Ptant Pathotogy (1994) 43, 554-561

Biological control of peach twig blight {Monilinia laxa) with

Epicoccum nigrum

C. MADRIGAL, S. PASCUAL andF. MELGAREJO.

Departamento de Proteccion Vegetat, CIT-INIA, Apartado 8.111, Madrid 28080, Spain.

Various preparations of spores and mycelium of the antagonist Epicoccum nigrum, alone or in
combination with Captan, were applied in four different field trials to peach trees inoculated with
Monitinia taxa. the cause of twig blight. Biocontrol obtained after application of £^. nigrum was variable
each year, depending on the releative disease severity in the first 2-3 weeks after infection and the
climatic conditions. In three out of four trials, treatments with E. nigrum were comparable with Captan
in reducing disease severity and combinations of E. nigrum and Captan gave a level of disease
suppression similar to that given by the antagonist or chemical alone. In the remaining trial,
combinations of E. nigrum and Captan were necessary to obtain successful disease suppression.

INTRODUCTION peach shoots. The pathogenicity of this isolate to

Twig blight, caused by Monitinia taxa. can be
severe in peach-growing areas of Spain under
certain conditions (M.-Sagasta, 1977). Control
of the disease is not always satisfactory, and new
alternative control methods are being sotight.
Biological control using Penicittium frequentans
has recently been reported (De Cal et ai, 1990).
The potential of Epicoccum nigrum. a compo-
nent ofthe resident mycofiora of peach twigs and
flowers (Melgarejo et ai. 1985), for biological
control of M. laxa in field orchards has been
demonstrated elsewhere (Melgarejo et ai, 1986).
E. nigrum has been used previously in experi-
ments on biocontrol of other diseases (Boland &
Inglis, 1988; Zhou & Reeleder, 1989, 1990). This
paper describes attempts to develop an effective
and practical method of controlling M. taxa in
peach orchards by using E. nigrum alone or in
combination with the fungicide Captan or with
certain nutrients. A glasshouse experiment to
determine the infiuence of M. taxa inoculum type
on control of twig blight by E. nigrum is
presented.
MATERIALS A N D METHODS
Isolates and inoculum preparation
A monosporic isolate of M. taxa, strain ZA-I
(stock culture in ATCC), recovered from apricot
(Zaragoza, Spain) was used in inoculations of
peach was shown in inoculation experiments
(Melgarejo et ai, 1986). An isolate of £. rugnim
(isolate 282), obtained from healthy peach twigs
in Madrid, Spain, and shown to be antagonistic
to M. taxa in vitro and in vivo (Melgarejo et al.,
198S; Madrigal et ai, 1991), was used in all
experiments. Both fungi were stored on potato-
dextrose agar (PDA) slants at 5°C.
Inoculum of M. laxa consisted of disks of 1 cm
diameter from 7-day-old PDA cultures. In the
glasshouse test, suspensions of 10^, 10* or lO*
conidia in Czapek's solution were also used.
Conidial stispensions of M. laxa were obtained
by gently washing 7-day-old PDA cultures with
Czapek's salt solution and adjusting the solu-
tions to the above-mentioned concentrations.
Three preparations with £. nigrum were used:
mycelial plugs, conidia + mycehum (prepared
with Tween 80 or Nu-Fihn-17, a commercial
preparation of 96% di-menthane from Miller
Chemical and Fertilizer Co., Hanover, PA,
USA), and conidia + mycelium + nutrients
(three different nutrients were used). Disks of
E. nigrum on PDA were obtained in the same
way as those of M. ta.\a (mycelial plug).
The conidia + mycelium preparation of
E. nigrum was obtained from 10-day-old flask
cultures ofthe fungus grown on potato-dextrose
broth (PDB) at 2O-2S°C. PDB was decanted to
separate out the mycelium and spores, then
sterile sand and 10 ml of 0 1 % Tween 80 or
0 06% Nu-Film-17 were added. Flasks were then
 Biological control of peach twig blight
shaken by hand. The fungal suspension was
separated from sand by filtration through two
layers of sterile cheesecloth and was adjusted to
lO' conidia (plus mycelial fragments) per ml by
adding 0 1 % Tween 80 or 0 06% Nu-Film-17. A
cotiidia + mycelium + nutrients preparation
was made by adding nutrients to the solutions of
conidia + mycelitun in 0-1% Tween 80 or in
006% Nu-Fihn-17. Three different nutrient
solutions were used. N| consisted of malt extract
(10 g/1) and yeast extract (3 g/1) prepared in 0-1 %
Tween 80. N2 consisted of malt extract (10 g/1),
yeast extract (1 g/1), vitamin C (0 01 g/1), KNO3
(2 g/1) and trace element solution (1 tnl/l) pre-
pared in 0 06% Nu-Film-17. The trace element
solution consisted of Na2B4O7 • 10 HjO (100 mg/
1), (NH4)6MO7O2-4 H2O (10 mg/l), ZnSO4-7
H2O (70 mg/l), FeSO4-7 H2O (50 mg/l),
MnSO4-4 H2O (lOmg/1), CuSO4-5 H2O
(lOmg/1) and Nu-Film-17 (0 06%). N3 consisted
of lactose (20g/1), KNO3 (lOg/l) prepared in
0 06% Nu-Film-17. The N3 nutrient mix
enhances the growth and sporulation of E.
nigrum but not that of M. taxa (De Cal et ai,
1993).
Glasshouse experiment
One-year-old peach trees, cv. Baby Gold, were
grown in sand: peat: soil (1:1:3 by volume) in
25 X 20 X 35-cm pots in a glasshouse.
Inoculation with M. laxa was performed as
described by De Cal et ai (1990). Inoculum was
applied to a small incision (3-4 mm long, 1 mm
wide, 0-5 mm deep) in the bark, 2-3 cm from the
base of the shoot and the shoot was then
wrapped in moist cotton wool and aluminum
foil, which were removed after 2 days. Seven
shoots of approximately the same diameter
(0 5 cm) and length (10-15 cm) on each of 25
trees were inoculated with one of four different
inoculum types of M. laxa on 28 March (total of
100 trees). Inoculum types were allocated at
random to the trees. Inoculum types were: PDA
culture disks (as above), or suspensions of 10^
10* or 10' conidia/ml in Czapek's salt solution.
Five additional trees were uninoculated with
M. laxa (treatment 16, see Table 1). Treatments
1, 3, 7 and 15 (Table I) and treatment 17
(inoculation of a culture disk of E. nigrum to the
wounded section of each shoot that was then
wrapped in moist cotton wool and aluminium
foil for 7 days) were applied to five trees in each
of the inoculation groups described above.
Treatments were allocated at random. Treat-
555
ments 1,3 and 7 were applied by spraying to run-
off to shoots inoculated with M. taxa.
Treatments with E. nigrum or Captan were
first applied before inoculation with M. laxa and
this was followed by four similar applications
made weekly, except in the case of treatment 17.
All treatments were applied at sunset to favour
the development of E. nigrum. During the
experiment, trees received no other crop protec-
tion treatments.
At 10, 20 and 40 days after inoculation with
M. laxa. the length of lesions on shoots was
measured. The extent of Af. laxa colonization on
shoots was detertnined at the end of the
experiment by cutting shoots into 5-cm pieces
from the inoculation point to the end of the
shoot. Pieces were surface-sterilized as described
by Sauer & Burroughs (1986) and plated on
PDA. After incubation in the dark at 25°C for 1
week, the presence or absence of the pathogen in
each piece was recorded. The 5-cm piece from
which the pathogen was recovered and was the
farthest away from the inoculation point indi-
cated the extent of fungal colonization in each
shoot.
Field trials
Field trials were carried out over four growing
seasons from 1988 to 1991 (trials 1-4) in a peach
orchard, cv. Baby Gold, planted in 1984 and
located in Madrid, Spain. Shoot bark was
wounded as described above and then inocu-
lated with M. taxa on 20 May (trial 1). 23 May
(trial 2) and 17 May (trials 3 and 4). After
inoculation the wounded section of each shoot
was wrapped in moist cotton wool and alumi-
nium foil for 5 days before further treatment.
Seven shoots of approximately the same dia-
meter (0 5cm) and length (10-15cm) on each of
six trees were exposed to each treatment.
Treatments were first applied the day before
inoculation, with M. taxa, and applications then
were repeated seven times at 4-day intervals in
trials 1 and 2, four times at 7-day intervals in trial
3, and five times at 7-day intervals in trial 4 All
treatments were applied at sunset to favour the
development of E. nigrum. The orchards did not
receive any other treatment during these experi-
ments.
Shoots inoculated with M. laxa were treated
by spraying to run-off. Treatments were as
indicated in Table 1.
Length of lesions induced by M. laxa was
measured at 6, 12, 21, 35 and 50 days after
556 C. Madrigal et aL
Table 1. Treatments applied to shoots in field experiments*

Control group Treatment No. Trial Description

Biological 1 2 Conidia + mycelium + Tween 80

2 3,4 Conidia + mycelium + Nu-
film-17
3 1,2 Conidia + mycelium + nutrient
solution 1
4 2 Alternating treatment 3/no
treatment
5 3,4 Conidia + mycelium + nutrient
solution 2
6 4 Conidia + mycelium + nutrient
solution 3
Chemical 1 1,2,3,4 Captan (1 3 ga.i./l)
8 4 Only one application of treatment 7
9 2 Captan (0 65ga.i./l)
Integrated 10 1 Alternating treatment 3 and 7
11 2 Alternating treatment 3 and 8
12 3 One application of treatment 7
followed by applications of
treatment 2
13 4 One application of treatment 7
followed by applications of
treatment 5
Others 14 2 Nutrient solution 1
15 1,2,3,4 No treatment
16 1,2,3.4 Uninoculated and untreated

"Some treatments were also applied in glasshouse experiments.

inoculation with the pathogen. The extent of RESULTS

shoot colonization by M. taxa mycelium was
determined at the end of each experiment as
described above.
Statistical analyses
Data obtained from the glasshouse experiment
were subjected to a factorial analysis of variance
and means were compared by a least significant
difference (LSD) test at P = 005 when treat-
ment effects were found to be significant.
Treatment 7 (Captan) and treatment 16 (unin-
oculated and untreated control) were excluded
from analyses.
Results {"rom each trial ofthe field were analysed
independently at the end of the experiment by
contrast with the Ftest at significance levels of 0-1,
005 and 001 (Snedecor & Cochran, 1980).
Treatment 17 (uninoculated and untreated con-
trol) was excluded from analyses.
Glasshouse experiment
Disease development, estimated by lesion length
data, was rapid on shoots inoculated with
mycelial plugs of M. laxa for 10 days after
inoculation, and then slowed during the next 30
days. However, with cotiidial inoculum (10* and
10 conidia/ml) disease development was more
rapid from 10 to 40 days after inoculation than
during the first 10 days.
Data presented in Table 2 are from assess-
ments of the extent of pathogen colonization at
40 days. The analysis of variance resulted in
significant interaction (P = 0 05) between treat-
ments and type of inoculum.
The mycelial plug and lO' conidia ml inocula
resulted in less pathogen colonization than 10* or
lO' conidia/ml inocula when there was no further
treatment (control treatment). However, the type
of M. laxa inoculum did not affect the extent of
 Biological control of peach twig btight 557
Table 2. Effects of various methods of inoculating with Monitinia taxa and of treatments with shools on potted trees
in the greenhouse'

Epicoccum/Hgri/m/Captan treatments

conidia + mycelium +
M. taxa inoculum none conidia + mycelium nutrient solution mycelial plugs Captan

Mycelial plugs 46 ±6 33±4 26 ±5 21±5 7

lO'conidia/ml 45 ±4 25 ±5 24 ±7 29 ±7 0
10' conidia/ml 164 ± 18 45 ± 7 26±7 15±5 0
10' conidia/ml 96 ± 14 41 ± 8 20±6 35 ±6 0
LSD [P = 0 05) for all comparisons = 2

' Data are the mean extent (35 observations) of pathogen colonization on shoots (mm) evaluated by isolating M
laxa from 5<m sections of shoots, 40 days after inoculation with M. laxa ± standard deviation ofthe mean Shoots
were inoculated with M. taxa on 27 March, and were first treated with E. nigrum or Captan the day before, and
thereafter four times at weekly intervals.

pathogen colotiization in the treatments with the
antagonist (comparison by column in Table
2). When comparing different treatments in
shoots inoculated with the same type of
inoculum (comparison by row in Table 2), all
ofthe treatments reduced pathogen colonization,
except the following combinations: mycehal plug
of E. nigrum-lO^ spores of M. taxa/ml. and
conidia + mycelium of £. n/g/-«w-mycelial plugs
of M. taxa. Captan treatment gave the best
disease control overall and was unaffected by the
type of M. taxa inoculum applied.
Field trials
Inoculation of shoots with M. taxa caused
blight symptoms sitnilar to natural infections and
to those described in previous experiments
(Melgarejo et ai, 1986; De Cal et ai. 1990).
The inoculation procedure without M. taxa
caused only a localized brown discoloration of
the wood in the area of inoculation.
In trials 1 and 2, lesion development in
untreated shoots was faster from 21 to 50 days
after inoculation with M. taxa than during the
first 21 days (Fig. 1). Lesion length for control
treatment at the end of the trial was greater in
trials 1 and 2 than in trials 3 and 4. In trials 3 and
4, the development of lesions was extremely
rapid from 6 to 12 days, and in the case of trial 3
this rate of development continued until 21 days
after inoculation; in both cases the development
then slowed progressively, reaching a total lesion
length less than that in trials 1 and 2.
The effect of different treatments on lesion
lengths is illustrated in Fig. 1. For clarity, only
representative biological, chemical and inte-
grated (biological + chemical) treatments were
represented in each trial. The extent of pathogen
colonization, as determined by isolations from
5-cm shoot pieces 50 days after inoculation with
M. taxa is presented in Table 3.
Table 4 shows the contrast analysis of different
methods of control (biological, chemical and
integrated made by grouping the corresponding
groups of treatments) m the four trials (Table 1).
When data from biological, chemical or inte-
grated treatments were compared with the
untreated control, significant differences were
observed in trials 1, 2 and 4. as estimated by
lesion lengths. These differences were also clear
when estimated by the extent of pathogen
colonization in trials 1 and 2. In trial 3.
significant differences were found when compar-
ing integrated treatments with untreated when
estimated by this parameter.
No significant differences were observed in an>
ofthe four trials when comparing biological and
chemical treatments. Integrated treatments
where less effective than chemical and biological
treatments in trials 1 and 2.
The comparison of biological treatments with
nutrients and biological treatments without
nutrients showed a significant result onl\ in
trial 3, when disease was estimated by lesion
lengths. Nutrients alone significantly reduced
lesion lengths and the extent of pathogen
colonization in trial 2. This reduction was not
significantly different from that obtained with
biological, chemical and integrated treatments.
558 C. Madrigal et al.
120 r 120
Trial 1 Trial 2

100 - 100

80 80

t 60 60

40 40

20 20

Ol—1-
6 12 18 24 30 36 42 48 6 12 18 24 30 36 42 48
120r 120-
Trial3 Trial 4

100 100 -

80 80

60
c
o

40 40

20 20

6 12 18 24 30 36 42 48 6 12 18 24 30 36 42 48
Days after inoculation
Fig. I. Lesion lengths (mm) induced by MoniUnia taxa in peach shoots treated with various combinatioiis of
Epicoccum nigrum and Captan in 1988-91. Biological treatments indicated as O O are treatment 3 (conidia -*-
mycelium + nutrient solution I) in trials 1 and 2. treatment 5 (conidia + mycelium + nutrient solution 2) ill trial 3
and treatment 6 (conidia + mycelium + nutrient solution 3) in trial 4. Chemical treatments indicated as • - - • - B
are treatment 7 (Captan I 3g a.i./l) Integrated treatments indicated as A - - - - A are treatment 10 (altematint
treatment 3 and 7) in trial I, treatment 11 (alternating treatment 3 and 8: only one application of tieatment 7) in trial
2, treatment 12 (one application of treatment 7 followed by applications of treatment 2) in trial 3 and treatment 13(1
application of treatment 7 followed by applications of treatment 5) m trial 4. Treatment 16 (uninoculated aad
untreated) is indicated • • Points are the mean of 42 replicates.
 Biological control of peach twig blight 559

Table 3. The effects of Epicoccum nigrum (applied in various forms) and Captan
on the colonization of peach shoots by MoniUnia taxa under field conditions

Mean extent of colonization (mm)*

Treatment*" Trial 1 Trial 2 Trial 3 Trial 4

1 _ 25-5±4 5 _
2 - - 76 0 ± 9 6 48 7 ± 5 9
3 83±31 32 0±5 0 - -
4 - 26 8 ± 4 7 - -
5 - - 65 8 ±121 41 6 ± 5 1
6 - - 30 0 ± 5 6
7 9 7±3 3 21 7 ± 5 3 78 5±9 5 33 7±5 2
8 - - _ 49 7±7 3
9 29 0 ± 5 4 _ 47 0 ± 7 0
10 I6 7 ± 5 0 - _ _
It 45 7 ± 5 1
12 - 613±I3 5 _
13 - - - 39 7±6 9
14 - 34 4 ± 5 6
15 57 8±8 6 63 9 ± 5 1 87 8 ± 9 1 40 0 ±4 9

'Evaluated 50 days after inoculation with M. laxa by isolating M. taxa from 5cm
sections of shoots. Shoots were inoculated with M. taxa on 20 May (Trial 1). 23
May (Trial 2), and 17 May (Trials 3 and 4). Data are the means of 42 observations
± standard error of the mean.
''See Table 1 for details of treatments.

DISCUSSION The glasshouse experiment provided useful

The potential of several fungal antagonists
(Penicillitmt freqitentans, P, purpurogenum, Asper-
gillus flavus and E. nigrum) for biocontrol of M.
laxa twig blight on peach was demonstrated in
field experiments (Melgarejo et ai, 1986) using the
procedure described here for inoculating shoots
with mycelial plugs of £. nigrum. Disease was also
reduced by the application of P. frequentans in
orchard trials, although the commerical aptplica-
tion ofthe method needs further work (I>e Cal et
ai, 1990). The results of the present study show
that E. nigrum can reduce twig blight under field
conditions, although incomplete control of
M, laxa was obtained in some years. These
results are useful, as are those from previous
studies where disease suppression was higher.
The results indicate the conditions under which
the biocontrol agent is unable to perform well
and therefore which factors should be modified
in order to improve biocontrol performance.
Bioeontrol strategies may be possible alternatives
to other control methods, but improved
performance is required for better disease
control.
information for selecting the best artificial
inoculation method for field trials. A method
was needed that induced moderate but not severe
disease, as is the case for the mycelial plug
inoculation method (Table 2). Twig blight in
peach trees occurs mostly through movement of
the pathogen into the stem following blossom
infection, or from other previously infected parts
ofthe tree (Byrde & Willetts, 1977). The artificial
inoculation method chosen and used in this study
simulates natural infections.
Control obtained from the application of
E. nigrum to diseased twigs in the field was
variable, depending on several factors. Relative
disease severity in untreated controls indicated
that conditions favouring rapid disease develop-
ment in the first 2-3 weeks after infection have a
negative influence on disease control (trials 3 and
4). The 2 3-week period following inoculation
appears to be crucial in obtaining successful
control of the disease, and future control
strategies should be centred on this period.
Climatic conditions may also influence the
development of antagonists and therefore the
suppression of disease. Mean relative humidity in
560 C. Madrigal et al.
Table 4. Contrast analysis of different methods of control of MoniUnia laxa in the field experiments*

Trials

Trial 1 Trial 2 Trial 3 Trial 4

Methods of control LL" EC LL" EC LL" EC LL" EC

Biological versus untreatd *** *** «*• ••* ns ns ns

Chemical versus untreated *** **« ••* *«« ns ns ••• ns
+ **
Integrated versus untreated *•* *** *** ns * •• ns
Biological versus chemical ns ns ns ns ns ns ns ns
Biological versus integrated * *** *« •« ns •* ns ns
Chemical versus integrated *** *** ** **• ns ns ns ns
Biological with nutrients ns ns * ns ns ns
versus biological without
nutrients
Nutrients versus untreated *** *** - - -
Nutrients versus biological ns ns - - -
Nutrients versus chemical ns ns - - - -
Nutrients versus integrated — ns ns — — — —

• = /"test significant at P = 0 1; *• = /"test significant alP = 0 05; **• = f test significant at /• = 0.01; NS = not
significant; - , not tested.
"Lesion length evaluated 50 days after inoculation with M. laxa.
"^ Extent of pathogen colonization on twigs evaluated 50 days after inoculation with M. laxa by isolating M. laxa
from 5-cm sections on shoots.

1988 and 1989 was 55-60% in the 2-3-week
period after inoculation with M. laxa, while in
1990 and 1991 the mean relative humidity
oscillated from 40 to 33%. These low humidity
values may have contributed to differences in
suppression of disease, as E. nigrum develops
best in conditions of high humidity. All treat-
ments were less effective in years with lower
relative humidity (see trials 3 and 4). The greater
number of treatment applications and shorter
interval between applications in trial 2 possibly
contributed to the differences in disease suppres-
sion.
Differences in disease suppression are not
related to the nutrients added to the inoculum
of E. nigrum. Although the importance of
nutrients in successful biological control has
been demonstrated in the control of twig blight
with P. frequentans (De Cal et ai, 1990), as well
as in other systems (Dickinson, 1976; Bashi &
Fokkema, 1977; Fokkema et ai, 1979;
Sundheim, 1982; Cullen et ai, 1984), it appears
not to be an important factor in Ihis case. Disease
suppression obtained in trial 2 with the applica-
tion of nutrient solution I to twigs could be due
to the development of an antagonistic microflora
on twig surfaces. Fokkema et al, (1979) also
observed that the application of yeast extract and
saccharose to wheat leaves increased populations
of antagonist yeasts.
A further reduction of twig blight severity may
be possible by increasing the concentration of
inoculum of E, nigrum in sprays. Nelson &
Powelson (1988) reduced pod rot of snap beans
(Botrytis cinerea) by 77% by applying 42c.f.u. of
Trichoderma harzianum per blossom and by 97V»
by applying 233cfu. per blossom. Disease
control could also be increased by adding
additional adjuvants to spray preparations of
E, nigrum, by using isolates with improved
adaptation to the phyllosphere, and applying
E. nigrum in mixtures with other antagonists,
such as P. frequentans (De Cal et ai, 1990).
Studies on industrial production of inoculum are
now in progress.
Bollen (1982) pointed out that differences
between pathogen and antagonist sensiti\ity to
fungicides should be considered in the develop-
ment of new integrated control strategies.
Although M. laxa is about 20 times more
sensitive to Captan than E, nigrum (De Cal,
unpublished data), combinations of £. nigrum
and Captan did not improve control when
compared to that obtained with £. nigrum or
Captan alone, except in trial 3. Neverthdess,
alternating treatments o f f . nigrum with Captan
 Biological control of peach twig blight
have advantages over the application of antago-
nists or chemical alone: the chemical acts as a
safegtiard for those years in which the weather
conditions do not favour the activity of
E, nigrum. Another advantage is that the total
fungicide application is reduced. The possibility
of exploiting other isolates of £, nigrum resistant
to Captan or other fungicides needs to be
examined further,
ACKNOWLEDGEMENTS
We wish to thank A, Iglesias, University of
Columbia, New York, for manuscript revision.
Funding was provided by INIA (Proyecto n
8561), C, Maddgal was in receipt of a scholarship
from Ministerio de Educacion y Ciencia, Spain,
REFERENCES
Bashi E, Fokkema NJ, 1977. Environmental factors
limiting growth of Sporobolomyees rosetis, an
antagonist of Coehliobolus sativtis, on wheat leaves.
Transactions of the British Myeologieal Soeiety 6S,
17-25.
Boland GJ, Inglis GD, 1988. Antagonism of white
mold {Selerotinia seleratiorum) of bean by fungi
from bean and rapeseed flowers. Canadian Journalof
Botany 61, 1775-81.
Bollen GJ, 1982. Fungicide resistance and microbial
balance. In: £>ekker J, Georgopoulos SG, eds.
Fungicide Resistanee in Crop Proteetion, Wagenin-
gen, The Netherlands: Centre for Agricultural
Publishing and Documentation, 161-76.
Byrde RJ, Willetts HG, 1977. The Brown Rot Fungi of
Fruit, Their Biology and Control, Oxford, UK:
Pergamon Press.
Cullen D, Berbee FM, Andrews JH, 1984. Chaetomium
globosum antagonizes the apple scab pathogen,
Venturia inaequalis, under field conditions. Cana-
dian Journal of Botany 62, 1814-18.
De Cal A, M.-Sagasta E, Melgarejo P, 1990. Biological
control of peach twig blight {Monilinia laxa) with
Penicillium frequentans. Plant Pathology 39, 612-18.
561
De Cal A, Pascual S, Melgarejo P, 1993. Nutritional
requirements of antagonists lo peach twig blight,
Monilinia laxa, in relation to biocontrol. Myeo-
pathologia 121, 21-6.
Dickinson CH, 1976. Fungi on the aerial surfaces of
higher plants. In: Dickinson CH, Preece TF, eds.
Mierobiology of Aerial Plant Surfaees, London, UK:
Academic Press, 293-324.
Fokkema NJ, Den Houter JG, Kosterman YJC, Nellis
AL, 1979. Manipulations of yeasts on field-grown
wheat leaves and their antagonistic effect of
Coehliobolus sativus and Septoria nodorum, Transae-
tions ofthe British Myeologieal Soeiety 72, 19-29.
Madrigal C, Tadeo JL, Melgarejo P, 1991. Relation-
ship between flavipin production by Epieoeeum
nigrum and antagonism against Monilinia taxa.
Myeologieal Researeh 95, 1375-81.
Melgarejo P, CarriUo R, M.-Sagasta E, 1985. Mycoflora
of peach twigs and flowers and its possible significance
in biological control of Monilinia laxa, Transaetions of
the British Myeologieal Soeiety 85, 307-17.
Melgarejo P, Carrillo R, M.-Sagasta E, 1986. Potential
for biological control of Monilinia laxa in peach
twigs. Crop Proteetion 5, 422-6.
M.-Sagasta E, 1977. Monitia disease. EPPO Bulletin 7,
105-16.
Nelson ME, Powelson ML, 1988. Biological control of
grey mold of snap beans by Triehoderma hamatum.
Plant Disease 72, 727-9.
Sauer DB, Burroughs R, 1986. Desinfectation of seed
surfaces with sodium hypochlorite (NaOCl). Phyto-
pathology 76, 745-9.
Snedecor GW, Cochran WG, 1980. Statistical Methods
17th Edn. Ames, Iowa, USA: The Iowa State
University Press.
Sundheim L, 1982. Control of cucumber powdery
mildew by hyperparasite Ampelomyces quisqualis
and fungicides. Plant Pathology 31, 209-14.
Zhou T, Reeleder RD, 1989. Application e>f Epieoeeum
purpuraseens spores to control white mold of snap
bean. Plant Disease 73, 639-42.
Zhou T, Reeleder RD, 1990. Selection of strains of
Epieoeeum purpuraseens for tolerance to fungicides
and improved biocontro! of Sclerotirtia selerotiorum.
Canadian Journat of Mierobiology 36, 754-9.