INTRODUCTION

Sea asparagus (Salicornia neei Lag.) is a perennial species distributed in salt marshes and mangroves of the Atlantic and Pacific coasts of South America, as well as in the saline lowlands of central-south Argentina (^{Costa et al., 2019}), characterized as a gourmet vegetable with high nutritional quality. Its shoots and seeds have a high mineral content (syn. Sarcocornia ambigua - ^{Bertin et al., 2014}; ^{Doncato & Costa, 2018}; ^{Alves et al., 2020}), as well as chemical characteristics (e.g. polyunsaturated fatty acids, such as linoleic acid and oleic acid), which can potentially be used for consumption (Costa, 2006; Bertin et al., 2014; Costa et al., 2014a), biofuel production and in the pharmaceutical industry (EPAGRI, 2008; Bertin et al., 2014; Costa et al., 2014a). Such aspects can be maximized from the selection of progenies with certain characteristics of agroeconomic interest, as it has been done with other species of the genus Amaranthaceae, such as the commercial species Salicornia bigelovii (^{Zerai et al., 2010}). In 2010, a selection program was started at the Laboratory of Biotechnology of Halophytes (BTH) linked to the Federal University of Rio Grande (FURG), through the identification and crossing of two biotypes of natural populations of S. neei from southern Brazil by pure lines, described in Doncato & Costa (2018).

This species is being experimentally cultivated in the field under irrigation with saline water (^{Costa et al., 2014b}; ^{Alves et al., 2020}) and with aquaculture water or effluent (Costa, 2006; Costa et al., 2014a; ^{Doncato & Costa, 2018}, 2022). Recirculating waters and effluents of aquaculture are rich in nutrients that essential for plant nutrition. Previous studies have shown that Biofloc Technology (BFT) system of marine shrimp farming as water source for halophytes, like S. neei, can provide all the macronutrients and micronutrients required for mineral nutrition of plants without the need of supplementation (Doncato & Costa, 2021, 2023a, 2023b). To maximize the production and nutritional composition of plants, irrigation is one of the management practices that must be adapted to the growing condition. According to ^{Lieth & Mansoom (1993}), saline irrigation must be performed frequently to maintain a constant salinity level, adapting the irrigation frequency to the growing condition. For example, in sandy soils with low water retention rates and dry or seasonally dry climates, daily irrigation is required. Soil salinity level must be monitored to maintain yield and avoid reduced productivity (^{Glenn et al., 1999}). Under surface irrigation with high saline water, halophytes often require a leaching percentage of 30% above their water use, so that excess salt accumulated near the roots during periods between irrigations is removed (Glenn et al., 1997, 2013).

The chemical composition of plants can also be markedly affected by saline conditions (^{Costa, 2006}; ^{Ventura et al., 2011}; ^{Duarte et al., 2014}). For example, with increasing irrigation salinity, sodium accumulates in the plant vacuole while nitrogen osmolytes are produced and stored in the cytosol in order to stabilize protoplasmic structures (^{Glenn et al., 1999}; ^{Davy et al., 2006}; ^{Flowers et al., 2010}; Duarte et al., 2014; ^{Alves et al., 2020}). According to ^{Lieth & Mansoom (1993}), the concentration of minerals in halophytes usually increases by 20-40% with the transfer of mixohaline waters to sea level salinity, with sodium being the largest accumulated cation. Ventura et al. (2011) observed that Salicornia irrigated with seawater diluted in different percentages, showed a noticeable increase in sodium and subtle potassium, in 100% of seawater. Furthermore, at this saline concentration, there was an increase in the percentage of total polyphenols (^{Bertin et al., 2014}), which are recognized as antioxidant substances.

This increase in essential minerals and bioactive substances in plant tissue represents an improvement in the nutritional composition of plants under irrigation with high saline concentration. Several studies (^{Luque et al., 1999}; ^{Curado et al., 2014}; ^{Smillie, 2015}) reported that Salicornia has great relevance in the bioaccumulation of metallic elements such as Al, Fe, Zn, Mn and Cu. A better understanding of the effects of saline irrigation on the nutritional qualities of S. neei progenies is needed. In our previous reports, we investigated the influence of saline irrigation on vegetative growth, flowering and biomass production of S. neei progenies (^{Doncato & Costa, 2018}, 2022). The present study aimed to investigate variations in the mineral composition of S. neei progenies subjected to irrigated field cultivation with two saline irrigation schedules from marine shrimp farming (BFT system).

MATERIALS AND METHODS

The selection of S. neei progenies and the experimental design were described in detail in our previous studies, respectively (^{Doncato & Costa, 2018}, 2022). In brief, from November 2014 to April 2015, F₃ and F₄ progenies of the lineages BTH1 and BTH2 of S. neei were grown in two field plots (6.5 × 3.5 m) with sandy soil, irrigated with two distinct schedules, with 375 L per plot of saline effluent from a tank of Litopenaeus vannamei shrimp cultivated in a BFT system. It is a minimal water exchange system where bacteria convert ammonia to nitrate, allowing a low-toxicity nitrogen compound to accumulate without reducing water quality (Doncato & Costa, 2023a). The irrigation schedule treatments were named T2 (irrigation every two days) and T4 (irrigation every four days), which corresponded to 189% (T2) and 94% (T4) of the monthly potential evapotranspiration (Eto; calculated by the Penman-Monteith method; ^{Allen et al., 1998}) of the study period (135 mm per month). In each plot, the four progenies were randomly assigned to the subplots, and each plant represented a replicate of the progeny treatment (n = 20 plants per progeny). The experimental design was in completely randomized blocks (F₃ and F₄ progenies of the lineages BTH1 and BTH2) with no replication for the factor irrigation schedule.

At the end of the experiment, five dry shoot samples (vegetative segments only) from each progeny were ground in a pestle mortar, subsequently subjected to nitric-perchloric and sulfuric digestion (for nitrogen analysis), according to the methodology described by ^{Tedesco et al. (1995}). With the extracts obtained in the digestions, nitrogen (N) was determined by distillation and titration (Tedesco et al., 1995), phosphorus (P) with the ultraviolet visible spectrophotometer, potassium (K) by flame photometry, as well as calcium (Ca), magnesium (Mg), copper (Cu), zinc (Zn), iron (Fe) and manganese (Mn) were determined by atomic absorption spectrophotometer. Only essential plant mineral microelements important for human consumption were analyzed, since S. neei is cultivated for biomass aimed at food. These chemical analyses were performed by the soil laboratory of the Federal University of Pelotas (UFPel; Brazil).

Data was analyzed by two-way nested analyses of variance (ANOVAs) with irrigation schedule as the fixed factor and progeny as the random factor nested in the irrigation schedule. The ANOVAs were followed with a comparison by Tukey’s HSD test at 5% significance.

RESULTS

The different irrigation schedules significantly affected only the macroelements N (18% higher in T4) and Ca (16% higher in T2) (Table 1). Concerning microelements, only Mn and Cu were affected by irrigation schedules, with the contents of both elements being higher in T2. For instance, BTH2-F₃ plants showed 67% more Mn levels in the treatment T2 than in the treatment T4. The overall Cu average of plants from T2 were 54% higher than the one from T4 (Table 1). The BTH2 lineage had a significantly higher content of P (F₄) and Mg (F₃) than BTH1 (Table 1). These differences were 37.2% higher for P concentration than BTH1-F₄ and 17-18% higher for Mg content compared to the BTH1 lineage. Regarding to microelements, differences between progenies occurred for Fe and Cu only, but with the higher values occurring in BTH1 plants, which contained an average of 48% more Fe (F₃) and 39% more Cu (F₄) than BTH2-F₃ (Table 1).

DISCUSSION

Throughout the experiment, it became evident that S. neei was easily cultivated in the field plots with sandy soil under different irrigation schedules with saline shrimp effluent, which was the only source of nutrients available to the plants. This different irrigation schedules significantly influenced the mineral composition (e.g. N, Ca, Cu and Mn). Additionally, the lineages maintained most of the differences in terms of irrigation schedules and BTH2 had great mineral content, with emphasis on the F₄ progeny. This great mineral content under prolonged irrigation periods in also supported by a high vegetative performance (^{Doncato & Costa, 2022}).

Irrigation schedule effects

Few nutritional characteristics of S. neei progenies were affected by the irrigation experiment, demonstrating the high degree of adaptation of this species to water/salt stress. Most glycophyte species and even many halophytes when subjected to the same conditions of soil electrical conductivity and high NaCl contents would lead to an ionic imbalance (^{Flowers et al., 2010}; ^{Ventura et al., 2011}; ^{Rozema & Schat, 2013}). Extended period between irrigations (i.e. T4) resulted in significantly lower concentrations of the minerals Ca, Cu and Mn in S. neei shoots, but particularly of BTH2-F₃. Additionally, N shoot content increased with prolonged irrigation.

The lower moisture and higher saline contents in the T4 field plot (e.g. summer-autumn; ^{Doncato & Costa, 2022}) may have led to the differences in N and Ca contents in the shoots between irrigation treatments. Under salt and hydric stresses, both annual (^{Davy et al., 2001}) and perennial (Davy et al., 2006) Salicornia species produce and accumulate low molecular weight nitrogen compounds (markedly glycine-betaine), mainly in chloroplasts (^{Duarte et al., 2014}), allowing an intracellular osmotic balance with the vacuole, where a large amount of NaCl is compartmentalized (^{Glenn et al., 1999}; ^{Zheng et al., 2009}; ^{Ventura et al., 2011}; Duarte et al., 2014; ^{Alves et al., 2020}). Salicornia neei also accumulated soluble amino-acids in their shoots under water stress conditions (Alves et al., 2020). Consequently, the increase in N content in S. neei shoots from the field plot subjected to greater water stress (i.e. T4) was expected. On the other hand, Zheng et al. (2009) highlighted that the levels of Ca and K in Salicornia europaea shoots decreased with the increasing of salinity in the cultivation. These elements were maintained in plant roots, leading these authors to conclude that Ca and K are strongly involved in root cell ionic homeostasis. In a previous study (Doncato & Costa, 2018), BTH2 plants maintained high levels of Ca in their roots, suggesting a translocation mechanism of Ca into roots under more saline soil, and reduction of this element in S. neei shoots may also occur.

As for the Cu and Mn shoot contents, lower values in T4 plot may be resulted from lower input (via saline effluent loading) and lesser accumulation of these elements than in T2 plot. ^{Smillie (2015}) showed that the Cu content in Salicornia spp. shoots is directly related to its content in the sediment. Additionally, these same authors point out that, during the period of rapid pre-fruiting vegetative growth, less essential or non-essential metals, such as Cu, Fe and Mn, could be diluted by tissue expansion. Due to the higher growth of S. neei in T4 (^{Doncato & Costa, 2022}), this dilution process may have been more marked, leading to differences between irrigation treatments.

Comparison of the mineral content

BTH2 lineage showed significantly higher levels of P and Mg than BTH1 lineage, while the microelements Fe and Cu remained more concentrated in the shoots of BTH1 lineage. Except for P, previously found in higher content in BTH1 lineage than in BTH2 lineage, these results were similar those described by ^{Doncato & Costa (2018}). Shoot contents of P and Mg in the plants that showed the best performance were higher than that observed in S. neei from salt marshes (syn. Sarcocornia perennis -^{Medina et al., 2008}; ^{Bertin et al., 2014}) and S. bigelovii (^{Lu et al., 2010}). The content of Mg in S. neei shoots can be 2-6 times higher than in gourmet vegetables such as A. officinalis (2.0 g Kg^-1 DW; ^{Makus, 1994}) and S. oleracea (4.3 g Kg^-1 DW; ^{Sheikhi & Ronaghi, 2012}).

In general, high but not toxic content of microelements were found in shoots of all S. neei progenies cultivated with saline effluent. Tissue content of these elements in annual and perennial species of Salicornia are directly dependent on soil/irrigation water concentrations, and their species are important bioindicators or bioaccumulators of metals (^{Luque et al., 1999}; ^{Curado et al., 2014}; ^{Smillie, 2015}). In this way, great variations in the levels of metals are cited in the literature and toxic levels of these elements can be observed in plants of some localities.

The average Fe contents in S. neei shoots were 102-304% higher than those mentioned for S. perennis, Salicornia stricta (50-90 mg Kg^-1 DW; ^{Gorham & Gorham, 1955}), S. bigelovii (86.4 mg Kg^-1 DW; ^{Lu et al., 2010}) and A. officinalis (99.9 mg Kg^-1 DW; ^{Makus, 1994}). Average of Mn concentration in BTH2-F₄ (71.68 mg Kg^-1 DW) was higher than in S. stricta shoots from eastern England (60 mg Kg^-1 DW; Gorham & Gorham, 1955), S. perennis (20-81.2 mg Kg^-1 DW; Gorham & Gorham, 1955; ^{Luque et al., 1999}) and the vegetable A. officinalis (21.4 mg Kg^-1 DW; Makus, 1994). The contents of Cu and Zn were lower than those mentioned for cultivated plants of S. bigelovii (7.9 and 35.0 mg Kg^-1 DW, respectively; Lu et al., 2010), A. officinalis (18 and 77.3 mg Kg^-1 DW; Makus, 1994) and S. oleracea (9.9 and 108.6 mg Kg^-1 DW; ^{Sheikhi & Ronaghi, 2012}), as well as Salicornia species grown in soils contaminated with metals in European salt marshes (Luque et al., 1999; ^{Curado et al., 2014}; ^{Smillie, 2015}).

The contents of N, K and Ca were similar among the progenies of S. neei. Their average values were in the same range observed in previous studies with saline effluent irrigation (^{Bertin et al., 2014}; ^{Doncato & Costa, 2018}). However, S. neei progenies had higher values than those observed in shoots of S. neei in the salt marshes of Venezuela (^{Medina et al., 2008}), and irrigated field with lateritic soil in the semiarid region of Brazil (^{Alves et al., 2020}). The concentration of these three minerals ranked in the mid-upper range of values in other species of the subfamily Salicornioideae (^{Gorham & Gorham, 1955}; ^{Lu et al., 2010}) and gourmet vegetables (^{Makus, 1994}; ^{Sheikhi & Ronaghi, 2012}).

CONCLUSIONS

The mineral content of S. neei progenies are influenced by irrigation schedule. The extension of the irrigation regime (i.e. T4) provided nitrogen accumulation in shoots, probably associated with the increment of soluble amino-acids under salt/water stress conditions. Lower shoot contents of Ca, Mn and Cu in T4 than T2 were related to element translocation to roots, dilution of elements in larger plant biomass and/or lower loading of saline effluent in T4. Although both tested irrigation schedules might be applied for the cultivation of S. neei, extended period between irrigations (i.e. T4) provided the best mineral nutrition profile for human consumption (i.e. gourmet food), since the halophytes presented shoot contents richer in N and with less content of Cu and Mn than T2.