Bioavailability and -accessibility of subsoil allocated 33P-labelled hydroxyapatite to wheat under different moisture supply

Soil status, plant development and root architecture

Measurements of the gravimetric soil water content of the subsoils in both variants of the soil rhizotrons showed clear differences depending on the irrigation scenario: For the variants with top-irrigation, the water availability in the topsoil corresponded to a pF value of 2.0 at the beginning of the experiment, while for the variants with sub-irrigation the pF value was 2.2 (Fig. 2).

The initial pF value of the subsoils was approximately 2.1 in both variants. Irrigation affected the time course of pF values: It remained within the range of the field capacity (pF 2.1–2.2 at day 44) in the irrigated top- and subsoils, respectively (Fig. 2a). The other, non-irrigated complementary soil layers dried out and the pF values increased to 2.8–2.9 from approximately day 20 onwards. Changes in gravimetric water content reflected these scenarios: gravimetric soil moisture remained at 5% in the irrigated topsoil but dropped to 2% (the matric potential declined by − 53 kPa) in the variants with subsoil irrigation. Also, the irrigation of the subsoil almost maintained a constant water content (the matric potential declined by − 5 kPa, only), while the subsoil dried out upon top-irrigation (the matric potential declined by − 61 kPa). Consequently, our setup allowed a comparison of plant growth and related P acquisition from soil with either sufficient water supply in top- or subsoil, respectively.

The 10 cm thick layer of topsoil, which was implemented in all rhizotron types, supported similar developments of wheat plants in all rhizotrons. Progressing plant developmental changes in both the aboveground plant parts and the root architecture were observed once the sand was accessed by roots: Since then, plant growth was significantly reduced in the sandy rhizotrons compared with those filled with soil as illustrated by the measured plant parameters after 44 days (Table 1, quantitatively evaluated only for the end of the experiment).

The root architecture within the sand rhizotrons was characterized by two thicker primary roots, which grew strictly towards the bottom of the rhizotrons without exploration of the remaining areas of the rhizotron (Fig. S1; Supplementary Information). The poor growth of the aboveground plant structures in combination with a pronounced root growth within both sand rhizotron types could be attributed to nutrient stress, as plant resources are allocated to root growth under nutrient stress and thus represent a significant metabolic cost factor^9,32. As soon as the roots of the sub-irrigated sand rhizotrons reached the fleece, they started proliferating and rooting within it, thus increasing their water uptake as indicated by increased need for irrigation after 44 days (Table 1).

In all rhizotrons, roots reached the apatite hotspots. Nevertheless, only the roots of the rhizotron variants filled with soil as well as the sub-irrigated sand rhizotrons proliferated within the hydroxyapatite, thus prioritizing the deeper located hotspots (Fig. S2; Supplementary Information). A study of Lynch and Brown⁹ described this behavior as a reaction of the plant to P stress, which they try to overcome by excessive root proliferation within hotspots in the soil once they have encountered them. Noteworthy, this process was only pronounced upon sub-irrigation, with reduced water stress near the hydroxyapatite hotspots⁸. The water availability in the topsoil was of minor importance in this regard. In general, low availability of water can have a drastic effect on root physiology and, in combination with nutrient stress, it can exacerbate root costs⁸.

The root architecture of the soil rhizotrons showed a different pattern to those filled with sand: The entire soil column and the hotspots were interwoven with finer roots (Fig. S1, SI). These optical differences were confirmed by differences in the biomass development of the wheat plants. Many parameters, i.e., the weight (p = 0.006), total height (p < 0.001), length of the ears (p < 0.001), and number of stems, were significantly larger in the soil than in the sand rhizotrons (Table 1). In contrast, differences among the irrigation treatments were not apparent for the soil rhizotrons, whereas plant growth was additionally suppressed by top-irrigation of the sand rhizotrons (Table 1). With an average of 12.4 leaves per plant, the wheat from the soil rhizotrons, for instance, produced 63% more leaves than the wheat from the sand rhizotrons, also the weight to leaf ratios were 86% above those of the two sand rhizotrons variants. The results are consistent with previous studies showing that water stress may reduce leaf area while P deficiency may reduce the rate of leaf development, the number of simultaneous emerging leaves, and thus the final number of leaves^21,33,34. In addition, the number and length of internodes also differed. While wheat plants from soil rhizotrons formed three longer internodes, wheat plants from sand rhizotrons formed five short internodes. This morphological characteristic in the sand rhizotrons is an additional indication of both nutrient and primary water stress, known to lead to a reduction in internodal elongation^35,36.

Nutrient content analyses

The total elemental concentrations of P, N, Ca, Mg and K within all plant parts and soils of the different rhizotron setups are shown in Table 2. In general, nutrient acquisition was most pronounced in the sub-irrigated soil rhizotrons, followed by the top-irrigated trials and the sub- and top-irrigated sand rhizotrons (Table 2).

The trend to improved development of the wheat plants in sub-irrigated treatments (Table 1) was thus frequently accompanied by elevated nutrient concentrations within the plants of the soil rhizotrons (Table 2), and thus by elevated nutrient stocks within the plant compartments (Table 3, see also discussion below). The data thus confirms that uptake of P strongly depends on soil moisture availability^20,37,38. The acquired P was ultimately concentrated in the ears (Tables 2, 3), because stress induced by restricted nutrient and water availability leads to an increased shift of nutrients into the grains of ears³⁹.

Noteworthy, the remaining P in the surface soil hardly contributed to increased uptake of nutrients from the sand rhizotrons. This indicates that the effort of finding nutrients and water in the subsoil was greater than the investment in a common strategy called “topsoil foraging”, which describes an increased root growth in the topsoil due to higher soil P accumulations^11,16,40. We assume that the plants somehow recognized that the topsoil cannot provide sufficient plant-available P in the long-term, making an investment in greater depth more profitable.

The concentration patterns of Ca, Mg and K in the plants only partly resembled those of P, with Ca and K being partly enriched in some plant compartments of the sand rhizotrons relative to the soil rhizotrons (Table 2). These differences likely result from both different priorization by the plants and by different mobility of these nutrients within the plants^41,42. The overall stock of these nutrients in the plants was lower in the sand rhizotrons, due to lower plant biomass development (Table S1, Supplementary Information). Differences in nutrient uptake were also reflected by remaining nutrient concentrations in the seeds. In the comparison of the two soil rhizotron variants, the concentrations of P, Ca, Mg and K in the seeds were about 71 to 75% larger in the sub-irrigated soil rhizotrons than in the top-irrigated ones; relative differences in the sand rhizotrons also being larger (Table 2), simply because utilization of nutrients from the seeds was less required at elevated plant growth conditions in the sub-irrigated trials.

Multiplying biomass weights with their P concentrations yielded the total P_(tot) contents per plant organ (Table 3). The data generally confirmed the information from the above-mentioned tables that P accrual was best in sub-irrigated soil rhizotrons, followed by the top-irrigated ones and the two sand rhizotron variants (Table 3). No significant difference could be determined between the sand rhizotron variants themselves. Leaves and stems acquired most P as intermediate storage organs. Our data is thus in line with observations of Wang et al.²² that higher root proliferation within the subsoil due to superior subsoil moisture conditions improves P uptake from the subsoil.

The P stocks in the ears contributed between 11 and 23% to the total shoot P, reflecting that the grain filling phase was not finished in the soil rhizotrons after 44 days, with grains becoming the major P-sink at maturity, containing up to 89% of the total shoot P⁴³. For the sand rhizotrons, accumulation of P in the ears was more pronounced, reaching up to 63% of the total shoot P in the sub-irrigation trials and indicating a premature grain filling induced by stress. According to Raghothama², this phenomenon can be related to P-starvation, which can significantly increase the uptake and translocation of P through the roots to the ears. Besides, this finding may support Gutiérrez-Boem and Thomas²¹, who hypothesized that P and water availability do not interact in regard to biomass production except when it comes to the allocation of nutrients within the plant.

Distribution of ³³P within plant and soil

Digital autoradiography was used to analyze both the plants and the soil/sand. Due to the high ³³P activity chosen at the beginning of the experiment, there was still a sufficient ³³P activity to generate distinct images after 44 days (Fig. 3).

The images of the wheat plants supported our hypothesis that wheat plants were able to access and take up P from hydroxyapatite hotspots. Furthermore, they supported the conclusions of the different P uptake patterns drawn from the elemental analysis data. These visible differences in the P uptake patterns between the different rhizotron variants can be evaluated semi-quantitatively on the basis of the displayed intensities. The images of the plants from the two soil rhizotron variants indicated a fairly homogeneous distribution of the ³³P in the plants, with higher activities in the plants grown with sub-irrigation (Fig. 3a,b). The respective plants from the sand rhizotrons confirmed the ³³P accrual inside the ears, whereas in the sand rhizotrons with top-irrigation, little, if any, ³³P was detected in the plants (Fig. 3c,d, respectively).

The differences in P uptake patterns between the rhizotron types were also reflected by the ³³P activities within the roots, i.e., they truly resulted from P acquisition from the hydroxyapatite: Roots from sub-irrigated rhizotrons (Fig. 4a,c) displayed higher activities than from the top-irrigated ones (Fig. 4b,d). As we detected higher radioactive intensities in the roots at the bottom of the rhizotrons, surrounding the lower hydroxyapatite hotspots, we concluded that ³³P was primarily acquired from the deeper hotspots at 30 cm depth, likely promoted by better subsoil moisture supply for P uptake provided via sub-irrigation. Potentially, root exudates and the resulting acidification of apatite surfaces were responsible for the release of P from apatite²⁸, even if no overall change in soil pH was detected (data not shown), as the remaining apatite likely buffered all gross pH changes.

For the quantitative assessment of ³³P activities, we performed LSC measurements of digested plants and soils. The results confirmed the visual impressions. Clearly, higher ³³P activities were found in wheat plants when grown under conditions of sub-irrigation than for plants grown under top-irrigation (Fig. 5a). The consistent pattern resulted from both, elevated P uptake and thus elevated ³³P activity within the plants (Fig. S3; Supplementary Information), as well as from the improved overall plant growth in the variants with sub-irrigation (Table 1). Between the two soil rhizotron types, an increase of the ³³P stock activity by a factor of 2.2 (54.9%) could be determined within the wheat plants, promoted by increased subsoil moisture contents, in comparison to the sand rhizotrons, where the increase was equal to a factor of 231.7 (99.6%) (Fig. 5a). These results underline the importance and beneficial effect of soil moisture for the accessibility and acquisition of subsoil P originally bound in hydroxyapatite.

When summing up the percentage distribution of the ³³P radiotracer within the different plant segments, the activity was rather equally distributed within the wheat organs of the soil rhizotrons, whereas in the sand rhizotrons a gradient of increasing ³³P accrual from the lowest internode to the ears was found (Fig. 5b). This data illustrates once more that P allocation in the plant is variable for this nutrient and water stress promotes the redistribution of nutrients from the roots to the ear.

Source: Ecology - nature.com