Natural Occurrence of Fusarium Species Associated with Root and Stalk Rot of Maize in Kermanshah Province, Iran
The objective of this study was to identify Fusarium species associated with root and stalk rot of maize and their pathogenicity on root and stalk of maize in Kermanshah Province, Iran. In this survey 480 Fusarium strains were isolated and identified from maize ears collected from different geographic regions in Kermanshah province, Iran during 2006-2009. All these 480 strains belonged to 22 Fusarium species. This Fusarium strains were evaluated for their pathogenicity on maize and observed that F. verticillioides, F. subglutinans and F. pseudoanthophilum are the most pathogenic on stalk. Foot rot assessment revealed that F. verticillioides as the most damaging species. This is the first comprehensive report on identity and distribution of Fusarium spp. from maize in west of Iran while F. pseudonygamai was reported for the first time from west of Iran.
November 21, 2010; Accepted: January 13, 2011;
Published: February 26, 2011
Maize (Zea mays L.) is one of the most important cereal crops being
cultivated in Iran. Worldwide reports have been shown that maize is one of the
most prevalent crops contaminated by Fusarium (Fandohan
et al., 2003). Fusarium spp. can cause a variety of diseases
in many agricultural, horticultural and forestry crops. Due to their significant
annual losses, studies on different aspects of its life cycle, biology, ecology,
taxonomy and its pathogenicity has been regarded by plant pathologists. Fusarium
species are a serious threat to susceptible crops worldwide, particularly
maize (Nicolaisen et al., 2009). They cause root,
stalk and ear rot, with severe reductions in crop yield, often estimated between
10 and 30% annually (Logrieco et al., 2002).
There are two types of maize ear rot, called red ear rot and pink ear rot. Red
ear rot universally caused by Discolor section and pink ear rot caused
by Liseola section (Aliakbari et al., 2007).
Fusarium spp. can also produce mycotoxins which relevant to animal and
human health (Woloshuk and Shim, 2000; Velluti
et al., 2001). F. graminearum, F. culmorum, F. cerealis, F. poae
are the major producers of trichothecenes (Logrieco et
al., 2002). Maize grains may be contaminated with different Fusarium
toxins such as trichothecenes, zearalenone, fumonisins and moniliformin
in grains that are harmful to both human and animals (IARC,
2002). Some of the strains of F. verticillioides do not produce disease
in maize and are an endophyte, because it does not cause visible damage to the
plant (Stone et al., 2000). The aim of this study
is to identification of Fusarium species associated with maize and their
pathogenicity assay on root and stalk of maize in Kermanshah province of Iran.
MATERIALS AND METHODS
Isolation and Identification of Fusarium spp.: During 2006-2009,
diseased maize tissue sample were collected from 120 maize fields from different
regions of Kermanshah province from West of Iran. Roots and stalks of diseased
samples were washed under tap water. Then the samples were surface sterilized
with 70% ethanol and cut into small blocks. The blocks were soaked in 1% sodium
hypochlorite solution for 3 min and rinsed in several changes of sterile distilled
water. All the sterilized samples were placed on Peptone-Pentachloronitrobenzene
Agar (PPA) plates, a selective medium for Fusarium (Nash
and Snyder, 1962). The plates were incubated under standard incubation conditions
(Salleh and Sulaiman, 1984) for 24 h. The resulting
single-spore Fusarium colonies were transferred to fresh Potato Dextrose
Agar (PDA) plates and maintained at 4°C for further studies. To study the
growth rates and pigment production of Fusarium spp. all the strains
were transferred onto PDA plates and incubated at ambient temperature. Ten replications
were maintained for each Fusarium strain. For microscopic observations,
all the strains of Fusarium were transferred to Carnation Leaf Agar (CLA)
(Fisher et al., 1982), Spezieller Nahrstoffarmer
Agar (SNA) (Nirenberg, 1976) and Potassium Chloride
Agar (KClA) (Fisher et al., 1983) medium. The
species were identified on the basis of macroscopic characteristics such as
pigment production and growth rates on PDA plates, as well as their microscopic
features including size of macroconidia, presence of microconidia and its production
in chains or false heads, type of conidiogenous cells (monophialidies and polyplialidies
conidiophores) and also absence or presence of chlamydospores (Gerlach
and Nirenberg, 1982). Identification of species was based on species description
of Gerlach and Nirenberg (1982), Nelson
et al. (1983) and Leslie and Summerell (2006).
Pathogenicity assay: All of the identified Fusarium species were
tested for their pathogenicity on apparently healthy maize and individual plants
were used for inoculation tests on root and stalk. The root and stalk of the
maize were washed and surface sterilized before inoculation. For inoculation,
each strain of Fusarium species were grown on PDA plates as described
by Salleh and Sulaiman (1984). Conidial suspension of
each strain was prepared by scrapping the mycelium with sterile distilled water
onto 7 day-old cultures, shaken thoroughly and the concentration was adjusted
to 2x106 conidia mL-1 using haemocytometer. Twenty milliliter
of the spore suspension of each Fusarium species was sprayed on the root
and stalk of the plants. Control plants were sprayed with 20 mL of sterile distilled
water. Three replications were maintained for each strain and the experiment
was repeated twice.
RESULTS AND DISCUSSION
In this study, 480 Fusarium isolates were isolated from diseased plants and based on morphological characteristics, these isolates were identified as 22 Fusarium species including F. avenaceum, F. acuminatum, F. anthophilum, F. culmorum, F. chlamydosporum, F. equiseti, F. globosum, F. graminearum, F. lateritium, F. nygamai, F. oxysporum, F. poae, F. proliferatum, F. pseudoanthophilum, F. subglutinans, F. pseudonygamai, F. solani, F. semitectum, F. sambucinum, F. sporotrichioides, F. tricinctum and F. verticillioides (Table 1, 2). Of the Fusarium isolates collected in Kermanshah province, Iran, F. verticillioides was the most prevalent with a frequency of 51.04 % (245 of 480), followed by F. proliferatum with a frequency of 30.42% (146 of 480). Among isolates, 25% from root and 75% from stalk tissue were recovered. All of the species except F. oxysporum and F. lateritium were recovered from stalk rot of maize. The species identified from root rot of maize were F. verticillioides, F. oxysporum and F. lateritium. In stalk tissues F. verticillioides, F. proliferatum, F. subglutinans, F. chlamydosporum, F. pseudoanthophilum, F. poae and in root tissue F. verticillioides were predominant species. In this survey F. pseudonygamai is reported for the first time from Iran.
Eight days after inoculation, disease symptoms were observed on the roots. Initially, the roots illustrated brownish discoloration and eventually turned to dark color, which indicates rotting of the root tissues. Twelve days after inoculation, the first symptoms on maize stalks on yellowish and then degrees of rotting were observed on the stalk base. The results in pathogenicity tests indicated that F. verticillioides, F. subglutinans and F. pseudoanthophilum are the most pathogenic on stalk and that only F. verticillioides is pathogenic on the root.
In this study, the association was observed between maize plant and the species
of Gibberella fujikuroi species complex (Nirenberg
and ODonnell, 1998). In the pathogenicity test, F. verticillioides
was recovered from the inoculated root and stalk which indicated that this species
was able to cause root and stalk rot of maize. F. verticillioides, F.
proliferatum and F. subglutinans are the common pathogens of maize
in different climates (Lew et al., 1991; Miller,
2001; Logrieco et al., 2002; Munkvold,
2003). The results in this study are in accordance with other findings in
Iran (Ghiasian et al., 2004), Dawodee
(2002), Mohammadi et al. (2002) and Naderpour
(2004) had reported that F. verticillioides and F. proliferatum
as the most comon species isolated from maize in Iran. This fungus can be
seed borne internally in symptomless, apparently healthy maize kernels (Thomas
and Buddenhagen, 1980). Also some strains of this fungus can produce potent
mycotoxins associated with serious animal and human diseases (Wilson
et al., 1985). Mycotoxin-producing Fusarium species such
as F. verticillioides, F. graminearum and F. poae were
isolated at high levels from disease samples in this study. This indicates that
the presence of high infections of stalk rot on the ears thus supporting the
farmers information on the presence of the disease in their fields. Thus, identification
of the different species of Fusarium, including saprobe, pathogenic and
toxin producing species, is of vital importance (Nelson
et al., 1994).
|| Morphological characters of Fusarium spp. associated
|+: Presence, -: Absence, F: False heads (chains absent), M:
Medium to long chains (>15), S: Short chains (<15), P: Pyriform, G:
Globose, C: Clavate, Ob: Obovoid, Ova: Oval to allantoids/fusiod, Poly:
Polyphialidic, Mono: Monophialidic
|| Macroconidia characters of Fusarium spp. associated
|Pdfs: Poorly developed foot shape, Nfs: Notch or foot shape,
Fs: Foot shape
Our results indicated that a section Liseola or complex of Gibberella
fujikuroi species (Nirenberg and ODonnell, 1998)
could be pathogenic to the maize and suggested that F. verticillioides
could be the main causal agent of stalk and root rots. However, strains of F.
subglutinans and F. pseudoanthophilum were the main causal agents
of stalk rot as well. In this survey, the widespread of species especially in
Gibberella fujikuroi species complex offers the opportunity for relatively
easy collection of large numbers of species by phylogenetically-based classification
(ODonnell, 1996). The identification of the Fusarium
species contaminating maize in different areas is evidence for a need, not only
for studying the levels of the interaction between Fusarium pathogens
and maize, but also to obtain a precise picture of the toxicological risks related
to the maize consumption by humans and animals (Logrieco
et al., 2002). The present study demonstrates the importance of Fusarium
stalk and root rot of maize in Iran. We believe that this study will serve as
a foundation for the further identification of Fusarium species, particularly
molecular level from Iran. This is the first comprehensive report on identification
of many Fusarium spp. from maize in west of Iran.
Author acknowledges the Islamic Azad University, Sanandaj Branch, Iran and University Sains Malaysia, Penang, Malaysia for providing necessary facilities to carry out this research.
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