CHAPTER 5: Evaluation of resistance of sugarcane varieties to Chilo partellus
5.2 Materials and methods
5.2.2 Pot trials used to evaluate damage on sugarcane by Chilo
This experiment took place in a glasshouse at the South African Sugar Research Institute (SASRI, KwaZulu-Natal, South Africa). Stalks from the selected sugarcane varieties were collected from established field trials at SASRI and used for planting.
Stalks were cut into single budded setts using secateurs, and then hot water treated in a water bath (Lasec, Durban, South Africa) at 50oC for 30 minutes. Setts were
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planted into 25 Litre PVC pots (30 cm diameter and 45 cm deep) (Grovida, Durban, South Africa) containing cleaned, coarse Umgeni river sand (Chain sands, Durban, South Africa). Pots were placed into troughs (Wardkiss, Durban, South Africa) filled with water to ensure that they would not dry out and that ants could not predate inoculated larvae. The experiment was laid out in a completely randomized design, with a total of five replications (Appendix 5.1). Each replication consisted of 20 pots representing all 20 sugarcane varieties (Figure 5.1b). Pots were arranged in double rows of 10 pots each for each replication, and spaced so that they did not come into contact with each other, to avoid contact between neighbouring plants. Planting was done over three weeks, with one sett being planted in each pot per week, until a total of three setts of the same variety were planted in each pot (Figure 5.1a). Planting was done in this manner for inoculation purposes. Pots were labelled with the variety name using pot labels (Wardkiss, Durban, South Africa). They were watered
manually on a daily basis until inoculation of larvae took place, after which troughs were maintained to always be filled with water. Plants were fertilized monthly with 4:1:1 (44) N:P:K fertilizer (16 g/pot) (Grovida, Durban, South Africa).
When each plant reached 21 days of age, inoculations were performed. This is the stage when female stem borer moths lay eggs on plants under natural conditions (Kumar et al., 2007). Neonate larvae of C. partellus were provided by the established colony at SASRI. Inoculations took place over a period of three weeks, with the first- planted plant per pot being inoculated in the first week, the second in the second week, and the third plant in the third week until all the plants in the pot trial had been inoculated (Figure 5.1a). Each plant was inoculated with 10 neonate C. partellus (Nibouche and Tibere, 2010) larvae, and they were released directly into the central whorl of each plant with a camel hair brush (Winsor and Newton, London, UK).
According to Sharma (1997), five to seven larvae usually causes a sufficient amount of damage to susceptible sugarcane varieties. Inoculated plants were labelled with the date using white labels (Wardkiss, Durban, South Africa). Inoculations took place in the early morning between 07h00 and 11h00 to avoid larval death due to high temperatures.
Thirty days after inoculating, damage assessments were made. Plants were cut at their base and used for analysis. The number of entry/exit holes (borings) due to larvae was counted (Figure 5.2a). Stalks were split open throughout their entire
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length and the total length of tunnels in each stem was recorded (Figure 5.2b). Upon dissection of the plants, the number of larvae and pupae found were counted. Live larvae that were found on the outside of the plant were considered established.
Larval weight of recovered larva was also recorded. The number of damaged leaves and the number of shotholes or lesions per leaf were counted and recorded (Figure 5.2c). Two different leaf feeding damage rating scales were used. One of the rating scales was on a scale of 0 to 4 (Figure 5.3), where 0 = no visible leaf injury, 1 = few shotholes or lesions observed on leaves, 2 = medium damage, 3 = heavy damage, and 4 = extensive and severe damage to leaves. The second rating scale was based on a method used by Conlong et al. (2004), which uses the number of damaged leaves and the number of feeding holes as part of a rating system on a scale of 1 to 9. A stalk damage rating system was also used, based on the same scale as that of the leaf feeding damage rating on a scale of 0 to 4 (Figure 5.4). An overall rating was given to each variety, based on the total number of borings, total number of larvae recovered and mean number of shotholes/lesions over the two pot trials that were carried outThis was a subjective rating whereby R = resistant, IR = intermediate- resistant, I = intermediate, IS = intermediate-susceptible, and S = susceptible After the first pot trial was complete, a second pot trial was conducted in the exact same way. However, the Variety R574 was replaced with Variety NCo376 as there was no R574 planting material available from the field. The glasshouse was fogged using Doom Fogger (Checkers Hyper, Durban, South Africa) to kill off any predators or moths that may have an effect on the trial.
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a.
Figure 5.1 Layout of (a) individual pots and (b) a replication from the pot trial used for Chilo partellus resistance studies
Figure 5.2 Damage parameters used to assess Chilo partellus damage to sugarcane varieties (a) entry/exit holes (borings) (b) tunnel length and (c) Shotholes or lesions
b.
Sett 1 Sett 3 Sett 2
Planted at week 1 1st inoculated Planted at week 3 3rd inoculated Planted at week 2 2nd inoculated
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Figure 5.3 Examples of damage caused to sugarcane leaves by Chilo partellus used as a visual leaf rating system on a scale of 0 to 4
Figure 5.4 Examples of damage caused to sugarcane stalks by Chilo partellus used as a visual stalk rating system on a scale of 0 to 4
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