Agronomic management of the vineyard

The plants are drip irrigated (0.75 m between emitters) applying a flow of 2.1 LPH. The plants are watered 2 h daily during the crop growth production cycle. Each section received the same crop management (pruning, irrigation, fertilization and plant health control) following the norms established by the San Lorenzo wine company.

Treatments applied

On March 14 2014 to one of the sections of the vineyard, 1 L ha^-1 of extract of the seaweed Sargassum spp. (Algaenzims^®) was applied to the soil. Later, on April 29 the same dose of the extract was applied to the plant leaves.

Chlorophyll content

The difference in leaf chlorophyll content between the plants with and without the application of seaweed extract were evaluated with a completely randomized statistical design with three treatments: without the biofertilizer, with application only to the soil and with application to soil and plant leaves and 10 replications, where the experimental unit was the average value of six reading of chlorophyll per leaf of 10 leaves per plant of a group of three plants, so that each replica corresponded to the mean of 180 readings. The leaves chlorophyll content was obtained with a portable gauge (SPAD 502 Plus, Spectrum, Technologies, Inc.). The sampled leaves where from the middle of each shoot under shade conditions avoiding the exposition of the SPAD to direct solar radiation. Treatment means were compared with the Tukey test (α ≤ 0.05).

Net ecosystem exchange of carbon dioxide

The net ecosystem exchange of carbon dioxide (NEE) between the vineyard canopy and the atmosphere was obtained with the next relation (^{Marterns et al., 2004}):

Where: FCO₂ is the carbon dioxide flux between the vineyard canopy and the atmosphere (measured with the eddy covariance method), ΔρCO₂ is the change in the CO₂ density measured to the height Δz, Δt is the time interval (30 min), Δz is the height above the soil surface where the flux measurements are made (3 m), the FCO₂ was obtained with eddy covariance measurements (^{Ham y Heilman, 2003}). With the relation:

Where w is the vertical wind speed, ρCO₂ is the carbon dioxide density. The variables with the prime symbol are deviations from the mean and the horizontal line above two variables denotes the covariance between the variables for a given time segment (30 min). For that, an eddy system with the corresponding sensors was installed (Figure 1). The vertical wind speed was measure with a three dimensional sonic anemometer (CSI-CSAT3, Campbell, Scientific, Inc., Logan, Utah, USA); to obtain ρCO₂ an open path CO₂/ H₂O infrared gas analyzer was used (Open Path CO₂/H₂O analyzer, LI-7500. LI-COR, Lincon, Nebraska USA).

Figure 1 Sensors of an eddy system above a grapevine plantation (cv Shiraz) to measure the net ecosystem exchange of CO₂ (NEE) between the vineyard canopy and the atmosphere. 

The sensors were mounted on a pole 3 m above the soil surface (1.2 m above the vineyard canopy), that was placed at the center of the west edge of each vineyard section. The 3-D sonic anemometer was oriented to the east for the wind to have at least 200 m of contact with the vegetated surface in the east-west direction and 125 m in the north-south direction, before contact with the sensors. Winds form the west that impacted in the back of the 3-D sonic anemometer were no considered for the flux determinations. The vertical wind speed and the CO₂ density were measured at a frequency of 10 Hz with a datalogger CR1000 (Campbell, Scientific, Inc., Logan, Utah, USA). The corresponding covariance were calculated every 30 min. The statistical differences in the NEE in the two sections (with and without the application of the seaweed extract) were evaluated with the non-parametric Wilcoxon test for paired populations (α< 0.05).

Fruit yield and quality

The effect of the seaweed application in the fruit yield was evaluated using a completely randomized statistical design with three treatments (with no application of the seaweed extract, with application only to the soil and with application to the soil and foliage) and five replications where the experimental unit was de average of the yield of 20 plants.

The fruits quality was determined measuring the Brix grades (using a reflectometer), pH (with a potentiometer) and the juice acidity (by the volumetric method). For these parameters, a completely randomized statistical design was also applied with the same treatments and five replications, the experimental unit corresponded to the juice of 40 fruits gathered from the plants of each treatment. Treatment means were compared using the Tukey test (α≤ 0.05).

Leaves chlorophyll content

For all sampled dates, the application of the seaweed extract to the soil and foliage, resulted in a leaf chlorophyll content equal or higher than the application only to the soil and with no extract application (Tukey, α≤ 0.05) (Table 1).This result was probably due because the seaweed extract contain substances as betains, micro elements as iron that are required in the synthesis of chlorophyll of the leaves. It was observed that the leaves chlorophyll content of the three treatments increased from the sample date of May 15, to the one of June 11, this was due to the increase of the leaves chlorophyll pigments because of the leaves maturation. From this date to July 25, a small variation in the chlorophyll content was noted and form August 15 to October 3 a decreasing tendency was observed, due to the loss of pigments because of the progression of the leaves senescence (^{Ahmed y Shalaby, 2012}; ^{Nagy y Pintér, 2015}).

Table 1 Leaves chlorophyll content in a grapevine plantation (cv Shiraz) with and without application of the seaweed extract Sargassum spp. San Lorenzo wine company, Parras, Coahuila. 

Medias con letra diferente dentro de la misma columna son estadísticamente diferentes (Tukey. α≤ 0.05).

The average chlorophyll content of the time evaluated (May to October) in the vineyard section with seaweed application to the soil and foliage was 40.92, while in the section with no application was 39.85. This corresponded to an increase of 2.68%. Previous studies have reported increments of the leaves chlorophyll content. For example, ^{Spinelli et al. (2010)} observed that the application of the seaweed extract Ascophillum nodosum in a concentration of 2% to a strawberry crop (Fragaria moschata) increased 11% the leaves chlorophyll content and 27% the photosynthesis rate. Other studies have reported that foliage application of seaweed extract of Ascophyllum nodosum, Laminaria spp. and Sargassum spp. at a dose of 1 and 2 % respectively increased the leaf chlorophyll content of a garlic crop (Allium sativum L.) that may be related to the betains, a substance present in the liquid fertilizers made of seaweed extracts which increases the leaves chlorophyll content (^{Shehata et al., 2011}). Similarly, ^{Spinelli et al. (2009)} observed an increment of 12% of the leaves chlorophyll content of the apple-tree (Malus Domestica) with the application of a biofertilizer drived from the seaweed extract Ascophillum nodosum at a concentration of 3%, iron is an essential element for the biosynthesis of chlorophyll that was present in applied extract.

Net ecosystem exchange of carbon dioxide

The application of the seaweed extract had no effect in the net ecosystem exchange of carbon dioxide (NEE) between the vineyard canopy and the atmosphere (Table 2) (Wilcoxon, α≤ 0.05). The average daily rate of each month (April to September) of NEE was higher in the vineyard section with no application of the seaweed extract. The daily average of the six months was 274.57 and 205.09 mmol m^-2 in the section without and with the extract application respectively; this represented a difference of 38.87% that was probably due to the difference on the soil type of soil of the vineyard sections because the vineyard section with no seaweed extract application has a soil of higher soil water holding capacity, and the plants had a bigger soil water availability, this had a bigger impact in the photosynthesis rate than the effect of the seaweed extract application.

Table 2 Daily average (of each month) of net CO₂ exchange (mmol m^-2) of a grapevine plantation (cv Shiraz), 2014, without and with the application of the seaweed extract Sargassum spp. San Lorenzo wine company, Parras, Coahuila. 

Medias con letra diferente dentro de la misma columna son estadísticamente diferentes (Tukey. α≤ 0.05).

Previous studies have shown a strong relation between the soil water availability and the photosynthesis rate. On this regard, ^{Castañeda et al. (2006)} observed a reduction of up to 72% of the photosynthesis rate of a bean crop (Phaseolus vulgaris) due to a long lasing stress of water deficit. Other study reported the photosynthesis rate of a corn crop (Zea mays) decreased from 35 to 5.5 µmol CO₂ m^-2 s^-1 due to interruption of irrigation (^{Zarco et al., 2005}). In a similar way, ^{Hayat et al. (2008)} observed a reduction of 75% of the photosynthesis rate of a tomato crop (Lycopersicon esculentum) under water stress.

It was also observed that the CO₂ assimilation rate increased from April to May (due to the vegetative development of the plants) during June and July the assimilation decreased due to the pruning of branches (that was performed the first week of June); the assimilation rate decreased in August and September due to the senescence of the plants (Table 2)

The highest CO₂ assimilation rate was 417.41 mmol m^-2 d^-1 observed in the vineyard section with no application of the seaweed extract in the May month (Table 2). Because in the grapevine plantations, the area covered by the rows of plants is a small part of the total surface, the CO₂ assimilation rate by the vegetation (mmol m^-2 d^-1) is smaller than the observed in crops of full coverage. For example, ^{Vote et al. (2015)} observed an assimilation rate of 1 775 in a corn crop (Zea mays) and 725 in a crop rice (Oryza sativa). For sugar cane (Saccharum officinarum L.) the assimilation rate during fall time was 571.08 (^{Zermeño-González et al., 2012})

Yield and fruit quality

The application of the seaweed extract had no effect in the fruits yield (Tukey, α≤ 0.05)(Table 3), since the highest yield was observed in the vineyard section without the extract. The average plant yield of the sections with and without the extract application were 9.09 kg and 10.32 kg respectively, that corresponded to a difference of 13.5 % this was due to as it was mentioned before, the impact of a bigger soil water holding retention, had a greater effect in the leaves chlorophyll content and consequently in the fruits yield. However, for homogeneous soil humidity conditions, previous studies have reported an increase in the yield for the application seaweed extracts in different crops (^{Fornes et al., 2005}; ^{Zodape et al., 2011}; ^{Pramanick et al., 2014}). Regarding fruit quality, the application of the seaweed extract had no effect in the Brix grades of the fruit juice; however, the higher grade of acidity was obtained with the application of the seaweed extract only to the soil and the lower pH with application to the soil and foliage (Tukey, α≤ 0.05) (Table 3).

Table 3 Yield and fruit quality of a grapevine plantation (cv. Shiraz) without and with the application of the seaweed extarct Sargassum spp. San Lorezo wine company, Parras, Coahuila. 

Medias con letra diferente dentro de cada columna son estadísticamente diferentes (Tukey. α≤ 0.05).

The recommended values for the red wine elaboration from the cv. Cabernet Sauvignon are a pH of 3.28 and a total acidity of 4.92, while for the cv. Chardonnay it is recommended a pH of 3.31 and acidity of 4.04 (^{Ortega et al., 2002}).For the cv. Shiraz the values are a pH of 3.78 and the acidity of 5.5 (^{Gil et al., 2013}). The pH and acidity observed in this study are in the recommended range, while the acidity values are slightly smaller.