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Hydroponic culture of the mycorrhizal fungus Glomus mosseae with Linum

usitatissimum L., Sorghum bicolor L. and Triticum aestivum L

Article  in  Plant and Soil · September 1997

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Hydroponic culture of the mycorrhizal fungus Glomus mosseae with Linum usitatissimum L.,
Sorghum bicolor L. and Triticum aestivum L.
Author(s): H.-J. Hawkins and E. George
Source: Plant and Soil, Vol. 196, No. 1 (September 1997), pp. 143-149
Published by: Springer
Stable URL: http://www.jstor.org/stable/42948158 .
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PlamandSoil196:143-149,
1997. 143
© 1997KluwerAcademic
Publishers.
Printed
intheNetherlands.

Hydroponie culture of the mycorrhizal fungus Glomus mosseae with Linum

usitatissimum L., Sorghum bicolor L. and Triticum aestivum L.

H.-J. Hawkins and E. George1

Institute
ofPlantNutrition
(330), Hohenheim , 70593 Stuttgart
University , Germany author*
; Corresponding

Received
3 April
1997. inrevised
Accepted form
5 August
1997

Keywords:arbuscular
mycorrhiza, wheat
linseed,sorghum,
hydroponics,

Abstract

Linumusitatissimum , Sorghumbicolorand Triticum aestivumplantswerefurther colonisedby the arbuscular

mycorrhizal fungus,Glomusmosseae, duringa fourweekperiodofhydroponie cultureaftera pre-culture period
of threeweekswiththefungusin perlitesubstrate. The viabilityof mycorrhizal colonisationof T aestivumwas
indicatedby an initialexperiment whereG. mosseaefrommycorrhizal plantscolonisednon-mycorrhizal plants
whentheplantswere growntogether in the same hydroponie containerusingmodifiedLong Ashtonnutrient
solution.Intermittantaerationoftheplantroots(2 h periods,fourtimesperday)provideda compromise between
adequate aeration and minimaldisturbance of thefungus.In a secondexperiment, twonutrient media,modified
Long Ashtonandmodified KnopplusHoaglandmediumwerecomparedforculturing G. mosseaeon T. aestivum .
A significantly
higher root dryweight was found for the mycorrhizal versus thenon-mycorrhizal wheat plantsin
modifiedLong Ashtonnutrient medium,whichcontained10 ¡¿M P and an organicbuffer. ModifiedKnop plus
Hoaglandnutrient mediumcontaineda highP concentration (0.9 mM) and did notproduceviable culturesof
mycorrhizal colonisation. In a third experiment, modified Long Ashtonmediumwas used forhydroponie culture
of mycorrhizal L. usitatissimum , S. bicolor and T. aestivum.The root colonisation percentages for T. aestivum
(73%), 5. bicolor(36%) and L. usitatissimum (65%) werewithintherangeof colonisationratesobtainedwith
solid substrate
culturein perlite.Viabilityof themycorrhizal structures in hydroponie culturewas assessedby
monitoring of
activity fungal succinate dehydrogenase and found to be similarto culturesin perlite.No difference
in theP concentration of mycorrhizal and non-mycorrhizal plants was observed,possiblyowingto thelack of
diffusionlimitsforP in hydroponie solution.This reportdescribesa systemfortheviablecultureof G. mosseae
withdifferentplantspecieswherea highmycorrhizal colonisationratewas producedunderconditions of a short
cultureperiodusingintermittent aeration, a low concentration ofP supplyandan organicbuffer.

KHNS - modified
Abbreviations: Knop andHoaglandnutrient solution,LANS - modified
Long Ashtonnutrient
MES - /3-morpholino-ethane
solution, sulfonicacid,SDH - succinatedehydrogenase.

Introduction (Ruiz-Lozanoet al., 1995) andphytohormone balance

(Duan et al., 1996). The mechanismsinvolvedare
Therootsofmostplantspeciesareoftencolonisedwith to elucidatedue to limitations
difficult in theculture
endomycorrhizal fungi when grown in natural soils. of
systems mycorrhizas.
Thissymbiotic relationship has existedsincetheearly Therehas beenno successin cultivating endomy-
Devonian(Tayloret al., 1995).Endomycorrhizas play corrhizalfungiindependently of thehostplantunder
a rolenotonlyin nutrient transfer toplants(Georgeet axenicconditions. It is possiblehowever,to produce
al., 1995) butalso inplantresistance againstsoil-borne largeamounts of sporesandmycelium fromcolonised
pathogens (Newsham et al., 1995),drought resistance shearedroots (Diop et al., 1994) and colonised trans-
* FAXNo:+4971 formed root cultures (St. Arnaud et al., 1996). These
14593295.E-mail:george@uni-hohenheim.de

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144

methodswill contribute substantially to understand- colonisedplantswereplaced either2 to 4 cm apart

ingtheregulation of thesymbiosisbutare at present or theplanthypocotyls wereplacedtogether in foam
restrictedto a fewhostplant-fungal isolatecombina- rubber.Colonisation ofthemycorrhizal andoriginally
tions.To date,mostphysiologicalstudiesof mycor- non-mycorrhizal plantswas determined at theend of
rhizashave come fromsoil-based,wholeplantsys- thehydroponie cultureperiod.In the secondexper-
tems.These studiesare limitedin thatratesof nutri- iment,mycorrhizal and non-mycorrhizal plantswere
entsupplyat therootsurfacecannotbe manipulated culturedinseparatehydroponie containerswhileinthe
easily and the rootand funguscannotbe harvested thirdexperiment onlymycorrhizal plantswereused.
without significant damage.Also,suchrootsareoften The secondexperiment testedtwo nutrient solutions
unsuitableformolecularstudies,whichrequiresoil- (LANS and modified Knop plus Hoagland nutrient
less preparations, or forstudying nutrient metabolism solution,KHNS, see below)in termsof suitability for
ofroots. hydroponie culture of T. aestivum with G. mosseae.
Mycorrhizal plantscanalso be cultured inaeropon- The nutrient solutionsdiffered withrespectto P con-
ics (HungandSylvia,1988; SylviaandHubbel,1986) centration,micronutrients and buffering source.The
and in a nutrient flow system(Mosse and Thomp- thirdexperiment testedthe system threedif-
culture on
son, 1984). Aeroponiccultureof mycorrhizal plants ferentplantspecies colonised with G. mosseaeusing
has beencommercially used in nurseries (Jarstferand LANS: Linumusitatissimum L. cv. Atlante(linseed),
Sylvia, 1995). While requiring some investment in Sorghum bicolorL. cv.Tesa (sorghum) andT.aestivum
materials,both of thesesystems can be useful forstudy- L. cv.Hano.The plantsandfungusweregrownunder
ingnutrient utilisation by mycorrhizal roots.In addi- the same conditionsin all experiments as described
tionto aeroponicand nutrient flowculturesystems, below.
hydroponie culture of mycorrhizal plantsprovidesa
controlled nutrient environment andallowstheharvest Plantandfungusculture
of mycorrhizal plantsfreefromsoil forthepurpose
of physiologicaland molecularstudies.Hydroponics Seeds were pre-germinated overnightin 7.5 mM
are,however,infrequently referred to as a methodfor CaSC>4andplanteddirectly into150 ml potscontain-
and
growing studying mycorrhizal plants. ingperliteandinoculum( 10% ofpotvolume)orperlite
Hydroponiecultureof mycorrhizalfungi was andinoculumfiltrate paper,Schleich-
(Blue Bandfilter
reportedfirstby Peuss (1958) forGlomusmosseae er and SchuellLtd.,Dassel, Germany)and grownfor
withNicotiana tabacum. Mycorrhizalplantswere threeweeks.Perlite(1-3 mm)was preparedbywash-
grownalso in nutrient solutionculturebyCresset al. ingin tapwaterovera 1 mmmeshsieve,sun-drying
(1979; 1986) and Karunaratne et al. (1986). Recent- and thenheattreating for24 h at 110°C. Perlitewas
ly,Dugassa et al. (1995) reported a cultureofGlomus mixeduniformly withtheinoculum(formycorrhizal
intraradices withLinumusitatissimum. However,G. plants)or theinoculumfiltrate (fornon-mycorrhizal
mosseaeremainsuncharacterised withrespecttonutri- plants). The mycorrhiza inoculumcomprisedloess
entsupplyandfungalviability in hydroponie culture. soil containing G. mosseaesporesandZea maysroot
pieces,where50% oftherootlengthwas colonisedby
G. mosseae.The funguswas originally isolatedbyE.
Materialsand methods Sieverdingfroma Luvisol nearGöttingen, Germany
(51° 30' N, 9° 55' E) andpropagated severaltimeson
Experiments maize grownon loess soil in a greenhouseforeight
weeks.Plantsin potswereirrigated dailywith40 ml
Threeexperiments were conducted.The firsttested LANS (see below).Afterthreeweeks,therootlength
andviability
theinfectivity ofGlomusmosseae(Nicol, andpercentage colonisedrootweredetermined inaddi-
andGerd.)Gerd,andTrappewhencolonisingTriticum tionaltestplants.Plants were transferred
thereafter into
aestivumL. cv.Hano(wheat)inhydroponics. Thiswas hydroponics tobe cultivatedforanotherfourweeks. In
accomplishedby growingcolonisedplantstogether orderto transplant, thepotsweresubmerged in deion-
withinitiallyuncolonisedplants(froma pre-culture ized water,therootscarefully separatedfromtheper-
periodin perlite,see below) in thesame hydropon- liteand theplantsplaced intoblack 5 L hydroponie
ie containercontainingmodifiedLong Ashtonnutri- containers (fiveplantspercontainer). Rootlengthand
entsolution(LANS, see below).The uncolonisedand percentage colonised rootwere determinedagainatthe

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 145

timeofharvest.Rootsoftheindividualplantscouldbe everythreedays. Aerationof the nutrient solutions

easilyattheendofthegrowth
separated period,except was suppliedfroman air pumpvia aquariumstones
inthecase ofsorghum which
roots(thirdexperiment), thatproduceda finestreamof bubbles.The airpump
wereseparatedwitha needle.Shootsand rootswere was connectedto a timeswitchso as to aeratefor2 h
harvestedfordryweightand P concentration deter- periods,fourtimesperday.
minationaftersamplingformycorrhizal colonisation.
The experiments werecarriedoutin a growthcham- determination
Phosphorus
berwitha PAR 500-600/zmol m-1 s-1 (Mercury
of
halidelamps,OsramPowerstar HQI-T-2000W/D),a Phosphorusconcentrationwas determinedusingthe
day/nighttemperatureof 25 °C/ 18°C and a relative molybdo-P blue assay (Murphy Riley,1962) on
and
humidityof60%. milled
dried, shoot thathadbeendry
androotmaterial
ashed.
Nutrient
media
and rootlength
rootcolonisation
Mycorrhizal
The plants in perliteculturewere supplied with
1/100LANS (Hewitt,1966) forthefirstthreedays Percentage ofrootlengthcolonisedbyG. mosseaeand
in all experiments and thereafter with a modified totalrootlengthweredetermined usingthegridline-
half strengthLANS, which consistedof the fol- intersect method(Giovannetti andMosse, 1980) with
lowing macronutrients (mM): Ca(NC>3)2.4H20(2); rootsstainedeitherin trypanblue (Koske and Gem-
NaH2P04.2H20 (0.0094); Na2HP04.12H20 (0.006); ma, 1989) or for succinatedehydrogenase activity
K2S04 (1); MgS04.7H20 (0.75); CaCl2.2H20 (2) (Schaffer and Peterson, 1993). The lattermethod is
andmicronutrients (ļiM): H3BO3 (69); MnS04.4H20 a counterstaining procedureinvolving first
staining for
(10.4); ZnS04.7H20 (1.2); CuS04.5H20 (1.7); fungalsuccinatedehydrogenase (SDH) activityand
NaMo04.2H20 (0.13) and FeEDDHA (0.3). A pH of thenforall fungalstructures using0.01% (w/v)acid
6.0 was maintainedby 0.15 mM MES-KOH. Long fuchsin in75% lacticacidand5% glycerol(Kormanik
Ashtonnutrient solutionhas beenusedpreviously as a and McGraw,1984). The purpleformazanpigment,
nutrientsolutionforsandculture ofmycorrhizal plants formedwhenelectronsare donatedto thenitro-blue
(Cressetal., 1979;Davis et al., 1992). tetrazolium saltin theassaymediumbySDH activity,
PlantsweresuppliedwitheitherLANS or KHNS is visibleagainstthepinkfuchsin background andindi-
(Dugassa et al., 1995) forhydroponie culturein the cates whererespiratory activityhas occurred.In this
secondexperiment. Molybdenum was added toKHNS way themethod,herereferred to as counterstaining,
since Mo is requiredfor fungalgrowth(Epstein, distinguishes betweenviable and non-viable mycor-
1972). The KHNS nutrient solutionconsistedof the rhizalstructures (Schafferand Peterson,1993). Sam-
following macronutrients (mM): Ca(N03)2.4H20 (2); ples forthedetermination ofmycorrhizal colonisation
KNO3 (1.2); KH2P04 (0.9); KCl (0.8); MgS04. 7H20 wererepresentative ofthewholeroot.
(0.5) and micronutrients (/iM): A12(S04)3(0.18); KI
(0.2); KBr (0.25); Ti02 (0.75); SnCl2.2H20 (0.13); Statistics
LiCl (0.71); MnCl2.2H20(2.0); H3BO3 (9.7); ZnS04
(0.21); CuS04.5H20 (0.24); NiS04.6H20 (0.23); Tenreplicatespertreatment wereusedin thefirstand
Co(N03)2.6H20 (0.21); NaMo04.2H20 (0.13) and secondexperiments andeightin thethirdexperiment.
FeEDTA (0.1). The pH of thesolutionwas 6.5. The Plantcontainers wereplacedrandomly in thegrowth
pH was checkedeverythirddayinbothofthenutrient chamberand thepositionof thecontainerswas var-
solutions(LANS andKHNS) andifnecessary, adjust- ied eachtimeafterreplacingthenutrient solution.The
ed with2 N HCl. The nutrient solutionswerechanged Student's andone-wayanalysis
t-test of variance were
wheneithertheN concentration ortheP concentration appliedto determine differencesdue to typeof nutri-
droppedbelow0.6 mM or5 ¡jlMrespectively inLANS entsolutioninthesecondexperiment andmycorrhizal
and 1 mM or 0.45 mM respectively in KHNS. The N colonisationoftheplantspeciesinthethird experiment
and P concentrations weremeasuredusingtheNC>3~ respectively.Percentagerootcolonisationdata were
Quiektest(Merck)and molybdo-Pblue assay (Mur- normalised byarcsinsquareroottransformation before
phyand Riley,1962) respectively. The nutrientsolu- performing one-wayanalysis of variance. Results in
tionshad to be changedafterone weekandthereafter thetablesand textare given± standard deviation.

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146

Results Afterfourweeksof hydroponie culturein LANS

medium, the total root colonisation percentage
Firstexperiment remained highcomparedto theinitialpercentage root
colonisedinperlite(Table1). Plantswhichweregrown
Therootlengthofmycorrhizal wheatafterthreeweeks inhydroponics withKHNS mediumhada muchlower
in perliteculturewas 3.3 ± 0.6 m. The averageini- totalrootcolonisation percentage. ofthis
The viability
tial colonisationof mycorrhizal in
plants perlite cul- root colonisation on KHNS medium was essentially
turewas 59.6 ± 6% forboththe firstand second zero as determined by SDH activity(formazanpre-
experiments, theplantsof whichweregrownat the cipitation)in thecounterstaining procedure(Table 1).
same time(Table 1). Non-mycorrhizal wheatplants In comparison,theviablerootcolonisationpercent-
whichwereplaced in close associationwithmycor- age on LANS accountedforoverhalfofthetotalroot
rhizalplantsin hydroponics had a totalcolonisation colonisation.
percentage of 8.8 ± 3.6% afterfourweeks.Morethan Colonisationof plantswas testedusingthecom-
halfofthetotalcolonisation was viableas determined monlyappliedtrypanblue methodto comparewith
bythecounterstaining method(4.87 ± 0.27%). Those thecounterstaining method(Table 1). The significant
non-mycorrhizal plantsculturedinthe same hydropon- difference between resultsusingthetwo methodsin
ie tankbutspaced2 to 4 cm fromthenearestmycor- the perliteculture(p = 0.0395, Studentt-test)was
rhizalplantwerenotcolonised. mostlikelya resultof theslightoverestimation due
to background staining, which can occur when using
Secondexperiment thecounterstaining method.
Shoot and root P concentrations were not sig-
The shoot dry weightsof mycorrhizaland non- nificantlydifferent between mycorrhizaland non-
mycorrhizal plantsgrown in eitherLANS or KHNS mycorrhizal plantswhen plantsweregrowninthesame
werenotsignificantly (Table2). Mycorrhizal nutrient
different medium(Table2). The P concentration in the
plantsgrown in LANS medium had a significantly shoots and rootsof mycorrhizal and non-mycorrhizal
higherrootdryweightcomparedto thecorrespond- plantsgrownon KHNS mediumwas greater compared
ing non-mycorrhizal plants(p < 0.001). No signif- to thosein LANS.
icantdifference in rootdryweightbetweenmycor-
rhizalandnon-mycorrhizal plantswas observedwhen Thirdexperiment
KHNS mediumwas used.Afterfourweeksin LANS
and KHNS media,therootlengthsofthemycorrhizal In thethirdexperiment whereonlymycorrhizal plants
plantswere13.6db3.9 m and 11.0 ± 2.6 mrespective- wereused,theplantdryweightdecreasedintheorder:
ly. S. bicolor> T. aestivum> L. usitatissimum (Table 3).

Table1. Viable
andtotal ofT.aestivum
rootcolonisation with
colonised G.mosseaeas
determined method
bya counterstaining andtotal bythe
as determined
rootcolonisation
bluemethod.
trypan Theplantsweregrowninitially forthree
inperlite weeks in
andthen
for
hydroponics weeks.
four Letters statistical
indicate between
difference media
hydroponie
(p< 0.05,Student's Dataarethemean
t-test). deviation
± standard
of10observations
Culture (%root
Colonisation length)
method
Counterstaining Trypanblue
method
Viable Total : non- Total
Viable
ratio colonisation
colonisation colonisation viable
(initial 31.18± 7.70 67.98± 8.46 0.97±0.52 59.61± 6.09
Perlite
colonisation)
Hydroponics
LANSmedium36.00a ±5.19 1.19a
± 3.60 66.36a ±0.11 56.11a
±13.06
KHNSmedium 0.60b±0.46 10.70" ± 5.64 0.05"±0.03 8.66b±3.32

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 147
Table
2. Dryweight
andphosphorus ofT.aestivwn
concentration after
plants three
weeks inperlite
ofpre-culture andsubsequent
culture
for wee
four
intwodifferent nutrient
hydroponie media(LANSandKHNS)with (+AM)orwithout(-AM)arbuscular colonisation
mycorrhizal byG.mossec
indicate
Letters statistical
difference +AMand-AMshoots
between androots
within
eachnutrient (p < 0.05,Student's
treatment Dataareť
t-test).
mean ± standard
of10and8observations for
deviation dry andP concentration
weight respectively
~
Dryweight
(g) (mgg 1dry
P concentration weight)
Shoot Root Shoot Root
Medium -AM +AM -AM +AM -AM +AM -AM +AM
LANS 0.43± 0.15a 0.54± 0.12a 0.09± 0.02a 0.17±0.04b 0.27± 0.12a 0.24± 0.06a 0.17± 0.09a 0.16± 0.07a
KHNS 0.58± 0.15a 0.67± 0.16a 0.13± 0.04a 0.17± 0.06a 1.70± 0.35a 1.39± 0.45a 1.30± 0.33a 0.94± 0.53a

S. bicolorhad a higherroot:shootratiocomparedto thirdexperiments and (c) by colonisationof initially

T. aestivumandL. usitatissimum. The averageinitial roots
non-mycorrhizal by adjacentmycorrhizal roots
colonisationof mycorrhizal plantsin perliteculture in the firstexperiment. Only thosenon-mycorrhizal
was 48 to 76% (Table 3). Viable colonisationwith rootswhichwerein directcontactwiththerootsys-
G. mosseaewas maintained overa fourweekperiodin temsofmycorrhizal plantswerecolonisedwhilethose
hydroponics forall threespecieswithtotalcolonisation non-mycorrhizal plantswhichwere2 to 4 cm away
percentages between 36% and73% (Table3). A higher frommycorrhizal plantsinthesamecontainer werenot
percentage of viable infective was foundin
structures colonised.Thisindicatesthatexternal hyphaegrowing
T. aestivumcomparedto S. bicolorand L. usitatissi- in nutrientsolutiondid notgrowaway fromtheroot
mum, whilethetotalrootcolonisationpercentages in butwereattachedto therootsurfaceor thosehyphae
T. aestivumandL. usitatissimum werehigherthanin growingawayfromtherootcouldnotcoloniseother
S. bicoloras determined bythecounterstaining proce- roots.
dure. The colonisationofwheatwithG. mosseaeresult-
ed in a higherroot dryweightcomparedto non-
mycorrhizal plantswhengrownon LANS mediumin
Discussion thesecondexperiment (Table2). The P concentration
in non-mycorrhizal and mycorrhizal plantsgrownin
Thefungalstructures inthemycorrhizal rootsofwheat LANS was similar(Table 2). Therefore, thegrowth
secondandthirdexperiments),
(first, sorghum andlin- stimulation in therootsof themycorrhizal plantsrel-
seed (thirdexperiment) were active. This is shown ativeto thenon-mycorrhizal rootswas probablynot
ratesofrootsevenafter
by (a) hightotalcolonisation relatedto increasedP uptakebut to non-nutritive
vigorousrootgrowth innutrient
solution,(b) measure- fungus-mediated effectson, forexample,shoot:root
mentsof viable rootcolonisationin the secondand carbohydrate allocation.Glomusmosseaemaintained

3.Drymassandviable
Table plustotal
colonisation method)
(counterstaining ofS.bicolor
, T.
aestivwn
andL. usitatissimum
colonisedwith
G.mosseae grown inperlite
initially forthree
weeksandtheninhydroponics
forfourweeksinLANSmedium. Letters
indicate
statistical
difference
between (p< 0.05,pair-wise
thespecies range
multiple Tukey Dataarethe
Test).
mean ± standard
of8 observations deviation
Colonisation
(%rootlength)
Culture S.bicolor T.aestivum L. usitatissimum
Perlite total
(initial 48± 6.02
colonisation) 75±2.13 76± 5.45
Hydroponics
Viable
colonisation 23.24a
± 11.02 48.33b± 16.80 22.07a±18.18
Total
colonisation ± 10.20 73.13b
36.34a ± 14.40 65.46b± 5.88
Viable:non-viable
ratio 1.49a±0.78 1.80a±0.88 0.72b± 0.70

Plant
dry
weight
(g) 1.16a±0.42 1.04a± 0.20 0.54b±0.16

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148

a colonisation of66% in wheatplantsgrowninhydro- but also sorghumand linseedin hydroponie culture

ponics using LANS but a muchlowercolonisation (Table 3). Wheat had a higher rootcolonisation per-
whenKHNS mediumwas used (Table 1). The KHNS centagecomparedto sorghum andlinseed.The range
mediumwas used in thecultureof L. usitatissimum ofcolonisation ratesinall threeplants(36 to73%) was
withG. intraradices(Dugassa et al., 1995). The P within therangeofreported colonisationsinsolid,low-
concentration in theKHNS mediumwas as described P substrates. Considering that G. mosseae colonised
in theoriginalKnop plus Hoaglandnutrient solution threedifferent in
plantspecies hydroponics, itis like-
et
(Urbach al., 1983), thisbeing 0.9 m M P, which rep- ly that the method described here would be usefulas
resentsa 90-foldincreasecomparedtoLANS medium a basis forthecultivation of otherplantspecieswith
( 10 ļiM). ThesolutionP concentration wasmeasured at G. mosseae.The suitability of thismethodforgrow-
regular intervals (see Materials and methods) to avoid ingmycorrhizal fungi other than G. mosseaemustbe
depletionof P below a certainthreshold leadingto testedin furtherexperiments.
uncontrolled periods of P deficiency (McDonaldetal., A difference comparedto previousreportsof the
1996). A lower P concentration was used inLANS and cultureofmycorrhiza in hydroponics, besidestheuse
thisP concentration is also within therangeofconcen- oflow P concentration andthebuffer MES, is theuse
trations (1 to 50 (lM P) usuallyfoundin soil solutions of a longeraerationperiod.The totalaerationtime
(Barber,1995). The organicbufferMES (buffering of 8 h was longerthantheperiodsuggestedbyPeuss
range:5.8-6.5) was used to maintaina stablepH due (1958) and Dugassa et al. (1995), i.e. 10 and 30 min
to thelow P concentration usedin LANS. The P con- daily,respectively.The latterauthorsaeratedforshort
centration in KHNS mediumwas sufficient to buffer periods in orderto reducetheamountof movement
pH at 6.5 (50 mM to 0.5 mM KH2PO4 solution plus in the nutrient mediumwhichwould disturbfungal
NaOH buffers betweenpH 5.8 and 8.0) and as a P growth. The aerationconditions describedhere,which
source.Most likely,thepoor colonisationof wheat werechosenas a compromise betweengood aeration
withG. mosseae usingKHNS mediumunderthese andreducedwatermovement, apparently had no neg-
experimental conditions was due totherelatively high ativeeffect on rootcolonisation bythefungusrelative
P concentration in thismedium,whichinhibited root to previousreports. Thoroughaerationof thenutrient
colonisation. solutionmaybe necessary toprevent plantinjuriesdue
There was no statisticaldifference betweenthe to hypoxia(MorardandSilvestre,1996).
P concentration in non-mycorrhizal and mycorrhizal In conclusion,theseresultsoutlinea methodfor
wheatwhengrownin eitherLANS or KHNS media thecultureof threeplantspecieshavingvaryingroot
(Table2). Sinceplantsgrownin KHNS nutrient medi- morphologies in a dilutednutrient mediumwithinter-
umwerepractically freeofactivemycorrhiza afterfour mittent aerationandpH control usingan organicbuffer
weeksin hydroponics (Table 1), a lack of difference (inLANS). Viablecolonisation oftherootswas shown
intheP concentration ofplantswithandwithout myc- to be greaterin LANS comparedto KHNS medium.
orrhizalinoculation was to be expected.Plantsgrown The colonisationlevels and viabilityof culturesin
in LANS mediumwere,however,heavilycolonised. LANS compared favourably tosolidsubstrate cultures.
Presenceof mycorrhizal colonisationin hydroponics Hydroponie cultureofendomycorrhizas is ofpotential
was,in contrast to manyobservations in soil,without use forinvestigating theeffectsof endomycorrhizas
consequencefortheP acquisitionof theplants,tis- on nutrient metabolism and energybalancein plants,
sue P concentration beinglow in bothcontrolandtest as well as forthe purposesof harvesting rootsand
treatments. Thereasonforthismaybe that,incontrast fungalpropaguleswithout contamination froma solid
to thesituation in soil,P in nutrient solutionsis freely substrate.
available.Thiswouldmake,forexample,thegrowth of
external hyphaeredundant intermsofP acquisition of
themycorrhizal root.A particular drawbackofhydro- Acknowledgements
ponics(and aeroponics)is thatthenutrients thatare
relatively immobilein thesoil arefreelyavailableand The firstauthorthankstheFoundationforResearch
so themechanismby whichmycorrhizal hyphaenor- and Development(SouthAfrica)fortheawardof a
mallyimpartbenefit to thehostwouldbe neutralised. doctoralbursary
during ofwhichthepresent
thetenure
UsingtheLANS mediumin thethirdexperiment, was
investigation out.
carried
G. mosseae was shownto colonisenot onlywheat

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