氏名TANG THUY MINH授与した学位博士

専攻分野の名称農学

学位授与番号

博甲第学位授与の日付

平成２９年 ９月２９日

学位授与の要件

環境生命科学研究科

５６３０号

農生命科学専攻（学位規則第４条第１項該当）

学位論文の題目

Robustness and resilience of gut microbiota in ruminants in relation to feeding and nutritionalmanagement（給与飼料及び飼養管理との関連からみた反芻動物腸内細菌叢の堅牢性及び順応性）

論文審査委員

教授

齋藤昇教授

舟橋弘晃

教授

西野直樹

学位論文内容の概要
The divergence of gut microbiota in ruminants is affected by many factors including genotype, aging, farmmanagement, geographical locations, and diet formulation. Likewise, differences can be seen even when animals
consume the same diet and show similar physiological profiles. Because microbial activity and metabolism in the gut isa key to secure health and productivity of ruminants, understanding robustness and resilience of the gut microbiota is ofgreat importance. Therefore, in this thesis, three experiments were carried out to examine variability, stability, andadaptability of gut microbiota in relation to feeding and nutritional management.
In the first experiment, fecal microbiota of seven Holstein heifers was examined during growing stage, wherein
weaning and feeding of silage, a lactic acid bacteria-enriched fermented food, were practiced. Populations of total
bacteria and Bacteroides-Prevotella-Porphyromonas, Clostridium XIVa, and Clostridium I-II groups were relatively
stable during the growing stage, and those of Lactobacillus and Bifidobacterium groups decreased after weaning.
Regarding bacterial composition, distinctive changes were shown after weaning in all bacterial taxa examined, whereas
no changes were observed thereafter.
In the second experiment, the microbiota of whole crop corn silage and feces of 18 silage-fed lactating cows were
examined in Japan and China. Although fecal microbiota composition was similar in most samples, relative abundancesof several families, such as Ruminococcaceae, Christensenellaceae, Turicibacteraceae, and Succinivibrionaceae varied
between farms and countries. In addition to the geographical location, differences in feeding management between total
mixed ration feeding and separate feeding appeared to be involved in the variations. Moreover, a cow-to-cow variation
for concentrate-associated families was demonstrated at the same farm.
In the third experiment, digesta- and mucosa-associated microbiota were examined across the gastrointestinal tract
using goats. The bacterial populations varied across gut segments and between bacterial taxa. In digesta-associatedsamples, the populations of total bacteria, Bacteroides-Prevotella-Porphyromonas group, Faecalibacterium prausnitzii,
and Ruminococcus albus were low at duodenum, whereas the numbers appeared similar between rumen and large
intestine. The population of Lactobacillus group was highly stable across the gut segments. In mucosa-associatedsamples, variations of bacterial numbers between gut segments were small regardless of bacterial taxa, whereas thenumbers of Enterobacteriaceae, Lactobacillus group, F. prausnitzii, and R. albus were higher at duodenum than others.
It is concluded that diet formulation and feeding management showed substantial influence on shaping the gut
microbiota in ruminants, whereas food-borne bacteria like silage lactic acid bacteria had little ability of modulation.
Weaning had a great impact regardless of bacterial taxa. Variations with regard to farms, countries, feeding regimes, andindividual cows and differences between gut segments and between digesta- and mucosa-associated samples were also
clarified. Because most of the findings to date are derived from rectum samples, there is still a large room for
improvement on understanding the roles and functions of gut microbiota in ruminants.

論文審査結果の要旨

腸内細菌の構成と機能を理解することは，反芻動物の栄養，健康及び生産性を制御するうえで重要である。
給与飼料，月齢，遺伝，管理衛生，地理的環境などが腸内細菌叢を変動させると考えられているが，同じ環境及び飼料で飼育されている個体同士でも違いがある。TANG Thuy Minh 氏は，反芻動物の腸内細菌叢がもつ多様性，安定性及び順応性について，飼養管理との関連を中心に調査研究を行った。
最初の実験では，ホルスタイン種育成牛（雌）を対象として，成長過程における腸内細菌叢の変化を調べた。
離 乳 後 に Lactobacillus spp. 及 び Bifidobacterium spp. は 減 少 し た が ， Bacteroides-Prevotella-Porphyromonas,Clostridium XIVa 及び Clostridium I-II の菌数は，離乳前後でも大きく変化しなかった。
発酵飼料には Lactobacillusspp.が数多く存在したが，その給与で腸内細菌叢が変化することはなかった。
続 く 実 験 で は ， 日 本 と 中 国 で 泌 乳 牛 か ら 糞 便 を 採 取 し ， MiSeq に よ る 腸 内 細 菌 叢 解 析 を 行 っ た 。
Ruminococcaceae, Christensenellaceae, Turicibacteraceae 及び Succinivibrionaceae は，農場，調査国及び分離給与と TMR（混合飼料）給与という飼養管理法による違いが認められた。一方，穀類消化に関わる細菌群が同一農場でも個体によって異なる構成を示すなど，腸内細菌叢には一定の適応性が認められた。
最後の実験では，腸内容物と腸粘膜に存在する細菌群を分けて解析した。発酵飼料を含む TMR をシバヤギに給与し，第 1 胃，第 4 胃，小腸，盲腸及び直腸からサンプルを採取して腸内細菌叢を調べた。腸内容物に存在する Bacteroides-Prevotella-Porphyromonas, Faecalibacterium prausnitzii 及び Ruminococcus albus は第 4 胃で菌数が低下したが，第 1 胃と直腸ではほぼ同様の高値を示した。腸粘膜に存在する細菌群は採取部位に関わらず安定した菌数を示し，Lactobacillus spp.は給与飼料の Lactobacillus spp.にほとんど影響されなかった。
これらの知見は，反芻動物の腸内細菌叢に給与飼料を含めた飼養管理技術が強く影響することを示している。
飼料として与えた Lactobacillus spp.が腸内細菌叢を改善しないことも意義ある知見であり，TANG Thuy Minh 氏は博士（農学）の学位を授与するに値すると判断した。