1. Introduction
Pastures are used worldwide to feed livestock; however, the anatomy and digestive physiology of these animals means that methane is generated as an end product of this digestive process, the greenhouse effect of which is more harmful than other gases, and the state of maturity of the pasture influences methane production [
1]. It is essential, therefore, to recognise that pastures serve to control soil carbon cycling [
2] and nutrient recycling, mainly of nitrogen [
3]; nutrients that remain accumulated in soils for up to decades [
4], forming a soil-pasture interaction suitable for the grass-legume association environment [
5]. Italian ryegrass-white clover pastures, introduced in the northern highlands of Peru from 1950 onwards, still constitute the basis of the diet of dairy cattle, the main economic activity of the rural population in this area. This grass-legume association, characterised by complementing each other very well in terms of yield and nutritional value [
6,
7,
8], has remained in most of the farms in this area for several decades, without being renewed, and is therefore considered a perennial life cycle grass and identified as
Lolium multiflorum L. "cajamarquino ecotype". The adaptation and persistence of this grass in the northern highlands of Peru is due to the low frequency of grazing or cutting that is practised, which is done during the reproductive stage. In some of the high Andean plains of our region, it is grazed 90 days after regrowth [
9].
The coexistence between grasses and legumes, in combination with the frequency of cutting, affects the morphological and productive behaviour of Italian ryegrass [
10,
11], as well as its botanical composition, particularly when cutting or harvesting is done after 28 days; it is recommended that this frequency be done at 42 days [
12] when its yield is positively correlated with the mass of stolons. Long intervals between cuts allow for advanced pasture maturity, reflected in increased fibre [
11] higher aerial biomass yield. Still, low protein concentration, digestibility and energy [
13,
14,
15], due to wall thickening and reduced cell content [
16,
17].
In addition to pasture management, the climatic characteristics of the time of year also affect the productive performance of the ryegrass-white clover association [
18,
19,
20], with ryegrass showing more limitations in its growth during drought events [
21] compared to white clover, which thanks to its variability responds with visible phenological changes to new environmental conditions, competing successfully against other plants [
22,
23]. White clover is characterised by its essential contribution of protein in the diet of animals [
16] and by its capacity to fix nitrogen and improve the productivity of this association [
23]; however, it has been found that values lower than 20 % of white clover in associated pastures seriously affect these indicators, in addition to restricting the total daily intake of nutrients. The presence of white clover in pastures should not be above 50 % to meet the nutritional requirements of animals and the release of nitrogen to the environment [
24,
25]
For Tilus et al. (2022) [
11], forage yield increases linearly as the time of cutting is prolonged thus yields have been obtained in this association of 1470 kg DM ha
−1 when this is done around 10 cm in height with a remnant greater than 4 cm [
6]; 2000 kg DM ha
−1 if the height is 13 cm and 2781 kg DM ha
−1 if the ryegrass is 29.8 cm [
26]; 3974 kg DM ha
−1 when the height is 20 cm high and post-grazing remnant of 5 cm [
27]. For white clover, heights of 22 cm have been found when cutting every 60 days [
28]. Cumulative yields ranged from 13039 to 27290 kg DM ha
−1 year
−1 [
6,
20,
27,
29].
Taking into account that the chemical composition presents a high variability due to the time of harvest or maturity stage [
16], when evaluating ryegrass independently, values of 10.01 to 14.67 % CP; 37.64 to 47.52 % NDF; 67.65 to 71.04 % DIVMS and 2.36 to 2.49 Mcal kg
−1 DM of ME have been obtained [
30,
31]. In white clover, concentrations of 24.8 % CP and 40.9 % NDF were found for five cutting frequencies and three cuts 20.6 % CP and 42.9 % NDF were obtained [
14]; when Ladino white clover was cut between 16 to 18 cm in height, Vallejos et al. (2021) [
28] found 23.06 % CP, 11.6 % ash; 25.8 % NDF and 75.04 % DIVMS; in this same species Fonseca-López et al., (2020) [
32], determined 10.9 % ash.
As part of the agronomic characteristics of this association, Ganderats and Hepp (2003) [
33] obtained 3,368 tillers m
2 and Balocchi et al. (2003) [
34], 5,800 to 7,000 tillers m
2; this quantity probably influences the greater basal diameter. In this regard Han et al. (2022) [
35] add that to increase the number of tillers and basal diameter of ryegrass, the soil must be well managed so that the microbial population develops well. In white clover, Doussoulin et al. (2018) [
36] found no difference in stolon elongation rate (0.3 - 0.5 cm day
−1); Lluga-Rizani et al. (2021) [
22] obtained values from 9 to 60 flower heads. When the mowing frequency is between 56 and 84 days, a marked reduction of white clover is observed in the floristic composition [
12], probably due to the shading generated by the ryegrass [
6,
8]. Conversely, high mowing frequencies favour ryegrass and make white clover more competitive [
37]. Although Vallejos (2009) [
38] finds no difference between mowing frequencies about weeds, these can vary between 17.6 and 36.3 % [
39]. The objective of this work was to determine the sustainability for production yield, forage biomass, plant height, floristic composition (basically ryegrass, clover and weeds), morphology of ryegrass and white clover in association and nutritive value, at different phenological stages of associated ryegrass-clover pastures in the northern highlands of Peru.
2. Materials and Methods
2.1. Location
The research was carried out at the Centro de Investigación y Promoción Pecuaria "Huayrapongo" of the Universidad Nacional de Cajamarca, Peru (Latitude 07º09’49" "S", Longitude 78º30’00" "W") located at 2,718 masl; it covered the period from May 2018 to April 2019, in an area of 10 hectares. The pastures were installed approximately 40 years ago, and reseeding is done every five years for ryegrass and clover. A randomized complete block experiment was carried out, where three ages of poly pasture cut (30, 45 and 60 days old); for this, 12 plots of 30 m2 each were selected for each treatment, which were evaluated for a whole year in the two agro-meteorological stations present in the study area.
2.2. Soil Characteristics and Weather Conditions
The pasture corresponded to an association of Italian ryegrass (
Lolium multiflorum L.) "cajamarquino ecotype" - white clover (
Trifolium repens L.) var. Ladino has a marked presence of Kikuyu (
Pennisetum clandestinum), chicory, and wild plantain. Soil analysis indicated clay texture, neutral pH 6.8, organic matter 8.04 %, phosphorus 2.5 ppm and 320 ppm potassium. The ambient temperature and rainfall conditions during the months of evaluation are detailed in
Figure 1.
As shown in
Figure 1, the dry season from May to October and the rainy season from November to April are considered. With this, the two climatic seasons have been differentiated and are associated with fodder availability for livestock.
2.3. Sample Collection
One square metre of quadrats was placed inside each plot, obtaining three representative samples through the cut, 5 cm above the ground. The samples were placed in plastic bags, identified and weighed on an OHAUS electronic balance (± 0.5 g), then transported to the Soil, Water, Fertilizer and Pasture Service Laboratory of the National Institute for Agrarian Innovation (INIA); 100 g of each sample were used to determine the percentage of Dry Matter, in a forced air cooker (cooker equipment, MRC) at 105 °C for 24 h and constant weight, 300 g of the samples of each treatment were separated for the analysis of the nutritional value.
2.4. Parameters Assessed
From the weights obtained from the green forage, the yield in kg of dry matter (DM) per cut and per year in one hectare was determined according to the cutting frequency. Twelve cuts were made during the year for 30-day frequencies, eight for 45-day frequencies and six for 60-day frequencies. From these values and the time elapsed in each cutting, the growth rate was expressed in kg DM ha-1 day-1. Plant height was also measured for ryegrass and white clover; the Height at the time of cutting or harvesting was measured within each quadrant, taking as a reference the ground level up to the maximum Height at which the most significant number of leaves were concentrated (more than 70 %). A 70 cm metal ruler was used.
The floristic composition was quantified using the same samples obtained for yield, separating and classifying them according to the species present (ryegrass, clover and weeds). Considering the green forage weights of each of the species considered. For ryegrass morphology, the number of tillers and ears of ryegrass per square metre was considered, and these values were obtained by counting each one of them for each ryegrass plant contained in the 1 square metre quadrant in three representative places within the subplot. To determine the basal diameter, a tape measure (1.5 m) was used and placed around the base of the ryegrass plant. Depending on the treatments, stolon length and internode length were measured using a ruler for white clover morphology. The growing points per square metre were counted from where the leaves emerged along the stolons, and the number of flowers (flower heads) contained in the quadrants was counted.
2.5. Nutritional Value
The determination of crude protein was carried out at the Soil, Water, Fertilizer and Pasture Service Laboratory of INIA, Cajamarca, according to AOAC 928.08 [
41]; NDF according to AOAC 962.09 [
42] and ash according to AOAC 942.05 [
43]. The determination of neutral detergent fibre [
44] was carried out with a fibre analyzer kit, FIWE, VELP and the
in vitro digestibility of dry matter - DIVMS with the digester kit (ANKOM, USA) [
45]. These analyses were conducted at the Laboratory of Animal Nutrition and Food Bromatology of the National University Toríbio Rodriguez de Mendoza of Amazonas.
2.6. Statistical Analysis
An exploratory analysis of the data was carried out to determine normality and homogeneity of variances, using the Shapiro-Wilks (
p < 0.05) and Levene (
p < 0.05) tests, respectively. The Analysis of Variance (ANOVA) was carried to determine the differences in dry matter yield, plant height and other morphogenetic characteristics corresponding to Italian ryegrass ecotype cajamarquino and white clover variety Ladino, as well as their nutritional value, out using the GNU of the RStudio platform of R Project [
46]. For the comparison of means, Tukey’s multiple range test was used (
p < 0.05).
Author Contributions
Conceptualization, L.V.F. and R.F.C.; methodology, R.F.C., L.V.F. and W.Y.A.G; software, W.Y.A.G., E.A.T.A. and S.S.O.; validation, C.E.Q.P., R.V.C. and L.V.F; formal analysis, W.Y.A.G.; C.E.Q.P.; E.A.T.A. and L.A.V.F; investigation, R.V.C., L.A.V.F., S.S.O. and E.A.T.A; resources, C.E.Q.P., L.A.V.F.; data curation, R.V.F., W.Y.A.G. and E.A.T.A.; writing—original draft preparation, R.V.C., L.A.V.F. and W.Y.A.G.; writing—review and editing, L.A.V.F., W.Y.A.G and R.V.F.; visualization, W.Y.A.G; supervision, L.A.V.F. and C.E.Q.P.; project administration, R.V.C., L.A.V.F. and C.E.Q.P.; funding acquisition, L.A.V.F. and C.E.Q.P. All authors have read and agreed to the published version of the manuscript.