General management of the meadow

We worked with an alfalfa meadow, variety San Miguelito, planted in October 2004. The sowing was broadcast, and the study area was divided into 32 plots of 30 m² (6x5 m), comprising four cut-off intervals: 3, 4, 5 and 6 weeks for spring-summer and 4, 5, 6 and 7 weeks for autumn-winter, with four repetitions. The sowing density was 30 kg of viable pure seed per ha; was adjusted with the percentage of purity and germination. At the beginning of the study, a uniformity cut was made at 5 cm height, with a pruning tractor (New Holland Model 6610, USA). It was not fertilized in all the research and irrigation was provided every 15 days at field capacity.

Dry matter yield

The dry matter yield of alfalfa was estimated with two fixed tables of 0.25 m², previously located randomly in each treatment and repeated at the beginning of the experiment. The forage present within each frame was harvested one day before the cut was washed and deposited in labeled paper bags and dried in a forced air oven at 55 °C to constant weight. Subsequently, the dry weight was recorded, to determine the yield per unit area (kg of DM ha^-1) for each season of the year.

Leaf-stem relationship

From the samples collected in the field, to record the fresh weight, 100 g of forage was separated approximately. From this sample, leaf and stem were separated to calculate the leaf-stem ratio (RHT), which was obtained with the following formula.

RHT=H/T

Where: H= leaf dry weight (g); T= stem dry weight (g).

Protein in leaves and stems

From the samples for dry matter yield, a subsample of approximately 100 g was taken in each season and cut interval and dried in a forced air oven at 55 °C to constant weight, later the total protein content of leaves was determined and stems by the micro Kjeldahl method (^{AOAC, 1990}).

In situ digestibility of leaves and stems

With a digestion time of 72 h, in situ digestibility of the dry matter (DISMS) of the stem and leaf of the forage harvested in each season and interval between cut was determined (^{Meherez and Orskov, 1977}).

Climatic data

The maximum monthly temperature ranged between 25 and 32 °C, while the minimum monthly temperature ranged between -3 and 8 °C. The maximum temperatures were recorded from February to June 2007 exceeding 26 °C, while the minimum temperatures were recorded from November 2006 to February 2007 with temperatures below 0 °C. The highest precipitation was recorded from August to November 2006 and from May to August 2007 with an accumulated rainfall of 479 mm. The months with the lowest precipitation were recorded from December 2006 to April 2007 with 74 mm. Obtaining a cumulative annual rainfall of 553 mm (Figure 1).

Figure 1 Maximum and minimum monthly average temperatures and monthly accumulated precipitation during the study period (2006-2007). 

Statistical analysis

To compare the effect of the cut-off intervals, an analysis of variance was performed with the Mixed Model procedure (^{SAS, 2011}). The treatments consisted of four cut-off intervals, randomly distributed under a completely random design with four repetitions. The comparison of means was performed using the adjusted Tukey test ((= 0.05).

Dry matter yield

The lowest seasonal average yield of alfalfa was obtained in the winter season regardless of the interval between cut with 2 438 kg DM ha^-1 and the highest value in summer and spring with an average of 3 357 kg DM ha^-1 (p> 0.05; Table 1).

As the interval between cuts increased, the average seasonal yield increased regardless of the seasons of the year, taking the lower and higher yield in the cutoff range of 4-3 and 7-6 with 1 501 and 4 393 kg DM ha^-1, respectively (p> 0.05).

The tendency of reduction in the yield to lower cut-off intervals, evidences a possible depletion of the reserves located in the roots of the plants, since it has been demonstrated that, at short intervals, there is a depletion of the reserve substances and a rapid disappearance of forage species (^{Matthew et al., 1996}; ^{Valentine and Matthew, 1999}; ^{Villegas et al., 2006}; ^{Teixeira et al., 2008}).

However, cut-off intervals similar to those studied may mean lower or higher yields depending on environmental conditions, the variety of alfalfa and agronomic management (^{Rivas et al., 2005}; ^{Villegas et al., 2004}). The results obtained by these researchers indicate a distribution of performance, similar to that observed in the present study.

The differences in the average yield of forage (Table 1) for each cut interval and season (p< 0.05) show a frank combined effect of the cut interval and season, probably due to the effect of the minimum temperatures in autumn and winter. Although, this assertion requires considering a specific harvest interval, for each season of the year; definitely the lowest forage yields are obtained with the combination of cut-off intervals of 4-3 and 5-4 weeks in any season of the year (p< 0.05). Therefore, it is suggested to cut at intervals of 6-5 and 7-6 weeks regardless of the season of the year.

For the variety Jupiter of alfalfa ^{Rojas et al. (2017)} recorded the highest accumulation of dry forage in spring, with harvest intervals of four weeks. On the other hand, ^{Rojas et al. (2016)} when evaluating different varieties of alfalfa in the Valley of Mexico, they reported the highest forage yield in the stations with higher temperature and vice versa, managing a cut interval of 6, 5 and 4 weeks for winter, autumn and spring-summer.

Leaf-stem relationship

The index calculated for the leaf-stem relationship reveals that more alfalfa leaf was harvested in intervals of 4 to 3 weeks, in autumn-winter and spring-summer, than in intervals of 7 to 6 weeks, in autumn-winter and spring-summer (p< 0.05, Table 2).

That is, a high proportion of leaves is associated with a low yield of dry matter, at short intervals and in autumn and winter (p< 0.05).

With the increase in the cut interval, leaf/stem ratio decreased (p< 0.05) from 1.86 (4 to 6 weeks in winter) to 0.60 (7 to 6 weeks in summer). This has been previously reported (^{Mendoza et al., 2010}), as demonstrated by the results of ^{Villegas et al. (2006)} for Valencian variety alfalfa, with leaf-stem ratio with the highest and lowest value of 1.29 and 0.72 in autumn and spring at cut-off intervals of 5 and 7 weeks, respectively (p< 0.05).

On average, leaf/stem ratio was 50% higher (1.47 vs 0.74), at cutoff intervals 4-3 than at 7-6. A value lower than unity, similar to that obtained in this work, was reported by different researchers in alfalfa in monoculture (^{Morales et al., 2006}) and associated with different grasses (^{Camacho and García, 2003}; ^{Zaragoza et al., 2009}). ^{Rivas et al. (2005)} obtained in five varieties of alfalfa an average leaf/stem ratio of 0.79, with the highest and lowest values of 1.05 and 0.62 in January and November (p< 0.05). On the other hand, ^{Montes et al. (2016)} reported a leaf-stem relationship in Alfalfa cv Oaxaca creole similar to that indicated in this research, obtaining the highest value when the interval between cuts is smaller and vice versa.

Protein in leaf and stem

The protein content, expressed as a percentage of dry matter, in leaves and stems, was influenced by the cut-off intervals and by the season (p< 0.05; Table 3).

The alfalfa leaves harvested every 4 weeks, in the winter, had the highest protein content with 38% and less every 6 weeks with 23.4% in summer (p< 0.05). The cut interval that produced the lowest protein content of leaves was 7-6 weeks with an average of 34.1, 32.4, 31.6 and 28.4%, for the winter, autumn, spring and summer seasons, respectively.

The leaves with high protein content were harvested with cut intervals of 4-3 weeks, while leaves with low protein content were harvested by cutting the alfalfa for 7-6 weeks. This was probably due to the state of maturity of the leaves, which with cuts of 4-3 weeks were younger (^{Valentine and Matthew, 1999}; ^{Brock and Tilbrook, 2000}).

In the case of stems, in all cut intervals, the highest protein content was recorded when alfalfa was harvested in winter with 16.1% and lower in summer with 12.7% (p< 0.05, Table 3). As in the leaves, in all the seasons, at a shorter cutting interval, higher protein concentration of stems (p< 0.05). The shorter cut intervals (4 to 3 weeks) produced higher protein content in stems, with respect to the other intervals (p< 0.05). In descending order, the protein content of stems was 18.1, 16.4, 16.8 and 14.7%, for short intervals (4-3 weeks) and 13.5, 12.1, 11.2 and 10.3% for longer intervals (7-6 weeks), in winter, autumn, spring and summer, respectively.

The highest protein content in alfalfa stems is attributed to the lower age of the harvested plants at short intervals (^{Valentine and Matthew, 1999}). The results reveal that increasing the age of the plants decreases the protein content of leaves and stems (^{Avice et al., 1997}). Similar results reported by ^{Chocarro et al. (2001)} as the interval between cuts decreased the same as the yield of dry matter, however, increased the amount of protein in alfalfa.

Digestibility of leaf and stem

The amounts of digestible dry matter contained in leaves and stems of alfalfa plants are presented in Table 4.

In both cases, an increase in digestibility is clearly observed with the reduction of the cut interval (p< 0.05), in all seasons. In winter, the numerical values ​​were higher, digestible dry matter of leaves and stems with an average of 86 and 74.4%, while in summer the lowest values were 80.8 and 65.1%, respectively (p< 0.05). These results may be due to the low temperatures registered in winter that stop the physiological maturity of the forage species (^{Brock and Tilbrook, 2000}, ^{Sage and Kubein, 2007}), making them more digestible (^{Avice et al., 1997}).

The digestibility differences between winter-summer were 4.1, 3.7, 4.8 and 8.2% for leaves and 5.2, 11.9, 9.4 and 10.8% and for stems, with the intervals of 4-3, 5-4, 6-5 and 7- 6 respectively. In general, the greater interval between cuts affects more the stems and leaves digestibility than the seasons of the year (Table 4). Similar results reported by ^{McMahon et al. (1999)} in alfalfa digestibility with the highest and lowest value from 89.8 to 69%.