Microbiological activities in moonmilk monitored using isothermal microcalorimetry (Cave of Vers chez le Brandt, Neuchatel, Switzerland)

Abstract

Studies of the influence of microbial communities on calcium carbonate deposits mostly rely on classical or molecular microbiology, isotopic analyses, and microscopy. Using these techniques, it is difficult to infer microbial activities in such deposits. In this context, we used isothermal microcalorimetry, a sensitive and nondestructive tool, to measure microbial activities associated with moonmilk ex-situ. Upon the addition of diluted LB medium and other carbon sources to fresh moonmilk samples, we estimated the number of colony forming units per gram of moonmilk to be 4.8 3 10 5 6 0.2 3 10 5 . This number was close to the classical plate counts, but one cannot assume that all active cells producing metabolic heat were culturable. Using a similar approach, we estimated the overall growth rate and generation time of the microbial community associated with the moonmilk upon addition of various carbon sources. The range of apparent growth rates of the chemoheterotrophic microbial community observed was between 0.025 and 0.067 h 21 and generation times were between 10 and 27 hours. The highest growth rates were observed for citrate and diluted LB medium, while the highest carbon-source consumption rates were observed for low molecular weight organic acids (oxalate and acetate) and glycerol. Considering the rapid degradation of organic acids, glucose, and other carbon sources observed in the moonmilk, it is obvious that upon addition of nutrients during snow melting or rainfall these communities can have high overall activities comparable to those observed in some soils. Such communities can influence the physico-chemical conditions and participate directly or indirectly to the formation of moonmilk.

Microcalorimetric assays for measuring cell growth and metabolic activity: Methodology and applications

“…In solid medium or in solid substrates, the peaks in the thermal power curve (i.e., the metabolic activity) tend to become smaller but broader with decreasing inoculum (or bacterial density). This has been observed in agar medium, fermented milk products, and speleothems (i.e., secondary mineral deposit formed in a cave) for example [67][68][69]. This phenomenon can easily be explained by the limitation in substrates availability introduced by the medium.…”

“…In soil, one might add a specific substrate such as oxalate [64] to measure the activity of oxalate degrading bacteria. In speleothems, antibiotics have been used to inhibit bacterial activity and investigate fungal activity instead [69], however the effect is often limited in time and some groups of bacteria might be rather resistant to specific antibiotics. In sediment, sodium molybdate and bromoethanesulfonic acid (BES) are commonly used to inhibit sulfate reduction and trimethylamine is used as a specific substrate for methanogenesis [106].…”

Magnetic fingerprint of southern Portuguese speleothems and implications for paleomagnetism and environmental magnetism

“…Both aquifers are essentially developed in Jurassic carbonates and are separated by~40 km (Figure 1a). Weathering in the SPA stalactite is visible at the naked eye by the presence of moonmilk, a disintegration product of calcite that is generally interpreted to result from microbial activity (Figures 1c and 1d) [Barton and Northup, 2007;Braissant et al, 2012;Canaveras et al, 2006;Cañaveras et al, 1999]. We also collected cave sediments (samples SG1) and terra rossa soils, developed on Jurassic limestones and located above the Excentricas cave (samples TR_1, TR_2, and TR_3; Figure 1a).…”

“…Under SEM microscope, these altered areas are consisting of spiky calcite (a destructive fabric formed by etching) and microcrystalline calcite partially associated to biocrystalline assemblages constituted of anhedral calcite crystals and needle-fiber calcite . Spiky calcite and needle-fiber calcite are common in moonmilk deposits and are generally interpreted to result from microbial activity [Barton and Northup, 2007;Braissant et al, 2012;Canaveras et al, 2006;Cañaveras et al, 2001Cañaveras et al, , 1999.…”

Biogenicity and Characterization of Moonmilk in the Grotta Nera (Majella National Park, Abruzzi, Central Italy)

“…Biotic and abiotic processes have been postulated (Gradziñ sky et al, 1997;Borsato et al, 2000;Forti, 2001;Northup and Lavoie, 2001). Some evidences of microbial activities related with calcite moonmilk deposits have been reported (Bertouille, 1972;James et al, 1982;Callot et al, 1985;Gradziñ ski et al, 1997;Barton and Northup, 2007;Braissant et al, 2012). Moonmilk is the cave deposit most commonly associated with biogenic calcite precipitation, either by direct precipitation by microorganisms (fungi, algae, bacteria, and archaea) (Castanier et al, 1999;Barton and Northup, 2007;Ercole et al, 2012) or by passive precipitation in which microorganisms themselves act as nucleation surfaces on which minerals precipitate (Jones and Kahle, 1993;Blyth and Frisia, 2008).…”