Isolation of mycobacteria from acidic forest soil samples: comparison of culture methods

Abstract

To evaluate which combination of decontamination method and medium is most reliable when examining acidic, organic forest soils for mycobacteria, three decontamination methods and five media supplemented with cycloheximide were compared.Before decontamination, the samples were incubated at 37°C for 5 h to allow germination of microbial spores. T h e recovery of mycobacteria was significantly influenced both by the method and by medium. Decontamination with N a O H or H2S04 both combined with malachite green and cycloheximide yielded higher viable counts of mycobacteria than decontamination with N a O H followed by oxalic acid. Egg media at p H 5.5 resulted in lower mycobacterial counts than egg media at p H 6.5 or Mycobacteria 7H11 agar. T h e numbers of slopes totally free of contaminants revealed Mycobacteria 7H11 agar medium to be more prone to contamination than the four egg media tested. T h e highest counts of mycobacteria and a low rate of contamination were obtained when decontamination with NaOH-malachite green-cycloheximide was combined with culture on glycerol and cycloheximide supplemented egg medium at p H 6.5.

A longitudinal study of environmental mycobacteria on a farm in south-west England

“…1984a; Portaels et nl. 1988;Iivanainen 1995). In the present study, the methods recommended by Portaels et nl.…”

Influence of Environmental Gradients on the Abundance and Distribution of Mycobacterium spp. in a Coastal Lagoon Estuary

“…Therefore, the findings of this study provide a greater understanding of how human activities influence coastal ecosystem Of the few efforts to examine environmental mycobacterial ecology, this is the first to rely strictly on a molecular approach (1,20,21,43). It is well known that mycobacterial culture is not optimal for all species and that decontamination procedures required to accommodate slow-growing species from environmental samples can dramatically reduce the abundance and diversity of what is cultured (18,19). Molecular techniques such as qPCR have become very common in studies of the diversity and abundance of aquatic bacterioplankton (5,9,15,38,40).…”

“…Because of the slow growth of many mycobacteria, culture from environmental samples requires decontamination, which can severely impact both the quantity and diversity of species recovered (18,19). Recently, quantitative PCR (qPCR) has gained favor as a means of rapidly enumerating organisms or genes in environmental samples (5,15,38,40).…”

Tetracycline Resistance and Presence of Tetracycline Resistance Determinants <i>tet</i>(V) and <i>tap</i> in Rapidly Growing Mycobacteria from Agricultural Soils and Clinical Isolates

“…Soils were kept at 4°C during transport to the laboratory and prior to the accompanying physicochemical and microbial analyses (Table S1). RGM were isolated from soil using the NaOH/malachite green/cycloheximide decontamination method of Iivanainen (27) or the olive oil/SDS decontamination method (with 10 mg SDS per plate) of Yamamura and Harayama (53) or directly on Tryptic-Soy agar plates with 25 mg L −1 chlortetracycline (2 isolates from 2007) (Table S2). …”