scholarly journals Lipidomics of facial sebum in the comparison between acne and non-acne adolescents with dark skin

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Obumneme Emeka Okoro ◽  
Adebomi Adenle ◽  
Matteo Ludovici ◽  
Mauro Truglio ◽  
Federico Marini ◽  
...  
Keyword(s):  
Relative Abundance ◽  
Building Blocks ◽  
Carbon Chain ◽  
Gas Chromatography Mass Spectrometry ◽  
Lower Percentage ◽  
Void Space ◽  
Straight Chain ◽  
Dark Skin ◽  
Cross Sectional ◽  
Weight Percentage

AbstractLipidomics is advantageous in the study of sebum perturbations occurring in acne. An extended evaluation of the sebum lipid profiles in acne-prone sebaceous areas is lacking in dark skin. Yet, there is a void space in understanding how the building blocks of sebum lipids, i.e. individual fatty acids (FAs), are intertwined with acne-prone skin. We aimed to determine the sebum lipidome in facial areas of adolescents with and without acne in Nigeria. A cross-sectional analytical study was conducted in 60 adolescents/young adults divided in 30 acne patients (15F, 15M) and 30 age and sex-matched controls. Sebum samples obtained from foreheads and cheeks were analysed separately by gas chromatography–mass spectrometry (GCMS) and thin layer chromatography (HPTLC). Distributions of sebum components were investigated with multivariate ANOVA-simultaneous component analysis (ASCA). Sebum incretion in acne was paralleled by significantly higher abundance of triglycerides, wax esters, and squalene together with monounsaturated FAs (MUFAs), and straight chain saturated FAs (SFAs), especially those with odd-carbon chain, i.e. C13:0, C15:0, and C17:0. Profiling weight/weight percentage of individual components revealed that, in acne, the free FAs (FFAs) array was shifted towards higher relative abundance of the SFAs C15:0, C16:0, and C17:0 and lower percentage of the anteiso-branched FFAs with 12, 14, 16, and 18 carbons. In acne patients, MUFAs and PUFAs were quantitatively increased and decreased on foreheads and cheeks, respectively. Relative abundance of fatty alcohols was decreased in acne independent on the site. The results indicated that acne associates with site-specific derangement of the pathways regulating the balance among odd straight-chain and branched-chain SFAs, MUFAs, which included sapienate (C16:1n-10), PUFAs, and squalene.

scholarly journals Lipidomics of Facial Sebum in the Comparison Between Acne and Non-Acne Adolescents with Dark Skin

2021 ◽  
Author(s):  
Emeka Okoro ◽  
Adebomi Adenle ◽  
Matteo Ludovici ◽  
Mauro Truglio ◽  
Federico Marini ◽  
...  
Keyword(s):  
Relative Abundance ◽  
Building Blocks ◽  
Carbon Chain ◽  
Gas Chromatography Mass Spectrometry ◽  
Lower Percentage ◽  
Void Space ◽  
Straight Chain ◽  
Dark Skin ◽  
Cross Sectional ◽  
Weight Percentage

Abstract Lipidomics is advantageous in the study of sebum perturbations occurring in acne. An extended evaluation of the sebum lipid profiles in acne-prone sebaceous areas is lacking in dark skin. Yet, there is a void space in understanding how the building blocks of sebum lipids, i.e. individual fatty acids (FAs), are intertwined with acne-prone skin. We aimed to determine the sebum lipidome in facial areas of adolescents with and without acne in Nigeria. A cross-sectional analytical study was conducted in 60 adolescents/young adults divided in 30 acne patients (15F, 15M) and 30 age and sex-matched controls. Sebum samples obtained from foreheads and cheeks were analysed separately by gas chromatography–mass spectrometry (GCMS) and thin layer chromatography (HPTLC). Distributions of sebum components were investigated with multivariate ANOVA-simultaneous component analysis (ASCA). Sebum incretion in acne was paralleled by significantly higher abundance of triglycerides, wax esters, and squalene together with monounsaturated FAs (MUFAs), and straight chain saturated FAs (SFAs), especially those with odd-carbon chain, i.e. C13:0, C15:0, and C17:0. Profiling weight/weight percentage individual components revealed that in acne the free FAs (FFAs) array was shifted towards higher relative abundance of the SFAs C15:0, C16:0, and C17:0 and lower percentage of the anteiso-branched FFAs with 12, 14, 16, and 18 carbons. In acne patients, MUFAs and PUFAs were quantitatively increased and decreased on foreheads and cheeks, respectively. Relative abundance of fatty alcohols was decreased in acne independent on the site. The results indicated that acne associates with site-specific derangement of the pathways regulating the balance among odd straight-chain and branched-chain SFAs, MUFAs, including sapienate (C16:1n-10), PUFAs, and squalene.

Physical restraint to patients with dementia in acute physical care settings: effect of the financial incentive to acute care hospitals

2017 ◽  
Vol 30 (7) ◽  
pp. 991-1000 ◽  
Author(s):  
Miharu Nakanishi ◽  
Yasuyuki Okumura ◽  
Asao Ogawa
Keyword(s):  
Acute Care ◽  
Financial Incentive ◽  
Physical Restraint ◽  
Acute Care Hospitals ◽  
Dementia Care ◽  
Lower Percentage ◽  
Public Healthcare ◽  
Cross Sectional ◽  
Insurance Program ◽  
Healthcare Insurance

ABSTRACTBackground:In April 2016, the Japanese government introduced an additional benefit for dementia care in acute care hospitals (dementia care benefit) into the universal benefit schedule of public healthcare insurance program. The benefit includes a financial disincentive to use physical restraint. The present study investigated the association between the dementia care benefit and the use of physical restraint among inpatients with dementia in general acute care settings.Methods:A national cross-sectional study design was used. Eight types of care units from acute care hospitals under the public healthcare insurance program were invited to participate in this study. A total of 23,539 inpatients with dementia from 2,355 care units in 937 hospitals were included for the analysis. Dementia diagnosis or symptoms included any signs of cognitive impairment. The primary outcome measure was “use of physical restraint.”Results:Among patients, the point prevalence of physical restraint was 44.5% (n= 10,480). Controlling for patient, unit, and hospital characteristics, patients in units with dementia care benefit had significantly lower percentage of physical restraint than those in any other units (42.0% vs. 47.1%; adjusted odds ratio, 0.76; 95% confident interval [0.63, 0.92]).Conclusions:The financial incentive may have reduced the risk of physical restraint among patients with dementia in acute care hospitals. However, use of physical restraint was still common among patients with dementia in units with the dementia care benefit. An educational package to guide dementia care approach including the avoidance of physical restraint by healthcare professionals in acute care hospitals is recommended.

scholarly journals Taxonomic and Functional Characteristics of the Gill and Gastrointestinal Microbiota and Its Correlation with Intestinal Metabolites in NEW GIFT Strain of Farmed Adult Nile Tilapia (Oreochromis niloticus)

2021 ◽  
Vol 9 (3) ◽  
pp. 617
Author(s):  
Zhenbing Wu ◽  
Qianqian Zhang ◽  
Yaoyao Lin ◽  
Jingwen Hao ◽  
Shuyi Wang ◽  
...  
Keyword(s):  
Relative Abundance ◽  
Oreochromis Niloticus ◽  
Nile Tilapia ◽  
High Throughput Sequencing ◽  
Gas Chromatography Mass Spectrometry ◽  
Gastrointestinal Mucosa ◽  
Gastrointestinal Microbiota ◽  
Linear Discriminant ◽  
Intestinal Microbes ◽  
Nile Tilapia Oreochromis Niloticus

The gill and gastrointestinal tract are primary entry routes for pathogens. The symbiotic microbiota are essential to the health, nutrition and disease of fish. Though the intestinal microbiota of Nile tilapia (Oreochromis niloticus) has been extensively studied, information on the mucosa-associated microbiota of this species, especially the gill and gastrointestinal mucosa-associated microbiota, is lacking. This study aimed to characterize the gill and gastrointestinal mucosa- and digesta-associated microbiota, as well as the intestinal metabolite profiles in the New Genetically Improved Farmed Tilapia (NEW GIFT) strain of farmed adult Nile tilapia by high-throughput sequencing and gas chromatography/mass spectrometry metabolomics. The diversity, structure, composition, and predicted function of gastrointestinal microbiota were significantly different across gastrointestinal regions and sample types (Welch t-test; p < 0.05). By comparing the mucosa- and digesta-associated microbiota, linear discriminant analysis (LDA) effect size (LEfSe) analysis revealed that Pelomonas, Ralstoniapickettii, Comamonadaceae, and Staphylococcus were significantly enriched in the mucosa-associated microbiota, whereas many bacterial taxa were significantly enriched in the digesta-associated microbiota, including Chitinophagaceae, Cetobacterium, CandidatusCompetibacter, Methyloparacoccus, and chloroplast (LDA score > 3.5). Furthermore, Undibacterium, Escherichia–Shigella, Paeniclostridium, and Cetobacterium were dominant in the intestinal contents and mucosae, whereas Sphingomonasaquatilis and Roseomonasgilardii were commonly found in the gill and stomach mucosae. The Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt2) analysis revealed that the predictive function of digesta-associated microbiota significantly differed from that of mucosa-associated microbiota (R = 0.8152, p = 0.0001). In addition, our results showed a significant interdependence between specific intestinal microbes and metabolites. Notably, the relative abundance values of several potentially beneficial microbes, including Undibacterium, Crenothrix, and Cetobacterium, were positively correlated with most intestinal metabolites, whereas the relative abundance values of some potential opportunistic pathogens, including Acinetobacter, Mycobacterium, Escherichia–Shigella, Paeniclostridium, Aeromonas, and Clostridiumsensustricto 1, were negatively correlated with most intestinal metabolites. This study revealed the characteristics of gill and gastrointestinal mucosa-associated and digesta-associated microbiota of farmed Nile tilapia and identified a close correlation between intestinal microbes and metabolites. The results serve as a basis for the effective application of targeted probiotics or prebiotics in the diet to regulate the nutrition and health of farmed tilapia.

scholarly journals Gut Microbiome Composition Remains Stable in Individuals with Diabetes-Related Early to Late Stage Chronic Kidney Disease

Biomedicines ◽  
2020 ◽  
Vol 9 (1) ◽  
pp. 19
Author(s):  
Ashani Lecamwasam ◽  
Tiffanie M. Nelson ◽  
Leni Rivera ◽  
Elif I. Ekinci ◽  
Richard Saffery ◽  
...  
Keyword(s):  
Chronic Kidney Disease ◽  
Kidney Disease ◽  
Relative Abundance ◽  
Cross Sectional Study ◽  
Early Event ◽  
Sectional Study ◽  
Cross Sectional ◽  
Microbiome Composition ◽  
Stool Samples ◽  
Late Stages

(1) Background: Individuals with diabetes and chronic kidney disease display gut dysbiosis when compared to healthy controls. However, it is unknown whether there is a change in dysbiosis across the stages of diabetic chronic kidney disease. We investigated a cross-sectional study of patients with early and late diabetes associated chronic kidney disease to identify possible microbial differences between these two groups and across each of the stages of diabetic chronic kidney disease. (2) Methods: This cross-sectional study recruited 95 adults. DNA extracted from collected stool samples were used for 16S rRNA sequencing to identify the bacterial community in the gut. (3) Results: The phylum Firmicutes was the most abundant and its mean relative abundance was similar in the early and late chronic kidney disease group, 45.99 ± 0.58% and 49.39 ± 0.55%, respectively. The mean relative abundance for family Bacteroidaceae, was also similar in the early and late group, 29.15 ± 2.02% and 29.16 ± 1.70%, respectively. The lower abundance of Prevotellaceae remained similar across both the early 3.87 ± 1.66% and late 3.36 ± 0.98% diabetic chronic kidney disease groups. (4) Conclusions: The data arising from our cohort of individuals with diabetes associated chronic kidney disease show a predominance of phyla Firmicutes and Bacteroidetes. The families Ruminococcaceae and Bacteroidaceae represent the highest abundance, while the beneficial Prevotellaceae family were reduced in abundance. The most interesting observation is that the relative abundance of these gut microbes does not change across the early and late stages of diabetic chronic kidney disease, suggesting that this is an early event in the development of diabetes associated chronic kidney disease. We hypothesise that the dysbiotic microbiome acquired during the early stages of diabetic chronic kidney disease remains relatively stable and is only one of many risk factors that influence progressive kidney dysfunction.

scholarly journals Gut Microbiome and Metabolome Profiles Associated with High-Fat Diet in Mice

Metabolites ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 482
Author(s):  
Jae-Kwon Jo ◽  
Seung-Ho Seo ◽  
Seong-Eun Park ◽  
Hyun-Woo Kim ◽  
Eun-Ju Kim ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Relative Abundance ◽  
High Fat Diet ◽  
Amplicon Sequencing ◽  
Gas Chromatography Mass Spectrometry ◽  
Control Group ◽  
Rrna Gene ◽  
High Fat ◽  
Underlying Mechanisms ◽  
Insight Into

Obesity can be caused by microbes producing metabolites; it is thus important to determine the correlation between gut microbes and metabolites. This study aimed to identify gut microbiota-metabolomic signatures that change with a high-fat diet and understand the underlying mechanisms. To investigate the profiles of the gut microbiota and metabolites that changed after a 60% fat diet for 8 weeks, 16S rRNA gene amplicon sequencing and gas chromatography-mass spectrometry (GC-MS)-based metabolomic analyses were performed. Mice belonging to the HFD group showed a significant decrease in the relative abundance of Bacteroidetes but an increase in the relative abundance of Firmicutes compared to the control group. The relative abundance of Firmicutes, such as Lactococcus, Blautia, Lachnoclostridium, Oscillibacter, Ruminiclostridium, Harryflintia, Lactobacillus, Oscillospira, and Erysipelatoclostridium, was significantly higher in the HFD group than in the control group. The increased relative abundance of Firmicutes in the HFD group was positively correlated with fecal ribose, hypoxanthine, fructose, glycolic acid, ornithine, serum inositol, tyrosine, and glycine. Metabolic pathways affected by a high fat diet on serum were involved in aminoacyl-tRNA biosynthesis, glycine, serine and threonine metabolism, cysteine and methionine metabolism, glyoxylate and dicarboxylate metabolism, and phenylalanine, tyrosine, and trypto-phan biosynthesis. This study provides insight into the dysbiosis of gut microbiota and metabolites altered by HFD and may help to understand the mechanisms underlying obesity mediated by gut microbiota.

scholarly journals Comparing dental plaque microbiome diversity of extrinsic black stain in the primary dentition using Illumina MiSeq sequencing technique

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Lulu Chen ◽  
Qiong Zhang ◽  
Yan Wang ◽  
Keke Zhang ◽  
Jing Zou
Keyword(s):  
Relative Abundance ◽  
Dental Plaque ◽  
Illumina Miseq ◽  
Primary Dentition ◽  
Tooth Surface ◽  
Oral Microbiota ◽  
Illumina Miseq Sequencing ◽  
Miseq Sequencing ◽  
Cross Sectional ◽  
Sequencing Technique

Abstract Background Extrinsic black stain (EBS) is characterized by discrete dark dots or lines on the tooth surface. The relationship between EBS and oral microbiota in children remains elusive. The aim of this study was to compare dental plaque microbiome in EBS children with that in EBS-free children in the primary dentition. Methods The Illumina MiSeq sequencing technique was utilized in the cross-sectional pilot study to investigate the diversity and composition of the supragingival plaque microbiota from 10 EBS-positive and 10 EBS-free children. The results were analysed with nonparametric Mann-Whitney U test, Pearson Chi-Square test, Fisher’s Exact test and one-way ANOVA tests. Results We identified 13 different phyla, 22 classes, 33 orders, 54 families, 105 genera, and 227 species from a total of 52,646 high-quality sequences. Between two groups, no statistical differences were observed in the estimators of community richness and diversity at 97% similarity, as well as in the Unweighted Unifrac principal co-ordinates analysis (PCoA). At the species level, higher level of relative abundance of Actinomyces naeslundii and lower level of relative abundance of a species belonging to Candidate_division_TM7 was observed in dental plaque of EBS-positive subjects, compared to dental plaque of EBS-free subjects (P < 0.05). This indicated that some species might be involved in the EBS process. Conclusion Changes in dental plaque microbiota is possibly relevant to the process of EBS in the primary dentition.

scholarly journals Dysbiosis of Gut Microbiota and Its Relationship with the Regulatory T Cells in Patients with Aplastic Anemia 

2021 ◽  
Author(s):  
Mengxiao Ren ◽  
Yongqin Ge ◽  
Jindan Qi ◽  
Shengli Xue ◽  
Miao Miao ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Aplastic Anemia ◽  
Treg Cell ◽  
Relative Abundance ◽  
Treg Cells ◽  
Cross Sectional Survey ◽  
Cross Sectional ◽  
Cell Counts ◽  
And Control ◽  
Healthy Participants

Abstract Background: The characteristics of gut microbiota (GM) and its relationship with the Regulatory T Cells (Treg) remains unclear in patients with aplastic anemia (AA). Methods: This study was a cross-sectional survey which included 12 AA patients consisted of 6 with severity aplastic anemia (SAA) and 6 with non-severity aplastic anemia (NSAA) and 6 healthy participants. The GM and its relationship with the Treg cells of AA patients were analyzed. Results: The results showed that the presence of compositional differences in the GM structure between the AA and Control groups. The bacterial communities were depleted of Clostridia class (e.g., Lachnospiraceae ND3007, Lachnospiraceae XPB1014, Lachnolostridium, Ruminococcaceae UCG 013 and Butyricicoccus genus) in AA group, especially in SAA group. Inversely, the relative abundance of Lactobacillus and Streptococcus genus from Bacilli class were increased significantly in patients with SAA. The relative abundance of Lachnospiraceae (r=0.663, p=0.029), Clostridiaceae 1 (r=0.619, p=0.042) and Clostridiales vadinBB60 group family (r=0.674, p=0.023) which from Clostridia class, were positively correlated with the Treg cell counts. Conclusion: We speculated that the decrease of some bacteria from Clostridia class may participate in the pathophysiological process of AA through reducing the Treg cell counts. Notwithstanding the low sample size, our data provided some clues that the treatment strategy of AA could start by adjusting the imbalance of GM, increasing Treg cell counts to improve the suppression of bone marrow hematopoiesis.

scholarly journals The estimated effect of graphic warning labels on smoker’s intention to quit in Shanghai, China: a cross-sectional study

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Ruiping Wang ◽  
Yan Qiang ◽  
Yan Zhu ◽  
Xiangjin Gao ◽  
Qiong Yang ◽  
...  
Keyword(s):  
Smoking Cessation ◽  
Logistic Regression ◽  
Control Measures ◽  
Intention To Quit ◽  
Lower Percentage ◽  
Warning Labels ◽  
Cross Sectional ◽  
Smoking Intensity ◽  
Graphic Warning ◽  
Cigarette Packs

Abstract Background Tobacco consumption is the leading cause of death worldwide. Overwhelming studies demonstrate graphic warning labels (GWLs) on cigarette packs are effective in eliciting negative response to tobacco smoking, modifying beliefs about tobacco dangers, and increasing reported intention to quit, but the estimated effect of GWLs on smoking cessation intention among smokers is still limited in China. In this study, we aim to understand the smoking intensity, smoking duration and smoking cessation intention among current smokers, and to explore how their smoking cessation intention would be influenced by the GWLs in Shanghai. Methods From January to June 2021, we totally recruited 1104 current smokers in Songjiang district and Fengxian district of Shanghai by multistage sampling design. We used Android pad assisted electronic questionnaire for data collection, and then implemented logistic regression for odds ratio (OR) and 95% confidence interval (CI) calculation to explore how smoking cessation intention would be influenced by the GWLs among current smokers. Results One thousand one hundred four current smokers included 914 males (82.79%), with an average age of 43.61 years. 58.06% of current smokers reported smoking cessation intention due to GWLs. Logistic regression indicated a higher percentage of smoking cessation intention due to GWLs was among female smokers [OR = 2.41, 95% CI (1.61–3.59)], smokers with smoking intensity < 20 cigarette/day [OR = 1.92, 95% CI (1.44–2.55)], smokers with tobacco burden < 20% [OR = 1.94, 95% CI (1.35–2.79)], and among smokers had plan to quit in a year [OR = 6.58, 95% CI (4.71–9.18). Smokers with higher individual monthly income had lower percentage of smoking cessation intention (OR were 0.35, 0.46 and 0.41). Meanwhile, among 642 current smokers without plan to quit in a year, approximately 40% of them reported smoking cessation intention due to GWLs. Conclusions Smoking cessation intention due to the assumed GWLs on cigarette packs is high among current smokers in Shanghai, especially in female smokers, smokers with light tobacco burden and mild nicotine dependence. Incorporating smoking intensity as well as smoking burden into the implementation of GWLs as tobacco control measures would discourage smoking in China.

Can we reduce psychosomatic symptoms and work presenteeism with physical activity?

2021 ◽  
pp. 1-11
Author(s):  
Andreja Petrovčič ◽  
Vanja Erčulj ◽  
Eva Boštjančič
Keyword(s):  
Physical Activity ◽  
Cost Effective ◽  
Cross Sectional Study ◽  
Lower Percentage ◽  
Cross Sectional ◽  
Effective Measure ◽  
Psychosomatic Symptoms ◽  
General Evaluation ◽  
Patient Health ◽  
The Relationship

BACKGROUND: Past studies have tended to consider the relationships between presenteeism, physical activity and psychosomatic symptoms in a partial manner and not as a whole. OBJECTIVE: This cross-sectional study investigated the relationship between physical activity, psychosomatic symptoms and presenteeism, in order to increase the visibility of presenteeism and promote workplace health. METHODS: Two questionnaires (Patient Health Questionnaire and Work Limitations Questionnaire) and a set of physical activity questions were administered on the sample of 514 employees. RESULTS: The results showed a statistically significant relationship between presenteeism and some variables of physical activity, as well as between presenteeism and psychosomatic symptoms. Further results showed a statistically significant association between psychosomatic symptoms and some variables of physical activity. General evaluation (adequacy) of physical activity, and general, aerobic physical activity proved to be important predictors of presenteeism, while general, aerobic physical activity also predicted psychosomatic symptoms. Analysis of the results revealed that psychosomatic symptoms moderate the relationship between physical activity and presenteeism. CONCLUSIONS: The article points out that employees with health problems have a lower percentage of productive time than healthy employees. However, developing an active lifestyle can be a cost-effective measure to cope with presenteeism and promote psychophysical health among employees.

scholarly journals Associations of tobacco smoking with body mass distribution; a population-based study of 65,875 men and women in midlife

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Sidsel Graff-Iversen ◽  
Stephen Hewitt ◽  
Lisa Forsén ◽  
Liv Grøtvedt ◽  
Inger Ariansen
Keyword(s):  
Cardiovascular Risk ◽  
Smoking Status ◽  
Health Indicators ◽  
Population Based ◽  
Lower Percentage ◽  
Body Region ◽  
Cross Sectional Survey ◽  
Cross Sectional ◽  
Men And Women ◽  
Never Smokers

Abstract Background Studies indicate an effect of smoking toward abdominal obesity, but few assess hip and waist circumferences (HC and WC) independently. The present study aimed to assess the associations of smoking status and volume smoked with HC and WC and their ratio in a population with low prevalence of obesity together with high prevalence of smoking. Methods We used cross-sectional survey data from 11 of a total 19 Norwegian counties examined in 1997–99 including 65,875 men and women aged 39–44 years. Analysis of associations were adjusted for confounding by socioeconomic position, health indicators, and additionally for BMI. Results Compared with never-smokers, when adjusting for confounders and in addition for BMI, mean HC remained lower while mean WC and waist-hip-ratio (WHR) were higher in current smokers. The finding of a lower HC and higher WHR level among smokers was consistent by sex and in strata by levels of education and physical activity, while the finding of higher WC by smoking was less consistent. Among current smokers, BMI-adjusted mean HC decreased whereas WC and WHR increased by volume smoked. Compared with current smokers, former smokers had higher BMI-adjusted HC, lower WHR and among women WC was lower. Conclusions The main finding in this study was the consistent negative associations of smoking with HC. In line with the hypothesis that lower percentage gluteofemoral fat is linked with higher cardiovascular risk, our results suggest that smoking impacts cardiovascular risk through mechanisms that reduce the capacity of fat storage in the lower body region.

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