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Effects of Sapindus saponaria fruits on ruminal fermentation and duodenal nitrogen flow of sheep fed a tropical grass diet with and without legume¹

A. Abreu^*, J. E. Carulla^*, C. E. Lascano, T. E. Díaz, M. Kreuzer and H. D. Hess^,2

^* Department of Animal Production, National University of Colombia, Bogotá, Colombia; and Tropical Grass and Legume Project, CIAT, Cali, Colombia; and National Program of Animal Physiologyand Nutrition, Corpoica, Bogotá, Colombia; and and Institute of Animal Science, Animal Nutrition, Swiss Federal Institute of Technology (ETH) Zürich, CH-8092 Switzerland

Abstract

Six adult African-type hair sheep (BW = 40.3 ± 6.3 kg) fitted with ruminal and duodenal cannulas were subjected to four treatments. Sheep were offered basal diets at a rate of 80 g of DM/kg of metabolic BW (equivalent to ad libitum access) consisting either of a low-quality grass hay (Brachiaria dictyoneura, 3.7% CP, DM basis) alone or in combination with a forage legume (Cratylia argentea, 18.6% CP, DM basis) in a 3:1 ratio (DM basis). In addition, 0 or 8 g of DM of Sapindus saponaria fruits (12.0% crude saponins, DM basis) per kilogram of metabolic BW was administered intraruminally. Supplementation of C. argentea increased intakes of OM (+21%; P < 0.01) and CP (+130%; P < 0.001), as well as ruminal fluid ammonia N concentrations (from 2.40 to 8.43 mg/dL; P < 0.001). Apparent OM and N digestibilities were not affected by legume addition, but ADF digestibility decreased by 10% (P < 0.01). Total ruminal VFA concentration was unchanged, but acetate:propionate was lower (P < 0.01) and isobutyrate proportion was greater (P < 0.001) with the legume addition. Legume supplementation increased duodenal flows of total N (+56%; P < 0.001), nonammonia N (+52%; P < 0.001), ruminal escape N (+80%; P < 0.001), and microbial N (+28%; P < 0.05). Microbial efficiency was not affected by legume addition. Supplementation of S. saponaria increased (P < 0.05) dietary OM intake by 14%, but had no effect on CP intake and ruminal fluid ammonia concentration or on OM and N digestion. Digestibility of ADF was decreased (P < 0.01) by 10% with S. saponaria as was acetate:propionate (P < 0.001) and the isobutyrate proportion (P < 0.001). Ruminal protozoa counts increased (P < 0.01) by 67% with S. saponaria. Duodenal N flows were not significantly affected by S. saponaria supplementation, except for microbial N flow (+34%; P < 0.01). Microbial efficiency was greater (P < 0.05) by 63% with the addition of S. saponaria. Few interactions between legume and S. saponaria supplementation were observed. The NDF digestibility was decreased with S. saponaria in the grass-alone diet, but not in the legume-supplemented diet (interaction; P < 0.05). Interactions were absent in ruminal fermentation measures and duodenal N flow, indicating that effects were additive. Results suggest that, even when not decreasing ruminal protozoa count, supplementation of S. saponaria fruits is a beneficial way to improve ruminal VFA profile, microbial efficiency, and duodenal flow of microbial protein in sheep fed tropical grass-alone or grass-legume diets.

Key Words: Brachiaria dictyoneura • Cratylia argentea • Protein • Protozoa • Saponins • Tropical Forage

Introduction

Tropical ruminant feeding systems heavily rely on pastures and crop residues. These forages usually have a low digestibility and are deficient in N, which restricts the efficiency of feed utilization (Leng, 1990). Utilization could be improved by manipulation of ruminal fermentation either through changes in diet composition (e.g., legume supplementation) or through manipulation of key ruminal microbial groups (Domínguez Bello and Escobar, 1997). Removing ruminal ciliate protozoa has been proposed as one promising approach because their presence decreases bacterial count and the total amount of microbial protein leaving the rumen (Leng, 1990). A range of techniques for defaunation has been tested, but suitable methods for defaunation under normal farm conditions are still lacking (Teferedegne, 2000). Recently, plant secondary metabolites with an inherent capability to partially defaunate the rumen have been described. The incorporation of pure saponins or saponin-rich feeds, such as Sapindus saponaria fruits, into the diet decreased the ruminal ciliate population, whereas bacterial and fungal biomass was increased (Díaz et al., 1993; Navas-Camacho et al., 1993; Makkar and Becker, 1997). This was associated with improved feed conversion efficiency (Domínguez Bello and Escobar, 1997; Navas-Camacho et al., 1997). Supplementing grass-alone and grass-legume diets with S. saponaria fruits resulted in favorable ruminal changes in vitro, but effects occasionally depended on the basal diet (Hess et al., 2003b). Little is known about the effects of S. saponaria supplementation on duodenal N flow in these diet types. The hypothesis tested in the present study was that fruits of S. saponaria supplemented to sheep fed tropical grass and grass-legume diets would change ruminal fermentation toward increased duodenal flows of microbial and total nonammonia N independent of the extra N supplied by the legume, thereby facilitating a broad applicability of both feeding measures.

Materials and Methods

Experimental Procedure
The experiment was conducted at the Quilichao Research Station (3°60' N, 76°31' W, 990 m elevation, 23°C annual mean temperature, 1,772 mm annual rainfall) of the Centro Internacional de Agricultura Tropical (CIAT) in the Cauca Valley of Colombia. Six adult, castrated African-type hair sheep (in good body condition and with 40.3 ± 6.3 kg BW; mean ± SD), fitted with permanent ruminal and duodenal cannulas from Ankom (distributed by Bar Diamond Inc., Parma, ID) were used in this experiment. The animals were randomly allotted to four treatments in different sequences in an unbalanced block crossover design with four experimental periods, and were housed in individual metabolic crates. Treatments consisted of two basal diets, a grass-alone and a grass-legume diet (3:1, DM basis), offered at 80 g/kg BW^0.75, equivalent to ad libitum access (>130% of effective intake). Basal diets were fed alone or supplemented intraruminally with 8 g/kg BW^0.75 of dried and ground (3-mm sieve) S. saponaria fruits. The test grass, offered as hay of Brachiaria dictyoneura cv. Llanero (CIAT accession No. 6133) was harvested from a pasture with a 6-mo-old regrowth, and represented a poor-quality feed, as was obvious from the very low CP (<4% DM basis) and high fiber concentrations, which gave low IVDMD values (Table 1). Dried foliage of a tropical multipurpose shrub, Cratylia argentea (Desvaux) O. Kuntze (CIAT accession No. 18516/18668), served as the legume source. The leaves of C. argentea were harvested from a 5-mo-old regrowth and were high in CP (>18% DM basis) but were similar to the grass in fiber concentration and IVDMD. After harvest, the grass and legume forages were chopped to 5 cm, sun-dried for 3 d, and stored indoors until feeding. The fruits of the S. saponaria (soapberry) tree were collected in the Northern Coastal region of Colombia from several individual plants during the dry season and contained 120 g of saponins and 221 g of total sugar/kg of DM. Fiber concentrations were lower and IVDMD greater in the fruit than in the forages (Table 1). The legume was offered to the animals from 0800 to 0900 and from 1400 to 1500, and the grass was offered during the remainder of the day. Ground S. saponaria fruits were administered at 0900 every day directly into the rumen via the ruminal cannula to avoid refusals and to guarantee a supply of the total allocated amount. Animals had free access to a commercial mineralized salt mix (125 g of Ca; 80 g of P; 157 g of Na; 243 g of Cl; 3 g of Mg; 40 g of S; 7 g of Zn; 0.1 g of I; and 0.05 g of Co/kg). Fresh water was supplied three times per day.

Laboratory Analyses
Feeds, refusals, lyophilized duodenal digesta, and feces were analyzed for DM (105°C for 24 h), OM (500°C for 3 h), total N (micro Kjeldahl; CP = 6.25 x N; AOAC, 1975), NDF (without the addition of -amylase and sodium sulfite), ADF (Van Soest et al., 1991), and indigestible ADF (Waller et al., 1980). Additionally, feeds were analyzed for ADL (Van Soest et al., 1991) and IVDMD (Moore and Mott, 1974). The crude saponin concentration of S. saponaria fruits was determined by the butanol extraction method as described by Hess et al. (2003a). The total sugar concentration in the fruits was measured after hot extraction with ethanol (80%), subsequent filtration, and the addition of an orcinol/sulphuric acid reagent (Shannon, 1972), which gave a yellow color. Absorbance at 420 nm was obtained using a continuous-flow autoanalyzer (AutoAnalyser 2, Bran and Luebbe GmbH; Norderstedt, Germany).

Ciliate protozoa counts were determined using a 0.1-mm depth Bürker counting chamber (Bürker Blau Brand; Wertheim, Germany). Before counting, samples were fixed by the addition of 0.8 mL/mL of formaldehyde-saline solution (37% [vol/vol] formaldehyde and 0.9% [wt/vol] NaCl). Ruminal fluid pH was recorded using a pH meter (MP 120, Mettler Toledo; Greifensee, Switzerland). Ruminal fluid samples for VFA determination were first deproteinized with 25% (wt/vol) metaphosphoric acid (0.2 mL/mL of ruminal fluid), centrifuged at 9,000 x g for 10 min, and then analyzed according to Hess et al. (2003b) using a gas chromatograph (XL GC, Perkin Elmer Instruments; Shelton, CT) equipped with a flame ionization detector and a capillary column (Elite-FFAP, Perkin Elmer Instruments, 30 m long, 0.32 mm i.d., 0.25 µm film thickness). In ruminal fluid and duodenal digesta, concentrations of ammonia N were determined after centrifugation (14,000 x g, 5 min) using the indophenol method (McCullough, 1967). Total N concentration of ruminal fluid was determined by micro-Kjeldahl (AOAC, 1975). The purine concentration of lyophilized duodenal digesta and ruminal bacteria was determined using the technique described by Zinn and Owens (1986) with yeast RNA (RNA Type II–C from Torula yeast; Sigma Chemical Co., St. Louis, MO) used as the standard. Bacteria were isolated from ruminal fluid using differential centrifugation (Smith and McAllan, 1974). In the first step, ruminal fluid was centrifuged twice at 200 x g for 5 min (4°C) to separate bacteria from protozoa and feed particles. The supernatant was centrifuged at 30,000 x g (15 min, 4°C). The resulting supernatant was discarded. The pellet was resuspended using a 0.9% (wt/vol) NaCl solution and centrifuged as before. The supernatant was again discarded, the pellet was resuspended using distilled water, and then lyophilized.

Blood plasma urea N (BUN) concentration was analyzed by means of a commercial kit (Sera-pak, Bayer Diagnostics Manufacturing S.A., Tournai, Belgium) and reading at 546 nm with a spectrophotometer (Spectronic 601, Milton Roy Co., Rochester, NY). Plasma activities of -glutamyl transferase and aspartate amino transferase were analyzed with colorimetric kits (BioSystems S.A.; Barcelona, Spain), and readings were done at 340 and 405 nm, respectively, with another spectrophotometer (QuitKLab2, Ames, distributed by Bayer; Leverkusen, Germany).

Calculations and Statistical Analysis
Indigestible ADF was used as an internal particulate digesta marker to determine duodenal nutrient and DM flow (Waller et al., 1980), and values were corrected for marker recovery in feces, which averaged 83%. The N:purine ratio of isolated ruminal bacteria and the purine concentration of the duodenal digesta were used to estimate duodenal flow of microbial N (Zinn and Owens, 1986). Ruminal escape N was calculated as total duodenal N flow minus microbial N flow and endogenous N flow (Barahona et al., 1997). For that, endogenous N was considered to be proportional to DMI (2.2 g of N/kg of DMI; Hart and Leibholz, 1990). The amount of N apparently absorbed in the lower gut was computed by the difference of total duodenal N and N recovered in feces. Microbial efficiency was estimated as the amount of microbial N reaching the duodenum per kilogram of OM apparently fermented in the rumen. Data of ruminal fluid and blood plasma traits obtained on different days within a measurement period and times within day were combined for further data evaluation.

Data were subjected to ANOVA for an unbalanced block crossover design with four experimental periods and a 2 x 2 factorial arrangement of the four treatments, using the GLM procedure of SAS (SAS Inst., Inc., Cary, NC). Basal diets (grass-alone and a grass-legume mixture), S. saponaria treatment, and basal diet x S. saponaria treatment interaction were considered as main effects. Experimental period and animal were treated as additional sources of variation. In case of significant interactions, multiple comparisons of the least squares means were done using the Scheffé method. Because the interactions were mostly nonsignificant, only the least squares means of the main factors are presented in the tables and interactions are described in the text.

Results

Total intakes of OM, NDF, and ADF/kg of BW^0.75 were greater (P < 0.05) with the legume-supplemented basal diet than with the grass-alone diet (Table 2). Intake of CP was 2.3-fold greater (P < 0.001) with the legume diet than with grass hay alone. Intraruminal supplementation of S. saponaria fruits did not decrease the intake of basal diets, thus increasing (P < 0.05) total OM supply. No effects of legume and S. saponaria supplementation were observed on apparent ruminal degradation of OM and NDF, or on apparent total-tract digestibility of OM, N, and NDF. Digestibility of ADF was lower (P < 0.01) in the grass-legume diet compared with the grass-alone diet and was decreased (P < 0.01) by S. saponaria fruits. No interaction of legume and S. saponaria supplementation on forage intake or on digestibility was observed, except for NDF digestibility, which was decreased (P < 0.05) from 58.0 to 51.0% by S. saponaria in the grass-alone diet, but which remained unchanged by S. saponaria (51.8 and 53.7% without and with S. saponaria, respectively) in the grass-legume diet (data not shown). The same trend (interaction; P < 0.10) was obvious for ADF digestibility.

Effects of Cratylia argentea Supplementation
The foliage of C. argentea used in this experiment presented high CP (18.6% DM basis) and fiber concentrations, and a low digestibility (IVDMD of 46%). The values agreed well with those reported previously for C. argentea cultivated under similar climatic and soil conditions (Fässler and Lascano, 1995; Wilson and Lascano, 1997). Also similar to these reports, the main effect was the increase in N intake when supplementing C. argentea to the low-quality Brachiaria dictyoneura hay. In the present study, the increase was 2.3-fold compared with that recorded with the grass-alone diet. A similar effect of the legume was also observed with almost all variables related to N flow to the duodenum, which is in accordance with the observations made by Wilson and Lascano (1997). However, ruminal N degradation was considerably lower than the values reported by Wilson and Lascano (1997) for diets of similar botanical composition, which could be related to the much lower N concentration in the B. dictyoneura hay used in our experiment. The increased N intake not only resulted from the greater N concentration of the forage, but also from greater feed intake. Improved forage intake resulting from the supplementation of low-quality roughages with foliage from tree legumes has been reported (Norton, 1994; Poppi and Norton, 1995; Osuji and Odenyo, 1997). Forage intake in ruminants consuming fibrous feeds, when not affected by palatability, may be primarily determined by the level of ruminal fill, which in turn is directly related to the rate of digestion and passage of fibrous particles from the rumen (Van Soest, 1994). Therefore, thus maximizing the rate of ruminal fermentation can increase intake. In low-quality forages, however, microbial fermentation activity is often limited by the shortage of one or more essential nutrients, typically N, which results in low concentrations of ruminal fluid ammonia and, consequently, low OM degradation rates (Mehrez et al., 1977; Leng, 1990). In the present experiment, ruminal ammonia N concentration for the grass-alone diet was less than half of the 5 mg/dL reported to be necessary for optimal microbial growth in vitro (Satter and Slyter, 1974) and less than one-eighth of the concentration recommended to achieve the maximum rate of fermentation (Mehrez et al., 1977). This suggests that ruminal fermentation and intake could have been limited by the low N content of the grass-alone diet, and the legume supplementation would be expected to improve ruminal OM and fiber degradation. However, this was not the case, and ruminal fluid VFA concentration did not increase with legume supplementation. This is contrary to results of a recent in vitro study (Hess et al., 2003b), where the replacement of one-third of the B. dictyoneura in the diet by C. argentea drastically enhanced OM degradation, fiber degradation, and VFA production. In that study, however, ammonia concentration of the ruminal fluid from the grass-alone diet was even lower than in the present experiment. Therefore, we suggest that in the current experiment, fiber degradation was not seriously limited by a deficiency of fermentable N in the grass-alone diet and that the threshold level for the minimum ruminal ammonia value reported by Mehrez et al. (1977) for maximum fermentation is probably not applicable for the type of low-quality grass used in our experiment. Accordingly, Erdman et al. (1986) suggested that the minimum ammonia concentration is not constant but rather is a function of the fermentability of the diet. The decrease in ADF digestibility found when the grass diet was supplemented with the legume agrees with the observations of Fässler and Lascano (1995) and Wilson and Lascano (1997), and could be related to the greater lignin (ADL) concentration of C. argentea compared with B. dictyoneura.

Supplementing the legume decreased acetate:propionate in ruminal fluid, which would improve the efficiency of VFA use for productive purposes (Leng, 1990; France and Siddons, 1993). The increased isobutyrate proportion observed when the legume was supplemented could be the result of an enhanced degradation of branched-chain AA.

A major part of the additional N intake with legume supplementation was recovered in total N and nonammonia N arriving at the duodenum, which agrees with results of Wilson and Lascano (1997). The increase in total N flow was due to a greater flow of both microbial N and ruminal escape N. Increased forage intake and greater amounts of nonammonia N escaping the rumen are likely to be the most important nutritional advantages of supplementing low-quality grass-alone diets with foliage of C. argentea.

Effects of Supplementation of Sapindus saponaria Fruits
A major effect expected from supplementation of S. saponaria was a decreased ruminal ciliate protozoa population. Many authors have suggested that total and partial defaunation (removal of protozoa from the rumen) are favorable to improve the utilization of low-quality tropical forages and to increase animal productivity (Leng, 1990; Domínguez Bello and Escobar, 1997; Teferedegne, 2000). This hypothesis is based mainly on the often-found increase in the net efficiency of microbial protein synthesis in the rumen due to the decrease in bacterial protein degradation by the protozoa, followed by an increased flow of total microbial protein to the duodenum (Williams and Coleman, 1992). Foliage (Rosales et al., 1989; Navas-Camacho et al., 1994; Newbold et al., 1997) and fruits (Díaz et al., 1993; Navas-Camacho et al., 1994, 1997) of several tropical and subtropical multipurpose shrubs and trees have been reported to adversely affect ruminal ciliate protozoa. The protozoa-suppressing effect of S. saponaria fruits has been demonstrated in vitro (Hess et al., 2003a,b) and in vivo (Díaz et al., 1993; Navas-Camacho et al., 1994). Contrary to those results, the ciliate protozoa count was increased by supplementation of S. saponaria fruits in the present experiment. Variable effects on ruminal protozoa have also been reported from the use of foliage of Sesbania sesban, a saponin-containing forage from an African multipurpose legume tree (Newbold et al., 1997; Odenyo et al., 1997; Teferedegne et al., 1999). Sesbania sesban depressed protozoa count in the ruminal fluid of sheep kept in Great Britain (Newbold et al., 1997; Teferedegne et al., 1999) and also depressed protozoa count when administered directly into the rumen of sheep stationed in Ethiopia, but had no effect on protozoa number when fed orally to sheep in Ethiopia (Odenyo et al., 1997). Therefore, Teferedegne (2000) suggested that, in sheep adapted to S. sesban, saponins could be degraded in the saliva before reaching the rumen. This cannot explain the absence of antiprotozoal activity of S. saponaria in the present experiment, because the fruits were intraruminally supplied. Adaptation to saponins cannot be excluded in the present experiment either, because sheep were freely browsing native shrubs and grazing native and improved pastures prior to the experiment and could have had access to saponin-containing feeds. Also, Wallace et al. (2002), reviewing current literature, suggested adaptation of mixed microbial population of the rumen as one factor contributing to the variability of the antiprotozoal activity of saponins or saponin-containing plants. Additionally, results of Hess et al. (2003b) suggest that effects of S. saponaria on ruminal protozoa could be diet-dependent, because protozoa numbers were decreased by S. saponaria fruits only when added to a diet supplemented with a high-quality legume (Arachis pintoi), but not with C. argentea. Diet dependency of saponin effects and adaptation of ruminal microbial populations to saponins could be responsible for the absence of antiprotozoal activity of S. saponaria in this study, but cannot explain the highly significant increase in protozoa counts. Besides fiber, total sugars were the most abundant nutrient fraction in the S. saponaria fruits, representing over 200 g/kg of DM, thus providing a unique extra supply of readily fermentable carbohydrates available for ruminal microbes. This could have enhanced protozoa populations (Williams and Coleman, 1992).

Forage intake was not affected by the supplementation of ground S. saponaria fruits. This agrees with results from previous studies (Díaz et al., 1993; Navas-Camacho et al., 1997), which showed that forage intake of sheep did not change when the pericarp of S. saponaria fruits was supplemented to the diet at rates between 3.5 to 4.5 g/kg BW^0.75 (corresponding to approximately 7 to 9 g/kg BW^0.75 of complete fruits). However, at greater levels (9 g/kg BW^0.75 of pericarp), a drastic decrease in feed intake was observed (Navas-Camacho et al., 1997).

Contradictory results on the effect of saponins or saponin-containing plants on nutrient digestion in ruminants are described in the literature. Lu and Jorgensen (1987) reported a general decrease of fermentative activity and lower ruminal cellulose degradation rates when alfalfa saponins were supplied to sheep. However, apparent digestibility coefficients of OM and cellulose in the total tract were increased. In contrast, Goetsch and Owens (1985) found that the ruminal degradation of OM was increased, whereas degradation of ADF was not affected when sarsaponin was added to diets with medium and low concentrate proportion. In both studies, effects of saponins were highly diet-dependent, an observation also made by Wang et al. (2000), who reported that a saponin-containing extract of Yucca schidigera enhanced in vitro fermentation of barley grain but not of alfalfa hay. In the present study, apparent total-tract digestibilities of OM, N, and NDF were not affected by supplementation of S. saponaria. These results are in agreement with those obtained by Navas-Camacho et al. (1997), who did not observe effects on in sacco DM, N, and NDF degradabilities when supplementing the pericarp of S. saponaria to sheep. In contrast, a significant decrease in ADF digestibility was observed when supplementing S. saponaria fruits in the present experiment. Wallace et al. (1994) and Wang et al. (2000) emphasized that differences may exist in the susceptibility of individual ruminal bacterial species to saponins. From in vitro results, Wang et al. (2000) concluded that cellulolytic bacteria are more susceptible to saponins than other bacteria. If so, it is likely that the degradation of cellulose (a major component of ADF) was depressed by the saponins. Also, the shift in VFA profile from acetate to propionate (and butyrate) at constant total VFA concentration in ruminal fluid underlines this assumption. Similarly, Hristov et al. (1999) reported a greater propionate concentration and decreased acetate:propionate ratio in heifers receiving Yucca schidigera powder as a saponin source. These shifts would be expected when a massive decrease in protozoa count takes place simultaneously, which was the case in the study of Hristov et al. (1999), but not in the present investigation. However, when using defaunated ruminal fluid, Wang et al. (2000) again found similar saponin effects on VFA profile in vitro. This indicates that the decrease in acetate:propionate ratio by the supplementation of saponins is not necessarily mediated by a suppression of ruminal protozoa. Additionally, the VFA profile could have been affected by the sugars provided through the fruits of S. saponaria. Sugars may reduce the proportion of acetic acid and increase the proportion of propionic and butyric acid (Chamberlain et al., 1993; Friggens et al., 1998), which agrees well with the observations made in the present study.

The effects of S. saponaria on duodenal N flow were less pronounced than those of legume supplementation. The effects of saponins on duodenal N flow vary considerably between saponin sources and application levels. For example, Hristov et al. (1999) found no significant effect on microbial N flow when Y. schidigera powder was given to heifers at doses of 20 and 60 g/d. Similarly, Goetsch and Owens (1985) found no effect on total and microbial N entering the duodenum when sarsaponin of Y. schidigera origin was included in the diet of mature dairy cows at 44 mg/kg. Lu and Jorgensen (1987) even demonstrated that alfalfa saponins at levels of 20 and 40 g/kg dietary DM can be detrimental to ruminal microorganisms, and might decrease microbial protein synthesis in the rumen and microbial N flow to the duodenum of sheep. In contrast, Klita et al. (1996) reported an increased duodenal flow of microbial N when sheep received daily doses of 200, 400, or 800 mg/kg of BW of alfalfa root saponins. This increase in microbial N flow was associated with a significantly decreased ruminal protozoa count. However, in the present study, ruminal protozoa count was increased by S. saponaria. On the other hand, it seems unlikely that the protozoa themselves might have largely contributed to the estimated increased duodenal flow of microbial N because absorption and excretion of purines, which were used to estimate microbial N in this study, are generally decreased with increasing protozoa counts (Fujihara et al., 2003). Growth of bacteria could have been enhanced by the sugars present in the S. saponaria fruits. Chamberlain et al. (1993) reported an increased flow of microbial protein to the small intestine (+40%) in sugar-supplemented sheep. Overall, supplementation of S. saponaria improved efficiency of ruminal fermentation via an increased efficiency of microbial protein synthesis and decreased acetate:propionate.

Interactions of Cratylia argentea and Sapindus saponaria Supplementation
It is unclear why NDF digestion was depressed by S. saponaria in the grass-alone diet but not in the grass-legume diet. The major difference between the two basal diets was in the N concentration and the resulting differences in ruminal ammonia concentration. Supplementation with S. saponaria did not significantly affect the ammonia concentration in ruminal fluid, but did decrease isobutyrate proportion of total VFA, an essential growth factor for certain fibrolytic ruminal bacteria (France and Siddons, 1993), which could have contributed to the contrasting effects of S. saponaria on fiber digestibility in the two diets.

The absence of significant interactions for all other variables evaluated is relevant for two reasons. Firstly, this suggests that the potential of S. saponaria to improve efficiency of ruminal fermentation through increased microbial protein synthesis is not dependent on the type of basal diet fed. This greatly extends the range of diets and production systems to which this supplement can be applied. Secondly, it indicates that the effects of the legume and S. saponaria could be additive. This was especially obvious in duodenal flow of microbial N, which was increased by 28% and 34% with legume and S. saponaria supplementation alone, respectively, whereas in combination, the two supplements increased microbial N flow by 80%.

Implications

Supplementing 8 g/kg of BW^0.75 of Sapindus saponaria fruits to sheep improved efficiency of ruminal fermentation and microbial N flow to the duodenum, despite showing no antiprotozoal activity. This tree species could be grown in most tropical agroecological zones from sea level up to over 1,500 m. In addition, results confirmed that the shrub legume Cratylia argentea is an effective protein supplement for ruminants fed low-quality grasses. Because the extremely low nitrogen concentration of tropical grasses often seems to be first limiting, supplementation of C. argentea can improve common tropical diets. Combining the two feeding alternatives provides tropical livestock producers with two promising and additive measures to improve low-quality grass diets. Further research is needed to show the applicability and the implications for productivity and products resulting from these improved tropical feeding systems.

Footnotes

¹ This study was supported by the Swiss Agency for Development and Cooperation (SDC) through their Research Fellow Partnership Program. The authors are grateful to P. Avila and G. Ramirez for their assistance in this study.

² Correspondence: ETH Center/LFW (phone: +41-1-632-5683; fax: +41-1-632-1128; e-mail: dieter.hess{at}inw.agrl.ethz.ch).

Received for publication September 24, 2003. Accepted for publication February 4, 2004.

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