DOI: 10.5061/dryad.44j0zpcjj
title: Long-term seed bank persistence in Arctomecon californica

description: Seed banks, the collection of viable seeds in the soil, are particularly
 important determinants of population survival in highly variable
 environments. Predictions of increased stochasticity in the amount and
 timing of precipitation in desert environments raise the question of how
 seed banks of desert species will respond to climate change, and
 ultimately, whether these species will persist. Here, we present data from
 our long-term studies of germination requirements and seed bank dynamics
 in a rare desert gypsophile perennial, Arctomecon californica (Las Vegas
 bearpoppy). Arctomecon californica is a relatively short-lived plant that
 recruits from seed in sequences of unusually favorable years. We used
 germination experiments, an in situ seed bank study, and a 15-year field
 seed retrieval study to examine factors affecting seed bank persistence.
 In the germination study, a majority of seeds remained dormant, despite a
 wide variety of treatments, suggesting that a large proportion of the seed
 dispersed each year has cue-nonresponsive dormancy. Our seed bank study
 showed that seed density varied widely between sites, among transects, and
 among samples within a transect. The patchiness of seeds in the soil
 highlights the importance of protecting large areas where A. californica
 populations are known to have existed in the past. The seed retrieval
 study provided strong evidence that this species has a long-lived seed
 bank in which only a small fraction of seeds (roughly 5%) become
 nondormant each year, allowing seed banks of this species to last up to 20
 years without a seed production event. Whether this impressive life
 history strategy can maintain the species in the face of climate change
 depends on the future frequency of the well-timed precipitation that
 allows for establishment of new cohorts of adult plants.
description: Study organism and habitat
 <em>Arctomecon californica</em>, the Las Vegas
 bearpoppy, is a rare herbaceous perennial plant of the Mojave Desert.  The
 species is restricted to barren ‘badlands’ soils high in gypsum or with
 other unusual chemical attributes. It has lost much of its habitat,
 primarily due to urbanization, and it is limited to fragmented, remnant
 populations in the Las Vegas Valley, larger populations around Lake Mead
 in Nevada and Arizona, and a disjunct population in the Lower Grand
 Canyon. The species was recently petitioned for listing under the
 Endangered Species Act. We chose two sites in different
 parts of the species range for the field experiments reported here. The
 first site was a small Las Vegas bearpoppy preserve at the northeastern
 corner of the North Las Vegas Air Terminal in the Las Vegas Valley, while
 the second site was within a bearpoppy population on the road to the St.
 Thomas historic townsite near the north end of the Lake Mead National
 Recreation Area. <em>Arctomecon
 californica</em> is a relatively short-lived plant (known maximum
 life span of seven years) that recruits from seed in sequences of
 unusually favorable years. It has been thought to depend on a long-lived
 seed bank for population persistence through sometimes long periods
 unfavorable for recruitment and survival. Seeds of
 <em>A. californica</em> are known to exhibit
 morphophysiological dormancy (i.e., the embryo is immature at seed
 dispersal and also requires a physiological cue for germination). Wide
 variation among species exists in the conditions required to break
 morphophysiological dormancy. In some species, warm conditions break the
 physiological dormancy of the embryo, and embryo growth occurs at cold
 temperatures; however, some seeds with this morphophysiological dormancy
 can complete both dormancy break and embryo growth during moist chilling.
 Germination of nondormant seeds in <em>A. californica</em>
 depends on winter/spring precipitation, which is highly variable among
 years. Seed germination study The
 seed germination study was carried out in 1995 with seeds collected from
 ripe capsules at the North Las Vegas Air Terminal (NLVAT), at Saint Thomas
 Road (STHOM), and at Rainbow Gardens (RBG) on BLM land in June 1995. The
 germination experiment was carried out in the laboratory by placing
 recently harvested seeds in Petri dishes on the surface of water-saturated
 germination blotters, with subsequent incubation in temperature-controlled
 chambers. The original experimental design included a total of 1032
 experimental units (Petri dishes) and 462 treatment combinations. Each
 treatment included either four replicate dishes of 25 seeds or two
 replicate dishes of 50 seeds as detailed below. We
 determined in earlier experiments with a single 1985 seed population from
 Rainbow Gardens that a single warm moist stratification at 20°C plus a
 single short to intermediate cold stratification (4–12 wks) at 2°C did not
 result in significant dormancy break for <em>A.
 californica</em> seeds previously stored under laboratory conditions
 for three years (germination ≤2%). For this reason, we did not include a
 formal warm moist stratification pretreatment in the 1995 experimental
 design. Our treatments included three seed populations (NLVAT, RBG, and
 STHOM), a series of 11 dry storage temperature by duration treatments,
 seven chilling duration by temperature combinations, and two post-chilling
 incubation temperatures.  Our initial design included a treatment that
 included high-temperature dry-after-ripening (50°C) followed by moist
 chilling on the logic that this combination was most likely to result in
 first-year dormancy break in this spring-emerging species.
 The laboratory seed germination experiment was carried out in two
 steps: an original experiment (data not shown), and an additional
 experiment that added a re-chill treatment as a continuation. The original
 experimental design was factorial, with three seed populations
 (accessions), 11 storage duration-temperature combinations, four chilling
 durations (0, 4, 8, and 12 weeks), two chilling temperatures (1°C, 5°C)
 and two post-chilling incubation temperatures (5/15°C or 10/20°C for
 12h:12h). Treatment combinations included either four Petri dish
 replicates (early treatments) or two replicates (remaining
 treatments). For the storage treatments, seeds were
 stored at 20, 30, 40, and 50°C under dry conditions for 0 to 18 weeks.
 Shorter maximum durations were used at the two warmer temperatures based
 on data from other species showing that prolonged storage at constant high
 temperature could damage the seeds. Storage duration and storage
 temperature were therefore not combined in a full factorial design. After
 each chilling treatment, seeds were incubated at one of the two incubation
 temperatures for 2 weeks and scored for germination. In the earliest
 treatments, seeds were then immediately subjected to viability evaluation;
 viability was consistently extremely high (≥99%). After
 observing essentially no germination in any of the initial treatments, we
 decided to subject the later treatments to a second chilling experience
 after the 2-week germination period. We chose a 12-week cold
 stratification period for the re-chilling treatment. This likely exceeds
 the maximum chilling duration that would be experienced in the field.
 After this second 12-week cold stratification at the same temperatures as
 the original chilling treatments, the seeds were again incubated at the
 two original incubation temperatures for 2 weeks and scored for
 germination and viability. This second round of experimental treatments
 resulted in some dormancy break as reported below. We
 have found that long-term viability in dry storage is usually a necessary
 but not sufficient indicator of the ability of seeds to persist across
 years in field seed banks. We evaluated long-term viability ex post facto
 for the seed lots used in germination and retrieval studies reported here
 by performing tetrazolium tests on four replicates of 50 seeds for each
 seed lot following 27 years of unsealed laboratory storage at room
 temperature (ca. 20°C). Seeds lose viability gradually over time in dry
 storage and this is reflected in a gradual diminution in respiration and
 resulting red color after tetrazolium staining. As individual seeds lose
 viability at different rates, this results in a range of embryo color
 intensity. We took this into account by scoring seeds in three categories;
 viable (red or dark pink), marginal (pale pink), or nonviable (white).
 Seeds in the marginal category would probably not be able to produce
 seedlings, but the fact that they are still respiring indicates that they
 maintained viability longer than seeds in the nonviable
 category. In situ seed bank study
 Plots were established in bearpoppy habitat at NLVAT and at STHOM
 in March 2005. Seed bank plots were set up within seedling establishment
 transects that were part of a separate study, therefore, seedling data are
 not presented here. Seedling emergence transects were located within areas
 where living or dead adult bearpoppies and gypsum soils were
 visible. At NLVAT, 10 transects used for seed bank
 sampling were established across the study site. Quadrats were located at
 distances along the transect lines determined by random numbers generated
 using a stopwatch and were designated only if seedlings were found within
 the randomly located 4 dm<sup>2</sup> quadrat. At STHOM, the
 search protocol was different due to lower seedling densities. The entire
 area within 2 m on one side of each of five transects was searched for
 seedlings. When seedlings were located, 4 dm<sup>2</sup>
 quadrats were designated around them, and these plots were used for both
 the seedling study and the seed bank study.  The transects were laid out
 in parallel at each site, but spacing and length of transects at each site
 were constrained by the requirement that there be suitable soils and
 evidence of bearpoppy occupation within each transect. This resulted in
 variable inter-transect spacing and transect length. Paired seed bank
 samples were taken just outside of each seedling quadrat using a
 cylindrical soil can 6 cm in diameter and 4 cm in depth. Samples were
 lifted into labeled paper collection bags with the aid of a mason trowel.
 Sample size was 247 for NLVAT and 294 for STHOM. Samples were returned to
 the lab for sieving and seed extraction. It was relatively easy to
 visually identify the 2 to 3 mm-long jet black seeds of <em>A.
 californica</em> in the white soils where they occur. Seeds in each
 sample were counted and tested for viability using tetrazolium
 staining. Seed retrieval study Our
 seed retrieval study was conducted with seeds collected from ripe capsules
 from the same three populations described above, at two sites, over 15
 years (1995–2010). Seeds were separated into sets of approximately 100
 using a volume estimate and placed along with identification tags in 3x3
 cm flat packets made of nylon mesh, which were folded over and stapled.
 The seeds were returned to the field within 3 days of harvest. The
 installations were carried out at NLVAT and STHOM. For each replicate,
 packets of each seed collection were buried as a set at approximately 1 cm
 depth by lifting and then replacing the surface lichen crust with a mason
 trowel. Packet groups were installed in three linear blocks at each site
 with 20 packet sets at approximately 0.5 m spacing per block.
 Starting in early summer 1996, seed packets at NLVAT were
 retrieved yearly through 2010, for a total of 15 retrieval dates.
 Retrievals at STHOM were discontinued in 2004, when ground squirrels
 destroyed the seed packets. On each retrieval date except for the final
 date, one set of seed packets from each block was excavated and taken to
 the laboratory for processing. On the final date, all remaining seed
 packets at NLVAT were retrieved. For each packet, any recently germinated
 seeds were counted and removed, and remaining seeds were incubated at a
 near-optimum temperature (5/15 °C) for 4 weeks. Seeds that germinated
 during this period were scored as germinable (i.e., non-dormant). Seeds
 remaining ungerminated at the end of the incubation period were then
 evaluated for viability using tetrazolium and were scored as nonviable,
 viable and dormant, or empty and presumed germinated in the
 field.