In Vitro Propagation of Isoëtes sabatina (Isoetaceae): A Key Conservation Challenge for a Critically Endangered Quillwort
Abstract
:1. Introduction
2. Results
2.1. Spore Germination
2.2. Sporeling Development
2.2.1. Effect of the Substrate on Sporeling Emergence
2.2.2. Effect of the Megaspores Developmental Stage on Sporeling Emergence
2.3. Transplants
3. Discussion
3.1. In Vitro Sporeling Production
3.2. The Conservation Challenges for I. sabatina
4. Materials and Methods
4.1. Study Area
4.2. Plant Material
4.3. Spore Collection Ex Situ
4.4. In vitro Mixed Culture of Mega- and Microspores
- (1)
- Water-agar (1%) medium, as described by [31];
- (2)
4.5. Evaluation of Spore Germination and Sporeling Development
4.6. Calculation of Sporeling Emergence Indices
- -
- Sporeling emergence percentage (SE, in percent), calculated on the number of sown megaspores, and the number of germinated megaspores (SEG, in percent);
- -
- Time for first sporeling emergence (T1, in days);
- -
- Mean daily emergence (MDE, in percent), calculated by the expression: , where Tn is the total number of days required to reach the final sporeling emergence percentage (SE);
- -
- Mean emergence time (MET, in days) indicates the average length of time required for maximum sporelings emergence and was calculated according to the following formula: , where ni is the number of sporelings newly emerged at time ti;
- -
- Mean emergence rate (MER, in day−1) is the average number of sporelings emerged per day, calculated as the reciprocal of MET;
- -
- Coefficient of variation of emergence time (CVt), calculated by the expression , where SDt is the standard deviation of the emergence time and MET is the mean emergence time;
- -
- Emergence Index (EI, number of sporelings per day), calculated by the expression , where ni is the number of sporelings emerged at time ti;
- -
- Peak value (PV, sporelings/day) is the accumulated number of sporelings at the point on the emergence curve at which the rate of emergence starts to decrease. It is calculated as the maximum quotient obtained by dividing successive cumulative emergence values by the respective incubation time;
- -
- Uncertainty of the sporeling emergence process (U, in bit), i.e., the uncertainty associated with the distribution of the relative frequency of emergence (fi), calculated through the formula: , being , where ni is the number of sporelings emerged at time ti. It can assume values spanning from 0 and log2n, with n being the total number of sporelings emerged.
4.7. Transplants
- (1)
- Lake sediment was homogeneously distributed at the bottom of glass containers (20 × 10 × 10 cm), covered with a thin layer of sterilized quartzite sand to avoid the spread of debris according to [18], and then covered with distilled water up to a height of ca. 3 cm;
- (2)
- Quartzite sand was homogeneously distributed at the bottom of the glass container and covered with distilled water up to a height of ca. 3 cm;
- (3)
- Water-agar (1%) medium (4 cm high) in plant culture containers MagentaTM GA7 (7.6 × 7.6 × 10.2 cm), wetted with only 5 mL of distilled water.
4.8. Statistical Analysis
5. Conclusions
Author Contributions
Funding
Conflicts of Interest
References
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Substrates | Distilled Water | Water-Agar Medium | ||
---|---|---|---|---|
Megaspore Developmental Stage | Mature (n = 104) | Mature (n = 361) | Immature (n = 205) | |
G (%) | 80.1 ± 5.0 a | 82.1 ± 13.1 a | 65.6 ± 16.1 b | * |
TG (days) | 41.7 ± 1.5 a | 48.3 ± 9.3 a | 77.0 ± 6.7 b | ** |
Sporeling Emergence Indices | Distilled Water | Water-Agar Medium | ||
---|---|---|---|---|
Mature Megaspores | Mature Megaspores | Immature Megaspores | ||
SE (%) | 64.5 ± 8.3a | 69.6 ± 15.5a | 11.6 ± 7.9b | ** |
SEG (%) | 80.4 ± 5.4a | 85.0 ± 12.7a | 16.7 ± 7.9b | ** |
T1 (day) | 52.7 ± 3.1a | 59.5 ± 8.5a | 85.0 ± 6.7b | * |
MDE (% sporelings/day) | 0.8 ± 0.2a | 0.6 ± 0.1b | 0.1 ± 0.1c | *** |
MET (day) | - | 82.9 ± 6.7 | 87.9 ± 5.0 | ns |
MER (day−1) | - | 0.012 ± 0.001 | 0.011 ± 0.001 | ns |
CVt (%) | - | 12.1 ± 1.8 | 6.9 ± 4.3 | * |
EI (n. sporelings/day) | - | 0.8 ± 0.4 | 0.1 ± 0.1 | * |
PV (day−1) | - | 0.70 ± 0.11 | 0.12 ± 0.07 | *** |
U (bit) | - | 3.1 ± 0.3 | 1.7 ± 1.1 | * |
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Magrini, S.; Azzella, M.M.; Bolpagni, R.; Zucconi, L. In Vitro Propagation of Isoëtes sabatina (Isoetaceae): A Key Conservation Challenge for a Critically Endangered Quillwort. Plants 2020, 9, 887. https://doi.org/10.3390/plants9070887
Magrini S, Azzella MM, Bolpagni R, Zucconi L. In Vitro Propagation of Isoëtes sabatina (Isoetaceae): A Key Conservation Challenge for a Critically Endangered Quillwort. Plants. 2020; 9(7):887. https://doi.org/10.3390/plants9070887
Chicago/Turabian StyleMagrini, Sara, Mattia M. Azzella, Rossano Bolpagni, and Laura Zucconi. 2020. "In Vitro Propagation of Isoëtes sabatina (Isoetaceae): A Key Conservation Challenge for a Critically Endangered Quillwort" Plants 9, no. 7: 887. https://doi.org/10.3390/plants9070887