Wang Xiaofan, Howe Samantha, Wei Xiaoyuan, Deng Feilong, Tsai Tsungcheng, Chai Jianmin, Xiao Yingping, Yang Hua, Maxwell Charles V, Li Ying, Zhao Jiangchao
Department of Animal Science, Division of Agriculture, University of Arkansas, Fayetteville, Arkansas, USA.
Guangdong Provincial Key Laboratory of Animal Molecular Design and Precise Breeding, College of Life Science and Engineering, Foshan University, Foshan, China.
mSystems. 2021 Aug 31;6(4):e0047721. doi: 10.1128/mSystems.00477-21. Epub 2021 Jul 20.
Despite the substantial progress made in human gut culturomics, little is known about the culturability of the swine gut microbiota. In this study, we cultured swine gut microbiota using 53 bacterial cultivation methods with different medium and gas combinations from three pigs at four different growth stages. Both culture-dependent (CD; colony mixtures from each method) and culture-independent (CI; original fecal suspensions) samples were subjected to 16S rRNA gene amplicon sequencing. Increasing microbial diversities were observed in both CI and CD samples from successive growth stages. While a total of 378, 482, 565, and 555 bacterial amplicon sequence variants (ASVs) were observed in the CI samples, higher microbial diversities (415, 675, 808, and 823 observed ASVs) were detected using the CD methods at the lactation, nursery, growing, and finishing stages, respectively. We constructed reference culture maps showing the preferred cultivation conditions for specific bacterial taxa and examined the effects of culturing factors such as oxygen, medium, donor pig age, antibiotics, and blood culture preincubation on swine gut microbiota cultivation. We focused on a wide range of beneficial bacteria, chose 1,299 colonies based on the reference map, and Sanger sequenced their 16S rRNA genes. These isolates clustered into 148 different bacterial taxa covering 28 genera. We observed 11, 19, 33, and 25 pairs of cooccurring ASVs in both CD and CI samples at four successive growth stages. This study provides guidance in culturing the swine gut microbiota of interest, which is critical when characterizing their functions in this important animal species. The swine gut microbiome has been the focus of many investigations due to the fact that pigs serve as both an excellent biomedical model for human diseases and an important protein source. Substantial progress has been made in swine gut microbiome studies using next-generation sequencing-based culture-independent approaches, but little is known about the culturability of the swine gut microbiota. To understand their roles in swine production, it is critical to culture bacterial strains of interest. In this study, we cultured the gut microbiota from pigs at different growth stages using 53 bacterial cultivation methods with different medium and gas combinations. This study provides evidence that the swine gut microbiota is much more diverse based on a culture-dependent approach than previously known. It provides preliminary guidance for isolating certain bacteria of interest from pigs, which is critical in establishing causal relationships between the gut microbiota and the health status of pigs.
尽管人类肠道培养组学已取得重大进展，但对于猪肠道微生物群的可培养性却知之甚少。在本研究中，我们使用53种不同培养基和气体组合的细菌培养方法，对来自三头处于四个不同生长阶段的猪的肠道微生物群进行培养。对基于培养法（CD；每种方法的菌落混合物）和非培养法（CI；原始粪便悬液）的样本进行16S rRNA基因扩增子测序。在连续生长阶段的CI和CD样本中均观察到微生物多样性增加。虽然在CI样本中总共观察到378、482、565和555个细菌扩增子序列变体（ASV），但在哺乳、保育、生长和育肥阶段分别使用CD方法检测到更高的微生物多样性（观察到的ASV分别为415、675、808和823个）。我们构建了参考培养图谱，显示特定细菌类群的优选培养条件，并研究了培养因素如氧气、培养基、供体猪年龄、抗生素和血培养预孵育对猪肠道微生物群培养的影响。我们关注多种有益细菌，根据参考图谱选择了1299个菌落，并对其16S rRNA基因进行桑格测序。这些分离株聚为148个不同的细菌类群，涵盖28个属。在四个连续生长阶段的CD和CI样本中，我们分别观察到11、19、33和25对共现ASV。本研究为培养感兴趣的猪肠道微生物群提供了指导，这在表征其在这种重要动物物种中的功能时至关重要。由于猪既是人类疾病的优秀生物医学模型又是重要的蛋白质来源，猪肠道微生物组一直是许多研究的重点。使用基于下一代测序的非培养方法在猪肠道微生物组研究中已取得重大进展，但对于猪肠道微生物群的可培养性却知之甚少。为了解它们在猪生产中的作用，培养感兴趣的细菌菌株至关重要。在本研究中，我们使用53种不同培养基和气体组合的细菌培养方法，对不同生长阶段猪的肠道微生物群进行培养。本研究提供的证据表明，基于培养法的猪肠道微生物群比以前所知的更加多样。它为从猪中分离某些感兴趣的细菌提供了初步指导，这对于建立肠道微生物群与猪健康状况之间的因果关系至关重要。
