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Simultaneous Metabarcoding and Quantification of Neocallimastigomycetes from Environmental Samples: Insights into Community Composition and Novel Lineages

Young, Diana ; Joshi, Akshay; Huang, Liren; Munk, Bernhard; Wurzbacher, Christian; Youssef, Noha H.; Elshahed, Mostafa S.; Moon, Christina D.; Ochsenreither, Katrin ORCID iD icon 1; Griffith, Gareth W.; Callaghan, Tony M.; Sczyrba, Alexander; Lebuhn, Michael ; Flad, Veronika
1 Institut für Bio- und Lebensmitteltechnik (BLT), Karlsruher Institut für Technologie (KIT)

Abstract (englisch):

Anaerobic fungi from the herbivore digestive tract (Neocallimastigomycetes) are primary lignocellulose modifiers and hold promise for biotechnological applications. Their molecular detection is currently difficult due to the non-specificity of published primer pairs, which impairs evolutionary and ecological research with environmental samples. We developed and validated a Neocallimastigomycetes-specific PCR primer pair targeting the D2 region of the ribosomal large subunit suitable for screening, quantifying, and sequencing. We evaluated this primer pair in silico on sequences from all known genera, in vitro with pure cultures covering 16 of the 20 known genera, and on environmental samples with highly diverse microbiomes. The amplified region allowed phylogenetic differentiation of all known genera and most species. The amplicon is about 350 bp long, suitable for short-read high-throughput sequencing as well as qPCR assays. Sequencing of herbivore fecal samples verified the specificity of the primer pair and recovered highly diverse and so far unknown anaerobic gut fungal taxa. As the chosen barcoding region can be easily aligned and is taxonomically informative, the sequences can be used for classification and phylogenetic inferences. ... mehr


Verlagsausgabe §
DOI: 10.5445/IR/1000150342
Veröffentlicht am 05.09.2022
Originalveröffentlichung
DOI: 10.3390/microorganisms10091749
Scopus
Zitationen: 11
Web of Science
Zitationen: 8
Dimensions
Zitationen: 18
Cover der Publikation
Zugehörige Institution(en) am KIT Institut für Bio- und Lebensmitteltechnik (BLT)
Publikationstyp Zeitschriftenaufsatz
Publikationsjahr 2022
Sprache Englisch
Identifikator ISSN: 2076-2607
KITopen-ID: 1000150342
Erschienen in Microorganisms
Verlag MDPI
Band 10
Heft 9
Seiten Art.-Nr.: 1749
Vorab online veröffentlicht am 30.08.2022
Schlagwörter anaerobic gut fungi; Neocallimastigomycetes; environmental screening; Quantitative Real-Time PCR; large ribosomal subunit; barcoding; high-throughput sequencing; phylogenetic analysis
Nachgewiesen in Web of Science
Dimensions
Scopus
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