The study was carried out in the south region of the state of Yucatan, Mexico at the Uxmal Experimental Station of INIFAP located in the municipality of Muna (20˚29'08.1" north latitude and 89˚24'39" west longitude) at an altitude of 50 meters above sea level. The experimental plots were established on red soils (Figure 1) classified as rhodic Luvisols differentiated by their low (Lot 1) and high (Lot 2) intensive agricultural use.

Both lots have important chemical differences as shown in Table 1. Even though, both lots have neutral pH, the salinity of Lot 1 is low according to the Electrical Conductivity (EC) of 0.66 mS・cm⁻¹) whilst in Lot 2 the salinity is medium with an EC of 1.53 mS・cm⁻¹.

The organic matter (MO), as the main source of Nitrogen (N), is satisfactory in both Lots, but the content is higher in Lot 1.

Regarding to phosphorus (P), it is in the optimal range (17 ppm) in Lot 1 but excessive (80 ppm) in Lot 2. It seems that frequent application of fertilizers in Lot 2 has induced toward an increment of residual effects of P in the soil. Potassium (K) is excessive in both Lots as Calcium (Ca) and Magnesium (Mg) are.

The comparison of the results were made considering, as reference, the Nom-021-SEMARNAT-2002 [8] .

The excessive contents of Ca, in the soil, are due to its intrinsic genetic formation. In these soils, the Calcium Carbonate (CaCO₃) is a dominant factor. During its dilution, Ca as an ion (Ca²⁺) form released continuously to the soil solution. In that way, Ca can interfere in the plant absorption of other essential elements such as potassium (K) in an antagonistic process.

The soil reference critical levels, for comparison purposes, were taken from the Official Mexican Standard that establishes specifications for fertility, salinity and soil classification, studies, sampling and analysis suggested (Table 1) by SEMAR-NAT, (2002) [9] .

The free pollination Quality Protein Maize (QPM) varieties so called Sac Beh (Sac) and Chichen Iza (Chich) were the phytometers used. Both with white and yellow grain respectively.

Three treatments were studied as follow: 1) the Control (00-00-00), 2) the Chemical (Chem) fertilization with the formula (60-80-00) of N, P₂0₅, K₂O where Potassium (K) was not applied due to the high levels in the soil, and 3) the Chemical fertilization with Bio-fertilizers (Chem-Bio) (60-80-00 + Mycorrhizal fungi + Azospirillum bacteria) expecting a good synergism between roots and bio-fertilizers.

The treatments were distributed in a completely random block design with three replications in experimental units of 5 m × 4 m (20 m²) consisting of 4 rows of maize 5 m long, separated by 1.0 m and with distances of 0.40 m between strains of 2 plants in order to have an equivalent population density of 50,000 plants ha⁻¹.

The sowing was in the spring-summer of 2017 under well-distributed rainfed conditions. The unique chemical fertilizer was applied 15 days after planting, incorporated into the soil manually while the Bio-fertilizers were added to the seed at planting time.

Bio-fertilizers were applied to the seeds (Figure 2) with a mixture (1:1 ratio) of both: 1) INIFAP™ brand biofertilizer with Rhizophagus intraradices (Mycorrhizae fungus) at a concentration of ≥60 spores and 2) Azospirillum brasilense (Bacterium) at a concentration of 1 × 10⁻⁶ Colony Forming Units (CFU) mL⁻¹. The seeds were mixed (Figure 3) with the Bio-fertilizers and dried at room temperature for 8 hours before planting to the experimental plots. The fertilizer was applied 15 days after sowing and buried 10 cm from the stem in the form of Urea (46% N) and Triple Calcium Superphosphate (46% P₂O₅) in a single application.

For foliar sampling, five opposite leaves from the ears were taken at silk stage in each experimental unit. The five leaves, from the same number of plants randomly selected, were mixed together to make a composite sample to be sent to the laboratory. The samples were dried at 70˚C during 72 hours and the content of N, P, and K was determined in the laboratory [10] in percentage (%); and the critical levels were those reported by Jones and Eck (1973) [11] as reference values.

In both lots, the two varieties showed N deficiencies in all treatments; however, in the case of Sac the general average of N from all treatments was higher in the Lot 1 (2.02%) than in the Lot 2 (1.81%). In contrast, the Chich variety, showed an opposite trend since the average N content was lower in Lot 1 (1.80%) than in Lot 2 (1.86%).

In the extreme cases like the Control and the high intensive soil use of Lot 2, it was observed that Sac had a higher N content (1.86%) than Chich (1.72%); suggesting a better efficiency of Sac to extract N from the soil and export it into the plant.

The N content in plants increased when chemical fertilizer (Chem) and its combination with bio-fertilizer (Chem-Bio) was applied as compared with the Control. However, this trend occurred only in Sac when it was planted in Lot 1 (Table 2) and only in Chich when planted in Lot 2 (Table 3).

The added Bio effect, is detected when contrasting the Chem treatment with

the Chem-Bio one. In Sac, the difference was 2.04% vs 2.17% (Lot 1) whilst in Chich the difference was 1.89% vs 1.99%.

The behavior of N deficiencies was according to Remache et al. [12] who mentioned that in most tropical soils, the main limiting nutrient is Nitrogen (N) followed by Phosphorus (P); N is the most relevant nutrient in crop nutrition and some physiological processes of maize depend on its availability. Accompanying nutrients, which ultimately tend to affect crop yield is another factor to be considered.

Another factor, to explain the differentiated result between both varieties is highlighted by Aguilar-Carpio et al. 2015 [13] when observing genotypic differences due to the effect of Bio-fertilizer and N on the production of dry matter (DM) and yield (GY).

It was determined that only the maize variety VS-535 presented the best agronomic efficiency of nitrogen with the application of Bio-fertilizer (nitrogen-fixing bacteria Azospirillum and Mycorrhizal fungi Glomus sp.); however, when nitrogen fertilization was reduced (80 kg・ha⁻¹ N), the Agronomic Nitrogen Efficiency (ANE) was higher, which indicates the potential of the genotype in the assimilation of nitrogen [13] due to the bio-fertilizer [14] .

On the other hand, it seems that the rate of N applied (60 k・ha⁻¹) as UREA was not good enough to supply N to the plant since maize of all treatments had N deficiencies. Even with the application of Bio-fertilizers, in combination with the chemical fertilizers, the N deficiencies persisted.

Related to this study, it has been found [15] that application of N (180 kg・ha⁻¹) and P (120 kg∙ha⁻¹) significantly increased fodder yield of maize. The N was a limiting nutrient factor and there was a positive interaction with P. The uptake of N increased by N at higher application rates and so was the biomass component. Nitrogen losses to the environment is highly reduced.

The same trend, as N in Lot 1, was for P content in Sac since it increased from the Chem treatment (0.24%) until the Chem-Bio one (0.28%) as compared to the Control with 0.21% (Table 2). The added effect of Bio was only noticeable in Sac when the foliar P content (0.28%) was in the satisfactory reference value considered as 0.27% [11] . In the case of Chich, in Lot 1, no differences were noted between treatments, and the values varied very little from 0.24% to 0.25%, just below the optimal one of 0.27%. The effect of residual P of the soil was noticed in Lot 2, for both varieties.

With one exception, as in Chem applied to Chich in Lot 2, all treatments had P contents above the critical level of 0.28% but opposite to the findings in Lot 1, the Control, in both varieties, had higher P content than Chem and Chem-Bio (Table 3). This suggests that the higher the P in the soil the lower the plant response to chemical fertilizers and Bio-fertilizers will be.

However, the added effect of Bio was detected when comparing Chem vs

Chem-Bio in both varieties of Lot 2. The Sac had 0.29% in Chem and 0.31% in Chem-Bio whilst for Chich the Pcontent was 0.24% for Chem and 0.28% for Chem-Bio.

One of the most important functions of Mycorrhizae is to improve the absorption of phosphorus (P) in plants, especially in soils low in P [16] . However, even though in Lot 1 with high content of P in the soil (80 ppm) the Bio-fertilizers worked efficiently; indicating that residual P, due to constant fertilizer applications, can be activated and introduced in to the plant regardless of the variety. This is a typical trend in intensive agriculture of developing countries in recent decades due to high application rates of phosphate fertilizers [17] [18] .

In the soil analysis, prior to the establishment of the experiment, it was determined that the Potassium (K) in the soil is excessive in both experimental Lots (Table 1) with more than 1000 parts per million (ppm) while critical levels are 117 to 234 ppm.

No maize plant showed nutritional deficiencies of K in the Chem and Chem-Bio treatments since they were above the critical value of 1.70%. However, in the Control (00-00-00) both varieties showed deficiencies depending on the Lot. Sac showed deficiencies in Lot 1 (1.60%) but not in Lot 2 (1.86%) while Chich showed deficiencies in Lot 2 (1.56%) but not in Lot 1 (1.70%).

There is a clearly trend observed in Lot 1, about the positive influence of Bio in the absorption of native K in both varieties. In the Chem treatment, the Sac variety obtained 1.76% of K, while when applying Chem-Bio the K increased to 1.93%. The same happened to Chich. This positive response and activation of soil K is due to the influence of Chemical fertilizers to solubilize native K of the soil. It seems that the antagonistic effect of Calcium (Ca) and Magnesium (Mg) on K is neutralized by fertilizers.

The application of Bio encouraged K uptake since the content in the leaves generally exceeded the chemical treatment applied alone. Consequently, more attention are to be paid on the effects of nitrogenous fertilizers on the reactivation of K in the soil and the synergism it causes when applied with Bio-fertilizers.

The Sac, under extreme conditions of intensive soil use and null application of Chem and Bio (Lot 2) can be more efficient in absorbing K than Chich. This is noted (Table 3) by having a higher content of K (1.86%) as compared to that of Chich (1.56%).

In Lot 1, no significant differences were found between treatments. Regardless of the treatments, the average yields of both genetic materials were higher in Lot 1 with the low agricultural use than in Lot 2.

The yield of Sac in Lot 1 was 6.42 t・ha⁻¹ against that of Lot 2 with 5.10 t・ha⁻¹, a difference of 1.32 t/ha. Meanwhile, Chich obtained 6.86 t・ha⁻¹, in Lot 1, and 5.68 t・ha⁻¹ in Lot 2. There was a general trend of yields associated with N in Sac. Thehigher the yield (Lot 1) the higher the general average of N in leaves (2.02%) and the lower the yield (Lot 2) the lower the general average N (1.81%). This was not the case of Chich which showed a higher general average of N in Lot 2 (1.86%) than in Lot 1 (1.80%) but grain yields were in the opposite trend.

There was not a direct proportional relationship of foliar P and yields for any variety. The general average P content of Sac in Lot 2 (0.31%) was higher than in Lot 1 (0.24%) whilst for Chich the same trend happened with 0.28% in Lot 2 and 0.24% in Lot 1. The same behavior happened with K, whose contents in leaves do not present a defined tendency to be associated with yields.