Development of three specific PCR-based tools to determine quantity, cellulolytic transcriptional activity and phylogeny of anaerobic fungi

Dollhofer, Veronika; Callaghan, Tony M.; Dorn‐In, Samart; Bauer, Johann; Lebuhn, Michael

doi:10.1016/j.mimet.2016.05.017

Cited by 21 publications

(34 citation statements)

References 47 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Sizes of other regions are based on a Genbank reference sequence ( AJ864475 ). Primer references: 1 White et al ( 1990 ) and 2 Dollhofer et al ( 2016 ).…”

Section: Barcode Markers For Rapid Phylotyping Of Anaerobic Fungimentioning

confidence: 99%

PCR and Omics Based Techniques to Study the Diversity, Ecology and Biology of Anaerobic Fungi: Insights, Challenges and Opportunities

et al. 2017

Self Cite

View full text Add to dashboard Cite

Anaerobic fungi (phylum Neocallimastigomycota) are common inhabitants of the digestive tract of mammalian herbivores, and in the rumen, can account for up to 20% of the microbial biomass. Anaerobic fungi play a primary role in the degradation of lignocellulosic plant material. They also have a syntrophic interaction with methanogenic archaea, which increases their fiber degradation activity. To date, nine anaerobic fungal genera have been described, with further novel taxonomic groupings known to exist based on culture-independent molecular surveys. However, the true extent of their diversity may be even more extensively underestimated as anaerobic fungi continue being discovered in yet unexplored gut and non-gut environments. Additionally many studies are now known to have used primers that provide incomplete coverage of the Neocallimastigomycota. For ecological studies the internal transcribed spacer 1 region (ITS1) has been the taxonomic marker of choice, but due to various limitations the large subunit rRNA (LSU) is now being increasingly used. How the continued expansion of our knowledge regarding anaerobic fungal diversity will impact on our understanding of their biology and ecological role remains unclear; particularly as it is becoming apparent that anaerobic fungi display niche differentiation. As a consequence, there is a need to move beyond the broad generalization of anaerobic fungi as fiber-degraders, and explore the fundamental differences that underpin their ability to exist in distinct ecological niches. Application of genomics, transcriptomics, proteomics and metabolomics to their study in pure/mixed cultures and environmental samples will be invaluable in this process. To date the genomes and transcriptomes of several characterized anaerobic fungal isolates have been successfully generated. In contrast, the application of proteomics and metabolomics to anaerobic fungal analysis is still in its infancy. A central problem for all analyses, however, is the limited functional annotation of anaerobic fungal sequence data. There is therefore an urgent need to expand information held within publicly available reference databases. Once this challenge is overcome, along with improved sample collection and extraction, the application of these techniques will be key in furthering our understanding of the ecological role and impact of anaerobic fungi in the wide range of environments they inhabit.

show abstract

“…Sizes of other regions are based on a Genbank reference sequence ( AJ864475 ). Primer references: 1 White et al ( 1990 ) and 2 Dollhofer et al ( 2016 ).…”

Section: Barcode Markers For Rapid Phylotyping Of Anaerobic Fungimentioning

confidence: 99%

PCR and Omics Based Techniques to Study the Diversity, Ecology and Biology of Anaerobic Fungi: Insights, Challenges and Opportunities

et al. 2017

Self Cite

View full text Add to dashboard Cite

show abstract

“…First the concentration of anaerobic fungal 18S rDNA gene copies in the samples was quantified by qPCR with primer and probe combination AF-SSU (see 2.4 PCR and qPCR assays). AF 18S rDNA was detected in seven of the sampled biogas plants(Figure 1); data for biogas plant PB 25 was previously published(Dollhofer et al, 2016) but is presented again here for comparison. The three other sampled biogas plants (PB 15, PB 17 and PB 19) showed no evidence for the presence of AF.…”

mentioning

confidence: 68%

“…transcripts/ml and 1.8x10 2 GH5 transcripts/ml were found. Transcript numbers were thus lower than in the rumen of cattle, where 1.88x10 3 to 2.83x10 5 AF GH5 endoglucanase transcripts/ml were detected using the same method (Dollhofer et al, 2016).…”

Section: 6mentioning

confidence: 83%

“…Isolation of anaerobic fungi ( Figure 1). AF 18S rDNA gene copies were thus detected in most examined agricultural biogas plants, but at levels lower than in cattle rumen fluid (1.69x10 10 copies/ml; SD=3.88x10 9 ) and cattle slurries used as substrate in PB 14 (1.88x10 9 copies/ml; SD=3.3x10 8 ) and PB 22 (6x10 9 ; SD=1.16x10 9 copies/ml) (Dollhofer et al, 2016). An exception was the digester of PB 22 in which 1.65x10 9 copies/ml sludge were found, which is close to the values measured in cattle slurry.…”

Section: 6mentioning

confidence: 90%

“…One PCR and two qPCR assays, including standards for quantification, had been designed, optimized and validated in a previous study (Dollhofer et al 2016). Reagents from the Platinum ® Taq DNA…”

Section: Pcr and Qpcr Assaysmentioning

confidence: 99%

See 2 more Smart Citations

Presence and transcriptional activity of anaerobic fungi in agricultural biogas plants

Dollhofer¹,

Callaghan

Griffith

et al. 2017

Bioresource Technology

Self Cite

View full text Add to dashboard Cite

Bioaugmentation with anaerobic fungi (AF) is promising for improved biogas generation from lignocelluloses-rich substrates. However, before implementing AF into biogas processes it is necessary to investigate their natural occurrence, community structure and transcriptional activity in agricultural biogas plants. Thus, AF were detected with three specific PCR based methods: (i) Copies of their 18S genes were found in 7 of 10 biogas plants. (ii) Transcripts of a GH5 endoglucanase gene were present at low level in two digesters, indicating transcriptional cellulolytic activity of AF. (iii) Phylogeny of the AF-community was inferred with the 28S gene. A new Piromyces species was isolated from a PCR-positive digester. Evidence for AF was only found in biogas plants operated with high proportions of animal feces. Thus, AF were most likely transferred into digesters with animal derived substrates. Additionally, high process temperatures in combination with long retention times seemed to impede AF survival and activity.

show abstract

Biogas from Manure: The Future of Renewable Natural Gas and Its Implications

Nwuche¹,

Gupta²,

Akor³

et al. 2023

Climate Changes Mitigation and Sustainable Bioenergy Harvest Through Animal Waste

View full text Add to dashboard Cite

Development of three specific PCR-based tools to determine quantity, cellulolytic transcriptional activity and phylogeny of anaerobic fungi

Cited by 21 publications

References 47 publications

PCR and Omics Based Techniques to Study the Diversity, Ecology and Biology of Anaerobic Fungi: Insights, Challenges and Opportunities

PCR and Omics Based Techniques to Study the Diversity, Ecology and Biology of Anaerobic Fungi: Insights, Challenges and Opportunities

Presence and transcriptional activity of anaerobic fungi in agricultural biogas plants

Biogas from Manure: The Future of Renewable Natural Gas and Its Implications

Contact Info

Product

Resources

About