Abstract
Biolog® EcoPlates™ (Biolog Inc., Hayward, CA, USA) were developed to analyse the functional diversity of bacterial communities by means of measuring their ability to oxidize carbon substrates. This technique has been successfully adopted for studying bacterial soil communities from different soil environments, polluted soils and soils subjected to various agronomic treatments. Unfortunately, Biolog® EcoPlates™ assay, especially working on soil, can be difficult to reproduce and hard to standardize due to the lack of detailed procedures and protocols. The main problems of this technique mainly regard soil preparation, bacterial inoculum densities and a correct definition of blank during the calculation of the diversity indices. On the basis of our previous research on agricultural soils, we here propose a standardized and accurate step-by-step method for estimating the functional diversity of a soil bacterial community by Biolog® EcoPlatesTM assay. A case study of soils sampled in a Mediterranean olive orchard managed accordingly to sustainable/conservation practices was reported for justifying the standardized method here used. The results of this methodological paper could be important for correctly evaluating and comparing the microbiological fertility of soils managed by sustainable/conservation or conventional/non-conservation systems.
Highlights
Microorganisms are present in all ecosystems and due to their rapid responses to physical and chemical changes, they can be used as bioindicators of environmental quality
In our case the dilutions examined were 10−5, corresponding to ~104 colony forming units (CFUs) mL−1 measured by plate counting, 10−4 and 10−6
It appears that the dilution with 10−6 dilution contained a relatively low number of bacteria (~103 CFUs mL−1 ) causing an increasing average well colour development (AWCD)
Summary
Microorganisms are present in all ecosystems and due to their rapid responses to physical and chemical changes, they can be used as bioindicators of environmental quality. Physiological Profiling (CLPP) is a rapid and relatively inexpensive technique to relate microbial functional diversity over space and time to changes in the environment [1,2,3,4]. Biolog® EcoPlatesTM (Biolog Inc., Hayward, CA, USA) were developed to analyse the functional diversity of bacterial communities by means of measuring their ability to oxidize carbon substrates. An EcoPlate is a 96-well microplate that contains 31 common carbon sources from altogether five compound groups—that is, carbohydrates, carboxylic and ketonic acids, amines and amides, amino acids and polymers—plus a blank well as a control, all these replicated thrice to control variation in inoculum densities. Each EcoPlate is filled with a dilution of one soil suspension, representing one soil sample. The turbidity of dilutions is due to clay and humic particles in soil colloidal suspension
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