Due to deficiency of quality feed and fodder resources, use of crop residues in livestock feeding especially for ruminants seem to be indispensable. Paddy straw is the main crop-residue available in Jammu & Kashmir state which farmers usually store for use as a ruminant feed during winter months (Anonymous, 2013). It is a good source of cellulose and hemi-cellulose but does not provide enough nutrients when fed as a sole source of roughage due to its low nutritive value, poor palatability, high lignin content, limited ruminal degradation and the presence of some anti-nutritional factors like silicates and oxalates (Ganai and Teli, 2010). Efforts are therefore directed towards assisting the animals to digest these low grade feedstuffs more efficiently for enhancing livestock productivity. Use of feed additives in ruminant ration has shown promising results for enhanced growth and production due to increased nutrient digestibility and utilization. However, ban on synthetic feed additives like antibiotics, hormones, arsenicals etc. due to residues in the animal products, has increased consumer awareness, environmental issues, health concerns and safety further encouraged exploration and use of natural feed additives in both human and animal foods.

Saccharomyces cerevisiae, a unicellular fungus, commonly used as additive in ruminant diet as it stabilizes the pH of rumen favoring the growth of cellulolytic bacteria sensitive to low pH. Oxygen scavenger property of yeast helps to protect obligate anaerobes from the air ingested in rumen along with feed intake. Improvement in digestibility and production of animals following feeding of Saccharomyces cerevisiae is well documented (Garg et al., 2009; Ganai et al., 2015). Urtica dioica a perennial herb commonly called as nettle, is a rich source of protein, fiber, vitamins A, B1, B2, E, K and C, and also provides minerals viz. iron, calcium, magnesium and trace elements like Cu, Zn, Mn and Co (Krystofova et al., 2010; Rafajlovska et al., 2013). High protein content (26%) and calcium (5.09%) present in leaves, stem and roots makes nettle a good source of nourishment (Rafajlovska et al., 2013). Keeping in view the wild nature, easy availability, nutritional and health benefits of Urtica dioica and the beneficial effects of Saccharomyces cerevisiae, the study was undertaken to use these two natural sources as feed additives at different levels on In vitro nutrient degradability of paddy straw and fermentation pattern in paddy straw-based complete feed using rumen liquor of sheep.

Materials and Methods                               

The proximate constituents of paddy straw, yeast, herb and complete feed was done as per AOAC (2000) and fibre fractions as per Van Soest et al. (1991). The In vitro studies were carried as per the procedure of Tilley and Terry (1963) in two steps. During first experiment five different level of feed additives were used to reach at optimum level of feed additive with highest in vitro dry matter (IVDMD) and neutral detergent fibre digestibility (IVNDFD). In experiment II the optimum level of feed additives were selected for study of digestibility and fermentation parameters.

Three adult male Corriedale sheep were given 20 days adoption time to complete feed containing paddy straw at 50% level to meet nutrient requirements as per ICAR (2013). Rumen fluid was collected from these sheep before morning feeding (Solvia and Hess, 2007) by perforated tubing device under negative pressure and squeezed through four layers of muslin cloth to get inoculums (SRL), then transferred to pre-warmed flask (39ºC) and flushed with CO2. SRL (10 ml) was dispensed into pre-warmed (39ºC) fermentation vessels (Jakhmola et al., 2010) containing 500 mg finely ground (1 mm) paddy straw and complete feed containing paddy straw and concentrate mixture in 50:50 ratio (concentrate mixture contained maize 6, wheat bran 7.6, deoiled rice bran 9, mustard oil cake 10, soyabean 15, molasses 0.8, mineral mixture 0.8 and salt 0.4 parts) without or with five levels of each feed additives viz., yeast (Sacchromyces cerevisiae) @  1g (T₁), 2g (T₂), 3g (T₃), 4g  (T₄), 5g (T₅) per kg dry matter and herb (Urtica dioica) @ 1g (T₁), 2g (T₂), 3g (T₃), 4g  (T₄), 5g (T₅) percent of dry matter. Paddy straw without supplementation of feed additives served as control (T₀). 40 ml of McDougall buffer is flushed with oxygen free CO₂ in each fermentation vessels. The experiment was conducted in completely randomized block design and each treatment run in triplicate with negative controls (SRL + buffer alone). The controls were used to correct for fermentation residues resulting directly from SRL. Each vessel cork fitted with control value was kept in incubator at 39ºC for 48 hr. At the end of 48hr incubation two drops of saturated HgCl2 was added in each vessel to stop microbial activity. Contents of each vessel were transferred to 1 L spout-less beakers. The vessels were thoroughly washed with neutral detergent solution and final volume made to 150 ml. The contents were refluxed for 1 hr at 100ºC, filtered and washed through preweighed Gooch crucible (Grade 1, 50 ml capacity). This undigested residue (NDF) was oven-dried at 100ºC for 24 hr, cooled in desiccator and weighed. Loss in DM and NDF was digested dry matter and digested NDF. The crucibles containing residue were ignited in muffle furnace at 500ºC and the ash left in crucible after ignition was subtracted from residual dry matter to get the organic matter content. In experiment II In vitro studies at 72 hr incubation of samples was done in same way as for 48 hr incubation. The reaction was stopped by adding 2 ml of 6 N HCl and 0.1 g pepsin powder (1:3000) to each vessel at the end of 48 hr incubation. Then the vessels were incubated for another 24 hr and procedure repeated as for 48 hr incubation, except the addition of HgCl2 at 48 hr. The optimum level of feed additives, chosen from first step, was mixed with complete feed containing paddy straw and concentrate mixture in 50:50 ratio. The experimental procedures for degradability of nutrients in complete feed were similar to experiment I. The pH in rumen liquor was determined immediately after the termination of incubation at 48 hr using portable digital pH meter. Total volatile fatty acids was determined as per method of Barnett and Reid (1957) using Markham still distillation apparatus, total rumen nitrogen as per AOAC (2000) and ammonia nitrogen by spectrophotometer method (Chaney and Marbach, 1962). The data were analyzed statistically as per Snedecor and Cochran (1994) to draw the inference.

Result and Discussion

Chemical Composition of Feed Ingredients, Supplements and Experimental Complete Feeds

The chemical composition of paddy straw and complete feed reported in this study is present in Table 1.  Results for chemical composition were in close agreement with observations of Van Soest (2006), Ganai and Teli (2010), Akinfemi and Ogunwole (2012), Khattab et al. (2013) and Sheikh et al. (2017). The chemical composition of yeast was in accordance to the existing reports of El-Ghani et al. (2004), Ganai et al. (2015) and Sheikh et al. (2017). The chemical composition of urtica dioica is in accordance with earlier reports of Andualem et al. (2016a; 2016b).

Table 1: Chemical composition of paddy straw, complete feed and feed additives

In vitro Nutrient Degradability and Fermentation Parameters of Paddy Straw

The results of In vitro nutrient degradability of paddy straw and complete feed are presented in Table 2. There was significant (P<0.01) improvement in degradability of dry matter and  NDF as an effect of supplementation of herb and probiotic at various levels, with optimum results at, 3g/kg  of yeast and 3% DM of Urtica dioica levels.

Table 2: In vitro DM and NDF digestibility of paddy straw and complete feed at 48 hrs of incubation (%DM basis)

^abcd Means superscripted with different letters in a row for a particular data differ significantly from each other **(P<0.01)

These results are comparable with the earlier report of Ganai et al. (2015) and El-Waziry et al. (2007). Sheikh et al. (2017) also reported that probiotic mix containing Saccharomyces cerevisiae and lactobacillus improved in vitro digestibility of DM, OM and NDF in Corriedale lambs fed paddy straw based complete feed. Supplementation of paddy straw and paddy straw based complete feed with Urtica dioica as feed additive resulted an increase in IVDMD and IVNDFD have been observed which is in close conformity with reports of Ganai et al. (2011b) for bajra straw, Afzal (2017) for oat straw and Sachan et al. (2014) for wheat straw based complete diet. A linear increase in in vitro nutrient degradability with the increase in the level of Urtica dioica herb supplementation upto 3% level (T₃) was observed and thereafter further inclusion of the herb had no additional effect on improvement of in vitro degradability. These results suggested that addition of Urtica dioica herb as feed additive upto 3% level might have increased the population of fibre degrading bacteria and/or their activity in the rumen (McDonald et al., 2002) as nettle leaves are rich in vitamins, minerals and trace elements. These results are in agreement with Kumar et al. (2006) who reported significantly improvement in IVDMD of paddy straw supplanted with different herbs including Urtica dioica at 72 h of incubation. Andualem et al. (2016a) also reported that stinging nettle (Urtica simensis) leaf and flowers have the potential to be used as feed supplement for ruminants based on its better effect on DMD and IVOMD.

In vitro Nutrient Degradability of Experimental Complete Feeds

The results of in vitro study of complete feed supplemented with optimum levels of feed additives (3g/kg of yeast and 3% DM of Urtica dioica) indicated significant increase in IVDMD, IVNDFD and fermentative metabolites on supplementation of herb and yeast alone and the effect was more pronounced in combination group (Table 3).

Table 3: In vitro digestibility and rumen metabolites of complete feed at 72 h of incubation (%DM basis)

^abcdMeans superscripted with different letters in a row for a particular data differ significantly from each other **(P<0.01)

These results are in line with the earlier report of Ganai et al. (2015), Malik and Singh, 2009 and Sheikh et al. (2017) who also reported higher digestibility of DM, OM and NDF values at supplementation of probiotics mix containing yeast to paddy straw based complete feed using sheep rumen liquor. Kumar et al. (2006) reported that IVDMD of paddy straw also improved significantly supplemented with Urtica dioica and can be incorporated in the diet of ruminants as feed additive for better utilization of nutrients. Similar results were observed by Wadhwa and Bakshi (2006), Mir et al. (2010), Ganai et al. (2011b) and Meel et al. (2015) who reported improved IVDMD and IVOMD on herb supplementation to different substrates.

Regarding fermentation parameters, the results suggested that addition of yeast alone as well as in combination with Urtica dioica modify rumen pH, increase TVFA, total rumen N and TCA-ppt N with non-significant effect on concentration of NH₃-N. These results are in accordance with reports of Garg et al.(2009) and Sheikh et al.(2017) suggested that supplementation of yeast causes an elevation in pH possibly due to utilization of lactic acid from ruminal contents, there by stabilizing pH (Dawson and Tricarica, 2002). An increase in TVFA concentration upon supplementation of yeast has been reported (Ganai et al., 2015 and Sheikh et al., 2017). The higher (P<0.01) level of total N upon yeast supplementation alone and in combination with herb may be attributed to possibly higher proteolytic activity. Such higher proteolytic activity in the rumen has been reported by Yoon and Stern (1996). There was non-significant effect of feed additives on concentration of NH₃-N, as reported by Oeztuerk (2009). Humphries   and Reynolds (2014) reported that adding stinging nettle (Urtica dioica) haylage to a total mixed ration in lactating dairy cows resulted better rumen pH. With higher values of TVFA, total-N as well as TCA-perceptible-N concentrations, which indicates stimulatory effect of the Urtica dioica herb on fibre degrading rumen microbes that leads to more energy availability to the animals by herb supplementation.

Conclusion

The study revealed a significant improvement in In-vitro dry matter and nutrient detergent fibre digestibility with improved in vitro rumen fermentation parameters as an effect of supplementation of yeast and Urtica dioica on paddy straw based complete feed with maximum values at 3g/kg DM and 3% respectively. However, In vivo trial is required before advocating these findings to sheep raisers.

Acknowledgements

The authors acknowledge the support of Directorate of Research, SKUAST-K, for financial help and Incharge Mountain Research Centre for Sheep and Goat for providing experimental animals.

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