Browsing by Author "Adams, Hannah M."
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Item A Type I Restriction-Modification System Associated with Enterococcus Faecium Subspecies Separation(Amer Soc Microbiology, 2019-01-09) Huo, Wenwen; Adams, Hannah M.; Trejo, Cristian; Badia, Rohit; Palmer, Kelli L.; 0000-0002-7343-9271 (Palmer, KL); Huo, Wenwen; Adams, Hannah M.; Trejo, Cristian; Badia, Rohit; Palmer, Kelli L.The gastrointestinal colonizer Enterococcus faecium is a leading cause of hospital-acquired infections. Multidrug-resistant (MDR) E. faecium isolates are particularly concerning for infection treatment. Previous comparative genomic studies revealed that subspecies referred to as clade A and clade B exist within E. faecium. MDR E. faecium isolates belong to clade A, while clade B consists of drug-susceptible fecal commensal E. faecium isolates. Isolates from clade A are further grouped into two subclades, clades A1 and A2. In general, clade A1 isolates are hospital-epidemic isolates, whereas clade A2 isolates are isolates from animals and sporadic human infections. Such phylogenetic separation indicates that reduced gene exchange occurs between the clades. We hypothesize that endogenous barriers to gene exchange exist between E. faecium clades. Restriction-modification (R-M) systems are such barriers in other microbes. We utilized a bioinformatics analysis coupled with second-generation and third-generation deep-sequencing platforms to characterize the methylomes of two representative E. faecium strains, one from clade A1 and one from clade B. We identified a type I R-M system that is clade A1 specific, is active for DNA methylation, and significantly reduces the transformability of clade A1 E. faecium. Based on our results, we conclude that R-M systems act as barriers to horizontal gene exchange in E. faecium and propose that R-M systems contribute to E. faecium subspecies separation. IMPORTANCE Enterococcus faecium is a leading cause of hospital-acquired infections around the world. Rising antibiotic resistance in certain E. faecium lineages leaves fewer treatment options. The overarching aim of this work was to determine whether restriction-modification (R-M) systems contribute to the structure of the E. faecium species, wherein hospital-epidemic and non-hospital-epidemic isolates have distinct evolutionary histories and highly resolved clade structures. R-M provides bacteria with a type of innate immunity to horizontal gene transfer (HGT). We identified a type I R-M system that is enriched in the hospital-epidemic clade and determined that it is active for DNA modification activity and significantly impacts HGT. Overall, this work is important because it provides a mechanism for the observed clade structure of E. faecium as well as a mechanism for facilitated gene exchange among hospital-epidemic E. faecium isolates.Item Mutations Associated with Reduced Surotomycin Susceptibility in Clostridium Difficile and Enterococcus Species(American Society for Microbiology, 2015-05-04) Adams, Hannah M.; Li, X.; Mascio, C.; Chesnel, Laurent; Palmer, Kelli L.; Adams, Hannah M.; Li, X. Chesnel, Laurent; Palmer, Kelli L.Clostridium difficile infection (CDI) is an urgent public health concern causing considerable clinical and economic burdens. CDI can be treated with antibiotics, but recurrence of the disease following successful treatment of the initial episode often occurs. Surotomycin is a rapidly bactericidal cyclic lipopeptide antibiotic that is in clinical trials for CDI treatment and that has demonstrated superiority over vancomycin in preventing CDI relapse. Surotomycin is a structural analogue of the membrane-active antibiotic daptomycin. Previously, we utilized in vitro serial passage experiments to derive C. difficile strains with reduced surotomycin susceptibilities. The parent strains used included ATCC 700057 and clinical isolates from the restriction endonu-clease analysis (REA) groups BI and K. Serial passage experiments were also performed with vancomycin-resistant and vancomycin-susceptible Enterococcus faecium and Enterococcus faecalis. The goal of this study is to identify mutations associated with reduced surotomycin susceptibility in C. difficile and enterococci. Illumina sequence data generated for the parent strains and serial passage isolates were compared. We identified nonsynonymous mutations in genes coding for cardiolipin synthase in C. difficile ATCC 700057, enoyl-(acyl carrier protein) reductase II (FabK) and cell division protein FtsH2 in C. difficile REA type BI, and a PadR family transcriptional regulator in C. difficile REA type K. Among the 4 enterococcal strain pairs, 20 mutations were identified, and those mutations overlap those associated with daptomycin resistance. These data give insight into the mechanism of action of surotomycin against C. difficile, possible mechanisms for resistance emergence during clinical use, and the potential impacts of surotomycin therapy on intestinal enterococci.Item Streptococcus mitis and S. oralis Lack a Requirement for CdsA, the Enzyme Required for Synthesis of Major Membrane Phospholipids in Bacteria(Amer Soc Microbiology, 2018-06-01) Adams, Hannah M.; Joyce, Luke R.; Guan, Ziqiang; Akins, Ronda L.; Palmer, Kelli L.; 0000-0002-7343-9271 (Palmer, KL); Adams, Hannah M.; Joyce, Luke R.; Akins, Ronda L.; Palmer, Kelli L.Synthesis and integrity of the cytoplasmic membrane are fundamental to cellular life. Experimental evolution studies have hinted at unique physiology in the Gram-positive bacteria Streptococcus mitis and S. oralis. These organisms commonly cause bacteremia and infectious endocarditis (IE) but are rarely investigated in mechanistic studies of physiology and evolution. Unlike in other Gram-positive pathogens, high-level (MIC ≥ 256 mu g/ml) daptomycin resistance rapidly emerges in S. mitis and S. oralis after a single drug exposure. In this study, we found that inactivating mutations in cdsA are associated with high-level daptomycin resistance in S. mitis and S. oralis IE isolates. This is surprising given that cdsA is an essential gene for life in commonly studied model organisms. CdsA is the enzyme responsible for the synthesis of CDP-diacylglycerol, a key intermediate for the biosynthesis of all major phospholipids in prokaryotes and most anionic phospholipids in eukaryotes. Lipidomic analysis by liquid chromatography-mass spectrometry (LC-MS) showed that daptomycin-resistant strains have an accumulation of phosphatidic acid and completely lack phosphatidylglycerol and cardiolipin, two major anionic phospholipids in wild-type strains, confirming the loss of function of CdsA in the daptomycin-resistant strains. To our knowledge, these daptomycin-resistant streptococci represent the first model organisms whose viability is CdsA independent. The distinct membrane compositions resulting from the inactivation of cdsA not only provide novel insights into the mechanisms of daptomycin resistance but also offer unique opportunities to study the physiological functions of major anionic phospholipids in bacteria.