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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

Escherichia coli O157:H7 (Sakai) on leaves. The numbers of E. coli O157:H7 (Sakai) recovered from the leaves of roundhead lettuce [L. sativa; A), spinach (Spinacia oleracea; B), wild prickly lettuce (Lactuca serriola; C), and vining green pea (Pisum sativum; D], following inoculation of the abaxial (black triangles) or adaxial (black circles) surfaces with a starting inoculum of 2 × 106 CFU (Log10 6.3). The average and the standard error of the mean for six replicates for each of the time points is expressed as the number of CFU 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.
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Figure 7: Escherichia coli O157:H7 (Sakai) on leaves. The numbers of E. coli O157:H7 (Sakai) recovered from the leaves of roundhead lettuce [L. sativa; A), spinach (Spinacia oleracea; B), wild prickly lettuce (Lactuca serriola; C), and vining green pea (Pisum sativum; D], following inoculation of the abaxial (black triangles) or adaxial (black circles) surfaces with a starting inoculum of 2 × 106 CFU (Log10 6.3). The average and the standard error of the mean for six replicates for each of the time points is expressed as the number of CFU 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: An E. coli O157:H7 (Sakai) inoculum of 6.3 log10 CFU was applied to the adaxial (upper) and abaxial (lower) surface of the leaves of four different plant species and the bacteria enumerated over 10 days. There was a decrease in bacterial numbers compared to the starting inoculum for all four species, on both leaf surfaces. However, in each case, a higher average number of E. coli O157:H7 (Sakai) was recovered from the abaxial than adaxial surface after 10 days (Figure 7), although the difference was not significant at the 95% confidence level. The average number of E. coli O157:H7 (Sakai) on both leaf surfaces of both species of lettuce (L. sativa and L. serriola) decreased over the time tested, although the numbers recovered at d10 were significantly different: 1.66/2.84 (adaxial/abaxial) log10 CFU for L. sativa and at the limit of detection (0.15/0.63 log10 CFU, adaxial/abaxial) for L. serriola (p < 0.05), with bacteria only recovered from 22% of the samples for L. serriola for this time point. The number of E. coli O157:H7 (Sakai) on spinach also decreased from the starting inoculum and although higher counts were obtained from the abaxial side of the leaf at d2, by d10 they had reached similar levels, stabilizing at 0.69 adaxial and 1.99 abaxial log10 CFU. Pea was the only plant where the numbers increased between d2 and d10, from 1.05 to 3.08 log10 CFU (abaxial). By d10, significantly higher numbers were recovered found pea than L. serriola (adaxial, p < 0.01; abaxial, p < 0.05).


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 leaves. The numbers of E. coli O157:H7 (Sakai) recovered from the leaves of roundhead lettuce [L. sativa; A), spinach (Spinacia oleracea; B), wild prickly lettuce (Lactuca serriola; C), and vining green pea (Pisum sativum; D], following inoculation of the abaxial (black triangles) or adaxial (black circles) surfaces with a starting inoculum of 2 × 106 CFU (Log10 6.3). The average and the standard error of the mean for six replicates for each of the time points is expressed as the number of CFU 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 7: Escherichia coli O157:H7 (Sakai) on leaves. The numbers of E. coli O157:H7 (Sakai) recovered from the leaves of roundhead lettuce [L. sativa; A), spinach (Spinacia oleracea; B), wild prickly lettuce (Lactuca serriola; C), and vining green pea (Pisum sativum; D], following inoculation of the abaxial (black triangles) or adaxial (black circles) surfaces with a starting inoculum of 2 × 106 CFU (Log10 6.3). The average and the standard error of the mean for six replicates for each of the time points is expressed as the number of CFU 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: An E. coli O157:H7 (Sakai) inoculum of 6.3 log10 CFU was applied to the adaxial (upper) and abaxial (lower) surface of the leaves of four different plant species and the bacteria enumerated over 10 days. There was a decrease in bacterial numbers compared to the starting inoculum for all four species, on both leaf surfaces. However, in each case, a higher average number of E. coli O157:H7 (Sakai) was recovered from the abaxial than adaxial surface after 10 days (Figure 7), although the difference was not significant at the 95% confidence level. The average number of E. coli O157:H7 (Sakai) on both leaf surfaces of both species of lettuce (L. sativa and L. serriola) decreased over the time tested, although the numbers recovered at d10 were significantly different: 1.66/2.84 (adaxial/abaxial) log10 CFU for L. sativa and at the limit of detection (0.15/0.63 log10 CFU, adaxial/abaxial) for L. serriola (p < 0.05), with bacteria only recovered from 22% of the samples for L. serriola for this time point. The number of E. coli O157:H7 (Sakai) on spinach also decreased from the starting inoculum and although higher counts were obtained from the abaxial side of the leaf at d2, by d10 they had reached similar levels, stabilizing at 0.69 adaxial and 1.99 abaxial log10 CFU. Pea was the only plant where the numbers increased between d2 and d10, from 1.05 to 3.08 log10 CFU (abaxial). By d10, significantly higher numbers were recovered found pea than L. serriola (adaxial, p < 0.01; abaxial, p < 0.05).

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