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Evaluation of the Most Probable Number Method for Enumerating Denitrifying Bacteria¹

ABSTRACT

Most probable number (MPN) enumerations of denitrifying bacteria were conducted on an agricultural soil (pH 6.6) and a forest soil (pH 5.6), in both neutral (pH 6.8) and acid (pH 5.6) media. The neutral medium yielded higher enumerations for both soils and, therefore, appears to be optimal, even for denitrifiers present in acid soils. Enzyme activity assays and MPN enumerations of seven soil samples were weakly correlated (r = 0.71; p = 0.07). One soil exhibited a high MPN enumeration, but had no detectable denitrifying enzyme activity. While the enzyme assay indexes denitrification potential of soils under the environmental conditions at the time of sampling, the MPN enumeration indexes the denitrifying capability of the populations inhabiting soils. A population's denitrifying capability is related to its size, but other factors which might also influence MPN enumerations are discussed. Precise incubation time was critical in the MPN procedure. Delays of 2h affected the scoring results. Populations in some of the incubation tubes appeared to be in a dynamic growth phase at the end of the 14-d incubation period. Inconsistencies in incubation duration could confound comparisons among soils. Despite inherent flaws and disadvantages, the MPN procedure may be appropriate for certain research objectives as an index of a population's denitrifying capacity. Recommendations and cautions are offered.

¹ Contribution from the Dep. of Forestry, North Carolina State Univ., Raleigh, NC 27695-8002. Paper no. 9415 of the journal series of the North Carolina Agric. Res. Serv., Raleigh, NC 27695-7601.

² Graduate Research Fellow, Dep. of Forestry, North Carolina State Univ., Raleigh.

³ Undergraduates, Dep. of Biology, Oberlin College, Oberlin, OH.

Received for publication July 25, 1984. Accepted for publication October 31, 1984.