Metabolomic and lipidomic characterization of Oxalobacter formigenes strains HC1 and OxWR by UHPLC-HRMS
Diseases of oxalate, such as nephrolithiasis and primary hyperoxaluria, affect a significant portion of the US population and have limited treatment options. Oxalobacter formigenes, an obligate oxalotrophic bacterium in the mammalian intestine, has generated great interest as a potential probiotic or therapeutic treatment for oxalate-related conditions due to its ability to degrade both exogenous (dietary) and endogenous (metabolic) oxalate, lowering the risk of hyperoxaluria/hyperoxalemia. Although all oxalotrophs degrade dietary oxalate, Oxalobacter formigenes is the only species shown to initiate intestinal oxalate secretion to draw upon endogenous, circulating oxalate for consumption. Evidence suggests that Oxalobacter regulates oxalate transport proteins in the intestinal epithelium using an unidentified secreted bioactive compound, but the mechanism of this function remains elusive. It is essential to gain an understanding of the biochemical relationship between Oxalobacter and the host intestinal epithelium for this microbe to progress as a potential remedy for oxalate diseases. This investigation includes the first profiling of the metabolome and lipidome of Oxalobacter formigenes, specifically the human strain HC1 and rat strain OxWR, the only two strains shown thus far to initiate net intestinal oxalate secretion across native gut epithelia. This study was performed using untargeted and targeted metabolomics and lipidomics methodologies utilizing ultra-high-performance liquid chromatography-mass spectrometry. We report our findings that the metabolic profiles of these strains, although largely conserved, show significant differences in their expression of many compounds. Several strain-specific features were also detected. Discussed are trends in the whole metabolic profile as well as in individual features, both identified and unidentified.
KeywordsMetabolomics Lipidomics Oxalobacter formigenes Mass spectrometry LC-MS Oxalate Nephrolithiasis Hyperoxaluria
The authors would like to acknowledge Dr. Cory A. Leonard, Department of Pathology, Immunology and Laboratory Medicine, University of Florida, for her assistance with sample generation for this experiment, including media preparation and cell culture, harvest, and lysis. Also to be acknowledged are Dr. Jeremy P. Koelmel, Department of Pathology, Immunology and Laboratory Medicine, University of Florida, and Vanessa Y. Rubio, Department of Chemistry, University of Florida, for their contributions with figure generation for the lipidomics analysis and graphical abstract, respectively.
This work was funded by the National Institutes of Health grant 2R01DK088892-05A1.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflicts of interest.
- 9.Dawson KA, Allison MJ, Hartman PA. Isolation and some characteristics of anaerobic oxalate-degrading bacteria from the rumen. Appl Environ Microbiol. 1980;40(4):833–9.Google Scholar
- 12.Hatch M, Gjymishka A, Salido EC, Allison MJ, Freel RW. Enteric oxalate elimination is induced and oxalate is normalized in a mouse model of primary hyperoxaluria following intestinal colonization with Oxalobacter. Am J Physiol Gastrointest Liver Physiol. 2011;300(3):G461–9. https://doi.org/10.1152/ajpgi.00434.2010.CrossRefGoogle Scholar
- 13.Liu H, Garrett TJ, Tayyari F, Gu L. Profiling the metabolome changes caused by cranberry procyanidins in plasma of female rats using (1) H NMR and UHPLC-Q-Orbitrap-HRMS global metabolomics approaches. Mol Nutr Food Res. 2015;59(11):2107–18. https://doi.org/10.1002/mnfr.201500236.CrossRefGoogle Scholar
- 14.Folch J, Lees M, Sloane Stanley GH. A simple method for the isolation and purification of total lipides from animal tissues. J Biol Chem. 1957;226(1):497–509.Google Scholar
- 21.Koelmel JP, Kroeger NM, Ulmer CZ, Bowden JA, Patterson RE, Cochran JA, et al. LipidMatch: an automated workflow for rule-based lipid identification using untargeted high-resolution tandem mass spectrometry data. BMC Bioinformatics. 2017;18(1):331. https://doi.org/10.1186/s12859-017-1744-3.CrossRefGoogle Scholar
- 24.Holm S. A simple sequentially rejective multiple test procedure. Scand J Stat. 1979;6(2):65–70.Google Scholar
- 25.Hatch M, Allison MJ, Yu F, Farmerie W. Genome sequence of Oxalobacter formigenes strain HC-1. Genome Announc. 2017;5(27). https://doi.org/10.1128/genomeA.00533-17.
- 26.Hatch M, Allison MJ, Yu F, Farmerie W. Genome sequence of Oxalobacter formigenes strain OXCC13. Genome Announc. 2017;5(28). https://doi.org/10.1128/genomeA.00534-17.
- 32.Allen F, Pon A, Wilson M, Greiner R, Wishart D. CFM-ID: a web server for annotation, spectrum prediction and metabolite identification from tandem mass spectra. Nucleic Acids Research. 2014;42:(W1)W94–W99.Google Scholar
- 38.Jackowski S, Murphy CM, Cronan JE, Rock CO. Acetoacetyl-acyl carrier protein synthase. A target for the antibiotic thiolactomycin. J Biol Chem. 1989;264(13):7624–9.Google Scholar
- 39.Khandekar SS, Gentry DR, Van Aller GS, Warren P, Xiang H, Silverman C, et al. Identification, substrate specificity, and inhibition of the Streptococcus pneumoniae beta-ketoacyl-acyl carrier protein synthase III (FabH). J Biol Chem. 2001;276(32):30024–30. https://doi.org/10.1074/jbc.M101769200.CrossRefGoogle Scholar
- 40.Reynolds CM, Kalb SR, Cotter RJ, Raetz CR. A phosphoethanolamine transferase specific for the outer 3-deoxy-D-manno-octulosonic acid residue of Escherichia coli lipopolysaccharide. Identification of the eptB gene and Ca2+ hypersensitivity of an eptB deletion mutant. J Biol Chem. 2005;280(22):21202–11. https://doi.org/10.1074/jbc.M500964200.CrossRefGoogle Scholar
- 46.Goldfine H. Bacterial-membranes and lipid packing theory. J Lipid Res. 1984;25(13):1501–7.Google Scholar