Temperature Influences the Production and Transport of Saxitoxin and the Expression of sxt Genes in the Cyanobacterium Aphanizomenon gracile
Abstract
:1. Introduction
2. Results
2.1. Ecophysiology of Aphanizomenon Gracile
2.2. Effects of Temperature on Saxitoxin Production and Release
2.3. Effects of Temperature on the Expression of sxt Genes
2.3.1. Dynamics of STX-Biosynthesis Gene sxtA
2.3.2. Dynamics of STX-Transporter Genes sxtM and sxtPer
2.4. Effects of Nitrogen Depletion at 23 °C
3. Discussion
4. Conclusions
5. Materials and Methods
5.1. Culture Conditions
5.2. Growth Dynamics
5.3. Alterations in Membrane Integrity and Membrane Potential by Flow Cytometry
5.4. Analysis of the Expression of Genes sxtA, sxtM, and sxtPer by qPCR
5.4.1. Total RNA Extraction and cDNA Synthesis
5.4.2. Real Time qPCR
5.5. Quantification of Intracellular and Extracellular Saxitoxins
5.6. Statistical Analysis
Supplementary Materials
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Temperature (°C) | Growth Rate (day−1) | Chl a Content (mg g−1 DW) |
---|---|---|
12 | 0.10 ± 0.02 a | 8.6 ± 1.6 a |
23 | 0.17 ± 0.03 b | 13.6 ± 2.4 b |
30 | 0.07 ± 0.01 a | 9.9 ± 3.1 a |
Temperature (°C) | Total STX Content (µg mg−1 DW) | Total STX Content (µg mg−1 Chl a) | Extracellular STX (% of Total STX) |
---|---|---|---|
12 | 0.08 ± 0.03 a | 8.7 ± 2.2 a | 16.9 ± 10.2 a |
23 | 0.20 ± 0.08 b | 14.0 ± 4.3 a | 11.8 ± 6.1 a |
30 | 0.25 ± 0.07 b | 28.6 ± 13.7 b | 53.8 ± 15.5 b |
Culture Medium | Ecophysiology | Toxin Production/Release | Relative Expression | ||||
---|---|---|---|---|---|---|---|
Growth Rate (day−1) | Chl a Content (mg g−1 DW) | Total STX Content (µg mg−1 DW) | Extracellular STX (%) | sxtA | sxtM | sxtPer | |
BG11 | 0.17 ± 0.03 | 13.6 ± 2.4 | 0.20 ± 0.08 | 11.8 ± 6.1 | 1.0 ± 0.01 | 1.0 ± 0.02 | 1.0 ± 0.01 |
BG110 | 0.05 ± 0.01 * | 7.6 ± 1.9 * | 0.15 ± 0.04 | 51.2 ± 20.2 * | 1.5 ± 0.5 | 1.2 ± 0.5 | 1.1 ± 0.3 |
Organism | Factor | Effect on Extracellular STX Release | Effect on the Expression of STX Transporters | Reference | |
---|---|---|---|---|---|
sxtM | sxtPer | ||||
Aphanizomenon gracile | Temperature (12°C and 30°C vs. 23 °C) | 4-fold higher at 30 °C than at 23 °C | Upregulation (2.9–4.3X) at 30 °C Upregulation (3.6–3.8X) at 12 °C | Upregulation (2.1X) at 30 °C | This study |
Nitrate (absence—BG110- vs. presence—BG11-) | 4-fold higher without nitrate (BG110) | NS | NS | This study | |
Anabaena circinalis | Ph (9 vs. 7) | Higher at pH 9 (exact value not provided) | Downregulation at pH 9 (59X) vs. pH 7 | NP | [18] |
Na+ (10 mM vs. 1.3 mM 1) | Higher at 10 mM (exact value not provided) | Downregulation at 10 mM (2.7X) vs. 1.3 mM | NP | [18] | |
Cylindrospermopsis raciborskii | pH (9 vs. 7) | Higher at pH 9 (exact value not provided) | Upregulation at pH = 9 (24X) vs. pH 7 | NP | [18] |
Na+ (10 mM vs. 1.3 mM 1) | Higher at 10 mM (exact value not provided) | Upregulation at 10 mM (2.7X) vs. 1.3 mM | NP | [18] |
Gene | Primer | Sequence (5’–3’) | Amplicon Size (bp) | Slope; R2 | Efficiency (%) 1 | Source |
---|---|---|---|---|---|---|
16S rRNA | q16grF q16grR | GAGAGACTGCCGGTGACAAA TGCCCTTTGTCCGTAGCATT | 106 | −3.24 0.995 | 103 | This study |
sxtA | jrtPKSF jrtPKSR | GGAGTGGATTTCAACACCAGAA GTTTCCCAGACTCGTTTCAGG | 147 | −3.38 0.999 | 98 | [18] |
sxtM | qMgrF qMgrR | GAAGCACGAGTCAGCCTACA CAAAGCACCACCAGCCAAAA | 129 | −3.29 0.998 | 101 | This study |
sxtPer | qPERgrF qPERgrR | CTGGGCGAGACATTTGAGA GCACAGAGACAGGCGAACTA | 116 | −3.37 0.993 | 98 | This study |
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Cirés, S.; Delgado, A.; González-Pleiter, M.; Quesada, A. Temperature Influences the Production and Transport of Saxitoxin and the Expression of sxt Genes in the Cyanobacterium Aphanizomenon gracile. Toxins 2017, 9, 322. https://doi.org/10.3390/toxins9100322
Cirés S, Delgado A, González-Pleiter M, Quesada A. Temperature Influences the Production and Transport of Saxitoxin and the Expression of sxt Genes in the Cyanobacterium Aphanizomenon gracile. Toxins. 2017; 9(10):322. https://doi.org/10.3390/toxins9100322
Chicago/Turabian StyleCirés, Samuel, Adrián Delgado, Miguel González-Pleiter, and Antonio Quesada. 2017. "Temperature Influences the Production and Transport of Saxitoxin and the Expression of sxt Genes in the Cyanobacterium Aphanizomenon gracile" Toxins 9, no. 10: 322. https://doi.org/10.3390/toxins9100322