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