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High-Throughput Screen for Inhibitors of Klebsiella pneumoniae Virulence Using a Tetrahymena pyriformis Co-Culture
Angela L Woods1, David Parker2, Meir M Glick3
1Infectious Diseases, Novartis Institutes for Biomedical Research Inc, 250 Massachusetts Avenue, Cambridge, Massachusetts 02139, United States.
Abstract:
The continuing emergence of antibacterial resistance reduces the effectiveness of antibiotics and drives an ongoing search for effective replacements. Screening compound libraries for antibacterial activity in standard growth media has been extensively explored and may be showing diminishing returns. Inhibition of bacterial targets that are selectively important under in vivo (infection) conditions and, therefore, would be missed by conventional in vitro screens might be an alternative. Surrogate host models of infection, however, are often not suitable for high-throughput screens. Here, we adapted a medium-throughput Tetrahymena pyriformis surrogate host model that was successfully used to identify inhibitors of a hyperviscous Klebsiella pneumoniae strain to a high-throughput format and screened circa 1.2 million compounds. The screen was robust and identified confirmed hits from different chemical classes with potent inhibition of K. pneumoniae growth in the presence of T. pyriformis that lacked any appreciable direct antibacterial activity. Several of these appeared to inhibit capsule/mucoidy, which are key virulence factors in hypervirulent K. pneumoniae. A weakly antibacterial inhibitor of LpxC (essential for the synthesis of the lipid A moiety of lipopolysaccharides) also appeared to be more active in the presence of T. pyriformis, which is consistent with the role of LPS in virulence as well as viability in K. pneumoniae.
Insights
This study adapted a Tetrahymena pyriformis model for high-throughput screening of antibacterial compounds. The screen identified novel inhibitors of Klebsiella pneumoniae virulence factors, offering a new avenue for antibiotic discovery.
Area of Science:
- Microbiology
- Drug Discovery
- Antimicrobial Resistance
Background:
- Antibiotic resistance necessitates novel therapeutic strategies.
- Conventional in vitro screening methods may miss essential antibacterial targets active during infection.
- Surrogate host models offer an alternative for identifying infection-specific antibacterial compounds.
Purpose of the Study:
- To adapt a Tetrahymena pyriformis surrogate host model for high-throughput screening.
- To identify novel antibacterial compounds targeting Klebsiella pneumoniae virulence in a host-associated environment.
- To discover inhibitors of bacterial targets missed by standard in vitro assays.
Main Methods:
- Adapted a medium-throughput Tetrahymena pyriformis model to a high-throughput format.
- Screened approximately 1.2 million compounds for inhibition of Klebsiella pneumoniae growth in the presence of T. pyriformis.
- Assessed direct antibacterial activity and identified key virulence factors targeted by confirmed hits.
Main Results:
- Identified potent inhibitors of Klebsiella pneumoniae growth in the T. pyriformis model, with minimal direct antibacterial activity.
- Several hits demonstrated inhibition of bacterial capsule/mucoidy, crucial virulence factors.
- A lipopolysaccharide (LPS) synthesis inhibitor (LpxC) showed enhanced activity in the presence of T. pyriformis.
Conclusions:
- The high-throughput Tetrahymena pyriformis model is effective for discovering novel antibacterial compounds targeting virulence.
- This approach can identify inhibitors of bacterial mechanisms important during infection, not detected by conventional screens.
- Findings support targeting capsule and LPS biosynthesis as strategies against hypervirulent Klebsiella pneumoniae.
