The overall goal of PHILMARINE-BLI is to identify promising BLI lead compounds from marine sediment-derived Actinomycetes from Tubbataha Reefs stored in the University of San Agustin biobank. We will combine the actinomycete extracts with common beta-lactams and discover combinations with activity against well-characterized, beta-lactam-resistant, Gram-negative pathogens circulating in the Philippines. The PHILMARINE-BLI project will be structured as described below and in the graphical abstract.
Characterization of BLA-inactivation genes of Gram-negative clinical isolates at a tertiary hospital in the Philippines. Different genes that encode for beta-lactamases from unique strains of Escherichia coli, Klebsiella pneumoniae, Acinetobacter baumannii, and Pseudomonas aeruginosa will be identified. This part of the project will produce an exclusive panel of beta-lactam-resistant, genetically characterized, clinical isolates with Filipino origin.
Primary screen for inhibitory activity of crude extracts from Philippine marine actinomycetes when used in combination with commonly used beta-lactam antibiotics against Gram-negative clinical isolates at a tertiary hospital in the Philippines. These in vitro experiments in bacterial cell culture will select promising crude extracts with inhibitory activity when used in combination therapy (with no antibacterial activity when used alone).
Secondary screen for enzyme-targeted inhibitory activity of BLI-containing actinomycete crude extracts using commercially available purified serine- and metallo-beta-lactamases. These in vitro studies using pure enzymes will test promising crude extracts with inhibitory activities against serine- or metallo-beta-lactamases (or both).
Bioactivity-guided isolation of beta-lactamase inhibitors from the chosen extracts and determination of their enzyme binding via in silico docking and molecular dynamics simulation. This section will provide the identity of BLI lead compounds in the screened extracts, elucidate molecular interactions with serine- or metallo-beta-lactamases, and gain an understanding of possible mechanisms-of-inhibition.
