ORCID Profile
0000-0002-2593-9642
Current Organisation
University of Southampton
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Publisher: Wiley
Date: 04-05-2018
DOI: 10.1002/PPUL.24037
Abstract: The study of the community of microorganisms (the microbiota) in the lower airways in children is restricted to opportunistic s ling in children undergoing elective general anaesthetic. Here we tested the hypothesis that induced sputum is a valid alternative to directly s ling the lower airways to study lower airway microbiota. Children scheduled for elective operations were recruited. Pre-operatively a s le of induced sputum was obtained. After anaesthesia was induced, a bronchial brushing and swabs of the upper respiratory tract were obtained. Bacterial community analysis was performed by lification of the V3-V4 16S rRNA gene region. Twenty children were recruited, mean age 10.7 years. Induced sputum s les were obtained from 12 children, bronchial brushing from 14 and nasal, mouth, and throat s les in 15, 16, and 17 children. The profile of bacterial communities was similar in the mouth, throat, and sputum s les with the nose and bronchial s les being different. Actinobacteria species dominated the nose and mouth, Fusobacteria were the dominant species in the throat and sputum while Proteobacteria species dominated in bronchial s les. Forty-one percent of detected bacteria in bronchial s les were unclassified. Bacterial communities from the mouth, throat, and induced sputum were tightly clustered and were distinct from nose and those found in bronchial communities. Induced sputum may not be a valid surrogate for microbiome assessment of the lower airways in all in iduals. Many bacteria in bronchial s les were not recognized by standard testing, suggesting that our understanding of the lower airway microbiota in children remains rudimentary.
Publisher: Oxford University Press (OUP)
Date: 03-2022
DOI: 10.1111/LAM.13618
Abstract: The human oral cavity is host to a erse microbiota. Much of what is known about the behaviour of oral microbes derives from studies of in idual or several cultivated species, situations which do not totally reflect the function of organisms within more complex microbiota or multispecies biofilms. The number of validated models that allow examination of the role that biofilms play during oral cavity colonization is also limited. The CDC biofilm reactor is a standard method that has been deployed to study interactions between members of human microbiotas allowing studies to be completed during an extended period under conditions where nutrient availability, and washout of waste products are controlled. The objective of this work was to develop a robust in vitro biofilm-model system from a pooled saliva inoculum to study the development, reproducibility and stability of the oral microbiota. By employing deep sequencing of the variable regions of the 16S rRNA gene, we found that the CDC biofilm reactor could be used to efficiently cultivate microbiota containing all six major phyla previously identified as the core saliva microbiota. After an acclimatisation period, communities in each reactor stabilised. Replicate reactors were predominately populated by a shared core microbiota variation between replicate reactors was primarily driven by shifts in abundance of shared operational taxonomic units. We conclude that the CDC biofilm reactor can be used to cultivate communities that replicate key features of the human oral cavity and is a useful tool to facilitate studies of the dynamics of these communities.
Publisher: Springer Science and Business Media LLC
Date: 02-01-2020
DOI: 10.1186/S12866-019-1672-7
Abstract: Interactions between transcription factors and DNA lie at the centre of many biological processes including DNA recombination, replication, repair and transcription. Most bacteria encode erse proteins that act as transcription factors to regulate various traits. Several technologies for identifying protein–DNA interactions at the genomic level have been developed. Bind-n-seq is a high-throughput in vitro method first deployed to analyse DNA interactions associated with eukaryotic zinc-finger proteins. The method has three steps (i) binding protein to a randomised oligonucleotide DNA target library, (ii) deep sequencing of bound oligonucleotides, and (iii) a computational algorithm to define motifs among the sequences. The classical Bind-n-seq strategy suffers from several limitations including a lengthy wet laboratory protocol and a computational algorithm that is difficult to use. We introduce here an improved, rapid, and simplified Bind-n-seq protocol coupled with a user-friendly downstream data analysis and handling algorithm, which has been optimized for bacterial target proteins. We validate this new protocol by showing the successful characterisation of the DNA-binding specificities of YipR (YajQ interacting protein regulator), a well-known transcriptional regulator of virulence genes in the bacterial phytopathogen Xanthomonas c estris pv. c estris ( Xcc ). The improved Bind-n-seq approach identified several DNA binding motif sequences for YipR, in particular the CCCTCTC motif, which were located in the promoter regions of 1320 Xcc genes. Informatics analysis revealed that many of these genes regulate functions associated with virulence, motility, and biofilm formation and included genes previously found involved in virulence. Additionally, electromobility shift assays show that YipR binds to the promoter region of XC_2633 in a CCCTCTC motif-dependent manner. We present a new and rapid Bind-n-seq protocol that should be useful to investigate DNA-binding proteins in bacteria. The analysis of YipR DNA binding using this protocol identifies a novel DNA sequence motif in the promoter regions of target genes that define the YipR regulon.
Location: United Kingdom of Great Britain and Northern Ireland
Location: United Kingdom of Great Britain and Northern Ireland
Location: United Kingdom of Great Britain and Northern Ireland
No related grants have been discovered for Shi-qi An.