Carlos E. Lascano

Evidence for biological nitrification inhibition in Brachiaria pastures

Nitrification, a key process in the global nitrogen cycle that generates nitrate through microbial activity, may enhance losses of fertilizer nitrogen by leaching and denitrification. Certain plants can suppress soil-nitrification by releasing inhibitors from roots, a phenomenon termed biological nitrification inhibition (BNI). Here, we report the discovery of an effective nitrification inhibitor in the root-exudates of the tropical forage grass Brachiaria humidicola (Rendle) Schweick. Named “brachialactone,” this inhibitor is a recently discovered cyclic diterpene with a unique 5-8-5-membered ring system and a γ-lactone ring. It contributed 60–90% of the inhibitory activity released from the roots of this tropical grass. Unlike nitrapyrin (a synthetic nitrification inhibitor), which affects only the ammonia monooxygenase (AMO) pathway, brachialactone appears to block both AMO and hydroxylamine oxidoreductase enzymatic pathways in Nitrosomonas. Release of this inhibitor is a regulated plant function, triggered and sustained by the availability of ammonium (NH4+) in the root environment. Brachialactone release is restricted to those roots that are directly exposed to NH4+. Within 3 years of establishment, Brachiaria pastures have suppressed soil nitrifier populations (determined as amoA genes; ammonia-oxidizing bacteria and ammonia-oxidizing archaea), along with nitrification and nitrous oxide emissions. These findings provide direct evidence for the existence and active regulation of a nitrification inhibitor (or inhibitors) release from tropical pasture root systems. Exploiting the BNI function could become a powerful strategy toward the development of low-nitrifying agronomic systems, benefiting both agriculture and the environment.

The extractable and bound condensed tannin content of leaves from tropical tree, shrub and forage legumes

The extractable, protein-bound and fibre-bound condensed tannin (CT) concentrations in the leaves of tropical legumes grown in both Colombia and Northern Australia were determined by the butanol–HCl method, whilst extractable CT was also determined by the vanillin–HCl method. With the exception of Senna siameaall species contained CT. The very high CT concentration found in many plants growing in Colombia may have been partly due to soil fertility being much lower at the Colombian than the Northern Australian site.Acacia boliviana,Arachis pintoi,Centrosema latidens,Senna velutinaandGliricidia sepiumcontained <55 g total CT kg−1 DM, which suggests that they could comprise a reasonable proportion of ruminant diets. All other species grown in South America contained 100–240 g CT kg−1 DM, which suggests that they should only be fed in small amounts as supplements to dilute the CT concentration.Leucaenaspecies andCalliandra calothyrsusgrown in Northern Australia contained intermediate concentrations of total CT (60–90 g kg−1 DM). Most species contained 70–95% of total CT as extractable CT, with the exception ofFlemingia macrophylla, where 60% was extractable and 40% bound, andGliricidia sepium, where almost all the CT was bound to protein. Values forFlemingia macrophylladiffered between accessions. Extractable CT determined with vanillin–HCl was generally higher than extractable CT determined with butanol–HCl. Three accessions showed negligible (<1 g kg−1 DM) extractable CT with butanol–HCl but 10–12 g extractable CT kg−1 DM with vanillin–HCl. Two accessions showed undetectable levels of extractable CT but substantial levels of protein-bound CT, illustrating the importance of using a bound CT method for identifying forages containing CT. Relative to freeze drying, oven drying ofLeucaenaspecies reduced the concentration of extractable CT and increased concentrations of bound CT. The significance of the results for the nutrition of ruminant livestock are discussed, including the possible roles of protein-bound and fibre-bound CT.

Effect of the tropical tannin-rich shrub legumes Calliandra calothyrsus and Flemingia macrophylla on methane emission and nitrogen and energy balance in growing lambs

The objective of this study was to test whether the use of tannin-rich shrub legume forage is advantageous for methane mitigation and metabolic protein supply at unchanged energy supply when supplemented in combination with tannin-free legumes to sheep. In a 6 × 6 Latin-square design, foliage of two tannin-rich shrub legume species (Calliandra calothyrsus and Flemingia macrophylla) were used to replace either 1/3 or 2/3, respectively, of a herbaceous high-quality legume (Vigna unguiculata) in a diet composed of the tropical grass Brachiaria brizantha and Vigna in a ratio of 0.55 : 0.45. A Brachiaria-only diet served as the negative control. Each experimental period lasted for 28 days, with week 3 serving for balance measurement and data collection inclusive of a 2-day stay of the sheep in open-circuit respiration chambers for measurement of gaseous exchange. While Vigna supplementation improved protein and energy utilisation, the response to the partial replacement with tannin-rich legumes was less clear. The apparent total tract digestibilities of organic matter, NDF and ADF were reduced when the tannin-rich plants partially replaced Vigna, and the dose-response relationships were mainly linear. The tannin-rich plants caused the expected redistribution of more faecal N in relation to urinary N. While Flemingia addition still led to a net body N retention, even when fed at the higher proportion, adding higher amounts of Calliandra resulted in body protein mobilisation in the growing lambs. With respect to energy, supplementation of Vigna alone improved utilisation, while this effect was absent when a tannin-rich plant was added. The inclusion of the tannin-rich plants reduced methane emission per day and per unit of feed and energy intake by up to 24% relative to the Vigna-only-supplemented diet, but this seems to have been mostly the result of a reduced organic matter and fibre digestion. In conclusion, Calliandra seems less apt as protein supplement for ruminants while Flemingia could partially replace a high-quality legume in tropical livestock systems. However, methane mitigation would be small due to associated reductions in N and energy retention.

A Method for Isolating Condensed Tannins from Crude Plant Extracts with Trivalent Ytterbium

A method to precipitate condensed tannin from crude plant extracts using trivalent ytterbium has been developed. The new method requires less time and resources than the condensed isolation procedure using Sephadex LH-20 recommended by Hagerman (1991, Tannin Analysis, Miami University, Oxford, OH, USA). The absorbance of the preparations obtained by precipitation with trivalent ytterbium was similar to the preparations obtained with the original isolation procedure, when the acid butanol method (Porter et al 1986, Phytochemistry1 223–230) was used to measure condensed tannins. Condensed tannins were isolated from crude plant extracts of three plant species, Desmodium ovalifolium, Gliricidia sepium and Manihot esculenta, and the condensed tannin content of the lyophilised leaf tissue was determined. For each plant species, the amounts of the soluble, insoluble and fibre-bound condensed tannins were estimated using five different standards. These standards included two tannin preparations obtained either by (1) isolation with Sephadex LH-20, or (2) by the precipitation with trivalent ytterbium and three external standards: (3) cyanidin, (4) delphinidin and (5) purified quebracho (Schinopsis balansae). When external standards were used (cyanidin, delphinidin, purified quebracho), it was likely that the condensed tannin content of the plant tissue would be under- or overestimated. When an internal standard based on the isolated tannin from the respective plant species was used, accurate estimates were obtained.

Comparison of the Precipitation of Alfalfa Leaf Protein and Bovine Serum Albumin by Tannins in the Radial Diffusion Method

The precipitation of protein by condensed and hydrolysable tannins was evaluated with the radial diffusion method of Hagerman (1987) using bovine serum albumin (BSA) and isolated leaf protein from fresh alfalfa (Medicago sativa). Alfalfa leaf protein (AALP) was included at two concentrations, 25 and 156 mg N litre-1, at pH 6·8 and 39°C to simulate rumen conditions. The condensed tannins were purified from lyophilised samples of Arachis pintoi, Desmodium ovalifolium, Gliricidia sepium, Manihot esculenta and quebracho (Schinopsis balansae). Hydrolysable tannins from tannic acid (TA) were used as well. There was a significant interaction (P<0·001) between tannin and protein source, and protein level on protein precipitation. Most purified condensed tannins (CTs) precipitated more AALP than BSA when protein was included at the same level. Purified CT from quebracho and hydrolysable tannin from TA failed to precipitate AALP at both protein levels. In a second experiment, tannins from crude plant extracts were studied in the radial diffusion method using BSA and two levels of AALP. The crude plant extracts were obtained from lyophilised plant samples of A pintoi, Centrosema macrocarpum, Clitoria ternatea, D ovalifolium, Erythrina berteroana, E poepigiana, G sepium, M esculenta, Pueraria montana and P phaseoloides. The protein precipitated by soluble tannins in the plant samples was correlated to the total phenolic content and to the soluble CT estimated by the acid butanol assay or by the radial diffusion method. Tannins from different plant species precipitated different amounts of BSA and AALP. Therefore, the measures of the biological activity of tannins based on BSA precipitation may not reflect the ability of tannins to precipitate proteins of plant origin such as those commonly found in the diets of herbivores. The present study offers the possibility of using the radial diffusion method with plant proteins at precipitation conditions similar to those in the rumen.

Phenological, agronomic and forage quality diversity among germplasm accessions of the tropical legume shrub Cratylia argentea

SUMMARYCratylia argentea (Desv.) O. Kuntze is a drought-tolerant tropical shrub legume that can helpto ensure continuity of forage supply in smallholder systems either through direct grazing or as acut-and-carry plant for fresh foliage or silage. A collection of 38 accessions was characterized agro-nomically and nutritionally. High diversity was detected between accessions. Time to floweringranged from 217–329 days after transplanting seedlings to the field and from 129–202 days aftercutting. Flowering is probably induced by reduction of day length. Seed production was high butvariable. Dry matter production ranged from 190–382 g/plant in the rainy and from 124–262 g/plantin the dry season, content of in vitro digestible dry matter from 589–690 g/kg, crude protein contentfrom184–237g/kgandfibrecontentfrom403–528 g/kg(neutraldetergent fibre,NDF),240–335 g/kg(acid detergent fibre, ADF), and 9–13 g/kg (acid detergent fibre-bound nitrogen, N-ADF).Accessions CIAT 18674 and CIAT 22406 were identified as promising for further study. They weresuperior to the commercial cultivar in terms of dry matter (DM) production, particularly in the dryseason. Further research is required to determine the prevailing reproduction strategy of C. argenteaand to quantify outcrossing-rates. Multilocational trials with a selected set of accessions should beconducted under different environmental conditions.INTRODUCTIONLivestock productivity in the tropics is severelyaffected in the dry season by low availability andquality of fodder (NAS 1979; Ranjhan 1986;Enri´quez et al. 2003). High-protein legumes, in par-ticular shrub species, can contribute to improvedproductivity of livestock during dry seasons and atthe same time maintain and even restore soil fertility(Brewbaker 1986; Schultze-Kraft & Peters 1997;Peters et al. 2001; Shelton 2001). Cratylia argentea(Desv.) O. Kuntze (syn. C. floribunda Benth., Diocleaargentea Desv.) is a shrub legume which was selectedas promising for dry-season supplementation, par-ticularly in regions with acid soils and extended dryseasons(Argel MArgelLPeters & Schultze-Kraft 2002).Cratylia argentea belongs to the familyLeguminosae, subfamily Papilionoideae, and is themostwidely distributed ofthe fivespecies in the genus(de Queiroz & Coradin 1996; Pizarro et al. 1997). Theleafy shrub usually reaches 1.5–3 m, but there aretrees up to 10 m tall. It is found in a broad range ofhabitats from Western Peru to the state of Ceara´ inBrazil, and is well adapted to acid soils of low tomedium fertility and altitudes up to 1200 m asl(Xavieretal.1995;Maass1996;Schultze-Kraft1996;Peters & Schultze-Kraft 2002). Its nutritive value ishigher than thatof most other shrublegumes adaptedto acid soils. Plants contain only trace amounts oftannins(Lascano1996;Shelton2001).Ithasexcellentregrowthcapacityaftercuttingandcanbeusedassoilcover, mulch and green manure. The species is verydrought tolerant and remains green and productive

Detecting bacterial endophytes in tropical grasses of the Brachiaria genus and determining their role in improving plant growth

Sorghum is a staple food grain in many semi-arid and tropic areas of the world, notably in Sub-Saharan Africa because of its good adaptation to hard environments and its good yield of production. Among important biochemical components for sorghum processing are levels of starch (amylose and amylopectin) and starch depolymerizing enzymes. Current research focus on identifying varieties meeting specific agricultural and food requirements from the great biodiversity of sorghums to insure food security. Results show that some sorghums are rich sources of micronutrients (minerals and vitamins) and macronutrients (carbohydrates, proteins and fat). Sorghum has a resistant starch, which makes it interesting for obese and diabetic people. In addition, sorghum may be an alternative food for people who are allergic to gluten. Malts of some sorghum varieties display

Linking research on forage germplasm to farmers: the pathway to increased adoption—a CIAT, ILRI and IITA perspective

Abstract The aim of most publicly funded research and development of forages in the subtropics and tropics of the developing world is to improve the livelihoods of smallholder farmers. In order to achieve this goal, technical options are sought which not only contribute to alleviation of poverty and improved food security but also protect natural resources. This paper argues that in order to enhance adoption of multipurpose forages by small farmers, there is a need to utilise participatory methods and to invest in the development of a range of forage alternatives for different environments and production systems. Approaches linking on-station research to farmer participation are described and examples for pathways to adoption presented.

The effect of mixing prunings of two tropical shrub legumes (Calliandra houstoniana and Indigofera zollingeriana) with contrasting quality on N release in the soil and apparent N degradation in the rumen

Lack of synchronization between N released from prunings applied to the soil as green manures and crop uptake as well as optimization of protein digestibility for ruminants, remain major research objectives for the selection of multipurpose tree and shrub legumes (MPT) for mixed smallholder systems in the tropics. Prunings of the high tannin, low quality MPT Calliandra houstoniana CIAT 20400 (Calliandra) and the tannin free, high quality MPT Indigofera zollingeriana (Indigofera) were mixed in the proportions 100:0, 75:25, 50:50, 25:75, and 0:100 (w/w) in order to measure the aerobic rate and extent of N release in a leaching tube experiment, and the anaerobic extent of N degradation in an in vitro gas production experiment. Parameters measured in Calliandra:Indigofera mixtures were compared to theoretical values derived from single species plant material (i.e. 100:0 and 0:100). Aerobic N release and apparent anaerobic N degradation increased with increasing proportion of the high quality legume (Indigofera) in the mixture. While N release in the soil was lower than theoretical values in the mixture 50% Calliandra/50% Indigofera, this was not the case with apparent anaerobic N degradation with the same mixture. Aerobic N immobilization was more pronounced for the mixture 75% Calliandra/25% Indigofera than for 100% Calliandra and negative interaction was observed with apparent anaerobic N degradation in the mixture 75% Calliandra/25% Indigofera. Plant quality parameters that best correlated with aerobic N release and apparent anaerobic N degradation in the rumen were lignin + bound condensed tannins (r=−0.95 and −0.95 respectively, P<0.001). In addition, a positive correlation (r=0.89, P<0.001) was found between aerobic N release in the leaching tube experiment and apparent N degradation in the in vitro anaerobic gas production experiment. Results show that mixing prunings of MPT materials with contrasting quality is an effective way to modify aerobic N release pattern as well as apparent anaerobic N degradation and could possibly be applied to minimize N losses in the rumen and in the soil. In addition, apparent anaerobic N degradation was identified as good predictor of aerobic N release in the soil, which has resource saving implications when screening MTP to be used as green manures.