Bivariate analysis and binarylogistic regression were used to identify the factors associated with theuse and reuse of masks. 3,981 (100%) individuals participated in the study. In total, 95.5% (CI 95%:94.8-96.1) reported using masks. Fabric masks were more frequently reported, followed by surgical masks . The percentage of reuse was 71.1% (CI 95%: 69.7-72.5). Most of those exclusively wearing surgical masksreported its reuse. Being a woman and having had contact with individualspresenting respiratory symptoms increased the likelihood of wearing masks(p≤0.001). Additionally, being a woman decreased the likelihood of reusingsurgical masks (p≤0.001). virtually all the participants reported the use of masks, most frequentlyfabric masks. The findings draw attention to a risky practice, that ofreusing surgical and paper masks. Therefore, guidelines, public policies,and educational strategies are needed to promote the correct use of masks tocontrol and prevent COVID-19. Brazil ranks third in the world in the number ofdeaths; more than 680,456 cases and 36,151 deaths were confirmed up to the same datein the country-2.The COVID-19 pandemic, a disease caused by the novel human coronavirus (SARS-CoV-2),has affected people in all continents, with 6,912,751 cases confirmed up to June8.Symptoms among those infected with SARS-CoV-2 range from no symptoms up to severecases of pulmonary disease. The incubation period that involves the onset of thefirst symptoms may range from two to 14 days after the infection, though some peopledo not present any symptoms-4. Transmission mainly occurs fromperson to person through respiratory droplets, when an individual coughs, sneezes orspeaks near other people, while contaminated surfaces or objects may also be sourcesof transmission.Note that, even if asymptomatic, individuals infected with SARS-CoV-2 have thepotential to transmit the virus. Hence, to self-protect againstdiseases and also prevent the transmission of pathogens between healthy and sickindividuals, masks have been used as a popular public health intervention.Therefore, the general population is advised to wear face masks as a mechanicalbarrier to prevent the dispersion of droplets, in addition to frequent handsanitization using soap and water or alcohol-based solutionsnd, 2020, to make and wear theirhomemade masks. It also advises that masks made of vacuum bags, cotton, orantimicrobial pillowcases ensure an effective physical barrier, provided they areproperly designed and sanitized.In Brazil, given a shortage of personal protective equipment (PPE), especiallysurgical masks, the Ministry of Health advised the population, through aninformative note published on April 2. Surgical masks are disposable and should not be sanitizedbecause they lose their filtering capacity. Similarly, N95 respirators should not bewashed and the number of times it may be reused is informed bymanufacturers.Note that to ensure masks are effective for what they are proposed, that is, toimpede the dissemination of expelled droplets, masks need to completely cover mouthand nose and adjust well to the face, without side clearances. Homemade masks arerecommended to be sanitized before reuse.How frequently the population wears masks increases with the onset of localepidemics. It is, however, necessary to better investigate factors such as theduration of protection and how this piece of PPE is used and reused, consideringthat inappropriate use, especially of disposable masks, may compromise itsprotective effect and even raise the risk of infectionIn this sense, considering that the use of masks is not a common practice in thepopulation in general, in times of pandemic, this measure has been stronglyencouraged and represents a new reality experienced by the Brazilian people.Therefore, identify how people have used and reused this protective piece ofequipment is extremely important to discuss the potential effects of not wearing thedifferent types of masks or wearing them inappropriately in the face of the COVID-19pandemic. Hence, this study’s objective was to identify the factors associated withthe use and reuse of masks among Brazilian individuals during the COVID-19pandemic.th and27th in the North, Northeast, Southeast, Midwest, and South ofBrazil. This study is part of a multinational project addressing the practice ofreusing masks in the population in general during the COVID-19 pandemic in Braziland is part of an international study “Practice of face mask use among generalpublic during the outbreak of COVID-19: a multi-country cross-sectional study”, inpartnership with Squina International Centre for Infection Control, Hong KongPolytechnic University.This cross-sectional study was conducted between April 16Instituto Brasileiro de Geografia e Estatística - IBGE)concerning the 4th trimester of 2019, the population of Brazilianindividuals aged over 18 years old is approximately 159,095,000. A representativesample of this population group was used. A finite population sample was calculatedwith a confidence interval of 98%, sampling error of 2%, prevalence of 50%, and testpower of 80% and a minimum sample of 3,393 individuals was found. The inclusioncriterion was being 18 years old or older, while the exclusion criterion was being aforeigner living in Brazil or a health worker.According to data made available by the Brazilian Institute of Geography andStatistics ; 2- Information concerning theuse of masks: type of mask used , frequency of reuse (number of times a mask is reused).Considering that it is a multinational study, the questions were translated fromEnglish to Portuguese by two independent translators and were content validated by apanel composed of five experts in the subject. No changes were suggested. A pilottest was conducted prior to the data collection with 20 individuals convenientlyselected from five different Brazilian regions to verify the instrument’ssensitiveness and applicability. Only adaptations in terms of the form’s onlinepresentation and layout were suggested and implemented. The instrument was convertedto the online format using Google Forms.Invitation to participate in the study was sent through messages disseminated insocial media, containing a link to the survey. The researchers also published thelink to access the instrument in their social media profiles. By accessing the link,the Internet user was directed to the Google platform with access to the form. Afree and informed consent form was presented on the initial page and participantsonly accessed the instrument of data collection after providing their consent.The outcome (dependent) variables were mask use and surgical mask reuse. Theindependent variables for both were: sex, age group, educational level, maritalstatus, and contact with people presenting respiratory symptoms.® and later to IBM® SPSS, version 20.0. Descriptivestatistical analysis included absolute and relative frequencies, to characterize theparticipants and data concerning the use and reuse of masks, in addition to centraltendency  and dispersion measures (standarddeviation) for age.Data were exported from Google Forms directly to a spreadsheet in Microsoft OfficeExcelPrevalence concerning the use and reuse of masks was estimated considering aconfidence interval of 95% (CI 95%). The Chi-square test was adopted to analyze theassociation between the outcome and independent variables. Associations withp<0.20 were verified using binary logistic regression considering the dependentvariables were dichotomous, the probabilities concerning the use of face masks andreuse of surgical masks in the population were verified using Odds Ratio (OR) andits respective confidence intervals of 95%. Associations were consideredstatistically significant when p<0.05.Comissão Nacional de Ética em Pesquisa - CONEP) . All ethical aspects were complied with inBrazil according to Resolutions 466/2012 and 510/2016 of the National HealthCouncil. Financial support: Conselho Nacional de Desenvolvimento Científicoe Tecnológico (CNPq), Brazil, grant #401371/2020-4.The project was submitted to and approved by the National Committee of EthicsResearch  individuals participated in the study. Regardingsociodemographic characteristics, the regions with the largest number ofparticipants were the Southeast with 1,874 (47.1%) and Northeast with 1,348 (33.9%),followed by the Midwest with 320 (8.0%), North with 265 (6.7%), and South with 174(4.4%). Most, 2,893 (72.7%) were women, 2,057 (51.7%) were married, aged 40 yearsold on average (SD=13.9); minimum 18 and maximum 86 years old. Concerning theeducational level, most participants, 3,280 (82.4%), reported holding a collegeeducation degree. Most, 2,622 (65.9%), reported no close contact with individualswith respiratory symptoms; 1,359 (34.1%) mentioned at least one contact, 224 (5.6%)of whom had daily and continuous contact with people presenting respiratory symptoms.per region, the region with the highest rateof use was the South with 172 , followed by the North with259 , Northeast with 1,290 ,Southeast with 1,779  and the Midwest with 303 .Most of the participants, 3,803  reported the use of masks;only 178 individuals  reporting not wearing masks. When thefrequency is verified Regarding the type of mask used, the most frequently reported type was fabric/cottonhomemade masks, used exclusively or in combination with other masks, 2,895 , followed by surgical masks, reported by 1,108  individuals, N95 respirators reported by 335 ,paper or gauze reported by 293 , active carbon by 38 , industrial masks by 14 , and other types ofmasks were reported by 10  participants.Note that most of the 3,803 (100%) individuals who reported the use of masks, 3,002, reported wearing only one type of mask, followed by 716 who reported two types of masks; 80  individuals reported three types; three individuals reported wearing four different types of masks; and two individuals  reported five types of masks. The combination of masks most frequentlyreported was fabric/cotton homemade masks in combination with surgical masks, 371.The variables that presented p<0.20 in the association between variables were: sex(p=0.000), age (p=0.084), educational level (p=0.047), and contact with individualspresenting respiratory symptoms (p=0.001) . These vAmong the 3,803 (100%) who reported the use of masks, 2,832  reported reusing masks, while 1,149  did notreuse masks. Among the 2,832 (100%) who reuse masks, 947 reported reusing 1-2 times; 717  reported masks were used3-4 times; 661  reported reusing masks more than 7 times;and 507  reported reusing masks 5-6 times .Note that most of the 551 (100%) participants who report exclusively wearing surgicalmasks, 308  reuse masks according to the followingfrequencies: 150 (27.2%) rarely/1-2 times reuse masks, 91 (16.5%) reusesometimes/3-4 times, 44 (8.0%) frequently/5-6 times, and 23 (4.2%) participantsreuse always/7 times or more. Of those, 131 (100%) who wear only paper or gauzemasks, 60  participants reuse according to the followingfrequencies: 24 (18.3%) reported using masks rarely/1-2 times; 23 (17.6%)sometimes/3-4 times, 8 (6.1%) reported always/7 times or more, and 5 participants(3.8%) reported reusing masks frequently/5-6 times. Among the 2,152 (100%) whoexclusively wear fabric/cotton masks, 1,685  reuseaccording to the following frequencies: 513 (23.8%) reported rarely/1-2 times; 483(22.4%) reported always/7 times or more; 371 (17.2%) reported sometimes/3-4 times,and 318 (14.8%) reported frequently/5-6 times. The confidence intervals concerningthe prevalence of reuse according to types of masks are presented in In the association between variations concerning the reuse of surgical masks, sex(p=0.001) was the variable that presented p<0.20 and was thus included in thebinary logistic regression model for the outcome variable “reuse of mask” .After the binary logistic regression, being a woman  continued as a factor associated with the more frequent use of masks,presenting approximately twice the likelihood of wearing masks, while having contactwith people presenting respiratory symptoms increased approximately 1.8 times the likelihood of wearing masks. Being a woman also remained a factor associated with the reuseof surgical masks, though, the reuse of surgical masks was two times less likelyamong women than in the population .-11.This study’s findings reveal the pattern of use and reuse of masks in the Brazilianpopulation. Virtually all the participants reported the use of this piece of PPE andfabric masks were the most frequently reported. As the COVID-19 pandemicincreasingly aggravated worldwide, the use of masks has been recommended to thepopulation as a non-pharmacological intervention that plays a vital role incontrolling the spread of the disease, mainly because the dissemination of theSARS-CoV-2 from human to human mainly occurs through respiratory droplets.The recommendation for the population to wear homemade masks during epidemics ofrespiratory infectious diseases such as the Severe Acute Respiratory Syndrome (SARS)and COVID-19, has been successful as a public health intervention at the globallevel. In China, for instance, this practice has been efficacious in combinationwith other protective strategies to prevent the dissemination of viruses causingthese two diseases.The populations of many countries in all the continents have adhered to the use ofmasks due to the rapid dissemination of the SARS-CoV-2. People in China, includingHong Kong, and Japan, Thailand, and South Korea, have used different types of masks.The growth of cases of COVID-19 in the Czech Republic, where the government has madeits use mandatory, has been slower than in other European countriesnd, 2020,the Brazilian Ministry of Health recommended the population at a national level tostart wearing masks. The adoption of this recommendation is confirmed in a studyconducted in the second half of April of the same year, which reports that theprevalence of the use of masks was almost absolute in all Brazilian regions; theSouth presented the highest prevalence. Being a woman doubles the likelihood ofwearing masks. One study conducted in China, addressing 10,304 participants duringthe period of the COVID-19 pandemic, also reports greater adherence to protectivemeasures among women, indicating that sex influences adherence to officialrecommendations related to health.On April 2. One Chinese study, however,verified that this use, regardless of the presence of respiratory symptoms, wasassociated with lower levels of anxiety and depression, that is, the use of maskshas potential psychological benefits, giving a sense of safety. Even though masks promote asense of safety, despite having contact with asymptomatic individuals or those withsymptoms, the use of masks cannot be the only measure adopted at the expense ofother essential protective measures to prevent COVID-19. Social distancing and handsanitation with soap and water, or with an antiseptic solution of at least 60%alcohol, are highlighted.In this study, the likelihood of wearing masks almost doubled among those who hadcontact with individuals with respiratory symptoms, which is in agreement with theWorld Health Organization’s recommendation to those with a confirmed diagnosis orsuspected infection and their respective caregivers to wear masks, reinforcingthe need for educational strategies directed to the entire population, especiallythose with fewer years of schooling.Another aspect to take into account is the participants’ educational level, as mostreported a higher education degree, suggesting they had a better understanding ofthe importance of wearing masks to prevent the COVID-19. This finding corroborateswith a literature review addressing the use of masks to prevent respiratoryinfections, which revealed that higher education was positively associated with thepractice. As current results show, the useof this type of mask among Brazilian individuals has been prevalent during theCOVID-19 pandemic.Regarding the type of masks, those made of fabric predominated in this population,emerging as an alternative in the face of a shortage of surgical masks. Even thoughfabric masks are not as effective as those recommended for hospital use, adherenceto fabric masks reduces the risk of disseminating respiratory infections.Additionally, they are particularly popular in developing countries given theiravailability, low cost, and the possibility to be reused after washing. Therefore, investment is needed,especially on the part of public policymakers, to provide educational guidelines onhow to reuse masks to the population in general.The reuse of this piece of PPE was a practice mentioned by a large portion of theindividuals addressed in this study. The reuse of masks requires attention andshould be better discussed with the population adopting this practice. Fabric masksneed to be washed after being used. The circumstances in which individuals considerit necessary to wash masks were not addressed in this study, but it became apparentthat a good portion of the participants reused masks, at least five times. Washing,drying, and stretching gradually increase the porosity of fabrics and therefore,impair filtration efficiency, as a study conducted in Nepal shows. Additionally,more than half of the participants who reported the exclusive use of surgical masksreuse them more than once, revealing a risky practice that demands the immediateintensification of public intervention, using educational strategies to ensure thecorrect use of this piece of protective equipment.The study shows that, even though surgical masks are recommended to be discardedafter use, reuse reports were considerably high. Surgical masks are disposable andshould not be cleaned or disinfected for later use as they lose their filtrationcapacity when wet; studies have not reportedconclusive evidence. Therefore, the reuse of surgical masks is not recommended tothe population without an effective possibility to safely decontaminate them. Infact, the reuse of surgical masks may even increase the risk of contamination byCOVID-19.Researchers have tested alternatives to decontaminate masks, including surgicalmasks, however, some of these alternatives may damage the masks’ blocking structurethrough physical or chemical action, or incompletely inactivate pathogens. Moreover,these alternatives may not be available to the population in general as they requirespecific instruments or material. Additionally, further studies are needed to assessappropriate measures to decontaminate N95 respirators and surgical masks. Hence, educational actions arenecessary to address preventive measures in the context of the COVID-19 among themale population.Another noteworthy finding in this study is the considerable number of people whouse, and reuse, paper or gauze masks as respiratory protection, even though, thistype of masks have not been recommended in Brazil. Another factor is that the reuseof masks is less frequent among women than among men, reinforcing that sex isassociated with adherence or lack of adherence to protective practices. These aresituations that should be carefully assessed by health managers in Brazil to devisemore assertive plans and protocols.This study only addressed the use and reuse of masks in the Brazilian population,however, it is important to stress that masks must be adopted together with otherpreventive measures. This research draws attention to two concerning facts: theinappropriate use of masks and/or the possibility that people become careless withother preventive measures when wearing masks. Therefore, the set of measures thatcan decrease the spread of SARS-CoV-2 and flatten the pandemic curve, include socialdistancing, respiratory etiquette, and handwashing, together with the appropriateuse of masks.The current COVID-19 pandemic remains severe worldwide and is an international sourceof concern, considering that it is a disease with high contagious potential, so theimportance of sensitizing the population, reinforcing control measuresThis study’s limitations include the fact that digital illiterates were excluded, theimpossibility to clarify doubts when participants did not understand a question andlack of knowledge regarding the circumstances in which the form was completed.The results show that most of this study’s participants reported the use of masks andthat fabric masks were more frequently reported. Being a woman and prior contactwith individuals with respiratory symptoms were associated with the use of masks,increasing its likelihood. Being a woman was also associated with the reuse ofsurgical masks though, as women are twice less likely to reuse this type of mask inthe population.This study’s findings draw attention to a risk practice, that of reusing paper andsurgical masks, which increases the chance of transmission given inefficaciousrespiratory protection. In this sense, this study presents important contributionsin the health field as it provides new scientific evidence regarding the use andreuse of masks, which can be used to support the establishment of guidelines, publicpolicies and educational strategies to promote the adoption of correct practices tocontrol and prevent the COVID-19 in the Brazilian territory.


Dialister genus was significantly lower and Blautia, Desulfovibrio, and Succinivibrio were significantly higher in abundance in CHD patients compared with the HT group, and the changes were significantly correlated with physiological indexes, such as increased lipopolysaccharides. Moreover, enrichment of genes decreased in four pathways of amino acid metabolism, such as arginine biosynthesis and histidine metabolism, although two lipid metabolism pathways, including fatty acid degradation and arachidonic acid metabolism, increased in the CHD group. Additionally, occupation and a family history of CHD were shown to be risk factors and affected the gut microbiota in Tibetans. Our study will provide insights into the understanding of CHD, leading to better diagnosis and treatment of Tibetan patients.Coronary heart disease (CHD) is closely related to gut microbiota, which may be significantly affected by ethnicity and the environment. Knowledge regarding the gut microbiome of Tibetan CHD patients living in the Qinghai–Tibet Plateau is very limited. In this study, we characterized the physiological parameters and gut microbiota from 23 healthy Tibetans (HT), 18 CHD patients, and 12 patients with non-stenosis coronary heart disease (NCHD). We analyzed the alterations of the gut microbiome in CHD patients and investigated the relationship between these alterations and the pathological indicators. We found no changes in trimethylamine N-oxide, however, a significant increase in lipopolysaccharides and white blood cells, and a decrease in high-density lipoprotein were observed in the blood of CHD patients, compared to that in the HT group. The gut microbiota of the NCHD group had a significantly higher Shannon index than that of the HT group. Adonis analysis showed that both microbial compositions and functions of the three groups were significantly separated. The  China Cardiovascular Diseases Report 2018 -induced inflammation, which may be a new strategy for the prevention and treatment of CHD , 18 Tibetan inpatients with CHD, and 12 Tibetan inpatients with non-stenosis coronary heart disease (NCHD) were profiled by 16S ribosomal DNA (rDNA), and physiological parameters were measured in the three groups. Metagenomic sequencing and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional analysis of 39 samples  were performed. Bacterial community structures, co-occurrence networks, and functional differences between the samples were assessed. We also explored the relationship between the gut microbiome and physiological parameters to better explain the pathogenesis. In addition, we performed a preliminary examination of the effects of occupation and family genetic background on the gut microbiota of Tibetan patients with CHD. Our findings provide insights for the prevention and treatment of CHD in this population.The study was approved on 09 March 2017 by the Ethics Committee of Qinghai Province Cardiovascular and Cerebrovascular Disease Specialist Hospital, China. All subjects provided written informed consent in accordance with the Declaration of Helsinki. Informed consent was obtained after written introductions were provided to all 53 Tibetan volunteers from the Tibetan Autonomous County in the Qinghai Province of China. None of the respondents used antibiotics, probiotics, or prebiotics for at least 3 months before sampling, and their ages ranged from 40 to 70 years.The diagnostic records of the Tibetan patients were collected from August 2017 to February 2018 from the cardiac center, Qinghai Cardio-cerebrovascular Disease Specialist Hospital in Xining City. Thirty Tibetan patients completed questionnaires that included information. Subjects without CHD and acute and chronic inflammatory diseases that may affect gut microbiota, including diseases of the digestive system, tumors, renal failure, and other serious diseases were excluded.The 30 Tibetan patients were divided into two groups according to their coronary angiography results. 18 patients with coronary artery stenosis >50% were placed in the CHD group and 12 patients with coronary artery stenosis <50% were placed in the non-stenosis coronary heart disease (NCHD) group. The patients in the NCHD group had coronary risk factors, including hypertension, dyslipidemia, arrhythmia, and ischaemic changes evident on an electrocardiogram (ECG) .Twenty three HTs, most of whom were patients' accompanying nursing professionals, were included in the control group. The individuals in the HT group were family members of the patients and so their diets were similar to the patients. Exclusion criteria for the HT group included those with a history of cardiovascular disease, diabetes, and patients with treated hyperlipidaemia. All participants in the HT group were Tibetan native residents, had similar living habits, and were required to complete the same questionnaires as the patients. The ECG and biochemical indexes in the blood were investigated in the HT group.Fecal samples were collected at the hospital while the patients consumed the hospital diet and the samples were immediately frozen at −80°C before being further processed. Next, the samples were submerged in 30 mL of 97% ethanol for 24–36 h. The remaining solid materials were transferred to 50 mL tubes with silica beads . Blood samples were collected in the morning following overnight fasting conditions. Plasma was collected by centrifugation and stored at −80°C until use.Biochemical parameters in plasma, renal function parameters, blood lipids, clotting factors, and blood cell counts were determined using a model AU2700 fully automatic blood biochemical analyser . The 17 parameters assessed were: triglyceride (TG), total cholesterol (TC), high-density lipoprotein (HDL), low-density lipoprotein (LDL), apolipoprotein A1 (APO-A1), apolipoprotein B (APO-B), lipoprotein(a) [LP(a)], glucose (GLU), blood urea nitrogen/creatinine (BUN/Cr), uric acid (UA), white blood cell (WBC) count, red blood cell (RBC) count, hemoglobin (HGB), haematocrit (HCT), platelet (PLT) count, homocysteine (Hcy), and C-reactive protein (CRP).Metabolites of choline, betaine, creatinine, and l-carnitine, which are associated with TMAO, were selected for preparation of the standard solution. Choline, betaine, TMAO, creatinine, and l-carnitine were accurately weighed, and a high concentration of the standard solution was prepared as an internal solution with 1% formate-acetonitrile. The standard solution was stored at −20°C for short-term use.In 20 μL of the sample or standard solutions, 10 μL of internal standard solution was added, followed by 750 μL of 1% formic acid-acetonitrile solution, mixed, and centrifuged. The supernatant was collected.A high-performance liquid chromatography-tandem mass spectrometry method was applied to explore the metabolites of choline, TMAO, betaine, creatinine, and l-carnitine in plasma. Chromatographic separation was performed on an ACQUITY UPLC® BEH HILIC 2.1 column , with 5 μL of the sample, mobile phase A-acetonitrile, B-water, including 10 mm ammonium acetate, and a flow rate of 0.4 mL/min at 40°C. The gradient elution program was as follows: 0~1 min, 80% A; 1~2 min, 80~70% A; 2~2.5 min, 70% A; 2.5~3 min, 70~50% A; 3~3.5 min, 50% A; 3.5~4 min, 50~80% A; and 4~6 min, 80% A. Mass spectrometry elution was performed with an electrospray ionization source in the positive ion mode. Multiple response monitoring scans and quantitative analyses were performed on all plasma samples.A double-antibody one-step sandwich enzyme-linked immunosorbent assay kit was used for LPS evaluation. The samples, standards, and horseradish peroxidase-labeled detection antibodies were loaded into the coated micropores with precoated human LPS-captured antibodies and incubated thoroughly before washing. The 3, 3′, 5, 5′-tetramethylbenzidine (TMB) substrate was converted to blue in the presence of peroxidase and to the final yellow color with the addition of an acid stop solution. To quantify LPS, absorbance was measured on a microplate reader at 450 nm and the sample concentration was calculated by comparison to a standard curve.DNA from fecal material was extracted using a QIAamp DNA Stool Mini Kit  following the standard protocol. The DNA concentration was determined using a NanoDrop ND-1000 . The V3 and V4 regions of 16S rDNA from all samples were amplified using the 341F (5′-CCTAYGGGRBGCASCAG-3′) and 806R (5′-GGACTACNNGGGTATCTAAT-3′) primers. The polymerase chain reaction (PCR) products were quantified and purified using a QuantiFluor™ fluorometer . Negative controls included no template controls for DNA extraction and PCR amplification. PCR products were mixed in equidensity ratios. The mixture of PCR products was purified with a Gel Extraction Kit (QIAGEN). Sequencing libraries were generated using the TruSeq® DNA PCR-Free Sample Preparation Kit  following the manufacturer's recommendations, and index codes were added. The library quality was assessed on a Qubit@ 2.0 Fluorometer (Thermo Fisher Scientific) and the Bioanalyzer 2100 system . Finally, the library was sequenced on a HiSeq2500 platform (Illumina), and 250 bp paired-end reads were generated.DNA from 39 samples  was sequenced using the HiSeq platform (Illumina) and paired-end reads were generated. The raw data were collected using Readfq V8 to acquire clean data for subsequent analysis, producing an average of 540 million metagenomic jointed reads (12 Gb) per sample.The reads of 16S rDNA were trimmed based on the cut-off of Q-value >20, missing bases >10%, and relative abundance <40%. The high-quality-paired reads were merged into a single tag based on the overlapping region using FLASH v1.2.11  was used to blast Unigenes to a functional database with the parameter setting of blastp, and the best blast hit was used for subsequent analysis , Bacteroides (9.4%), and Faecalibacterium (5.6%). The HT group had a higher abundance of Prevotella (23.7%) and Dialister genera (2.6%) in comparison to the CHD (13 and 0.9%) and NCHD (7.6 and 0.8%) groups  groups .Faecalibacterium was the most abundant network of the HT group, with lower abundances in the CHD and NCHD groups. In particular, Prevotella was the most abundant genus in the CHD group and Bacteroides was most frequently observed in the NCHD group, but these were barely detectable in the HT group. These three genera were more negatively correlated with the OTUs of other genera.We found that the network structures in some patients were more complex, as the NCHD group had the most nodes and links, while the HT group had the least . The genTo gain an insight into the functional changes associated with the patients' gut microbiome, metagenomic sequencing data were processed and the relative abundance of the KEGG orthology (KO) and gene pathways were obtained. The richness of KOs, defined as the number of KOs detected within each subject, was significantly higher in the CHD group than in the HT group, while they were not significantly different in terms of KO Shannon diversity . Functiot-tests using STAMP to compare the level-2 and level-3 KEGG pathways between the groups. Compared with the HT group, two level-2 pathways, including amino acid metabolism and global and overview maps, and 13 level-3 pathways, such as valine, leucine, and isoleucine biosynthesis, starch and sucrose metabolism, 2-oxocarboxylic acid metabolism, and thiamine metabolism, were decreased, while three level-3 pathways, including ascorbate and aldarate metabolism, arachidonic acid metabolism, and fatty acid degradation, were increased in the CHD group , and other factors. A correspondence analysis (CA) of the survey data revealed lifestyle differences among the three groups . The firLachnospira and Dialister genera and a greater abundance of the Oscillospira and Blautia genera were found in herdsmen in comparison to other professions  of CHD than in those without . The monthly intake of beef and mutton by each adult exceeds 30 kg Liu, . The lonBacteroides and Prevotella, in the microbiota of Asians and Americans (Ley, Prevotella in the network (Kehrmann et al., Faecalibacterium revealed it as the dominant bacterium, while other genera were more balanced. Faecalibacterium is considered an anti-inflammatory bacterium that is deficient in many metabolic disorders (Machiels et al., Lachnospira did not occupy a notable position in the network structure, while the genera Ruminococcaceae were important and may have been decreased in Dutch cardiovascular patients (Koopen et al., There were more lines in the network structure of the CHD and NCHD groups than in the HT group, showing that the relationship and interactions between intestinal flora in patients with CHD were more complex than in HTs. A similar antagonistic relationship has been described between the two dominant groups, ans Ley, . HoweverWe also found that occupation and family history of CHD were the most important lifestyle and genetic factors associated with CHD in Tibetans. Over the past 40 years, rapid urbanization in China has led to Tibetans discovering jobs and settling in towns or cities, leading to the westernization of their gut microbiota (Li et al., Blautia, Dialister, and Succinivibrio genera, between Tibetan patients with CHD and NCHD and healthy Tibetans. These findings suggest a combined effect from the gut microbial community. The pathology of CHD is accompanied by significantly increased plasma and fecal LPS, rather than TMAO. The bacterial alterations resulted in an appreciable reduction in intestinal amino acid metabolism pathways, and they were significantly related to LPS and HDL levels and numbers of WBCs, which may lead to low inflammatory coronary atherosclerosis. Finally, occupation and FMH of CHD were important risk factors in Tibetan individuals.In conclusion, we discovered risk biomarkers and a significant difference in the presence of the gut microbiome, including The measured 16S rDNA sequences have been uploaded to the National Center for Biotechnology Information using accession number: PRJNA550301. The metagenomics data are available from the corresponding author on reasonable request. Other data can be found in the article/The studies involving human participants were reviewed and approved by Ethics Committee of Qinghai Province Cardiovascular and Cerebrovascular Disease Specialist Hospital. The patients/participants provided their written informed consent to participate in this study.FL and YZ conceived the study, directed the project, designed the experiments, interpreted the results, and wrote the manuscript. CF and LZ performed the computational and 16S rDNA microbiota analysis and wrote the results and methods. HH, YM, YT, and JF collected the samples and clinical data. YL supervised the fecal microbiota collection and detected the biochemistry parameters. SJ supervised the data analysis and revised the manuscript. TW revised the manuscript. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Interest in the involvement of the cholinergic system at the non-neuronal level emerged in recent years, thereby allowing to identify different physiological and pathological processes that acetylcholine and its receptors modulate in different tissues. The cholinergic system can also take part in the activation and maintenance of pathological processes such as neurodegenerative diseases and cancer. This review summarizes studies concerning the involvement of the cholinergic system in different tumor isotypes, with a special focus on melanoma, the most lethal form of skin cancer.In the last two decades, the scientific community has come to terms with the importance of non-neural acetylcholine in light of its multiple biological and pathological functions within and outside the nervous system. Apart from its well-known physiological role both in the central and peripheral nervous systems, in the autonomic nervous system, and in the neuromuscular junction, the expression of the acetylcholine receptors has been detected in different peripheral organs. This evidence has contributed to highlight new roles for acetylcholine in various biological processes, . In addition, growing evidence in recent years has also demonstrated new roles for acetylcholine and its receptors in cancer, where they are involved in the modulation of cell proliferation, apoptosis, angiogenesis, and epithelial mesenchymal transition. In this review, we describe the functional characterization of acetylcholine receptors in different tumor types, placing attention on melanoma. The latest set of data accessible through literature, albeit limited, highlights how cholinergic receptors both of muscarinic and nicotinic type can play a relevant role in the migratory processes of melanoma cells, suggesting their possible involvement in invasion and metastasis. Acetylcholine (ACh) was the first molecule identified as a neurotransmitter operating through two subgroups of receptors, namely muscarinic receptors (mAChRs) and nicotinic receptors (nAChRs) . The preThe importance of ACh as a modulator of different biological processes and the concept itself of non-neuronal acetylcholine function, has been growing since the early 1970s, capitalizing on the discoveries of functional cholinergic receptors in non-neural cells . In part+, Ca2+ and K+, with α7 nAChR displaying the highest permeability for the Ca2+. In fact, the binding of nAChRs to acetylcholine or nAChRs agonists leads to membrane depolarization and opening of voltage-gated ion channels, resulting in ion influx and efflux [The nAChRs are fast ionotropic cationic acetylcholine receptors that mediate the fast-synaptic transmission, whose activation is extremely quick (msec to sub-msec) ,32. Theyd efflux . The chad efflux .As previously described, the nicotinic acetylcholine receptors are expressed not only in the neuronal system, but also in numerous non-neuronal tissues such as skin, pancreas, and lung, thus suggesting a role in other biological processes in addition to synaptic transmission. Specifically, these receptors were found to play a role in the regulation of cellular processes such as cell proliferation and cell death .It is known that a dysregulation of nAChRs is associated with neurological disorders such as Alzheimer, Parkinson’s disease, and schizophrenia . MoreoveThe other class of acetylcholine receptors is represented by muscarinic receptors, which belong to the class of heptaelical G protein-coupled receptors and consist of five subtypes (M1–M5). Like all the G protein-coupled receptors, they share a structure composed of seven transmembrane helices (TM1–TM7) connected by three intracellular (i1–i3) and three extracellular (e1–e3) loops. The receptor activation occurs always via coupling to heterotrimeric guanine nucleotide-binding proteins and subsequent kinases activation or ion channel activity regulation ,44.+ channels. Muscarinic receptor subtypes can also activate small G proteins, such as Rho GTPase and recruit new effectors including IP3K and MAPK/ERK kinases. These last signaling pathways appear to play important roles in the control of cell growth and proliferation, as suggested by studies on glial cell primary cultures [The odd muscarinic receptors, M1, M3, and M5, are coupled to the Gq11 protein, which stimulates the IP3 hydrolysis and the intracellular calcium mobilization. They also modulate phospholipase A2 and phospholipase D in different cell lines and primary cultures ,46,47. Mcultures ,52,53,54cultures ,56. Genecultures ,58,59,60cultures . These rcultures . M3R medcultures ,64,65. Tcultures ,67. M4R cultures ,69. Dysrcultures . The lascultures . The M5Rcultures ,73,74,75cultures ,77,78.The distribution of all mAChRs in different tissues, such as nervous system , gastroiThe M1 receptor is involved in the progression of the prostate cancer, where it regulated cell migration and invasion through hedgehog signaling regulation ,92,93,94An inhibitory effect on tumor progression was demonstrated for M2 receptor subtype. In particular in human glioblastoma, the M2 receptor stimulation by the preferential orthosteric agonist arecaidine propargyl ester, was reported to inhibit cell proliferation and survival in stable cell lines and in glioblastoma cancer stem cells isolated from human biopsies ,96,97,98Several data were reported on the role of the M3 muscarinic receptor in promoting tumor cell growth, invasion, migration and angiogenesis in different tumor types such as lung, breast, ovarian, and brain cancers . In partIn 2012, 67,753 people were diagnosed with melanoma and 9251 individuals died from this type of tumor . At the Melanoma arises from genetic mutations in melanocytes, the cells producing the cutaneous pigment, which can be found in the skin, eye, inner ear, and leptomeninges ,115.In relation to clinical and histological features, melanoma can be divided into three main subtypes: superficial spreading melanoma (SSM), nodular melanoma (NMM), and lentigo malignant melanoma (LMM) .Superficial spreading melanoma is the most common type that accounts for 70% of the cases approximately. SSM may arise de novo or in association with a nevus . NodularThe nicotinic receptors have been studied in melanoma cells, highlighting the presence of multiple subunits such as α5/α9 / α3/ β4 ,126,127.Microarray studies and database analysis have suggested in melanoma a co-expressed upregulation of α9-nAChR and PD-L1, a protein overexpressed in melanoma cells and involved in in the regulation of EMT, cancer stemness, tumor development, metastasis formation, and resistance to therapy . The silwww.cBioPortal.org).Albeit α9-nAChR receptor was not indicated as a prognostic factor for melanoma, different missense mutations causing an altered function of this receptor have been identified. At least 82 different mutations  have been described in various kinds of melanoma tumors  and acetylcholinesterase (AChE) , demonstrating that melanoma cells may synthesize and degrade ACh in an autonomous manner. Moreover, M3R expression at protein level was evidenced by the use of a specific antibody for M3 receptor. In addition, the presence of mRNA for M3 and M5 subtypes was reported, indicating that melanoma cells express mainly M3 subtype and probably also M5 receptor .2+ concentration, indicating the functional activity of these receptors [Calcium is the second messenger associated with the transduction pathway downstream from the cholinergic activation of the M1, M3, and M5 receptor subtypes. In melanoma cells, fluorometric measurements with Fura-2 have shown that in the presence of the ACh mimetic carbachol (CCh), there was an increase of intracellular Caeceptors .A chemotaxis measurement assay was also performed on these cells using the Boyden chamber. The melanoma cells were placed in the upper chamber in the absence or in the presence of carbachol and/or atropine (selective muscarinic antagonist), while the chemotactic agent fibronectin was placed in the lower chamber. The time taken by the cells to migrate from the upper to the lower chamber was measured as a chemotactic power of the agent used. It was observed that the addition of carbachol in the upper chamberm increased chemotaxis by 30%, while atropine inhibited it . The migAll these evidences clearly suggest that the cholinergic activation of mAChRs on melanoma cells modulates cell migration and that, in general, their expression may be required to control invasion and metastasis formation. This may prompt the potential use of M3 antagonists to inhibit melanoma growth and metastasis, albeit this remains to be further determined. Although the M3 muscarinic receptor subtype function has been well characterized, the presence and role of other muscarinic receptor subtypes have not been thoroughly investigated in melanoma. Whilst a faint expression of M5 subtype has been described in the melanoma cells, its function remains at least unknown .The data presented in this review clearly indicate the relevant roles played by cholinergic receptors in the cancer . Compreh


Worldwide, more than 75% of people with acquired brain injury (ABI) experience communication disorders. Communication disorders are impairments in the ability to communicate effectively, that is, sending, receiving, processing, and comprehending verbal and nonverbal concepts and symbols. Such disorders may have enduring impacts on employment, social participation, and quality of life. Technology-enabled interventions such as mobile apps have the potential to increase the reach of speech-language therapy to treat communication disorders. However, ensuring that apps are evidence-based and of high quality is critical for facilitating safe and effective treatment for adults with communication disorders.The aim of this review is to identify mobile apps that are currently widely available to adults with communication disorders for speech-language therapy and to assess their content and quality using the validated Mobile App Rating Scale (MARS).Google Play Store, Apple App Store, and webpages were searched to identify mobile apps for speech-language therapy. Apps were included in the review if they were designed for the treatment of adult communication disorders after ABI, were in English, and were either free or for purchase. Certified speech-language pathologists used the MARS to assess the quality of the apps.language, 17 apps for speech, 8 apps for cognitive communication, 6 apps for voice, and 5 apps for oromotor function or numeracy. The overall MARS mean score was 3.7 out of 5, SD 0.6, ranging between 2.1 and 4.5, with functionality being the highest-scored subscale , followed by aesthetics , information , and engagement . The top 5 apps were Naming Therapy (4.6/5), Speech Flipbook Standard (4.6/5), Number Therapy (4.5/5), Answering Therapy, and Constant Therapy (4.4/5).From a total of 2680 apps identified from Google Play Store, Apple App Store, and web searches, 2.61% (70/2680) apps met the eligibility criteria for inclusion. Overall, 61% (43/70) were available for download on the iPhone Operating System (iOS) platform, 20% (14/70) on the Android platform, and 19% (13/70) on both iOS and Android platforms. A content analysis of the apps revealed 43 apps for To our knowledge, this is the first study to systematically identify and evaluate a broad range of mobile apps for speech-language therapy for adults with communication disorders after sustaining ABI. We found a lack of interactive and engaging elements in the apps, a critical factor in sustaining self-managed speech-language therapy. More evidence-based apps with a focus on human factors, user experience, and a patient-led design approach are required to enhance effectiveness and long-term use. Acquired brain injury (ABI) is a life-changing health condition that can result from trauma, cerebrovascular events, and brain tumors . The popEvidence supports the delivery of rehabilitation by speech-language pathologists (SLPs) for adults after sustaining ABI to treat language , motor sDespite the availability and potential benefits of mobile apps for adults with communication disorders after ABI, there is little published evidence regarding their quality beyond the app star rating allocated by some consumers in app store platforms and web-based reviews. A recent analysis of apps for children with speech-language disorders found that most apps were of average quality and that app cost did not always correlate with therapeutic quality . A systeIn recent years, the term gamification has become increasingly popular in digital health technologies as an underlying element to enhance individuals’ engagement with mobile health technologies. Gamification has been described by Detering et al  as the “This study aims to (1) identify the available apps designed for adults with communication disorders and (2) evaluate the quality of the available apps using the Mobile App Rating Scale (MARS) , a validThe PRISMA  guidelines were adopted for this study. These have been used successfully for evaluating speech-language therapy apps in children with speech disorders .Apps were included in this review if they were primarily designed for adults (18 years or older) with communication disorders secondary to ABI, were for the provision of speech-language therapy , were in English, were either free or for purchase, were compatible with Android or iPhone Operating System (iOS), and were available on mobile phones and/or tablets.Apps were excluded if they were primarily designed for children, were not designed for speech-language therapy , provided assessment only without speech-language therapy, and were speech-to-text/text-to-speech apps or other augmentative and alternative communication apps.aphasia,” “apraxia,” “dysarthria,” “dyspraxia,” “dysphasia,” “speech, articulation,” “speech therapy,” “speech pathology,” “language therapy,” “speech-language pathology,” “speech rehabilitation,” and “language rehabilitation”. The search terms were generated in consultation with 4 certified and experienced SLPs. One researcher entered the terms into the search fields of both the Google Play Store and Apple App Store. Boolean operators were not used as they were not supported by these platforms. In addition, webpages reviewing multiple apps for adults with communication disorders after ABI were reviewed for apps that had not been identified previously. The titles, app platform , and marketing description of all resulting apps were extracted into a spreadsheet and duplicates were removed. The last search was conducted in November 2019.To identify mobile apps, a search on Google Play Store (74.6% of the phone and 41.4% tablet market) and Apple App Store (24.8% of the phone and 58.5% of tablet market) was conducted using the following keywords: “App screening was performed by an Australian certified SLP using the marketing description of the apps against a list of eligibility criteria. If the SLP could not decide the eligibility, a second reviewer was consulted. The included apps were downloaded to either an Android  or iOS (iPhone/iPad) device, depending on compatibility, for further evaluation and quality appraisal.To identify the functionality of the apps for adults with communication disorders following ABI, a coding sheet was developed to categorize the apps based on the description of the primary therapeutic function contained in the app description on the Apple App Store or Google Play Store . The coding process was a two-step analysis in which 2 independent researchers reviewed the coding sheet and extracted relevant categories.engagement, including individual items for entertainment, interest, customization, interactivity, and whether the app was engaging for the target users; (2) functionality, including performance, ease of use, navigation, and gestural design; (3) aesthetics, including layout, graphics, and visual appeal; and (4) information quality, including accuracy of app description; whether the app had specific, measurable, and achievable goals; quality of information; quantity of information; visual information; credibility; and whether the app was evidence-based. Furthermore, 2 additional categories, subjective quality and perceived impact, captured the rater’s overall impressions. Subjective quality included whether the rater would recommend the app (1=definitely not to 5=definitely yes); time predicted by the rater during which the app would be used in the next 12 months, if it was relevant (1=none to 5=>50 times); rater willingness to pay for the app (1=definitely not to 5=definitely yes); and the overall star rating (1=one of the worst apps I have used to 5=one of the best apps I have used). Perceived impact was also rated using a 5-point Likert scale (1=strongly disagree to 5=strongly agree) and addressed the app’s perceived potential to increase users’ awareness, knowledge, attitudes, and motivation to complete the desired health behavior, encourage help-seeking, and improve user communication.A total of 3 certified SLPs scored the quality of the included apps using the MARS . The MARAs recommended by the MARS developers, each rater viewed the training video by Stoyanov and Hides ,31. The A MARS mean score was calculated for each app in each category, and then a total MARS mean score was calculated from the first 4 categories, with 5 being the highest score possible. The final MARS score was the average score of the 2 raters for each app. The overall MARS score demonstrated a good  level ofP=.76). From a total of 2680 apps identified from Google Play Store, Apple App Store, and web searches, 70 apps met the eligibility criteria. language therapy, defined as therapy for comprehending and producing written and spoken words and sentences. Among these language apps, 19 targeted spoken language expression, 15 targeted spoken language comprehension, 13 targeted reading, and 8 targeted writing. A total of 17 apps were designed for speech therapy, defined as therapy to improve perception and production of speech sounds and speech segments. This included 10 apps for articulation , 6 for motor speech (addressing problems with motor speech planning), and 2 for speech rate. Furthermore, 6 apps were used for voice therapy, including 5 addressing vocal loudness and 3 addressing vocal pitch. Overall, 8 apps were designed for cognitive communication therapy or cognitive skills underlying communication, such as problem solving, reasoning, inferencing, and executive functions. Finally, 5 apps were designated as others, including 3 apps for numbers/numeracy and 1 app for oromotor function .The apps identified as primarily targeting adults with communication disorders after ABI were organized by therapeutic purposes . Overalltalk around me apps were rated together once, as they differed only with regard to the vocabulary items used. A summary of the MARS scores is included in Of the 70 apps, 66 (94%) were rated using the MARS tool. A total of 5 The average MARS rating was 3.7 out of 5 . Overall, 27 apps achieved ratings between 4.0 and 4.9 , 30 apps were rated between 3.0 and 3.9 , whereas 11% (7/66) apps received ratings between 2.0 and 2.9. No app received a rating of 1 (inadequate). Overall, 86% (57/66) apps were rated as acceptable (3 out of 5) or higher, indicating favorable perceptions of the apps as perceived by certified SLPs.Naming Therapy, Speech Flipbook Standard (4.6/5), Number Therapy (4.5/5), Answering Therapy, and Constant Therapy (4.4/5). The lowest-rated apps were Aphasia: Start Talking Again (2.2/5), Aphasia Speech Therapy (2.3/5), HelpMeTalk, and Speech therapy Logopedic Free (2.5/5). Across all apps identified in the review, functionality was the highest-rated subscale , followed by aesthetics , information , and engagement .The top 5 apps were Naming Therapy (4.4/5), as it had a progress bar to introduce an element of gamification, a range of interesting cues, and a number of customization options . Naming Therapy also allowed the recording and emailing of the audio recording, provided self-rating options and cues on request, and utilized a simple layout and visual icons suitable for adults with communication disorders. In contrast, the lowest-rated app regarding engagement was Aphasia Speech Therapy (1.6/5), which lacked entertaining and engaging elements, did not have a setting page or method for customizing targets, and relied on written targets with no other visual elements to support adults with communication disorders. In addition, the voice recognition function of the Aphasia Speech Therapy app did not work at the time of rating .The top-rated app on the engagement subscale was Aphasia: Start Talking Again (2.1/5), as the speech recognition function was poor and failed to recognize many accurate speech productions. Therefore, the accuracy of the recorded data was poor. In addition, the feedback for speech evaluations was very general in nature and did not provide specific feedback.A total of 13 apps received an excellent (5) rating on the functionality subscale for timely app performance, absence of technical issues, being easy and intuitive to use, employing a logical and clear layout, and consistent and intuitive gestural design. The lowest-rated app regarding functionality was Think Therapy, which had a simple layout, high-quality graphics, a very attractive color scheme and visual motifs, and seamless animations. Speech therapy Logopedic Free was the lowest-rated app on this subscale (2/5). This rating was given because of the unusual layout involving menu items distributed across the screen, activities embedded within activities, its low quality, stylistically inconsistent graphics, and garish color scheme.A total of 2 apps received an excellent (5) rating on the aesthetic subscale. An example of this was Naming Therapy (4.5/5) and Speech Flipbook Standard (4.4/5). These contained detailed information about how to use the app that incorporated visual information (images of icons and buttons). The information contained in the Naming Therapy app is linked to evidence-based techniques, for example, cueing hierarchies and semantic feature analysis [Naming Therapy, Reading Therapy, Writing Therapy, and Comprehension Therapy and one trial of Constant Therapy [Constant Therapy was the only app to describe the use of a big data approach to collect patient data to personalize therapy and optimize outcomes based on what worked for patients with similar demographics and diagnoses [Aphasia: Start Talking Again, HelpMeTalk, and Cognitive Rehabilitation 1-3 were rated lowest on this subscale because of poor description of purpose and goals.The top 2 apps on the information subscale were analysis -36. Only Therapy ,37. It iiagnoses . In contNumber Therapy, received an excellent subjective quality rating. Other highly rated apps were Apraxia Therapy, Category Therapy, Comprehension Therapy, Naming Therapy, and Talk Around It (4.75/5). These apps share a combination of appealing qualities, including multiple ways of interacting with the app , clear intuitive layout, navigation and gestural design, high-quality attractive visuals, and easy ways to track progress and obtain accurate feedback.Only 1 app, Category Therapy and Cognifit–Test and Brain Games, received an excellent perceived impact rating. Other highly rated apps included Naming Therapy, Advanced Comprehension Therapy, Advanced Naming Therapy, Advanced Reading Therapy, and Asking Therapy (4.67/5). With regard to perceived impact, in general, apps did not receive ratings for awareness, attitudes, or help-seeking. Rather than building public awareness of communication disorders, changing user attitudes, or encouraging help-seeking, most apps were designed to assist individuals in working on specific communication skills and gaining awareness of their own strengths and weaknesses. Furthermore, apps designed to facilitate repetitive practice of specific communication skills appeared best suited for use as part of a more holistic therapy program directed by an SLP.Only 2 apps, perceived impact generally functioned well, accurately rewarded target performance, provided multiple cues/prompts, and offered multiple options for presentation and response modalities. For example, while using Advanced Comprehension Therapy, the user can select auditory stimuli, written stimuli, or both; request hints; or request that the stimulus be repeated slowly. In contrast, low-rated apps provided inaccurate feedback  or rewarded incorrect productions (Speech therapy Logopedic Free). Other low-rated apps offered features that were not functional on the device used at the time of the review .Apps that were rated highly on speech, language, voice, cognitive communication,oromotor function, and/or numeracy. A high proportion of the reviewed apps had an average MARS rating of 3.7 out of 5, which is similar to that from previous research by Furlong et al [This study was designed to identify speech-language therapy mobile apps available in English for adults with communication disorders and evaluate their content and quality using the MARS tool . A totalng et al  who reviNaming Therapy, Number Therapy, and Apraxia Therapy). Despite these claims, only 2 published studies were found that evaluated the clinical effectiveness of these apps. A small pilot, crossover design study evaluated the use of 4 Tactus Therapy apps  in patients with chronic expressive aphasia. This study found small but significant improvements in language outcomes as measured by a standardized aphasia battery and a narrative discourse measure. However, the number of participants who completed the study was small (n=10), and participants differed significantly in terms of aphasia severity at baseline [Constant Therapy, was trialed to ascertain its effects on specific therapy tasks and overall language and cognitive skills. Both the experimental group (n=42) and the control group (n=9) received individual face-to-face clinic sessions once a week for 10 weeks, which involved clinician-assisted delivery of Constant Therapy tasks. The experimental group was also asked to use Constant Therapy to practice language activities at home, whereas the control group did not do home practice. Small but significant improvements on a standardized aphasia battery were reported in both participant groups, with more significant gains evident in the experimental group. However, as the experimental group spent more time on therapy tasks, it is not clear whether the improvement was the result of more opportunities to use Constant Therapy or simply the result of more time spent practicing language. In addition, as the participant groups were not matched for severity level at baseline or with regard to time after onset of stroke/TBI, the results may have been confounded. Typically, a degree of improvement over time is expected, especially for individuals still within the first 12 months after onset. Therefore, none of the apps identified in this review had high-level evidence of clinical effectiveness. These findings have important implications for further research with a focus on evaluating the clinical effectiveness of mobile apps. In particular, with the recent worldwide COVID-19 pandemic, it is crucial to offer health care support remotely via digital health technologies [This review revealed a lack of clear evidence to demonstrate the clinical benefits of speech-language therapy mobile apps currently available in Google Play Store or Apple App Store for adults with communication disorders. At instances where claims were made for clinical effectiveness, there was a lack of high-quality clinical trials to support these assertions. Overall, 3 of the top 10 apps claimed to be designed and developed based on evidence-based therapy techniques  over aesthetics , information  and engagement . This was surprising given the long-term engagement in speech-language therapy commonly required to gain real health benefits . The ideproaches ,41. Althproaches -44, this factors  to achie factors , which c factors .A multidisciplinary approach should be taken from the early stages of speech-language therapy app design and development and should involve SLPs, human-computer interaction researchers, user experience researchers, developers, and individuals with communication disorders. This approach would ensure the consideration of both human-centered design ,48 and eThis review identified limited gamification elements  in the included apps. Further research should be undertaken to investigate other game design elements to enhance engagement with rehabilitation technologies by adults with communication disorders. In particular, attention must be paid to inclusive and accessible game design for people with different communication needs and to the inclusion of game elements that are relevant for users of different ages , experience with technology, cultures, languages, and regional dialects. Future research should also include non-English apps that could be used for bilingual/multilingual patients who should have the opportunity to be treated in their primary or preferred language, irrespective of its popularity. Finally, this review found a lack of relationship between the MARS rating and the in-app consumer star ratings. Similar findings were reported in the study by Knitza et al , where MThe findings of this study have number of recommendations and practical implications.More research using a co-design process involving all stakeholders  who will use or prescribe the end product is essential for developing apps targeting specific speech-language and communication impairments.It is essential to tailor apps to suit targeted patient group profiles by incorporating the capabilities to customize or adjust the content of the apps to simplify or increase complexity to match patient skill levels. For example, incorporation of gamification elements may be a priority to engage young patients with TBI, whereas for older patients affected by stroke, it may be more important to present age-appropriate content in an aphasia-friendly way through the use of visually distinct and appealing pictorial information and simplified language/text and formatting .Subject matter expert ratings for mobile health apps must be included to provide a more reliable measure of the app’s quality, which could include ratings of engagement, functionality, aesthetics, information, and subjective quality.Owing to the increasing number of new app releases in Google Play Store and Apple App Store globally, it is recommended that professional bodies such as Speech-Language Pathology associations establish a database where app developers could register their apps. This could assist with the long-term management of mobile apps and support SLPs in decision making to recommend particular apps to their patients.A limitation of this review is that details regarding the number of app downloads and demographics of current users were not available, which would have been beneficial for in-depth analysis. Another limitation of this review is that the content and quality of the apps were evaluated solely from the perspective of SLPs and did not include consumers. However, most of these apps were designed to deliver speech-language therapy, where the guidance of SLPs would be instrumental in influencing user uptake. Further research should evaluate the usability of speech-language therapy app and the level of adoption by the target population. In addition, this paper did not review experimental apps currently reported only on journal articles, conference proceedings, or developer websites. Although such apps are not widely available, they may have more theory-driven designs and could show more promise of efficacy for use by adults with communication disorders. Future research focusing on these experimental mobile apps may be worthwhile.The results of this study revealed a limited evidence base for speech-language therapy apps and a lack of highly engaging elements such as gamification techniques. Therefore, there is a need for apps to be developed to complement traditional speech-language therapy, utilizing interactive and engaging design elements to enhance user experience and optimize sustainable technology uptake. Furthermore, from the early stages, apps should be designed and developed by a multidisciplinary team of experts, including speech pathologists, human-computer interaction experts, user experience designers, and app developers to ensure a balance between clinically suitable content and positive user experience. Future studies should also ensure the design and development of speech-language therapy apps using patient-led co-design principles by involving adults with communication disorders as codevelopers. If this is done, mobile apps can have the potential to positively enhance the effectiveness and reach of long-term speech-language therapy for adults with communication disorders. Finally, adequately powered randomized controlled trials are needed to assess the efficacy of apps developed for long-term use in the management of adult communication disorders.


Students have been highly vulnerable to mental health issues during the COVID-19 pandemic, and researchers have shown that perceived stress and mental health problems have increased during the pandemic. The aim of this study was to reveal the prevalence of perceived stress and mental health among students during the pandemic and to explore predictors of stress levels. A cross-sectional study was conducted on a sample of 358 undergraduates from 14 universities in Turkey, including 200 female students (56%). The measurements used in the study were the Generalized Anxiety Disorder 7-item (GAD-7) scale, Patient Health Questionnaire (PHQ-8), Satisfaction with Life Scale (SWLS), Perception of COVID Impact on Student Well-Being (CI), Perceived Stress Scale (PSS-10), Physical Activity Scale (PA), and a sociodemographic survey. Students reported high perceived stress, mild generalized anxiety, and low satisfaction with life. More than half of the students met the diagnostic criteria of GAD (52%) and depression (63%). Female and physically inactive students had higher PSS-10 levels. A hierarchical linear regression model showed that after controlling for gender and negative CI, anxiety and physical inactivity significantly predicted high perceived stress. The study shows that students’ mental health during the pandemic is at high risk.  The coronavirus disease COVID-19) pandemic is a global experience that is unique in modern world history. Since it affects widely disparate areas of life on the social and individual levels, it is classified as an existential experience 9 pandemi,3,4,5,6.Psychological stress emerges from an imbalance between an individual’s perception and external demands. Perceived stress refers to the assessment of the degree to which the situation in one’s life is seen as stressful; therefore, it is related to the subjective assessment of life events . It is rPublic mental health during COVID-19 deserves special attention; based on previous experiences, pandemic outbreaks are related to generalized fear that can impede infection control and lead to depression, anxiety, and post-traumatic stress . PreviouA national study among Chinese people showed that 35% of participants experienced psychological distress during the COVID-19 pandemic. Psychological distress was higher in women, people between 18 and 30 years or over 60 years of age, and those with higher education . Higher Students are particularly vulnerable to mental health problems, as a study before the COVID-19 pandemic showed . Among vThe impact of the COVID-19 pandemic on mental health and education is expected to be highly noticeable . A compaThe prevalence of stress during the COVID-19 outbreak among first-year French students was 61.6% , while iPhysical activity (PA) is strongly related to mental ,47; therThere are several studies on mental health during the pandemic among Turkish , medicalConsidering the novelty of the coronavirus pandemic, it is still unclear how factors related to COVID-19 (perception of its impact on students’ well-being), sociodemographic variables, physical activity, and mental health (anxiety and depression) are associated with perceived stress levels in students. Therefore, the aims of this study are to reveal the prevalence of mental health and perceived stress levels and to explore perceived stress predictors in Turkish university students. Considering previous research showing the role of sociodemographic variables such as place of residence and gender, year of study, physical activity, satisfaction with life, and mental health indicators such as anxiety and depression symptoms, we examined those variables as predictors of perceived stress among Turkish students during the COVID-19 pandemic.A cross-sectional study was conducted online between 16 May and 10 June 2020. The first confirmed case of COVID-19 in Turkey was reported on 10 March 2020, and the first COVID-19-related death in the country was announced on 17 March. The number of daily coronavirus cases was 5000–5100 per day, reaching a peak between 11–21 April. From the beginning of June the number decreased to 1000–1500 cases per day and stayed stable until October 2020 . After tThe survey was created by means of Google Forms. The invitation to the online study was sent to students by researchers via email and social media. For 95% of the sample, academics at three main universities sent invitations to students . Within this sample across the three main universities, the invitation was send by academics to about 4500, 410, and 150 students, respectively. The remaining 5% of the sample was collected by both, academics and students, and the invitation was shared with 8000 students; nevertheless, only 19 students responded. The survey was usually completed within 20 min. The study adhered to ethical requirements pertaining to the anonymity and voluntariness of participation. An informed consent question was added to the survey, and each participant consented to participate in the study. Initially, 361 persons responded to the invitation. However, ultimately, three respondents did not agree to participate in the study. Sampling was purposive. The selection criterion was being a Turkish student. To assess the sample size, we used the sampling technique presented by . The calThe study encompassed 358 students from 13 Turkish universities located in 10 cities: Bingol University, Bingöl ; Atatürk University, Erzurum ; Muğla Sıtkı Koçman University, Muğla ; Başkent University, , Beykoz University , Boğaziçi University , and Istanbul University , İstanbul; Ağrı İbrahim Çeçen University, Ağrı ; Adnan Menderes University, Aydın ; Fırat University, Elazığ ; Akdeniz University, Antalya ; Dicle University, Diyarbakır ; and Kırıkkale University, Kırıkkale . Predominantly, students represented the bachelor level of studies, and the most frequent fields of study represented were social sciences and health sciences. Nearly 70% of participants lived in a city.Women constituted 56% of the sample (n = 200). The participants’ age ranged from 19 to 40 years, and the average age was 23. Detailed descriptive statistics on gender, place of residence, level of study, field of study, year of study, and type of study are presented in To measure whether the respondents’ appraised the situation in their life as stressful, the Perceived Stress Scale (PSS-10)  in the TThe 7-item generalized anxiety disorder (GAD-7) scale  in the TThe Patient Health Questionnaire (PHQ-8) was used to measure depression symptoms among Turkish students. The PHQ-8 consists of eight items, conforming with DSM-IV diagnostic criteria . ParticiPerception of COVID-19 impact (PCI) on well-being  was measExposure to COVID-19  was measPhysical activity (PA) during the coronavirus-related lockdown  was assePA level was divided into two groups: inactive (less than 150 min weekly) and active (150 or more minutes weekly) in relation to World Health Organization (WHO)  recommenp < 0.05), a further analysis of distribution based on skewness and kurtosis coefficients indicated sufficient symmetry and similarity to the Gaussian curve since the absolute values of the obtained coefficients did not exceed 1, which indicates good psychometric properties [The study used SPSS.25  software. The analysis encompassed Pearson’s r coefficient correlations, one-way analysis of variance (ANOVA) to evaluate significance of differences, and hierarchical multiple linear regression. In addition, descriptive statistics of the surveyed variables were presented. Although the analysis did not prove a normal distribution of variables  was mild, and depression symptoms (PHQ) were moderate, while satisfaction with life among Turkish students during the pandemic was low. Even though average anxiety in the sample was mild, over half of the students presented symptoms according to the GAD (52%) and PHQ (62%). One-third of students met neither depression nor anxiety diagnostic criteria, whereas nearly half were characterized by a dual diagnosis (45%). Almost every fifth university student (23%) met the criteria for at least one diagnosis (anxiety or depression).Perception of the negative impact of COVID-19 on students’ well-being was the most pronounced regarding their financial situation, job search, and completion of the semester. The least negative impact was perceived in the area of relationships with loved ones, family, and friends. When analyzing results for the perception of the impact of COVID-19 on well-being as a five-item scale, it can be noted that the grand mean was 3.75. It can be interpreted that, in general, students perceived the impact of the pandemic as negative. Before the pandemic, students were physically active for 189 min weekly, and during the pandemic for 75 min weekly. Referring to the variables related to the COVID-19 pandemic, the percentages of physical activity (PA) before and during the pandemic and exposure to the virus were analyzed. Before the pandemic, 38% of students were physically active and met the WHO criterion (over 150 min per week), while during the pandemic only 13% remained active. The effect of exposure to COVID-19 was the strongest in the aspect of declining economic status (65%) and experiencing job loss . Although 6% of students experienced symptoms of coronavirus infection, only one student was hospitalized. Of the 12% of students tested for COVID-19, 5% were in a strict 14-day quarantine. Every fifth student (22%) experienced COVID-19 infection among close relatives. There were 19 cases of coronavirus-related deaths among students’ families.Referring to psychological variables, only 13% of students were characterized by a normal level of generalized anxiety disorder (GAD), whereas over half of them (52%) presented clinical symptoms of GAD. Low satisfaction with life was perceived by 56% of students, and high satisfaction by only 19%. A majority of students (71%) reported high levels of perceived stress, while only 6% reported low levels. The results are presented in Pearson’s r coefficient was applied in order to verify correlations among variables . The corp > 0.005), whereas gender and PA were significant. For statistical purposes, six participants who did not want to describe themselves as male or female were excluded from the analysis. Gender turned out to significantly diversify PSS: F(1350) = 15.79, p < 0.001. Female students were characterized by significantly higher PSS levels  compared to male students , although the effect size was small; Cohen’s d = 0.43. PA significantly diversified PSS: F(1356) = 4.04, p = 0.045. Inactive students had significantly higher PSS levels  compared to active students . The effect size was medium; Cohen’s d = 0.55.One-way ANOVA was introduced to assess the significance of differences in perceived stress in relation to gender, place of residence, year of study, and physical activity (PA) during the pandemic. Place of residence and year of study turned out to be insignificant  = 13.85, p < 0.001). Being a physically inactive female student explained 7% of the variance. The perception of a negative impact of COVID-19 on students’ well-being was introduced to model 3. The additional variable positively predicted PSS. Model 3 was statistically significant (F(3348) = 13.28, p < 0.001) and explained 9% of the variance. Psychological variables satisfaction with life and generalized anxiety disorder were introduced as predictors to model 4, which also turned out to be significant (F(5346) = 33.81, p < 0.001) and explained 32% of the variance. Nevertheless, only two predictors remained statistically significant; high generalized anxiety disorder and being physically inactive during the pandemic explained 32% of the variance. Therefore, after controlling for gender and the perception of a negative impact of COVID-19, mostly generalized anxiety disorder, along with physical inactivity, predicted high perceived stress. The results are shown in detail in For the first block analysis, the predictor variable gender was analyzed. The results of the hierarchical analysis of model 1 turned out to be statistically significant: F(1350) = 15.79, Our study evidenced that students reported high perceived stress, mild generalized anxiety, and low satisfaction with life. More than half of the students met the diagnostic criteria of generalized anxiety disorder (52%) and depression (63%). Female and physically inactive students had higher perceived stress levels. A hierarchical linear regression model showed that after controlling for gender and negative COVID-19 impact on well-being, anxiety and physical inactivity significantly predicted high perceived stress. The study shows that students’ mental health during the pandemic is at high risk. In this study, students were characterized in general by high perceived stress, mild anxiety, and moderate depression, with 5.6% reporting low perceived stress, 23% medium, and 71.2% high. The results differed among Indian students, i.e., 4%, 82.7%, and 13.3%, respectively ; by compCompared to stress levels of Turkish students before the pandemic , currentTurkish students had mild anxiety and moderate depression symptoms. Among those with anxiety, 35.7% reported mild symptoms, 28.8% moderate, and 22.9% severe; 12.6% of students had no anxiety symptoms. Among those with depression, 27.7% reported mild symptoms, 24% moderate, 23.7% moderately severe, and 15.1% severe symptoms; 9.5% of students had no depression symptoms. Depression symptoms were more frequent than anxiety symptoms (62.8% vs. 51.6%). One-third of students (31%) did not meet any diagnostic criteria for anxiety or depression, but almost half (45.5%) had a dual diagnosis. Therefore, the prevalence of comorbid depression encompasses a large number of Turkish university students, although there were more students with severe anxiety (23%) than severe depression (15%). Previous research evidenced higher anxiety in students during the pandemic , althougDepression levels were moderate in Turkish students, whereas severe depression symptoms were reported by 15%, which is in agreement with a Chinese student sample (16%) . On the These differences might be due to the different measurement periods during the COVID-19 pandemic. Therefore, although high anxiety and depression are related to the pandemic in general, cross-cultural differences should be taken into account when explaining perceived stress in student populations. On the other hand, the results are in agreement with other research revealing higher stress, anxiety, and depression in students compared to the general population .Direct exposure to COVID-19 was relatively low among Turkish students in terms of hospitalization . At the same time, every fifth student experienced COVID-19 infection among family or friends, and 5.3% experienced the death of close relatives. By the end of data collection (10 June) there were 4747 deaths due to COVID-19, and while the curve of new cases was decreasing, the number of total deaths was rising. Therefore, it would seem that the student sample was overexposed to the bereavement experience. Nevertheless, there were concerns regarding the openness and clarity of COVID-19 data in Turkey, as it seemed that the prevalence of the disease  might be underreported by governments of other countries ,71.Turkish students were most affected by the COVID-19 pandemic in the financial aspect. Among the students, 65% noted a deterioration of their economic status, and half of them experienced losing a job (themselves or among family members). This means that the coronavirus pandemic affects healthy people mostly by way of economic consequences. The beginning of the pandemic brought high uncertainty among students regarding academic life  .Even though such problems were related to the abrupt introduction of remote learning and the uncertainty arising at universities seemed to be the strongest source of stress in students , it turnWe have shown that perceived stress relates positively with a negative perception of COVID-19 impact (PCI) on students’ well-being in the areas of education, economy, and relationships. Perceived stress was strongly linked to generalized anxiety disorder and negatively with satisfaction with life. Place of residence and year of study turned out to be insignificant for stress level differentiation. Previous research showed an inconsistent but significant association between living in a rural or urban area and mental health ,45 but nOur findings confirm the importance of physical activity for decreasing perceived stress levels . The stuThe study revealed significant differences in stress levels due to gender. In relation to other studies, female students reported significantly higher stress levels compared to male students ,74,75,76We have shown that being female, being physically inactive, and having a negative perception of the impact of COVID-19 on students’ well-being are significant predictors of perceived stress in students. However, after introducing satisfaction with life and anxiety to the model, gender turned out to be an insignificant predictor. Additionally, satisfaction with life turned out to be insignificant with regard to stress levels during the pandemic. It might be due to the fact that the Satisfaction with Life Scale (SWLS) is a generalized cognitive assessment of subjective well-being that is quite stable throughout the course of life ,78. TherThe perception of the COVID-19 impact (PCI) on students’ well-being turned out to be a significant predictor of perceived stress in the model of regression together with gender and physical activity. In this model (without anxiety and satisfaction with life), negative PCI, female gender, and physical inactivity predicted high stress levels in students. The PCI components were related to problems with graduation and completion of the semester, job search, financial situation, and concerns about relationships with family and friends. Therefore, unless anxiety is disregarded, a negative perception of the impact of COVID-19 in those areas of students’ lives strongly predicts higher stress levels.The pandemic situation affects students’ well-being, as the results of a negative PCI (including concerns about academic future and completing the semester) show. Considering a strong relationship between perceived stress, mental health, and problems with academic life with burnout syndrome ,80,81,82Future research should focus on comparing student populations to other young adults and adults. The second stage of this study will be carried out in October–November 2020 to determine the longitudinal effects of COVID-19 on mental health and well-being. As most of the pandemic research relates to negative consequences, an analysis of positive outcomes will be introduced in the second wave of the study. There are several limitations to this study. One is its cross-sectional character. A longitudinal study design would have enabled establishing the impact of the COVID-19 pandemic on perceived stress and the direct cause-and-effect relationship between examined variables and perceived stress. Also, the lack of random sampling and the representation of a student population limited to eastern Turkey seem to be a burden in generalizing the results. Using globally validated standardized tools allowed for discussion with regard to perceived stress and mental health. Nevertheless, the discussion and literature review showed great differences in student populations between countries, and an international study design could help to explain those differences and understand students’ perceived stress levels during the COVID-19 pandemic. In addition, we did not control the number of students with a previous clinical diagnosis of generalized anxiety disorder. Controlling this variable could prove the effect of COVID-19 on anxiety and perceived stress, so as to fully reflect the severity of depression and anxiety symptoms among students.The biggest limitation to this study is very low response rate (2.7%). The response rates were 3.4% (151/4500) at Bingöl University, 37.3% (153/410) at Atatürk University, 23.3% (35/150) at Muğla Sıtkı Koçman University, and 0.2% (19/8000) at all other 10 universities. The highest response rate at Atatürk University could be due to the form of dissemination of the invitation to the study participation. It was spread directly via the university’s online platform, i.e., a highly formal manner. That could cause a strong feeling of obligation to fulfill the survey by students. The lower response rate in other cases could be due to the use of many communication procedures  such that they could have missed the survey or were not willing to fill it out. Nevertheless, research with low response rates may generate prevalence estimates that are biased by selective non-response ; therefoinfodemic (an overabundance of information) and misinformation during the pandemic can help to improve the mental health of students.The state of perceived stress, mental health, and well-being among Turkish students is alarming and requires systemic and dedicated action. Universities together with authorities should collaborate to create an efficient system, of both psychological and financial help for students. Considering the high number of students experiencing a deterioration of economic status, special programs dedicated to financial support during the pandemic should emerge, i.e., scholarships or student loans. Also, fighting an To decrease stress levels, online psychological counseling services should be provided. Even though female students presented higher anxiety and stress levels, decreasing anxiety levels and boosting physical activity for all students seem critical. Providing mental health support systems for students and promoting physical activity on a regular basis could decrease perceived stress levels.