A high-throughput and low-waste viability assay for microbes.

Counting viable cells is a universal practice in microbiology. The colony-forming unit (CFU) assay has remained the gold standard to measure viability across disciplines, but it is time-intensive and resource-consuming. Here we describe the geometric viability assay (GVA) that replicates CFU measurements over 6 orders of magnitude while reducing over 10-fold the time and consumables required. GVA computes a sample's viable cell count on the basis of the distribution of embedded colonies growing inside a pipette tip. GVA is compatible with Gram-positive and Gram-negative planktonic bacteria (Escherichia coli, Pseudomonas aeruginosa and Bacillus subtilis), biofilms and fungi (Saccharomyces cerevisiae). Laborious CFU experiments such as checkerboard assays, treatment time-courses and drug screens against slow-growing cells are simplified by GVA. The ease and low cost of GVA evinces that it can replace existing viability assays and enable viability measurements at previously impractical scales.

High Throughput Viability Assay for Microbiology.

Counting viable cells is a universal practice in microbiology. The colony forming unit (CFU) assay has remained the gold standard to measure viability across disciplines; however, it is time-intensive and resource-consuming. Herein, we describe the Geometric Viability Assay (GVA) that replicates CFU measurements over 6-orders of magnitude while reducing over 10-fold the time and consumables. GVA computes a sample's viable cell count based on the distribution of embedded colonies growing inside a pipette tip. GVA is compatible with gram-positive and -negative planktonic bacteria, biofilms, and yeast. Laborious CFU experiments such as checkerboard assays, treatment time-courses, and drug screens against slow-growing cells are simplified by GVA. We therefore screened a drug library against exponential and stationary phase E. coli leading to the discovery of the ROS-mediated, bactericidal mechanism of diphenyliodonium. The ease and low cost of GVA evinces it can accelerate existing viability assays and enable measurements at previously impractical scales.

Photoactivated antibiotics to treat intracellular infection of bacteria.

Antibiotic resistance combined with pathogen internalization leads to debilitating infections. Here we test novel superoxide producing, stimuli-activated quantum dots (QDs), to treat an intracellular infection of Salmonella enterica serovar Typhimurium in an osteoblast precursor cell line. These QDs are precisely tuned to reduce dissolved oxygen to superoxide and kill bacteria upon stimulation (e.g., light). We show QDs provide tunable clearance at various multiplicities of infection and limited host cell toxicity by modulating their concentration and stimuli intensity, proving the efficacy of superoxide producing QDs for intracellular infection treatment and establishing a framework for further testing in different infection models.

Light-activated quantum dot potentiation of antibiotics to treat drug-resistant bacterial biofilms.

CdTe-2.4 eV quantum dots (QDs) show excellent efficacy due to their tunability and photo-potentiation for sterilizing drug-resistant planktonic cultures without harming mammalian cells but this QD fabrication has not been tested against biofilms. While the QD attack mechanism-production of superoxide radicals-is known to stimulate biofilm formation, here we demonstrate that CdTe-2.4 eV QD-antibiotic combination therapy can nearly eradicate Escherichia coli, methicillin-resistant Staphylococcus aureus, and Pseudomonas aeruginosa biofilms. CdTe-2.4 eV QD versatility, safety, and ability to potentiate antibiotics makes them a potential treatment strategy for biofilm-associated infections.