Microbial oxidation is the only biological sink for atmospheric methane. We assessed seasonal changes in atmospheric methane oxidation and the underlying methanotrophic communities in grassland near Giessen (Germany), along a soil moisture gradient. Soil samples were taken from the surface layer (0-10 cm) of three sites in August 2007, November 2007, February 2008 and May 2008. The sites showed seasonal differences in hydrological parameters. Net uptake rates varied seasonally between 0 and 70 μg CH(4) m(-2) h(-1). Greatest uptake rates coincided with lowest soil moisture in spring and summer. Over all sites and seasons, the methanotrophic communities were dominated by uncultivated methanotrophs. These formed a monophyletic cluster defined by the RA14, MHP and JR1 clades, referred to as upland soil cluster alphaproteobacteria (USCα)-like group. The copy numbers of pmoA genes ranged between 3.8 × 10(5)-1.9 × 10(6) copies g(-1) of soil. Temperature was positively correlated with CH(4) uptake rates (P<0.001), but had no effect on methanotrophic population dynamics. The soil moisture was negatively correlated with CH(4) uptake rates (P<0.001), but showed a positive correlation with changes in USCα-like diversity (P<0.001) and pmoA gene abundance (P<0.05). These were greatest at low net CH(4) uptake rates during winter times and coincided with an overall increase in bacterial 16S rRNA gene abundances (P<0.05). Taken together, soil moisture had a significant but opposed effect on CH(4) uptake rates and methanotrophic population dynamics, the latter being increasingly stimulated by soil moisture contents >50 vol% and primarily related to members of the MHP clade.