Phvsioloaical and aenetical -- ---- aspects ...
Phvsioloaical and aenetical
-- ----
aspects of mycorrhizae
SI
a
;
Aspects physiologiques
,
et _aér,étiaues
.-
‘
des mvcorhizes
Proceedings of the 1st European Symposium
on Mycorrhizae,
Dijon, l-5 July 1985.
Actes du Ier Symposium Européen
sur ies Mycorhizes,
Diion, l-5 juilie t 1985.
Editors/Editeurs
V. GIANINAZZI-PEARSON
S. GIANINAZZI
CNRS - INRA, Dijon
INSTITUT NATIONAL DE LA RECHERCHE AGRONOMIQUE
145-l 47 rue de l’Université, 75007 Paris
Mycorrhizae
: physiology andgenetics
- les mycorhizes : physiologie et g&&ique.
I*I ESMIl- SEM, Dijon, l-5 July 1985.- IIVRA, Paris, 1966
The development of extraradical hyphae in relation
to the response of cowpea to VA mycorrhizal infection
H.G. DIEM “‘, M. GUEYE and Y. DOMMERGUES “’
CNRS-ORSTOM, BP 1386, Dakar, SBndgal
CNRA-ISRA, Bambey, Sth5gal
INTRODtlCTlON
The principal way in which VA mycorrhizal fungi benefit plant
,mlo,dth is by increasing the volume of soi1 explored. Consequently an
< ffec! ive VA mycorrhizal fungus should be able to form an
extensive
:vtwoik of hyphae,called external or extraradical hyphae, in
t h e soi1
. n and beyond the rhizosphere. Thus Lt would be most useful to know the
suaLia distribution of VA mycorrhizal hyphae in soi1 or at least to
bave a measure of the weight or length of these hyphae. Very feu authors
have attempted to assess ths amount of extraradical hyphae and relate
it to the beneficial effect of mycorrhizal infection on the plant. Compa-
ring four mycorrhizal fungi, SANDERS et oi .(1977) found that three of
ihem with similar excernal mycelium equally innroved niant ,r,rowth and one
with little external mycelium produced no growth increase (onion). They
1
oncluded their study by stressing the vital role of the external hyphae.
ihich enhance nutrient uptake and transfer
to the system. Using soybean
~itTHI,ENFZILVA’f 6?r Ul. (1982) found that the progress of extraradical
Qïphoe reflected the plant growth enhancement to a much greater extent
. ban ‘bat of internal hyphae.
in the present communication we report observations on the time c o u r s e
C~I the intra- and extraradical development of ,$lomus mosseae infecting
t.owpea (C’igna unguiculata) and the simultaneous effect of the VA mycor-
rhizal infe&-tion on Ng fixation, growth, F and water content of the host.
We also attempt to reappraise the interpretation of the results obtained
vith the different methods of assessment of VA mycorrhizal infection.
MATERZAL ANQ METHOUS
Cowpea,
cv 58-185 from the National Centre of Agronomie Research
kambe y, Senegal,
was grown in pots filled with 1 kg of a sterile typical
Psamment soi1 (DECK soil). Details about the growth conditions a n d inocu-
lation are reported elsewhere (GUEYE et ol ., 1985). Two treatments were
compared: inoculation with Rhizobium (ORS407) alone (R) and dual inocu-
lation with Rhizobium and Clornus mosseae
(RN).
“--
(1) New address: BSSFT laboratory (CNRS/CTFT)
45 bis Avenue de la Belle Gabrielle 94130 Nogent-sur-Marne
each harvest, nodules, shoot dry weight,P and water content of shoots and
acetylene reduction activity were assessed. Root samples were removed
for infection assessment after clearing and staining with trypan blue in
lactophenol. Frequency (percentage of infected root pieces 0.3 mm in
length) and intensity (percentage of infected root volume) were then
measured according to OLLIVER et ~2. (1983). The length of extraradical
hyphae per cm of root was estimated as follows. After the entire root
system of each plant had been gently washed from the soil, randomly aelec-
ted samples of roots (ca 50 cm in total length) were stained with trypan
blue as above. Al1 the extraradical hyphae from the stained
roots were collected under a dissecting microscope and homogenized in
5ml of water at high speed. Then five 25 ~1 aliquots were removed and
deposited onto a thin layer of 1.5% sterile water agar in a petri dish.
After the liquid was completely absorbed, the agar layer bearing the
mycelium from a 25 ~1 aliquot was microscopically observed at low magni-
fication. Total length of the hyphae was evaluated using an eyepiece
micrometer and expressed as cm of hyphae per cm of root.
RESULTS
Intraradical infection of CLornus mosseae. The frequency percen-
tage was ;iil at day 5, but it increased rapidly at day 10 (39%) and was
close to maximal at day 15 (87%); then it remained at the highest level
(100%) up to the end of the experiment. The intensity percentage was ni1
at day 5; it increased rapidly from day 10 (6%) to day 15, remained at
the same level (41 - 53%) between day 15 and 3.5, then reached a plateau
(ca 75%) where ir stayed up to day 50 (Table 1).
?$traradicaldeuelopment
of Glomus mosseae. The pattern of extra-
. - -
radical hyphae length was quite differeni, The extraradical hyphae
started to develop significantly only at day 20, then they rapidly
reached a plateau (5 - 6 cm/cm of root) up to day 50 (Table 1).
( J
Nodulation and acetylene reduction activity. Nodule dry weight
)
of dually-inoculated plants was only ca 6 mg
per plant on day 15 but
it increaseed sharply to 35 mg on day 20 and reached a plateau (ca 100 w)
where it stayed up to day 50. Nodule dry weight of plants inoculated
.i
“:
with Rhizobium alone increased more slowly: from 5 mg on day 15 lt
reached only 15 mg on day 25 and then remained more or less constant
i’
around 50 .- 60 mg (Table 1). Similarly the increase of N -fixing activitp ”
expressed as ARA (acetylene reduction activity) was relagively more rapid
in dually inoculated plants than in non-mycorrhizal ones between day 15 ,’
and 25. Thus in mycorrhizal plants, nodulation and N
fixation appeared to
be markedly increased from day 20, which coincided &th the onset of thé:
extension of extraradical hyphae (Fig. 1). There was no evident
relation between the N -fixing activity or nodulatlon and the developmeat
of intraradical infect 2 on as measured by frequency or intensity index.
Plant grouth.
-
Surprisingly the beneficial effect of mycorrhizalx
infection was significant only from day 45 whereas the effect on nodu-‘.’
lation had occurred much earlier. The delayed response of plant growth ‘*
to mycorrhizal infection could be attributed to the large requirements
the nodules for P and perhaps to the C drain by Glomus mosaeue and
:
/
Rhizobium.
f.
sch t r e a r -
zrvals. A t
nhoot s and
removed
pan blue in
3 mm In
x t h e n
:r-aradical
ire root
Idomly selec-
gi.th t r y p a n
Ijzed i n
)ved and
!tri d i s h .
iag t h e
low magni-
repiece
icy percen-
:) and was
lest level
rge was ni1
lained a t
a plateau
.n o f extra-
*p’ha e
ridly
I.
I~:V raelght
iy 15 but
(ca 100 mg)
.ulated
5 tt
Instant
.ng activity
*
i,f P ContC
i,f non-my<
derlined 1
1 t rematnc
,,f *nycorr~
phases:
- a f i r s t
- a secont
enrichma
t:ion o f
- a third
a stabi
Figuu 1.
Jimc! CO~LS~ 04 actiylene kedution aca%ity (ARAI expuded
Fig 3 si-1
in rlanomolen CfH4 put pLant pti h in mycow&zaI and non-myc
ï,,ntent i
thizd cowpeoA.
mycorrhiz
pïobably
consider,
1980) hav
- MYCORRHIAL RANIS
scment o f
P inflow
t
------ NON WCORRMZAL PIANlS
extraradi
proceeds
. a first
level C
.
.
.
.
'
.
-
.
b
0
5
15
25
35
45
52
approx:
DAYS AFTER PlANTlNli
author:
- a sec0
Figwle 2.
Jime cowa 06 P content (%l 06 bhooti in mycowkizd and
ir.tcns
non-myc.ohhkizd
coupe.a.
indica
net t h
“intra
a t h i r
hqphae
cannot h
CO the e
the part
authors
wsentia
Thus i t
o f extr;
sssessmc
l-8
.
.
.
.
1
4
!5
1 5
25
35
4.5
52
presentt
DAM AFIER PLMlffi
- evalu.
F.igwte. 3.
T.i.me couhbe 06 wa;teh wnte.nt 06 nhooa2, i.tt mycot&izaR and
e t cil
non-mycotizd cowpeab.
- micro
or fr
2 3 1
P and water content of shoots
----~--_--. - - - - Fig. 2 shows that the time course
<,f P content
(P %l i n m y c o r r h i z a l planls d i f f e r e d strikingly
f r o m t h a t
. _
i> i non-mycorrhizal plants. In the non-mycorrhizal plants shoot P %
declined f o r 2 5 d a y s a f t e r planting,
then reached a low level at which
tt rcmained up t o d a y 5 0 . Instead o f t h e s e t w o p h a s e s , t h e time c o u r s e
of mycorrhizal plants exhibited a very different pattern with three
phases:
- a first phase (day 0 ta 20) duri.ng which P % decreased;
- a second phase (day 20 to 25) d ur ng which P
i
% rapidly increased, this
enrichment phase being concomitant and probably related
ta t h e exten-
tion of extraradical hyphae;
- a t h i r d p h a s e (day 2 0 ta 5 0 ) o f moderate decline o f P %, followed b y
a stabilization o f P % a t a relatively h i g h l e v e l .
F i g . 3 s h o w s that,during
the time interval between day 20 and 30,the water
cent ent i II mycorrhizal plants was significant ly higher than that of non-
mycorrhizal plants. The higher water content during this period is
prnbahly related ta the concurrent enhancement of P inflow, if we
consider,as a number of authors (e.g. SAFIR et aZ., 1972; LEVY and KRIKUN,
1980) bave, rhat water uptake and transfer are facilitatcd by the impro-
vement o f P n u t r i t i o n . We have already ment ioned thar the enhancement of
P inflow was probably the consequence of the extension of the netwnrk of
ext raradical h y p h a e o f CZonit~ moss~(le.
The data presented here suggests that VA mycorrhizal infection
proceeds in three steps:
- a f i r s t scep during Uhich t h e roots a r e p r o g r e s s i v e l y infected, t h e
l e v e l o f i n f e c t i o n
being measured by our “frequency” index, which
approximates the “infected root length” index used by a number of
aut hors;
- a second step during which the hyphae pervade the root cortex, the
intensity of thls process being measured by our “ i n t e n s i t y ” index.which
indicates the percentage of the cortex volume invaded by the hyphae and
not rhe root infected length. Our intensity index would approximate the
“intraradical biomass” as defined by BETHLENFALVAY et aZ. (1982);
- a third step during which the VA fungus develops its extraradical
hyphae throughout the soi1 and around the roots.
One should be aware of the fact that this succession of events
cannot b e generalized since t h e r e a r e p r o b a b l y l a r g e v a r i a t i o n s a c c o r d i n g
to the environmenral conditions and to the specific characteristics of
the partners of the symbiosis. However. there is one point on which most
a u t h o r s agree. a t l e a s t i n t h e o r y . that the extraradical hyphae are
essential i n promoting t h e g r o w t h o f m y c o r r h i z a l p l a n t s (TINKER, 19801.
Thus it is surprising that SO few papers have dealt with the evaluation
of extraradical hyphae. The main reason is probably that this type of
assessment i s very difficult.
A feu techniques have been proposed. In addition to the one
Presented here, let us quote the following:
- evaluation of the chitin content oE the rooting medium (BETHLENFALVAY
et aZ., 1982);
- microscopic measurement of hyphae extracted from the rhizosphere soi1
or from the roots (KUCEY and PAUL, 1982);
232
fb4ycorrhi~ae
’ physiol
131 ESM/î* SEM. Dijor
- weighing ?hc hyphae çollected by stripping the roots and wet-sieving
the rhizosphere soi.1 (SANDERS et ~2.) 1977).
Since these methods are time consuming, it is tempting to use
the regression analysis approach to obtain correlation between the amount
of extraradical hyphae and any other parameter whose assessment would be
Developmer
casier (e.g. KUCEY and PAUL, 1982). However, even if a relationship is
shown to bc relevant in a given situationit is not necessarily applicable
in sehi-nat&
in other circumstances.
Thus SANDERS et oZ.(1977)found that the dry weights
of extraradical hyphae were proportional to the total infected root
length in their experiment on onion. The results of the study we present
C.P.D. EiIRCH
here show that such a relation is not appropriate. Consequently it is
advisable ta assess the amount of extraradical hyphae using direct
Universify of Shefl
methods.
Sheffield, Si0 2Tb
It is interesting to compare the amount of extraradical hyphae as calcu-
lated by differents authors. If this characteristic is expressed as cm of
hyphae per cm of root, the figures published range between 80 cm (SANDERS
and TINKER, 1973) and 1.5 cm (SANDERS et ~2.. 1977). Thus the figure
obtained in our experiment falls within this range, even though it must be
an underestimation as a11 the extraradical mycelium cannot be recovered by
the method used. Nevertheless,our
data supports the idea that extraradical
hyphae play a major role by promoting the uptake of P and consequently
In the I
improving the water transport to the plant and markedly enhancing N
sent of VA myco
fixation in the case of N2-fixi’g plants. However,we have seen that 2there
environment diifc
may b e a delay between the Initiation of these processes and the initia-
b e expeqted t o
tion of the stimulation of the plant growth itself, which indicates that
roots of a number
other phenomena may interfer?, such a s the C and/or P drain by the
myceliw is pre.
endophytes.
new host roots.
the formation of
Finally it appears that thr study of VA fungal infection and of
of the progrers c
irs consequenccs o n Lhe hüst shou~d be carried olut: over a p e r i o d , i f
i n a n investigz
possi.ble throughout a g,rowth cycle, in order to give a sufficient insight
mycorrhiaal infec
into the symbiotic system.
BIBLIUGRAPW
In earl:
BETHLENFALVAY G.J., PACOVSKY R.S. and BROWN M.S. 1982. Phytopathology, 72, ‘I
LC s 1 h.2 .zinn,
F
894-897.
.Q
during t h e d a y
Carboniferous lj
GUEYE M., DIEM H.G. and DOMMERGUES Y.R. 1985. Mircen J. (in press).
Derbyshire, UK.
.t
species @as e x c a
KUCEY R.M.N. and PAUL E..A. 1982. Soi2 BioZ. Biochem., 14, 413-414.
days after plantil
examined microsc
LEVY Y. and KRIKUN J. 1980. New PhytoZ., 85, 25-31.
‘4
zones that are de:
:a
first 3 harvests
OLLIVIER B., BERTHEAU Y., DIEM H.G. and GIANINAZZI-PEARSON V. 1983.
:iE
length o f uninfeci
L'an. J. B o t . , 6 1 , 3 5 4 - 3 5 8 .
!
were recorded, so
SAFIR C.R.,
BOYER J.S. and GERDEMAN J.W. 1972. Plant Physiol., 49,
q
*a,
SANDERS F.E. and TINKER P.B. 1973. Pestic. Sci.. 4, 385-395.
Infectic
s p e c i e s except i
SANDERS F.E., TINKER P.B., , BLACK R.L.B. and PALMERLY S.M. 1977.
,L
developed rapidly
New Phytol., 78, 257-268.
; kY
days 1 the averagc
TINKER P.B. 1980. In: The role of phosphorus in agriculture, 617-654
little variation
(F.E. KAMPRATH et al. eds) American Society of Agronomy,
.F’‘i
P l a n t s a l r e a d y h
;is
-- ----
aspects of mycorrhizae
SI
a
;
Aspects physiologiques
,
et _aér,étiaues
.-
‘
des mvcorhizes
Proceedings of the 1st European Symposium
on Mycorrhizae,
Dijon, l-5 July 1985.
Actes du Ier Symposium Européen
sur ies Mycorhizes,
Diion, l-5 juilie t 1985.
Editors/Editeurs
V. GIANINAZZI-PEARSON
S. GIANINAZZI
CNRS - INRA, Dijon
INSTITUT NATIONAL DE LA RECHERCHE AGRONOMIQUE
145-l 47 rue de l’Université, 75007 Paris
Mycorrhizae
: physiology andgenetics
- les mycorhizes : physiologie et g&&ique.
I*I ESMIl- SEM, Dijon, l-5 July 1985.- IIVRA, Paris, 1966
The development of extraradical hyphae in relation
to the response of cowpea to VA mycorrhizal infection
H.G. DIEM “‘, M. GUEYE and Y. DOMMERGUES “’
CNRS-ORSTOM, BP 1386, Dakar, SBndgal
CNRA-ISRA, Bambey, Sth5gal
INTRODtlCTlON
The principal way in which VA mycorrhizal fungi benefit plant
,mlo,dth is by increasing the volume of soi1 explored. Consequently an
< ffec! ive VA mycorrhizal fungus should be able to form an
extensive
:vtwoik of hyphae,called external or extraradical hyphae, in
t h e soi1
. n and beyond the rhizosphere. Thus Lt would be most useful to know the
suaLia distribution of VA mycorrhizal hyphae in soi1 or at least to
bave a measure of the weight or length of these hyphae. Very feu authors
have attempted to assess ths amount of extraradical hyphae and relate
it to the beneficial effect of mycorrhizal infection on the plant. Compa-
ring four mycorrhizal fungi, SANDERS et oi .(1977) found that three of
ihem with similar excernal mycelium equally innroved niant ,r,rowth and one
with little external mycelium produced no growth increase (onion). They
1
oncluded their study by stressing the vital role of the external hyphae.
ihich enhance nutrient uptake and transfer
to the system. Using soybean
~itTHI,ENFZILVA’f 6?r Ul. (1982) found that the progress of extraradical
Qïphoe reflected the plant growth enhancement to a much greater extent
. ban ‘bat of internal hyphae.
in the present communication we report observations on the time c o u r s e
C~I the intra- and extraradical development of ,$lomus mosseae infecting
t.owpea (C’igna unguiculata) and the simultaneous effect of the VA mycor-
rhizal infe&-tion on Ng fixation, growth, F and water content of the host.
We also attempt to reappraise the interpretation of the results obtained
vith the different methods of assessment of VA mycorrhizal infection.
MATERZAL ANQ METHOUS
Cowpea,
cv 58-185 from the National Centre of Agronomie Research
kambe y, Senegal,
was grown in pots filled with 1 kg of a sterile typical
Psamment soi1 (DECK soil). Details about the growth conditions a n d inocu-
lation are reported elsewhere (GUEYE et ol ., 1985). Two treatments were
compared: inoculation with Rhizobium (ORS407) alone (R) and dual inocu-
lation with Rhizobium and Clornus mosseae
(RN).
“--
(1) New address: BSSFT laboratory (CNRS/CTFT)
45 bis Avenue de la Belle Gabrielle 94130 Nogent-sur-Marne
each harvest, nodules, shoot dry weight,P and water content of shoots and
acetylene reduction activity were assessed. Root samples were removed
for infection assessment after clearing and staining with trypan blue in
lactophenol. Frequency (percentage of infected root pieces 0.3 mm in
length) and intensity (percentage of infected root volume) were then
measured according to OLLIVER et ~2. (1983). The length of extraradical
hyphae per cm of root was estimated as follows. After the entire root
system of each plant had been gently washed from the soil, randomly aelec-
ted samples of roots (ca 50 cm in total length) were stained with trypan
blue as above. Al1 the extraradical hyphae from the stained
roots were collected under a dissecting microscope and homogenized in
5ml of water at high speed. Then five 25 ~1 aliquots were removed and
deposited onto a thin layer of 1.5% sterile water agar in a petri dish.
After the liquid was completely absorbed, the agar layer bearing the
mycelium from a 25 ~1 aliquot was microscopically observed at low magni-
fication. Total length of the hyphae was evaluated using an eyepiece
micrometer and expressed as cm of hyphae per cm of root.
RESULTS
Intraradical infection of CLornus mosseae. The frequency percen-
tage was ;iil at day 5, but it increased rapidly at day 10 (39%) and was
close to maximal at day 15 (87%); then it remained at the highest level
(100%) up to the end of the experiment. The intensity percentage was ni1
at day 5; it increased rapidly from day 10 (6%) to day 15, remained at
the same level (41 - 53%) between day 15 and 3.5, then reached a plateau
(ca 75%) where ir stayed up to day 50 (Table 1).
?$traradicaldeuelopment
of Glomus mosseae. The pattern of extra-
. - -
radical hyphae length was quite differeni, The extraradical hyphae
started to develop significantly only at day 20, then they rapidly
reached a plateau (5 - 6 cm/cm of root) up to day 50 (Table 1).
( J
Nodulation and acetylene reduction activity. Nodule dry weight
)
of dually-inoculated plants was only ca 6 mg
per plant on day 15 but
it increaseed sharply to 35 mg on day 20 and reached a plateau (ca 100 w)
where it stayed up to day 50. Nodule dry weight of plants inoculated
.i
“:
with Rhizobium alone increased more slowly: from 5 mg on day 15 lt
reached only 15 mg on day 25 and then remained more or less constant
i’
around 50 .- 60 mg (Table 1). Similarly the increase of N -fixing activitp ”
expressed as ARA (acetylene reduction activity) was relagively more rapid
in dually inoculated plants than in non-mycorrhizal ones between day 15 ,’
and 25. Thus in mycorrhizal plants, nodulation and N
fixation appeared to
be markedly increased from day 20, which coincided &th the onset of thé:
extension of extraradical hyphae (Fig. 1). There was no evident
relation between the N -fixing activity or nodulatlon and the developmeat
of intraradical infect 2 on as measured by frequency or intensity index.
Plant grouth.
-
Surprisingly the beneficial effect of mycorrhizalx
infection was significant only from day 45 whereas the effect on nodu-‘.’
lation had occurred much earlier. The delayed response of plant growth ‘*
to mycorrhizal infection could be attributed to the large requirements
the nodules for P and perhaps to the C drain by Glomus mosaeue and
:
/
Rhizobium.
f.
sch t r e a r -
zrvals. A t
nhoot s and
removed
pan blue in
3 mm In
x t h e n
:r-aradical
ire root
Idomly selec-
gi.th t r y p a n
Ijzed i n
)ved and
!tri d i s h .
iag t h e
low magni-
repiece
icy percen-
:) and was
lest level
rge was ni1
lained a t
a plateau
.n o f extra-
*p’ha e
ridly
I.
I~:V raelght
iy 15 but
(ca 100 mg)
.ulated
5 tt
Instant
.ng activity
*
i,f P ContC
i,f non-my<
derlined 1
1 t rematnc
,,f *nycorr~
phases:
- a f i r s t
- a secont
enrichma
t:ion o f
- a third
a stabi
Figuu 1.
Jimc! CO~LS~ 04 actiylene kedution aca%ity (ARAI expuded
Fig 3 si-1
in rlanomolen CfH4 put pLant pti h in mycow&zaI and non-myc
ï,,ntent i
thizd cowpeoA.
mycorrhiz
pïobably
consider,
1980) hav
- MYCORRHIAL RANIS
scment o f
P inflow
t
------ NON WCORRMZAL PIANlS
extraradi
proceeds
. a first
level C
.
.
.
.
'
.
-
.
b
0
5
15
25
35
45
52
approx:
DAYS AFTER PlANTlNli
author:
- a sec0
Figwle 2.
Jime cowa 06 P content (%l 06 bhooti in mycowkizd and
ir.tcns
non-myc.ohhkizd
coupe.a.
indica
net t h
“intra
a t h i r
hqphae
cannot h
CO the e
the part
authors
wsentia
Thus i t
o f extr;
sssessmc
l-8
.
.
.
.
1
4
!5
1 5
25
35
4.5
52
presentt
DAM AFIER PLMlffi
- evalu.
F.igwte. 3.
T.i.me couhbe 06 wa;teh wnte.nt 06 nhooa2, i.tt mycot&izaR and
e t cil
non-mycotizd cowpeab.
- micro
or fr
2 3 1
P and water content of shoots
----~--_--. - - - - Fig. 2 shows that the time course
<,f P content
(P %l i n m y c o r r h i z a l planls d i f f e r e d strikingly
f r o m t h a t
. _
i> i non-mycorrhizal plants. In the non-mycorrhizal plants shoot P %
declined f o r 2 5 d a y s a f t e r planting,
then reached a low level at which
tt rcmained up t o d a y 5 0 . Instead o f t h e s e t w o p h a s e s , t h e time c o u r s e
of mycorrhizal plants exhibited a very different pattern with three
phases:
- a first phase (day 0 ta 20) duri.ng which P % decreased;
- a second phase (day 20 to 25) d ur ng which P
i
% rapidly increased, this
enrichment phase being concomitant and probably related
ta t h e exten-
tion of extraradical hyphae;
- a t h i r d p h a s e (day 2 0 ta 5 0 ) o f moderate decline o f P %, followed b y
a stabilization o f P % a t a relatively h i g h l e v e l .
F i g . 3 s h o w s that,during
the time interval between day 20 and 30,the water
cent ent i II mycorrhizal plants was significant ly higher than that of non-
mycorrhizal plants. The higher water content during this period is
prnbahly related ta the concurrent enhancement of P inflow, if we
consider,as a number of authors (e.g. SAFIR et aZ., 1972; LEVY and KRIKUN,
1980) bave, rhat water uptake and transfer are facilitatcd by the impro-
vement o f P n u t r i t i o n . We have already ment ioned thar the enhancement of
P inflow was probably the consequence of the extension of the netwnrk of
ext raradical h y p h a e o f CZonit~ moss~(le.
The data presented here suggests that VA mycorrhizal infection
proceeds in three steps:
- a f i r s t scep during Uhich t h e roots a r e p r o g r e s s i v e l y infected, t h e
l e v e l o f i n f e c t i o n
being measured by our “frequency” index, which
approximates the “infected root length” index used by a number of
aut hors;
- a second step during which the hyphae pervade the root cortex, the
intensity of thls process being measured by our “ i n t e n s i t y ” index.which
indicates the percentage of the cortex volume invaded by the hyphae and
not rhe root infected length. Our intensity index would approximate the
“intraradical biomass” as defined by BETHLENFALVAY et aZ. (1982);
- a third step during which the VA fungus develops its extraradical
hyphae throughout the soi1 and around the roots.
One should be aware of the fact that this succession of events
cannot b e generalized since t h e r e a r e p r o b a b l y l a r g e v a r i a t i o n s a c c o r d i n g
to the environmenral conditions and to the specific characteristics of
the partners of the symbiosis. However. there is one point on which most
a u t h o r s agree. a t l e a s t i n t h e o r y . that the extraradical hyphae are
essential i n promoting t h e g r o w t h o f m y c o r r h i z a l p l a n t s (TINKER, 19801.
Thus it is surprising that SO few papers have dealt with the evaluation
of extraradical hyphae. The main reason is probably that this type of
assessment i s very difficult.
A feu techniques have been proposed. In addition to the one
Presented here, let us quote the following:
- evaluation of the chitin content oE the rooting medium (BETHLENFALVAY
et aZ., 1982);
- microscopic measurement of hyphae extracted from the rhizosphere soi1
or from the roots (KUCEY and PAUL, 1982);
232
fb4ycorrhi~ae
’ physiol
131 ESM/î* SEM. Dijor
- weighing ?hc hyphae çollected by stripping the roots and wet-sieving
the rhizosphere soi.1 (SANDERS et ~2.) 1977).
Since these methods are time consuming, it is tempting to use
the regression analysis approach to obtain correlation between the amount
of extraradical hyphae and any other parameter whose assessment would be
Developmer
casier (e.g. KUCEY and PAUL, 1982). However, even if a relationship is
shown to bc relevant in a given situationit is not necessarily applicable
in sehi-nat&
in other circumstances.
Thus SANDERS et oZ.(1977)found that the dry weights
of extraradical hyphae were proportional to the total infected root
length in their experiment on onion. The results of the study we present
C.P.D. EiIRCH
here show that such a relation is not appropriate. Consequently it is
advisable ta assess the amount of extraradical hyphae using direct
Universify of Shefl
methods.
Sheffield, Si0 2Tb
It is interesting to compare the amount of extraradical hyphae as calcu-
lated by differents authors. If this characteristic is expressed as cm of
hyphae per cm of root, the figures published range between 80 cm (SANDERS
and TINKER, 1973) and 1.5 cm (SANDERS et ~2.. 1977). Thus the figure
obtained in our experiment falls within this range, even though it must be
an underestimation as a11 the extraradical mycelium cannot be recovered by
the method used. Nevertheless,our
data supports the idea that extraradical
hyphae play a major role by promoting the uptake of P and consequently
In the I
improving the water transport to the plant and markedly enhancing N
sent of VA myco
fixation in the case of N2-fixi’g plants. However,we have seen that 2there
environment diifc
may b e a delay between the Initiation of these processes and the initia-
b e expeqted t o
tion of the stimulation of the plant growth itself, which indicates that
roots of a number
other phenomena may interfer?, such a s the C and/or P drain by the
myceliw is pre.
endophytes.
new host roots.
the formation of
Finally it appears that thr study of VA fungal infection and of
of the progrers c
irs consequenccs o n Lhe hüst shou~d be carried olut: over a p e r i o d , i f
i n a n investigz
possi.ble throughout a g,rowth cycle, in order to give a sufficient insight
mycorrhiaal infec
into the symbiotic system.
BIBLIUGRAPW
In earl:
BETHLENFALVAY G.J., PACOVSKY R.S. and BROWN M.S. 1982. Phytopathology, 72, ‘I
LC s 1 h.2 .zinn,
F
894-897.
.Q
during t h e d a y
Carboniferous lj
GUEYE M., DIEM H.G. and DOMMERGUES Y.R. 1985. Mircen J. (in press).
Derbyshire, UK.
.t
species @as e x c a
KUCEY R.M.N. and PAUL E..A. 1982. Soi2 BioZ. Biochem., 14, 413-414.
days after plantil
examined microsc
LEVY Y. and KRIKUN J. 1980. New PhytoZ., 85, 25-31.
‘4
zones that are de:
:a
first 3 harvests
OLLIVIER B., BERTHEAU Y., DIEM H.G. and GIANINAZZI-PEARSON V. 1983.
:iE
length o f uninfeci
L'an. J. B o t . , 6 1 , 3 5 4 - 3 5 8 .
!
were recorded, so
SAFIR C.R.,
BOYER J.S. and GERDEMAN J.W. 1972. Plant Physiol., 49,
q
*a,
SANDERS F.E. and TINKER P.B. 1973. Pestic. Sci.. 4, 385-395.
Infectic
s p e c i e s except i
SANDERS F.E., TINKER P.B., , BLACK R.L.B. and PALMERLY S.M. 1977.
,L
developed rapidly
New Phytol., 78, 257-268.
; kY
days 1 the averagc
TINKER P.B. 1980. In: The role of phosphorus in agriculture, 617-654
little variation
(F.E. KAMPRATH et al. eds) American Society of Agronomy,
.F’‘i
P l a n t s a l r e a d y h
;is