Herbarium specimens reveal a significantly earlier flowering trend for Cypripedium acaule (Orchidaceae) in North Carolina (U.S.A.)

Cypripedium acaule, the pink lady’s slipper, is an orchid native to eastern North America, ranging from central and eastern Canada to the southeastern United States. It is a rather common and striking spring wildflower in North Carolina (NC), extending from the mountains to the coastal plain region. A preliminary survey of herbarium specimens at Duke University (DUKE) suggested that flowering times for C. acaule collected in NC in recent years were notably earlier than for those from the last century. Here we set out to investigate this phenological hypothesis by accessing the Southeast Regional Network of Expertise and Collections (SERNEC) portal to extract the metadata from 57 herbaria for 502 herbarium records of C. acaule from NC. Of these, only 193 herbarium specimens from 55 counties, spanning the years 1886–2022, had been collected when the plant was in flower. Each of these “time-stamped” records was manually georeferenced to include latitude, longitude, and elevation coordinates, using tools in Google Earth Pro. Because NC varies by over 2037m in elevation, nearly 3° in latitude, and 10° in longitude, the flowering times recorded for C. acaule spanned 83 spring days. To control for the effect of location on flowering time, we implemented Hopkins’ Bioclimatic Law to normalize flowering times across the state. A linear regression of the 193 normalized flowering dates suggests an overall shift in blooming that has advanced by 21 days since 1886. A mixed-effects regression was performed to determine the relationship of elevation, latitude, average winter temperature, total winter precipitation, and the total number of winter frost days from the year and location of collection (as fixed variables) on the effect of flowering day of year (DOY). The most significant effect on the flowering DOY was from average winter temperature: for each increase in 1° C, the flowering DOY was 3.23 days earlier. If this trend continues, the flowering time of C. acaule could become decoupled from the peak activity of its pollinators, increasing the risk of reproductive failure. This study highlights the vital role of herbarium specimens in understanding the effects of climate change on shifts in phenological patterns.