Foliar Micronutrient Application for High-Yield Maize

: Nebraska soils are generally micronutrient su ﬃ cient. However, critical levels for current yields have not been validated. From 2013 to 2015, 26 on-farm paired comparison strip-trials were conducted across Nebraska to test the e ﬀ ect of foliar-applied micronutrients on maize ( Zea mays L.) yield and foliar nutrient concentrations. Treatments were applied from V6 to V14 at sites with 10.9 to 16.4 Mg ha − 1 yield. Soils ranged from silty clays to ﬁne sands. Soil micronutrient availability and tissue concentrations were all above critical levels for deﬁciency. Signiﬁcant grain yield increases were few. Micronutrient concentrations for leaf growth that occurred after foliar applications were increased 4 to 9 mg Zn kg − 1 at 5 of 17 sites with application of 87 to 119 g Zn ha − 1 , 12 to 16 mg kg − 1 Mn at 2 of 17 sites with application of 87 to 89 g Mn ha − 1 , and an average of 8.1 mg kg − 1 Fe across 10 sites showing signs of Fe deﬁciency with application of 123 g foliar Fe ha − 1 . Foliar B concentration was not a ﬀ ected by B application. Increases in nutrient concentrations were not related to grain yield responses except for Mn (r = 0.54). The mean, signiﬁcant grain yield response to 123 g foliar Fe ha − 1 was 0.4 Mg ha − 1 for the 10 sites with Fe deﬁciency symptoms. On average, maize yield response to foliar Fe application can be proﬁtable if Fe deﬁciency symptoms are observed. Response to other foliar micronutrient applications is not likely to be proﬁtable without solid evidence of a nutrient deﬁciency.


Introduction
Nebraska soils are generally micronutrient sufficient with few sites with soil and/or plant tissue samples below critical values for maize production. However, critical levels have not been validated for current yield levels. Production of 12.0 Mg ha −1 of maize grain requires 1.4 kg Fe, 0.5 kg Mn, 0.5 kg Zn, and 0.08 kg B ha −1 [1]. For most Nebraska soils, agronomic crops are most likely constrained from reaching their genetic and environmental potential by insufficient N and water availability [2] but other growth factors become more likely to be limiting as these factors are exceedingly met according to Julius von Liebig and Carl Sprengel's "Law of the Minimum" [3]. Thus, the likelihood of a micronutrient deficiency being the yield limiting factor then increases [4]. Maize, especially when high yielding, has high rates of nutrient uptake during the V4 to VT stage during which demand may exceed supply. Foliar application of micronutrients during this period to complement soil nutrient supply is commonly

Field Sites
During 2013 to 2015, 26 paired comparison strip trials were performed across Nebraska (Table 1 and Figure 1) in order to test the effect of commercial foliar micronutrient products on grain yield. Sites were selected to be representative of Nebraska soils and encompassed 23 soil series ( Table 1). The selected sites had no history of micronutrient or manure applications in the previous 10 years. Nebraska soils are generally micronutrient fertile so finding micronutrient deficient sites proved to be relatively unobtainable [18] as confirmed by a state-wide survey of soil and plant tissue samples and data obtained from Nebraska agronomic testing laboratories [8]. Preseason N applications varied by source, application time, and rate but all sites applied N at a rate sufficient for at least 12.0 Mg ha −1 maize grain production [19].

Treatments
Sites had between three and 20 replications (i.e., strips) depending on farmer equipment and willingness ( Table 2). Two of the 26 trials were on popcorn (Zea mays everta) ( Table 1). Each of the foliar micronutrient applications were selected by the producer and their local agronomist as to best fit the micronutrient needs of each field and based on prior soil and plant tissue recommendations from commercial laboratories. No initial soil micronutrient levels were below critical levels (Ward, 2015) and no plant tissue micronutrient concentrations were below reported critical values for maize prior to tasseling [20]. Maize leaves at sites 20-24 had interveinal chlorosis (Figure 2), which can be an indication of micronutrient deficiency though plant tissue samples were above critical levels. prior to tasseling [20]. Maize leaves at sites 20-24 had interveinal chlorosis (Figure 2), which can be an indication of micronutrient deficiency though plant tissue samples were above critical levels. Products ranged in micronutrient application rate, method and time, adjuvant formulation, and crop growth stage ( Table 2). Management practices varied according to the producers' preferences (Table 1) and soil properties (Table 3). Applications were applied aerially or with a high-clearance applicator in strips with and without the foliar applications and split into adjacent pairs for analysis. Strips ranged in size from approximately 27 to 34 m wide × 715 m long. The high-clearance applicator did not pass through control strips. Products ranged in micronutrient application rate, method and time, adjuvant formulation, and crop growth stage ( Table 2). Management practices varied according to the producers' preferences (Table 1) and soil properties (Table 3). Applications were applied aerially or with a high-clearance applicator in strips with and without the foliar applications and split into adjacent pairs for analysis. Strips ranged in size from approximately 27 to 34 m wide × 715 m long. The high-clearance applicator did not pass through control strips.

Sampling and Laboratory Analyses
Plant tissue samples were collected once after application and were composed of 10 unsprayed upper most fully collared leaves [21] and a soil sample of 0-20 cm depth was collected and composed of 14 cores per strip. Grain yield was determined for each strip from 10 m 2 strip −1 by hand harvest or with a weigh wagon or yield monitor for one combine pass. Grain weight was adjusted to 155 g kg −1 water content. Corresponding plant tissue, soil samples, and grain yield were all collected from the same strip.
Laboratory analysis of plant tissue P, K, S, Fe, Mn, Zn, and B were completed using microwave nitric acid digestion and concentrations determined using inductively-coupled plasma emission spectroscopy (ICP-ES) [22]. Tissue N concentration was determined using the Dumas Method with a Leco FP-428, St. Joseph, MI, USA [23]. Soil samples were dried at 40 • C and ground to pass through a 2-mm sieve prior to analysis. Extractions were with DPTA (diethylenetriaminepentaacetic acid) for B, Mn, and Zn and were extracted with 0.1 N HCl for Fe and Cu and measured with ICAP detection [24] or with sorbitol ICAP detection for B.

Statistical Design and Analysis
The experimental design was a paired comparison strip trial (26 paired comparisons). This design was chosen to allow maize producers to use conventional practices to apply (i.e., high clearance and aerial applicators) and select their foliar micronutrient applications based on their relationship with their local agronomist, the availability of foliar micronutrient formulations, and their knowledge of which micronutrient formulation best fits their field site's demand. Site data was analyzed separately and data from sites with identical treatments (i.e., fertilizer source) and crop were combined and analyzed across sites and years. Yield and plant tissue nutrient concentration data were analyzed using the ANOVA procedure in Statistix 10.0 Analytical Software (Analytical Software Tallahassee, FL, USA) with the assumption of fixed treatment and site effects. A mean comparison test using Tukey's HSD was used to compare treatment effects. Data were managed using Microsoft Excel (Microsoft Excel 2013, Microsoft Corp. Santa Rosa, CA, USA) and plotted to test correlations of plant tissue and soil properties with grain yield response using the scatterplot and Pearson correlation coefficient (r) function.

Field Sites
Soil test values for micronutrients were above published critical levels at all sites [25] (Table 3) though visual signs of Fe deficiency were observed at some locations. The soil B, Fe, Mn, and Zn concentrations (mg kg −1 ) ranged from 0.3 to 1.0 for B, 4.0 to 71.7 for Fe, 2.0 to 23.2 for Mn, and 1.0 to 6.7 for Zn. Soil organic matter ranged from 10 to 33 g kg −1 and pH ranged from 5.6 to 8.2. Sites included clays, loams, and sands (Table 3). Plant tissue micronutrient concentrations were also above published critical levels at all sites according to Bryson et al. (2014). Maize tissue B, Fe, Mn, and Zn concentrations (mg kg −1 ) ranged from 3.7 to 16 for B, 59 to 213 for Fe, 38 to 109 for Mn, and 18 to 57 for Zn in untreated plots prior to tassel. The only visual signs of deficiency were at sites 20-24. These sites had high soil pH and plant leaves showed interveinal chlorosis ( Figure 2) in upper leaves indicating Fe deficiency although soil and foliar Fe analysis did not indicate deficiency.

Effects of Foliar Micronutrients on Grain Yield
The effect of foliar micronutrient application at sites 1-19 was not significant for leaf micronutrient concentrations and grain yield with grain yield effects ranging from −0.38 to 0.75 Mg ha −1 (Table 4). There were no significant yield increases for any micronutrient product when averaged over the three years (Table 5). In conditions with visual signs of deficiency, although above critical plant tissue levels, foliar Fe supplementation was effective at increasing grain yield by an average of 0.4 Mg ha −1 (Figure 3). There were several significant (p < 0.05) treatment*site interactions of the treatments on grain yield. These interactions were likely due to unique site conditions such as soil parameters or various hybrids varying in their micronutrient responsiveness.
Overall, there was no significant yield difference across all sites receiving micronutrient application. In conditions without confirmed micronutrient deficiency, yield responses were unpredictable (i.e., as based on prior soil and plant nutrient concentrations) with significant yield increases for sites 1, 14, and 23 and significant decreases for sites 10 and 13 (Table 4). These sites had some combination of foliar B, Mn, Zn, and Fe applied. Site 23 received foliar Fe only ( Table 1). The soil pH for the responsive sites ranges from 5.6 to 7.8 and likely affected micronutrient availability and the likelihood of response. Overall, maize grain yield was 0.69 Mg ha −1 greater when micronutrients were applied though not significant. The average of significant yield decreases was −0.38 Mg ha −1 , which was also not significant. The negative responses only occurred with high-clearance application at V8 and V11 and not by aerial application, which contributed to the uncontrolled variability related to on-farm field trials. This suggests that the yield reduction may be associated with damage incurred during treatment application, as the high-clearance applicator did not pass through the control strips. Additionally, none of the applied micronutrients from the blends led to toxic levels in plant tissue.     (Figure 3). There were several significant (p < 0.05) treatment*site interactions of the treatments on grain yield. These interactions were likely due to unique site conditions such as soil parameters or various hybrids varying in their micronutrient responsiveness.  (Table 5). Additionally, visual observations of treated strips showed small regreening patterns where foliar Fe droplets contacted the leaf surface on the V5−6 leaf ( Figure 2). No regreening appeared in upper untreated leaves (Figure 2a). Iron and most micronutrients are considered relatively immobile within plant tissue (Bryson et al., 2014). Thus, no regreening in upper untreated new growth leaves was expected nor observed, confirming previously described deficiency correction theory [20]. However, plant and soil Fe concentrations alone were not predictive indicators of grain yield response to foliar Fe. This is consistent with the findings from a hydroponics study evaluating foliar Fe [26]. Micronutrient supplementation must be both predictable for grain yield response and lead to an increase that is great enough to cover the added cost, which is not evident in these trials. It is of note that the lack of significant yield differences may also be due to high levels of variability as in the case of on-farm field trials (i.e., type ll error).

Effects of Foliar Micronutrients on Leaf Tissue Concentrations
Foliar micronutrient applications were more consistent in increasing their respective micronutrient concentrations in leaf tissue than in increasing grain yield, especially in the case of Zn (Figures 3-6) ( Table 4). Of the sites that received foliar Zn, 47% had a nearly significant increase (p < 0.10) of an average of 4 mg Zn kg −1 in the leaf tissue ( Figure 4) (Table 5) which is consistent with other results from a hydroponic greenhouse study (Stewart et al., 2019). In general, the primary nexus for deciding to apply Zn was high, yielding maize locations aiming to boost yields with low plant or soil Zn concentrations but otherwise not below critical levels. Foliar application also affected plant N and P concentrations. The combined analysis of sites 1, 2, and 3 which received 116, 87, 87, 87, and 7 g of N, S, Mn, Zn, and B ha −1 , respectively, increased plant N by 1.3 g kg −1 and plant P by 0.2 g kg −1 (p < 0.05) ( Table 5). The increase in N would be expected with the addition of foliar N, and sulphur application also has synergistic impact on N uptake in plant [27]. The increase in P may have been due to the interaction of P with Fe, Mn, or Zn [28]. Agronomy 2020, 10, x FOR PEER REVIEW 13 of 19    For the combined year and site analysis, foliar applications of 87 to 119 g Zn ha −1 at 17 sites, regardless of product formulation, significantly increased Zn concentration (p < 0.05) in plant tissue by 2.9 to 6.1 mg kg −1 ; however, these increases were not consistently associated with an increase in grain yield (Table 4). Foliar applications of 87 to 89 g Mn ha −1 at 17 sites also had increased plant tissue Mn concentrations by 9.3 (p < 0.05) to 4.2 (p < 0.10) mg kg −1 , with a positive correlation with increased grain yield r = 0.54 but without a significant three-year average yield increase in any product formulation ( Figure 5; Table 5). One site (i.e., site 10) had a significant decrease in Mn and Fe due to their application from 109 to 96 g Mn ha −1 and 141 to 128 g Fe ha −1 ., which corresponded with a decrease in grain yield by 0.57 kg ha −1 (p = 0.06). Foliar applications of B at a rate of 3 to 235 g B ha −1 did not affect plant tissue B concentrations nor grain yield at any of the 15 sites receiving foliar B (Table 4; Figure 6). Though there was a direct linkage between the application of a micronutrient blend and the increase in the corresponding nutrient in the plant tissue, we cannot establish a direct relationship between any singular nutrient and the lack of grain yield response.
Application of 123 g foliar Fe ha −1 at four sites increased plant tissue Fe concentration (p = 0.1) with correlated yield increases that averaged 0.4 Mg ha −1 (p = 0.04) ( Table 5). This response of plant tissue concentrations and grain yield to the foliar Fe application was consistent with the deficiency correction theory in that there were visual signs of Fe deficiency identified at sites 20-23 prior to foliar Fe application and thus, there was an increase in grain yield due to the application of Fe. However, neither the soil nor the plant tissue concentrations were below reported critical values for Fe and thus were not predictive of the associated increase in grain yield. This may also suggest that the critical levels for plant tissue and soil Fe may need to be updated and that precision agriculture technologies may be valuable to match deficiency symptoms with application [29]. Additionally, sites receiving foliar B, Mn, and Zn applications under conditions with no respective deficiency identified by plant tissue samples, soil samples, or visual symptoms, did not have a yield increase for a three-year average.

Relationship between Higher Grain Yield and Response to Foliar Micronutrients
Due to increased micronutrient demand of higher yielding sites, we hypothesized that sites with higher yields and lower micronutrient plant or soil concentrations but not necessarily below critical levels, may be more likely to have increased yield response due to foliar micronutrient supplementation. The combined analysis for maize production sites showed no relationship (r = 0.03) between yield level and yield response to foliar micronutrient supplementation ( Figure 7). Additionally, sites with micronutrient concentrations in soil or plant tissue near critical levels but not below were not consistently associated with an increased likelihood of increased grain yield (r < 0.1). Neither soil organic matter, pH, nor soil/plant P was correlated (r < 0.01) with a positive increase in grain yield.
Agronomy 2020, 10, x FOR PEER REVIEW 17 of 19 Figure 7. Relationship between the control maize grain yield and the grain yield response to foliar micronutrient application for 26 sites. Our hypothesis was that higher yielding sites would have a greater yield response but this relationship did not occur (r = 0.03).

Conclusions
The objective of these on-farm strip-trials was to evaluate the effect of foliar-applied micronutrients on grain yield and plant tissue nutrient status under current management practices using commercially available foliar micronutrients. This research was conducted across a wide range of irrigated maize production conditions, and the results are widely applicable to high yield maize production with similar soil and plant micronutrient status. These data are largely supportive of the deficiency correction theory and refute the use of foliar micronutrient applications for high-yield situations. Higher yielding locations were not associated with an increased likelihood of response to Change in yield (Mg ha -1 ) due to foliar application Figure 7. Relationship between the control maize grain yield and the grain yield response to foliar micronutrient application for 26 sites. Our hypothesis was that higher yielding sites would have a greater yield response but this relationship did not occur (r = 0.03).

Conclusions
The objective of these on-farm strip-trials was to evaluate the effect of foliar-applied micronutrients on grain yield and plant tissue nutrient status under current management practices using commercially available foliar micronutrients. This research was conducted across a wide range of irrigated maize production conditions, and the results are widely applicable to high yield maize production with similar soil and plant micronutrient status. These data are largely supportive of the deficiency correction theory and refute the use of foliar micronutrient applications for high-yield situations. Higher yielding locations were not associated with an increased likelihood of response to foliar micronutrients, thus rejecting the first hypothesis.
Soil test results and plant tissue nutrient concentrations were found to not be predictive of yield response under high-yielding conditions. However, no sites reported soil or plant tissue micronutrient concentrations below critical levels [20]. Nonetheless, there were some significant yield increases indicating a need for better means of predicting yield response to foliar micronutrient application if such treatments are to be profitable. These results are not sufficient to revise previously established critical values but do suggest that sites with plant tissue concentration or soil availability for Fe near to critical levels may still be candidates for foliar Fe application, especially if associated deficiency symptoms and calcareous soil. The mean grain yield response to 123 g foliar Fe ha −1 was 0.4 Mg ha −1 for the four sites with Fe deficiency symptoms. Without visual signs of deficiency or other confirmation of a micronutrient deficiency, yield response to foliar applications of micronutrients remains elusive, thus accepting the second hypothesis for visual signs of deficiency being associated with an increased likelihood of response but rejecting the hypothesis for critical levels being associated with an increased likelihood of response.
Though the use of foliar micronutrient blends made it challenging to decipher which nutrient had the causal relationship with changes in grain yield, the causal linkage of the applied micronutrient to changes in leaf tissue concentration were clearer. Micronutrient concentrations for leaf growth that occurred after foliar applications were significantly increased for Zn, Mn, and Fe but not for B. Thus, the third hypothesis is accepted that applied micronutrients will increase their corresponding foliar concentration for Zn, Mn, and Fe and reject for B.