Towards optimized viral metagenomes for double-stranded and single-stranded DNA viruses from challenging soils
Soils impact global carbon cycling and their resident microbes are critical to their biogeochemical processing and ecosystem outputs. Based on studies in marine systems, viruses infecting soil microbes likely modulate host activities via mortality, horizontal gene transfer, and metabolic control. Ho...
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| Format: | Article |
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PeerJ Inc.
2019-07-01
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| Online Access: | https://peerj.com/articles/7265.pdf |
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| author | Gareth Trubl Simon Roux Natalie Solonenko Yueh-Fen Li Benjamin Bolduc Josué Rodríguez-Ramos Emiley A. Eloe-Fadrosh Virginia I. Rich Matthew B. Sullivan |
| author_facet | Gareth Trubl Simon Roux Natalie Solonenko Yueh-Fen Li Benjamin Bolduc Josué Rodríguez-Ramos Emiley A. Eloe-Fadrosh Virginia I. Rich Matthew B. Sullivan |
| author_sort | Gareth Trubl |
| collection | DOAJ |
| description | Soils impact global carbon cycling and their resident microbes are critical to their biogeochemical processing and ecosystem outputs. Based on studies in marine systems, viruses infecting soil microbes likely modulate host activities via mortality, horizontal gene transfer, and metabolic control. However, their roles remain largely unexplored due to technical challenges with separating, isolating, and extracting DNA from viruses in soils. Some of these challenges have been overcome by using whole genome amplification methods and while these have allowed insights into the identities of soil viruses and their genomes, their inherit biases have prevented meaningful ecological interpretations. Here we experimentally optimized steps for generating quantitatively-amplified viral metagenomes to better capture both ssDNA and dsDNA viruses across three distinct soil habitats along a permafrost thaw gradient. First, we assessed differing DNA extraction methods (PowerSoil, Wizard mini columns, and cetyl trimethylammonium bromide) for quantity and quality of viral DNA. This established PowerSoil as best for yield and quality of DNA from our samples, though ∼1/3 of the viral populations captured by each extraction kit were unique, suggesting appreciable differential biases among DNA extraction kits. Second, we evaluated the impact of purifying viral particles after resuspension (by cesium chloride gradients; CsCl) and of viral lysis method (heat vs bead-beating) on the resultant viromes. DNA yields after CsCl particle-purification were largely non-detectable, while unpurified samples yielded 1–2-fold more DNA after lysis by heat than by bead-beating. Virome quality was assessed by the number and size of metagenome-assembled viral contigs, which showed no increase after CsCl-purification, but did from heat lysis relative to bead-beating. We also evaluated sample preparation protocols for ssDNA virus recovery. In both CsCl-purified and non-purified samples, ssDNA viruses were successfully recovered by using the Accel-NGS 1S Plus Library Kit. While ssDNA viruses were identified in all three soil types, none were identified in the samples that used bead-beating, suggesting this lysis method may impact recovery. Further, 13 ssDNA vOTUs were identified compared to 582 dsDNA vOTUs, and the ssDNA vOTUs only accounted for ∼4% of the assembled reads, implying dsDNA viruses were dominant in these samples. This optimized approach was combined with the previously published viral resuspension protocol into a sample-to-virome protocol for soils now available at protocols.io, where community feedback creates ‘living’ protocols. This collective approach will be particularly valuable given the high physicochemical variability of soils, which will may require considerable soil type-specific optimization. This optimized protocol provides a starting place for developing quantitatively-amplified viromic datasets and will help enable viral ecogenomic studies on organic-rich soils. |
| first_indexed | 2024-03-09T06:54:56Z |
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| id | doaj.art-36ba41b4e0d44090970f5e501b79f697 |
| institution | Directory Open Access Journal |
| issn | 2167-8359 |
| language | English |
| last_indexed | 2024-03-09T06:54:56Z |
| publishDate | 2019-07-01 |
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| spelling | doaj.art-36ba41b4e0d44090970f5e501b79f6972023-12-03T10:05:01ZengPeerJ Inc.PeerJ2167-83592019-07-017e726510.7717/peerj.7265Towards optimized viral metagenomes for double-stranded and single-stranded DNA viruses from challenging soilsGareth Trubl0Simon Roux1Natalie Solonenko2Yueh-Fen Li3Benjamin Bolduc4Josué Rodríguez-Ramos5Emiley A. Eloe-Fadrosh6Virginia I. Rich7Matthew B. Sullivan8Department of Microbiology, The Ohio State University, Columbus, OH, United States of AmericaUnited States Department of Energy Joint Genome Institute, Lawrence Berkeley National Laboratory, Walnut Creek, CA, United States of AmericaDepartment of Microbiology, The Ohio State University, Columbus, OH, United States of AmericaDepartment of Microbiology, The Ohio State University, Columbus, OH, United States of AmericaDepartment of Microbiology, The Ohio State University, Columbus, OH, United States of AmericaDepartment of Microbiology, The Ohio State University, Columbus, OH, United States of AmericaUnited States Department of Energy Joint Genome Institute, Lawrence Berkeley National Laboratory, Walnut Creek, CA, United States of AmericaDepartment of Microbiology, The Ohio State University, Columbus, OH, United States of AmericaDepartment of Microbiology, The Ohio State University, Columbus, OH, United States of AmericaSoils impact global carbon cycling and their resident microbes are critical to their biogeochemical processing and ecosystem outputs. Based on studies in marine systems, viruses infecting soil microbes likely modulate host activities via mortality, horizontal gene transfer, and metabolic control. However, their roles remain largely unexplored due to technical challenges with separating, isolating, and extracting DNA from viruses in soils. Some of these challenges have been overcome by using whole genome amplification methods and while these have allowed insights into the identities of soil viruses and their genomes, their inherit biases have prevented meaningful ecological interpretations. Here we experimentally optimized steps for generating quantitatively-amplified viral metagenomes to better capture both ssDNA and dsDNA viruses across three distinct soil habitats along a permafrost thaw gradient. First, we assessed differing DNA extraction methods (PowerSoil, Wizard mini columns, and cetyl trimethylammonium bromide) for quantity and quality of viral DNA. This established PowerSoil as best for yield and quality of DNA from our samples, though ∼1/3 of the viral populations captured by each extraction kit were unique, suggesting appreciable differential biases among DNA extraction kits. Second, we evaluated the impact of purifying viral particles after resuspension (by cesium chloride gradients; CsCl) and of viral lysis method (heat vs bead-beating) on the resultant viromes. DNA yields after CsCl particle-purification were largely non-detectable, while unpurified samples yielded 1–2-fold more DNA after lysis by heat than by bead-beating. Virome quality was assessed by the number and size of metagenome-assembled viral contigs, which showed no increase after CsCl-purification, but did from heat lysis relative to bead-beating. We also evaluated sample preparation protocols for ssDNA virus recovery. In both CsCl-purified and non-purified samples, ssDNA viruses were successfully recovered by using the Accel-NGS 1S Plus Library Kit. While ssDNA viruses were identified in all three soil types, none were identified in the samples that used bead-beating, suggesting this lysis method may impact recovery. Further, 13 ssDNA vOTUs were identified compared to 582 dsDNA vOTUs, and the ssDNA vOTUs only accounted for ∼4% of the assembled reads, implying dsDNA viruses were dominant in these samples. This optimized approach was combined with the previously published viral resuspension protocol into a sample-to-virome protocol for soils now available at protocols.io, where community feedback creates ‘living’ protocols. This collective approach will be particularly valuable given the high physicochemical variability of soils, which will may require considerable soil type-specific optimization. This optimized protocol provides a starting place for developing quantitatively-amplified viromic datasets and will help enable viral ecogenomic studies on organic-rich soils.https://peerj.com/articles/7265.pdfSoil virusesViromesDNA extractionOrganicsMicrobiologyssDNA viruses |
| spellingShingle | Gareth Trubl Simon Roux Natalie Solonenko Yueh-Fen Li Benjamin Bolduc Josué Rodríguez-Ramos Emiley A. Eloe-Fadrosh Virginia I. Rich Matthew B. Sullivan Towards optimized viral metagenomes for double-stranded and single-stranded DNA viruses from challenging soils PeerJ Soil viruses Viromes DNA extraction Organics Microbiology ssDNA viruses |
| title | Towards optimized viral metagenomes for double-stranded and single-stranded DNA viruses from challenging soils |
| title_full | Towards optimized viral metagenomes for double-stranded and single-stranded DNA viruses from challenging soils |
| title_fullStr | Towards optimized viral metagenomes for double-stranded and single-stranded DNA viruses from challenging soils |
| title_full_unstemmed | Towards optimized viral metagenomes for double-stranded and single-stranded DNA viruses from challenging soils |
| title_short | Towards optimized viral metagenomes for double-stranded and single-stranded DNA viruses from challenging soils |
| title_sort | towards optimized viral metagenomes for double stranded and single stranded dna viruses from challenging soils |
| topic | Soil viruses Viromes DNA extraction Organics Microbiology ssDNA viruses |
| url | https://peerj.com/articles/7265.pdf |
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