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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

Gene expression overview. Heatmap of Escherichia coli O157:H7 (Sakai) total gene expression changes in response to different temperature and plant extract treatments. Changes in gene expression for E. coli O157:H7 (Sakai) grown for 1 h at 18°C are compared to cultures grown similarly at 37°C (37_MM), or at 18°C containing spinach (S. olercera) extracts of leaf lysates (Spin_LL) or root exudates (Spin_RE; A). Changes in gene expression for exposure to 1 h exposure to medium at 18°C containing polysaccharide extracts from spinach (S. olercera; Spin_PS) or lettuce (Lactuca sativa; Lett_PS) are compared to a baseline for E. coli O157:H7 (Sakai) in medium containing a no-plant control extract (‘Media’; B). Significant changes in expression of at least twofold are shown for induced (red) or repressed (green) genes.
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Figure 1: Gene expression overview. Heatmap of Escherichia coli O157:H7 (Sakai) total gene expression changes in response to different temperature and plant extract treatments. Changes in gene expression for E. coli O157:H7 (Sakai) grown for 1 h at 18°C are compared to cultures grown similarly at 37°C (37_MM), or at 18°C containing spinach (S. olercera) extracts of leaf lysates (Spin_LL) or root exudates (Spin_RE; A). Changes in gene expression for exposure to 1 h exposure to medium at 18°C containing polysaccharide extracts from spinach (S. olercera; Spin_PS) or lettuce (Lactuca sativa; Lett_PS) are compared to a baseline for E. coli O157:H7 (Sakai) in medium containing a no-plant control extract (‘Media’; B). Significant changes in expression of at least twofold are shown for induced (red) or repressed (green) genes.

Mentions: As expected, gene expression of E. coli O157:H7 (Sakai) grown for 1 h at 18°C was markedly different from that of the culture grown at 37°C (Figure 1). A total of 1,127 genes were differentially expressed in response to incubation temperature, representing 20.6% of E. coli O157:H7 Sakai ORFs. Of these, 500 genes were induced and 627 genes (9.16 and 11.48% of Sakai ORFs) were downregulated (Supplementary Table S1). Notable changes in expression of specific genes at 18°C (cf. 37°C) included repression of a subset of genes in the locus of enterocycte effacement (LEE). These included ler (130-fold repression; which encodes the master regulator of the lee genes), several type III secretion (T3SS) genes (ECs4583, escC, escJ, escS, and espF: repressed by 15-, 10-, 12-, 20-, and 10-fold respectively; Supplementary Table S1). The control of ler expression by low temperature is likely caused by H-NS silencing, which is known to suppress A/E lesion formation below 37°C (Umanski et al., 2002). Motility genes were also repressed, particularly in the flg and fli loci (e.g., flgBCDE, 26–59-fold repressed; fliE, 26-fold repressed. Three hypothetical genes in an apparent operon of unknown function (ECs2623-2625) were amongst those most strongly repressed (∼200-fold), as were a series of prophage CP-933T genes (coxT, Z2971-4; 46–276-fold) possibly in response to QseA control (Kendall et al., 2010). The major class of genes subject to induction at 18°C were those involved in various aspects of stress resistance: acid resistance (e.g., ECs2098, gadABCE; 38–121-fold induced), heavy-metal resistance (e.g., cusBX; 56–81-fold induced), putrescine metabolism (e.g., ygjG, ECs3955; 52–65-fold induced), multidrug efflux (e.g., sugE; 33-fold induced) and osmotic stress (proVW; ∼25-fold). In addition, a cluster of genes (ECs1653-1655; 14–28-fold) of unknown function was strongly induced as were several genes involved in biofilm formation (Z2229, ECs2085, bdm, c_1914; 51–59-fold; Supplementary Table S1). In summary, the expression data suggest that growth at ambient rather than body temperature causes reduces motility and increases sessile behavior, reduced ability to colonize the mammalian gut and suppresses some prophage, but raises ability to resist a range of environmental stresses (a possible adaptation to slower growth at lower temperature). Such temperature-dependent changes would be expected to confound interpretation of expression data obtained in previous studies on bacterial plant colonization where a temperature change was included along with plant exposure – a complication that was avoided within the research reported 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)

Gene expression overview. Heatmap of Escherichia coli O157:H7 (Sakai) total gene expression changes in response to different temperature and plant extract treatments. Changes in gene expression for E. coli O157:H7 (Sakai) grown for 1 h at 18°C are compared to cultures grown similarly at 37°C (37_MM), or at 18°C containing spinach (S. olercera) extracts of leaf lysates (Spin_LL) or root exudates (Spin_RE; A). Changes in gene expression for exposure to 1 h exposure to medium at 18°C containing polysaccharide extracts from spinach (S. olercera; Spin_PS) or lettuce (Lactuca sativa; Lett_PS) are compared to a baseline for E. coli O157:H7 (Sakai) in medium containing a no-plant control extract (‘Media’; B). Significant changes in expression of at least twofold are shown for induced (red) or repressed (green) genes.
© Copyright Policy
Related In: Results  -  Collection

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

Figure 1: Gene expression overview. Heatmap of Escherichia coli O157:H7 (Sakai) total gene expression changes in response to different temperature and plant extract treatments. Changes in gene expression for E. coli O157:H7 (Sakai) grown for 1 h at 18°C are compared to cultures grown similarly at 37°C (37_MM), or at 18°C containing spinach (S. olercera) extracts of leaf lysates (Spin_LL) or root exudates (Spin_RE; A). Changes in gene expression for exposure to 1 h exposure to medium at 18°C containing polysaccharide extracts from spinach (S. olercera; Spin_PS) or lettuce (Lactuca sativa; Lett_PS) are compared to a baseline for E. coli O157:H7 (Sakai) in medium containing a no-plant control extract (‘Media’; B). Significant changes in expression of at least twofold are shown for induced (red) or repressed (green) genes.
Mentions: As expected, gene expression of E. coli O157:H7 (Sakai) grown for 1 h at 18°C was markedly different from that of the culture grown at 37°C (Figure 1). A total of 1,127 genes were differentially expressed in response to incubation temperature, representing 20.6% of E. coli O157:H7 Sakai ORFs. Of these, 500 genes were induced and 627 genes (9.16 and 11.48% of Sakai ORFs) were downregulated (Supplementary Table S1). Notable changes in expression of specific genes at 18°C (cf. 37°C) included repression of a subset of genes in the locus of enterocycte effacement (LEE). These included ler (130-fold repression; which encodes the master regulator of the lee genes), several type III secretion (T3SS) genes (ECs4583, escC, escJ, escS, and espF: repressed by 15-, 10-, 12-, 20-, and 10-fold respectively; Supplementary Table S1). The control of ler expression by low temperature is likely caused by H-NS silencing, which is known to suppress A/E lesion formation below 37°C (Umanski et al., 2002). Motility genes were also repressed, particularly in the flg and fli loci (e.g., flgBCDE, 26–59-fold repressed; fliE, 26-fold repressed. Three hypothetical genes in an apparent operon of unknown function (ECs2623-2625) were amongst those most strongly repressed (∼200-fold), as were a series of prophage CP-933T genes (coxT, Z2971-4; 46–276-fold) possibly in response to QseA control (Kendall et al., 2010). The major class of genes subject to induction at 18°C were those involved in various aspects of stress resistance: acid resistance (e.g., ECs2098, gadABCE; 38–121-fold induced), heavy-metal resistance (e.g., cusBX; 56–81-fold induced), putrescine metabolism (e.g., ygjG, ECs3955; 52–65-fold induced), multidrug efflux (e.g., sugE; 33-fold induced) and osmotic stress (proVW; ∼25-fold). In addition, a cluster of genes (ECs1653-1655; 14–28-fold) of unknown function was strongly induced as were several genes involved in biofilm formation (Z2229, ECs2085, bdm, c_1914; 51–59-fold; Supplementary Table S1). In summary, the expression data suggest that growth at ambient rather than body temperature causes reduces motility and increases sessile behavior, reduced ability to colonize the mammalian gut and suppresses some prophage, but raises ability to resist a range of environmental stresses (a possible adaptation to slower growth at lower temperature). Such temperature-dependent changes would be expected to confound interpretation of expression data obtained in previous studies on bacterial plant colonization where a temperature change was included along with plant exposure – a complication that was avoided within the research reported 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