Effects of maternal type 1 diabetes and confounding factors on neonatal microbiomes.

AIMS/HYPOTHESIS: Body niche-specific microbiota in maternal-neonatal dyads from gravidae with type 1 diabetes have not been quantitatively and functionally examined. Similarly, the impact of pregnancy-specific factors, such as the presence of comorbidities known to occur more frequently among gravidae with type 1 diabetes, including Caesarean delivery, as well as antibiotic prophylaxis, level of glycaemic control during each trimester of pregnancy and insulin administration, has not been adequately considered. The aims of this study were to characterise the maternal and neonatal microbiomes, assess aspects of microbiota transfer from the maternal microbiomes to the neonatal microbiome and explore the impact of type 1 diabetes and confounding factors on the microbiomes. METHODS: In this observational case-control study, we characterised microbiome community composition and function using 16S rRNA amplicon sequencing in a total of 514 vaginal, rectal and ear-skin swabs and stool samples derived from 92 maternal-neonatal dyads (including 50 gravidae with type 1 diabetes) and in-depth clinical metadata from throughout pregnancy and delivery. RESULTS: Type 1 diabetes-specific microbiota were identified among gravidae with type 1 diabetes and their neonates. Neonatal microbiome profiles of ear-skin swabs and stool samples were established, indicating the taxa more prevalent among neonates born to mothers with type 1 diabetes compared with neonates born to control mothers. Without taking into account the type 1 diabetes status of mothers, both delivery mode and intrapartum antibiotic prophylaxis were found to have an influence on neonatal microbiota composition (both p=0.001). In the logistic regression analysis involving all confounding variables, neonatal ear-skin microbiome variation was explained by maternal type 1 diabetes status (p=0.020) and small for gestational age birthweight (p=0.050). Moreover, in women with type 1 diabetes, a relationship was found between HbA1c levels >55 mmol/mol (>7.2%) measured in the first trimester of pregnancy and neonatal ear-skin microbiota composition (p=0.008). In the PICRUSt (Phylogenetic Investigation of Communities by Reconstruction of Unobserved States) assessment, pathways concerning carbohydrate biosynthesis were predicted as key elements of the microbial functional profiles dysregulated in type 1 diabetes. Additionally, in SourceTracker analysis, we found that, on average, 81.0% of neonatal microbiota was attributed to maternal sources. An increase in the contribution of maternal rectum microbiota and decrease in the contribution of maternal cervix microbiota were found in ear-skin samples of vaginally delivered neonates of mothers with type 1 diabetes compared with neonates born to control mothers (83.2% vs 59.5% and 0.7% vs 5.2%, respectively). CONCLUSIONS/INTERPRETATION: These findings indicate that, in addition to maternal type 1 diabetes, glycaemic dysregulation before/in the first trimester of pregnancy, mode of delivery and intrapartum antibiotic prophylaxis may contribute to the inoculation and formation of the neonatal microbiomes. DATA AVAILABILITY: The BioProject (PRJNA961636) and associated SRA metadata are available at http://www.ncbi.nlm.nih.gov/bioproject/961636 . Processed data on probiotic supplementation and the PICRUSt analysis are available in the Mendeley Data Repository ( https://doi.org/10.17632/g68rwnnrfk.1 ).

Gender Influences Gut Microbiota among Patients with Irritable Bowel Syndrome.

Irritable bowel syndrome (IBS) is a chronic functional gastrointestinal disease that affects approximately 11% of the general population. The gut microbiota, among other known factors, plays a substantial role in its pathogenesis. The study aimed to characterize the gut microbiota differences between patients with IBS and unaffected individuals, taking into account the gender aspect of the patients and the types of IBS determined on the basis of the Rome IV Criteria, the IBS-C, IBS-D, IBS-M, and IBS-U. In total, 121 patients with IBS and 70 unaffected individuals participated in the study; the derived stool samples were subjected to 16S rRNA amplicon sequencing. The gut microbiota of patients with IBS was found to be more diverse in comparison to unaffected individuals, and the differences were observed primarily among Clostridiales, Mogibacteriaceae, Synergistaceae, Coriobacteriaceae, Blautia spp., and Shuttleworthia spp., depending on the study subgroup and patient gender. There was higher differentiation of females' gut microbiota compared to males, regardless of the disease status. No correlation between the composition of the gut microbiota and the type of IBS was found. Patients with IBS were characterized by more diverse gut microbiota compared to unaffected individuals. The gender criterion should be considered in the characterization of the gut microbiota. The type of IBS did not determine the identified differences in gut microbiota.

Influence of TGFBR2, TGFB3, DNMT1, and DNMT3A Knockdowns on CTGF, TGFBR2, and DNMT3A in Neonatal and Adult Human Dermal Fibroblasts Cell Lines.

Dermal fibroblasts are responsible for the production of the extracellular matrix that undergoes significant changes during the skin aging process. These changes are partially controlled by the TGF-ß signaling, which regulates tissue homeostasis dependently on several genes, including CTGF and DNA methyltransferases. To investigate the potential differences in the regulation of the TGF-ß signaling and related molecular pathways at distinct developmental stages, we silenced the expression of TGFB1, TGFB3, TGFBR2, CTGF, DNMT1, and DNMT3A in the neonatal (HDF-N) and adult (HDF-A) human dermal fibroblasts using the RNAi method. Through Western blot, we analyzed the effects of the knockdowns of these genes on the level of the CTGF, TGFBR2, and DNMT3A proteins in both cell lines. In the in vitro assays, we observed that CTGF level was decreased after knockdown of DNMT1 in HDF-N but not in HDF-A. Similarly, the level of DNMT3A was decreased only in HDF-N after silencing of TGFBR2, TGFB3, or DNMT1. TGFBR2 level was lower in HDF-N after knockdown of TGFB3, DNMT1, or DNMT3A, but it was higher in HDF-A after TGFB1 silencing. The reduction of TGFBR2 after silencing of DNMT3A and vice versa in neonatal cells only suggests the developmental stage-specific interactions between these two genes. However, additional studies are needed to explain the dependencies between analyzed proteins.

Are dietary supplements containing plant-derived ingredients safe microbiologically?

Dietary supplements, particularly those containing ingredients of natural origin, may contain microbiological contaminants, both bacterial and fungal. The present study evaluated the microbiological purity of selected dietary supplements containing plant-based ingredients before their release to the market, as well as raw materials of plant origin which are used in the production of such supplements. A total of 122 samples of supplements and 30 materials of plant origin were studied, with 92.1% exhibiting different degrees of bacterial contamination. Eight samples (5.3%) were contaminated by aerobic bacteria in amounts exceeding 105 CFU/g. Five (3.3%) of the studied supplements were found to contain bacteria from the family Enterobacteriaceae at a level exceeding 103 CFU/g. Furthermore, a considerable proportion of the studied samples (86.8%) contained fungal contamination. Microbiological contamination may contribute to a deterioration in quality and stability of dietary supplements. In addition, high levels of pathogenic bacteria and microorganisms may pose a risk to consumers.

Genomic strategies to understand causes of keratoconus.

Keratoconus (KTCN) is a degenerative disorder of the eye characterized by the conical shape and thinning of the cornea. The abnormal structure of KTCN-affected cornea results in loss of visual acuity. While many studies examine how environmental factors influence disease development, finding the genetic triggers has been a major emphasis of KTCN research. This paper focuses on genomic strategies that were implemented for finding candidate genes, including linkage and association studies, and presents different approaches of mutation screening. The advantages and limitations of particular tools are discussed based on literature and personal experience. Since etiology underlying KTCN is complex, numerous findings indicating heterogeneity of genetic factors involved KTCN etiology are presented.

CRISPR/Cas9-mediated deletion of lncRNA Gm26878 in the distant Foxf1 enhancer region.

Recent genome editing techniques, including CRISPR mutagenesis screens, offer unparalleled opportunities to study the regulatory non-coding genomic regions, enhancers, promoters, and functional non-coding RNAs. Heterozygous point mutations in FOXF1 and genomic deletion copy-number variants at chromosomal region 16q24.1 involving FOXF1 or its regulatory region mapping ~300 kb upstream of FOXF1 and leaving it intact have been identified in the vast majority of patients with a lethal neonatal lung disease, alveolar capillary dysplasia with misalignment of pulmonary veins (ACDMPV). Homozygous Foxf1 -/- mice have been shown to die by embryonic day 8.5 because of defects in the development of extraembryonic and lateral mesoderm-derived tissues, whereas heterozygous Foxf1 +/- mice exhibit features resembling ACDMPV. We have previously defined a human lung-specific enhancer region encoding two long non-coding RNAs, LINC01081 and LINC01082, expressed in the lungs. To investigate the biological significance of lncRNAs in the Foxf1 enhancer region, we have generated a CRISPR/Cas9-mediated ~2.4 kb deletion involving the entire lncRNA-encoding gene Gm26878, located in the mouse region syntenic with the human Foxf1 upstream enhancer. Very recently, this mouse genomic region has been shown to function as a Foxf1 enhancer. Our results indicate that homozygous loss of Gm26878 is neonatal lethal with low penetrance. No changes in Foxf1 expression were observed, suggesting that the regulation of Foxf1 expression differs between mouse and human.

Halloysite nanotubes as carriers of vancomycin in alginate-based wound dressing.

The influence of an inorganic support - halloysite nanotubes - on the release rate and biological activity of the antibiotic encapsulated in alginate-based dressings was studied. The halloysite samples were loaded with approx. 10 wt.% of the antibiotic and then encapsulated in Alginate and Gelatin/Alginate gels. The material functionalized with aliphatic amine significantly extended the release of vancomycin from alginate-based gels as compared to that achieved when silica was used. After 24 h, the released amounts of the antibiotic immobilized at silica reached 70%, while for the drug immobilized at halloysite the released amount of vancomycin reached 44% for Alginate discs. The addition of gelatin resulted in even more prolonged sustained release of the drug. The antibiotic was released from the system with a double barrier with Higuchi kinetic model and Fickian diffusion mechanism. Only the immobilized drug encapsulated in Alginate gel demonstrated very good antimicrobial activity against various bacteria. The inhibition zones were greater than those of the standard discs for the staphylococci and enterococci bacteria tested. The addition of gelatin adversely affected the biological activity of the system. The inhibition zones were smaller than those of the reference samples. A reduction in the drug dose by half had no significant effect on changing the release rate and microbiological activity. The in vivo toxicity studies of the material with immobilized drug were carried out with Acutodesmus acuminatus and Daphnia magna. The material studied had no effect on the living organisms used in the bioassays. The proposed system with a double barrier demonstrated high storage stability.

Unrevealed mosaicism in the next-generation sequencing era.

Mosaicism refers to the presence in an individual of normal and abnormal cells that are genotypically distinct and are derived from a single zygote. The incidence of mosaicism events in the human body is underestimated as the genotypes in the mosaic ratio, especially in the low-grade mosaicism, stay unrevealed. This review summarizes various research outcomes and diagnostic questions in relation to different types of mosaicism. The impact of both tested biological material and applied method on the mosaicism detection rate is especially highlighted. As next-generation sequencing technologies constitute a promising methodological solution in mosaicism detection in the coming years, revisions in current diagnostic protocols are necessary to increase the detection rate of the unrevealed mosaicism events. Since mosaicism identification is a complex process, numerous examples of multistep mosaicism investigations are presented and discussed.

Synthesis, structure and antimicrobial evaluation of a new gossypol triazole conjugates functionalized with aliphatic chains and benzyloxy groups.

Synthetic limitations in the copper-catalyzed azide alkyne cycloaddition (CuAAC) on gossypol's skeleton functionalized with alkyne (2) or azide (3) groups have been indicated. Modified approach to the synthesis of new gossypol-triazole conjugates yielded new compounds (24-31) being potential fungicides. Spectroscopic studies of triazole conjugates 24-31 have revealed their structures in solution, i.e., the presence of enamine-enamine tautomeric forms and π-π stacking intramolecular interactions between triazole arms. Biological evaluation of the new gossypol-triazole conjugates revealed the potency of 30 and 31 derivatives, having triazole-benzyloxy moieties, comparable with that of miconazole against Fusarium oxysporum. The results of HPLC evaluation of ergosterol content in different fungi strains upon treatment of gossypol and its derivatives enabled to propose a mechanism of antifungal activity of these compounds.

The effect of complexation of 3-formylrifamycin SV macrocyclic ether derivatives with metal cations and small nitrogen-containing organic molecules on antibacterial activity against S. aureus and S. epidermidis.

Spectroscopic studies of ether rifamycins (1-9) have shown that all these compounds tend to be zwitterions with different localizations of intramolecularly transferred proton, which influences their solubility and logP values. According to ESI MS studies, rifamycins 3 and 4 form complexes with Li(+) or Na(+), while the other ones (7-9) coordinate small organic molecules, which can be further replaced by Na(+) cation. Biological assays revealed that the use of 7-9 in the form of complexes with small organic molecules improves their antibacterial potency as a result of changed: logP, solubility and binding mode with bacterial RNA polymerases.

A Combined Metabolomic and Proteomic Analysis of Gestational Diabetes Mellitus.

The aim of this pilot study was to apply a novel combined metabolomic and proteomic approach in analysis of gestational diabetes mellitus. The investigation was performed with plasma samples derived from pregnant women with diagnosed gestational diabetes mellitus (n = 18) and a matched control group (n = 13). The mass spectrometry-based analyses allowed to determine 42 free amino acids and low molecular-weight peptide profiles. Different expressions of several peptides and altered amino acid profiles were observed in the analyzed groups. The combination of proteomic and metabolomic data allowed obtaining the model with a high discriminatory power, where amino acids ethanolamine, L-citrulline, L-asparagine, and peptide ions with m/z 1488.59; 4111.89 and 2913.15 had the highest contribution to the model. The sensitivity (94.44%) and specificity (84.62%), as well as the total group membership classification value (90.32%) calculated from the post hoc classification matrix of a joint model were the highest when compared with a single analysis of either amino acid levels or peptide ion intensities. The obtained results indicated a high potential of integration of proteomic and metabolomics analysis regardless the sample size. This promising approach together with clinical evaluation of the subjects can also be used in the study of other diseases.

Recurrence risks for different pregnancy outcomes and meiotic segregation analysis of spermatozoa in carriers of t(1;11)(p36.22;q12.2).

Cumulative data obtained from two relatively large pedigrees of a unique reciprocal chromosomal translocation (RCT) t(1;11)(p36.22;q12.2) ascertained by three miscarriages (pedigree 1) and the birth of newborn with hydrocephalus and myelomeningocele (pedigree 2) were used to estimate recurrence risks for different pregnancy outcomes. Submicroscopic molecular characterization by fluorescent in situ hybridization (FISH) of RCT break points in representative carriers showed similar rearrangements in both families. Meiotic segregation patterns after sperm analysis by three-color FISH of one male carrier showed all possible outcomes resulting from 2:2 and 3:1 segregations. On the basis of empirical survival data, we suggest that only one form of chromosome imbalance resulting in monosomy 1p36.22→pter with trisomy 11q12.2→qter may be observed in progeny at birth. Segregation analysis of these pedigrees was performed by the indirect method of Stengel-Rutkowski and showed that probability rate for malformed child at birth due to an unbalanced karyotype was 3/48 (6.2±3.5%) after ascertainment correction. The risk for stillbirths/early neonatal deaths was -/48 (<1.1%) and for miscarriages was 17/48 (35.4±6.9%). However, the probability rate for children with a normal phenotype at birth was 28/48 (58.3±7.1%). The results obtained from this study may be used to determine the risks for the various pregnancy outcomes for carriers of t(1;11)(p36.22;q12.2) and can be used for genetic counseling of carriers of this rearrangement.

Differentially expressed microRNAs targeting genes in key pathways in keratoconus.

Introduction: Keratoconus (KTCN) is a corneal ectasia, characterized by a progressive thinning and protrusion of the cornea, with a complex etiology involving genetic, behavioral, lifestyle, and environmental factors. Previous studies indicated that microRNAs (miRNAs) could be involved in KTCN pathogenesis. This in silico study aimed to identify precursor microRNAs (pre-miRNAs) differentially expressed in KTCN corneas and to characterize mature miRNAs and their target genes. Materials and methods: Expression levels of pre-miRNAs were retrieved from our previously obtained RNA sequencing data of 25 KTCN and 25 non-KTCN human corneas (PMID:28145428, PMID:30994860). Differential expression with FDR ≤0.01 and ≥1.5-fold changes were considered significant. Lists of target genes (target score ≥90) of mature miRNAs were obtained from miRDB. Revealed up-/downregulated miRNAs and their target genes were assessed in databases and literature. Enrichment analyses were completed applying the DAVID database. Results: From a total of 47 pre-miRNAs, six were remarkably upregulated (MIR184, MIR548I1, MIR200A, MIR6728, MIR429, MIR1299) and four downregulated (MIR6081, MIR27B, MIR23B, MIR23A) in KTCN corneas. Out of the 1,409 target genes, 220 genes with decreased and 57 genes with increased expression levels in KTCN samples vs non-KTCN samples were found. The extracellular matrix (ECM) organization, response to mechanical stimulus, regulation of cell shape, and signal transduction processes/pathways were identified as distinctive in enrichment analyses. Also, processes associated with the regulation of transcription and DNA binding were listed. Conclusion: Indicated miRNAs and their target genes might be involved in KTCN pathogenesis via disruption of crucial molecular processes, including ECM organization and signal transduction.

Macrolide and lincosamide resistance of Streptococcus agalactiae in pregnant women in Poland.

Knowing about the antibiotic resistance, serotypes, and virulence-associated genes of Group B Streptococcus for epidemiological and vaccine development is very important. We have determined antimicrobial susceptibility patterns, serotype, and virulence profiles. The antibiotic susceptibility was assessed for a total of 421 Streptococcus agalactiae strains, isolated from pregnant women and neonates. Then, 89 erythromycin and/or clindamycin-resistant strains (82 isolates obtained from pregnant women and seven isolates derived from neonates) were assessed in detail. PCR techniques were used to identify the studied strains, perform serotyping, and assess genes encoding selected virulence factors. Phenotypic and genotypic methods determined the mechanisms of resistance. All tested strains were sensitive to penicillin and levofloxacin. The constitutive MLSB mechanism (78.2%), inducible MLSB mechanism (14.9%), and M phenotype (6.9%) were identified in the macrolide-resistant strains. It was found that macrolide resistance is strongly associated with the presence of the ermB gene and serotype V. FbsA, fbsB, fbsC, scpB, and lmb formed the most recurring pattern of genes among the nine surface proteins whose genes were analysed. A minority (7.9%) of the GBS isolates exhibited resistance to lincosamides and macrolides, or either, including those that comprised the hypervirulent clone ST-17. The representative antibiotic resistance pattern consisted of erythromycin, clindamycin, and tetracycline resistance (71.9%). An increase in the fraction of strains resistant to macrolides and lincosamides indicates the need for monitoring both the susceptibility of these strains and the presence of the ST-17 clone.

Sequence variants contributing to dysregulated inflammatory responses across keratoconic cone surface in adolescent patients with keratoconus.

Background: Keratoconus (KTCN) is the most common corneal ectasia resulting in a conical shape of the cornea. Here, genomic variation in the corneal epithelium (CE) across the keratoconic cone surface in patients with KTCN and its relevance in the functioning of the immune system were assessed. Methods: Samples from four unrelated adolescent patients with KTCN and two control individuals were obtained during the CXL and PRK procedures, respectively. Three topographic regions, central, middle, and peripheral, were separated towards the whole-genome sequencing (WGS) study embracing a total of 18 experimental samples. The coding and non-coding sequence variation, including structural variation, was assessed and then evaluated together with the previously reported transcriptomic outcomes for the same CE samples and full-thickness corneas. Results: First, pathway enrichment analysis of genes with identified coding variants pointed to "Antigen presentation" and "Interferon alpha/beta signaling" as the most overrepresented pathways, indicating the involvement of inflammatory responses in KTCN. Both coding and non-coding sequence variants were found in genes (or in their close proximity) linked to the previously revealed KTCN-specific cellular components, namely, "Actin cytoskeleton", "Extracellular matrix", "Collagen-containing extracellular matrix", "Focal adhesion", "Hippo signaling pathway", and "Wnt signaling" pathways. No genomic heterogeneity across the corneal surface was found comparing the assessed topographic regions. Thirty-five chromosomal regions enriched in both coding and non-coding KTCN-specific sequence variants were revealed, with a most representative 5q locus previously recognized as involved in KTCN. Conclusion: The identified genomic features indicate the involvement of innate and adaptive immune system responses in KTCN pathogenesis.

Non-allergic eye rubbing is a major behavioral risk factor for keratoconus.

Since the environmental, behavioral, and socioeconomic factors in the etiology of keratoconus (KTCN) remain poorly understood, we characterized them as features influencing KTCN phenotype, and especially affecting the corneal epithelium (CE). In this case-control study, 118 KTCN patients and 73 controls were clinically examined and the Questionnaire covering the aforementioned aspects was completed and then statistically elaborated. Selected KTCN-specific findings were correlated with the outcomes of the RNA-seq assessment of the CE samples. Male sex, eye rubbing, time of using a computer after work, and dust in the working environment, were the substantial KTCN risk factors identified in multivariate analysis, with ORs of 8.66, 7.36, 2.35, and 5.25, respectively. Analyses for genes whose expression in the CE was correlated with the eye rubbing manner showed the enrichment in apoptosis (TP53, BCL2L1), chaperon-related (TLN1, CTDSP2, SRPRA), unfolded protein response (NFYA, TLN1, CTDSP2, SRPRA), cell adhesion (TGFBI, PTPN1, PDPK1), and cellular stress (TFDP1, SRPRA, CAPZB) pathways. Genes whose expression was extrapolated to the allergy status didn't contribute to IgE-related or other inflammatory pathways. Presented findings support the hypothesis of chronic mechanical corneal trauma in KTCN. Eye-rubbing causes CE damage and triggers cellular stress which through its influence on cell apoptosis, migration, and adhesion affects the KTCN phenotype.

The Impaired Wound Healing Process Is a Major Factor in Remodeling of the Corneal Epithelium in Adult and Adolescent Patients With Keratoconus.

Purpose: Keratoconus (KTCN) is the most common corneal ectasia, characterized by pathological cone formation. Here, to provide an insight into the remodeling of the corneal epithelium (CE) during the course of the disease, we evaluated topographic regions of the CE of adult and adolescent patients with KTCN. Methods: The CE samples from 17 adult and 6 adolescent patients with KTCN, and 5 control CE samples were obtained during the CXL and PRK procedures, respectively. Three topographic regions, central, middle, and peripheral, were separated toward RNA sequencing and MALDI-TOF/TOF Tandem Mass Spectrometry. Data from transcriptomic and proteomic investigations were consolidated with the morphological and clinical findings. Results: The critical elements of the wound healing process, epithelial-mesenchymal transition, cell-cell communications, and cell-extracellular matrix interactions were altered in the particular corneal topographic regions. Abnormalities in pathways of neutrophils degranulation, extracellular matrix processing, apical junctions, IL, and IFN signaling were revealed to cooperatively disorganize the epithelial healing. Deregulation of the epithelial healing, G2M checkpoints, apoptosis, and DNA repair pathways in the middle CE topographic region in KTCN explains the presence of morphological changes in the corresponding doughnut pattern (a thin cone center surrounded by a thickened annulus). Despite similar morphological characteristics of CE samples in adolescents and adults with KTCN, their transcriptomic features were different. Values of the posterior corneal elevation differentiated adults with KTCN from adolescents with KTCN and correlated with the expression of TCHP, SPATA13, CNOT3, WNK1, TGFB2, and KRT12 genes. Conclusions: Identified molecular, morphological, and clinical features indicate the effect of impaired wound healing on corneal remodeling in KTCN CE.

Differences in the composition of the bacterial element of the urinary tract microbiome in patients undergoing dialysis and patients after kidney transplantation.

Introduction: The development of molecular biology methods and their application in microbial research allowed the detection of many new pathogens that cause urinary tract infections (UTIs). Despite the advances of using new research techniques, the etiopathogenesis of UTIs, especially in patients undergoing dialysis and patients after kidney transplantation, is still not fully understood. Methods: This study aimed to characterize and compare the composition of the bacterial element of the urinary tract microbiome between the groups of patients undergoing dialysis (n = 50) and patients after kidney transplantation (n = 50), with positive or negative urine culture, compared to healthy individuals (n = 50). Results: Asymptomatic bacteriuria was observed in 30% of the urine cultures of patients undergoing dialysis and patients after kidney transplantation, with Escherichia coli as the most dominant microorganism (73%) detected with the use of classical microbiology techniques. However, differences in the bacterial composition of the urine samples between the evaluated patient groups were demonstrated using the amplicon sequencing. Finegoldia, Leptotrichia, and Corynebacterium were found to be discriminative bacteria genera in patients after dialysis and kidney transplantation compared to the control group. In addition, in all of urine samples, including those without bacteriuria in classical urine culture, many types of bacteria have been identified using 16S rRNA sequencing. Discussion: The revealed microbial characteristics may form the basis in searching for new diagnostic markers in treatment of patients undergoing dialysis and patients after kidney transplantation.

Prenatal Detection of a FOXF1 Deletion in a Fetus with ACDMPV and Hydronephrosis.

Alveolar capillary dysplasia with misalignment of pulmonary veins (ACDMPV) is a lethal lung developmental disorder caused by the arrest of fetal lung formation, resulting in neonatal death due to acute respiratory failure and pulmonary arterial hypertension. Heterozygous single-nucleotide variants or copy-number variant (CNV) deletions involving the FOXF1 gene and/or its lung-specific enhancer are found in the vast majority of ACDMPV patients. ACDMPV is often accompanied by extrapulmonary malformations, including the gastrointestinal, cardiac, or genitourinary systems. Thus far, most of the described ACDMPV patients have been diagnosed post mortem, based on histologic evaluation of the lung tissue and/or genetic testing. Here, we report a case of a prenatally detected de novo CNV deletion (~0.74 Mb) involving the FOXF1 gene in a fetus with ACDMPV and hydronephrosis. Since ACDMPV is challenging to detect by ultrasound examination, the more widespread implementation of prenatal genetic testing can facilitate early diagnosis, improve appropriate genetic counselling, and further management.

Differential methylation of microRNA encoding genes may contribute to high myopia.

Introduction: High myopia (HM), an eye disorder with a refractive error ≤-6.0 diopters, has multifactorial etiology with environmental and genetic factors involved. Recent studies confirm the impact of alterations in DNA methylation and microRNAs (miRNAs) on myopia. Here, we studied the combined aspects evaluating to the role of methylation of miRNA encoding genes in HM. Materials and Methods: From the genome-wide DNA methylation data of 18 Polish children with HM and 18 matched controls, we retrieved differentially methylated CG dinucleotides localized in miRNA encoding genes. Putative target genes of the highest-ranked miRNAs were obtained from the miRDB and included in overrepresentation analyses in the ConsensusPathDB. Expression of target genes was assessed using the RNA sequencing data of retinal ARPE-19 cell line. Results: We identified differential methylation of CG dinucleotides in promoter regions of MIR3621, MIR34C, MIR423 (increased methylation level), and MIR1178, MIRLET7A2, MIR885, MIR548I3, MIR6854, MIR675, MIRLET7C, MIR99A (decreased methylation level) genes. Several targets of these miRNAs, e.g. GNAS, TRAM1, CTNNB1, EIF4B, TENM3 and RUNX were previously associated with myopia/HM/refractive error in Europeans in genome-wide association studies. Overrepresentation analyses of miRNAs' targets revealed enrichment in pathways/processes related to eye structure/function, such as axon guidance, transcription, focal adhesion, and signaling pathways of TGF-ß, insulin, MAPK and EGF-EGFR. Conclusion: Differential methylation of indicated miRNA encoding genes might influence their expression and contribute to HM pathogenesis via disrupted regulation of transcription of miRNAs' target genes. Methylation of genes encoding miRNAs may be a new direction in research on both the mechanisms determining HM and non-invasive indicators in diagnostics.