Pracownie naukowe

Badania

Najważniejsze osiągnięcia badawcze

• Bacteriocin recognition and molecular mode of action. We established the role of the mannose phosphotransferase system (Man-PTS) as a receptor for structurally diverse class II bacteriocins, characterized non-pediocin-like Man-PTS-targeting peptides and identified determinants of receptor specificity. Work on garvicin C revealed a distinctive V-shaped bacteriocin and an insertion-based mode of Man-PTS engagement (PNAS Nexus, 2024; Biophysical Journal, 2026).
• Mechanisms of resistance to antimicrobial peptides. We identified molecular determinants of bacteriocin resistance and cross-susceptibility to membrane-active antibiotics, including the roles of lipid metabolism (dgkB, dxsA), the YsaCB-KinG-LlrG regulatory system in Lactococcus lactis and the LiaFSR-LiaX response system in Enterococcus faecium (International Journal of Molecular Sciences, 2021; Antimicrobial Agents and Chemotherapy, 2021; subsequent studies).
• Molecular biology of dairy bacteriophages. Our bacteriophage research, led by Agnieszka Szczepankowska, identified and functionally characterized Gp21 as an SF4-type DNA helicase of the lytic Lactococcus lactis phage 94p4 and established functional interactions with its Gp22 and Gp23 accessory proteins. These findings define components of a phage replication module with relevance to the biology of phages affecting dairy starter cultures (International Journal of Biological Macromolecules, 2025; Journal of Dairy Science, 2026).
• Genome-to-function profiling of candidate probiotic strains. We integrated comparative genomics, phenotypic and antimicrobial screening, extracellular metabolomics and selected epithelial-model measurements to investigate strain-specific functional properties. Recent studies examined candidate strains in gut-skin-axis-oriented in vitro screening and compared the metabolite profiles of Lactiplantibacillus plantarum LP140 and Bifidobacterium breve BB010 in the context of gut-brain-axis-related hypotheses (Probiotics and Antimicrobial Proteins and BMC Microbiology, 2026).
• Applied microbial biotechnology and knowledge transfer. Our research contributed to the experimental characterization of lactic acid bacteria capable of reducing the immunoreactivity of gliadin peptides and to work underpinning the commercially available Gluticon formulation and patent application P.447563. We also contributed as co-authors to a published multicentre, randomized, placebo-controlled study of LP140 and LPC100 in postmenopausal women (2024); the clinical trial was sponsored and led by external collaborators. Further research addresses functional strain selection, bacterial metabolic interactions, propionate-related fermentation strategies and preclinical/translational projects with external partners. The laboratory's role in these projects is specified according to its actual research contribution; product efficacy is not inferred from in vitro data.

Opis badań

LABORATORY PROFILE AND RESEARCH STRATEGY

The Laboratory of Applied Microbiology (PN-39) is led by Tamara Aleksandrzak-Piekarczyk and comprises two researchers: Tamara Aleksandrzak-Piekarczyk and Agnieszka Szczepankowska. Their complementary interests bring together the molecular basis of bacteriocin activity and resistance, functional and applied microbiology, and fundamental studies of bacteriophage DNA replication. The laboratory builds on long-standing expertise in Lactococcus lactis, lactic acid bacteria, microbial genetics and biotechnology.

BACTERIOCINS: RECEPTOR RECOGNITION AND MODE OF ACTION

We study how ribosomally synthesized antimicrobial peptides recognize and inhibit Gram-positive bacteria. A central model is the mannose phosphotransferase system (Man-PTS), which functions both as a sugar transporter and a receptor for multiple class II bacteriocins. By combining genetic and microbiological tests, resistance mapping, sequence and structure analyses, and targeted mutagenesis, we investigate specificity at the peptide–receptor interface. Our work established an expanded group of non-pediocin-like Man-PTS-binding bacteriocins and explored their structural diversity. Recent research on the V-shaped bacteriocin garvicin C uncovered an atypical mode of receptor interaction through insertion between Man-PTS domains.

BACTERIOCIN RESISTANCE AND CELL-ENVELOPE ADAPTATION

We analyze resistant bacterial mutants to determine how changes in peptide receptors, lipid metabolism and cell-envelope stress response pathways influence antimicrobial susceptibility. Work on Lactococcus lactis has implicated dgkB and dxsA as well as the four-component YsaCB–KinG–LlrG system. Related studies in Enterococcus faecium identified roles for LiaFSR–LiaX-associated processes. These investigations help explain bacterial adaptation, cross-resistance and reciprocal susceptibility to bacteriocins and membrane-active peptide antibiotics; they support evidence-based assessment of possible applications and limitations of antimicrobial peptides.

BACTERIOPHAGES, DNA HELICASES AND DAIRY STARTER CULTURE BIOLOGY

A major research line led by Agnieszka K. Szczepankowska focuses on lytic bacteriophages of Lactococcus lactis and Lactococcus cremoris. Phage infections are an important cause of disruption in dairy fermentations using lactococcal starter cultures. Our recent studies addressed the replication machinery of the prolate-headed Ceduovirus phage 94p4. We characterized Gp21 as an SF4-family helicase, examining its DNA-binding and ATP-dependent unwinding properties and conserved functional motifs. We also identified functional relationships between Gp21 and the phage proteins Gp22 and Gp23, thereby defining a replication-associated module. This line links molecular mechanisms of phage genome replication to the broader problem of phage-host interactions and fermentation reliability. The key findings were reported in International Journal of Biological Macromolecules (2025) and Journal of Dairy Science (2026).

FUNCTIONAL GENOMICS, PHENOTYPIC MICROARRAYS AND MICROBIAL METABOLISM

We investigate lactic acid bacteria, bifidobacteria and selected other microorganisms using complementary molecular, phenotypic and biochemical approaches. An important capability is Biolog Phenotype MicroArray analysis, enabling systematic comparison of substrate utilization, growth phenotypes and cellular responses under defined conditions. Depending on the experimental system and available validated panels, this platform supports microbial phenotyping and specialized analyses of other cell types. The approach is combined with growth kinetics, stress-tolerance assays, comparative genome analysis and interpretation of metabolic pathways. We also investigate metabolic interactions in bacterial combinations, including cross-feeding and propionate-oriented fermentation strategies. Whole-genome sequencing, advanced mass-spectrometric measurements and selected specialized cell assays may be performed in collaboration with relevant facilities or partner laboratories; these are not presented as instruments operated exclusively by PN-39.

PROBIOTIC CANDIDATES, MULTI-OMICS AND HOST-INTERACTION MODELS

In collaboration with academic, clinical and industrial partners, we examine the strain-specific metabolic and biological properties of candidate probiotics and related microorganisms. Our studies combine antagonistic activity against indicator organisms, extracellular metabolite measurements and selected in vitro model assays. We investigate bacterial adhesion to biotic and abiotic surfaces and explore its molecular basis by linking adhesion phenotypes with candidate adhesins, cell-surface structures and genetic determinants. In epithelial cell-line models, we examine barrier integrity using transepithelial electrical resistance (TEER), together with tight-junction-associated endpoints where experimentally assessed. A 2026 multi-criterion study evaluated candidate strains for gut–skin-axis-oriented research through antimicrobial profiles, targeted LC-MS/MS metabolomics, a preliminary surface-binding test and TEER measurements in an intestinal epithelial model. A complementary 2026 study compared genome-based metabolic predictions with experimentally measured extracellular metabolites of L. plantarum LP140 and B. breve BB010 in the context of gut-brain-axis-associated hypotheses. These results support candidate prioritization and further mechanistic research; they do not establish clinical benefit. In the PIVOT project, the laboratory contributes microbial and multi-omics expertise to collaborative investigations involving probiotic and postbiotic candidates and colorectal-cancer-oriented experimental models.

RESEARCH INFRASTRUCTURE AND COLLABORATIVE SERVICES

The laboratory provides scientific oversight for selected research infrastructure supporting in-house, collaborative and contract R&D. The Biolog OmniLog Phenotype MicroArray platform enables systematic profiling of microbial metabolism and stress responses; Bioscreen C supports automated growth-kinetic assays. The laboratory uses a CO₂ incubator for mammalian cell culture and epithelial cell-line models to investigate bacterial–host interactions, including selected co-culture and epithelial-barrier assays. These capabilities may be offered through collaborative or commissioned studies under IBB PAS procedures.

APPLIED RESEARCH, INDUSTRIAL PARTNERSHIPS AND TRANSLATION

The laboratory conducts research and development activities with industrial and academic partners, including functional screening, comparative characterization of bacterial strains, antimicrobial testing and analysis of selected bioactive metabolites. Our published work on bacterial peptidase activities and reduction of gliadin-peptide immunoreactivity contributed to research underlying the Gluticon formulation, which is publicly available as a commercial product; related intellectual property is described in a Polish patent application. Members of the laboratory co-authored a published randomized, placebo-controlled clinical study involving LP140 and LPC100 in postmenopausal women (2024), with experimental contributions documented in the article; the trial sponsor and clinical leadership were external. The laboratory also participates in the Polish-Turkish LAMPAF project, integrating microbial genomics, functional strain characterization and metabolite profiling with industrial-scale fermentation and the development of functional food additives. We are open to scientifically defined collaborations involving Biolog Phenotype MicroArrays, strain characterization and applied microbial biotechnology.

Metodologia

• Bacterial genetics and molecular microbiology: cultivation and genetic manipulation of lactic acid bacteria and selected other microorganisms; PCR/qPCR, cloning, sequence-based identification, gene-expression and genetic-function studies, including molecular determinants of surface-associated phenotypes.
• Bacteriocins and antimicrobial susceptibility: activity screening, indicator-strain assays, resistance-mutant selection and characterization, receptor–peptide interaction analysis, membrane-associated phenotypes.
• Bacteriophages and proteins: cultivation and functional study of lactococcal phages, identification and characterization of phage replication proteins, recombinant protein expression/purification, DNA-binding, helicase and ATP-dependent activity assays, and structure-function analyses.
• Phenotypic profiling and kinetic growth analysis: Biolog OmniLog Phenotype MicroArrays for high-throughput microbial metabolism, substrate use and stress-response profiling; Bioscreen C for automated growth curves; yeast/fungal and specialized cellular assays where validated. Analyses may support research collaborations and agreed contract studies.
• Genome-to-phenotype analysis: comparative and functional genomics, genomic annotation, pathway reconstruction, safety-related gene screening, integration of genomic, phenotypic and metabolite datasets; sequencing may be outsourced or conducted with partner facilities.
• Bacterial adhesion and biofilms: adhesion and surface-binding assays on biotic and abiotic substrates; selected biofilm tests; comparison of adhesion phenotypes with genomic and molecular determinants, including candidate adhesins and cell-surface structures.
• Cell culture and host-interaction studies: cultivation of mammalian epithelial cell lines in a CO₂ incubator; selected bacterial–epithelial co-culture models and functional screening; TEER measurements and tight-junction-related assessments where experimentally performed; specialized biochemical and metabolomic measurements with collaborators.
• Structural and analytical collaborations: peptide/protein modelling, sequence analysis and experimental structure-function testing; advanced NMR, LC-MS/MS and other specialized measurements are performed with collaborators and shared facilities as applicable.

Wybrane publikacje

• Tymoszewska A., Aleksandrzak-Piekarczyk T. Subclass IId bacteriocins targeting mannose phosphotransferase system: Structural diversity and implications for receptor interaction and antimicrobial activity. PNAS Nexus. 2024;3(9):pgae381. DOI: 10.1093/pnasnexus/pgae381.
• Tymoszewska A., Steczkiewicz K., Aleksandrzak-Piekarczyk T. An atypical V-shaped garvicin C bacteriocin targets Man-PTS via domain insertion. Biophysical Journal. 2026;125(1):99–112. DOI: 10.1016/j.bpj.2025.11.010.
• Santo A., Chmielewska-Jeznach M., Steczkiewicz K., Aleksandrzak-Piekarczyk T., Szczepankowska A.K. Structural and functional insights into Gp21 as a new SF4 helicase of prolate-headed Lactococcus lactis phage 94p4. International Journal of Biological Macromolecules. 2025;320(Pt 2):145668. DOI: 10.1016/j.ijbiomac.2025.145668.
• Aleksandrzak-Piekarczyk T., Tymoszewska A., Radkiewicz M., Szczepankowska A.K., Samborowska E., Koryszewska-Bagińska A., Cukrowska B. Multi-Criterion In Vitro Profiling of Probiotic Strains for Preliminary Gut–Skin-Axis-Oriented Candidate Prioritization. Probiotics and Antimicrobial Proteins. Published online 27 October 2026. DOI: 10.1007/s12602-026-11269-3.
• Aleksandrzak-Piekarczyk T., Szczepankowska A.K., Radkiewicz M., Samborowska E., Majsiak E., Cukrowska B. Comparative genomic and extracellular metabolomic profiling of Lactiplantibacillus plantarum LP140 and Bifidobacterium breve BB010 reveals distinct gut–brain-axis-associated metabolite profiles. BMC Microbiology. Published online 30 October 2026. DOI: 10.1186/s12866-026-05842-7.

Współpraca

• Katarzyna Stadnicka, Collegium Medicum, Nicolaus Copernicus University, Bydgoszcz, Poland.
• Edyta Szymańska, Department of Gastroenterology, Hepatology, Feeding Disorders and Paediatrics, The Children's Memorial Health Institute, Warsaw, Poland.
• Lidia Stasiak-Różańska, Warsaw University of Life Sciences (SGGW), Warsaw, Poland.
• John C. Vederas, University of Alberta, Edmonton, Canada.
• Djamel Drider, University of Lille, France.
• Ömer Şimşek, Department of Food Engineering, Yıldız Technical University, Istanbul, Türkiye.
• Nordic Biotic Ltd., Warsaw, Poland.
• Propharma Science Sp. z o.o., Warsaw, Poland.
• JHJ Sp. z o.o., Nowa Wieś (Gizałki), Poland.
• Olimp Laboratories Sp. z o.o., Pustynia (Dębica), Poland.
• MAYSA GIDA SANAYİ VE TİCARET A.Ş., Istanbul, Türkiye.