Background: Patients with heart failure (HF) often display dyspnea associated with pulmonary congestion, along with intravascular congestion, both may result in urgent hospitalization and subsequent death. A combination of radiographic pulmonary congestion and plasma volume might screen patients with a high risk of in-hospital mortality in the emergency department (ED).Methods: In the pathway of dyspneic patients in emergency (PARADISE) cohort, patients admitted for acute HF were stratified into 4 groups based on high or low congestion score index  and estimated plasma volume status (ePVS) calculated from hemoglobin/hematocrit.Results: In a total of 252 patients , CSI and ePVS were not correlated . High CSI/high ePVS was associated with poorer renal function, but clinical congestion markers  were comparable across CSI/ePVS categories. High CSI/high ePVS was associated with a four-fold higher risk of in-hospital mortality  compared with low CSI/low ePVS, whereas neither high CSI nor ePVS alone was associated with poor prognosis . High CSI/high ePVS improved a routine risk model , resulting in high prediction of risk of in-hospital mortality .Conclusion: In patients hospitalized for acute HF with relatively old age and comorbidity burdens, a combination of CSI and ePVS was associated with a risk of in-hospital death, and improved prognostic performance on top of a conventional risk model. Congestion in heart failure (HF) is defined as a high left ventricular filling pressure associated with signs and symptoms of HF such as dyspnea and edema –3. PatieThe congestion score index (CSI) is a semi-quantitative method for assessing pulmonary congestion on chest radiography (CXR) –11. PlasThe main objectives of the present study are to (i) assess the interplay between CSI and ePVS in patients admitted for AHF in the ED and (ii) explore the association of CSI and/or ePVS with in-hospital mortality.The Pathway of dyspneic patients in Emergency (PARADISE) cohort is a retrospective cohort study including consecutive acute dyspneic patients aged 18 years or older who admitted for the ED of the Nancy University Hospital (France) between January 1, 2015 and December 31, 2015 as previously reported , 17, 18.Clinicaltrials.gov” (NCT02800122).Among 1,589 patients included in the PARADISE cohort, 252 AHF patients with available data on CXR and hemoglobin/hematocrit were studied. Those with length of hospital stay > 30 days were excluded since they likely had either atypical degrees of congestion or non-congestion related problem . DemograCSI is assessed to quantify the severity of pulmonary congestion on CXR as previously described , 10. AftCXR was analyzed by a single emergency physician (AD), blinded to clinical data and outcome, with no prior training in CSI. After a short training (~3 h) using a 20-patient sample with a radiographic CSI expert (MK), when blinded to clinical data and outcome, intra-class correlation coefficients showed good reproducibility .ePVS was calculated from the Strauss-derived Duarte formula using hematocrit and hemoglobin , 20 as f2 test, as appropriate. Inter-observer and intra-observer agreements of CSI were assessed with intra-class correlation coefficients.Patients were divided into four groups according to sex-specific median ePVS  and median CSI (2.20); low CSI/low ePVS, high CSI/low ePVS, low CSI/high ePVS, and high CSI/high ePVS. Categorical variables are presented as frequencies (percentages) and continuous variables as means ± standard deviation or median (25th and 75th percentiles), according to variable distribution. Comparisons of demographic, clinical and biological parameters across CSI/ePVS categories were analyzed using variances, Kruskal-Wallis test and χLogistic regression analysis was used to study the associations of CSI/ePVS categories with in-hospital mortality. The relevant covariates as previously shown were followings , 9: age,Furthermore, net reclassification improvement (NRI) was used to assess the increase in discriminative value of the addition of high CSI/high ePVS on the top of the aforementioned covariates including clinical model  plus biological model . The continuous NRI method was implemented in the survIDINRI package of the R software .p-value < 0.05 was considered significant. No imputation was performed. Statistical analyses were performed using the R statistical software (3.4.0).A 2 and 22.2% had a prior admission for HF. Median BNP was 516  pg/ml (p = 0.44).Approximately a half of the 252 included patients were male (46.8%), mean age was 81.9 ± 9.8 years, mean body mass index was 26.7 ± 6.4 kg/m1) pg/ml . Median When considering subsets of patients divided according to the median of CSI and ePVS , high CSN = 45). By univariable logistic models, neither high admission CSI nor ePVS was associated with a higher risk of in-hospital mortality   .p-value for interaction = 0.03). The association of CSI/ePVS-derived subsets with outcome was consequently investigated. In univariable analysis, patients with high CSI/high ePVS were associated with a higher risk of in-hospital mortality [OR [95%CI] = 2.60 [1.10-6.51], p = 0.03] compared with those with low CSI/low ePVS. After adjustment for potential clinical covariates including BNP and lactate, patients with high CSI/high ePVS still had a four-fold increase in the risk of in-hospital mortality   . In contp-value for interaction > 0.05; Similarly, the prognostic value of high CSI/ high ePVS was consistent across subgroups of age, sex and comorbidities  or associated diagnosis  or quality of chest radiogram (p = 0.07), but not before 7 days   .p < 0.001) (p = 0.02). The combination of clinical, biological models and high ePVS/high CSI resulted in a high level of prediction of the risk of in-hospital mortality (AUC [95%CI] = 0.85 [0.82-0.89]) ; the addition of biological variables on top of the clinical model significantly improved reclassification  . Importa2-0.89]) .The main findings of the present study are that (1) patients with both high CSI and high ePVS had similar clinical congestion and BNP concentrations than other categories of CSI/ePVS; (2) these patients had a four-fold increased risk of in-hospital mortality, whereas patients with either high CSI or high ePVS alone did not display a poorer prognosis compared with those with low CSI/low ePVS and; (3) the addition of high levels of CSI and ePVS on top of the clinical plus biological prediction models  was associated with a significant improvement in reclassification for in-hospital mortality. Our results highlight that radiographic pulmonary congestion scoring and PV (estimated from hemoglobin/hematocrit) further refined patient prognosis on top of well-known confounders , thus helping emergency physicians in triaging high-risk acute heart failure patients admitted in the emergency department.This study included patients with relatively old age (mean age 82 years) and/or a comorbidity burden , which may partly explain a high rate of in-hospital death when compared with other cohorts –25. PatiOur results showed that only the simultaneous presence of pulmonary and intravascular congestion (namely high CSI/high ePVS) conferred a higher risk of in-hospital mortality, whereas patients with only one severe congestion (pulmonary congestion or intravascular congestion) marker had a comparable risk of adverse outcome when compared with those without. In previously published data, a uniform definition of congestion not acknowledging the distinction between pulmonary and intravascular congestion provided a limited predictive value of congestion for in-hospital mortality , 7. Our The management of AHF in the ED limits available tools of assessing the severity of congestion. Several prognostic markers  are widely used for triage assessment, allowing a determinant of decision-making in patients admitted for AHF in the ED , 32. HowMore than 80% of AHF patients expressing dyspnea initially present to the ED , 34. Curp-value of 0.07 for high risk of outcome. Further research with a larger sample size is needed. Second, CSI is a semi-quantitative tool with some subjective assessment. However, in our experience, a short training period of ~3 h is sufficient to achieve accurate and reproducible scoring. The prognostic value of CSI/ePVS, in addition, was not significantly influenced by the quality of the chest radiographic images, suggesting that our findings may apply to a broad spectrum of clinical settings. Third, chest radiogram was generally assessed as soon as possible after ED admission, but the exact time at which chest radiogram was performed is lacking. We, thus, cannot ascertain that initial therapeutic interventions  may not impact CSI evaluation, although this would be the case in routine ED practice in most centers. Fourth, in addition to lack of direct measured PV, there is no accepted normal value of ePVS; however, PV estimated from hemoglobin/hematocrit had a good correlation with measured PV in a recent report  with a high level of comorbidity burdens, leading to limited statistical power and possible residual confounding. Particularly, prognostic value of ePVS/CSI interplay was corroborated by results in logistic regression model, but not in time-to-event analyses in which high CSI/ high ePVS subgroup had a nominal t report , and ePVt report , 14, 38.t report . Howevert report , 39.In relatively old (mean age 82 years) patients hospitalized for acute HF with a high level of comorbidity burdens, pulmonary/intravascular congestion scoring categories as assessed by CXR and ePVS had similar severity of congestion assessed by physical examination/natriuretic peptides. However, high CSI and high ePVS identified patients at a high-risk of in-hospital death, improving patient-risk stratification beyond clinical plus biological prognostic models. These results suggest that this readily and widely available congestion assessment, integrating chest radiography and hemoglobin/hematocrit, may represent a valuable triage tool for patients admitted for AHF with dyspnea in the ED.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by CNIL (Number R2016-08). Written informed consent for participation was not required for this study in accordance with the national legislation and the institutional requirements.MK and AD drafted the manuscript. NG supervised the manuscript. All authors edited the paper and/or provided constructive comments.MK was granted by the RHU Fight-HF, a public grant overseen by the French National Research Agency (ANR) as part of the second “Investissements d'Avenir” program (ANR-15RHUS-0004). MK, PR, KD, NG, and FZ are supported by the RHU Fight-HF, a public grant overseen by the French National Research Agency (ANR) as part of the second “Investissements d'Avenir” program (ANR-15-RHUS-0004) and by the FrenchPIA project “Lorraine Universite' d'Excellence” (ANR-15-IDEX-04-LUE).NG receives honoraria from Novartis and Boehringer. TC receives fees from Novartis for scientific board. FZ and PR are the cofounders of CardioRenal. FZ reports personal fees from Boehringer Ingelheim, Janssen, Novartis, Boston Scientific, Amgen, CVRx, AstraZeneca, Vifor Fresenius, Cardior, Cereno pharmaceutical, Applied Therapeutics, Merck, Bayer and Cellprothera, and is a founder of Cardiovascular Clinical Trialists. PR reports grants and personal fees from AstraZeneca, Bayer, CVRx, Fresenius, and Novartis, personal fees from Grunenthal, Servier, Stealth Peptides, Vifor Fresenius Medical Care Renal Pharma, Idorsia, NovoNordisk, Ablative Solutions, G3P, Corvidia and Relypsa. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


The use of plant ash as a sustainable cementitious material in concrete composition is a widely researched subject in the construction domain. A plant studied so far more for its thermal insulation properties, sunflower, was analyzed in this study with regard to its ash effects on the concrete composition. The present research aimed to analyze the effects of a 2.5%, 5%, 7.5%, 10%, 15%, 20%, or 30% volume replacement of cement by sunflower stalk ash (SA), a sustainable cementitious material, on the concrete compressive strength at 28 days and three months, the flexural and splitting tensile strengths, the resistance to repeated freeze–thaw cycles, and the resistance to chemical attack of hydrochloric acid. The elementary chemical composition of the SA and the composites was included also. According to the experimental results, SA decreased the values of the compressive and tensile strength of the concrete, but it improved the concrete behavior under repeated freeze–thaw cycles and under the action of hydrochloric acid. A percent of 10% of SA led to a much more pronounced development of compressive strength over time than conventional concrete (26.6% versus 12%). Yearly, 3.0–3.6 Gt of cement is produced, leading to around 3.24 billion tons of CO2  [Sustainability became an important preoccupation of the twenty-first century in all industries, including the building materials industry. The building industry is the third largest COissions) . These kThe development of concrete compositions based on renewable raw materials is a measure that diminishes the negative effects of conventional concrete made with Portland cement on the environment. Regarding the use of plant ashes in cement-based composites, various studies have been conducted, a few examples being rice husk ash ,8,9, sug6 ha, with a yield of 2.07 tons/ha [A plant of interest for the building materials sector, widely spread across the world, is the sunflower, studied until now mostly as raw material for developing sustainable materials with insulation characteristics ,20,21,22 tons/ha . It repr tons/ha . Sunflow tons/ha . It does tons/ha .Sunflower stalks can be a raw material for the construction industry, being an annually renewable resource, cheap, and widely available. To the best of our knowledge, in the scientific literature, there are very few studies on the use of ash from sunflower stalks to make cement-based composites. These studies have revealed that this sustainable raw material has pozzolanic qualities, leading to improved concrete durability and good resistance to sodium sulfate. Aksoğan et al.  studied Through this study, we wanted to make new concrete compositions with improved performance compared to those of classic concrete with Portland cement, but to be adapted to the new requirements related to energy efficiency in their production process or their durability. The aim of this research was to develop several variants of environmentally friendly concrete, by partially replacing cement with an easily renewable raw material, namely, sunflower stalk ash. Analyses performed on the developed cement-based compositions included the elementary chemical composition of SA and of the composites, mechanical tests  and durability tests for the action of repeated freeze–thaw cycles and the action of hydrochloric acid.-A reference composition of microconcrete, RC, with cement, sand, and river gravel aggregates up to 8 mm in diameter; the water/cement (w/c) ratio used was 0.5.-Seven cement-based composite mixes with SA as a partial replacement material for cement, in volume proportions of 2.5%, 5%, 7.5%, 10%, 15%, 20%, and 30%, the notations applied being CSA2.5, CSA5.0, CSA7.5, CSA10, CSA15, CSA20, and CSA30. The w/c ratio applied was increased every 10% step of used SA: In the CSA2.5, CSA5.0, CSA7.5, and CSA10 mixes, a 0.5 w/c ratio was used; in CSA15 and CSA20, a 0.518 w/c ratio was used; and in CSA30, a 0.536 w/c ratio was used. These w/c ratio differences were applied in order to maintain the same workability level for the fresh composite material.The analyzed compositions in this study were the following:3; sand (sort 0–4 mm), 673 kg/m3; river gravel (sort 4–8 mm), 432 kg/m3; water, 2365 L superplasticizer additive based on polycarboxylateter, 2% of binder.The reference composition, RC, was realized by: Portland cement CEM II/A-LL42.5R type , 473 kg/m▪CEM II: Portland-composite cement.▪3) of the total mass of calcium oxides, clay a maximum of 1.20 g/100 g of limestone, and a maximum of 0.20% of total organic carbon of the total mass of limestone.A-LL: Portland limestone cement that contains 80–94% clinker of the total mass, 6–20% limestone, and 0–5% additional constituents; limestone consists of a maximum of 75% calcium carbonate   studied According to Bahuradeen et al. (2015) , the pozThe ash that passed through the 300 µm (0.3 mm) sieve after 5 min of sieving was subjected to a grinding process for 120 min in a ball dust crusher. The obtained sunflower ash (SA)  was usedThe composition and aspect of SA was analyzed using scanning electron microscopy  coupled with an energy dispersive X-ray spectrometer detector .In this research, cement-based composites with 0%, 2.5%, 5.0%, 7.5%, 10%, 15%, 20%, and 30% sunflower stalk ash as a cement substitute were developed. The mixes were made according to NE 012/1-2007 . For the3 volume) were mechanically cut and prepared for analyses.The composition and aspect of the specimens were analyzed using scanning electron microscopy  coupled with an energy dispersive X-ray spectrometer detector . A low-vacuum mode was necessary to analyze the samples based on the fact that all of the samples gassed quite hard. Cubes samples , which leads to smaller permeability of the cement matrix and, consequently, to a better resistance to chemical attack . The C, The Si content was almost the same in the RC and CSA2.5 (20.8 ± 0.1 wt%), and slightly increased in CSA7.5 (22.12 wt%), CSA5 (24.86 wt%), CSA10 (25.22 wt%). CSA15 and CSA20 presented the highest Si content of all of the mixes: 28.48 wt% and 33.66 wt%, respectively. CSA30 had a much smaller Si content of 13.73 wt%.The C content was smaller in all CSA mixes, comparative to RC, although it was missing in CSA20 and was higher in CSA30, with 2.73 wt%, compared to RC.The Ca content was quite similar in CSA7.5 and RC, namely, 10.4 wt%. In CSA2.5 and CSA30, it was higher: 12.13 wt% and 16.32 wt%, respectively. In the other CSA mixes, it decreased up to 5.44 wt%.Al was present in the highest amount in CSA5: 4.04 wt%. In CSA10 and CSA15, the Al content decreased to 2.6–2.7 wt%, and in CSA7.5 and CSA20 to around 2 wt%. The smallest quantities of Al were found in the RC and CSA30: Around 1.1%.Regarding the Fe content, the highest level was found in CSA20 and CSA5, at 2.39 wt% and 2.02 wt%, respectively. The other mixes had smaller Fe contents, with around 50%.Na and K elements were present in small quantities (between 0.7 and 1.4 wt%) and only in a few mixes. Mg was found only in CSA30, in a very small rate of around 1 wt%.In In The compressive strength determined at 28 days is presented in If we take into account the optimum rate between the cement quantity replaced and the compressive strength value, the best result was obtained by CSA7.5 regarding the compressive strength at 28 days.In this research, the evolution of the compressive strength over time was studied, at three months after casting , given tThe SA used in the concrete composition led to a decrease in the flexural tensile strength . The repThe best results in terms of the flexural tensile strength were obtained by CSA15, if the aim is to use as much SA ash as possible.The SA determined decreases in the case of splitting tensile strength as well, but its influence in the concrete composition was not a predictable one . The obtThe composition that can be considered optimal in terms of the splitting tensile strength, taking into account the highest possible replacement rate, is CSA7.5.The SA positively influenced the concrete resistance to freeze–thaw when up to 15% of the cement volume was used . After sIn terms of the best ratio of freeze–thaw resistance versus percentage of SA used, CSA7.5 can be considered the optimal composition.Following the action of the 18% HCl solution , it coulThe composition that can be considered optimal in terms of durability to the action of hydrochloric acid versus the cement replacement rate is CSA10.The research consisted of the analysis of the influence of sunflower stalk ash as a substitute for 2.5%, 5%, 7.5%, 10%, 15%, 20%, and 30% of the cement volume in the composition of a microconcrete on the compressive strength, flexural and splitting tensile strengths, freeze–thaw resistance, and chemical resistance to a hydrochloric acid solution.As a partial replacement of cement, SA led to a decrease in the mechanical performance of concrete, and the obtained results did not follow a predictable logic, in accordance with the replacement step applied. On the contrary, SA determined the improvement of the concrete resistance to freeze–thaw.As chemical composition, CSA mixes contain, in general, supplementary chemical elements compared to RC, with the presence of potassium and magnesium being assumed to be due to the SA composition. The chlorine found in the SA composition was not detected in any of the CSA mixes.Taking into account the results obtained in all of the tests performed in this study, CSA7.5 seems to be the optimum mix.As a result, this sustainable raw material, SA, could be part of the concrete used to make prefabricated elements used outdoors, such as pavements, curbs, drains, gutters, irrigation systems , and the like. The realization of such elements with increased durability would lead to a decrease in their replacement rate due to cyclical deterioration of natural causes; therefore, it would also lead to obtaining economic gains over time. If we take into account the quantities used to make pavements for pedestrian alleys, for example, and their replacement rate, we can largely predict the economic benefit obtained, and the use of bio-based construction composites with SA could be a solution to be considered in the future.To achieve improved resistance to the attack of hydrochloric acid, it is recommended to use SA as a raw material in the composition of concrete to be used in the construction of buildings intended for production activities of dyes, organic fertilizers, galvanized sheet, leather processing, etc., as these activities involve the use of hydrochloric acid; therefore, these buildings must be protected from the action of this substance.Given that SA has reduced mechanical strength but improves durability, it is considered that research should be continued in terms of using SA as an sustainable additive in the composition of cement-based composites and not as a substitute for cement, in order to obtain a material with improved performance in terms of mechanical characteristics as well, not just as durability.


The highest antibiotic residue concentrations reported in the chicken liver were TYL, GEN, OTC, SMZ, and ENR, respectively. The lowest residual antibiotic concentrations observed in the chicken breast were TYL, GEN, OTC, SMZ, and ENR, respectively. Furthermore, the Kruskal–Wallis statistical test revealed a significant difference between the five antibiotic concentrations in both breast (H (4) = 54.69, p < 0.05) and liver (H (4) = 44.36, p < 0.05). A follow up of this finding by Bonferroni correction for both breast and liver samples revealed a significant difference for the breast sample between the concentration of ENR residue, and the concentration of residues for of both OTC and TYL (p < 0.05). These data show that not all tissues incorporate antibiotics at the same concentration. The results of this study could support regulatory bodies in adopting, monitoring, and enforcing guidelines pertinent to safety levels of different antibiotic residue concentrations in poultry meat when antibiotics are used for different indications.The aim of this study was to estimate the residue levels of five commonly used antibiotics in poultry tissue samples using enzyme linked immunosorbent assay (ELISA). A total of 200 samples that comprised breast and liver (100 each) were collected from five poultry farms randomly selected from Muscat regions. The samples were analyzed for enrofloxacin (ENR), gentamicin (GEN), oxytetracycline (OTC), sulfamethazine (SMZ), and tylosin (TYL) residue concentrations. Comparisons of antibiotic residues between breast and liver of chickens under investigations showed a significant difference of ENR, GEN, OTC, SMZ, and TYL residue concentrations ( There is growing concern over the use of antibiotics in livestock production in the Middle East countries, including Oman. Other countries such as the USA and countries in Europe have prevented such use, except for indicated treatment of diseases, considering that uncurbed use of antibiotics is a major problem in food safety. These tally with the fact that antibiotic residues are key drivers of antibiotic resistance, with consequent risk to human health. In practice, antibiotics can be used for treatment and prevention of several diseases, as well as for improving feed efficiency, promoting growth, and most important, for preventing economic losses to growers ,2,3, as Antibiotics pose a threat to consumer health, such as allergy, gastrointestinal disorders, and development of bacterial resistance. Antibiotics can penetrate and remain in an animal’s tissues, especially if the initial antibiotic dosage is high or withdrawal time is not sufficient before slaughtering ,9. ThereMacrolides  are bacteriostatic antimicrobial agents. Macrolides are metabolized in the liver, and the highest tissue concentrations for chickens and turkeys are also found in the liver . TylosinSeveral screening and confirmatory analytical methods for the detection of antibiotic residues have been developed . The immThe study was approved by the Animal Research Ethics Board at Sultan Qaboos University . All experimental work was conducted at the Poultry Research Unit at the Agricultural Experiment Station in accordance with the experimental unit policy on animal welfare, and the requirements of the procedures involving animals/birds and their care were conducted in conformity with international laws and policies  at Sultan Qaboos UniversityThe antibiotic competitive enzyme immunosorbent assay kits were purchased from Shenzhen Lvshiyuan Biotechnology Company, China. SMZ , ENR , TYL , GEN , and OTC . Each kit was specified for a different antibiotic. Two extractions were performed for each sample. Sensitivity, detection limit, and recovery rate for each kit were as follows: SMZ ; ENR ; TYL ; GEN ; OTC . Each immunoassay kit contained sufficient material for 96 measurements. Each microtiter consisted of 96 wells, coated with capture antibodies against each antibiotic.Given the frame and plan of the study and facilities available, it was not feasible at the time of collection to gather detailed information relevant to management in each farm, noting that all just aimed at marketing chicken at a certain age.Five privately owned poultry farms from the Muscat region were randomly selected for the study. A total of 100 live birds (20 birds from each farm) were collected in the study area and sacrificed by standard slaughtering process to collect a total of 200 samples, which comprised breast and liver, 100 each. Each sample was placed into a separate plastic zipper bag, numbered, and then transferred to the laboratory in an iced plastic container and stored at −20 °C until extraction. The samples were subjected later to grinding before the extraction procedure.For extraction, samples were homogenized according to the manufacture instruction, as we followed the procedure specified for each antibiotic kit  level of significance.The data were analyzed statistically using IBM SPSSThe ELISA technique was used to detect different antibiotic residue concentrations in breast and liver of broiler chickens. The quantity of antibiotic residue concentrations in broiler meat and liver samples are presented in p < 0.05; The Mann–Whitney test  was used to assess the significance of differences among broiler chicken tissues; i.e., breast and liver. A significant difference between enrofloxacin, gentamicin, oxytetracycline, sulfamethazine, and tylosin residue concentrations were observed between the breast and liver samples (H (4) = 54.69, p < 0.05) and (H (4) = 44.36, p < 0.05), respectively . The difference between tylosin residue concentration and concentrations of both gentamicin and sulfamethazine residues was significant (p < 0.05), with tylosin being lower. However, there was no significant difference between other residue antibiotic concentrations (p > 0.05)  .p < 0.05). The sulfamethazine residue concentration was also significant compared with that of gentamicin (p < 0.05). However, there was no significant difference between the other residue antibiotic concentrations (p > 0.05)  .The poultry industry is a growing sector in Oman, with several brands and companies having been launched in recent years. There are large corporate producers, but as well there are small- and medium-size farms scattered around the country, readily supplying consumers. Poultry feed constitutes a major cost component in production, and feed suppliers in the Middle East incorporate varying levels of antibiotics as supplements to enhance growth and for prophylaxis. Therefore, this study represents an overarching stage to counteract the bacterial resistance resulting from some antibiotic residues in poultry meat, and also an attempt to better align with standards pertaining to human health. It should also be noted that our study reflects the prevailing characteristics of chicken meat in the local market.To probe this, we sampled chicken for this study based on a diligent search for information on use of antibiotics by farmers in their privately owned farms. Nonetheless, the presence of antibiotics, whether in water or feed, could hardly be ascertained since farmers here just collectively acknowledged obtaining feed that was prevailing in the market, with no additional treatments throughout the growing period. This should be considered since competitiveness dictates taking into account the additional cost of antibiotics incurred by farmers in that region, should they apply antibiotics on their own. Moreover, this study is the first that paves the way into further investigation of food safety and the presence of residual antibiotics in poultry products in Oman, as well as raising awareness of the indiscriminate use of antibiotics in food producing species. Pertinently, the chickens from all farms used in the study were all about the same slaughter age (about 7 weeks of age), which is important for the assessment of the level of antibiotics that would be indicative of a threshold for safe consumption of such products, given the impact of the residual effect of antibiotics on the emergence of antibiotic-resistant bacterial strains. It also sets the stage for regulatory bodies to reconsider and enforce MRL in the country that are in line with procedures that benefit public health.The present study, using ELISA, showed that samples from the liver and breast of broiler chicken, these being organoleptically favored in Middle East countries, contain various levels of residues of different antibiotics. Antibiotics can penetrate and remain in an animal’s tissues, especially if the initial dosage of the antibiotic is high or if the withdrawal time is not sufficient before slaughtering .In this study, ENR residues in both breast and liver samples from farm five were above the MRL. ENR, a kind of amphoteric of the quinolone (fluoroquinolone) antimicrobial family, is a potent antibiotic with a broad spectrum of activity against Gram-positive and Gram-negative micro-organisms . QuinoloAlthough the residual levels of GEN, OTC, SMZ, and TYL in both breast and liver samples were below the MRL, their hazard for human health still exists. In this study TYL residue concentrations were the highest recovered from liver (mean rank 150.33) compared with breast (50.67). Our results are nearly consistent with Mossad et al. , in whicIt is well recognized that the steps in food processing should be kept under continuous monitoring for preventing overexpressed levels of drug residues. This work was a prelude to unraveling the extent of residual antibiotics in poultry chicken in Oman. As it revealed residual antibiotic levels of a number of antibiotics in chicken meat samples from randomly selected farms, it must be a reminder to heed the recommended use of antibiotics, lest the risk of development of antibiotic resistance should escalate. This should encourage regulatory bodies to adopt and enforce guidelines for antibiotics use in poultry based on specifically identified indications. This study furnishes the base for further studies on the regulated use of veterinary drugs in Oman.The results indicate the presence of antibiotic residues and that ENR residue concentrations were above the MLR. The consumption of this contaminated meat can result in antibiotic resistance in consumers and poses a public health threat. In Oman, most of the privately owned poultry farms and farmers do not engage the services of veterinarians for disease diagnosis and drug prescription. They claim to recognize disease conditions based on their knowledge and experience of clinical signs and postmortem findings. Therefore, there is a need to educate farmers and encourage them to seek laboratory confirmation of disease diagnosis and to conduct antibiotic sensitivity tests for bacterial isolates. National authorities should also adopt more judicious approaches to ensure the prudent use of antibiotics in food animals.


The pMECs were used as an in vitro model and were treated with various concentrations of Se-Met . Cells were analyzed for cell viability, selenoprotein transcriptome, selenoprotein expression, and antioxidant enzyme activities. The results showed that, with increasing Se-Met concentrations, cell viability first increased and then decreased at 24, 48, or 72 h posttreatment with maximum values at 0.5-μM Se-Met. As the Se-Met concentrations increased, the mRNA expression of 17 selenoproteins first upregulated and then downregulated, with maximum values at 0.5-μM Se-Met. The 17 selenoproteins included SEPHS2, SELENOP, GPX1, GPX2, GPX3, GPX6, TXNRD1, SELENOK, SELENOW, DIO1, DIO2, DIO3, SELENOF, SELENOS, SELENOH, SELENOI, and SELENOT. Additionally, the protein expression levels of SEPHS2, SELENOP, GPX1, and TXNRD1 and the activities of glutathione peroxidase and thioredoxin were highest at 0.5-μM Se-Met. In conclusion, 0.5-μM Se-Met promotes cell viability partially by improving selenoprotein expression and antioxidant function in pMECs, which provides evidence for the potential ability of Se-Met to improve mammary gland health in sows.This study investigated the effects of selenomethionine (Se-Met) on the cell viability, selenoprotein expression, and antioxidant function of porcine mammary epithelial cells (pMECs) to reveal the underlying molecular mechanism of Se-Met on the lactation performance and antioxidant capacity of sows  CSN1S2, LALBA, WAP, SAA2, and BTN1A1, and the transcriptional regulators SREBP1 and XBP1 are activated  . Once soctivated , and thectivated . Howevervia milk (via selenoproteins (It has been demonstrated that selenium (Se) promotes the development of the mammalian mammary gland  and affevia milk –17. The via milk , 19 becavia milk . Seleniuvia milk , and it via milk . Se statvia milk . The Se via milk . The Se proteins . Se phosproteins . SEPHS2 proteins , which iproteins . Glutathproteins . There aproteins . Thioredproteins . Howeverin vitro.Therefore, this study aimed to investigate the effect of selenomethionine (Se-Met) on selenoprotein expression and antioxidant function in pMECs to reveal the underlying molecular mechanism of Se-Met on the lactation performance and antioxidant capacity of sows 2. Cells were cultured in a complete medium according to the formula of Jaeger et al. . At 24, 48, and 72 h posttreatment, 20-μL CCK-8 was added to each well and incubated for 4 h at 37°C and then measured using a microplate reader at a wavelength of 450 nm.Cell viability was assessed using the CCK-8 assay  according to the instructions of the manufacturer. Briefly, pMECs were seeded into 96-well microplates at 200 μL/well at 2 × 104 cells/mL and cultured in a complete medium at 37°C and 5% CO2 for 48 h. Then, the cells were treated with different levels of Se-Met  for 48 h. After that, total RNA was extracted from pMECs using TRIzol  according to the instructions of the manufacturer. The quality and the quantity of RNA were analyzed by an Agilent Bioanalyzer 2100 using an RNA 6000 Labchip kit. Potential DNA contamination in the extraction was eliminated using a DNA-free kit , and the RNA quality was verified by both agarose gel (1%) electrophoresis and spectrometry (A260/A280). First-strand cDNA synthesis was performed by using a PrimeScript RT reagent kit with a gDNA eraser . cDNA was synthesized from 1 μg of total RNA using SuperScript III reverse transcriptase according to the instructions of the manufacturer. The mRNA levels of 25 selenoprotein genes were analyzed by qPCR using SYBRR Green PCR Master Mix according to the instructions of the manufacturer . Primers for the 25 selenoprotein genes (see ACTB) were from our previous study  for 48 h. After that, the cells were collected and homogenized in a RIPA lysis buffer . Western blot analysis was performed according to the procedures described in our previous study  for 48 h. After that, the cells were collected for glutathione peroxidase (GPX) activity and thioredoxin reductase (TRX) activity analysis. GPX activity (nmol NADPH/min/mL) was measured in the supernatant using a cellular glutathione peroxidase assay kit (Beyotime Institute of Biotechnology) that measures the coupled oxidation of NADPH during glutathione reductase (GR) recycling of oxidized glutathione from GPX-mediated reduction of t-butyl peroxide. For this assay, excess GR, glutathione, and NADPH were added according to the instructions of the manufacturer. The protein concentration of splenocyte lysate was measured using a Bicinchoninic Acid assay (Beyotime Institute of Biotechnology). Protein concentrations were used to correct the GPX activity of the cell lysates. GPX activity was expressed as mU/mg.Porcine mammary epithelial cells were seeded into six-well plates at 2 mL/well at 5 × 102−), which generates a strong yellow color (λmax = 412 nm). Since other enzymes, such as glutathione reductase and glutathione peroxidase, can also reduce DTNB in crude biological samples, a TRX-specific inhibitor was utilized to determine the TRX-specific activity. Two assays were performed: the first measurement was the total DTNB reduction by the sample, and the second was the DTNB reduction by the sample in the presence of the TRX specific inhibitor. The difference between the two results represented the DTNB reduction by TRX.Thioredoxin reductase activity was measured using a thioredoxin reductase activity colorimetric assay kit . In this assay, TRX catalyzes the reduction of 5, 5′-dithiobis (2-nitrobenzoic) acid (DTNB) with NADPH to 5-thio-2-nitrobenzoic acid , respectively.Statistical analysis was conducted using SPSS 22.0 . Data were analyzed using one-way ANOVA, followed by Duncan's multiple comparison test. The results are presented as mean and SEM. p < 0.05), respectively, while 4.μM Se-Met did not affect cell viability (p > 0.05). After 48 h of incubation, compared with the control group, 0.5- and 1.-μM Se-Met enhanced cell viability by 15.26 and 15.36% (p < 0.05), respectively, but 2.- or 4.-μM Se-Met did not influence cell viability (p > 0.05). When cells were treated for 72 h, Se-Met did not affect cell viability in comparison to the control group (p > 0.05), but 0.5-μM Se-Met improved cell viability compared with the 4.-μM Se-Met group by 12.55% (p < 0.05). Therefore, we selected 48 h as the incubation time for the subsequent experiments.As shown in A heat map of the effects of Se-Met supplementation on the selenoprotein transcriptome of pMECs is shown in SEPHS2 and SELENOP first increased and then gradually decreased, reaching a maximum value at 0.5-μM Se-Met, which was higher than that of the 4.-μM Se-Met group (p < 0.05) . However, the mRNA expression of GPX1, GPX2, GPX3, and GPX6 first increased and then decreased with the increase of Se-Met concentration, reaching a plateau value at 0.5-μM Se-Met  (MSRB1 (p > 0.05). As the Se-Met concentration increased, the mRNA expression of SELENOK and SELENOW first increased and then decreased, peaking at 0.5-μM Se-Met (p < 0.05). As displayed in DIO1, DIO2, and DIO3 first increased and then decreased, reaching a maximum value at 0.5-μM Se-Met (p < 0.05). As shown in SELENOM (p > 0.05). With the increasing Se-Met concentrations, the mRNA expression of SELENON first increased and then increased, reaching a minimum value at 0.5-μM Se-Met. With the increasing Se-Met concentration, the mRNA expressions of SELENOF and SELENOS mRNA were increased firstly and then decreased, and the expression was the highest at 0.5-μM Se-Met (p < 0.05). As shown in SELENOV (p > 0.05). With the increasing Se-Met concentration, the mRNA expression of SELENOO first increased and then increased. With the increasing Se-Met concentration, the mRNA expression of SELENOH, SELENOI, and SELENOT first increased and then decreased, and the expression was the highest at 0.5-μM Se-Met. The mRNA expressions of SELENOH, SELENOI, and SELENOT were higher in the 0.5-μM Se-Met group than that in the 4.-μM Se-Met group (p < 0.05).With increasing Se-Met concentration, the mRNA expression of  < 0.05) . For theM Se-Met . Regardi < 0.05) . As presp < 0.05). Additionally, the protein expression of SEPHS2 in the 1.-μM Se-Met group was increased compared with that in the 4.-μM Se-Met group (p < 0.05). As the Se-Met concentration increased, the protein expression of SELENOP first increased and then decreased; the expression was the highest at 0.5-μM Se-Met, and the expression of SELENOP in the 0.5-μM Se-Met group was higher than that in the 2.- and 4.-μM Se-Met groups (p < 0.05) (P < 0.05), while the 1.- and 2.-μM Se-Met groups had higher GPX1 protein expression than the control group (p < 0.05). As represented in p < 0.05), while the 1.-μM Se-Met group was higher than that in the 2.- and 4.-μM Se-Met groups (p < 0.05). The original images for the blots are provided in the As displayed in  < 0.05) . As presp < 0.05), while GPX activity in the 1.-μM Se-Met group was elevated compared with that in the 4.-μM Se-Met group (p < 0.05). With the increasing Se-Met concentration, TRX activity first increased and then decreased, peaking at 0.5 μM. The TRX activity in the 0.5-μM Se-Met group was higher than that in the 0-, 1.-, 2.-, and 4.-μM Se-Met groups (p < 0.05).As presented in in vitro. Yan et al.  is the primary transporter of Se in milk . The Se d by 65% . In othefsprings . Hill etfsprings  also repSEPHS2, SELENOP, GPX1, GPX2, GPX3, GPX6, TXNRD1, SELENOK, SELENOW, DIO1, DIO2, DIO3, SELENOF, SELENOS, SELENOH, SELENOI, and SELENOT, was first increased and then decreased, reaching a maximum at 0.5-μM Se-Met. It has been reported that Se can affect the selenoprotein transcriptome in mouse ATDC5 chondrocytes and human C28/I2 cells  production in mice, inhibit cadmium-induced mitochondrial membrane collapse, prevent cytochrome C release, and inhibit caspase death receptor activation  can be found in the article/WG and SZ: conceptualization. FC: methodology and supervision. JY: software. YZ and MT: validation. YZ: formal analysis. JC: investigation and writing—original draft preparation. WG: resources, writing—review and editing, project administration, and funding acquisition. YL: data curation. SZ: visualization. All authors have read and agreed to the published version of the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Patients treated with antibiotics have worse outcomes from cancer immunotherapy. While antibiotics are known to affect intestinal flora, it is not clear how they affect overall immune function. The aim of our study was to identify differences in immune parameters according to antibiotic exposure. Among 251 total patients, the 135 (54%) who received antibiotics had lower response rates and shorter survival. We identified significant differences in multiple antibodies according to antibiotic exposure, including antibodies specific for nucleolin, MDA5, c-reactive protein, LC1, heparin sulfate, Matrigel, and CENP.B. In lung cancer patients, antibiotics were associated with differences in IFN-γ, IL-8, and macrophage inflammatory protein cytokines. Administration of antibiotics to patients receiving cancer immunotherapy is associated with changes in circulating antibodies and cytokines, although it is not clear if antibiotics cause these differences. Given the frequency of antibiotic use in cancer populations and potentially detrimental effects on immunotherapy outcomes, more research in this area may guide patient management.p < 0.05 and false discovery rate (FDR) < 0.2 considered significant. A total of 251 patients were included, of whom the 135 (54%) who received antibiotics had lower response rates and shorter survival. Patients who received antibiotics prior to ICI initiation had modestly but significantly lower baseline levels of nucleolin, MDA5, c-reactive protein, and liver cytosol antigen type 1 (LC1) antibodies, as well as higher levels of heparin sulfate and Matrigel antibodies. After ICI initiation, antibiotic-treated patients had significantly lower levels of MDA5, CENP.B, and nucleolin antibodies. Although there were no clear differences in cytokines in the overall cohort, in the lung cancer subset (53% of the study population), we observed differences in IFN-γ, IL-8, and macrophage inflammatory proteins. In ICI-treated patients, antibiotic exposure is associated with changes in certain antibodies and cytokines. Understanding the relationship between these factors may improve the clinical management of patients receiving ICI.Antibiotic administration is associated with worse clinical outcomes and changes to the gut microbiome in cancer patients receiving immune checkpoint inhibitors (ICI). However, the effects of antibiotics on systemic immune function are unknown. We, therefore, evaluated antibiotic exposure, therapeutic responses, and multiplex panels of 40 serum cytokines and 124 antibodies at baseline and six weeks after ICI initiation, with  Immune checkpoint inhibitors (ICI) have transformed the treatment landscape of many advanced cancers and have substantially improved outcomes for a subset of patients. Despite the widespread use of ICI, the identification of patients most likely to benefit from these costly and potentially toxic therapies remains challenging. Established tumor-based predictive biomarkers for ICI efficacy include PD-L1 expression, microsatellite instability, and tumor mutational burden ,3,4,5. BAdditionally, a number of clinical characteristics appear to predict ICI efficacy. In patients with lung cancer, the extent of prior or current smoking is associated with tumor mutation burden and ICI efficacy . ExposurAmong the most striking clinical associations with ICI outcomes are antibiotic exposure. Numerous cohort studies have demonstrated that patients receiving antibiotics around the time of ICI initiation experience inferior progression-free survival (PFS) and overall survival (OS) ,24,25,26Although numerous studies have established an association between antibiotics and ICI clinical outcomes, little is known about the effects of antibiotics on systemic immune parameters. We, therefore, analyzed cytokine and antibody profiles near the time of ICI initiation according to antibiotic exposure, recognizing that such observations imply association and not necessarily causation. In order to determine clinical benefit in this period, we included tumor radiographic response, which may be less prone to confounders (such as infection) than other endpoints.This study was approved by the UT Southwestern Institutional Review Board (IRB #STU 082015-053). In this prospective cohort study, patients planned to initiate ICI for cancer therapy underwent clinical data and serial biospecimen collection. Clinical data included age, sex, race, ethnicity, cancer type and stage, ICI type, date of ICI initiation, progression-free survival (PFS), overall survival (OS), and best radiographic response . We abstracted Eastern Cooperative Oncology Group (ECOG) performance status from the medical records by searching for “performance status” and “ECOG” keywords and documenting patient performance status within one month prior to or after starting ICI treatment. If performance status was not documented in this timeframe, it was designated as missing. For the current analysis, we performed a retrospective medical records review to identify antibiotic exposure. Based on earlier studies in this area, we defined antibiotic exposure as an antibiotic prescription between six weeks before and six weeks after ICI initiation ,33,34. Wg at 4 °C for 15 min to obtain plasma.After patients were enrolled in the cohort study, peripheral blood was collected before and 6 weeks after ICI initiation; these time-points were selected based on prior studies of baseline and post-treatment circulating immune parameters . At eachCytokine levels were measured using a Bio-Plex Pro Human Chemokine 40-plex Panel  on the Luminex 200 System . The conWe previously developed and manufactured a custom protein array panel of 124 antigens, including nuclear antigens, cytosolic/matrix antigens, and tissue/organ-specific antigens , and havAntibody panels were run in six batches containing between 42 and 345 unique samples. For each antigen and batch combination, we required that at least 10% of samples had a signal-to-noise ratio ≥3. Antigens with less than 99% of values available across all samples were then dropped from analysis. Antibody data was further normalized using variance stabilizing normalization (VSN) and batch-corrected using ComBat . Any valp values using Fisher’s exact test for categorical variables and t-tests for continuous variables. We generated heat maps for baseline and 6-week cytokine and antibody values based on the z-score of each sample relative to all other samples for the given cytokine or antibody. False discovery rates (FDR) were evaluated using the Benjamini–Hochberg procedure. We produced Kaplan–Meier curves and cox regressions using R survival package (v3.1-8). We defined overall survival (OS) as the interval between ICI initiation and date of death. We defined progression-free survival (PFS) as the interval between ICI initiation and either (a) clinical progression, (b) radiographic progression, or (c) death. In the event a particular case had an unidentified endpoint, the endpoint was censored at the date of last known follow-up. For antibiotic timing comparisons, we compared individuals that received antibiotics pre-ICI to the rest of the cohort, regardless of post-ICI antibiotic exposure, and we compared individuals that received post-ICI antibiotics to the rest of the cohort, regardless of pre-ICI antibiotic exposure. Antibiotics received after ICI initiation and the cumulative number of distinct antibiotics received were modeled as time-dependent variables in Cox regressions to avoid immortality bias. Because cytokine patterns and effects differ by cancer type [p values were determined using the Pearson product-moment correlation coefficient. All computation was performed with R (v3.6.3). ComBat batch correction was applied using the sva package (v3.34.0), and VSN was applied using the vsn package (v3.54.0).For demographic characteristics, we computed cer type , we perfn = 133, 53%) and melanoma . Cancer stage distribution was as follows: Stage I/II ; Stage III ; Stage IV ; Unknown . ECOG performance status distribution was as follows: ECOG 0 , ECOG 1 , ECOG 2–4 , and ECOG missing . Overall, 135 patients (54%) received an antibiotic prescription between six weeks before and six weeks after ICI initiation as follows: 44 (18%) only prior to ICI, 57 (23%) only after ICI, and 34 (14%) both before and after ICI.A total of 251 patients were included in the analysis. Median age was 68 years and 98 (39%) were female. The most common cancer types were non-small cell lung cancer (NSCLC) (n = 481), prophylaxis (n = 116), genitourinary (n = 94), unknown (n = 86), skin and soft tissue (n = 74), gastrointestinal (n = 35), sepsis (n = 27), other (n = 7), and fever (n = 6).Indications for antibiotic prescriptions included the following: respiratory (p = 0.09), with NSCLC accounting for more than 60% of cases receiving antibiotics post-ICI initiation . This difference is driven primarily by cases receiving antibiotics after ICI initiation (p = 0.02).To establish the association between antibiotic exposure and clinical outcomes in the study cohort, we examined OS and PFS. Because these parameters may be determined well after the period of antibiotic exposure and may be subject to confounding factors, we also determined radiographic response according to response evaluation criteria in solid tumors (RECIST) . Evaluatp = 0.63) , there was no association with antibiotic exposure ( = 0.63) . AlthougConsistent with other reports, patients who received antibiotics had worse OS and PFS . Similarn = 180), six weeks (n = 108), both time-points (n = 107). Availability of cytokine data was as follows: baseline (n = 229), six weeks (n = 133), both time-points (n = 128). After applying our statistical quality control procedure to analytes across samples, a total of 31 cytokines and 71 antibodies were suitable for further analysis.After removing samples that did not meet specified quality thresholds, the availability of antibody data was as follows: baseline (p < 0.05): nucleolin (FDR 0.11), C-reactive protein (CRP) (FDR 0.11), liver cytosol antigen type 1 (LC1) (FDR 0.17), and melanoma differentiation-associated protein 5 (MDA5) (FDR 0.17). Baseline levels of heparin sulfate (FDR 0.1) and Matrigel (FDR 0.16) antibodies were significantly higher in patients who received pre-ICI antibiotics. At the 6-week time-point, levels of antibodies specific for MDA5, centromere protein B (CENP-B), and nucleolin were significantly lower  in patients who received antibiotics before starting ICI. We observed no significant differences in changes in antibody levels between the baseline and 6-week time-point. In patients receiving antibiotics after ICI initiation, there were no significant differences in antibody levels for any time permutation. p < 0.05) grouped by antibiotic exposure without incorporating the FDR cutoff. We observed no significant differences in cytokine profiles according to antibiotic exposure for any time permutation.As displayed in p = 0.09) and was significantly worse with post-ICI antibiotic exposure (p = 0.02) (p = 0.5) . In our study cohort, NSCLC was the most prevalent type of cancer and had the highest rate of antibiotic exposure. Consistent with data from the overall cohort, we observed worse OS and a trend towards worse PFS with antibiotic exposure . Further = 0.02) . As withp = 0.5) .p < 0.01, FDR 0.01). When examining the fold-change from baseline to six weeks, the following cytokines had significantly greater changes in patients with pre-ICI antibiotic exposure: interferon-gamma (IFN-γ) , interleukin-8 (IL-8)/chemokine (C-X-C motif) ligand 8 (CXCL8) , macrophage inflammatory protein-1a (MIP-1a)/CCL3 , and I309/CCL1 . The fold-change for macrophage inflammatory protein-1d (MIP-1d)/CCL15 was significantly lower in the pre-ICI antibiotic exposure group .We analyzed systemic immune parameters in the NSCLC cohort, displaying those with significant association with antibiotic exposure in We then analyzed the association between clinical outcomes and the systemic immune parameters identified as associated with antibiotic exposure. Among these, no biomarkers identified in the overall cohort were associated with outcomes, while IL-8/CXCL8 and IFN-γ were significantly associated with outcomes in the NSCLC cohort. Specifically, higher baseline IL-8/CXCL8 and fold change in IL-8/CXCL8 from baseline to six weeks was associated with inferior OS; higher fold change in IL-8/CXCL8 from baseline to six weeks was associated with inferior PFS; higher 6-week IL-8/CXCL8 was associated with inferior OS, PFS, and radiographic response; and greater positive fold-change in IFN-γ from baseline to six weeks was associated with inferior OS .Along with smoking history, steroid exposure, and body mass index, antibiotic exposure has emerged as a clinical factor influencing ICI efficacy. Specifically, patients receiving antibiotics have consistently been found to have inferior disease control and survival, an effect attributed to changes in the gut microbiome. In the present study, we confirmed this finding and examined systemic immune parameters across ICI patient populations according to antibiotic status. We observed significant differences in levels of approximately five percent of evaluated antibodies according to antibiotic exposure.Heparan sulfate antibody levels were higher in patients treated with antibiotics. Notably, heparan sulfate is a crucial mediator for various infections, including SARS-CoV-2 , suggestAlthough we did not identify differences in cytokine profiles in the overall study cohort, in NSCLC patients—who represented almost 55% of the total study population—we noted numerous differences. We performed this tumor-specific subset analysis because cytokine patterns and effects may differ according to cancer type . Among NIt is well established that antibiotics alter the gastrointestinal microbiome. Antibiotic-induced dysbiosis is associated with decreased microbiome diversity, which may take years to recover ,54. ThisWithout clear systemic markers of antibiotic effects on anti-tumor immunity, the use of antibiotics in cancer patients undergoing immunotherapy presents a clinical conundrum. Although antibiotics have been linked to numerous adverse events, including allergic reactions, gastrointestinal toxicity, drug–drug interactions, development of antimicrobial resistance, tendinopathies, and rupture, and bleeding due to effects on colonic bacteria vitamin K production ,57,58, aIn the present study, antibiotic exposure prior to ICI initiation had the strongest association with systemic immune parameters, while antibiotic exposure after ICI initiation had the strongest association with clinical outcomes. While the reasons for this discrepancy cannot be determined from available data, there are some potential explanations. First, the cohort receiving antibiotics prior to ICI initiation was smaller than the antibiotics post-ICI initiation cohort. While this cohort demonstrated a trend toward worse clinical outcomes, a small sample size may have limited statistical power. Another potential factor is the nature of the cohort receiving antibiotics post-ICI initiation. Because of the growing awareness of antibiotics’ detrimental effects on clinical outcomes in patients receiving checkpoint inhibitor therapy, it is possible that clinicians may now be hesitant to prescribe the agents, resulting in their use only in more clinically severe cases, which in turn might have worse outcomes. Lastly, the effects of post-ICI antibiotics on systemic immune parameters may not be apparent during the assessment period. The median time to maximal effect of antibiotics on the gut microbiome is nine days . If micrThis study has a number of limitations. First, this is a retrospective study from a single institution. Second, data for additional factors that may alter the gastrointestinal microbiome, such as diet, country of origin, medications other than antibiotics, and genetic predisposition, were not available. We also recognize that our review of clinical data may have missed antibiotic prescriptions. Conversely, patients may have been prescribed but never taken antibiotics. While we were able to identify the clinical indication for most antibiotic prescriptions, it is not possible to determine the association between indication, systemic immune parameters, or clinical outcomes because patients may have received multiple antibiotics. Importantly, we do not have stool samples to assess the gut microbiome in study subjects. Finally, in this study, we are unable to determine whether the observed associations between antibiotic exposure and immune parameters represent causative effects or surrogates for other phenomena such as clinical infections or tumor-associated inflammatory changes. Such distinction might require a prospective, randomized trial of antibiotics in patients receiving ICI, which is unlikely to be feasible.Despite these limitations, we note certain strengths, including the availability of pre- and post-treatment plasma samples, a cohort large enough to confirm the established effect of antibiotics on ICI outcomes, and extensively characterized clinical data.To our knowledge, our assessment of cytokines and antibodies represents the first interrogation of systemic immune parameters according to antibiotic exposure in a population treated with ICI. Although antibiotics were associated with inferior clinical outcomes, including radiographic response, they were associated with few differences in cytokine and antibody profiles. Furthermore, these differences did not reveal any patterns with clear clinical or biological significance. Mechanistic studies, including stool composition and immune cell population analysis, might further elucidate the molecular effects of antibiotic exposure on immune physiology. Increased understanding of the relationship between these factors may allow clinicians to improve the clinical management of patients receiving ICI.Drs. Khan, Fattah, Park, Xie, Li, Wakeland, and Gerber report a U.S. patent application .


In this article, the sum secure degrees-of-freedom (SDoF) of the multiple-input multiple-output (MIMO) X channel with confidential messages (XCCM) and arbitrary antenna configurations is studied, where there is no channel state information (CSI) at two transmitters and only delayed CSI at a multiple-antenna, full-duplex, and decode-and-forward relay. We aim at establishing the sum-SDoF lower and upper bounds. For the sum-SDoF lower bound, we design three relay-aided transmission schemes, namely, the relay-aided jamming scheme, the relay-aided jamming and one-receiver interference alignment scheme, and the relay-aided jamming and two-receiver interference alignment scheme, each corresponding to one case of antenna configurations. Moreover, the security and decoding of each scheme are analyzed. The sum-SDoF upper bound is proposed by means of the existing SDoF region of two-user MIMO broadcast channel with confidential messages (BCCM) and delayed channel state information at the transmitter (CSIT). As a result, the sum-SDoF lower and upper bounds are derived, and the sum-SDoF is characterized when the relay has sufficiently large antennas. Furthermore, even assuming no CSI at two transmitters, our results show that a multiple-antenna full-duplex relay with delayed CSI can elevate the sum-SDoF of the MIMO XCCM. This is corroborated by the fact that the derived sum-SDoF lower bound can be greater than the sum-SDoF of the MIMO XCCM with output feedback and delayed CSIT. K-user single-input single-output (SISO) interference channel with confidential messages (ICCM), the sum-SDoF was characterized in [The deployment of 5G and all of the connections to 6G around the world have all exacerbated the concerns for information-theoretic security in mobile communication networks ,2,3. Therized in . For therized in . The X nrized in . The sumrized in .For the fast-fading wireless channel, the CSIT can be delayed, that is, mismatching with current CSI but matching with past CSI. Under this imperfect setting, i.e., delayed CSIT, the research of SDoF was stemmed from , where uK-user SISO interference channel. In [L multiple-antenna full-duplex relays enhanced the sum-DoF of the Even with delayed CSI, it is shown in ,20,21,22shown in  addresseTo derive the sum-SDoF lower bound, under arbitrary antenna configurations, we propose three relay-aided transmission schemes, whose achievable sum-SDoF serves as the sum-SDoF lower bound. Specifically, we propose the relay-aided jamming scheme, and the relay-aided jamming and one-receiver IA scheme, and the relay-aided jamming and two-receiver IA scheme, where each scheme corresponds to one case of antenna configurations. In each scheme, the security and decoding are analyzed.To obtain the sum-SDoF upper bound, which does not appear in the existing literature and is non-trivial, we first treat two transmitters and the relay as a co-located transmitter, which is an enhanced scenario. Thereafter, we apply the results of the existing SDoF region of two-user MIMO BCCM, which is proven in , into thOur results show that if the full-duplex relay has double antennas of the receiver, the proposed sum-SDoF lower bound is not less than the sum-SDoF in existing two-user interference networks with delayed feedback in ,15,16,17However, to summarize, none of existing works considered the SDoF of the relay-aided MIMO XCCM, where there is no CSI at two transmitters and a delayed CSI at the relay, such as the CSI model in ,22. For Notations: The identity matrix of dimensions m is denoted by Organizations: The rest of this paper is organized as follows: The system model is defined in R-antennas full-duplex  with confidential messages t, a stochastic encoder t, a stochastic encoder n, a decoder A code rding to , the secThe sum of the secure channel capacity is defined as Theorem 1.(sum-SDoF lower bound under arbitrary antenna configurations). For the relay-aided MIMO XCCM with delayed CSI at the multiple-antenna full-duplex relay and no CSI at two transmitters, the sum-SDoF lower bound under arbitrary antenna configurations is given as follows:whereProof. Please refer to Remark 1.(sum-SDoF lower bound under symmetric antenna configurations). For the relay-aided MIMO XCCM with delayed CSI at the multiple-antenna full-duplex relay and no CSI at two transmitters, the sum-SDoF lower bound under symmetric antenna configurations is given as follows by setting Proposition 1.(sum-SDoF upper bound under arbitrary antenna configurations). For the relay-aided MIMO XCCM with delayed CSI at the multiple-antenna full-duplex relay and no CSI at two transmitters, the sum-SDoF upper bound under arbitrary antenna configurations is given by the following:Proof. We enhance the channel by supposing that the two transmitters and the relay constitute a co-located transmitter. Based on . For the relay-aided MIMO XCCM with delayed CSI at the multiple-antenna full-duplex relay and no CSI at two transmitters, the sum-SDoF is characterized by the following:Proof. N for Following Theorem 1 and Proposition 1, we can verify the match of the sum-SDoF upper and lower bounds for Remark 2.. To illustrate the proposed upper and lower bounds for arbitrary antenna configurations, we have shown four examples in When Phase I (Jamming by the Relay): This phase aims at sending R antennas. Denote the artificial noise vector sent by R antennas in Phase II Chain rule of mutual information.(b)Applying the data processing inequality for the Markov chain (c)When the input is the circularly symmetric complex Gaussian, according to , rewrited using The rank of matrix We shall ensure alently,  is zero.(a)Chain rule of mutual information.(b)Applying the data processing inequality for the Markov chain (c)When the input is the circularly symmetric complex Gaussian, according to , and rewd using The rank of matrix Secondly, we only need to verify the zero information leakage at the We shall ensure alently,  is zero.Decoding analysis: Firstly, we need to verify the decoding of transmitted data symbols Secondly, we need to verify the decoding of transmitted data symbols Achievable sum-SDoF analysis: As shown in the decoding analysis, two receivers can decode a total of If bstitute  and 19)N2<R, we If τ2=0 and  into (M1If The achievable sum-SDoF of the proposed relay-aided jamming scheme is given in  for M1+MWhen Phase I (jamming by the relay): This phase aims at sending R antennas. Denote the artificial noise vector sent by R antennas in The CSI matrices of Phase I return to the relay at the end of Phase I.Phase II  at the relay, the received signals of Phase II at the relay are expressed as follows:R observations at the relay, the relay can immediately decode all transmitted Since there are Phase III : In order to provide lacking equations for decoding, we aim at sending At the same time, two transmitters keep silent. The received signals of Phase IV at two receivers are written as follows:Security Analysis: As the received signals in Phase IV can be constructed by the received signals in Phase II and CSI matrices, the security analysis is similar to that in the relay-aided jamming scheme. Therefore, it can be checked that to ensure zero information leakage at d follow  and 19)Security Decoding analysis: Firstly, we need to verify the decoding of transmitted data symbols Then, multiplying  by 19),,19), we Secondly, we need to verify the decoding of transmitted data symbols At each time slot, there are Achievable sum-SDoF analysis: As shown in the decoding analysis, two receivers can decode a total of If bstitute , 19), a, aN2<R, bstitute  with equIf bstitute , τ2=0, aIf The achievable sum-SDoF of the proposed relay-aided jamming and one-receiver IA scheme is given in  for N1<MWhen Phase I (jamming by the relay): This phase aims at sending R antennas in R antennas in The CSI matrices of Phase I return to the relay at the end of Phase I.Phase II : In order to provide lacking equations for decoding, we aim at sending At the same time, two transmitters keep silent. The received signals of Phase IV at two receivers are written by the following:Security Analysis: As the received signals in Phase IV can be constructed by the received signals in Phase II, Phase III, and CSI matrices, the security analysis is similar to that in the relay-aided jamming scheme. Therefore, it can be checked that to ensure zero information leakage at d follow  and 19)Security Decoding analysis: Firstly, we need to verify the decoding of transmitted data symbols If 2≤N1+N2,  simplifiOtherwise, if 2<M1+M2,  simplifiCombining  and 51)51), we hThen, multiplying  by 19),,19), we Secondly, we need to verify the decoding of transmitted data symbols If 2≤N1+N2,  simplifiOtherwise, if 2<M1+M2,  simplifiCombining  and 57)57), we hThen, multiplying  by 17),,17), we Achievable sum-SDoF analysis: As shown in the decoding analysis, two receivers can decode a total of If bstitute , 19), a, aN2<R, bstitute  with equbstitute  with equIf bstitute , τ2=0, a2=0, and  with equIf The achievable sum-SDoF of proposed relay-aided jamming and two-receiver IA scheme is given in (We studied the sum-SDoF of the multiple-antenna full-duplex relay-aided MIMO XCCM with arbitrary antenna configurations, where there is no CSI at two transmitters and delayed CSI at the relay. To establish a sum-SDoF lower bound, we designed three transmission schemes. For the proposed schemes, full-duplex relay was utilized to receive the data symbol signals meanwhile transmit jamming signals, as this method can increase the transmission efficiency in contrast to half-duplex systems. We also derived a sum-SDoF upper bound. We characterized the sum-SDoF for


Children show important developmental and maturational changes, which may contribute greatly to pharmacokinetic (PK) variability observed in pediatric patients. These PK alterations are further enhanced by disease-related, non-maturational factors. Specific to the intensive care setting, such factors include critical illness, inflammatory status, augmented renal clearance (ARC), as well as therapeutic interventions . This narrative review illustrates the relevance of both maturational and non-maturational changes in absorption, distribution, metabolism, and excretion (ADME) applied to antibiotics. It hereby provides a focused assessment of the available literature on the impact of critical illness—in general, and in specific subpopulations —on PK and potential underexposure in children and neonates. Overall, literature discussing antibiotic PK alterations in pediatric intensive care is scarce. Most studies describe antibiotics commonly monitored in clinical practice such as vancomycin and aminoglycosides. Because of the large PK variability, therapeutic drug monitoring, further extended to other antibiotics, and integration of model-informed precision dosing in clinical practice are suggested to optimise antibiotic dose and exposure in each newborn, infant, or child during intensive care. Antibiotics are commonly used in hospitalised children ,2,3. A oThe decision on the timing of prescription (when), the choice of the antibiotic(s) (what), and appropriate dosing (how much) to achieve sufficient exposure are crucial determinants for successful antibiotic therapy . PharmacChildren show important developmental or maturational changes, which may contribute greatly to PK variability observed in pediatric patients ,9. StandMost drugs—also antibiotics—have been developed in and for adults, with subsequent extrapolation of dosing regimens to children. Pediatric antibiotic trials are seldom performed due to ethical, practical, and economic considerations . This isThe aim of this narrative review is to illustrate the relevance of maturational changes in absorption, distribution, metabolism, and excretion (ADME) applied to antibiotics, and to perform a focused assessment of the available literature on the impact of critical illness—in general, and in specific subpopulations —on PK and potential underexposure in children and neonates. Such information on the relevant PK covariates is crucial to implement precision medicine for antibiotics in NICU and PICU populations. We, therefore, conducted a PubMed search to identify all relevant English-language articles and abstracts reporting antibiotic PK data in critically ill children and neonates from inception until July 2021. Two authors performed the literature review (M.G. and K.A.).Developmental pharmacology describes the impact of maturation on drug disposition (PK) and drug effects (pharmacodynamics [PD]) throughout the pediatric age range (0–18 y) . There iA recent ‘throughout-life’ PK modelling effort on vancomycin hereby illustrated that the current dosing recommendations (Food and Drug Administration label) commonly resulted in underexposure, with the highest probability in children and adolescents (70%), except in newborns, while the label remains very descriptive on dosing in preterm neonates ,22.Absorption: the impact of gastrointestinal variability on oral drug absorption and the relevance of developmental changes  has recently been reviewed . A systeDistribution: throughout infancy, the extracellular and total body water proportions are higher, and this results in higher volumes of distribution (Vd) and lower (peak) concentrations for water-soluble antibiotics  when administered on an mg/kg basis. 0.75) was used [Elimination: the main route of elimination of most antibiotics is by renal excretion, determined by both glomerular filtration rate (GFR) and renal tubular activities  so that pharmacometric approaches to personalise the use of primarily renally eliminated antibiotics in a pediatric and neonatal ICU setting should focus on renal models and renal (patho)physiology . A maturwas used . When bawas used . Along twas used ,35. Whenwas used . In contwas used .Such semi-physiological functions are relevant to truly assess the impact of non-maturational factors. Besides aspects related to the critical illness that will be discussed below, this also includes co-administration of nephrotoxic drugs, like non-steroidal anti-inflammatory drugs , commonly administered to induce closure of a patent ductus arteriosus. Using the impact of those drugs on aminoglycoside or vancomycin CL, the transient reduction in GFR during ibuprofen or indomethacin administration is −20 and −40%, respectively, in the newborn ,39.Pathophysiological changes and resulting PK alterations have been demonstrated for many antibiotics in critically ill children. Nevertheless, pediatric PK data remain relatively scarce and are lagging behind the plethora of PK studies reporting antibiotic PK in critically ill adults. Increased Vd is frequently observed due to fluid resuscitation and inflammation-induced capillary leak, further enhanced by hypoalbuminemia. Alterations in antibiotic CL have also been reported repeatedly in pediatric antibiotic studies, and have been related to both extremes of renal function  ,41. DespHigher Vd (L/kg) have been reported for most beta-lactam antibiotics in pediatric patients with severe bacterial infections compared with adult ICU patients. However, it should be noted that there is still large variability in PK between and within the different pediatric studies ,41. BodyDespite >50 published antibiotic PK studies in critically ill pediatric patients, PK data remain scarce and for most antibiotics, PK knowledge is based on only a few studies with small sample sizes ,41. For It should be noted that most PK studies report antibiotics that are subjected to routine TDM in clinical practice . Vancomycin is by far the most frequently reported antibiotic in critically ill children, with studies covering its PK over the whole pediatric age range in more than 1000 critically ill children ,66,67,68Aminoglycosides are also commonly monitored in clinical practice, which is reflected by several studies describing the PK of gentamicin ,74,75,76Beta-lactam PK studies in critically ill children show diverse results and are mostly limited to a few studies or cases reports for a specific beta-lactam antibiotic ,100,101.Finally, we also found a few PK studies describing linezolid , ciprofl2 at 1 month of postnatal age to 150 mL/min/1.73 m2 for all children aged 2 years and older [Although body weight and age-related covariates are most consistently identified in antibiotic PK studies in children, renal function is also frequently retained as a significant predictor of antibiotic exposure and target attainment. While decreased renal function is generally taken into account in drug dosing guidelines, increased renal function was rarely considered until recently. ARC, referring to the enhanced renal elimination of solutes , has only been identified as a major predictor of altered drug CL over the past decade . The vasnd older .Schwartz or the SCr concentration [Similar to previous findings in adults, there is an increasing number of studies demonstrating subtherapeutic exposure to antibiotics with predominant renal clearance in pediatric patients with ARC. The majority of these studies have been performed in critically ill children, and consistently reveal increased antibiotic CL associated with increased renal function, mostly measured by means of the estimated GFRntration . An overMost antibiotic PK studies in ARC patients have been performed with beta-lactams ,95,97,98Several studies have also reported subtherapeutic vancomycin exposure in critically ill children with ARC ,114,115.0–24) and target attainment for gentamicin in critically ill children with ARC vs. those without ARC [Interestingly, even for concentration-dependent antibiotics like gentamicin, ARC has been shown to contribute to subtherapeutic exposure. A recent study demonstrated decreased exposure  are suboptimal markers of renal function in this population ,108. SCrExtracorporeal membrane oxygenation (ECMO) is a life-saving technique used as a bridge-to-recovery for pediatric and adult patients with severe respiratory and/or cardiac failure . ECMO haThe potential impact of ECMO on antibiotic PK can be related to the direct interaction of the antibiotic with the ECMO circuit  ,118,119.An increase in Vd is observed for most hydrophilic antibiotics during ECMO in children. The impact of ECMO on Vd is inversely related to age. Due to the smaller amount of circulating blood volume, ECMO-induced hemodilution will have a larger impact in younger children than in older children or adults. Additionally, neonates and infants have a proportionally higher body water composition, which further enhances the impact of ECMO on the Vd of hydrophilic antibiotics ,119.As presented in Recent neonatal studies showed conflicting data concerning the impact of ECMO on vancomycin CL ,122. NevFor gentamicin, a hydrophilic drug with a low Vd, a few studies reported an increased Vd in neonates due to ECMO, which is a logical consequence of more pronounced hemodilution in younger children ,131. On Increased Vd was also observed during ECMO for cefotaxime , cefepimFinally, one popPK study in 63 critically ill children did not find a significant difference in linezolid PK in two patients supported with ECMO as compared to the other (non-ECMO) patients .Overall, the effect of ECMO on antibiotic PK in children appears to be limited. Changes in CL can mostly be related to renal function. Furthermore, increased Vd is mainly observed in neonates due to a proportionally higher hemodilution effect of the priming volume associated with ECMO.A few studies describe the PK of the most commonly used antibiotics during cardiopulmonary bypass (CPB). CPB is a technique very similar to ECMO, though only applied for a short period of time, typically the duration of cardiac surgery. Hence, PK alterations similar to those encountered with ECMO may be relevant. A significant effect from CPB on cefazolin  and cefuApproximately 25% of the children admitted to the ICU will develop acute kidney injury, and 4–6% will need RRT ,146. CurIt should be noted that the vast majority of PK studies investigating CRRT were performed in adults. Accordingly, none of the previously published reviews describing PK alterations and antibiotic dosing in CRRT patients provided recommendations for antibiotic therapy in children or neonates receiving CRRT. As a result, recommendations concerning antibiotic dosing in children during CRRT are largely extrapolated from adult studies. It is generally assumed that hydrophilic antibiotics with small molecular weight, and low plasma protein binding are most susceptible to CRRT induced PK alterations ,148.The effect of CRRT on antibiotic PK has only been investigated in a few pediatric studies with relatively small sample sizes, as shown in the overview of antibiotic PK studies during CRRT in Most pediatric PK studies during CRRT have been performed with meropenem ,137,138.One case report described the PK of piperacillin during CRRT and molecular adsorbent recirculating system (MARS) therapy . PiperacOne pediatric ICU study investigating linezolid PK included a subset of children with CRRT . CRRT waFor cefepime, a case series of 4 patients receiving CVVHDF showed that standard 50 mg/kg q12h might not be sufficient to reach predefined PK/PD targets . Dosing Finally, a recent case series of three critically ill children with different renal functions  treated with ceftolozane/tazobactam described the PK differences amongst these three patients . The CL Overall, in children treated with CRRT, antibiotic PK, and consequently antibiotic dosing, appears to be dictated by the effluent flow rate .WBH has a neuroprotective effect in (near)term neonates with moderate to severe hypoxic ischemic hypothermia and is applied to reach a target temperature of 33–34 °C for 72 h shortly (<6 h) after birth. However, hypoxic ischemic asphyxia and lowering the core body temperature affects PK, up to the level that dosing regimens for this specific setting should be adapted. A significant decrease in CL is observed in neonates during therapeutic hypothermia, be it that the extent differs between routes of elimination and compounds, but is most pronounced for renal elimination ,158. BasBased on the pediatric study decision tree, the maturational physiology throughout pediatric life results in more complex antibiotic dosing regimens to attain similar target exposure in children as compared to adults ,20. As hIn a first step (For the second step (Our review rather intended to summarise the available evidence on (non-)maturational covariates in the NICU and PICU setting. The next step to convert this information to attain precision medicine, based on Bayesian and MIPD software techniques, has been recently reviewed by others so that we refer the interested reader to this review .Han et al. recently identified and assessed the Bayesian software programs applicable to neonatal and pediatric patients for vancomycin (AUC over 24 h exposure) in such an MIPD context. Of the Bayesian programs retrieved, DoseMeRx, InsightRx, and PrecisePK utilised clinically validated neonatal and pediatric models. The authors came to the conclusion that clinicians should focus on selecting a model that best fits their patient population characteristics and utilise Bayesian estimation tools for therapeutic AUC-targeted dosing and monitoring of vancomycin .Finally, the third step (Obviously, such an approach necessitates additional multidisciplinary collaboration as the skills related to MIPD should be integrated into clinical care, while further evidence on efficacy and cost-benefit analysis of MIPD in healthcare is needed before MIPD will become common clinical practice . Further


When using either aid, there was no contamination of the bottle tip, which occurred in 46% of subjects when no aid was used (p = 0.0005). Fewer drops were expelled when using the conventional bottle as compared to the aids ; (4) Conclusions: Autodrop and Opticare can assist patients with eye drop placement. These aids completely prevented bottle tip contamination, which was frequently observed when the conventional bottle was used alone.(1) Background: Eye drops are the most common route of administration for ophthalmic medications. Administering drops can be a major hurdle for patients, potentially resulting in noncompliance and treatment failure. The purpose of this study is to compare the efficacy and safety of two different aids and the conventional bottle for eye drop instillation; (2) Methods: An interventional crossover study involving standard eye drop bottle, Opticare aid and Autodrop aid. The study included healthy subjects without a history of regular eye drop use; (3) Results: Twenty-six subjects were enrolled. Of those subjects, 96% and 92% were able to assemble the eye drop bottle into the Autodrop and the Opticare aids, respectively. Subjective assessment indicated that Autodrop was significantly easier to assemble than Opticare (95% CI: −1.6802 to −0.1659,  Eye drops are by far the most common route of administration for medications to treat ophthalmic diseases. Most drops are supplied in a small plastic bottle that is designed for direct instillation into the eye.The ability of patients to use eye drops independently and successfully can present a major challenge, potentially leading to noncompliance and treatment failure ,2,3. NonDifferent aids to administer eyedrops, aimed to help patients to self-administer drops, are readily available for purchase ,11,12,13The Autodrop  and Opticare  devices are two of the aids commercially available . Both arTo the best of our knowledge, no study that directly compared the efficacy and safety of different aids has been published . Therefohttps://clinicaltrials.gov/ct2/show/NCT03417453 (accessed on 21 November 2021).This was an interventional, randomized, crossover study. Initially, multiple readily available dispensing aids were evaluated by the local institutional review board committee, including by the hospital’s medical engineer and an infectious disease specialist. These aids included the Opticare, the Opticare Arthro 5, the Opticare Arthro 10  and the Autodrop . Each dispensing aid was tested and evaluated by the committee, with an emphasis on ergonomics, reliable application, and patient safety. As the Opticare Arthro 5 and Opticare Arthro 10 seemed to be more challenging to use and could fit only small, rounded bottles, the Autodrop and the Opticare aids were approved by the committee for use in this study. The study was registered in the NIH clinicaltrials.gov website: For use, both devices are placed on the orbit. Subject inclusion criteria were: age over 18 years, no previous regular eye drop use , and no motor disability that could negatively affect the ability to self-instill eye drops, as was defined in prior studies  ,10.http://www.randomization.com, accessed on 21 November 2021) and the instillation sequences were recorded in sealed envelopes labelled with the patient number. Each subject underwent a trial of all three instillation methods  in a randomized order, obtained from the envelope. Artificial tears  were used.For each subject, the drops instillation order was randomized using Randomization.com  and by counting the drops by the study personal who visually assessed the instillation; 3. If no accumulation of fluid in the eye was seen and the subject reported drop instillation, the investigator concluded that only one drop was instilled. Otherwise, the investigator concluded that no drops at all were expelled. After the use of each device, subjects completed a questionnaire  concerniTo estimate the minimum sample size, we used the difference in the number of drops dispensed in two recent studies. The first was a study of 23 patients that compared the instillation of eye drops with and without an assisting delivery device . The secp-value < 0.05 was considered statistically significant. Data are presented as mean ± standard deviation (SD).Clinical parameters distribution was tested for normality by the Shapiro–Wilk test. Independent and paired t-tests were conducted for continuous variables with a normal distribution. The Wilcoxon signed-rank test and the Mann–Whitney U test were used for variables with a non-normal distribution. The McNemar test for paired proportions was used. Statistical analysis was performed using the SPSS software (version 25.0). A There were 26 subjects  were enrolled in the study with a mean age of 51.4 ± 21.6 years.p = 0.02, p = 0.05). In 35–38% of the patients, the eye drops fell either on the eyelids or on the cheek when using the two aids or the conventional bottle.The objective and subjective measurements of the two aids and the conventional bottle for eye drop instillation are shown in p = 0.0005). More subjects preferred Autodrop over Opticare (42% vs. 27%), but this was not significant (Regarding the risk of contamination (defined as touch of the bottle tip with the eyelids or an eye structure), no subjects using either the Autodrop or the Opticare had the bottle tip touch their eye, while 46% of subjects had contact with their canthus, eyelid, cornea or conjunctiva with the tip when using the conventional bottle without an aid  . They haThe participants in the current study were healthy people without arthritis, neurological disabilities or other diseases that might compromise drop instillation. Accordingly, our results may not be applicable to patients with systemic diseases that might compromise drop instillation. Future studies should include patients with physical limitations and also compare experienced versus nonexperienced patients in eye drop instillation. In addition, in this study, the participants had two attempts to use the eye drops aids before the experiment initiation. It is reasonable to assume that with continued use of the devices, the patients will better use them, with enhanced accuracy in administration. Our results can be utilized to assess the ease of acquiring the skill of using the eye drop aids but cannot be easily generalized for all future uses of the aids. Repeating the experiment either immediately or after a set amount of time and examining whether the results have changed would help make the results more generalizable. Based on previous studies ,20, the In conclusion, aids to administer eye drops such as Autodrop and Opticare were found to reduce the risk of bottle tip contamination. Assemble of the eye drops bottle into the aid was easy in both aids. These aids could have an advantage, especially in patients who had subconjunctival hemorrhage or corneal erosion from the bottle tip or had an eye infection that was attributed to the tip touching the eye.