Neeta Agarwal

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.

Diurnal variation in ciliate protozoa in the rumen of black buck (Antilope cervicapra) fed green forage

The protozoa in the rumen of a black buck were a B‐type population with numbers varying between 0·31 and 0·61 times 106cells/ml rumen liquor, when the animal was fed either vegetative green oat or third cut berseem. The total protozoa, total holotrichs, Dasytricha, total spirotrichs and small spirotrichs were significantly higher (P < 0·01) on berseem feeding than those on oat feeding, while the numbers of Isotricha and large spirotrichs were unaffected by change of diet. Numerically the most important group of protozoa was small spirotrichs (74·4–75·6% of total population) which accounted for only 9·85–13·61% of protozoal cell mass in the rumen.

Effects of extracts of spices on rumen methanogenesis, enzyme activities and fermentation of feeds in vitro

BACKGROUND An experiment was conducted to study the effects of boiling water, methanol and ethanol extracts (0, 0.25 and 0.50 mL) of seeds of Foeniculum vulgare (fennel), flower buds of Syzygium aromaticum (clove), bulbs of Allium sativum (garlic), bulbs of Allium cepa (onion) and roots of Zingiber officinalis (ginger) on rumen methanogenesis, fibrolytic enzyme activities and fermentation characteristics in vitro. RESULTS Ethanol and methanol extracts of fennel, clove and garlic at 0.50 mL and clove at 0.25 mL inhibited (P < 0.05) methane production. Carboxymethylcellulase activity was reduced (P < 0.05) by ethanol and methanol extracts (0.50 mL) of fennel and clove (0.25 and 0.50 mL). The extracts of clove reduced (0.25 and 0.50 mL) xylanase and acetylesterase activities, and the fennel extract (0.50 mL) reduced (P < 0.05) xylanase activity. However, the extracts of garlic (0.50 mL) increased (P < 0.05) acetylesterase activity. Concentrations of volatile fatty acids were reduced (P < 0.05) by the extracts of garlic and onion. The extracts of garlic caused a decrease (P < 0.05) in acetate:propionate ratio (A:P) at 0.50 mL, whereas A:P was increased (P < 0.05) by the inclusion of 0.50 mL extracts of clove. Methanol and ethanol extracts of clove decreased (P < 0.05) in vitro organic matter degradability. Extracts (0.50 mL) of clove decreased (P < 0.05) the numbers of total protozoa, small entodiniomorphs and holotrichs, whereas extracts of onion, ginger and garlic enhanced (P < 0.05) protozoal numbers (both entodiniomorphs and holotrichs). CONCLUSION Ethanol and methanol extracts of fennel and garlic have potential to inhibit rumen methanogenesis without adversely affecting rumen fermentation.

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.

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.

Effect of Dietary Supplementation of Probiotics on Growth Performance, Nutrients Digestibility and Faecal Microbiology in Weaned Piglets

In order to determine the effects of dietary supplementation of two different probiotics (Saccharomyces cerevisiae and Lactobacillus acidophilus) on growth performance, nutrient digestibility and faecal microbial count in weaned piglets (28d of age), 36 crossbred (Landrace x Local) piglets were allocated into three treatments on the basis of the BW in a completely randomized design. Each treatment was comprised of four replicates with three piglets in each. Dietary treatments were basal diet without any probiotic (Control), or with 10% of the basal diet replaced by feed fermented with either S. cerevisiae NCDC49 (SC; 3-5x106 cfu/g) or L. acidophilus NCDC-15 (LA; 2-3x109 cfu/g). Supplementation of SC or LA to piglets improved (P 0.05) among the groups. However, piglets in SC and LA groups showed better (P<0.05) FCR compared to Control. Yeast count and lactobacillus counts were higher (P<0.01) in both SC and LA groups as compared to Control accompanying a concomitant reduction (P<0.01) in coliform count. The results indicated that dietary supplementation of S. cervisiae or L. acidophilus has a positive effect on the performance of weaned piglets.

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.