Research

Pathogenicity and Management of Olpidium bornovanus,

a Root Pathogen of Melons

M. E. Stanghellini, D. M. Mathews, and I. J. Misaghi, Department of Plant Pathology and Microbiology, Univer-
sity of California, Riverside 92521

(20 to 35°C) greenhouse containing 18

ABSTRACT drip-irrigated recirculating hydroponic
Stanghellini, M. E., Mathews, D. M., and Misaghi, I. J. 2010. Pathogenicity and management of units (Jetstream, American Agritech,
Olpidium bornovanus, a root pathogen of melons. Plant Dis. 94:163-166. Tempe, AZ) (Fig. 1). Each drip-irrigated
unit consisted of two fiberglass troughs
Greenhouse studies document, for the first time, that Olpidium bornovanus, an obligate, holo- (each 110 × 21.5 × 12 cm) connected to a
carpic, root-inhabiting zoosporic fungus heretofore regarded as a nonpathogenic parasite, is a common reservoir containing 25 liters of
root pathogen. Significant browning of the roots and reductions in shoot and root growth were aerated nutrient solution, which was dis-
recorded within 28 days following inoculation of melons with the fungus. Amending the recircu-
tributed continuously to each plant via 7.2
lating nutrient solution with either a nonionic surfactant (Agral 90) or a strobilurin fungicide
(azoxystrobin) resulted in efficacious management of the disease caused by the fungus.
liters/h emitters. Four potted plants were
placed in each trough, eight plants per unit
(Fig. 1A). Eight days after placement of
potted plants in troughs, 0.5 g (fresh
Vine decline of melons (Cucumis melo 1979) raised questions regarding the weight) of cantaloupe roots colonized by
L.), a disease characterized by a sudden pathogenicity of O. bornovanus. The O. bornovanus was excised from the ISP
and field-wide collapse of mature fruit- latter fungus, in addition to two other and placed in the nutrient solution in res-
bearing plants 1 to 2 weeks before harvest, species of the genus that are associated ervoirs of appropriate treatments. Treat-
has historically been attributed to Mono- with plant roots (i.e., O. brassicae and O. ments included four noninoculated and
sporascus cannonballus, a soilborne, root- virulentus), is commonly regarded as a four inoculated hydroponic units. Root
infecting ascomycete (6,15,19). However, nonpathogenic obligate parasite (10,17,23). colonization by O. bornovanus was as-
Melon necrotic spot virus (MNSV), which However, a possible pathological effect sessed as follows: at 2-day intervals after
is well documented as a destructive viral was suggested from a greenhouse study inoculation, a small section of the potting
disease of greenhouse-grown melons following the observation of a brown substrate was excised from one potted
(1,5,7,11,14,26), as well as field-grown discoloration of melon roots colonized by plant from each replication of the inocu-
melons (12,13,24), has recently been re- O. bornovanus (3). lated treatments. Roots were extracted
ported as a probable cause of vine decline The primary objective of this study was from the potting medium and examined
of melons in the field in Guatemala (8). to assess the pathogenic capabilities of O. microscopically (×40) for the presence of
This virus is seedborne and transmitted (in bornovanus on melons. Additionally, the sporangia in epidermal cells. The shoot
a vector-assisted manner; 4) by Olpidium efficacy of a surfactant and a fungicide on portions of two plants randomly selected
bornovanus (Sahtiyanci) Karling (=Olpidium control of this root-inhabiting, zoosporic from each treatment replication (N = 8)
radicale), an obligate, soilborne, root- fungus was examined. A preliminary report were excised at the surface of the potting
inhabiting, zoosporic fungus (3,25). Ac- has been published (20). medium at 14, 21, and 28 days after inocu-
cording to Campbell and Sim (3), there are lation, and their fresh shoot weights were
three host-specific strains (i.e., melon, MATERIALS AND METHODS recorded (Fig. 1B). Additionally, the root
cucumber, and squash) of O. bornovanus. Pathogen and host. All experiments systems of two plants from each treatment
Our preliminary studies showed that the were conducted using cantaloupe (Cucu- replication were removed from the pots,
melon strain of O. bornovanus is common mis melo L. cv. Caravelle) as a host. A washed, and visually assessed for root
in soil samples collected from commercial single sporangium isolate of O. borno- discoloration. At the final harvest date (i.e.,
melon fields (4 from Arizona and 10 from vanus (IP-1) recovered from soil from a 28 days after inoculation), ten 1-cm-long
California) with a history of vine decline commercial melon field (Imperial Valley, root segments (0.2 to 0.4 mm in diameter)
caused by M. cannonballus. The fungus CA) was maintained on cantaloupe seed- from each plant were randomly selected
was isolated from soil using melon seed- lings (hereafter referred to as the inocu- and examined microscopically (×40) for
lings as bait (8). The presence of the fun- lum-source-plant [ISP]) growing in ver- the presence of sporangia and resting
gus and the absence of the virus (which miculite in a growth chamber. spores of O. bornovanus, and their num-
has not been reported to occur in the Cantaloupe seeds were surface sterilized bers were recorded. The remaining root
United States since its original discovery by soaking in 0.6% commercial sodium system of each plant was blotted dry and
[9] in a greenhouse in Riverside, CA in hypochlorite for 3 min, rinsed in distilled fresh weights recorded. Data were sub-
water for 1 min, and germinated asepti- jected to analysis of variance (ANOVA).
cally on 1.5% water agar in petri plates at Means were compared by Fisher’s pro-
Corresponding author: M. E. Stanghellini 30°C. The seedlings (3 days old) were then tected least significant difference (LSD) at
E-mail: [email protected] transplanted into rockwool cubes (1.5 × P = 0.05. The experiment was repeated
1.5 × 1.5 cm) and grown for 10 days in a two more times over a 12-month period,
Accepted for publication 28 October 2009. growth chamber at 30°C with a 12-h pho- and results from a single representative
toperiod. They were then transplanted into experiment were chosen for presentation.
doi:10.1094 / PDIS-94-2-0163 vermiculite in 11 × 11 × 12 cm plastic pots Additionally, the experiment was repeated
© 2010 The American Phytopathological Society and relocated to a temperature-controlled once with another single sporangium iso-

Plant Disease / February 2010 163

late of O. bornovanus recovered from a fungicide was added to the reservoir at the also used for double-stranded RNA
commercial melon field soil in Arizona following rate: 10 µg a.i./ml 3 days prior to (dsRNA) extraction and analysis by 6%
(HQ-1). inoculation and then repeated at 7-day polyacrylamide gel electrophoresis (16).
Chemical control studies. Cantaloupe intervals. At 28 days after inoculation, the Finally, the cantaloupe samples were
seedlings were reared and grown in the shoot and root systems of all plants in each ground in 0.1 M potassium phosphate
greenhouse as described above. There treatment replication were harvested and buffer, pH 7.2 (1:2 wt/vol), and the extracts
were four treatments with two replications data recorded as described above. Data were used to inoculate Chenopodium qui-
per treatment, and the experiment was were subjected to analysis of variance noa, Nicotiana glutinosa, Gomphrena
repeated once. Treatments included non- (ANOVA). Means were compared by globosa, and cantaloupe plants. Plants
inoculated hydroponic units, inoculated Fisher’s protected least significant differ- were observed for 18 days for the devel-
units, and inoculated units in which the ence (LSD) at P = 0.05. The experiment opment of symptoms.
nutrient solution in the reservoir was was conducted two times, and results from
amended with either a nonionic surfactant a single representative experiment were RESULTS
(Agral 90, ICI, United Plant Protection chosen for presentation. Pathogenicity studies. Sporangia of O.
Division, UK) or a systemic azoxystrobin Virus screening. Leaves and roots from bornovanus were first observed in epider-
fungicide (Quadris, 2.08 SC, Syngenta). randomly selected cantaloupe seedlings mal cells of roots of melon plants 6 days
The surfactant was chosen because it had and mature experimental plants were har- after artificial infestation of the recirculat-
previously been documented to control this vested throughout the above trials and used ing nutrient solution. At that time, ap-
zoosporic fungus on hydroponically grown to test for the presence of plant viruses by proximately 5% of the roots examined
cucumbers (25) and zoosporic root- several methods. They were specifically were colonized, and the mean number of
infecting oomycetes (18,21,22). The sur- tested for the presence of MNSV and Cu- sporangia per millimeter of root (0.2 to 0.4
factant was applied at the following rate: cumber mosaic virus (CMV) by double- mm in diameter) was 0.04 (range = 0 to
20 µg a.i./ml of the nutrient solution in the antibody sandwich, direct enzyme-linked 28). Significant reductions (38 to 58%) in
reservoir 3 days prior to inoculation and immunosorbent assay (DAS-direct ELISA) fresh shoot weights in inoculated treat-
then 20 µg a.i./ml at 7-day intervals. The (Agdia, Elkhart, IN). These tissues were ments, relative to the noninoculated treat-
ments, were recorded at all sampling dates
(Fig. 2A). Roots of cantaloupe plants in
inoculated treatments appeared slightly
discolored (tan coloration) 14 days after
inoculation. By 28 days following inocula-
tion, the root systems of all inoculated
plants were brown (Fig. 3), and all roots
examined were extensively colonized by
O. bornovanus. Both sporangia and resting
spores were microscopically observed in
colonized root cells (Fig. 4), and the mean
number of propagules of O. bornovanus
per millimeter of root was 215 (range = 41

Fig. 2. Effect of Olpidium bornovanus on A,

fresh shoot weight of cantaloupe plants at vari-
ous time intervals after inoculation and B, fresh
root weight 28 days after inoculation. Each
column is the mean weight from eight plants,
and different letters indicate significant differ-
Fig. 1. Potted cantaloupe plants in recirculating hydroponic units in a temperature-controlled green- ences according to Fisher’s protected least sig-
house A, 5 days prior to and B, 21 days after inoculation with Olpidium bornovanus. nificant difference (P = 0.05).

164 Plant Disease / Vol. 94 No. 2

to 632). A significant reduction (40%) in treated plants (Fig. 5A and B). Roots of common melon-infecting viruses. Based
fresh root weights of inoculated plants, inoculated melon plants were brown in upon these findings, the extent of crop loss
relative to the noninoculated ones, was color, and almost all epidermal root cells due to O. bornovanus in cantaloupe plants
recorded (Fig. 2B). contained either sporangia or resting grown commercially in greenhouse envi-
No sporangia or resting spores were mi- spores of O. bornovanus. ronments (1,5,7,11,14,26), which was
croscopically observed in roots of non- Virus detection. All plants tested were previously attributed primarily, if not
inoculated plants. Repeat experiments with negative for MNSV and CMV by DAS- solely, to the virus, needs to be reassessed.
both the IP-1 and HQ-1 isolates of O. bor- direct ELISA (data not shown). No dsRNA Our studies also indicate that amending the
novanus gave similar results (data not segments were observed on gels used to recirculating nutrient solution with either
shown). analyze extracts from experimental canta- azoxystrobin (Quadris) or Agral 90 (which
Chemical control studies. Amending loupe plants. No symptoms developed on confirms an earlier study [25] on its effi-
the nutrient solution with either the non- inoculated leaves or in systemic leaves of cacy in control of the cucumber strain of
ionic surfactant or the strobilurin fungicide indicator hosts after inoculation with canta- the fungus) provided excellent manage-
resulted in complete control of the patho- loupe extracts from the various experiments. ment of the disease on greenhouse grown
gen. No sporangia or resting spores of O. melons caused by the cantaloupe strain.
bornovanus were observed in roots of DISCUSSION However, the role of O. bornovanus as a
inoculated–chemically treated plants. While O. bornovanus is well docu-
Fresh weights of roots of the latter plants mented as the vector of MNSV (2,9,14),
at the end of the experiment were statisti- our data indicate that this fungus is also a
cally similar to or greater than root weights virulent pathogen capable of causing ex-
of noninoculated plants. In contrast, root tensive browning of the roots and growth
and shoot fresh weights of inoculated reductions in infected cantaloupe plants.
plants were significantly reduced (22 and We ruled out the possibility of viral infec-
45%, respectively) relative to the non- tions causing any of these symptoms dur-
inoculated and inoculated–chemically ing this study, either due to MNSV or other

Fig. 5. Effect of chemical amendments to the

recirculating nutrient solution on A, fresh shoot
weight and B, fresh root weight of hydroponi-
cally grown cantaloupe plants 28 days after
inoculation with Olpidium bornovanus (Ob). Ck
= control, S = a nonionic surfactant (Agral 90),
and Q = a strobilurin fungicide (Quadris). Each
column is the mean weight from 16 plants, and
different letters indicate significant differences
Fig. 3. Comparison of the root system of a healthy and an Olpidium bornovanus–colonized plant 28 according to Fisher’s protected least significant
days after inoculation. difference (P = 0.05).

Fig. 4. A, Sporangia and B, resting spores of Olpidium bornovanus in cantaloupe roots 28 days after inoculation.

Plant Disease / February 2010 165

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166 Plant Disease / Vol. 94 No. 2