Limits...
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

Escherichia coli O157:H7 (Sakai) on roots. The numbers of E. coli O157:H7 (Sakai) recovered from the roots of roundhead lettuce (L. sativa; A), spinach (S. oleracea; B), wild prickly lettuce (L. serriola; C), and vining green pea (P. sativum; D), following inoculation of the compost (triangles) or hydroponics liquid media (circles). P. sativum was not grown under hydroponics conditions. Bacteria were not recovered from the hydroponic media-only control (squares). The average and the standard error of the mean for each of the time points is expressed as the number recovered per gram of fresh tissue (Log10). Data is generated from nine replicate samples and was analyzed by one-way ANOVA with the Tukey multiple comparison test.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4940412&req=5

Figure 8: Escherichia coli O157:H7 (Sakai) on roots. The numbers of E. coli O157:H7 (Sakai) recovered from the roots of roundhead lettuce (L. sativa; A), spinach (S. oleracea; B), wild prickly lettuce (L. serriola; C), and vining green pea (P. sativum; D), following inoculation of the compost (triangles) or hydroponics liquid media (circles). P. sativum was not grown under hydroponics conditions. Bacteria were not recovered from the hydroponic media-only control (squares). The average and the standard error of the mean for each of the time points is expressed as the number recovered per gram of fresh tissue (Log10). Data is generated from nine replicate samples and was analyzed by one-way ANOVA with the Tukey multiple comparison test.

Mentions: Colonization of roots was compared for plants grown in compost or hydroponics medium, to partly account for any potential effect from native compost-associated microbiota. Inoculation of compost-grown plants was achieved by partially immersing the plant pots in a bacterial suspension at 7.3 log10 CFU/ml, which resulted in the recovery of between 2.0 and 4.0 log10 CFU/g E. coli O157:H7 (Sakai) from the roots at the initial time point (1 h post inoculation; Figure 8). Despite some variation between plant species, the bacterial populations remained relatively stable and did not decrease as observed on leaves. E. coli O157:H7 (Sakai) recovered from P. sativum roots decreased marginally at day 2 but increased again by d10. Highest recovery at d10 occurred from spinach, followed by L. serriola, L. sativa and pea (3.4, 3.35, 2.76, and 2.24 log10 CFU, respectively). For the colonization potential of E. coli O157:H7 (Sakai) on roots of plants grown under hydroponics (liquid) conditions, the inoculum (7 log10 CFU/ml) was introduced into the medium adjacent to the roots. The number of E. coli O157:H7 (Sakai) recovered at the first time point was ∼2 orders of magnitude higher than that for compost-grown plants. The levels of E. coli O157:H7 (Sakai) recovered after 10 days were at least as high, or higher, than the initial inoculum (Figure 2). Greater recovery of bacteria occurred from L. serriola and spinach than L. sativa at d10 (7.04, 6.36, and 5.88 log10 CFU, respectively). No proliferation of E. coli O157:H7 (Sakai) occurred in the hydroponics medium in the absence of plant roots, with the population at 4.46 log10 CFU at d10, significantly different to E. coli O157:H7 (Sakai) from the three plants (p < 0.001). In our hands, it was not possible to remove surface-associated fungi from P. sativum seeds sufficiently well to allow its growth under aseptic hydroponics conditions; therefore, this combination was not tested. These experiments demonstrate that E. coli O157:H7 (Sakai) was able to either stabilize or increase its population on leaf and root, but that there were plant, tissue and growth media specific differences that affected colonization potential.


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)

Escherichia coli O157:H7 (Sakai) on roots. The numbers of E. coli O157:H7 (Sakai) recovered from the roots of roundhead lettuce (L. sativa; A), spinach (S. oleracea; B), wild prickly lettuce (L. serriola; C), and vining green pea (P. sativum; D), following inoculation of the compost (triangles) or hydroponics liquid media (circles). P. sativum was not grown under hydroponics conditions. Bacteria were not recovered from the hydroponic media-only control (squares). The average and the standard error of the mean for each of the time points is expressed as the number recovered per gram of fresh tissue (Log10). Data is generated from nine replicate samples and was analyzed by one-way ANOVA with the Tukey multiple comparison test.
© Copyright Policy
Related In: Results  -  Collection

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

Figure 8: Escherichia coli O157:H7 (Sakai) on roots. The numbers of E. coli O157:H7 (Sakai) recovered from the roots of roundhead lettuce (L. sativa; A), spinach (S. oleracea; B), wild prickly lettuce (L. serriola; C), and vining green pea (P. sativum; D), following inoculation of the compost (triangles) or hydroponics liquid media (circles). P. sativum was not grown under hydroponics conditions. Bacteria were not recovered from the hydroponic media-only control (squares). The average and the standard error of the mean for each of the time points is expressed as the number recovered per gram of fresh tissue (Log10). Data is generated from nine replicate samples and was analyzed by one-way ANOVA with the Tukey multiple comparison test.
Mentions: Colonization of roots was compared for plants grown in compost or hydroponics medium, to partly account for any potential effect from native compost-associated microbiota. Inoculation of compost-grown plants was achieved by partially immersing the plant pots in a bacterial suspension at 7.3 log10 CFU/ml, which resulted in the recovery of between 2.0 and 4.0 log10 CFU/g E. coli O157:H7 (Sakai) from the roots at the initial time point (1 h post inoculation; Figure 8). Despite some variation between plant species, the bacterial populations remained relatively stable and did not decrease as observed on leaves. E. coli O157:H7 (Sakai) recovered from P. sativum roots decreased marginally at day 2 but increased again by d10. Highest recovery at d10 occurred from spinach, followed by L. serriola, L. sativa and pea (3.4, 3.35, 2.76, and 2.24 log10 CFU, respectively). For the colonization potential of E. coli O157:H7 (Sakai) on roots of plants grown under hydroponics (liquid) conditions, the inoculum (7 log10 CFU/ml) was introduced into the medium adjacent to the roots. The number of E. coli O157:H7 (Sakai) recovered at the first time point was ∼2 orders of magnitude higher than that for compost-grown plants. The levels of E. coli O157:H7 (Sakai) recovered after 10 days were at least as high, or higher, than the initial inoculum (Figure 2). Greater recovery of bacteria occurred from L. serriola and spinach than L. sativa at d10 (7.04, 6.36, and 5.88 log10 CFU, respectively). No proliferation of E. coli O157:H7 (Sakai) occurred in the hydroponics medium in the absence of plant roots, with the population at 4.46 log10 CFU at d10, significantly different to E. coli O157:H7 (Sakai) from the three plants (p < 0.001). In our hands, it was not possible to remove surface-associated fungi from P. sativum seeds sufficiently well to allow its growth under aseptic hydroponics conditions; therefore, this combination was not tested. These experiments demonstrate that E. coli O157:H7 (Sakai) was able to either stabilize or increase its population on leaf and root, but that there were plant, tissue and growth media specific differences that affected colonization potential.

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