An isolated chemolithoautotrophic ecosystem deduced from environmental isotopes: Ayyalon cave (Israel)

Amos Frumkin*, Ariel D. Chipman, Israel Naaman

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review


The stable isotopes composition of chemolithoautotrophic cave ecosystems is known to differ from epigenic caves. Here we show that in addition, dead carbon (devoid of 14C), is utilized and transferred throughout this ecosystem, rendering it unsuitable for radiocarbon dating. The connectivity of the Ayyalon Cave ecosystem with the surface is studied, along with its sources of energy and carbon, as well as the interconnections between its constituents. We use isotopic evidence to show that its ancient resilient ecosystem is based on an underground food web depending on rich biomass production by chemolithoautotrophic nutrient supplies, detached from surface photosynthesis. Carbon isotopic values indicate that: (1) the microbial biota use bicarbonate from the groundwater (23.34 pMC [% of modern carbon]) rather than the atmospheric CO2 above the water (71.36 pMC); (2) the depleted 14C signal is transferred through the entire ecosystem, indicating that the ecosystem is well-adapted and based on the cave biofilm which is in turn based on groundwater-dissolved inorganic carbon. Incubation of Ayyalon biofilm with 14C-labelled bicarbonate indicates uptake of the radio-labeled bicarbonate by sulfur-oxidizing proteobacteria Beggiatoa, suggesting that these sulfur-oxidizing microorganisms use the water-dissolved inorganic carbon for chemolithoautotrophic carbon fixation. Organic matter in the cave is much lighter in its stable nitrogen and carbon isotopes compared with respective surface values, as expected in chemolithoautotrophic systems. This evidence may be applicative to subsurface voids of ancient Earth environments and extraterrestrial systems.

Original languageAmerican English
Article number1040385
JournalFrontiers in Ecology and Evolution
StatePublished - 26 Jan 2023

Bibliographical note

Funding Information:
The Israel Hydrological Service and Israel Nature and Parks Authority financed the study (grant #0321869).

Funding Information:
We are grateful for the following assistance: Elisabetta Boaretto of the Kimmel Center for Archaeological Science, Weizmann Institute of Science, helped with the laboratory analysis of Ayyalon carbon isotopes. Chanan Dimentman of the Hebrew University separated faunal samples and sent them for definition. Aharon Oren of the Hebrew University helped with the incubation of Ayyalon’s biofilm with radio-labeled 14C bicarbonate. Boaz Luz and Eugeni Barkan helped with C and N isotopic analysis. Ronen Ron of the Hebrew University performed the statistical analysis. Oren Kolodny and Efrat Gavish-Regev of the Hebrew University, Serban Sarbu of California State University and Amitai Cooper of Intel provided useful advice.

Publisher Copyright:
Copyright © 2023 Frumkin, Chipman and Naaman.


  • arthropod adaptation
  • biofilm
  • cave ecosystem
  • chemolithoautotrophy
  • environmental isotopes
  • radiocarbon
  • stable isotopes
  • sulfidic cave


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