Human organoid biofilm model for assessing antibiofilm activity of novel agents

Abstract

Bacterial biofilms cause 65% of all human infections and are highly resistant to antibiotic therapy but lack specific treatments. To provide a human organoid model for studying host-microbe interplay and enabling screening for novel antibiofilm agents, a human epidermis organoid model with robust methicillin-resistant Staphylococcus aureus (MRSA) USA300 and Pseudomonas aeruginosa PAO1 biofilm was developed. Treatment of 1-day and 3-day MRSA and PAO1 biofilms with antibiofilm peptide DJK-5 significantly and substantially reduced the bacterial burden. This model enabled the screening of synthetic host defense peptides, revealing their superior antibiofilm activity against MRSA compared to the antibiotic mupirocin. The model was extended to evaluate thermally wounded skin infected with MRSA biofilms resulting in increased bacterial load, cytotoxicity, and pro-inflammatory cytokine levels that were all reduced upon treatment with DJK-5. Combination treatment of DJK-5 with an anti-inflammatory peptide, 1002, further reduced cytotoxicity and skin inflammation.

Multifunctional Antibiotic–Host Defense Peptide Conjugate Kills Bacteria, Eradicates Biofilms, and Modulates the Innate Immune Response

“…We established the skin model based on N/TERT cells as recently described . In brief, N/TERT cells in DermaLife K Keratinocyte Complete Medium were seeded at a concentration of 3 × 10 5 per 400 μL on filter inserts (ThinCert Cell culture insert, Greiner bio-one, Kremsmünster, Austria) in a 12-well plate (ThinCert, Greiner bio-one).…”

Calcium carbonate mineralization is essential for biofilm formation and lung colonization

“…To further explore the role of biomineralization during lung infection by P. aeruginosa , we set up a lung ex vivo model. Organ cultures derived from lung and colon tissues have been applied previously to study biofilm infections and viral tropism ( Kolodkin-Gal et al., 2007 , 2008 ; Massler et al., 2011 ; Wu et al., 2021 ) and thus can potentially be used to study lung infections. We cultured mice lungs and then infected them with P. aeruginosa constitutively expressing GFP.…”

Competition between Pseudomonas aeruginosa and Staphylococcus aureus is dependent on intercellular signaling and regulated by the NtrBC two-component system

“…A human air liquid interface organoid model 42 was modified by using Ker-CT human keratinocytes (ATCC CRL_4048) that were routinely cultured in Keratinocyte-SFM medium (Gibco) at 37 °C, 5% CO 2 . Human skin-equivalent organoids were formed by seeding cells on Transwell filter inserts (0.4 μm pore size) in deep 12-well ThinCert™ plates containing DermaLife K Keratinocyte Complete Medium (Lifeline Cell Technology) prepared according to the manufacturer’s specifications.…”

“…Human skin-equivalent organoids were formed by seeding cells on Transwell filter inserts (0.4 μm pore size) in deep 12-well ThinCert™ plates containing DermaLife K Keratinocyte Complete Medium (Lifeline Cell Technology) prepared according to the manufacturer’s specifications. After growth to confluency the medium in the Transwells above the keratinocyte layer was removed for 2–3 days to initiate multi-structured skin formation 42 . Prior to infection, K0 medium (Dulbecco’s Modified Eagle Medium supplemented with Ham’s F-12 (hydrocortisone, isopreterenol, insulin, selenious acid, L-serine and L-carnitine; Gibco) was added to the wells.…”