Antimicrobial resistance is a global problem of increasing proportions that we cannot afford to look away from. This World Antimicrobial Awareness Week, we shine a light on the crisis and ways we can all help to address it. Antimicrobial resistance is a global problem of increasing proportions that we cannot afford to look away from. Our Editorial shines a light on the crisis and ways we can all help to address it. Antimicrobial resistance (AMR) is considered by the World Health Organization (WHO) to be multi-drug-resistant gonorrhea worldwide. World Antimicrobial Awareness Week is a global campaign observed annually on 18–24 November to promote awareness and understanding of AMR and to inform of best practices to reduce its emergence and spread. This year, we are participating in this effort to bring some needed attention to the AMR crisis and how it can be tackled, including the basic and applied research that PLOS Biology would be happy to give a voice to.As we end year three of the COVID-19 pandemic, our response to the AMR crisis—already insufficient before the pandemic—needs urgent action, coordination and funding. The reallocation of resources, overwhelmed hospitals and doctors, and over-prescription of antibiotics during these years have set us back. According to the Centers for Disease Control and Prevention, in the United States alone AMR increased by 15% between 2019 and 2020 [In a Perspective in this issue , DolecekCandida auris, that are already resistant to available antifungals [Even further from the spotlight are fungal and parasite AMR, despite being a threat to both public health and food security. Resistance to antifungals can occur in previously susceptible fungi, but there are also new emerging species, such as the yeast ifungals . With feifungals —in the fAlthough it is a global problem, low- and middle-income countries bear a disproportionate amount of the burden of AMR, irrespective of pathogen. The maxim oft repeated during the COVID-19 pandemic “nobody is safe until we are all safe” also applies here—it is a global responsibility to help all countries deal with the burden of AMR. As Dolecek et al. discuss , infectiantibiotic prescription would be devolved to pharmacists to shorten doctor waiting lists. While the aim was good, the means might prove disastrous.The WHO developed a roadmap to address AMR that was internationally ratified at the 2015 World Health Assembly . This plAMR surveillance is crucial to detect the emergence and spread of resistance in different ecological settings and could, if combined with phenotypic assays, point to new resistance mechanisms . WPLOS Biology has and will continue to cover [CARB-X and GARDP. In one such partnership, GlaxoSmithKline (GSK) and the US Government BARDA office have reported that a new, first-in-class antibiotic, gepotidacin, has been so successful in a phase III clinical trial against urinary tract infections that the trial was halted and GSK will soon file for drug approval. They are also studying efficacy against gonorrhea in another phase III study. Because gepotidacin has a new mechanism of action, it could be effective against drug-resistant bacteria. These encouraging results showcase the potential of these partnerships to drive antimicrobial drug discovery.Development of rapid diagnostic tests, especially those that can be easily deployed in resource-limited settings, and renewed drug discovery efforts are also pillars of the response to the AMR crisis and topics that to cover ,13. DeveIn addition to drugs, investment and research should be geared towards finding vaccines, looking beyond viruses to include those against bacteria, fungi and parasites. The resounding success of the mRNA vaccines against COVID-19 has given new impetus to the possibility of using new technology to find effective vaccines against pathogens for which slow progress had previously thwarted vaccine development.The pandemic threw us off track, and the current world situation with war in Ukraine, drought and famine in large regions of Africa, flooding and other devastating effects of climate change, as well as a global economic recession, pulls our collective attention in many directions. However, we cannot ignore the pressing need to address the AMR crisis. Let us use this World Antimicrobial Awareness Week to gain momentum and discuss ways we can collectively tackle AMR.


Antimicrobial resistance (AMR) and the increase in multi-resistant bacteria are among the most important threats to public health worldwide, according to the World Health Organisation (WHO). Moreover, this issue is underpinned by the One Health perspective, due to the ability of AMR to be transmitted between animals and humans living in the same environment. Therefore, since 2014 different surveillance and control programmes have been established to control AMR in commensal and zoonotic bacteria in production animals. However, public health authorities’ reports on AMR leave out companion animals, due to the lack of national programmes and data collection by countries. This missing information constitutes a serious public health concern due to the close contact between companion animals, humans and their surrounding environment. This absence of control and harmonisation between programmes in European countries leads to the ineffectiveness of antibiotics against common diseases. Thus, there is a pressing need to establish adequate surveillance and monitoring programmes for AMR in companion animals and further develop alternatives to antibiotic use in this sector, considering the impact this could have on the gut microbiota. In this context, the aim of this review is to evaluate the current control and epidemiological situations of AMR in companion animals in the European Union (EU), as well as the proposed alternatives to antibiotics. Antimicrobial resistance (AMR) is currently one of the main concerns worldwide, signalled by the World Health Organisation (WHO) as one of the top 10 global public health threats in 2019 . In thisDue to the importance of AMR, governments worldwide have been involved in the search for solutions and have established surveillance programmes to monitor AMR prevalence and evolution in zoonotic and commensal bacteria, considered potential reservoirs of resistant genes . FinallyIn the EU, the European Food Safety Authority (EFSA) coordinates surveillance programmes for commensal bacteria in food-producing animals, along with the monitoring of zoonotic agents and their AMR . In 2009However, the focus has always been on food-producing animals because of their importance as transmitters of AMR through the food chain. These programmes have achieved a significant reduction in antibiotic consumption, so the next step is to focus all efforts on companion animals . CurrentIn this context, the EU plans to launch the European Antimicrobial Resistance Surveillance Network in Veterinary Medicine (EARS-Vet) to get an overview of the current situation and establish European AMR monitoring systems . TherefoAn exhaustive literature search was conducted in which different sources of information were reviewed and evaluated to address the current status of AMR surveillance programmes in companion animals. The search was performed through search engines such as PubMed, Google Scholar and Scopus. In addition, reports and publications of health organisations such as the WHO, OIE , EFSA, ECDC or EMA were reviewed. This review included articles written and published in English and German, as the annual AMRs report submitted by Germany was written in their native language. The search terms that provided most of the information were: “Antimicrobial resistant”, “Companion animals”, “AMR surveillance programmes in Europe”, “AMR monitoring programmes in Europe”, “EARS-Vet”, “One Health”. Moreover, all articles were filtered to be no older than 2018, except for one article about a specific clinical case from 2009. The current imbalance between AMR programmes and reported data limit the full understanding of an integrated One Health global AMR surveillance system . In 2015In the global scope, there are different programmes to control AMR in both production and companion animals. On the one hand, there are AMR Surveillance Programmes that study bacterial species present in animal infections and their AMR ,19. On tIn the EU, the EFSA coordinates the AMR Surveillance Programmes in food-producing animals, in accordance with Directive 2003/99/EC and Commission Implementing Decision (EU) 2020/1729 . MoreoveFinally, the JIACRA brings together all available data on AMR and AMC, comparing animal and human results. In the last report, AMC was lower in food-producing animals than in humans, with 108 mg/Kg and 130 mg/Kg of estimated biomass, respectively. These results show the effectiveness of the current surveillance and monitoring programmes in place in the animal production sector . Moreover, the EMA categorised antimicrobials according to the risk they pose to public health and the need for their use in veterinary medicine. These are divided into four categories: Category D, which should be used with caution and as a first line of treatment whenever possible; Category C, with antimicrobials that should be used with caution and only when Category D antibiotics fail clinically; Category B, including antimicrobials that are critically important in human medicine and whose use should be restricted in veterinary medicine and adopted only when all therapeutic alternatives (D and C) have been exhausted; and finally, Category A, whose use is limited in human medicine and is not authorised in the European Union (EU) in veterinary medicine and therefore for the treatment of production animals . HoweverAntimicrobial use (AMU) in companion animals is also not included in the annual reports conducted by the EMA (at a European level)  and by tStaphylococcus pseudintermedius as well as Staphylococcus aureus are coagulase-positive species of Staphylococcus. These bacterial species are opportunistic pathogens found in the mucous membranes of animals as part of the commensal bacteria and have been reported to infect humans ..10].FINRES stands for Finnish Veterinary Antimicrobial Resistance Monitoring and Consumption of Antimicrobial Agents . The samAMU data for companion animals in Finland are not available. The report only includes information on food-producing animals .S. aureus and S. pseudintermedius isolates, which indicated the presence of methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant Staphylococcus pseudintermedius (MRSP), respectively [Staphylococcus canis, all the isolates analysed were sensitive to penicillin [Regarding the main results obtained in 2020, it is important to highlight the presence of oxacillin, resistance to which was found in ectively ,28, as onicillin  , which includes public, private and university laboratories ,31. In the latest report, AMU showed a continuous decrease to 670 tonnes of total antimicrobials sold. It is important to highlight that the sales of compounds of the fluoroquinolones group and of third- and fourth- generation cephalosporins reached the lowest value since 2011, but all these active substances cannot be linked to any specific species, as the use of some antimicrobials is approved for many animal species . E. coli strains isolated from urinary tract infections (UTI) in cats were resistant to ampicillin and amoxicillin–clavulanic acid, whereas for dogs, the figures were below 20% [The results are shown in elow 20% .NORM-VET reports on the use of antimicrobial agents and occurrence of AMR in Norway. This programme was launched in 2000, coordinated by the Norwegian Veterinary Institute commissioned by the Norwegian Food Safety Authority .In this report, AMU is expressed as kg of active substance sold for companion animals (cats and dogs) and corresponds to 360 kg. The most active sold substances were from the penicillin group, along with first-generation cephalosporins and trimethoprim, of which the most sold antimicrobial was the combination of amoxicillin and clavulanic acid .E. coli isolates was to aminopenicillins. However, for S. pseudintermedius, the highest AMR percentage was observed to fusidic acid [As seen in dic acid ,33.SWEDRES-SVARM reports on Swedish Antibiotic Sales and Resistance in Human Medicine (SWEDRES) and Swedish Veterinary Antibiotic Resistance Monitoring (SVARM). Data for this programme are provided by the Public Health Agency of Sweden and the National Veterinary Institute . In Sweden, referring to companion animals, AMU data are only related to dogs . In addiANRESIS/ARCH-Vet is the Swiss Centre for Antibiotic Resistance, which has drafted the Antibiotic Resistance Report in this country since 2019 . Sales of antimicrobials in the last report were increased from the previous year to 775 kg of substances. Switzerland follows the trends in AMU of the rest of the European countries, with the penicillin group being the most consumed .S. pseudintermedius showed high AMR level to some antibiotics, such as gentamicin (~38%) and erythromycin (nearly 30%). However, in E. coli, higher AMR to cephalothin was observed in dogs and cats [and cats ,35 , and the other 3 were in the process of setting up these programmes . The shared information about the future scope of Belgium, Greece and Spain was defined through relevant national experts . In thisE. coli, Actinobacillus pleuropneumoniae, Klebsiella pneumoniae, Pasteurella multocida, Mannheimia haemolytica, S. aureus, S. pseudintermedius, Staphylococcus hyicus, Streptococcus suis, Streptococcus uberis and Streptococcus dysgalactiae to be monitored [Finally, EARS-Vet chose 6 animal species, i.e., cat, cattle, chicken (broiler and laying hen), dog, swine and turkey 11 bacterial species, i.e., onitored . Moreoveonitored .As reported above, AMR represents one of the greatest threats to animal, human and environment health. In the past, antibiotics have been used not only to control infections, but also to prevent pathologies and improve the health status of the animal, without limits on their use or AMR prevalence control . TherefoLactobacillus spp., Streptococcus spp., Lactococcus spp. and Bifidobacterium spp. [Probiotics are live microorganisms that confer benefits to host health when administered in adequate doses . There aium spp. ,38. Probium spp. ,39.Bifidobacterium animalis, in dogs with acute idiopathic diarrhoea showed that the use of the probiotic combined with the improvement of nutritional management reduced the need to administer metronidazole to the diseased dogs [One study evaluating the benefits of a probiotic, based on canine-derived sed dogs .Prebiotics are non-digestible compounds that are metabolised by gut microorganisms and modulate the composition and activity of the gut microbiota, providing benefits to the host’s physiological bacteria .According to the results reported by De Souza et al. (2019), in dogs fed a mixture of fibre and prebiotics, no negative effect on nutrient digestibility and faecal quality was observed, and increased digestibility of food was reported. In addition, beneficial changes were observed in the faeces, which may indicate support of gut health. While the test substances caused slight changes in faecal microbial populations in adult healthy dogs, they had a significant effect on faecal metabolite physiology, demonstrating a possible microbial improvement in dogs fed diets supplemented with prebiotics. However, further research is needed to establish the optimal doses according to the age of the animals and the disease stages and to understand what conditions can be prevented or treated with prebiotic supplements .Symbiotics can strengthen the beneficial effects that probiotics and prebiotics have on their own, as probiotics use prebiotics as food sources to extend their survival in the digestive tract, increasing the digestibility and availability of certain nutrients such as vitamins, minerals, and proteins . A study performed in dogs with acute diarrhoea, comparing the therapeutic effect of nutraceuticals and antibiotics on clinical activity, showed that the role of symbiotics in the positive effect seen in patients is unclear. Thus, more studies are needed both in vitro and in vivo, in companion animals, to further investigate the real effect of these supplements .Bifidobacterium , Bacteroides fragilis, E. coli Nissle 1917 and Faecalibacterium prausnitzii, among others [Postbiotics are metabolites generated by the fermentation of probiotic bacteria in the gut . In the g others , and it g others ,45.Lactobacilli-derived factors in vitro, postbiotics were shown to have beneficial properties in relation to pathogen-induced inflammation and altered cytokine release [In a study evaluating the postbiotic activities of  release .However, as AMR is present in animal diseases and antibiotic treatment options sometimes do not work properly, efforts are being made to develop alternative treatments for ongoing infections . Faecal microbiota transplantation (FMT) involves the transfer of faeces from a healthy donor to the intestine of a diseased recipient with the aim of adjusting the gut microbiome of the diseased subject. The gut microbiota is seriously affected by the use of antibiotics or by inflammatory gastrointestinal diseases treated with antibiotics, which further aggravates a pathology . Therefore, FMT is sometimes the only and last viable option ,47. AlthBacteriophages (or “phages”) are viruses that possess the natural characteristic of specifically targeting and killing bacteria . One of Pseudomonas aeruginosa, with a mixture of bacteriophages [Although most studies have been conducted in vitro, a clinical trial was carried out to study the treatment of otitis in dogs, caused by iophages . The outiophages .All the information collected in this section is summarised in colonisation resistance [The biotic family explained above has played an essential role in complementing the prophylaxis and treatment of different diseases ,52. Howesistance . This tesistance .The gut microbiota lives in symbiosis with the host and is involved in its health status. The bacteria that make up the microbiota protect themselves from pathogens by competing for essential metabolites and nutrients and inducing intestinal immune responses, as studies show that there is increased immune system activity in the gut than in all other lymphoid tissues combined . The gutFusobacteria, Bacteroidetes, Actinobacteria, Firmicutes and Proteobacteria, which represent more than 99% of the gut microbiota of dogs and cats [Chloroflexi, Tenericutes, Spirochaetes, Cyanobacteria, Verrucomicrobia and a few unclassified bacterial groups [Helicobacter spp. [The microbiota of companion animals is dominated by the phyla and cats ,40,54. Tter spp. ,54. In conclusion, dogs and cats are considered an important potential source of AMR, posing a risk to public health. However, there are no harmonised control and monitoring programmes established in the EU, highlighting the need to create a joint plan to combat the evolution of AMR in companion animals, encompassing this concern under the One Health concept. For this reason, the EARS-Vet network is currently being developed, with the aim of establishing the bacterial species to be monitored, the complete antibiotic panels for these bacteria and the type of procedure to be followed to analyse collected samples. Moreover, alternatives to the use of antibiotics in veterinary clinic practice are being developed, with the aim of modulating the intestinal microbiota and prevent the onset of diseases , also considering treatments such as faecal transplantation and bacteriophages.


Given the stigma of eating disorders (EDs), anonymous online communities of individuals with EDs can play a critical role in their treatment. In our study, we aimed to identify prevalent topics related to EDs in one community. To discover latent topics in an online community dedicated to EDs, we applied an automated text-mining approach to topic modeling after collecting data from Reddit created between January 2011 and September 2020. As a result of topic modeling, topics regarding emotional support, informational support, and inquiries about EDs were discovered. In frequently asked questions and advice about EDs in the online community, community members empathized more with topics soliciting emotional support rather than informational support. Despite the importance of online communities, studies on this topic, especially those analyzing online conversations, have remained limited. By demonstrating the role of online communities in supplementary interventions, our findings can support clinicians in caring for patients with EDs. Eating disorders (EDs) involve bulimia and anorexia nervosa, binge-eating disorder, and specified feeding . Eating Social media platforms have increasingly become a popular space for individuals to share similar ideas, provide advice, seek information, discuss content, and assess themselves regarding their body and EDs through photos and videos . VariousMany online pro-ED communities exist on social media platforms  , increasInterest in trends within social networks and the forms of social support that they can offer, especially regarding the impact of health conditions, has increased considerably. Social support is a meaningful element in the maintenance and development of EDs, and members of ED online communities tend to participate in community activities because they offer direct and indirect social support ,12. In pAgainst this background, we investigated the U.S.-based online platform, Reddit, to identify and analyze salient topics actively discussed in a representative ED online community, which influences the perceptions of its members. Reddit is a widely used social media platform for stigmatized illnesses . Reddit,Some studies related to Reddit have been focused on and conducted recently . PreviouReddit was selected for data collection. In our study, we were particularly interested in the subreddit concerning EDs (“r/EatingDisorders”), which, as of July 2021, had more than 55,000 members. In April 2021, we used Reddit’s Python-based application programming interface to collect all texts associated with threads  and comments in this subreddit, along with the creation dates and scores of each thread. As a result, we collected 11,484 documents generated between January 2011 and September 2020 .Before conducting text mining, we performed text-cleaning procedures, including tokenization, lowercase conversion of all characters, stop word removal, lemmatization, and bigram creation. Once the documents were cleaned, we retained those with more than three words for topic modeling analysis, which amounted to a final dataset of 10,050 documents . The number of original threads and comments for each time period is displayed in To discover salient topics in Reddit’s ED online community, we followed an R-based probabilistic topic modeling approach called structural topic modeling (STM) . Unlike Topic modeling algorithms require investigators to input the optimal topic number (K) for the dataset. Thus, the quantitative index  was compared to evaluate the topic modeling performance for the models with consecutive topic numbers, from 3 to 50. Based on the quantitative index results and topic modeling results, researchers manually reviewed the topic modeling results between 10 and 15 topics, and agreed that 14 topics were appropriate for our dataset. Over the course of multiple iterations, STM automatically built the model and generated a list of words that constitute each topic (β) and the probability of each topic being associated with each document (θ), as shown in Of all topic modeling algorithms, STM has been widely applied because of its strength in covariate estimation ,25,26. AUsing the results of topic modeling, we performed a topic network analysis in order to determine the hierarchical structure of the 14 extracted topics. STM offers a function of “topicCorr,” which enables the estimation of correlations with the document-topic proportion distributions. The topic network was generated through topic correlation estimations, and community detection was applied to the topic network. In topic network analysis, topics that share characteristics can be grouped to create communities, which we accomplished with igraph package from R software. Following Girvan and Newman , who proMembers of a subreddit can “upvote” or “downvote” a thread to express their approval or disapproval of the thread, and the score of each thread represents the net score of votes . Thus, the threads that resonate with more users are likely to score higher, whereas threads that address controversial issues are likely to score lower. It is assumed that the net score of votes for each thread represents how much other users agreed with not only the original thread, but also the discussions in the comments. Hence, all documents belonging to each thread were assigned the same net score as the original thread. To identify which threads tended to have higher scores, we performed multiple regression analysis between topic proportions (θ) and scores in SPSS 21.0. In that analysis, we used document-level topic proportion as an independent variable and the net score of votes as a dependent variable. Because the sum of topic proportions per document is always 1, using raw topic proportion scores can cause serious multicollinearity in the regression model. To counter this risk, we recorded the topic proportion scores. Antons et al.  proposedSTM estimates the associations  between 14 topics and vocabulary in the corpus. The top five words that are most closely associated with each topic with the highest β are listed in The most popular topic, communication (Topic 8), accounted for 13.1% of the total topic proportion; the top words associated with the topic were “talk,” “tell,” “want,” “friend,” and “support,” thereby indicating that the importance of communication as social support in the ED online community. The topic consisted of discussions about issues related to communicating about EDs. More specifically, members actively shared their opinions about whether seeking help by communicating their own or close ones’ ED symptoms with others was necessary and about the issues that they need to consider when communicating those symptoms.Another popular topic in the ED online community was binge eating (Topic 13), which accounted for 10.5% of the total topic proportion. The top words associated with Topic 13 were “eat,” “food,” “binge,” “body,” and “feel,’ which indicates members’ concerns about binge eating and its impact on their body’s conditions and emotions. The documents most closely associated with binge eating offered practical tips and advice to overcome the ED and to establish healthy eating routines. The topic of professional help (Topic 10), with the top words “help,” “therapist,” “need,” “doctor,” and “therapy,” accounted for 9.5% of the total topic proportion. The documents associated with the topic mostly concerned the importance of finding professional help to overcome ED. In particular, many community members advised others to access specialists in EDs.Using the results of topic modeling, we implemented a fast greedy algorithm that automatically detected four communities that maximized the modularity scores. In Community 1, we grouped two topics related to emotional support in order to boost self-esteem, namely self-esteem for existence (Topic 1) and self-esteem for health (Topic 5). In Community 2, we grouped five topics related to informational support, meaning a question about or recommendations for managing ED-related issues, including ED-related health issues (Topic 7), communication about EDs with others (Topic 8), and professional help (Topic 10). Another topic in Community 2 represented the fact that individuals sometimes seek cooperation from the ED online community for their research (Topic 2). Next, the topics belonging to Community 3 were feeling in control (Topic 3), recovering from EDs (Topic 4), snacking behavior (Topic 6), binge eating (Topic 13), and exercise routine (Topic 14). Community 2 and Community 3 are both related to social support for ED community members. However, whereas the topics in Community 2 were more applicable to individuals at the initial stage of admitting ED symptoms, those in Community 3 were more applicable to individuals already in the process of overcoming EDs. Last, Community 4 concerned inquiries about ED in topics related to EDs (Topic 9) and ED diagnosis (Topic 12). Topic 9 also included factors or aspects of personal background that may be associated with the occurrence of EDs or challenges in overcoming EDs. On that topic, many community members shared research findings or their own experiences, as illustrated in the following three documents:“(…) My ED tendencies started immediately following having jaw surgery, && having my jaw wired shut for a few weeks. (…)”[Comment on a post written in March 2019]“(…) I studied the link between eating disorders ED and autoimmune diseases AD, which is growing more and more evident to the point that many doctors will screen for an ED after a AD diagnosis. (…)”[Comment on a post written in March 2019]“(…) Another man with an eating disorder here. For your reference, I am in a support group for men with EDs, and there are 50/50 straight/gay men in it. (…)”[Comment to a post written in April 2014]Two topics were more prevalent among the original threads than comments: research requests and resource seeking (Topic 2) and recovering from EDs (Topic 4). Because researchers have used the online community to request members’ participation in their research, the topic proportion of Topic 2 was relatively high among the original threads. The same dynamic occurred in Topic 4 because many people requested other members’ advice on recovering from EDs.Three topics were mentioned more frequently in comments than in original threads: self-esteem for existence (Topic 1), various issues related to ED (Topic 9), and feeling in control (Topic 3), thereby demonstrating the most popular comments or recommendations in response to those ED-related questions. As shown in various topics offering practical information to overcome EDs, providing useful information plays a critical role in the online community. Additionally, comments about having control over self-esteem and feelings were also prevalent. The popularity of topics regarding various issues related to ED indicates that many members of the ED community have acknowledged that EDs have complex symptomology involving various situations or factors that trigger ED.Topic content was compared to identify potential gaps between the original threads and comments. Words skewed to the left side  were more frequently used in original posts, whereas words skewed to the right side  were more frequently used in comments. The words placed in the middle were commonly used in both original posts and comments, and the word size represents the frequency of the terms in the dataset. Regarding ED-related health issues (Topic 7), original threads offer brief explanations about health conditions  that trigger EDs or physiological complications due to the disorders . Some coFor the topic of communication (Topic 8), the term “boyfriend” was frequently used in original threads, whereas “family” and “friends” were frequently used in comments . The oriAmong original threads related to the topic of exercise routine (Topic 14), the terms “want,” “need,” and “help” appeared frequently, thereby indicating that community members have acknowledged the importance of exercise in overcoming EDs . Even soTo identify which topics were largely supported by members of the online ED community, we performed regression analysis to gauge the effects of topical structures on net scores . That isp < 0.001), feeling in control , and self-esteem for health , suggesting that many members of the ED community supported the underlying ideas of those topics. In general, all these topics are related to emotional support. In contrast, five topics had a negative significant regression: research requests and resource seeking , snacking behavior , various issues related to EDs , professional help , and ED diagnosis . These topics, all related to informational support, tended to have negative coefficients. Finally, four topics had insignificant coefficients: ED recovery , communication , binge eating , and exercise routine . Except for exercise routine (Topic 4), all three topics had topic weights greater than the median topic weights, indicating that many ED members were interested in these topics. Hence, for these three topics, it is plausible that those topics had insignificant coefficients because they are controversial. That is, they have high upvotes and downvotes, which reduced their net scores. However, the exercise routine topic (Topic 4) had a low topic weight, indicating that the discussion about this topic was not active among the community members. Therefore, it is possible that members of the ED online community are not interested enough in Topic 4 to vote on them.Three topics had significantly positive regression coefficients: self-esteem for existence , they sPrevious studies have focused on the nature and content of pro-ED communities that promote EDs as a lifestyle on different social media using content or network analyses ,32. ReleThis study is grounded on “social support and computer-mediated communication theory”, which indicates online media can serve as a tool to help individuals with physical and mental health . Based oWe also analyzed the topical contents of both original threads and comments. Original threads are mostly questions regarding EDs written by those who are suffering from the symptoms while comments tend to be advice for the corresponding questions written by those who are going through or overcame EDs. By comparing original threads and comments, we discovered the most frequently questioned concerns and types of advice. EDs are mental illnesses at the base, advice related to self-esteem, feelings, and subjective self-evaluations had great weight and value in the ED online community, and the relevant topics were popularly mentioned and advised.Finally, we were able to identify which topics were approved or disapproved by other members by using the vote score on each post. Voting behaviors can be considered as an intervention or an indirect participation in the communication to solve the issues and support the members with the problems. Such an approach is useful to capture the perceptions of members who actively participate in the online community by creating posts and leaving comments, as well as inactive members who browse posts, comments, and votes.Popularity and voting patterns were used as criteria to classify the topics into the following categories: popular and upvoted, popular and downvoted, unpopular and upvoted, unpopular and downvoted, popular and controversial, and unpopular and uninterested. These findings demonstrate ED-related topics that many people are interested in and group-based opinions toward topics shaped by online community members. For instance, the topic related to self-esteem in relation to existence was shared in comments and received many upvotes, indicating that many ED members agreed on this idea and were willing to share their opinions through active participation in commenting to original threads. On the other hand, the topic regarding professional help was frequently mentioned in the ED online community, but this topic tended to be downvoted. Despite the popularity of this topic, ED members disapproved posts or comments about the topic regarding professional help because they believe it is not appropriate to seek professional help in the ED community, which mostly consists of non-experts. In other words, ED members agreed that questions about professional help should be discussed with health care professionals or experts rather than the ED community.The current study revealed the role of ED online communities by examining how ED online community members communicate with each other to deal with EDs. Many community members sympathized more with topics soliciting emotional support rather than those soliciting informational support. Therefore, the role of ED online communities should be emphasized as a means of gaining emotional support instead of searching for factual information. Moreover, people who use ED online communities should carefully consider that most community members are not professionals but merely people who are interested in EDs or who have or have had EDs. It is plausible, therefore, that some posts in the community may contain incorrect information, which demands caution from members when reviewing the information.Our findings can help individuals with symptoms of ED and clinicians who treat them. For one, we found that many people freely share their symptoms and situations related to EDs in online spaces, where anonymity is guaranteed. Because online communities are regarded as protected from the judgment of others, they are open to anyone with concerns about EDs, including pre-eating disorders. Thus, online communities can be a medium to inform patients and facilitate changes in their thoughts and behaviors before their symptoms progress to a serious degree.Another benefit of the online communities is that they are easily available to those suffering from EDs as well as those who are close to the patients. We discovered that friends and family members of people with ED symptoms commonly used online communities to seek for advice. As a result, clinicians can utilize online platforms to understand difficulties affecting ED patients and those close to them. Despite the benefits of the online communities outlined, most online community contents are emotional support from non-experts rather than informational support from professionals. Hence, in the future, online service through readily accessible media will be required so that people suffering from EDs can receive credible information and guidance from experts without too much difficulties.Based on the limitations of the current study, we propose the following tasks for future research. The current study is limited to ED communities on Reddit with a limited number of original threads, so the results cannot be generalized to other communities. Therefore, future study may target different types of online discussion at different social media platforms, such as Twitter and Facebook. Since this study analyzed Reddit posts generated over several years, people’s perceptions may have changed, which requires future studies to compare the differences in people’s perceptions and opinions. For the regression analysis, this study assigned the same net scores of the original threads to the comments based on the assumption that other users vote the thread after reading both the original threads and comments. However, some people may disregard the comments and vote on the threads solely based on the original thread content. Future studies are necessary to overcome such limitations.


E. coli phage ID52 lysis protein E was introduced to generating BGs for the first time. Above all, we compared the lysis activity of lysis protein of E. coli phage φX174 and E. coli phage ID52 as well as the effects of promoters on the lysis activity of ID52-E, which shown that the lysis activity and BGs formation rate of protein ID52-E was significantly higher than protein φX174-E. Further, the lysis activity of ID52-E was significantly improved under the control of L-arabinose inducible promoter which initial induction OD600 reached as high as 2.0. The applicability of lysis protein ID52-E induced by L-arabinose was proved by preparing probiotic E. coli Nissle 1917 BGs and pathogenic Salmonella typhimurium BGs in mass production. This paper introduced a novel and highly efficient method for BGs preparation depending on recombinant expression of E. coli phage ID52-E under eco-friendly and reasonable price inducer L-arabinose.Bacterial ghosts (BGs) are nonliving empty bacterial shells without cytoplasm retaining original morphology and identical antigenicity of natural bacteria, making them high potential and promising vaccine candidates and delivery vehicles. However, the low yield of commonly used BGs preparation methods limits its mass production and widely application. In order to improve BGs production,  Bacterial ghosts (BGs) mainly refer to the inactive hollow cell envelopes derived from Gram-negative bacteria without cytoplasmic contents, which retain the intact cellular morphology and natural surface antigenic structures such as pathogen-associated molecular patterns (PAMPs), which include lipopolysaccharides (LPS), peptidoglycan, monophosphoryl lipid A (MPL), flagella, adhesins and other immune-related stimulating elements of natural bacteria . TherefoE. coli phage phiX174 lysis protein gene E (φX174-E), which under the control of the temperature-inducible expression plasmid pBV220 without adding other chemicals. Contrary to lysis proteins derived from other phages, gene φX174-E has no inherent enzymatic function, which encodes a 91-amino acid membrane protein with the competence to fuse the inner and outer membranes of Gram-negative bacteria, giving rise to forming an E-specific transmembrane tunnel structure with a size in diameter between 40~400 nm [BGs have so many advantages that they have broad application prospect and great research value, while the low yield of BGs limits the large-scale production and application of BGs. In the short term, BGs were mainly prepared by inducing the expression of 0~400 nm  by oligo0~400 nm . Scannin0~400 nm ,20.600) approximated 0.2~0.6, host bacteria can be lysed effectively by a temperature elevation to 42 °C [600 can be improved during the production of BGs, not only can medium components be fully utilized, but also can the yield of BGs be significantly increased. In order to improve the yield of BGs, Yu et al. [600 was up to 1.1. Further, Zhu et al. [Staphylococcus aureus nuclease A gene, which could effectively induce host bacteria to lysis with the intracellular degradation of genetic material of host cells, and initial induced lysis OD600 could reach as high as 1.2.However, lysis activity of protein φX174-E depended on the growth phase of bacteria . In a geto 42 °C , resultiu et al.  obtainedu et al.  construcM by site-directed mutation of φX174-E, and the initial induced OD600 could reach 2.0 with the joint control of T7 promoter and pLysS plasmid, achieving efficient preparation of E. coli and Salmonella enteritidis. However, the inducer IPTG had a certain toxic effect, and the high price and cost of IPTG were not conducive to the large-scale production and application of BGs. Furthermore, Ma et al. [M without screening the mutant protein with high lysis activity. Therefore, it can be seen that the possibility of improving the yield of BGs by mutating lysis gene φX174-E was low, and this method has been in a bottleneck period. Some chemical agents such as NaOH, H2O2 and SDS have been proved to be an alternative method to commonly method involving protein E for producing BGs [However, the BGs yield of above preparation methods was still far from large scale production. Based on this, Ma et al.  screeneda et al.  continuecing BGs . But thecing BGs . TherefoE. coli Nissle 1917 (EcN), a widely used probiotic, not only possesses the special lipopolysaccharide (LPS) in outer membrane, which exhibits a good immunomodulating property, but also can colonize and replicate in tumors efficiently [Salmonella typhimurium (ST), an important food-borne non-host-adapted Gram-negative intracellular pathogen, has competence to infect a variety of animal hosts and humans around the world. Vaccination is one of the effective methods to reduce the infection of ST in animals [The iciently . Therefo animals , and it  animals . HoweverE. coli phage phiX174, E. coli phage ID52 also belongs to the Microvirus family, and it also contains lysis protein gene gpE (ID52-E), which encodes a 103-amino acid [E. coli phage ID52 lysis protein ID52-E to detect whether it can be applied to produce BGs and improve the yield of BGs. First, the lysis gene φX174-E and lysis gene ID52-E were cloned into pBV220-Chl plasmid to compare the lysis effects of two lysis proteins, whose results showed that the lysis effect of lysis protein ID52-E was significantly higher than that of φX174-E. In order to test whether the yield of BGs could be further improved, the temperature inducible promoter of pBV220 was replaced with L-arabinose inducible promoter and the results manifested that the OD600 initiation can reach as high as 2.0, while the minimum OD600 can be reduced to 0.3~0.4 after lysis. Finally, in order to test the feasibility of the efficient BGs preparation system by protein ID52-E under control of L-arabinose inducible promoter, the system was applied to prepare EcN ghosts and ST ghosts, respectively. And the results manifested that protein ID52-E could be applied to the high-efficiency and high-yield production of EcN ghosts And ST ghosts, whose initial induced OD600 could be as high as 2.5.Like ino acid . Accordi+), and other reagents used in this study were purchased from Sangon Biotech  unless otherwise specified. The final concentrations of chloramphenicol, kanamycin and ampicillin in LB were 25 μg/mL (Chl+), 50 μg/mL (Kan+) and 100 μg/mL (Amp+), respectively.The Bacterial strains, plasmids, and primers used in this research were listed in φX174-E and lysis gene ID52-E were amplified by PCR with primer pairs of φX174-E-F/φ174-E-R and ID52-E-F/ID52-E-R, respectively. Then PCR products were detected by 1% agarose gel and purified and recovered by DNA Gel Extraction Kit . Next, using the purified lysis gene φX174-E and the purified lysis gene ID52-E fragment as a pair of primers in a linear amplification reaction around a circular plasmid pBV220-sGFP-Chl, respectively. Later, the methylated parental plasmid was digested by treating the linear amplification reaction products with Dpn I enzyme. Then the reaction mixture was transformed into E. coli DH5α competent cells. Positive clones were confirmed by DNA sequencing . The resulting plasmids were designated as pBV220-φX174-E and pBV220-ID52-E.Plasmid pBV220-φX174-E and pBV220-ID52-E were constructed by restriction-free cloning technology (RF cloning) ,32 Figu. First, araC-ParaBAD was amplified from the plasmid pKD46 using primers “araC-ParaBAD-F/araC-ParaBAD-R”. Finally, seamless Assembly Cloning kit was used for seamless assembly cloning of linearized fragment pBV220-ID52-E and L-arabinose inducible promoter araC-ParaBAD. Then the reaction mixture was transformed into E. coli DH5α competent cells. Positive clones were confirmed by DNA sequencing . The resulting plasmids were designated as araC-ParaBAD-ID52-E. Primers required to the above vectors were listed in Plasmid araC-ParaBAD-ID52-E was constructed by seamless assembly cloning with Seamless Assembly Cloning kit  . FirstlyE. coli BL21(DE3) competent cells, respectively. Next, the recombinant strains containing different plasmid were cultured to different OD600 values, and the expression of lysis proteins in bacteria containing plasmid pBV220-φX174-E or pBV220-ID52-E was activated by shift the temperature to 42 °C, while L-arabinose with final concentration of 0.5 mg/mL was added to activate the expression of lysis protein ID52-E in bacteria containing plasmid araC-ParaBAD-ID52-E.The three aforementioned plasmids were transformed into 600 value is one of the indicators to evaluate the lysis activity of different lysis proteins E in E. coli BL21(DE3). The values of OD600 were detected at initial induction OD600 approximately 0.8, 1.2, 1.6, 2.0 and detected every 30 min after inducing the expression of lysis protein E. And the OD600 decrease degree was calculated by the following formula: OD600 decrease degree % = (1 − post-lysis OD600/pre-lysis OD600) × 100%. On the other hand, the lysis efficiency of different lysis proteins E was evaluated by detecting viable cell counts before and after lysis, which was conducted by serial dilution and plate count methods. And the lysis efficiency was calculated by the following formula: lysis efficiency % = (1 − post-lysis CFU/pre-lysis CFU) × 100%.On the one hand, the decrease degree of ODE. coli BL21(DE3) harboring pBV220-φX174-E, pBV220-ID52-E and araC-ParaBAD-ID52-E plasmid respectively was cultured to OD600 approximately 1.6 to induce the expression of lysis protein. Then 2 mL bacterial liquid before and after lysis were collected respectively and centrifuged at 5000 rpm for 20 min to collect the bacterial deposits whose genome were extracted by Rapid Bacterial Genomic DNA Isolation Kit  and the genome concentration was measured. The degree of genome leakage was calculated by the following formula: genome leakage degree % = (1 − post-lysis genome concentration/pre-lysis genome concentration) × 100%. All experiments were conducted with three biological replicates.Furthermore, the outflow of genome and intracellular protein after lysis were another indicator to evaluate the lysis activity of lysis protein. In order to detect the outflow of the genome after lysis, E. coli BL21(DE3) harboring pBV220-φX174-E, pBV220-ID52-E and araC-ParaBAD-ID52-E plasmid respectively was cultured to OD600 approximately 1.6 to induce the expression of lysis protein. Then 5 mL bacterial liquid before and after lysis were collected respectively and centrifuged at 5000 rpm for 20 min to collect the bacterial deposits and culture supernatant. The bacterial deposits were re-suspended with 500 μL PBS and ultrasonic crushing was performed. Then the bacterial deposits sample after crushing and the culture supernatant sample were detected by SDS-PAGE.At the same time, the protein leakage after lysis was detected by SDS-PAGE. E. coli BL21(DE3) harboring different plasmids respectively was cultured to OD600 approximately 1.6 to induce the expression of lysis protein Then 10 mL bacterial liquid was collected before lysis and after lysis and the bacteria cells were pelleted by centrifuging at 5000 rpm for 20 min at 4 °C, respectively. Then the precipitate was washed three times with sterile phosphate-buffered saline (PBS) to fully remove the medium. Next, the pellets were resuspended in 5 mL 2.5% glutaraldehyde special electron microscopic fixative and fixed at 4 °C for 8 h. After fixation, cells were washed 3 times with deionized water and eventually resuspended in 1 mL deionized water. Drop samples to the dried cell strip and embed them in filter paper. Subsequently, samples were dehydrated with 70%, 85% and 95% ethanol step by step for 15 min each time, and soaked with 100% ethanol for 15 min (repeat this step 3 times). Next, samples were dried with critical point dryer named Autosamdri-815 and following were coated by a gold sputter coater. Finally, the SEM images were acquired using Merlin SEM .E. coli BL21(DE3) harboring different plasmids respectively was cultured to OD600 approximately 1.6 to induce the expression of lysis protein Then 10 mL bacterial liquid was collected before lysis and after lysis and the bacteria cells were pelleted by centrifuging at 5000 rpm for 20 min at 4 °C, respectively. Same as SEM sample pretreatment, the pellets were washed three times with sterile PBS and fixed with 5 mL 2.5% glutaraldehyde special electron microscopic fixative at 4 °C for 8 h. Then samples were washed three times with deionized water to get rid of fixative. Eventually, 1 mL sterile ddH2O was used to resuspend the sample. Placed a clean piece of filter paper in the culture dish and placed the carbon coated copper mesh (face up) on top of the filter paper. Added 20 µL bacterial droplets to the carbon film copper mesh for 5~10 min at room temperature. Gently moved the copper mesh with tweezers so that the excess bacteria droplets on the copper net were absorbed by the filter paper. Then dropped 20 µL of 3% phosphotungstic acid staining solution to the carbon coated copper mesh with bacteria. Finally, the samples were visualized under a transmission electron microscope .g for 90 min at 4 °C to obtain supernatant and precipitation. Added 5× loading buffer to the whole liquid, supernatant and precipitation in proportion, and boiled for 10 min at 100 °C, respectively. 10 µL protein samples of E. coli BL21(DE3) was separated by 16% tricine-SDS-PAGE. After electrophoresis, proteins bands on the gel were blotted onto a nitrocellulose (NC) membrane  using the Bio-Rad Transblot Turbo system at 80 V for 1.5 h. Following washed with TBST buffer  for 10 min, the NC membrane was incubated with blocking buffer (5% (w/v) skimmed milk powder in TBST) for 2 h to block the reactive sites of the NC membrane. Then the NC membrane was washed with TBST for three times to get rid of superfluous milk, 10 min each time. Later, the NC membrane was incubated with Anti-Strep-Tag II Monoclonal Antibody 8C12 produced in mouse  at 1:5000 in TBST at 4 °C overnight. The following day, the NC membrane was washed with TBST for three times to remove surplus primary antibody, 10 min each time. The NC membrane was incubated subsequently with horseradish peroxidase (HRP)-conjugated Goat Anti-Mouse IgG H&L  at 1:3000 in TBST at room temperature for 2 h. Following repeated washing with TBST, the Super Signal West Pico Chemiluminescent Substrate  was used to visualize the labeled bands on the NC membrane.Western blot analysis in this research was performed according to an established procedure with slight modifications . In brieE. coli Nissle 1917 and Salmonella Typhimurium engineering strain with araBAD gene deletion were constructed by using λ Red homologous recombination system. Firstly, the pKD46 plasmid was transformed into EcN or ST competent cells by electroporation with 2.0 kV and the recombinant positive clones were screened by ampicillin and colony PCR by P1/P2. Then the FRT-kanamycin-FRT linear fragment was amplified by using P3/P4 (EcN) and P5/P6 (ST) containing 56-nt homology arms targeting the araBAD gene as primers and pKD4 plasmid as PCR template following purified by DNA Gel Extraction Kit . The EcN or ST recombination clones containing pKD46 plasmid was inoculated into LB liquid medium with 100 µg/mL Amp+ at 30 °C. The next day, the cells were transferred to fresh LB broth and cultured at 30 °C and 220 rpm until OD reached 0.3~0.4, then L-arabinose with final concentration of 0.5 mg/mL was added to continue culturing until OD reached 0.5~0.6. Subsequently, the cells were made into electrocompetent cells and were transformed with 500 ng purified FRT-kanamycin-FRT linear fragment. Colony PCR was performed to verify whether araBAD gene was successfully replaced by kanamycin fragment by primers P7/P8 (EcN) and P9/P10 (ST) on the colonies grown on ampicillin and kanamycin double resistant plates, respectively. Nextly, the strain EcNΔaraBAD::kanamycin and the strain STΔaraBAD::kanamycin were obtained by culturing at LB broth at 42 °C for 12 h to remove the pKD46 plasmid. Then the plasmid pCP20 was transformed into the strain EcNΔaraBAD::kanamycin and STΔaraBAD::kanamycin competent cells by electroporation with 2.0 kV, respectively, which could express FLP recombinase at 42 °C to remove kanamycin resistance genes inserted into the genome and disappeared with the cell division and proliferation. The successful transfer of pCP20 plasmid was achieved by colony PCR using P11/P12 as primers. Finally, the EcN engineering strain and ST engineering strain with araBAD gene deletion named EcNΔaraBAD::FRT and STΔaraBAD::FRT were identified by PCR using primers P7/P8 and P9/P10 according to the nucleotide sequence of EcN and ST available in NCBI . The lysis curves of EcNΔaraBAD:FRT engineering bacteria and STΔaraBAD::FRT engineering bacteria were plotted by measuring OD600. The morphological observation and identification of EcNΔaraBAD:FRT BGs and STΔaraBAD::FRT BGs were performed by SEM and TEM.All statistical analyses were performed with GraphPad Prism 8.0.2 software and OriginPro 2021. The Data were expressed in the format of mean ± S.D, and the significance level was analyzed by ordinary one-way ANOVA with Tukey multiple comparison.The specific primers were used to conduct colony PCR on the positive clones obtained from the transformation products to verify whether the construction of lysis protein E expressing plasmid was successful. Bands of approximately 343 bp and bands of approximately 376 bp were visualized in positive clones harboring pBV220-φX174-E and pBV220-ID52-E by PCR using φX174-E-F/φX174-E-F and ID52-E-F/ID52-E-R primers, respectively . Bands o600 value decrease degree. Therefore, different initial induction OD600 values (low OD and high OD) were controlled to be the same before inducing the expression of lysis proteins E. When the initial induction OD600 value was low, there was no significant difference between lysis protein φX174-E and lysis protein ID52-E under the control of thermo-inducible lambda pL/pR-cI857 promoter, while there were significant differences between lysis protein ID52-E under the control of L-arabinose inducible promoter and the other two  containing araC-ParaBAD-ID52-E plasmid had the least residual amount of genome and the highest degree of genome leakage, while E. coli containing pBV220-φX174-E plasmid had the most residual amount of genome and the lowest degree of genome leakage. These results indicated that lysis protein ID52-E under the control of L-arabinose inducible promoter had the best lysis effect, which resulted in more genome flow to external media (E. coli BL21(DE3) containing araC-ParaBAD-ID52-E plasmid had the most intracellular protein leakage  showed uniform dark color due to containing DNA, protein and other cell contents  containing different plasmid (E. coli BL21(DE3) containing pBV220-φX174-E was significantly higher than E. coli BL21(DE3) containing pBV220-ID52-E or araC-ParaBAD-ID52-E plasmid, while the expression levels of different plasmids containing lysis protein ID52-E were almost the same. It was speculated that the molecular weight of the lysis protein φX174-E (11.64 kDa) was lower than that of ID52-E (12.77 kDa), so the expression quantity of lysis protein φX174-E was higher in the same induction time. Further, there is possible that the amount of protein required by lysis protein ID52-E to lyse host bacteria was far less than that required by lysis protein φX174-E to lyse host bacteria. When the number of lysis protein ID52-E synthesis reached the threshold of lysis host bacteria, lysis pores formed on the cell membrane, and the host bacteria began to lyse and die, and the lysis protein ID52-E synthesis was terminated, resulting that the expression of ID52-E was lower than φX174-E. And contrary to the expression levels of the two lysis proteins E, the lysis activity of protein ID52-E with lower expression level was significantly higher than those of φX174-E with higher expression level, indicating that the lysis activity of protein ID52-E was so stronger that lysis bacteria required less protein ID52-E.Western blot of the samples manifested that the expression levels of different lysis protein E were significantly different in  plasmid A–C. The Microviridae , EcN and ST have a faster consumption rate of inducer L-arabinose, so it was easy to cause insufficient supply of inducer and thus reduced the lysis rate. At the same time, a certain amount of L-arabinose can promote the growth of EcN and ST, and finally leaded to the phenomenon that the growth rate was faster than the lysis rate and the lysis curve OD600 value rised, resulting in a diminished output of BGs. Therefore, in order to reduce the use of inducers and increase the yield of BG, it was necessary to knock out araBAD gene related to L-arabinose consumption. And the λ Red homologous recombination system was used for gene knockout.In order to test the application scope of the recombinant expression of protein ID52-E under the control of L-arabinose inducible promoter to prepare BGs, the system was applied to EcN and ST. However, different from araBAD gene was amplified and then was identified by 1% agar electrophoresis analysis  ghosts, EcN ghosts and ST ghosts with high efficiency and high yield, but the immune effect was not performed, which still needs to be further expanded. And the lysis efficiency of protein ID52-E under control of L-arabinose inducible promoter was lower than that of protein φX174-E under control of temperature-inducible promoter, but the death of host bacteria didn’t represent the formation of BGs. As observed by TEM (And it has been reported that protein φX174-E mediated cell lysis mechanism was related to cell division, and its kinetics was species-specific, which was speculated to be the same for protein ID52-E ,24. Therd by TEM , proteind by TEM , but mosd by TEM . TherefoFinally, inspired by other scholars’ research on targeted tumor therapy with EcN BGs , becauseE. coli phage ID52-E under the control of L-arabinose inducible promoter, which provided the possibility for the efficient preparation of Enterobacteriaceae BGs and significantly improved the yield of BGs, laying a technical foundation for the large-scale production and application of BGs.In summary, this study not only described a novel and excellent lysis protein ID52-E for BGs preparation for the first time, but established an efficient high-yield BGs preparation system with the recombinant expression of


Electrospun porous nanofibers have large specific surface areas and abundant active centers, which can effectively improve the properties of nanofibers. In the field of photocatalysis, electrospun porous nanofibers can increase the contact area of loaded photocatalytic particles with light, shorten the electron transfer path, and improve photocatalytic activity. In this paper, the main pore−forming mechanisms of electrospun porous nanofiber are summarized as breath figures, phase separation  and post−processing . Then, the application of electrospun porous nanofiber loading photocatalytic particles in the degradation of pollutants  in water is introduced, and its future development prospected. Although porous structures are beneficial in improving the photocatalytic performance of nanofibers, they reduce their mechanical properties. Therefore, strategies for improving the mechanical properties of electrospun porous nanofibers are also briefly discussed. As a “top−down” membrane preparation technology, electrospinning technology can produce continuous fibers with diameters ranging from microns to nanometers . It alsoThe process and technical principle of electrospinning is driven by the propelling pump while the spinning solution is extruded from the spinneret of the syringe. When no external electric field force is applied, a spherical droplet is formed under the action of surface tension and gravity. When an electric field force is applied, the droplet is connected to a high−voltage power source, and its surface is instantly covered by the same charge. The repulsive force and surface tension between these charges cancel each other out, and the droplet becomes unstable. If the repulsive force is strong enough to overcome the surface tension, the droplet will become conical (Taylor cone) and jet down from the spinneret. As the diameter of the jet continues to decrease until it bends, the solvent in the polymer jet evaporates rapidly in a short period of time, and finally solidifies into fibers with diameters ranging from nanometers to micrometers. The structure and morphology of the prepared fibers are affected by three sets of parameters: (1) system parameters ; (2) process parameters ; and (3) Environmental parameters (such as ambient temperature and humidity). These parameters will have a certain impact on the final morphology of the fiber. For example, increasing the voltage within a certain range will increase the charge density on the jet surface and increase the repulsive force, thus forming fibers with smaller diameters. However, if the voltage is too high, the formation of the Taylor cone will be hindered, resulting in the inability to spin normally. If the voltage and polymer solution concentration are too low, this may result in the production of some beads, which do not form continuous fibers of uniform diameter. If the voltage and polymer concentration are too high, the spinning process will be difficult, and the selection of appropriate experimental parameters can electrospin continuous, smooth, and uniform nanofibers.The porous structure is beneficial to the increase of the specific surface area of fibers, increase of ion transport channels, shortening ion transport paths, and enrichment of the active sites on the fiber surface and the internal space ,26,27. T2O and O2 to generate highly oxidizing hydroxyl radicals (OH) and superoxide anions (O2−), which can directly oxidize organic matter into inorganic small molecules such as CO2 and H2O, without producing intermediate products, which is relatively clean and fast. During photocatalysis, the photocatalyst itself does not participate in the reaction but can accelerate the chemical reaction rate by promoting the separation of electron–hole pairs under the excitation of photons, thus improving the photocatalytic efficiency [Photocatalysis was discovered in 1967 when Akira Fujishima decomposed water into oxygen and hydrogen through titanium dioxide single crystals under light . It is aficiency . Howeverficiency ,38. In tIn the electrospinning process, some solvents with high volatility  and high environmental humidity. As shown in The circular holes are mainly formed in the later stage of the electrospinning process. Otherwise, these holes will be pulled into elliptical holes due to the electrostatic force. This phenomenon usually occurs when highly volatile solvents and hydrophobic polymers , dimethylacetamide (DMAC), and dimethyl sulfoxide (DMSO) are suitable solvents for pore formation. For example, Huang et al.  preparedThe phase separation caused by the increase in polymer concentration during evaporative cooling and jet drying is a key step in the formation of pores in composite nanofiber sheaths, and if the evaporative drying process of the solvent is slow, then the phase separation process will decelerate, which will reduce the pore distribution density. In coaxial electrospinning, the core and sheath liquid flow rates are different, which affects the pore distribution density. For example, the core feed rate is high, slowing the evaporation of the solution jet and the polymer drying and reducing the pore density.Unlike BFs, which can only form pores on the surface of fibers, VIPS can form pores on the surface of and inside the fibers, and both require only one solvent.NIPS requires a highly evaporative good solvent and a low evaporative non−solvent, which is miscible with the good solvent but cannot dissolve the polymer or can only dissolve a very small amount of the polymer; the non−solvent has a high boiling point and the dielectric constant for the good solvent and the choice of polymer is wide.A certain amount of non−solvent to form a polymer/solvent/non−solvent ternary solution must be added to form pores in NIPS. The choice of solvent/non−solvent combination is very important. To achieve phase separation, the volatile order of solvent and non−solvent should be different: high volatility of good solvent and low volatility of non−solvent. As shown in 2O [For example, Qi et al.  used a P2O , and DCM2O ,85.2O ternary system at room temperature. The composition of the PAN solution approaches the equilibrium point of the ternary system as the non−solvent H2O content increases, indicating that strong phase separation may occur and result in nanofibers with porous structures on the surface and inside during electrospinning.Yu et al.  used eleThuy et al.  preparedSong et al.  preparedUnlike BFs and VIPS, which require only one solvent for the fiber to have a porous structure, non−solvent induced separation usually requires two or more solvents, and the selection of solvents is also different from that of the former two.The pore−forming mechanism of TIPS is shown in McCann et al.  electrosThus, the porous fiber membranes prepared by TIPS require harsh conditions and are unsuitable for industrial mass production.Unlike the above pore induction mechanisms, SR is a two−step process requiring post−treatment, as shown in 2S4 and MXene, which has high electrical conductivity and is a good capacitive material. MXene, which was first reported by Gogotsi’s team in 2011, has ultra−high electrical conductivity, a hydrophilic surface, and high chemical and mechanical stability [PVP and PEO are commonly used polymer materials and can be used as sacrificial components, which are removed by post−treatment (deionized water rinsing or high temperature extraction). We can control the proportion and distribution of pores by changing the sacrificial component content, but when the content exceeds 50%, the fibers removed will no longer be stable, so we must select the right proportion of polymer when preparing the precursor solution. For example, Yan et al.  used PANtability ,97. PoroMethaapanon et al.  preparedPorous structures can be obtained by the selective removal of the sacrificial phase in the post−treatment process, but the requirements for the components of the sacrificial phase and the post−treatment are strict.2 [2O3, electrons leap from the VB side of ZnO to the VB side of In2O3, and holes leap from the CB side of In2O3 to the CB side of ZnO, inhibiting the complexation of photogenerated electron−hole pairs [Photocatalysis is a simple and efficient technology for photocatalysts to remove pollutants by stimulating its oxidation−reduction ability through light. The common photocatalytic particles include CdS ,101, SnO2 ,103, AgC2 , and ZnO2 ,106,107.2 ,109. Thi2 ,114,115.2 . In the 2 . The typle pairs .Photocatalytic particles are inevitably lost during the catalytic process, which affects the catalytic performance in the cycle, and sufficient photocatalyst must be added to ensure the catalytic performance, thus increasing the cost. By loading photocatalytic particles onto nanofiber membranes through electrospinning, the loss of catalysts during recycling can be greatly avoided, saving costs and increasing the recycling capacity. Organic pollutants are natural organic substances in the form of carbohydrates, proteins, fats, and other biodegradable synthetic organic substances as constituent pollutants. Organic pollutants widely exist in water, air, and soil, and only a few can spontaneously degrade. However, the degradation rate of organic pollutants is very slow. This is because they generally have macromolecular chains or benzene rings, which inhibit the growth of organisms and are resistant to light, chemical and biodegradation, so they are difficult to spontaneously degrade in nature. The photocatalytic degradation of organic pollutants is a green and clean technology, which has great application prospects.16H18ClN3S, and its solubility in water is very large. This is because it contains benzene ring, cyano group and methyl group, its structure is relatively stable, and it contains sulfur ions, which can inhibit the growth of bacteria, and its spontaneous degradation is extremely slow [2 nanoparticles by electrospinning. TiO2, as a common photocatalytic material, has been widely used because of its advantages of low cost, good stability, and low toxicity [2 has the highest photocatalytic activity. By adding water−soluble polymer PVP and backbone polymer PAN to the electrospinning precursor and removing PVP by washing the fiber membrane with deionized water, a nano−PAN fiber membrane with high porosity on the surface can be obtained, exposing more TiO2 to the fiber surface, which not only increases the accessibility of TiO2 to light and generates more electrons and holes but also increases the contact between the TiO2 and the MB and the reaction active sites, thus improving the photocatalytic efficiency. The prepared pore size of 5–40 nm was analyzed by BJH. The pore size not only increased the adsorption of organic dyes but also immobilized the TiO2 particles to a great extent and prevented the shedding of catalysts in the photocatalytic process. The successful preparation of this porous fiber membrane loaded with photocatalytic particles provides a new direction for the design and development of a promising environmental application with adsorption−photocatalysis. The molecular formula of RhB is C28H31ClN2O3. It has more benzene rings, carboxyl groups and methyl groups than MB, which means that its structure is very stable and will not degrade spontaneously. Long−term exposure will bring huge harm to human health [6Fe2Ti3O18 (BFTO) nanofibers with diameters of 100–150 nm by electrostatic spinning and post−treatment calcination. BFTO has a certain absorption intensity in visible light due to its lamellar structure with a proper band gap (2.5 eV). To improve the photocatalytic performance of BFTO, it was uniformly dispersed onto the fiber membrane by combining BFTO with electrostatic spinning. Through the subsequent calcination, many pores appeared on the fiber surface to increase the contact area of the BFTO with the pollutants. Au nanoparticles were also loaded onto the mesoporous BFTO fiber membrane by a simple photodeposition method because the plasma effect on the surface of Au nanoparticles improves the absorption of visible light by the BFTO nanofibers, and Schottky defects are generated at the Au and BFTO interface to promote the separation of electron–hole pairs, thus significantly improving the photocatalytic efficiency. To further investigate the effect of the iron polarization of BFTO nanofibers, a series of corona polarization experiments were conducted, and the structures proved that the photocatalytic efficiency of RhB was significantly improved after corona polarization. Therefore, porous Au/BFTO nanofiber membranes subjected to corona polarization have a wide range of application prospects for future industrial wastewater treatment. As shown in 2 porous fiber membranes by electrospinning. Although TiO2 possesses the advantages of low price and good stability, it has poor reusability and may cause secondary pollution. Therefore, porous PCL fiber membranes with a large specific surface area were loaded with TiO2 to enhance the capacity and dispersion of TiO2 and achieve photocatalytic efficiency.The organic dye compounds in general industrial wastewater are not only very stable under normal conditions but also highly toxic, mutagenic, and carcinogenic to any organisms. Therefore, these toxic organic dyes must be effectively removed from industrial wastewater to prevent them from harming the ecosystem . Porous ely slow ,131. MB ely slow  preparedtoxicity ,134. TiOn health ,136. Gu n health  prepared6H5NO3, and it has functional groups with stable structures such as benzene ring, nitro group, and hydroxyl group, so it is difficult to degrade spontaneously [3O4 and CuO by electrostatic spinning using PVP and PEG as polymer templates. By removing the polymers through calcination, nanofibers with mesoporous structures can be constructed, thus increasing the specific surface area of the photocatalyst and the reactive sites, while mesopores shorten the electron transport path and increase the reactivity. The surface of the nanofibers prepared by electrostatic spinning contains many hydroxyl groups (−OH), which can improve the photocatalytic efficiency of p−nitrophenol and phenol.Phenolic compounds are also an important class of wastewater pollutants produced by the chemical, paint, pesticide, biotechnology, and textile industries that can invade the human body through the digestive tract, the respiratory tract, and the skin and bind to proteins in the cellular protoplasm to cause cell inactivation. Nitrophenol is an important organic pollutant with an anti−degradation ability, carcinogenic substances, and mutagenic agents. The molecular formula of nitrophenol is Caneously ,139. Oncaneously  prepared6H5ClO. Because it has a benzene ring and contains chloride ions, it can inhibit bacterial growth and is not easy to spontaneously degrade [3N5 nanofibers by electrostatic spinning−calcined nitridation and deposited Bi2MoO6 on their surface by in situ growth to form heterojunctions with Ta3N5, which inhibited the complexation of photogenerated electron–hole pairs and improved the photocatalytic activity for RhB and p−chlorophenol.As shown in  degrade . Li et a degrade  prepared22H24N2O8, and it has many structurally stable functional groups such as benzene ring and carbonyl group, which are difficult to degrade spontaneously [3N5 porous nanofibers by electrostatic spinning−calcined nitridation and deposited Ag2O on their surface by in situ growth, forming p−n type heterojunctions, which are favorable for the separation and jumping of photogenerated electrons and holes and can mineralize organic pollutants and exhibit excellent stability, showing great application potential in TC contamination treatment in the environment.Since the introduction of antipenicillin into medical therapy in 1942, hundreds of other antibiotics have been isolated or synthesized for the treatment of human and animal infections . Howeveraneously ,146. TC aneously  prepared17H18FN3O3. It has structurally stable groups such as benzene ring, cyclopropyl group, carbonyl group, and contains fluorine atoms. It can inhibit bacterial growth and is difficult to spontaneously degrade [3/CdWO4 graded porous structures (the inner and outer surfaces of the sheath layer and the outer surface of the core layer have porous structures) by electrostatic spinning. Given that WO3 and CdWO4 have different band gap properties, the formed Z−scheme heterojunction can increase the total wavelength of light absorption, make photogenerated electron holes migrate to the photocatalyst with the lowest CB/highest VB, induce the separation of electron holes, inhibit their compounding, and increase the substances that can participate in redox reactions, thus improving the photocatalytic efficiency of TC and CIP. Such nanofibers in tubes containing both Z−scheme heterojunctions and special graded porosity have more void fraction, larger specific surface area, more reactive active sites, and shorter ion transport paths, which can maximize the efficiency of each component and achieve high photocatalytic efficiency.Ciprofloxacin (CIP) is a widely used fluoroquinolone antibiotic . Its mol degrade ,150. Due degrade . As show degrade  preparedThe inorganic pollutants in water are continuously increasing with the rapid development of modern industry. These pollutants affect the pH in water, inhibit the growth of microorganisms, and hinder the self−purification of water. Among many inorganic pollutants, hexavalent Cr(VI) is an extremely dangerous heavy metal ion . Cr(VI) 2− and ·OH produced in the photocatalytic process can decompose organic substances such as proteins on the cell membrane of E. coli, and destroy the cell membrane. Because E. coli lacks the isolation and protection of the cell membrane, active substances can invade the cells, resulting in inactivation of E. coli until death [Sewage often contains pathogens that are harmful to human and animal health , porous il death ,161. As il death  preparedUnlike the BF pore−forming mechanism, which can only prepare nanofibers with porous surface, the nanofibers prepared by the VIPS, NIPS, TIPS and SR pore−forming mechanism have porous structures on the surface and inside of the fibers, which can reveal more photocatalytic particles, increase the contact area with organic pollutants, shorten the ion transport path, and show stronger adsorption capacity and photocatalytic efficiency. However, compared with the nanofibers with porous surface prepared by the BF pore−forming mechanism, they also show greater disadvantages, such as significantly reducing the mechanical properties of the fibers. As a result, the electrospun porous nanofibers are easy to fracture in the process of degrading the pollutants in water. This disadvantage may reduce the activity and reusability of photocatalyst , even wo2 skeleton by a combination of self−assembly and electrospinning. The core layer of the porous TiO2 skeleton and sheath layer can significantly improve the mechanical properties of porous nanofiber films. Changing the distribution of pores can also be used to improve the mechanical properties of nanofibers.The mechanical properties can be improved by preparing core−sheath nanofiber. Xu et al.  preparedSome nanofibers with special structures, such as beads−on−the−string nanofibers or Janus nanofibers, also have good mechanical properties. For example, Wang et al.  preparedThe mechanical properties of electrospun nanofibers can also be improved by coating the cast film on the electrospun nanofiber film. For example, Liu et al.  combinedIn recent years, electrospinning has become a common method of preparing nanofibers with various structures due to its advantages of simple operation and controllability, and nanofibers with various structures can be prepared by changing the number of polymer fluids, the type of electrospinning head, and the working parameters. This paper reviews the formation mechanism of porous fiber membranes prepared by electrospinning: BF, VIPS, NIPS, TIPS, SR. The porous structure can increase the specific surface area of nanofibers and the area in contact with the light source while shortening the electron transport path and increasing the reaction activity, thus showing a wide range of application prospects in photocatalysis. From the perspective of the environment and the economy, loading photocatalytic particles on porous nanofibers can not only effectively remove organic pollutants, heavy metal ions, and harmful bacteria in water and reduce water pollution but also greatly avoid the loss of photocatalysts in recycling and save costs.To improve the application prospects of porous nanofibers loaded with photocatalysts in the catalytic degradation of pollutants in water, the following aspects should be emphasized: (1) more effective methods are needed to construct regular pore structures; (2) the presence of many porous structures on fibers will inevitably have an impact on the mechanical properties of fibers and affect the recyclability of fibers, and the study of a highly effective method for increasing the mechanical properties of fibers will widen the range of applications; and (3) the effective and stable large−scale preparation of porous nanofibers loaded with photocatalysts is also a major challenge in industrial applications at present. With the in−depth research on photocatalysts and electrostatic spinning, these difficulties can be overcome in the near future in order to solve various environmental problems and achieve a sustainable environment.


Outcome assessment in stroke patients is essential for evidence-based stroke care planning. Computed tomography (CT) is the mainstay of diagnosis in acute stroke. This study aimed to investigate whether CT-derived cervical fat-free muscle fraction (FFMF) as a biomarker of muscle quality is associated with outcome parameters after acute ischemic stroke.t test and Mann-Whitney U test were used for statistical analysis.In this retrospective study, 66 patients  with acute ischemic stroke in the anterior circulation who underwent CT, including CT-angiography, and endovascular mechanical thrombectomy of the middle cerebral artery between August 2016 and January 2020 were identified. Based on densitometric thresholds, cervical paraspinal muscles covered on CT-angiography were separated into areas of fatty and lean muscle and FFMF was calculated. The study cohort was binarized based on median FFMF  to compare clinical variables and outcome data between two groups. Unpaired P = 0.297) and modified Rankin scale (mRS)  at admission, and pre-stroke mRS  were similar between groups with high and low FFMF. NIHSS and mRS at discharge were significantly better in patients with high FFMF compared to patients with low FFMF . 90-day mRS was significantly better in patients with high FFMF compared to patients with low FFMF .National Institute of Health Stroke Scale (NIHSS) (12.2 ± 4.4 vs. 13.6 ± 4.5, Cervical FFMF obtained from routine clinical CT might be a new imaging-based muscle quality biomarker for outcome prediction in stroke patients. Stroke remains the 2nd leading cause of death and the 3rd leading cause of death and disability-adjusted life years combined worldwide –3. HowevSkeletal muscle quality is an established target and determinant of functional recovery and thereby outcome in patients with acute severe illness . Fatty iTherefore, the purpose of our explorative study was to investigate whether the fat-free muscle fraction (FFMF) derived from diagnostic stroke workup CT might be useful as a tool for outcome prediction in patients after acute ischemic stroke due to middle cerebral artery occlusion.This retrospective study was approved by the local institutional review board, which waived the need for patient informed consent as all examinations were performed as a part of the routine clinical workup. Between August 2016 to January 2020 patients with acute ischemic stroke from emergent large-vessel occlusion in the anterior circulation who underwent endovascular treatment with a mechanical thrombectomy were identified. Inclusion criteria were: (1) complete initial and follow-up clinical examinations and scores; (2) interpretable CT examinations, including supraaortic CT angiography; (3) acute middle cerebral artery occlusion; (4) endovascular thrombectomy of middle cerebral artery occlusion with/without prior systemic thrombolytic therapy. A flow chart demonstrating the patient recruitment process is presented in the Fig. All CT examinations were performed on clinical multidetector CT scanners . CT examinations included non-enhanced CT, perfusion CT  and CT angiography after injection of intravenous contrast agent  followed by 50 ml isotonic saline solution in bolus technique with bolus tracking in a single setting. Separate contrast media boluses in the same technique were used when both CT perfusion and CT angiography were performed. Standard parameters of image acquisition were as follows: slice thickness 1 or 2 mm, tube voltage 120 kVp, tube current (exposure time product) 82 mAs. Additionally, computer-assisted automated ASPECTS (Alberta Stroke Program Early CT Score) measurements were performed for each patient as a supportive tool for the assessment of early ischemic changes.For each patient, a single-slice cross-sectional image at the level of cervical intervertebral disc space C3/4 was extracted and used for opportunistic muscle composition analysis. This level was chosen because it was always covered on the supraaortic CT angiography. Image analysis was conducted by a radiology resident  as well as a postgraduate student (L.K.) blinded to clinical data. An experienced board-certified radiologist  supervised the labeling process and was always available in cases of uncertainty. Muscle composition analysis was performed using an in-house software written in MATLAB as previously described . For segt test and Mann-Whitney U test were used for parametric and non-parametric data, respectively. Dichotomous variables were compared using x2 test (with a cell count greater than five) and Fischer test . Spearman correlation coefficient was used for a correlation analysis between FFMF and outcome data. Additionally, a univariable ordinal regression model was created, where FFMF is the predictor (independent) variable and the 90-day mRS is the outcome (dependent variable). In addition, power analysis using Cohen’s effect size d of the current study cohort has been performed. According to the power analysis from an effect size of d = 0.7, a reasonable power of 80% can be achieved with the available number of cases. The level of statistical significance was set on P < 0.05.Prism 9 (GraphPad Software) was used for statistical analysis. The whole study cohort was divided into two groups with patients with low and high FFMF based on the median FFMF . Comparisons of clinical variables and outcome data was performed between these two groups. Data was checked for normal distribution using the Shapiro-Wilk test. Data is given as mean ± standard deviation or as absolute frequencies, as appropriate. Unpaired In total, 66 patients  with complete clinical examination and CT data who underwent endovascular mechanical thrombectomy with or without prior systemic thrombolysis after acute ischemic stroke due to middle cerebral artery occlusion were identified  and high FFMF (≥71.6% n = 33). Age (P = 0.378) and sex (P = 0.999) did not differ significantly between the groups with low and high FFMF. With the only exception of atrial fibrillation (P < 0.001) all other possible stroke risk factors were similar between both groups . The percentage of patients with atrial fibrillation was significantly higher in the group of patients with low FFMF . There were no significant differences in pre-stroke mRS between patients with low and high FFMF . On admission, there were no significant differences in neurologic deficit severity and global disability between patients with low and high FFMF as assessed by NIHSS  and mRS . NIHSS on discharge was significantly higher in patients with low FFMF compared to that of patients with high FFMF . The mRS on discharge was significantly higher in the group of patients with a low FFMF compared to the patients with high FFMF . The 90-day mRS differed also in both groups with significantly higher scale values in the group of patients with a low FFMF compared to the group of patients with a high FFMF . There were 11/33 (33%) and 12/33 (36%) patients in the group with low and high FFMF, respectively, who received rehabilitation after 90 days (P = 0.768). All basic demographic and clinical parameters of the study cohort are presented in the Table Based on median FFMF the study cohort was binarized in patients with low FFMF  and corresponding skeletal muscle area  between patients with low and high FFMF.There were no significant differences in mean total skeletal muscle compartment area (40.9 ± 7.1 cmP < 0.001). Mean muscle attenuation was significantly lower in patients with low compared to patients with high FFMF . The fatty muscle area in the first group was significantly higher in patients with low compared to patients with high FFMF . According to the correlation analysis outcome data  correlated significantly with FFMF. Univariable ordinal regression analysis revealed an association between outcome data (the 90-day mRS) and CT-derived FFMF . Parameters of opportunistic muscle composition analysis are presented in the Table Mean FFMF was 60.1 ± 10.8% in the lower median group and 79.5 ± 5.3% in the higher median group  all other major risk factors for stoke under investigation  were also similar in the both groups. This might have an influence on our study results, especially in patients who underwent systemic thrombolysis prior to endovascular thrombectomy [P = 0.671). In fact, in the group of patients with high FFMF, recanalization failed in slightly more cases (12% vs. 9%), which, however, had also no influence on general outcome in the whole group. These findings are consistent with the existing literature but in case of sarcopenia. Sarcopenia is proven to be related to adverse outcomes in stroke patients [Our study results show that regardless of similar baseline anthropometric and demographic characteristics, pre-stroke disability, and comparable severity of global and neurological disability on admission, patients with low FFMF demonstrated a worse clinical outcome and were more likely to have persistent neurological disabilities on discharge and after 3 months. It is worth mentioning that with the only exception of atrial fibrillation . However, this was beyond the scope of our explorative study and assessment of myosteatosis is no standard of care in patients with acute ischemic stroke Another limitation of our study is that we did not perform body composition analysis of other muscle regions. The loss of some other muscle types and groups  are especially known to be associated with aging and first affected by sarcopenia . However, whole-body CT scans for assessment of other muscles groups were not available for body composition analysis in our stroke cohort.In conclusion, this is the first study indicating an association between CT-derived cervical FFMF and patient outcome after acute ischemic stroke. In the future, FFMF might be helpful in the assessment of body composition in the clinical setting of acute ischemic stroke, particularly as a prognostic marker for functional recovery. This study may also motivate further prospective and longitudinal studies on a larger population to investigate the role of CT-derived cervical FFMF as a potential new imaging-based biomarker for outcome prediction in stroke patients. Another possible important point for future studies could be the investigation of associations between neurological outcome scores and muscle quality markers of other muscle groups.