Published in Crop Sci 39:1830-1834 (1999)
© 1999 Crop Science Society of America
677 S. Segoe Rd., Madison, WI 53711 USA
Crop Science 39:1830-1834 (1999)
© 1999 Crop Science Society of America
CROP ECOLOGY, PRODUCTION & MANAGEMENT
Irrigated Hybrid Maize Crop Yield Losses Due to Barley Yellow Dwarf Virus-PAV Luteovirus
M. Beuvea,
B. Naïbob,
L. Foulgocqc and
H. Lapierrea
a Unité de pathologie végétale, INRA, Route de Saint-Cyr, 78026 Versailles Cédex, France
b AGPM, 21 Chemin de Pau, 64121 Montardon, France
c AGPM. 91720 Boigneville, France
lapierre{at}versailles.inra.fr
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ABSTRACT
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PAV, one of the luteoviruses inducing the barley yellow dwarf disease, greatly reduces yield of cereal crops and has been reported to cause reddening in several varieties of maize (Zea mays L.). However, there have been no reports that the virus causes significant reductions in size or yield of irrigated maize. Trials to assess the effect of PAV on maize yields were conducted for 3 yr with two hybrids (`Déa' and `Nobilis') and two isolates of PAV (2t and L14). These two isolates differed in geographic origin and virulence on barley (Hordeum vulgare L.), but they multiplied similarly to wild isolates in leaves of maize hybrids. The virus was detected in maize leaves during the same period after inoculation in two out of three years. The reason for low levels of virus in leaves in the 1996 season may have been high temperatures (>30°C) following inoculation. Under these conditions, the virus was restricted to the roots for long periods. This blocking of virus migration and/or foliar multiplication was accompanied by less reddening, and there was almost no decrease in grain yield. High early leaf infection of the virus had little effect on the vegetative development of the hybrids (<10% reduction in plant height), but grain yield was between 15 and 20% lower for a production of 10 to 12 t ha-1. This drop in yield was due to fewer kernels of maize per ear, while the 1000 kernel mass was unaffected. The loss in yield was not affected by either the virus isolate or the maize hybrid, although the intensity of symptoms differed between hybrids. The data show that there are potential losses incurred by PAV on maize yield. Therefore, these findings justify the establishment of a breeding program.
Abbreviations: BYDV, barley yellow dwarf virus • DAS-ELISA, double antibody sandwich enzyme linked immunosorbent assay • ELISA, enzyme linked immunosorbent assay • OD, optical densities • TAS-ELISA, triple antibody sandwich enzyme linked immunosorbent assay
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INTRODUCTION
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BARLEY YELLOW DWARF VIRUSES
(BYDVs) are frequently encountered in maize crops in temperate areas. Those viruses of similar biology designated GPV, MAV, PAV, RMV, RPV, and SGV have been detected on many species of the Poaceae family. D'Arcy and Mayo (1997) suggested grouping these viruses into two genera: Luteovirus, which would include PAV and MAV and probably SGV, and Polerovirus, for RPV and probably RMV (Geske et al., 1996) and GPV (Cheng et al., 1996). Only MAV, PAV, RMV, and SGV have been clearly identified on maize crops either singly or in combination. The RPV serotype has been detected in maize (Comas et al., 1993; Webby et al., 1993), but the extent to which RPV infects this species is unclear (Beuve and Lapierre, 1993). SGV and RMV are found mainly in regions with a continental climate, such as central Europe (Pocsaï et al., 1995). MAV also infects maize and, in a few cases, is the most frequently encountered BYDV virus in maize (Comas et al., 1993). PAV is present in most growing regions and is often the prevalent BYDV detected in maize (Brown et al., 1984; Beuve and Lapierre, 1993).
Many maize hybrids show symptoms of leaf reddening when infected with PAV (Brown et al., 1984). Although maize crops may be highly infected in some years, maize has long been considered to be simply a reservoir of virus responsible for the infestation of cereals in the northern hemisphere (Brown et al., 1984; Stoner, 1977). However, little is known about the effect of the BYDVs on maize yields in field conditions. Sterility of some lines is associated with infection by BYDVs (Reffati et al., 1990), and Loi et al. (1994) reported BYDV causes a 27% drop in the yield of maize hybrid B73. D'Arcy et al. (1995) estimated a 25% yield loss for two sweet corn hybrids infected with an RMV isolate. In contrast, a 2-yr trial with PAV-inoculated plants in Germany showed no yield drop in maize hybrids (Grüntzig et al., 1997).
The aim of the study was to investigate the effect of PAV infection on the yield of two high-potential maize hybrids under irrigation.
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Materials and methods
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Field Conditions
The experiment was conducted on maize under irrigation on one site at Desmonts (48.1°N, 2.3°E) in the Paris basin, across three seasons (1995, 1996, and 1997). The two maize hybrids (Déa and Nobilis) have similar yield potentials. The planting dates were 4 May 1995, 26 Apr. 1996, and 30 Apr. 1997. The experiment was a randomized split-plot design with four replicates in 1995 and 1997. The two hybrids were applied to main plots and the two isolates of PAV (2t and L14) to subplots. In 1996, it was a randomized complete-block design. Each plot consisted of six 10-m-long rows with 72 plants per row. Inter-row spacings were 0.8 m and plant stand was 90000 plants ha-1. Fertilizers were applied at 160 kg N ha-1, 105 kg P ha-1, and 105 kg K ha-1. Weeds were controlled with atrazine (2-chloro-4-ethylamino-6-isopropylamino-s-triazine) (Ciba-Geigy, Reuil-Malmaison, France) applied at 1 kg ha-1 preplanting and a mixture of 0.5 kg ha-1 atrazine and 0.48 kg ha-1 pyridate (0-(6-chloro-3-phenyl-4-pyridazinyl)-S-octyl-carbonothioate) (Sandoz-Agro, St Germain en Laye, France) at the two- to three-leaf stage. Air temperature was recorded at a weather station near the experiment site.
Inoculum
Populations of virus-infected bird cherry-oat aphid (Rhopalosiphum padi L.) were reared in a growth chamber (at 16–20°C with a 15-h photoperiod, 280 µmol m-2 s-1) on barley cultivar Vixen infected with PAV-2t or PAV-L14 isolates (Table 1)
. In 1995, both isolates were used to inoculate plants. In 1996 and 1997, only isolate PAV-2t was inoculated. Fifty maize plants in the two central rows of each plot were inoculated at the three- to five-leaf stage on 22 May 1995, 28 May 1996, and 26 May 1997 by transferring five to ten apterous virus-infected aphids with a fine paintbrush. Aphids were killed by spraying insecticide (lambda-cyhalothrine; Zeneca, Velizy, France) 48 h later. Two other insecticide treatments with the same insecticide were applied 2 and 4 wk later to both the control and inoculated plants to protect against natural infection, which occured at this time.
Enzyme Linked Immunosorbent Assay
Maize was assessed for virus content by enzyme linked immunosorbent assay (ELISA) 3 and 5 wk after inoculation in 1995 and 1, 5, and 7 wk after inoculation in 1996 and 1997. The maize plants in the two central rows of each plot were sampled. Until the flower stage, the uppermost fully expanded leaf was sampled. Later, leaves were chosen from any part of the shoot where symptoms appeared (at ear level or higher). Extracts from shoots and roots were prepared with a roller press, with 
0.2 g tissue mL-1 in 10% polyvinylpyrrolidone and 0.1% bovine serum albumin in phosphate buffer. They were tested by double antibody sandwich enzyme linked immunosorbent assay (DAS-ELISA) in 1995 and by triple antibody sandwich enzyme linked immunosorbent assay (TAS-ELISA) in 1996 and 1997. Duplicate wells in Greiner flat microplates were used for each sample. Anti-PAV polyclonal antiserum was produced in rabbit. Monoclonal antibody PM63 (specific for an epitope common to PAV and MAV) was produced in mouse. Conjugated alkaline phosphatase anti-mouse IgGs from rabbit were provided by Biosys (Compiègne, France). Microplates were coated by incubation with PAV polyclonal antibody (1 µg mL-1) for 2 h at 37°C. Plant extracts were added to the plates and incubated overnight at 4°C. Conjugated polyclonal antibodies (1 µg mL-1) were added and the plates incubated for 3 to 4 h at 37°C (DAS-ELISA). Monoclonal 63 and labelled anti-mouse antibodies (1 and 0.25 µg mL-1, respectively) were added successively and incubated for 2 h at 37°C (TAS-ELISA). Absorbance at 405 nm was recorded 2 h after adding the enzyme substrate (1 mg mL-1 of p-nitrophenylphosphate) using a Molecular Devices Emax reader (Molecular Devices, Palo Alto, CA).
Other Analyses
Infected plants were identified by ELISA and were marked accordingly with a colored tag. Symptoms (only tips and edges of leaves turning red) were noted regularly every 15 d from July to October for each year. In 1997 only, the frequency of symptoms was recorded on the top eight leaves of 100 infected plants for the two hybrids on 10 September. Also in 1997 only, plant height was measured 9 wk after inoculation with a graduated measuring rod on 29 July. Harvests were done on 25 Sept. 1995, 30 Sept. 1996, and 29 Sept. 1997. Ears were hand picked and the number of kernels immediately counted (number of rows and number of kernels per row for each ear). Both noninfected and infected plants had only one ear each. The mass of 1000 kernels (15% moisture) and the number of kernels per ear were determined for infected and noninfected plants. The experiments were analyzed using analyses of variance for split-plot or complete-block designs. Treatments effects were tested with Fisher test (F test) (Cochran and Cox, 1950). Comparisons between treatments were carried out using the Newman-Keuls test (Newman, 1939; Keuls, 1952). Statistical analysis was performed using STATITCF (Tranchefort et al., 1991) or Genstat (Genstat 5 Committee, 1993).
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Results
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Barley Yellow Dwarf Virus Infection
The percentages of infected plants in field maize were estimated across 3 yr (1995, 1996, and 1997; Table 2)
. Natural infection of PAV was high in 1995: 53% in the first week of July (13–14 leaf stage). In contrast, no natural infection was detected for the same period in 1996 and 1997. The level of infection over the same period following artificial inoculation was 90% in 1995, 2% in 1996, and 72% in 1997. The level was very low in mid July 1996, reaching a maximum of 40% and remained at that value. In 1996 only, high temperatures (>30°C) occurred during the 15 d following inoculation (Table 2). During this period in 1996, when found, the distribution of the virus in maize plants was very uneven (Table 3)
, contrasting with the uniform distribution in 1997 (data not shown). Virus was not recovered at all from most plants. In some plants, the virus was detected only in the roots, while in other plants the virus was present in the roots and the youngest leaf only. More frequently, when present, the virus was detected in the four to seven uppermost leaves and the roots. The mean optical densities obtained by ELISA for naturally and artificially infected plants in 1995 were compared (Fig. 1)
. No significant differences were found, but optical densities for natural infection were slightly lower than those for isolate PAV-L14.
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Table 3 Optical densities (OD) enzyme linked immunosorbent assay (ELISA) values for roots and leaves of the maize hybrid Déa, 42 d after inoculation with PAV-2t in 1996
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Fig. 1 Mean optical densities (OD) enzyme linked immunosorbent assay (ELISA) values from leaf extract for natural and artificial PAV infection of two maize hybrids in 1995. Error bars indicate one standard deviation
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Symptoms
Symptoms appeared at the beginning of July both in 1995 and 1997 and affected several leaves (up to eight). Symptoms detected at the end of August in 1996 were very mild and present only on the uppermost one to three leaves. Each season reddening was more severe on Nobilis than on Déa. In 1997, the frequency of symptoms on the uppermost eight leaves of infected plants was 69% for Nobilis and 49% for Déa.
Plant Height
In 1997, there were significant differences between treatments in maize plant height. Infected plants were 9.3 and 9.9 cm shorter than noninfected plants for Nobilis and Déa, respectively (Table 4)
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Grain Yield
In 1995, the mean grain yield of infected plants was 11.14 t ha-1 with isolate PAV-L14 and 11.54 t ha-1 with isolate PAV-2t. This was 15 and 12% less, respectively, than the grain yield of noninfected maize (Table 5)
. In 1996, infected and noninfected plants did not differ for yield, averaging 9.58 t ha-1 (Table 6) . In 1997, noninfected-plant yields were 17% higher than the yields from infected plants (Table 7)
. The 1000 kernel mass was not significantly affected by PAV but was consistently 4 to 5% lower in infected plants than in the control. In contrast, the number of kernels per ear of the plants infected with PAV was 10% lower than the controls in 1995 and 12% lower than the controls in 1997.
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Discussion
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The grain yield of hybrid maize crops infected with PAV was reduced by nearly 13.8%. The number of ears per tiller was unaffected and the 1000 kernels mass was only slightly affected by infection. The decrease in yield was due mainly to a reduction in the number of kernels per ear and not a large reduction in flowering, as suggested by Reffati et al. (1990) for certain genotypes. The decreases in plant size and yield were smaller than those reported by Pearson and Robb (1984), who studied plants infected with virus in the greenhouse and then planted out in the field.
High levels of natural infection by PAV and other BYDVs have been found in other environments (Comas et al., 1993; Webby et al., 1993; Loi et al., 1994). Rates of infection after controlled inoculation were very high in 1995 and 1997, whereas they were low in 1996, the virus being difficult to detect in some cases. As in our 1996 trials, Grüntzig et al. (1997) found no symptoms and no effect on yield for two consecutive years. However, no data on temperature conditions during the two seasons were reported. Eweida et al. (1983) showed that temperature has a large effect on the infection of greenhouse-grown maize by two BYDVs. The frequency of infection with the viruses was seven times higher for plants grown at 18 to 20°C than for those grown at 23 to 29°C. The growing conditions at the start of our trial in 1996 probably partially reflected this, the higher temperatures resulting in lower infection. In that year, temperatures rose above 30°C on the days following inoculation. Several steps in the virus life cycle may be affected by high temperature, including inoculation, migration to the roots, migration to the leaves, and virus multiplication. None of these early steps is specifically affected if barley is inoculated at 30°C in a growth chamber (data not shown). Therefore, this resistance to virus at high temperature seems to be specific to maize and results in a wide variety of virus cycle stages being found in field-grown inoculated plants. The lack of virus migration to the aerial organs in the field (although temporary) may have resulted in there being fewer foliar symptoms in 1996. Some maize hybrids express few foliar symptoms (Lorenzoni et al., 1990) and others are resistant to local PAV isolates (Stoner, 1977; Panayotou, 1977; Brown et al., 1984; Eweida et al., 1983). Some of these resistant genotypes are sensitive to infection by other isolates of the virus (Pearson and Robb, 1984). In our study, the effect of PAV on maize was assessed using only two virus isolates of the same cpA serotype (Mastari et al., 1998). Therefore, some PAV isolates may be more virulent for maize than those used in this study. This may also account for the environmental and genotype differences and the virulence of PAV reported by Panayotou (1977), Pearson and Robb (1984), and Loi et al. (1994).
PAV affects the yield of hybrid maize grown under irrigation. This effect of PAV should be evaluated for maize grown with limited water input, and the effects of the other Luteovirus species, Polerovirus species, and complexes of them that infect maize should also be assessed.Genstat 5
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ACKNOWLEDGMENTS
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The authors wish to thank B. Le Tarnec, L. Coudard, L. Maurice, and L. Marbot for technical assistance and O. David for statistical analysis.
Received for publication November 20, 1998.
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ABSTRACT
INTRODUCTION
