FINAL PROJECT REPORT Project Title: Mechnicl pollintion for yield security PI: Mtthew Whiting Orgniztion: WSU Telephone: 59 786-926 Emil: mdwhiting@wsu.edu Address: 2416 N. Bunn Rod City/Stte/Zip: Prosser/WA/9935 Coopertors: OnTrget Spry Systems, Firmn Pollen Co., Olsen Brothers, Hyden Frms, Russ LeSge; Json Mtson; Alln Brothers Totl Project Request: Yer 1: 3,123 Yer 2: 31,86 Percentge time per crop: Apple: 25% Per: 25% Cherry: 5% Stone Fruit: % Other funding sources Notes: In kind support is provided y Firmn Pollen Co (pollen dontions) nd OnTrget Spry Systems (dontion of spryer nd technicin for support) Budget 1 Orgniztion Nme: WSU Contrct Administrtor: Crrie Johnston Telephone: 59 335-4564 Emil ddress: crriej@wsu.edu Item 214 215 Slries 13,28 13,549 Benefits 1,173 1,219 Wges 9,36 9,734 Benefits 562 584 Equipment Supplies 3, 3, Trvel 2,5 2,5 Plot Fees 5 5 Miscellneous Totl 3,123 31,86 Footnotes:
OBJECTIVES: 1. Ensure consistent fruiting in sweet cherry, pple, nd per through the development of n effective mechnicl pollintion system. 2. Pursue further funding to uild upon this project s outcomes SIGNIFICANT FINDINGS Artificil pollintion with pollen suspensions pplied electrostticlly is cple of setting fruit in pple, per, nd sweet cherry We documented the deposition of mechniclly-pplied pollen to the stigmtic surfce in sweet cherry incresing pollen density per stigm y out 3-fold compred to open pollinted A single supplementry ppliction of pollen suspension cn improve fruit set Replcement pollintion tests (i.e., no pollen pplied other thn vi our system) re promising in pple nd sweet cherry tree yield ws similr from two pplictions of pollen compred to open pollinted Inconsistent nd very wek correltion found etween flower density, fruit set, shoot length, shoot ngle nd shoot dimeter Including sucrose nd oron in pollen suspension medi cn improve pollen viility for more thn 1 hr Pure pollen is required for our pollintion system Pure pollen cn e prepred on commercil scle for rtificil pollintion systems Percentge of pollen germintion vried for sme sucrose concentrtion in medium, cross the genotypes used in current study Miniml vrition of pollen deposition vrition y flower loction in the tree ws noticed, receiving electrosttic pollen suspension spry It is fesile to incorporte growth regultor ReTin to id ginst premture ovule senescence in sweet cherry, without further ffecting stisfctory germintion rte in pollen suspension Sweet cherry stigms exhiit high receptivity on the second dy fter loom RESULTS AND DISCUSSION This project hs evluted the potentil to pollinte nd fertilize tree fruit flowers with pplictions of liquid pollen suspensions through commercil electrosttic spryers. This reserch project hs comined smll-scle l/reserch plot trils with lrger-scle field trils. Our proposed precision pollintion system is comprised of 3 steps: 1. Collect nd purify pollen 2. Suspend pollen in liquid 3. Apply pollen through electrosttic ppliction system This reserch project hs ddressed the second nd third spects directly, nd we hve worked in collortion with Firmn Pollen Compny to ddress pollen collection nd purifiction. 1. Pollen collection nd purifiction.
Through our field trils nd l nlyses of suspension mterils, we lerned tht it is necessry to utilize pure pollen in our pollintion system. Field trils in 214 reveled prolems with rehydrtion of non-pollen florl prts (e.g., filment, nthers) tht cused filters in the spryer to clog rpidly. The stndrd pollen tht is used commercilly for hive inserts or to e pplied through dusting mens contins too mny of these non-pollen mterils to e useful for field ppliction through spryer. We investigted the potentil to modify filtrtion systems in the On Trget Spry Systems spryer ut it ecme cler tht further pollen purifiction would e necessry. Firmn Pollen Compny hs developed n dditionl filtrtion step tht yields pure pollen. Our field tests in 215 utilized this purified pollen product, nd we hd no difficulties with filtrtion. Pollen collection systems will need to e improved to keep up with demnd. The current process of hnd-hrvesting flowers in commercil orchrds is too lorious nd time-consuming. This is lrgely due to the pickers inility to hrvest ll ville flowers the process is selective ecuse flower must remin for the grower to hrvest crop. This criticl issue is ddressed in our new proposl. Orchrds should e plnted strictly for pollen collection this will require investigtion of new trining systems (or modifiction of existing systems) to optimize pollen yield. 2. Suspend pollen in liquid In 214 we conducted l studies on pollen viility nd the role of suspension medi components. From our tests of three sweet cherry pollen genotypes (Lpins, Rinier, Sweethert), we found tht we could improve pollen viility (i.e., germintion) when pollen ws suspended in modified medium, compred to wter. For ech genotype, the following suspension mediums were evluted: solution 1: 5% (w/v) sucrose +.15% (w/v) pollen grin, solution 2: 1% sucrose (w/v) +.15% (w/v) pollen grin, solution 3: 15% (w/v) sucrose +.15 % (w/v) pollen. Pollen germintion declined over time in the control tretment with only wter. This occurred for ll cultivrs similrly, losing out 2% germintion from initil suspension to 6 minutes lter. In contrst, the suspension solutions improved pollen germintion nd extended viility over time. There ppers to e enefit to higher rtes of sucrose, with viility improved more thn 6% fter one hour in suspension. Suspension medium contining 15% sucrose (w/v) improved germintion significntly in ll pollen genotype irrespective of how long the pollen ws in suspension. This is likely due to the fvorle osmotic lnce in the medium with sucrose. In contrst, we documented pproximtely 15-2% decrese in the viility of pollen grins in control etween the first nd lst smpling time. The highest pollen germintion ws oserved with Sweethert (67%) followed y Rinier (59%) fter 4 minute suspension time in medi enriched with 15% sucrose (w/v). In 215, pollen suspension development continued with multiple experiments crried out to investigte the role of cndidte suspension components on pollen viility. For our in vitro pollen germintion experiments, we utilized one sweet cherry, pple nd per genotype (Rinier, Red Delicious nd Brtlett). Cultivr selections were sed on most suitle nd comptile pollen genotypes to source pollen for our field tril pollen receiving cultivrs. In every cse, pure pollen ws utilized. Cndidte suspension components were selected from literture review of relevnt scholrly rticles. Initilly we evluted pollen viility fter incution of 1 hour t concentrtion grdient of ech the components. Afterwrds, we ssessed comintions of components on pollen viility. In ech cse we used.18% w/v of pollen for the experiments. Replicte tues of suspension mterils were creted, pollen ws dded, nd the tues were gitted riefly (<5 sec) efore incuted t room temperture. Suspension liquots were withdrwn fter 5, 3, nd 6 minutes nd pollen germintion ws evluted to ssess viility. Following suspension, our modified medium mintined or significntly improved pollen germintion percentge compred to only wter, irrespective of pollen genotypes nd cultivrs. Recent findings reveled tht, using sucrose t lower rte could mintin fvorle osmotic lnce retention with optiml pollen
Germintion (%) Pollen germintion (%) viility, either lone or in comintion of other components in suspension, cross ll three genotypes. 6 5 4 3 2 1 7 6 5 4 3 2 1 Control Suspension medi + Regulid Suspension medi + Ad- Wet 9 Control Suspension medi w/o surfctnt Suspension medi + Regulid Suspension medi + Ad Wet 9 Figure 1: Effects of suspension medi on sweet cherry (left) nd pple (right) in vitro pollen germintion fter 6 minutes in suspension (oth p-vlues:.). Pollen + wter lone is not n effective comintion. First, pollen is not esily wettle (i.e., difficult to suspend); secondly, pollen losses viility in wter. Germintion rte of pollen suspended in wter consistently decresed over the incution periods, nd reched minimum fter 6 minutes. We evluted cndidte suspension components individully nd in comintion for their effect on pollen germintion. After 5 minutes in suspension with wter + single suspension ingredient, severl suspension components improved germintion rtes were 151% nd 144% of control for Red Delicious pollen. Similr results were found with cherry nd per pollen. We then comined suspension constituents t their optimum concentrtion nd found improved pollen germintion (26% of control). After 3 minutes of pollen in suspension, we found gret improvements in pollen germintion. Severl suspension components incresed germintion y 22% to 23% of control. The gretest improvements in pollen viility were seen fter 6 minutes of pollen incution, when our suspension improved pollen germintion rtes tht were more thn three-fold greter tht the control of wter + pollen. Our comined suspension medi improve pollen germintion y more thn 4x compred to the control fter 6 minutes (Figure 1). Future work is needed to trnsition from l-scle tests to lrge-scle suspensions in commercil ppliction equipment. L tests need to e repeted in the field, collecting pollen tht hs een loded in suspension in the spry tnk, nd spryed through the system. We do not know how the ppliction system will ffect pollen viility, nor how esily suspensions will e mintined under constnt gittion, nor the effect of gittion on the suspension nd pollen viility. In ddition, suspension cretion in the l is firly stright forwrd using l gittion tools (e.g., vortex system), in our field trils of 215 we encountered some difficulty in creting uniform suspension in the spryer tnk. This process needs further development.
Pollen depositon (%) In vivo pollen deposition We lso evluted pollen deposition to the stigm using cut lims of Rinier sweet cherry in the l. In vivo pollen deposition tests were crried out on emsculted flowers ws crried out with two pollen rtes pplied t 2 nd 3 dys fter loom (DAB) under greenhouse conditions. Flowering lims were positioned verticlly in uckets of wter. Using single-nozzle electrosttic ppliction 3 25 2 15 1 1x rte 2DAB 1x rte 3DAB 2x rte 2DAB 2x rte 3DAB 5 Fcing spryer 9 degee right 9 degee left 18 degree fcing wy Figure 2: Percent pollen deposition fter mechnicl pollintion only, ccording to the totl numer of pollen collected from emsculted sweet cherry flower stigm t 2 nd 3 dys fter loom under controlled condition. Pollen deposition represented s the verge of 5 flowers from ech of five replicted flowering lims, i. e. for ech tretment n=25/direction, per smpling dy. setup we pplied two rtes of pollen in suspension to replicte lims moving the nozzle pst the lims t out 2 mph (to mimic ppliction in the field). Post ppliction, smple flowers from two different loom dtes were collected nd the deposition of pollen on the stigmtic surfces ws ssessed microscopiclly on flowers tht fced the spryer s well s those tht were 9 degrees wy, nd those tht were 18 degrees from the spryer (i.e., on the ck of the lim). Pollen deposition ws consistently higher when pollen ws pplied t 2DAB, t oth pollen rtes (Figure 2). Highest pollen deposition on flower stigms fcing nozzles ws found from doule rte (3g/cre) of pollen pplied t 2DAB (279%), nd lowest from single rte (15g/cre) pplied t 3DAB t similr ngle. Miniml differences oserved in pollen deposition on flowers stigms locted t 9 o from the direction of the spry t oth 2DAB nd 3DAB pplictions, seprtely, with either pollen rte. However, t this ngle flowers from 2DAB hs shown 125% higher deposition compred to 3DAB. Flowers locted t stright ngle (18 o fcing wy from the spry) received lowest pollen deposition (18%), when single rte pollen pplied t 3DAB. These results support our l s previous studies of stigmtic receptivity which reveled mximum receptivity of sweet cherry stigms on flowers tht were open for two dys. Field trils In 214 we conducted severl field trils to evlute the potentil for improving fruit set with supplementl pplictions of pollen s well s replcing the use of pollenizers nd pollintors (i.e., replcement pollintion). Cherry field trils were estlished in n Erly Roin orchrd (Psco) nd two Tieton orchrds (Roosevelt nd Benton City). We re unle to collect dt from the Erly Roin nd Roosevelt Tieton orchrds due to difficulties with the ppliction system. We discovered tht our pollen mixture ws clogging the spryer s filter nd we hd troule with the nozzles getting
plugged too. We ttriuted this to two fctors: the qulity of the pollen used nd the chrcteristics of the pollen suspension medi used. We hve lerned tht only pure pollen will work with the current ppliction system ecuse nthers nd filment mteril commonly present in stndrd offthe-shelf pollen mixes will hydrte rpidly nd plug the filters nd nozzles. In ddition, the pollen suspension medi we used ws too viscous nd contriuted to the clogging of the spryer. This medi ws provided y PollenTech, strtup compny trying to develop mechnicl pollintion systems. We did not use their proprietry slurry in 215 trils. In our third field tril, we were le to use pure pollen, in the Tieton orchrd in Benton City, nd the system performed well. Our supplementl pollen pplictions t out 5% nd 9% of full loom incresed fruit set y 15% compred to openpollinted trees (Fig. 1). The ppliction system ws clerly effective t plcing pollen in suspension on the stigmtic surfce we documented nerly 3-fold increse in pollen deposition on treted stigms compred to open-pollinted stigms (Figure 3). We lso conducted full replcement pollintion (i.e., pollen pplied through ee exclusion netting) tril, pplying pollen suspension twice (out 25% nd 75% full loom) to Bing trees. The rtificil pollintion system ws s effective s nturl open pollintion (Fig. 2). Interestingly, yield ws less vrile mong trees tht were pollinted rtificilly compred to those open pollinted trees. Furthermore, in Gl pple tril of replcement pollintion to lims covered with netting, we recorded very high fruit set from electrosttic pollen ppliction. We evluted two pollen suspension solutions nd recorded pproximtely 56% nd 75% higher fruit set over nturl pollintion (see continuing report from 215 for these results). In 215 we estlished four sweet cherry trils were designed t multiple loctions including Tieton (Grndview), Erly Roin (Psco), Benton (Prosser) nd Regin (Brewster). We lso conducted one pple tril nd one per tril (D Anjou ner Nches). Ech of these trils ws supplementl pollintion we pplied pollintion tretments to trees or entire rows in orchrds where oth pollenizers nd pollintors were present. In 9 th lef Tieton orchrd trined to verticl UFO rchitecture, supplementl pollen ppliction using single pollen rte of 15g/cre, with two pplictions (once t 5% nd gin t 1% loom, incresed fruit set y 1% compred to nturl pollintion. In this orchrd we recorded 2 to 3 fold increse in pollen deposition (Figure 1) on flower stigm spryed with our liquid pollen suspension compred to nturlly pollinted stigms. We hve lso noticed tht pollen deposition ws superior in treted thn non-treted stigms, irrespective of flower position nd trining systems of cultivrs under investigtion.
Fruit set (%) Pollen deposition (%) 6 5 4 214 215 18 16 14 12 Supplementry-Deposition Nturl-Deposition 3 2 1 8 6 1 4 2 Supplementl pollintion Nturl pollintion F R L O Figure 3. Effect of supplementl pollintion (two pplictions) on fruit set in Tieton sweet cherry (oth yer p-vlue:.41) (left) nd pollen grin deposition on the stigm (yer 215; p-vlue:.) (right). F=Fcing spryer, R nd L= 9 o from the direction of spryer right nd left, respectively, nd O= 18 o from the direction of spryer. In Y-trellised Erly Roin orchrd we evluted pollintion efficcy of two ppliction timings nd two rtes of pollen. Supplementl pollintion ws mde t out 5% nd 1% full loom. Ech supplementl pollintion tretment improved fruit set. The gretest response ws in response to pplying the high pollen rte (3g/cre) pplied twice this tretment incresed fruit set y 65% compred to nturl pollintion. Including the growth regultor ReTin in the spry did not improve fruit set (our l tests reveled no toxic effect of ReTin on pollen viility). Single pplictions of pollen were similrly effective the timing of ppliction nd the rte of pollen mde no difference. In ech cse, fruit set ws increse y out 4%.
Fruit set (%) 1 9 8 7 6 5 4 3 2 1 c Nturl pollintion 1x rte 5%FB 2x rte 5%FB 1x rte 1%FB 2x rte 1%FB 2x rte 5% FB + 1%FB 2x rte ReTin 5% FB + 1%FB Figure 4. Fruit set in Erly Roin sweet cherry upon two supplementl pollen suspension pplictions (t 5% nd 1% loom) with single nd doule rte nd nturl pollintion (nonsupplementl) (p-vlue:.). In Regin orchrd ner Brewster single ppliction of 15g pollen ws pplied t out 75% full loom. This supplementl pollintion tretment improved fruit set y out 2% (Figure 5). The improvement in fruit set (nd susequently yield) from our supplementl pollintion tretment ws enough to convince the orchrdist to keep the orchrd (this lock ws set to e removed due to poor production). Figure 5. Fruit set (% of ville flowers) for open-pollinted trees nd those treted with supplementl pollintion t c. 75% full loom nd 15 g/cre of pollen.
Figure 6. Fruit set (% of ville flowers) in D Anjou per s ffected y pollintion tretment. 1x rte = 15 g/cre, 2x = 3 g/cre. Supplementl pollintion tretment to D Anjou per resulted in significnt improvements in fruit set. Nturl fruit set in this orchrd ws less thn 15% of ville flowers (Figure 6). Two pplictions of pollen t roughly 5% nd 1% full loom improved fruit set y 2% nd 15% for single nd doule pollen rtes, respectively. Interestingly, our ppliction of 1x rte (15 g/cre) ws more effective thn the doule rte. This underscores the importnce of further investigtion into the pollen rte response. It will e prudent to utilize s little pollen s necessry to set the desired crop since pollen will e in short supply. We propose to evlute fertiliztion rte response to pollen rte in sweet cherry nd pple in new proposl. In Jzz/M9 lock we treted trees with two pollen rtes (15 or 3 g/cre) t roughly 5% nd 1% full loom. We lso treted rows where mnul lossom thinning ws complete, leving only one flower per cluster in comprison to rows tht were unthinned. Both unthinned nd thinned fruiting lims of Jzz pple receiving replcement pollintion hd fruit set despite the sence of ny eemedited pollintion (Figure 7). We documented no fruit set in untreted nd gged lims wheres lims treted with our pollintion system exhiited fruit set rtes etween out 5% nd 65% (unthinned lims) or 4% nd 55% (thinned lims). The doule rte induced 17% higher fruit set compred to single rte. Percent of open flower t pollen ppliction ws sme in thinned nd unthinned lock. This is further evidence tht we re le to fertilize flowers successfully with our pollintion system. This my e prticulrly importnt when growers go to the expense of hndthinning flowers. It will e criticl to fertilize ech remining flower, nd this my e ccomplished with rtificil pollintion. Our vision of precision pollintion systems in the sence of pollintors nd pollenizers hs the potentil to revolutionize crop lod mngement if we re le to successfully pollinte only portion of the flowers.
Fruit set (%) Fruit set (%) 8 7 6 5 4 3 2 1 Unthinned 1 Thinned 1 6 5 4 3 2 1 Control Single Doule Control Single Doule Figure 7. Fruit set in Jzz /M 9 pple rnches upon pollen suspension tretment with two pollen rtes- single (15g/cre) nd doule (3g/cre), pplied to unthinned (left) nd thinned (right) flowering lims t out 5% nd 1% full loom. All treted lims including control were enclosed y ee oxes throughout flowering (p-vlue:.). 1 Percent of open flower t pollen ppliction ws sme in thinned nd unthinned lock.
EXECUTIVE SUMMARY This reserch project hs provided preliminry dt nd proof of concept dt on the possiility to pollinte tree fruit crops rtificilly with pollen suspensions pplied with commercilly ville electrosttic spryers. Herein we report on successful pollintion of pple, per, nd sweet cherry using pollen suspension tht is cple of mintining or improving pollen viility for up to 1 hour. We hve proven tht one cn tke previously hrvested pollen, incorporte it in suspension, nd spry it through commercilly ville spryers to effect pollintion in tree fruit crops. The implictions re considerle ensuring consistent fruiting in the fce of declining ee popultions, incresingly vrile spring wether conditions, nd the perennil chllenges growers fce with oth pollintors nd pollenizers. Further, the ility to rtificilly pollinte tree fruit holds the potentil to revolutionize crop lod mngement. Our vision for precision pollintion systems tht do not include pollenizers nor pollintors ppers plusile. Clerly these results should e considered promising yet preliminry. More reserch is needed into the role of pollen rte, ppliction timing, nd suspension development to extend pollen viility. We hve collorted with interested growers cross Wshington, Firmn Pollen Compny, nd On Trget Spry Systems ll to fcilitte the commercil doption of our reserch. As result of this reserch, there is significnt interest in precision pollintion systems. Every grower we worked with in 215 is interested to collorte further, seeing promising results this underscores the importnce of pollintion/fertiliztion in commercil fruit production s well s the confidence our collortors hve in our reserch pproch nd vision.