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Increased soil frost associated with winter climate change could have immediate and carry-over effects on biological processes in high-altitude forest soils, but the nature of these processes remain poorly understood. We conducted a snow-exclusion experiment to investigate the immediate and cross-seasonal effects of increased soil frost on soil CO2 efflux and biological activity in a subalpine spruce forest on the eastern Tibetan Plateau, China. The increased frost reduced soil CO2 efflux by similar to 15 and similar to 19% in the winters of 2015/2016 and 2016/2017, respectively. Increased frost also tended to decrease soil basal respiration, the amount of microbial phospholipid fatty acids and the activities of enzymes involved in soil carbon cycling during the winters. Winter soil nitrogen availabilities were higher in the snow-exclusion treatment than in the control plots. However, these effects did not carry over to the following growing season. Our results suggest that increased frost reduces winter soil respiration by direct environmental effects (e.g. soil temperature) and indirect biological processes (e.g. microbial biomass and activity), whereas increased frost did not induce any cross-seasonal effects. These findings underscore the ecological importance of seasonal snowpack and microbe-associated carbon processes in sub alpine forests where winter snowfall is decreasing substantially.