Progesterone and Its Metabolites Play a Beneficial Role in Affect Regulation in the Female Brain.

Premenstrual dysphoric disorder is a female affective disorder that is defined by mood symptoms. The condition is linked to unstable progesterone concentrations. Progestin supplementation is given in cases of threatened or recurrent miscarriage and for luteal phase support. Progesterone is essential for implantation, immune tolerance, and modulation of uterine contractility. For a long time, the administration of progestins was associated with an unfavorable impact on mood, leading to negative affect, and, therefore, was contraindicated in existing mood disorders. Establishing the role of the natural progesterone derivative allopregnanolone in advances in the treatment of postpartum depression has shed new light on the general pathophysiology of mood disorders. Allopregnanolone directly interacts with gamma-aminobutyric acid type A (GABA-A) receptors even at nanomolar concentrations and induces significant anti-depressant, anti-stress, sedative, and anxiolytic effects. Postpartum depression is caused by a rapid drop in hormones and can be instantly reversed by the administration of allopregnanolone. Premenstrual dysphoric disorder can also be considered to result from insufficient neuroactive steroid action due to low progesterone derivative concentration, unstable hormone levels, or decreased receptor sensitivity. The decrease in progesterone levels in perimenopause is also associated with affective symptoms and an exacerbation of some psychosomatic syndromes. Bioidentical progesterone supplementation encounters several obstacles, including limited absorption, first-pass effect, and rapid metabolism. Hence, non-bioidentical progestins with better bioavailability were widely applied. The paradoxical, unfavorable effect of progestins on mood can be explained by the fact that progestins suppress ovulation and disturb the endocrine function of the ovary in the luteal phase. Moreover, their distinct chemical structure prevents their metabolism to neuroactive, mood-improving derivatives. A new understanding of progesterone-related mood disorders can translate the study results from case series and observational studies to cohort studies, clinical trials, and novel, effective treatment protocols being developed.

The Urogynecology Section of the Polish Society of Gynecologists and Obstetricians Guideline for the diagnostic assessment of stress urinary incontinence in women.

OBJECTIVES: The aim of the Urogynecology Section of the Polish Society of Gynecologists and Obstetricians (PSGO) was to develop an updated Guideline for the diagnostic assessment of stress urinary incontinence (SUI) in women. MATERIAL AND METHODS: Earlier PSGO guidelines and the literature about the diagnostic assessment of SUI, including current international guidelines, were reviewed. RESULTS: As in the earlier guidelines, the diagnostic process was subdivided into the initial and the specialized diagnostics. Patients who required specialized diagnostic testing were identified. Functional diagnostic tests, performed by physiotherapists, were included. Attention was paid to new diagnostic possibilities. CONCLUSIONS: Initial diagnostic assessment is sufficient to devise the optimal treatment plan in a number of patients. It also allows to identify which patients will require specialized diagnostics, whose scope is individually tailored to the patient needs and depends on symptom complexity, surgical history, treatment plan, experience of the physician, availability of the equipment, and cost-effectiveness ratio.

New Kid on the Block: The Efficacy of Phytomedicine Extracts Urox® in Reducing Overactive Bladder Symptoms in Rats.

The aim of the current study was to determine if phytomedicine (Urox®) would reverse retinyl acetate (RA)-induced changes characteristic of bladder overactivity. There were 60 rats divided into the following 4 groups: I-control, II-received RA to induce detrusor overactivity (DO), III-received Urox (840 mg daily for 14 days), and IV-received combination of RA and Urox®. The cystometry was performed 2 days after the last dose of Urox®. Next, urothelium thickness and biochemical parameter measurements were performed. In group IV, a decrease in basal pressure and detrusor overactivity index was noted when compared to group II. Furthermore, in group IV the following parameters were increased: threshold pressure, voided volume, intercontraction interval, and bladder compliance in comparison with group II. There were significant elevations in c-Fos expression in the neuronal voiding centers in group II, while the expression of c-Fos in group IV was normalized. No significant changes in the values of the analyzed biomarkers in group III were found, while in group II, an elevation in BDNF, NGF, CGRP, ATP, Rho kinase, malondialdehyde, 3-nitrotyrosine, TRPV1, OCT-3, and VAChT and then a decrease in E-cadherin and Z01 were found. A successful restoration of all the abovementioned biomarkers' levels was observed in group IV. Phytomedicine extracts (Urox®) were found to be potent in reversing RA-induced changes in several cystometric and biochemical parameters that are determinants of overactive bladder (OAB). The actions of Urox® were proved to be dependent on several factors, such as growth factors and several OAB biomarkers but not pro-inflammatory cytokines.

Urine Nerve Growth Factor May Not Be Useful as a Biomarker of Overactive Bladder in Patients with Pelvic Organ Prolapse.

(1) Background: Overactive bladder (OAB) symptoms are frequently present in women with pelvic organ prolapse (POP). Although urinary nerve growth factor (NGF) is a promising biomarker of OAB, little is known about its role in patients with OAB secondary to POP. The aim of the study was to evaluate urinary NGF levels in patients with POP involving the anterior vaginal wall and check if it may serve as a predicting factor for postoperative resolution of OAB symptoms. (2) Methods: Eighty-three Caucasian women included in the study were divided into three groups: pure OAB, one associated with POP (POP&OAB) and a control group composed of healthy volunteers. The urine NGF and creatinine were assessed with ELISA tests to calculate the NGF/creatinine ratio. (3) Results: The NGF/creatinine ratio was significantly higher in patients with pure OAB in comparison with other groups; however, it did not differ between the control group and the POP&OAB group. There was no correlation between NGF/creatinine ratio and age, menopausal status, BMI, parity or urodynamic findings. The NGF/creatinine ratio was not a prognostic factor for OAB symptoms' resolution after surgical treatment of POP. (4) Conclusions: Urinary NGF excretion is not increased in women with OAB secondary to POP; thus, it may not serve as an OAB biomarker in these patients.

Urogynecology Section of the Polish Society of Gynecologists and Obstetricians Guidelines on the management of recurrent pelvic organ prolapse.

OBJECTIVES: The aim of the publication was to present the Guideline of the Urogynecology Section of the Polish Society of Gynecologists and Obstetricians (PSGO) for the management of recurrent pelvic organ prolapse, based on the available literature, expert knowledge and opinion, as well as everyday practice. MATERIAL AND METHODS: In 2005, 2006 and 2010, the panel of PSGO experts published guidelines for the diagnosis and treatment of patients with lower urinary tract symptoms (LUTS). This publication presents an update of those recommendations and concerns recurrent POP treatment. MAIN CONCLUSION: The analysis of data revealed that sacrocolpopexy with the use of commercial sets or polypropylene hernia mesh is the method of choice for the surgical repair of recurrent vaginal vault prolapse. However, a significantly higher risk of surgical and postoperative complications after sacrocolpopexy, as compared to vaginal surgeries, should be considered when making treatment decisions. In other types of recurrent POP, the choice of surgery method should be tailored to the individual needs of each patient and may depend on the medical center.

Urogynecology Section of the Polish Society of Gynecologists and Obstetricians guidelines on the management of stress urinary incontinence in women.

OBJECTIVES: The aim was to present an interdisciplinary Guideline of the Urogynecology Section of the Polish Society of Gynecologists and Obstetricians (PSGO) for the management of stress urinary incontinence (SUI). MATERIAL AND METHODS: A review of the literature, including current international guidelines and earlier recommendations of the PSGO Urogynecology Section, about the treatment of SUI was conducted. RESULTS: Management of SUI is presented. Four lines of therapy were identified: line 1 - the so-called 'conservative treatment', which should always be attempted, regardless of SUI symptom severity; line 2 - surgical intervention; lines 3 and 4 - reoperations after unsuccessful surgeries from line 2. The literature reports which provided supporting evidence for this Guideline, including the practical aspects, were discussed. CONCLUSIONS: A systematic review of the guidelines and an analysis of SUI management were conducted. The need for an individualized approach was emphasized.

Psychiatric History and Overactive Bladder Symptom Severity in Ambulatory Urogynecological Patients.

INTRODUCTION AND HYPOTHESIS: A link between psychiatric comorbidities and overactive bladder symptomatology has been suggested by preclinical and clinical studies. Given this, we hypothesized that a psychiatric history and current treatment with psychotropic medications could be related to the severity of overactive bladder and incontinence symptoms in patients referred to a tertiary care urogynecological center. METHODS: One hundred and twenty-seven female patients diagnosed with an overactive bladder were screened for a lifetime history of psychiatric disorders and the type and number of psychotropic medications currently taken. The overall severity of overactive bladder symptoms was assessed using the Indevus Urgency Severity Scale. The severity and impact of urinary incontinence on the quality of life were quantified with the International Consultation on Incontinence Questionnaire-Urinary Incontinence Short Form. Urinary incontinence was further quantified with the aid of the Urinary Distress Inventory-6. The patients were screened for stress urinary incontinence using the Stamey Incontinence Score. RESULTS: A psychiatric history, as well as current use of at least two psychotropic medications, was associated with increased severity of overactive bladder symptoms. A history of depression and current treatment with any selective serotonin reuptake inhibitor was associated with increased severity of stress urinary incontinence symptoms. Current treatment with other psychotropic medications, including sedative-hypnotics and drugs with anticholinergic properties was not related to the severity of overactive bladder and incontinence symptoms.

The Potential of Asiatic Acid in the Reversion of Cyclophosphamide-Induced Hemorrhagic Cystitis in Rats.

The purpose of this study was to determine if asiatic acid may act efficiently in the model of cyclophosphamide (CYP)-induced cystitis in rats. We performed experiments after administration of CYP (single dose 200 mg/kg, intraperitoneally), asiatic acid (30 mg/kg/day for 14 consecutive days, by oral gavage), or CYP plus asiatic acid, during which conscious cystometry, measurements of urothelium thickness and bladder edema, as well as selected biomarkers analyses were conducted. In rats that received asiatic acid together with CYP, a drop in bladder basal pressure, detrusor overactivity index, non-voiding contraction amplitude, non-voiding contraction frequency, and the area under the pressure curve were observed, when compared to the CYP group. Furthermore, a significant increase in threshold pressure, voided volume, intercontraction interval, bladder compliance, and volume threshold to elicit NVC were found in that group accordingly. Administration of the asiatic acid successfully restored concentrations of biomarkers both in bladder urothelium (BDNF, CGRP, OCT-3, IL-1ß, IL-6, NGF, nitrotyrosine, malondialdehyde, TNF-α, SV2A, SNAP23, SNAP25, PAC-1, ORM1, occludin, IGFBP-3, HB-EGF, T-H protein, Z01, and HPX) and detrusor muscle (Rho kinase and VAChT) in CYP-treated rats. Finally, asiatic acid significantly decreased urothelium thickness and bladder oedema. Asiatic acid proved to be a potent and effective drug in the rat model of CYP-induced cystitis.

Bacterial Colonization as a Possible Source of Overactive Bladder Symptoms in Pediatric Patients: A Literature Review.

Overactive Bladder (OAB) is a common condition that is known to have a significant impact on daily activities and quality of life. The pathophysiology of OAB is not completely understood. One of the new hypothetical causative factors of OAB is dysbiosis of an individual urinary microbiome. The major aim of the present review was to identify data supporting the role of bacterial colonization in overactive bladder symptoms in children and adolescents. The second aim of our study was to identify the major gaps in current knowledge and possible areas for future clinical research. There is a growing body of evidence indicating some relationship between qualitative and quantitative characteristics of individual urinary microbiome and OAB symptoms in adult patients. There are no papers directly addressing this issue in children or adolescents. After a detailed analysis of papers relating urinary microbiome to OAB, the authors propose a set of future preclinical and clinical studies which could help to validate the concept in the pediatric population.

The Urogynecology Section of the Polish Society of Gynecologists and Obstetricians Guideline on the use of urodynamic testing in gynecological practice.

OBJECTIVES: The aim was to present an interdisciplinary Guideline of the Urogynecology Section of the Polish Society of Gynecologists and Obstetricians (PSGO) for the use of urodynamics (UDS) in the diagnostic process of patients with lower urinary tract symptoms (LUTS) based on the available literature, expert knowledge, and everyday practice. MATERIAL AND METHODS: A review of the literature concerning the use of UDS in women, including current international guidelines and earlier recommendations of the PSGO Urogynecology Section, was conducted. RESULTS: Urodynamic testing allows to make the urodynamic diagnosis which, nevertheless, remains to be the preliminary diagnosis. Medical history, physical examination, and detailed analysis of the previous test results (laboratory, imaging, endoscopic) need to be taken into consideration before making the final diagnosis. Urodynamic testing before surgical treatment of SUI is allowable, but the decision remains at the discretion of the physician. Urodynamic testing is not necessary before primary surgical treatment of uncomplicated SUI, but it has been demonstrated to optimize the therapeutic methods in complicated SUI. The significance of UDS in the diagnostic process of patients with overactive bladder symptoms, voiding dysfunction, and bladder outlet obstruction was discussed. CONCLUSIONS: Urodynamic testing is a vital element of the urogynecological diagnostic process. The scope of UDS should reflect the individual needs and symptoms of each patient and be based on the current guidelines, expert knowledge and experience of the physician, indications, and eligibility, as well as additional test results of the affected patients. Due to formal and legal requirements, PSGO, in this Guideline, wishes to emphasize the need for an individualized approach to both, test performance and result interpretation.

The Urogynecology Section of the Polish Society of Gynecologists and Obstetricians guidelines on the management of non-neurogenic overactive bladder syndrome in women.

OBJECTIVES: The aim of the publication was to present the interdisciplinary guidelines of the Urogynecology Section of the Polish Society of Gynecologists and Obstetricians (PSGO) for the treatment of overactive bladder (OAB) syndrome based on the available literature, expert knowledge, and everyday practice. MATERIAL AND METHODS: A review of the literature, including current recommendations for the treatment of overactive bladder syndrome, urinary incontinence, urgency and mixed urinary incontinence, as well as the earlier recommendations of the PSGO Urogynecology Section, was conducted. RESULTS: Management of the patients with OAB is presented. Four lines of therapy were identified: 1) educating the patient, behavioral therapy with pelvic floor muscle training, 2) pharmacotherapy, 3) botulinum toxin injection and tibial nerve stimulation; and sacral nerve stimulation even though so far it has been used only in selected populations, 4) surgical intervention. The literature reports which provided supporting evidence and presented various aspects of the therapy were discussed. OAB pharmacotherapy-related issues which are vital in everyday clinical practice were presented. CONCLUSIONS: A systematic review of the available guidelines and an analysis of OAB (including urgency urinary incontinence) management were conducted. The Polish Society of Gynecologists and Obstetricians issued the guidelines for the therapeutic management of OAB patients. The need for an individualized approach was emphasized.

Risk Factors for Surgical Treatment of Endometrial Cancer Using Traditional and Laparoscopic Methods.

Surgical treatment is the most important part of therapy for endometrial cancer. The aim of the study was to define factors having the most significant impact on surgical treatment of endometrial cancer when using traditional and laparoscopic methods. In the study, we evaluated 75 females who were treated for endometrial cancer via laparoscopic surgery in 2019 and used a historical control of 70 patients treated by laparotomy in 2011. The evaluated risk factors included the method of surgery, type of lymphadenectomy, patient's age, various obesity parameters, histological grading, cancer clinical staging, pelvic dimensions, previous abdominal surgeries, comorbidities, and number of deliveries. The duration of hospitalization, operation time, loss of hemoglobin, and procedure-related complications were used as parameters of perioperative outcomes. Multivariable linear regression analysis confirmed the following factors as being predictors of worse perioperative outcomes: laparotomy, abdominal obesity (waist circumstance and waist-to-hip ratio), range of lymphadenectomy, prior abdominal surgeries, and larger pelvic dimensions. Abdominal obesity is a significant risk factor in the treatment of endometrial cancer. Laparotomy continues to be utilized frequently in the management of endometrial cancer in Poland as well as elsewhere, and adopting a minimally invasive approach is likely to be beneficial for patient outcome.

Efficacy and safety of the Calistar and Elevate anterior vaginal mesh procedures.

OBJECTIVES: The pelvic organ prolapse (POP) surgery with implantation of anterior transvaginal mesh (e.g. Elevate or Calistar) may provide objective and subjective improvement as compared to traditional POP repair without mesh. Given differences between the Elevate and the Calistar mesh and their different placement methods, some variation inlong-term clinical outcomes of these anterior vaginal mesh procedures can be expected. STUDY DESIGN: The purpose of the study was to compare the 18-month operative success in patients who had undergone anterior POP surgery with either the Calistar (n = 54) or Elevate mesh (n = 50). RESULTS: There were no between-group differences in objective measures of operative efficacy, including POP-Q anterior stage 0 or I (94% for Calistar, 92% for Elevate) and "no descent beyond the hymen" (98% for Calistar, 94% for Elevate). The proportion of patients with subjective measure of operative efficacy (no vaginal bulge symptoms) did not differ between the groups (91% for Calistar, 78% for Elevate). There were no between-group differences in the proportion of women suffering from vaginal exposure, de novo stress urinary incontinence (SUI), de novo overactive bladder (OAB) symptoms, pelvic floor pain or dyspareunia. The operative cure of OAB symptoms was similar in the groups. The proportion of patients with the operative cure of SUI symptoms was significantly higher in the Calistar as compared to the Elevate group. CONCLUSIONS: The results suggestthat the Calistar system offers similar efficacy in the treatment of anterior and both anterior and apical POP as compared to the Elevate. The use of anterior Calistar is associated with some additional benefits, i.e. SUI treatment in patients with concomitant anterior and both anterior and apical POP and SUI symptoms.

Anti-Müllerian hormone level is associated with vitamin D receptor polymorphisms in women with polycystic ovary syndrome.

OBJECTIVE: Our study aimed to investigate the relationship between polymorphisms (Apa1, Bsm1, Fok1, and Cdx2) in the VDR gene as well as AMH and AMHR2 genes and their influence on AMH and 25(OH)D levels in PCOS women. STUDY DESIGN: Seventy-five patients with PCOS and 23 control women were included. Serum AMH and 25(OH)D levels in patients and controls were measured by enzyme-linked immunosorbent assay (ELISA). Polymorphisms in VDR gene Fok1 C/T (rs2228587), Bsm1 A/G (rs1544410), Apa1 A/C (rs7975232), and Cdx2 A/G (rs11568820) polymorphisms as well as AMH G/T (rs10407022) and AMHR2 A/G (rs2002555) were analyzed using real-time PCR. RESULTS: Analysis of the VDR Cdx2 polymorphism showed a significantly higher frequency of the homozygous GG (mutant) genotype in the PCOS group as compared with the control group (p < 0.05). The analysis revealed a statistically significant correlation between the presence of FokI and ApaI polymorphisms and AMH levels in PCOS women (p < 0.05). The presence of mutant genotypes (CT, TT) in the Fok1 and (CA, CC) in the Apa1 polymorphisms were associated with higher AMH level in PCOS women (p < 0.05). No statistically significant correlations between AMH and AMHR2 polymorphisms and AMH level were found. Moreover, there was no correlation between AMH and 25(OH)D levels in the PCOS or in the control group. CONCLUSION: It seems that the elevated AMH level is associated with VDR Fokl and Apal polymorphisms, but not with 25(OH)D levels in PCOS women. Further research is needed to determine the role of VDR polymorphism in AMH level in PCOS.

Determination of Fructan (Inulin, FOS, Levan, and Branched Fructan) in Animal Food (Animal Feed, Pet Food, and Ingredients): Single-Laboratory Validation, First Action 2018.07.

Traditional enzyme-based methods for measurement of fructan were designed to measure just inulin and branched-type (agave) fructans. The enzymes employed, namely exo-inulinase and endo-inulinase, give incompletely hydrolysis of levan. Levan hydrolysis requires a third enzyme, endo-levanase. This paper describes a method and commercial test kit (Megazyme Fructan Assay Kit) for the determination of all types of fructan (inulin, levan, and branched) in a variety of animal feeds and pet foods. The method has been validated in a single laboratory for analysis of pure inulin, agave fructan, levan, and a range of fructan containing samples. Quantification is based on complete hydrolysis of fructan to fructose and glucose by a mixture of exo-inulinase, endo-inulinase, and endo-levanase, followed by measurement of these sugars using the PAHBAH reducing sugar method which gives the same color response with fructose and glucose. Before hydrolysis of fructan, interfering sucrose and starch in the sample are specifically hydrolyzed and removed by borohydride reduction. The single-laboratory validation (SLV) outlined in this document was performed on commercially available inulin (Raftiline) and agave fructan (Frutafit®), levan purified from Timothy grass, two grass samples, a sample of legume hay, two animal feeds and two barley flours, one of which (Barley MAX®) was genetically enriched in fructan through plant breeding. Parameters examined during the validation included working range, target selectivity, recovery, LOD, LOQ, trueness (bias), precision (repeatability and intermediate precision), robustness, and stability. The method is robust, quick, and simple.

Comprehensive functional characterization of the glycoside hydrolase family 3 enzymes from Cellvibrio japonicus reveals unique metabolic roles in biomass saccharification.

Lignocellulose degradation is central to the carbon cycle and renewable biotechnologies. The xyloglucan (XyG), ß(1→3)/ß(1→4) mixed-linkage glucan (MLG) and ß(1→3) glucan components of lignocellulose represent significant carbohydrate energy sources for saprophytic microorganisms. The bacterium Cellvibrio japonicus has a robust capacity for plant polysaccharide degradation, due to a genome encoding a large contingent of Carbohydrate-Active enZymes (CAZymes), many of whose specific functions remain unknown. Using a comprehensive genetic and biochemical approach, we have delineated the physiological roles of the four C. japonicus glycoside hydrolase family 3 (GH3) members on diverse ß-glucans. Despite high protein sequence similarity and partially overlapping activity profiles on disaccharides, these ß-glucosidases are not functionally equivalent. Bgl3A has a major role in MLG and sophorose utilization, and supports ß(1→3) glucan utilization, while Bgl3B underpins cellulose utilization and supports MLG utilization. Bgl3C drives ß(1→3) glucan utilization. Finally, Bgl3D is the crucial ß-glucosidase for XyG utilization. This study not only sheds the light on the metabolic machinery of C. japonicus, but also expands the repertoire of characterized CAZymes for future deployment in biotechnological applications. In particular, the precise functional analysis provided here serves as a reference for informed bioinformatics on the genomes of other Cellvibrio and related species.

Complex pectin metabolism by gut bacteria reveals novel catalytic functions.

The metabolism of carbohydrate polymers drives microbial diversity in the human gut microbiota. It is unclear, however, whether bacterial consortia or single organisms are required to depolymerize highly complex glycans. Here we show that the gut bacterium Bacteroides thetaiotaomicron uses the most structurally complex glycan known: the plant pectic polysaccharide rhamnogalacturonan-II, cleaving all but 1 of its 21 distinct glycosidic linkages. The deconstruction of rhamnogalacturonan-II side chains and backbone are coordinated to overcome steric constraints, and the degradation involves previously undiscovered enzyme families and catalytic activities. The degradation system informs revision of the current structural model of rhamnogalacturonan-II and highlights how individual gut bacteria orchestrate manifold enzymes to metabolize the most challenging glycan in the human diet.

Systems analysis in Cellvibrio japonicus resolves predicted redundancy of ß-glucosidases and determines essential physiological functions.

Degradation of polysaccharides forms an essential arc in the carbon cycle, provides a percentage of our daily caloric intake, and is a major driver in the renewable chemical industry. Microorganisms proficient at degrading insoluble polysaccharides possess large numbers of carbohydrate active enzymes (CAZymes), many of which have been categorized as functionally redundant. Here we present data that suggests that CAZymes that have overlapping enzymatic activities can have unique, non-overlapping biological functions in the cell. Our comprehensive study to understand cellodextrin utilization in the soil saprophyte Cellvibrio japonicus found that only one of four predicted ß-glucosidases is required in a physiological context. Gene deletion analysis indicated that only the cel3B gene product is essential for efficient cellodextrin utilization in C. japonicus and is constitutively expressed at high levels. Interestingly, expression of individual ß-glucosidases in Escherichia coli K-12 enabled this non-cellulolytic bacterium to be fully capable of using cellobiose as a sole carbon source. Furthermore, enzyme kinetic studies indicated that the Cel3A enzyme is significantly more active than the Cel3B enzyme on the oligosaccharides but not disaccharides. Our approach for parsing related CAZymes to determine actual physiological roles in the cell can be applied to other polysaccharide-degradation systems.

Corrigendum: Complex pectin metabolism by gut bacteria reveals novel catalytic functions.

This corrects the article DOI: 10.1038/nature21725.

The Mechanism by Which Arabinoxylanases Can Recognize Highly Decorated Xylans.

The enzymatic degradation of plant cell walls is an important biological process of increasing environmental and industrial significance. Xylan, a major component of the plant cell wall, consists of a backbone of ß-1,4-xylose (Xylp) units that are often decorated with arabinofuranose (Araf) side chains. A large penta-modular enzyme, CtXyl5A, was shown previously to specifically target arabinoxylans. The mechanism of substrate recognition displayed by the enzyme, however, remains unclear. Here we report the crystal structure of the arabinoxylanase and the enzyme in complex with ligands. The data showed that four of the protein modules adopt a rigid structure, which stabilizes the catalytic domain. The C-terminal non-catalytic carbohydrate binding module could not be observed in the crystal structure, suggesting positional flexibility. The structure of the enzyme in complex with Xylp-ß-1,4-Xylp-ß-1,4-Xylp-[α-1,3-Araf]-ß-1,4-Xylp showed that the Araf decoration linked O3 to the xylose in the active site is located in the pocket (-2* subsite) that abuts onto the catalytic center. The -2* subsite can also bind to Xylp and Arap, explaining why the enzyme can utilize xylose and arabinose as specificity determinants. Alanine substitution of Glu68, Tyr92, or Asn139, which interact with arabinose and xylose side chains at the -2* subsite, abrogates catalytic activity. Distal to the active site, the xylan backbone makes limited apolar contacts with the enzyme, and the hydroxyls are solvent-exposed. This explains why CtXyl5A is capable of hydrolyzing xylans that are extensively decorated and that are recalcitrant to classic endo-xylanase attack.