INTRODUCTION

The genus Salicornia belongs to the family Chenopodiaceae, constituted by the species S. subterminalis; S. virginica; S. maritima.; S. borealis; S. rubra; S. pacifica and S. bigelovii (^{Lutts and Lefèvre, 2015}). This last species usually grows in saline coasts and habitats in Europe, Africa, and North America (^{Munns and Tester, 2008}). The studies performed demonstrated that the species is highly tolerant to salinity, considering it a facultative halophyte that can grow at different salt concentrations with an annual life cycle (10-12 months) (^{Bagwell et al., 2001}; ^{Beltrán-Burboa et al., 2017}).

Cultivation of halophyte plants has some advantages because their growth does not depend totally on fresh water, and some species are suitable for human consumption in salads or flour. They can be used as high protein content forage for animal consumption and livestock, particularly in arid and semiarid regions where traditional forage is not possible due to lack of water and adverse environmental conditions (^{Rueda-Puente et al., 2017}). Moreover, halophyte plants have an interesting potential as industrial components because of the extraction of vegetal oil content in their seeds, which can be used in cosmetology (^{Slama et al., 2015}). A recent study demonstrated that halophyte plant oil, such as that of S. bigelovii, can be used to produce biofuel (^{Al-Rashed et al., 2016}), which suggests that its cultivation could have the advantage of allowing using coastal land and others with high salinity. In this manner, agricultural land would not be used where non-halophyte species suitable for human or animal consumption could be cultivated. Under this circumstances, these authors argued that cultivation of S. bigelovii would allow producing forage and alternative products without affecting food security.

In Mexico, S. bigelovii can be found frequently in the coasts of the Gulf of México, also in Sonora and the Baja California peninsula, particularly in habitats subjected to periodical flooding-drying processes by tidal effect (^{Rueda-Puente et al., 2011}). The state of Baja California Sur is possibly the region with the most possibilities in the country of developing halophyte cultivation because of its semiarid conditions and large extension of salinity soil not suitable for traditional cultivations besides the low availability of quality irrigation water. The cultivation of S. bigelovii could be a viable alternative to restore vitality to these areas, generating jobs and producing nutritional biomass for human or animal consumption or as raw matter to obtain oil for the industry. To achieve these goals, optimizing seed germination is a priority since seeds have a naturally strong dormancy that makes initial germinal tissue development difficult, and consequently, massive, uniform and systematic production of seedlings as raw matter for technified cultivation systems. Different research studies have revealed important results in this sense, from thermal seed treatments to comparative studies, to define the optimum temperature to improve the germination process (^{Rivers and Weber, 1971}) and the application of sodium nitrate for optimization (^{Wong-Corral et al., 2010}). The use of growth promoting or nitrogen-fixing probiotic bacteria is a more natural alternative to optimize the germination process in Salicornia (^{Rueda-Puente et al., 2009} and ^{Hernández-Perales et al., 2016}). The use of homeopathic medicines has been tested successfully in several cultivations, demonstrating its mode of action as plant growth promoters (^{Pinto et al., 2014}; ^{Meneses-Moreno, 2017}). On the other hand, some homeopathic medicines for use in humans, registered in the Health Ministry of México (^{SSA, 2015}), have been applied successfully as plant growth promoters (^{Mazón-Suástegui et al., 2019}) for the control of pathogen organisms (^{Narváez-Martínez et al., 2014}) and as attenuators of negative effects of abiotic stress (^{Giardini-Bonfim et al., 2012}). Given that the homeopathic medicines Natrum muriaticum and Phosphoricum acidum contain nanoparticles of the active ingredient, they can activate diverse physiological processes in plants that allow a better cellular function and a more efficient development of their tissues and organs (^{Mazón-Suástegui et al., 2018}; ²⁰¹⁹). With this background and considering to our knowledge that no references were available for halophyte plants, the objective of this study was to evaluate the effect of homeopathic medicines in initial germination and growth of S. bigelovii, under controlled conditions.

Materials and Methods

The experiment was performed at Centro de Investigaciones Biológicas del Noroeste (CIBNOR) in the Laboratorio de Fisiotecnia Vegetal (Physical-technical Plant Laboratory). Salicornia bigeloovii seeds were used, previously selected by the methodology proposed by the International Rules for Seed Testing (^{ISTA 2010}).

Results and Discussion

The results showed a significant difference (P = 0.0000) in germination percentage (PG), of which treatment NaM-7CH had the greatest response with 86% of germinated seeds, 44% more than those in the control treatment (AD). No significant differences were found between PhA-13CH and NaM-31CH treatments, but both were statistically different from the control group (Figure 1). Germination is a complex process where a group of physiological and biochemical factors take part and give rise to a new plant. This process depends greatly on external agents to reach their maximum expression, in this case in S. bigelovii, the presence of marine salt stimulates germination because it is a facultative halophyte plant and needs salt to optimize its physiological processes (^{Beltrán-Burboa et al., 2017}). It explains the results obtained with NaM-7CH since this medicine is made from sea salt, which agrees with one of the basic assumptions of homeopathy: the denominated “Law of Similars” (Similia Similibus Curentur: ‘Like Cures Like’ or ‘Treat likes by likes’), its main philosophical and conceptual foundation from the biophysical point of view (^{Mazón-Suástegui et al., 2018b}). However, from a chemical point of view and taking as reference only Avogadro’s theory, the sea salt content in NaM as an active ingredient in nanoparticles, would be greater in the seventh centesimal dynamization (7CH) than in the 31^st centesimal dynamization (31CH), something that could also justify such results (^{Mazón-Suástegui et al., 2019}). Other authors have obtained similar results applying sodium nitrate to stimulate germination (^{Wong-Corral et al., 2010}).

Figure 1: Effect of homeopathic medicines on the germination percentage of Salicornia bigelovii. Different letters show significant statistical differences (P = 0.05). 

In relation to the effect of homeopathic medicines in germination rate, a significant difference was found (P = 0.0000) among the homeopathic treatments applied, of which NaM-7CH stood out as the one with greater TG (Figure 2), which made a positive effect evident in germination speed of S. bigelovii seeds treated with NaM-7CH. This finding can also be associated to the presence of nanoparticles of the active ingredient that have the capacity of stimulating germination efficiently, increasing speed significantly with this physiological process (^{Mazón-Suástegui et al., 2019}). The treatments with PhA-13CH and NaM-31CH, also favored the germination rate although in lesser proportion; however, it is convenient to highlight that the three homeopathic treatments applied stimulated the germination rate of S. bigelovii when compared with the control treatment (Figure 2).

Figure 2: Effect of homeopathic medicines on the germination rate of Salicornia bigelovii. Different letters show significant statistical differences (P = 0.05). 

When stem length (LT) was analyzed, a significant difference of this variable could be observed with respect to the homeopathic treatments applied (P = 0.000), noticing again a greater response in plants treated with NaM-7CH (Figure 3). With this homeopathic medicine, it was possible to increase LT in 63.6% with respect to the control treatment (DW), such as with the other response variables, all treatments stimulated growth in LT compared with the control but in a lesser percentage to NaM-7CH (PhA-13CH, 45% and NaM-31CH, 27.2%).

Figure 3: Effect of homeopathic medicines on the length of the stem of Salicornia bigelovii. Different letters show significant statistical differences (P = 0.05). 

The results observed in LT for the treatment NaM-7CH could be explained by the presence of oligoelements that constitute the active ingredient (sea salt) of the medicine. It could be the case of the magnesium (Mg) oligoelement chemically present in NaM-7CH, which is essential for the development of chlorophyll molecules, thus, its vital importance in photosynthesis, the main plant production process starting from nutrients and luminous energy. Moreover, magnesium has a predominant role in the enzymatic activity associated with carbohydrate metabolism (^{Xiao et al., 2014}). Similar results were obtained by ^{Lippert et al. (2007)} when they studied the effect of homeopathic sulfur dynamizations in sorghum (Sorghum bicolor L. Moench) cultivation growth; a stimulating effect was evident on the LT variable, associated to the beneficial effect of the treatments used, concluding that they contained plant growth stimulating oligoelements.

With respect to the radicle length (LR) variable a significant difference was recorded among treatments (P = 0.000), with a favorable response to all the homeopathic dilutions applied but greater LR in those treated with NaM-7CH (Figure 4). Nonetheless, it is worth to point out that the rest of the treatments (PhA-13CH and NaM-31CH) also showed positive responses in this variable. These results could be attributed to the presence of nanoparticles of the active NaM ingredient (^{Mazón-Suástegui et al., 2018}), which in this case they could be oligoelments, for example, calcium whose function in plants is vital to maintain the integrity of the cell wall membrane besides its importance in strengthening roots in regulating the physiological processes related to nutrient absorption (^{Hojjat- Nooghi and Mozafari, 2012}).

Figure 4: Effect of homeopathic medicines on the length of the radicle of Salicornia bigelovii. Different letters show significant statistical differences (P = 0.05). 

With respect to plant biomass production, significant differences were observed for BFPA (P = 0.000), BSPA (P = 0.023), BFR (P = 0.0000) and for BSR (P = 0.00), highlighting once again treatment NaM-7CH as the one with the greatest response for these variables (Table 1). The results obtained demonstrated, without any doubt, a positive effect of all the homeopathic treatments used in this study. Additionally, these results suggested that low dilutions, in other words, those that were found below Avogadro’s (1 × 10^-23) limit, which is approximately equivalent to a 12-Centesimal Hahnemanian (12CH) dynamization, ensured a better response of S. bigelovii.

Applying the criteria based only in the chemical presence of the elements contained in the initial concentrate or Mother Tincture (MT), from which the homeopathic medicine was prepared by a serial dilution/agitation process, may guarantee the presence of the “active principle” in NaM-7CH but not in NaM-31CH. However, the basic principle of homeopathy does not reside only in the presence of molecules and/or nanoparticles of the so called MT “active principle but in complex processes of physical and electromagnetic nature that maintain its activity, including in ultradilution beyond the limit established by Avogadro’s theory. Therefore, all the previously mentioned should be investigated in depth because dynamizations beyond the 12CH dilution maintain the capacity of inducing biologically measurable effects, unchaining positive responses at cellular, physiological, genomic, transcriptomic and metagenomic levels, among others (^{Mazón-Suástegui et al., 2018b}, ²⁰¹⁹).

Conclusions

All the homeopathic medicines applied in this study increased germination rate and percentage of S. bigelovii significantly, as well as stem length, radicle length and biomass production. However, the best results and with the greatest productive applicability were obtained with the treatment NaM-7CH, with which the variables PG, TG, RL and BFPA were highly stimulated with the prophylactic application of the homeopathic medicine. These results confirmed that agricultural homeopathy is a viable and low cost alternative to optimize S. bigelovii cultivation, and that Natrum muriaticum favored seed germination and biomass production in an eco-friendly manner of this halophyte species that has an excellent agricultural, productive and marketing potential.