Experiences in the molecular genetic and histopathological evaluation of calpainopathies.

Calpainopathy is mainly characterized by symmetric and progressive weakness of proximal muscles. Several reports showed that the most common LGMD subtype is LGMDR1 or calpainopathy, which had previously been defined as LGMD2A. Until now, more than 500 likely pathogenic/pathogenic variants in the CAPN3 gene have been reported. However, a clear genotype-phenotype association had not yet been established and this causes major difficulties in predicting the prognosis in asymptomatic patients and in providing genetic counseling for prenatal diagnosis. In this report, we aimed to add new data to the literature by evaluating 37 patients with likely pathogenic/pathogenic variants for the detected variants' nature, patients' phenotypes, and histopathological features. As a result, the general clinical presentation of the 23 different variants was presented, the high frequency of NM_000070.3:c.550delA mutation in Exon 4 was discussed, and some novel genotype-phenotype associations were suggested. We have underlined that calpainopathy can be misdiagnosed with inflammatory myopathies histopathologically. We have also emphasized that, in young or adult patients with mild to moderate proximal muscle weakness and elevated CK levels, calpainopathy should be the first suspected diagnosis.

Impact of next-generation sequencing panels in the evaluation of limb-girdle muscular dystrophies.

INTRODUCTION: Limb-girdle muscular dystrophy (LGMD) is the fourth most common muscular dystrophy, with progressive proximal muscle weakness. However, a large number of neuromuscular conditions are similarly presented. Because of this, the use of high-throughput methods such as next-generation sequencing (NGS) is important in the evaluation of LGMD. METHODS: In this report, we applied a custom target capture-based NGS panel covering 31 LGMD-associated genes (MYOT, LMNA, CAV3, DES, DNAJB6, FLNC, CAPN3, DYSF, SGCG, SGCA, SGCB, SGCD, TCAP, TRIM32, FRKP, TTN, POMT1, ANO5, FKTN, POMT2, POMGnT1, DAG1, PLEC, GAA, GMPPB, HNRNPDL, TNPO3, LIMS2, POMK, TRAPPC11, ISPD) in 74 patients suspected of LGMD. RESULTS: In 25 (33.8%) out of 74 patients analyzed, one or more pathogenic/likely pathogenic variants in 13 different genes were detected. Six of the patients had the variants that were not found in databases and literature; thus, they were interpreted as novel pathogenic variants. DISCUSSION: The diagnosis rate achieved (33.8%) is consistent with previous literature reports and underlines the efficiency and importance of NGS technology in the molecular genetic evaluation of LGMD.

Molecular genetic evaluation of NLRP3, MVK and TNFRSF1A associated periodic fever syndromes.

Periodic fever syndromes (PFSs) are a family of clinical disorders, which are characterized by recurrent episodes of fever in the absence of microbial, autoimmune or malign conditions. Most common types of PFSs are associated with four genes: MEFV, MVK, TNFRSF1A and NLRP3. This paper aims to add new data to the genotype-phenotype association of MVK-, TNFRSF-1A- and NLRP3-associated PFSs. A total number of 211 patients were evaluated. Two different approaches were used for the molecular genetic evaluation of MVK-, TNFRSF-1A- and NLRP3-associated PFSs. For the first 147 patients, Sanger sequence analysis of selected exons of MVK, TNFRSF1A and NLRP3 genes was done. For subsequent 64 patients, targeted NGS panel analysis, covering all exons of MVK, TNFRSF1A and NLRP3 genes, was used. A total number of 48 variants were detected. The "variant detection rate in index patients" was higher in the NGS group than Sanger sequencing group (19% vs. 15,1%). For the variant positive patients, a detailed genotype-phenotype table was built. In PFSs, lack of correlation exists between genotype and phenotype in the general population and even within the families. In some cases, mutations behave differently and yield unexpected phenotypes. In this study, we discussed the clinical effects of eight different variants we have detected in the MVK, TNFRSF1A and NLRP3 genes. Four of them were previously identified in patients with PFS. The remaining four were not reported in patients with PFS. Thus, we had to interpret their clinical effects by analysing their frequencies and in silico analysis predictions. We suggest that new studies are needed to evaluate the effects of these variants more clearly. To be able to demonstrate a clearer genotype-phenotype relationship, all PFS-related genes should be analysed together and the possibility of polygenic inheritance should be considered.

The Association of Serum Galectin-3 Levels with Atrial Electrical and Structural Remodeling.

INTRODUCTION: Left atrial (LA) interstitial fibrosis is known to have a role in the initiation and maintenance of atrial fibrillation (AF). The role of galectin-3 in the pathogenesis of cardiac fibrosis has been demonstrated in previous studies. We aimed to determine whether serum galectin-3 level is associated with markers of atrial remodeling, including the extent of LA fibrosis detected by delayed enhancement magnetic resonance imaging (DE-MRI) and atrial electromechanical delay (AEMD) in paroxysmal AF patients with preserved left ventricular (LV) functions. METHODS AND RESULTS: Thirty-three patients (58 [28-74] years, 51.5% male) with paroxysmal AF who underwent DE-MRI prior to cryoballoon-based AF ablation were included in the study. Serum galectin-3 levels were measured with ELISA. LA volume index (B ± SE: 0.424 ± 0.504, 95% CI: 0.560-2.627, P = 0.004) and serum galectin-3 levels (B ± SE: 0.549 ± 7.745, 95% CI: 16.874-47.550, P < 0.001) were found to be independently correlated with extent of LA fibrosis detected with DE-MRI in paroxysmal AF patients with preserved LV function. Correlation analysis between AEMD parameters and baseline characteristics showed that galectin-3 was significantly correlated with intra-left (ρ = 0.432, P = 0.012) and inter-AEMD (ρ = 0.395, P = 0.023). Duration of AF, LAD, and extent of LA fibrosis were also found to be significantly correlated with AEMD parameters. CONCLUSION: This is a hypothesis-generating study pointing out that serum galectin-3 level is significantly associated with atrial remodeling in paroxysmal AF patients with preserved LV function. Further studies are necessary to provide exact pathophysiological mechanisms.

Association of dietary and gut microbiota-related metabolites with calcific aortic stenosis.

BACKGROUND: Histopathological changes in calcific aortic stenosis (CAS) resemble changes in coronary atherosclerosis. Concerning recent evidence on dietary and gut microbiota-related metabolites representing players in atherosclerosis, we aimed to investigate the link between dietary and gut microbiota-derived metabolites and CAS. METHODS: We consecutively recruited eligible subjects with moderate-severe CAS (n = 60), aortic sclerosis (ASc) (n = 49) and age and gender-matched control subjects (n = 48) in May 2016-December 2016. Plasma dietary and gut microbiota-related metabolite levels, namely choline, betaine, and trimethylamine N-oxide (TMAO), were measured using ultra-performance liquid chromatography-tandem mass spectroscopy method. Histopathological examinations were performed in patients that underwent aortic valve surgery. RESULTS: Prevalence of traditional cardiovascular risk factors or co-morbidities did not differ among groups (all p > 0.05). CAS patients had higher plasma choline levels compared to both control (p < 0.001) and ASc (p = 0.006). Plasma betaine and TMAO levels were similar (both p > 0.05). Compared to the lowest quartile choline levels (<11.15 µM), patients with the highest quartile choline levels (≥14.98 µM) had higher aortic valvular (p < 0.001) and mitral annular (p = 0.013) calcification scores. Plasma choline levels were independently associated with aortic peak flow velocity (B ± SE:0.165 ± 0.060, p = 0.009). Choline levels were elevated in subjects who had aortic valves with denser lymphocyte infiltration (p < 0.001), neovascularization (p = 0.011), osseous metaplasia (p = 0.004), more severe tissue remodelling (p = 0.002) and calcification (p = 0.002). CONCLUSION: We found a significant association between choline levels and CAS presence and severity depicted on imaging modalities and histopathological examinations. Our study may open new horizons for prevention of CAS.

Cholesterol efflux promoting function of high-density lipoproteins in calcific aortic valve stenosis.

Background and aims: Cholesterol efflux capacity is a functional property of high-density lipoproteins (HDL) reflecting the efficiency of the atheroprotective reverse cholesterol transport process in humans. Its relationship with calcific aortic valve stenosis (CAVS) has not been fully assessed yet. Methods: We evaluated HDL-CEC in a patient population with varying degrees of aortic valvular calcific disease, assessed using echocardiography and cardiac computed tomography. Measurement of biomarkers that reflect osteogenic and tissue remodeling, along with dietary and gut microbiota-derived metabolites were performed. Results: Patients with moderate-severe CAVS had significantly lower HDL-CEC compared to both control and aortic sclerosis subjects (mean: 6.09%, 7.32% and 7.26%, respectively). HDL-CEC displayed negative correlations with peak aortic jet velocity and aortic valve calcium score, indexes of CAVS severity (ρ = -0.298, p = 0.002 and ρ = -0.358, p = 0.005, respectively). In multivariable regression model, HDL-CEC had independent association with aortic valve calcium score (B: -0.053, SE: 0.014, p < 0.001), GFR (B: -0.034, SE: 0.012, p = 0.007), as well as with levels of total cholesterol (B: 0.018, SE: 0.005, p = 0.002). Conclusion: These results indicate an impairment of HDL-CEC in moderate-severe CAVS and may contribute to identify potential novel targets for CAVS management.

The Efficiency of SNP-Based Microarrays in the Detection of Copy-Neutral Events at 15q11.2 and 11p15.5 Loci.

Prader-Willi, Angelman, Beckwith-Wiedemann, and Russell-Silver are imprinting syndromes. In this study, we aimed to compare the efficiency of single nucleotide polymorphism (SNP) microarray analysis with methylation-specific Multiplex ligation-dependent probe amplification (MS-MLPA) in the detection of uniparental disomy in these syndromes. The patient samples with regions of loss of heterozygosity (LOH), covering 15q11.2 and 11p15.5 critical loci, were analyzed with MS-MLPA to demonstrate the efficiency of SNP microarray in the detection of uniparental disomy (UPD). In a total of seven patients, LOH covering 15q11.2 and 11p15.5 critical loci was detected. Two (28.6%) of these seven patients showed aberrant methylation (suggesting UPD) in MS-MLPA. SNP microarray is a useful tool in the detection of LOH; however, it should be used with caution, since false-positive or false-negative LOH results can be obtained. Although methylation analysis is recommended as the first tier test in the diagnosis of most of the imprinting disorders, combining methylation analysis with SNP microarray can enhance our evaluation process.

Are there any clinical and electrocardiographic predictors of heart rate reduction in relapsing- remitting multiple sclerosis patients treated with fingolimod?

BACKGROUND: Fingolimod, a sphingosine-1-phosphate receptor agonist, is used for treatment of relapsing-remitting multiple sclerosis (RRMS). S1P receptors that fingolimod acts upon have also been shown to be expressed on atrial myocytes. This expression pattern has been linked with the drug's cardiovascular effects, such as bradycardia. We aimed to evaluate the clinical and electrocardiographic predictors of heart rate (HR) reduction in patients receiving first-dose fingolimod. METHODS: We retrospectively analyzed subjects diagnosed with RRMS who were allocated to fingolimod treatment. HR, systolic and diastolic blood pressure values and electrocardiography during the first dose of fingolimod were accessed. RESULTS: A total of 114 RRMS patients (65.8% female, 33.58 ±â€¯8.63 years) were included. After the initial dose of fingolimod, the heart rate decreased significantly at each hour (each p < 0.001). Nadir heart rate was reached at 4 h. The multivariate binary logistic regression analysis revealed that BMI (OR: 0.878, p = 0.045), optic nerve involvement (OR: 3.205, p = 0.018), baseline HR (OR: 1.079, p = 0.002) and T-peak-T-end interval (OR: 1.046, p = 0.030) were independent predictors of greater HR reduction. During 6-h monitorization, none of the patients had relevant adverse reactions. CONCLUSION: Our findings provide an insight on clinical and electrocardiographic predictors of HR reduction that occurs in RRMS patients receiving first dose of fingolimod.

cfDNA in exhaled breath condensate (EBC) and contamination by ambient air: toward volatile biopsies.

Exhaled breath is a source of volatile and nonvolatile biomarkers in the body that can be accessed non-invasively and used for monitoring. The collection of lung secretions by conventional methods such as bronchoalveolar lavage, induced sputum collection, and core biopsies is limited by the invasive nature of these methods. Non-invasive collection of exhaled breath condensate (EBC) provides fluid samples that are representative of airway lining fluids. Various volatile and nonvolatile biomarkers can be detected in volatile condensates, such as H2O2, nitric oxide, lipid mediators, cytokines, chemokines, DNA, and microRNAs. Studies have examined cell-free DNA (cfDNA) in plasma samples from non-small-cell lung cancer patients, offering to new insights and fostering development of the liquid biopsy. However, few studies have examined cfDNA in EBC samples. This study examined whether EBC is an appropriate source of cfDNA using housekeeping-gene-specific primer probes and quantitative real-time polymerase chain reaction in healthy subjects. Ambient (room) air is contaminated with DNA, so caution is needed. Preliminary studies indicated that volatile biopsies are becoming an important diagnostic tool in lung cancer.

Targeted fetal cell-free DNA screening for aneuploidies in 4,594 pregnancies: Single center study.

BACKGROUND: Next-generation sequencing (NGS) and discovery of fetal cell-free DNA (cfDNA) in the maternal circulation render possible prenatal screening for trisomy 21 (Down syndrome), trisomy 18, trisomy 13, and sex chromosome aneuploidies. The approach is called "fetal cfDNA screening" and in contrast to noninvasive conventional serum screening, it provides the identification of 98%-99% of fetuses with Down syndrome. METHODS: Retrospective analysis of targeted noninvasive prenatal testing (NIPT) (Clarigo Test) pregnancies with moderate risk, which we have reported between 2016 and 2018 years is presented. Two separate laboratory workflows and NGS platforms are used for the same targeted NIPT analysis. RESULTS: In total, 4,594 pregnant women were investigated. Initial 3,594 cases are studied by MiSeq platform, the last 1,000 cases by NextSeq. Failure rate for MiSeq platform is 10.9% and for NextSeq is 8.7%. Automatically reported cases constitute 75% of the MiSeq group and 87% of the NextSeq group. CONCLUSIONS: Targeted NIPT results suggest that MiSeq platform could be used for NIPT which would be an essential option particularly for laboratories with low sample flow. And, the NextSeq platform has easier wet lab process and also increased success rate in automatic reporting which is suitable for centers with high number of NIPT cases.

Targeted next generation sequencing in patients with maturity-onset diabetes of the young (MODY).

Background Maturity-onset diabetes of the young (MODY) is a common form of monogenic diabetes. Fourteen genes have been identified, each leading to cause a different type of MODY. The aims of this study were to reveal both known and novel variants in MODY genes in patients with MODY using targeted next generation sequencing (NGS) and to present the genotype-phenotype correlations. Methods Mutation analysis of MODY genes (GCK, HNF1A, HNF4A, HNF1B, ABCC8, INS and KCNJ11) was performed using targeted NGS in 106 patients with a clinical diagnosis of MODY. The variants were evaluated according to American College of Medical Genetics and Genomics (ACMG) Standards and Guidelines recommendations. Results A total of 18 (17%) variants were revealed among all patients. Seven variants in GCK, six in HNF4A, four in HNF1A and one in ABCC8 genes were found. Eight of them were previously published and 10 of them were assessed as novel pathogenic or likely pathogenic variants. Conclusions While the most frequent mutations are found in the HNF1A gene in the literature, most of the variants were found in the GCK gene in our patient group using the NGS method, which allows simultaneous analysis of multiple genes in a single panel.

Pharmacokinetics of imipenem in sheep with special reference to its hepato-renal effects.

BACKGROUND: Imipenem is a carbapenem antibiotic with a broad spectrum of activity. The aim of this study was to determine the pharmacokinetics of imipenem after single intravenous and intramuscular injections and the effect of repeated intramuscular injections on hepato-renal functions in sheep. METHODS: Imipenem concentrations in plasma and urine were determined by a microbiological agar plate assay. RESULTS: Following intramuscular injection, imipenem was rapidly absorbed and the peak plasma concentration was 9.99 microg ml(-1) and the systemic bioavailability was 65.97%. Urine concentrations of imipenem were much higher than in plasma. Light and electron microscopy evidenced little changes in the kidney and liver. CONCLUSION: Imipenem is likely to be efficacious in most urinary tract infections and has no adverse effects on hepato-renal structures and functions.

Elevated Serum Beta-Trace Protein Levels are Associated With the Presence of Atrial Fibrillation in Hypertension Patients.

Beta-trace protein (BTP) has emerged as a novel biomarker of cardiovascular risk. In this study, the authors aimed to assess the relationship between BTP levels and presence of atrial fibrillation in patients who had controlled hypertension (HTN) and normal renal function. A total of 80 controlled HTN patients with paroxysmal atrial fibrillation (PAF) and 80 age- and sex-matched controls with controlled HTN were enrolled. Serum BTP levels were measured by enzyme-linked immunosorbent assay. BTP levels were found to be significantly higher in patients with PAF (P<.001). Other parameters including mean systolic and diastolic blood pressure values, serum creatinine levels, and glomerular filtration rate were similar between the two groups. Along with left atrial diameter (odds ratio, 1.504; P<.001), BTP levels (odds ratio, 1.015; P<.001) were independently associated with the presence of PAF. BTP levels were increased in controlled HTN patients with PAF compared with controls, and this association was observed within normal renal functions as reflected by normal glomerular filtration rate.

Elevated M2-muscarinic and ß1-adrenergic receptor autoantibody levels are associated with paroxysmal atrial fibrillation.

BACKGROUND: Pathophysiologic mechanisms underlying lone atrial fibrillation (AF) have not been clearly demonstrated yet. Emerging evidence has indicated that autoimmunity may play a role in the development of AF. Relationship between serum anti-M2-muscarinic receptor autoantibody (anti-M2-R) and anti-ß1-adrenergic receptor autoantibody (anti-ß1-R) levels and lone paroxysmal atrial fibrillation (PAF) has not been investigated. We aimed to compare anti-M2-R and anti-ß1-R levels between lone PAF patients and healthy control subjects. METHODS AND RESULTS: 75 patients with lone PAF (age: 52.80 ± 6.80 years, 53 % male) and 75 healthy control subjects (age: 53.30 ± 6.80 years, 54 % male) were enrolled in the study. Serum anti-M2-R and anti-ß1-R levels were measured by ELISA and compared between two groups. Anti-M2-R [142.30 (77.65-400.00) vs. 69.00 (39.48-299.04) ng/mL; p < 0.001) and anti-ß1-R [102.56 (65.18-348.41) vs. 44.17 (30.89-158.54) ng/mL; p < 0.001] levels were significantly higher in patients with lone PAF compared to healthy controls. Multivariate regression analysis showed that left atrial diameter (OR: 1.471, p < 0.001), hs-CRP(OR: 1.940, p < 0.001), anti-M2-R (OR: 1.158, p < 0.001) and anti-ß1-R (OR: 1.296, p < 0.001) levels were independent predictors for the presence of lone PAF. Using a cut-off level of 101.83 ng/mL, anti-M2-R levels predicted presence of lone PAF with a sensitivity of 94.68 % and specificity of 81.33 %. Anti-ß1-R levels predicted presence of lone PAF with a sensitivity of 92.00 % and specificity of 73.30 %, using a cut-off level of 72.16 ng/mL. CONCLUSION: Our results demonstrated that higher serum anti-M2-R and anti-ß1-R levels are associated with lone PAF. Autoantibodies related to autonomic system may play an important role in the development of lone AF.

L-trans-2,3-pyrrolidine dicarboxylate: characterization of a novel excitotoxin.

This study investigated the in vitro and in vivo excitotoxic properties of a novel conformationally constrained analogue of L-glutamate, L-trans-2,3-pyrrolidine dicarboxylate (L-trans-2,3-PDC). When tested for excitotoxic activity in rat cortical cultures, L-trans-2,3-PDC mimicked the action of NMDA in both acute (30 min) and chronic (24 h) exposure paradigms. This neurotoxicity was attenuated by co-addition of MK-801 (10 microM). Microinjections of L-trans-2,3-PDC into the dorsal hippocampus of male rats also induced a selective pattern of pathology indicative of an NMDA receptor excitotoxin. In contrast to the equipotency observed in vitro, 100 nmol of L-trans-2,3-PDC were needed to produce cellular damage comparable to that induced by 25 nmol of NMDA. Consistent with an action at NMDA receptors, L-trans-2,3-PDC-induced damage could be significantly reduced by co-administration of MK-801 (3 mg/kg i.p.), but not by NBQX (25 nmol). In radioligand binding assays L-trans-2,3-PDC inhibited the binding of 3H-L-glutamate to NMDA receptors (IC50 1 microM), although it also exhibited some cross reactivity with KA and AMPA receptors. L-trans-2,3-PDC was also identified as a competitive inhibitor (Ki = 33 microM) of 3H-D-aspartate uptake into rat forebrain synaptosomes. In contrast to the action of a transported substrate, such as L-glutamate, L-trans-2,3-PDC did not exchange with 3H-D-aspartate that had been previously loaded into the synaptosomes.

Commissurotomy and bileaflet pericardial augmentation-resuspension for bicuspid aortic valve stenosis.

We describe a valve reconstruction technique for congenital bicuspid aortic valve stenosis employing a commissurotomy, resection of raphe between conjoint leaflets, and bileaflet augmentation-resuspension using a triangular strip of glutaraldehyde-preserved autologous pericardium. This maneuver relieves aortic valve stenosis, preserves the native valve leaflets, reproduces the natural trileaflet scalloping of the aortic valve annulus, and improves cusp coaptation.

Repair of congenital pulmonary incompetence by bicuspidization of the pulmonary valve.

We report the successful repair of pulmonary incompetence in an adult due to the congenital absence of the posterior leaflet of the pulmonary valve. The repair consisted of bicuspidization of the pulmonary valve, which achieved competence and eliminated the symptoms and echocardiographic manifestations of right ventricular overload while preserving the native valve.

The efficiency of various solvents in the extraction of chemical mutagens from living tissues: a comparative study.

The aim of this study is to find out the best solvent to be used in extraction of chemical pollutants from living organisms. Ethanol, benzene, chloroform, pentane, 1,4-dioxan, acetone, cyclohexane and ether were used in extraction of shrimp and clam samples collected from a Fahaheel site. The tissue extracts were tested for their mutagenic activity using E. coli and Salmonella on tester strains. The results revealed that ethanol was more efficient than other solvent in extraction of chemical mutagens concentrated in clams and shrimps.

Survival and prognosis following resection of primary non small cell bronchogenic carcinoma.

A prospective follow up was carried out on 479 consecutive patients who underwent lung resection for non small cell primary bronchogenic carcinoma between 1980 and 1987 under the care of one surgeon at Guy's Hospital and Brook Hospital, London. The mean age of patients was 61.8 years; 16.9% were aged 70 years or over. Of the 479,237 patients had stage I disease, 108 patients stage II disease, and 134 patients stage III. Lobectomy was performed in 280 patients, pneumonectomy in 191, and wedge resection in 8. Operative mortality was 5% overall, 6.8% following pneumonectomy and 3.9% following lobectomy. There was no operative mortality following wedge resection. Old age did not affect operative mortality. Overall actuarial survival was 76.2% and 39.8% at 1 year and 5 years postoperatively, respectively (stage I: 86% and 55%; stage II: 77.8% and 35.5%; stage III: 57.5% and 16.2%). There were statistically significant differences in survival between the stages. Five-year actuarial survival was 45% for squamous cell carcinoma, 36.3% for adenocarcinoma, 31.9% for dimorphic carcinoma and a 21% for undifferentiated carcinoma. There were statistically significant differences in survival between undifferentiated carcinoma and each of the other cell types. The favourable survival in stage I disease lends weight to the concept that there is hope for cure in patients with early non small cell lung cancer.

Biodegradation of anthracene and fluoranthene by fungi isolated from an experimental constructed wetland for wastewater treatment.

Pilot-scale constructed wetlands were used to treat water contaminated by polycyclic aromatic hydrocarbons (PAHs), particularly fluoranthene, and the possible role of fungi present in these ecosystems was investigated. A total of 40 fungal species (24 genera) were isolated and identified from samples (gravel and sediments) from a contaminated wetland and a control wetland. All of them were assayed for their ability to remove anthracene (AC) and fluoranthene (FA) from liquid medium. FA was degraded efficiently by 33 species while only 2 species were able to remove AC over 70%. A selection of 10 strains of micromycetes belonging to various taxonomic groups was further investigated for FA and AC degradation, toxicity assays and phenoloxidases (POx) detection. Interesting and not previously reported species were revealed (Absidia cylindrospora, Cladosporium sphaerospermum, and Ulocladium chartarum). They were all able to highly degrade the PAH-model compounds chosen. An interesting inducibility was noted for Ulocladium chartarum. Degradative ability of fungi was not related to their extracellular POx activity. This study may contribute to the improvement of constructed wetlands for water treatment, which may be enriched in efficient fungi.