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Evidence of genetic transmission of antibiosis and antixenosis resistance of sorghum to the spotted stemborer, Chilo partellus (Lepidoptera: Pyralidae)
Crop Protection 31 (2012) 21e26
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Evidence of genetic transmission of antibiosis and antixenosis resistance of sorghum to the spotted stemborer, Chilo partellus (Lepidoptera: Pyralidae) P.G. Padmaja*, C. Aruna, J.V. Patil Directorate of Sorghum Research, Rajendranagar, Hyderabad 500 030, Andhra Pradesh, India
a r t i c l e i n f o
a b s t r a c t
Article history: Received 22 January 2011 Received in revised form 20 September 2011 Accepted 20 September 2011
Spotted stemborer, Chilo partellus (Swinhoe) (Lepidoptera: Pyralidae), is the most important pest of sorghum in Asia and south and eastern Africa. Host plant resistance is an important control tactic for controlling this pest. Two breeding lines 27B PB 15881-3 and 463B PB 15881-3 with their parents, resistant and susceptible genotypes were evaluated in the field, glasshouse and laboratory for different resistance parameters. Breeding lines and genotypes varied significantly in foliar damage ratings, percentage of stem length tunneled, percentage of plants with deadhearts, larval survival, larval and pupal weights, larval and pupal duration, and percentage pupation and adult emergence in diets amended with leaf powder of different sorghum genotypes. The breeding lines 27B PB 15881-3 and 463B PB 15881-3 showed antixenosis and antibiosis to C. partellus in terms of reduced eggs per plant, larval survival and development. The levels of antixenosis and antibiosis of both breeding lines were similar to their resistant parents. Results indicate that transmission of characteristics responsible for resistance to the progeny from the resistant parent occurred. Ó 2011 Elsevier Ltd. All rights reserved.
Keywords: Chilo partellus Leaf damage Growth and development Antibiosis Sorghum
1. Introduction Several species of stemborers attack sorghum (Sorghum bicolor L. Moench) in different growing regions (Nwanze, 1997), of which the spotted stemborer, Chilo partellus (Swinhoe) (Lepidoptera: Pyralidae) is the predominant species in the Indian subcontinent, and south and eastern Africa, causing serious damage to sorghum, maize (Zea mays L.), and pearl millet (Pennisetum glaucum (L.) R. Br.) (Ingram, 1958; Jotwani and Young, 1972; Singh and Rana, 1989). It causes US$ 334 million annual loss to sorghum alone in the semi-arid tropics (Sharma, 2006). It attacks sorghum from two weeks after germination until crop harvest and affects all plant parts except the roots. The first symptom of attack is the irregular shaped pinholes, caused by the early instar larvae feeding in the whorl, which later become elongated lesions on the leaves. The older larvae leave the whorl, bore into the stem and reach the growing point, where feeding results in a characteristic “deadheart” symptom. In older plants, where internode elongation has started and the growing point has moved upwards, the larva feeds inside the stem, causing extensive tunneling. It also tunnels the peduncle and moves up the earhead. Feeding and stem tunneling
* Corresponding author. Tel.: þ91 40 24015349; fax: þ91 40 20416378. E-mail address: [email protected] (P.G. Padmaja). 0261-2194/$ e see front matter Ó 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.cropro.2011.09.011
by C. partellus larvae in sorghum results in crop losses as a consequence of reduction in foliage, destruction of the growing point, early leaf senescence, interference with translocation of metabolites and nutrients that result in under development of the grain, stem breakage, reduced plant vigor, lodging, direct damage to panicles and loss in grain yield (Sharma et al., 2007). Several insect control strategies such as crop rotation, field sanitation, introduction of parasitoids and use of synthetic pesticides such as furadan have been employed, but have not fully curtailed the problem, suggesting a need for developing more sustainable and effective control measures (Sharma et al., 2007). Systematic screening of the world germplasm collection for resistance to spotted stemborer was undertaken in the Indian national sorghum improvement program (Singh et al., 1968; Pradhan, 1971; Jotwani, 1978). A number of genotypes with resistance to C. partellus have been identified, but the levels of resistance are low to moderate (Sharma et al., 2003). Amongst the identified sources, a number of mechanisms contribute to sorghum resistance to the stemborer, including non-preference for oviposition, reduced feeding by the first instars on young leaves, low deadheart formation, reduced tunneling, and tolerance to leaf damage and stem tunneling (Chapman et al., 1983; Dabrowski and Kidiavai, 1983; Woodhead and Taneja, 1987; Sharma and Nwanze, 1997). These sources can be used as parents in a resistance breeding programme to increase the levels of resistance in elite genotypes and diversify the bases of resistance to C. partellus.
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Knowledge of the resistance mechanisms and associated factors is essential for effective utilization of resistant sources in breeding programs. Antibiosis resistance affects the biology of the insect so pest abundance and subsequent damage is reduced compared to that which would have occurred on a susceptible variety. Antibiosis resistance often results in increased mortality or reduced longevity and reproduction of the insect. Antixenosis resistance affects the behavior of an insect pest and usually is expressed as non-preference of the insect for a resistant plant compared to a susceptible plant. A complex interaction of several factors determines the resistance/susceptibility of sorghum to stemborer attack. Since the available resistant sources typically have low yields, there is a need to incorporate resistance into high yielding varieties (Aruna and Padmaja, 2009). The objective of the present study was to demonstrate antibiosis and antixenosis in different sorghum genotypes and the transfer/transmission of resistance in the breeding lines. 2. Material and methods 2.1. Plant material The experimental material consists of two breeding lines, 27B PB 15881-3 and 463B PB 15881-3, with their parents’ 27B (female parent of the nationally released hybrid, CSH 16), 463B (female parent of the dual purpose hybrid, SPH 1148), PB 15881-3 (improved line for stemborer resistance) and a resistant (IS 2205) and susceptible genotype (DJ 6514) (Anonymous, 2011). To develop the breeding lines in the present study, initial crosses were made between the elite lines (27B and 463B) and the stemborer-resistant line PB 15881-3 during 2004, and F1 and F2 generations were raised. Single plant selections were made in F2, and were advanced to F3, F4, F5 and F6 based on the resistance parameters (leaf damage, deadheart formation and stem tunneling). In each generation, the progenies were planted along with their parents, resistant and susceptible genotypes. Since the two breeding lines used in this study are the derivatives of the cross between the susceptible (27B and 463B) and the resistant parents (PB 15881-3), the study of resistance mechanisms in these lines and their parents would give a clear picture of genetic transmission of the antibiosis and antixenosis mechanisms. 2.2. Maintenance of C. partellus colony The C. partellus colony was maintained in the laboratory on semi-synthetic diet (Sharma et al., 1992) under controlled conditions (27 C, 70% RH, 12:12 lightedark photoperiod). The colony was initiated from field collected larvae that were maintained in a laboratory for two generations before use in this experiment. The larvae were collected from infested sorghum fields in the RangaReddy district of AndhraPradesh, India during 2007. A volume of 250 ml of diet were dispensed while warm into 1000 ml capacity wide mouthed plastic jars (14.5 12.5 cm diameter) and the containers were left open for 2 h to allow escape of the excess moisture and then covered using a clean white cloth. The surface of the diet in each jar was first punctured in several places using a sterilized plastic rod to facilitate larval penetration. Diet infestation used surface sterilized blackhead stage eggs (dipped in 10% formaldehyde for 5 min). After diet infestation, the larvae were allowed to continue feeding undisturbed within the jars until pupation. The pupae were transferred to petri dishes and put in an emergence cage consisting of a plastic container (14.5 12.5 cm) ventilated at the top with fine mesh. The emerged moths were collected using a glass vial (5.5 2.0 cm diameter) and transferred to an oviposition cage lined with butter paper folded to form
several pleats. The moths were fed on water from a water-soaked cotton wool in a petri dish placed in each cage. Eggs oviposited on the butter papers were collected daily from the cage and fresh butter paper replaced. The C. partellus colony reared in the laboratory was used for field, greenhouse and laboratory studies. 2.3. Field experiments 2.3.1. Evaluation of resistance to C. partellus under field conditions Experiments were conducted at the Directorate of Sorghum Research, Rajendranagar, India (Latitude 17 190 28.5” N and Longitude 78 240 13.4” E) at an altitude of 524 MSL (mean above sea level) during the years 2007 and 2008. The evaluation of individual plants for stemborer resistance was conducted on the basis of leaf feeding score and stem tunneling during 2005e2007. The plants showing the least damage were selected and advanced to the next generation. The two most resistant F5 plants were selected and tested in the F6 generation during the years 2007 and 2008. The two breeding lines with their parents and resistant and susceptible genotypes were sown in two row plots of 4 m row length, 60 cm row spacing. Three replications were planted in a randomized complete block design. One week after seedling emergence, thinning was carried out to maintain a spacing of 10 cm between plants. No insecticide was applied. The recommended cultivation practices were followed to maintain good crop stand. 2.3.2. Artificial infestation of C. partellus Plants at 21 DAE were infested with five neonate larvae of C. partellus in the field. One day before infestation, paper strips with blackhead stage egg masses were transferred from the refrigerator into plastic jars containing a carrier of 80 g of poppy (Papaver sp.) seed in the evening. On the subsequent morning, the freshly hatched neonate larvae were gently mixed with the carrier, and transferred into plastic bottles fixed to the bazooka applicator (applicator developed at the International Maize and Wheat Improvement Center (CIMMYT) for field infestation was modified to suit sorghum). Plants were individually infested by placing the nozzle of the bazooka onto the leaf whorl. In each stroke, five larvae were released in the morning between 0700 h and 0900 h into the whorls to cause an optimum level of leaf damage and deadheart formation. Generally 5e7 larvae per plant are sufficient to cause appreciable leaf feeding and deadhearts (>90% damage) in susceptible genotypes (Sharma et al., 1997). Data were recorded on leaf feeding, deadheart formation due to stemborer, stem length, stem tunneling, peduncle damage, number of tunnels and number of exit holes per plant. Leaf feeding was assessed two weeks after artificial infestation on a 1 to 9 rating scale (1 ¼ no visible leaf injury or a small number of pin/shot hole type of injury on a few leaves, 2 ¼ small amount of shot hole type lesions on a few leaves, 3 ¼ shot hole injury common on several leaves, 4 ¼ several leaves with shot hole and elongated lesions, 5 ¼ several leaves with elongated lesions (<2.5 cm), 6 ¼ several leaves with elongated lesions (>2.5 cm), 7 ¼ long lesions common on about onehalf of the leaves, 8 ¼ long lesions common on about two-third of the leaves, and 9 ¼ most leaves with long lesions) based on the type and amount of feeding (Sharma et al., 1992). The number of plants with stemborer deadhearts was recorded at 35 days after seedling emergence (DAE) and expressed as a percentage of the total number of plants. Stalk length was measured from five randomly selected plants per replication from the base of the plant to the last node from where panicle peduncle starts, and the data were denoted as stem length (cm). From the infested plots, five randomly selected plants were cut at the base before harvest, and exit hole counts were recorded per plant after removing the sheath leaves (Sharma et al., 2007). The stems of five randomly selected plants were split open
P.G. Padmaja et al. / Crop Protection 31 (2012) 21e26
to determine the tunnel length per plant (cm) caused by C. partellus, and represented as tunneling percentage in relation to stem length. Data on leaf feeding score, deadhearts, stemtuneling, peduncle tunneling, number of tunnels and exit holes per plant were pooled for both years (2007 and 2008).
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the resistant, IS 2205 and susceptible, DJ 6514 genotypes. The sorghum seedlings were grown under greenhouse conditions. The plants were raised on medium sized pots (60 cm diameter 30 cm deep) in the greenhouse. At 10 days after seedling emergence, four healthy seedlings were retained in each pot. Antibiosis to C. partellus in different sorghum genotypes was studied by incorporating dried leaf powder of the genotypes into the diet. For this purpose, leaves of the test genotypes were collected from 21 day old plants raised under greenhouse conditions. Two to three whorl leaves were removed with scissors at the growing point from each plant on which the larvae feed under natural conditions. The leaves were washed and then shade dried. The dried leaves were powdered in a blender for use in the artificial diet. One set of diet was prepared for each genotype and was replicated thrice in a completely randomized design. Diet was poured into 100 ml capacity plastic cups and each cup had 50 ml diet. After pouring the diet into the cups, it was allowed to cool for 2e3 h on the laboratory table. Ten first instar larvae were released into each cup, using a fine camel hairbrush. The cups were kept in the rearing room in the dark for as the neonate larvae have a strong photosensitive behavior, and settle better on the diet in darkness. In the rearing room, temperature was maintained at 27 C, 70% RH, and 12 h photoperiod. Observations were recorded on larval survival and larval weight at 10 days after releasing the larvae into the artificial diet, pupation and adult emergence. Pupal weight was recorded for each sex separately on the second day after pupation. The pupae were sexed on the basis of their relative size and genital openings (Sithanantham and Subramaniam, 1975).
2.4. Greenhouse experiments 2.4.1. Oviposition preference of C. partellus Oviposition preference was studied under controlled conditions (27 C, 70% RH, and 12 h photoperiod). Seven genotypes were grown in 21 pots (30 cm diameter and 45 cm deep) in the greenhouse. Four seeds were sown in each pot and watered immediately. Twenty day old plants, the stage at which the stemborer females lay eggs on plants under natural conditions, were placed on a table in a randomized complete block design with three replications, and covered with a nylon net (2.0 m 1.0 m 0.6 m). Twenty pairs of newly emerged adults were released inside the nylon net. The moths were provided with water in a cotton swab, and allowed to oviposit on the test genotypes for three days. The number of egg masses and number of eggs laid were recorded on each genotype. 2.4.2. Growth and development of C. partellus under greenhouse conditions Survival and development of neonate larvae of C. partellus were studied on 21-day old seedlings under greenhouse conditions. Four plants were grown in each pot. The two breeding lines with their parents and resistant and susceptible genotypes were infested with 10 neonate larvae per plant at 21 days after seedling emergence with a bazooka applicator. A randomized complete block design was used with three replications. Larval survival was recorded 10 days after infestation, as this is the time larvae successfully bore into the stem after feeding on the whorls. Moreover, at 25 days after infestation, most of the larvae will start pupating in the susceptible genotypes. At 25 days after infestation, the plants were dissected to remove the larvae/pupae, which were then placed in a plastic jar along with 15 cm pieces of sorghum stems of the same genotype. Data were recorded on pupal weights and post-embryonic development period. Pupal weights were recorded separately for the males and females one day after pupation.
2.6. Statistical analysis The data obtained from field, greenhouse and laboratory experiments were subjected to analysis of variance using the statistical software Windostat (Indostat, 2004). Genotypes were used as fixed effects and blocks and years as random effects. The data on percentages were subjected to arcsine transformation. The data on number of tunnels, peduncle tunneling percentage, exit holes, number of egg masses and number of eggs per plant and larval weights were subjected to square root transformations. LSD was used to compare the treatment means. 3. Results
2.5. Laboratory experiment 3.1. Evaluation of resistance to C. partellus under field conditions 2.5.1. Growth and development of C. partellus on artificial diet The survival and development of C. partellus was studied on artificial diet with leaf powders of 27B PB 15881-3-1 and 463B PB 15881-3, their parents (27B, 463B and PB 15881-3) and
There were significant differences among genotypes for leaf feeding injury, percentage deadhearts, number of holes and tunnels and percentage tunneling (Table 1). The breeding lines, 27B PB
Table 1 Performance of breeding lines for resistance to spotted stemborer, C. partellus, in sorghum. Genotype
Leaf feeding score (1e9)
27B 463B PB 15881-3 27B PB 15881-3 463B PB 15881-3 IS 2205 DJ 6514
8.3 7.4 5.2 5.8 5.7 5.9 8.0
Mean SEM CV F ratio P
6.6 0.22 5.9 30.71 <0.0001
0.2 0.2 0.2 0.5 0.2 0.1 0.1
Treatment means were compared using LSD ddf ¼ 6,14.
Deadhearts (%)
Stem tunneling (%)
45.3 41.4 32.8 28.9 28.9 27.8 48.3
45.0 50.8 27.8 27.2 27.8 25.3 58.9
2.5 1.2 4.3 2.6 1.6 1.4 1.9
36.4 2.08 9.9 16.6 <0.0001
1.6 6.8 1.4 1.1 1.3 2.9 1.2
37.5 3.0 13.9 20.9 <0.0001
Peduncle tunneling (%) 3.6 3.4 2.8 2.5 2.6 3.0 3.6
0.1 0.1 0.1 0.1 0.1 0.1 0.2
3.1 0.05 2.7 91.3 <0.0001
Tunnels plant1 (no.) 2.4 2.0 1.6 1.7 1.7 1.4 2.1
0.1 0.1 0.1 0.0 0.1 0.0 0.1
1.8 0.04 4.2 53.2 <0.0001
Exit holes plant1 (no.) 3.0 3.0 2.0 2.2 2.3 1.7 2.8
0.0 0.2 0.1 0.1 0.1 0.2 0.1
2.4 0.06 4.1 80.2 <0.0001
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15881-3 and 463B PB 15881-3 were as good as the resistant genotype IS 2205 and their resistant parent PB 15881-3 for stemborer resistance (leaf feeding score 5.2e5.8), whereas the leaf feeding was greater on the susceptible parents, 27B and 463B (7.4e8.3). The genotypes 27B PB 15881-3 and 463B PB 15881-3 had significantly fewer deadhearts had reduced stem and peduncle tunneling compared to their parents 27B and 463B (Table 1). 3.2. Oviposition preference The oviposition preference of C. partellus differed significantly according to genotype. Significantly more eggs masses and total number of eggs were laid on 27B, 463B and DJ 6514 than on IS 2205 and PB 15881-3. The genotypes 27B PB 15881-3-1 and 463B PB 15881-3, IS 2205 and PB 15881-3 were least preferred for oviposition. These genotypes had 131 to 146 eggs per plant compared to 320 eggs on 27B (Table 2). 3.3. Growth and development There were significant differences in larval survival, pupal weights, and post-embryonic development periods of C. partellus on different sorghum genotypes. Larval survival at ten days after infestation varied from 51 to 76% (Table 3). Larval survival was >70% on 27B, DJ 6514 and 463B, 51% on IS 2205, as compared to 54e56% on breeding lines. The weights of both male and female pupae were lower in insects reared on IS 2205, breeding lines and PB 15881-3 as compared to those reared on 27B, 463B and DJ 6514. Post-embryonic development period was >50 days for males and >60 days for females when the larvae were reared on 27B PB 15881-3 and 463 PB 15881-3, compared to 42.0 days for the males and 43.2 days for the females on 27B. Pupal weights were greatest when larvae were reared on 27B, DJ 6514 and 463B. A better consumption and utilization of the cultivars 27B and DJ 6514 was shown as expressed by the higher weights attained by pupae grown on these two than on PB 15881-3 and IS 2205. 3.4. Survival and development of C. partellus on artificial diet Larval survival and larval weight of C. partellus reared on artificial diet with leaf powder showed significant differences between the genotypes tested. Larval survival was >70% in diets with leaf powder of 27B and DJ 6514 compared to 50% survival in artificial diet amended with leaf powder of 27B PB 15881-3 and 463B PB 15881-3 (Table 4). The larval weight was 5.1e5.6 mg on artificial diets with 27B PB 15881-3, 463B PB 15881-3 and PB 15881-3 Table 2 Oviposition preference by the spotted stemborer, C. partellus, toward sorghum genotypes under multi-choice cage conditions. Genotype
No. of egg masses per plant
27B 463B PB 15881-3 27B PB 15881-3 463B PB 15881-3 IS 2205 DJ 6514
6.0 5.0 3.5 2.0 2.3 3.0 5.0
Mean SEM CV F ratio P
3.83 0.25 11.33 37.04 <0.0001
0.3 0.5 0.3 0.2 0.2 0.3 0.4
Treatment means were compared using LSD ddf ¼ 6,14.
No. of eggs per plant 320 285 128 131 146 125 301
12.7 21.4 6.9 9.1 15.0 4.0 13.8
205.67 14.03 11.82 41.97 <0.0001
leaf powder compared to 7.4e7.9 mg on 463B, DJ 6514 and 27B. Low larval survival and larval weight at 10 DAI were recorded on IS 2205, PB 15881-3, 27B PB 15881-3 and 463B PB 15881-3. Pupal weight varied from 51.9 to 71.6 mg for the males and from 96.4 to 142.5 mg for the females. Significantly lower pupal weight was observed in male and female pupae from the larvae reared on diets with leaf powder of 27B PB 15881-3 and 463B PB 15881-3 as compared to those insects reared on their parents. Percentage pupation varied from 41.5 to 62.3%, while the adult emergence varied from 30.9% to 44.3%. Comparatively lower pupation (45e48%) and adult emergence (32e33%) were recorded in artificial diets impregnated with leaf powder of 27B PB 15881-3 and 463B PB 15881-3 as compared to those reared on diets with leaf powder of 27B, 463B and DJ 6514 (52e62% pupation and 37e44% adult emergence). The larval period varied from 24.6 to 37.1 days for males, and from 25.1 to 39.4 days for the females on artificial diet impregnated with leaf powder from different sorghum genotypes (Table 4). Larval duration for the males was prolonged by 12 days on 27B PB 15881-3 and 463B PB 15881-3 as compared to 27B. Duration of larval period for the females was significantly longer on 27B PB 15881-3 and 463B PB 15881-3 as compared to the larvae reared on 27B. Pupal period for the males was 8.0 days on 27B PB 15881-3 and 8.8 days on 463B PB 15881-3, compared to 7.1 days on 27B. 4. Discussion Selections for resistance to C. partellus based on leaf injury, deadhearts and stem tunneling were made in the F3eF6 generations of two sorghum crosses. The plants selected in each generation were then subject to further screening. Both the breeding lines were significantly more resistant than the respective susceptible parents and DJ 6514 based on leaf injury, deadhearts and stem tunneling. Both the lines were found to be close to their resistant parent PB 15881-3 with regard to these parameters. Thus, it was considered that continuous screening for leaf injury, deadhearts and stem tunneling over several generations would prove to be more effective in improving the level of resistance. Stem tunneling had earlier been reported to be the major plant character associated with resistance to stemborer (Kishore, 1991). Stem tunneling rather than leaf feeding and deadhearts is the primary cause of yield loss (Alghali, 1986). However, these are not the only damage parameters responsible for yield reduction in sorghum (Pathak and Olela, 1983; Singh et al., 1983; Taneja and Leuschner, 1985). The number of exit holes per plant is another criterion of resistance, which indicates suitability of the genotype for the larva to complete development and successfully emerge from the stalk for pupation. Significant differences in oviposition were observed among the sorghum genotypes, with the breeding lines being relatively less preferred for oviposition. Oviposition preference has earlier been identified to be one of the components of resistance to C. partellus in sorghum (Singh and Rana, 1984; Alghali, 1985; Saxena, 1990; van den Berg and van der Westhuizen, 1997; Kishore Kumar et al., 2007). Most of these studies were aimed at identifying sorghum lines with resistance to stemborer under field or greenhouse conditions. Comparison of the biology of C. partellus when reared on different lines showed that rearing on the 27B PB 15881-3 and 463B PB 15881-3 resulted in lower larval and pupal weights, a longer pupal period, fewer larvae completing development, and smaller, fewer females. Antibiosis in terms of reduced larval survival and prolongation of post-embryonic development period of C. partellus has earlier been reported by Jotwani et al. (1978), Woodhead and Taneja (1987), van den Berg and van der Westhuizen (1997) and Kishore Kumar et al. (2007). Adverse
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Table 3 Development of neonate larvae of spotted stemborer, C. partellus, on seedlings of sorghum genotypes under greenhouse conditions. Genotype
Larval survival (%) at 10 DAI
Pupal weight (mg) Males
27B 463B PB 15881-3 27B x PB 15881-3 463B x PB 15881-3 IS 2205 DJ 6514
76.0 71.6 53.8 54.3 56.2 51.4 73.0
4.5 0.5 1.7 1.4 0.7 1.4 3.0
Mean SEM CV F ratio P
62.3 1.62 4.5 42.9 <0.0001
47.4 44.6 39.2 32.2 34.3 36.7 46.2
Post-embryonic development period (days) Females
2.2 2.5 4.7 0.7 0.2 3.1 2.4
96.4 87.4 71.2 71.2 75.3 70.6 92.8
40.09 2.76 11.90 4.80 <0.0001
3.6 1.6 4.0 1.6 2.6 0.9 1.2
80.7 1.89 4.07 34.75 0.0101
Males 42.0 44.0 50.1 55.7 52.6 53.8 44.3
Females 0.6 1.2 1.3 1.4 1.8 2.4 2.4
43.2 48.5 59.9 61.8 60.4 59.1 49.3
48.92 1.79 6.33 9.30 <0.0001
1.4 3.5 4.3 2.1 1.9 4.4 0.4
54.61 1.74 5.53 18.02 0.0006
DAI ¼ Days after infestation. Treatment means were compared using LSD ddf ¼ 6,14.
effects of stemborer-resistant lines on survival and development of C. partellus under field conditions have been reported by Lal and Pant (1980), Singh and Verma (1988), and Woodhead and Taneja (1987). Genotypic differences in larval establishment in the field have been reported on different sorghum genotypes (Jotwani et al., 1978; Woodhead and Taneja, 1987; Singh and Rana, 1989; van den Berg and van der Westhuizen, 1997). However, expression of resistance to C. partellus under field conditions can be quite variable due to variation in environmental conditions, differential growth of the test genotypes, and the nutrient status of the soil. Dried leaf powder amended into the artificial diet is helpful to overcome the variation in borer infestation observed under field conditions to allow a comparison of the test genotypes under uniform conditions. There was considerable variation in larval survival, larval and pupal weight, larval and pupal development period, and percentage pupation and adult emergence in diets amended with leaf powder of different sorghum genotypes. Low larval survival, larval and pupal weights, low pupation and adult emergence were recorded in artificial diets amended with dried leaf powder of 27B PB 15881-3, 463B PB 15881-3, PB 15881-3 and IS 2205. Larval, pupal, and the total development period is also prolonged (Jotwani et al., 1978; Lal and Sukhani, 1979, 1982; Singh and Rana, 1984, 1989; Saxena, 1990, 1992; Verma et al., 1992). Antibiosis can also be expressed in terms of reduced pupal weight (Lal and Sukhani, 1982; Singh and Rana, 1984; Singh and Verma, 1988; Verma et al., 1992; Kishore Kumar et al., 2006) and low pupation and adult emergence (Singh and Verma, 1988). The antibiotic effects of the resistant genotypes on the development of
C. partellus may be caused by secondary plant substances in the leaves and/or poor nutritional quality of the food. Reduced survival and establishment will reduce the insect population and the resultant crop damage. Prolongation of development period will also result in reduction of number of generations in a season/year. These effects of the resistant varieties will have a constant and cumulative effect on the stemborer population over the season (Sharma, 1993), and can be utilized as an environmental friendly method to reduce the damage by this pest under subsistence farming conditions. Results from this study indicated significant differences between resistant and susceptible genotypes with regard to reduction in foliar damage ratings, percentage stem length tunneled, and percentage of plants showing deadhearts, oviposition preference, larval survival, larval and pupal mass and percentage pupation and adult emergence. It has been concluded that slow development of larvae on resistant lines is due to antibiosis. The possibility of selection of parents to breed should therefore be based on antixenosis and antibiosis to increase the levels of resistance to C. partellus in sorghum. Therefore, although a complex interaction of different factors affect resistance/susceptibility of sorghum to stemborer attack, differences in level of preference and performance among resistant and susceptible genotypes strongly support antixenosis and antibiosis as major mechanisms of resistance to C. partellus. The study clearly indicates the genetic transmission of antixenosis and antibiosis resistance mechanisms from resistant parent to the progeny. It also shows that by utilizing the resistant lines in the crossing program and consciously selecting resistant
Table 4 Growth and development of spotted stemborer, C. partellus, larvae on semi-synthetic diet amended with leaf powder of different sorghum genotypes. Genotypes
Larval survival (%) 10 DAI
27B 463B PB 15881-3 27B PB 15881-3 463B PB 15881-3 IS 2205 DJ 6514
72.8 71.0 52.2 50.8 51.0 51.5 71.6
Mean SEM CV F ratio P
60.1 3.32 9.6 10.9 0.0003
2.4 7.6 1.7 0.7 0.2 1.4 1.1
Larval weight (mg) 10 DAI 7.9 7.4 5.0 5.1 5.6 4.6 7.6
0.1 0.2 0.5 0.3 0.2 0.4 0.3
6.17 0.33 9.15 18.57 <0.0001
Larval development time
Pupal weight (mg)
Male
Males
24.6 25.1 36.4 37.0 35.1 37.1 27.3
Female
31.77 2.57 14.01 5.21 0.0074
1.8 0.5 2.4 2.8 2.9 4.0 1.0
25.2 25.9 37.9 39.0 36.4 39.4 25.1
2.9 2.6 4.5 4.6 1.0 0.4 2.9
32.7 2.22 11.76 9.65 0.0005
DAI Days after infestation. Treatment means were compared using LSD ddf ¼ 6,14.
71.6 62.1 53.5 52.7 58.4 51.9 68.1
59.76 3.49 10.10 5.05 0.0084
Females 4.9 1.8 2.0 1.6 4.8 3.9 1.2
Pupation (%)
142.5 1.4 127.4 7.4 98.6 1.8 104.1 1.8 108.7 5.0 96.4 3.6 131.4 1.8
62.3 52.3 44.1 45.0 48.2 41.5 57.8
115.59 3.99 5.97 20.39 <0.0001
50.3 0.72 2.5 113.7 <0.0001
3.6 2.0 2.4 2.7 2.4 2.9 1.2
Pupal development time
Adult emergence (%)
Males
Females
7.1 0.1 7.3 0.2 9.5 0.2 8.0 0.1 8.8 0.2 10.7 0.7 7.8 0.2
7.0 0.0 7.4 0.2 9.2 0.1 8.1 0.1 8.9 0.5 10.4 0.2 8.2 0.2
44.3 37.1 32.0 32.2 33.1 30.9 42.1
8.46 0.23 4.78 31.15 <0.0001
8.45 0.25 5.12 21.15 <0.0001
36.0 1.74 8.4 9.5 0.0005
1.4 2.1 1.0 2.4 2.1 1.1 1.4
26
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progenies in the subsequent generations, there is every possibility of obtaining improved lines with resistance on par with the resistant source. Thus, isolating, characterizing and developing marker systems for the identified resistant factors will greatly aid rapid screening. References Alghali, A.M., 1985. Insect-host plant relationships. The spotted stalk-borer, Chilo partellus (Swinhoe) (Lepidoptera: Pyralidae) and its principal host, sorghum. Insect Sci. Appl. 6, 315e322. Alghali, A.M., 1986. Effect of cultivar, time, and amount of Chilo partellus Swinhoe Lepidoptera: Pyralidae infestation on sorghum yield components in Kenya. Trop. Pest Manage. 32, 126e129. Anonymous (AICSIP, All India co-ordinated sorghum improvement project) 2010. Progress report: Entomology section. In XXXXI Annual group meeting held at University of Agricultural Sciences, Dharwad, India. April 15e17, 2011. Aruna, C., Padmaja, P.G., 2009. Evaluation of genetic potential of shoot fly resistant sources in sorghum (Sorghum bicolor (L.) Moench). J. Agric. Sci. 147, 71e80. Chapman, R.F., Woodhead, S., Bernays, E.A., 1983. Survival and dispersal of young larvae of Chilo partellus (Swinhoe) (Lepidoptera: Pyralidae) in two cultivars of sorghum. Bull. Entomol. Res. 73, 65e74. Dabrowski, Z.T., Kidiavai, E.L., 1983. Resistance of some sorghum lines to the spotted stalk borer, Chilo partellus, under western Kenyan conditions. Insect Sci. Appl. 4, 119e126. Indostat services, 2004. Windostat. Hyderabad. Indostat services, India. Ingram, W.R., 1958. The lepidopterous stalk borers associated with Gramineae in Uganda. Bull. Entomol. Res. 49, 367e383. Jotwani, M.G., 1978. Investigations on Insect Pests of Sorghum and Millets with Special Reference to Host Plant Resistance. Final Technical Report (1972e1977). Research Bulletin of the Division of Entomology. Indian Agricultural Research Institute, New Delhi, India, p. 114. Jotwani, M.G., Young, W.R., 1972. Recent developments in chemical control of insect pests of sorghum. In: Rao, N.G.P., House, L.R. (Eds.), Sorghum in Seventies. Oxford & IBH Publishing, New Delhi, India, pp. 251e256. Jotwani, M.G., Chaudhari, S., Singh, S.P., 1978. Mechanism of resistance to Chilo partellus (Swinhoe) in sorghum. Indian J. Entomol. 40, 273e276. Kishore, P., 1991. Relationship between parameters for damage (leaf injury and stem tunnelling) caused by the stem borer, Chilo partellus (Swinhoe) on grain yield of different sorghum cultivars. J. Entomol. Res. 15, 236e241. Kishore Kumar, V., Sharma, H.C., Dharma Reddy, K., 2006. Antibiosis mechanism of resistance to spotted stem borer, Chilo partellus in sorghum, Sorghum bicolor. Crop Prot. 25, 66e72. Kishore Kumar, V., Dharma Reddy, K., Sharma, H.C., 2007. Expression of antixenosis and antibiosis components of resistance to spotted stem borer Chilo partellus in sorghum under greenhouse conditions. SAT eJournal 3 (1). ejournal.icrisat.org. Lal, G., Pant, J.C., 1980. Laboratory and field testing for resistance in maize and sorghum varieties to Chilo partellus (Swinhoe). Indian J. Entomol. 42, 606e610. Lal, G., Sukhani, T.R., 1979. Development of sorghum stem borer Chilo partellus (Swinhoe) larvae on some resistant lines of sorghum. Bull. Entomol. 20, 67e70. Lal, G., Sukhani, T.R., 1982. Antibiotic effects of some resistant lines of sorghum on postlarval development of Chilo partellus Swinhoe. Indian J. Agric. Sci. 52, 127e129. Nwanze, K.F., 1997. Integrated management of stemborers of sorghum and pearl millet. Insect Sci. Appl. 17, 1e8. Pathak, R.S., Olela, J.C., 1983. Genetics of host plant resistance in food crops with special reference to sorghum stem borers. Insect Sci. Appl. 4, 127e134.
Pradhan, S., 1971. Investigations on Insect Pests of Sorghum and Millets. Final Technical Report (1965e1970). Division of Entomology. Indian Agricultural Research Institute, New Delhi, India, p. 157. Saxena, K.N., 1990. Mechanisms of resistance/susceptibility of certain sorghum cultivars to the stem borer Chilo partellus: role of behaviour and development. Entomol. Exp. Appl. 55, 91e99. Saxena, K.N., 1992. Larval development of Chilo partellus (Swinhoe) (Lepidoptera: Pyralidae) on artificial diet incorporating leaf tissues of sorghum lines in relation to their resistance or susceptibility. Appl. Entomol. Zool. 27, 325e330. Sharma, H.C., 1993. Host plant resistance to insects in sorghum and its role in integrated pest management. Crop Prot. 12, 11e37. Sharma, H.C., 2006. Integrated Pest Management Research at ICRISAT. Present Status and Future Priorities. International Research Institute for the semi-Arid Tropics, Patancheru. 502324, AP, India, p. 48. Sharma, H.C., Nwanze, K.F., 1997. Mechanisms of Resistance to Insects in Sorghum and Their Usefulness in Crop Improvement. Information Bulletin No. 45. International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), Patancheru, Andhra Pradesh, India, p. 56. Sharma, H.C., Singh, Faujdar, Nwanze, K.F., 1997. Plant Resistance to Insects in Sorghum. International Crops Research Institute for the Semi-Arid Tropics, Patancheru 502 324, Andhra Pradesh, India, p. 216. ISBM 92-9066-382-0. Sharma, H.C., Taneja, S.L., Leuschner, K., Nwanze, K.F., 1992. Techniques to Screen Sorghums for Resistance to Insect Pests. Information Bulletin No. 32. Int. Crops Res. Inst. forernational Crops Research Institute for the Semi-Arid Tropics, Patancheru, Andhra Pradesh, India, p. 48. Sharma, H.C., Taneja, S.L., Kameshwara Rao, N., Prasada Rao, K.E., 2003. Evaluation of Sorghum Germplasm for Resistance to Insects. International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), Patancheru 502 324, Andhra Pradesh, India. Information Bulletin No. 63p. 184. Sharma, H.C., Dhillon, M.K., Pampapathy, G., Reddy, B.V.S., 2007. Inheritance of resistance to spotted stem borer, Chilo partellus in sorghum, Sorghum bicolor. Euphytica 156, 117e128. Singh, B.U., Rana, B.S., 1984. Influence of varietal resistance on oviposition and larval development of stalk-borer Chilo partellus Swinhoe and its relationship to field resistance in sorghum. Insect Sci. Appl. 5, 287e296. Singh, B.U., Rana, B.S., 1989. Varietal resistance in sorghum to spotted stem borer, Chilo partellus (Swinhoe). Insect Sci. Appl. 10, 3e27. Singh, S.P., Verma, A.N., 1988. Antibiosis mechanism of resistance to stem borer, Chilo partellus (Swinhoe) in sorghum. Insect Sci. Appl. 9, 579e582. Singh, B.U., Rana, B.S., Reddy, B.B., Rao, N.G.P., 1983. Host plant resistance to stalkborer, Chilo partellus Swinhoe in sorghum. Insect Sci. Appl. 4, 407e414. Singh, S.R., Vedamoorthy, G., Thobbi, V.V., Jotwani, M.G., Young, W.R., Balan, J.S., Srivastava, K.P., Sandhu, G.S., Krishnananda, N., 1968. Resistance to stem borer, Chilo zonellus (Swinhoe) and stem fly, Atherigona varia soccata Rond. in the world sorghum collection in India. Mem. Entomol. Soc. India 7, 1e79. Sithanantham, S., Subramaniam, T.R., 1975. Sex determination of pupae of sorghum stalk borer, Chilo zonellus (Swinhoe). Madras Agri. J. 62, 62e63. Taneja, S.L., Leuschner, K., 1985. Methods of rearing, infestation and evaluation for Chilo partellus resistance in sorghum. In: Proceedings of International Symposium on Sorghum Entomology July 15e21, 1984. Texas A and M University, College Station, Texas, USA. International Crops Research Institute for the SemiArid Tropics (ICRISAT), Patancheru, Andhra Pradesh, India, pp. 175e188. van den Berg, J., van der Westhuizen, M.C., 1997. Chilo partellus (Lepidoptera: Pyralidae) moth and larval response to levels of antixenosis and antibiosis in sorghum inbred lines under laboratory conditions. Bull. Entomol. Res. 87, 541e545. Verma, O.P., Bhanot, J.P., Verma, A.N., 1992. Development of Chilo partellus (Swinhoe) on pest resistant and susceptible sorghum cultivars. J. Insect Sci. 5, 181e182. Woodhead, S., Taneja, S.L., 1987. The importance of the behaviour of young larvae in sorghum resistance to Chilo partellus. Entomol. Exp. Appl. 45, 47e54.
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Evidence of genetic transmission of antibiosis and antixenosis resistance of sorghum to the spotted stemborer, Chilo partellus (Lepidoptera: Pyralidae) P.G. Padmaja*, C. Aruna, J.V. Patil Directorate of Sorghum Research, Rajendranagar, Hyderabad 500 030, Andhra Pradesh, India
a r t i c l e i n f o
a b s t r a c t
Article history: Received 22 January 2011 Received in revised form 20 September 2011 Accepted 20 September 2011
Spotted stemborer, Chilo partellus (Swinhoe) (Lepidoptera: Pyralidae), is the most important pest of sorghum in Asia and south and eastern Africa. Host plant resistance is an important control tactic for controlling this pest. Two breeding lines 27B PB 15881-3 and 463B PB 15881-3 with their parents, resistant and susceptible genotypes were evaluated in the field, glasshouse and laboratory for different resistance parameters. Breeding lines and genotypes varied significantly in foliar damage ratings, percentage of stem length tunneled, percentage of plants with deadhearts, larval survival, larval and pupal weights, larval and pupal duration, and percentage pupation and adult emergence in diets amended with leaf powder of different sorghum genotypes. The breeding lines 27B PB 15881-3 and 463B PB 15881-3 showed antixenosis and antibiosis to C. partellus in terms of reduced eggs per plant, larval survival and development. The levels of antixenosis and antibiosis of both breeding lines were similar to their resistant parents. Results indicate that transmission of characteristics responsible for resistance to the progeny from the resistant parent occurred. Ó 2011 Elsevier Ltd. All rights reserved.
Keywords: Chilo partellus Leaf damage Growth and development Antibiosis Sorghum
1. Introduction Several species of stemborers attack sorghum (Sorghum bicolor L. Moench) in different growing regions (Nwanze, 1997), of which the spotted stemborer, Chilo partellus (Swinhoe) (Lepidoptera: Pyralidae) is the predominant species in the Indian subcontinent, and south and eastern Africa, causing serious damage to sorghum, maize (Zea mays L.), and pearl millet (Pennisetum glaucum (L.) R. Br.) (Ingram, 1958; Jotwani and Young, 1972; Singh and Rana, 1989). It causes US$ 334 million annual loss to sorghum alone in the semi-arid tropics (Sharma, 2006). It attacks sorghum from two weeks after germination until crop harvest and affects all plant parts except the roots. The first symptom of attack is the irregular shaped pinholes, caused by the early instar larvae feeding in the whorl, which later become elongated lesions on the leaves. The older larvae leave the whorl, bore into the stem and reach the growing point, where feeding results in a characteristic “deadheart” symptom. In older plants, where internode elongation has started and the growing point has moved upwards, the larva feeds inside the stem, causing extensive tunneling. It also tunnels the peduncle and moves up the earhead. Feeding and stem tunneling
* Corresponding author. Tel.: þ91 40 24015349; fax: þ91 40 20416378. E-mail address: [email protected] (P.G. Padmaja). 0261-2194/$ e see front matter Ó 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.cropro.2011.09.011
by C. partellus larvae in sorghum results in crop losses as a consequence of reduction in foliage, destruction of the growing point, early leaf senescence, interference with translocation of metabolites and nutrients that result in under development of the grain, stem breakage, reduced plant vigor, lodging, direct damage to panicles and loss in grain yield (Sharma et al., 2007). Several insect control strategies such as crop rotation, field sanitation, introduction of parasitoids and use of synthetic pesticides such as furadan have been employed, but have not fully curtailed the problem, suggesting a need for developing more sustainable and effective control measures (Sharma et al., 2007). Systematic screening of the world germplasm collection for resistance to spotted stemborer was undertaken in the Indian national sorghum improvement program (Singh et al., 1968; Pradhan, 1971; Jotwani, 1978). A number of genotypes with resistance to C. partellus have been identified, but the levels of resistance are low to moderate (Sharma et al., 2003). Amongst the identified sources, a number of mechanisms contribute to sorghum resistance to the stemborer, including non-preference for oviposition, reduced feeding by the first instars on young leaves, low deadheart formation, reduced tunneling, and tolerance to leaf damage and stem tunneling (Chapman et al., 1983; Dabrowski and Kidiavai, 1983; Woodhead and Taneja, 1987; Sharma and Nwanze, 1997). These sources can be used as parents in a resistance breeding programme to increase the levels of resistance in elite genotypes and diversify the bases of resistance to C. partellus.
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Knowledge of the resistance mechanisms and associated factors is essential for effective utilization of resistant sources in breeding programs. Antibiosis resistance affects the biology of the insect so pest abundance and subsequent damage is reduced compared to that which would have occurred on a susceptible variety. Antibiosis resistance often results in increased mortality or reduced longevity and reproduction of the insect. Antixenosis resistance affects the behavior of an insect pest and usually is expressed as non-preference of the insect for a resistant plant compared to a susceptible plant. A complex interaction of several factors determines the resistance/susceptibility of sorghum to stemborer attack. Since the available resistant sources typically have low yields, there is a need to incorporate resistance into high yielding varieties (Aruna and Padmaja, 2009). The objective of the present study was to demonstrate antibiosis and antixenosis in different sorghum genotypes and the transfer/transmission of resistance in the breeding lines. 2. Material and methods 2.1. Plant material The experimental material consists of two breeding lines, 27B PB 15881-3 and 463B PB 15881-3, with their parents’ 27B (female parent of the nationally released hybrid, CSH 16), 463B (female parent of the dual purpose hybrid, SPH 1148), PB 15881-3 (improved line for stemborer resistance) and a resistant (IS 2205) and susceptible genotype (DJ 6514) (Anonymous, 2011). To develop the breeding lines in the present study, initial crosses were made between the elite lines (27B and 463B) and the stemborer-resistant line PB 15881-3 during 2004, and F1 and F2 generations were raised. Single plant selections were made in F2, and were advanced to F3, F4, F5 and F6 based on the resistance parameters (leaf damage, deadheart formation and stem tunneling). In each generation, the progenies were planted along with their parents, resistant and susceptible genotypes. Since the two breeding lines used in this study are the derivatives of the cross between the susceptible (27B and 463B) and the resistant parents (PB 15881-3), the study of resistance mechanisms in these lines and their parents would give a clear picture of genetic transmission of the antibiosis and antixenosis mechanisms. 2.2. Maintenance of C. partellus colony The C. partellus colony was maintained in the laboratory on semi-synthetic diet (Sharma et al., 1992) under controlled conditions (27 C, 70% RH, 12:12 lightedark photoperiod). The colony was initiated from field collected larvae that were maintained in a laboratory for two generations before use in this experiment. The larvae were collected from infested sorghum fields in the RangaReddy district of AndhraPradesh, India during 2007. A volume of 250 ml of diet were dispensed while warm into 1000 ml capacity wide mouthed plastic jars (14.5 12.5 cm diameter) and the containers were left open for 2 h to allow escape of the excess moisture and then covered using a clean white cloth. The surface of the diet in each jar was first punctured in several places using a sterilized plastic rod to facilitate larval penetration. Diet infestation used surface sterilized blackhead stage eggs (dipped in 10% formaldehyde for 5 min). After diet infestation, the larvae were allowed to continue feeding undisturbed within the jars until pupation. The pupae were transferred to petri dishes and put in an emergence cage consisting of a plastic container (14.5 12.5 cm) ventilated at the top with fine mesh. The emerged moths were collected using a glass vial (5.5 2.0 cm diameter) and transferred to an oviposition cage lined with butter paper folded to form
several pleats. The moths were fed on water from a water-soaked cotton wool in a petri dish placed in each cage. Eggs oviposited on the butter papers were collected daily from the cage and fresh butter paper replaced. The C. partellus colony reared in the laboratory was used for field, greenhouse and laboratory studies. 2.3. Field experiments 2.3.1. Evaluation of resistance to C. partellus under field conditions Experiments were conducted at the Directorate of Sorghum Research, Rajendranagar, India (Latitude 17 190 28.5” N and Longitude 78 240 13.4” E) at an altitude of 524 MSL (mean above sea level) during the years 2007 and 2008. The evaluation of individual plants for stemborer resistance was conducted on the basis of leaf feeding score and stem tunneling during 2005e2007. The plants showing the least damage were selected and advanced to the next generation. The two most resistant F5 plants were selected and tested in the F6 generation during the years 2007 and 2008. The two breeding lines with their parents and resistant and susceptible genotypes were sown in two row plots of 4 m row length, 60 cm row spacing. Three replications were planted in a randomized complete block design. One week after seedling emergence, thinning was carried out to maintain a spacing of 10 cm between plants. No insecticide was applied. The recommended cultivation practices were followed to maintain good crop stand. 2.3.2. Artificial infestation of C. partellus Plants at 21 DAE were infested with five neonate larvae of C. partellus in the field. One day before infestation, paper strips with blackhead stage egg masses were transferred from the refrigerator into plastic jars containing a carrier of 80 g of poppy (Papaver sp.) seed in the evening. On the subsequent morning, the freshly hatched neonate larvae were gently mixed with the carrier, and transferred into plastic bottles fixed to the bazooka applicator (applicator developed at the International Maize and Wheat Improvement Center (CIMMYT) for field infestation was modified to suit sorghum). Plants were individually infested by placing the nozzle of the bazooka onto the leaf whorl. In each stroke, five larvae were released in the morning between 0700 h and 0900 h into the whorls to cause an optimum level of leaf damage and deadheart formation. Generally 5e7 larvae per plant are sufficient to cause appreciable leaf feeding and deadhearts (>90% damage) in susceptible genotypes (Sharma et al., 1997). Data were recorded on leaf feeding, deadheart formation due to stemborer, stem length, stem tunneling, peduncle damage, number of tunnels and number of exit holes per plant. Leaf feeding was assessed two weeks after artificial infestation on a 1 to 9 rating scale (1 ¼ no visible leaf injury or a small number of pin/shot hole type of injury on a few leaves, 2 ¼ small amount of shot hole type lesions on a few leaves, 3 ¼ shot hole injury common on several leaves, 4 ¼ several leaves with shot hole and elongated lesions, 5 ¼ several leaves with elongated lesions (<2.5 cm), 6 ¼ several leaves with elongated lesions (>2.5 cm), 7 ¼ long lesions common on about onehalf of the leaves, 8 ¼ long lesions common on about two-third of the leaves, and 9 ¼ most leaves with long lesions) based on the type and amount of feeding (Sharma et al., 1992). The number of plants with stemborer deadhearts was recorded at 35 days after seedling emergence (DAE) and expressed as a percentage of the total number of plants. Stalk length was measured from five randomly selected plants per replication from the base of the plant to the last node from where panicle peduncle starts, and the data were denoted as stem length (cm). From the infested plots, five randomly selected plants were cut at the base before harvest, and exit hole counts were recorded per plant after removing the sheath leaves (Sharma et al., 2007). The stems of five randomly selected plants were split open
P.G. Padmaja et al. / Crop Protection 31 (2012) 21e26
to determine the tunnel length per plant (cm) caused by C. partellus, and represented as tunneling percentage in relation to stem length. Data on leaf feeding score, deadhearts, stemtuneling, peduncle tunneling, number of tunnels and exit holes per plant were pooled for both years (2007 and 2008).
23
the resistant, IS 2205 and susceptible, DJ 6514 genotypes. The sorghum seedlings were grown under greenhouse conditions. The plants were raised on medium sized pots (60 cm diameter 30 cm deep) in the greenhouse. At 10 days after seedling emergence, four healthy seedlings were retained in each pot. Antibiosis to C. partellus in different sorghum genotypes was studied by incorporating dried leaf powder of the genotypes into the diet. For this purpose, leaves of the test genotypes were collected from 21 day old plants raised under greenhouse conditions. Two to three whorl leaves were removed with scissors at the growing point from each plant on which the larvae feed under natural conditions. The leaves were washed and then shade dried. The dried leaves were powdered in a blender for use in the artificial diet. One set of diet was prepared for each genotype and was replicated thrice in a completely randomized design. Diet was poured into 100 ml capacity plastic cups and each cup had 50 ml diet. After pouring the diet into the cups, it was allowed to cool for 2e3 h on the laboratory table. Ten first instar larvae were released into each cup, using a fine camel hairbrush. The cups were kept in the rearing room in the dark for as the neonate larvae have a strong photosensitive behavior, and settle better on the diet in darkness. In the rearing room, temperature was maintained at 27 C, 70% RH, and 12 h photoperiod. Observations were recorded on larval survival and larval weight at 10 days after releasing the larvae into the artificial diet, pupation and adult emergence. Pupal weight was recorded for each sex separately on the second day after pupation. The pupae were sexed on the basis of their relative size and genital openings (Sithanantham and Subramaniam, 1975).
2.4. Greenhouse experiments 2.4.1. Oviposition preference of C. partellus Oviposition preference was studied under controlled conditions (27 C, 70% RH, and 12 h photoperiod). Seven genotypes were grown in 21 pots (30 cm diameter and 45 cm deep) in the greenhouse. Four seeds were sown in each pot and watered immediately. Twenty day old plants, the stage at which the stemborer females lay eggs on plants under natural conditions, were placed on a table in a randomized complete block design with three replications, and covered with a nylon net (2.0 m 1.0 m 0.6 m). Twenty pairs of newly emerged adults were released inside the nylon net. The moths were provided with water in a cotton swab, and allowed to oviposit on the test genotypes for three days. The number of egg masses and number of eggs laid were recorded on each genotype. 2.4.2. Growth and development of C. partellus under greenhouse conditions Survival and development of neonate larvae of C. partellus were studied on 21-day old seedlings under greenhouse conditions. Four plants were grown in each pot. The two breeding lines with their parents and resistant and susceptible genotypes were infested with 10 neonate larvae per plant at 21 days after seedling emergence with a bazooka applicator. A randomized complete block design was used with three replications. Larval survival was recorded 10 days after infestation, as this is the time larvae successfully bore into the stem after feeding on the whorls. Moreover, at 25 days after infestation, most of the larvae will start pupating in the susceptible genotypes. At 25 days after infestation, the plants were dissected to remove the larvae/pupae, which were then placed in a plastic jar along with 15 cm pieces of sorghum stems of the same genotype. Data were recorded on pupal weights and post-embryonic development period. Pupal weights were recorded separately for the males and females one day after pupation.
2.6. Statistical analysis The data obtained from field, greenhouse and laboratory experiments were subjected to analysis of variance using the statistical software Windostat (Indostat, 2004). Genotypes were used as fixed effects and blocks and years as random effects. The data on percentages were subjected to arcsine transformation. The data on number of tunnels, peduncle tunneling percentage, exit holes, number of egg masses and number of eggs per plant and larval weights were subjected to square root transformations. LSD was used to compare the treatment means. 3. Results
2.5. Laboratory experiment 3.1. Evaluation of resistance to C. partellus under field conditions 2.5.1. Growth and development of C. partellus on artificial diet The survival and development of C. partellus was studied on artificial diet with leaf powders of 27B PB 15881-3-1 and 463B PB 15881-3, their parents (27B, 463B and PB 15881-3) and
There were significant differences among genotypes for leaf feeding injury, percentage deadhearts, number of holes and tunnels and percentage tunneling (Table 1). The breeding lines, 27B PB
Table 1 Performance of breeding lines for resistance to spotted stemborer, C. partellus, in sorghum. Genotype
Leaf feeding score (1e9)
27B 463B PB 15881-3 27B PB 15881-3 463B PB 15881-3 IS 2205 DJ 6514
8.3 7.4 5.2 5.8 5.7 5.9 8.0
Mean SEM CV F ratio P
6.6 0.22 5.9 30.71 <0.0001
0.2 0.2 0.2 0.5 0.2 0.1 0.1
Treatment means were compared using LSD ddf ¼ 6,14.
Deadhearts (%)
Stem tunneling (%)
45.3 41.4 32.8 28.9 28.9 27.8 48.3
45.0 50.8 27.8 27.2 27.8 25.3 58.9
2.5 1.2 4.3 2.6 1.6 1.4 1.9
36.4 2.08 9.9 16.6 <0.0001
1.6 6.8 1.4 1.1 1.3 2.9 1.2
37.5 3.0 13.9 20.9 <0.0001
Peduncle tunneling (%) 3.6 3.4 2.8 2.5 2.6 3.0 3.6
0.1 0.1 0.1 0.1 0.1 0.1 0.2
3.1 0.05 2.7 91.3 <0.0001
Tunnels plant1 (no.) 2.4 2.0 1.6 1.7 1.7 1.4 2.1
0.1 0.1 0.1 0.0 0.1 0.0 0.1
1.8 0.04 4.2 53.2 <0.0001
Exit holes plant1 (no.) 3.0 3.0 2.0 2.2 2.3 1.7 2.8
0.0 0.2 0.1 0.1 0.1 0.2 0.1
2.4 0.06 4.1 80.2 <0.0001
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15881-3 and 463B PB 15881-3 were as good as the resistant genotype IS 2205 and their resistant parent PB 15881-3 for stemborer resistance (leaf feeding score 5.2e5.8), whereas the leaf feeding was greater on the susceptible parents, 27B and 463B (7.4e8.3). The genotypes 27B PB 15881-3 and 463B PB 15881-3 had significantly fewer deadhearts had reduced stem and peduncle tunneling compared to their parents 27B and 463B (Table 1). 3.2. Oviposition preference The oviposition preference of C. partellus differed significantly according to genotype. Significantly more eggs masses and total number of eggs were laid on 27B, 463B and DJ 6514 than on IS 2205 and PB 15881-3. The genotypes 27B PB 15881-3-1 and 463B PB 15881-3, IS 2205 and PB 15881-3 were least preferred for oviposition. These genotypes had 131 to 146 eggs per plant compared to 320 eggs on 27B (Table 2). 3.3. Growth and development There were significant differences in larval survival, pupal weights, and post-embryonic development periods of C. partellus on different sorghum genotypes. Larval survival at ten days after infestation varied from 51 to 76% (Table 3). Larval survival was >70% on 27B, DJ 6514 and 463B, 51% on IS 2205, as compared to 54e56% on breeding lines. The weights of both male and female pupae were lower in insects reared on IS 2205, breeding lines and PB 15881-3 as compared to those reared on 27B, 463B and DJ 6514. Post-embryonic development period was >50 days for males and >60 days for females when the larvae were reared on 27B PB 15881-3 and 463 PB 15881-3, compared to 42.0 days for the males and 43.2 days for the females on 27B. Pupal weights were greatest when larvae were reared on 27B, DJ 6514 and 463B. A better consumption and utilization of the cultivars 27B and DJ 6514 was shown as expressed by the higher weights attained by pupae grown on these two than on PB 15881-3 and IS 2205. 3.4. Survival and development of C. partellus on artificial diet Larval survival and larval weight of C. partellus reared on artificial diet with leaf powder showed significant differences between the genotypes tested. Larval survival was >70% in diets with leaf powder of 27B and DJ 6514 compared to 50% survival in artificial diet amended with leaf powder of 27B PB 15881-3 and 463B PB 15881-3 (Table 4). The larval weight was 5.1e5.6 mg on artificial diets with 27B PB 15881-3, 463B PB 15881-3 and PB 15881-3 Table 2 Oviposition preference by the spotted stemborer, C. partellus, toward sorghum genotypes under multi-choice cage conditions. Genotype
No. of egg masses per plant
27B 463B PB 15881-3 27B PB 15881-3 463B PB 15881-3 IS 2205 DJ 6514
6.0 5.0 3.5 2.0 2.3 3.0 5.0
Mean SEM CV F ratio P
3.83 0.25 11.33 37.04 <0.0001
0.3 0.5 0.3 0.2 0.2 0.3 0.4
Treatment means were compared using LSD ddf ¼ 6,14.
No. of eggs per plant 320 285 128 131 146 125 301
12.7 21.4 6.9 9.1 15.0 4.0 13.8
205.67 14.03 11.82 41.97 <0.0001
leaf powder compared to 7.4e7.9 mg on 463B, DJ 6514 and 27B. Low larval survival and larval weight at 10 DAI were recorded on IS 2205, PB 15881-3, 27B PB 15881-3 and 463B PB 15881-3. Pupal weight varied from 51.9 to 71.6 mg for the males and from 96.4 to 142.5 mg for the females. Significantly lower pupal weight was observed in male and female pupae from the larvae reared on diets with leaf powder of 27B PB 15881-3 and 463B PB 15881-3 as compared to those insects reared on their parents. Percentage pupation varied from 41.5 to 62.3%, while the adult emergence varied from 30.9% to 44.3%. Comparatively lower pupation (45e48%) and adult emergence (32e33%) were recorded in artificial diets impregnated with leaf powder of 27B PB 15881-3 and 463B PB 15881-3 as compared to those reared on diets with leaf powder of 27B, 463B and DJ 6514 (52e62% pupation and 37e44% adult emergence). The larval period varied from 24.6 to 37.1 days for males, and from 25.1 to 39.4 days for the females on artificial diet impregnated with leaf powder from different sorghum genotypes (Table 4). Larval duration for the males was prolonged by 12 days on 27B PB 15881-3 and 463B PB 15881-3 as compared to 27B. Duration of larval period for the females was significantly longer on 27B PB 15881-3 and 463B PB 15881-3 as compared to the larvae reared on 27B. Pupal period for the males was 8.0 days on 27B PB 15881-3 and 8.8 days on 463B PB 15881-3, compared to 7.1 days on 27B. 4. Discussion Selections for resistance to C. partellus based on leaf injury, deadhearts and stem tunneling were made in the F3eF6 generations of two sorghum crosses. The plants selected in each generation were then subject to further screening. Both the breeding lines were significantly more resistant than the respective susceptible parents and DJ 6514 based on leaf injury, deadhearts and stem tunneling. Both the lines were found to be close to their resistant parent PB 15881-3 with regard to these parameters. Thus, it was considered that continuous screening for leaf injury, deadhearts and stem tunneling over several generations would prove to be more effective in improving the level of resistance. Stem tunneling had earlier been reported to be the major plant character associated with resistance to stemborer (Kishore, 1991). Stem tunneling rather than leaf feeding and deadhearts is the primary cause of yield loss (Alghali, 1986). However, these are not the only damage parameters responsible for yield reduction in sorghum (Pathak and Olela, 1983; Singh et al., 1983; Taneja and Leuschner, 1985). The number of exit holes per plant is another criterion of resistance, which indicates suitability of the genotype for the larva to complete development and successfully emerge from the stalk for pupation. Significant differences in oviposition were observed among the sorghum genotypes, with the breeding lines being relatively less preferred for oviposition. Oviposition preference has earlier been identified to be one of the components of resistance to C. partellus in sorghum (Singh and Rana, 1984; Alghali, 1985; Saxena, 1990; van den Berg and van der Westhuizen, 1997; Kishore Kumar et al., 2007). Most of these studies were aimed at identifying sorghum lines with resistance to stemborer under field or greenhouse conditions. Comparison of the biology of C. partellus when reared on different lines showed that rearing on the 27B PB 15881-3 and 463B PB 15881-3 resulted in lower larval and pupal weights, a longer pupal period, fewer larvae completing development, and smaller, fewer females. Antibiosis in terms of reduced larval survival and prolongation of post-embryonic development period of C. partellus has earlier been reported by Jotwani et al. (1978), Woodhead and Taneja (1987), van den Berg and van der Westhuizen (1997) and Kishore Kumar et al. (2007). Adverse
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25
Table 3 Development of neonate larvae of spotted stemborer, C. partellus, on seedlings of sorghum genotypes under greenhouse conditions. Genotype
Larval survival (%) at 10 DAI
Pupal weight (mg) Males
27B 463B PB 15881-3 27B x PB 15881-3 463B x PB 15881-3 IS 2205 DJ 6514
76.0 71.6 53.8 54.3 56.2 51.4 73.0
4.5 0.5 1.7 1.4 0.7 1.4 3.0
Mean SEM CV F ratio P
62.3 1.62 4.5 42.9 <0.0001
47.4 44.6 39.2 32.2 34.3 36.7 46.2
Post-embryonic development period (days) Females
2.2 2.5 4.7 0.7 0.2 3.1 2.4
96.4 87.4 71.2 71.2 75.3 70.6 92.8
40.09 2.76 11.90 4.80 <0.0001
3.6 1.6 4.0 1.6 2.6 0.9 1.2
80.7 1.89 4.07 34.75 0.0101
Males 42.0 44.0 50.1 55.7 52.6 53.8 44.3
Females 0.6 1.2 1.3 1.4 1.8 2.4 2.4
43.2 48.5 59.9 61.8 60.4 59.1 49.3
48.92 1.79 6.33 9.30 <0.0001
1.4 3.5 4.3 2.1 1.9 4.4 0.4
54.61 1.74 5.53 18.02 0.0006
DAI ¼ Days after infestation. Treatment means were compared using LSD ddf ¼ 6,14.
effects of stemborer-resistant lines on survival and development of C. partellus under field conditions have been reported by Lal and Pant (1980), Singh and Verma (1988), and Woodhead and Taneja (1987). Genotypic differences in larval establishment in the field have been reported on different sorghum genotypes (Jotwani et al., 1978; Woodhead and Taneja, 1987; Singh and Rana, 1989; van den Berg and van der Westhuizen, 1997). However, expression of resistance to C. partellus under field conditions can be quite variable due to variation in environmental conditions, differential growth of the test genotypes, and the nutrient status of the soil. Dried leaf powder amended into the artificial diet is helpful to overcome the variation in borer infestation observed under field conditions to allow a comparison of the test genotypes under uniform conditions. There was considerable variation in larval survival, larval and pupal weight, larval and pupal development period, and percentage pupation and adult emergence in diets amended with leaf powder of different sorghum genotypes. Low larval survival, larval and pupal weights, low pupation and adult emergence were recorded in artificial diets amended with dried leaf powder of 27B PB 15881-3, 463B PB 15881-3, PB 15881-3 and IS 2205. Larval, pupal, and the total development period is also prolonged (Jotwani et al., 1978; Lal and Sukhani, 1979, 1982; Singh and Rana, 1984, 1989; Saxena, 1990, 1992; Verma et al., 1992). Antibiosis can also be expressed in terms of reduced pupal weight (Lal and Sukhani, 1982; Singh and Rana, 1984; Singh and Verma, 1988; Verma et al., 1992; Kishore Kumar et al., 2006) and low pupation and adult emergence (Singh and Verma, 1988). The antibiotic effects of the resistant genotypes on the development of
C. partellus may be caused by secondary plant substances in the leaves and/or poor nutritional quality of the food. Reduced survival and establishment will reduce the insect population and the resultant crop damage. Prolongation of development period will also result in reduction of number of generations in a season/year. These effects of the resistant varieties will have a constant and cumulative effect on the stemborer population over the season (Sharma, 1993), and can be utilized as an environmental friendly method to reduce the damage by this pest under subsistence farming conditions. Results from this study indicated significant differences between resistant and susceptible genotypes with regard to reduction in foliar damage ratings, percentage stem length tunneled, and percentage of plants showing deadhearts, oviposition preference, larval survival, larval and pupal mass and percentage pupation and adult emergence. It has been concluded that slow development of larvae on resistant lines is due to antibiosis. The possibility of selection of parents to breed should therefore be based on antixenosis and antibiosis to increase the levels of resistance to C. partellus in sorghum. Therefore, although a complex interaction of different factors affect resistance/susceptibility of sorghum to stemborer attack, differences in level of preference and performance among resistant and susceptible genotypes strongly support antixenosis and antibiosis as major mechanisms of resistance to C. partellus. The study clearly indicates the genetic transmission of antixenosis and antibiosis resistance mechanisms from resistant parent to the progeny. It also shows that by utilizing the resistant lines in the crossing program and consciously selecting resistant
Table 4 Growth and development of spotted stemborer, C. partellus, larvae on semi-synthetic diet amended with leaf powder of different sorghum genotypes. Genotypes
Larval survival (%) 10 DAI
27B 463B PB 15881-3 27B PB 15881-3 463B PB 15881-3 IS 2205 DJ 6514
72.8 71.0 52.2 50.8 51.0 51.5 71.6
Mean SEM CV F ratio P
60.1 3.32 9.6 10.9 0.0003
2.4 7.6 1.7 0.7 0.2 1.4 1.1
Larval weight (mg) 10 DAI 7.9 7.4 5.0 5.1 5.6 4.6 7.6
0.1 0.2 0.5 0.3 0.2 0.4 0.3
6.17 0.33 9.15 18.57 <0.0001
Larval development time
Pupal weight (mg)
Male
Males
24.6 25.1 36.4 37.0 35.1 37.1 27.3
Female
31.77 2.57 14.01 5.21 0.0074
1.8 0.5 2.4 2.8 2.9 4.0 1.0
25.2 25.9 37.9 39.0 36.4 39.4 25.1
2.9 2.6 4.5 4.6 1.0 0.4 2.9
32.7 2.22 11.76 9.65 0.0005
DAI Days after infestation. Treatment means were compared using LSD ddf ¼ 6,14.
71.6 62.1 53.5 52.7 58.4 51.9 68.1
59.76 3.49 10.10 5.05 0.0084
Females 4.9 1.8 2.0 1.6 4.8 3.9 1.2
Pupation (%)
142.5 1.4 127.4 7.4 98.6 1.8 104.1 1.8 108.7 5.0 96.4 3.6 131.4 1.8
62.3 52.3 44.1 45.0 48.2 41.5 57.8
115.59 3.99 5.97 20.39 <0.0001
50.3 0.72 2.5 113.7 <0.0001
3.6 2.0 2.4 2.7 2.4 2.9 1.2
Pupal development time
Adult emergence (%)
Males
Females
7.1 0.1 7.3 0.2 9.5 0.2 8.0 0.1 8.8 0.2 10.7 0.7 7.8 0.2
7.0 0.0 7.4 0.2 9.2 0.1 8.1 0.1 8.9 0.5 10.4 0.2 8.2 0.2
44.3 37.1 32.0 32.2 33.1 30.9 42.1
8.46 0.23 4.78 31.15 <0.0001
8.45 0.25 5.12 21.15 <0.0001
36.0 1.74 8.4 9.5 0.0005
1.4 2.1 1.0 2.4 2.1 1.1 1.4
26
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