Project description:Previously, we have shown that broad spectrum antibiotic treatment reduces reactive astrocyte phenotypes in the APPPS1-21 model of AD-related amyloidosis. We have also found that antibiotics selectively increases propionate levels and exogenous propionate treatment recapitulates phenotypes observed in antibiotic treated mice. In the current study, we wanted to assess astrocyte transcriptional state using bulk RNA sequencing. To accomplish this we used translating ribosome affinity purification (TRAP) sequencing, a ribosomal protein L10a is fused to eGFP under the control of a cell type specific promoter in a transgenic mouse model. We crossed APPPS1-21 mice to the Aldh1l1-eGFP/Rpl10a bacTRAP mouse model and progeny were treated with antibiotics, propionate, or VHL and performed bulk TRAPseq.
Project description:The inappropriate use of antibiotics is a severe public health problem worldwide, contributing to the emergence of multidrug-resistant (MDR) bacteria. To explore the possible impacts of the inappropriate use of antibiotics on the immune system, we use Klebsiella pneumoniae (K. pneumoniae) infection as an example and show that imipenem increases the mortality of mice infected by MDR K. pneumoniae. Further studies demonstrate that imipenem enhances the secretion of outer membrane vesicles (OMVs) with significantly elevated presentation of GroEL, which promotes the phagocytosis of OMVs by macrophages that depends on the interaction between GroEL and its receptor LOX-1. OMVs cause the pyroptosis of macrophages and the release of proinflammatory cytokines, which contribute to exacerbated inflammatory responses. We propose that the inappropriate use of antibiotics in the cases of infection by MDR bacteria such as K. pneumoniae might cause damaging inflammatory responses, which underlines the pernicious effects of inappropriate use of antibiotic.
Project description:The goal of this project is to develop a new class of urea-depsipeptide (UDEP) antibiotics to treat prosthetic joint infections (PJI). UDEPs kill bacteria through activation of the ClpP protease, causing cells to self-digest. This unique activating mechanism allows UDEPs to kill biofilms and non-growing persister cells, which are prevalent in PJI and explain why current antibiotics are largely ineffective. Current therapies involve weeks to months of antibiotic treatment, debridement surgeries, and medical device replacement. UDEPs have the potential to minimize surgical interventions due to PJI and improve patient care. PJI are primarily caused by the Gram-positive pathogens Staphylococcus aureus and epidermidis and the UDEPs are potently active against these pathogens, including multi-drug resistant strains. A recent advance in our UDEP medical chemistry program yielded a new compound which has improved safety, solubility, and bone penetration compared to first generation UDEPs. A preliminary study found that the compound was effective in a K-wire femur medullary canal implant model of PJI, which is known to be difficult to treat. In this project, we will evaluate if the compound is an acceptable pre-clinical candidate for PJI by testing it in a series of in vitro and in vivo studies focused on this indication. Specifically, the aims are to 1) scale up the compound; 2) determine the microbiological and biofilm killing effect against the main pathogens isolated from PJI; and 3) determine the efficacy of the compound in mouse and rabbit models of PJI.
Project description:Understanding how M. tuberculosis survives during antibiotic treatment is necessary to rationally devise more effective tuberculosis chemotherapy regimens. Using genome-wide mutant fitness profiling and the mouse model of TB, we identified genes that alter antibiotic efficacy specifically in the infection environment.
Project description:Deeper understanding of antibiotic-induced physiological responses is critical to identifying means for enhancing our current antibiotic arsenal. Bactericidal antibiotics with diverse targets have been hypothesized to kill bacteria, in part, by inducing production of damaging reactive species. This notion has been supported by many groups, but recently challenged. Here we robustly test the hypothesis using biochemical, enzymatic and biophysical assays along with genetic and phenotypic experiments. We first used a novel intracellular hydrogen peroxide (H2O2) sensor, together with a chemically diverse panel of fluorescent dyes sensitive to an array of reactive species, to demonstrate that antibiotics broadly induce redox stress. Subsequent gene expression analyses reveal that complex antibiotic-induced oxidative stress responses are distinct from canonical responses generated by supra-physiological levels of H2O2. We next developed a method to dynamically quantify cellular respiration and found that bactericidal antibiotics elevate oxygen consumption, indicating significant alterations to bacterial redox physiology. We further show that catalase or DNA mismatch repair enzyme overexpression, as well as antioxidant pre-treatment limit antibiotic lethality, indicating that reactive oxygen species causatively contribute to antibiotic killing. Critically, the killing efficacy of antibiotics was diminished under strict anaerobic conditions, but could be enhanced by exposure to molecular oxygen or addition of alternative electron acceptors, suggesting that environmental factors play a role in killing cells physiologically primed for death. This work provides direct evidence that bactericidal antibiotics, downstream of their target-specific interactions, induce complex redox alterations that contribute to cellular damage and death, thus supporting an evolving, expanded model of antibiotic lethality. Here, we used microarrays to analyze oxidative stress responses to bactericidal antibiotic treatment in wildtype and mutant E coli
Project description:Non-typeable Haemophilus influenzae (NTHi) is a common acute otitis media pathogen, with an incidence that is increased by previous antibiotic treatment. NTHi is also an emerging causative agent of other chronic infections in humans, some linked to morbidity, and all of which impose substantial treatment costs. In this study we explore the possibility that antibiotic exposure may stimulate biofilm formation by NTHi bacteria. We discovered that sub-inhibitory concentrations of beta-lactam antibiotic (i.e., amounts that partially inhibit bacterial growth) stimulated the biofilm-forming ability of NTHi strains, an effect that was strain and antibiotic dependent. When exposed to sub-inhibitory concentrations of beta-lactam antibiotics NTHi strains produced tightly packed biofilms with decreased numbers of culturable bacteria but increased biomass. The ratio of protein per unit weight of biofilm decreased as a result of antibiotic exposure. Antibiotic-stimulated biofilms had altered ultrastructure, and genes involved in glycogen production and transporter function were up regulated in response to antibiotic exposure. Down-regulated genes were linked to multiple metabolic processes but not those involved in stress response. Antibiotic-stimulated biofilm bacteria were more resistant to a lethal dose (10µg/mL) of cefuroxime. Our results suggest that beta-lactam antibiotic exposure may act as a signaling molecule that promotes transformation into the biofilm phenotype. Loss of viable bacteria, increase in biofilm biomass and decreased protein production coupled with a concomitant up-regulation of genes involved with glycogen production might result in a biofilm of sessile, metabolically inactive bacteria sustained by stored glycogen. These biofilms may protect surviving bacteria from subsequent antibiotic challenges, and act as a reservoir of viable bacteria once antibiotic exposure has ended.