PMID- 29773905
OWN - NLM
STAT- MEDLINE
DCOM- 20190415
LR  - 20190415
IS  - 1750-2799 (Electronic)
IS  - 1750-2799 (Linking)
VI  - 13
IP  - 6
DP  - 2018 Jun
TI  - Establishing anaerobic hydrocarbon-degrading enrichment cultures of 
      microorganisms under strictly anoxic conditions.
PG  - 1310-1330
LID - 10.1038/nprot.2018.030 [doi]
AB  - Traditionally, the description of microorganisms starts with their isolation from 
      an environmental sample. Many environmentally relevant anaerobic microorganisms 
      grow very slowly, and often they rely on syntrophic interactions with other 
      microorganisms. This impedes their isolation and characterization by classic 
      microbiological techniques. We developed and applied an approach for the 
      successive enrichment of syntrophic hydrocarbon-degrading microorganisms from 
      environmental samples. We collected samples from microbial mat-covered 
      hydrothermally heated hydrocarbon-rich sediments of the Guaymas Basin and mixed 
      them with synthetic mineral medium to obtain sediment slurries. Supplementation 
      with defined substrates (i.e., methane or butane), incubation at specific 
      temperatures, and a regular maintenance procedure that included the measurement 
      of metabolic products and stepwise dilutions enabled us to establish highly 
      active, virtually sediment-free enrichment cultures of actively 
      hydrocarbon-degrading communities in a 6-months to several-years' effort. Using 
      methane as sole electron donor shifted the originally highly diverse microbial 
      communities toward defined mixed cultures dominated by syntrophic consortia 
      consisting of anaerobic methane-oxidizing archaea (ANME) and different 
      sulfate-reducing bacteria. Cultivation with butane at 50  degrees C yielded consortia of 
      archaea belonging to Candidatus Syntrophoarchaeum and Candidatus Desulfofervidus 
      auxilii partner bacteria. This protocol also describes sampling for further 
      molecular characterization of enrichment cultures by fluorescence in situ 
      hybridization (FISH), and transcriptomics and metabolite analyses, which can 
      provide insights into the functioning of hydrocarbon metabolism in archaea and 
      resolve important mechanisms that enable electron transfer to their 
      sulfate-reducing partner bacteria.
FAU - Laso-Perez, Rafael
AU  - Laso-Perez R
AUID- ORCID: 0000-0002-6912-7865
AD  - Max-Planck Institute for Marine Microbiology, Bremen, Germany.
AD  - Alfred Wegener Institute, Helmholtz Centre for Polar and Marine Research, 
      Bremerhaven, Germany.
FAU - Krukenberg, Viola
AU  - Krukenberg V
AUID- ORCID: 0000-0001-8369-8114
AD  - Max-Planck Institute for Marine Microbiology, Bremen, Germany.
AD  - Alfred Wegener Institute, Helmholtz Centre for Polar and Marine Research, 
      Bremerhaven, Germany.
FAU - Musat, Florin
AU  - Musat F
AUID- ORCID: 0000-0002-4240-3495
AD  - Helmholtz Centre for Environmental Research - UFZ, Leipzig, Germany.
FAU - Wegener, Gunter
AU  - Wegener G
AUID- ORCID: 0000-0002-6819-373X
AD  - Max-Planck Institute for Marine Microbiology, Bremen, Germany.
AD  - Alfred Wegener Institute, Helmholtz Centre for Polar and Marine Research, 
      Bremerhaven, Germany.
AD  - MARUM, Center for Marine Environmental Sciences, University of Bremen, Bremen, 
      Germany.
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, Non-P.H.S.
DEP - 20180517
PL  - England
TA  - Nat Protoc
JT  - Nature protocols
JID - 101284307
RN  - 0 (Hydrocarbons)
SB  - IM
MH  - Anaerobiosis
MH  - Archaea/growth & development/*metabolism
MH  - Bacteria, Anaerobic/growth & development/*metabolism
MH  - Biotransformation
MH  - *Environmental Microbiology
MH  - Hydrocarbons/*metabolism
MH  - *Microbial Consortia
MH  - Microbiological Techniques/*methods
MH  - Temperature
EDAT- 2018/05/19 06:00
MHDA- 2019/04/16 06:00
CRDT- 2018/05/19 06:00
PHST- 2018/05/19 06:00 [entrez]
PHST- 2018/05/19 06:00 [pubmed]
PHST- 2019/04/16 06:00 [medline]
AID - nprot.2018.030 [pii]
AID - 10.1038/nprot.2018.030 [doi]
PST - ppublish
SO  - Nat Protoc. 2018 Jun;13(6):1310-1330. doi: 10.1038/nprot.2018.030. Epub 2018 May 
      17.