Darja Žgur Bertok

Background/Objectives: Gestational diabetes mellitus (GDM) is associated with altered maternal gut microbiota and increased risk of large-for-gestational age (LGA) births. The contribution of gut microbiota to fetal overgrowth in GDM, independent of glycemic control, remains unclear. Methods: In this pilot longitudinal study, the gut microbiota of 18 women with GDM was followed from the second (2T) to the third trimester (3T). Maternal fecal samples were analyzed by 16S rRNA gene sequencing, and associations between microbial profiles and infant birth weight were examined. In addition, these associations were adjusted for pre-pregnancy body mass index (BMI) and gestational weight gain (GWG). Results: Maternal gut microbiota of LGA infants exhibited consistently lower microbial diversity, a reduced Bacillota/Bacteroidota ratio, and enrichment of pro-inflammatory taxa including Prevotella, Sutterella, and Bilophila. Short-chain fatty acids (SCFAs)-producing genera such as Acinetobacter, Odoribacter, Faecalibacterium, and Lachnoclostridium were depleted. Although Bilophila was identified as a third-trimester biomarker with LEfSE approach, its association with LGA disappeared after adjusting for BMI and GWG. Conversely, Nitrospirota, Polaromonas, Acinetobacter, and Aeromonas correlated negatively with LGA even after BMI and GWG adjustment. Conclusions: These findings suggest that specific maternal microbiota signatures, together with pre-pregnancy adiposity, influence fetal overgrowth in GDM and may serve as early biomarkers or targets for preventive interventions.

Background: Short-chain fatty acids (SCFAs), which are produced by the microbial fermentation of undigested carbohydrates, play an important role in the metabolism and physiology of the host. SCFAs are involved in the regulation of maternal metabolism during pregnancy and influence weight gain, glucose metabolism, and metabolic hormones. Methods: In 2017, women who were treated for gestational diabetes mellitus (GDM) at the University Medical Centre Ljubljana were invited to participate in a longitudinal study. A total of 45 women were included in this study and comprehensively phenotyped. During the second and third trimester of pregnancy, the women with GDM provided fecal samples for SCFA analysis. The samples were analyzed by high-performance liquid chromatography for the simultaneous determination of acetate, propionate, and butyrate. Results: SCFA concentrations in feces differed between overweight/obese and normal-weight women with GDM. Acetate and propionate concentrations were significantly higher in pregnant women who were overweight or obese before pregnancy compared to normal-weight women but butyrate concentrations were not. Butyrate was elevated in the third trimester in the group with excessive gestational weight gain. Conclusions: The relationship between SCFAs and obesity is complex, and the association between SCFAs and GDM remains to be clarified. Regardless of the conflicting publications on the role of SCFAs, our study showed that higher acetate and propionate levels were associated with the weight categories of overweight or obesity before pregnancy and higher butyrate levels were associated with excessive gestational weight gain.

Short-chain fatty acids (SCFAs), which are produced by the microbial fermentation of undigested carbohydrates, play an important role in the metabolism and physiology of the host. SCFAs are involved in the regulation of maternal metabolism during pregnancy and influence weight gain, glucose metabolism and metabolic hormones. In 2017, women who were treated for gestational diabetes mellitus (GDM) at the University Medical Centre Ljubljana were invited to participate in a longitudinal study. A total of 45 women were included in the study and comprehensively phenotyped. During the second and third trimester of pregnancy, the GDM women provided fecal samples for SCFA analysis. The samples were analyzed by high-performance liquid chromatography for the simultaneous determination of acetate, propionate, and butyrate. SCFA concentrations in feces differed between overweight/obese and normal weight GDM women. Acetate and propionate concentrations were significantly higher in pregnant women who were overweight or obese before pregnancy compared to normal weight women, but butyrate concentrations were not. Butyrate was elevated in the third trimester in the group with excessive gestational weight gain. Regardless the conflicting publications on the role of SCFAs, our study showed that higher acetate and propionate levels were associated with overweight or obesity before pregnancy and higher butyrate levels were associated with excessive gestational weight gain. The relationship between SCFAs and obesity is complex, and the association between SCFAs and GDM remains to be clarified.

Escherichia coli is known to be an important uropathogenic agent. Several models were developed for investigating the uropathogensis of E. coli, including the recent biomimetic porcine urothelial in vitro model. The aim of this study was to assess the cytokine response of the cells of the biomimetic porcine urothelial model to different E. coli strains. The production of nine different cytokines in response to E. coli infection was evaluated using the commercial pre-configured immunoassay multiplex Cytokine & Chemokine 9-Plex Porcine ProcartaPlex™ Panel 1 kit. Our results showed that cells of the biomimetic porcine urothelial model reacted to the presence of all the employed different E. coli strains, albeit with some differences in levels and types of cytokines produced. Increased production of IL-10, IL-8, TNF-α, IL-1β, IL-4 and IL-12p40 was observed. Statistical analysis (Fisher’s exact test) revealed a correlation between the high fold change in the immune response and the presence of the cnf1 gene that encodes the cytotoxic necrotizing factor. Our results shed light on the cytokine response of normal urothelial cells to different E. coli strains and have the potential to fuel the search for understanding the mechanisms behind the different cytokine responses to different E. coli strains.

Urinary tract infections can be severe, sometimes fatal, diseases whose etiological pathogens are predominantly uropathogenic strains of E. coli (UPEC). To investigate the UPEC pathogenesis, several models have already been established with minor or major disadvantages. The aim was to develop a simple, fast, and inexpensive biomimetic in vitro model based on normal porcine urothelial (NPU) cells that are genetically and physiologically similar to human bladder urothelium and to perform basic studies of E. coli pathogenicity. Initially, the model was tested using a set of control E. coli strains and, subsequently, with human E. coli strains isolated either from patients with urinary infections or from the feces of healthy individuals. A drop in viability of NPU cells was used as a measure of the pathogenicity of the individual strain tested. To visualize the subcellular events, transmission and scanning electron microscopy was performed. The strains were tested for the presence of different virulence-associated genes, phylogroup, type of core lipid, O-serotype, and type of lipopolysaccharide and a statistical analysis of possible correlations between strains’ characteristics and the effect on the model was performed. Results showed that our model has the discriminatory power to distinguish pathogenic from non-pathogenic E. coli strains, and to identify new, potentially pathogenic strains.

The SOS response including two main proteins LexA and RecA, maintains the integrity of bacterial genomes after DNA damage due to metabolic or environmental assaults. Additionally, derepression of LexA-regulated genes can result in mutations, genetic exchange and expression of virulence factors. Here we describe the first comprehensive description of the in silico LexA regulon in Clostridium difficile, an important human pathogen.

The aim of the presented study was to estimate the prevalence, distribution and associations of different pathogenicity islands (PAI I536 to PAI IV536, PAI IJ96, PAI IIJ96, PAI ICFT073 and PAI IICFT073), iron uptake systems (genes iutA, iucD, iroN, iroCD, fyuA, irp2, iha, ireA, and hbp) and plasmids among extraintestinal pathogenic Escherichia coli (ExPEC) strains isolated from Slovenian patients. Twenty-nine ExPEC isolates obtained from the Institute of Microbiology and Immunology, Medical Faculty, University of Ljubljana were investigated for the presence of different pathogenicity islands and iron uptake systems with PCR, the plasmid content of the investigated strains was determined by molecular biology techniques. The significance of the found associations of the studied PAIs and iron uptake systems was analyzed with the Fisher’s exact test. PAI IV536 was found in 19, PAI II CFT073 in 6, PAI ICFT073 in 4, and PAI IIJ96 in one of the studied isolates. PAI I536, PAI II536, PAI III536 and PAI IJ96 were not detected in any studied isolate. In 19 of the studied isolates plasmids were detected. The irp2 was found in 20, fyuA in 19, iucD and iutA in 12, iha in 9, iroN in 8, iroCD in 7, ireA in 7 and hbp in 4 of studied isolates. PAI IV536 was statistically significantly associated with the yersiniabactin siderophore system and PAI ICFT073 was statistically significantly associated with the aerobactin siderophore system as well as Iha. To our knowledge this is the first report on PAIs and iron uptake systems among Slovenian ExPEC isolates, as well as a first report on PAIs, iron uptake systems and plasmids among isolates from skin and soft tissue infections.

The aim of the presented study was to identify the similarity of the plasmid pRK100 RepFIIA replicon (replication region) with similar replicons of other known plasmids of Enterobacteriaceae. For this purpose, within the determined nucleotide sequence of pRK100, theRepFIIA replicon rep genes/regions were identified. The nucleotide sequences of the pRK100 determined rep genes/regions were subsequently compared with the nucleotide sequences of other RepFIIA replicon rep genes/regions deposited in GenBank. Further, the nucleotide divergence between them was calculated. The obtained results clearly demonstrated, that the individual pRK100 rep regions are the same/most similar to rep regions from different plasmids. RepA2 of pRK100 is most similar to repA2 of pCP301, pINV_F6_M1382, pWR501 and R1, copA is the same as copA of plasmids pC15-1a and R100, repA6 of pRK100 is the same as repA6 in plasmids pC15-1a, pCP301, pINV_F6_M1382, pWR501, R1 and R100, repA1 is most similar to repA1 of the plasmid p1658/79, and repA4 of pRK100 is most similar to repA4 of pC15-1a. Hence, the composition of the pRK100 RepFIIA replicon is mosaic and unique among the plasmids.