L. C. Chaudhary

Microbial status and rumen enzyme profile of crossbred calves fed on different microbial feed additives

Aims: To test various microbial cultures as cattle feed additives.

Effect of plant extracts on methanogenesis and microbial profile of the rumen of buffalo: a brief overview

Plants rich in secondary metabolites (saponins, tannins, essential oils, etc.) have antimicrobial activity which can be exploited for selective inhibition of a particular group of microbes in the rumen. We have screened a large number of plant extracts for their potential to inhibit methanogenesis and ciliate protozoa in an in vitro gas production test using buffalo rumen liquor as the inoculum. Out of 93 plant extracts tested, 11 inhibited in vitro methanogenesis to the extent of 25–50% and nine plant extracts inhibited methanogenesis more than 50%. Among 20 extracts exhibiting antimethanogenic activity, nine were ethanol extracts, 10 were methanol extracts and only one was a water extract. Some of these plant extracts inhibited ciliate protozoa as tested by microscopic examination and 14C-labelled radioisotopic technique, but the protozoa inhibition was not correlated with methane inhibition, indicating that the methanogens sensitive to plant secondary metabolites may or may not be having any symbiotic relationship with ciliate protozoa. Methane inhibition was accompanied by a drastic fall in the number of methanogens as determined by real time PCR. Plants that appeared to have some potential as feed additives to control methanogenesis by the ruminants are: (i) seed pulp of Sapindus mukorossi (rich in saponins) and Terminalia chebula (rich in tannins); (ii) leaves of Populus deltoides, Mangifera indica and Psidium guajava (rich in tannins and essential oils); and (iii) flower buds of Syzygium aromaticum and bulb of Allium sativum (rich in essential oils). Some of the plants reported in literature exhibiting antimethanogenic activity include Equisetum arvense, Lotus corniculatus, Rheum palmatum, Salvia officinalis, Sapindus saponaria, Uncaria gambir and Yucca schidigera.

Selection of Saccharomyces cerevisiae strains for use as a microbial feed additive

Saccharomyces cerevisiae ITCCF 2094, NCIM 3052, 1031, 1032, NCDC 42, 45, 47, 49 and 50 were screened for their tolerance to pH 2·0–7·0, various concentrations (0·00, 0·10, 0·25 0·50 and 1·0%) of a mixture of acetic, propionic and butyric acids (70:20:10), and bile salts (0·00, 0·30, 0·60 and 0·90%). Low pH (2·0–4·0) and addition of organic acids or bile salts in the medium inhibited the growth of all the strains tested, but the percentage of inhibition was variable in the different strains of yeast. Two of the strains showing maximum tolerance, 42 and 49, were further tested for in vitro dry matter degradability (IVDMD) using green berseem, wheat straw and oat hay as substrates. Saccharomyces cerevisiae 49 enhanced the IVDMD of berseem and wheat straw whereas S. cerevisiae 42 was ineffective. Based on the results of the present experiment, S. cerevisiae NCDC 49 can be considered as the best strain which might tolerate the adverse conditions in the gastrointestinal tract when used as a live microbial feed supplement in the diet of the animals.

Effect of Sapindus mukorossi Extracts on in vitro Methanogenesis and Fermentation Characteristics in Buffalo Rumen Liquor

Abstract Agarwal, N., Kamra, D.N., Chaudhary, L.C. and Patra, A.K. 2006. Effect of Sapindus mukorossi extracts on in vitro methanogenesis and fermentation characteristics in buffalo rumen liquor. J. Appl. Anim. Res., 30: 1–4. The berries of Sapindus mukorossi (soap nut) extracted in water, methanol and ethanol were tested for their effects on methanogenesis and fermentation of feed with buffalo rumen liquor in in vitro gas production test. The degression in methane and gas production was 96% and 39.4%, 20% and 11.5%, 22.7% and 0% with ethanol, water and methanol extracts, respectively, as compared to respective controls. The extracts also exhibited antiprotozoal activity resulting in 70–90% lesser protozoa count in the treated samples. The proportion of acetate was lower (P<0.05) and that of propionate higher (P<0.05) resulting in a decrease in acetate/propionate ratio with all the three extracts tested. The pH of incubation medium containing extract was significantly lower as compared with the medium without any extract. There was a significant depression in in vitro dry matter degradability of feed with all the three extracts. The data indicated that soapnut extracts appear to have a potential to be used as antimethanogenic and antiprotozoal agents.

Diurnal variations in the activities of hydrolytic enzymes in different fractions of rumen contents of Murrah buffalo.

Abstract Agarwal, N., Agarwal, I., Kamra, D.N. and Chaudhary, L.C. 2000. Diurnal variations in the activities of hydrolytic enzymes in different fractions of rumen contents of Murrah buffalo. J. Appl. Anim. Res. 18: 73–80. The hydrolytic enzymes like carboxymethylcellulase, α-amylase, xylanase, microcrystalline cellulose and filter paper degrading activities were estimated in various fractions of rumen contents of Murrah buffalo. About 80–92% of the enzyme activities were associated with the particulate material, while only a negligible portion (1–4%) was present as extracellular fraction in the liquid portion of rumen contents. Rest of the activities (8–15%) were associated with freely suspended microbial cells in the liquid portion of the rumen contents. Maximum activities of carboxymethylcellulase, α-amylase, xylanase and micro-crystalline cellulase were observed at 2h post feeding while in the case of filter paper degrading activity, there was no difference in the activities in the first 4 hours of offering feed to the animals.

Rumen fermentation pattern and digestion of structural carbohydrates in buffalo (Bubalus bubalis) calves as affected by ciliate protozoa

Four male murrah buffalo calves (average weight 188 kg), each fitted with a rumen and abomasal cannula were subjected to treatments in which they were defaunated (Period I) or refaunated (Period II). The animals were given same wheat straw and concentrate ration in both periods. The pH and concentrations of ammonia nitrogen and total volatile fatty acids were lower concomitant with higher rumen volume and bacterial counts in rumen liquor of defaunated vs. refaunated animals. The molar proportion of propionate increased, and butyrate decreased, with defaunation. Intake of organic matter, neutral detergent fibre, acid detergent fibre, cellulose and hemicellulose did not differ between treatments although their forestomach and whole tract digestibilities were reduced with defaunations. It was concluded that defaunation reduces digestibility of structural carbohydrates in buffalo calves.

Effect of Sodium Nitrate and Nitrate Reducing Bacteria on In vitro Methane Production and Fermentation with Buffalo Rumen Liquor

Nitrate can serve as a terminal electron acceptor in place of carbon dioxide and inhibit methane emission in the rumen and nitrate reducing bacteria might help enhance the reduction of nitrate/nitrite, which depends on the type of feed offered to animals. In this study the effects of three levels of sodium nitrate (0, 5, 10 mM) on fermentation of three diets varying in their wheat straw to concentrate ratio (700:300, low concentrate, LC; 500:500, medium concentrate, MC and 300:700, high concentrate, HC diet) were investigated in vitro using buffalo rumen liquor as inoculum. Nitrate reducing bacteria, isolated from the rumen of buffalo were tested as a probiotic to study if it could help in enhancing methane inhibition in vitro. Inclusion of sodium nitrate at 5 or 10 mM reduced (p<0.01) methane production (9.56, 7.93 vs. 21.76 ml/g DM; 12.20, 10.42 vs. 25.76 ml/g DM; 15.49, 12.33 vs. 26.86 ml/g DM) in LC, MC and HC diets, respectively. Inclusion of nitrate at both 5 and 10 mM also reduced (p<0.01) gas production in all the diets, but in vitro true digestibility (IVTD) of feed reduced (p<0.05) only in LC and MC diets. In the medium at 10 mM sodium nitrate level, there was 0.76 to 1.18 mM of residual nitrate and nitrite (p<0.01) also accumulated. In an attempt to eliminate residual nitrate and nitrite in the medium, the nitrate reducing bacteria were isolated from buffalo adapted to nitrate feeding and introduced individually (3 ml containing 1.2 to 2.3×106 cfu/ml) into in vitro incubations containing the MC diet with 10 mM sodium nitrate. Addition of live culture of NRBB 57 resulted in complete removal of nitrate and nitrite from the medium with a further reduction in methane and no effect on IVTD compared to the control treatments containing nitrate with autoclaved cultures or nitrate without any culture. The data revealed that nitrate reducing bacteria can be used as probiotic to prevent the accumulation of nitrite when sodium nitrate is used to reduce in vitro methane emissions.

Phenotypic and phylogentic characterisation of tannin degrading/tolerating bacterial isolates from the rumen of goats fed on pakar (Ficus infectoria) leaves

The tannin degrading/tolerating bacteria were isolated from goat rumen fed on tannin rich diet on a medium containing 1% tannic acid. One hundred and twenty five isolates were picked up and screened for their potential to grow on a medium containing tannic acid as one of the ingredients and eight best isolates were selected for further characterisation. The isolates were coccoid to cocco-bacillary. Two out of eight isolates were Gram negative and the rest were Gram positive. All the isolates exhibited tannase activity with maximum of 29.0 units in isolate numbers 5, 6 and 7 and were able to grow on glucose, cellobiose, galactose, fructose, starch and sucrose but none of the isolates was able to grow on rahmnose. The isolates were tolerant to the presence of gallic acid in the medium up to 20mM, but pyrogallol, ferulic acid and coumaric acid were toxic at higher concentrations (20mM). True degradability of pakar leaves was increased by 9.7% by inclusion of live cultures of isolate number 6 (Accession no. HM771331) in the incubation medium. The phylogenetic analysis of 16S rRNA sequences of the isolates made a tight cluster with Streptococcus gallolyticus showing similarity ranging from 97.4% to 99%. The goat isolate number 6 (Accession no. HM771331) was tolerant to the phenolic monomers, exhibited tannase activity and improved in vitro true digestibility of tannin rich pakar leaves. The isolate appears to have a potential to be used as microbial feed additive to improve the utilisation of poor quality tannin containing feeds by the ruminants.

Garlic as a rumen modifier for eco-friendly and economic livestock production

Allium sativum (garlic) has been very popular in Asian kitchen as one of the major spices. The strong smelling juice of bulb contains volatile oils composed of sulphur-containing compounds: allicin, diallyl disulfide and diallyl trisulfide, which are responsible for anti-microbial activity. Recently garlic, its oil, extracts in different organic and aqueous solvents and individual components of essential oils have been examined as a feed additive to manipulate rumen function for controlling enteric methane emission and improving feed conversion efficiency. The results reported so far indicate that garlic and its essential oils inhibit methanogenesis significantly accompanied with a lower acetate to propionate ratio indicating a diversion of fermentation in a favourable direction. As methanogenesis is the major hydrogen sink in the rumen, its inhibition requires the disposal of reducing equivalents produced during fermentation of feed. The propionate synthesis serves as an alternate hydrogen sink. The anti-microbial compounds present in garlic appear to be selective inhibitors of methanogenesis, as there is no adverse effect on feed degradation in the rumen. Garlic and its oil adversely affect the protein degrading bacteria and deamination activity of the rumen contents. Only a few in vivo experiments have been conducted using garlic as a feed additive, and it appears to have good potential for rumen manipulation for eco-friendly (with minimum methane emission) and economic livestock production.

Manipulation of Rumen Microbial Ecosystem for Reducing Enteric Methane Emission in Livestock

Rumen has a complex consortium of microorganisms comprising of bacteria, protozoa, fungi, archaea and bacteriophages, which synergistically act upon the lignocellulosic feeds consisting of cereal straws and stovers, green forages and hays, oil cakes, etc., and produce a mixture of short-chain volatile fatty acids and microbial proteins which the animals can use as a source of nutrients. During this bioconversion process, hydrogen is generated in large quantities which combine with carbon dioxide to generate methane by the activity of methanogenic archaea. Depending upon the composition of diet, the animals might lose 5–12 % of gross energy intake in the form of methane, which leads to poor feed conversion efficiency. To avoid this loss of energy in the form of methane, several methods are technically available (e.g. methane analogues, inorganic terminal electron acceptors, ionophore antibiotics, organic unsaturated fatty acids, microbial intervention like use of probiotics and selective removal of ciliate protozoa and plant secondary metabolites), but each one of them has its own merits and demerits. In many cases, the results are based upon only in in vitro experiments. In this chapter, a few of the feed supplements which have a potential to inhibit methanogenesis and have been tested in in vivo experiments will be discussed.