Karolina Bielskė defended her thesis entitled "Immunodetection of factors conferring bacterial resistance to antibiotics" for the degree of Doctor of Science in Chemical Engineering.
Scientific supervisor: Prof. Dr. Aurelija Žvirblienė (Vilnius University, Technological Sciences, Chemical Engineering).
Composition of the Dissertation Defense Board: Chairperson - Prof. Dr. Edita Sužiedėlienė (Vilnius University, Natural Sciences, Biochemistry); Dr. Rima Budvytytė (Vilnius University, Technological Sciences, Chemical Engineering), Prof. Dr. Rimantas Daugelavičius (Vytautas Magnus University, Technological Sciences, Chemical Engineering), Dr. Algirdas Grevys (Thermo Fisher Scientific, Technological Sciences, Chemical Engineering), Prof. Dr. Modestas Ružauskas (Lithuanian University of Health Sciences, Agricultural Sciences, Veterinary).
Bacterial resistance to antimicrobial agents is one of the most significant global threats to human health in this century. Due to the widespread use of antimicrobial compounds in society, healthcare institutions, and livestock farming, bacteria are rapidly acquiring new mechanisms of antibiotic resistance. Currently, β-lactam antibiotics account for approximately 60% of all antibiotics used in the healthcare system; therefore, bacterial antibiotic resistance factors that hydrolyze these antibiotics – specifically, β-lactamases – can be selected as diagnostic targets. Monoclonal antibodies (mAbs) specific to β-lactamases can be employed for the development of advanced diagnostic systems.
In this work, the diagnostic targets AmpC and metallo-β-lactamases were selected based on their relevance and importance. Two strategies were applied for mAb development. Using the first strategy – recombinant β-lactamases together with hybridoma technology – four mAb collections were generated against ACT-14, NDM-1, PDC-195, and CMY-34 β-lactamases. Using the second strategy, a collection of monoclonal antibodies (mAbs) with broad reactivity to AmpC β-lactamases was developed. The immunogen used was a chimeric, nanotube-forming bacteriophage protein containing an inserted 17-amino-acid sequence from the DHA-1 β-lactamase, which is characterized by high conservation among AmpC enzymes.
The developed mAb collections were thoroughly characterized, and the mAbs demonstrating high affinity for their antigens were tested in various immunochemical assay formats. Their diagnostic potential was confirmed by testing them with bacterial isolates that naturally produce β-lactamases.