Yield Performance of Forage Shrubs and Effects on Milk Production and Chemical Composition under the Tropical Climatic Conditions of Peru

: Forage shrubs have the potential to substantially contribute to pasture and increase the milk production of cows in tropical environments. The yield performance of forage shrubs and its effects on the production and chemical composition of milk in Bos indicus and Bos taurus crossbred cows in the tropics of Peru were studied. Fifteen cows were divided into M. alba , L. leucocephala , M. oleifera , and C. argentea treatments and only one of B. brizantha (control). Analysis of variance ( p < 0.05) and comparison of means with Tukey’s test were performed. The highest plant height, stem diameter, fresh forage, and dry matter were observed in L. leucocephala and M. oleifera . The highest milk production was observed in cows fed B. brizantha with M. alba , and the highest milk production was in the rainy season. The highest concentration of fat and total solids was observed in milk from cows fed B. brizantha with L. leucocephala . The highest utility was observed in cows fed B. brizantha with M. alba ; however, the highest operational proﬁtability was observed in the treatment of only B. brizantha and B. brizantha with L. leucocephala . The use of forage shrubs can contribute to cattle feeding, especially in the dry season when there is a shortage of pastures, and possibly contribute to improving the soil and overcoming climate change.


Introduction
The growth of the human population and the increase in income in underdeveloped countries have stimulated the increase in the demand for animal protein. In 2020, it was estimated that these countries consumed 223 million metric tons of milk, more than in 1993 [1], and by 2050, an increase of 58% in consumption of dairy products is estimated compared to current values [2]. In this sense, livestock activity plays an important role in the economy of producers in Latin America [3], but until now, production indicators have continued to remain unchanged, having negative repercussions on the economy of producers [4]. The negative repercussions are due to the disadvantages that occur in old livestock systems, such as monocultures that provide a reduced quantitative and qualitative offer of grasslands, constant droughts, and the loss of organic matter and soil biodiversity [4,5].
A challenge of sustainable livestock is to improve the existing trend with the implementation of efficient and sustainable systems over time that allow us to cover the nutritional deficit of livestock and reduce the production of greenhouse gases [6,7]. All A challenge of sustainable livestock is to improve the existing trend with the implementation of efficient and sustainable systems over time that allow us to cover the nutritional deficit of livestock and reduce the production of greenhouse gases [6,7]. All tropical grasses of the grass family have protein contents of less than 12% that do not gather the nutritional requirements of the animal; however, this can be balanced by supplying forages and shrubs in the diet that have a protein content greater than 15% [8]. A high number of forage woody species have the potential to produce plant biomass that serves as feed for cattle in silvopastoral systems and could help mitigate the effects of climate change and nutritional deficiencies in dry areas [9,10]. Tree forages are used in cattle diets to dilute the starch content in the ration, prevent acidosis, and regulate methane production in the rumen [11]. Feeding ruminants in intensive production systems for dairy production requires the supply of very high levels of energy and protein [12]. Most tree forage species have ecological plasticity because they are found in different soil conditions, precipitation, and temperature [13]. Therefore, its use in the diet of dairy cows in silvopastoral systems could improve the balance and use of the energy contained in it and, consequently, optimize milk production and quality [14].
Cattle raising is one of the main economic activities in the San Martín region (Peru), with the use of natural and introduced grasses, with good initial performance, but which decreases due to inadequate management of overgrazed soils. The development of livestock activity using woody, weedy, and climbing species in association with pastures is an alternative that should be promoted in the different livestock areas, especially in the tropics, due to its great plant biodiversity [15]. Over the years, research has been carried out on efficient and sustainable alternatives, such as the identification of species such as grasses, legumes, and weeds, among others, that have good agricultural potential, production, and nutritional quality [15]. However, these species have a different behavior according to the geographical area and agroclimatic conditions, among other factors [16]. Under this perspective, it was proposed (i) to analyze the yield performance of four forage shrub species for animal feed, (ii) to determine milk production in Gyr with Brown Swiss crossbred cows fed with four forage shrub species, (iii) to analyze the lactic acid, fat, and total solids of milk from Brown Swiss cows fed with four forage shrub species at two times of the year, and (iv) to analyze the economic and profitability indicators.

Site and Experimental Design
The experiment was carried out in the paddocks of the National Institute of Agrarian Innovation, Fernando Belaunde Sur Highway Km 14-Juan Guerra District, San Martín Province, Peru (6°35′42″ S, 76°18′24″ W; 205 m altitude). The temperature and precipitation are detailed in Figure 1. January to June was the rainy season, and July to December was the dry season. The temperature and precipitation values were taken from SENAMHI (https://www.senamhi.gob.pe/?p =seasons (accessed on 14 May 2022)). leucocephala), (iii) Moringa (Moringa oleifera), (iv) Cratylia (Cratylia argentea), and (v) a noforage shrub, grass only control (B. brizantha). Three replicates were established for each treatment for a total of 15 experimental plots in a randomized blocked experimental design. In each replica plot of L. leucocephala and M. oleifera containing a woody treatment, a total of 100 woody forages were arranged in three 1 × 2 m plots, and for M. alba and C. argentea containing a woody forages treatment, a total of 65 woody forages were arranged in three 1.5 × 2 m alleys with pasture grass. Stands of L. leucocephala, M. oleifera, and C. argentea. were established from locally collected seeds and M. alba planted from 40 cm stakes 2 years prior to the experiment ( Figure 2). Each treatment contained only one forage shrub species and the pasture Brachiaria brizantha cv. The treatments were as follows: (i) Morera (Morus alba), (ii) Leucaena (Leucaena leucocephala), (iii) Moringa (Moringa oleifera), (iv) Cratylia (Cratylia argentea), and (v) a no-forage shrub, grass only control (B. brizantha). Three replicates were established for each treatment for a total of 15 experimental plots in a randomized blocked experimental design. In each replica plot of L. leucocephala and M. oleifera containing a woody treatment, a total of 100 woody forages were arranged in three 1 x 2 m plots, and for M. alba and C. argentea containing a woody forages treatment, a total of 65 woody forages were arranged in three 1.5 x 2 m alleys with pasture grass. Stands of L. leucocephala, M. oleifera, and C. argentea. were established from locally collected seeds and M. alba planted from 40 cm stakes 2 years prior to the experiment ( Figure 2). Fifteen Gyr with Brown Swiss crossbred cows that were visually estimated to have the same size, age, and body condition were selected for the experiment. The cows were initially placed in a training corral where they had a 30-day period of acclimatization [17]. The cows were housed according to treatment, the forage shrubs were cut and offered twice a day (6:00 a.m. and 16:00 p.m.), and water was supplied ad libitum. The experiment began on January 2018 and concluded on December 2018.

Forage Measurements and Milk Production
Plant height was measured from the base of the stem to the top of the forage shrubs with a winch, and stem diameter was measured using a forestry millimeter tape. For the fresh matter, 10 plants per treatment were randomly selected, and only the edible matter (leaves and succulent stems) was weighed. For the dry matter, 250 g were weighed and brought to 60 °C in an oven until a constant weight was obtained [18]. Fifteen Gyr with Brown Swiss crossbred cows that were visually estimated to have the same size, age, and body condition were selected for the experiment. The cows were initially placed in a training corral where they had a 30-day period of acclimatization [17]. The cows were housed according to treatment, the forage shrubs were cut and offered twice a day (6:00 a.m. and 16:00 p.m.), and water was supplied ad libitum. The experiment began on January 2018 and concluded on December 2018.

Forage Measurements and Milk Production
Plant height was measured from the base of the stem to the top of the forage shrubs with a winch, and stem diameter was measured using a forestry millimeter tape. For the fresh matter, 10 plants per treatment were randomly selected, and only the edible matter (leaves and succulent stems) was weighed. For the dry matter, 250 g were weighed and brought to 60 • C in an oven until a constant weight was obtained [18].
Milk production was recorded for each treatment. Milking was performed at 2:00 a.m. and 12:00 p.m. In the milk register, the breed, age, parents, and date of birth of each individual were described. For the analysis of total solids, fat, and acidity, individual samples were taken from each treatment and determined according to the AOAC [19]. The profitability was determined by following the method of Cacep et al. [20]. All procedures respected international standards for animal experimentation and animal welfare (Peruvian National Law No. 30407: "Animal Protection and Welfare").

Data Analysis
The data was processed in a completely randomized design, and data were checked for normality with Shapiro-Wilk and variance with Levene's test. Analysis of variance (p < 0.05) and comparison of means with Tukey's test were performed. All statistical tests were carried out in IBM ® SPSS vs. 26.

Productive Performance of Forage Shrub Species
The plant height, stem diameter, fresh forage, and dry matter significantly varied across treatments (p < 0.05) ( Table 1). For plant height, L. leucocephala and M. oleifera were similar in height, but they were superior to C. argentea and M. alba. The stem diameter varied according to forage shrub species; M. oleifera showed a greater diameter compared to the other species. L. leucocephala and M. alba were statistically similar in stem diameter but higher than M. oleifera. Fresh forage production varied significantly according to forage shrub species (p < 0.05). L. leucocephala produced approximately 5.1 t/ha more than C. argentea and M. alba and approximately 2.0 t/ha more than M. oleifera (Table 1). A similar effect was observed in dry matter production; L. leucocephala showed superiority in dry matter production compared to the other forage shrub species (p < 0.05). In the dry season, the forage shrubs L. leucocephala and M. oleifera were taller than C. argentea and M. alba The production of fresh forage and dry matter was higher for L. leucocephala, and production was lower for M. alba (Table 1).
In the rainy season, the plant height, stem diameter, fresh forage, and dry matter varied between forage shrub species. L. leucocephala and M. oleifera showed similar growth but were superior to C. argentea and M. alba. M. oleifera had a higher diameter, and C. argentea had a smaller diameter. L. leucocephala doubled the production of fresh forage and dry matter compared to M. alba and C. argentea (Table 1). Finally, in the rainy season, higher productions of fresh forage and dry matter per hectare were observed (p < 0.05).

Milk Production
The average values of milk production according to treatment and by season are shown in Figure 3. The treatment of B. brizanta with M. alba achieved higher milk production compared to the cows that were fed B. brizanta and B. brizanta with C. argentea (p < 0.05). The cows with the lowest milk production were those that were fed only with B. brizantha and B. brizantha with C. argentea. The cows with the highest milk production were those that were fed B. brizantha with M. alba ( Figure 3A). The highest milk production was observed in the rainy season compared to the dry season (p < 0.05, Figure 3B).

Milk Production
The average values of milk production according to treatment and by season are shown in Figure 3. The treatment of B. brizanta with M. alba achieved higher milk production compared to the cows that were fed B. brizanta and B. brizanta with C. argentea (p < 0.05). The cows with the lowest milk production were those that were fed only with B. brizantha and B. brizantha with C. argentea. The cows with the highest milk production were those that were fed B. brizantha with M. alba ( Figure 3A). The highest milk production was observed in the rainy season compared to the dry season (p < 0.05, Figure 3B).

Milk Components
Milk lactic acid levels varied between treatments (p < 0.05), and the average values are detailed in Table 2. Milk from cows fed B. brizantha with M. oleifera and B. brizantha with L. leucocephala presented higher levels of acidity. Milk fat concentration was higher in cows fed B. brizantha with L. leucocephala, and the lowest concentration was recorded in milk from cows fed B. brizantha with C. argentea. The lowest concentration of solids was recorded in the milk of cows fed B. brizantha with C. argentea compared to the other treatments. However, lactic acid, fat, and total solids were not affected when analyzed in the dry season compared to the rainy season (p > 0.05).

Economic and Profitability Indicators
The higher utility was observed in the treatment of cows fed B. brizantha with M. alba, followed by the treatment of B. brizantha with L. leucocephala, and the least useful treatment was B. brizantha with C. argentea. However, the highest operational profitability was observed in the treatment of only B. brizantha and B. brizantha with L. leucocephala, and in these two treatments, a higher benefit/cost ratio was also observed compared to the other treatments (Table 3).

Discussion
The height of M. oleifera and L. leucocephala were higher than the values of C. argentea and M. alva. These forage shrubs species (M. oleifera and L. leucocephala) exceeded 200 cm in height at 12 months of age [21,22]. The height of L. leucocephala was higher than that obtained by Anguiano et al. [23], who obtained a value of 138.28 cm under conditions of Colima, Mexico, with high planting densities under coconut woody forages at the height of 59 m above sea level. The diameter varied according to the forage shrub species, and a range from 1 cm to 6 cm was observed. Studies report stem diameters of 0.92 cm for M. oleifera and 0.62 cm for L. leucocephala at 14 weeks of age [24]. These dasometric indicators in forage shrub species are influenced by environmental parameters such as temperature. The height and diameter of the L. leucocephala forage shrub decreased at temperatures below 20 • C [25]. The production of fresh forage and dry matter varied according to the forage shrub species and season. Higher values of fresh forage were obtained in the rainy season compared to the dry season. The highest values were reported for L. leucocephala compared to the other species. Our records of fresh forage based on M. oleifera are lower than those obtained by Navas [26], who obtained fresh forage production of 12.08 t/ha. However, forage production can vary depending on humidity, temperature, and wind speed, as well as the type of fertilization, variety, density, and cutting height [27,28].
L. leucocephala produced more dry matter (2.67 t/ha) than the other forage shrub species. The values obtained in this investigation were lower than the 8.28 t/ha reported by Benítez-Bahena et al. [29] for L. leucocephala plus B. brizantha at a planting density of 2500 plants/ha. M. oleifera produced 1.54 t/ha of dry matter; however, this value is lower than that obtained by ref. [30], who obtained dry matter values of M. oleifera of 7.3 t/ha.
The analysis of plant height, stem diameter, fresh forage, and dry matter in the dry season varied between forage shrub species. L. leucocephala and M. oleifera were the two species that showed superiority over C. argentea and M. alba. Due to the limited forage production or inadequate handling of gramineous pastures, the sowing of alternative species such as L. leucocephala and M. oleifera would become alternatives to supply the lack of food for cattle. For a high biomass production of L. leucocephala, it is necessary to prune regularly, producing palatable, digestible, and nutritious foliage for ruminants and increasing feed intake and rumen fermentation [31]. The forage shrub L. leucocephala was used in the dry season [32]. It is dried to the foliage of L. leucocephala and used as hay to feed dairy cows in the dry season when there is a shortage of grass [33]. On the other hand, in the rainy season, L. leucocephala and M. oleifera also showed superiority in plant height, stem diameter, fresh forage, and dry matter compared to C. argentea and M. alba. Rengsirikul et al. [34] reported 139 cm plant height, 1.10 cm stem diameter, 3.75 t/ha of fresh forage, and 1.68 t/ha of dry matter in L. leucocephala plantations at two years of age. The establishment of L. leucocephala in tropical pastures is a good source of food for livestock, allowing greater availability of food and competing with weeds, which are problems in livestock systems. In the tropics and the rapid growth of the stem diameter of L. leucocephala is an advantage for producers who would cut these legumes at an early stage of development to use them as cattle feed [34,35].
Livestock productivity in the tropics is greatly affected in the dry season by low forage availability and quality [36]. In this study, in the analyses according to season, we found higher fresh forage and dry matter in the rainy compared to dry seasons. C. argentea is a legume that shows a potential source of supplementation in the dry season, especially in acidic soils and prolonged dry seasons [37]. Here, we evaluate L. leucocephala, M. oleifera, C. argentea, and M. alba, which demonstrate good dry matter production in both seasons.
Milk production varied according to the feed source the cows received. We observed higher milk production in cows fed with M. alba, lower production with C. argentea, and higher production in the rainy season. Our findings were lower (5.72 kg/cow per day, cows fed B. brizantha with C. argentea) than those found by Romero and Gonzáles [38], who reported milk yields of 10.9 kg/cow per day in Jersey cows fed Fresh C argentea. In addition, cows fed M. oleifera with B. brizantha produced 7.47 kg/cow per day, whose values exceed those of Reyes et al. [39], who recorded 5.07 kg/cow per day fed B. brizantha with M. oleifera. The forage of M. oleifera contains high amounts of protein, which is reflected in the milk production of cows [40,41]. In crossbred Bos taurus with Bos indicus cows, they obtained 14.1 L/cow per day when fed L. leucocephala [42]. Lamela et al. [43] used M. alba and L. leucocephala to improve milk production in Bos taurus and Bos indicus crossbred cows and obtained an average of 10 L/cow per day. M. alba and M. oleifera have higher nutritional value compared to L. leucocephala; in particular, supplement protein degraded within the rumen [44]. Our findings on milk production are acceptable and are above the average values for Latin America in cows fed in grazing systems (milk production ranges from 2 to 5 kg/cow per day) [45,46]. We found a variation in milk components according to the food source of the cows. Higher acidity was recorded in milk from cows fed M. oleifera and L. leucocephala. According to the NTP 202.001 of Peru, the minimum specification of acidity as lactic acid is 0.13 g/100 g, and the maximum is 0.17 g/100 g of milk [47]. The content of fat and total solids in this research was shown to be higher in cows fed B. brizantha with L. leucocephala compared to the other groups. Regarding the three milk components evaluated, similar results were found in previous studies with similar experimental conditions [38,48,49]. In milk from Jersey cows fed fresh C. argentea, 3.69% fat and 12.47% total solids were observed [38]. In Jersey crossbred cows with Central American Milking Creole grazed in silvopastoral systems of Erythrina poeppigiana associated with Cynodon niemfuensis, Brachiaria rusisiensis, Axonopus compresus, and Paspalum conjugatum pastures, and supplemented with sorghum, milk fat production was 42.2 g/kg, and total solids was 129 g/kg [48]. In Brown Swiss-Zebu crossbred cows grazed in intensive systems of L. leucocephala with Cynodon nlemfuensis, the concentration of fat in the rainy season was 3.3% and in dry was 3.7%, and total solids were 11.3% in rainy and 11.8% in dry [49].
The economic and profitability indicators of this research provide evidence of the economic benefits associated with the use of four forage shrub species (B. brizantha with L. leucocephala, B. brizantha with M. alba, B. brizantha with C. argentea. and B. brizantha with M. oleifera). The tendency to adopt and implement alternative protein banks and silvopastoral systems with forage shrub species in tropical regions of Latin America is increasing due to the economic benefits they generate in livestock due to the increase in the availability and quality of forage throughout the year [50,51]. However, in the Northern Peruvian Amazon, adoption of these systems has been slow despite low-cost food sources. Cow feeding provides additional benefits that are often unseen and unrecorded.

Conclusions
The highest plant height, stem diameter, fresh forage, and dry matter were observed in L. leucocephala and M. oleifera. The highest milk production was observed in cows fed B. brizantha with M. alba, and the highest milk production was in the rainy season. Lactic acid was higher in milk from cows fed B. brizantha with M. oleifera, but a higher concentration of fat and total solids was observed in milk from cows fed B. brizantha with L. leucocephala. The highest utility was observed in cows fed B. brizantha with M. alba; however, the highest operational profitability was observed in the treatment of only B. brizantha and B. brizantha with L. leucocephala. Funding: This research was funded by Project 039_PI "Incrementar los rendimientos de carne y leche a través del mejoramiento de la nutrición del ganado bovino con cuatro especies arbóreas forrajeras en la Región San Martín" of Programa Nacional de Innovación Agraria-PNIA; Laboratorio de Suelos Aguas y Foliares-LABSAF, Proyecto de Inversión en Suelos y Aguas with CUI N • 2487112 and Proyecto "Mejoramiento de la disponibilidad y acceso del material genético mediante el uso de técnicas de biotecnología reproductiva en ganado bovino tropical en las regiones de San Martín, Loreto y Ucayali-PROMEG-Tropical" with CUI N 2338934.

Institutional Review Board Statement: Not applicable.
Informed Consent Statement: Not applicable.

Data Availability Statement:
The data that support the findings of this study are available from the corresponding author, J.A.S.-U., upon reasonable request.