Graduate School of Agricultural Science, Tohoku University, Osaki, Miyagi 989-6711, Japan.
Department of Biotechnology, Faculty of Bioresource Sciences, Akita Prefectural University, Akita, Akita 010-0195, Japan.
J Biosci Bioeng. 2020 Aug;130(2):137-141. doi: 10.1016/j.jbiosc.2020.03.010. Epub 2020 Apr 21.
We had developed a new pretreatment system using cow rumen fluid to improve the methane production from lignocellulosic substrates. However, the pretreatment conditions differ from the in-situ rumen environment, therefore different microbes may be involved in plant cell wall decomposition. In the current study, shotgun metagenomic analysis using MiSeq platform was performed to elucidate the bacteria which produce cellulase and hemicellulase in this pretreatment system. The rumen fluid which contained waste paper pieces (0.1% w/v) were incubated at 37°C during 120 h. The fluid samples were collected from the reactor at each time-point and analyzed for chemical properties. Rumen microbial DNA was extracted from 0-h and 60-h samples and subjected to shotgun-metagenomic analysis. After pretreatment, approximately half of cellulose and hemicellulose contents of the waste paper were decomposed and some volatile fatty acids were accumulated. Clostridia (e.g., Ruminococcus and Clostridium) were the predominant bacteria before and after 60-h pretreatment, and their relative abundance was increased during pretreatment. However, Prevotella and Fibrobacter, one of the most dominant bacteria in-situ rumen fluid, were observed less than 3% before incubation and they were decreased after pretreatment. Genes encoding cellulase and hemicellulase were mainly found in Ruminococcus, Clostridium, and Caldicellulosiruptor. Calicellulosiruptor, which had not been previously identified as the predominant genus in lignocellulose decomposition in in-situ rumen conditions, might be considered as the main fibrolytic bacterium in this system. Thus, this study demonstrated that the composition of fibrolytic bacteria in this system was greatly different from those in the in-situ rumen.
我们开发了一种新的预处理系统，使用牛瘤胃液来提高木质纤维素底物的甲烷产量。然而，预处理条件与原位瘤胃环境不同，因此可能涉及不同的微生物参与植物细胞壁的分解。在本研究中，使用 MiSeq 平台进行了 shotgun 宏基因组分析，以阐明在该预处理系统中产生纤维素酶和半纤维素酶的细菌。将含有废纸碎片（0.1%w/v）的瘤胃液在 37°C 下孵育 120 小时。在每个时间点从反应器中收集流体样品并分析其化学性质。从 0 小时和 60 小时的样品中提取瘤胃微生物 DNA，并进行 shotgun 宏基因组分析。预处理后，废纸中约一半的纤维素和半纤维素含量被分解，一些挥发性脂肪酸积累。梭菌（如 Ruminococcus 和 Clostridium）是预处理前后的主要细菌，其相对丰度在预处理过程中增加。然而，普雷沃氏菌和纤维菌（一种原位瘤胃液中最主要的细菌之一）在孵育前观察到不到 3%，预处理后减少。纤维素酶和半纤维素酶的基因主要存在于 Ruminococcus、Clostridium 和 Caldicellulosiruptor 中。Calicellulosiruptor 以前未被鉴定为原位瘤胃条件下木质纤维素分解的主要属，在该系统中可能被认为是主要的纤维分解细菌。因此，本研究表明该系统中纤维分解细菌的组成与原位瘤胃中的组成有很大的不同。
