• Asian-Australasian Journal of Animal Sciences
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• v.29 no.6
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• pp.807-813
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• 2016

Two experiments were conducted assessing the effects of presence or absence of rumen protozoa and dietary nitrate addition on rumen fermentation characteristics and in vitro methane production in Brahman heifers. The first experiment assessed changes in rumen fermentation pattern and in vitro methane production post-refaunation and the second experiment investigated whether addition of nitrate to the incubation would give rise to methane mitigation additional to that contributed by defaunation. Ten Brahman heifers were progressively adapted to a diet containing 4.5% coconut oil distillate for 18 d and then all heifers were defaunated using sodium 1-(2-sulfonatooxyethoxy) dodecane (Empicol). After 15 d, the heifers were given a second dose of Empicol. Fifteen days after the second dosing, all heifers were allocated to defaunated or refaunated groups by stratified randomisation, and the experiment commenced (d 0). On d 0, an oral dose of rumen fluid collected from unrelated faunated cattle was used to inoculate 5 heifers and form a refaunated group so that the effects of re-establishment of protozoa on fermentation characteristics could be investigated. Samples of rumen fluid collected from each animal using oesophageal intubation before feeding on d 0, 7, 14, and 21 were incubated for in vitro methane production. On d 35, 2% nitrate (as $NaNO_3$) was included in in vitro incubations to test for additivity of nitrate and absence of protozoa effects on fermentation and methane production. It was concluded that increasing protozoal numbers were associated with increased methane production in refaunated heifers 7, 14, and 21 d after refaunation. Methane production rate was significantly higher from refaunated heifers than from defaunated heifers 35 d after refaunation. Concentration and proportions of major volatile fatty acids, however, were not affected by protozoal treatments. There is scope for further reducing methane output through combining defaunation and dietary nitrate as the addition of nitrate in the defaunated heifers resulted in 86% reduction in methane production in vitro.

• Asian-Australasian Journal of Animal Sciences
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• v.31 no.11
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• pp.1738-1746
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• 2018

Objective: The aim of the experiment was to assess the effect of increasing amounts of Leucaena leucocephala forage on dry matter intake (DMI), organic matter intake (OMI), enteric methane production, rumen fermentation pattern and protozoa population in cattle fed Pennisetum purpureum and housed in respiration chambers. Methods: Five crossbred heifers (Bos taurus${\times}$Bos indicus) (BW: $295{\pm}6kg$) were fed chopped P. purpureum grass and increasing levels of L. leucocephala (0%, 20%, 40%, 60%, and 80% of dry matter [DM]) in a $5{\times}5$ Latin square design. Results: The voluntary intake and methane production were measured for 23 h per day in respiration chambers; molar proportions of volatile fatty acids (VFAs) were determined at 6 h postprandial period. Molar concentration of VFAs in rumen liquor were similar (p>0.05) between treatments. However, methane production decreased linearly (p<0.005), recording a maximum reduction of up to ~61% with 80% of DM incorporation of L. leucocephala in the ration and no changes (p>0.05) in rumen protozoa population were found. Conclusion: Inclusion of 80% of L. leucocephala in the diet of heifers fed low-quality tropical forages has the capacity to reduce up to 61.3% enteric methane emission without affecting DMI, OMI, and protozoa population in rumen liquor.

• Asian-Australasian Journal of Animal Sciences
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• v.15 no.10
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• pp.1439-1444
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• 2002

An objective of this study was to determine the effects of increasing contents of rumen undegradable protein (RUP) from formalin treated soy bean (FSBM) on rumen functions. Four rumen canulated non-lactating cows were randomly allocated to total mixed rations (TMR) containing different proportions of soy bean meal (SBM) and FSBM. Of rumen fermentation characteristics, concentrations of ruminal fluid ammonia and molar proportions of isoacids decreased with increasing contents of RUP in diets (p<0.01). The animals on TMR containing only SBM gained less weight and had smaller rumen volume than those on TMR containing RUP from FSBM (p<0.05). Organic matter and neutral detergent fiber digestibility in sacco were not different (p>0.05). The density of protozoa particularly small Entodinium sp. in ruminal fluid was higher in animal fed TMR containing SBM:FSBM (34:66) and FSBM than those fed TMR containing SBM:FSBM (66:34) and SBM (p<0.01). Total viable count, and net microbial protein synthesis as indicated by purine derivatives in urine increased with increasing contents of RUP from FSBM (p<0.01). It can be concluded that a reduction in net microbial protein synthesis in the rumen with increasing contents of RUP in the diet can be due to the reduction of preformed protein available for microbial growth as well as an increased turnover rate of microbial cells by predatory activity of protozoa.

• Asian-Australasian Journal of Animal Sciences
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• v.2 no.3
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• pp.475-476
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• 1989

• Asian-Australasian Journal of Animal Sciences
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• v.18 no.1
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• pp.38-41
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• 2005

An experiment was conducted on 3 male rumen fistulated adult buffaloes fed on wheaten straw and concentrate mixture in a Latin square design to study the impact of niacin supplementation on rumen metabolites. Three animals were fed wheaten straw+concentrate mixture (group I, control), wheaten straw+concentrate mixture+100 ppm niacin (group II), and wheaten straw +concentrate mixture+200 ppm niacin (group III). After 21 days feeding, rumen liquor was drawn for 3 consecutive days at different time intervals (0, 2, 4, 6 and 8 h) to study the various rumen metabolites i.e., rumen pH, ammonia-N, total-N, trichloroacetic acid precipitable-N, non-protein nitrogen, total volatile fatty acids, their fractions and number of protozoa. Mean pH values in strained rumen liquor (SRL) of animals in 3 groups were 6.64, 6.71 and 6.67, indicating no statistically significant difference. Results revealed a significant (p<0.01) increase in TVFA concentration among the supplemented groups (group II and III) in comparison to control group. Mean TVFA concentration (meq/dl) was 9.75, 10.97 and 11.44 in 3 groups respectively. The highest concentration of TVFA was observed at 4 h and minimum at 0 h in all the 3 groups. The percentage of acetic, propionic, butyric and isobutyric acid was statistically similar among the three groups. The mean ammonia-N concentration (mg/dl SRL) was significantly (p<0.01) lower in group II (16.38) and group III (15.42) than group I (18.14). Ammonia-N concentration was higher (p<0.01) at 4 h as compared to all the time intervals. The mean total-N concentration (mg/dl SRL) was higher (p<0.01) in group II (74.16) and group III (75.47) as compared to group I (62.04). Total-N concentration was higher (p<0.01) at 4 h as compared to other time intervals and lowest value was recorded at 0 h.Concentration of TCA-ppt-N (mg/dl SRL) was significantly (p<0.01) lower in control group as compared to niacin supplemented groups. Mean value of NPN (mg/dl SRL) was significantly (p<0.01) lower in group III (23.21) as compared to group I (25.71), whereas groups I and II, and groups II and III were similar to each other. Total protozoa number (${\times}10^4$/ml SRL) ranged from 18.06 to 27.41 in group I, 20.89 to 38.44 in group II and 27.61 to 39.45 in group III. The mean protozoa number was significantly (p<0.01) higher in SRL of group II (27.60) and III (30.59) as compared to group I (22.48). It can be concluded from the study that supplementation of niacin in the diet of buffaloes had improved the rumen fermentation by decreasing the concentration of ammonia-N and increasing protein synthesis.

• Asian-Australasian Journal of Animal Sciences
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• v.11 no.6
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• pp.661-672
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• 1998

Two experiments were conducted to examine the effects of a range of concentrations of ruminal fluid ammonia ($NH_3$-N) on forage digestibility, microbial growth efficiency and the mix of microbial species. Urea was either continuously infused directly into the rumen of sheep fed 33.3 glh of oaten chaff (Exp. I) or sprayed onto the oaten chaff (750 g/d) given once daily (Exp. 2). Concentrations of $NH_3$-N increased with incremental addition of urea (p < 0.01). Volatile fatty acids (VFA) concentrations and 24 h in sacco organic matter digestibility in the rumen were higher when supplemental urea was given (p < 0.01). The (C2 + C4) : C3 VFA ratio was lower (p < 0.05) when $NH_3$-N was above 200 mgN/I. The fungal sporangia appearing on oat leaf blades were significantly higher when urea was supplemented, indicating that $NH_3$-N was a growthlimiting nutrient for fungi at levels of $NH_3$-N below 30 mgN/l. The density of protozoa was highest when $NH_3$-N concentrations were adjusted to 30 mgN/I for continuously fed ($4.4{\times}10^5/ml$) and to 168 mgN/1 for once daily feeding ($2.9{\times}10^5/ml$). Thereafter increasing concentrations of $NH_3$-N, were associated with a concomitant decline in protozoal densities. At the concentration of $NH_3$-N above 200 mgN/l, the density of protozoa was similar to the density of protozoa in ruminal fluid of the control sheep ($1.8{\times}10^5/ml$). The efficiency of net microbial protein synthesis in the rumen calculated from purine excretion was 17-47% higher when the level of $NH_3$-N was above 200 mgN/1. The possibilities are that 1) there is less bacterial cell lysis in the rumen because of the concomitant decrease in the protozoal pool and/or 2) microbial growth per se in the rumen is more efficient with increasing $NH_3$-N concentrations.

• Asian-Australasian Journal of Animal Sciences
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• v.20 no.8
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• pp.1196-1200
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• 2007

Eight feeds (mixture of wheat straw and oil seed cakes in 3:1 ratio) were evaluated for methane emission and fermentation pattern with buffalo rumen liquor as inoculum in an in vitro gas production test. The cakes tested were groundnut cake (GNC), soybean cake (SBC), mustard seed cake (MSC), cotton seed cake (CSC), karanj seed cake expeller extracted (KCEE), karanj seed cake solvent extracted (KCSE), caster bean cake expeller extracted (CBCEE) and caster bean cake solvent extracted (CBCSE). The gas production (ml/g dry matter) was significantly higher with SBC and MSC followed by CSC, GNC, KCSE, KCEE, CBCSE and was the lowest with CBCEE. Methane emission was significantly lower with KCEE, KCSE, CBCEE, CBCSE (20.32- 22.43 ml/g DM) than that with SBC, GNC, CSC (27.34-31.14 ml/g DM). Mustard seed cake was in-between the two groups of oil cakes in methane production. In vitro true digestibility was highest with SBC followed by GNC, CSC, MSC, KCSE, KCEE, CBCSE and CECEE. Ammonia nitrogen level was positively correlated with the amount of protein present in the cake. Total holotrich protozoa were significantly higher with SBC, whereas, large spirotrich protozoa tended to be lower than with other cakes. The counts of small spirotrich and total protozoa were similar with all the cakes. Total volatile fatty acid production and acetate to propionate ratio were significantly higher with SBC and significantly lower with KCEE as compared to the other cakes. Among the conventional oil cakes tested in the present experiment (GNC, SBC, MSC and CSC), mustard seed cake-based feed produced the minimum methane without affecting other fermentation characteristics adversely.

• Korean Journal of Veterinary Research
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• v.35 no.2
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• pp.327-336
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• 1995

This study was carried out to develop the animal protein(feed for fry) that was isolated, purified and lyophilized the rumen ciliates from the healthy rumen contents which have $10^5-10^6/g$ ciliates and were discarded in abattoirs. The rumen ciliates are non-pathogenic, anaerobic and the weight of this protozoa is 2% of rumen content. The rumen protozoan and bacterial proteins both have a biological value for rats of 80-81, which is higher than the 72 of brewer's yeasts. Furthermore, the true digestibility and net protein utility of the protozoan protein are 91 and 73, much higher than those of bacterial(74 and 60) or yeast(84 and 60) proteins. The amino acids of rumen protozoa is nutritionally superior than the others. The size of rumen ciliates is $30-200{\times}20-110{\mu}m$ and so we had isolated and purified the rumen ciliates from the rumen contents by the physical methods. The purified rumen protozoa was lyophilized with freezing dryer. The results of this experiment were as follows : 1. Population dynamics of protozoan ciliates in slaughtered rumens; % of samples which small ciliates were predominated was 82.5%(52/63) and that of large ciliates was 17.5%(11/63). 1) predominant species of small ciliates were Entodinium ovinum and E nanellum. 2) predominant species of large ciliates were Epidinium ecaudatum and Diploplastron affine. 2. The lyophilized rumen ciliates which were isolated and purified from 1 kg of rumen content at the pH 6.2-6.8 was about 7.0 gram. 3. The nutrient analysis of lyophilized rqmen ciliates(LRC) was as follows: 1) Proximate analysis of the LRC and the composition of fry feed; moisture 8.05%(below 10.0), protein 35.37%(45), fat 5.39%(4.5), fiber 1.23%(below 2.5), ash 2.25%(below 15.0), Ca 0.26%(below 2.0), P 0.14%(below 1.1), energy 4,608.11(fish meal 5000 cal/g) 2) Amino acids (% in crude protein) of the LRC and the rotifer(Brachionus plicatilis); Arg 5.19%(4.50), His 2.50%(1.55), Ile 5.29%(3.45), Leu 8.11%(5.85), Lys 10.34%(6.15), Met 2.25% (0.85), Phe 5.66%(3.80), Thr 5.14% (3.45), Val 4.18%(3.90), Ala 4.13%(3.35), Asp 13.26%(8.25), Glu 16.62%(9.20), Gly 4.23%(3.10), Pro 3.25%(5.05), Ser 4.85%(3.85), Tyr 5.04%(3.05) 3) Fatty acids(% in fat) of the LRC and the rotifer(biological feed ; Brachionus plicatilis); myristic acid(C14:0) 3.27%(0.3), myristoleic acid(C14:1) 0.83%(-), palmitic acid(C16:0) 39.11% (23.5), palmitoleic acid(C16:1) 2.81%(2.0), stearic acid(C18:0) 9.36%(5.6), oleic acid(C18:1) 25.54%(3.5), linoleic acid(C18:2) 15.05%(32.9), linolenic acid(C18:3) 1.74%(9.8). Judging from the above investigated results, the analytical data of proximate analysis, amino acids, fatty acids of the purified and lyophilized rumen protozoa are reasonable for the feed of freshwater fishes(fry and fingerling). But it was disappointed of our expectation that the crude protein of lyophilized rumen ciliates contains low percentage, it was thought that because of the small ciliates(starch digester) in beef cattle rumens which were administered the concentrated feed, is much difficult to isolate and purify than the large ciliates(fiber digester).

• Asian-Australasian Journal of Animal Sciences
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• v.8 no.3
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• pp.249-254
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• 1995

The effects of bentonite (B) on rumen protozoal population and rumen fluid characteristics of sheep fed palm kernel cake (PKC) were studied for a period of 21 days. Two groups, each comprising two sheep were fed either PKC or PKC + B ad libitum A third group was left at pasture. Rumen fluid was sampled through a rumen cannula three times daily from all animals. Palm kernel cake contained 16% crude protein, 1 % crude fat and high amounts of copper, zinc, iron and manganese. Protozoal population in the rumen fluid decreased significantly (p < 0.05) after the onset of feeding PKC or PKC + B. However, sheep given bentonite supplementation at 2% of the dietary dry matter, maintained higher protozoal densities ($15{\times}10^4/ml$) when compared to animals fed only PKC ($8{\times}10^4/ml$). With both diets, the protozoa were mainly of the small entodinia species. Animals at pasture had higher protozoal population ($47{\times}10^4/ml$) with varying species of entodiniomorphids and holotrichs. Rumen fluid pH and ammonia concentration was significantly (p < 0.05) higher in animals at pasture compared to animals fed PKC or PKC + B. Volatile fatty acid concentration was significantly (p < 0.05) lower in animals fed PKC when compared to animals at pasture. There was a shift in fermentation pattern in animals fed PKC or PKC + B towards a lower acetate; and higher propionate, isovalerate and valerate. Studies in vitro also showed the positive effect of bentonite on protozoal numbers.

• Asian-Australasian Journal of Animal Sciences
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• v.12 no.6
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• pp.988-1001
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• 1999

The rumen ecosystem is increasingly being recognized as a promising source of superior polysaccharide-degrading enzymes. They contain a wide array of novel enzymes at the levels of specific activities of 1,184, 1,069, 119, 390, 327 and $946{\mu}mol$ Reducing sugar release/min/mg protein for endoglucanase, xylanase, polygalactouronase, amylase, glucanase and arabinase, respectively. These enzymes are mainly located in the surface of rumen microbes. However, glycoside-degrading enzymes (e.g. glucosidase, fucosidase, xylosidase and arabinofuranosidase, etc.) are mainly located in the rumen fluid, when detected enzyme activities according to the ruminal compartments (e.g. enzymes in whole rumen contents, feed-associated enzymes, microbial cell-associated enzymes, and enzymes in the rumen fluid). Ruminal fungi are the primary contributors to high production of novel enzymes; the bacteria and protozoa also have important functions, but less central roles. The enzyme activities of bacteria, protozoa and fungi were detected 32.26, 19.21 and 47.60 mol glucose release/min/mL mediem for cellulose; 42.56, 14.96 and 64.93 mmol xylose release/min/mL medium after 48h incubation, respectively. The polysachharide-degrading enzyme activity of ruminal anaerobic fungi (e.g. Neocallimastix patriciarum and Piromyces communis, etc.) was much higher approximately 3~6 times than that of aerobic fungi (e.g. Tricoderma reesei, T. viridae and Aspergillus oryzae, etc.) used widely in industrial process. Therefore, the rumen ecosystem could be a growing source of novel enzymes having a tremendous potential for industrial applications.