• Asian-Australasian Journal of Animal Sciences
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• v.14 no.6
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• pp.880-884
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• 2001

Rumen microbiology research has undergone several evolutionary steps: the isolation and nutritional characterization of readily cultivated microbes; followed by the cloning and sequence analysis of individual genes relevant to key digestive processes; through to the use of small subunit ribosomal RNA (SSU rRNA) sequences for a cultivation-independent examination of microbial diversity. Our knowledge of rumen microbiology has expanded as a result, but the translation of this information into productive alterations of ruminal function has been rather limited. For instance, the cloning and characterization of cellulase genes in Escherichia coli has yielded some valuable information about this complex enzyme system in ruminal bacteria. SSU rRNA analyses have also confirmed that a considerable amount of the microbial diversity in the rumen is not represented in existing culture collections. However, we still have little idea of whether the key, and potentially rate-limiting, gene products and (or) microbial interactions have been identified. Technologies allowing high throughput nucleotide and protein sequence analysis have led to the emergence of two new fields of investigation, genomics and proteomics. Both disciplines can be further subdivided into functional and comparative lines of investigation. The massive accumulation of microbial DNA and protein sequence data, including complete genome sequences, is revolutionizing the way we examine microbial physiology and diversity. We describe here some examples of our use of genomics- and proteomics-based methods, to analyze the cellulase system of Ruminococcus flavefaciens FD-1 and explore the genome of Ruminococcus albus 8. At Illinois, we are using bacterial artificial chromosome (BAC) vectors to create libraries containing large (>75 kbases), contiguous segments of DNA from R. flavefaciens FD-1. Considering that every bacterium is not a candidate for whole genome sequencing, BAC libraries offer an attractive, alternative method to perform physical and functional analyses of a bacterium's genome. Our first plan is to use these BAC clones to determine whether or not cellulases and accessory genes in R. flavefaciens exist in clusters of orthologous genes (COGs). Proteomics is also being used to complement the BAC library/DNA sequencing approach. Proteins differentially expressed in response to carbon source are being identified by 2-D SDS-PAGE, followed by in-gel-digests and peptide mass mapping by MALDI-TOF Mass Spectrometry, as well as peptide sequencing by Edman degradation. At Ohio State, we have used a combination of functional proteomics, mutational analysis and differential display RT-PCR to obtain evidence suggesting that in addition to a cellulosome-like mechanism, R. albus 8 possesses other mechanisms for adhesion to plant surfaces. Genome walking on either side of these differentially expressed transcripts has also resulted in two interesting observations: i) a relatively large number of genes with no matches in the current databases and; ii) the identification of genes with a high level of sequence identity to those identified, until now, in the archaebacteria. Genomics and proteomics will also accelerate our understanding of microbial interactions, and allow a greater degree of in situ analyses in the future. The challenge is to utilize genomics and proteomics to improve our fundamental understanding of microbial physiology, diversity and ecology, and overcome constraints to ruminal function.

• Asian-Australasian Journal of Animal Sciences
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• v.33 no.7
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• pp.1126-1137
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• 2020

Objective: Pellet durability, particle size distribution, growth response, tibia bone characteristics and energy retention were measured to evaluate cassava as an alternative energy source to replace maize in broiler diets with or without Ronozyme (A+VP) enzyme composites. Methods: A total of 480 one-day broiler chicks were randomly assigned to 8 treatments in a 4×2 factorial arrangement. Four levels of cassava: (0%, 25%, 50%, 75%) and 2 levels of enzymes (0 and 500 g/tonne) were used. Each treatment was replicated six times, with ten birds per replicate. Results: The particle size distribution in the diets showed an increasing trend of small particles with increase in cassava level. Pellet durability decreased (p<0.05) with cassava inclusion. Feed intake was highest in birds fed diets with medium cassava level at 1 to 24 d and 1 to 35 d of age. The body weight gain of birds reduced (p<0.037) as cassava level increased, but it increased (p<0.017 when enzymes were added. The feed conversion ratio was high (p<0.05) when cassava level was increased, but it reduced (p<0.05) when enzymes were added. The dressing percentage (DP), and weight of drumsticks reduced (p<0.05) with increasing cassava level. Enzyme supplementation increased (p<0.05) DP, and weight of breast, thighs and drumsticks. Ash content, weight, length, width, and bone strength decreased (p<0.05) when cassava level was increased, however, they were increased with enzyme addition. The contents of Ca, K, and Zn were raised (p<0.001) with increasing cassava level. Enzyme inclusion increased (p<0.001) all mineral contents in tibia bones. Body fat and energy retained as fat decreased (p<0.001) as cassava level increased. Enzyme inclusion increased (p<0.05) body protein content and energy retained as protein. Conclusion: Although broiler performance was depressed by high levels of cassava inclusion, it was not affected by low levels, which further improved by enzyme supplementation.

• Asian-Australasian Journal of Animal Sciences
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• v.33 no.7
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• pp.1148-1155
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• 2020

Objective: The objective was to determine standardized ileal digestibility (SID) of crude protein (CP) and amino acids (AA) in cereal grains and various co-products fed to growing pigs. Methods: Ten feed ingredients tested were barley (9.3% CP), lupin kernels (31.1% CP), and wheat (11.3% CP) as cereal grains, and 2 sources of corn gluten feed produced in China (21.6% CP) and Korea (24.6% CP), corn gluten meal (65.3% CP), lupin hulls (11.6% CP), rice bran (14.5% CP), soybean meal (44.8% CP), and wheat bran (15.4% CP) as co-products. Ten experimental diets were formulated to contain each ingredient as a sole source of N and an N-free diet was used to correct basal endogenous losses of CP and AA. All diets also contained 0.5% Cr₂O₃ as an indigestible index. A replicated 11×6 incomplete Latin square design with 11 dietary treatments, 6 periods, and 22 animals was employed. Twenty-two barrows with an initial body weight of 64.6±4.9 kg were equipped with a T-cannula in the distal ileum. An experimental period consisted of a 4-d adaptation period and a 2-d collection period. Results: The SID of CP in the barley, lupin kernels, wheat, 2 sources of corn gluten feed, corn gluten meal, lupin hulls, rice bran, soybean meal, and wheat bran were 84.7%, 90.5%, 90.4%, 77.4%, 74.6%, 89.5%, 90.4%, 74.4%, 86.9%, and 63.4% (standard error of the mean [SEM] = 5.3, p = 0.006), respectively. The respective SID values of Lys were 75.5%, 88.4%, 83.9%, 74.7%, 62.4%, 80.3%, 83.9%, 78.5%, 88.0%, and 71.2% (SEM = 3.3, p<0.001), and the SID values of Met were 83.6%, 88.7%, 89.4%, 85.7%, 78.3%, 88.9%, 89.4%, 85.3%, 91.1%, and 77.0% (SEM = 2.4, p<0.001), respectively. Conclusion: The ileal digestibility of protein and amino acids varies among the feed ingredients fed to pigs.

• Microbiology and Biotechnology Letters
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• v.40 no.2
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• pp.92-97
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• 2012

Antimicrobial peptides (AMPs) are important components of living organisms acting against Gram-negative and Gram-positive bacterial and fungal pathogens. Cathelicidin human peptides have a variety of biological activities that can be used in clinical applications. AMPs are not produced naturally in large quantities, and chemical synthesis is also economically impractical, especially for long peptides. Therefore, as an alternative, heterologous expression of AMPs by recombinant techniques has been studied as a means to reduce production costs. E. coli is an excellent host for the expression of AMPs, as well as other recombinant proteins, because of the low cost involved and its easy manipulation. However, overexpression of AMPs in E. coli has been shown to cause difficulties resulting from the toxicity of the subsequently produced AMPs. Therefore, fusion expression was theorized to be a solution to this problem. In this study, AMPs were expressed as fused proteins with the glutathione S-transferase (GST) binding protein to protect against the toxicity of AMPs when expressed in E. coli. The LL37, and hybrid gaegurin and LL37 (GGN4(1-16)-LL37(17-32), which we designated as GL32, peptides were expressed as GST-fusion proteins in E. coli and the fusion proteins were then purified by affinity columns. The purified peptides were obtained by removal of GST and were confirmed by western blot analysis. The purified antimicrobial peptides then demonstrated antimicrobial activities against Gram-negative and Gram-positive bacterial strains.

• Tuberculosis and Respiratory Diseases
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• v.75 no.6
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• pp.244-249
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• 2013

Background: Conventional biomarkers cannot always establish the cause of pleural effusions; thus, alternative tests permitting rapid and accurate diagnosis are required. The primary aim of this study is to assess the ability of pentraxin-3 (PTX3) in order to diagnose the cause of pleural effusion and compare its efficacy to that of other previously identified biomarkers. Methods: We studied 118 patients with pleural effusion, classified as transudates and exudates including malignant, tuberculous, and parapneumonic effusions (MPE, TPE, and PPE). The levels of PTX3, C-reactive protein (CRP), procalcitonin (PCT) and lactate in the pleural fluid were assessed. Results: The levels of pleural fluid PTX3 were significantly higher in patients with PPE than in those with MPE or TPE. PTX3 yielded the most favorable discriminating ability to predict PPE from MPE or TPE by providing the following: area under the curve, 0.74 (95% confidence interval, 0.63-0.84), sensitivity, 62.07%; and specificity, 81.08% with a cut-off point of 25.00 ng/mL. Conclusion: Our data suggests that PTX3 may allow improved differentiation of PPE from MPE or TPE compared to the previously identified biomarkers CRP and PCT.

• Korean Journal of Acupuncture
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• v.21 no.1
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• pp.113-127
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• 2004

Objectives : The usage of acupuncture has gained popularity as an alternative method of treatment for certain chronic pain conditions. However, the efficacy of acupuncture in various diseases has not been fully established and the underlying mechanism is not clearly understood. In the present study, the effect of electroacupuncture (EA) applied to foot samli$(ST_{36})$ on the carrageenan-induced knee arthritic pain was examined. Methods : A common source of persistent pain in humans is the knee arthritis. Knee arthritis was induced by injection of 2 % carrageenan $50\;{\mu}l$ into the knee joint cavity. When rats developed pain behaviors, EA was applied for 30 min. under enflurane anesthesia with repeated train stimuli at the intensity of 10X of muscle twitch threshold. The weight bearing force of the hind limb was measured for an indicator of pain level after each manipulation. Results : The average weight borne by the hind limb during normal gait was 55% of total body weight, which was reduced to less than 10% after knee arthritis. EA improved the weight bearing of the arthritic hind limb significantly for the duration of 4 hr. EA applied to $ST_{36}$ point produced a significant improvement of stepping force of the arthritic foot lasting for at least 4 h. However, $GB_{31}$ point did not produce any significant increase of weight bearing force. The analgesic effect was specific to the acupuncture point since the analgesic effect on the knee arthritis model could not be mimicked by EA applied to a nearby point, $GB_{31}$. The relations between EA-induced analgesia and endogenous nitric oxide(NO) and inducible NO synthase(iNOS)/neuronal NOS was also examined. Results were turned out that both NO production and nNOS/iNOS protein expression which is increased by arthritis were suppressed by EA stimulation applied to $ST_{36}$ point. Conclusions : The data suggest 1) that EA produces a potent analgesic effect on the rat model of chronic knee arthritis pain in a point specific manner and 2) that EA-induced analgesia modulate endogenous NO through the suppression of nNOS/iNOS protein expression.

• Korean journal of applied entomology
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• v.55 no.1
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• pp.1-9
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• 2016

Edible insects are an interesting alternative global food resource. The purpose of this study was to investigate whether ingestion of silkworm (Bombyx mori) pupae powder with and without resistance exercise training (isometric contraction training) increased muscle mass in ICR mice. To achieve this, 28 ICR mice were grouped into control (CON), resistance exercise training (EX), silkworm pupae powder ingested control (SP), and silkworm powder ingestion with resistance exercise training (SPEX) groups. The change in body weight ratio was significantly decreased in the EX and SPEX groups compared to the CON and SP groups. Total blood protein levels were the highest in SPEX mice compared to those in other groups. The albumin concentration increased only in the EX group. Blood GOT and GPT levels were not significantly affected. Changes in Akt and Gsk-$3{\beta}$ protein expression were not significant but there was a tendency for Akt to increase and for Gsk-$3{\beta}$ to increase following the ingestion of the powder. The size of the gastrocnemius muscle increased significantly in response to resistance exercise training only. Furthermore, the ingestion of silkworm pupae powder tended to increase muscle mass without significance. These results suggested that the ingestion of silkworm pupae powder with resistance exercise training might enhance muscle mass without hepatotoxicity. However, future study may be needed to obtained detailed results and practical suggestions.

• Asian-Australasian Journal of Animal Sciences
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• v.24 no.2
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• pp.285-294
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• 2011

The abatement of methane emission from ruminants is an important global issue due to its contribution to greenhouse gas with carbon dioxide. Methane is generated in the rumen by methanogens (archaea) that utilize metabolic hydrogen ($H_2$) to reduce carbon dioxide, and is a significant electron sink in the rumen ecosystem. Therefore, the competition for hydrogen used for methanogenesis with alternative reductions of rumen microbes should be an effective option to reduce rumen methanogenesis. Some methanogens parasitically survive on the surface of ciliate protozoa, so that defaunation or decrease in protozoa number might contribute to abate methanogenesis. The most important issue for mitigation of rumen methanogenesis with manipulators is to secure safety for animals and their products and the environment. In this respect, prophylactic effects of probiotics, prebiotics and miscellaneous compounds to mitigate rumen methanogenesis have been developed instead of antibiotics, ionophores such as monensin, and lasalocid in Japan. Nitrate suppresses rumen methanogenesis by its reducing reaction in the rumen. However, excess intake of nitrate causes intoxication due to nitrite accumulation, which induces methemoglobinemia. The nitrite accumulation is attributed to a relatively higher rate of nitrate reduction to nitrite than nitrite to ammonia via nitroxyl and hydroxylamine. The in vitro and in vivo trials have been conducted to clarify the prophylactic effects of L-cysteine, some strains of lactic acid bacteria and yeast and/or ${\beta}$1-4 galactooligosaccharide on nitrate-nitrite intoxication and methanogenesis. The administration of nitrate with ${\beta}$1-4 galacto-oligosaccharide, Candida kefyr, and Lactococcus lactis subsp. lactis were suggested to possibly control rumen methanogenesis and prevent nitrite formation in the rumen. For prebiotics, nisin which is a bacteriocin produced by Lactococcus lactis subsp. lactis has been demonstrated to abate rumen methanogenesis in the same manner as monensin. A protein resistant anti-microbe (PRA) has been isolated from Lactobacillus plantarum as a manipulator to mitigate rumen methanogenesis. Recently, hydrogen peroxide was identified as a part of the manipulating effect of PRA on rumen methanogenesis. The suppressing effects of secondary metabolites from plants such as saponin and tannin on rumen methanogenesis have been examined. Especially, yucca schidigera extract, sarsaponin (steroidal glycosides), can suppress rumen methanogenesis thereby improving protein utilization efficiency. The cashew nutshell liquid (CNSL), or cashew shell oil, which is a natural resin found in the honeycomb structure of the cashew nutshell has been found to mitigate rumen methanogenesis. In an attempt to seek manipulators in the section on methane belching from ruminants, the arrangement of an inventory of mitigation technologies available for the Clean Development Mechanism (CDM) and Joint Implementation (JI) in the Kyoto mechanism has been advancing to target ruminant livestock in Asian and Pacific regions.

• Herbal Formula Science
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• v.25 no.1
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• pp.1-10
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• 2017

Objectives : The fruit of Prunus mume Siebold & Zucc. has been used as an alternative medicine and functional food in Korea and Japan for preventive and therapeutic purposes. However, its molecular actions and mechanism on anti-inflammatory activity have not been clearly investigated. The aim of this study was to clarify the anti-inflammatory activity of the ethanol extract of P. mume fruit (EEPM) in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells, and sought to understand the associated molecular mechanisms. Methods : Cytotoxicity was assessed by an MTT assay. The amount of nitric oxide (NO) production was determined by nitrite assay. The mRNA expression of inducible nitric oxide synthase (iNOS) was analyzed by RT-PCR. In addition, expression levels of iNOS, nuclear factor-erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) protein were detected by Western blotting. Results : Our data indicated that EEPM inhibited NO production in LPS-stimulated RAW264.7 cells in a concentration-dependent manner. At the mRNA and protein levels, EEPM suppressed LPS-induced iNOS expression. On the other hand, EEPM markedly enhanced HO-1 expression, which was associated with an induction and nuclear translocation of Nrf2. Moreover, the inhibitory effect of EEPM against LPS‑induced NO production was significantly enhanced by hemin, a HO-1 inducer; however, EEPM's effect on the production of NO was abolished by zinc protoporphyrin IX, a HO-1 inhibitor. Conclusion : The results suggest that EEPM can act as a suppressor agent on NO production through an activation of Nrf2/HO-1 signaling pathway, and may be a promising candidate for the treatment of inflammatory diseases.

• Asian-Australasian Journal of Animal Sciences
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• v.30 no.12
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• pp.1684-1688
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• 2017

Objective: From a review of published information on genetic association studies, a meta-analysis was conducted to determine the influence of the genes ${\kappa}-casein$ (CSN3) and ${\beta}-lactoglobulin$ (LGB) on milk yield traits in Holstein, Jersey, Brown Swiss, and Fleckvieh. Methods: The GLIMMIX procedure was used to analyze milk production and percentage of protein and fat in milk. Models included the main effects and all their possible two-way interactions; not estimable effects and non-significant (p>0.05) two-way interactions were dropped from the models. The three traits analyzed used Poisson distribution and a log link function and were determined with the Interactive Data Analysis of SAS software. Least square means and multiple mean comparisons were obtained and performed for significant main effects and their interactions (p<0.0255). Results: Interaction of breed by gene showed that Holstein and Fleckvieh were the breeds on which CSN3 ($6.01%{\pm}0.19%$ and $5.98%{\pm}0.22%$), and LGB ($6.02%{\pm}0.19%$ and $5.70%{\pm}0.22%$) have the greatest influence. Interaction of breed by genotype nested in the analyzed gene indicated that Holstein and Jersey showed greater influence of the CSN3 AA genotype, $6.04%{\pm}0.22%$ and $5.59%{\pm}0.31%$ than the other genotypes, while LGB AA genotype had the largest influence on the traits analyzed, $6.05%{\pm}0.20%$ and $5.60%{\pm}0.19%$, respectively. Furthermore, interaction of type of statistical model by genotype nested in the analyzed gene indicated that CSN3 and LGB genes had similar behavior, maintaining a difference of more than 7% across analyzed genotypes. These results could indicate that both Holstein and Jersey have had lower substitution allele effect in selection programs that include CSN3 and LGB genes than Brown Swiss and Fleckvieh. Conclusion: Breed determined which genotypes had the greatest association with analyzed traits. The mixed model based in Bayesian or Ridge Regression was the best alternative to analyze CSN3 and LGB gene effects on milk yield and protein and fat percentages.