J. Poultry Sci. 和文要旨 60巻2号　2023年

ニワトリヒナにおける感染時の摂食行動の低下に関わるメカニズムの解明

橘哲也1・マーク A クライン2

1愛媛大学農学部　松山市　790-8566
2バージニア工科大学神経科学　ブラックスバーグ　24061

ニワトリヒナの摂食行動は体内外の様々な因子の影響を受ける。細菌、真菌およびウイルスなどの病原体に由来する物質もその因子の一つであり、それらの多くはニワトリヒナの摂食行動を抑制する。病原体による感染はニワトリヒナの摂食行動を抑制することが知られているが、そのメカニズムはほとんど明らかにされていない。我々の研究グループは感染時のニワトリヒナにおける摂食行動低下機構の解明に取り組んでいる。その成果として、Toll様受容体のリガンドである病原体関連分子パターンが感染時の摂食行動の低下に関与する可能性を見出した。さらに、インターロイキン-1βおよび腫瘍壊死因子様サイトカイン1Aなどの炎症性サイトカイン、プロスタグランジン、および一酸化窒素が感染時の摂食行動低下に関わる生理活性物質である可能性を見出している。この総説は感染時の摂食行動低下機構に関する最近の知見をまとめたものである。
キーワード ：ニワトリヒナ、摂食、感染、病原体関連分子パターン、Toll様受容体

DOI： https://doi.org/10.2141/jpsa.2023009

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Biomolecules Triggering Altered Food Intake during Pathogenic Challenge in Chicks

Tetsuya Tachibana, Mark A. Cline
Food intake is regulated by several complicated synergistic mechanisms that are affected by a variety of internal and external influences. Some of these factors include those that are released from pathogens such as bacteria, fungi, and viruses, and most of these factors are associated with suppression of the chick’s food intake. Although chicks are well-known to decrease their food intake when they experience a pathogenic challenge, the mechanisms that mediate this type of satiety are poorly understood. One of the goals of our research group has been to better understand these mechanisms in chicks. We recently provided evidence that pathogen-associated molecular patterns, which are recognized by pattern-recognition receptors such as Toll-like receptors, likely contribute to satiety in chicks that are experiencing a pathogenic challenge. Additionally, we identified several inflammatory cytokines, including interleukin-1β, tumor necrosis factor-like cytokine 1A, prostaglandins, and nitric oxide, that likely contribute to satiety during a pathogenic challenge. This review summarizes the current knowledge on pathogen-induced satiety in chicks mainly accumulated through our recent research. The research will give good information to improve the loss of production during infection in poultry production in the future.

DOI： https://doi.org/10.2141/jpsa.2023009

Regulatory Effects of the Probiotic Clostridium butyricum on Gut Microbes, Intestinal Health, and Growth Performance of Chickens

Min Liu, Victoria Anthony Uyanga, Xikang Cao, Xinyu Liu, and Hai Lin College of Animal Science and Technology, Shandong Agricultural University, Key Laboratory of Efficient Utilization of Non-grain Feed Resources (Co-construction by Ministry and Province), Ministry of Agriculture and Rural Affairs, Shandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, Tai’an City, Shandong Province 271018, China
抄録
Clostridium butyricum is an important probiotic for chickens and exerts various biological activities, including altering the composition of the intestinal microbiota, competing with other microorganisms for nutrients, improving the integrity of the intestinal mucosal system, changing the intestinal barrier, and improving overall host health. Intestinal microbes also play vital roles in maintaining the intestinal barrier, regulating intestinal health, and promoting chicken growth. During chicken production, chickens are vulnerable to various stressors that have detrimental effects on the intestinal barrier with significant economic consequences. C. butyricum is a known probiotic that promotes intestinal health and produces the short-chain fatty acid butyric acid, which is beneficial for the growth performance of chickens. This review elucidates the development and utilization of C. butyricum to improve intestinal barrier function and growth performance in chickens through its probiotic properties and interactions with intestinal microbes.

DOI： https://doi.org/10.2141/jpsa.2023011

日本における家禽の歴史的展開

田島淳史
筑波大学生命環境系

日本の家禽産業は、第二次世界大後に生じた家禽製品に対する旺盛な需要に応えるべく急速に発展した。しかし、この第二次世界大後の日本の養鶏業の隆盛は、二十世紀当初から数十年をかけて築き上げた強固な学術・教育基盤の上に築かれたことを忘れてはらない。さらに、家禽は日本社会において特別な文化的役割を果たしている。 そこで本稿では、1) 日本における家禽産業の歴史的な発展過程、2）日本の教育・研究成果が養鶏産業に果たした役割、3) 日本社会に深く根付いた家禽の儀礼的・神話的・芸術的役割の3点について概観する。

DOI： https://doi.org/10.2141/jpsa.2023015

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Historical Overview of Poultry in Japan

Atsushi Tajima
Faculty of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki 305-8572, Japan.

To meet the strong quantitative demands for poultry products with high sanitary standards soon after World War II, the Japanese poultry industry experienced a rapid expansion period. However, it should be remembered that the post-war flourishment of the Japanese poultry industry was built on a solid academic and educational platform that took several pre-war decades to construct. In addition, poultry play a special cultural role in Japanese society. In this review, poultry in Japan is illustrated from the following three historical viewpoints: 1) development of the Japanese poultry industry; 2) academic and educational contributions to the poultry industry in Japan; and 3) ritualistic, mythic, and artistic attributes of poultry that are deeply embedded in Japanese society.

DOI： https://doi.org/10.2141/jpsa.2023015

暑熱ストレスがニワトリの腸管の健全性、炎症および細菌叢におよぼす影響とその機構

喜久里基・豊水正明　

東北大学・大学院農学研究科

鶏肉と鶏卵の生産は他の動物性タンパク質に比べ、製造時の二酸化炭素排出量や飼料、水の消費量が少ないという利点がある。このため、鶏肉の持続可能な生産の維持は主要タンパク質源としての同肉の世界的な需要を満たすために重要である。夏期や熱帯・亜熱帯地域における家禽の暑熱ストレスは、生産性や健康に悪影響をおよぼすことが古くから知られているが、その悪影響は地球温暖化の加速とともに増大すると予測されている。暑熱ストレスはニワトリの生理機能、代謝、免疫反応に影響をおよぼし、電解質の不均衡や酸化ストレス、内分泌疾患、炎症、免疫異常を引き起こす。これらの生体応答は独立して発生するものではなく、体系的・連鎖的な機構が存在することが示されている。最近、腸管の恒常性が栄養素吸収や全身性炎症の進行に重要な役割を果たしていることが明らかにされ、その作用機序には、神経内分泌シグナル伝達、抗酸化反応、食餌中の酸化物質および微生物叢の組成が関与すると考えられている。本総説では暑熱ストレスによるニワトリの腸管の機能不全とその根本的な原因に焦点を概説し、家禽の健康と持続可能な生産を実現するための戦略を構築するための情報を提供する。

DOI： https://doi.org/10.2141/jpsa.2023021

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Mechanisms underlying the Effects of Heat Stress on Intestinal Integrity, Inflammation, and Microbiota in Chickens

Motoi Kikusato and Masaaki Toyomizu

Animal Nutrition, Life Sciences, Graduate School of Agricultural Science, Tohoku University, Sendai, Japan/p>

Poultry meat and egg production benefits from a smaller carbon footprint, as well as feed and water consumption, per unit of product, than other protein sources. Therefore, maintaining a sustainable production of poultry meat is important to meet the increasing global demand for this staple. Heat stress experienced during the summer season or in tropical/subtropical areas negatively affects the productivity and health of chickens. Crucially, its impact is predicted to grow with the acceleration of global warming. Heat stress affects the physiology, metabolism, and immune response of chickens, causing electrolyte imbalance, oxidative stress, endocrine disorders, inflammation, and immunosuppression. These changes do not occur independently, pointing to a systemic mechanism. Recently, intestinal homeostasis has been identified as an important contributor to nutrient absorption and the progression of systemic inflammation. Its mechanism of action is thought to involve neuroendocrine signaling, antioxidant response, the presence of oxidants in the diet, and microbiota composition. The present review focuses on the effect of heat stress on intestinal dysfunction in chickens and the underlying causative factors. Understanding these mechanisms will direct the design of strategies to mitigate the negative effect of heat stress, while benefiting both animal health and sustainable poultry production.

DOI： https://doi.org/10.2141/jpsa.2023021

Effects of Dietary Tryptophan on Growth Performance, Plasma Parameters, and Internal Organs of 1–28-Day-Old Sichuan White Geese

Yang Fu1*, Bo Liu2*, Hui Lei1, Zhenping Lin3, JunPeng Chen3, Yongwen Zhu1, Hui Ye1, Lin Yang1 and Wence Wang1

1College of Animal Science and National Engineering Research Center for Breeding Swine Industry, South China Agricultural University, Guangzhou 510642, China 2Changsha Sanwang Feed Co. Ltd, Changsha 410000, China 3Shantou Baisha Research Institute of Original Species of Poultry and Stock, Shantou 515041, China
抄録
Although the nutrient requirements of geese during the growing stage are known, the dietary requirement of amino acids during the starting period remains unclear. Optimum nutrient supplementation during the starting period is crucial for improved survival rates, body-weight gain, and marketing weight in geese. Our study focused on the effect of dietary tryptophan (Trp) supplementation on the growth performance, plasma parameters, and internal-organ relative weights in 1–28-day-old Sichuan white geese. A total of 1080 1-day-old geese were divided randomly into six Trp-supplemented(0.145%, 0.190%, 0.235%, 0.280%, 0.325%, and 0.370%) groups. Average daily feed intake (ADFI), average daily gain(ADG), and duodenal relative weight were highest in the 0.190% group, brisket protein level and jejunal relative weight in the 0.235% group, and plasma total protein and albumin levels in the 0.325% group (P < 0.05). Dietary Trp supplementation did not significantly affect the relative weights of the spleen, thymus, liver, bursa of Fabricius, kidneys, and pancreas. Moreover, the 0.145% – 0.235% groups showed significantly decreased liver fat (P < 0.05). Based on the non-linear regression analysis of ADG and ADFI, the dietary Trp levels between 0.183% and 0.190% were estimated to be optimal for 1–28-day-old Sichuan white geese. In conclusion, optimal dietary Trp supplementation in 1–28-day-old Sichuan white geese resulted in increased growth performance (0.180% – 0.190%) along with improved proximal intestinal development and brisket protein deposition (0.235%). Our findings provide basic evidence and guidance for optimal levels of Trp supplementation in geese.

DOI： https://doi.org/10.2141/jpsa.2023008

Characterization of Chicken-Derived Genotype VII Newcastle Disease Virus Isolates from Northwest China

Xinxin Qiu1,2, Yanqing Jia2, Zhencang Zhang2, Xianglin Fo2 and Wenhui Wang1

1College of Veterinary Medicine, Gansu Agricultural University, Lanzhou, Gansu 730070, China
2Department of animal engineering/Shaanxi engineering research center of the prevention and control for animal disease, Yangling Vocational & Technical College, Yangling, Shaanxi Province 712100, China
抄録
Newcastle disease virus (NDV) threatens global poultry production, with genotype VII the most prevalent strain in China. However, little information is available regarding viral multiplication and pathogenicity based inoculation route. The objectives of this study were to sequence NDV VII isolates and to analyze their biological characteristics in detail. A total of 86 oral and cloacal swabs were collected from Shaanxi and Gansu provinces in northwest China. Identification of genotype VII NDV based on the M gene was performed by qPCR. Viral multiplication and pathogenicity were assessed as a function of route of infection. We observed increased morbidity and mortality using intravenous injection, whereas intranasal, intraocular, and cloacal infections resulted in slower progression and milder clinical disease, with viral proliferation obvious in different tissues. These results provide an important basis for the clinical control and prevention of NDV epidemics in poultry.
Key words: infection, inoculation route, NDV, phylogenetics

DOI： https://doi.org/10.2141/jpsa.2023010

RNA-sequencing Reveals Differentially Expressed Genes of Laying Hens Fed Baihu Decoction Under Heat Shock

Tingting Lu, Lihong Li, Yuwei Li and Xianghui Li

Institute of Veterinary Medicine, Henan University of Animal Husbandry and Economy, Zhengzhou, Henan Province 450002, China
抄録
Egg production, an important economic trait in the poultry industry, is sensitive to heat stress. The hypothalamus is a crucial center for thermoregulation by detecting temperature changes and regulating the autonomic nervous system in poultry. Baihu decoction (BH), which contains four ingredients (Rhizoma Anemarrhenae, Gypsum Fibrosum, Radix Glycyrrhizae, and Semen Oryzae Nonglutinosae), is a traditional Chinese medicinal formula for clearing heat. Our study aimed to investigate the changes in gene transcription levels in the hypothalamus of laying hens treated with heat stress with and without BH using RNA sequencing. A total of 223 differentially expressed genes (DEGs) were identified in the heat-treated group compared with the control group and 613 DEGs were identified in the BH group compared with the heat-treated group. Heat shock led to significant changes in the expression of multiple genes involved in the “neuroactive ligand-receptor interaction” pathway. Moreover, feeding BH led to significant upregulation in the expression of eight genes encoding heat shock proteins (HSPs), which were highlighted as candidates to control the “protein processing in the endoplasmic reticulum (ER)” pathway. These results provide the novel insight that BH responds to heat stress by participating in regulation of the ER signaling pathway and HSPs expression.
Key words: Baihu decoction, chicken, heat stress, hypothalamus, RNA-seq

DOI： https://doi.org/10.2141/jpsa.2023012

DNA免疫法とタンパク質免疫法により調製した抗狂犬病ウイルス核タンパク質特異的IgYの特性
久保 七彩1,2・朴 天鎬3・井上 智3,4・八田 一1,2 1（株）NBL 鶏と卵の研究所　京都府京都市東山区　605-8501
2京都女子大学　京都府京都市東山区　605-8501
3北里大学獣医学部　青森県十和田市　0341-8628
4国立感染症研究所 獣医科学部　東京都新宿区　162-8640
鶏卵卵黄から狂犬病ウイルス核タンパク質（RV-N）に対する特異的IgY抗体を調製することを目的に、産卵鶏にアジュバントとしてλ-カラギーナンまたはFCAの事前注射（免疫前刺激）を行い、局所の免疫活性を上昇させた後にDNA免疫を実施する手法でRV-N特異的IgY抗体の調製を実施した。また、産卵鶏に対し従来からのIgY抗体調製法であるタンパク質免疫法も実施し、各群の最もIgY抗体価が上昇した個体の卵黄からIgYを精製した。従来法であるタンパク質免疫法では、すべての個体で特異的IgYが誘導された。一方で、DNA免疫実施前に免疫前刺激を行うことで、各群3羽中1羽ずつではあるもののELISAにてタンパク免疫と同等の反応性（ELISA値）を有するIgY抗体が得られた。
続いて、これらのIgYを一次抗体として、イヌ等の動物に対する狂犬病の判定に利用される免疫組織化学染色（IHC）を実施した。タンパク質免疫法で調製した抗体はウイルス感染犬の脳切片中に存在するウイルス抗原を検出できたが、DNA免疫法（＋免疫前刺激）で調製したIgY抗体では検出できなかった。その理由として、市販の狂犬病ワクチン（不活化ウイルス）を10％ホルマリン処理や加熱処理し、濃度を振って固相抗原として用いたELISAの結果から、DNA免疫法で調製したIgY抗体はタンパク質免疫法で調製したIgY抗体と比較して変性抗原に対する結合力が低く、また微量の未変性抗原に対しても反応性が低いためであると考えられた。IHC等で抗原検出に用いることができる特異的IgYをDNA免疫法で調製するには、変性抗原に対しても強く結合するIgY抗体の誘導法の開発が必要となる。また、特に未変性抗原を認識させることを目的とした抗体試薬や中和抗体としての活用に注目すべきであると考えられる。
キーワード: DNA immunization, FCA, IgY, IHC, λ-carrageenan, Rabies virus

DOI： https://doi.org/10.2141/jpsa.2023014

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Comparison of Anti-rabies Virus Nucleoprotein IgY Prepared by DNA Immunization and Protein Immunization

Nanase Kubo1,2, Chun-ho Park3, Satoshi Inoue3,4 and Hajime Hatta1,2

1 Egg & Poultry Research Center, NBL Co., Ltd., Kyoto 605-8501, Japan
2 Kyoto Women’s University, Kyoto 605-8501, Japan
3 Department of Veterinary Pathology, School of Veterinary Medicine, Kitasato University, Aomori 0341-8628, Japan
4 Laboratory of Transmission Control of Zoonosis, Department of Veterinary Science National Institute of Infectious Diseases, Tokyo 162-8640, Japan
抄録
Immunization of egg-laying hens with viral antigens efficiently produces large amounts of virus-specific IgY antibodies from egg yolks. A supply of practical and economical antibodies against the rabies virus is being desired worldwide. We immunized hens with the antigen gene DNA of the rabies virus, purified specific IgY antibodies from the egg yolk, and characterized the immuno-protein chemistry for use as a diagnosis. To prepare specific IgY antibodies against rabies virus nucleoprotein (RV-N) by DNA immunization, laying hens were pre-injected with λ-carrageenan or Freund’s complete adjuvant to increase local immune activity (pre-immune stimulation), and then immunized with RV-N recombinant plasmid DNA. RV-N-specific IgY antibodies were prepared from egg yolks of immunized hens. For comparison, conventional protein antigen immunization was also used to induce the production of RV-N-specific IgY antibodies. Laying hens were immunized with an RV-N protein antigen and RV-N-specific IgY was purified from egg yolks. The binding activity against RV-N antigens was examined using IgY samples prepared by DNA (with pre-immune stimulation) and protein immunization. Immunohistochemical staining showed that IgY antibodies prepared by protein immunization strongly detected viral antigens in the brain sections of dogs infected with the virus, whereas IgY antibodies prepared by DNA immunization did not. Enzyme-linked immunosorbent assay was performed using a commercially available rabies vaccine (inactivated virus) treated with 10% formalin and heating (60°C, 30 min and 90°C, 5 min). IgY prepared by DNA immunization had weaker reactivity with denatured antigens and lower antigen concentrations than IgY prepared by protein immunization. These results suggest that it is necessary to develop a DNA immunization method for inducing IgY antibodies against the rabies virus that strongly bind to native and denatured antigens to prepare specific IgYs that can be used for antigen detection in clinical tests.
Key words: DNA immunization, FCA, IgY, IHC, λ-carrageenan, Rabies virus

DOI： https://doi.org/10.2141/jpsa.2023014

Immune-enhancing Effects of Chitosan-fermented Feed Additive on Broiler Chickens and Subsequent Protection Conferred against Experimental Infection with Salmonella Gallinarum

Bo Mi Park1,*, Jina Lee1, 2,*, Young Kyu Park3, Young Cheol Yang3, Bock Gie Jung1, 4 and Bong Joo Lee1
1Department of Veterinary Infectious Diseases, College of Veterinary Medicine, Chonnam National University, Gwangju 61186, Korea.
2Current address: Biology Department at Morrissey College of Arts and Sciences, Boston College, Chestnut Hill, MA 02467, U.S.A.
3Korea Beneficial Insects Lab. Co., Ltd., Soryong-ri, Okgwa-myeon, Gokseong-gun, Jeollanam-do 57507, Republic of Korea.
4Current address: Department of Pulmonary Immunology, Center for Pulmonary and Infectious Diseases Control, University of Texas Health Science Center at Tyler, Tyler, TX 75708, U.S.A.
抄録
Benefits chitosan-fermented feed additives (CFFAs) particularly in the regulation of the immune system and antimicrobial activity. Therefore, we investigated the immune-enhancing and bacterial clearance effects of CFFA (fermented by Bacillus licheniformis) on broiler chickens Salmonella Gallinarum challenge. We administered 2% or 4% CFFA evaluated its immune-enhancing effects using several immunological experiments, including examination of lysozyme activity, lymphocyte proliferation, and expression of cytokines. We also evaluated the bacterial clearance effects of CFFA against S. Gallinarum. CFFA administration markedly enhanced lysozyme activity, lymphocyte proliferation, and the expression of interleukin (IL)-2, IL-12, tumor necrosis factor alpha, and interferon gamma in the spleen. In broilers challenged with S. Gallinarum, the clinical signs of S. Gallinarum infection and the number of viable bacterial colonies in the feces and tissues decreased in both CFFA groups. Therefore, CFFAs could be good candidates for feed additive to improve nonspecific immune responses and bacterial clearance.
Key words: broilers, fermentation, immune response, insect, Salmonella Gallinarum

DOI： https://doi.org/10.2141/jpsa.2023016

安定同位体希釈分析を用いたLC-MS/MSによるNτ-methylhistidineおよびNπ-methylhistidineにおける分析条件の検討および妥当性の比較
白石純一1・井尻大地2,3・片渕歩美2・友永省三4・島元紗希5・都ハンウル4・石原慎矢1・大塚彰2,3
1 日本獣医生命科学大学応用生命科学部、武蔵野市境南町1-7-1 180-8602
2 鹿児島大学農林水産学研究科、鹿児島市郡元1-21-24 890-0065
3 鹿児島大学大学院連合農学研究科、鹿児島市郡元1-21-24 890-0065
4 京都大学大学院農学研究科、京都市左京区北白川追分町, 606-8502
5 新潟大学自然科学研究科、新潟市西区五十嵐2の町8050 950-2181
ニワトリの血中のNτ-methylhistidine濃度は、骨格筋タンパク質分解の指標として利用されてきた。本研究では、安定同位体希釈分析を用いたLC-MS/MSによるNτ-methylhistidineおよびNπ-methylhistidineメチルヒスチジンの分析条件の検討および妥当性の評価を実施した。内部標準は、安定同位体標識したNτ-methylhistidineを用いた。LC-MS/MSを用いて、1.56-50.00 μmol/Lおよび0.78-25.00 μmol/Lの範囲でNτ-methylhistidineとNπ-methylhistidineを分析できた。続いて、絶食および再給餌を行ったニワトリの血漿中のNτ-methylhistidineおよびNπ-methylhistidineを分析した結果、絶食に対するニワトリ血漿中のNτ-methylhistidineおよびNπ-methylhistidine濃度の変動を検出可能であった。以上の結果より、本研究で検討したLC-MS/MSによるNτ-methylhistidineおよびNπ-methylhistidineの安定同位体希釈分析法は、ニワトリ血漿の定量分析に適用可能であることが示唆された。
キーワード: Nτ-methylhistidine, Nπ-methylhistidine, LC-MS/MS, method validation

DOI： https://doi.org/10.2141/jpsa.2023017

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Quantification of Nτ -Methylhistidine and Nπ-Methylhistidine in Chicken Plasma by Liquid Chromatography–Tandem Mass Spectrometry
Jun-ichi Shiraishi1, *, Daichi Ijiri2, 3, *, Ayumi Katafuchi2, Shozo Tomonaga4, Saki Shimamoto5,Hanwool Do4, Shinya Ishihara1 and Akira Ohtsuka2, 3
1 Graduate School of Applied Life Science, Nippon Veterinary and Life Science University, 1-7-1 Kyonan-cho, Musashino, Tokyo 180-8602, Japan
2 Graduate School of Agriculture, Forestry and Fisheries, Kagoshima University, 1-21-24 Korimoto, Kagoshima 890-0065, Japan
3 The United Graduate School of Agricultural Sciences, Kagoshima University, 1-21-24 Korimoto, Kagoshima 890-0065, Japan
4 Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan
5 Graduate School of Science and Technology, Niigata University, 8050 Ikarashi 2-no-cho, Nishi-ku, Niigata 950-2181, Japan
抄録
The concentration of Nτ-methylhistidine in plasma provides an index of skeletal muscle protein breakdown. This study aimed to establish a quantitative method for measuring the concentrations of Nτ-methylhistidine and its isomer Nπ-methylhistidine in chicken plasma, using liquid chromatography–tandem mass spectrometry with stable isotope dilution analysis. The acceptable linear ranges of detection were 1.56–50.00 μmol/L for Nτ-methylhistidine and 0.78–25.00 μmol/L for Nπ-methylhistidine. The proposed method detected changes in the plasma levels of Nτ-methylhistidine and Nπ-methylhistidine in response to fasting and re-feeding. These results suggest that the method developed in this study can be used for the simultaneous measurement of Nτ-methylhistidine and Nπ-methylhistidine in chicken plasma.
Key words: LC–MS/MS, method validation, Nτ-methylhistidine, Nπ-methylhistidine

Effects of Polysaccharides-Rich Extract from Gracilaria lemaneiformis on Growth Performance, Antioxidant Capacity, Immune Function, and Meat Quality in Broiler Chickens

Shengwang Jiang1, Chaoyun Yang1, Yintao Xiao2, Saizhen Zheng2, Qian Jiang2, and Jiashun Chen2

1College of Animal Science, Xichang University, Xichang, Sichuan 615013, P.R. China
2College of Animal Science and Technology, Hunan Agricultural University, Changsha, Hunan 410128, P.R.China
抄録
This study investigated the effects of dietary supplementation with Gracilaria lemaneiformis polysaccharides (GLPs) on the growth performance, antioxidant capacity, immune function, and meat quality of broiler chickens. A total of 320 one-dayold Arbor Acres broiler chicks were individually weighed and randomly assigned to four groups of eight replicate cages (10 broilers per cage). Birds were fed a basal diet supplemented with 0 (control), 1,000, 2,000, or 4,000 mg/kg GLPs. Compared to that of the control group, dietary supplementation with 2,000 mg/kg GLPs linearly increased the average daily weight gain during days 0-42 (P < 0.05) and linearly decreased the feed to gain ratio during days 1–21 and 22–42 (P < 0.05). Broilers fed GLP-supplemented diets showed linear (P < 0.05) and quadratic (P < 0.05) increases in serum superoxide dismutase (P < 0.05), glutathione peroxidase, and catalase activities in the liver, whereas GLP supplementation decreased serum and liver malondialdehyde concentrations (P < 0.05). A linear increase in serum catalase activity was observed following supplementation with 2,000 or 4,000 mg/kg GLPs (P < 0.05). Broilers fed GLP-supplemented diets showed linear (P < 0.05) and quadratic (P < 0.05) increases in serum immunoglobulin (Ig) A, IgG, interleukin (IL)-6, IL-1β, IL-10, and interferon-γ concentrations (P < 0.05), and a trend towards linear improvement in IL-4 levels (P = 0.089). Dietary GLP supplementation increased the Lactobacillus spp. population compared to that of the control group (P < 0.05) and 2,000 and 4,000 mg/kg of GLPs nearly decreased the population of E. coli in the cecum (P = 0.056). Therefore, dietary GLP supplementation may improve broiler growth performance by altering antioxidant capacity, immune function, and the gut microbiota composition. Considering the effects of different doses of GLP on the above parameters, 2,000 mg/kg of GLPs was identified as the best dose.
Key words: antioxidant capacity, broilers, Gracilaria lemaneiformis polysaccharides, growth performance, immune function

DOI： https://doi.org/10.2141/jpsa.2023018

A Novel Bacterial 6-Phytase Improves Productive Performance, Precaecal Digestibility of Phosphorus, and Bone Mineralization in Laying Hens Fed a Corn-Soybean Meal Diet Low in Calcium and Available Phosphorus

Maamer Jlali1, Clémentine Hincelin2, Maria Francesch3, Tania Rougier1, Pierre Cozannet1, Sarper Ozbek1, Marcio Ceccantini2, Baris Yavuz2, Aurélie Preynat1 and Estelle Devillard1

1Adisseo France S.A.S., Center of Expertise in Research and Nutrition, 03600 Malicorne, France
2Adisseo France S.A.S., 92160 Antony, France
3IRTA, Animal Nutrition, Ctra. Reus-El Morell km. 3.8, E-43120 Constantí, Tarragona, Spain
抄録
Exogenous phytases are commonly added to low-phosphorus and low-calcium diets to improve P availability and reduce P excretion by poultry. This study investigated the effect of supplementation with a novel bacterial 6-phytase on egg production, egg quality, bone mineralization, and precaecal digestibility of P in laying hens fed corn-soybean meal-based diets. A total of 576 Hy-Line brown laying hens were used in a completely randomized block design at 25–45 weeks of age (woa). The three treatments included a positive control (PC) adequate-nutrient diet with 2840 kcal metabolizable energy/kg, 0.77% digestible lysine, 3.5% Ca, and 0.30% available P (avP); a negative control (NC) diet with 0.16% points less Ca and avP; and an NC diet supplemented with a novel bacterial 6-phytase at 300 phytase units/kg diet. Hen performance and the percentage of damaged eggs were measured every 4 weeks. Body weight, precaecal digestibility of P, and bone parameters at 45 woa were also measured. The reduction in avP and Ca in the NC diet did not compromise performance or egg quality. However, it decreased (P < 0.001) body weight, tibial dry matter, tibial ash and P content, and precaecal digestibility of P. Importantly, all these parameters were significantly improved (P < 0.001) and essentially restored to the levels measured in PC diet-fed hens upon supplementation with phytase. In summary, the present study demonstrates that the new bacterial 6-phytase could effectively counteract the negative effects of P and Ca deficiencies on body weight, bone mineralization, and P availability, thereby supporting high productivity without compromising the welfare of laying hens.
Key words: Keywords:available phosphorus, calcium, digestibility, laying hens, mineralization, phytase

DOI： https://doi.org/10.2141/jpsa.2023019

卵黄嚢膜における糖新生：採卵鶏の発生過程における酵素活性，遺伝子発現および代謝産物について
柴田光浩・岩澤　淳・八代田真人
岐阜大学大学院連合農学研究科 岐阜市柳戸1-1　501-1193

採卵鶏の卵黄嚢膜を孵卵3日目から孵化後3日目まで毎日採取し（n=7-9），糖新生の鍵酵素（グルコース-6-ホスファターゼ，フルクトース-1，6-ビスホスファターゼ，細胞質およびミトコンドリアのホスホエノールピルビン酸カルボキシキナーゼ，ピルビン酸カルボキシラーゼ）のmRNA発現および酵素活性を定量した。卵黄嚢膜の乳酸，トリグリセリド，遊離脂肪酸，グリコーゲン，グルコース，および血糖値も測定した。調査したすべての酵素のmRNA発現と活性が認められた。発現量や酵素活性の違いは，胚の発生環境と発生段階ごとの生理的要求を反映していると思われた。すなわち，孵卵 1 週目から 2 週目半ばまでは，糖新生酵素の発現量と活性および乳酸濃度が高く，漿尿膜の毛細血管が完成する前の胚の低酸素状態を反映して，乳酸からの糖新生が活発であることが示唆された。胚が好気的状態にある第 2 週半ばから第 3 週半ばにかけて，卵黄嚢内のトリグリセリドと遊離脂肪酸の含量が増加した。卵黄脂質のトリグリセリドは加水分解されてエネルギー源となる遊離脂肪酸を生成し，グリセロール骨格は糖新生に利用される可能性がある。胚が再び嫌気状態になると考えられる3週目後半には，グルコース-6-ホスファターゼのみのmRNA発現と酵素活性が高くなり，卵黄嚢内のグリコーゲン量が減少していることが確認された。したがって，この時期は糖新生の活性が低く，卵黄嚢膜に貯蔵された糖質が血液中に分泌され，孵化のためのエネルギーとなっていることが示唆される。本研究では，卵黄嚢膜が孵卵中のニワトリ胚の重要な糖新生器官であることを確認できた。
キーワード：ニワトリ胚，糖新生，乳酸，PEPCK，卵黄嚢

DOI： https://doi.org/10.2141/jpsa.2023020

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Gluconeogenesis in the Yolk Sac Membrane: Enzyme Activity, Gene Expression, and Metabolites During Layer Chicken Development

Mitsuhiro Shibata1, Atsushi Iwasawa2 and Masato Yayota2

1 1The United Graduate School of Agricultural Science, Gifu University, Gifu 501-1193, Japan
2 2Faculty of Applied Biological Sciences, Gifu University, Gifu 501-1193, Japan

抄録
Yolk sac membranes of layer eggs were collected daily (n = 7–9) from day three of incubation to day three post-hatch, and mRNA expression and activities were quantified for key gluconeogenesis enzymes (glucose-6-phosphatase, fructose-1,6- bisphosphatase, cytosolic and mitochondrial phosphoenolpyruvate carboxykinases, and pyruvate carboxylase). Lactate, triglycerides, non-esterified fatty acids, glycogen, and glucose in the yolk sac membrane, and blood glucose levels were also measured. The mRNA expression and activity were detected for all enzymes. Differences in expression levels and enzyme activities seemed to reflect the embryo’s developmental environment and physiological demands at different developmental stages. During the first week to the mid-second week of incubation, the expression and activity of gluconeogenic enzymes and lactate concentrations were high, suggesting an active period of gluconeogenesis from lactate, reflecting possible hypoxia in the embryo before completed formation of the chorioallantoic capillaries. From the mid-second week to mid-third week, when embryos were in an aerobic state, the triglyceride and non-esterified fatty acid contents increased in the yolk sac. Triglycerides from yolk lipids are typically hydrolyzed to produce non-esterified fatty acids as an energy source, whereas the glycerol skeleton is used for gluconeogenesis. In the late third week, when embryos were considered to re-enter an anaerobic state, the mRNA expression and enzyme activity of only glucose-6-phosphatase were high and the amount of glycogen in the yolk sac was reduced. Therefore, it is suggested that gluconeogenesis activity is low during this period, and the carbohydrates stored in the yolk sac membrane are secreted into the blood as energy for hatching. This study confirmed the role of the yolk sac membrane as a vital gluconeogenic organ during chicken egg incubation.
Key words: chicken embryo, gluconeogenesis, lactate, PEPCK, yolk sac

DOI： https://doi.org/10.2141/jpsa.2023020

ミトコンドリアの酸化的リン酸化脱共役が鶏培養筋管細胞のオートファジーに及ぼす影響
中島一喜・石田藍子
農研機構 畜産研究部門　茨城県つくば市池の台2　305-0901

オートファジーは、異化的条件下で細胞内タンパク質をバルクに分解するシステムである。骨格筋において、細胞内エネルギーの枯渇がタンパク質分解を促進することが知られているが、鶏骨格筋においては、ミトコンドリアのエネルギー産生とオートファジーとの関係は十分に検討されていない。そこで、本研究では、ミトコンドリアの酸化的リン酸化脱共役が、鶏筋管細胞のオートファジーに及ぼす影響を調べた。鶏筋管細胞をミトコンドリア脱共役剤であるカルボニルシアナイド−メタクロロフェニルヒドラゾン(CCCP, 25 M）を添加した培地で3時間培養した。オートファジーの指標としてサイトゾルLC3-II量を測定し、また、マイトファジーの指標として、ミトコンドリア分画のLC3-II量を測定した。さらに、オートファジー関連遺伝子（LC3B、GABARAPL1およびATG12）の発現量を測定した。また、mTOR活性ならびにタンパク質合成についても測定した。鶏筋管細胞において、CCCPにより細胞内ならびにミトコンドリア分画のLC3-II量が増加した。オートファジー関連遺伝子発現には、影響は見られなかった。mTOR活性ならびにタンパク質合成はCCCPにより減少した。以上の結果から、鶏骨格筋のミトコンドリアにおける酸化的リン酸化脱共役は、オートファジーを促進する可能性が示唆された。
Key words: autophagy, carbonyl cyanide m-chlorophenyl hydrazone (CCCP), chick myotubes, mitophagy, protein synthesis

DOI： https://doi.org/10.2141/jpsa.2023022

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Regulation of autophagy in chick myotube cultures: Effect of uncoupling mitochondrial oxidative phosphorylation

Kazuki Nakashima and Aiko Ishida

Institute of Livestock and Grassland Science, NARO, Tsukuba 305-0901, Japan

抄録
Abstracts: Skeletal muscles have a high demand for ATP, which is met largely through mitochondria oxidative phosphorylation. Autophagy is essential for the maintenance of skeletal muscle mass under catabolic conditions. This study investigated the effect of uncoupling mitochondrial oxidative phosphorylation on autophagy in chicken skeletal muscle. Chick myotubes were incubated with the mitochondrial uncoupler carbonyl cyanide m-chlorophenyl hydrazone (CCCP) at 25 μM for 3h. CCCP prevented the phosphorylation of p70 ribosomal S6 kinase 1 (Thr389), S6 ribosomal protein (Ser240/244), and eukaryotic translation initiation factor 4E-binding protein 1 (Thr37/46), which are the measures of the mechanistic target of rapamycin complex 1 (mTORC1) activity. CCCP significantly increased cytoplasmic and mitochondrial LC3-II content, which act as indices of index for autophagosome formation and mitophagy, respectively, but did not influence the expression of autophagy-related genes LC3B, GABARAPL1, and ATG12. Finally, surface sensing of translation method revealed that protein synthesis, a highly energy consuming process, was significantly decreased upon CCCP treatment. These results indicate that the uncoupling of mitochondrial oxidative phosphorylation stimulates autophagy and inhibits protein synthesis through mTORC1 signaling in chick myotube cultures.
Key words: autophagy, carbonyl cyanide m-chlorophenyl hydrazone (CCCP), chick myotubes, mitophagy, protein synthesis

DOI： https://doi.org/10.2141/jpsa.2023022

Effect of Dietary Gelsemium elegans Benth. Extract on the Growth, Slaughter Performance, Meat Quality, Intestinal Morphology, and Microflora of Yellow-Feathered Chickens

Yu-Hang Cao1, Ting-Ting Chen1, Xiong Peng1, Rong-Rong Wu1, Xiang Li2, Gao-Feng Liu2, Li-Xia Shen1, Xiao-Jun Chen1, Zi Yang3, Zhao-Ying Liu1, Zhi-Liang Sun1 and Yong Wu1

1College of Veterinary Medicine, Hunan Agricultural University, Changsha 410128, China
2Hunan Canzoho Biological Technology Co., Ltd., Liuyang 410329, China
3Academician Workstation, Changsha Medical University, Changsha 410219, China
抄録
The plant species Gelsemium elegans Benth. (GEB) promotes pig and sheep growth; however, little is known about its effects in chickens. In this study, a GEB extract (GEBE) was prepared, and its effects on the growth, slaughter, antioxidant performance, meat quality, serum biochemical indices, intestinal morphology, and microflora of yellow-feathered chickens were evaluated. In total, 600 chickens aged 15 days were randomly divided into four groups with five replicates each and fed a basal diet containing 0% (control), 0.25% (0.25 GEBE), 0.75% (0.75 GEBE), or 1.25% (1.25 GEBE) GEBE until 49 days of age. Chickens were then killed, and their meat, organs, and serum and cecal contents were collected. GEBE reduced the feed conversion ratio, particularly in the 0.75 and 1.25 GEBE groups. Furthermore, the GEBE diet improved meat tenderness and reduced the meat expressible moisture content and liver malondialdehyde content, indicating high meat quality. Whereas the 0.25 GEBE diet increased the level of Lactobacillus acidophilus in the cecum, the 0.75 GEBE diet decreased the Escherichia coli level therein. These findings demonstrate that GEBE may improve the meat quality and cecal microbiota of yellow-feathered chickens, providing a basis for identifying candidate alternatives to conventional antibiotics as growth promoting feed additives.
Key words: feed conversion ratio, Gelsemium elegans Benth., intestinal morphology, meat quality, slaughter performance, yellow-feathered chickens

DOI： https://doi.org/10.2141/jpsa.2023023

Effects of Artemisia annua L. Water Extract on Growth Performance and Intestinal Related Indicators in Broilers

Shiwei Guo, Jiaxin Ma, Yuanyuan Xing, Yuanqing Xu, Xiao Jin, Sumei Yan, Lulu Shi, Linghui Zhang and Binlin Shi

College of Animal Science, Inner Mongolia Agricultural University, Hohhot, 010018, P. R. China
抄録
Artemisia annua L. is a natural herb with a variety of bioactive substances, which can play a variety of biological functions such as anti-inflammatory, antioxidant, antibacterial and antiviral, and can be used as a potential feed additive. The purpose of this study was to investigate the effects of different doses of Artemisia annua L. water extract (AAWE) on growth performance and intestinal related indicators in broilers. A total of 200 one-day-old Arbor Acre broilers were selected and randomly divided into five treatment groups, with five replicates in each group and eight birds per replicate. The control group was fed a basal diet, whereas the other groups were fed a basal diet supplemented with 0.5, 1.0, 1.5, or 2.0 g/kg AAWE. On d21, with the increase in AAWE dose, final body weight and feed efficiency showed a quadratic increase effect, whereas feed intake showed a linear reduction effect; however, the apparent metabolic rate of dry matter, crude protein, and ether extract increased quadratically on d 42. In addition, the activity of duodenal chymotrypsin and trypsin, and of jejunal lipase quadratically increased, whereas the intestine crypt depth linearly decreased on d 42. The number of total anaerobic bacteria increased quadratically, whereas the number of Escherichia coli decreased quadratically. The number of Lactobacillus increased linearly, whereas H2S emission linearly decreased on d 21; moreover, NH3 emission (24 h) quadratically decreased on d 42. In conclusion, AAWE promoted the growth performance and intestinal related indicators of broilers.
Key words: apparent metabolic rate, Artemisia annua L. water extract, digestive enzyme activity, growth performance, harmful gas, intestinal flora

DOI： https://doi.org/10.2141/jpsa.2023024

ウズラの卵胞におけるリラキシン3の発現

ホアン スアン コイ1、松崎芽衣2、高坂哲也3、笹浪知宏1,4

1岐阜大学大学院連合農学研究科
2広島大学大学院統合生命科学研究科
3大阪物療大学　保健医療学部
4静岡大学農学部

哺乳類のリラキシンは生殖器官で発現し、例えば分娩時の恥骨靭帯の弛緩に関与することが知られている。ニワトリにおいても卵巣での発現が報告されているが、その生理的な意義は不明である。本研究ではウズラを用いて、リラキシンの組織発現および発現のホルモン制御を調べた。半定量または定量RT-PCR解析の結果から、リラキシンは卵巣の顆粒膜細胞および卵胞膜細胞で発現し、顆粒膜細胞のリラキシンは卵胞発育と共に発現が上昇することがわかった。顆粒膜細胞の初代培養実験の結果から、リラキシンの発現はエストラジールの添加で上昇し、プロゲステロンの添加で減少することがわかった。また、リラキシン発現を卵胞の部位別に測定したところ、スティグマ部分でもっとも高く、スティグマ部分では、排卵が近づくと発現が激減することがわかった。以上の結果より、ウズラにおけるリラキシンは、顆粒膜細胞で主に作られ、排卵が近づくと発現量が低下することから、何らかの形で排卵に関わっている可能性が示唆された。

DOI： https://doi.org/10.2141/jpsa.2023025

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Expression of Relaxin 3 in the Ovarian Follicle of Japanese Quail

Khoi X. Hoang1, Mei Matsuzaki 2, Tetsuya Kohsaka3 and Tomohiro Sasanami1,4

1United Graduate School of Agricultural Science, Gifu University, 1-1 Yanagido, Gifu 501-1193, Japan
2Program of Food and AgriLife Science, Graduate School of Integrated Sciences for Life, Hiroshima University, 1-4-4 Kagamiyama, Higashi-Hiroshima City, Hiroshima 739-8528, Japan
3Faculty of Health Science, Butsuryo College of Osaka, 3-33 Otorikita-machi, Sakai, Osaka 593-8328, Japan
4Department of Applied Life Sciences, Faculty of Agriculture, Shizuoka University, 836 Ohya, Shizuoka, Shizuoka 422-8529, Japan

The relaxin (RLN) gene is expressed in the reproductive tracts, such as the ovary and uterus, of mammalian species. Although RLN expression is detected in the chicken ovary, detailed clarification of the physiological role of RLN has not yet been reported. To address this issue, in the present study we aimed to examine the spatiotemporal expression and hormonal control of RLN in Japanese quail. By performing semi-quantitative and quantitative reverse transcription-polymerase chain reaction analysis, we found that RLN mRNA was mainly expressed in the granulosa and theca layers of the ovary. The expression level in the granulosa layer increased with the stage of follicular development. Results from granulosa layer culture experiments revealed that RLN mRNA expression increased with the addition of estradiol-17β, whereas the addition of progesterone suppressed RLN transcription. More detailed analysis indicated that RLN expression was highest in the stigma region of the follicle but significantly decreased as the time of the expected luteinizing hormone (LH) surge approached. Together, our findings demonstrated that the granulosa cells in the mature preovulatory follicles constitute the main source of RLN in the Japanese quail. Because RLN expression was highest in the stigma region and the expression dramatically decreased following the LH surge, the results further suggest that RLN may be related to tissue remodeling for the ovulation process in birds. Key words: follicle, granulosa layer, Japanese quail, relaxin, stigma

DOI： https://doi.org/10.2141/jpsa.2023025

シャモの皮膚におけるコラーゲン含有量とコラーゲン線維の構築特性

西村正太郎・荒井彩也香・保坂善真

九州大学大学院農学研究院　福岡市西区元岡744 813-0395

シャモの皮膚に含まれるコラーゲン量とコラーゲン線維の構築特性について雌雄間並びに体の部位間で比較した。頚部，胸部，背部，大腿部及び下腿部より皮膚を採取し，コラーゲン含有量を測定した。また，アルカリ浸軟法により作製した皮膚のコラーゲン線維標本を，走査型電子顕微鏡を用いて観察した。皮膚のコラーゲン含有量の雌雄差は成鶏では大腿部においてのみ認められたが，10週齢の若鶏ではいずれの部位においても認められなかった。コラーゲン含有量は体の部位間で異なっており，雌雄ともに大腿部及び下腿部の皮膚で他の部位よりも高い値を示した。走査型電子顕微鏡による観察の結果，コラーゲン線維の構築像は真皮の表在層と深層で全く異なり，表在層では帯状のコラーゲン線維が緩く絡み合っていたのに対し，深層では厚く密なコラーゲン線維層が幾重にも重なっていた。真皮表在層におけるコラーゲン線維の幅は，背部，大腿部及び下腿部の皮膚において雌雄間で異なっていた。これらの結果より，シャモ成鶏の大腿部皮膚におけるコラーゲン含有量の雌雄差は性ホルモンの影響によるものではなく，日常の活動における機械的刺激など他の要因によることが考えられる。また，真皮表在層におけるコラーゲン線維の太さは，雌雄間並びに部位間におけるコラーゲン含有量の違いに関与しているのかもしれない。 キーワード：コラーゲン，SEM，シャモ，皮膚

DOI： https://doi.org/10.2141/jpsa.2023026

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Collagen Content and Collagen Fiber Architecture in the Skin of Shamo Chicken, a Japanese Game Fowl
Shotaro Nishimura1, Sayaka Arai2 and Yoshinao Z Hosaka1

1 Faculty of Agriculture, Kyushu University, Fukuoka, Japan
2 Graduate School of Bioresource and Bioenvironmental Sciences, Kyushu University, Fukuoka, Japan

抄録
Collagen content and collagen fiber architecture in the skin of Shamo chickens were compared between sexes and body parts. Cervical, thoracic, dorsal, femoral, and crural skin samples were collected and their collagen content was analyzed. Collagen fiber specimens were prepared for scanning electron microscopy using the cell maceration method with a NaOH solution. Sex differences in collagen content were only observed in the femoral skin of mature chickens, but not in 10-weekold chicks. The difference in collagen content between body parts was obvious; femoral and crural skin had higher collagen content than those of other parts in both sexes. Scanning electron microscopy indicated that the collagen fiber architecture was quite different between the superficial and deep layers in the dermis, with the former consisting of loosely tangled band-like collagen fibers, and the latter composed of thick and dense layers of collagen bundles in a parallel arrangement. The width of collagen fibers in the superficial layer of the dermis differed between sexes in the dorsal, femoral, and crural skin. From these results, it is likely that the difference in collagen content in the femoral skin is not due to sex hormones but other factors, such as mechanical stimulation in daily activity. Additionally, collagen fiber width in the superficial layer is likely related to the difference in collagen content between sexes and between body parts.
Key words: collagen, SEM, Shamo chicken, skin

DOI： https://doi.org/10.2141/jpsa.2023026

微酸性電解水の浸漬頻度の違いが生の骨付き鶏モモ肉の品質に及ぼす影響

Muliasari Kartikawati1・北村豊1・粉川美踏1・濵谷希人2・副島 隆志2

1 筑波大学大学院生命環境科学研究科, 2 森永乳業株式会社

微酸性電解水（SAEW）は，現在主に鶏生肉の殺菌に使用されている。ただし，一度の浸漬で使用される水量が肉の重さの10倍にもなるため，大量の廃水が発生している。一方，興味深いことに，浸漬の頻度が増加すると微生物汚染が減少する可能性があると考えられている。よって本研究の目的は，SAEWの浸漬を異なる頻度で行うことが，生の骨付き鶏モモ肉の殺菌と品質に及ぼす影響を詳細に調査し，これによってSAEWの使用を制限する可能性を検証することにある。浸漬の頻度は1回，3回，5回で，SAEW：肉の比率は7：1，浸漬時間は計15分に設定された。肉の品質は，総好気性細菌，腸内細菌，総揮発性塩基性窒素，チオバルビツール酸反応性物質，および色に基づいて評価された。実験では，浸漬の頻度が高い場合，好気性細菌と腸内細菌科細菌の総数が減少した。さらに，SAEW：肉の比率が4：1で，総浸漬時間が６分の2回浸漬は，SAEW：肉の比率が7：1で同15分の1回の浸漬と同程度，効果的に細菌量を減少させた。また，SAEWの浸漬頻度が高いほど，保存後の総揮発性塩基性窒素と脂質の酸化も低かった。ただし，肉の色調は変化し，結果として肉はより鮮やかになった。総じて，2～5回の浸漬によるSAEW処理は，生の骨付き鶏モモ肉の品質を向上させつつ，廃水の発生を減少させる殺菌方法となる可能性が示唆された。

DOI： https://doi.org/10.2141/jpsa.2023027

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Effect of Slightly Acidic Electrolyzed Water Immersion at Different Frequencies on Quality of Raw Chicken Legs

Muliasari Kartikawati1, Yutaka Kitamura1, Mito Kokawa1, Mareto Hamatani2 and Takashi Soejima2

1Graduate School of Life and Environmental Sciences University of Tsukuba, Tennodai, Tsukuba 305-0005, Japan
2Morinaga Milk Industry Co., LTD., 33-1, Shiba 5-Chome, Minato-ku, Tokyo, 108-0014, Japan

抄録
Slightly acidic electrolyzed water (SAEW) is used as a disinfectant for raw chicken meat. Because its volume for a single immersion exceeds 10 times the weight of meat, a large amount of wastewater is generated. Importantly, a higher frequency of immersion is believed to reduce microbial contamination. The objective of this study was to investigate the effect of SAEW immersion at different frequencies on the disinfection and quality of raw chicken legs, thereby possibly limiting the usage of SAEW. Immersion for 1, 3, and 5 times, with a 7:1 SAEW:meat ratio, and duration of 15 min was tested. Meat quality was evaluated based on total aerobic bacteria, Enterobactericeae, total volatile basic nitrogen, thiobarbituric acid reactive substances, and color. A higher immersion frequency lowered the numbers of total aerobic bacteria and Enterobacteriaceae. Moreover, two immersions with a SAEW:meat ratio of 4:1 and a total immersion time of 6 min reduced the bacterial load as effectively as a single 15-min immersion with a SAEW:meat ratio of 7:1. Higher frequencies of SAEW immersion also resulted in lower total volatile basic nitrogen and lipid oxidation after 0 or 3 days of storage. They did, however, magnify the change in color, resulting in brighter meat. Overall, SAEW treatments with two to five immersions can improve the quality of raw chicken legs and reduce wastewater generation.
Key words: Keywords:chicken, frequencies, immersion, microorganism, quality, SAEW

DOI： https://doi.org/10.2141/jpsa.2023027

ニワトリ7日胚の生殖巣生殖細胞における移住・増殖の解析

中島友紀1,2・田上貴寛2・田島淳史1

1筑波大学生命環境系、茨城県つくば市天王台1-1-1
2農業・食品産業技術総合研究機構畜産研究部門、茨城県つくば市池の台2

本研究はニワトリ7日胚生殖巣に存在する生殖巣生殖細胞（GGC）の生殖巣への移住および増殖能の解析を目的とした。まず、GGCsの移住能を解析するため、蛍光標識した雌雄のGGCs 50個を2.5日胚に移植した。移植5日後の宿主胚生殖巣で観察された蛍光陽性GGCsの割合は生殖巣外より有意に高かった（P<0.05）。また、生殖巣外のGGCsのほとんどは生殖巣周辺領域である腸間膜および中腎で観察された。次に、GGCsの宿主胚生殖巣とその周辺領域での移動・増殖様式を解析するため、蛍光標識した雌雄のGGCs 50個を2.5日胚に移植し、移植3および6日後の宿主胚生殖巣および中腎における蛍光陽性GGC数を比較した。その結果、宿主胚生殖巣で観察されたGGCsは移植6日後に有意に増加したのに対し、中腎で観察されたGGCsは6日後に有意に減少した（P<0.05）。さらに、GGCの均質性を解析するため、1胚あたり1細胞の蛍光標識GGCを移植した。その結果、移植5日後に宿主胚生殖巣へ生着したGGCの割合は、雌雄で有意差は認められなかったが、生殖巣で増殖能を示すGGCの割合は、雄の方が雌よりも有意に高かった（P<0.05）。また、GGCの増殖率は雌および雄でそれぞれ2-19倍および2-12倍であり、細胞ごとのばらつきが大きかった。以上より、本研究において7日胚GGCは胚内ではほとんどが生殖巣領域に移住し、増殖能はドナーの性別および細胞間で異なることが明らかとなった。
キーワード：ニワトリ、生殖巣生殖細胞、移住、増殖、7日胚

DOI： https://doi.org/10.2141/jpsa.2023028

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Gonadal Germ Cell Migration and Proliferation after Transfer in Developing Chicken Embryos

Yuki Nakajima1,2, Takahiro Tagami2 and Atsushi Tajima1

1 Faculty of Life and Environmental Sciences, University of Tsukuba, Ten-noh Dai 1-1-1, Tsukuba, Ibaraki 305-8572,Japan
2 Division of Meat Animal and Poultry Research, NARO Institute of Livestock and Grassland Science, 2 Ikenodai, Tsukuba, Ibaraki 305-0901, Japan

抄録
A germline chimera is a useful model for developing and differentiating germ cells in vivo. Gonadal germ cells (GGCs) collected from chicken embryonic gonads may be used to produce germline chimeras as donor cells. However, the migratory and proliferative abilities of GGCs after transfer into recipient embryos are unclear. Here, the migratory and proliferative abilities of GGCs collected from 7-day-old White Leghorn embryos and fluorescently labeled were analyzed following transfer into the dorsal aorta of 2.5-day-old Rhode Island Red (RIR) embryos. Five days after transfer, the numbers of male and female GGCs were significantly higher in the RIR gonads than those in non-gonadal RIR organs when 50 GGCs were transferred per embryo. To analyze the temporal migration of GGCs in intermediate mesoderm, 50 GGCs were again transferred. The numbers of male and female GGCs in RIR gonads increased significantly from days 3 to 6 after transfer. To analyze GGC migration and proliferation in the gonads, a single GGC was transferred into 100 male and 100 female embryos. Five days after transfer, the frequencies of settled and proliferated GGCs were 37% (37/100) and 24% (24/100) in males, and 23% (23/100) and 8% (8/100) in females, respectively. Thus, GGCs are a heterogeneous cell population that may or may not have migratory and proliferative abilities. The heterogeneity of GGCs may be greater in females than that in males. When 50 GGCs were transplanted, almost all those present in embryos had settled and proliferated in the gonads and mesonephros. The migratory and proliferative abilities of GGCs in recipient gonads were considerably diverse in individual GGCs or between donor sexes.
Key words: 7-day-old embryo, chicken, gonadal germ cells (GGCs), migration, proliferation

DOI： https://doi.org/10.2141/jpsa.2023028

孵化前後におけるニワトリ腸管内分泌細胞の動態

エムディ アル アミン1・エムディ サラフディン2・今井裕理子3・平松浩二3

1信州大学大学院総合医理工学研究科　長野県上伊那郡南箕輪村8304　399-4598
2プレイリー・ビューA＆M大学　アメリカ合衆国
3信州大学学術研究院農学系　長野県上伊那郡南箕輪村8304　399-4598

本研究は、孵化前後のニワトリ腸内分泌細胞の組織学的特徴を明らかにすることを目的とした。孵卵18日目、19日目、20日目、21日目の胚及び3日齢のヒナから十二指腸、空腸と回腸の近位部と遠位部、盲腸と結直腸を組織サンプルとして採取した。グルカゴン様ペプチド(GLP)-1、ソマトスタチン(SST)、ニューロテンシン(NT)の発現をストレプトアビジン-ビオチン法で検出し、これらのペプチド間の共局在を二重免疫蛍光法で示した。すべてのホルモンは、孵卵18日目にはすでに腸管細胞で発現していた。GLP-1免疫反応陽性細胞は空腸と回腸でのみ観察され、その数は孵卵ステージが進むにつれて徐々に増加した。NT-免疫反応陽性細胞はすべてのステージにおいてすべての腸管部位で検出され、3日齢のヒナでは結直腸で最も発現が高かった。SST免疫反応陽性細胞は結直腸を除く十二指腸から回腸にかけて観察された。二重免疫蛍光法により、GLP-1とNTは空腸と回腸の同じ内分泌細胞に共局在することが示された。GLP-1とNTの共局在比は3日齢のヒナで最も高かった。一方、SSTはGLP-1ともNTとも共局在しなかった。これらの結果は、ニワトリの腸内分泌細胞は孵化前後でその密度と共局在比が著しく変化することを示している。

DOI： https://doi.org/10.2141/jpsa.2023029

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Characteristics of Enteroendocrine Cells of White Leghorn Chickens, Gallus gallus, Before and After Hatching

Md. Al amin, Md Salahuddin, Yuriko Imai, Kohzy Hiramatsu

The aim of this study was to identify the histological features of chicken enteroendocrine cells before and after hatching. Tissue samples from the duodenum, proximal and distal parts of the jejunum and ileum, cecum and colorectum were collected from the embryos at days 18, 19, 20, and 21 of incubation, and from 3-day-old chicks. The expression of glucagon-like peptide (GLP)-1, somatostatin (SST), and neurotensin (NT) in the enteroendocrine cells was detected using the streptavidin-biotin method, and the colocalization of these peptides was revealed using the double immunofluorescence method. All of assessed peptides were expressed in the enteroendocrine cells at day 18 of incubation. GLP-1-immunoreactive cells were only observed in the jejunum and ileum. The cell numbers gradually increased as incubation progressed. NT-immunoreactive cells were detected in all intestinal parts at all incubation days, and the highest expression was observed in the colorectum of 3-day-old chicks. SST-immunoreactive cells were observed from the duodenum to the ileum, excluding the colorectum. The double immunofluorescence method revealed that GLP-1 and NT colocalized in the same endocrine cells of the jejunum and ileum. The colocalization ratio of GLP-1 with NT was the highest in the distal ileum of 3-day-old chicks. However, neither GLP-1 nor NT colocalized with SST. These results indicate that chicken enteroendocrine cells markedly change their density and colocalization ratios before and after hatching.

DOI： https://doi.org/10.2141/jpsa.2023029

D-ガラクトサミンはリポポリサッカライドと相乗的にニワトリヒナの肝臓の損傷を引き起こす

高橋真紀1・千貫晃1・寺岡慧1・シャキル カーン2・牧野良輔1・マーク A クライン3・橘哲也1

1愛媛大学農学部　松山市　790-8566
2大分大学医学部　由布市　879-5593
3バージニア工科大学神経科学　ブラックスバーグ　24061

哺乳類において、グラム陰性菌由来のリポポリサッカライド（LPS）や真菌由来のザイモサンなどの病原体関連分子パターン（PAMPs）は自然免疫を活性化するとともに、肝臓や消化管などの臓器に損傷をもたらす。このPAMPsによる損傷はD-ガラクトサミン（D-GalN）の同時投与によって増強されることがげっ歯類を用いた研究で明らかにされている。しかし、鳥類においてはPAMPsやD-GalNが協調して臓器損傷を引き起こすかは明らかにされていない。そこで、本研究ではD-GalNとLPSまたはザイモサンの同時腹腔内投与がニワトリヒナの血漿中肝臓損傷マーカーに与える影響を調査した。D-GalNはそれ自体では血漿中アスパラギン酸アミノトランスフェラーゼ（AST）、アラニンアミノトランスフェラーゼ（ALT）、および乳酸脱水素酵素（LDH）活性に影響を与えなかったものの、LPSと同時投与した場合にはこれらの血漿中活性を増加させた。一方、ザイモサンおよびザイモサンとD-GalNの同時投与はこれらの血漿中肝臓損傷マーカーに影響を与えなかった。LPSとD-GalNの同時投与は肝臓中インターロイキン-6（IL-6）および誘導型一酸化窒素合成酵素（iNOS）の遺伝子発現量を増加させた。一方、ザイモサンとD-GalNの同時投与は肝臓損傷マーカーや肝臓中IL-6遺伝子発現量に影響を与えなかったが肝臓中iNOS遺伝子発現量を増加させた。したがって、IL-6がLPSとD-GalNの同時投与による肝臓の損傷に関与している可能性を示した。
キーワード ：ニワトリヒナ、D-ガラクトサミン、損傷、リポポリサッカライド、肝臓、ザイモサン

DOI： https://doi.org/10.2141/jpsa.2023031

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D-Galactosamine Causes Liver Injury Synergistically with Lipopolysaccharide but not Zymosan in Chicks

Maki Takahashi, Akira Senga, Kei Teraoka, Sakirul Khan, Ryosuke Makino, Mark A. Cline, Tetsuya Tachibana

The pathogen-associated molecular patterns (PAMPs) lipopolysaccharide (LPS) and zymosan, derived from gram-negative bacteria and fungi, respectively, activate the innate immune system and cause injury to multiple organs, including the liver and intestine, in mammals. In rodents, PAMP-induced injury has been demonstrated to be potentiated by co-administration of D-galactosamine (D-GalN) in rodents. However, whether PAMPs and D-GalN collectively cause organ injury in birds remains unclear. The present study aimed to measure the effects of intraperitoneal injection of D-GalN with LPS or zymosan on parameters related to hepatic injury in chicks (Gallus gallus). Plasma aspartate aminotransferase (AST), alanine aminotransferase (ALT), and lactate dehydrogenase (LDH) activities were not affected by intraperitoneal injection of D-GalN alone. Although these activities were not affected by LPS injection alone, they were increased by combining LPS with D-GalN. In contrast, plasma AST, ALT, and LDH activities were not affected by zymosan, both alone and with D-GalN. The expression of mRNAs for interleukin-6 (IL-6) and inducible nitric oxide synthase (iNOS) in the liver was significantly increased by the combination of LPS and D-GalN. In contrast, combining zymosan with D-GalN significantly increased iNOS mRNA expression, irrespective of hepatic injury. These results suggest that IL-6 may be the cause and/or result of hepatic injury in chicks. Additionally, chicks are tolerant to the hepatic effects of D-GalN, LPS, or zymosan alone.

DOI： https://doi.org/10.2141/jpsa.2023031

初生ヒナへの飼料給与開始の遅延が暑熱環境下で飼育した肉用鶏の血漿中代謝物に及ぼす影響

片渕歩美１・島元紗希２・川口真奈3・友永省三4・中島一喜5・石原慎矢6・大塚彰1,3,7・井尻大地1,3,7

1 鹿児島大学農林水産学研究科、鹿児島市郡元1-21-24 890-0065
2 新潟大学自然科学研究科、新潟市西区五十嵐2の町8050 950-2181
3 鹿児島大学農学部生物資源化学科、鹿児島市郡元1-21-24 890-0065
4 京都大学大学院農学研究科、京都市左京区北白川追分町 606-8502
5 農研機構畜産研究部門、つくば市池ノ台2 300-1240
6 日本獣医生命科学大学応用生命科学部、武蔵野市境南町1-7-1 180-8602

肉用鶏初生ヒナへの飼料給与開始が遅れると，増体量が減少し，鶏肉の収量や肉質に負の影響が現れる。本研究では, 孵化後の飼料給与の遅延が肉用鶏の血漿中の代謝物に及ぼす影響について, ノンターゲット法とターゲット法を組み合わせたメタボローム解析により調べた。0日齢の肉用鶏ヒナ（Ross308）12羽を対照区と遅延区に分けた。対照区には0日齢から給餌し，遅延区には2日齢から給餌し, 13日齢まで飼育した．高速液体クロマトグラフ-蛍光検出器によりOPA誘導体化したアミノ酸のターゲット分析を行い, ガスクロマトグラフ質量分析計により有機酸, 糖などの低分子代謝物のノンターゲット分析を行った。飼料給与の遅延は血漿中のスクロース, マルトース, ラクチトール, ゲンチオビオース, キシリトール, セロトニン, スレオニン酸およびアスパラギンを増加させ, クレアチニン, アスパラギン酸およびグルタミン酸を減少させた。また, 飼料給与の遅延により可溶性無窒素物の消化率および盲腸内の酪酸濃度が増加した。以上の結果より, 2日間（48時間）の飼料給与の遅延は, 消化管内の炭水化物の消化率や盲腸内の酪酸濃度の増加を介して, 生体内の炭水化物, 酪酸, アミノ酸ならびに窒素に関連する代謝経路を変動させることが示唆された。
キーワード: 肉用鶏, 初期栄養, 代謝物, メタボローム解析, 初生ヒナ

DOI： https://doi.org/10.2141/jpsa.2023032

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Effects of Delaying Post-hatch Feeding on the Plasma Metabolites of Broiler Chickens Revealed by Targeted and Untargeted Metabolomics

Ayumi Katafuchi, Saki Shimamoto, Mana Kawaguchi, Shozo Tomonaga, Kazuki Nakashima, Shinya Ishihara, Akira Ohtsuka, Daichi Ijiri

Exogenous nutrients are essential for body and skeletal muscle growth in newly hatched chicks, and delaying post-hatch feeding negatively affects body growth, meat yield, and meat quality. The aim of this study was to investigate the effects of delayed post-hatch feeding on the metabolic profiles of broiler chickens using a combination of targeted and untargeted metabolomics. Newly hatched chicks had either immediate free access to feed (freely fed chicks) or no access to feed from 0 to 2 days of age (delayed-fed chicks); both groups were subsequently provided feed ad libitum until 13 days of age. Untargeted metabolomic analysis was performed using gas chromatography-mass spectrometry, whereas targeted metabolomic analysis of amino acids was performed using high-performance liquid chromatography with ortho-phthalaldehyde derivatization. Delayed feeding increased the plasma levels of sucrose, maltose, serotonin, lactitol, gentiobiose, xylitol, threonic acid, and asparagine, and decreased the plasma levels of creatinine, aspartic acid, and glutamic acid. In addition, the digestibility of the nitrogen-free extract (starch and sugar) and the cecal butyric acid concentration increased in chicks subjected to delayed feeding. In contrast, delayed feeding did not affect muscle protein degradation or digestibility in chicks. Taken together, our results indicate that delaying feeding until 48 h post-hatch alters multiple metabolic pathways, which are accompanied by changes in intestinal carbohydrate digestion and cecal butyric acid content in broiler chickens.

DOI： https://doi.org/10.2141/jpsa.2023032

Identification of Polymorphisms in the HERC2-OCA2 Gene Locus and their Association with Feather Color in Quail

Xiaohui Zhang1, Yanxia Qi1,2, Youzhi Pang1,2, Bingjie Yuan1 and Xiaolong Li1

1College of Animal Science, Henan University of Science and Technology, Luoyang, 471003 He’nan, P.R. China
2Luoyang Key Laboratory of Animal Genetics and Breeding, Luoyang 471003, P.R. China

抄録
Mutations in the HERC2 and OCA2 genes have the potential to affect pigment deposition and alter feather color in birds. Therefore, in this study, we evaluated HERC2-OCA2 gene locus polymorphisms in Korean and Beijing white quails using RNA-Seq and KASP technology. The expression levels of HERC2 and OCA2 mRNA in skin tissues were analyzed using RT-qPCR. Ten single nucleotide polymorphisms were identified by RNA-Seq, of which three (n.117627564T>A,n.117674275T>G, n.117686226A>C) exhibited significant association with feather color in quail. The expression of OCA2 mRNA was significantly lower in the skin of Beijing white quails than that in the skin of Korean quails. These results suggested that variants in HERC2-OCA2 intergenic region could influence the expression of OCA2, which may underlie diluted feather color in the Beijing white quail.
Key words: gene expression, HERC2, OCA2, polymorphism, quail

DOI： https://doi.org/10.2141/jpsa.2023013

雄鶏の鳴き声と存在が名古屋種産卵鶏の抱卵行動に及ぼす影響

中村明弘1・小林慧三1・宮川博充1・神作宜男2

1愛知県農業総合試験場畜産研究部　愛知県長久手市岩作三ケ峯　480-1193
２麻布大学獣医学部動物資源育種学研究室　神奈川県相模原市中央区淵野辺　252-5201

名古屋種の抱卵行動は産卵の急激な低下を突然に引き起こすことから，産業上の問題となっている。本研究では，名古屋種産卵鶏の抱卵行動の発生率が同じ鶏舎内の雄鶏の鳴き声や存在によって影響するか調査した。雄鶏の存在による影響を確認するため，孵化時期と雄鶏の提示の順序を変えた4つの試験が採卵用の名古屋種実用鶏を用いて行われた。試験1では，供試鶏を飼育するペンの隣のペンに雄鶏が存在する時の抱卵行動の発生率は51.3%と，雄鶏の不在時(15.9%)あるいは鳴き声のみの提示時(23.8%)と比べて高かった。さらに，試験2, 3および4では，雄鶏が存在する時の抱卵行動の発生率(47.3%, 33.3%および37.9%)は不在時(33.3%, 17.4%および25.6%)より高かった。すべての試験において雄鶏の不在時に観察された抱卵鶏の約70％は，存在時でも抱卵行動を示した。したがって，名古屋種産卵鶏の抱卵行動は，雄鶏の存在によって増進する可能性がある。
キーワード：抱卵行動，産卵鶏，名古屋種，雄鶏

DOI： https://doi.org/10.2141/jpsa.20230230

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Effects of the Calls and Presence of Roosters on Egg Incubation Behavior of Nagoya Laying Hens

Akihiro Nakamura, Keizou Kobayashi, Hiromitsu Miyakawa, Norio Kansaku

The incubation behavior of the Japanese Nagoya chicken breed is a commercial issue because it often causes a sudden and sharp drop in egg production. In this study, whether the incidence of incubation behavior in Nagoya laying hens was associated with calls and the presence of roosters in the same laying house was investigated. Four experiments were conducted using commercial layer-type Nagoya hens where the hatching time of the experimental birds and the treatment order in the presence of males were changed . In Experiment 1, the proportion of incubation behavior in the presence of roosters kept in another pen located between pen-rearing hens (51.3%) was higher than that in their absence (15.9%) or with only rooster calls (23.8%). In Experiments 2, 3, and 4, the proportion of incubation behavior in the presence of roosters (47.3%, 33.3%, and 37.9%, respectively) was higher than that in their absence (33.3%, 17.4%, and 25.6%, respectively). In all experiments, approximately 70% of the incubating hens observed in the absence of roosters exhibited incubation behavior, even in the presence of roosters. Therefore, the presence of roosters may enhance egg incubation behavior in Nagoya laying hens.

DOI： https://doi.org/10.2141/jpsa.2023030