Effect of ad libitum tree leaves feeding with varying levels of concentrate on intake, microbial protein yield and growth of lambs

M K Tripathi, S A Karim, O H Chaturvedi and V K Singh

Division of Animal Nutrition, Central Sheep and Wool Research Institute, Avikanagar (via-Jaipur) Rajasthan, 304 501 India
mktripathi@gmail.com

Abstract

This experiment studied effects of free choice tree leaves feeding with restricted or ad libitum concentrate supplementation on intake, nitrogen utilisation and performance of weaner lambs. Tree leaves offered to lambs contained khejri (Prosopiscineraria) and siris (Albizialebback) in 50:50 ratio. Sixty weaner (90 d) lambs, were divided into three equal groups and in addition to free choice of tree leaves lambs were supplemented with concentrate either 1.5 (C1.5) or 2.5 % (C2.5) of body weight (BW), while the third group (CAL) was fed ad-libitum. The experiment lasted for 90 days.

Lambs on C1.5 or CAL consumed similar amount of dry matter (4.2 kg 100 kg^-1 BW) but C2.5 lambs had higher (P < 0.01) dry matter intake (4.9 kg 100 kg^-1 BW). Digestibility of dry matter (DM), intake of digestible crude protein (DCP) and digestible organic matter (DOM), digestible organic matter fermented in rumen (DOMR) and microbial protein yield (MPY) increased linearly (P<0.01) with increasing concentrate feeding. The mean ADG during 0- 90 d feeding increased linearly with higher concentrate feeding but was not statistically different between C1.5 (81 g/ day) and C2.5 (106 g /day) animals but CAL animals had significantly (p<0.01) higher (170 g / day) ADG. The FCR was higher (P< 0.01) in CAL, while C1.5 and C2.5 had similar FCR. Lambs under all the three feeding regimen had positive N-balance, which was significantly different (p<0.01) among the three groups. The N balance was 3.7, 7.7 and 12.2 g / day respectively in C1.5, C2.5 and CAL lambs. Condensed tannins (CT) intake 5.8 g day^-1 or 6.9 g kg^-1 DMI improved MPY in CAL lambs and 24.3 g day^-1 or 24.8 g kg^-1 DMI CT intake on moderate concentrate feeding (C2.5) did not impaired N utilization and MPY, but a CT intake of 30.8 g kg/ kg DMI impaired N utilisation and MPY in C1.5. DOMR had linear and positive relationship with daily gain (ADG, g/d= 0.535DOMR-11.486, R²=0.65, SE=21.8, P<0.003) and N retention (N retention, g/d = 0.0663DOMR-6.468, R²=0.82, SE=1.727, P<0.001).

It is concluded that with high level of tree leaves feeding,  ad libitum concentrate feeding is desirable for intensive lamb production.

Key words: Concentrate, growth, lamb, nitrogen utilization, tannins, tree leaves

Introduction

Quality and quantity of feed and fodders are the major constraints in increasing ruminant's productivity under tropical conditions. Existing feedstuffs in tropical countries often provide inadequate energy, protein, minerals and vitamins to support optimum animal productivity (Reed et al 1990). Tree and shrub fodders are an important source of supplementary protein, vitamins and minerals in developing countries. Tree leaves, which are rich in nitrogen and widely used in the tropics (Le Houerou 1980; Baumer 1992), offer the opportunity for use as N supplements to ruminant livestock. Supplementation of Leucaena leucocephala and Sesbania sesban to Menz sheep provided higher concentrations of rumen metabolites, which  improved rumen function and feed digestibility (Bonsi et al 1995). Moreover, multipurpose trees of the genera Leucaena and Sesbania were reported to improve the efficiency of microbial N synthesis and N retention when supplemented to sheep fed teff straw (Umunna et al 1995). Tessema and Baars (2004) recommended the inclusion of Sesbania leaves at 250 g/kg diet DM with hay grass-based feeding, which improved DMD, OMD and CPD. Similarly, supplementation of dried Elaeis guineense leaves at 25% or less was found to be suitable in hay-based diets for sheep (Osakwe et al 2004). However, Melaku et al (2004) reported that increasing supplementation of tree fodder levels reduced nutrient digestibility due to increased passage rate and recommended low levels of tree leaves supplementation.

Many tree leaves contain various levels of antinutritional factors that have an affinity for carbohydrates, amino acids and minerals, rendering them unavailable for rumen microflora and the host animal (Makkar 2003), thereby, decreasing livestock production and reproductive performance (Waghorn et al 1999). Silanikove et al (1997) confirmed the assumption that the high tannin content of browses negatively affects the utilization of protein in supplementary feed. The principal negative effect of tannins is on protein utilization (Silanikove et al 2001); however, the inclusion of a limited quantity of tree leaves in animal feed is recommended to improve rumen function and productivity (Osakwe et al 2004).

Improved livestock production could be achieved through cultivation of high quality forage adapted to local conditions as well as feeding concentrate. Concentrate feeds promote rapid growth of sheep and cattle (McDonald et al 1996), reduce ruminal methane production and increase ruminal propionate production, thereby lowering energy losses and contributing to higher overall efficiency of utilization of dietary energy for body weight gain (Mandebvu and Galbraith 1999). Therefore, tree foliage or forage based ruminant feeding with an appropriate level of concentrate may provide optimum nutrient balance to improve animal productivity. Growing weaner lambs supplemented with 250 g concentrate in addition to grazing in semi-arid regions of India (Karim et al 2004) is the prevailing lamb production system, however their growth performance is not optimum. Khejri (Prosopis cineraria) and Siris (Albizzia lebbek) are the major fodder trees in semi-arid regions of India, but less is known about their use in animal feeding. The study was, therefore, conducted to study the effect of tree leaves feeding as source of roughage with restricted or ad-libitum concentrate feeding on intake, nitrogen utilisation pattern and growth performance of lambs.

Materials and methods

The experiment was conducted at the Central Sheep and Wool Research Institute, Avikanagar (Rajasthan, India) located at 26⁰ 17'N latitude and 75⁰ 28'E longitude and 320 m above sea level. The climate is hot and semi-arid. The experiment was initiated in April and ended in July (2003). During the experiment, minimum and maximum ambient temperature ranged from 23⁰ to 35⁰C and 27⁰ to 49⁰C, respectively. Relative humidity varied from 24 to 96%.

Animals and feeding management

Sixty male weaner lambs (13.9 ±1.94 kg body weight, BW) were divided into three equal groups and were penned in well-ventilated enclosures for the experiment. The animals were allowed to walk in an open yard for two hr daily in the morning. Deworming was done at the beginning of the experiment using 'Albendazole' (WOCKhARDT India Ltd. Bombay) @ 10 mg/ kg BW. Animals were fed for 90 d under three feeding regimes comprising of ad-libitum tree leaves (Prosopis cineraria and Albizialebback, in 50: 50 ratio) feeding with restricted (15, "C1.5" or 25 g " C2.5" kg^-1 live weight) or ad libitum "CAL" concentrate. The tree leaves and concentrate sources contained crude protein 125 and 186 g kg^-1 DM, respectively (Table 1).

Feed was offered once daily at 09:00 h, after discarding the previous day's residue, for an excess of 10 %. Feed samples were collected weekly for DM determination and three or four-week samples were pooled for chemical analysis. Water was available twice a day in the morning (10:00 to 11:00 h) and evening (16:00 to 17:00 h). Feed intake was recorded daily. Lamb BW's were recorded for 2 consecutive days, every 7 d immediately before offering feed and water and these values were used to determine BW gain and feed conversion ratio (FCR).

Metabolism trial and sample collection

A metabolism trial was conducted after 75 d of experimental feeding on 6 randomly selected lambs from each treatment. The metabolism trial lasted for 10 d (i.e., 3 d adaptation followed by 7 d of sample collection) during which daily feed intake and output of faeces and urine were collected and recorded. Samples of feed, orts, faeces and urine were collected every morning. Faeces and urine were collected using total collection method in which urine was collected into acidified containers. The air DM of feeds, faeces and orts was determined by drying to a constant weight in a forced air oven at 70 ⁰C. Dried samples for each day of the 7 d collection were pooled, ground to pass a 1 mm screen and preserved for chemical analysis.

Chemical analysis and calculations

Feed, orts and faeces were analysed for DM by drying at 100 ⁰C for 24 h. The OM was determined by ashing at 550 ⁰C for 4 h and CP by a Kjeldahl technique (AOAC 1995). Neutral detergent fiber (NDF) was determined by procedure of Van Soest et al (1991) without sodium sulfite or a-amylase, whereas acid detergent fiber (ADF) and acid detergent lignin (ADL) were determined according to the method described by Robertson and Van Soest (1981). NDF and ADF are expressed with residual ash. Extractable condensed tannins and total phenols were estimated as per procedures described in the manual of IAEA (2000) . Digestible organic matter fermented in the rumen (DOMR) was calculated following the equation (DOMR= DOMI * 0.65; Chen et al 1992). Rumen microbial protein yield (MPY, g/d) was calculated using Muia et al (2001) equations as, Total MPY= (FOM/1000) * 150, FOM: fermentable OM (g/d).

Calculations and Statistical analysis

Data on intake, nitrogen utilization pattern, gain and feed conversion efficiency were subjected to analysis of variance by the mathematical model of Harvey (1975) as:

Y_ij = m + T_i + e_ij

where:

m = General mean,
T_i = Effect of i^th treatment (i = 1, 4),
e_ij= Random error

Group means were separated using Duncan's multiples range test (Duncan 1955). Regression analysis was also done to assess the linear and quadratic effects (Snedecor and Cochran 1994).

Results and discussion

Nutrient intake and digestibility

Dry matter intake (DMI) was significantly (p<0.01) higher in C2.5 lambs than in C1.5 or CAL lambs (Table 2). Intake of tree leaves did not differ between C1.5 and C2.5  but was much less on CAL.  The digestibility of DM increased with increasing concentrate allowances and showed both linear and quadratic relationships. Digestible crude protein (DCP), digestible organic matter (DOM) and DOMR increased linearly with increasing concentrate feeding.

Incorporation of concentrate in ruminant diets is intended to optimise the efficiency of feed utilization for growth and production. However, concentrate supplementation may reduce digestibility in forage containing diets by cattle and sheep (Archimede et al 1995). The depression of digestion is related to a decrease in ruminal pH, a preference by rumen microbes for readily fermentable carbohydrates (Harrison and McAllan 1980). The extent of effect of concentrate on digestion depends on the nature and proportion of the concentrate as well as the quality of the forage species (Archimede et al 1995) and supplementation of concentrate did not affect digestibility of high quality hay fed to lambs (Matejousky and Sanson 1995). Therefore, high quality forage is less susceptible to negative associated effects when concentrate is incorporated in the diet than are low quality forages. In the present study, animals with high concentrate feeding (CAL) had higher digestibility consequently higher digestible crude protein and organic matter intake. Higher concentrate intake and low tree leaves intake possibly synchronized better nutrient availability for optimum rumen fermentation and microbial growth, which in turn improved intake and DM digestibility. Increased total tract digestibility coefficients were also noticed by Cerrillo et al (1999) on higher incorporation of sorghum grain in to the hay diets in goats. Tree leaves used in present experiment were of high quality. Further the concentrate and forage fed to animals contained higher proportions of ash that might have helped in maintaining ruminal pH near neutral. Moreover, negative influence of concentrate feeding on ruminal digestion coefficient is postulated when the proportion of grain in ruminant diets increases to more than 30 % of the DM (Mould et al 1983; Beck et al 1992). Concentrate of the present experiment contained only 29 % maize which was below the critical level that reduces ruminal digestion.

Average daily gain (ADG) and feed conversion ratio (FCR)

ADG during 0-30 days feeding was not different among the three groups but it linearly increased with increasing concentrate feeding from 31^st day onward. The lamb of C1.5 group showed variable ADG but C2.5 lambs stabilised their ADG during 31-90 d feeding while CAL animals continuously improved ADG during 0-90 d feeding (Table 2). The mean ADG during 0- 90 d feeding increased linearly with higher concentrate feeding but was not statistically different between 1.5 (C1.5) and 2.5 % (C2.5) concentrate fed animals. The mean FCR was higher (P< 0.01) in CAL, while C1.5 and C2.5 had similar FCR. But during 61-90 d feeding FCR was similar among the three groups. Lower growth of C1.5 lambs was the cumulative effect of lower feed and nutrient intake and reduced microbial growth. The improvement in growth and feed conversion efficiencies of C2.5 and CAL animals was the influence of better nutrient density and quality of nutrients available for utilisation. Higher DOMR availability, greater microbial protein yield and proportionate N intake in relation to DOMI and DOMR together positively contributed to daily gains and N utilisation (Figures 1 and 2).

Possibly better protein availability and low tannin intake further improved the efficiency of animals fed higher levels of concentrate. Different concentrate allowances potentially have different impact on the rumen microbial protein synthesis and growth (Slyter et al 1970; McAllister et al 1990). Present findings are in accordance with the findings of Cerrillo et al (1999), Piwonka et al (1994) and Grigsby et al (1993) who reported higher incorporation of consumed N into bacterial-N as well as greater flow and availability of microbial-N in the duodenum on feeding greater proportions of concentrate plus forage to goats, steers and heifers, respectively. Therefore, better nutrient availability and utilisation, higher N retentions and microbial protein yield improved growth performance of lambs on higher concentrate feeding to animals.

Nitrogen (N) utilisation pattern and microbial protein yield (MPY)

N intake was lower (P< 0.008) in C1.5 compared to C2.5 and CAL animals. Nitrogen excretion in faeces was higher in C1.5 and C2.5 lambs than in CAL (Table 3). Nitrogen utilisation improved with increased concentrate feeding and had linear effects.

The N excretion pattern revealed that faecal N excretion accounted for 61.4 % and 73.5 % of total N exertion, whereas urinary N execration accounted for 17.8 to 20.7 % of total N excretion. Total MPY (g/day) was higher (p<0.01) in C2.5 and CAL lambs compared to C1.5 and had linear (p<0.01) effects with concentrate feeding. Concentrate feeding with tree leaves increased DOMR in lambs and DOMR improved ADG (Figures 1) and N retention (Figure 2) in a linear manner. The CT intake reflected the amount of tree leaves consumed by animals. C1.5 animals consumed 31 g CT whereas C2.5 and CAL lambs consumed 25 and 7 g CT per kg dry matter intake. Low nutrient intake and DM digestibility in C1.5 lambs may have been due to high tannin intake through tree leaves and restricted concentrate supplement. The Prosopis ceneraria leaves contain high amount of condensed tannins (91 g kg^-1 DM), low hydrolysable tannins (3.4 g) and have a greater protein precipitation capacity 111 g kg^-1 DM (Bhatta et al 2004). Tannins are known to reduce dry matter digestion in the rumen causing a reduction in the passage rate of digesta (Silanikove et al 1996) or post ingestive malaise (Silanikove et al 2001). Tannins may bind to cell wall and cell solubles and reduce the digestion of protein and microbial fermentation (Makkar et al 1995), which negatively affected nutrient digestion in the rumen.

The N retention is considered as the most common index of the protein nutrition status of ruminants (Owen and Zinn 1988). However, the differences in the quantities of N excretion in faeces and its influences on N retention reflected the differences in N intake and metabolism. Lower N retention and higher faecal N excretion in C1.5 and C2.5 animals could be due to higher Prosopis tannin intake. Higher N balance in CAL animals could have been due to increased protein intake and degradation in the rumen. The higher degradation of N and dry matter promotes higher microbial N yield and contributes to higher efficiency of microbial N supply in ruminants (Umunna et al 1995). In the present study 24 g daily CT intake through tree leaves at lower level of concentrate feeding reduced N utilization and microbial protein yield but similar tannin intake at moderate concentrate feeding did not show negative influence.

Conclusions

• Feeding tree leaves ad libitum with increased concentrate feeding improved DM intake and digestibility.

• The level of 2.5 % concentrate feeding optimised the DM intake, digestible OM intake and DOMR.

• Increased concentrate feeding linearly improved DM digestibility, DCP and digestible OM intake, DOMR and ADG, and N retention and simultaneously reduced faecal and urinary N losses.

• Ad lib  feeding of concentrates and tree leaves appears to be the most desirable combination for intensive lamb production programmes

Acknowledgements

Funds provided  for the experiment under JAI VIGYAN Project are acknowledged.

References

AOAC 1995: Official methods of analysis. 16th ed., Association of Official Analytical Chemists Washington, DC.

Archimede H, Sauvant D, Herview J, Poncet C and Dorleans M 1995 Digestive interactions in the ruminant: relationships between whole tract and stomach evaluation. Animal Feed Science and Technology.54:327-340.

Baumer M 1992 Trees as browse and to support animal production. In: Speedy, A., Pugliese, P.L. (Eds.), Legume Trees and Other Fodder Trees as Protein Sources for Livestock. FAO Animal Production and Health Paper No. 102. FAO (Food and Agriculture Organization), Rome, Italy, pp. 1-10.

Beck T J, Simms D D, Cochran R C, Brandt Jr R T, Vanzant E S and Kuhl G L  1992  Supplementation of ammoniated wheat straw: performance and forage utilization characteristics in beef cattle receiving energy and protein supplements.  Journal of Dairy Science. 70:349-357.

Bhatta R, Shinde A K, Verma D L, Sankhyan S K and Vaithiyanathan S 2004 Effect of supplementation containing polyethylene glycol (PEG)-6000 on intake, rumen fermentation pattern and growth in kids fed foliage of Prosopis cineraria. Small Ruminant Research. 52 :45-52.

 

Bonsi  M L K, Osuji P O and Tuah AK 1995 Effect of supplementing tef straw with different levels of Leucaena or Sesbania leaves on the degradabilities of teff straw, sesbania, leucaena, tagasaste and vernonia and on certain rumen and blood metabolites in Ethiopian Menz sheep. Animal Feed Science and Technology, Volume 52, Number 1, March 1995, pp. 101-129(29)

 

Cerrillo M A, Russell J R and Crump M H 1999  The effect of hay maturity and forage to concentrate ratio on digestion kinetics in goats. Small Ruminant Research. 32 :51-60.

 

Chen X B, Chen Y K, Franklin M F, Orskov E R and Shand W J 1992 The effect of feed intake and body weight on  purine derivatives excreation and microbial protein supply to sheep. Journal of Animal Science. 70 :1535-1542.

Duncan D B 1955 Multiple range and multiple F tests. Biometrics. 11: 1-2. 

Grigsby KN, Kerley MS, Peterson J A and Weigel J C 1993 Combinations of starch and digestible fiber in supplements for steers consuming a low quality brome grass hay diet. Journal of Animal Science. 71: 1057-1064.

Harrison D G and McAllan A B 1980  Factors affecting the microbial growth yields in the reticulum rumen.  In: Ruckebush, Y., Thivend, P.(Eds.), Digestive Physiology and Metabolism in Ruminants, Proc.. 5^th Int.  Symposium on Ruminant Physiology, MTP Press, Lancaster, England, pp. 205-226.

Harvey W R 1975 Least Square Analysis. United States Department of Agricultural Research Services (ARS) Washington, DC.

IAEA 2000 Quantification of tannins in tree foliage. A laboratory manual for the FAO/IAEA Co-ordinated research project “Use of nuclear and related techniques to develop simple tannin assay for predicting and improving the safety and efficacy of feeding ruminants on tanniferous tree foliage” FAO/IAEA working document, IAEA, Vienna. pp 26.

Le Houerou H N 1980 Browse in Northern Africa. In: Le Houerou, H.N. (Ed.), Browse in Africa, The current State of Knowledge. ILCA (International Livestock Centre for Africa), Addis Ababa, Ethiopia, pp. 55-80.

 

Makkar H P S 2003 Effects and fate of tannins in ruminant animals, adaptation to tannins, and strategies to overcome detrimental effects of feeding tannin-rich feeds. Small Ruminant Research. 49:241–256.

 

Makkar H P S, Blummel M and Becker K 1995 Formation of complexes between polyvinyl pyrrolidones or polyethyleneglycols and tannins, and their implications in gas production and true digesstibility in in vitro techniques. British Journal of  Nutrition 73:897-913.

Mandebvu P and Galbraith H 1999 Effect of sodium bicarbonate supplementation and variation in the proportion of barley and sugar beet pulp on growth performance and rumen, blood and carcass characteristics in young entire lambs. Animal Feed Science and Technology 82:37-49. 

Matejousky K M and Sanson D W 1995  Intake and digestion of low-, medium-, and high- quality grass hays by lambs receiving increasing levels of corn supplementation.  Journal of Animal Science 73:2156-2163.

McAllister T A, Rode L M, Major D J, Cheg K J and Buchanan S 1990  Effect of ruminal microbial colonization on cereal grain digestion. Canadian Journal of Animal Science 70:571-579.

McDonald P, Edward R A, Greenhalgh J F D F and Morgan C A 1996 Animal Nutrition. Logman Scientific and Technical, Harlow, UK.

Melaku S, Peters K J and Tegegne A 2004 Microbial nitrogen supply, nitrogen retention and rumen function in Menz sheep supplemented with drid leaves of multipurpose three, their mixture or wheat bran. Small Ruminant Research 52:25-36.

Mould F L, Ørskov E R and Mann O S 1983 Associative effect of mixed feeds. I. Effect of type and level of supplementation and its influence on the rumen fluid pH on cellulolysis in vivo and dry matter digestion of various tree leaves. Animal Feed Science and Technology 10:15-30.

Muia M K, Tamminga S, Mbugua P N and Kariuki J N 2001 Rumen degradation and estimation of microbial protein yield and intestinal digestion of napier grass (Pennisetum purpureum) and various concentrates. Animal Feed Science and Technology 93:177-192.

Osakwe I I, Steingass H and Drochner W 2004 Effect of dried Elaeis guineense supplementation on nitrogen and energy partitioning of WAD sheep fed a basal hay diet. Animal Feed Science and Technology 117:75-83.

 

Owen F N and Zinn R 1988. Protein metabolism in ruminant animals. In : Church, D.C. (Ed), The Ruminant Animal Digestive Physisology and nutrition. Waveland Press Inc., Prospects Hights, IL, USA, pp. 227-249.

Piwonka E J, Firkins J L and Hull B L 1994 Digestion in the rumen and total tract of forage based diets with starch or dextrose supplements fed to Holstein heifers.  Journal of Dairy Science 77:1570-1579.

Reed J D, Soller H and Woodward A 1990 Fodder tree and strover diets for sheep : intake, growth, digestibility and effects of phenolics on nitrogen utilisation. Animal Feed Science and Technology 30:39-50.

Robertson J B and Van soest  P J 1981 The detergent system of analysis and its application to human foods. Cornell University, Ithaca, New York.

Silanikove N, Giloba N and Nitsan Z 1997 Interactions among tannin, supplementation and polyethylene glycol in goat fed oak leaves. Animal Science 64:479-483.

Silanikove N, Giloba N, Perevolotsky A and Nitsan Z 1996 Effect of daily supplementation of polyethylene glycol on intake and digestion of tannin containing leaves (Quercus calliprinos, Pistacia lenticus and Ceratonia siliqua) by goats. Journal of Agriculture and Food. Chemistry 44:199-205.

Silanikove N, Perevolotsky A and Provenza F D 2001 Use of tannin binding chemicals to assay for tannin and their negative post-ingestive effects in ruminants. Animal Feed Science and Technology 91:69-81.

Slyter L L, Oltjen R R, Kern DL and Blank F C 1970 Influence of type and level of grain and diethylstilbesterol on the rumen microbial population of steers fed all concentrate diets. Journal of Animal Science 31:996-1002.

Snedecor W G and Cochran G W 1994  Statistical Methods. 8^th Edn. Oxford and IBH Publication Co. Calcutta, India.

 

Tessema Z and Baars R M T 2004 Chemical composition, invitro dry matter digestibility and ruminal degradation of Napier grass (Pennisetum purpureum (L.) Schumach.) mixed with different levels of Sesbania seban (L.) merr. Animal Feed Science and Technology 117:29-41.

 

Umunna N N, Osuji P O, Nsahlai I V, Khalili H and Mohamed-Saleem M A 1995 Effect of supplementing oat hay with lablab, sesbania, tagasate or wheat middlings on voluntry ontake, N utlisation and weight gain of Ethopian Menz sheep. Small Ruminant Research 18:113-120.

 

Van Soest P J, Robertson J B and Lewis B A 1991 Methods for dietary fiber, neutral detergent fiber and nonstarch polysaccharides in relation to animal nutrition. Symposium: Carbohydrate methodology, metabolism and nutritional implications in dairy cattle. Journal of Dairy Science 74:3583-3597.

 

Waghorn G C, Reed J D and Ndlovu  L R 1999 Condensed tannins and herbivore nutrition. In: Buchanan-Smith, J G Bailey and L D McCaughy (Eds.), Proceedings of the 18th International Grassland Congress, vol. 3. Association Management Centre, Calgary, pp. 153–166.

Received 15 July 2006; Accepted 5 September 2006; Published 6 December 2006

Go to top