MinION™ Nanopore Sequencing of Skin Microbiome 16S and 16S-23S rRNA Gene Amplicons

Abstract

Human skin microbiome dysbiosis can have clinical consequences. Characterizing taxonomic composition of bacterial communities associated with skin disorders is important for dermatological advancement in both diagnosis and novel treatments. This study aims to analyze and improve the accuracy of taxonomic classification of skin bacteria with MinION™ nanopore sequencing using a defined skin mock community and a skin microbiome sample. We compared the Oxford Nanopore Technologies recommended procedures and concluded that their protocols highly bias the relative abundance of certain skin microbiome genera, most notably a large overrepresentation of Staphylococcus and underrepresentation of Cutibacterium and Corynebacterium. We demonstrated that changes in the amplification protocols improved the accuracy of the taxonomic classification for these three main skin bacterial genera. This study shows that MinION™ nanopore could be an efficient technology for full-length 16S rRNA sequencing; however, the analytical advantage is strongly influenced by the methodologies. The suggested alternatives in the sample processing improved characterization of a complex skin microbiome community using MinION™ nanopore sequencing.

Nanopore Sequencing Using the Full-Length 16S rRNA Gene for Detection of Blood-Borne Bacteria in Dogs Reveals a Novel Species of Hemotropic Mycoplasma

“…NGS using the Illumina platform has been one of the most widely adopted methods for bacterial community analysis to date, due to the high depth of sequencing achievable and excellent raw read accuracy ( 24 , 25 ). Nonetheless, such methods are hindered by a maximum sequence length of about 500 bp and long library preparation and sequencing times ( 19 , 24 , 25 ).…”

“…NGS using the Illumina platform has been one of the most widely adopted methods for bacterial community analysis to date, due to the high depth of sequencing achievable and excellent raw read accuracy ( 24 , 25 ). Nonetheless, such methods are hindered by a maximum sequence length of about 500 bp and long library preparation and sequencing times ( 19 , 24 , 25 ). The arrival of third-generation sequencing methods such as nanopore-based sequencing by Oxford Nanopore Technologies (ONT) has obviated many of these limitations, including the ability to provide theoretically uncapped sequencing lengths with usable results obtained in as little as 5 min ( 19 , 26 , 27 ).…”

“…For microbiome analysis, the longer read lengths attainable through ONT platforms permit sequencing of the entire 16S rRNA gene or the full rrn operon, providing greatly improved taxonomic resolution ( 25 , 30 , 31 ). Such species- or even strain-level characterization is of crucial importance for veterinarians and clinicians, given the significant interspecific differences in pathogenicity and treatment approaches that exist between pathogens ( 31 , 32 ).…”

Developing Strategies to Help Bee Colony Resilience in Changing Environments

“…Over the last two decades, the microbiota has been largely studied in the context of human health and is linked with numerous pathological situations [ 8 , 11 , 17 , 22 , 23 , 24 , 28 , 32 , 33 , 35 , 43 , 48 , 50 , 59 , 60 , 61 , 62 , 63 , 64 , 65 , 66 , 67 , 68 , 69 , 70 , 71 , 72 , 73 , 74 , 75 , 76 , 77 , 78 , 79 , 80 , 81 , 82 , 83 , 84 , 85 , 86 , 87 ]. Interestingly, the microbiota is also seen as a cornerstone to honeybee health, as for so many other living organisms.…”

MinION Whole-Genome Sequencing in Resource-Limited Settings: Challenges and Opportunities

“…With the continued development of sequencing chemistry and software improvements, the capacity of MinION WGS has improved. MinION WGS has lower base call accuracy during the This article is part of the Topical Collection on Mycology * Kirsten Nielsen knielsen@umn.edu sequencing reaction than first-(Sanger sequencing) and secondgeneration (short-read WGS) sequencing [11,12]. However, various software updates that bioinformatically account for base miscalls have improved the MinION WGS sequencing accuracy to 90% [9].…”