Development of a New Most-probable-number Method for Enumerating Methanotrophs, Using 48-well Microtiter Plates

Abstract

We developed a new most-probable-number (MPN) method for enumeration of methanotrophs, using 48-well microtiter plates. With this method, a microtiter plate which has been inoculated with serial dilutions of a sample is placed in a plastic desiccator containing methane and shaken. The proposed method gave estimates of population densities of a pure culture of Methylocystis sp. strain M and of methanotrophs in a paddy soil sample similar to those obtained by a conventional MPN method using serum bottles. The new microtiter plate MPN method with an additional replica-plating technique made it possible to enumerate small populations of methanotrophs in cornfield and forest soils. It also requires less equipment, labor, and time than the conventional serum bottle MPN method.