Materials and methods Plant and beetle collection and census. Gender sorting of the adult beetles was accomplished by finding copulating pairs in embrace for at least 1 minute and assuming the male to be mounted on the dorsal side of the female. Verification of this technique was confirmed by aedeagus dissection. Daily census of beetles on host milkweed populations in western Nevada took place around noon on 14 randomly chosen patches of A. speciosa in Douglas county, Nevada since 1992. Beetles, herbivory patterns, and egg cases ovideposited, along with total host milkweed plants, stems, and leaves, were counted and assessed during each visitation at each patch site.
Chemical analyses of whole beetles and egg cases.
The whole beetles and egg cases stored at -20 C were thawed and dried in an oven at 40 C for 48 hours and then placed in 2 ml spectral grade methanol for 24 hrs to extract cardenolide. The total cardenolide content was determined on three or more beetles or samples of eggs using the methods of TNDP spectroassay reported in Isman et al. Egg cases analyzed included the fecal covering and the eggs within, usually between 8 to 10 eggs per clutch. Approximately 12 live female adult C. colbaltinus beetles collected in summer of 1995 on a host of A. speciosa in Douglas county, Nevada were placed in an empty 250 ml amber headspace chamber at 20 C without food for 1 week. GC-MS analysis of the headspace atmosphere of a live male beetle confined to a 20 ml headspace vial showed the same two GC peaks for the two fatty acids given above for female beetles.