Annls of Botny 102: 255 264, 2008 doi:10.1093/o/mcn087, ville online t www.o.oxfordjournls.org Interction Between Seed Dormncy-relese Mechnism, Environment nd Seed Bnk Strtegy for Widely Distriuted Perennil Legume, Prkinsoni culet (Ceslpincee) RIEKS D. VAN KLINKEN 1,2, *, BERT LUKITSCH 3 nd CARLY COOK 1 1 CSIRO Entomology, Indooroopilly, Queenslnd, Austrli, 2 CRC for Austrlin Weed Mngement nd 3 Weed Mngement Brnch, Berrimh, Northern Territory, Austrli Received: 14 Mrch 2008 Returned for revision: 24 April 2008 Accepted: 12 My 2008 Pulished electroniclly: 16 June 2008 Bckground nd Aims Prkinsoni culet (Ceslpincee) is perennil legume with seeds tht hve hrdseeded ( physicl) dormncy nd re potentilly very long-lived. Seed dormncy is chrcteristic tht cn oth help mximize the proility of seedling estlishment nd spred the risk of recruitment filure cross yers (et-hedging). In this study, dormncy-relese ptterns re descried cross the diverse environments in which this species occurs in order to test whether wet het (incution under wet, wrm-to-hot, conditions) lone cn explin those ptterns, nd in order to determine the likely ecologicl role of physicl dormncy cross this species distriution. Methods A seed uril tril ws conducted cross the full environmentl distriution of P. culet in Austrli (rid to wet-dry tropics, uplnds to wetlnds, soil surfce to 10 cm deep). Key Results Wet het explined the pttern of dormncy relese cross ll environments. Most seeds stored in the lortory remined dormnt throughout the tril (t lest 84 %). Dormncy relese ws quickest for seeds uried during the wet seson t reltively high rinfll, uplnd sites (only 3 % of seeds remined dormnt fter 35 d). The longest-lived seeds were in wetlnds (9 % remined dormnt fter lmost 4 yers) nd on the soil surfce (57 % fter 2 yers). There ws no consistent correltion etween incresed ridity nd rte of dormncy relese. Conclusions The results suggest tht physicl dormncy in P. culet is mechnism for mximizing seedling estlishment rther thn et-hedging strtegy. However, seed persistence cn occur in environmentl refuges where dormncy-relese cues re wek nd conditions for germintion nd estlishment re poor (e.g. under dense vegettion or in more rid micro-environments) or unsuitle (e.g. when seeds re inundted or on the soil surfce). Risks of recruitment filure in suoptiml environments could therefore e reduced y inter-yer fluctutions in microclimte or seed movement. Key words: Bet-hedging, dormncy-relese mechnisms, environmentl refuges, legume, Prkinsoni culet, physicl dormncy, seed nk persistence, seed uril depth, seed dormncy, tropicl wetlnds, wet het, vrile environment. INTRODUCTION Dormncy is chrcteristic of the seed tht prevents it from germinting in conditions tht otherwise would hve een fvourle (Vleeshouwers et l., 1995; Bskin nd Bskin, 1998). Seed dormncy cn serve t lest two roles. First, it cn restrict opportunities for germintion to smll windows in time (e.g. time of yer) or spce (e.g. seed uril depth) when seedling estlishment is most likely to occur (Vleeshouwers et l., 1995; Benech-Arnold et l., 2000; Bker et l., 2005). In this cse seeds my e expected to e short-lived in environments where suitle conditions for recruitment re frequent. Second, it cn serve s et-hedging strtegy y delying germintion, even when suitle conditions for germintion nd seedling estlishment re encountered, nd thus spreding the risk of recruitment filure cross yers (Fcelli et l., 2005; Lmers et l., 2005; Evns et l., 2007; Simons nd Johnston, 2007). Bet-hedging, lthough rrely tested (Evns et l., 2007), is expected to ecome incresingly importnt s the proility of * For correspondence. E-mil rieks.vnklinken@csiro.u Present ddress: School of Nturl nd Rurl Systems Mngement, The University of Queenslnd, St Luci, Queenslnd, Austrli recruitment filure for prticulr cohort increses, for exmple with incresing inter-yer environmentl vrince. Seed dormncy nd the mechnisms for dormncy relese cn therefore hve n importnt role in modulting the distriution nd undnce of plnt species (Bskin nd Bskin, 1998; Cin et l., 2000; Hndley nd Dvy, 2005). Hrdseededness, or physicl dormncy, is very common type of seed dormncy imposed y hving hrd test (or endocrp), especilly mong shrus nd trees (Bskin nd Bskin, 1998; Morrison et l., 1998; Bskin et l., 2000). Seeds with physicl dormncy re typiclly very long-lived (.100 yers) when stored under dry, cool conditions (Fenner, 1995), nd re generlly considered to e long-lived in the field (Bskin nd Bskin, 1998; Morrison et l., 1998; Holmes nd Newton, 2004). In contrst to mny physiologicl dormncy mechnisms, the timing of relese from physicl dormncy is prticulrly criticl s seeds lck secondry seed-dormncy mechnisms or cyclicl dormncy (Bskin nd Bskin, 2004; Bker et l., 2005). Seeds will therefore either germinte or die (rot) once the hrd test is compromised nd there is sufficient moisture for seeds to imie wter. Physicl dormncy my therefore e expected to e importnt for optimizing the timing of germintion events to mximize # The Author 2008. Pulished y Oxford University Press on ehlf of the Annls of Botny Compny. All rights reserved. For Permissions, plese emil: journls.permissions@oxfordjournls.org
256 vn Klinken et l. Seed Dormncy Relese in Prkinsoni culet seedling estlishment. The reltive importnce of et-hedging, where proportion of seeds would e expected to remin dormnt even when conditions for seedling estlishment were idel, is less cler lthough it is commonly ttriuted to seeds with physicl dormncy (Thompson et l., 2003). A rnge of mechnisms hs een proposed for dormncy relese of seeds with physicl dormncy, ut they generlly remin poorly understood (Bskin nd Bskin, 1998; Morrison et l., 1998; vn Klinken nd Flck, 2005). However, recent study hs shown tht wet het (incution under wet, wrm-to-hot conditions) is n importnt dormncy-relese mechnism for the perennil, tropicl, leguminous tree Prkinsoni culet (Ceslpincee; vn Klinken nd Flck, 2005). The reltionship etween temperture nd dormncy relese pproximtes sigmoidl curve for seed popultions under wet conditions, with the most temperture-sensitive prt of the reltionship (pprox. 80 % of fresh seeds remin dormnt under wet conditions t 25 8C ut only 20 % t 40 8C) coinciding with tempertures typiclly encountered in tropicl regions (Nix, 1981). Conditions required for dormncy relese re therefore tighter thn those required for germintion, which is optiml etween 25 nd 35 8C (vn Klinken nd Flck, 2005), nd cn commence t lower moisture levels (L. Anderson, Pilr Mesquite Mngement Committee, Austrli, unpul. res.) thn dormncy relese. In the wet-dry tropics this dormncy-relese mechnism ws demonstrted to result in low rtes of dormncy relese in situtions where the environment ws uffered from temperture extremes, such s under dense herceous or folir cover, or deep uril (vn Klinken et l., 2006). However, it is not known whether other mechnisms might lso e involved in other climtic conditions such s dry het, which hs een reported for some legumes in Mediterrnen nd rid environments (Normn et l., 2002). Indirect evidence suggests tht wet het might e reltively common mechnism for seeds with physicl dormncy (Benech-Arnold et l., 2000; vn Klinken et l., 2006), lthough results from most studies re difficult to interpret ecuse dormncy relese nd germintion (which is lso temperture-sensitive) processes re often confounded (Benech-Arnold et l., 2000). Studies on the interction etween dormncy relese nd the environment re typiclly restricted to one or few sites (Hesse et l., 2007), which potentilly limits our understnding of the ecologicl significnce of prticulr dormncy-relese mechnisms. For exmple, the seed nk strtegies of plnt species re often ctegorized s either trnsient or persistent (Csontos nd Tms, 2003), ut how these strtegies might e influenced y environmentl context is rrely considered. We therefore conducted seed-uril tril in Austrli cross the ntionl distriution of P. culet. Prkinsoni culet is ntive to the Americs (Hwkins et l., 2007), ut it is most widespred nd undnt s n invsive in Austrli where it occurs cross wide rnge of environments, from the rid zone to the wet-dry tropics, nd in uplnds nd wetlnds (Fig. 1). Seeds were uried cross the full rnge of environments in which this species occurs in order to test three things. First, whether wet het lone explins the dormncy-relese ptterns oserved cross the full rnge of nturl environments encountered y this species (ctegorized y climte, hitt nd uril depth). Second, whether single dormncy-relese mechnism cn e sufficiently roust to ensure tht seeds re relesed from dormncy t times nd plces when seedling estlishment is most likely to occur (with respect to seed uril depth, temperture, soil moisture nd competitors). Finlly, whether there is ny evidence tht seed dormncy lso cts to spred recruitment nd therefore the risk of recruitment filure cross yers, even when idel conditions for recruitment re experienced. If et-hedging occurs, then we would expect significnt proportion of seeds to remin dormnt following conditions tht re idel for estlishment. The documented etween-seed vrition in response to wet het (vn Klinken nd Flck, 2005) would e one mechnism for this to occur. MATERIALS AND METHODS Seeds Two seed cohorts of Prkinsoni culet L. were used, one from the Victori River District () of the Northern Territory nd the other from Centrl Queenslnd (Temus; Tle 1, Fig. 1), ecuse smples from these two regions hd previously een shown to hve contrsting sensitivities to wet het (vn Klinken nd Flck, 2005) nd therefore my respond differently in seed uril tril. Seeds were collected on 7 Novemer 2001 () nd 13 Jnury 2002 (Temus), to correspond with pek pod lods on trees in ech region (vn Klinken nd Flck, 2008). In ech cse, fully mture (strw-coloured) pods were collected from trees within single popultion nd irdried in the lortory upon collection. They were then shelled mnully nd undmged, fully-formed seeds were stored in pper gs under mient lortory conditions (25 + 5 8C) together with insecticide strips to protect ginst seed-feeders. One week prior to seed uril, undmged seeds (9 mm long, 4 mm wide) were rndomly su-smpled into lots of 50 seeds. Seed lots were plced in 15 15 cm pckets mde from plstic shde cloth (1.5 1.5 mm holes), folded over nd stpled shut, for susequent uril. Tretments Seed uril sites were locted throughout the Austrlin distriution of P. culet so s to represent the rnge of climtic conditions nd hitts in which it occurs (Tle 1, Fig. 1). Climtic conditions rnged from rid to wet-dry tropics. Hitt comprisons t some loctions contrsted either inundtion ptterns or soil type (Tle 1). Hitt ws defined y inundtion regimes: uplnds were never inundted (flooded); riprin sites were locted on river nks nd were suject to pulse inundtion; wetlnds were sesonlly inundted for long durtions (months); nd lowlnds were periodiclly inundted for long durtions. However, only wetlnd hitts were inundted during the course of the study. Soil type ws included s
vn Klinken et l. Seed Dormncy Relese in Prkinsoni culet 257 N Betrice Hill Millstrem Alcoot Temus Leur 500 km Annul rinfll (mm) 99 300 301 600 601 900 901 1200 1201 8000 FIG. 1. The seven seed uril loctions mpped over the men nnul rinfll cross Austrli (see Tle 1 for detils). tretment t s red (cly-lom) nd lck (hevy crcking cly) soils re the two most common ut contrsting soils in tht region (Brkly Region). Seed pckets were uried t depth of 2 cm t ll sites, which is likely to e n optiml depth for germintion (Cox et l., 1993). In ddition, the effect of seed uril depth (0 cm, on the soil surfce; 2 cm; 10 cm) ws tested t two loctions (Tle 1). Seeds were uried t the time of the yer tht corresponded with the end of pek pod-drop in ech loction (vn Klinken nd Flck, 2008). Seeds sourced from were uried from Decemer 2001 to Jnury TABLE 1. Site nd tretment detils for the seed uril tril Buril depth (retrievl events) Loction Climte Hitt seed Temus seed Betrice Hill 12 839 0 S 131 819 0 E Wet-dry tropics (wrm winters) Uplnd 2 cm (5) 2 cm (5) Wetlnd 2 cm (9) 15 826 0 S 130 820 0 E Semi-humid wet-dry tropics (wrm winters) Uplnd 0 cm (5) 2 cm (6) 2 cm (5) 10 cm (5) Wetlnd 2 cm (7) 19 818 0 S 136 804 0 E Semi-rid wet-dry tropics (wrm winters) Uplnd (lck soil) 2 cm (6) 2 cm (4) Uplnd (red soil) 2 cm (5) Temus 21 800 0 S 146 822 0 E Semi-humid wet-dry tropics (cool winters) Uplnd 0 cm (5) 2 cm (6) 10 cm (5) Lowlnd 2 cm (6) Leur 23 810 0 S 149 834 0 E Semi-humid wet-dry tropics (cool winters) Uplnd 2 cm (6) Millstrem 21 835 0 S 117 802 0 E Arid (wrm winters) Uplnd 2 cm (4) 2 cm (5) Lowlnd 2 cm (5) Alcoot 22 849 0 S 134 827 0 E Arid (cool winters) Uplnd 2 cm (5) Riprin 2 cm (7)
258 vn Klinken et l. Seed Dormncy Relese in Prkinsoni culet 2002 in loctions with wrm winters, nd those sourced from Centrl Queenslnd were uried t ll loctions from Mrch to April 2002. Ten to 16 seed pckets of ech seed source (depending on the numer of plnned retrievl events) were wired to steel pickets nd uried t three replicte sites in the vrious loction hitt uril depth comintions (Tle 1). Replictes within loction were plced etween 100 nd 5000 m prt. Where possile, hitt tretments were pired, with tretments eing plced 50 200 m prt (, Temus nd Alcoot). Sites were plced wy from the cnopy of shrus nd trees to minimize potentil shding effects. Two (or one, on only 10 % of occsions) rndomly selected pckets were periodiclly retrieved from ech site etween 35 nd 1281 d fter uril. Results from duplicte pckets were verged in the nlysis. The exct retrievl schedule vried etween tretments, depending on the numer of pckets uried, the expected rte of seed dormncy relese t prticulr site, nd the pttern of sesonl chnges t ech loction. Control seed pckets (one per replicte) from ech seed source were stored under mient lortory conditions (25 + 5 8C) nd processed with ech tch of retrieved seeds (four replictes). Seed evlution All intct seeds were ir-dried in the lortory within 2 d of retrievl nd their viility nd dormncy were determined, together with those of their controls. Seeds tht were no longer intct hd imied nd either germinted or died (rotted) within the uril pcket, lthough there ws no wy of telling which t the time of retrievl. A stndrd protocol (vn Klinken nd Flck, 2005; vn Klinken et l., 2006) ws used to determine the proportion of seeds in ech smple tht were dormnt, nd the proportion of nondormnt nd dormnt seeds tht ws vile. Dormncy ws defined s those seeds tht hd not imied following 4 d sumerged in wter t 20 8C. Rtes of dormncy relese re constnt elow pprox. 25 8C, nd most seeds tht will imie t 20 8C will do so within 4 d (vn Klinken nd Flck, 2005). Imied seeds were removed dily nd llowed to germinte in Petri dishes lined with pper nd kept moist t 25 8C, which is within the optiml rnge for germintion (25 35 8C; vn Klinken nd Flck, 2005). Seeds clssed s dormnt hd their seed cots individully nicked, were sumerged in wter t 20 8C for further 24 h nd, once imied, processed for germintion. Seeds were considered to e vile once the rdicle ws hlf the seed length. Any imied seeds tht hd not germinted fter 2 weeks were invrily mouldy nd were clssed s unvile. The effects of seed source, census dte nd their interction on the dormncy relese of seeds in the control tretment were nlysed using repeted-mesures generlised liner model (SAS MIXED procedure). In the model, seed retrievl dte ws pplied s fixed, repeted effect with n utoregressive covrince structure, seed source s fixed effect, nd replicte s rndom effect. Testing the effect of wet het The effect of wet het on seed dormncy relese over time ws tested using proportionl dily chnges in dormncy relese, which stndrdized the effect of vrile smpling intervls. This ws clculted y dividing the difference in seed dormncy etween two successive seed retrievl dtes (verged for the two gs retrieved from ech site on ech dte) y the numer of dys tht hd elpsed since the previous census. The resultnt vlue ws slightly negtive on 46 of the 244 collections used in the nlyses. This ws ttriuted to smpling error, s the process of dormncy relese cn not e reversed in this species nd, consequently, these vlues were truncted t zero. Dormncy levels of control seeds (95.5 % t dy zero) were used s n estimte of seed dormncy t the time of uril. Two types of nlyses were conducted: n nlysis of vrince to investigte the effect of moisture nd het comintions (i.e dry nd wet, nd cool nd wrm), nd multiple regression to more explicitly exmine the reltionship etween soil moisture nd temperture on dormncy relese (using SAS relese 8.2; SAS Institute, Cry, NC, 1999). Climte dt for ech loction ws otined from SILO, dtse of historicl climte dt interpolted t 0.05 degree intervls cross Austrli (Jeffrey et l., 2001). Soil moisture ws modelled using dily tipping ucket model for the top 200 mm of soil, ssuming run-off ove sturtion (200 mm), 25 mm d 21 dringe etween sturtion nd field cpcity (150 mm) nd 3.2 mm evpotrnspirtion etween field cpcity nd dry soil (110 mm; vn Klinken et l., 2006). Anlyses only included seeds uried t 2 cm s it ws not possile to model microclimte t the soil surfce, nd microclimte t 2 nd 10 cm ws expected to e qulittively similr (vn Klinken nd Flck, 2006). Seeds uried in the wetlnd hitts t oth nd Betrice were excluded ecuse the seeds were sujected to prolonged inundtion nd microclimte could therefore not e modelled with the ville dt. Dt were lso excluded for intervls with too few seeds remining t the strt of the intervl (10 %) to relisticlly ssess rtes of dormncy relese. For the nlysis of vrince, environmentl conditions experienced y the uried seeds etween seed retrievl dtes were ctegorized s wet nd wrm, dry nd wrm, wet nd cool, or dry nd cool. Time periods were ctegorized s wet nd wrm if ny 3 dys in tht period were wet (soil moisture greter thn 130 mm) nd wrm (verge temperture t lest 25 8C), or ctegorized in one of the remining groups if one or more of those criteri were not met. Wet nd wrm conditions were defined to cpture conditions under which dormncy relese is sensitive to wet het (vn Klinken nd Flck, 2005). Only one site on single occsion ws ctegorized s wet nd cool (Leur, 0 113 d fter uril), so those three replicte dt points were excluded from the nlysis. A three-fctor ANOVA (SAS MIXED procedure) ws used to exmine whether the dily rte of dormncy relese ws ffected y het type (three types), loction (seven types) nd uril durtion (uried less thn or more thn 12 months). Het type nd uril durtion were pplied s fixed
vn Klinken et l. Seed Dormncy Relese in Prkinsoni culet 259 effect nd loction s rndom effect. Interction terms could not e modelled due to the unlnced nture of the design (with respect to het types in ech region). Dormncy relese dt were trnsformed prior to nlysis to stisfy the ssumptions of normlity nd homoscedsticity, nd we found tht log (x þ 0.000025) ws more suitle thn rcsine trnsformtion. A multiple liner regression (SAS REG procedure) ws performed to exmine the reltionships etween temperture, soil moisture nd their interction on rte of dormncy relese. Averge temperture nd verge soil moisture conditions over the period since the lst census dte were modelled s continuous vriles. No dt trnsformtion ws required to stisfy the ssumptions of the nlysis. Effect of hitt nd depth The effects of seed depth (two loctions) nd hitt (six loctions; Tle 1) on dormncy relese were tested using one-wy ANOVA on rnk-trnsformed dt for ech dte tht seeds from different depth or hitt tretments were retrieved t the sme time from prticulr loction. RESULTS Seed dormncy nd viility Controls. Most seeds in the controls were dormnt when tested through the course of the tril (91 %, n ¼ 3200). Seed dormncy ws significntly ffected y time (F 9,42 ¼ 4.92, P ¼ 0.0002), with seed dormncy dropping pproximtely 6 % over 1200-d period. However, there ws no effect of seed source (F 1,6 ¼ 0.60, P ¼ 0.4685) nd no interction etween seed source nd time (F 5,42 ¼ 1.95, P ¼ 0.1061). Seed source ws therefore not considered s fctor in susequent nlyses. Most dormnt (96 %, n ¼ 2771) nd non-dormnt (83 %, n ¼ 278) seeds were vile. Buried seeds. Approximtely 34 % of the 32 050 uried seeds were intct upon retrievl of seed pckets. Of the intct seeds, 81 % were still dormnt nd 16 % were nondormnt nd vile. Of the dormnt seed, 89 % were vile. The remining nlysis is of seed dormncy s proportion of totl seeds uried. Reltionship etween wet het nd dormncy relese The nlysis of vrince showed tht het type (F 2,139 ¼ 30.7, P, 0.0001) hd n effect on seed dormncy relese, ut region (F 6,139 ¼ 2.0, P ¼ 0.07) nd uril durtion (uried less thn or more thn 12 months; F 1,139 ¼ 1.5, P ¼ 0.22) did not. Pooled cross the vrious loctions nd uril durtions, greter reduction in dily proportionl dormncy relese occurred when environmentl conditions were wet nd wrm (men + s.e., 0.0063 + 0.0009, n ¼ 63), compred with dry nd wrm (0.0016 + 0.0003, n ¼ 72) (t 139 ¼ 6.7, P, 0.0001) nd dry nd cool (0.0009 + 0.0004, n ¼ 15; t 139 ¼ 6.0, P, 0.0001), nd when conditions were dry nd wrm compred with dry nd cool (t 139 ¼ 2.0, P ¼ 0.048). Visul ssessment of these dt lso shows cler reltionship etween dormncy relese nd wet nd wrm conditions (Figs 2 5; see elow). However, there were three exceptions for Temus-sourced seeds: t uplnd A Proportion dormnt Proportion dormnt Proportion dormnt Proportion dormnt Temus Time since tril commencement (d) B seeds uried dy 0 (5 Dec. 2001) Temus seeds uried dy 97 (12 Mr. 2002) C D seeds uried dy 0 (19 Dec. 2001) Temus seeds uried dy 107 (20 Mr. 2002) Temus 900 Time since tril commencement (d) Betrice seeds uried dy 0 (19 Dec. 2001) Temus seeds uried dy 91 (20 Mr. 2002) Temus 900 Time since tril commencement (d) Millstrem seeds uried dy 0 (10 Jn. 2002) Temus seeds uried dy 91 (11 Apr. 2002) Temus 900 100011001200 Time since tril commencement (d) FIG. 2. The proportion of P. culet seeds (men + s.e., three replictes) tht remined dormnt throughout the tril tht were uried erly in the wet seson (seeds sourced from ) nd lte in the wet seson (seeds sourced from Temus) t uplnd sites t (A), (B), (C) Betrice, nd (D) Millstrem. Dormncy t time of uril is circled for ech seed source. Horizontl rs indicte when conditions were wet nd wrm.
260 vn Klinken et l. Seed Dormncy Relese in Prkinsoni culet A Proportion dormnt B Proportion dormnt C Proportion dormnt D Proportion dormnt Temus Dy 0 = 12 Mr. 2002 Uplnd Lowlnd Time since uril (d) Leur Dy 0 = 23 Apr. 2002 Time since uril (d) Dy 0 = 19 Dec. 2001 Uplnd, lck soil Uplnd, red soil Time since uril (d) Millstrem Dy 0 = 10 Jn. 2002 Uplnd Wetlnd 900 1000 1100 Time since uril (d) FIG. 3. The proportion of seeds (men + s.e., three replictes) tht remined dormnt throughout the tril t vrious hitts t (A) Temus, (B) Leur (uplnds only), (C) nd (D) Millstrem. There were no significnt hitt ffects on dormncy relese t these loctions. Horizontl rs indicte when conditions were wet nd wrm. Lowlnd nd wetlnd sites were not inundted during the course of this study. etween dy 0 nd 89 (Fig. 2B); t uplnd nd lowlnd Temus etween dy 0 nd 38 (Fig. 3); nd t uplnd Millstrem etween dy 182 nd 465 fter uril (or 273 556 d from commencement of the tril; Fig. 2D). A Proportion dormnt B Proportion dormnt C Proportion dormnt Alcoot Uplnd Riprin This unexplined dormncy relese my e rel, or could e due to ssumptions in the soil moisture model nd the interpolted rinfll dt it relied upon (e.g. 111 mm of rin fell over the relevnt period t Millstrem, ut ws not sufficient to meet our criteri for wet nd wrm ccording to the soil moisture model). The multiple liner regression showed tht the dily rte of seed dormncy relese ws strongly dependent on the comined effects of verge temperture, soil moisture nd their interction (F 7,247 ¼ 107.9, P, 0.0001, r 2 ¼ 0.68; Tle 2). There ws cler reltionship etween dormncy Dy 0 = 25 Fe. 2002 Time since uril (d) Betrice Time since uril (d) 0 200 400 600 800 1000 1200 Time since uril (d) Dy 0 = 5 Dec. 2001 Uplnd Wetlnd Dy 0 = 19 Dec. 2001 Uplnd Wetlnd FIG. 4. The proportion of seeds (men + s.e., three replictes) tht remined dormnt throughout the tril in contrsting hitts t (A) Alcoot, (B) nd (C) Betrice. Vlues represented y different letters indicte tht dormncy relese within the census dtes vried significntly etween hitt types (P, 0.05). Horizontl rs indicte when conditions were wet nd wrm (thick rs) nd when wetlnd sites were inundted (thin rs).
vn Klinken et l. Seed Dormncy Relese in Prkinsoni culet 261 A Proportion dormnt B Proportion dormnt Time since uril (d) Temus c 0 cm 2 cm 10 cm c Time since uril (d) relese nd incresingly wet nd wrm conditions (Fig. 6). However, the microclimte dt used in this nlysis were verged over ech seed uril intervl, ut in relity fluctuted considerly from dy to dy (especilly soil moisture). If the extreme moisture vlues re driving relese from dormncy, this focus on verges will underestimte the ctul moisture nd het required for dormncy relese. Reltionship etween climte, uril dte, hitt nd depth nd dormncy relese Most seeds were relesed from dormncy within the first yer of uril (Fig. 7). Of seeds sourced from nd uried in Decemer to Jnury, the gretest proportion of TABLE 2. Multiple liner regression on the reltionship etween the dily rte of seed dormncy relese, the effects of verge temperture, soil moisture, nd their interction Coefficient Estimte s.e. t P Dy 0 = 5 Dec. 2001 Intercept 0.378 0.067 5.63,0.0001 Temperture 0.0142 0.0022 6.36,0.0001 Soil moisture 0.00344 0.00061 5.66,0.0001 Temperture Soil moisture 0.00013 0.00002 10.12,0.0001 Dy 0 = 12 Mr. 2002 F IG. 5. The effect of seed uril depth on the proportion of seeds tht remin dormnt t uplnd sites t (A) nd (B) Temus. Vlues represented y different letters indicte tht dormncy relese within the census dtes vried significntly etween uril depths (P, 0.05). Dt for 0 d since uril refers to the control seeds. Horizontl rs indicte when conditions were wet nd wrm. < 1 Dormncy relese (% seeds d 1 ) 1 3 3 5 150 145 140 135 130 125 120 115 25 26 27 28 29 30 31 32 33 34 35 110 Temperture (ºC) FIG. 6. A contour-surfce plot of the reltionship etween rte of dormncy relese nd temperture nd soil moisture (verged over the uril durtion). The prmeter estimtes were tken from the multiple regression model (Tle 2). them (11 %) ws still dormnt fter the first yer t the most rid site (Fig. 7). However, there ws greter vrition in seed dormncy levels etween soil types within single climtic region (t ) thn etween climte regions. There ws no reltionship etween dormncy relese nd decresing ridity for seeds sourced from Temus nd uried lter in the wet seson. More seeds sourced from Temus were still dormnt fter 1 yer cross ll climte zones, when compred with those sourced from (Fig. 7). The response of seeds from the two seed sources to immersion t 20 8C ws identicl under controlled conditions. However, it is possile tht seeds uried lter in the wet seson were longer-lived ecuse the herceous cover hd more chnce to regrow nd uffer the microenvironment (see Discussion). The pttern of dormncy relese through time ws gretly ffected y time of seed uril (Fig. 2), s might e expected y the reltionship with wet het (Fig. 6). Most seeds uried in uplnd hitts t depth of 2 cm erly in the wet seson were no longer dormnt within few months, while most of those uried lte in the wet seson remined dormnt until the following wet seson. The interction etween hitt (inundtion ptterns nd/ or soil type) nd dormncy relese of seeds uried t 2 cm ws tested t six of the seven loctions (Tle 1). Hitt hd little or no effect t three loctions, Temus (uplnd vs lowlnds), Millstrem (uplnds vs wetlnds) nd Alroy Downs (lck soil vs red soil; Fig. 3). However, mesic hitts t Temus nd Millstrem were never inundted during the tril. This contrsted with Alcoot, where dormncy relese ws slower in the riprin hitt, despite never eing inundted (Fig. 4). Wetlnds t Betrice nd were the only sites tht were inundted, nd they were lso the only sites where there were contrsting dormncy-relese ptterns Soil moisture (mm)
262 vn Klinken et l. Seed Dormncy Relese in Prkinsoni culet 0 5 source seeds (uried Dec. Jn.) Temus source seeds (uried Fe. Apr.) 341 mm Proportion dormnt 0 3 0 1 1300 mm 805 mm 450 mm 278 mm 631 mm 1512 mm 560 mm 291 mm 191 mm 231 mm Millstrem Alcoot Temus (lck soil) Leur Betrice Millstrem (red soil) (lck soil) Betrice F IG. 7. Seed dormncy levels (seed persistence) fter pprox. 1 yer for seeds sourced from nd Temus nd uried 2-cm deep t uplnd sites in ech region. Regions re ordered y incresing ridity during the uril period (totl rinfll for ech uril period is given for ech loction). Detiled time series dt is provide in Figs 3 5. etween hitts (Fig. 4). Less thn 5 % of seeds were still dormnt t uplnd sites within the first 3 months. In contrst, most wetlnd seeds were still dormnt fter the first wet seson t, nd in Betrice 12 % were still dormnt t the finl retrievl dte during the fourth wet seson (1094 dys fter seed uril). The gretest drops in the proportion of dormnt seeds in wetlnd hitts coincided with the strt of the rins nd/or with the wetlnds drying out. However, there ws little reduction t Betrice fter the second wet seson. The interction etween uril depth nd dormncy relese ws tested in uplnd hitts in two regions (Tle 1; Fig. 5). Dormncy relese ws significntly (P, 0.05) ffected y seed depth cross oth regions on eight of the 11 dtes exmined. There ws little difference in dormncy relese etween seeds uried t 2 cm nd 10 cm. In contrst, seeds on the soil surfce persisted t much higher levels. For exmple, t less thn 3 % of seeds were still dormnt fter 37 d t 2 cm, ut 30 % of seeds were still dormnt fter 736 d on the soil surfce (Fig. 5). DISCUSSION The results suggest tht wet het is the primry mechnism for dormncy relese for P. culet cross ll environmentl conditions tht it is likely to encounter. Furthermore, the ville evidence suggests tht wet het provides n elegnt mechnism for ensuring tht germintion coincides with conditions tht re optiml for seedling estlishment cross diverse rnge of environments. In ddition, seeds cn e long-lived ut there is little evidence tht seed dormncy cts expressly s mechnism to del with inter-yer vrition in recruitment proilities, s most or ll uried seeds lose dormncy within yer when suitle recruitment conditions re encountered. Ech of these three points is ddressed in turn elow. Wet het s dormncy-relese mechnism The strong, empiriclly determined reltionship etween wet het nd dormncy relese for P. culet explined the pttern of dormncy relese of uried seed cross the full rnge of climte zones nd micro-environments in its Austrlin distriution. This reltionship therefore provides predictle nd strongly grduted reltionship etween environment nd seed longevity. This extends the conclusions drwn from physiologicl studies (vn Klinken nd Flck, 2005) nd seed uril tril conducted in the wet-dry tropics (vn Klinken et l., 2006) on the sme species. It lso suggests tht physicl dormncy mechnisms re specilized dpttions (Morrison et l., 1998; Bskin et l., 2000). For exmple, there ws no evidence tht dry het during hot dry summer periods resulted in dormncy relese in rid environments despite it eing commonly recorded dormncy-relese mechnism for other legume species (Normn et l., 2002). The physiologicl mechnisms underlying wet het s dormncyrelese mechnism hve not een determined, ut is likely to involve ctive, temperture-dependent chnges in seed-cot porosity elow threshold (pprox. 33.6 8C) overlying liner physicl reltionship etween temperture nd wter uptke (vn Klinken nd Flck, 2005). Wet het vried considerly cross the environments nd microclimtes tht we exmined, nd resulted in very different dormncy-relese responses. Inundted wetlnds provided the most stle micro-environment for uried seed (Hndley nd Dvy, 2005), with those in the wet-dry tropics typiclly eing pprox. 31 8C (R. vn Klinken, unpul. res.). Lortory results suggested tht pprox. 60 % of P. culet seeds will remin dormnt when
vn Klinken et l. Seed Dormncy Relese in Prkinsoni culet 263 inundted for long periods t this temperture (vn Klinken nd Flck, 2005) nd this prediction ws supported t oth of the wetlnd sites tht were inundted for long periods. Seeds of other species with physicl dormncy hve lso een reported to e very long-lived under wter, only eing relesed from dormncy once wters recede (Vitelli et l., 2006). In contrst, micro-environment vries considerly s wetlnds flood nd dry. For exmple, seeds cn e exposed to very high dily peks in temperture when locted for extended periods in shllow wter or sturted, re soils prior to the wetlnds filling or s the wetlnds dry out. Reduction in seed dormncy ws pprent s wetlnds filled nd dried out, lthough dt were not collected t sufficient temporl resolution to test this explicitly. Dormncy relese differed qulittively in comprisons etween other hitts tht were not inundted during the course of the study. For exmple, dormncy relese ws slower in riprin sites thn uplnd sites (which hd less herceous cover) t Alcoot, nd in red soil thn lck soil t Alroy. However, hitt differences were reltively sutle s were, presumly, the contrsts in wet het exposure resulting from differences in vegettion cover nd soil type. Timing of dormncy relese to mximize seedling estlishment Dormncy relese ws consistently timed to coincide with conditions tht re likely to e optiml for seedling germintion nd estlishment. Although germintion nd estlishment were not mesured in this study, this conclusion ws strongly supported y the situtions under which dormncy relese occurred. Rtes of dormncy relese were gretest when seeds were uried (survivl of seedlings tht germinte t the soil surfce is miniml; R. vn Klinken, pers. os.) ut not inundted (P. culet seedlings cn not germinte under wter; vn Klinken nd Flck, 2005), when conditions re wet nd wrm-to-hot (optiml conditions for germintion nd susequent recruitment; vn Klinken nd Flck, 2005) nd when there is little herceous or folir cover (Prkinsoni seedlings re poor competitors; S.D. Cmpell, Queenslnd Deprtment of Primry Industries, Austrli, unpul. res.). Furthermore, wet het s dormncy-relese mechnism ws very roust, ensuring tht dormncy relese occurred under optiml conditions cross very diverse climtes, environments nd ltitudes. This contrsts with other plnt species where seed dormncy mechnisms hve plced geogrphic or environmentl constrints on plnt distriutions (Bskin nd Bskin, 1998; Hndley nd Dvy, 2005). Seeds commonly hve mechnisms to prevent germintion when deeply uried nd to mximize germintion when there re gps in the folige. Three previously suggested mechnisms for this re inhiition of germintion y light with low red-to-fr-red rtio, stimultion y fluctuting (or high) tempertures, nd dormncy termintion cused y nitrte or other chemicls (Fenner, 1995; Benech-Arnold et l., 2000; Bskin nd Bskin, 2006). Our results suggest fourth mechnism, wet het, which hs the dded dvntge of limiting dormncy relese when seeds re still on the soil surfce. Seedling estlishment rtes re often low for seeds on the soil surfce ecuse of difficulties in penetrting soil crusts nd exposure to high tempertures nd desicction (Cox et l., 1993). Prkinsoni culet seeds were longest-lived on the soil surfce, which is the opposite result from previous study conducted in the wet-dry tropics (vn Klinken et l., 2006). This ws lmost certinly the consequence of seed pckets eing plced on top of the soil in the current tril (therefore llowing seeds to dry out rpidly following rinfll) s compred with eing covered with thin film of soil (vn Klinken et l., 2006). Seed uril depth (2 vs 10 cm) did not result in much chnge in dormncy relese, suggesting tht microclimtic differences etween the two depths were limited. This compres with the study in the wet-dry tropics, where seeds uried t 20 cm were exposed to tempertures tht were severl degrees cooler thn those uried t 3 cm, nd lso remined dormnt for much longer (vn Klinken et l., 2006). Bet-hedging Few or no seeds remined fter 1 yer in high rinfll uplnd hitts, despite considerle within-popultion vrition etween seeds in their sensitivity to wet het (vn Klinken nd Flck, 2005). There ws therefore no evidence for trde-off etween germinting when conditions re optiml nd delying some seed germintion in order to protect ginst high inter-yer vriility in recruitment proilities or with reproductive filure (et hedging; Evns et l., 2007). This compres with other species where most seeds cn still persist fter 1 yer (Bker et l., 2005; Thompson et l., 2003), nd is contrry to common perceptions tht P. culet, nd wide rnge of other species with hrd-seeded dormncy, hs long-lived or persistent seed nks (Bskin nd Bskin, 1998). However, it does support the prediction tht iteroprous perennils will gin little dvntge from dormncy mechnisms tht spred dormncy relese cross yers, ecuse risk is lredy spred through nnul seed production (Evns et l., 2007). Persistent seed nks were, however, pprent in some environments. This ws most notle in wetlnds t Betrice (wet-dry tropics) where 9 % of seeds were still dormnt fter lmost 4 yers, with little reduction in seed dormncy in the prior yer despite n intervening wet seson. However, herceous regrowth following seed uril ws not removed during our tril. The oserved dormncy-relese pttern is therefore likely to hve een the consequence of the micro-environment ecoming incresingly uffered y herceous regrowth, which ws especilly dense in the Betrice wetlnd fter the first yer. Similrly, greter persistence fter 1 yer of seeds uried in utumn (Mrch to April) is likely to e the result of herceous regrowth prior to the onset of the following wet seson. Seed uril trils rrely control for such chnges in microclimte through time, nd my therefore frequently overestimte seed persistence under optiml conditions for germintion.
264 vn Klinken et l. Seed Dormncy Relese in Prkinsoni culet The results suggest tht wet het represents n excellent dormncy-relese mechnism for mximizing seedling estlishment proilities, t lest for dry-tropic species such s P. culet. Persistent seed nks do occur, ut only in circumstnces where there is insufficient wet het for dormncy relese, which is lso likely to correspond with conditions tht re unsuitle or suoptiml for seedling estlishment. This includes sites where tempertures re highly uffered, such s under wter, in hevy shde or deep in the soil, or where conditions dry out rpidly, such s on the soil surfce (lthough most seeds generlly ecome incorported into the soil within 1 yer; R. vn Klinken, unpul. res.). Dormncy relese of persistent seeds therefore requires the microclimte to chnge either temporlly (e.g. through drying of wetlnds or the deth of prent trees) or sptilly (through secondry seed dispersl). Such environmentl seed refuges potentilly offer n lterntive mechnism to et hedging, which is property of the seed nd relies on trde-off etween men fitness nd vrince of fitness (Evns et l., 2007), to cope with nnul vrition in recruitment proilities. Aville evidence suggests tht such environmentl seed refuges re reltively common mong plnts (Fenner, 1995; Lmers et l., 2005; Bskin nd Bskin, 2006). Cution is clerly required when clssifying species ccording to seed nk strtegy (Csontos nd Tms, 2003) nd when considering the evolutionry significnce of dormncy mechnisms (Evns et l., 2007), without due considertion of the full rnge of ecologicl contexts in which those species my occur. 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