RESEARCH WORK

 

Ecosystem with Leucaena leucocephala: its effect on the rumen microbial population in fattening bulls

 

 

Juana L. Galindo-Blanco, Idalmis Rodríguez-García, Niurca González-Ybarra, Roberto García- López and Magaly Herrera-Villafranca

Instituto de Ciencia Animal, Carretera Central km 47 ½, San José de las Lajas, Mayabeque, Cuba CP 32700
E-mail: [e-mail]

 

 

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ABSTRACT

A trial was conducted in order to evaluate the effect of Leucaena leucocephala cv. Peru on the rumen microbial population of Zebu crossbred bulls, in the two stages of the fattening cycle. The system, integrated by five strips of grasses associated with L. leucocephala and five without association, consecutively alternated in 25 ha, allowed the fattening of 80 animals, from which eight were used for the microbiological studies. The samplings of rumen liquid were made bimonthly (February-March, April-May, June-July) and four-monthly (August-November). The cellulolytic bacteria reached values between 15,94 and 46,13 10⁶ CFU.mL^-1 when the bulls grazed L. leucocephala associated with grasses. The highest population of rumen cellulolytic fungi was found in the rumen of animals in the system with L. leucocephala, in the August-November samplings. The concentration of rumen ammonia had values of 8,76 and 24,09 mmol.L^-1 in the system with grasses and the one of L. leucocephala associated with grasses, respectively (p < 0,001). The pH in rumen was significantly higher, the population of total viable bacteria was 2,9 times higher and the protozoans decreased when L. leucocephala participated in the ration. It is concluded that the system with L. leucocephala associated with grasses is advantageous, because it propitiates increases in the populations of total viable bacteria and fiber-degrading microorganisms; this, along with the depressing effect of protozoans, guarantees a favorable ecological environment for rumen cellulolysis, which would contribute in a sustained way to the productive increase of the animal husbandry system.

Keywords: bacterium, metabolites, protozoans.

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INTRODUCTION

L. leucocephala is one of the most widely utilized legumes in Cuba (Ruiz, 2005), and among the qualities that distinguish it is its high protein content (Ku-Vera, 2013). In works conducted by Galindo et al. (2007) it was proven that L. leucocephala can be included in 100 % of a grassland area, which increases rumen cellulolysis as consequence of a higher population of fibrolytic organisms. Likewise, Galindo (2013) reported that L. leucocephala reduces the methane production, mainly due to the presence of secondary metabolites. Prieto-Manrique (2017), when using 14-16 % of L. leucocephala in grazing cows, considered that these quantities were insufficient to reduce the methane production at rumen level.

Iraola et al. (2016), when implementing an agroecological strategy of transformation in a system of degraded pasturelands with a silvopastoral arrangement with L. leucocephala for cattle fattening, found increases in the plant biomass of 60 % and a mean daily gain (MDG) between 0,648 and 0,769 kg/animal/day, with which economically viable systems in Cuban pesos were achieved.

In previous studies conducted by Iraola et al. (2014) in a silvopastoral system with L. leucocephala, the effect of management or season on the indicators related to the rumen microbial ecology was not studied. For such reason, the objective of this study was to evaluate the effect of Leucaena leucocephala cv. Peru on the rumen microbial population of Zebu crossbred bulls, in the two stages of the fattening cycle.

 

MATERIALS AND METHODS

The trial was conducted in the Ayala Feedlot of the Institute of Animal Science Mayabeque, Cuba, in an agroecosystem of pasturelands integrated by a mixture of natural and cultivated pastures with L. leucocephala cv. Peru, in a physical surface of 25 ha on a Brown soil with carbonates.

The design of the system corresponded to the strategy of ecological redesign for the recovery of degraded pastures in an agrosilvopastoral system, according to the report by Iraola et al. (2014), and its purpose was to improve the stocking rate capacity with fattening cattle. The system was structured according to the methodology proposed by Ruiz (2005), with L. leucocephala densities between 5 000 and 5 700 trees per hectare. Once L. leucocephala reached between 1,00 and 1,20 m of height, it began to be utilized.

In the experiment 80 Zebu crossbred bulls were used, of 199 ± 3 kg of average live weight (LW). From them eight animals were randomly selected to conduct the rumen fermentation studies. All the animals grazed in a unique group and passed successively through the five strips of grasses associated with L. leucocephala and the five strips of grasses without association.

The samplings of rumen liquid were made with bimonthly (February-March, April-May, June-July) and four-monthly frequency (August-November) for finishing fattening, through the esophageal pathway and with the aid of a probe, which was introduced through the mouth to the rumen. The animals entered the system 15 days before the first sampling, that is, in the second fortnight of January, in order to guarantee the adequate adaptation period.

Microbiological and fermentative analyses. The culture technique proposed by Hungate (1950) was used, in roll tubes and under strict anaerobiosis conditions. The sowing of total viable bacteria, proteolytic and cellulolytic, was carried out in Caldwell and Bryant (1966) culture media; the fungi were analyzed through the technique suggested by Joblin (1981); and the protozoans were counted under microscope, in Neubauer chamber.

The pH was determined in an EIL model 22-A pH meter, and the ammonia concentration, according to Conway (1957). The analysis of the chemical composition of the pastures was made through the techniques described by AOAC (1995); while the fibrous fractions were analyzed through the procedure proposed by Goering and Van Soest (1970). The chemical composition of the species is shown in table 1.

The secondary metabolites against the qualitative tests of the phytochemical sieving of L. leucocephala indicated high presence (+++) of tannins, alkaloids and anthocyanidins; and moderate quantities (++) of saponins, reducers, flavonoids, triterpenes and steroids.

Experimental design and statistical analysis. Multifactorial variance analysis was carried out. The considered factors were: 1) strips or grazing in grass associated with L. leucocephala and in mixture of grasses, and 2) sampling bimester (February-March, April-May, June-July) and four-month period (August-November).

In the case of the indicators proteolytic, cellulolytic bacteria and cellulolytic fungi a split plot design was used, where the main plot was the strip (L. leucocephala or mixture of grasses), and the subplot, the sampling bimesters. In the case of the indicators total viable bacteria, protozoans, pH and NH₃ a linear model was used for the effects of the strips and sampling bimesters.

It was verified that the data fulfilled the assumptions of normal distribution and variance homogeneity. The counts of viable microorganisms were transformed according to Ln N, to guarantee the normality conditions. For the analysis the formula (K+N).10^x was applied, where: K is the constant that represents the logarithm of the dilution in which the microorganism was inoculated; N is the logarithm of the count of colonies determined as CFU/mL, TFU/mL, or cells/mL; 10 is the basis of the logarithm; and x is the dilution at which the inoculation was carried out.

For the comparison between means Duncan's test (p < 0,05) was applied in the necessary cases. The statistical package INFOSTAT version 2001, by Balzarini et al. (2001) was used.

 

RESULTS AND DISCUSSION

L. leucocephala acquires particular importance in agroforestry systems because, besides its good growth in tropical environments, it shows high contents of protein, minerals, vitamins and energy, which guarantees an adequate nutritional value when it is used in the diet of animals.

In this study significant interaction was found between the grazing system and the sampling moment on the populations of proteolytic, cellulolytic bacteria and fungi of the rumen (table 2). The proteolytic bacteria were more abundant when the animals grazed L. leucocephala. Such results confirm the presence of factors that stimulate the growth of these microbial groups when L. leucocephala is present, due to its high content of protein and minerals (Rodríguez et al., 2013). However, when the animals grazed the grasses during the two-month period February-March and the four-month period August-November, there were no statistical differences in this indicator, aspect that should be object of study in later stages.

L. leucocephala produced a beneficial effect on the populations of cellulolytic bacteria and fungi of the rumen. In all the samplings higher population of cellulolytic bacteria was found when the animals grazed in the system with this plant, with the highest values (p < 0,001) in the samplings comprised between April and November, and within them, the four-month period August-November showed the highest populations. These high populations of cellulolytic organisms can lead to reach higher rumen cellulolysis, due to a higher population of microorganisms capable of excreting the complex of cellulase enzymes; likewise, provide the rumen of necessary metabolites for the synthesis of microbial proteins (Galindo et al., 2017); and, consequently, increases should be expected in the degradation of the fibrous fraction, given the interaction that appears at rumen level among the different microorganisms in the fiber degradation process (Gerber et al., 2013).

The experimental animals were exposed to better nutritional conditions in these months (four-month period August-November), because it has been reported that L. leucocephala is three times richer in protein (22,3-30,0 %) than tropical grasses in the same time of the year (Rivera et al., 2015).

No significant interaction was found among the treatments and the sampling moment on the population of total viable bacteria, protozoans, pH and ammonia concentration in the rumen.

When evaluating the effect of the system on pH and NH₃ (table 3), it was found that in the animals that grazed L. leucocephala/grass the ammonia concentration was higher, and, subsequently, also the rumen pH. This effect is due to the action exerted by the microbial deaminases excreted by proteolytic bacteria on the proteins contained in L. leucocephala.

In this regard, Rodríguez et al. (2014) stated that the protein contained in this plant is degraded between 56 and 59 % at rumen level; but, because its content is high, there will always be remarkable quantities of free ammonia. This effect can be attenuated with the use of energy supplementation to animals that graze L. leucocephala.

In works conducted by Iraola et al. (2017) it was reported that one of the elements which limit animal productivity in silvopastoral systems (SPS) with L. leucocephala is the contribution of energy and its synchronization with the available nitrogen sources for rumen microorganisms, limiting the microbial protein synthesis and the productive performance of fattening animals. On the other hand, L. leucocephala intake tends to increase the crude protein content in the rumen and can cause excesses in the nitrogen balance, situation that produces additional energy cost for its elimination as urea.

In this regard, there are different forms of manipulating the SPS with L. leucocephala and obtaining productive advantages; within them are the use of easily fermented carbon hydrates, such as the ones contributed by sugarcane (Saccharum officinarum), and the utilization of biologically activated beneficial microorganisms, produced in the Institute of Animal Science of Cuba (Elías, 2014). This product is composed by lactobacilli, yeasts and short-chain organic acids, and constitutes an activator of rumen fermentation (Garcia and Garcia-Curbelo, 2015). Its use as microbial additive in the finishing of cattle in silvopastoral system with L. leucocephala, complemented with sugarcane during the rainy season, generated a contrasting productive effect on the mean daily gain (MDG).

Figure 1 shows the effect of grazing in L. leucocephala associated with grasses or in the mixture of grasses, on the populations of total viable bacteria and protozoans of the rumen. As can be observed, when the animals grazed L. leucocephala, the populations of total viable bacteria were higher (p < 0,001); while the protozoans reduced their presence.

The reduction in the population of protozoans when the animals grazed L. leucocephala corroborates the anti-protozoan properties of this plant (Galindo, 2013); and represents an advantage that is ascribed to its high content in tannins (4,02 % DM) and other secondary metabolites, such as saponins, which exert similar effect at rumen level.

With regards to the role of saponins in the population of protozoans, Zhou et al. (2011) and Galindo et al. (2016) reported that this metabolite can cause depressive effect on the population of protozoans, because changes occur in the permeability of the cell membrane, which can facilitate lysis at rumen level.

Summarizing, the presence of the above-mentioned antinutritional factors or secondary metabolites causes modifications in the microbial population of the rumen, and sometimes they could be toxic (McSweeney et al., 1999).

Likewise, it is important to consider that defauning is one of the likely ways to reduce the enteric emission of CH₄, due to the flow of microbial cells from the rumen and to the reduction in the acetate/propionate ratio (Leng, 2014).

The results explain the productive responses reported by Iraola et al. (2015a), between 0,682 and 0,769 kg/animal/day, when studying the effect of L. leucocephala on cattle fattening; as well as higher animal comfort given by the fauna and ecological development represented by the presence of trees and the increase of biomass production in the system (Iraola et al., 2015b). This last aspect is indirectly related to the population of microorganisms of the rumen, while it contributes to the accurate decision making about the rational use of technical and economic resources that guarantee higher sustainability and profitability of animal husbandry systems for beef production.

It is concluded that the management system with L. leucocephala associated with grasses is advantageous, because it propitiates increases in the populations of total viable bacteria and fiber-degrading microorganisms; which along with the protozoan depressive effect, guarantees a favorable ecological environment for rumen cellulolysis. Likewise, it is recommended to study, in later stages, the effect of the two-month period February-March and the four-month period August-November in the populations of rumen proteolytic bacteria.

 

 

Received: August 12, 2017
Accepted: March 29, 2018

 

 

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