Streptococcus thermophilus and Lactobacillus delbrueckii; S2) vs. three  strain-supplemented yogurt.The consumption of probiotics may influence children’s gut microbiome and metabolome, which may reflect shifts in gut microbial diversity composition and metabolism. These potential changes might have a beneficial impact on health. However, there is a lack of evidence investigating the effect of probiotics on the gut microbiome and metabolome of children. We aimed to examine the potential impact of a two  probiotic strains for 10 days. Nevertheless, we observed a significant increase (Day 0 to Day 10) in the relative abundance of the two and three probiotics administered in the S2 and S2 + BB12 groups, respectively, indicating the intervention had a measurable impact on the bacteria of interest in the gut microbiome. Future research using longer probiotic intervention durations and in children at risk for gastrointestinal disorders may elucidate if functional metabolite changes confer a protective gastrointestinal effect.In conclusion, there were were no significant differences in the global metagenomic or metabolomic profiles between healthy children receiving two (S2)  Metagenomic analysis captures a comprehensive summary of the microbiome, unction) . Gut colunction) . Severalunction) , type ofunction) , and envunction) , shape tunction) . The humunction) . Pre-cliunction) . Disruptunction) .Probiotics are defined by the International Scientific Association for Probiotics and Prebiotics as “live microorganisms that, when administered in adequate amounts, confer a health benefit on the host .” ProbioBifidobacterium genus promote the growth of beneficial bacteria, inhibit pathogenic microorganisms by secreting antibacterial factors  is among the most common probiotic supplements and has previously been demonstrated to be well-tolerated by healthy children   .Streptococcus thermophilus and Lactobacillus delbrueckii [S2] vs.Streptococcus thermophilus, Lactobacillus delbrueckii, and Bifidobacterium animalis subsp. lactis strain BB-12 [S2 + BB12]) in children is an important first step to design future research evaluating the efficacy of probiotics in the prevention of dysbiosis–associated GI disorders.Exploring the effect of probiotics on the gut microbiome and metabolome in healthy children may provide more extensive insight into the relationship between probiotics, gut microbiota, metabolites, and human health. This may aid in developing more effective methods of assessing gut health by simultaneously characterizing the gut microbiome and functional impacts of microbiome community changes on the metabolome. Furthermore, GI disorders are among the most common ailments reported in pediatric primary care. Therefore, utilizing our approach including microbiome and metabolome analyses to characterize structural and functional responses of gut microbiota to two probiotics combinations  in healthy children aged one to five years old. This study builds upon findings by . The yogurt was administered to the participants for ten consecutive days. Fecal samples were analyzed at baseline (Day 0), ten (Day 10), and 30 (Day 30) days from both S2 + BB12  and S2  groups  were included in the study . The yog) groups .n = 28 males and n = 31 females). Most of the participants included were White (n = 40). Additional demographic information from the included participants is displayed in Participants were relatively divided equally between males and females , but Shannon diversity was significantly lower in the S2 + BB12 group compared to the S2 group . These findings agree with previously reported results by The administration of BB-12, in addition to B. animalis in the S2 + BB12 group. B. animalis abundance was significantly impacted by the administration of the BB12 supplemented yogurt . S. thermophilus’s relative abundance were significantly different in time and group × time interaction  , S. thermophilus (p < 0.001), and L. delbrueckii (p = 0.018) abundances. Similarly, Day 10 vs. Day 30 comparison showed a statistically significant decrease in BB-12 (p < 0.001), S. thermophilus (p = 0.001), and L. delbrueckii (p = 0.004) abundances (vs. Day 10 comparison of S. thermophilus (p < 0.001) and L. delbrueckii (p = 0.005) abundances. Interestingly, BB-12 increased comparing Day 0 to Day 30 in the same group (p = 0.016).We built upon these findings by using shotgun metagenomics, which allowed us to observe differential abundant taxa at the species level. As expected, the metagenomic analysis showed an increase of = 0.020) ,F. As exundances –F. In thvs. S2 + BB12) and were healthy children. The 25 taxa with the greatest change in relative abundance between Day 0 and Day 10 were evaluated to observe the effect on individual microbial taxa following a ten-day administration of two (S2) vs. three (S2 + BB12) strains of probiotics. S. thermophilus, one of the intervention bacteria, was significantly increased in the S2 group at Day 10 compared to Day 0  were significantly increased at Day 10 , but the S2 + BB12 intervention did not significantly influence the abundance of any other taxa when FDR correction was applied  and B. pseudocatenulatum was decreased in non-Hispanic children , msbA; ATP-binding cassette, subfamily B, bacterial MsbA, sstT; serine/threonine transporter, and chondroitin-sulfate-ABC endolyase/exolyase (p = 0.025) and aspartyl-tRNA synthetase increased  probiotic strains would influence the abundance other known probiotics. Most of the identified probiotic-associated taxa belonged to Lactobacillus, Bacteroides, and Bifidobacterium genera, and were commonly used as probiotic supplements in the food industry or clinical trials . After ten days of yogurt consumption, we found an increase of N-acetylvaline  and a decrease of arachidoylcarnitine (C20) * . Day 0 vs. Day 30 comparison exhibited several changes in lipids decreased at Day 30 belonging to diacyglicerol metabolism: linoleoyl-linoleoyl-glycerol (18:2/18:2) , palmitoyl-linoleoyl-glycerol (16:0/18:2) , palmitoyl-linoleoyl-glycerol (16:0/18:2) , oleoyl-linoleoyl-glycerol (18:1/18:2) , palmitoyl-oleoyl-glycerol (16:0/18:1) , and oleoyl-linoleoyl-glycerol  . The glycerolipids 2-palmitoyl-galactosylglycerol (16:0)*  and 1-palmitoyl-galactosylglycerol * (FDR p = 0.166 FC = 2.319), were increased together with 3-hydroxybutyrate (BHBA) .After excluding xenobiotics, we conducted exploratory metabolomic analyses on 601 biochemicals. We analyzed the differences between Day 0 2 groups . After c30 in S2 ,C,E and 30 in S2 ,D,F grou:2/18:2) * (FDR p :0/18:2) * (FDR p :0/18:2) * (FDR p :1/18:2)  (FDR p =:0/18:1) *   (FDR p =vs. Day 10)  and 20 days after yogurt consumption discontinuation (Day 0 vs. Day 30) . Lactate and indolelactate were also decreased looking at Day 0 vs. Day 30 comparison . PCA of untargeted metabolites showed no separation between the time points in any group -salsolinol was decreased after ten days of intervention (Day 0 ny group .p = 0.004), 3-methyl-2-oxovalerate (p = 0.025), alanine (p = 0.001), glutamate (p = 0.034), isoleucine (p = 0.020), and valine (p = 0.004), among others. Conversely, glycine (p = 0.025), indolelactate (p = 0.034), N-acetylserine (p = 0.049), and pyroglutamine (p = 0.007) had significant group × time effects as a result of the S2 and S2 + BB12 probiotic interactions (vs. baseline (H), while fecal cysteine s-sulfate levels increased back to baseline levels after probiotic discontinuation.Next, we focused on evaluating differences in specific metabolites associated with amino acid metabolism or biosynthesis, as we hypothesized that they would be impacted by probiotic intervention and associated with the gut microbiome in both groups. There were several amino-acid associated metabolites that differed as a result of time in both groups  includinractions . In the ractions –N. Alanibaseline ,D,H, whiMicrobe-metabolite interactions were tested through correlation matrices and visualized with network graphs. After filtering, 26 microbial taxa and 79 metabolites were tested for associations , S8.Enterobacteriaceae and Escherichia coli included trimethylamine N-oxide (TMAO), ursodeoxycholate, 7-ketodeoxycholate, and glycine , pantothenate (vitamin B5) correlated positively with Clostridiales, R. hominis, F. prausnitzii, and E. eligens . Uracil was positively associated with Clostridiales, A. finegoldii, and E. rectale. Uracil was also negatively correlated with B. dorei, F. plautii, Enterobacteriaceae and E. coli.Several amino acids correlated negatively with and TMAO . Lactatevs. three (S2 + BB12) probiotic strains in healthy children. We studied probiotics consumption effects on the fecal microbiome and metabolome comparing baseline, post-ten days (Day 10), and post-30 days (Day 30) following yogurt administration. There were no significant differences in the global metagenomic or metabolomic profiles between healthy children receiving two (S2) vs. three (S2 + BB12) probiotic strains for 10 days. Nevertheless, we observed a significant increase (Day 0 to Day 10) in the relative abundance of the two and three probiotics administered in the S2 and S2 + BB12 groups, respectively, indicating the intervention had a measurable impact on the bacteria of interest in the gut microbiome. Interestingly, in the S2 group, S. thermophilus species appear to maintain a sustained increase over time, maintaining the significant increase in relative abundance from Day 0 vs. Day 30 As both cohorts of children received a probiotic cocktail and the intervention lasted over a short period of time, the lack of strong association between groups in global microbiome changes is not surprising and suggests a resilient and stable gut microbiota in this cohort of healthy children ages one to five. Nevertheless, existing literature reports about infant’s and children’s gut microbiota stability are conflicting  flicting . Infant flicting  our dataflicting .Most studies examining the health benefits of probiotics have focused on people with pathologies. However, few studies have examined the effects of probiotics on healthy individuals and even fewer have examined the use of probiotics in children. A review of probiotic supplementation in healthy adults found that probiotic supplementation led to a transient increase in the concentration of supplement-specific bacteria but failed to support the ability of probiotics to cause persistent changes in gut microbiota . This isBifidobacterium and Lactobacillus genera, commonly used probiotics. The combined effects of these bacteria could have more functional implications and protective effects than individual bacterial changes supporting the use of probiotics to maintain a healthy gut ecosystem in hopes to prevent gastrointestinal disorders in children, although more research is needed to directly test and validate this notion. Importantly, as this study was conducted on healthy children, these findings should not be extrapolated to children with gastrointestinal disorders or subjects seeking intervention for other diseases or disorders.As yogurts and other dairy products commonly supplemented with probiotics often possess other beneficial characteristics, such as a high calcium content, these properties of probiotics could allow consumers to benefit from the nutritional components without risking disruption to their microbiota and health. This potential beneficial shift to probiotic-associated bacteria that we demonstrated in the S2 + BB12 group  supportsBifidobacterium genus were highly abundant in both groups at the three time points. Bifidobacteria are highly represented in children, as they use milk oligosaccharides as a carbon source and to restrict human milk oligosaccharides availability to other microorganisms  and in the control group S2 . Additionally, this metagenomic approach adds to the existing literature of the genomic potential of the microbial community underlying microbiome-host interactions.Species belonging to rganisms . In addiA. finegoldii, B. animalis, and E. rectale. B. dorei, F. plautii, Enterobacteriaceae family, and E. coli were negatively correlated with uracil. During infections, the immune response triggered by uracil promotes pathogen bacteria elimination, intestinal cell repair, and host homeostasis  and microorganisms. Uracil was positively associated with Clostridiales family, eostasis . Thymine in vivo . Deoxycaeability . This suA strength of our study is that it was a randomized, blinded controlled trial conducted on children ages one to five years old, an age group that is rarely studied in probiotic research. Additionally, we incorporated and integrated both metagenomic and metabolomic analyses to characterize the effect of BB-12 supplemented yogurt on the children’s gut-microbiome and metabolism. This unique integration allowed us to test more system-wide and functional effects on the gut microbiome as a result of two probiotic interventions, providing more comprehensive data on gut health. However, this study examined short-term changes of probiotics over a 10-day period, and more studies should be conducted to investigate the long-term effects on probiotic consumption in this age group. An additional limitation is that we did not incorporate information of dietary intake in the current analysis, as it was not collected in the original study procedures as described by More studies are needed to elucidate the mechanistic pathways by which probiotics such as BB-12 can affect mucosal barrier functions and innate immunity. Future studies should expand upon the findings presented in this double-blinded, randomized controlled trial and examine the interplay of diet and probiotics on metabolites and microbiota in children. Additionally, further research is needed to investigate environmental factors that influence the impacts of probiotics on children’s health status and behavior. The long-term supplementation BB-12 on this population and its longitudinal effects during development should also be examined. This would allow for more in-depth knowledge of the impact that probiotics have on gut microbial communities in developing children as they age. Additionally, future research in children at risk for gastrointestinal disorders may elucidate if these functional metabolite changes as a result of S2 + BB12 probiotic intervention confer a protective gastrointestinal effect.B. animalis and S. thermophilus) in study subjects receiving the based S2 probiotic intervention + BB12, but no individual taxonomic changes occurred in the S2 only group. Finally, although we did not see global fecal metabolome response to either probiotic, several fecal metabolites were increased in the S2 + BB12 group, indicating a net functional impact of the addition of BB12 to the probiotic intervention. Future research replicating these results across different patient populations will confirm the therapeutic use of BB12 as a probiotic intervention to exert beneficial impacts on the pediatric gastrointestinal system.In conclusion, the results from this deep metagenomic and metabolomic characterization of the gut microbiome and metabolome of children following BB-12 consumption did not show statistically significant differences between the groups, although net positive emergent property effects were witnessed in the S2 + BB12 group over time. The functional redundancy in healthy microbial systems and metabolic stability reflects no changes in the microbial diversity, although we did observe a separation effect in the S2 + BB12 group as a result of the three-strain intervention when we focused on probiotic-associated and beneficial bacteria reported the literature. Our study validated previous results from ® or control yogurt drink by family cluster. The study protocol was approved by the Georgetown University Institutional Review Board . The independent Data Safety Monitoring Board reviewed the protocol before study initiation and checked adverse event data at approximately 33, 50 and 66% data completion. Additional monitoring was conducted by the FDA/CBER, under IND#13691 and the National Institutes of Health (NIH), National Center for Complementary and Integrative Health (NCCIH), including its Office of Clinical and Regulatory Affairs. Participants’ eligibility criteria are described in Streptococcus thermophilus and Lactobacillus delbrueckii probiotics [referred to as the two strain [S2] yogurt group (n = 31) were administered 112 g of the base yogurt beverage (containing Streptococcus thermophilus and Lactobacillus delbrueckii only) with no additions to the drink. In contrast, participants in the S2 + BB12 group (n = 28) were administered the base yogurt beverage that was supplemented with an additional 1 × 1010 colony-forming units of BB12 per serving per day. BB12 was chosen as the interventional probiotic of interest as it has been widely studied in infants and children . Participants included in this study were children whose parents/caregivers were able to read, write, and speak either English or Spanish and had access to a telephone and refrigerator. Eligible participants provided written informed consent were enrolled and randomized as described by Tan et al. to either the BB-12rt group ], as deschildren , and is children . The yogn = 169) for microbiome analysis and another 100 mg were sent to Metabolon Inc.  for metabolomic analyses.Fecal samples were collected at baseline (Day 0), following 10 days of yogurt consumption (Day 10), and 20 days following discontinuation of yogurt administration (Day 30), and immediately stored after collection at −80°C. Samples from days 0, 10 and 30 were then thawed, and approximately 100 mg of the samples were sent to Microbiome Center of the Children’s Hospital of Philadelphia  and quantified with the Quant-iT PicoGreen Assay Kit (Molecular Probes). Shotgun libraries were generated from 0.5 ng DNA using the Nextera XT Library Prep Kit  and libraries were sequenced on an Illumina HiSeq 2500 in High Output mode to produce paired-end 125 bp sequence reads. Extraction blanks and nucleic acid-free water were processed along with experimental samples to empirically assess environmental and reagent contamination. A laboratory-generated mock community consisting of DNA from The metabolomic analysis was performed using untargeted ultra-performance liquid chromatography-tandem mass spectrometry , as described previously . Brieflyi.e., L. delbrueckii, S. thermophilus, and B. animalis). Differential abundance analysis was performed using linear models of Log10 transformed relative abundances. Reads were mapped to the KEGG database  and 20 days post probiotic discontinuation (Day 30). Shotgun metagenomic data were analyzed using Sunbeam . The abudatabase  using DIdatabase  to estim10 transformed relative abundance for the given comparison. Probiotic-associated bacteria, previously demonstrated to be short chain fatty acid (SCFA)-producers and beneficial for GI health  analyses (FC threshold = 2), and Principal Component Analysis (PCA) were performed to analyze differences between Day 0 vs. Day 10, Day 0 vs. Day 30, and Day 0 vs. Day 30 within the S2 and S2 + BB12 groups and between groups within each time point.Exploratory and hypothesis-driven metabolite analyses were performed with untargeted metabolite data processed by Metabolon Inc. using MetaboAnalyst 5.0i.e., glycine, serine and threonine metabolism, alanine and aspartate metabolism, etc.), and SCFAs, as these metabolites are strongly associated with gut microbial community characteristics (post hoc pairwise testing (when appropriate) and Tukey’s correction were performed in JMP statistical analysis platform . Spearman correlation testing was then performed on each microbe-metabolite pair with FDR correction applied to p-values. Network diagrams of bacteria and metabolites were generated using Cytoscape v3.9.1 . RJ-L is supported by the Center of Compulsive Behaviors Fellowship, National Institutes of Health. Funding from Dannon to TT and DM, the Department of Family Medicine, Georgetown University Medical Center, Washington, DC. KM is supported by intramural research funds at the National Institutes of Health, Clinical Center.DM previously served as a paid expert at Howard University and Bayer. DM has done legal work for Visniome VSL#3, Golo for Life, and President of the International Scientific Association for Probiotics and Prebiotics (ISAPP) board.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 identification of effective dementia prevention strategies is a major public health priority, due to the enormous and growing societal cost of this condition. Consumption of a Mediterranean diet (MedDiet) has been proposed to reduce dementia risk. However, current evidence is inconclusive and is typically derived from small cohorts with limited dementia cases. Additionally, few studies have explored the interaction between diet and genetic risk of dementia.We used Cox proportional hazard regression models to explore the associations between MedDiet adherence, defined using two different scores (Mediterranean Diet Adherence Screener [MEDAS] continuous and Mediterranean diet Pyramid [PYRAMID] scores), and incident all-cause dementia risk in 60,298 participants from UK Biobank, followed for an average 9.1 years. The interaction between diet and polygenic risk for dementia was also tested.Higher MedDiet adherence was associated with lower dementia risk . There was no significant interaction between MedDiet adherence defined by the MEDAS continuous and PYRAMID scores and polygenic risk for dementia.Higher adherence to a MedDiet was associated with lower dementia risk, independent of genetic risk, underlining the importance of diet in dementia prevention interventions.The online version contains supplementary material available at 10.1186/s12916-023-02772-3. There is also currently no gold standard assessment of MedDiet adherence, and some variability in study findings may therefore be due to different dietary assessment methods and scoring systems [Preventing dementia is a global public health priority due to the enormous and growing societal cost of this condition . A key s systems . TherefoAPOE genotype as the sole measure of genetic risk [A healthy diet might also mitigate individual genetic risk for dementia. Previous studies exploring gene-diet interactions are limited, have reported inconsistent results, and, typically, focus on tic risk , 18–20. tic risk , 22 and The purpose of this study was to investigate associations between MedDiet adherence and dementia incidence in a large prospective cohort study, and to explore the interaction between diet and genetic risk for dementia.The UK Biobank is an ongoing, multi-centre prospective cohort study of over half a million participants, that provides a resource for investigating the determinants of disease in middle and older age . The desThe Oxford WebQ is a web-based, self-administered 24-h dietary assessment tool, validated for use in large-scale observational studies , 26. ThiWe quantified MedDiet adherence using two separate scores: the MedDiet Adherence Screener (MEDAS) score, and the MedDiet PYRAMID score. These scores define MedDiet adherence in different ways  and therefore may differ in terms of their association with dementia.The MEDAS is a 14-point score developed as part of the Prevención con Dieta Mediterránea (PREDIMED) trial  that hasBoth the MEDAS and MEDAS continuous scores award points for use of olive oil as the main culinary fat and, separately, for consumption of a target amount (4 or more tablespoons per day) of olive oil. Although we were able to determine use of olive oil as a culinary fat and to award points for consumption (1 point) or non-consumption (0 points) accordingly, it was not possible to determine the amount of olive oil consumed from the available dietary data, limiting the maximum possible scores for the MEDAS and MEDAS continuous to 13 points in this study.The PYRAMID score is a 15-point MedDiet score used widely in epidemiological studies , 17, 27.To estimate genetic risk of dementia, we used the polygenic risk score developed by Lourida and colleagues, who demonstrated that higher values of this score are associated with higher all-cause dementia risk in the UK Biobank cohort . The scost March, 2021 for England and Scotland, and 28th February, 2018 for Wales. The mean (SD) and median (interquartile range) follow up was 9.1 (1.7) and 9.3 (8.8–9.7) years, respectively. Follow up time was calculated from the most recent eligible dietary report used for MedDiet score creation and either the date of first dementia diagnosis, death, loss to follow-up, or censoring date, whichever was the earliest.All-cause incident dementia cases were ascertained using data linkage to hospital inpatient records and death registries. Diagnoses were recorded using the International Classification of Diseases (ICD) coding system . ParticiAll analyses were conducted in SPSS version 27. Baseline characteristics of the analytic sample, stratified by dementia status, were summarised as mean ± SD for continuous variables and as percentages for categorical variables. Cox proportional hazard regression models were used to examine the association between MedDiet adherence and time to incident all-cause dementia, with the duration of follow-up in years used as the timescale. We also explored the association between the polygenic risk score and dementia incidence, to confirm the previously reported associations between these variables in this cohort . The posAnalyses were adjusted simultaneously for: age, sex, socioeconomic status , education , smoking status , typical sleep duration , physical activity , energy intake , third-degree relatedness of individuals in the sample, and the first 20 principal components of ancestry. Models which included the polygenic risk score were additionally adjusted for the number of alleles included in the score, to account for SNP-level variation 2). Seventh, as an alternative method of exploring whether associations between MedDiet adherence and dementia risk were influenced by polygenic risk score, we conducted stratified analyses exploring associations between MedDiet adherence and dementia risk in low, medium and high genetic risk categories. Eighth, we investigated the interaction between MedDiet adherence and genetic risk, with genetic risk defined by Apolipoprotein E (APOE) genotype only . APOE ε4 carriers were defined as higher risk, whilst non-carriers were defined as lower risk. Nineth, to evaluate the influence of missing data, we repeated analyses following imputation of missing dietary and covariate data using multiple imputations by chained Eqs.  [Sensitivity analyses were performed to test the robustness of associations between MedDiet adherence and dementia incidence. First, we used the conventional binary MEDAS score. Secondly, we included participants with a minimum of two, 24-h diet recalls to provide a more stringent measure of habitual dietary intake . Thirdlyrations) . We incl2), have a higher educational level, and be more physically active than those with lower MedDiet adherence. The mean MEDAS continuous and PYRAMID scores in this cohort were 6.1 ± 1.7 and 7.5 ± 1.8, respectively.A total of 502,536 participants underwent baseline assessment as part of the UK Biobank study, of whom 60,298 participants were included in this analysis  and PYRAMID  scores. When divided into tertiles, relative to low MedDiet, high but not moderate adherence was associated with lower dementia risk .Higher adherence to the MedDiet was associated with 4.2–6.9% lower risk for dementia for the MEDAS continuous  and 0.55% (95% CI: 0.32–0.79%) lower absolute risk of dementia than those in the low MedDiet adherence group. Meanwhile, the absolute risk of dementia in the low, medium and high MedDiet adherence groups defined by the PYRAMID score was 1.67%, 1.53%, and 1.21%, respectively. Moderate and high MedDiet adherence groups had 0.14% (95% CI: -0.1–0.39%) and 0.46% (95% CI: 0.23–0.69%) lower absolute risk of dementia than those in the low MedDiet adherence group.p < 0.001). There was no significant interaction between polygenic risk for dementia and MedDiet adherence defined by the MEDAS continuous  or PYRAMID  scores.A higher polygenic risk score was associated with greater risk for dementia  and, when split into tertiles, high  but not moderate  MedDiet adherence was associated with lower dementia risk versus low MedDiet adherence . Results were similar when we repeated analyses for participants with a minimum of 2 dietary reports  rather than continuous  MEDAS score, there was a similar, albeit slightly attenuated, association between MedDiet adherence and dementia risk. Specifically, each one-point increase in MEDAS score was associated with 4.5% lower risk of dementia . When analyses were stratified by polygenic risk category, higher MedDiet adherence according to the MEDAS continuous scores was associated with lower dementia incidence in individuals in the lower genetic risk category only  HR: 0.985; 95% CI: 0.913–1.064; p = 0.706; PYRAMID HR: 1.054; 95% CI: 0.978–1.136; p = 0.167). Likewise, when analyses were stratified by APOE genotype, there was a similar pattern of response  in APOE ε4 carriers/non-carriers , which was equivalent to an absolute risk difference (reduction) of 0.55%. We found no significant interaction between MedDiet adherence, defined by both the MEDAS continuous and PYRAMID scores, and polygenic risk for dementia. In addition, we found that a continuous MEDAS score was a more sensitive predictor of dementia risk when compared with a binary MEDAS or PYRAMID scores.Previous studies exploring associations between MedDiet adherence and dementia risk have produced inconsistent findings. Indeed, a systematic review by Limongi and colleagues  reportedAPOE ε4 carriers but not non-carriers in one study [APOE ε4 non-carriers. In the present study, we found no significant interaction between polygenic risk for dementia and MedDiet adherence defined by the MEDAS continuous or PYRAMID scores in our primary analyses. Likewise, when we explored the interaction between MedDiet adherence and genetic risk defined by APOE genotype, there was a similar pattern of response for both APOE ε4 carriers/non-carriers. Thus, our findings suggest similar associations between MedDiet adherence and dementia risk irrespective of genetic risk for this condition. Nevertheless, we acknowledge a degree of uncertainty in this conclusion, given that findings were not consistent across all sensitivity analyses. Further research into the interaction between diet and genetics on dementia risk is therefore warranted.There is limited and inconclusive evidence about the interaction between diet (defined by MedDiet adherence or another dietary index) and genetic risk on dementia incidence , 18–20. ne study . In contne study  are moren = 1000–6000) with limited dementia cases (n = 20–400) and may have lacked statistical power [n =  ~ 60,000) with more dementia cases (n = 882) than most previous investigations. We defined genetic risk for dementia using a comprehensive polygenic risk score whereas most previous studies have explored gene-diet interactions for individual genetic variants  [This study has several strengths. The majority of previous studies exploring associations between MedDiet adherence and dementia risk have involved relatively small numbers of participants  , 18–20. enotype)  and Barcenotype)  cohorts enotype) . Althougs n = 20–0 and mayenotype) . The meaenotype) , to derienotype) , and manenotype) . Based oenotype) , suggestenotype) , 45. Howenotype)  but thisenotype) . NeverthIn this large population-based prospective cohort study, higher adherence to a MedDiet was associated with reduced dementia risk. A continuous MEDAS score was the most sensitive predictor of dementia risk when compared with a binary MEDAS or PYRAMID score and could therefore be prioritised as a tool for defining MedDiet adherence in future observational studies. There was no clear evidence for an interaction with genetic risk. These results underline the importance of dietary interventions in future dementia prevention strategies regardless of genetic predisposition.Additional file 1: Text S1. Dietary assessment and creation of the MedDiet scores. Table S1. Components and scoring of the MEDAS and MEDAS Continuous Mediterranean diet scores. Table S2. Components and scoring of the PYRAMID Mediterranean diet adherence score. Table S3. ICD-9 and ICD-10 codes for dementia diagnosis. Figure S1. Participant flowchart. Table S4. Risk of incident dementia according to Mediterranean diet adherence, with analyses restricted to individuals with a minimum of 2 dietary reports. Table S5. Risk of incident dementia according to Mediterranean diet adherence excluding participants with extreme energy intakes. Table S6. Influence of each component of the MEDAS and MEDAS Continuous scores on dementia risk. Table S7. Influence of each component of the PYRAMID score on dementia risk. Table S8. Risk of incident dementia according to Mediterranean diet adherence, excluding participants with less than 2 years and 5 years of follow up. Table S9. Sensitivity analyses adjusting for potential effect mediators . Table S10. Association between MedDiet adherence and risk of dementia in analyses stratified by polygenic risk score tertiles. Table S11. Associations between MedDiet adherence and dementia incidence in APOE ε4 non-carriers and carriers. Table S12. Associations between MedDiet adherence and dementia incidence in analyses where missing data were imputed. Table S13. Risk of incident dementia according to Mediterranean diet adherence restricted to fatal and non-fatal dementia cases. Table S14. Risk of incident dementia according to Mediterranean diet adherence for individuals with higher and lower education levels.


This study aimed to build and validate a nomogram model to predict the risk of incomplete immune reconstitution in people living with HIV (PLWH).N = 2678) and was validated using the remaining cases (N = 1105). Univariate and multivariate logistic regression analyses were performed to determine valuable predictors among the collected clinical and laboratory variables. The predictive model is presented in the form of a nomogram, which is internally and externally validated with two independent datasets. The discrimination of nomograms was assessed by calculating the area under the curve (AUC). Besides, calibration curve and decision curve (DCA) analyses were performed in the training and validation sets.Totally 3783 individuals with a confirmed diagnosis of HIV/AIDS were included. A predictive model was developed based on a retrospective set (The final model comprised 5 predictors, including baseline CD4, age at ART initiation, BMI, HZ and TBIL. The AUC of the nomogram model was 0.902, 0.926, 0.851 in the training cohort, internal validation and external cohorts. The calibration accuracy and diagnostic performance were satisfactory in both the training and validation sets.This predictive model based on a retrospective study was externally validated using 5 readily available clinical indicators. It showed high performance in predicting the risk of incomplete immune reconstitution in people living with HIV.The online version contains supplementary material available at 10.1186/s12889-023-16738-w. However” (INRs) , 7. Comp” (INRs) –10.Incomplete immune reconstitution is pathophysiologically thought to be associated with decreased bone marrow hematopoiesis, thymic dysfunction, residual viral replication, altered gut microbiota, and coinfections, particularly persistent inflammation and abnormal immune activation, significantly decreasing CD4 production and persistent CD4 destruction , 11–13. + T cells, especially in INRs. At present, it is particularly important to assess patient condition earlier, especially at the initial examination, and to adopt a timely and individualized treatment plan. It is commonly admitted that several factors can predict immunological function recovery and disease progression, e.g., CD4+ T cell count, CD4/CD8 ratio, viral load (VL) and IFN-γ [+ T cell activation and apoptosis during ART treatment; the higher the US/MS RNA ratio the higher the frequency of HIV-infected cells, leading to sustained immune activation and apoptosis, resulting in decreased immune response to ART.To date, no effective treatment could recover CD4nd IFN-γ –22. Furtnd IFN-γ  found a In clinic, a single index is often inadequate to independently predict disease progression with satisfactory results. However, the combination of several single indexes may greatly improve the predictive effect. Medical nomograms based on various markers have been increasingly used in oncology and other areas of medicine in recent years. In addition, multiple prognostic models for PLWH have been established , 25. HowThen, a unique scoring system was created using the primary predictive model's modified nomogram for easy clinical application. Additionally, in a retrospective analysis, we internally verified the diagnostic capabilities of the improved scoring model.+ T cell counts < 350 cells/µL at 2 years after cART initiation, with an undetectable plasma VL(< 50 copies/mL) [This was a retrospective study of data collected from the China’s for Disease Prevention and Control (CDC)’s Information System. Patients with HIV/AIDS treated at Wuhan Jinyintan Hospital from December 2006 to October 2020 were included for the purpose of model construction and internal validation. Those Participants were splited randomly into the training and internal validation sets in a ratio of 7:3. Besides, the external validation set was obtained from Huangshi and Jingzhou, covering the period from May 2015 to May 2016. Inclusion criteria were: (1) Complete laboratory test confirming HIV infection; (2) Treatment with a combination ART regimen containing at least three drugs; (3) With follow-up results after two years of ART; (4) Age > 15 years. Exclusion criteria were: (1) Previous exposure to ART; (2) VL ≥ 400 copies/mL after 24 months of ART, indicating virologic treatment failure . INR defpies/mL) .2 (kg/m2), infection route, marital status, interval from diagnosis to ART, WHO clinical stage of the HIV disease, opportunistic infection (OI), coinfection with other bacteria or virus, several clinical symptoms, tumors, ART regimens, CD4+ T cells, VL, white blood cells (WBC), platelets (PLT), hemoglobin (HB), alanine aminotransferase (ALT), aspartate transaminase (AST), total bilirubin (TBIL), serum creatinine (Scr), triglycerides (TG), serum total cholesterol (TC) and blood glucose (BG). These parameters were obtained by trained professionals every 3 months.Demographic characteristics, clinical data and laboratory indexes were collected, including age at the time of diagnosis, sex, body mass index (BMI) calculated as weight/heightThere is no straightforward way to determine the right sample size for a multivariate regression model. A predictive component requires at least 10 effective outcomes, according to previous reports, based on a cautious estimate .Multiple imputations were used to acquire suitable values for missing data before data analysis since directly discarding data with missing values might cause selection bias or decrease the power of a test. The missing values of the training set were analyzed Fig. . The preVariables in the training and validation sets were described as number (percentage) or median , as appropriate. Continuous variables among groups were compared by the Mann–Whitney U test. Meanwhile, categorical variables were compared by the chi-square test, the fisher’s exact test or Wilcoxon rank sum test.P < 0.1 were retained for further analysis. After multivariate logistic regression, 15 candidate predictors were retained. Variables were further selected considering statistically significant parameters and medically important parameters such as availability at first assessment and objectivity of the metric. Finally, five variables, extracted by experienced physicians, were included in the predictive model with the highest predictive performance.Univariate logistic regression analysis (ULRA) was carried out to select factors in the training set. Then, 34 potential variables with Based on the five most significant variables, a nomogram model with an appropriate predictive ability was developed. The discrimination and calibration of the predictive model was evaluated to test the effectiveness of the model. Among the training set, internal validation, and external validation sets, receiver operating characteristic (ROC) curve analysis was utilized to quantify the discriminative value of the model, and a calibration curve was used to evaluate the calibration. Furthermore, decision curve analysis (DCA) was used to evaluate the clinical utilities of the model.http://cran.r-project.org). Two-sided p < 0.05 was considered statistically significant.Data analysis used SPSS version 26.0  and R-Studio for windows (version 4.2.0)  were INRs, including 21.8% (583/2678) in the training set, 22.0% (243/1105) in the internal validation set, and 14.7% (129/878) in the external validation set.p < 0.1 were selected by experienced physicians for further multivariate logistic regression analysis. Finally, five predictors  were selected as independent risk factors for the INR status  for model construction and validation. Secondly, this model performed well in the validation set, which suggests its potential generalizability. Thirdly, this model can more accurately help clinicians make decisions, with a high AUC. Fourthly, we developed a user-friendly online calculator that only requires the input of a few parameters, and all data conversions and computations are built right into the system, in order to decrease the difficulties imposed by model complexity in clinical application.We developed and validated a model consisting of 5 clinical and laboratory variables for accurate prediction of the risk of poor immune reconstitution at the time of primary diagnosis. This model can help predict disease progression and regression, providing efficient and precise treatments to improve the life expectancy and quality of life of patients.+ T cell count was not included as a candidate predictor in our model due to its high percentage of missing values. This was largely caused by the inherent drawback of retrospective data collection. Next, there was a bias in the predicted accuracy. Even though the majority of indicators in the developed model may be assessed objectively, the route of HIV acquisition is reported by the patients themselves, which could be biased.This study also had several limitations. First, CD8In conclusion, A sophisticated nomogram model has been developed and externally validated for the prediction of the risk of poor immune reconstitution at the time of primary diagnosis in this study. we recommend the widespread application of the novel nomogram model to effectively and efficiently identify individuals who are at a heightened risk of INR.Additional file 1: Supplementary table 1. Sensitivity analysis in imputation for missing data. Supplementary table 2. Univariable and multivariable logistic regression analysis of the training set. Supplementary Figure 1. Flow diagram of participants selection. Abbreviations: INRs, immunologic non responders; VL, viral load ; ART, antiretroviral therapy.


The severe acute abdomen associated with Henoch-Schonlein purpura (HSP) is an acute intussusception (AI). There is no reliable specific marker for AI with abdominal-type HSP. The serum total bile acid (TBA) level is a new prognostic marker associated with the severity of intestinal inflammation. The purpose of this study was to identify the prognostic value of serum TBA levels for the diagnosis of AI in children with abdominal-type HSP.A retrospective study of 708 patients with abdominal-type HSP was conducted, with demographic data, clinical symptoms, hepatic function index, immune function markers, and clinical outcomes assessed. Patients were divided into two groups: HSP (613 patients) and HSP with AI (95 patients). The data were analysed using SPSS 22.0.P < 0.05). Logistic regression analysis showed that vomiting , haematochezia , TBA , and D-dimer  were independent risk factors for abdominal-type HSP with AI. Receiver operating characteristic (ROC) curve analysis showed that the optimal cut-off serum TBA value  was >3 μmol/L for predicting AI in children with abdominal-type HSP. In this group of HSP patients with AI, a serum TBA level ≥6.98 μmol/L was significantly associated with an increased incidence of operative treatment , intestinal necrosis , and length of hospital stay .Of the 708 patients, the serum TBA levels were higher in the HSP with AI group than in the HSP group (In children with HSP and AI, the serum TBA level was significantly higher. A novel but promising haematological indicator, the serum TBA level, helps identify HSP with and without AI and predicts intestinal necrosis in HSP with AI. It maniAI is the most common acute abdomen associated with HSP . HoweverBile acids (BAs) play an important role in the body's endogenous metabolites. BAs in the intestine are involved in lipid absorption and the regulation of intestinal motility, mucosal secretory function, and intestinal barrier function . BAs areIn this study, we assessed whether there was a connection between the level of serum TBA and the occurrence of AI in patients with abdominal type HSP.2.2.1.The data of children with abdominal-type HSP admitted to the Nephrology Department and General Surgery Department of Children's Hospital of Chongqing Medical University between January 2017 and December 2021 were retrospectively reviewed. All abdominal-type HSP patients fulfilled the diagnostic standards that were jointly established in 2010 by the Paediatric Rheumatology International Trials Organization and the European League Against Rheumatism . The inc2.2.The subject characteristics included (1) demographic data such as sex, age, first episode, and onset seasons and (2) clinical features such as gastrointestinal symptoms  and whether receiving glucocorticoid therapy within 72 h of the onset of gastrointestinal symptoms.The hepatic function index , coagulation function parameters , and immune function markers  were collected. When gastrointestinal symptoms were present, the serum TBA level was measured.2.3.P < 0.05 was regarded as statistically significant. The Shapiro Wilk test was used to determine whether the data were normally distributed. Values with a normal distribution are represented by the mean and standard deviation (mean ± SD), whereas values without a normal distribution are represented by medians and interquartile ranges. Numbers and percentages are used to represent categorical variables. The area under the curve and the cut-off value for the best predictor were determined using receiver operating characteristic (ROC) curve analysis (AUC). The mean and 95% confidence interval (95% CI) of continuous variables were compared using Student's t test. Chi-square or Fisher exact tests were used to compare categorical variables between the groups. Univariate and multivariate logistic regressions were also used to calculate odds ratios (ORs), 95% confidence intervals (CIs), and P values.For data entry, Excel software was used, and statistical analyses were carried out with SPSS version 22.0 . 3.3.1.In the end, 708 patients with abdominal-type HSP involvement were included in this study . Table 1P > 0.05). There were statistically significant differences between the groups in age at onset (P = 0.0198) and whether glucocorticoid therapy was started within 72 h of the onset of gastrointestinal symptoms (P = 0.0005). In terms of symptoms, there were statistically significant differences in vomiting and haematochezia rates between the two groups (P < 0.001) but not in haematemesis rates and abdominal pain (P > 0.05). The involvement of the kidneys was also not significantly different (P > 0.05). Blood analysis revealed no statistically significant difference in white blood cell count, haemoglobin, neutrophil count, platelet count, or C-reactive protein levels between the study groups (P > 0.05).Gender, first disease episode, and onset seasons had no statistically significant between-group differences  had successful procedures in ileocolic intussusceptions. The success rate of air enema for small intestine intussusception was only 13.51% (5/37). Air enema failed in 46 patients (56.10%) following surgery. Of the 59 patients who underwent surgery, 17 (28.81%) underwent intestinal resection due to intestinal necrosis. There were 9 cases of ileal intussusception, 7 cases of ileocolic intussusception, and 1 case of jejunal intussusception among the 17 cases of enterectomy. The average nested distance in intussusception was 29 ± 15 cm. There were 28 cases of ileocolic intussusception, 13 cases of ileal intussusception, and 1 case of ileal ileocolic intussusception among the 42 children who underwent manual reduction, with an average nested distance in intussusception of 13 ± 6 cm.3.2.P < 0.0001), and there were no differences between the two groups in TBIL, DBIL, γ-GGT, and albumin (P > 0.05). Coagulation function tests indicated significant differences between the study groups in D-dimer (P = 0.0146) and FIB levels (P = 0.0103) but not in TT, PT, or APTT (P > 0.05). Humoral immunity tests showed a statistically significant difference in serum total IgG (<0.0001) and IgM (P = 0.0002) between the study groups but not in IgE and IgA , haematochezia , TBA , and D-dimer  were independent risk factors for abdominal-type HSP with AI . The ROC analysis revealed that a serum D-dimer cut-off level of 4.7 mg/L provides the prediction for abdominal-type HSP with AI in this cohort, with a sensitivity of 93.68% but a specificity of only 20.07% (AUC = 57.8724%) .In this cohort, vomiting  and haematochezia  were the best clinical predictors for abdominal-type HSP with AI .3.5.P = 0.0181), intestinal necrosis , and length of hospital stay  (In this group of HSP patients with AI, the average serum TBA level was 6.98 μmol/L. According to the average level of TBA, the group comparison showed that a TBA level ≥6.98 μmol/L was significantly associated with an increased incidence of operative treatment (51.85% vs. 75.61%,  0.0001] .4.The current study found that serum TBA levels, fibrinogen, vomiting, and haematochezia were independently associated with abdominal-type HSP and AI. In terms of prediction, TBA had a sensitivity of 91.58% and a specificity of 84.67% in identifying children at high risk for AI. In addition, an average serum TBA level of 6.98 mol/L was significantly associated with an increased incidence of surgery, intestinal necrosis, and length of hospital stay in HSP patients with AI.Children with HSP experience widespread intestinal wall oedema and segmental bleeding of the subserous membrane and mucosa as a result of antigens and antibodies acting on intestinal wall capillaries, increasing capillary permeability . IntussuAlthough the above variable is discussed in both previous studies and our study, it is important to note that those factors are greatly influenced by the attending physician's experience and are subjective. Furthermore, objective variables obtained from blood samples are usually more reproducible and thus more valuable. In a previous report, GI symptoms of HSP were correlated with serum biomarkers such as D-dimer, white blood cell count, and C-reactive protein level , 22. DurTo diagnose HSP with intussusception in children, it is necessary to identify diagnostic noninvasive laboratory markers. Bile acids are important signalling molecules that regulate immunity as well as maintain and promote intestinal barrier function. Bile acids have been associated with colorectal cancer , irritabThe TBA level at admission could guide the severity of HSP and predict prognosis, with 3 μmol/L being the best cut-off point. When TBA reached 3 μmol/L, the probability of intussusception complicated by HSP increased significantly.Air enema and surgery are used to treat intussusception. The success rate of air enema is relatively low in children with HSP due to the serious secondary changes such as intestinal wall oedema and bleeding . The sucThere were several limitations to this study. This study used a retrospective case‒control design. In terms of research objects, a large number of HSP patients with incomplete data were excluded, which may have biased the findings. The subjects in this study had been hospitalized in the previous 5 years, and the descriptions of clinical indicators may be inconsistent. Although logistic regression analysis was used to some extent in the data analysis, the influence of clinical indicators remained significant, which may have interfered with the detection value between HSP with intussusception and serum TBA. In terms of assessing patient prognosis, the evaluation methods in this study are relatively one-sided, and there is still a lack of comprehensive evaluation data of intussusception injury, so the relationship between HSP with intussusception and serum TBA should be explored more comprehensively and accurately.5.We demonstrated that serum TBA levels can assist clinicians to predict the occurrence of AI with abdominal type HSP in children. The serum TBA level can be used as a noninvasive and easily applicable biomarker for the screening and predicting progression in HSP with AI.


Orchidaceae from Caquetá, Colombia is presented here. We recorded 98 genera and 418 species, exceeding a previous inventory by 276 species. The checklist is conservative in the number of genera and species by including only taxa that were fully and reliably identified and that are either linked to a corresponding herbarium voucher, a living collection specimen or a photo taken in the field and published in iNaturalist by one of the authors or a collaborator. The documented species diversity in the region could dramatically increase in the next few years with additional collecting efforts in the eastern slopes of the Andes nested in Caquetá. About 9% (418/4600) of all Orchidaceae species recorded for Colombia are reported for this area, showing the important contribution to orchid diversity of Andean-Amazonian foothills of Caquetá.A checklist of  Orchidaceae are one of the most diverse and widely distributed flowering plant families including 25,000–27,000 species and 880 genera , the Amazon Basin  or the Orinoco Basin  . These eOrchidaceae are diverse within Caquetá, few checklists and taxonomic studies focusing on this group are available. For example, the Catalogue of Plants and Lichens of Colombia . This programme aims to explore, reproduce and support orchid species recovery, while developing sustainable strategies for business, based on horticulture for vulnerable communities around Colombia. Data obtained here come from living collections, photographs taken from field and an extensive review of herbarium collections around the world.In this study, we generated a detailed species list of Caquetá is located in south-western Colombia, between latitudes 0.7°S–2.9°N and longitudes 71°–76°W. It comprises 16 municipalities including Florencia, Belén de los Andaquíes, El Paujil, Doncello and La Montañita, amongst others Table  Orchidaceae that occur in Caquetá, we carried out a total of 12 field expeditions between 2019 and 2023 for the project “Orquídeas para la Paz.” Two expeditions explored part of the Alto Fragua Indi Wasi National Natural Park in collaboration with the Park and ten expeditions explored montane, premontane forests and lowland Amazonian Forest of the Department. Fertile specimens were collected and prepared for herbaria according to techniques used for orchid collections, that includes the preservation of flowers in spirit collections, photographs and tissue collections for ongoing DNA analyses. The specimens were deposited at either theUniversidad de la Amazonia (HUAZ) or theUniversidad del Valle (CUCV) Herbaria ,Herbario del Instituto Chinoin (AMO), Berlin (B),Royal Botanic Gardens Kew Herbarium (KEW),University of California, Los Angeles Herbarium (LA), theReal Jardín Botánico de Madrid (MA),Naturalis Biodiversity Center (NL),Herbarium Utrech (U), New York Botanical Garden (NY), theMuséum national d´Histoire naturelle in Paris (P), Marie Selby Botanical Gardens (SEL), Tropicos (2002) of the Missouri Botanical Garden (MO),Gdansk University (UGDA),University of Florida Herbarium (USF), W-Reichenbach (Vienna) andUniversity of Wisconsin Herbarium (WIS).To carry out online consultations in herbaria, search criteria were considered using the keywords: “Caquetá”, “Instituto Amazónico de Investigaciones Científicas – SINCHI (COAH),Herbario Nacional Colombiano (COL),Herbario de la Universidad del Cauca (CAUP),Herbario de la Universidad del Valle (CUCV),Herbario de la Universidad de Caldas (FAUC),Herbario Federico Medem-Bogotá (FMB),Herbario de la Pontificia Universidad Javeriana (HPUJ),Herbario de la Universidad de Antioquia (HUA),Herbario Enrique Forero (HUAZ) de la Universidad de la Amazonia,Jardín Botánico José Celestino Mutis de Bogota (JBB),Universidad de los Llanos (LLANOS),Universidad de Nariño (PSO),Universidad Surcolombiana (SURCO),Herbario Forestal de la Universidad Distrital Francisco José de Caldas (UDBC) andUniversidad Pedagógica y Tecnológica de Colombia (UPTC). Other checked databases included GBIF , IDigBio (Integrated Digitized Biocollections) and BRAHMS at Marie Selby Botanical Gardens .Colombian herbaria included:Tubella (Luer) Archila, which was accepted as valid following www.inaturalist.com) or living collections in one of the local nurseries that supported their presence in Caquetá with reliable taxonomic determination.Correct scientific names of species were assigned, based on the World Flora Online , except All records obtained from herbaria, databases and literature were carefully curated regarding their scientific name, locality, collector and collection number. For localities, names of municipalities were verified and updated. To assign a name to duplicates with different identification, the provenance of each assigned name was investigated, paying particular attention to plants identified by experts, curators and recognised taxonomists, besides using the date of determination and its citation in a publication. Lastly, using photographs and a list of taxonomic groups within orchids, we reached out to as many specialists as possible for their species expertise (see Acknowledgements). This final dataset was used for analyses.Collection records were georeferenced as precisely as the information allowed, since in many records, the location is not clearly specified, a frequent situation in orchids, old collections and those made by amateurs. Due to the lack of standardised geographical coordinates, only the number of species in municipalities is reported for this study. Georeferenced records and species distribution maps were constructed for the 16 municipalities and are available upon request.Epidendrumnocturnum Jacq., E.lacustre Lindl., Maxillariadiscolor (G. Lodd. ex Lindl.) Rchb. f. and Scaphyglottisstellata Lodd. ex Lindl. Some of the rarest ones only observed in the field once were Cyrtochilumcaquetanum P. Ortiz, L. E. Álvarez & A. J. Carrillo, Masdevalliaensata Rchb. f. and Paphiniacristata (Lindl.) Lindl. In “Orquídeas para la Paz” expeditions, 98 species (collections made by Arias T. and Chaux-Varela J.) representing 29 genera were collected. Living individuals that were not flowering in the field were brought to El Manantial (300 m a.s.l.) and El Caraño (950 m a.s.l.) both located in the Municipality of Florencia. Herbaria collections are actively being made once orchids start flowering. A total of 55 species are available at El Manantial and 60 at El Caraño.A total number of 228 fertile specimens were collected in the field , 692 herbarium specimens were reviewed in different herbaria, 100 photographs and two living collections accessed for specimen identifications. The most abundant species in the field were Epidendrum L. (68 spp.), Maxillaria Ruiz & Pav. (59 spp.), Pleurothallis R.Br. (16 spp.), Elleanthus C. Presl (14 spp.), Sobralia Ruiz & Pav. (14 spp.) (Table Arundinagraminifolia (D. Don) Hochr. and Dendrobiumnobile Lindl.; these were not included in the species list.We report 418 species belonging to 98 genera represented in 744 herbarium collections including duplicates. Eighty-two species are new reports for Caquetá since they have not been vouchered until this study (Table .) Table –5. Most https://www.inaturalist.org/projects/orquideas-del-caqueta). Twenty-seven species including 15 in El Manantial and 12 in El Caraño, are part of our living collections. They were collected fertile in the field and identified, but have not been photographed or documented with herbarium vouchers to date.Eighty-one species are included as “tentative” because they have been accurately identified, but lack a herbarium voucher. Fifty-three out of these 81 were included as potentially distributed in the Caquetá because records produced by the authors and collaborators in the field through photographs were not documented with herbarium vouchers. Such photographic records were submitted to iNaturalist , Miltoniopsisphalaenopsis (Linden & Rchb. f.) Garay & Dunst. (VU) and Oncidiumalexandrae (Bateman) M. W. Chase & N. H. Williams (EN) were included in the Red List of Colombian orchid species  and Maxillaria (59/570) was expected because these are some of largest Neotropical Orchidaceae genera with regards to species number A total of 276 new species records of s region . The sigxillaria 9/570 wasndrum 68/000 and MDuring the construction of this list, we left out collections made by Werner Hopp  since thCAR) are the main environmental authority. They are responsible for implementing policies and plans from the Ministry of Environment and are granted administrative and financial autonomy. Entities responsible for the formulation of such conservation strategies might benefit more from having such information presented following Departmental divisions. By depositing 140 herbarium specimens at HUAZ (duplicates will be sent to other herbaria) in the framework of this study, we substantially increased its orchid collection to 210 species, positioning HUAZ as the first local herbarium with more herbarium collections from Caquetá than any other herbaria . This could be explained by the convenience of collecting around cities and the wide altitudinal gradient in this municipality. We present collections numbers by municipality because conservation strategies might differ between political boundaries in Colombia. Regional Autonomous Corporations  Lindl. and Trichocentrumlanceanum (Lindl.) M. W. Chase & N. H. Williams have been found only in La Laguna del Chaira in the Cartagena del Chaira Municipality. We doubt these species have a natural distribution there. Rather, we suspect they were introduced during the massive effort to bring orchids to La Laguna del Chaira during the 1980s, during which “uninformed” reintroductions of non-native species could have taken place.During our expeditions, two species th place, 62 genera). As for the ranking in species number for Colombia, Caquetá goes from position 17th (142 spp.) to position 9th (418 spp.) (This checklist places Caquetá as the eighth Department in Colombia in terms of genera diversity (98 genera) from its original position in the National Plan of Orchid Conservation (1518 spp.) . Caquetá18 spp.) . CaquetáCaquetá has lost approximately 30% of its original area due to human impacts, such as cattle ranching. National parks in Caquetá make up 65% of the protected remnants. In the last 50 years, expansion of the agricultural frontier for the establishment of grazing lands, wood extraction and illegal coca crops have destroyed many ecosystems, greatly impacting all national parks. Florencia, for example, is currently undergoing consistent expansion of farming lands ultimately leading to the decimation of natural ecosystems .Our floristic study is a needed contribution towards a better understanding of the diversity of Colombian orchids. The checklist provides a set of freely-available data on orchid diversity in Caquetá. Furthermore, our study is a baseline panorama of orchid species diversity in the Department, identifying groups of interest for further taxonomic work, especially those which have not been monographed. Lastly, the information provided could enhance local conservation strategies for endangered floristic elements in the Department by adding to a more complete overview of the high orchid diversity in the region.


Mucinous carcinoma (MC) is a histological subtype of ovarian cancer that has a worse prognosis at advanced stages than the most prevalent histological subtype, high-grade serous carcinomas. Invasive patterns have been recognized as prognostic factors for MCs. MCs with infiltrative invasion were more aggressive than those with expansile invasion. MC with an expansile pattern exhibited behavior similar to mucinous borderline tumors (MBT). However, genomic analysis of invasive patterns is insufficient. This study aimed to compare genetic information between groups with MC and infiltrative invasion (Group A) and those with MC with expansile invasion or MBT (Group B).Ten cases each of MC with infiltrative invasion, MC with expansile invasion, and MBT between 2005 and 2020 were identified. Deoxyribonucleic acid (DNA) extraction from formalin-fixed paraffin-embedded tissues was performed, and cases with DNA fragmentation or the possibility of DNA fragmentation were excluded. Mutant base candidates and tumor mutation burden (TMB) values (mutations/megabase) were calculated.p < 0.01) and died of disease (p = 0.03). Moreover, the TMB value was statistically higher in group A than in group B (p = 0.049). There were no statistical differences in the incidence of the mutations of KRAS, TP53, and CREBBP. KRAS, TP53, and CREBBP mutations were discovered in 8/15 (53.3%), 6/15 (40.0%), and 5/15 (33.3%) cases, respectively.After assessing the quality of purified DNA, seven cases of MC with infiltrative invasion, five cases of MC with expansile invasion, and three cases of MBT were included. More patients in group A experienced recurrence or progression (Genetic analysis revealed that Group A had higher TMB than Group B. Therefore, this result might be useful for future treatment. Epithelial ovarian carcinoma (EOC) is one of the most lethal gynecological diseases , 2. The According to the 2020 World Health Organization (WHO) classification, EOCs are morphologically classified into several histological subtypes. Among them, mucinous carcinoma (MC), has an incidence ranging from 3 to 11% , 5. The Based on histological evaluation of invasive patterns, MCs are classified into two types: infiltrative invasion and expansile invasion . CompareAccording to previous literature, MC is associated with genetic alterations such as KRAS mutations, tumor suppressor protein p53 (TP53) mutations, and human epidermal growth factor receptor 2 (HER2) amplifications, but an effective treatment for MCs has not been established in spite of these identified gene alterations . FurtherTherefore, this study aimed to evaluate the genetic information of MC with infiltrative invasion, MC with expansile invasion, and MBT.Ten patients with MC with infiltrative invasion, 10 with MC with expansile invasion, and 10 with MBT who underwent primary surgery between 2005 and 2020 at National Defense Medical College were included. Cases with a prior history of chemotherapy and with destructive deoxyribonucleic acid (DNA) fragmentation or the possibility of DNA fragmentation were excluded.The pathological diagnosis was conducted based on the 2020 WHO classification by experienced gynecological pathologist at National Defense Medical College . In caseStaging was performed using the 2014 FIGO criteria . Data onDNA was extracted from formalin-fixed paraffin-embedded (FFPE) tissues using NucleoSpin DNA FFPE XS  according to the manufacturer’s instructions  . The are> T:A mutations with an allelic frequency < 15% in coding regions, and statistical analyses were performed using Oncomine™ Tumor Mutation Load Assay USER GUIDE (Rev. C.0).Mutant base candidates and tumor mutation burden (TMB) values (Mutations/Megabase [Mut/Mb]) were detected by mapping the sequence results obtained using sequence analysis on the Torrent Server according to the Ion Reporter 5.12, according to the Oncomine™ Tumor Mutation Load Assay USER GUIDE (Rev. C.0) . Brieflyp < 0.05.Statistical analysis was performed using the JMP Pro 14 software . The chi-square test, Fisher’s exact test, Mann-Whitney U test, and Wilcoxon test were used to evaluate the statistical significance of the clinical and genetic factors. Group A was defined as patients with MC with infiltrative invasion and Group B was defined as patients with MC with expansile invasion and MBT. The level of statistical significance was set at After the analysis to check the quality of DNA, seven cases of MC with infiltrative invasion, five of MC with expansile invasion, and three of MBT were included Fig. . There wp < 0.01) and died of the disease (p = 0.03) than in Group B.Comparisons of the characteristics between Group A and Group B are shown in Table p = 0.049). Mutations in KRAS, TP53, and cyclic adenosine monophosphate response element-binding protein-binding protein (CREBBP) were recognized in 8/15 (53.3%), 6/15 (40.0%), and 5/15 (33.3%) cases, respectively. The other mutational bases listed included one case each. There were no statistically significant differences in gene mutation patterns between the two groups.The TMB value (Mut/Mb), and the mutation of bases according to MC with infiltrative invasion, MC with expansile invasion, and MBT are shown in Fig. In our study, mutations in KRAS, TP53, and CREBBP were identified. The TMB value of MC with infiltrative invasion was higher than that of MC with expansile invasion and MBT.The clinical features of MC, such as FIGO stage, response rate to adjuvant chemotherapy, recurrence rate, and prognosis, may vary according to the invasive pattern –11. MC wHistologically, EOCs are classified into type I and type II based on distinctive morphological and molecular genetic features, and MC are classified as type I , 21. TheThe amplification of HER2 occurs in 18.2–45.5% of MC cases, and a recent study proposed trastuzumab therapy as a treatment option for MC with HER2 amplification –31. HoweTMB is the number of somatic nonsynonymous mutations per coding area of the tumor genome. It is considered an important biomarker for predicting the response rate to immune checkpoint inhibitors (ICIs) , 33. ComThis study had some limitations. First, our study included small sample size, especially only 3 cases with MBT, from a single institution and retrospective nature. Second, our study did not perform individual cases with any combinations of mucinous ovarian carcinomas with infiltrative invasion, mucinous ovarian carcinomas with expansile invasion, and mucinous borderline tumors. Therefore, We could not yield answers regarding progression. Third, immunohistochemistry for mismatch repair proteins, germline genetic testing and investigating the status of major histocompatibility complex/human leukocyte antigen were not conducted. Further large-scale studies and further examination are needed to develop the new strategy for MC.Genetic analysis of MC with infiltrative invasion, expansile invasion, and MBT revealed that ICIs might be an effective therapy for MC with infiltrative invasion. Further studies are needed to consider these relationships.


To identify long-term trajectories of physical function and health-related quality of life (HRQoL) among people with hip and/or knee osteoarthritis (HKOA) and the sociodemographic, lifestyle, and clinical factors associated with different trajectories.p < 0.05).Participants with HKOA from the EpiDoC study, a 10-year follow-up (2011–2021) population-based cohort, were considered. Sociodemographic, lifestyle, and clinical variables were collected at baseline in a structured interview and clinical appointment. Physical function and HRQoL were evaluated with the Health Assessment Questionnaire (HAQ) and EuroQoL, respectively, at baseline and the three follow-ups. Group-based trajectory modeling identified physical function and HRQoL trajectories. Multinomial logistic regression analyzed the associations between the covariates of interest and trajectory assignment (We included 983 participants with HKOA. We identified three trajectories for each outcome: “consistently low disability” (32.0%), “slightly worsening moderate disability” (47.0%), and “consistently high disability” (21.0%) for physical function; “consistently high HRQoL” (18.3%), “consistently moderate HRQoL” (48.4%) and “consistently low HRQoL” (33.4%) for HRQoL. Age ≥ 75 years, female sex, multimorbidity, and high baseline clinical severity were associated with higher risk of assignment to poorer physical function and HRQoL trajectories. Participants with high education level and with regular physical activity had a lower risk of assignment to a poor trajectory. Unmanageable pain levels increased the risk of assignment to the “consistently moderate HRQoL” trajectory.Although the trajectories of physical function and HRQoL remained stable over 10 years, approximately 70% of people with HKOA maintained moderate or low physical function and HRQoL over this period. Modifiable risk factors like physical activity, multimorbidity and clinical severity were associated with poorer physical function and HRQoL trajectories. These risk factors may be considered in tailored healthcare interventions.The online version contains supplementary material available at 10.1186/s12889-023-16167-9. Osteoarthritis (OA) is the most common joint disease, affecting 519 million people worldwide in 2019 .People with HKOA experience acute and chronic pain and limitations on physical function as well as progressive negative consequences for their mental health, health-related quality of life (HRQoL), and participation in social, leisure, and occupational activities . OA is aFew studies have analyzed the long-term trajectories of physical function and HRQoL in people with HKOA. The systematic review by Wieczorek et al. (2020) has suggested that pain and physical function trajectories are stable over time, but some people might improve their symptoms 6). HRQoL has been a less studied outcome, and to our knowledge, only two studies from the Osteoarthritis Iniciative analyzed the trajectory of HRQoL in the long term, in people with knee OA, with similar results . Studies. HRQoL hThis study aimed to identify longitudinal trajectories of physical function and HRQoL over 10 years and identify the sociodemographic, lifestyle, and clinical variables associated with different trajectories. Secondarily, this study aimed to describe the patterns of specific dimensions of physical function and HRQoL in the 10-year period.N = 10,661) collected baseline data from September 2011 to December 2013; EpiDoC 2  started in March 2013 and ended in July 2015; EpiDoC 3  started in September 2015 and ended in July 2016; and the most recent wave, EpiDoC 4 , occurred from March to August 2021. The baseline evaluation (EpiDoC 1) aimed to estimate the prevalence of 12 rheumatic and musculoskeletal diseases (RMDs) in Portugal, including HKOA. EpiDoC 1 was performed in two phases. In the first phase, trained research assistants conducted face-to-face interviews with a structured questionnaire to collect data on socioeconomic status, chronic non-communicable diseases, HRQoL, and healthcare resource consumption and screen for RMDs. The second phase integrated a clinical appointment for all participants who screened positive for RMDs and 20% of those with negative RMD screenings who agreed to participate. Each appointment consisted of a structured evaluation with a rheumatologist — including laboratory and imaging exams, if needed — to validate the RMD diagnosis and evaluate the patient’s disease-related information  and 5 [b) and d)] trajectory groups.Table S6. Trajectory model diagnostic criteria. Table S7. Univariate Multinomial Logistic Regression models for theassociation of baseline characteristics and physical function (HAQ)trajectories. Table S8.Univariate Multinomial Logistic Regression models for the association of HKOApatients baseline characteristics and HRQoL (EQ-5D) trajectories.


Opa1 mutation from mixed C3H; C57BL/6 J to a pure C57BL/6 J background. We no longer observed retinal and optic nerve abnormalities; the findings indicated no degeneration, but rather a sex-dependent negative effect on RGC connectivity. This highlights the fact that RGC synaptic alteration might precede neuronal death, as has been proposed in other neurodegenerative diseases, providing new clinical considerations for early diagnosis as well as a new therapeutic window for DOA. Furthermore, our results demonstrate the importance of secondary genetic factors in the variability of DOA expressivity and offer a model for screening for aggravating environmental and genetic factors.Dominant optic atrophy (DOA) is mainly caused by OPA1 mutations and is characterized by the degeneration of retinal ganglion cells (RGCs), whose axons form the optic nerve. The penetrance of DOA is incomplete and the disease is marked by highly variable expressivity, ranging from asymptomatic patients to some who are totally blind or who suffer from multisystemic effects. No clear genotype–phenotype correlation has been established to date. Taken together, these observations point toward the existence of modifying genetic and/or environmental factors that modulate disease severity. Here, we investigated the influence of genetic background on DOA expressivity by switching the previously described DOA mouse model bearing the c.1065 + 5G → A  We evaluated the impact of this genetic change on retinal and optic nerve integrity. Overall, our data provide evidence that (1) genetic background contributes to the phenotypic variability observed in DOA; (2) alteration of the connectivity of the RGCs is involved in DOA pathogenesis; and (3) our “new” model offers an exciting opportunity to analyze genetic and environmental contributions to DOA pathogenesis.Three mouse models have been developed to better understand DOA pathophysiology, and in particular, to assess the impact of bilities . As in tbilities . Hence, lleagues , which c2.2.1.ad libitum water and food. All procedures were conducted in accordance with European Union policies (2010/63/EU). According to European regulations, only the use of animals in experimental procedures requires the approval of an ethics committee. Article R214-89 of the French Rural Code specifies that the killing of animals for the sole purpose of using their organs or tissues is not considered an experimental procedure.Mice were bred and reared in the animal facility at the CBI  under a standard 12/12 h light–dark cycle and were provided with Opa1 c.1065 + 5G → A mutation from Alavi and collaborators (329-355del C57BL/6 J mouse model). The genotype of the mice was systematically checked by PCR followed by restriction enzyme digestion analysis (as the mutation removes an AccI restriction site), after birth and after euthanasia of each mouse.Male mice carrying the borators  were cro2.2.Retinas were prepared from eyes freshly isolated from euthanized mice as previously described . These w2.3.Eyes were freshly isolated from euthanized mice as previously described . The corFor counting of RGCs, Brn3-a immuno-stained sections were scanned using a NanoZoomer 2.0 HT (Hamamatsu). Automatic counting was performed using the ORBIT Image Analysis software package. Three to four retinal sections were analyzed for each mouse of each genotype.GFAP, Iba1, and synaptophysin immuno-stained retinal sections were imaged using a Leica SP8 confocal microscope. GFAP staining (one section per mouse) was examined on z-stacked images . Iba1 staining was investigated on three-dimensional images using the Imaris XT software package (Bitplane AG). Iba1-positive cells were counted on six retinal sections per mouse. The total process length of 18–20 Iba1-positive cells per mouse was measured using the Imaris Filament Tracer module. For synaptophysin experiments, all the confocal microscope settings were kept the same across all compared samples. Labeling was quantified on z-stacked images using ImageJ. The analysis was performed for each section (one per mouse) on a defined ROI corresponding to the entire inner or outer plexiform layer. For all images, the same threshold was applied, and the mean gray value was measured for each ROI and normalized to the area of the corresponding ROI.2.4.Optic nerves were isolated from freshly euthanized mice , then fi2.5.t-tests or Mann–Whitney tests, depending on the normality of the distribution of the data, which was checked using the Kolmogorov–Smirnov normality test. The type of statistical test used in each case, as well as the corresponding p-values, is found in the relevant figure legend.Results are expressed in the form mean ± SEM. Statistical analyses were performed using the GraphPad Prism software package. For comparisons between OPA1-mutated and control mice, we performed either unpaired Student’s 3.3.1.329-355del C57BL/6 J (DOA) mice and their wild-type (WT) littermates were estimated by Western blot  loss in the ganglion cell layer (GCL) were evaluated by DAPI staining and immunodetection using antibodies against Brn3-a (a specific marker of RGCs) on retinal sections, respectively . There w3.3.329-355del C57BL/6 J mouse model.Inflammatory status in the retina was assessed by immunostaining of astrocytes and Müller cells (GFAP staining) as well as microglial cells (Iba1 staining) ,B. GFAP-3.4.329-355del C57BL/6 J mice.To evaluate synaptic integrity, we performed synaptophysin immunostaining, as an index of presynaptic compartments, on retinal sections . As expe3.5.329-355del C57BL/6 J mice.An evaluation of optic nerve integrity was conducted via transmission electron microscopy . We firs4.OPA1. These two pathologies present with incomplete penetrance and highly variable expressivity, suggesting that other genetic and/or environmental factors might influence the onset and the severity of the disease , which is primarily caused by mutations in three mtDNA-encoded subunits of the mitochondrial respiratory complex I, and dominant optic atrophy (DOA), mainly attributable to mutations of the nuclear gene  disease . For LHO disease . Thus fa disease .329-355del) previously described by Alavi and collaborators (Opa1 gene (c.1065 + 5G → A), resulting in OPA1 haploinsufficiency. In our study, the presence of the Opa1 mutation and the 50% reduction in protein levels were systematically confirmed by PCR and Western blot, respectively.To evaluate the influence of genetic background on DOA expressivity, we switched the DOA mouse model   . In contctively) , while wctively) . Howeverctively) . It shouctively)  and in tctively) . RGC losctively) , but evictively) , triggerctively) . InteresAG model , 2016, bOP model . LikewisOpa1 gene are present in each of the three DOA mouse models, all trigger OPA1 haploinsufficiency, suggesting similar pathological consequences. However, all these models were developed in the context of different genetic backgrounds: mixed C3H; C57BL/6 J in the case of the Opa1329-355del model, mixed 129/SvPas; C57BL/6 J for the Opa1delTTAG model, and C57BL/6 after outcrossing over at least 4 to 5 generations for the Opa1Q285STOP model. Hence, we assume that the occurrence or absence of retinal and optic nerve defects depends on the presence of aggravating or protective genetic co-factors. This hypothesis is largely supported by the results we obtained by changing the genetic background for the same mutation. Overall, the observed differences suggest the contribution of secondary modifying factors. Comparison of the C57BL/6 J reference genome with 36 other sequenced mouse strains indicates numerous coding genes whose loss-of-function mutations may contribute to phenotypic differences  than in male mice (11 months) (While most previous studies agree that both sexes are similarly affected in DOA (or do not consider sex), we report a sex-dependent synaptic phenotype. This is consistent with data recently obtained for the Opa1sruption . Moreove months) . Interes months) . Sex dif months) . It woulOpa1 mutation was found to lead to distinct phenotypes in different genetic environments. This parallels the heterogeneity observed in patients, given that some mutations have consequences only in some individuals (incomplete penetrance) or present a range of forms of symptomatic expression between individuals (variable expressivity). Recently, several genes whose depletion can counteract the impairments to mitochondrial dynamics induced by OPA1 deficiency have been identified and could thus be tested as genetic modifying factors  by staff with all the necessary qualifications.DA, VG, PP, FK, MA, and MS performed the experiments. DA and PB designed the experiments. BW provided the original DOA mice. MB was in charge of the new DOA mouse model. PB supervised the work. DA, MS, and PB wrote the article. All authors contributed to the article and approved the submitted version.This work was supported by CNRS, Université Toulouse III, Aviesan Neurosciences/Unadev , Retina France , and AFM (20277). DA was the recipient of a PhD fellowship from Fondation de France (00099527) and MA was the recipient of a Postdoctoral Fellowship-Becas Chile CONICYT and ANID Fondecyt Iniciación (11220120).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.