What's new is old: resolving the identity of Leptothrix ochracea using single cell genomics, pyrosequencing and FISH.
Leptothrix ochracea is a common inhabitant of freshwater iron seeps and iron-rich wetlands. Its defining characteristic is copious production of extracellular sheaths encrusted with iron oxyhydroxides. Surprisingly, over 90% of these sheaths are empty, hence, what appears to be an abundant populatio...
| Main Authors: | , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Public Library of Science (PLoS)
2011-03-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC3060100?pdf=render |
| _version_ | 1818500150428958720 |
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| author | Emily J Fleming Amy E Langdon Manuel Martinez-Garcia Ramunas Stepanauskas Nicole J Poulton E Dashiell P Masland David Emerson |
| author_facet | Emily J Fleming Amy E Langdon Manuel Martinez-Garcia Ramunas Stepanauskas Nicole J Poulton E Dashiell P Masland David Emerson |
| author_sort | Emily J Fleming |
| collection | DOAJ |
| description | Leptothrix ochracea is a common inhabitant of freshwater iron seeps and iron-rich wetlands. Its defining characteristic is copious production of extracellular sheaths encrusted with iron oxyhydroxides. Surprisingly, over 90% of these sheaths are empty, hence, what appears to be an abundant population of iron-oxidizing bacteria, consists of relatively few cells. Because L. ochracea has proven difficult to cultivate, its identification is based solely on habitat preference and morphology. We utilized cultivation-independent techniques to resolve this long-standing enigma. By selecting the actively growing edge of a Leptothrix-containing iron mat, a conventional SSU rRNA gene clone library was obtained that had 29 clones (42% of the total library) related to the Leptothrix/Sphaerotilus group (≤96% identical to cultured representatives). A pyrotagged library of the V4 hypervariable region constructed from the bulk mat showed that 7.2% of the total sequences also belonged to the Leptothrix/Sphaerotilus group. Sorting of individual L. ochracea sheaths, followed by whole genome amplification (WGA) and PCR identified a SSU rRNA sequence that clustered closely with the putative Leptothrix clones and pyrotags. Using these data, a fluorescence in-situ hybridization (FISH) probe, Lepto175, was designed that bound to ensheathed cells. Quantitative use of this probe demonstrated that up to 35% of microbial cells in an actively accreting iron mat were L. ochracea. The SSU rRNA gene of L. ochracea shares 96% homology with its closet cultivated relative, L. cholodnii, This establishes that L. ochracea is indeed related to this group of morphologically similar, filamentous, sheathed microorganisms. |
| first_indexed | 2024-12-10T20:38:59Z |
| format | Article |
| id | doaj.art-4545b2c8ec1a4efcb0bdeb2fe203a141 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-10T20:38:59Z |
| publishDate | 2011-03-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-4545b2c8ec1a4efcb0bdeb2fe203a1412022-12-22T01:34:27ZengPublic Library of Science (PLoS)PLoS ONE1932-62032011-03-0163e1776910.1371/journal.pone.0017769What's new is old: resolving the identity of Leptothrix ochracea using single cell genomics, pyrosequencing and FISH.Emily J FlemingAmy E LangdonManuel Martinez-GarciaRamunas StepanauskasNicole J PoultonE Dashiell P MaslandDavid EmersonLeptothrix ochracea is a common inhabitant of freshwater iron seeps and iron-rich wetlands. Its defining characteristic is copious production of extracellular sheaths encrusted with iron oxyhydroxides. Surprisingly, over 90% of these sheaths are empty, hence, what appears to be an abundant population of iron-oxidizing bacteria, consists of relatively few cells. Because L. ochracea has proven difficult to cultivate, its identification is based solely on habitat preference and morphology. We utilized cultivation-independent techniques to resolve this long-standing enigma. By selecting the actively growing edge of a Leptothrix-containing iron mat, a conventional SSU rRNA gene clone library was obtained that had 29 clones (42% of the total library) related to the Leptothrix/Sphaerotilus group (≤96% identical to cultured representatives). A pyrotagged library of the V4 hypervariable region constructed from the bulk mat showed that 7.2% of the total sequences also belonged to the Leptothrix/Sphaerotilus group. Sorting of individual L. ochracea sheaths, followed by whole genome amplification (WGA) and PCR identified a SSU rRNA sequence that clustered closely with the putative Leptothrix clones and pyrotags. Using these data, a fluorescence in-situ hybridization (FISH) probe, Lepto175, was designed that bound to ensheathed cells. Quantitative use of this probe demonstrated that up to 35% of microbial cells in an actively accreting iron mat were L. ochracea. The SSU rRNA gene of L. ochracea shares 96% homology with its closet cultivated relative, L. cholodnii, This establishes that L. ochracea is indeed related to this group of morphologically similar, filamentous, sheathed microorganisms.http://europepmc.org/articles/PMC3060100?pdf=render |
| spellingShingle | Emily J Fleming Amy E Langdon Manuel Martinez-Garcia Ramunas Stepanauskas Nicole J Poulton E Dashiell P Masland David Emerson What's new is old: resolving the identity of Leptothrix ochracea using single cell genomics, pyrosequencing and FISH. PLoS ONE |
| title | What's new is old: resolving the identity of Leptothrix ochracea using single cell genomics, pyrosequencing and FISH. |
| title_full | What's new is old: resolving the identity of Leptothrix ochracea using single cell genomics, pyrosequencing and FISH. |
| title_fullStr | What's new is old: resolving the identity of Leptothrix ochracea using single cell genomics, pyrosequencing and FISH. |
| title_full_unstemmed | What's new is old: resolving the identity of Leptothrix ochracea using single cell genomics, pyrosequencing and FISH. |
| title_short | What's new is old: resolving the identity of Leptothrix ochracea using single cell genomics, pyrosequencing and FISH. |
| title_sort | what s new is old resolving the identity of leptothrix ochracea using single cell genomics pyrosequencing and fish |
| url | http://europepmc.org/articles/PMC3060100?pdf=render |
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