Osamu Enishi

PCR Detection of DNAs of Animal Origin in Feed by Primers Based on Sequences of Short and Long Interspersed Repetitive Elements

PCR primers for the detection of materials derived from ruminants, pigs, and chickens were newly designed on the basis of sequences of the Art2 short interspersed repetitive element (SINE), PRE-1 SINE, and CR1 long interspersed repetitive element (LINE), respectively. These primers amplified the SINE or LINE from total DNA extracted from the target animals and from test feed containing commercial meat and bone meal (MBM). With the primers, detection of Art2, PRE-1, or CR1 in test feed at concentrations of 0.01% MBM or less was possible. This method was suitable for the detection of microcontamination of feed by animal materials.

Mitigation of methane production from cattle by feeding cashew nut shell liquid

The effects of cashew nut shell liquid (CNSL) feeding on methane production and rumen fermentation were investigated by repeatedly using 3 Holstein nonlactating cows with rumen fistulas. The cows were fed a concentrate and hay diet (6:4 ratio) for 4 wk (control period) followed by the same diet with a CNSL-containing pellet for the next 3 wk (CNSL period). Two trials were conducted using CNSL pellets blended with only silica (trial 1) or with several other ingredients (trial 2). Each pellet type was fed to cows to allow CNSL intake at 4 g/100 kg of body weight per day. Methane production was measured in a respiration chamber system, and energy balance, nutrient digestibility, and rumen microbial changes were monitored. Methane production per unit of dry matter intake decreased by 38.3 and 19.3% in CNSL feeding trials 1 and 2, respectively. Energy loss as methane emission decreased from 9.7 to 6.1% (trial 1) and from 8.4 to 7.0% (trial 2) with CNSL feeding, whereas the loss to feces (trial 1) and heat production (trial 2) increased. Retained energy did not differ between the control and CNSL periods. Digestibility of dry matter and gross energy decreased with CNSL feeding in trial 1, but did not differ in trial 2. Feeding CNSL caused a decrease in acetate and total short-chain fatty acid levels and an increase in propionate proportion in both trials. Relative copy number of methyl coenzyme-M reductase subunit A gene and its expression decreased with CNSL feeding. The relative abundance of fibrolytic or formate-producing species such as Ruminococcus flavefaciens, Butyrivibrio fibrisolvens, and Treponema bryantii decreased, but species related to propionate production, including Prevotella ruminicolla, Selenomonas ruminantium, Anaerovibrio lipolytica, and Succinivibrio dextrinosolvens, increased. If used in a suitable formulation, CNSL acts as a potent methane-inhibiting and propionate-enhancing agent through the alteration of rumen microbiota without adversely affecting feed digestibility.

Effect of cashew nut shell liquid on metabolic hydrogen flow on bovine rumen fermentation.

Effect of cashew nut shell liquid (CNSL), a methane inhibitor, on bovine rumen fermentation was investigated through analysis of the metabolic hydrogen flow estimated from concentrations of short-chain fatty acids (SCFA) and methane. Three cows were fed a concentrate and hay diet without or with a CNSL-containing pellet. Two trials were conducted using CNSL pellets blended with only silica (trial 1) or with several other ingredients (trial 2). Methane production was measured in a respiration chamber system, and energy balance and nutrient digestibility were monitored. The estimated flow of metabolic hydrogen demonstrated that a part of metabolic hydrogen was used for hydrogen gas production, and a large amount of it flowed into production of methane and SCFA in both trial 1 and 2, when CNSL was administered to the bovine rumen. The results obtained by regression analyses showed that the effect of CNSL supply on methane reduction was coupled with a significant (P < 0.01) decrease of acetate and a significant (P < 0.01) increase of propionate and hydrogen gas. These findings reveal that CNSL is able to reduce methane and acetate production, and to increase hydrogen gas and propionate production in vivo.

Improvement and validation of the method to determine neutral detergent fiber in feed

To improve the performance of the analytical method for neutral detergent fiber in feed with heat-stable α-amylase treatment (aNDFom), the process of adding heat-stable α-amylase, as well as other analytical conditions, were examined. In this new process, the starch in the samples was removed by adding amylase to neutral detergent (ND) solution twice, just after the start of heating and immediately after refluxing. We also examined the effects of the use of sodium sulfite, and drying and ashing conditions for aNDFom analysis by this modified amylase addition method. A collaborative study to validate this new method was carried out with 15 laboratories. These laboratories analyzed two samples, alfalfa pellet and dairy mixed feed, with blind duplicates. Ten laboratories used a conventional apparatus and five used a Fibertec(®) type apparatus. There were no significant differences in aNDFom values between these two refluxing apparatuses. The aNDFom values in alfalfa pellet and dairy mixed feed were 388 g/kg and 145 g/kg, the coefficients of variation for the repeatability and reproducibility (CV(r) and CV(R) ) were 1.3% and 2.9%, and the HorRat values were 0.8 and 1.1, respectively. This new method was validated with 5.8% uncertainty (k = 2) from the collaborative study.