Maciej ObręBski

I. Escherichia coli is one of the most versatile bacterial species associated with humans and animals. It is a frequent member of the intestinal microbiota, but selected lineages have acquired viru...

Uropathogenic Escherichia coli (UPEC) often causes recurrent urinary tract infections, where biofilms and persister cells can promote recalcitrance. This study evaluated whether secretion-system marker genes associate with antimicrobial susceptibility, biofilm formation, and biofilm-associated persistence in clinical urinary isolates. A total of 71 isolates were screened by PCR for fimH and markers of secretion systems: type I ( hlyA ), type II ( gspD ), type V ( ag43 ), and type VI ( hcp ). Minimum inhibitory concentrations (MICs) were measured by Etest, biofilm biomass by crystal violet assay, and persister survival in strong biofilm producers after antibiotic challenge. fimH was detected in 91.5% isolates; gspD and ag43 were common (71.8% and 80.3%), while hlyA and hcp occurred in 32.4% and 46.5%. Marker co-occurrence was frequent, with 19.7% carrying all four secretion markers, and 8.5% with no secretion markers detected. A total of 81.7% isolates formed biofilm, including 18.3% strong producers. Individual marker prevalence did not differ across biofilm categories. Carriage of hlyA and hcp was associated with lower MIC distributions for selected antibiotics (including fosfomycin, ciprofloxacin, and ampicillin-sulbactam), whereas gspD and ag43 showed no consistent MIC associations. The biofilm category was linked to non-susceptibility to ampicillin-sulbactam, but not the other agents tested. Strong biofilm producers showed biphasic killing consistent with persister formation, and hcp carriage tended to align with higher ciprofloxacin survival. These findings map secretion-system markers in clinical UPEC and suggest that a type VI secretion system may track with biofilm-associated persistence. IMPORTANCE: Recurrent urinary tract infections (rUTI) often relapse after antibiotic treatment because bacteria can generate persister cells—temporary survivors that tolerate antibiotics without being genetically resistant. Clinicians, therefore, lack markers that flag isolates likely to persist. In 71 clinical urinary Escherichia coli isolates, this study surveyed genes encoding several secretion systems, molecular machines that help bacteria interact with each other and with host tissues. These markers were widespread and frequently co-occurred, but they did not track with stronger biofilm formation. However, a type VI secretion system marker ( hcp ) was associated with higher survival during ciprofloxacin exposure in strong-biofilm isolates, linking virulence-associated machinery with biofilm-associated persistence. Genetic signatures that reflect persistence, rather than routine resistance, could help prioritize follow-up testing and support development of strategies that target hard-to-eradicate rUTI infections.

Helichrysum arenarium (L.) Moench is a European native species typically associated with dry, sunny, sandy, and nutrient-poor habitats. In European herbal practice, preparations from H. arenarium inflorescences are traditionally used for the symptomatic relief of digestive complaints. Polyphenolic constituents, particularly flavonoid and chalcone derivatives, are among the most characteristic compounds detected in the inflorescences. Intensive harvesting and degradation of natural habitats have increased pressure on wild populations. Since H. arenarium is legally protected in many countries, biotechnological approaches for plant biomass production represent a promising alternative. In the present study, shoot and callus cultures were established, and detailed extract profiling was performed using ultra-high-performance liquid chromatography coupled with diode-array detection and electrospray ionization multistage mass spectrometry (UHPLC-DAD-ESI-MS³). Cytotoxic, antioxidant, anti-inflammatory, wound-healing, anti-collagenase, and anti-hyaluronidase activities were also evaluated. In vitro-derived materials were compared with inflorescences and shoots of the maternal plant. The analyzed materials showed pronounced tissue-dependent metabolic differentiation. The highest callus growth rate was observed for the Ha-C-1 line, which also accumulated the highest levels of chlorogenic acid (25.80 ± 3.09 mg/g dry extract) and isochlorogenic acid A (55.83 ± 13.78 mg/g dry extract). Apigenin was detected in the inflorescence extract, reaching 3.55 ± 1.42 mg/g dry extract. The extracts did not markedly affect HaCaT viability at 15.63–500 µg/mL, while the strongest bioactivities were observed for inflorescence and maternal plant extracts.

Somatic hybridization represents a powerful tool for generating novel chemotypes with enhanced biosynthetic capabilities. This study provides the first comprehensive phytochemical characterization of interspecific somatic hybrids between Gentiana cruciata L. and Gentiana tibetica King ex Hook.f., two medicinally important yet regionally rare gentians. A total of 107 compounds were detected using UHPLC-DAD-ESI-MS³, of which 31 were identified as metabolites across eight phytochemical classes. Comparative profiling revealed that all hybrids retained a conserved core of iridoids and secoiridoids while integrating lineage-specific compounds and producing hybrid-specific metabolites not detected in either parent. Despite inheriting plastids from G. tibetica, hierarchical clustering showed that the phytochemical profiles of hybrid lines were more similar to G. cruciata, the donor of the nuclear genome. Quantitative analysis of the major secoiridoids, such as gentiopicroside, swertiamarin, and sweroside, demonstrated that several hybrid lines, particularly F30A-5 and F30A-6, matched or surpassed the biosynthetic output of G. tibetica, the more productive parent. These lines also exhibited elevated antioxidant capacity, underscoring their phytochemical and functional potential. Altogether, our findings show that somatic hybridization not only preserves but may amplify the secondary metabolite capacity of the parental genotypes, offering a viable platform for sustainable in vitro production of pharmacologically relevant secoiridoids.