Evaluation of media and techniques to enumerate heterotrophic microbes from karst and sand aquifer springs

Abstract

Several media and techniques were compared for their efficiency to enumerate viable heterotrophs from both a karst and sand aquifer spring. A medium designed to enumerate bacteria from nutrient-poor waters (HCFU) as well as R2A medium proved superior to tryptic soy agar; however, the difference was always less than one order of magnitude. Membrane filtration resulted in lower counts of microbes than the spread plate, multitube turbidity, or drop plate methods from samples of both sand and karst springs. The drop plate technique yielded higher viable counts from the sand spring and basin of the karst spring, with a precision of 21% (coefficient of variation) and a maximum plating efficiency of 3.4% (viable count/direct count × 100). Subsequently, 63% of isolates from drop plates were recovered on HCFU. Microcolonies were visible by epifluorescence microscopy, acridine orange staining, and subsequent examination of excised agar sections containing drops.

The Relationship between pH and Bacterial Communities in a Single Karst Ecosystem and Its Implication for Soil Acidification

“…Although most cave systems are predominately oligotrophic habitats (Barton and Jurado, 2007), heterotrophs, which fed on organic matter, account for ≥75% of microbial communities in many caves (Mikell et al, 1996). Our results showed that the closest relatives associated with Planctomycetes (Fuerst, 1995), Verrucomicrobia (Hedlund et al, 1997), and Acidobacteria (Ward et al, 2009) were either chemoorganotrophs or aerobic heterotrophs.…”

Bacterial diversity and ecosystem function of filamentous microbial mats from aphotic (cave) sulfidic springs dominated by chemolithoautotrophic “Epsilonproteobacteria”

“…Previous studies have shown that microorganisms in caves associated with surface input are not chemolithoautotrophs, but instead are translocated soil heterotrophs, chemoorganotrophs, or fecal coliform bacteria from contaminated surface water [10]. Mikell et al [12] estimate that P75% of microbial communities in most caves are heterotrophs. While we recognize that in the past the Bighorn River near the cave entrance may have had a role in inoculating the cave with microorganisms during previous flood stages, we believe that the LKC microbial communities are endemic to the cave and unaffected by surface hydrologic conditions because (1) the filamentous microbial biomass in LKC is significantly higher than the 10 2 to 10 4 cells ml À1 commonly found in other aquatic cave systems [112], and (2) the discharging springs contribute little to no allochthonous DOC or particulate organic carbon to the microbial community (Table 1).…”

Sustained Anthropogenic Impact in Carter Saltpeter Cave, Carter County, Tennessee and the Potential Effects on Manganese Cycling

“…As a result, indirect methods of water monitoring for microbial contaminants are frequently employed by researchers: culture-based studies of water to detect coliform bacteria (Mikell et al, 1996;Rusterholtz and Mallory, 1994), molecular techniques to identify the presence/absence of fecal indicators and endemic species in karst systems (Ahmed et al, 2008;Johnson et al, 2011;Johnston et al, 2012;Porter, 2007;Roslev and Bukh, 2011), and the use of fluorescent dyetracers and microspheres to model water flow and pathogen dispersal in conduit systems (Goeppert and Goldscheider, 2011). Direct study of karst environments is restricted to open conduits that are large enough to allow for human movement and environmental manipulation.…”