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Molecular and chemical dialogues in bacteria-protozoa interactions.

Song C, Mazzola M, Cheng X, Oetjen J, Alexandrov T, Dorrestein P, Watrous J, van der Voort M, Raaijmakers JM - Sci Rep (2015)

Bottom Line: Lipopeptide (LP) biosynthesis was induced in Pseudomonas upon protozoan grazing and LP accumulation transitioned from homogeneous distributions across bacterial colonies to site-specific accumulation at the bacteria-protist interface.Also putrescine biosynthesis was upregulated in P. fluorescens upon predation.This multifaceted study provides new insights in common and strain-specific responses in bacteria-protozoa interactions, including responses that contribute to bacterial survival in highly competitive soil and rhizosphere environments.

View Article: PubMed Central - PubMed

Affiliation: 1] Laboratory of Phytopathology, Wageningen University, 6708 PB Wageningen, the Netherlands [2] Microbial Ecology Department, Netherlands Institute of Ecology (NIOO-KNAW), 6708 PB Wageningen, the Netherlands.

ABSTRACT
Protozoan predation of bacteria can significantly affect soil microbial community composition and ecosystem functioning. Bacteria possess diverse defense strategies to resist or evade protozoan predation. For soil-dwelling Pseudomonas species, several secondary metabolites were proposed to provide protection against different protozoan genera. By combining whole-genome transcriptome analyses with (live) imaging mass spectrometry (IMS), we observed multiple changes in the molecular and chemical dialogues between Pseudomonas fluorescens and the protist Naegleria americana. Lipopeptide (LP) biosynthesis was induced in Pseudomonas upon protozoan grazing and LP accumulation transitioned from homogeneous distributions across bacterial colonies to site-specific accumulation at the bacteria-protist interface. Also putrescine biosynthesis was upregulated in P. fluorescens upon predation. We demonstrated that putrescine induces protozoan trophozoite encystment and adversely affects cyst viability. This multifaceted study provides new insights in common and strain-specific responses in bacteria-protozoa interactions, including responses that contribute to bacterial survival in highly competitive soil and rhizosphere environments.

No MeSH data available.


Related in: MedlinePlus

(A) MALDI imaging mass spectrometry (IMS) shows production of massetolide A and its derivatives during the P. fluorescens SS101-N. americana interaction. a.u. = arbitrary units. (B) Box plots depicting the production of massetolide A and its derivatives (m/z 1142, 1163, 1165 and 1179) in P. fluorescens SS101 alone (SS101), N. americana alone (Protozoa), P. fluorescens SS101-N. americana interaction (SS101_Protozoa) and massA mutant alone (ΔmassA). (B) The box plots represent the median intensity in arbitrary units after TIC normalization (horizontal line), the upper and lower quartiles (box layout, spectra in which the intensities are within a range of 25%–75% of the data), the upper and lower quantiles (dashed lines, spectra in which the intensities are within a range of 0%–99%) as well as the outliers (spectra with intensities greater than 99% of the data). The green box-line indicates N. americana alone; the red box-line indicates P. fluorescens SS101-N. americana interaction; the yellow box-line indicates P. fluorescens SS101 alone.
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f4: (A) MALDI imaging mass spectrometry (IMS) shows production of massetolide A and its derivatives during the P. fluorescens SS101-N. americana interaction. a.u. = arbitrary units. (B) Box plots depicting the production of massetolide A and its derivatives (m/z 1142, 1163, 1165 and 1179) in P. fluorescens SS101 alone (SS101), N. americana alone (Protozoa), P. fluorescens SS101-N. americana interaction (SS101_Protozoa) and massA mutant alone (ΔmassA). (B) The box plots represent the median intensity in arbitrary units after TIC normalization (horizontal line), the upper and lower quartiles (box layout, spectra in which the intensities are within a range of 25%–75% of the data), the upper and lower quantiles (dashed lines, spectra in which the intensities are within a range of 0%–99%) as well as the outliers (spectra with intensities greater than 99% of the data). The green box-line indicates N. americana alone; the red box-line indicates P. fluorescens SS101-N. americana interaction; the yellow box-line indicates P. fluorescens SS101 alone.

Mentions: Spatial segmentation analysis of the MALDI IMS data revealed four specific classes of metabolites indicated with different colours (light blue, green, orange and dark red), that were co-localized in the Pseudomonas-protozoa interaction zone (Fig. 3B; Table S2). There were 6 and 14 ions found to be co-localized within the light blue and green cluster, respectively, with correlation values greater than 0.5 (Fig. 3B; Table S2) including 8 ions with predicted masses ranging from 1136 m/z to 1201 m/z (Fig. 4A). These ions were not detected in the massetolide-deficient ∆massA mutant or N. americana alone (Fig. 4A). The absence of these ions in the ∆massA mutant suggests that they are massetolide derivatives. Box plots further confirmed that the intensities of these ions were higher in wild type strain SS101, and in the SS101-protozoa interaction than in the samples with N. americana or ∆massA alone (Fig. 4B). Masses of massetolide A and its derivatives also clustered together in the MS/MS network (Fig. 3C; Table S4). Tandem MS of the ion with a mass of 1163 m/z indicated a peptide sequence of leucine, serine, leucine, serine and isoleucine. These amino acids are identical to the C-terminal peptide sequence of massetolide A (Fig. 3C). Based on our previous study, the mass of massetolide A is 1140 and the masses of its derivatives range from 1112–1158 m/z50. The larger ion masses detected here are most likely due to a sodium (molecular weight: 22.989) gain during ionization. Although the intensities of the massetolides were not different between SS101 and SS101-protozoa interaction (Fig. 4B), we observed a striking difference in spatial distribution of massetolide A. In absence of the protozoa, the lipopeptide was more homogeneously distributed in the SS101 colony, whereas in presence of protozoa it localized predominantly in/around the bacterial cells at the interaction zone (Fig. 4A). This result was reinforced by the absence of massetolides in the interaction between the massA mutant and the protozoan predator (Figure S2). To our knowledge, this is the first report of the real time visualization and spatial distribution of LPs during bacteria-protozoa interactions.


Molecular and chemical dialogues in bacteria-protozoa interactions.

Song C, Mazzola M, Cheng X, Oetjen J, Alexandrov T, Dorrestein P, Watrous J, van der Voort M, Raaijmakers JM - Sci Rep (2015)

(A) MALDI imaging mass spectrometry (IMS) shows production of massetolide A and its derivatives during the P. fluorescens SS101-N. americana interaction. a.u. = arbitrary units. (B) Box plots depicting the production of massetolide A and its derivatives (m/z 1142, 1163, 1165 and 1179) in P. fluorescens SS101 alone (SS101), N. americana alone (Protozoa), P. fluorescens SS101-N. americana interaction (SS101_Protozoa) and massA mutant alone (ΔmassA). (B) The box plots represent the median intensity in arbitrary units after TIC normalization (horizontal line), the upper and lower quartiles (box layout, spectra in which the intensities are within a range of 25%–75% of the data), the upper and lower quantiles (dashed lines, spectra in which the intensities are within a range of 0%–99%) as well as the outliers (spectra with intensities greater than 99% of the data). The green box-line indicates N. americana alone; the red box-line indicates P. fluorescens SS101-N. americana interaction; the yellow box-line indicates P. fluorescens SS101 alone.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4542665&req=5

f4: (A) MALDI imaging mass spectrometry (IMS) shows production of massetolide A and its derivatives during the P. fluorescens SS101-N. americana interaction. a.u. = arbitrary units. (B) Box plots depicting the production of massetolide A and its derivatives (m/z 1142, 1163, 1165 and 1179) in P. fluorescens SS101 alone (SS101), N. americana alone (Protozoa), P. fluorescens SS101-N. americana interaction (SS101_Protozoa) and massA mutant alone (ΔmassA). (B) The box plots represent the median intensity in arbitrary units after TIC normalization (horizontal line), the upper and lower quartiles (box layout, spectra in which the intensities are within a range of 25%–75% of the data), the upper and lower quantiles (dashed lines, spectra in which the intensities are within a range of 0%–99%) as well as the outliers (spectra with intensities greater than 99% of the data). The green box-line indicates N. americana alone; the red box-line indicates P. fluorescens SS101-N. americana interaction; the yellow box-line indicates P. fluorescens SS101 alone.
Mentions: Spatial segmentation analysis of the MALDI IMS data revealed four specific classes of metabolites indicated with different colours (light blue, green, orange and dark red), that were co-localized in the Pseudomonas-protozoa interaction zone (Fig. 3B; Table S2). There were 6 and 14 ions found to be co-localized within the light blue and green cluster, respectively, with correlation values greater than 0.5 (Fig. 3B; Table S2) including 8 ions with predicted masses ranging from 1136 m/z to 1201 m/z (Fig. 4A). These ions were not detected in the massetolide-deficient ∆massA mutant or N. americana alone (Fig. 4A). The absence of these ions in the ∆massA mutant suggests that they are massetolide derivatives. Box plots further confirmed that the intensities of these ions were higher in wild type strain SS101, and in the SS101-protozoa interaction than in the samples with N. americana or ∆massA alone (Fig. 4B). Masses of massetolide A and its derivatives also clustered together in the MS/MS network (Fig. 3C; Table S4). Tandem MS of the ion with a mass of 1163 m/z indicated a peptide sequence of leucine, serine, leucine, serine and isoleucine. These amino acids are identical to the C-terminal peptide sequence of massetolide A (Fig. 3C). Based on our previous study, the mass of massetolide A is 1140 and the masses of its derivatives range from 1112–1158 m/z50. The larger ion masses detected here are most likely due to a sodium (molecular weight: 22.989) gain during ionization. Although the intensities of the massetolides were not different between SS101 and SS101-protozoa interaction (Fig. 4B), we observed a striking difference in spatial distribution of massetolide A. In absence of the protozoa, the lipopeptide was more homogeneously distributed in the SS101 colony, whereas in presence of protozoa it localized predominantly in/around the bacterial cells at the interaction zone (Fig. 4A). This result was reinforced by the absence of massetolides in the interaction between the massA mutant and the protozoan predator (Figure S2). To our knowledge, this is the first report of the real time visualization and spatial distribution of LPs during bacteria-protozoa interactions.

Bottom Line: Lipopeptide (LP) biosynthesis was induced in Pseudomonas upon protozoan grazing and LP accumulation transitioned from homogeneous distributions across bacterial colonies to site-specific accumulation at the bacteria-protist interface.Also putrescine biosynthesis was upregulated in P. fluorescens upon predation.This multifaceted study provides new insights in common and strain-specific responses in bacteria-protozoa interactions, including responses that contribute to bacterial survival in highly competitive soil and rhizosphere environments.

View Article: PubMed Central - PubMed

Affiliation: 1] Laboratory of Phytopathology, Wageningen University, 6708 PB Wageningen, the Netherlands [2] Microbial Ecology Department, Netherlands Institute of Ecology (NIOO-KNAW), 6708 PB Wageningen, the Netherlands.

ABSTRACT
Protozoan predation of bacteria can significantly affect soil microbial community composition and ecosystem functioning. Bacteria possess diverse defense strategies to resist or evade protozoan predation. For soil-dwelling Pseudomonas species, several secondary metabolites were proposed to provide protection against different protozoan genera. By combining whole-genome transcriptome analyses with (live) imaging mass spectrometry (IMS), we observed multiple changes in the molecular and chemical dialogues between Pseudomonas fluorescens and the protist Naegleria americana. Lipopeptide (LP) biosynthesis was induced in Pseudomonas upon protozoan grazing and LP accumulation transitioned from homogeneous distributions across bacterial colonies to site-specific accumulation at the bacteria-protist interface. Also putrescine biosynthesis was upregulated in P. fluorescens upon predation. We demonstrated that putrescine induces protozoan trophozoite encystment and adversely affects cyst viability. This multifaceted study provides new insights in common and strain-specific responses in bacteria-protozoa interactions, including responses that contribute to bacterial survival in highly competitive soil and rhizosphere environments.

No MeSH data available.


Related in: MedlinePlus