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

Abstract: Observations and hypotheses on the possible influence of unidentified calcifying bacteria on moonmilk speleothem formation in the Grotta Nera are reported for the first time. The Majella Massif hosts a complex karst system of several caves; the accessible Grotta Nera is the most interesting one. Despite its name, the cave is characterized by particularly abundant ivory-white deposits of moonmilk. Two samples of moonmilk were analyzed to determine the geochemistry, fabric, depositional setting, and extent of biogenicity. For this, we combined geochemical, scanning electron microscopic, microbiological, and in vitro precipitation studies. X-ray diffraction of the moonmilk deposits gave clear evidence for the presence of calcite. Scanning electron microscopy showed that moonmilk in the Grotta Nera consists of a network of calcite fibers oriented in all directions, resembling a felted mat. The cultivation on specific medium of moonmilk and drip-water samples showed the presence of fungi, actinomycetes, and other bacteria, but the dominant cultivable microorganisms were bacteria, which produced significant crystallization. Examination of Gram-stained smears taken from the fifteen different colony types showed that the majority (66.7%) of the bacterial isolates were Gram-negative. Single small rods and rod chains were the most common bacteria isolated from the Grotta Nera. None of the molds isolated from the Grotta Nera samples were able to precipitate CaCO3 crystals, suggesting a major bacterial contribution to moonmilk deposition in the cave. Bacteria were capable of precipitating CaCO3 on B-4 solid medium at 15 (cave temperature), 22, and 32 uC. The calcifying bacteria isolated from the Grotta Nera showed a greater capability to solubilize CaCO3 than those associated with consolidated stalactites sampled from previously studied caves. The electron microscopy and microbiological evidences, together with the geochemistry and environmental data, allowed us to postulate the biogenic nature of the moonmilk in the Grotta Nera