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Item Search for a Shared Genetic or Biochemical Basis for Biofilm Tolerance to Antibiotics across Bacterial Species(American Society for Microbiology, 2022-04) Stewart, Philip S.; Williamson, Kerry S.; Boegli, Laura; Hamerly, Timothy; White, Ben; Scott, Liam; Hu, Xiao; Mumey, Brendan M.; Franklin, Michael J.; Bothner, Brian; Vital-Lopez, Francisco G.; Wallqvist, Anders; James, Garth A.Is there a universal genetically programmed defense providing tolerance to antibiotics when bacteria grow as biofilms? A comparison between biofilms of three different bacterial species by transcriptomic and metabolomic approaches uncovered no evidence of one. Single-species biofilms of three bacterial species (Pseudomonas aeruginosa, Staphylococcus aureus, and Acinetobacter baumannii) were grown in vitro for 3 days and then challenged with respective antibiotics (ciprofloxacin, daptomycin, and tigecycline) for an additional 24 h. All three microorganisms displayed reduced susceptibility in biofilms compared to planktonic cultures. Global transcriptomic profiling of gene expression comparing biofilm to planktonic and antibiotic-treated biofilm to untreated biofilm was performed. Extracellular metabolites were measured to characterize the utilization of carbon sources between biofilms, treated biofilms, and planktonic cells. While all three bacteria exhibited a species-specific signature of stationary phase, no conserved gene, gene set, or common functional pathway could be identified that changed consistently across the three microorganisms. Across the three species, glucose consumption was increased in biofilms compared to planktonic cells, and alanine and aspartic acid utilization were decreased in biofilms compared to planktonic cells. The reasons for these changes were not readily apparent in the transcriptomes. No common shift in the utilization pattern of carbon sources was discerned when comparing untreated to antibiotic-exposed biofilms. Overall, our measurements do not support the existence of a common genetic or biochemical basis for biofilm tolerance against antibiotics. Rather, there are likely myriad genes, proteins, and metabolic pathways that influence the physiological state of individual microorganisms in biofilms and contribute to antibiotic tolerance.Item A High-Throughput Metabolic Microarray Assay Reveals Antibacterial Effects of Black and Red Raspberries and Blackberries against Helicobacter pylori Infection(MDPI AG, 2021-07) Goodman, Candace; Lyon, Katrina N.; Scotto, Aitana; Smith, Cyra; Sebrell, Thomas A.; Gentry, Andrew B.; Bala, Ganesh; Stoner, Gary D.; Bimczok, DianeHelicobacter pylori infection is commonly treated with a combination of antibiotics and proton pump inhibitors. However, since H. pylori is becoming increasingly resistant to standard antibiotic regimens, novel treatment strategies are needed. Previous studies have demonstrated that black and red berries may have antibacterial properties. Therefore, we analyzed the antibacterial effects of black and red raspberries and blackberries on H. pylori. Freeze-dried powders and organic extracts from black and red raspberries and blackberries were prepared, and high-performance liquid chromatography was used to measure the concentrations of anthocyanins, which are considered the major active ingredients. To monitor antibiotic effects of the berry preparations on H. pylori, a high-throughput metabolic growth assay based on the Biolog system was developed and validated with the antibiotic metronidazole. Biocompatibility was analyzed using human gastric organoids. All berry preparations tested had significant bactericidal effects in vitro, with MIC90 values ranging from 0.49 to 4.17%. Antimicrobial activity was higher for extracts than powders and appeared to be independent of the anthocyanin concentration. Importantly, human gastric epithelial cell viability was not negatively impacted by black raspberry extract applied at the concentration required for complete bacterial growth inhibition. Our data suggest that black and red raspberry and blackberry extracts may have potential applications in the treatment and prevention of H. pylori infection but differ widely in their MICs. Moreover, we demonstrate that the Biolog metabolic assay is suitable for high-throughput antimicrobial susceptibility screening of H. pylori.