First report on honeydew excretion by the melon thrips, Thrips palmi karny (Thysanoptera : Thripidae) and its biochemical analysis

Sap sucking insects such as thrips, aphids, whitefly, mealybugs, leafhoppers, psyllids etc. feed primarily on the phloem sap which is rich in sugars such as sucrose, fructose, trehalose, maltose, raffinos, meteoritose etc.; free amino acids such as asparagine, glutamine, glutamate and serine (Hijaz and Killiny, 2014). Feeding of sugar rich sap leads to differential osmolarity between the hemolymph and gut lumen. To maintain the osmolarity between the gut lumen and the haemolymph, phloem feeders have developed several adaptations such as filter-chamber for efficient water usage and to excrete excess sugars in the form of honeydew through different orifices such as cornicles in aphids and anus in many other sap sucking insects. Honeydew acts as a medium through which insecticides are excreted and thereby contribute to the development of resistance to insecticides. This excretion of copious amount of honeydew on crops serves as substrate for the development of many saprophytic fungi like Capnodium sp., which affects photosynthesis (Lin, 2006; Wallace, 2008; Neto, 2011). It is also reported to be involved in natural enemy calling, which is self-inimical and provide food for ants which ensures dispersal and protection from the predators (Leroy et al., 2011). It has also been documented that honeydew is also a source of food for parasitoids involved in biological control. Rate of honeydew excretion and its composition by aphids and whiteflies is well studied, but the information on honeydew excretion by thrips is not studied in detail. Hence, a study was conducted to understand the pattern of honeydew excretion by the melon thrips, T. palmi due to its significance as a polyphagous pest and an important vector of Watermelon bud necrosis virus and Groundnut bud necrosis virus in India.

Stock culture of T. palmi was maintained on French bean (Phaseolus vulgaris CV. Arka Komal) pods at a temperature of 25±2°C and 68±% RH (Rebijith et al., 2011). One hundred adults of T. palmi were released on fresh French bean pods of approximately equal size (3cm surface width; 13 cm length and 6 g in weight) which were harvested from the French bean plants grown under insect proof cages. The samples with thrips were placed inside a plastic container (10cm x10cm) and kept at the room temperature (25±2°C, 68±% RH). There were three replicates and same replicates of control (without inoculating T. palmi) were also maintained. Observations were made continually on the behavior of T. palmi adults and for the excretion of honeydew under the stereo-zoom microscope Stemi 305 (ZEISS, Germany).

Sugars were separated by following modified Steppuhn and Wackers (2004) method. After 24 h observation, the bean pods were washed with 10 mL of 80% ethanol, and the extract was evaporated and re-dissolved in mobile phase containing solvent A and solvent B in 1:1 ratio, filtered and injected to LC–MS/MS for sugar profiling.

Sugar standards viz. fructose, sucrose, galactose, glucose, maltose, fucose, rhamnose, xylose, arabinose, mannose, sorbitol, inositol, lactose, ribose and trehalose were purchased from Sigma Chemical Co., USA and calibration curve was prepared using different concentration of individual sugars. The mobile phase used was composed of solvent (A) 80:20 (Acetonitrile: Water) and solvent (B) 30:70 Acetonitrile: water with 0.1% ammonium hydroxide was filtrated through 0.2 µm nylon filter paper and separation was done using gradient elution. The initial gradient was composed of 100% solvent A for one min and at 8th min it was champed to 88% of solution A and 12% of solution B, which was held for 1 mint and a linear gradient was followed by 98% of solution A and 2% of solution B and at 15th mins it was held for 30 sec. The system had returned to initial settings at 19th min and equilibrated for 6 min. before the next injection and the flow rate was 0.1mL/min. The analytical column used was 2.1x10 mm UPLC BEH-Amide (Waters, USA) with 1.7 μm particle size and protected by vanguard BEH-Amide with particle size 1.7 μm. The column was maintained temperature of 25°C.

Study was conducted to understand the pattern of honeydew excretion by the melon thrips. Close observation under the stereo microscope revealed that T. palmi adults excrete honeydew which lasted for about 10 sec from initiation bending of abdomen to the release. These events were recorded in VLC format. Analysis of sugars in the honeydew revealed that there was a significant difference between the control samples and samples inoculated with thrips. Among the sugars estimated, inositol was the predominant sugar (130.95 ±0.47 µg/pod) in the honeydew followed by fructose (95.13 ±0.45 µg/pod); maltose (60.700 ±0.28µg/pod); glucose (54.22 ±0.40µg/pod); sorbitol (28.15 ±0.35g/pod) (Fig: 2) followed by less of lactose, mannose, galactose, arabinose, ribose and fucose (Table 1). It clearly indicated that the honeydew excreted by thrips is rich in soluble sugars. There is a single report on honeydew excretion by the red banded thrips, Selenothrips rubrocinctus (Giard) (Buss et. al., 2006). Wool et al. (2006) reported about 20 sugars in the honeydew excreted by aphids. Glucose and fructose are basic components of the honeydew of sap feeding insects (Fischer et al., 2005; Wool et al., 2006). These sugars are present in honeydew in different proportions depending on the insect species and host plants. Hendrix et al. (1992) also observed the differences in sugar composition of honeydew excreted by Trialeurodes vaporariorum (Westwood) and Bemisia tabaci (Gennadius) feeding on different host plants.

Fig. 1
Contents of major water soluble sugars in beans with or without thrips infestation

Table 1
Biochemical analyses of water-soluble sugars by LC-MS-MS in the bean pods with or without thrips infestation

Differences in chemical composition of honeydew secreted by aphids are explained inter alia by genetic variation between insect populations (Fischer and Shingleton, 2001). However, the host plant sap is a primary factor that influence the diversity in biochemical composition of honeydew. Honeydew composition is an important factor in tritropic interaction involving natural enemies and also mediating ant–homopteran mutualisms. However, further studies on sugar composition in relation to species of ants attracted and it s impact on predation/parasitism is required in order to have sustainable management of this pest.

The authorsare thankful to the Director, ICAR-IIHR, Bengaluru for support and infrastructure facilities