Seasonal total methane depletion in limestone caves

Methane concentration in caves is commonly much lower than the external atmosphere, yet the cave CH4 depletion causal mechanism is contested and dynamic links to external diurnal and seasonal temperature cycles unknown. Here, we report a continuous 3-year record of cave methane and other trace gases in Jenolan Caves, Australia which shows a seasonal cycle of extreme CH4 depletion, from ambient ~1,775 ppb to near zero during summer and to ~800 ppb in winter. Methanotrophic bacteria, some newly-discovered, rapidly consume methane on cave surfaces and in external karst soils with lifetimes in the cave of a few hours. Extreme bacterial selection due to the absence of alternate carbon sources for growth in the cave environment has resulted in an extremely high proportion 2–12% of methanotrophs in the total bacteria present. Unexpected seasonal bias in our cave CH4 depletion record is explained by a three-step process involving methanotrophy in aerobic karst soil above the cave, summer transport of soil-gas into the cave through epikarst, followed by further cave CH4 depletion. Disentangling cause and effect of cave gas variations by tracing sources and sinks has identified seasonal speleothem growth bias, with implied palaeo-climate record bias.

From laboratory measurements we find a proportionality between Rn and ion concentrations, 1 Bq m -3 ~ 10 ions cm -3 , from which typical total ions concentrations in Chifley cave are 10 2 -10 5 cm -3 . Fernandez-Cortes et al. found 4x10 4 -2x10 5 ions cm -3 in Ojo Guarena cave (ref 4). Only a small fraction of these ions are reactive primary ions such as N + , O + , N 2 + derived from N 2 and O 2 ; primary ions rapidly coagulate with water vapour and other trace gases to form complex ions which are much less reactive. For reactive primary ions the maximum possible second order rate constant 1 k is ~ 1x10 -9 cm -3 s -1 . Estimating at most 1% of total ions existing as reactive ions, a maximum value for k 1 is ~ 10 -9 * 10 3 = 10 -6 s -1 , corresponding to a minimum methane lifetime to ion-induced decay of 10 days. Thus the ion mechanism is too slow by at least an order of magnitude to account for the observed rate of methane loss in the cave with a lifetime of a few hours.
The CH 4 loss mechanism via OH • radicals produced from radon decay suggested by Fernandez-Cortes et al (ref 4) is also not kinetically feasible. The rate constant for the CH 4 + OH • reaction is ~ 6 x 10 -15 cm -3 s -1 at cave temperature 1 , requiring an OH • concentration of ~ 10 10 -10 11 cm -3 to explain the observed rate constant of 10 -4 s -1 . This is 10 6 times higher than mean daytime OH • concentrations in the atmosphere in sunlight. With the production rate of total HO x • = OH • +HO 2 • radicals from radon decay 2 of 4 x 10 5 Bq -1 , observed cave radon levels of ~ 1,000 Bq m -3 (10 -3 Bq cm -3 ), and a lifetime for OH • radicals 3 of ~1 s, the steady state total HO x • concentration from radon decay (only some of which is OH • ) would be of the order of 4 x 10 2 cm -3 , 8 orders of magnitude too low to explain the observed rate of CH 4 loss.

Assessment of potential mechanisms for seasonal CH 4 depletion pattern
In the absence of a significant seasonal temperature difference in Chifley Cave (Figure 2, Supplementary Figure 1) causing changes to methanotroph activity we consider other possible mechanisms to account for the strong seasonal CH 4 observed.

a Seasonal ventilation, -macro-convection
We ask, is a simple steady state model of in-situ methanotroph CH 4 depletion diluted with seasonally varied external air velocity compatible with observations? The simple answer is no, because convective air-flow through Chifley Cave is seasonally bi-directional with different sources. In winter Chifley Cave is diluted with external air alone, CH 4 is ~800 ppb. In summer soil air is also drawn into Chifley Cave to totally deplete methane.
Summer peak CO 2 observed in Lower Katies Bower, (~8,000 ppm, δ 13 C -24 ‰ PDB) matches soil derived microbial and root respiration labelled with high Rn (> 2,000 Bq m -3 ) and high N 2 O (> 1,000 ppb) indicating an exogenous karst soil origin for air in Lower Katies Bower. Winter air in LKB (CO 2 ~600 ppm) does not match the overlying winter soil CO 2 concentration, 3,000 -4,500 ppm indicating seasonally different sources of air in LKB caused by a sharp change in the dominant convective air-flow direction ( Figure 2). Any single day may have a temperature reversal between external and cave temperatures, causing air-flow reversal and different proportional mixing and residence time for gas source tracers. External soil CH 4 depletion to 600 -800 ppb, or exposure to cave surfaces (winter cave air CH 4 ~800 ppb) alone is insufficient to account for CH 4 approaching zero for the summer months ( Figure 2).

b Convective ventilation & measurement asymmetry causing seasonal path length bias
There is a bias in the cave path length and exposed surface area from the lower Grand Arch opening to LKB measurement point (116 m, 4,496 m 2 ) compared to the upper Elder Cave opening (268 m, 19,714 m 2 ) (Figure 1). Different path lengths and interactive cave surface area to the measurement point in LKB may partially explain seasonal CH 4 depletion pattern where the dominant seasonal convective air-flow is reversed. The magnitude of the path length bias (3:7) or cave surface area bias (2:8) produced by in-situ methanotrophy does not account for the extreme seasonal methane depletion pattern observed (> 99% summer, 55% winter) (Supplementary data Table 4).
c Cave micro-temperature environment Air path length bias may influence very minor diurnal anomalies to cave air temperature (< 1.5 °C) due to less areal interaction with the rock mass in winter. Summer air-flow in the opposite direction with a greater path length and time for thermal equilibration with the rock mass does not vary cave air temperature (Supplementary Figure 1). Thermal stratification of air in Katies Bower causes summer air flow to pass over the top of the chamber. Winter air-flow in the opposite direction may retain a small low temperature anomaly relative to the rock mass causing sinking into Lower Katies Bower, effectively flushing with external air. The greater residence time for in-situ methanotrophy in summer may also contribute to the pattern of extreme methane depletion.

d Methanotroph activity changes
The population of Type I methanotrophs was significantly higher in the summer months, suggesting that these may play a minor role in the complete disappearance of cave methane seen in this season.   Figure 2). External atmospheric temperature (gold) and soil temperature (red) shows strong diurnal and seasonal cycles. Precipitation (dark blue) short-term record over the preceding few weeks determines the soil water fraction. Soil CO 2 derived from plant root respiration and soil microbial activity is a proxy measure of primary biological productivity. Soil CO 2 concentration is sensitive to temperature and precipitation daily -weekly weather cycles. Figure 2b. Detailed soil function for 10 days in summer, December 2014. Soil CO 2 is measured by diffusion (IR sensor) and by low volume gas suction to a CRDS instrument. Gas extraction for measurement is replaced by ambient air causing a small offset. Soil CH 4 shows an inverse relationship with CO 2 and primary biological productivity. Soil CH 4 is low (~600 ppb) when methanotroph activity is high under warm moist conditions. Soil CO 2 δ 13 C -24 ‰ VPDB is invariant with respect to short-term weather conditions.     Table 3 Methane depletion rates in cave soils; rate of methane consumption from an initial 1,000 ppm incubated in closed vial (120 mL) with 5 g soil.  Table 4 Path length and area measured to either entrance.