High-throughput sequencing identifies salivary microbiota in Chinese caries-free preschool children with primary dentition / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

OBJECTIVES@#The study aimed at identifying salivary microbiota in caries-free Chinese preschool children using high-throughput sequencing.@*METHODS@#Saliva samples were obtained from 35 caries-free preschool children (18 boys and 17 girls) with primary dentition, and 16S ribosomal DNA (rDNA) V3-V4 hypervariable regions of the microorganisms were analyzed using Illumina MiSeq.@*RESULTS@#At 97% similarity level, all of these reads were clustered into 334 operational taxonomic units (OTUs). Among these, five phyla (Firmicutes, Proteobacteria, Actinobacteria, Bacteroidetes, and Candidate division TM7) and 13 genera (@*CONCLUSIONS@#Our results revealed the diversity and composition of salivary microbiota in caries-free preschool children, with little difference between male and female subjects. Identity of the core microbiome, coupled with prediction of gene function, deepens our understanding of oral microbiota in caries-free populations and provides basic information for associating salivary microecology and oral health.

Analysis of microbial community characteristics of Panax notoginseng based on high-throughput sequencing technology / 中草药

Objective: The purpose of this study was to obtain the bacterial communities structures of fresh and dry Panax notoginseng. Methods: The bacterial were sequenced by Illumina MiSeq high-throughput sequencing technology, and the biological information was analyzed. Results: A total of 637 OTUs were obtained, of which 15 were in common, 484 were from fresh P. notoginseng and 123 were from dry P. notoginseng respectively. The larger phylum of fresh sample was Proteobacteria (59.7%) and Firmicutes (40.1%) while the dry sample were Firmicutes (99.8%). The main genus of fresh sample was Enterobacter (37.4%) and Alkaliphilus (11.5%), while dry sample were Bacillus (54.6%) and Paenibaciiius (44.9%). The functional prediction of KEGG showed that the population of bacteria metabolizing terpenoids and polyketones of dry sample was higher than that of fresh sample. Conclusion: The diversity of microbiota in fresh P. notoginseng was higher than that of dry sample; The species with the metabolic function of interpenoids and polyketones of dry P. notoginseng was higher than that of fresh sample, which provides an important reference for screening the microorganism for biotransformation of saponins in P. notoginseng.

Benefits of procyanidins on gut microbiota in Bama minipigs and implications in replacing antibiotics

Several studies have reported the effect of absorption of procyanidins and their contribution to the small intestine. However, differences between dietary interventions of procyanidins and interventions via antibiotic feeding in pigs are rarely reported. Following 16S rRNA gene Illumina MiSeq sequencing, we observed that both procyanidin administration for 2 months (procyanidin-1 group) and continuous antibiotic feeding for 1 month followed by procyanidin for 1 month (procyanidin-2 group) increased the number of operational taxonomic units, as well as the Chao 1 and ACE indices, compared to those in pigs undergoing antibiotic administration for 2 months (antibiotic group). The genera Fibrobacter and Spirochaete were more abundant in the antibiotic group than in the procyanidin-1 and procyanidin-2 groups. Principal component analysis revealed clear separations among the three groups. Additionally, using the online Molecular Ecological Network Analyses pipeline, three co-occurrence networks were constructed; Lactobacillus was in a co-occurrence relationship with Trichococcus and Desulfovibrio and a co-exclusion relationship with Bacillus and Spharerochaeta. Furthermore, metabolic function analysis by phylogenetic investigation of communities by reconstruction of unobserved states demonstrated modulation of pathways involved in the metabolism of carbohydrates, amino acids, energy, and nucleotides. These data suggest that procyanidin influences the gut microbiota and the intestinal metabolic function to produce beneficial effects on metabolic homeostasis.

Study on high throughput sequencing identification of Fructus Arctii and five counterfeit species mix power / 中国中药杂志

Fructus Arctii is a traditional Chinese medicine. The main counterfeit species are the seeds of Arctium tomentosum, Onopordum acanthium, Silybum marianum, Saussurea costus, Amorpha fruticosa. Traditional identification methods or molecular barcoding techniques can identify Fructus Arctii and its counterfeit species. However, the identification of the mixture of it and its spurious species is rarely reported. In this paper, we sequenced the ITS2 sequences of Fructus Arctii and 5 kinds of spurious species mix powder by high-throughput sequencing to identify the mixed powder species and providing new ideas for the identification of Fructus Arctii mix powder. The total DNA in mixed powder was extracted, and the ITS2 sequences in total DNA was amplified. Paired-end sequencing was performed on the DNA fragment of the community using the Illumina MiSeq platform. The sequence was analyzed by the software FLASH, QIIME and GraPhlAn etc. The results showed that the high quality ITS2 sequences of 39910 mix samples were obtained from the mixed samples, of which the total ITS2 sequence of the samples genus was 34 935. Phylogenetic analysis showed that the samples contained Fructus Arctii, A. tomentosum, O. acanthium, S. marianum, S. costus and A. fruticosa. Using ITS2 sequences as DNA barcodes, high-throughput sequencing technology can be used to detect the Fructus Arctii and its spurious specie in mixed powder, which can provide reference for the quality control, safe use of medicinal materials of Fructus Arctii and the identification of mixed powder of traditional Chinese medicine.

Diversity of endophytic bacteria in Astragali Radix analyzed by Illumina MiSeq high-throughput sequencing technology / 中草药

Objective This study aimed to analyze the diversity of endophytic bacterial communities, and to provide reference to the oriented processing technology, which helps to screen bacteria fermentation and conversion in the active ingredients of Astragali Radix. Methods The 16S rDNA V3-V4 region of Astragali Radix was sequenced by Illumina MiSeq high-throughput sequencing technology, and the abundance of species and other biological information were analyzed. Results The results showed that the numbers of effective sequences and OTUs for sample were 40 051 and 967, respectively. The number of sequencing was close to saturation, and the sequencing data volume was reasonable. The endophytic bacteria of Astragali Radix mainly belonged to Anderseniella, Bacillus, Burkholderia, Acinetobacter, Stenotrophomonas, and Oceanobacillus. Conclusion The diversity of endophytic bacteria was low in Astragali Radix. The dominant population of endophytic bacteria in Astragali Radix belongs to Anderseniella and Bacillus.

Study on DNA molecular identification of mix samples of five species of Baitouweng medicinal materials based on high-throughput sequencing technology / 药学学报

Traditional Chinese medicine Baitouweng have a long history of application. The pharmacopoeia included dry roots of Pulsatilla chinensis (Bge.) Regel of Ranunculaceae. There are easily confused species in the market circulation, such as P. cernua (Thunb.) Bercht. et Opiz., P. dahurica (Fisch.) Spreng., P. turczaninovii Kryl. et Serg., and P. chinensis (Bge.) Regel var. kissii (Mandl) S. H. Li et Y. H. Huang, etc. In this study, using the method of metagenomics, based on high-throughput sequencing technology, the ITS2 sequence of mixed samples of five species of Baitouweng medicinal materials was sequenced. First, the total DNA extraction of medicinal materials mixing powder, and the ITS2 fragment of total DNA was amplified by PCR. Second, the Illumina MiSeq platform was used to carry out Paired-end sequencing for DNA fragments. Last, using FLASH, QⅡME and GraPhlAn software to arrange and analyze, and clustering analysis with the sequences of uploaded to GenBank by our group in the early stage. The results showed that a total of 53 024 sequences of ITS2 were obtained from the mixed samples, there are 52 295 effective sequences, there are a total of 49 079 of five species of medicinal materials of P. Miller. After the representative sequences and the sequence of uploaded to GenBank by our group in the early stage were clustering analysis, 5 species of Baitouweng medicinal materials were clustered into one branch separately, presenting monophyletic. The results showed that using the high-throughput sequencing technology, using ITS2 sequence as DNA barcode, the mix powder of 5 species of Baitouweng medicinal materials could be effectively identified. It provides a new method and thought for the origin identification of mixed Chinese medicinal materials.

Analysis on microbial community diversity in gallbladder stone and mucosa based on Illumina Miseq platform / 重庆医学

Objective To investigate the microbial community diversity in stone and mucosa from the patients with gallbladder stone.Methods The gall bladder mucosal samples in 60 cases(case group) of gallbladder stone treated by cholecystectomy amd 11 cases (control group)of liver right anterior lobe hemangioma in this hospital from January to December 2014 were selected for conducting the bacterial culture.The stone and mucosa in the patients with negative culture results were performed the Illumina Miseq sequencing analysis.Results A total of 71 cases were included in the study.The microbial community DNA in the control group samples was not detected out.The bacterial DNA detection rate of samples in the case group was 94.2％;in regardless of the type of sample,the most dominant was Phylum Proteobacteria.The composition proportion of stone and mucosal microbial community in the patients with same type of stone was relatively consistent,whereas different types of composition had large difference.Conclusion Using the Illumina Miseq sequencing analysis can effectively evaluate the gallbladder microbial community diversity in the patients with gallbladder stone.

Analysis of microbial community of heat resistant microorganisms in Chinese herbal pieces / 中国中药杂志

To investigate the composition and diversity of heat resistant microorganisms in contaminated Chinese herbal pieces. Matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) protein fingerprinting and 16S rRNA high-throughput sequencing of Illumina Miseq were used to analyze the heat resistant microbial community of 9 varieties of Chinese herbal pieces. Stem pieces (Spatholobi Caulis, Tetrapanacis Medulla, Stachyuri Medulla) showed highest detection rate and most species of contaminants; However fruit pieces (Schisandrae Sphenantherae Fructus) had the lowest detection rate and least species of contaminants; among root pieces, the detection rate and number of contaminants species were lower in Tuber Dioscoreae Persimilis and Rehmanniae Radix Praeparata. The heat resistant microbial community was mainly of Bacillaceae and Paenibacillaceae, and Bacillus showed the highest detection rate among them, followed by Brevibacillus, Paenibacillus, and Solibacillus. The rest genus in high-throughput sequencing analysis included Enterobacter, Brevundimonas, Leuconostoc, Methylobacterium, Dechloromonas, Pantoea, Klebsiella, and Erwinia. There were potential risk factors in heat resistant microbial community of Chinese herbal pieces, so we shall improve the microbial limit standard, strictly control the pathogenic bacteria in the product, and strengthen the supervision and management in production and circulation of Chinese herbal pieces.

Illumina-based analysis of endophytic bacterial diversity of tree peony (Paeonia Sect. Moutan) roots and leaves

ABSTRACT Diverse communities of bacteria inhabit plant tissues and those bacteria play a crucial role for plant health and growth. Tree peony (Paeonia Sect. Moutan) is known for its excellent ornamental and medicinal values as Chinese traditional plant, but little is known about its associated bacterial community under natural conditions. To examine how endophytic bacteria in tree peony vary across tissues and cultivars, PCR-based Illumina was applied to reveal the diversity of endophytic bacteria in tree peony. A total of 149,842 sequences and 21,463 operational taxonomic units (OTUs) were obtained. The OTU abundance of roots was higher than leaves across other three cultivars except for 'Kinkaku' and 'Luoyanghong'. The community was composed of five dominant groups (Proteobacteria, Firmicutes, Bacteroidetes, Acidobacteria and Actinobacteria) in all samples. Endophytic bacteria community structures had changed in leaves and roots. Sequences of Pseudomonas and Enterobacteriaceae were prevalent in root samples, whereas Succinivibrio and Acinetobacter were the dominant genus in leaf samples. Otherwise, the distribution of each dominant genus among the 5 cultivars was either varied. These findings suggested that both plant genotype and tissues contribute to the shaping of the bacterial communities associated with tree peony.