Predicting Colorectal Surgery Readmission Risk: a Surgery-Specific Predictive Model.

Most current predictive models for risk of readmission were primarily designed from non-surgical patients and often utilize administrative data alone. Models built upon comprehensive data sources specific to colorectal surgery may be key to implementing interventions aimed at reducing readmissions. This study aimed to develop a predictive model for risk of 30-day readmission specific to colorectal surgery patients including administrative, clinical, laboratory, and socioeconomic status (SES) data. Patients admitted to the colorectal surgery service who underwent surgery and were discharged from an academic tertiary hospital between 2017 and 2019 were included. A total of 1549 patients met eligibility criteria for this retrospective split-sample cohort study. The 30-day readmission rate of the cohort was 19.62%. A multivariable logistic regression was developed (C=0.70, 95% CI 0.61-0.73), which outperformed two internationally used readmission risk prediction indices (C=0.58, 95% CI 0.52-0.65) and (C=0.60, 95% CI 0.53-0.66). Tailored surgery-specific readmission models with comprehensive data sources outperform the most used readmission indices in predicting 30-day readmission in colorectal surgery patients. Model performance is improved by using more comprehensive datasets that include administrative and socioeconomic details about a patient, as well as clinical information used for decision-making around the time of discharge.

Intracellular reactive oxygen species trafficking participates in seed dormancy alleviation in Arabidopsis seeds.

Reactive oxygen species (ROS) release seed dormancy through an unknown mechanism. We used different seed dormancy-breaking treatments to decipher the dynamics and localization of ROS production during seed germination. We studied the involvement of ROS in the breaking of Arabidopsis seed dormancy by cold stratification, gibberellic acid (GA3 ) and light. We characterized the effects of these treatments on abscisic acid and gibberellins biosynthesis and signalling pathways. ROS, mitochondrial redox status and peroxisomes were visualized and/or quantified during seed imbibition. Finally, we performed a cytogenetic characterization of the nuclei from the embryonic axes during seed germination. We show that mitochondria participate in the early ROS production during seed imbibition and that a possible involvement of peroxisomes in later stages should still be analysed. At the time of radicle protrusion, ROS accumulated within the nucleus, which correlated with nuclear expansion and chromatin decompaction. Taken together, our results provide evidence of the role of ROS trafficking between organelles and of the nuclear redox status in the regulation of seed germination by dormancy.

Analysis of a Modified Two-Stage Approach to Ileal Pouch-Anal Anastomosis Without Fecal Diversion in Pediatric Patients.

BACKGROUND: Fecal diversion after ileal pouch anal anastomosis (IPAA) in children with ulcerative colitis (UC) remains controversial. We hypothesize that a modified two-stage IPAA omitting diverting ileostomy (DI) after IPAA, found to be safe in adults, would produce similar results in children. METHODS: Retrospective, single-institution study of children (≤18 years) undergoing staged total proctocolectomy with IPAA from 2014 to 2020. Traditional two-stage and three-stage approaches including DI after IPAA were compared to two-stage approach without DI. RESULTS: 32 patients were included; of these, 7 (22%), 14 (44%), and 11 (34%) patients underwent traditional two-stage, modified two-stage, or three-stage IPAA, respectively. Following IPAA, modified two-stage patients had shorter operative time, decreased opioid utilization, quicker return to regular diet, and shorter stoma duration. After IPAA, there was similar postoperative length of stay, complication rates, readmissions, visits to the emergency department, or unplanned return to the operating room (OR) within 30 days. Anastomotic leak occurred in 2 patients; both were managed nonoperatively without evidence of pouch dysfunction. CONCLUSION: Modified two-stage IPAA with omission of DI after the IPAA stage is safe to perform in pediatric UC patients. Prospective studies with larger sample sizes are needed to identify risk factors associated with operative complications.

A Comparison of Adult and Pediatric Enhanced Recovery after Surgery Pathways: A Move for Standardization.

BACKGROUND: Enhanced recovery protocols (ERP) are a multimodal approach to standardize perioperative care. To substantiate the benefit of a pediatric-centered pathway, we compared outcomes of children treated with pediatric ERP (pERP) versus adult (aERP) pathways. We aimed to compare components of each pathway to create a new comprehensive pERP to reduce variation in care. METHODS: Retrospective study of children (≤18 y) undergoing elective colorectal surgery from August 2015 to April 2019 at a single institution managed with pERP versus aERP. Multivariable linear and logistic regression, adjusting for demographics and operation characteristics, were used to compare outcomes. RESULTS: Out of 100 hospitalizations (72 patients) were identified, including 37 treated with pERP. pERP patients were, on average, younger (13 versus 16 y), more likely to be ASA III (70% versus 30%), and more likely to receive regional (32% versus 3%) or neuraxial (35% versus 8%) anesthesia. Epidural use was an independent risk factor for longer length of stay (P = 0.000). After adjustment, pERP patients had similar LOS and time to oral intake, but shorter foley duration. pERP patients used significantly fewer opioids and were less likely to return to the operating room within 30 d. 30-d readmissions and ED visits were also lower, but this was not statistically significant. CONCLUSIONS: At our institution, data from both ERPs contributed formation of a synthesized pathway and reflected the pERP approach to opioid utilization and the aERP approach to earlier enteral nutrition.

An enhanced recovery after surgery pathway in pediatric colorectal surgery improves patient outcomes.

INTRODUCTION: Enhanced recovery after surgery (ERAS) pathways in adult colorectal surgery are known to reduce complications, readmissions, and length of stay (LOS). However, there is a paucity of ERAS data for pediatric colorectal surgery. METHODS: A 2014-2018 single-institution, retrospective cohort study was performed on pediatric colorectal surgery patients (2-18 years) pre- and post-ERAS pathway implementation. Bivariate analysis and linear regression were used to determine if ERAS pathway implementation reduced total morphine milligram equivalents per kilogram (MME/kg), LOS, and time to oral intake. RESULTS: 98 (70.5%) and 41 (29.5%) patients were managed with ERAS and non-ERAS pathways, respectively. There was no statistical difference in age, sex, diagnosis, or use of laparoscopic technique between cohorts. The ERAS cohort experienced a significant reduction in total MME/kg, Foley duration, time to oral intake, and LOS with no increase in complications. The presence of an ERAS pathway reduced the total MME/kg (-0.071, 95% CI -0.10, -0.043) when controlling for covariates. CONCLUSION: The use of an ERAS pathway reduces opioid utilization, which is associated with a reduction in LOS and expedites the initiation of oral intake, in colorectal pediatric surgery patients. Pediatric ERAS pathways should be incorporated into the care of pediatric patients undergoing colorectal surgery. LEVEL OF EVIDENCE: Level III evidence. TYPE OF STUDY: Retrospective cohort study.

Decreased Colonic Activin Receptor-Like Kinase 1 Disrupts Epithelial Barrier Integrity in Patients With Crohn's Disease.

BACKGROUND & AIMS: Intestinal epithelial cell (IEC) barrier dysfunction is critical to the development of Crohn's disease (CD). However, the mechanism is understudied. We recently reported increased microRNA-31-5p (miR-31-5p) expression in colonic IECs of CD patients, but downstream targets and functional consequences are unknown. METHODS: microRNA-31-5p target genes were identified by integrative analysis of RNA- and small RNA-sequencing data from colonic mucosa and confirmed by quantitative polymerase chain reaction in colonic IECs. Functional characterization of activin receptor-like kinase 1 (ACVRL1 or ALK1) in IECs was performed ex vivo using 2-dimensional cultured human primary colonic IECs. The impact of altered colonic ALK1 signaling in CD for the risk of surgery and endoscopic relapse was evaluated by a multivariate regression analysis and a Kaplan-Meier estimator. RESULTS: ALK1 was identified as a target of miR-31-5p in colonic IECs of CD patients and confirmed using a 3'-untranslated region reporter assay. Activation of ALK1 restricted the proliferation of colonic IECs in a 5-ethynyl-2-deoxyuridine proliferation assay and down-regulated the expression of stemness-related genes. Activated ALK1 signaling increased colonic IEC differentiation toward colonocytes. Down-regulated ALK1 signaling was associated with increased stemness and decreased colonocyte-specific marker expression in colonic IECs of CD patients compared with healthy controls. Activation of ALK1 enhanced epithelial barrier integrity in a transepithelial electrical resistance permeability assay. Lower colonic ALK1 expression was identified as an independent risk factor for surgery and was associated with a higher risk of endoscopic relapse in CD patients. CONCLUSIONS: Decreased colonic ALK1 disrupted colonic IEC barrier integrity and was associated with poor clinical outcomes in CD patients.

Crohn's Disease Differentially Affects Region-Specific Composition and Aerotolerance Profiles of Mucosally Adherent Bacteria.

BACKGROUND: The intestinal microbiota play a key role in the onset, progression, and recurrence of Crohn disease (CD). Most microbiome studies assay fecal material, which does not provide region-specific information on mucosally adherent bacteria that directly interact with host systems. Changes in luminal oxygen have been proposed as a contributor to CD dybiosis. METHODS: The authors generated 16S rRNA data using colonic and ileal mucosal bacteria from patients with CD and without inflammatory bowel disease. We developed profiles reflecting bacterial abundance within defined aerotolerance categories. Bacterial diversity, composition, and aerotolerance profiles were compared across intestinal regions and disease phenotypes. RESULTS: Bacterial diversity decreased in CD in both the ileum and the colon. Aerotolerance profiles significantly differed between intestinal segments in patients without inflammatory bowel disease, although both were dominated by obligate anaerobes, as expected. In CD, high relative levels of obligate anaerobes were maintained in the colon and increased in the ileum. Relative abundances of similar and distinct taxa were altered in colon and ileum. Notably, several obligate anaerobes, such as Bacteroides fragilis, dramatically increased in CD in one or both intestinal segments, although specific increasing taxa varied across patients. Increased abundance of taxa from the Proteobacteria phylum was found only in the ileum. Bacterial diversity was significantly reduced in resected tissues of patients who developed postoperative disease recurrence across 2 independent cohorts, with common lower abundance of bacteria from the Bacteroides, Streptococcus, and Blautia genera. CONCLUSIONS: Mucosally adherent bacteria in the colon and ileum show distinct alterations in CD that provide additional insights not revealed in fecal material.

Microtubule self-organisation during seed germination in Arabidopsis.

BACKGROUND: Upon water uptake and release of seed dormancy, embryonic plant cells expand, while being mechanically constrained by the seed coat. Cortical microtubules (CMTs) are key players of cell elongation in plants: their anisotropic orientation channels the axis of cell elongation through the guidance of oriented deposition of load-bearing cellulose microfibrils in the cell wall. Interestingly, CMTs align with tensile stress, and consistently, they reorient upon compressive stress in growing hypocotyls. How CMTs first organise in germinating embryos is unknown, and their relation with mechanical stress has not been investigated at such an early developing stage. RESULTS: Here, we analysed CMT dynamics in dormant and non-dormant Arabidopsis seeds by microscopy of fluorescently tagged microtubule markers at different developmental time points and in response to abscisic acid and gibberellins. We found that CMTs first appear as very few thick bundles in dormant seeds. Consistently, analysis of available transcriptome and translatome datasets show that limiting amounts of tubulin and microtubule regulators initially hinder microtubule self-organisation. Seeds imbibed in the presence of gibberellic acid or abscisic acid displayed altered microtubule organisation and transcriptional regulation. Upon the release of dormancy, CMTs then self-organise into multiple parallel transverse arrays. Such behaviour matches the tensile stress patterns in such mechanically constrained embryos. This suggests that, as CMTs first self-organise, they also align with shape-derived tensile stress patterns. CONCLUSIONS: Our results provide a scenario in which dormancy release in the embryo triggers microtubule self-organisation and alignment with tensile stress prior to germination and anisotropic growth.

Effects of Gender Bias and Stereotypes in Surgical Training: A Randomized Clinical Trial.

Importance: Factors contributing to underrepresentation of women in surgery are incompletely understood. Pro-male bias and stereotype threat appear to contribute to gender imbalance in surgery. Objectives: To evaluate the association between pro-male gender bias and career engagement and the effect of stereotype threat on skill performance among trainees in academic surgery. Design, Setting, and Participants: A 2-phase study with a double-blind, randomized clinical trial component was conducted in 3 academic general surgery training programs. Residents were recruited between August 1 and August 15, 2018, and the study was completed at the end of that academic year. In phase 1, surveys administered 5 to 6 months apart investigated the association of gender bias with career engagement. In phase 2, residents were randomized 1:1 using permuted-block design stratified by site, training level, and gender to receive either a trigger of or protection against stereotype threat. Immediately after the interventions, residents completed the Fundamentals of Laparoscopic Surgery (FLS) assessment followed by a final survey. A total of 131 general surgery residents were recruited; of these 96 individuals with academic career interests met eligibility criteria; 86 residents completed phase 1. Eighty-five residents were randomized in phase 2, and 4 residents in each arm were lost to follow-up. Intervention: Residents read abstracts that either reported that women had worse laparoscopic skill performance than men (trigger of stereotype threat [A]) or had no difference in performance (protection against stereotype threat [B]). Main Outcomes and Measures: Association between perception of pro-male gender bias and career engagement survey scores (phase 1) and stereotype threat intervention and FLS scores (phase 2) were the outcomes. Intention-to-treat analysis was conducted. Results: Seventy-seven residents (38 women [49.4%]) completed both phases of the study. The association between pro-male gender bias and career engagement differed by gender (interaction coefficient, -1.19; 95% CI, -1.90 to -0.49; P = .02); higher perception of bias was associated with higher engagement among men (coefficient, 1.02; 95% CI, 0.19-2.24; P = .04), but no significant association was observed among women (coefficient, -0.25; 95% CI, -1.59 to 1.08; P = .50). There was no evidence of a difference in FLS score between interventions (mean [SD], A: 395 [150] vs B: 367 [157]; P = .51). The response to stereotype threat activation was similar in men and women (interaction coefficient, 15.1; 95% CI, -124.5 to 154.7; P = .39). The association between stereotype threat activation and FLS score differed by gender across levels of susceptibility to stereotype threat (interaction coefficient, -35.3; 95% CI, -47.0 to -23.6; P = .006). Higher susceptibility to stereotype threat was associated with lower FLS scores among women who received a stereotype threat trigger (coefficient, -43.4; 95% CI, -48.0 to -38.9; P = .001). Conclusions and Relevance: Perception of pro-male bias and gender stereotypes may influence career engagement and skill performance, respectively, among surgical trainees. Trial Registration: ClinicalTrials.gov Identifier: NCT03623009.

Chronic Antibiotic Dependent Pouchitis Is Associated With Older Age at the Time of Ileal Pouch Anal Anastomosis (J-pouch) Surgery.

BACKGROUND: Risk factors for the development of chronic antibiotic dependent pouchitis (CADP) are not well understood. METHODS: Using multivariable logistic regression, we compared clinical factors between 194 patients with acute antibiotic responsive pouchitis or CADP. RESULTS: Individuals with CADP were significantly older (40.9 vs 30.8 years, P < 0.001) and demonstrated a longer disease duration before IPAA (10.3 vs 7.0 years, P = 0.004). Age ≥55 years at the time of IPAA was significantly associated with CADP (adjusted odds ratio = 4.35, 95% confidence interval = 1.01-18.7). CONCLUSIONS: Although older age should not represent a barrier to IPAA, further studies evaluating etiologies of this association are warranted.

Management of Freely Perforated Diverticulitis.

CASE SUMMARY: A healthy 65-year-old woman presents to the emergency department with a 12-hour history of sudden-onset severe lower abdominal pain. This is her first episode. She reports nausea, vomiting, and anorexia. Her last colonoscopy was at age 60, and was normal, except for diverticulosis of the sigmoid colon. Physical examination is significant for fever, tachycardia, and generalized abdominal pain with rebound tenderness. Pertinent laboratory findings include a leukocytosis and metabolic acidosis. A CT scan is obtained and is consistent with freely perforated diverticulitis, including a thickened sigmoid colon, free fluid in the pelvis, and free air noted near the diaphragm (). The surgeon completes the patient evaluation, recommends initiation of intravenous fluid resuscitation and antibiotics, and plans to go immediately to the operating room for surgical resection.

Arabidopsis S2Lb links AtCOMPASS-like and SDG2 activity in H3K4me3 independently from histone H2B monoubiquitination.

BACKGROUND: The functional determinants of H3K4me3, their potential dependency on histone H2B monoubiquitination, and their contribution to defining transcriptional regimes are poorly defined in plant systems. Unlike in Saccharomyces cerevisiae, where a single SET1 protein catalyzes H3K4me3 as part of COMPlex of proteins ASsociated with Set1 (COMPASS), in Arabidopsis thaliana, this activity involves multiple histone methyltransferases. Among these, the plant-specific SET DOMAIN GROUP 2 (SDG2) has a prominent role. RESULTS: We report that SDG2 co-regulates hundreds of genes with SWD2-like b (S2Lb), a plant ortholog of the Swd2 axillary subunit of yeast COMPASS. We show that S2Lb co-purifies with the AtCOMPASS core subunit WDR5, and both S2Lb and SDG2 directly influence H3K4me3 enrichment over highly transcribed genes. S2Lb knockout triggers pleiotropic developmental phenotypes at the vegetative and reproductive stages, including reduced fertility and seed dormancy. However, s2lb seedlings display little transcriptomic defects as compared to the large repertoire of genes targeted by S2Lb, SDG2, or H3K4me3, suggesting that H3K4me3 enrichment is important for optimal gene induction during cellular transitions rather than for determining on/off transcriptional status. Moreover, unlike in budding yeast, most of the S2Lb and H3K4me3 genomic distribution does not rely on a trans-histone crosstalk with histone H2B monoubiquitination. CONCLUSIONS: Collectively, this study unveils that the evolutionarily conserved COMPASS-like complex has been co-opted by the plant-specific SDG2 histone methyltransferase and mediates H3K4me3 deposition through an H2B monoubiquitination-independent pathway in Arabidopsis.

Twelve Tips for Preparing a Surgical Discharge Summary: Enabling a Safe Discharge.

This article was migrated. The article was marked as recommended. The surgical discharge summary allows the perioperative care team to summarize a recent hospitalization and relay important information to a variety of invested parties including other healthcare providers, outpatient caregivers, and the surgical patient. The inpatient care team can promote a smooth transition of care and empower outpatient providers and the patient to foster a confident progression through recovery. We describe twelve tips for a streamlined, successful discharge summary geared towards the surgical intern. A successful surgical discharge summary begins with patient and caregiver collaboration, communication and teamwork, and culminates with concise documentation. These tips reflect a review of the current literature and rely on the clinical expertise of an interdisciplinary surgical team. Our aim is to empower surgical educators and trainees to understand the complexity of discharge planning, and to improve the efficiency with which it can be completed and the quality of the discharge process at their training hospitals.

Risk factors for 30-day readmission after colorectal surgery: does transfer status matter?

BACKGROUND: Rates of readmission after colorectal surgery (CRS) range from 9% to 25% and cost the US $300 million annually. The aim of this study was to identify risk factors for 30-d readmission after CRS. Our hypothesis was that transfer from an outside hospital before CRS increases incidence of 30-d readmission. METHODS: Using the Healthcare Cost and Utilization Project Nationwide Readmissions Database, a retrospective analysis of surviving adult patients who underwent inpatient colon and/or rectal resection from 2010 to 2014 was performed. Using multivariable logistic regression, we assessed the direct effect of potential risk factors for readmission, including demographics, hospital characteristics, comorbidities, indication for CRS, and transfer status to the index hospital where the CRS was performed. RESULTS: A total of 336,792 patients were included, and 13% (n = 43,546) were readmitted within 30 d. Overall, 1% of patients (n = 3652) were transferred from another hospital for surgery, and these patients were more likely to be older, have comorbidities, have public insurance, and have low income. After adjustment, age, insurance type, household income, comorbidities, and primary indication for CRS were all significant predictors of readmission. Transfer status did not meaningfully impact the incidence of readmission after accounting for patient and hospital characteristics (aOR 1.08, 95% CI 1.00, 1.18). Patients with primary indications of trauma (aOR 1.88, 95% CI 1.48, 2.38), inflammatory bowel disease (aOR 1.64, 95% CI 1.1.56, 1.71), and ischemia (aOR 1.77, 95% CI 1.1.59, 1.97) were most likely to be readmitted. Patients treated at a rural nonteaching hospital, compared with those at a urban teaching hospital, were significantly less likely to be readmitted (aOR 0.80, 95% CI 0.76, 0.83). CONCLUSIONS: Preoperative factors, such as patient comorbidities and primary indications for CRS, are important risk factors for postoperative readmission. Although transfer status does not independently predict readmission, it serves as a proxy for a high-risk group of patients that could be targeted for future interventions.

Colonic epithelial miR-31 associates with the development of Crohn's phenotypes.

BACKGROUND: Crohn's disease (CD) is highly heterogeneous, due in large part to variability in cellular processes that underlie the natural history of CD, thereby confounding effective therapy. There is a critical need to advance understanding of the cellular mechanisms that drive CD heterogeneity. METHODS: We performed small RNA sequencing of adult colon tissue from CD and NIBD controls. Colonic epithelial cells and immune cells were isolated from colonic tissues, and microRNA-31 (miR-31) expression was measured. miR-31 expression was measured in colonoid cultures generated from controls and patients with CD. We performed small RNA-sequencing of formalin-fixed paraffin-embedded colon and ileum biopsies from treatment-naive pediatric patients with CD and controls and collected data on disease features and outcomes. RESULTS: Small RNA-sequencing and microRNA profiling in the colon revealed 2 distinct molecular subtypes, each with different clinical associations. Notably, we found that miR-31 expression was a driver of these 2 subtypes and, further, that miR-31 expression was particularly pronounced in epithelial cells. Colonoids revealed that miR-31 expression differences are preserved in this ex vivo system. In adult patients, low colonic miR-31 expression levels at the time of surgery were associated with worse disease outcome as measured by need for an end ileostomy and recurrence of disease in the neoterminal ileum. In pediatric patients, lower miR-31 expression at the time of diagnosis was associated with future development of fibrostenotic ileal CD requiring surgeryCONCLUSIONS. These findings represent an important step forward in designing more effective clinical trials and developing personalized CD therapies. FUNDING: This work was supported by CCF Career Development Award (SZS), R01-ES024983 from NIEHS (SZS and TSF), 1R01DK104828-01A1 from NIDDK (SZS and TSF), P01-DK094779-01A1 from NIDDK (SZS), P30-DK034987 from NIDDK (SZS), 1-16-ACE-47 ADA Pathway Award (PS), UNC Nutrition Obesity Research Center Pilot & Feasibility Grant P30DK056350 (PS), CCF PRO-KIIDS NETWORK (SZS and PS), UNC CGIBD T32 Training Grant from NIDDK (JBB), T32 Training Grant (5T32GM007092-42) from NIGMS (MH), and SHARE from the Helmsley Trust (SZS). The UNC Translational Pathology Laboratory is supported, in part, by grants from the National Cancer Institute (3P30CA016086) and the UNC University Cancer Research Fund (UCRF) (PS).

Variant Two-Stage Ileal Pouch-Anal Anastomosis: An Innovative and Effective Alternative to Standard Resection in Ulcerative Colitis.

BACKGROUND: Ulcerative colitis patients have been historically treated with standard single, 2-, and 3-stage operative approaches. We perform a variant 2-stage procedure beginning with total abdominal colectomy and end ileostomy followed by completion proctectomy and ileal pouch-anal anastomosis (IPAA) without a diverting loop ileostomy. This study evaluates the effectiveness of this innovative alternative. STUDY DESIGN: Patients with ulcerative colitis, admitted to the University of North Carolina Hospital between 2003 and 2010 for IPAA, were eligible for inclusion. The 3-year cumulative incidence of pouch leaks among patients undergoing variant 2-stage were compared with those undergoing classic 2-stage, using inverse probability-of-treatment weighted Kaplan- Meier survival curves, and 95% CIs were estimated using nonparametric bootstrapping. RESULTS: There were 248 patients who underwent IPAA; 139 (56.1%) underwent classic 2-stage and 109 (43.9%) underwent variant 2-stage. After standardization, there was no significant difference in the 3-year cumulative incidence of pouch leaks between patients undergoing variant 2-stage, compared with the standard single- or 2-stage procedure (risk difference 0.01; 95% CI -0.08, 0.15). At the time of the first surgical procedure, patients undergoing a variant 2-stage were more likely to have lower BMIs (median 22.5 kg/m2 vs 26.7 kg/m2; p < 0.0001), an urgent/emergent procedure (56.9% vs 0.0%; p < 0.0001), biologic use within 2 weeks of surgery (32.1% vs 17.5%; p = 0.003), and high dose steroid use (60.4% vs 16.7%; p ≤ 0.0001). CONCLUSIONS: Variant 2-stage IPAA is a safe and effective operative approach with comparable outcomes in a more acute population based on BMI, steroid use, and urgency of operation.