NMR‐based metabonomics analysis of mouse urine and fecal extracts following oral treatment with the broad‐spectrum antibiotic enrofloxacin (Baytril)

The human gastrointestinal tract is home to hundreds of species of bacteria and the balance between beneficial and pathogenic bacteria plays a critical role in human health and disease. The human infant, however, is born with a sterile gut and the complex gastrointestinal host/bacterial ecosystem is only established after birth by rapid bacterial colonization. Composition of newborn gut flora depends on several factors including type of birth (Ceasarian or natural), manner of early feeding (breast milk or formula), and exposure to local, physical environment. Imbalance in normal, healthy gut flora contributes to several adult human diseases including inflammatory bowel (ulcerative colitis and Crohn's disease) and Clostridium difficile associated disease, and early childhood diseases such as necrotizing enterocolitis. As a first step towards characterization of the role of gut bacteria in human health and disease, we conducted an 850 MHz 1H nuclear magnetic resonance spectroscopy study to monitor changes in metabolic profiles of urine and fecal extracts of 15 mice following gut sterilization by the broad‐spectrum antibiotic enrofloxacin (also known as Baytril). Ten metabolites changed in urine following enrofloxacin treatment including decreased acetate due to loss of microbial catabolism of sugars and polysaccharides, decreased trimethylamine‐N‐oxide due to loss of microbial catabolism of choline, and increased creatine and creatinine due to loss of microbial enzyme degradation. Eight metabolites changed in fecal extracts of mice treated with enrofloxacin including depletion of amino acids produced by microbial proteases, reduction in metabolites generated by lactate‐utilizing bacteria, and increased urea caused by loss of microbial ureases. Copyright © 2009 John Wiley & Sons, Ltd.

[1]  Ian D. Wilson,et al.  HIGH RESOLUTION PROTON MAGNETIC RESONANCE SPECTROSCOPY OF BIOLOGICAL FLUIDS , 1989 .

[2]  Dawn B. Beaulieu,et al.  Impact of Clostridium difficile on inflammatory bowel disease. , 2007, Clinical gastroenterology and hepatology : the official clinical practice journal of the American Gastroenterological Association.

[3]  T. Cataldi,et al.  Quantitative determination of taurine in real samples by high-performance anion-exchange chromatography with integrated pulsed amperometric detection. , 2004, Talanta.

[4]  F. Bäckhed,et al.  Host-Bacterial Mutualism in the Human Intestine , 2005, Science.

[5]  John C. Lindon,et al.  Pattern recognition methods and applications in biomedical magnetic resonance , 2001 .

[6]  W. D. de Vos,et al.  Interactomics in the human intestine: Lactobacilli and Bifidobacteria make a difference. , 2008, Journal of clinical gastroenterology.

[7]  Julian L Griffin,et al.  Metabonomics: NMR spectroscopy and pattern recognition analysis of body fluids and tissues for characterisation of xenobiotic toxicity and disease diagnosis. , 2003, Current opinion in chemical biology.

[8]  E. Purdom,et al.  Diversity of the Human Intestinal Microbial Flora , 2005, Science.

[9]  W. Oh,et al.  Oral Probiotics Reduce the Incidence and Severity of Necrotizing Enterocolitis in Very Low Birth Weight Infants , 2005, Pediatrics.

[10]  J. Neu Perinatal and neonatal manipulation of the intestinal microbiome: a note of caution. , 2007, Nutrition reviews.

[11]  R. Goodacre Metabolomics of a superorganism. , 2007, The Journal of nutrition.

[12]  Michel Verleysen,et al.  Comparison of some chemometric tools for metabonomics biomarker identification , 2008 .

[13]  W. Whitman,et al.  Prokaryotes: the unseen majority. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[14]  Kieran Clarke,et al.  A combined 1H-NMR spectroscopy- and mass spectrometry-based metabolomic study of the PPAR-alpha null mutant mouse defines profound systemic changes in metabolism linked to the metabolic syndrome. , 2006, Physiological genomics.

[15]  O. Kremp,et al.  Does the intestinal bifidobacterial colonisation affect bacterial translocation? , 2008, Anaerobe.

[16]  R. Rastall Nature, Nurture, and the Case for Nutrition Bacteria in the Gut: Friends and Foes and How to Alter the Balance 1 , 2004 .

[17]  P. Karplus,et al.  Identification and Characterization of Bacterial Cysteine Dioxygenases: a New Route of Cysteine Degradation for Eubacteria , 2006, Journal of bacteriology.

[18]  I. Wilson,et al.  Rapid and noninvasive metabonomic characterization of inflammatory bowel disease. , 2007, Journal of proteome research.

[19]  A. Kimmoun,et al.  Acute decompensation of isovaleric acidemia induced by Graves’ disease , 2008, Intensive Care Medicine.

[20]  K. Karlinger,et al.  The epidemiology and the pathogenesis of inflammatory bowel disease. , 2000, European journal of radiology.

[21]  M. Butel,et al.  Clostridial pathogenicity in experimental necrotising enterocolitis in gnotobiotic quails and protective role of bifidobacteria. , 1998, Journal of medical microbiology.

[22]  D. Hooper,et al.  The fluoroquinolones: structures, mechanisms of action and resistance, and spectra of activity in vitro , 1985, Antimicrobial Agents and Chemotherapy.

[23]  John C. Lindon,et al.  Metabonomics: metabolic processes studied by NMR spectroscopy of biofluids , 2000 .

[24]  E. Furrie,et al.  A molecular revolution in the study of intestinal microflora , 2006, Gut.

[25]  J. Hampe,et al.  Reduction in diversity of the colonic mucosa associated bacterial microflora in patients with active inflammatory bowel disease , 2004, Gut.

[26]  Elaine Holmes,et al.  Metabonomic and microbiological analysis of the dynamic effect of vancomycin-induced gut microbiota modification in the mouse. , 2008, Journal of proteome research.

[27]  M. McCarthy,et al.  Metabolic profiling reveals a contribution of gut microbiota to fatty liver phenotype in insulin-resistant mice , 2006, Proceedings of the National Academy of Sciences.

[28]  I. Wilson,et al.  Understanding 'Global' Systems Biology: Metabonomics and the Continuum of Metabolism , 2003, Nature Reviews Drug Discovery.

[29]  F. Guarner,et al.  Gut flora in health and disease , 2003, The Lancet.

[30]  R. Mackie,et al.  Developmental microbial ecology of the neonatal gastrointestinal tract. , 1999, The American journal of clinical nutrition.

[31]  R. Kauppinen,et al.  Tumour metabolomics in animal models of human cancer. , 2007, Journal of proteome research.

[32]  D. N. Salter The influence of gut micro-organisms on utilization of dietary protein , 1973, Proceedings of the Nutrition Society.

[33]  Daniel B. DiGiulio,et al.  Development of the Human Infant Intestinal Microbiota , 2007, PLoS biology.

[34]  J. Cnota,et al.  Lactobacillus Sepsis Associated With Probiotic Therapy , 2005, Pediatrics.

[35]  H. Flint,et al.  Lactate-Utilizing Bacteria, Isolated from Human Feces, That Produce Butyrate as a Major Fermentation Product , 2004, Applied and Environmental Microbiology.

[36]  S. Himeno,et al.  Effect of pH on 1H-NMR spectroscopy of mouse urine. , 2007, Biological & pharmaceutical bulletin.

[37]  C. Tancrède Role of human microflora in health and disease , 1992, European Journal of Clinical Microbiology and Infectious Diseases.

[38]  J. Nicholson,et al.  Experimental and analytical variation in human urine in 1H NMR spectroscopy-based metabolic phenotyping studies. , 2007, Analytical chemistry.

[39]  L. Morelli Postnatal development of intestinal microflora as influenced by infant nutrition. , 2008, The Journal of nutrition.

[40]  D. Savage Microbial ecology of the gastrointestinal tract. , 1977, Annual review of microbiology.

[41]  Anthony F. P. Nash,et al.  A 1H NMR-based metabonomic study of urine and plasma samples obtained from healthy human subjects. , 2003, Journal of pharmaceutical and biomedical analysis.

[42]  Jeremy K Nicholson,et al.  NMR spectroscopic-based metabonomic studies of urinary metabolite variation in acclimatizing germ-free rats. , 2003, Chemical research in toxicology.

[43]  S. Nicolson,et al.  The effect of different oral antibiotics on the gastrointestinal microflora of a wild rodent (Aethomys namaquensis). , 2004, Comparative biochemistry and physiology. Part A, Molecular & integrative physiology.

[44]  I. Adlerberth,et al.  Establishment of the gut microbiota in Western infants , 2009, Acta paediatrica.

[45]  I. Wilson,et al.  Metabonomics, dietary influences and cultural differences: a 1H NMR-based study of urine samples obtained from healthy British and Swedish subjects. , 2004, Journal of pharmaceutical and biomedical analysis.

[46]  Elaine Holmes,et al.  Systemic multicompartmental effects of the gut microbiome on mouse metabolic phenotypes , 2008, Molecular systems biology.

[47]  W. Fetter,et al.  The intestinal bacterial colonisation in preterm infants: a review of the literature. , 2006, Clinical nutrition.

[48]  M. Reily,et al.  Metabonomic assessment of vasculitis in rats , 2007, Cardiovascular Toxicology.

[49]  M. Fairchok,et al.  Lactobacillus Sepsis Associated With Probiotic Therapy , 2005, Pediatrics.

[50]  F. Guarner The intestinal flora in inflammatory bowel disease: normal or abnormal? , 2005, Current opinion in gastroenterology.

[51]  J. Utzinger,et al.  Metabolic Profiling of an Echinostoma caproni Infection in the Mouse for Biomarker Discovery , 2008, PLoS neglected tropical diseases.

[52]  D. Kyle,et al.  Determination of 15N isotopic enrichment and concentrations of allantoin and uric acid in urine by gas chromatography/mass spectrometry. , 1998, Journal of mass spectrometry : JMS.

[53]  Ting Wang,et al.  The gut microbiota as an environmental factor that regulates fat storage. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[54]  E. Myers,et al.  Basic local alignment search tool. , 1990, Journal of molecular biology.

[55]  M. Tanner,et al.  Metabonomic investigations in mice infected with Schistosoma mansoni: an approach for biomarker identification. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[56]  T. Fan Metabolite profiling by one- and two-dimensional NMR analysis of complex mixtures , 1996 .

[57]  J. Gordon,et al.  How host-microbial interactions shape the nutrient environment of the mammalian intestine. , 2002, Annual review of nutrition.

[58]  W. D. de Vos,et al.  Molecular Monitoring of Succession of Bacterial Communities in Human Neonates , 2002, Applied and Environmental Microbiology.

[59]  M. Wyss,et al.  Creatine and creatinine metabolism. , 2000, Physiological reviews.

[60]  R. Knight,et al.  Evolution of Mammals and Their Gut Microbes , 2008, Science.

[61]  P. Raibaud,et al.  Hydrolysis of urea in the gastrointestinal tract of "monoxenic" rats: effect of immunization with strains of ureolytic bacteria , 1976, Infection and immunity.

[62]  R. Berg,et al.  The indigenous gastrointestinal microflora. , 1996, Trends in microbiology.

[63]  A. Torres,et al.  Host-microbe communication within the GI tract. , 2008, Advances in experimental medicine and biology.

[64]  D. Sinderen,et al.  Human gut microbiota and bifidobacteria: from composition to functionality , 2008, Antonie van Leeuwenhoek.

[65]  H. Gaskins,et al.  Impact of the intestinal microbiota on the development of mucosal defense. , 2008, Clinical infectious diseases : an official publication of the Infectious Diseases Society of America.

[66]  J. Neu,et al.  Alive and dead Lactobacillus rhamnosus GG decrease tumor necrosis factor-alpha-induced interleukin-8 production in Caco-2 cells. , 2005, The Journal of nutrition.

[67]  P. A. van den Brandt,et al.  Factors Influencing the Composition of the Intestinal Microbiota in Early Infancy , 2006, Pediatrics.

[68]  S. Zeisel,et al.  Conversion of dietary choline to trimethylamine and dimethylamine in rats: dose-response relationship. , 1989, The Journal of nutrition.

[69]  Hung-Chih Lin,et al.  Possible Confounding Factors in an Oral Probiotics Trial: Breast Milk: In Reply , 2005, Pediatrics.