps uncover misconceptions or misunderstandings that may pose challenges in altering human behavior and potentially act as barriers to change . The proIn the realm of livestock production, the main factors contributing to the development of AMR in animals include farmers’ tendency to self-prescribe medications, unauthorized antimicrobial use (AMU), and the absence of regulations governing AMU. To effectively mitigate AMR, several key measures should be taken, including promoting the responsible use of antimicrobials, monitoring the prevalence of AMR, raising awareness about AMR, encouraging efficient antimicrobial practices, enacting relevant legislation, and establishing regulatory mechanisms to curb AMR. While more data are yet to be reported regarding the economic and livestock health impacts of AMR in developing countries, it is evident that veterinarians, as integral components of the AMR solution, should undergo comprehensive training and operate under the supervision of veterinary regulatory bodies ,18. UndeThe questionnaires were structured to encompass various aspects, including demographic details, an assessment of AMU conditions, and an evaluation of KAP associated with AMU and AMR among farmers. The majority of the questions were in a multiple-choice format. Demographic information covered aspects such as age, gender, educational level, work experience, province, and ecozone. Questions related to AMU delved into various aspects of poultry management, including the type, frequency, source, and storage of antimicrobials, as well as farm management regarding flock size, bird health, farm biosecurity, mortality rates, clinical signs, and disease occurrence. Knowledge-related questions focused on topics such as AMU, withdrawal periods for antimicrobials, AMR transmission, and government policies regarding AMU. In terms of perceptions about AMU and AMR, the attitude section explored aspects such as the safety of AMU and the use of nonprescribed antimicrobials. Finally, the practice-related questions sought information on completing full courses of AMU, skipping doses, the frequency of AMU, and the verification of antimicrobial expiration dates.Before commencing the actual survey, a pre-test of the questionnaires was conducted, involving the random selection of five farmers. Following this, the questionnaire underwent validation by being submitted to three experts in the field. These experts assessed the questionnaire items for their congruence with the study objectives using the item-objective congruence (IOC) index. For each section of the questionnaires, the IOC point was assigned a rating. To assess the consistency and congruence of all elements, experts were presented with a three-scale rating, and they were required to provide a single rating from these options. The overall IOC value of this set of questionnaires was determined to be 0.9.This study involved 500 broiler poultry farmers (BPFs) selected from 40 districts across Nepal’s seven provinces, namely, Sudurpaschim, Karnali, Lumbini, Gandaki, Bagmati, Madhesh, and Koshi. The selection criteria were based on the areas with the highest broiler poultry population density in the country. At least three districts from each province and ecozone of Nepal were selected for this study. In total, the study area covered 88.1% of Nepal’s broiler population.Specifically, the selected districts, which were all included in the farmer questionnaire survey, included the following: Sudurpaschim province ; Karnali province ; Lumbini province ; Gandaki province ; Bagmati province ; Madhesh province ; and Koshi province  .In Nepal, there were 52.7 million broiler poultry distributed across 20,483 farms in the year 2017. Among the selected 40 districts, there were a total of 46.4 million broiler poultry and 17,601 farms [To calculate the total number of farmers/respondents required for the questionnaire survey, a single proportion calculation was performed using a 95% confidence interval and a desired precision of 5%, assuming a 50% estimated response rate. The minimum sample size for respondents was determined to be at least 392 farmers.http://five.epicollect.net, accessed on 2 October 2023). Prior to participating, each farmer received an explanation of the survey’s objectives and purpose. The inclusion in this study was restricted to those farmers who provided their consent. To validate the information regarding AMU supplied by the farmers, records, antimicrobial photos, and antimicrobial labels or packaging materials were used.Between May 2022 and May 2023, a cross-sectional study involving broiler poultry farmers was conducted. The data collection process involved face-to-face interviews with the BPF using a semi-structured questionnaire. These interviews were carried out by a team of 46 trained veterinarians. All participating veterinarians involved in the farmer survey received orientation, with some attending in-person sessions across various locations within the seven provinces, while others received their orientation through online resources, video tutorials, or phone consultations. Initially, the questionnaires were prepared in English and were subsequently translated into Nepali for data collection purposes. All the data were recorded using EpiCollect5 , and QGIS 3.4  was used to depict the spatial distribution of sampling locations on a map.Descriptive analysis was carried out to provide an overview of the AMU situation and KAP among farmers. Logistic regression analyses were used to identify associations between variables related to KAP and the outcome variables of AMU and AMR. Independent variables with a Out of the 500 participating BPF, their distribution across different regions of Nepal was as follows: Terai region (53.8%), Hill region (42.2%), and Mountain region (4.0%) of Nepal . The higConcerning farm size, 57.8% of the farms were categorized as small (<1500 birds), while 29.4% and 12.8% were classified as medium (1501–5000 birds) and large (>5000 birds), respectively . The aveIn Nepal, nearly all BPF had easy access to antimicrobials within the study area, primarily attributed to the availability of over-the-counter sales of veterinary drugs. The poultry production sector employed a total of 27 different types of antimicrobials belonging to 13 distinct classes. Among these, the most commonly used class of antimicrobials was tetracyclines (28.0%), followed by aminoglycosides (21.5%), quinolones (13.0%), and polymyxins (9.6%) . ConcernOn average, during each cycle of broiler production, antimicrobials were employed approximately 1.6 (0–4) times across 781 cases of 500 responses. Approximately 9.2% of the farmers reported never using antimicrobials throughout the production cycle, while the remaining 90.8% of farmers used antimicrobials at least once. Among the 908 multiple responses, the most used antimicrobials were doxycycline (23.5%), neomycin (17.1%), and colistin sulfate (9.6%) .Regarding the purpose for which antimicrobials were used in animal feed, approximately 72.7% of farmers used them for treating poultry diseases, followed by prevention and control (18.2%) and growth promotion (3.1%). Approximately half (50.1%) of the farmers obtained antimicrobials directly from drug sellers. In contrast, only 33.4% and 10.3% of farmers purchased antimicrobials with prescriptions from veterinarians and para-veterinarians, respectively. Based on frequently observed clinical signs, digestive system-related signs (39.6%) were the most commonly reported, followed by respiratory system-related signs (37.7%). The use of antimicrobials in broiler production predominantly occurred during the first 3 days (16.6%) and slightly during days 4–10 (12.3%). Antimicrobial usage peaked between days 10 and 20 (25.4%), gradually decreasing to 21.9% and 23.8% on days 21–30 and >30 days, respectively. When birds exhibited illness or signs of disease, 53.2% of farmers sought advice from veterinarians, while 21.6% consulted drug sellers, 20.0% consulted para-veterinarians, and 4.4% engaged in self-treatment .A significant portion of farmers (62.6%) demonstrated good knowledge concerning AMU and AMR. A substantial majority (74.8%) either completely agreed or agreed with the idea that different antimicrobials have varying curative effects on various poultry diseases. About three-quarters of BPF (72.0%) were aware that antimicrobials used in poultry production could potentially be transmitted to humans through the consumption of poultry meat and eggs. Furthermore, 71.6% of farmers acknowledged that the presence of antimicrobial residues in poultry meat could pose hazards to human health. Similarly, 70.6% of respondents recognized that poultry should be sold after adhering to the proper antimicrobial withdrawal period to prevent the occurrence of antimicrobial residue in the meat. The results indicated that the majority (50.4%) of BPF were aware that the use of antimicrobials in animal feed is inappropriate. However, only 41.2% of farmers were aware of the government’s policies and plans related to AMU and AMR. Interestingly, a small proportion of farmers (16.0%) did not administer treatment to the entire flock when only one or a few birds were affected .This study revealed that most farmers (91.6%) held a positive attitude toward AMR and AMU. Approximately three-quarters of BPF (73.4%) believed that expired antimicrobials should not be administered to birds. Most respondents (72.2%) had a positive view of vaccination as a potential strategy to reduce the reliance on antimicrobials in poultry farming. Furthermore, a significant proportion of respondents (68.0%) advocated for the use of antimicrobials solely for preventing severe illnesses. Similarly, 60.0% of farmers acknowledged that improper doses of antimicrobials could contribute to AMR. In contrast, 54.6% of respondents believed that antimicrobials were unnecessary for treating fever or cold in humans. Additionally, nearly half of them expressed that antimicrobials were not needed for birds during seasonal changes (46.6%), and 41.2% believed that antimicrobials were unnecessary for treating any animal disease. Lastly, 24.2% of the respondents expressed concerns about the adverse effects of AMU on animal health .This study revealed that 55.5% of farmers exhibited commendable practices when it came to AMU and AMR. Most farmers (93.2%) adhered to the complete course of antimicrobial treatment as prescribed by veterinarians. Additionally, almost all farmers (88.0%) routinely checked the expiration dates of antimicrobials before using them. Two-thirds of farmers (66.8%) did not practice the habit of skipping one or two doses of their prescribed antimicrobial courses. Approximately 43.0% of respondents did not resort to increasing the dosage and frequency of prescribed antimicrobials when clinical symptoms failed to disappear or subside, and a similar proportion of farmers (42.0%) did not continue AMU when symptoms had disappeared. However, 37.2% of farmers indicated a preference for not using antimicrobials as a preventive measure against disease .p = 0.008), Koshi , and Sudurpaschim provinces . Moreover, farmers in the Koshi province exhibited 2.9 times better AMU practices than farmers in the Bagmati province (p = 0.006). Farmers within the 31–40 age group displayed a notably more positive attitude, which was 4.2 times better than their counterparts in the 10–30 age group. Furthermore, farmers who used antimicrobials for treatment purposes exhibited practices that were 2.1 times better than those using antimicrobials for all purposes (p = 0.01).In the univariate analysis, several factors were associated with knowledge of AMU and AMR, including province, age group, purpose of antimicrobials, control of antimicrobial sales, belief in the role of vaccination in reducing AMU, and the source of advice for antimicrobials . Farmersp = 0.00001). Similarly, farmers who believed in the statement “vaccination can reduce AMU” displayed better knowledge (odds ratio (OR) 3.5, p = 0.002) and attitude  compared to farmers who did not share this belief. Additionally, farmers who sought advice on AMU from veterinarians exhibited practices that were 21.0 times better than those seeking advice from drug sellers (p = 0.0001).Farmers who agreed with the statement “control of antimicrobial sales contributes to AMR” demonstrated significantly higher knowledge levels, which were 14.5 times greater than those who disagreed (p = 0.008), Koshi , and Sudurpaschim  provinces showed significantly lower levels of knowledge concerning AMU and AMR compared to farmers in the Bagmati province. Furthermore, farmers in the Bagmati province had a fivefold (1/0.2) higher level of awareness regarding AMU and AMR in comparison to those in the Lumbini province (p = 0.03). Farmers in the Koshi province demonstrated significantly better practices compared to those in the Bagmati province . In terms of age, farmers in the 31–40 age group displayed attitudes toward AMU and AMR that were 4.2 times more favorable compared to those in the 18–30 age group (p = 0.02). The multiple logistic regression analysis identified several factors associated with AMU and KAP among BPF in Nepal . Variablp = 0.02). Additionally, farmers who used antimicrobials for prevention and control purposes exhibited a 5.9 times more favorable attitude toward AMU and AMR compared to farmers who used antimicrobials for all purposes in poultry (p = 0.02). Interestingly, farmers who used antimicrobials to treat their poultry exhibited better practices in AMU than those who used antimicrobials for all purposes (p = 0.01).In terms of the purpose of AMU in poultry production, farmers who were unaware of the specific purpose of AMU demonstrated 16.4 times more knowledge about AMU and AMR than farmers who used antimicrobials for all purposes (p < 0.0001). Similarly, those who had faith that vaccines could reduce AMU demonstrated significantly better knowledge and attitudes toward AMU and AMR than those who did not believe in the reduction of AMU through vaccination, with p = 0.002 and p < 0.0001, respectively. Regarding the sources of advice for poultry health, farmers who consulted with veterinarians when their poultry were sick exhibited better practices regarding AMU than those who consulted with drug sellers only . Farmers who practiced self-treatment displayed 8.7 times better practices in AMU than those who consulted with drug sellers (p = 0.001).Regarding the perception of antimicrobial control and AMR occurrence, farmers who believed that the lack of antimicrobial sales control could contribute to AMR had 14.5 times greater knowledge and 2.6 times more favorable attitudes about AMU and AMR than their counterparts (p < 0.0001). Furthermore, there was a direct association between the depth of knowledge of farmers regarding AMU and AMR and their actual practices . However, the association between farmers’ attitudes and practices was not found to be significant. This suggests that having a positive attitude toward AMU and AMR may not necessarily translate into corresponding practices.The results indicate a significant positive correlation between the level of knowledge regarding AMU and AMR among BPF and their corresponding attitudes and practices . SpecifiThis study provided significant insights into the KAP of poultry farmers regarding AMU and AMR in Nepal’s poultry production. The findings underscored the importance of implementing policies that promote the responsible use of antimicrobials in poultry farming and raising awareness among key stakeholders within the framework of the One Health concept to effectively address AMR issues. Additionally, the research highlighted the necessity of fostering sustainable practices and attitudes related to AMU and AMR to address ongoing challenges posed by AMR and ensure the long-term health and sustainability of poultry production.Regarding specific observations, this study noted that the average mortality rate among broiler poultry flocks was 9.3 ± 15.5%, slightly lower than in previous studies . AdditioThis study identified the prevalent classes and types of antimicrobials utilized in broiler poultry farms in Nepal, with tetracyclines being the most commonly used, followed by aminoglycosides, quinolones, polymyxin, penicillin, macrolides, and sulfonamides. These findings align with similar observations made in other low- and middle-income countries (LMICs). For example, in the Kathmandu and Chitwan districts, a prior study reported that the most frequently used antimicrobials were doxycycline 25.9%), tylosin (21.5%), colistin (18.8%), ciprofloxacin (13.4%), and neomycin (12.5%) . In Paki.9%, tyloNepal has implemented regulations governing AMU in food animals, encompassing the Drug Act of 1967, the National Drug Policy introduced in 1995, and the Drug Sales and Distribution Code from 2014. Additionally, they have established a National AMR Containment Action Plan in 2016 . NeverthSeveral factors influence AMU, including the availability of antimicrobials, prescription patterns by veterinarians, potential pressure from drug sellers to reduce excess drug inventory, lack of awareness, and the relatively low cost or easy accessibility of certain antimicrobials for farmers. Despite the Nepal government’s prohibition of colistin sulfate in food animals since August 2019, this antimicrobial continues to be used in poultry farming. One possible reason for the ongoing use of colistin could be the belief that these antimicrobials are effective against common bacterial infections in poultry, such as respiratory and gastrointestinal infections. An integrated surveillance approach encompassing Pakistan, Bangladesh, and Nigeria regarding colistin resistance revealed that the practice of using colistin as a growth promoter persists in LMICs, with no signs of decline, despite its critical classification by the WHO ,39. ThesAccording to this study, most individuals engaged in poultry farming had less than 4 years of experience. This trend may be attributed to experienced farmers leaving the poultry sector due to financial setbacks resulting from various poultry disease outbreaks, such as Newcastle disease, Low Pathogenic Avian Influenza infection, etc. Additionally, the COVID-19 pandemic led to an influx of labor into the poultry sector due to reverse labor migration. Notably, farmers in the age group of 31–40 exhibited a more favorable attitude compared to those aged 18–30. This difference could be due to older farmers being more self-reliant and having greater exposure to training and work experience than their younger counterparts. A previous study on KAP in Bangladesh also highlighted the influence of farmers’ age and experience on AMU and AMR in poultry production . ApproxiThis study has highlighted that farmers, despite having a positive attitude toward AMU and AMR, often lack a comprehensive understanding of the consequences associated with these practices in farm settings. The gap between their knowledge and attitude, resulting in improper practices, can be attributed to several factors, including insufficient education regarding infectious disease control and prevention, limited access to veterinarians, inadequate awareness of AMU and AMR, and less stringent regulations governing AMU in animals. While the majority of farmers demonstrated positive awareness toward the withdrawal periods, the transmission of zoonotic diseases, and the appropriate use of antimicrobials for severe diseases, some farmers did not possess the same level of awareness regarding these critical aspects of AMU and AMR. This suggests that there is room for improvement in education and awareness-raising efforts to ensure that all farmers have a comprehensive understanding of these important concepts and practices. Despite their generally positive attitudes, most farmers (91.6%) exhibited limited knowledge and engaged in improper practices related to AMU and AMR. These knowledge and practice gaps may be attributed to various factors, including the limited availability of veterinarians, diagnostic facilities for disease, and training on the prudent use of antimicrobials and AMR. Notably, a significant proportion of farmers (85.0%) expressed support for the prohibition of non-prescribed antimicrobials for sale. This support may stem from negative past experiences with these drugs and concerns about the impact of AMR on both human and animal health. This statement aligns with our findings on knowledge, with three-quarters of farmers demonstrating awareness of national AMR policies and the concept of withdrawal periods. Additionally, this study revealed that 72.2% of BPF consider vaccination as a potential strategy to reduce the use of antimicrobials in poultry farming. This finding underscores the BPF’s clear understanding of the importance of adopting alternative approaches to reduce AMU in animal production.In this study, it was observed that 50.1% of farmers purchased antimicrobials directly from the drug sellers, which is consistent with findings from previous studies ,40. In cp = 0.008), Koshi , and Sudurpaschim  provinces. Furthermore, the farmers in the Bagmati province demonstrated more positive attitudes than those in the Lumbini province  and better practices than those from the Koshi province . This variation could be attributed to several factors, including higher levels of education among farmers in the Bagmati province and their residence in areas where poultry farming is more commercialized. Moreover, they have increased exposure to veterinary research centers, academic institutes, and veterinary training centers, all of which are situated within the Bagmati province [p = 0.01). These findings underscore the importance of implementing targeted educational and training programs for farmers. These initiatives should prioritize enhancing farmers’ understanding of AMU, particularly regarding the distinction between different types of use and their specific purposes. By promoting the appropriate use of antimicrobials, farmers can contribute to a more responsible utilization of these substances in poultry production.Based on logistic regression analysis, the BPF in the Bagmati province exhibited comparatively higher knowledge regarding AMU and AMR compared to those in the Lumbini  and attitude  toward AMU and AMR. This perception may stem from their awareness of government-led vaccination campaigns aimed at combating major poultry diseases like Newcastle disease, infectious bursal disease, and fowlpox [p < 0.0001) exhibited better AMU practices than those who consulted with drug sellers. This difference may be attributed to the provision of accurate information and the establishment of more effective learning platforms through veterinary consultations, which ultimately result in improved AMU practices.A significant number of farmers believed that vaccines could contribute to the reduction of AMU, and this belief is significantly and positively associated with both their knowledge (OR = 3.5;  fowlpox . TherefoTherefore, this study underscores the critical importance of raising awareness among farmers about strategies to mitigate AMU and emphasizes the significance of consulting veterinarians for poultry health management. To promote responsible AMU practices and achieve more sustainable and effective poultry farming, it is recommended to implement targeted educational and training programs. These initiatives can play a vital role in addressing concerns related to AMR and ensuring the long-term viability of poultry production.This study underscores the critical challenges confronting the poultry production sector, including limited access to veterinary services, the unregulated use of veterinary medications, farmers prescribing antimicrobials directly, and erratic prescriptions by para-veterinarians. These collective factors, coupled with farmers’ lack of knowledge and inappropriate practices, have significantly contributed to the rise of AMR in poultry farming. To effectively combat this issue, a multifaceted strategy is imperative. It should encompass comprehensive education and training programs targeting all relevant stakeholders, the promotion of alternatives to AMU, stringent regulation of veterinary drugs, and the establishment of monitoring and surveillance mechanisms for both AMU and AMR in poultry farming. This proactive approach is essential for mitigating the problem of AMR within the poultry industry and safeguarding the health of both animals and humans.


In this study, an investigation was conducted to determine the influences of ultrasonic fluid pretreatment on the quality of resultant ASDs fabricated from the working fluids. SEM results demonstrated that nanofiber-based amorphous solid dispersions from the treated fluids treated amorphous solid dispersions exhibited better quality than the traditional nanofibers from untreated fluids in the following aspects: 1) a straighter linear morphology; 2) a smooth surface; and 3) a more evener diameter distribution. The fabrication mechanism associated with the influences of ultrasonic treatments of working fluids on the resultant nanofibers’ quality is suggested. Although XRD and ATR–FTIR experiments clearly verified that the drug ketoprofen was homogeneously distributed all over the TASDs and the traditional nanofibers in an amorphous state regardless of the ultrasonic treatments, the  Modern science and technology have greatly promoted rapid developments in pharmaceutics for an “efficacious, safe and convenient” drug delivery, with pharmaceutical techniques and excipients as the focus . New tecThe knowledge and techniques about solid dispersions (SDs) are always being updated due to the continuous increase of poorly water-soluble drugs from high-throughput chemistry and bioengineering . In geneElectrospinning (or electrostatic spinning), initially a textile method, is entering pharmaceutics, particularly the development of ASDs of poorly water-soluble drugs  a working fluid, mainly consisting of a functional ingredient  and a pharmaceutical polymer  in an organic solvent ; 2) elecIn the present investigation, a working fluid composed of keteprofen (KET) as a model poorly water-soluble drug and ethyl cellulose as a drug carrier and the filament-forming polymeric matrix was prepared. Ethyl cellulose (EC) is a derivative of cellulose, which is a popular source for other drug carriers to modify controlled drug release profiles . The worEC was obtained from Shanghai Haosheng Bioengineering Company . KET (white powders with a purity of 98%) was bought from Hengrui Pharmaceutical Company . The organic solvents of anhydrous ethanol and dichlorometahne (DCM) were purchased from Shanghai Merck Co., Ltd. . All other chemicals were of analytical grade, and water was double-distilled immediately before usage.A 6.0 g sample of KET was placed into a 100 mL solvent mixture of ethanol and DCM  to form a drug solution. Subsequently, 22.0 g EC powder was placed into the drug solution. Mechanical stirring was performed for over 10 h to promote the dissolution of EC and homogenization of the co-dissolving solution. Two batches of the solutions were prepared in parallel. One of them was directly electrospun into nanofibers. The resultant nanofibers were termed as the traditional nanofibers F1 or amorphous solid dispersions (ASDs). The other nanofibers were treated using ultrasonic waves in an ice-water bath for 5 min, and later were subjected to electrospinning. The resultant nanofibers were termed as nanofibers F2 or treated amorphous solid dispersions (TASDs).3. The samples were directly fixed in the bath with the water level higher than the level of electrospinnable fluids.A BKE-1004DHT Digital Display Ultrasonic Cleaning Machine  was exploited to conduct the ultrasonic treatment. The water bath had a size (lenght×width×hight) of 300 × 240 × 150 cmThe working fluids were carefully loaded into a 10 mL syringe, which was fixed on a Cole-Parmer fluid driver. A blunt needle (G21) was utilized as the spinneret. The electrospun nanofibers were deposited on a collector, which was 20 cm away from the nozzle of spinneret and composed of an aluminum foil around a hard cardboard. After some pre-experiments, the applied voltage and the fluid flow rate were fixed at 14 kV and 2.0 mL/h, respectively. All the preparations were conducted at the ambient conditions . The samples were cut from the deposited nanofiber mats, and bound on a conductive adhesive to receive the sputtering of gold. The images were recorded under an applied voltage of 10 kV at different magnifications. The average diameters were estimated using ImageJ software  by randomly selecting approximately 100 locations in the SEM images.The amorphous state was measured using the X-ray diffraction (XRD) patterns. The raw powders of EC, KET, and their F1 and F2 nanofibers were measured in the 2θ range of 5°–60°. A Bruker X-ray diffractometer  with CuKα radiation was operated in the conditions of 40 kV and 30 mA.-1 with a resolution of 2 cm-1.Attenuated total reflection–Fourier-transform infrared (ATR–FTIR) spectroscopy was utilized to determine the compatibility between EC and KET in the electrospun nanofibers. A Spectrum 100 FTIR Spectrometer  was employed at a range of 500–4,000 cmλmax = 260 nm with a UV–vis spectrophotometer . The KET contents in the nanofibers were calculated through the pre-determined calibration equation. Thus, the EE% can be determined with the following equation:m represents the measured KET contents in the electrospun nanofibers and Wc is the originally added KET for preparing working fluid. All measurements were repeated three times.The KET entrapment efficiency (EE%) was measured using the following steps: the medicated nanofibers were accurately weighed and extracted in anhydrous ethanol. The solution was diluted using distilled water. The absorbance was measured at Chinese Pharmacopoeia (2020 Ed.) was adapted. Approximately 200 mg nanofibers was placed into 900 mL PBS, and kept at 37 °C and a rotation rate of 50 rpm. At predetermined time points, a 5.0 mL aliquot was withdrawn, and 5.0 mL of fresh PBS was added. The absorbance at λmax = 260 nm was measured, and the amounts of KET could be calculated. All experiments were repeated six times.The KET release profiles from the medicated nanofibers were assessed in phosphate buffer saline (PBS). The basket method in the in vitro dissolution tests of F1 and F2 nanofibers were analyzed using one-way ANOVA. The threshold significance level was set at 0.05. Thus, p (probability) values lower than 0.05 were considered statistically significant.The results from the The electrospinning process is only one step, and the nanofibers are generated in a straightforward and direct manner . The worSubsequently, the working fluids were divided into two sections, as shown in In comparison, they exhibit the following differences: 1) F2 nanofibers have a smoother surface than F1 nanofibers, which were bumpy and rough; 2) F1 nanofibers had a broader diameter distribution  of F1 and F2 nanofibers were 101.31% and 99.58%, respectively. Just as anticipated, the ultrasonic treatments on the working fluid resulted in no drug loss.in vitro drug release characterizations of F1 and F2 nanofibers are presented in in vitro dissolution profiles of F1 and F2 nanofibers were significantly different. In another expression method, an interpolation technique was exploited to achieve the time points that were needed for releasing a certain percentage of the loaded drug. The results are included in The homogeneous distribution of KET molecules within the EC matrices can benefit a stable and continuous release, which is a key application of ASDs as sustained drug release materials. The The Peppas equation  is exploThese results indicate that KET was releases through a typical Fickian diffusion mechanism from both F1 and F2 nanofibers, as suggested by the power exponent values of 0.34 and 0.38, smaller than the judgement value of 0.45. Thus, the ultrasonic treatment had no influence on the drug release mechanisms from the final electrospun products.Thus, the key is that there are several factors of nanofibers’ morphologies, which can exert their influences on the release behaviors of drug molecules (diagrammed in in vitro dissolution tests verified that the F2 nanofibers could provide a better sustained drug release performance than nanofibers with a smaller initial burst release and a longer sustained release effect.With EC and KET as the drug carrier and active ingredient, respectively, two kinds of amorphous solid dispersions were prepared from the same working fluids using electrospinning. Before electrospinning, one of the working fluids was treated using ultrasonic waves for 5 min and later converted into F2 nanofibers; the other was directly electrospun into the traditional nanofibers F1. XRD and ATR–FTIR clearly demonstrated that both F1 and F2 nanofibers were amorphous solid dispersions and the drug and carrier had good compatibility. However, SEM results revealed that the F2 nanofibers from the fluids treated with ultrasonic waves were of better quality than those from the untreated fluids in terms of linear morphology, smooth surface, and diameter distribution. Furthermore, the In this study, one of the most common poorly water-soluble drugs, KET, and one of the most common polymeric excipients, EC, were selected as a model drug and a filament-forming polymeric matrix, respectively, to determine the profound influence of embedded bubbles in working fluids on the electrospinning processes, the resultant nanofibers, and their controlled drug release performance. ASDs are always a focus for resolving the dissolution of poorly water-soluble drug. The disclosed strategy based on these common materials should make it suitable and useful to develop many other sorts of novel ASDs for promoting the dissolution of poorly water-soluble drugs or modifying their release profiles. Meanwhile, the protocols reported here should be a good reference for how to optimize the experimental procedures for creating high-quality nanofibers, and how to evaluate potential nanofiber-based products in future.


Psychiatric patients may refer to concepts neither medically accepted nor easily understood to describe their experiences when seeking medical care. These concepts may lie outside the clinician’s cultural references and consequently hinder the diagnostic consultation. In the fifth version of the Diagnostic and Statistical Manual of Mental Disorders (DSM-5), the clinical instrument Cultural Formulation Interview (CFI) was included. The CFI aims to facilitate the gathering and synthesis of culturally relevant clinical information. The notion of Cultural Concepts of Distress (CCD) was also introduced in DSM-5. The CCD include the subterms of the cultural syndrome, cultural explanation, and cultural idiom of distress. No previous study has used CFI for conceptualizing a cultural notion as a CCD. This study aimed to approach the cultural notion of being a highly sensitive person (HSP) in patients with bipolar disorder (BD) by applying the CFI. The cultural notion of HSP has garnered great interest globally, although scientific evidence is limited. No direct correlation between BD and HSP was hypothesized before or during the study process.In this case study, three patients with BD who reported being HSP were interviewed using the CFI. Furthermore, the applicability of the CCD was examined based on the outcomes of the CFI using an interpretive approach.All three patients reported that the CFI facilitated the clinical consultation, and in one of the cases, it may also have increased the treatment engagement. Based on the synthesis of the CFI outcomes in these illustrative cases, HSP could be understood as a cultural syndrome, a cultural explanation, and a cultural idiom of distress.By applying a person-centered perspective, CFI was used for the conceptualization of a cultural notion as a CCD . Moreover, the cases highlight the complexity of illness insight in BD as a medical phenomenon when patients’ illness perspectives are taken into consideration. Future studies need to further examine the clinical relevance of the CFI in the management of BD. A common clinical challenge in the encounter between patient and clinician is grasping the patient’s narrative and the importance of her lifeworld in order to elicit a deeper understanding . During To address these challenges several changes were made in the fifth version of the Diagnostic and Statistical Manual of Mental Disorders DSM-5). A major change was the inclusion of the clinical instrument Cultural Formulation Interview (CFI) to facilitate the gathering and synthesis of culturally relevant clinical information . The cor. A majorCultural syndromes are patterns of symptoms that are recognized within a particular cultural group but may not fit neatly into existing diagnostic categories. They can be approached as local distress categories based on layman’s theories which are supported by collective experience and anecdotal evidence. Thus, these syndromes reflect culturally specific ways of experiencing distress. Cultural idioms of distress refer to culturally specific expressions or phrases that people use to describe their distress. They can include metaphors, proverbs, or symbolic language that captures their emotional or psychological state. The CCD also encompasses the cultural beliefs and explanations about the causes, nature, and meaning of mental health problems. These explanations may involve religious, spiritual, or supernatural beliefs, as well as social and environmental factors. Hence, cultural explanations of illness provide a framework for understanding distress and guide help-seeking behaviors within a cultural context.Another important change in the DSM-5 was the introduction of the Cultural Concepts of Distress (CCD) partly in response to the critique of the previous notion of culture-bounded syndromes . The CCDHighly sensitive person (HSP) is a Western concept coined by the American psychologist Elaine Aron in the late nineties  utilizing the CFI and testing the relevance of the CCD in a real-life clinical setting. BD is a severe chronic mental disorder with a high burden of disease and excess mortality , 29. Theth European Congress of Psychiatry  using the core CFI as part of the clinical consultation. The collected data were discussed and analyzed with other colleagues  as a part of an internal educational program on cultural competence. The interpretation and synthesis of the CFI outcomes were based on the theoretical framework of the cultural material available in the DSM-5 . No otheychiatry . The Sweychiatry , 34.The three cases are presented below. The patients were of Swedish origin or second-generation immigrants with Swedish as their native language. All three patients reported that the CFI facilitated the clinical consultation by eliciting a person-centered approach. In the second case, the use of CFI positively affected the treatment engagement. However, there was no reported change in treatment engagement in the first or the third case.A woman in her mid-twenties was referred to an outpatient clinic for a post-discharge appointment after hospital admission. She was diagnosed with BD type I during the hospital admission. The visit occurred two weeks after the involuntary admission for a manic episode with psychotic features. Lithium treatment had been initiated during the hospital admission as a long-term prophylaxis of new mood episodes. However, the patient had discontinued lithium after the hospital discharge but wanted to come to the follow-up visit to the outpatient clinic. The CFI was conducted as part of the first outpatient consultation. She wasn’t interested in taking any medication but curious about the outcomes of the diagnostic investigation. She had doubts about BD and psychiatry in general. Other family members had been diagnosed with BD, but she could not see any similarities between their behavioral patterns and her own. However, her family members’ experiences and stories negatively affected her expectations from psychiatry. She referred to herself as being a HSP who needed to take it easy. Many persons from her social network happened to identify as HSP. She felt well-understood and supported by the community of HSP. Indeed, she explained her hospital admission as necessary after a mental breakdown due to her high sensitivity. Stimulus reduction was namely a significant measure for her recovery, according to her. After two more follow-up visits, the patient discontinued her contact with the outpatient clinic.For this patient, HSP was an important part of her cultural identity. She gave meaning and understanding to her experienced distress through the lens of the collective experience of HSP. Self-identifying as HSP was incompatible with also having a mental disorder. The self-attribution of HSP was far more than communicating or explaining her distress. The concept of the cultural syndrome was found to conceptualize her narrative better than the terms of the cultural idiom of distress and cultural explanation.A man in his mid-fifties was admitted to the psychiatric hospital due to bipolar depression with psychotic symptoms. He had been diagnosed with BD type I two decades ago and had been on a long-term lithium treatment since then. He had a good treatment response to lithium. However, a few months before the psychiatric admission, he interrupted his lithium treatment and discontinued his contact with the outpatient clinic after reading a book on HSP. The CFI took place during the last phase of the hospital admission, where the patient gradually improved, but he was still unwilling to restart with lithium treatment. During the CFI, he could describe in detail how the HSP concept explained many of his psychiatric symptoms. He regarded HSP as the main cause of his bipolarity, and to find balance in his own life, he needed to engage with that sensitivity actively. During the interview, the patient argued for his theory of the causes behind his chronic mental health issues, and he was convinced that he needed to continue adjusting his life based on the general recommendations for HSP. The clinician tried to bridge the differences between the explanatory models . At the end of the interview, the patient accepted lithium as a complementary therapy in his HSP-related lifestyle interventions. The patient continued with lithium treatment and has been without mood episodes for at least two years.The patient, although skeptical of the psychopharmacological treatment, did not deny the relevance of the psychiatric diagnosis of BD. It remained uncertain to which extent he could accept a biomedical explanatory model of his mental distress except for his self-diagnosed genetic predisposition to sensory sensitivity. In a sense, HSP was perceived as the underlying pathophysiologic mechanism of his mood states. The concept of cultural explanation could conceptualize adequately the use of HSP in his illness narrative.A woman in her late thirties diagnosed with BD type II was interviewed with the CFI during an annual check-up visit at an outpatient clinic. During the CFI, she described how she experienced and communicated her distress to others. She found the diagnosis of BD stigmatizing and often a barrier to seeking understanding and support from her network. She preferred to describe herself as “highly sensitive” rather than “bipolar” or “vulnerable,” as her disorder was not all about distress. Her sensitivity could be a source of creativity, sympathy, and emotionality. She appreciated that the clinician was interested to know more about her interpretation and management of her disorder. If doctors wanted to refer to her as “bipolar,” she would accept it as long as it did not leave the room. She already had good medication compliance, which did not change after the CFI assessment.In this case, the concept of the cultural idiom of distress was found relevant when approaching her illness narrative. The patient showed a similar understanding of her illness as her psychiatrist, without a report of parallel explanatory models based on HSP traits. The patient had good adherence to treatment without any change after the use of CFI.In this study, we argue that the concept of HSP can be approached as a CCD. This argument is based on the synthesis of the CFI outcomes in the illustrative cases described above, where HSP could be understood as a cultural syndrome, a cultural explanation and as a cultural idiom of distress. To our knowledge, this is the first study using the clinical instrument of CFI to conceptualize a cultural notion as a CCD. Furthermore, this study demonstrates the potential clinical usefulness of CFI and the relevance of CCD, even when encountering patients with Western cultural representations. Lastly, this study also comments on the ongoing ¨looping effect¨  between science and local concepts of illness , 35.It has been suggested that several conditions need to be in place for a cultural representation to be popularized , 37. IndIt must be emphasized that we do not argue for a direct correlation between BD and HSP. The aim of the study was neither to examine the validity of HSP as a clinical phenotype nor its utility as a diagnostic concept. However, the relevance of emotional processing in BD has previously been discussed in the literature . In thisFurthermore, the presented cases highlight the complexity of illness insight in BD as a medical phenomenon when patients’ illness perspectives are taken into consideration. Indeed, illness insight is a well-debated topic, and it may be understood better as a continuous, dynamic, and multidimensional concept , 43. MorThis case-study has some limitations. The main conclusions are based on the synthesis of the CFI outcomes in three selected patients without applying a systematic case study analysis. Using this approach, a hypothesis may be generated but not examined in depth . MoreoveFuture studies need to further examine the clinical relevance of CFI in the management of BD. Three directions may be of interest. The first direction regards the diagnostic accuracy of BD. The diagnostic reliability of BD may be vulnerable from a sociocultural perspective , 53. BiaThe original contributions presented in the study are included in the article/supplementary material. The data collected in the study are presented as fully as possible without revealing individual identity and no additional data is available.The studies involving human participants were reviewed and approved by Swedish Ethical Review Authority, Sweden. Written informed consent for participation was not required for this study in accordance with the national legislation and the institutional requirements.MI and SS were involved in the conceptualization of the study. MI conducted the interviews, undertook administration of the project, and wrote the first draft of the article. SS supervised the study. All authors were involved in the review, and editing of the article, and approved the submitted version.


Autism spectrum disorder (ASD) is a complex neurodevelopmental disorder that is highly phenotypically and genetically heterogeneous. With the accumulation of biological sequencing data, more and more studies shift to molecular subtype-first approach, from identifying molecular subtypes based on genetic and molecular data to linking molecular subtypes with clinical manifestation, which can reduce heterogeneity before phenotypic profiling.In this study, we perform similarity network fusion to integrate gene and gene set expression data of multiple human brain cell types for ASD molecular subtype identification. Then we apply subtype-specific differential gene and gene set expression analyses to study expression patterns specific to molecular subtypes in each cell type. To demonstrate the biological and practical significance, we analyze the molecular subtypes, investigate their correlation with ASD clinical phenotype, and construct ASD molecular subtype prediction models.The identified molecular subtype-specific gene and gene set expression may be used to differentiate ASD molecular subtypes, facilitating the diagnosis and treatment of ASD. Our method provides an analytical pipeline for the identification of molecular subtypes and even disease subtypes of complex disorders.The online version contains supplementary material available at 10.1186/s12859-023-05278-0. Autism spectrum disorder (ASD) is a severe neurodevelopmental disorder that is characterized by deficits in social communication and the presence of repetitive and restricted patterns of behaviors and interests . ASD hasIn the last decades, researchers have attempted to define subtypes of ASD. Traditionally, researchers first identify different clinical phenotypes and then identify and compare the biomolecular factors that may explain the differences in disease manifestation. For example, Diagnostic and Statistical Manual for Mental Disorders, version 5 (DSM-5)  defines CHD8 [DYRK1A [With the accumulation of sequencing data, studies are shifting to a genetic and molecular data-first approach for subtype definition , known aCHD8  and DYRK [DYRK1A , have alRecently widely used single-cell RNA sequencing (scRNA-seq) and single-nucleus RNA sequencing (snRNA-seq) technologies have the advantage of detecting heterogeneity between cells and distinguishing different cell types. In 2019, Velmeshev et al. published snRNA-seq data of human brains from ASD patients and healthy controls and analyzed cell type-specific gene dysregulation in ASD . ASD is To this end, we utilize similarity network fusion (SNF)  to integhttps://autism.cells.ucsc.edu. The matrix of raw counts includes 104,559 nuclei of 15 ASD patients and 16 control individuals, involving 41 post-mortem tissue samples from anterior cingulate cortex and prefrontal cortex. These nuclei are classified into 17 cell types, including fibrous astrocytes (AST-FB), protoplasmic astrocytes (AST-PP), endothelial, parvalbumin interneurons (IN-PV), somatostatin interneurons (IN-SST), SV2C interneurons (IN-SV2C), VIP interneurons (IN-VIP), layer 2/3 excitatory neurons (L2/3), layer 4 excitatory neurons (L4), layer 5/6 corticofugal projection neurons (L5/6), layer 5/6 cortico-cortical projection neurons (L5/6-CC), microglia, maturing neurons (Neu-mat), NRGN-expressing neurons I (Neu-NRGN-I), NRGN-expressing neurons II (Neu-NRGN-II), oligodendrocytes and oligodendrocyte precursor cells (OPC). We preprocessed the raw data with R package scran [We collected single-nucleus RNA-seq data of ASD and controls  from thege scran , normalikcdf was set to Gaussian, and min.sz was set to 10.Gene set variation analysis (GSVA)  is a nonSimilarity network fusion (SNF)  is a comaffinityMatrix function to calculate a patient-patient affinity matrix for each cell type, setting the number of neighbors K = 3. Next, we used the SNF function to fuse affinity matrices of all cell types, setting the number of neighbors K = 3 and the number of iterations T = 100. Based on the fused affinity matrix, patients were clustered using spectral clustering. We used the optimumNumberOfClustersGivenGraph function in SNFtool, i.e., utilizing eigen-gaps [SNF function again, integrating the affinity matrix based on the gene expression matrix and that based on the gene set expression matrix.For the gene expression data of cells in each cell type, we first calculated the average expression of each patient across all cells. Then we used the gen-gaps , rotatiogen-gaps , and alsgen-gaps  to deterBased on the identified ASD molecular subtypes, we analyzed subtype-specific differentially expressed genes (DEGs) and differentially expressed gene sets (DEGSs). For each molecular subtype and each cell type, we firstly identified the DEGs for cells from patients belonging to the considered subtype relative to cells from patients belonging to the other subtypes, and also identified the DEGs for cells from patients belonging to the considered subtype relative to cells from normal controls. The intersection of these two sets of DEGs was taken as the subtype-specific DEGs. For this, MAST  was usedP-value between individual-level fold changes and combined clinical scores. Next, we determined the meta Pearson’s P-value by combining the P-values of all genes of subtype-specific DEGs/DEGSs using Fisher’s method [P-value was used as an approximation of how well the changes of genes correlate with the clinical severity of ASD. To analyze the different clinical ADI-R category in score ranks between one considered molecular subtype and the other subtypes, we performed t test for each category, i.e., A, B-verbal, B-nonverbal, C and D, and the t test P-values of all categories were corrected using Benjamini–Hochberg procedure [P-values.We used the Autism Diagnostic Interview-Revised (ADI-R) data of patients obtained from the ASD snRNA-seq study , includis method . Meta P-rocedure  to obtaiR package caret  was usedTo integrate gene expression and gene set expression of multiple human brain cell types, we performed similarity network fusion (SNF) analysis. Specifically, based on the snRNA-seq data of ASD patients, SNF was applied to integrate the patient-patient similarity networks obtained from gene expression data of multiple cell types. At the same time, for each cell type, we supplied gene expression data and curated gene sets for gene set variation analysis (GSVA) to obtain gene set expression data, and then SNF was applied to integrate the patient-patient similarity networks obtained from gene set expression data of multiple cell types. Next, we used SNF again to integrate the above patient-patient similarity networks to get a final patient-patient affinity matrix. Then we performed clustering based on the affinity matrix to determine the molecular subtypes of ASD patients Fig. A. Based Using the gene expression data of multiple cell types, we used SNF to obtain a patient-patient affinity matrix. Based on eigen-gaps , rotatioBased on the three recognized ASD molecular subtypes, we identified subtype-specific DEGs for each molecular subtype and each cell type , we used the human brain bulk RNA-seq data of 47 ASD samples  and 57 controls  [P-value < 0.05) in the vast majority of cells  (released on 22 July 2022), of which 429 genes are in our gene expression matrix. The molecular subtype-specific DEGs are significantly enriched with ASD genes , indicating that the identified Sn molecular subtypes are associated with clinical symptoms . There may be also a tendency that A  score rank of Sn molecular subtype II is lower than those of the other subtypes , though it is not significant. These observations may be helpful in the diagnosis of molecular subtypes of ASD, which then facilitates the treatment of ASD. The association analysis with ADI-R or other clinical data, linking ASD molecular subtypes with clinical manifestation, would be more practical when more clinical data become available in the future.As to the analysis of different clinical ADI-R category in score ranks between one considered Sn molecular subtype and the other subtypes, we performed rocedure  (AdditioConstructing machine learning-based disease prediction model can facilitate the clinical diagnosis of diseases –43. UsinIn addition, we also considered to constructed predictive models at the patient level. The gene expression average of all cells of one patient was used as the patient’s gene expression data. For each subtype, we randomly selected one patient as a test set and the left patients as a training set. Because the sample size is small and to avoid the effect of randomness, we repeated the data set partition and model construction ten times and then calculated the average prediction accuracy to assess the effectiveness. The average prediction accuracy values are 0.6 for IN-PV, and above 0.7 for all other cell types. This reflects that these three subtypes can be well distinguished even in a cell type with a small sample size.As to molecular subtype-specific DEGSs, we also used them to build ASD subtype prediction models. Because only in L2/3 all three molecular subtypes possess subtype-specific DEGSs, we built prediction model only in L2/3 using the genes contained in all subtype-specific DEGSs as features. First, when 70% of cells from all patients were selected as the training set and the other cells were used as the test set, macro F1, micro F1 and weighted F1 are 0.72, 0.69 and 0.71, respectively. Second, when all cells from one patient in each molecular subtype were selected as the test set and cells from other patients were used as the training set, macro F1, micro F1 and weighted F1 are 0.81, 0.79 and 0.80, respectively. Finally, when one patient from each molecular subtype was selected as the test set and the other patients as the training set, the average accuracy of 10 training/testing replicates is 0.73. When more patient sequencing data become available, the construction of predictive models at the patient level to predict the molecular subtype of a patient will hopefully be realized.The identification of ASD subtypes remains an unresolved challenge, which adds a barrier to the treatment of patients with ASD. Traditional methods of identifying ASD subtypes defined by clinical behaviors directly may just provide limited assistance in the genetic genesis and treatment of ASD. With the recent accumulation of biological sequencing data, studies are shifting to a genetic and molecular data-first approach to subtype definition, i.e., identifying molecular subtypes first. Considering the fact that ASD is characterized by cell type heterogeneity and the presence of ASD-associated gene sets may determine the manifestation of ASD in different cell types, our study innovatively integrated gene and gene set expression data of multiple human brain cell types using SNF to define ASD molecular subtypes.To explore the characteristics of the identified molecular subtypes, we analyzed their cell type-specific and subtype-specific DEGs. All three subtypes have a large number of subtype-specific DEGs in L2/3, L4 and L5/6-CC, which is similar to the result of the ASD snRNA-seq study , indicatExcept for subtype-specific DEGs, we also identified molecular subtype-specific DEGSs. The subtype-specific DEGSs of the three molecular subtypes are distinctive no matter in the number of identified DEGSs or in the regulatory orientation, with molecular subtype I having more down-regulated DEGSs in L4 and molecular subtype III having more up-regulated subtype-specific DEGSs in IN-VIP. Furthermore, some gene sets of molecular subtypes I and III show a high correlation with the clinical severity, such as GOMF_STRUCTURAL_CONSTITUENT_OF_RIBOSOM and GOCC_RIBOSOMAL_SUBUNI of subtype I.The subtype-specific DEGs/DEGSs unique to molecular subtypes and the common subtype-specific DEGs/DEGSs across all subtypes but with different regulatory orientations can be used as distinguishing genes/gene sets for different molecular subtypes. To demonstrate their prediction ability, we constructed prediction models. Also, using the differences in clinical scores of the identified molecular subtypes would be helpful to diagnose the molecular subtype of ASD. Our results may aid in the identification and diagnosis of ASD molecular subtypes, and even disease subtypes. Our method can be further practically applied in the future when more ASD scRNA-seq/snRNA-seq data and clinical data are available.In this study, we identified ASD molecular subtypes by performing similarity network fusion to integrate gene and gene set expression data of multiple human brain cell types. Then we applied subtype-specific differential gene and gene set expression analyses to study expression patterns of different molecular subtypes. The identified molecular subtype-specific genes and gene sets may be used as biomarkers to classify ASD molecular subtypes, facilitating the diagnosis and treatment of ASD. Our method can also be applied for the identification of molecular subtypes and even disease subtypes of other complex disorders.Additional file 1. Table S1: Molecular subtype-specific DEGs information, the enrichment with SFARI ASD genes, and the association with clinical scores.Additional file 2. Table S2: Molecular subtype-specific DEGs unique to subtypes along with the regulation direction and |logFC| relative to other molecular subtypes, and those differentially regulated among different molecular subtypes relative to controls.Additional file 3. Table S3: GO and KEGG enrichment analysis of molecular subtype-specific DEGs.Additional file 4. Table S4: Molecular subtype-specific DEGSs, the enrichment with SFARI ASD genes, and the association with clinical scores.Additional file 5. Table S5: Molecular subtype-specific DEGSs unique to subtypes along with the regulation direction and |logFC| relative to other molecular subtypes, and those differentially regulated among different molecular subtypes relative to controls.Additional file 6. Table S6: The clinical information of ASD patients in each molecular subtype, and the result of t test between each molecular subtype and the other two for each category of ADI-R scores.


Allogeneic stem cell transplantation  is the optimal treatment strategy for adult acute leukemia patients due to its ability to reduce the risk of relapse. The ELN group recommends the consideration of alloSCT in fit adults with AML in CR1 who have a predicted relapse risk of 35 to 40% and a suitable donor ; in this context, in a monocentric experience,  focused Baseline genomic features seem to have an important role in predicting survival regardless of the age of AML patients. On the same topic,  in a sinIn this context, the intensification of the conditioning regimen may be able to improve the prognosis in terms of better survival. In a retrospective matched pair study,  analyzed focused  focused In summary, the collection of articles on this Research Topic provides an update on the progress in the understanding of both biological characterizations of AML and therapeutical approaches before and after transplant in order to reduce relapse. Molecular stratification has become fundamental not only for risk stratification but also for the identification of druggable mutations that could be used not only in the phase preceding transplant in order to reduce minimal residual disease but also in the post-transplant period as a further consolidation of the remission obtained. From the review of  and the PC wrote the Editorial, FC and SS approved it. All authors contributed to the article and approved the submitted version.


Eponyms are commonplace in the medical vernacular, however, their use has become increasingly controversial amongst clinicians. Whilst some view them as an honour bestowed on those whose achievements deserve recognition, others see them as thwarted with problems due to confusion, imprecision and unwittingly applauding controversial figures. Nevertheless, the history and culture retained within eponyms define modern-day medicine. To identify current trends in understanding of eponyms, we presented a questionnaire of orthopaedic eponyms and their associated imaging to unspecialised trainees, specialist orthopaedic trainees, and qualified consultants. Eponymous terms were poorly understood at all levels of experience, with- third and fourth-year Orthopaedic trainees ) being outperformed (22.3%) by non-specialist postgraduate doctors with two or more years of experience (foundation year two (F2) and core surgery year two (CT2)) (29.3%). Based on these trends we present a further narrative review of the challenges eponyms present, whilst justifying their continued use to acknowledge the origins of our discipline, from the favourable to shameful. Eponyms are commonplace throughout medicine. Their roots lie in the historical figures after whom they are named, and to whom much of modern medicine owes tribute. One such example is the term Charcot foot or joint, described by Jean-Martin Charcot for neuropathic osteoarthropathy. The disease refers to the peripheral loss of sensation and proprioception resulting in the absence of protective feedback from repetitive microtrauma. This leads to damage to skin integrity, soft tissue and bone infection, bony destruction and deformity of the joint, and eventual loss of function or loss of limb . The rolClinicians are often required to make quick and accurate decisions in time-pressured environments. It would follow that improved efficiency in conveying information through the use of eponyms would be reasonable and perhaps invaluable. Descriptive methods can be cumbersome to communicate, which also may favour the use of eponyms, whose understanding conveys specific information relating to the condition, its causes, and its associations.However, this is only the case if eponyms are accurately understood. Arguments against the use of eponyms are that they are an outdated and archaic form of communication. Learning the vocabulary of medicine is like studying another language, there are already a multitude of medical conditions to learn and an ever-increasing number of classification systems . The necOthers would argue that many of the eponymous terms we use must be updated either due to inaccuracies of the original describer, who some have found may not have even been the first to identify the traits or pathology they describe . AnotherMedicine is a culture of international contribution and collaboration, and this is directly visible through our eponyms. In 1912, Hakaru Hashimoto of Japan first identified what he termed struma lymphomatosa, at the time a new form of lymphoid and plasma cell infiltration within the thyroid. At the time, Hashimoto was training in Germany, however, his findings were seen to be indistinct from Riedel’s thyroiditis, a form of chronic thyroid inflammation. It was not until 1931 when an American, Allan Graham, supported his assertions that the term Hashimoto’s thyroiditis was coined. Eponyms not only demonstrate global contributions to the field but are used in all corners of the globe .We hypothesise that level of understanding of eponymous terms among orthopaedic clinicians is generally poor, but that knowledge will peak as trainees approach consultancy and then decline as they become more specialised. We will further provide a narrative review of the current challenges with the use of eponymous terms and will argue that this should be an area for possible development and revival to improve clinical practice and efficiency, as well as the overall appreciation of our medical culture.This article was previously presented at the 2022 42nd SICOT Orthopaedic World Congress on 28-30 September, 2022 and the 2022 British Trauma Society Annual Scientific Meeting on23-24 November, 2022.Two authors (EP and AK) produced an orthopaedic questionnaire focussing on the correct matching of eponymous fractures and orthopaedic pathology with their respective radiographic imaging, or illustrations of pathology. The included topics were chosen via consensus decision to appropriately cover a diverse range of levels of knowledge, which would be expected of trainees at different stages in their training.We presented the questionnaires at live events through the medium of a Microsoft PowerPoint presentation  (see Appendices). A total of 25 eponymous conditions over 25 slides were assessed. Each slide consisted of five different radiographic or illustrative options with 15 seconds given per slide to review the images. Answers were gathered anonymously, with participants using a quick response (QR) code linked to a Google Forms answer sheet .We collected data at separate training events for foundation trainee doctors , senior house officers , senior trainees , and consultant orthopaedic surgeons within the London region.We used descriptive statistics , and percentages as appropriate) to summarise collected data and performed analysis with Microsoft Excel  by one author (EP) with the consultation of senior authors .Informed consent was obtained from all individual participants included in the study. Ethical approval was not deemed necessary by the local committee as the research involved the use of non-sensitive, completely anonymous educational tests and surveys where participants were not defined as “vulnerable” and participation would not induce any undue psychological stress or anxiety.A total of 34 clinicians completed the questionnaire. Of these, 12 were considered junior trainees (foundation trainees and senior house officers) and 22 were specialists . A total of 25 different eponymous terms were assessed, these were subdivided into the following sections: seven hand, wrist, and forearm; two upper arm and shoulder; four spine, hip and pelvis; three knee; and nine ankle. Amongst junior trainees, the most correctly identified eponymous term was the Chauffeur’s fracture (54.5%), and amongst specialist trainees, the most correctly identified orthopaedic eponymous terms were Lisfranc, Segond, and Pilon fractures (95.5%)  and was categorised as a consultant for purposes of subgrouping. Amongst junior trainees, the mean number of correct answers for the questionnaire was four 16%) with a standard deviation (SD) of 1.71 and a range from one (4%) to seven 28%) correct answers. Amongst specialist trainees and qualified orthopaedic surgeons, the mean number of correct answers for the questionnaire was 15.3 (61%) with an SD of 3.18 and a range from nine (36%) to 21 (84%) correct answers. As a group, foundation year one trainees performed the worst and ST3 trainees performed the best in the questionnaires  trainees performed the best as a group, with a gradual decline in knowledge of orthopaedic eponyms until post-qualification. Across both groups, the knowledge of hand, wrist, and forearm, and knee eponyms was the greatest; whilst juniors performed worst in foot and ankle eponyms and specialists performed worst in spine, hip, and pelvis eponyms  performs on average worse than a senior house officer (SHO), who has not secured an orthopaedic training post. It is possible that the focus of training on an academic basis for management means that trainees favour more scientific and evidence-based systems. From the distribution we observe, we might imply that as clinicians develop and mature within the specialty, they come to see eponyms as outdated and of little clinical use and begin to disregard the importance of eponymous terminology. However, preparation for examinations may lead trainees to discover or rediscover these eponymous terms. Whilst a second peak at the post-CCT fellow may suggest that as trainees look to organise the knowledge they reencounter these eponyms, we postulate that these orthopaedic eponyms may also represent a valuable learning tool in those much more senior approaching consultancy. We would argue that where trainees are more focused on scientific systems they may be missing out on the value and knowledge offered in eponyms, that may only be realised towards the end of their training.One such solution to this challenge is a universal classification to allow consistent understanding, which we see in the Müller Arbeitsgemeinschaft für Osteosynthesefragen (AO) classification of fractures first published in 1987 . This prAs for the AO classification system, it has its own drawbacks. A 23-A2  does not communicate the mechanism or deforming forces that lead to the fracture. In comparison the eponymous Colles' and Smith’s fracture inform us that the distal fragment is either dorsally or volarly angulated, and likely result from falling forward or backward onto an outstretched hand, respectively. The other argument against this form of classification as a rather cold and unemotional way in which the fracture is described, by simple numbers and letters and that “eponyms bring colour to medicine” .Nonetheless, this vibrancy and history that eponyms bring to our practice becomes redundant if it impedes or is counterproductive to providing concise and accurate communication of information between clinicians. Whilst our study suggested that eponyms of the hand, wrist, and forearms were better understood, whilst 86.4% and 90.9% of specialist correctly identified the Smith’s and Colles' fracture, the remaining mistook it for the other, which was also the case with the Monteggia (90.5%) and Galeazzi (90.9%) fractures. This is likely due to their relative opposition in terms of description with respect to dorsal versus volar angulation (Colles' versus Smith’s) and dislocation of radial head versus distal ulna . This is in agreement with the work of Waseem et al. who found that only 10.7% of clinicians were correctly performing the Finklestein’s test, even though 90.4% of those assessed stated they regularly used it [Another common criticism of eponymous terms is the association of terms with less than savoury historical figures, whose contributions to medicine would preferably not be applauded. In the rebranding of eponymous conditions, often Nazi supporters and sympathisers are used as examples . The welPernkopf’s Atlas of Topographical and Applied Human Anatomy. Pernkopf was a supporter of the Nazi party, being principally involved in the University of Vienna’s removal of Jewish staff and students. Pernkopf’s illustrators were seen to have incorporated Nazi symbols in their signatures to their illustrations and there has been postulation that victims of Nazi atrocities may have made their way into the illustrations of the Atlas, although evidence remains unclear [Whilst during our research we could not identify any specific orthopaedic eponyms with disreputable namesakes, it is possible that in the study of anatomy, orthopaedic surgeons may come across  unclear . The eth unclear , howeverDespite the aforementioned issues with eponymous names that have led some to question the longevity of their usage, we see the potential for their continued use . HoweverLimitationsWhilst our study has demonstrated generally poor level of understanding of orthopaedic eponyms amongst both specialists and non-specialised trainees, there is a lack of power of conclusions due to the limited sample size. Furthermore, conclusions drawn relating to overall group and subgroup performance may not be generalisable due to the inherent sampling bias occurring from selection of participants based on geographical location.Eponymous terminology is in general poorly understood by both junior non-specialised trainees and senior orthopaedic trainees, with a few exceptions. Senior trainees demonstrated some of the worst knowledge, which should be a marker of concern for educators and trainers. The history and culture retained within these eponymous names define much of modern-day medicine and we should ensure our training is as focused on where we have come from as on where we are going. Whether to acknowledge great contributions or to not forget those whose ethics and morality harmed innocents and shamed the profession, we believe eponyms still have their place in our practice.


In contrast to silicon-based transistors, single-moleculejunctionscan be gated by simple mechanical means. Specifically, charge canbe transferred between the junction’s electrodes and its molecularbridge when the interelectrode distance is modified, leading to variationsin the electronic transport properties of the junction. While thiseffect has been studied extensively, the influence of the molecularorientation on mechanical gating has not been addressed, despite itspotential influence on the gating effectiveness. Here, we show thatthe same molecular junction can experience either clear mechanicalgating or none, depending on the molecular orientation in the junctions.The effect is found in silver–ferrocene–silver breakjunctions and analyzed in view of ab initio and transport calculations,where the influence of the molecular orbital geometry on charge transferto or from the molecule is revealed. The molecular orientation isthus a new degree of freedom that can be used to optimize mechanicallygated molecular junctions. Consequentially, the electronic transport characteristicsof the junction may change. Mechanically gated molecular junctionshave been extensively studied both experimentally and theoretically,15 for example in the context of nanoscale image charge9 and optimization of thermoelectricity.14 However, the influence of the molecular orientation onmechanical gating has not been examined. Such an influence can bean attractive route for optimization and regulation of mechanicalgating with implications on charge, spin, and heat transport in molecularjunctions. Here, we show that the mechanical gating of molecular junctionscan be dramatically affected by the molecular orientation, where thesame molecular junction experiences either a clear mechanical gatingor the absence of such an effect, depending on the orientation ofthe molecule with respect to the electrodes. By comparison of experimentsand calculations, this behavior can be related to the orbital natureat the metal-molecule interfaces, allowing the identification of thenecessary conditions for mechanical gating. The reported findingsin this letter show that the orientation of the molecule is an importantfactor for the design of mechanically gated molecular junctions andemphasizes the importance of orbital orientation in the general processof metal-molecule charge transfer.One of the fascinating propertiesof molecular junctions is their ability to function as nanoscale electromechanicaldevices. In particular, single-molecule junctions allow the studyof coupling between mechanical and electronic degrees of freedom ina structure of a typical single-nanometer size that has a dominantquantum nature and a pronounced orbital character. This combinationhas been used to study diverse phenomena, including electron–phononinteraction,Supporting Information). Measurementsof current as a function of applied voltage across the junctions (I–V curves) for different junctionrealizations reveal two distinctive cases, denoted here as type 1and type 2  curves. As will be furtherdiscussed below with the aid of ab initio calculations, the peaksoriginate from the contributions of molecular orbitals to the conductance.Therefore, shifts in the voltage at which the peaks are observed correspondto shifts in the molecular energy levels, with respect to the Fermilevel of the electrodes. Supporting Information, sections 2 and 3 for more details on types 1 and 2, and the reversibilityof the observed level shifts).We studied molecular junctionsbased on suspended individual ferrocenemolecules between two silver (Ag) electrode tips. A break junctionsetup is used to form in situ these molecular junctions 1 in a crd type 2 2a,b. TheI–V characteristics in I/V2) as a function of 1/|V|.20 These plots are expected to have a minimum at a certain transitionvoltage, Vtrans, whenever the currentas a function of voltage evolves from a linear dependence to morethan a quadratic dependence.19 We notethat for metallocene molecules with direct contacts to the electrodes, the transition voltagedependence on the energetic difference between the electrodes’Fermi level and the closest molecular energy level(s) is to date unknown.However, regardless the exact dependence, shifts in the transitionvoltage serve as an indication for shifts in the energy of the molecularlevels that dominate electron transport and/or systematic variationsin their coupling to the continuum states of the electrodes .22 As seen belowusing calculations, we expect dominant level shifts and moderate variationsin electrode–molecule coupling. This is manifested as significantshifts in the calculated transmission peaks with generally modestvariations in the peak widths. The TVS plots in Supporting Information section 2 for similardata for other junction realizations). We can take advantage of thepresence of peaks in the dI/dV thatcorrespond to molecular levels to shed light on the relation betweentransition voltage shifts and molecular level shifts. When the interelectrodedistance is reduced by 0.6 Å, the transition voltage shifts by370 mV. Assuming for simplicity a symmetric voltage drop on each electrode–moleculecontact, which can be justified as a crude approximation by the roughlysymmetric locations of the positive and negative peaks in Supporting Information, section 5), the 690 mVshift in the examined peak location in I/dV curve appears . We can conclude that theobserved mechanical gating leads to shifts in the transition voltagethat are rather similar in magnitude to the roughly estimated shiftsin the molecular energy levels . Our findingsshow that TVS is a good indicator for level shifts in the examinedjunction. The rather linear relation between transition voltage shiftsand level shifts calls for further study into the mechanistic detailsbehind it, which are beyond the scope of this Letter.The same effect can also be presented using transitionvoltagespectroscopy (TVS) plots, where the Supporting Information for details), as presentedin 23To betterunderstand the nature of types 1 and 2, we turn to densityfunctional theory (DFT) and electron transport calculations (see 13 Below the Fermi energy (taken aszero), pairs of narrow resonances can be seen (HOMOs), while abovethe Fermi energy, a single broad resonance is found. Its peak transmissionis 2, pointing to a two-channel or two-orbital origin (LUMOs). Thetransmissions for the perpendicular configuration have a richer structurewith narrow peaks on both sides of the Fermi energy. Also, the evolutionof transmission as a function of stretching is more complex in thiscase than that for the parallel molecular configuration. However,in contrast to the former case, no clear shifts in the transmissionpeaks are observed when the interelectrode distance is modified inthe given range. Thus, mechanical gating is not found for the perpendicularconfiguration. We note that in both configurations the transmissionresonances are highly asymmetric due to quantum interference,24 similarly to the asymmetries reported for molecularjunctions based on a ferrocene derivative.8Supporting Information). When the molecule is oriented parallelto the junction axis, shifts of peaks are seen, as also found in theexperiments for type 1. However, for the perpendicular molecular orientation,a similar effect cannot be found, in agreement with the absence ofmechanical gating in the measurements of type 2. Note that the featuresin Supporting Information, section 5.To have a more transparent comparison with the measured data,thedescribed transmission can be directly converted to calculated differentialconductance, as presented in Supporting Information, section 6). The mechanical gatingresponse for the parallel molecule configuration and the absence ofthis effect for the perpendicular configuration can be ascribed tothe orientation of the molecular orbitals that dominate transportwith respect to the electrodes and their coupling to the frontierelectrode states. Supporting Information). As canbe seen, the junction LUMOs, which are associated with the broad transmissionpeak above the Fermi energy in the parallel configuration, and thetwo narrow peaks above the Fermi energy in the perpendicular configurationresemble the LUMOs of the isolated molecule. These orbitals have asignificant π-character on the carbon rings; i.e., the wavefunction spreads away from the ring plane perpendicularly.25 The remaining contribution to the LUMOs comesfrom the Fe 3d atomic orbitals. Therefore, the molecular LUMOs overlapwith the electrodes much more efficiently in the parallel configurationthan in the perpendicular configuration. This difference makes theLUMO coupling to the frontier electrode states in the parallel configurationmore sensitive to changes in the interelectrode distance , the elongation of Au junctions produces atomic chainsthat may complicate data analysis. Therefore, in this study, we preferredto use Ag electrodes. Finally, we note that our calculations do nottake into consideration the effect of electrode polarization.To conclude, we showed by experiments and calculations that mechanicalgating of molecular junctions depends on the orientation of the moleculein the junction. In the extreme demonstrated case, the same molecularjunction can either experience mechanical gating or not, dependingon the molecular orientation with respect to the electrodes, as wellas on the nature of the interaction between the molecular orbitalsand the continuum states of the electrodes. These findings emphasizethe importance of geometry and local orbital structure in the contextof charge transfer across metal-molecule interfaces and point towarda way to control mechanical gating of charge, spin, and heat transportin molecular junctions.


Antimicrobial resistance is a major threat to human health. Since the 2000s, computational tools for predicting infectious diseases have been greatly advanced; however, efforts to develop real-time forecasting models for antimicrobial-resistant organisms (AMROs) have been absent. In this perspective, we discuss the utility of AMRO forecasting at different scales, highlight the challenges in this field, and suggest future research priorities. We also discuss challenges in scientific understanding, access to high-quality data, model calibration, and implementation and evaluation of forecasting models. We further highlight the need to initiate research on AMRO forecasting using currently available data and resources to galvanize the research community and address initial practical questions. Escherichia coli, Staphylococcus aureus, Klebsiella pneumoniae, Streptococcus pneumoniae, Acinetobacter baumannii, and Pseudomonas aeruginosa. To limit the spread of antimicrobial-resistant organisms (AMROs) and reduce AMR-related disease burden, improved predictive intelligence is required to better estimate the emergence and spread of AMR within populations and healthcare facilities. However, efforts to operationally forecast the burden of AMROs  are not active as of January 2023. Antimicrobial resistance (AMR) is a leading threat to global health  and the evolution of resistance. If used in real time, those predictions would support situational awareness and inform public health policies such as antimicrobial drug stewardship and more targeted antimicrobial prescription guidelines to slow down AMR spread.At the facility level, the forecast target of interest might be the number of AMR infections with clinical symptoms within a hospital or hospital system. Such predictions would support control of nosocomial AMRO transmission and resource planning for equipment, medications, staffing, and space in response to potential patient surges. Depending on the clinical relevance, the forecast horizon might be days or months. Of note, predictive models connecting multiple healthcare facilities in a region could elucidate the risk for AMR introduction through interhospital patient transfer and support decision making for preemptive measures in facilities without ongoing transmission.Although models and data differ considerably for forecasts at various scales, some common challenges impede the development and operational use of predictive models for AMR. Here we summarize these issues and highlight several research priorities to address these challenges in future studies.For forecasting using mathematical and statistical models, it is critical to understand the key processes affecting AMR spread. Those processes are often represented as nonlinear effects in forecasting models and, if not properly specified, will produce forecasts that quickly diverge from the truth. As of 2023, many questions on AMR remain open  and the environment. In addition, individual-level causal relationships between the type and duration of therapy and resistance emergence remain unknown in most instances. The human microbiome serves as a reservoir of antimicrobial resistance  and, therefore, with illness and death rates. Those correlations make definition of the forecasting target  relatively straightforward. However, for bacterial or fungal species, relationships between pathogen load and clinical outcomes are unclear. Because many bacterial species are commensal with their human host and have varying probabilities of presence across body sites, it is challenging to definitively determine whether a person is colonized. Without accurate observation of colonization, AMR burden is not well resolved and, consequentially, is more difficult to forecast.Forecasting is fundamentally a data-driven task. Without sufficient data, predictive models cannot be properly trained and evaluated. As of 2023, data that can inform operationally useful forecasts of AMR remain scarce. At the population level, several surveillance systems do exist. For instance, the US National Antimicrobial Resistance Monitoring System for Enteric Bacteria tracks changes in antimicrobial susceptibility for certain enteric bacteria in ill persons, retail meats, and food animals . There are no guidelines on such implementation for AMRO forecasting, such as appropriate data collection and forecast targets. Questions remain open on the proper time scale of forecast horizon, the frequency at which models need to be updated, and the fundamental limit of predictability of models. For long-lead forecasting, evaluation requires data collection in a consistent manner over a long time period. In healthcare facilities, the practice of testing and reporting AMR infections may change over time, which further complicates using such data records and forecast evaluation. A collaborative effort that standardizes training datasets, forecast targets, forecast horizons, and proper scoring rules for evaluating forecast performance  or the community level , decision makers must make decisions that leverage uncertain information. This truism holds for observations as well as forecasts. Designing optimal decision frameworks and architectures that best use forecasts, given their uncertainty, is a needed long-term goal.Effective communications between modelers and stakeholders such as public health officials, healthcare institutions, and individual practitioners are critical to learn their practical needs from AMR modeling. However, formal reports recording such communications are lacking in scientific journals, which is another factor limiting the generation and use of operational forecasts in real-world settings.Staphylococcus aureus (MRSA) as a concrete example (S. aureus (MSSA) in affecting the dynamics of MRSA. Answering those questions would improve methods to reduce MRSA burden in both community and hospital settings.To illustrate the interconnected challenges faced by AMRO forecasting across scales, we use methicillin-resistant  example . SeveralIn this perspective, we focus on real-time forecasting of AMROs. A parallel line of research is scenario-based simulations that project AMR infections conditional on postulated changes in prescribed interventions or expected conditions. Previous studies for HIV and tuberculosis control show that such scenario-based projections can substantially affect health policies and save lives .Increasing the availability of existing data could also accelerate progress. Electronic health records contain a wealth of AMR data, each of which reflects a certain aspect of AMR-related processes. Synthesizing previously siloed datasets into mathematical models can potentially answer scientific questions that are otherwise challenging to address using each dataset separately. Privacy-preserved data sharing across facilities can increase the amount of data for modeling and support the development of generalizable methods. When data sharing is not practical, models and algorithms can be shared, trained, and implemented with defined standards and quality control.Given existing gaps in forecasting AMR, predictive models are still not mature enough for operational application. To push forward advances in this burgeoning area, several research directions should be prioritized. First, better communication among multiple sectors and stakeholders, including academic researchers, public health agencies, healthcare providers, and the public, will help identify key questions and the needs of end users of predictive models. Developing and applying AMR forecasting will be a collective effort that should address real-world questions in public health and patient care. Second, studies should make better use of existing data and guide the collection of new data that are essential to understand AMR. Investing in consistent surveillance and data collection is of utmost importance for improving understanding of the emergence, spread, and outcomes of AMR. Third, more effective, computationally efficient algorithms are needed to calibrate complex AMR models to multitype and multiscale data. Better interpretability of models can infuse confidence in clinicians when using those tools. Further, research on computational methods that are tailored to AMR prediction could help bridge theoretical models and real-world applications. Fourth, predictive AMR models should be implemented in real-world settings in real time so that operational utility can be assessed by validating real-time operational predictions, as is done for numerical weather predictions. Forecasting skill, including forecast accuracy and uncertainty, should be evaluated to confirm that predictive models can produce useful predictions despite noisy and incomplete data.In summary, despite lessons learned from recent advances in forecasting for other acute infectious diseases, AMRO prediction has its own set of challenges, including wide and prolonged asymptomatic carriage, longer time scales, continuing evolution due to strain competition and antimicrobial drug use, and poorly observed disease burden. It will be critical to set appropriate expectations for the performance of AMRO predictions and establish sensible criteria for successful forecasting.