Show simple item record

dc.contributor.authorCastelino, M
dc.contributor.authorEyre, S
dc.contributor.authorMoat, J
dc.contributor.authorFox, G
dc.contributor.authorMartin, P
dc.contributor.authorHo, P
dc.contributor.authorUpton, Mathew
dc.contributor.authorBarton, A
dc.date.accessioned2017-05-09T13:15:58Z
dc.date.available2017-05-09T13:15:58Z
dc.date.issued2017-12
dc.identifier.issn1471-2180
dc.identifier.issn1471-2180
dc.identifier.other23
dc.identifier.urihttp://hdl.handle.net/10026.1/9205
dc.description.abstract

BACKGROUND: The composition of the skin microbiome is predicted to play a role in the development of conditions such as atopic eczema and psoriasis. 16S rRNA gene sequencing allows the investigation of bacterial microbiota. A significant challenge in this field is development of cost effective high throughput methodologies for the robust interrogation of the skin microbiota, where biomass is low. Here we describe validation of methodologies for 16S rRNA (ribosomal ribonucleic acid) gene sequencing from the skin microbiome, using the Illumina MiSeq platform, the selection of primer to amplify regions for sequencing and we compare results with the current standard protocols.. METHODS: DNA was obtained from two low density mock communities of 11 diverse bacterial strains (with and without human DNA supplementation) and from swabs taken from the skin of healthy volunteers. This was amplified using primer pairs covering hypervariable regions of the 16S rRNA gene: primers 63F and 519R (V1-V3); and 347F and 803R (V3-V4). The resultant libraries were indexed for the MiSeq and Roche454 and sequenced. Both data sets were denoised, cleaned of chimeras and analysed using QIIME. RESULTS: There was no significant difference in the diversity indices at the phylum and the genus level observed between the platforms. The capture of diversity using the low density mock community samples demonstrated that the primer pair spanning the V3-V4 hypervariable region had better capture when compared to the primer pair for the V1-V3 region and was robust to spiking with human DNA. The pilot data generated using the V3-V4 region from the skin of healthy volunteers was consistent with these results, even at the genus level (Staphylococcus, Propionibacterium, Corynebacterium, Paracoccus, Micrococcus, Enhydrobacter and Deinococcus identified at similar abundances on both platforms). CONCLUSIONS: The results suggest that the bacterial community diversity captured using the V3-V4 16S rRNA hypervariable region from sequencing using the MiSeq platform is comparable to the Roche454 GS Junior platform. These findings provide evidence that the optimised method can be used in human clinical samples of low bacterial biomass such as the investigation of the skin microbiota.

dc.format.extent23-
dc.format.mediumElectronic
dc.languageen
dc.language.isoen
dc.publisherSpringer Science and Business Media LLC
dc.subjectSkin microbiome
dc.subjectBacterial microbiome
dc.subjectLow biomass
dc.subject16s rRNA gene
dc.subjectMiseq platform
dc.subjectNext generation sequencing
dc.subjectMethods
dc.subjectOptimisation
dc.subjectPrimer selection
dc.titleOptimisation of methods for bacterial skin microbiome investigation: primer selection and comparison of the 454 versus MiSeq platform
dc.typejournal-article
dc.typeJournal Article
plymouth.author-urlhttps://www.webofscience.com/api/gateway?GWVersion=2&SrcApp=PARTNER_APP&SrcAuth=LinksAMR&KeyUT=WOS:000393327500001&DestLinkType=FullRecord&DestApp=ALL_WOS&UsrCustomerID=11bb513d99f797142bcfeffcc58ea008
plymouth.issue1
plymouth.volume17
plymouth.publication-statusPublished online
plymouth.journalBMC Microbiology
dc.identifier.doi10.1186/s12866-017-0927-4
plymouth.organisational-group/Plymouth
plymouth.organisational-group/Plymouth/Faculty of Health
plymouth.organisational-group/Plymouth/Faculty of Health/School of Biomedical Sciences
plymouth.organisational-group/Plymouth/REF 2021 Researchers by UoA
plymouth.organisational-group/Plymouth/REF 2021 Researchers by UoA/UoA01 Clinical Medicine
plymouth.organisational-group/Plymouth/Research Groups
plymouth.organisational-group/Plymouth/Research Groups/Institute of Translational and Stratified Medicine (ITSMED)
plymouth.organisational-group/Plymouth/Research Groups/Institute of Translational and Stratified Medicine (ITSMED)/CBR
plymouth.organisational-group/Plymouth/Research Groups/Plymouth Institute of Health and Care Research (PIHR)
plymouth.organisational-group/Plymouth/Users by role
plymouth.organisational-group/Plymouth/Users by role/Academics
plymouth.organisational-group/Plymouth/Users by role/Researchers in ResearchFish submission
dc.publisher.placeEngland
dcterms.dateAccepted2017-01-09
dc.identifier.eissn1471-2180
dc.rights.embargoperiodNo embargo
rioxxterms.versionofrecord10.1186/s12866-017-0927-4
rioxxterms.licenseref.urihttp://www.rioxx.net/licenses/all-rights-reserved
rioxxterms.licenseref.startdate2017-12
rioxxterms.typeJournal Article/Review
plymouth.oa-locationhttps://bmcmicrobiol.biomedcentral.com/articles/10.1186/s12866-017-0927-4


Files in this item

Thumbnail
Thumbnail

This item appears in the following Collection(s)

Show simple item record


All items in PEARL are protected by copyright law.
Author manuscripts deposited to comply with open access mandates are made available in accordance with publisher policies. Please cite only the published version using the details provided on the item record or document. In the absence of an open licence (e.g. Creative Commons), permissions for further reuse of content should be sought from the publisher or author.
Theme by 
Atmire NV