NASA Astrobiology Institute (NAI)

1. Genomics of sulfidic cave extremophiles (Supplement to NNA04CC06A)

   Project Investigators: Jennifer Macalady

   Other Project Members

   Sharmishtha Dattagupta (Research Staff)

   Katherine Dawson (Doctoral Student)

   Daniel Jones (Doctoral Student)

   Irene Schaperdoth (Research Staff)

   Stephanie Spielman (Undergraduate Student)

   Jignasha Patel (Undergraduate Student)

   Alessandro Montanari (Collaborator)

   Rebecca McCauley (Doctoral Student)

   Summary

   We investigated the ecology and evolutionary relationships among extremely acid-loving bacteria and archaea living in biofilms called “snottites” in sulfidic caves in Italy and Mexico. The acid-loving microbes form the base of food chains cut off from the surface, and present rare examples of microbially dominated ecosystems (like ecosystems present for much of earth history and those potentially elsewhere in the universe). The snottites are also important because they help us learn how life adapts to environmental conditions much different from the ones that can be tolerated by our own species (pH 0-1). Future work based on the foundation presented here will reveal how subsurface microorganisms in geographically isolated “geochemical islands” are related to each other and to microorganisms living at the earth’s surface.

   Astrobiology Roadmap Objectives:

   • Objective 5.1: Environment-dependent, molecular evolution in microorganisms
   • Objective 5.3: Biochemical adaptation to extreme environments
   • Objective 6.2: Adaptation and evolution of life beyond Earth

   Project Progress

   PSARC funds fully or partially supported the publication of 4 manuscripts and 6 abstracts last year. Field expeditions allowed us to make geochemical measurements and sample subsurface microbial communities in sulfidic caves hosting extreme acidophiles (snottites), including the Frasassi cave system (13°C) and Grotta Nuova del Rio Garaffo (45°C) in Italy.

   
   

   Doctoral student Dan Jones measures sulfide fluxes reaching extreme acidophile (snottite) communities in the Frasassi cave system (Italy).

   
   

   Doctoral student Dan Jones and postdoc Sharmishtha Dattagupta measure gas concentrations at high-temperature snottite sample location in Grotta Nuova di Rio Garrafo (Italy).

   We used a combination of metagenomic, phylogenetic, lipid, and culture-based analyses to investigate the ecology, biogeochemistry, and biosignatures of the extreme acidophile (pH 0-2) communities. 16S rDNA clone libraries contained at most two archaeal and six bacterial species. Fluorescence in situ hybridization (FISH) of snottites from over 12 cave locations confirmed that snottites are among the simplest microbial communities known, and that they are dominated by Acidithiobacillus and Ferroplasma species, with smaller populations of Acidimicrobium species and other bacteria, filamentous fungi, and protists. As described previously, nearly complete genomic coverage of an Acidithiobacillus strain was obtained using a metagenomics approach (work of Ph. D. student Dan Jones, postdoc Dattagupta, technician Schaperdoth).

   
   

   Metabolic genes relevant to snottite geochemistry and microbiology retrieved in metagenomic data (Jones et al., in preparation).

   
   

   Comparative analysis of snottite community composition using metagenomic, FISH, and 16S rDNA cloning datasets (Jones et al., in preparation).

   Acidithiobacillus strains isolated from cave samples on autotrophic media confirm that they are primary producers and that they excrete abundant extracellular polymers similar to those in snottite matrices. Lipid analyses (work of Ph.D. students Heidi Albrecht and Kat Dawson) revealed that snottite Acidithiobacillus isolates are capable of synthesizing up to 10 or more membrane hopanoids, an unusually high number compared to the 0-2 hopanoid structures produced by most other bacteria. We hypothesize that the membrane stiffening hopanoid compounds are one of the important mechanisms of acid adaptation in this bacterium. We were also able to enrich extremely acidophilic organoheterotrophs from the snottites (work of undergraduate Jignasha Patel and astrobiology REU student Stephanie Spielman). The cave acidophile project was highlighted in an interview for MicrobeWorld Radio (American Society for Microbiology).

   A newly discovered (non-acidophile) microbial community was obtained from deep within the Frasassi cave system in the anoxic water of a stratified lake. This community is of strong interest due to the potentially extreme energy limitation of the environment, and because of the unusual form of the biofilms (thick vertical “ropes” several meters long). 14-carbon dating of the biofilm suggests that it is extremely slow growing and may be thousands of years old. Archaea retrieved in 16S rDNA clone libraries are not related to cultivated organisms and have unknown metabolic capabilities. The presence of mcrA-related genes amplified from the biofilm suggests that methane cycling may be a source of usable energy, but neither mcrA nor 16S rDNA sequences show close relationships with known methanogens or anaerobic methane oxidizers.

   Field Expeditions

   Name

   Central Italy Sulfide Caves

   Dates

   May 2008 - June 2008

   Location

   43°23'51 12°57'2

   Description

   Geochemical measurements, sample collection of extremely acidic snottite biofilms for genomic analysis; exploration.

   Cross-Team Collaborations

   Nothing formal. We periodically exchange information with scientists in the Spanish Astrobiology Center working on extreme acidophiles in the Rio Tinto (Ricardo Amils).

Publications

Dattagupta, S., Schaperdoth, I., Montanari, A., Mariani, S., Kita, N., Valley, J.W. & Macalady, J.L.  (2008).  A Recently Evolved Symbiosis Between Chemoautotrophic Bacteria and a Cave-dwelling Amphipod.  Science, submitted.

Ewing, S.A., McKay, C.P., Warren-Rhodes, K., Macalady, J.L. & Amundson, R.  (2008).  Changes in the soil C cycle at the arid-hyperarid transition in the Atacama Desert.  Journal of Geophysical Research-Biogeosciences, in press.

Macalady, J.L., Dattagupta, S., Schaperdoth, I., Jones, D.S., Druschel, G.K. & Eastman, D.  (2008).  Niche differentiation among sulfur-oxidizing bacterial populations in cave waters.  ISME J, 2(6):509-601  [Online].

Strapoc, D., Picardal, F.W., Turich, C., Schaperdoth, I., Macalady, J.L., Lipp, J.S., Lin, Y.-S., Ertefai, T.F., Schubotz, F., Hinrichs, K.-U., Mastalerz, M. & Schimmelmann, A.  (2008).  Methane-Producing Microbial Community in a Coal Bed of the Illinois Basin.  Appl. Environ. Microbiol., 74(8):2424  [Online].