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Whole-Transcriptome Analysis of Verocytotoxigenic Escherichia coli O157:H7 (Sakai) Suggests Plant-Species-Specific Metabolic Responses on Exposure to Spinach and Lettuce Extracts.

Crozier L, Hedley PE, Morris J, Wagstaff C, Andrews SC, Toth I, Jackson RW, Holden NJ - Front Microbiol (2016)

Bottom Line: Plant extracts were used to reduce heterogeneity inherent in plant-microbe interactions and remove the effect of plant immunity.Induction of stress-response genes reflected the apparent physiological status of the bacterial genes in each extract, as a result of glutamate-dependent acid resistance, nutrient stress, or translational stalling.A large proportion of differentially regulated genes are uncharacterized (annotated as hypothetical), which could indicate yet to be described functional roles associated with plant interaction for E. coli O157:H7 Sakai.

View Article: PubMed Central - PubMed

Affiliation: Cell and Molecular Sciences, The James Hutton Institute Dundee, UK.

ABSTRACT
Verocytotoxigenic Escherichia coli (VTEC) can contaminate crop plants, potentially using them as secondary hosts, which can lead to food-borne infection. Currently, little is known about the influence of the specific plant species on the success of bacterial colonization. As such, we compared the ability of the VTEC strain, E. coli O157:H7 'Sakai,' to colonize the roots and leaves of four leafy vegetables: spinach (Spinacia oleracea), lettuce (Lactuca sativa), vining green pea (Pisum sativum), and prickly lettuce (Lactuca serriola), a wild relative of domesticated lettuce. Also, to determine the drivers of the initial response on interaction with plant tissue, the whole transcriptome of E. coli O157:H7 Sakai was analyzed following exposure to plant extracts of varying complexity (spinach leaf lysates or root exudates, and leaf cell wall polysaccharides from spinach or lettuce). Plant extracts were used to reduce heterogeneity inherent in plant-microbe interactions and remove the effect of plant immunity. This dual approach provided information on the initial adaptive response of E. coli O157:H7 Sakai to the plant environment together with the influence of the living plant during bacterial establishment and colonization. Results showed that both the plant tissue type and the plant species strongly influence the short-term (1 h) transcriptional response to extracts as well as longer-term (10 days) plant colonization or persistence. We show that propagation temperature (37 vs. 18°C) has a major impact on the expression profile and therefore pre-adaptation of bacteria to a plant-relevant temperature is necessary to avoid misleading temperature-dependent wholescale gene-expression changes in response to plant material. For each of the plant extracts tested, the largest group of (annotated) differentially regulated genes were associated with metabolism. However, large-scale differences in the metabolic and biosynthetic pathways between treatment types indicate specificity in substrate utilization. Induction of stress-response genes reflected the apparent physiological status of the bacterial genes in each extract, as a result of glutamate-dependent acid resistance, nutrient stress, or translational stalling. A large proportion of differentially regulated genes are uncharacterized (annotated as hypothetical), which could indicate yet to be described functional roles associated with plant interaction for E. coli O157:H7 Sakai.

No MeSH data available.


Related in: MedlinePlus

Glycolysis superpathway gene expression profiles. Expression data for E. coli O157:H7 (Sakai) in response to different plant extracts was overlaid onto the metabolic pathway in EcoCyc (Keseler et al., 2013) to generate a color scale of expression from orange for induction to blue for repression and white for no change < ± twofold. Expression is provided for relevant genes in the pathways that were changed in at least one of the four plant conditions. Gene names are in italics and placed adjacent or close to their relevant substrates. The data for all four conditions are arranged in a grid, ordered as indicated in the Key: LL, leaf lysates and RE, root exudates for spinach; Spin_PS, spinach CWPSs; Lett_PS, lettuce CWPSs.
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Figure 4: Glycolysis superpathway gene expression profiles. Expression data for E. coli O157:H7 (Sakai) in response to different plant extracts was overlaid onto the metabolic pathway in EcoCyc (Keseler et al., 2013) to generate a color scale of expression from orange for induction to blue for repression and white for no change < ± twofold. Expression is provided for relevant genes in the pathways that were changed in at least one of the four plant conditions. Gene names are in italics and placed adjacent or close to their relevant substrates. The data for all four conditions are arranged in a grid, ordered as indicated in the Key: LL, leaf lysates and RE, root exudates for spinach; Spin_PS, spinach CWPSs; Lett_PS, lettuce CWPSs.

Mentions: As indicated above, metabolism encompasses the largest number of differentially expressed genes for all of the extracts tested, although there were major differences between extracts. Components of glycolysis and the Krebs cycle that were induced on exposure to lettuce CWPS included enzymes required for the conversion of oxoglutarate to succinyl-CoA (sucAB, 17- and 20-fold), succinate (sucCD, 8- and 29-fold) and fumarate (sdhABCD, 11–38-fold), and for malate oxidation (mqo, sixfold; Figure 4). This was coupled with a 20-fold induction of the gene encoding the DctA symporter, required for aerobic uptake of C4-dicarboxylates such as succinate (Davies et al., 1999; Supplementary Figure S2). The main gene associated with central metabolism that was induced on exposure to root exudates was acetyl-CoA synthetase (acs, fivefold). The gene (pdhR) encoding the pyruvate-dehydrogenase complex regulator (PdhR, an autoregulatory repressor responding to pyruvate) was induced on exposure to both spinach leaf lysate (fourfold) and lettuce CWPS (ninefold), as were the three genes in the PdhR-controlled aceEF-lpd operon (Supplementary Table S1). However, induction of lpdA was higher than that of aceEF in lettuce CWPS (23- cf. 4- to 6-fold) which reflects lpdA expression from an independent promoter and the involvement of lipoamide dehydrogenase (E3) component in both the pyruvate dehydrogenase and 2-oxoglutarate dehydrogenase multienzyme complexes (Cunningham et al., 1998). In contrast, pdhR was 15-fold repressed in root exudates. These findings indicate low cellular pyruvate levels upon exposure to root exudates, suggestive of low carbon source availability (see below).


Whole-Transcriptome Analysis of Verocytotoxigenic Escherichia coli O157:H7 (Sakai) Suggests Plant-Species-Specific Metabolic Responses on Exposure to Spinach and Lettuce Extracts.

Crozier L, Hedley PE, Morris J, Wagstaff C, Andrews SC, Toth I, Jackson RW, Holden NJ - Front Microbiol (2016)

Glycolysis superpathway gene expression profiles. Expression data for E. coli O157:H7 (Sakai) in response to different plant extracts was overlaid onto the metabolic pathway in EcoCyc (Keseler et al., 2013) to generate a color scale of expression from orange for induction to blue for repression and white for no change < ± twofold. Expression is provided for relevant genes in the pathways that were changed in at least one of the four plant conditions. Gene names are in italics and placed adjacent or close to their relevant substrates. The data for all four conditions are arranged in a grid, ordered as indicated in the Key: LL, leaf lysates and RE, root exudates for spinach; Spin_PS, spinach CWPSs; Lett_PS, lettuce CWPSs.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4940412&req=5

Figure 4: Glycolysis superpathway gene expression profiles. Expression data for E. coli O157:H7 (Sakai) in response to different plant extracts was overlaid onto the metabolic pathway in EcoCyc (Keseler et al., 2013) to generate a color scale of expression from orange for induction to blue for repression and white for no change < ± twofold. Expression is provided for relevant genes in the pathways that were changed in at least one of the four plant conditions. Gene names are in italics and placed adjacent or close to their relevant substrates. The data for all four conditions are arranged in a grid, ordered as indicated in the Key: LL, leaf lysates and RE, root exudates for spinach; Spin_PS, spinach CWPSs; Lett_PS, lettuce CWPSs.
Mentions: As indicated above, metabolism encompasses the largest number of differentially expressed genes for all of the extracts tested, although there were major differences between extracts. Components of glycolysis and the Krebs cycle that were induced on exposure to lettuce CWPS included enzymes required for the conversion of oxoglutarate to succinyl-CoA (sucAB, 17- and 20-fold), succinate (sucCD, 8- and 29-fold) and fumarate (sdhABCD, 11–38-fold), and for malate oxidation (mqo, sixfold; Figure 4). This was coupled with a 20-fold induction of the gene encoding the DctA symporter, required for aerobic uptake of C4-dicarboxylates such as succinate (Davies et al., 1999; Supplementary Figure S2). The main gene associated with central metabolism that was induced on exposure to root exudates was acetyl-CoA synthetase (acs, fivefold). The gene (pdhR) encoding the pyruvate-dehydrogenase complex regulator (PdhR, an autoregulatory repressor responding to pyruvate) was induced on exposure to both spinach leaf lysate (fourfold) and lettuce CWPS (ninefold), as were the three genes in the PdhR-controlled aceEF-lpd operon (Supplementary Table S1). However, induction of lpdA was higher than that of aceEF in lettuce CWPS (23- cf. 4- to 6-fold) which reflects lpdA expression from an independent promoter and the involvement of lipoamide dehydrogenase (E3) component in both the pyruvate dehydrogenase and 2-oxoglutarate dehydrogenase multienzyme complexes (Cunningham et al., 1998). In contrast, pdhR was 15-fold repressed in root exudates. These findings indicate low cellular pyruvate levels upon exposure to root exudates, suggestive of low carbon source availability (see below).

Bottom Line: Plant extracts were used to reduce heterogeneity inherent in plant-microbe interactions and remove the effect of plant immunity.Induction of stress-response genes reflected the apparent physiological status of the bacterial genes in each extract, as a result of glutamate-dependent acid resistance, nutrient stress, or translational stalling.A large proportion of differentially regulated genes are uncharacterized (annotated as hypothetical), which could indicate yet to be described functional roles associated with plant interaction for E. coli O157:H7 Sakai.

View Article: PubMed Central - PubMed

Affiliation: Cell and Molecular Sciences, The James Hutton Institute Dundee, UK.

ABSTRACT
Verocytotoxigenic Escherichia coli (VTEC) can contaminate crop plants, potentially using them as secondary hosts, which can lead to food-borne infection. Currently, little is known about the influence of the specific plant species on the success of bacterial colonization. As such, we compared the ability of the VTEC strain, E. coli O157:H7 'Sakai,' to colonize the roots and leaves of four leafy vegetables: spinach (Spinacia oleracea), lettuce (Lactuca sativa), vining green pea (Pisum sativum), and prickly lettuce (Lactuca serriola), a wild relative of domesticated lettuce. Also, to determine the drivers of the initial response on interaction with plant tissue, the whole transcriptome of E. coli O157:H7 Sakai was analyzed following exposure to plant extracts of varying complexity (spinach leaf lysates or root exudates, and leaf cell wall polysaccharides from spinach or lettuce). Plant extracts were used to reduce heterogeneity inherent in plant-microbe interactions and remove the effect of plant immunity. This dual approach provided information on the initial adaptive response of E. coli O157:H7 Sakai to the plant environment together with the influence of the living plant during bacterial establishment and colonization. Results showed that both the plant tissue type and the plant species strongly influence the short-term (1 h) transcriptional response to extracts as well as longer-term (10 days) plant colonization or persistence. We show that propagation temperature (37 vs. 18°C) has a major impact on the expression profile and therefore pre-adaptation of bacteria to a plant-relevant temperature is necessary to avoid misleading temperature-dependent wholescale gene-expression changes in response to plant material. For each of the plant extracts tested, the largest group of (annotated) differentially regulated genes were associated with metabolism. However, large-scale differences in the metabolic and biosynthetic pathways between treatment types indicate specificity in substrate utilization. Induction of stress-response genes reflected the apparent physiological status of the bacterial genes in each extract, as a result of glutamate-dependent acid resistance, nutrient stress, or translational stalling. A large proportion of differentially regulated genes are uncharacterized (annotated as hypothetical), which could indicate yet to be described functional roles associated with plant interaction for E. coli O157:H7 Sakai.

No MeSH data available.


Related in: MedlinePlus