ecology of phasmids - KLUEDO - Universität Kaiserslautern

Adult female feeding preference & nymph performance 53
distilled water. The phenolic content of the resulting solution was assessed by the Price and Butler
method (1977) as described above. In preliminary infiltration trials I assessed the amount of water that
could be infiltrated into leaves (27.4 ± 4.3 % fresh weight). In combination with dry weight and natural
total phenol contents of P. hispidum leaves I could then calculate the concentration of solution needed to
yield particular increases of % TAE in leaf discs.
Infiltration was carried out with a 50 ml syringe filled with approximately 25 ml of aqueous phenol or
tannin solution (or water for the control). Immediately after punching the disc was weighed and put in
the syringe. All remaining air was removed through the outlet. Then the outlet was closed and the piston
was forcefully pulled outward the depression leading to an evacuation of the internal air and the
intercellular space. Simultaneously the syringe was shaken to remove gas bubbles from the leaf surface
and prevent them from entering the leaf disk during the following infiltration process. By slowly
pushing the piston back into the syringe the solution (or water) column was set under pressure and
entered the leaf via open stomatal pores and via intercellular space at the cutting edges. After
infiltritation the leaf disc surface was dried carefully with a peace of paper towel and weighed again.
Weight gain was considered as infiltrated amount of solution (water). In combination with natural total
phenol contents of leaves and their dry weight infiltrated percent TAE in the leaf disc was calculated.
Until the start of the feeding trials leaf discs were kept on ice.
First instar nymphs of M. diocles were presented with two leaf discs: 1) infiltrated with water (control)
and 2) infiltrated with the according solution (treatment).
Nymphs were set in plastic food containers (10 cm wide, 5 cm deep, 5 cm high). Leaf discs were stuck
into a piece of cardboard to assure free access for the nymph. A piece of humid paper towel helped
minimizing desiccation. Climate chambers allowed for conditions similar to the tropics (27 o C average
temperature, 70 to 90 % relative humidity). Dual-choice feeding trials lasted 24 hours. This was a
sufficient time span to assess preference because a preliminary test on dual-choice feeding trials over
three days showed no differences in preference among days (Friedmans ANOVA χ 2 (N=38, FG=2) = 1.28,
P < 0.53). After finishing the trial consumed leaf area was measured with a transparent grid. Preference
was then calculated on the base of consumed dry weight (see Chapter 4.3.2).
To assure that infiltration did not alter feeding behavior I tested normal leaf discs against water
infiltrated leaf discs. Both, control and treatment were fed equally resulting in a relative preference of
0.5 indicating that infiltration did not affect feeding behavior (compare to control in Figure 4-12 &
Figure 4-13).
4.3 Data analysis
Multiple groups where compared in Analysis of Variance. ANOVA assumes normality of data and
homogeneity of variances. If not stated differently data transformation resulted in normally distributed
data and allowed minimizing differences in variances whereas homogeneity of variances could not be