Inclusion of Gliricidia Hay in Total Mixed Rations Silage Made from Giant Cactus Forage

Abstract

This study evaluated the effects of gliricidia hay inclusion in the total mixed rations made from giant forage cactus on the fermentative profile, losses, chemical composition, and aerobic stability. The completely randomized design was adopted with five treatments (0, 5, 10, 15, and 20% of gliricidia hay inclusion on a natural matter-NM basis) and five replications. Dry matter, ether extract, and crude protein exhibited quadratic effects with maximum peaks at 9.33%, 4.94%, and 15.38%. Linear increases were observed on the neutral and acid detergent fiber and hemicellulose, while non-fibrous carbohydrates decreased linearly. The pH showed a linear increase, while ammoniacal nitrogen, propionic, and lactic acids decreased linearly. Acetic acid displayed a quadratic effect with a maximum peak at 11.69%. The minimum silage temperature decreased linearly. Forage losses exhibited quadratic effects with a minimum peak at 8.15%. The effluent, gas, and total losses displayed quadratic effects with minimum peaks at 12.43%, 13.65%, and 11.19%, while dry matter recovery exhibited a maximum peak at 9.34%. The inclusion of up to 15% of gliricidia hay into total mixed rations silages made from giant forage cactus improved the chemical composition and fermentative profile, decreasing forage losses, without promoting changes in the aerobic stability.

1. Introduction

The shortage of feed for ruminants in the semi-arid region leads to lower growth performance in animals. This situation results in underfeeding, also limited by the availability of water, which ultimately leads to low net income for breeders. This issue is more pronounced during the dry season of the year [1]. In this scenario, forage cactus, an exotic plant adapted to the edaphoclimatic conditions of the semi-arid region, has morphophysiological characteristics that enable it to survive. In addition to its high production potential per unit of area, it has an acceptable nutritional value and water content and can even supply part of the water needs of the herds [2].

Forage cactus (Opuntia ficus indica), as mentioned by Alencar et al. [3], is a species of cactus rich in soluble carbohydrates, containing more than 200 g kg⁻¹ dry matter (DM) [4,5]. Furthermore, it is widely cultivated in the semi-arid region of Brazil, covering more than 550,000 ha [6]. Despite this, due to the high water content of forage cactus, combined with low levels of dry matter (8.20% natural matter—NM), fiber (20.6% DM), and protein (5.20% DM), it should be associated with other ingredients for silage production [7]. Therefore, it is possible to complement its nutritional composition and promote the proper functioning and maintenance of the rumen microbiota when used in ruminant diets.

One strategy that has been explored is the use of species resistant to the climatic conditions of the semi-arid region for hay or silage production, thus ensuring the supply of quality food during the period of forage scarcity [8]. The use of appropriate technologies, such as the preservation of forages in the form of silage, is an alternative to overcome the problems arising from food scarcity in arid regions [9]. Due to the low content of DM (8.20% NM), neutral detergent fiber (NDF: 25.97% DM), and crude protein (CP: 5.86% DM) in forage cactus, it is not recommended to use it in isolation in ruminant diets [10].

Given this scenario, over the years, studies have been conducted to assess the association of cactus forage with legumes or grasses for the production of mixed silage made from cactus forage. Among them, it is possible to highlight some forages proper for this process, which are considered crops adapted to semi-arid regions, such as pornunça or cassava (Manihot spp.) and buffel grass (Cenchrus ciliaris L.) [10,11].

Besides these forages, gliricidia (Gliricidia sepium) is a legume abundant in semi-arid regions and nutritionally complements forage cactus. Therefore, the combination of forage cactus with this species adapted to the semi-arid region in the form of hay, in the preparation of mixed silages, may represent an important alternative to improve the nutritional and fermentative characteristics of the ensiled material [12]. In this context, gliricidia hay can be used as an absorbent additive to increase the DM, CP, and fiber contents of forage cactus silages, improving the fermentative and nutritional characteristics of the silages and increasing the efficiency of the ensiling process [13].

In addition, according to Godoi et al. [10], a homogeneous mixture of forage cactus-based silage combined with these tropical forages can reduce the selection of components by the animals. Therefore, this association can increase the productive performance of the animals while also reducing costs compared to conventional diets, as food intake becomes more efficient. The use of the silage technique using gliricidia hay would be an alternative to resolve the issues related to giant forage cactus silage, as this technique aims to combine two or more crops to develop the nutritional and fermentative characteristics of silages.

It was hypothesized that the inclusion of up to 20% NM of gliricidia hay replacing ground corn and cottonseed cake to produce silage of total mixed rations based on giant forage cactus would maintain the nutritive value, fermentative profile, and aerobic stability. Furthermore, this inclusion is expected to promote the reduction in fermentative losses.

Given this scenario, this study aimed to evaluate the fermentative profile and losses, chemical composition, microbial population, and aerobic stability of gliricidia hay replacing ground corn and cottonseed cake to produce silage of total mixed rations based on giant forage cactus.

2. Materials and Methods

2.1. Experiment Location

The forage culture, harvest, and processing were carried out at the Instituto Federal de Educação, Ciência e Tecnologia Baiano, Campus Santa Inês—BA, a municipality that is 300 km from Salvador. According to the Köppen–Geiger climate classification, Santa Inês has a tropical savannah (Aw) climate. Temperatures usually range between 20 °C and 24 °C throughout the year, but they can rarely drop to 12 °C or can rise as high as 34 °C. The average annual rainfall is about 853 mm. Furthermore, in this location, the mini-silos were made and filled.

After filling, sealing, and weighing, the mini-silos were taken and stored in the Forage Laboratory at the Experimental Farm of São Gonçalo dos Campos, belonging to the School of Veterinary Medicine and Animal Science of the Federal University of Bahia. The farm is located at Km 174 of the BR 101 highway, Mercês city, Municipality of São Gonçalo dos Campos—BA (12°23′57.5″ south latitude and 38°52′44.6″ west longitude), located at a distance of 108 km from Salvador.

2.2. Experimental Design, Treatments, and Ensiling

The experiment was conducted using a completely randomized design with five treatments and five replications during the months of June and September 2023. The ingredients used to compound the silages as total mixed rations were giant forage cactus, gliricídia hay, cottonseed cake, and ground corn (

Table 1. Chemical composition of the ingredients used in the production of silage of total mixed rations based on giant forage cactus.

Item, % Dry Matter	Ingredients
	Giant Forage Cactus	Gliricidia Hay	Cottonseed Cake	Ground Corn
Dry matter, % NM	6.84	84.03	88.79	87.77
Ash	11.67	9.52	5.35	2.22
Crude protein	3.58	18.25	26.30	8.61
Ether extract	2.53	3.14	9.75	4.15
Neutral detergent fiber	31.96	42.50	17.19	21.13
Acid detergent fiber	3.10	22.61	9.18	12.11
Non-fibrous carbohydrates	50.26	26.58	41.41	63.89
Total carbohydrates	82.22	69.08	58.60	85.02

).

The forage cactus was harvested manually and processed in a stationary forage machine, providing particles of approximately 2 cm. As for the gliricidia hay, the material for production was harvested from an established plantation of gliricidia approximately 6 months earlier on a farm in Brejões-BA, which is around 45 km from Santa Inês. The material was taken to the IF Baiano, Santa Inês Campus, where it was produced using dehydration by sunlight. It was then ground and stored in an airy shed.

Treatments referred to the mixture of 80% giant forage cactus silage and inclusion levels of gliricidia hay (0, 5, 10, 15, and 20% NM), cottonseed cake, and ground corn. At the time of ensiling, the forage cactus was mixed with the ingredients in the respective proportions of each treatment. Thus, the material was manually homogenized and immediately compacted with the aid of cement rods until it reached a density of 700 kg NM m⁻³.

The experimental diets in the form of a complete total mixture ratio were formulated to meet the nutritional requirements of sheep with an average weight of 20 kg and an average daily gain of 200 g day⁻¹ according to NRC [14]. The ingredient proportions and chemical composition of the treatments are described in

Table 2. Proportions of ingredients used in the production of silage of total mixed rations based on giant forage cactus.

Ingredient, % NM	Gliricidia Hay Inclusion Level, % NM
	0	5	10	15	20
Forage cactus silage	80	80	80	80	80
Ground corn	13	10	6.5	3.5	0
Cottonseed cake	7	5	3.5	1.5	0
Gliricidia hay	0	5	10	15	20
Total	100	100	100	100	100

.

The silages were made in 25 experimental mini-silos of polyvinyl chloride (PVC) (diameter—10 cm and length—30 cm), which were equipped with a Bunsen valve adapted to the lid to allow the elimination of gases resulting from fermentation. At the bottom of each mini-silo, 0.6 kg of dry sand was added, the layer of which was separated from the chopped forage by a non-woven fabric (TNT) bag, for the capture and subsequent measurement of effluent losses. Then, all experimental mini-silos were closed with adhesive tape, weighed, and stored in a covered place at room temperature, and opened at 120 days of ensiling.

2.3. Opening of Mini-Silos, Fermentation Losses, and Dry Matter Recovery

At the time of opening, each closed mini-silo was weighed, as well as the mini-silo was weighed without the lid, and each one of them was weighed after the silage was removed. The dry matter losses in silages in the form of gases (GL), effluents (EL) and total (TL) were quantified using weight difference and the following equations described by Jobim et al. [15]:

GL = [(SWC − SWO)/(FMC × DMCC)] × 100,

where GL = gas losses (%); SWC = total silo weight at closure (kg); SWO = total silo weight at opening (kg); FMC = forage mass at closure (kg); and DMCC = forage DM at closure (%).

EL = [(ESWO − SW) − (ESWC − SW)] × 1000/FMC,

where EL = effluent losses (kg/ton NM); ESWO is the empty silo weight +  sand weight at opening (kg); SW is the empty silo weight (kg); ESWC is the empty silo weight  +  sand weight at closure (kg); and FMC is the forage mass at closure (kg).

TL = [(DMi − DMf)] × 100/DMi,

where TL = Total loss of DM (%); DMi = initial DM. Weight of the silo after filling − weight of the empty set, without forage, before filling (dry tare) × DM content of the forage in the silage and DMf = final DM. Weight of the silo before opening − weight of the empty set, without the forage, after opening the silos (wet tare) × DM content of the forage at the opening.

DMR = (FMO × DMO) × 100/(FMC × DMC)

where DMR = dry matter recovery rate (%); FMO = forage mass at opening (kg); DMO = DM at the opening (%); FMC = forage mass at closure (kg); and DMC = DM at closure (%).

2.4. Sample Collection and Assessment of the Chemical Composition

After evaluating the fermentation losses, silage samples were collected from each mini-silo and submitted to pre-drying in a forced ventilation oven at 55 °C for 72 h (INCT G-001/2). Then, the samples were ground in a knife mill (Tecnal^®, Piracicaba, São Paulo, Brazil) with a 1 mm mesh sieve, stored in sealed plastic bags, and identified for further evaluation of the chemical composition. The ingredients and silage samples were analyzed for evaluations of DM (G-003/1), ash (INCT-CA M-001/2), CP (INCT-CA N-001/2), ether extract (EE; INCT-CA G-004/1), NDF (INCT F-001/2), and acid detergent fiber (ADF: INCT F-003/2) contents following the recommendations described in Detmann et al. [16]. In addition, non-fibrous carbohydrates (NFC) and total carbohydrates (TC) contents were determined according to equations described by Weiss [17] and Sniffen et al. [18], respectively.

2.5. Evaluations of pH, Ammoniacal Nitrogen, and Organic Acids

In addition to the silage samples collected for the evaluations of the bromatological composition, materials were collected from each mini-silo for the evaluation of the fermentative profile. The pH in each mini-silo was measured with the aid of a benchtop pH meter (KASVI^®, São José dos Pinhais, Paraná, Brazil) as described by Bolsen et al. [19]. Thus, 25 g of samples were collected from each mini-silo, which were added to 100 mL of distilled water and submitted to subsequent pH readings after 1 h of rest.

Ammoniacal nitrogen (NH₃-N) concentrations were determined by distillation with potassium hydroxide (KOH) 2N, as proposed by Fenner [20]. Organic acids were quantified by high-performance liquid chromatography (HPLC), according to the methodology described by Canale et al. [21]. Prior to the analysis, the samples were submitted to processing with conservation in 20% metaphosphoric acid and then maintained at −20 °C. Propionic, acetic, lactic, and butyric acids were determined by injecting 20 μL of the samples into a liquid chromatograph equipped with SPD-M20A-UV DETECTOR (SHIMADZU) and AMINEX HP87-H column (Bio-Rad Laboratories Ltd., São Paulo, Brazil) (30 cm × 4.5 mm in diameter, flow rate of 0.6 mL min⁻¹, temperature of 39 °C for 30 min, and pressure of 6.0 MPa).

Although all the previously mentioned acids were evaluated, it was found that the concentrations of propionic and butyric acids were below the detectable levels in all silage samples. Thus, the results of these acids could not be mentioned in the table of results.

2.6. Microbiological Analyses of Silage

The microbial populations were evaluated at the opening of the mini-silos. Thus, the counts of lactic acid bacteria (LAB), molds, and yeasts were isolated using selective culture medium following the methodology described by González and Rodríguez [22].

The LAB populations were quantified using MRS agar (Man, Rogosa, and Sharpe; KASVI^® São José dos Pinhais, Paraná State, Brazil) with the addition of 0.1% acetic acid. Molds and yeasts were quantified using potato dextrose agar (PDA, KASVI^® São José dos Pinhais, Paraná, Brazil) acidified with 1% tartaric acid (weight/volume). Thus, the microbial populations were quantified from samples obtained from the homogenization of 10 g of silage sample, 90 mL of distilled water for 1 min.

Thus, the aqueous extract of the silage was filtered, and 9 mL were added to plastic flasks containing sterile distilled water. Subsequently, this material was subjected to serial dilutions (10⁻¹ to 10⁻⁶) and then plated in duplicate for each dilution performed. The plates were incubated in B.O.D (Biochemical Oxygen Demand) at 37 °C for 48 h for lactic acid bacteria (LAB) and at 37 °C for 72 h for mold and yeast populations. Molds and yeasts were differentiated with the aid of morphological characteristics of the colonies.

After the incubation period in B.O.D. at the times mentioned above, the colonies were counted. Thus, countable plates were those with values between 30 and 300 CFU (colony-forming units), and the averages of the plates of the selected dilution were calculated. After the counting, the values were converted to a logarithmic scale (base log10).

2.7. Assessment of Aerobic Stability

Aerobic stability was evaluated by returning 1.0 kg of the silage sample to its respective mini-silo in a closed environment with controlled temperature (25 °C). Thus, the samples were allocated without compacting the material and kept uncovered to allow air infiltration into the silage mass.

The internal temperatures of the silages were evaluated every 2 h with the aid of a skewer thermometer (KASVI^® São José dos Pinhais, Paraná, Brazil) inserted in the geometric center of the mass for 120 h. In addition, the ambient temperature was measured at the same time as the evaluation of the silage temperatures, using a digital laser thermometer. Thus, the following variables were determined: maximum and minimum temperatures (°C), thermal amplitude, forage losses (% NM) and aerobic stability (hour). The beginning of deterioration was considered when the internal temperature of the silages reached 2 °C above the ambient temperature [23].

2.8. Statistical Analysis

The experiment was conducted in a completely randomized design with five treatments and five replications, totaling 25 experimental units. Thus, the following statistical model was used:

Ŷij = μ + Ti + Ɛij,

where Ŷij = observed value of the dependent variable; μ = overall average; Ti = fixed effect of gliricidia hay inclusion; Εij = experimental random error associated with each assumption observation NID~(0, σ²).

The data were submitted to an analysis of variance and regression. Thus, the degrees of freedom were decomposed by orthogonal contrasts in linear and quadratic effects, according to the levels of inclusion of gliricidia hay in the silage of giant forage cactus. The significance of the regressions was determined at the 5% probability level for type-I error, using the PROC MIXED of the SAS 9.4 statistical package program.

3. Results

3.1. Chemical Composition of Silages

The concentrations of DM (p < 0.001), ash (p < 0.001), CP (p < 0.001), EE (p < 0.001), and TC (p = 0.004) were adjusted to the quadratic regression model (

Table 3. Chemical composition of the silage of total mixed rations based on giant forage cactus.

Ingredients	Gliricidia Hay Inclusion Level, % NM					SEM 1	p-Value
	0	5	10	15	20		Linear	Quadratic
Dry matter 2	18.7	26.2	22.2	21.9	19.3	1.25	0.043	<0.001
Organic matter 3	6.42	6.11	8.56	10.3	11.9	1.02	<0.001	<0.001
Crude protein 4	12.7	15.5	16.7	16.9	17.0	0.79	<0.001	<0.001
Ether extract 5	5.72	5.56	4.31	3.14	1.72	0.73	<0.001	<0.001
Neutral detergent fiber 6	16.3	21.6	29.7	34.0	39.7	3.77	<0.001	0.226
Acid detergent fiber 7	8.65	11.9	17.7	20.5	26.1	2.81	<0.001	0.490
Hemicellulose 8	7.68	9.65	12.1	12.3	13.6	1.13	<0.001	0.126
Non-fibrous carbohydrates 9	58.8	51.2	40.8	36.2	29.7	4.76	<0.001	0.041
Total carbohydrates 10	75.1	72.8	70.5	70.2	69.4	1.10	<0.001	0.004

¹ Standard error of the mean; equations: ² Ŷ = 19.937 + 0.0785x − 0.0471x² (R² = 0.57); ³ Ŷ = 6.0697 + 0.1256x + 0.009x² (R² = 0.96); ⁴ Ŷ = 12.887 + 0.5599x − 0.0182x² (R² = 0.98); ⁵ Ŷ = 5.8264 − 0.0691x − 0.0072x² (R² = 0.99); ⁶ Ŷ = 16.444 + 1.1818x (R² = 0.99); ⁷ Ŷ = 8.2816 + 0.8677x (R² = 0.99); ⁸ Ŷ = 8.1626 + 9.2897x (R² = 0.94); ⁹ Ŷ = 57.965 − 1.464x (R² = 0.98); ¹⁰ Ŷ = 75.138 − 0.5737x + 0.0146x² (R² = 0.98).

). Using the generated equations, there were maximum DM, EE, and CP concentrations of 24.3%, 5.66%, and 17.19% at the inclusion levels of 9.33%, 4.94%, and 15.38% of gliricidia hay in the giant forage cactus silages, respectively. On the other hand, a minimum ash concentration of 5.63% was estimated with the use of 6.98% of gliricidia hay in the silage.

Neutral detergent fiber (p < 0.001), acid detergent fiber (p < 0.001), and hemicellulose (p < 0.001) concentrations increased linearly as gliricidia hay was added to the giant forage cactus silage (Table 3). Thus, for each percentage unit of inclusion of gliricidia hay, there was an increase of 1.18, 0.87, and 0.29% in the NDF, ADF, and hemicellulose contents in giant forage cactus silage, respectively.

On the other hand, the inclusion of gliricidia hay resulted in a linear decrease (p < 0.001) in NFC concentrations in the giant forage cactus silage (Table 3). Thus, for each 1% inclusion of gliricidia hay, there was a reduction of 1.46% in the NCF concentrations in the silage.

3.2. Fermentation Profile and Microbial Population Counts in Silages

The inclusion of gliricidia hay in giant forage cactus silages resulted in a linear increase (p = 0.005) in pH (

Table 4. Fermentative profile and microbiology of the silage of total mixed rations based on giant forage cactus.

Ingredients	Gliricidia Hay Inclusion Level, % NM					SEM 1	p-Value
	0	5	10	15	20		Linear	Quadratic
pH 2	3.79	3.76	3.83	3.88	4.02	0.06	0.005	0.219
Ammoniacal nitrogen (% TN) 3	2.32	1.84	1.79	1.47	1.40	0.29	0.044	0.755
Organic acids (mg ml−1 DM)
Acetic 4	0.457	0.978	0.797	0.802	0.781	0.08	0.043	<0.001
Propionic 5	0.624	0.835	0.544	0.470	0.425	0.09	<0.001	0.411
Lactic 6	1.90	3.34	1.39	1.74	1.32	0.39	<0.001	0.071
LAB (log 10 CFU g−1 NM)	3.71	3.69	4.07	3.85	3.80	0.06	0.442	0.154

¹ Standard error of the mean; equations: ² Ŷ = 3.7388 + 0.0117x (R² = 0.83); ³ Ŷ = 2.2071 − 0.0442x (R² = 0.91); ⁴ Ŷ = 0.5407 + 0.0608x − 0.0026x² (R² = 0.57); ⁵ Ŷ = 0.7322 − 0.0153x (R² = 0.56); ⁶ Ŷ = 2.4902 − 0.0553x (R² = 0.28).

). Thus, for each 1% inclusion of gliricidia hay, there was an increase of 0.011 units in pH values. On the other hand, concentrations of ammonia nitrogen (p = 0.044), propionic acid (p < 0.001), and lactic acid (p < 0.001) decreased linearly by 0.04%, 0.02 mg ml⁻¹ DM, and 0.06 mg ml⁻¹ DM for each percentage unit of inclusion of gliricidia hay in the silages.

In addition, a quadratic effect was observed for acetic acid, with a maximum concentration of 0.896 mg ml⁻¹ DM at the inclusion level of 11.69% of gliricidia hay in the giant forage cactus silages (Table 4). However, the LAB counts were not significantly influenced (p > 0.05) by the addition of gliricidia hay in the giant forage cactus silage (Table 4).

3.3. Aerobic Exposure

Maximum temperature, thermal amplitude, and aerobic stability were not significantly influenced (p > 0.05) by the addition of gliricidia hay in the giant forage cactus silage (

Table 5. Aerobic stability of the silage of total mixed rations based on giant forage cactus.

Ingredients	Gliricidia Hay Inclusion Level, % NM					SEM 1	p-Value
	0	5	10	15	20		Linear	Quadratic
Temperature (°C)
Maximum	26.2	27.1	25.8	25.8	25.9	0.33	0.154	0.888
Minimum 2	23.2	23.2	22.7	22.9	22.8	0.16	0.017	0.196
Thermal amplitude	3.16	3.9	3.34	3.12	3.06	0.34	0.486	0.449
Forage losses (%NM) 3	21.3	12.13	10.1	8.92	9.09	2.17	<0.001	<0.001
Aerobic stability (hour)	120.0	120.0	120.0	120.0	120.0	-	-	-

¹ Standard error of the mean; equations: ² Ŷ=23.16 − 0.0228x (R² = 0.58); ³ Ŷ = 20.613 − 1.6662x + 0.0557x² (R² = 0.96).

). Therefore, it was observed that all silages showed aerobic stability with times greater than 120 h.

The minimum silage temperature decreased linearly (p = 0.017) by 0.02 °C for each 1% inclusion of gliricidia hay in the giant forage cactus silage (Table 5). In addition, forage losses (p < 0.001) were adjusted to the quadratic regression model. Thus, the lowest forage losses (14.96%) were observed with the inclusion of 8.15% of gliricidia hay in the giant forage cactus silages.

3.4. Fermentation Losses During Ensiling and DMR of Silages

The effluent (p < 0.001), gas (p < 0.001), and total (p < 0.001) losses showed negative quadratic effects as gliricidia hay was included in the silages (

Table 6. Fermentative losses of the silage of total mixed rations based on giant forage cactus.

Ingredients	Gliricidia Hay Inclusion Level, % NM					SEM 1	p-Value
	0	5	10	15	20		Linear	Quadratic
Losses
Effluents (kg/ton NM) 2	98.9	58.0	39.0	40.7	59.2	18.82	<0.001	<0.001
Gases (%) 3	4.12	0.99	1.19	0.91	1.03	0.59	<0.001	<0.001
Total (%) 4	13.32	6.79	6.84	6.40	10.1	2.06	0.115	<0.001
Dry matter recovery (% DM) 5	86.2	90.3	94.7	89.4	83.9	3.38	0.486	0.003

¹ Standard error of the mean; equations: ² Ŷ = 98.468 − 9.918x + 0.3991x² (R² = 0.99); ³ Ŷ = 3.764 − 0.4697x + 0.0172x² (R² = 0.85); ⁴ Ŷ = 12.909 − 1.2776x + 0.0571x² (R² = 0.93); ⁵ Ŷ = 85.834 + 1.553x − 0.0831x² (R² = 0.99).

). Therefore, minimum effluents, gas, and total losses of 36.85%, 0.56%, and 5.76% were estimated when 12.43%, 13.65%, and 11.19% of gliricidia hay were included in giant forage cactus silage, respectively. In contrast, the DMR presented a positive quadratic effect (p = 0.003), being the highest DMR of 93.09% estimated when 9.34% of gliricidia hay was included in the silage of total mixed rations based on giant forage cactus.

4. Discussion

4.1. Chemical Composition

The gliricidia hay inclusion promoted positive quadratic responses on the DM content in the silage of total mixed rations based on giant forage cactus, with higher values being observed in silages containing hay, compared to those without the use of the additive. Despite this, the silages presented DM levels lower than 30 to 35%, which were below the recommended values described by McDonald et al. [24].

This result is expected, since gliricidia hay has been used in silage as a moisture-absorbing additive, because it has high dry matter contents, resulting in an increase in the DM contents of silages. Furthermore, it is important to highlight that the forage cactus has mucilage, a hydrocolloid substance, in its composition, which is composed of glycoprotein and organic acids, promoting the water retention capacity, which avoids the extreme loss of water due to the production of an emulsifying gel [25].

As highlighted by Kung Jr. et al. [26], plants with DM contents lower than 25% tend to present development and clostridial activity during the fermentation process, which results in high concentrations of ammonia nitrogen in silages due to the breakdown of proteins into ammonia, in addition to the production of butyric acid. However, this did not happen in this study because all silages presented ammonia nitrogen levels below 10%, recommended by McDonald et al. [24], to obtain good quality silages and an adequate fermentation process.

The results of this study corroborate what was verified in other studies developed with forage cactus silages and based on forage cactus [27]. Therefore, the results may be associated with the adequate fermentation process of the mixed silages, which possibly inhibited clostridial or enterobacteria activities, promoting a reduction in proteolysis or few fermentative losses of DM in the silages [28].

The ash concentrations were quadratically influenced by the inclusion of gliricidia hay in giant forage cactus silages. This result may be associated with the behavior observed in the DM contents, which ended up influencing the mineral fraction of the silages in all the treatments evaluated, or because of the different proportions and levels of ash in the ingredients in the concentrates of the treatments. In addition, the ash contents may have been influenced due to the contents of this fraction in forage cactus and gliricidia hay, corroborating what was verified by De Sá et al. [29].

Crude protein contents were positively quadratically influenced by the inclusion of gliricidia hay. This behavior possibly corroborates what was expected, given that gliricidia hay has higher CP levels in relation to the other ingredients. In addition, the variations observed in the CP contents of the silages may be associated with the fermentation process and effects on the protein fraction, associated with the fact that different inclusions of ingredients in the concentrates were made in the treatments, which have different CP contents.

Despite the effects observed, all silages in this study presented CP levels higher than the minimum necessary to ensure rumen fermentation. As highlighted by the NRC [30], when the CP levels of the feed offered to small ruminants are less than 7%, they have low nitrogen availability. Therefore, because all silages had CP levels above 13%, they presented adequate values for a diet that meets the requirements for lambs, as indicated by the NRC [14], which recommends around 130 g CP kg⁻¹ DM for live weight gain of 200 g day⁻¹.

Similarly to the CP content, the EE content of the silages was quadratically influenced by the addition of gliricidia hay. The results corroborate the behaviors verified by Brito et al. [27] and De Sá et al. [30], who also observed changes in EE levels in forage cactus silages with gliricidia hay levels. This result was possibly due to the behavior also observed in the DM contents, which was quadratically influenced. In addition, it may have occurred due to changes in the proportions of ingredients used in concentrates in giant palm-based silages, which have different levels of EE in their chemical composition, in association with the effects of the fermentation process on nutrients.

Giant forage cactus silages without inclusion and with 5% inclusion of gliricidia hay presented EE contents slightly higher than 5%. As highlighted by Bionaz et al. [31] and Marques et al. [32], EE content below this value is considered beneficial so that feed intake by ruminants is not limited and there is toxicity to rumen microorganisms. Thus, in this study, forage cactus silages containing 10% of gliricidia hay or more were composed of satisfactory EE content to be offered to ruminants, as they presented energy concentration within the recommended levels so as not to limit ruminant intake in a chemostatic way.

The ensilage of giant forage cactus with gliricidia hay increased the contents of NDF, ADF, and hemicellulose compared to silages that did not use this additive. This result, as well as the other nutrients, was expected because the hay used in this study had higher NDF and ADF levels than forage cactus, which, consequently, contributed to influencing the levels of these nutrients in the silages, as well as the levels of hemicellulose. A similar behavior was found by Sá et al. [28], who mentioned that the higher levels of NDF and ADF in gliricidia hay contributed to the increase in these components in the silages evaluated. Despite the observed increases, the NDF and ADF values obtained in all the silages studied were below the maximum limits of these fibrous compounds recommended by Van Soest [33] for diets for small ruminants, which are 60% and 40%, respectively.

In contrast to NDF and ADF contents, which increased as gliricidia hay was added to giant forage cactus silage, NCF contents decreased linearly. This result can be justified by the increase in fibrous fractions, which, consequently, resulted in a reduction in NCF levels in silages in this study. Regarding TC, the contents were negatively quadratically influenced by the inclusion of gliricidia hay in giant forage cactus silages. These results may be associated with changes in the proportions of ingredients in the concentrates used in each treatment, which may have influenced silage levels.

In view of the results observed, it is possible to conclude that the chemical composition of the mixed forage cactus silages with gliricidia hay, in association with other ingredients in the concentrate, was influenced by the concentrations of the nutrients of the plants used, as well as the ingredients used, corroborating Brito et al. [27]. Therefore, there may have been an interaction between the nutrients used in each treatment, which may have resulted in changes in the development of microorganisms, and, in turn, changes in the fermentation process of the silages.

4.2. Fermentative Profile

Silage of giant forage cactus with gliricidia hay increased the pH values of silages compared to those without inclusion of the additive, probably because there was an increase in the buffering capacity. Thus, similar to all legumes, gliricidia has high levels of orthophosphate and salts of organic acids, in addition to the high protein content, which makes it difficult to lower the pH in the material with impacts on the buffering capacity of the silages [34].

The results of this study corroborate the same behaviors verified by Brito et al. [27] and De Sá et al. [29] when ensiling forage cactus with gliricidia hay and verified an increase in the pH of silages due to the use of this additive. Despite the increase in pH, the values can be considered close to or within the ideal for silages with an adequate fermentation pattern, as highlighted by McDonald et al. [24], varying from 3.8 to 4.2.

Furthermore, as highlighted by Brito et al. [27], legumes are generally resistant to pH reduction in silages due to their high buffering capacity, mainly due to the presence of cations (K⁺, Ca²⁺, and Mg²⁺). Pirhofer-Walzl et al. [35] also highlighted that these cations come into contact with the organic acids formed by fermentation, neutralizing them and preventing the occurrence of a drop in pH. According to Muck [36], the greater the buffering capacity, the greater the amount of lactic acid that must be formed so that the pH reaches sufficient levels to inhibit the clostridial and enterobacteria activities, which impair the quality of the silage.

On the other hand, NH₃-N concentrations reduced as gliricidia hay was included in giant forage cactus silages, behaviors similar to those described by Brito et al. [27] and De Sá et al. [29]. Thus, the highest values were observed in the silages with the inclusion of gliricidia hay, while the lowest values were observed in the silages with 20% of gliricidia hay. Therefore, the verified results can be considered satisfactory because in all treatments the values were less than 10%, as recommended by McDonald et al. [24], which is indicative of the lack of excessive breakdown of proteins in ammonia and characterizes the adequate fermentation of the silages.

According to Pacheco et al. [37], silage can be classified according to ammonia nitrogen content in relation to total nitrogen as very good (values below 10%), adequate (between 10 and 15%), acceptable (between 15 and 20%), and unsatisfactory (when values are above 20%). Thus, based on the results verified, it is possible to verify that the forage cactus silages in this study can be classified as very good.

The concentrations of NH₃-N in the silage, as highlighted by Sá et al. [38], are a parameter of great importance, as they indicate the loss of protein, an essential nutrient in the diet of ruminants. In addition, as mentioned by the authors, they indicate a higher intensity of proteolysis, mainly due to the degradation of amino acids by proteolytic clostridia. Thus, excessive values of NH₃-N (above 10%) cause low animal acceptability and, consequently, low animal performance. Therefore, the forage cactus silages in this study could be recommended for intake and would present acceptability by ruminants, since they did not present high levels of NH₃-N.

Several organic acids are produced during silage fermentation (lactic, acetic, butyric, isobutyric, propionic, valeric, isovaleric, succinic, and formic acids) [24]. Nevertheless, to evaluate the quality of fermentation, the most commonly used acids are acetic, propionic, lactic, and butyric. The ensiling of the giant forage cactus with increasing levels of gliricidia hay quadratically influenced the concentrations of acetic acid. In general, all silages presented AA concentrations below 2.0%, which, according to Roth and Undersander [39], is considered the maximum limit for the production of good quality silage.

As described by Muck and Bolsen [40], the acetic acid content is related to the lowest decrease rates and higher final pH values in silages, corresponding mainly to the prolonged action of heterofermentative lactic acid bacteria and enterobacteria. Therefore, this result may be associated with the predominance of the Weissella genus, which is a heterofermentative lactic bacteria that has been described in forage cactus silages [41]. In addition, this result may have occurred due to the inclusion of gliricidia, which is a legume and has high CP levels that can make it difficult to reduce the pH of silages, in relation to forage cactus silage without the use of this additive.

Propionic acid concentrations decreased linearly as gliricidia hay was added to giant forage cactus silages. In addition, it was found in this study that only giant forage cactus silages with up to 15% of gliricidia hay inclusion had propionic acid concentrations below the 0.5% recommended by Roth and Undersander [39]. According to these authors, these levels are indicative of well-fermented silages, with higher values indicative of lactic acid degradation. As described by Muck [36], propionic acid occurs on a smaller scale in silage and is produced by heterofermentative propionic bacteria and LAB with antifungal potential during the initial and final phases of silage. Therefore, the presence of this acid in silage has beneficial effects and can effectively contribute to the control of undesirable microorganisms, such as molds and yeasts.

Regarding the concentrations of lactic acid, it was also observed that there was a linear reduction as gliricidia hay was included in the giant forage cactus silage. Based on the classification criteria established by Roth and Undersander [39], silages with lactic acid values of 4.0 to 6.0% DM are considered of good quality. In this study, the values determined in giant forage cactus silages with increasing levels of gliricidia hay inclusion showed lactic acid concentrations ranging from 3.34 to 1.32 mg/mL of DM, which allows us to conclude that the silages can be classified as good quality.

Therefore, the values of lactic acid determined in this study were higher than those of acetic acid, which corroborates the statements of Kung Jr. et al. [26]. According to these authors, lactic acid should be the main acid in good quality silage and that the lactic acid content should be higher than the remaining acids (acetic, propionic, butyric) because although all the acids produced during fermentation contribute to reducing the pH of the silage, lactic acid plays a fundamental role in this process because it has a higher dissociation constant than other acids [42]. Therefore, the results of this study corroborate what is described in the scientific literature, as the concentrations of lactic acid in the forage cactus silages were higher than the concentrations of acetic acid and propionic acid, regardless of the inclusion or not of gliricidia hay in the silages, suggesting an adequate fermentation process.

Although in this study the concentrations of butyric acids in giant forage cactus silages were evaluated, significant levels in silages were not quantified, regardless of the levels of gliricidia hay inclusion. Thus, the low concentrations observed prove that there was no effective clostridic fermentation and indicate good quality of the silages, since low counts of mold and/or yeast development were detected in the microbiological analyses.

These results corroborate what was highlighted by Roth and Undersander [38], who mentioned that butyric acid levels lower than 1% indicate desirable fermentation. Thus, it is possible to conclude the low activity of bacteria of the Clostridium genus that present proteolytic activity.

4.3. Microbial Population Counts in Silages

Regarding the counts of microbial populations, it was observed that the inclusion of gliricidia hay was not sufficient to promote changes in the growth of LAB. Furthermore, although a microbiological evaluation of molds and yeasts was carried out, in none of the treatments evaluated were there sufficient counts of these microorganisms.

The results observed in this study may be associated with the adequate fermentation process in silages promoted by forage cactus, regardless of the inclusion of hay, since there was no difference between treatments in LAB growth, and there was no sufficient quantification of mold and/or yeast populations in the silages. In addition to the satisfactory production of lactic acid, it was also observed that there was adequate production of acetic acid in the silages. Possibly, the absence of mold and/or yeast growth may be associated with the production of this acid, which has antifungal characteristics, inhibiting the growth of these microorganisms and improving silage quality. The results of this study corroborate what was observed by Pereira et al. [41], who selected LAB strains isolated from the plant and forage cactus silage. From the results obtained, the authors observed a predominance of heterofermentative LABs, such as (Weissella cibaria, W. confusa, and W. paramesenteroides). As highlighted by Schmidt et al. [43], these microorganisms are capable of producing antifungal compounds, mainly acetic acid, and allow the associated growth of LAB populations that produce other organic acids, such as propionic acid.

The reduction in the population of these microorganisms may also be related to the buffering substances present in forage cactus, such as oxalic, malic, citric, malonic, succinic, and tartaric acids [44], which can prevent the abrupt drop in the pH of the ensiled mass, as well as control the growth of yeast populations during aerobic exposure of the silage. This effect was also observed by Brito et al. [27], who evaluated mixed silages of gliricidia and forage cactus, where the inclusion of forage cactus controlled the populations of fungi and yeasts.

4.4. Aerobic Stability

The inclusion of gliricidia hay in the production of cactus pear silage did not promote changes in maximum temperatures, thermal amplitude, or aerobic stability. Therefore, it is possible to conclude that all cactus pear silages, with or without the inclusion of gliricidia hay, showed the same propensity for aerobic deterioration up to 120 h.

Similarly to this study, Brito et al. [27] and Sá et al. [28] also did not observe a break in aerobic stability in silages based on cactus pear. This result may be related to the microbial characteristics of the silages because there was low or insignificant growth of molds or yeasts, which are considered the main microorganisms associated with silage deterioration.

Pereira et al. [41] identified in their research with forage cactus the predominance of lactic bacteria (LAB) of the genus Weissella. This microorganism is classified as an obligate heterofermentative LAB. The predominance of heterolactic fermentation has been attributed to the presence of buffering substances in forage cactus, which promote a less abrupt drop in pH, enabling the proliferation of heterofermentative LAB [28,41]. Obligate heterolactic species improve aerobic stability due to greater production of acetic acid [45]. Thus, these effects may have possibly occurred in the present study and enabled aerobic stability in the silages evaluated, regardless of the level of inclusion of gliricidia hay.

Therefore, since there was little mold growth in the silages, this may have contributed to the results observed. As highlighted by Woolford [46], molds are indicated as the first microorganisms associated with silage deterioration. Consequently, they make the environment favorable to the development of other aerobic microorganisms involved in the aerobic deterioration of the ensiled mass.

When the silo is opened, exposure to oxygen causes an increase in temperature due to exothermic reactions, such as respiration and multiplication of microorganisms, which degrade the silage [15]. On the other hand, there was a reduction in the minimum temperature of the silage in addition to a negative quadratic behavior of forage losses as gliricidia hay.

Although no aerobic deterioration was observed in the silages, forage losses were still observed during the 120 h test, demonstrating that the final quality of the material was modified and that there were possibly nutrient losses. Thus, greater DM losses during the aerobic stability test were observed in the forage cactus silage without the inclusion of gliricidia hay, compared to the others in which this additive was included.

Given these results, it is possible to conclude that gliricidia hay showed the potential to minimize forage losses in giant forage cactus silage, compared to those silages without the use of this additive. The same behavior was observed in fermentation losses, where better results were observed in silages with hay, in relation to silages without the use of this additive.

4.5. Fermentative Losses

The inclusion of gliricidia hay in total mixed rations silage made from giant cactus forage promoted quadratic effects on fermentative losses being observed, negative responses on effluent, gases, and total losses, whereas positive responses on the dry matter recovery.

De Sá et al. [29], based on the results found in studies of the inclusion of gliricidia hay in cactus silage, observed the reduction in effluent losses. Thus, it is inferred that gliricidia hay favors the fermentation process, reducing losses and favoring the recovery of dry matter from silages. Similarly, Brito et al. [27] reported a reduction in fermentation losses with greater dry matter recovery when including different levels of gliricidia in forage cactus silages.

Therefore, based on the fermentative profile and losses, chemical composition, and aerobic stability, mixed silages of cactus pear and gliricidia showed adequate fermentative, chemical composition, and aerobic stability response characteristics. Nevertheless, it is suggested to include up to a level of 15% in giant forage cactus silage to promote lower fermentative losses in the ensiled mass. Consequently, it is important that further studies be carried out to assess the intake, digestibility, growth performance, health parameters, carcass traits, and meat quality of feedlot lambs or goats fed with diets based on mixed silages of giant forage cactus and gliricidia hay.

5. Conclusions

The inclusion of up to 15% of gliricidia hay replacing corn and cottonseed cake (%NM) in the production of silage of total mixed rations based on giant forage cactus maintains the chemical composition and fermentative profile without promoting changes in the aerobic stability. Furthermore, its inclusion up to this level is suggested because it promotes lower forage losses in the production of silage of total mixed rations based on giant forage cactus.