Browsing by Author "Johnson, Christina"

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Dynamic gut microbiome changes to low-iron challenge
(American Society for Microbiology, 2020-11) Coe, Genevieve L.; Pinkham, Nicholas V.; Celis, Arianna I.; Johnson, Christina; DuBois, Jennifer L.; Walk, Seth T.
Iron is an essential micronutrient for life. In mammals, dietary iron is primarily absorbed in the small intestine. Currently, the impacts of dietary iron on the taxonomic structure and function of the gut microbiome and reciprocal effects on the animal host are not well understood. Here, we establish a mouse model of low-iron challenge in which intestinal biomarkers and reduced fecal iron reveal iron stress while serum iron and mouse behavioral markers indicate maintenance of iron homeostasis. We show that the diversity of the gut microbiome in conventional C57BL/6 mice changes dramatically during two-weeks on a low-iron diet. We also show the effects of a low-iron diet on microbiome diversity are long-lasting and not easily recovered when iron is returned to the diet. Finally, after optimizing taxon association methods, we show that some bacteria are unable to fully recover after the low-iron challenge and appear to be extirpated from the gut entirely. In particular, OTUs from the Prevotellaceae and Porphyromonadaceae families and Bacteroidales order are highly sensitive to low-iron conditions, while other seemingly insensitive OTUs recover. These results provide new insights into the iron requirements of gut microbiome members and add to the growing understanding of mammalian iron cycling. IMPORTANCE All cells need iron. Both too much iron and too little lead to diseases and unwanted outcomes. Although the impact of dietary iron on human cells and tissues has been well studied, there is currently a lack of understanding about how different levels of iron influence the abundant and diverse members of the human microbiome. This study develops a well-characterized mouse model for studying low-iron levels and identifies key groups of bacteria that are most affected. We found that the microbiome undergoes large changes when iron is removed from the diet but that many individual bacteria are able to rebound when iron levels are changed by to normal. That said, a select few members, referred to as “iron-sensitive” bacteria seem to be lost. This study begins to identify individual members of the mammalian microbiome most affected by changes in dietary iron levels.
Evaluation of portability and cost of a fluorescent PCR ribotyping protocol for Clostridium difficile epidemiology
(2015-01) Martinson, Jonathan N.V.; Broadaway, Susan C.; Lohman, Egan J.; Johnson, Christina; Alam, M. Jahangir; Khaleduzzaman, Mohammed; Garey, Kevin W.; Schlackman, Jessica; Young, Vincent B.; Santhosh, Kavitha; Rao, Krishna; Lyons, Robert H. Jr; Walk, Seth T.
Clostridium difficile is the most commonly identified pathogen among health care-associated infections in the United States. There is a need for accurate and low-cost typing tools that produce comparable data across studies (i.e., portable data) to help characterize isolates during epidemiologic investigations of C. difficile outbreaks and sporadic cases of disease. The most popular C. difficile-typing technique is PCR ribotyping, and we previously developed methods using fluorescent PCR primers and amplicon sizing on a Sanger-style sequencer to generate fluorescent PCR ribotyping data. This technique has been used to characterize tens of thousands of C. difficile isolates from cases of disease. Here, we present validation of a protocol for the cost-effective generation of fluorescent PCR ribotyping data. A key component of this protocol is the ability to accurately identify PCR ribotypes against an online database (http://walklab.rcg.montana.edu) at no cost. We present results from a blinded multicenter study to address data portability across four different laboratories and three different sequencing centers. Our standardized protocol and centralized database for typing of C. difficile pathogens will increase comparability between studies so that important epidemiologic linkages between cases of disease and patterns of emergence can be rapidly identified.
Pathways of Iron and Sulfur Acquisition, Cofactor Assembly, Destination, and Storage in Diverse Archaeal Methanogens and Alkanotrophs
(2021-08) Johnson, Christina; England, Alexis; Munro-Ehrlich, Mason; Colman, Daniel R.; DuBois, Jennifer L.; Boyd, Eric S.
Archaeal methanogens, methanotrophs, and alkanotrophs have a high demand for iron (Fe) and sulfur (S); however, little is known of how they acquire, traffic, deploy, and store these elements. Here, we examined the distribution of homologs of proteins mediating key steps in Fe/S metabolism in model microorganisms, including iron(II) sensing/uptake (FeoAB), sulfide extraction from cysteine (SufS), and the biosynthesis of iron-sulfur [Fe-S] clusters (SufBCDE), siroheme (Pch2 dehydrogenase), protoheme (AhbABCD), cytochrome c (Cyt c) (CcmCF), and iron storage/detoxification (Bfr, FtrA, and IssA), among 326 publicly available, complete or metagenome-assembled genomes of archaeal methanogens/methanotrophs/alkanotrophs. The results indicate several prevalent but nonuniversal features, including FeoB, SufBC, and the biosynthetic apparatus for the basic tetrapyrrole scaffold, as well as its siroheme (and F430) derivatives. However, several early-diverging genomes lacked SufS and pathways to synthesize and deploy heme. Genomes encoding complete versus incomplete heme biosynthetic pathways exhibited equivalent prevalences of [Fe-S] cluster binding proteins, suggesting an expansion of catalytic capabilities rather than substitution of heme for [Fe-S] in the former group. Several strains with heme binding proteins lacked heme biosynthesis capabilities, while other strains with siroheme biosynthesis capability lacked homologs of known siroheme binding proteins, indicating heme auxotrophy and unknown siroheme biochemistry, respectively. While ferritin proteins involved in ferric oxide storage were widespread, those involved in storing Fe as thioferrate were unevenly distributed. Collectively, the results suggest that differences in the mechanisms of Fe and S acquisition, deployment, and storage have accompanied the diversification of methanogens/methanotrophs/alkanotrophs, possibly in response to differential availability of these elements as these organisms evolved.
Proteomic Analysis of Methanococcus voltae Grown in the Presence of Mineral and Nonmineral Sources of Iron and Sulfur
(American Society for Microbiology, 2022-08) Steward, Katherine F.; Payne, Devon; Kincannon, Will; Johnson, Christina; Lensing, Malachi; Fausset, Hunter; Németh, Brigitta; Shepard, Eric M.; Broderick, William E.; Broderick, Joan B.; Dubois, Jen; Bothner, Brian
Clusters of iron and sulfur are key components of the active sites of enzymes that facilitate microbial conversion of light or electrical energy into chemical bonds. The proteins responsible for transporting iron and sulfur into cells and assembling these elements into metal clusters are not well understood.