High performance of growing lambs grazing Paspalum notatum INIA Sepé with energy-protein supplement including sorghum-DDGS

Sorghum dried distillers’ grain with soluble (S-DDGS) has not been utilized as supplement in lamb growing and finishing diets under grazing conditions in summer. The hypothesis was that the performance of post-weaning lambs grazing Paspalum notatum (PN) INIA Sepé supplemented with an energy-protein diet (whole sorghum grain [SG], and soybean meal [SM]) is not reduced by the inclusion of 40% S-DDGS. During 82 days, in summer, 42 Merino Dohne male lambs (130 ± 7.6 days of age, 32 ± 2.4 kg of body weight [BW], and 2.2 ± 0.2 body condition score [BCS]) were evaluated, in a completely random experimental design with three treatments and two replicates each (n = 14 each one). All lambs grazed PN INIA Sepé and in two treatments they were daily supplemented (1.7% BW) with different components. The treatments were: PN (control group, no supplementation), SGSM (70% SG, 30% SM) and DDGS (45% SG, 40% S-DDGS, and 15% SM). Supplemented lambs had higher (P<0.0001) body weight gain (BWG), final BW and wool growth (140 and 126 g/a/d, 43.5 and 42.4 kg; 1814 and 1892 μg/cm2/d for DDGS and SGSM, respectively) than control lambs (40 g/a/d; 35.5 kg; 1353 μg/cm2/d) (P<0.0001), without differences between supplemented groups. Gastrointestinal nematodes infestation (GIN) was affected by the treatments on day 69, where supplemented animals showed lower infestation levels than control lambs (P=0.0024). The inclusion of S-DDGS in the supplement did not reduce the productive performance of the lambs when the concentration of crude protein and energy remained constant.


Introduction
Weaning can be defined as a complete physical separation between dam and lamb, and is a stressful time for lambs, and a critical management period (1) . During post-weaning, the first year of life, lambs have relative higher energy and crude protein (CP) requirements than adult sheep (2) . Furthermore, grazing animals have even higher maintenance energy requirements than feedlot or housed animals, partly because of forage search and harvesting activities (3) . In the first year of life, important mortality rates have been reported in Merino lambs, which vary between 4.5% to 26.8%, and represent high costs for farmers and animal welfare concerns (4) .
The mortality risk is higher in lambs with less than 22 kg at weaning, and it decreases with increasing body weight (BW) (4) . Also, growth rate post-weaning should be considered, which ideally should not be less than 50 g/a/d (4) . However, lambs grazing native summer pastures might not reach a high growth rate and wool production (5) . Another concern when grazing lambs are gastrointestinal nematodes (GIN) infestations, which are a serious challenge to the health and performance of ruminants (6) . A GIN infestation decreases dry matter intake, increases endogenous protein loss into the gastrointestinal tract, as well as reorients the use of nutrients to repair damaged tissue instead of production functions. Of the several aspects of integrated control against GIN (7) , feeding strategy has a major impact. Therefore, inadequate post-weaning nutrition limits lamb growth and development, increasing their susceptibility to GIN and increasing morbidity.
The Rio de la Plata Grassland (28°S-38°S, 50°W-61°W) is characterized by the dominance of grasslands and is one of the largest natural ecoregions of the world. These grasslands in South America (campos in Portuguese, pastizales or pampa in Spanish) include three countries: Uruguay (fully included), central-eastern Argentina and part of southern Brazil (8) . Native pastures are highly diverse with the coexistence of C3 and C4 species. Growth during summer is dominated by C4 grasses, with Paspalum notatum (PN) as one of the most frequent species in native pastures. In line with this, the genetic variability of PN in Uruguay has been analyzed and valued. The product of this largescale germplasm collection and evaluation was Paspalum notatum INIA Sepé (9) . Similarities in chemical composition between native pastures and PN INIA Sepé can be observed, but not in the growth rate, where INIA Sepé is much higher. During summer, native pastures have low quality, decreasing the growth potential rate of lambs (10) . The crude protein (CP) content varies between 6 and 15% according to season, maximum in winter and early spring, and minimum in summer. The dry matter digestibility (DMD) varies between 50 and 61%, while metabolizable energy (ME) does so between 1.8 and 2.2 Mcal/kg DM (11) . The DMD and CP contents are related to ruminal degradability and function, digesta kinetics, mean retention time and level of intake; the nutritive values of summer can limit animal performance (12) (13) (14) . In this situation, supplemental feeding is necessary to complement nutrients, reduce or avoid protein deficiency, and enhance relative intake (15) .
The plant-based ethanol industry is growing, and a primary by-product called dried distillers' grain with soluble (DDGS) is an excellent energy and protein source for ruminants. Rations with 30% of corn DDGS resulted in the highest gross margin and economic efficiency with the lowest fed cost per kg of weight gain (16) . Traditional supplements like soybean meal (SM) or yellow corn can be replaced by DDGS (sorghum, corn or barley (17) ). Particularly, sorghum DDGS (S-DDGS) has a high nutritional value (30.2% CP; 9.4% ether extract; 2.8 Mcal ME/kg DM, and 46 % neutral detergent fibre), about 60% of the protein is rumen by-pass protein and it is highly palatable (18) (19) . Finishing steers fed with SM, S-DDGS and corn DDGS (200 g/kg DM) showed no differences in performance among diets (20) . Different studies evaluated dietary corn DDGS levels for growing and finishing lambs. Inclusion of corn DDGS from 20%, 30% up to 60% has been used as an energy and protein source (21) (22) , increasing about 25% of body weight gain (BWG) with level 30% corn DDGS (16) . Therefore, DDGS has a high nutritional value, can be used in lamb diets and is less costly for the farmer than other feed sources, such as corn or SM (22) . However, S-DDGS has not been utilized as a supplement in lamb growing and finishing diets when grazing.
This study aims to evaluate the effect of partial substitution of SG and SM by S-DDGS on the performance of post-weaning lambs grazing Paspalum notatum INIA Sepé, in summer. The working hypothesis was that the performance of post-weaning lambs grazing Paspalum notatum INIA Sepé supplemented with an energy-protein diet (SG and SM) is not reduced by the substitution of 40% of the supplement with S-DDGS.

Animals and management
Forty-two Dohne Merino male lambs were weaned on pasture (96 ± 6.9 days of age, 28.8 ± 2.8 kg of BW) and supplemented with commercial ration (at 1.5% of BW; CP 16%) until the start of the experiment. The average age of lambs at the beginning of the experiment (Day 0) was 130 ± 7.6 days, with an average BW of 32 ± 2.4 kg, and body condition score (BCS, score 1 to 5 (23) ) of 2.2 ± 0.2. The lambs had free access to clean water, mineral blocks and collective shade (1.8 m 2 /lamb) during the experiment. All lambs were drenched by a suppressive dose at the beginning of the experiment (Day 0) to GIN control (Naftalophos 15%; Vermkon®, Lab König, Buenos Aires, Argentina; 35 mg kg BW -1 ). Every 14 days, faeces were sampled from all lambs. Whenever the faecal egg count (FEC) was over 500 eggs per gram of faeces in 50% of animals/plot and anaemia score (Famacha®, 1 = optimal to 5 = fatal) was above 3 in 20% of animals/plot, the lambs were drenched. The Famacha® test was used as support tool when making the decision to drench. All the animals were drenched on Day 50 (03/3; Naftalophos 15%) and Day 78 (03/31; Ivermectina 0.2% plus Levamisol 8% plus Rafoxanide 7.5%; Ranizole®, Lab Cibeles, Montevideo, Uruguay; 1 mL kg BW -1 ).

Supplement management
The adaptation period lasted 13 days, until the target intake level was reached (1.70 % of BW). Every 14 days, adjustments of the amount of supplement were made, according to the average BW of each replicate. The lambs received half the daily ration twice a day at 9 am and 5 pm. The linear space of the feeder was more than 30 cm for each animal.

Forage management
The lambs grazed on Paspalum notatum INIA Sepé, which was sown in November 2015 (7 kg/ha, two fertilizations with granulated urea). The grazing system was continuous with a fixed stocking rate of 24 lambs/ha. The height of pasture was maintained at approximately 17.2 cm (ruler) and 12 cm (rising plate meter [RPM]), and five mechanical interventions with a mower were needed (Days 7, 14, 29, 43 and 60) to maintain pasture within the expected height.

Animal measurements
Every 14 days the lambs were weighed unfasted, and BCS and Famacha® were recorded by the same person throughout the study. Wool growth was measured with the mid-side patch method (24) using Herons area formula: = √s (s-a) (sb) (s-c) (25) . Wool growth rate was calculated with the following equation: [(wool weight (g) / patch's area (cm 2 ) / days of experiment) × 1000000]. Feed conversion of the supplement was calculated using the following formula: [(kg supplement (DM)) / (kg BW (SGSM or DDGS) -kg BW control (PN))]. Production of BW per hectare was calculated as [(BWG × total days of the experiment / 1000) × (stocking rate)].

Pasture measurements
Herbage mass dry matter (kg DM/ha) of each plot was estimated weekly within six quadrants of 20 cm × 50 cm (0.1 m 2 ). Pasture was cut with a manual scissor (above 3 cm), each sample was individually fresh weighed and then pooled. Two subsamples were fresh weighed (each 100 g) and dry weighed (after 72 h, at 60 °C) for dry matter content estimation. One of them was separated into fresh and dead forage and weighed. The average of these estimations was applied to each of the six fresh weights. Herbage mass (kg DM/ha) was estimated using the following equation: [(fresh weigh × % DM × 10000)/0.1/1000]. Pasture height was measured using RPM (50 times) and ruler (20 times), and normalized difference vegetation index (NDVI) was estimated through a 25 m diagonal per plot with a GreenSeeker® (Trimble®, Sunnyvale, USA). Botanic composition was estimated to register the species with the highest frequency (26) .

Pasture and supplement chemical composition
To estimate the pasture and supplement chemical composition, samples were sent to the Animal Nutrition Laboratory of INIA La Estanzuela (Colonia, Uruguay). For pasture, ground samples analysed CP according to the procedure described by AOAC (27) ; ME was calculated according to the ARC (28) (29) , and analytic dry matter was estimated according to AOAC (30) . For supplement chemical composition, 200 g of each component (SG, S-DDGS and SM) were sent to the laboratory. In addition to the same variables as the pasture, neutral detergent fibre and ashes were estimated according to the procedure described by AOAC (30) . Results are reported on a dry basis. The CP and ME data of each supplement were extracted from the laboratory analysis and from FEDNA table (Spanish Foundation for the Development of Animal Nutrition) for CP and ME, respectively, and the total value of CP and ME offered by the diet was calculated according to the proportion of each component. The information on forage and supplement offered is presented in Table 1.

Statistical analysis
Animal and forage data were analysed using a mixed model of the statistical package Infostat (Grupo Infostat Student, FCA, Universidad Nacional de Córdoba, Argentina) (31) . Animal model included 'replicates', 'diet' and their 'interaction' as fixed effects, while 'lamb' and 'plot' were random effects. Initial BW and initial BCS were used as covariates for subsequent analysis of those variables. Body weight gain and FEC were analysed as repeated measures over time, 'date' and their interaction with 'diet' were included in the animal model as fixed effects. Body weight gain was calculated based on the difference between BW measurements. For the analyses of forage related traits (herbage mass, sward height and NVDI) the model included 'diet' and 'replicates' as fixed effects, while 'plot' was a random effect. Differences were considered statistically significant when P<0.05.

Transparency of data
Data not available: The data set that supports the results of this study is not publicly available.

Results
An effect of supplementation (P<0.0001) but not on the type of supplement (P>0.05) was observed in some of the variables analysed: BW, BWG, wool growth and productivity per hectare (Table 2). On the other hand, differences in final BCS were observed among all the treatments ( Table 2). The use of one or another source of nutrients, SGSM or S-DDGS, does not alter animal performance (16) . The nutritional contribution in terms of CP and ME offered by both supplements was similar (Table 1), and animals did not differ in their productive performances. According to Grazfeed software (33) , the net energy gain efficiency for the supplemented treatments was the same, 0. 39  This partition of energy could be the basis for the difference observed in the performance of the animals. Supplementing lambs in the rearing stage during the summer period could be an appropriate tool to achieve higher animal performances.
Strategic supplementation is a tool used for animals grazing on pastures of different quality to add energy and protein to their diets, especially to categories with high requirements (34) . This also allows higher stocking rates in grazing-based production systems. The results of BW, BCS and BWG obtained in the present study were similar to those reported by Piaggio and others (35) , and Ramos and others (5) . Although it is worth noting the important difference in stocking rate. In this study, the use of PN contributed to being able to use higher stocking rate (24 lambs/ha) than other experiments evaluated in native pastures (10 lambs/ha). According to Grazfeed predictions (33) , there could have been a possible substitution of forage by supplements in the supplemented groups, because estimated total intake of control treatment (PN) and supplemented treatments (SGSM, DDGS) was similar (PN = 1.63; SGSM = 1.61; DDGS = 1.55 kg/DM/a/d). Therefore, the intake of protein and energy may have been higher for the supplemented lambs than the nonsupplemented ones, allowing the animals to achieve higher BWG. The assumed substitution would not only have allowed the animals to decrease their energy requirements (less grazing time) and improve their grass selectivity, but would also have given them the opportunity to create an improved diet in terms of ME, protein content, in favour of better animal performance. On the other hand, the supplementation allowed to triple the productivity per hectare. The results reported by Piaggio and others (35) are lower than those of the present study, 46 kg BW/ha at a rate of 10 lambs/ha in native pastures, while for the treatment of native pasture and supplementation with soybean expeller the value was 71 kg BW/ha. Additionally, the treatments ended with different stocking rates in BW terms (35.5*24=852 versus 43.5*24=1044), 20% higher in the supplemented treatments. Supplementation of animals with high requirements is a valid tool that allows adding energy and protein to diets of animals grazing on pasture of varying quality, and permits higher stocking rates, resulting in increased productivity per hectare.
Many studies indicate a linear relationship between BW and BCS in sheep, but the increase in BW for each BCS unit varies by breed (36) . In this experiment, the difference detected in BCS between control and supplemented lambs was 1.4 units and was associated with a change in BW of 7.45 kg. Our results agree with Coop and Sykes (37) , who say that higher weight gain rates lead to higher proportions of animal fat, improving body condition and body weight. In this way, the supplemented lambs would have the adequate BW and BCS for slaughter according to the Uruguayan meat industry requirements. The higher BW and BCS achieved by the supplemented lambs were due to higher daily gains of these lambs compared to control lambs. This better performance achieved by the supplemented lambs reveals a possible association between BWG, FEC and rainfall.
The weather conditions that occurred during the current experiment were beneficial for the development of the nematodes. These nematodes affected the animals differently depending on the treatment to which they were subjected, which can be seen in Figure 2 (FEC). Clearly, this different infestation, added to other factors, such as nutrition, resulted in different BWG between animals ( Figure 1).
The wool growth of grazing sheep depends largely on their genetic potential and their ability to meet their energy and amino acid requirements on pasture (38) . Lower wool growth was associated with lower quality diet (PN), while higher wool growth was observed in the supplemented lambs, with no differences between them (SGSM, DDGS). These results are in agreement with Kempton and others (39) , who reported that the availability of energy and protein determines the wool growth and the BWG, with the availability of protein as the limiting factor, which, when increased, stimulates wool growth and BWG. Wool growth trends were essentially similar to those of body weight, which were similar to the results found by Ramos and others (5) , who obtained the maximum wool growth with the treatment with 20% CP. The authors argued that the differences found in the protein/energy content of the diet, and the additional estimated intake of the supplement, would provide more protein and energy, and therefore increase wool production. In this sense, Reis and others (40) report that increases in the protein content of the diet favour the production of wool, while feeds with high energy content increase the production of wool only if the protein level is adequate. On the other hand, Nagorcka (41) mentions a directly proportional relationship between the estimated intake of digestible feeds and wool growth.
In susceptible categories such as weaners, sanitary control is key to achieve adequate animal performance, and FEC acquires practical importance, monitoring an epidemiological situation that may be unstable. The FEC varied during the experimental period, which could be due to various factors, such as the environmental conditions (temperature and rainfall), and the effect of diet, as the control treatment presented higher levels of GIN infestation. The results of higher levels of FEC in lambs of the control group could be due to a greater parasite challenge, as reported by Ramos and others (5) , as lambs fed exclusively on pastures spend more time grazing (47% extra) than supplemented animals. Higher pasture intake would be equal to higher larvae (L3) intake (5) . Additionally, there is a direct relationship between the number of animals per surface area and the number of larvae (L3), where a higher parasite load implies a greater number of infested animals (42) . In the present study this aspect would be relevant given the high stocking rate utilized (24 lambs/ha). This could be indicative of a relationship between the nutritional level and the susceptibility of the animal to parasite infestation, showing that a diet with a higher contribution of nutrients (energy and protein) leads to lower parasite infestation in animals. In this sense, Coop and Sykes (37) establish that protein supplementation has a positive effect on the expression of immunity against GIN; and Goldberg and others (43) mention that better-fed lambs will be less susceptible to parasitic infestations. The lambs presented differences in their productive performance at the same parasite load, which may be evidence for increased resilience. The epidemiological cycle of GIN is governed by contamination of the pasture and its transfer to the animal. Infestation of the animal will be seasonally limited by the supply of larvae from the pasture, with Haemonchus contortus being the most prevalent nematode in the summer if the weather conditions are favourable (42) . The high parasite load negatively and differentially affected the productive performance of the lambs. The diet had a clear effect on the FEC, allowing it to be lower in the supplemented treatments, associated with better nutritional balance and resilience (6) .

Conclusions
The partial substitution of the supplement (SGSM) with S-DDGS allowed similar animal performances maintaining the crude protein and energy concentrations in the final supplement. The productive performance of lambs, evaluated as body weight, body condition score, body weight gain, wool growth, and productivity per hectare, was affected by supplementation, as well as gastrointestinal nematodes infestation. Supplementation gave the animals a greater opportunity to resume productivity in the face of an adverse epidemiological situation.

Author contribution statement
ER: data collection, results interpretation and article writing; AIT: data collection, results interpretation and article writing; EvL: experiment design, results interpretation and article writing; RR: experiment design and results interpretation; IDB: experiment design, results interpretation and article writing.
Authors would like to indicate that Romaniuk and Tafernaberry are co-first authors, having contributed equally to the article.