Cyst hatching and nauplii vitality of <em>Artemia salina</em> treated with different potencies of Silicea terra
Authors: Von Ancken Adalberto C , Da Silva Rodrigo A , Ramos de Miranda Adriana , Dorfman Pierre , Bonamin Leoni 
Published: December 2024

This study is a preliminary step of the Terra Project, a multicentric study aiming to develop methods for improving agriculture and husbandry production through potentized high dilutions (dynamized products), popularly known as “agro-homeopathy.” The present data reflect the characterization of the biological effects of different potencies of Silicea terra on the Artemia salina model concerning cyst hatching, nauplii vitality, and heat shock proteins (HSPs) gene expression, following the standards previously established to study bio-resilience processes [1]. Later, a similar protocol will be tested in rice germination in greenhouses to compose the whole data framework. Herein, the following potencies of Silicea terra were used: 6, 9, 12, 15, 18, 21, 24, 27, and 30cH, all prepared in purified sterile water (PSW). PSW and succussed PSW were used as controls. The potencies and controls were poured into the seawater (10% of the volume) at the moment of cyst seeding using 96-well plates kept at room temperature (average 25oC). All experiment steps were run during the full moon phase due to the high sensitivity of Artemia spp to the circalunar cycle. Two protocols were tested: (a) the constant temperature for 48 hours of incubation and (b) the exposition to one hour of thermic stress (40oC) after 24 hours of incubation. The samples were tested in octuplicate, and each data point was defined as a well containing 5 to 8 cysts. After 24 and 48 hours, the following parameters were evaluated: cyst hatching rate, nauplii vitality (number of live nauplii/total of cysts per well), and nauplii lethality (number of dead nauplii/total of cysts per well). After 48 hours, the remaining biological material was pooled according to each treatment, including a supplementary series of no-treated wells (baseline control), and the mRNA was extracted for further assay of HSP gene expression (HSP40, HSP90, HSP26, and p26). The statistical analysis was made by two-way ANOVA and Tukey test to compare potencies and controls in the function of time, being α=0.05. Lagrangian statistics was used to evaluate the linear time-dependent response in the function of the potencies.  Among the different potencies, the 24cH showed a significant reduction in the lethality rate and a significant increase in the hatching and vitality rates when exposed to thermic stress. These data were similar to the baseline standards. The Lagrangian statistics showed a linear relation between time and potencies. The evaluation of HSP gene expression is still in course. This step concludes that Silicea terra 24cH is a good model for the embryo development of Artemia salina and bio-resilience. Both statistical approaches are valuable tools to characterize the mode of action of Silicea terra on biological systems, giving scientific plausibility for its further use in food production chains. Since Artemia salina are non-sentient animals, there is no need for ethical committee approval.