Hanne Vorwerk

Xcenda GmbH, Hannover, Germany

Campylobacter jejuni is one of the leading infectious causes of food-borne illness around the world. Its ability to persistently colonize the intestinal tract of a broad range of hosts, including food-producing animals, is central to its epidemiology since most infections are due to the consumption of contaminated food products. Using a highly satu...

Amino acid utilization by C. jejuni 81–176 determined by exometabolome analysis. The uptake of amino acids by C. jejuni 81–176 was examined by measuring their concentrations in the culture supernatants within 24 h of cultivation relative to their original concentration prior to bacterial inoculation, which was considered to be 100%. Shown are the m...

In vivo fitness of C. jejuni insertion mutants determined by INSeq analysis. Relative abundance of C. jejuni insertion mutants in the inoculum and the mouse cecum samples after infection for 4 (A), 7 (B) and 21 (C) days. In (A) and (B), each point represents the average abundance of read numbers of a single gene obtained from 13 or 5 mice, respecti...

Role of amino acid biosynthesis of C. jejuni 81–176 in mouse colonization. Illustrated is the impact of amino acid biosynthesis pathways in C. jejuni mouse intestinal colonization as determined by INSeq analysis. Numbers indicate the log2 value of fold change (intestine/inoculum) in the number of insertions in the indicated genes and are derived fr...

Role of methionine and S-adenosyl methionine (SAM) biosynthesis in C. jejuni 81–176 in vivo growth. Illustrated is the impact of methionine and SAM biosynthesis in C. jejuni mouse intestinal colonization as determined by INSeq analysis. Numbers indicate the log2 value of fold change (intestine/inoculum) in the number of insertions in the indicated...

Impact of the non-oxidative pentose phosphate pathway (PPP) on the growth of C. jejuni 81–176. Shown is the impact of the inactivation of genes encoding components of the PPP pathway in C. jejuni mouse intestinal colonization as determined by INSeq analyses. Numbers indicate the log2 value of fold change (intestine/inoculum) in the number of insert...

Core metabolic reactions of C. jejuni 81–176 that generate carbon dioxide. Selected metabolic reactions in C. jejuni that release carbon dioxide (CO2). Numbers indicate the log2 value of fold change (intestine/inoculum) in the number of insertions in the indicated genes and are derived from the raw data in S3 Table. Values below -6.2 indicate mutat...

INSeq raw data for each gene in mouse colonization experiments, which were normalized by median nomarlization method. (XLSX)

Summary of potassium channel and transporters in Epsilon-proteobacteria. (XLSX)

Contribution of a C. jejuni motility-associated gene cluster to mouse colonization. (A)Blue and red bars indicate the normalized read number of each insertion site within the different ORFs in the input and output pool, respectively. Motility assays (B) and growth curves (see S12 Table) (C) for wild-type C. jejuni 81–176 and the ΔCJJ81176_0479, ΔCJ...

Impact of the purine and pyrimidine biosynthesis pathways on the in vivo growth of C. jejuni 81–176. Shown is the impact of the inactivation of genes encoding components of the purine and pyrimidine biosynthesis pathways in C. jejuni mouse intestinal colonization as determined by INSeq analyses. Numbers indicate the log2 value of fold change (intes...

Analysis of potential polar effects of transposon insertions. (XLSX)

Strains and plasmids used in this study. (XLSX)

Raw data for Figs 6E, 7E, S2B, S7C and S10. (XLSX)

Growth properties of serine-biosynthesis defective C. jejuni mutants. Wild type C. jejuni 81–176 (WT) and the indicated isogenic mutant strains (serA: D-3-phosphoglycerate dehydrogenase; sdaA: L-serine ammonia-lyase/dehydratase) were grown in defined medium (DAAM) in the presence or absence of serine as indicated. Lactate or glutamate are provided...

13C-incorporation and isotopologue profiles of protein-derived amino acids in C. jejuni 81–176 after incubation with [5-13C1]-Glu in DMEM medium. Black columns on the left y axis represent percentage of 13C-excess (mol %) into the respective protein-derived amino acids. The colored columns on the right y axis depict the percentages of labeled isoto...

Role of the “acetate switch” in mouse colonization. Six mice were inoculated with an equal number of wild-type C. jejuni 81–176 and the ΔackA isogenic mutant strain via oral gavage. Competitive indices (CI) were calculated as the ratio of the CFU of the ΔackA mutant over wild type recovered from the ceca of infected mice (see S14 Table). (TIF)

List of genes that show similar mutant phenotype according to the Venn diagram depicted in Fig 1B and 1C. (XLSX)

Summary of INSeq data for Tlp and chemotaxis proteins in C. jejuni 81–176. (XLSX)

Raw data for Figs 2, 5inset, 6C, 6D, S1, S12 and S13. (XLSX)

Raw data for Figs 7D, S4 and S14. (XLSX)

Labeling of polar metabolites in C. jejuni 81–176 upon cultivation with [3-13C1]Ser. Shown are the 13C-incorporation (13C-excess) and the relative isotopologue distributions into polar metabolites isolated from the cytoplasm of C. jejuni after incubation with [3-13C1]Ser. Illustrated are the means ± SD of 6 measurements with the colored boxes indic...

Amino acid uptake ability of C. jejuni 81–176. (A) Scheme presenting the predicted amino acid uptake capacity of C. jejuni 81–176 according to the INSeq analysis. Viable mutants with transposon insertions in the indicated amino acid biosynthesis pathways have been identified in the screen suggesting the import of respective amino acids. No transpos...

Colony forming units of commensal bacteria in mice feces before and after antibiotic treatment. Shown is the intestinal bacterial load after antibiotic treatment of animals immediately prior to infection with C. jejuni. Collected feces samples from each mouse were dissolved and diluted serially with PBS buffer, then plated on blood agar in duplicat...

Log2 (output/input) ratio distribution of all insertions during mouse colonization. Histogram depicting the number of genes (y axis) that exhibited the indicated log2 [fold change (output/input)] change (x axis) in the numbers of transposon insertions recovered from infected mice relative to the number of transposon insertions in the original inocu...

Amino acid composition of all predicted proteins encoded by C. jejuni 81–176. The frequency of all amino acids in all predicted proteins are shown and branched-chain amino acids are highlighted in orange. Calculations were carried out using BacMap (http://wishart.biology.ualberta.ca/BacMap/cgi/getGraphs.cgi?accession=NC_008787&ref=index_2.html). (T...

Schematic overview of the 13C-flux into protein-derived amino acids after catabolism of [5-13C1]-Glu in C. jejuni 81–176. The positions of the 13C-label of the amino acid isotopologues are indicated by the colored dots. Due to the stereoisomerism of the TCA-cycle intermediates succinate and fumarate, it is not possible to distinguish between the C1...

INSeq raw data for each C. jejuni gene after growth under different in-vitro growth condition. (XLSX)

INSeq raw data for each gene in mouse colonization experiments. (XLSX)

Growth-promoting substrates utilized by Campylobacter jejuni during colonization of gastrointestinal tract. (DOCX)

Composition of a modified defined DMEM medium for Campylobacter jejuni 81–176 used in the indicated studies. (XLSX)

Raw data for S3 Fig. (XLSX)

Supplementary methods. (DOCX)

Thermophilic Campylobacter species colonize the intestine of agricultural and domestic animals commensally, but cause severe gastroenteritis in humans. In contrast to other enteropathogenic bacteria, Campylobacter have been considered to be non-glycolytic, a metabolic property originally used for their taxonomic classification. Contrary to this dog...

The non-glycolytic food-borne pathogen Campylobacter jejuni successfully colonizes the intestine of various hosts in spite of its restricted metabolic properties. While several amino acids are known to be used by C. jejuni as energy sources, none of these have been found to be essential for growth. Here we demonstrated through phenotype microarray...