A no-probe experiment Obeticholic Acid in vivo and the hybridization of an aposymbiotic ovariole was executed as a specifity control. Fitness effects To investigate the effect

of the endosymbionts on the fitness of M. pygmaeus, nymphal development and fecundity of the predator were compared between the infected laboratory-strain of M. pygmaeus and an endosymbiont-free M. pygmaeus population. The general procedure largely follows the method of Vandekerkhove et al. [48], with slight modifications. First instars (<24h) of the 39th generation of each population were individually caged in vented plastic cups (4 cm diameter and 2.5 cm high) containing a wax paper drenched in paraffin. A parafilm dome filled with water and E. kuehniella eggs were provided as a source of water and food, respectively. Water domes and eggs were replaced every two days. Nymphs which died on the first or second day of the experiment were replaced by new ones, assuming that their death was caused by handling. Nymphal development and survival were checked daily. Nymphs that successfully reached the

adult stage were sexed and weighed at emergence (i.e., within 24 h after moulting). Adult pairs were then transferred to a new plastic cup containing a tobacco leaf disc placed with the upper side on a 1 % agar layer. Two crosses were tested: infected males with infected females [I♂ x I♀] and uninfected males with uninfected females [U♂ x U♀]. Eggs of E. kuehniella were offered as a food source for the adult predators, whereas the tobacco leaf served as a source of RO4929097 supplier moisture and an oviposition substrate. After

7 days, females were dissected and oocytes were counted [28]: late vitellogenic to mature oocytes were scored 1; early to mid vitellogenic oocytes 0.5 and previtellogenic oocytes 0.25. Mature oocytes present in the oviducts were also scored as 1. The scores for all ovarioles were then summed providing a weighted sum of oocytes, which can reliably be used to predict the lifetime fecundity of M. pygmaeus [28]. Furthermore, the leaf discs were immersed in safranin and screened for oviposited eggs. Effects of infection status on nymphal development, adult weight and fecundity were 3-mercaptopyruvate sulfurtransferase statistically examined by a one-way analysis of variance (ANOVA) or a Mann-Whitney U Test using SPSS 17.0 [49]. Results Insect species collection and identification The Macrolophus populations from Greece and Italy were collected on the wild plants Solanum nigrum and Dittrichia viscosa which are considered to be conservation host plants for M. pygmaeus and M. caliginosus, respectively [50, 51]. Some M. pygmaeus populations were also collected on D. viscosa, although their survival is reported to be poor on this plant [50]. In Spain, M. pygmaeus was also collected on tomato, Solanum lycopersicum. The primer pairs CB1-CB2 and LAU1f-CB2, which both amplify a part of the cytochrome b gene, were used to elucidate the species identity of the collected insects. In accordance with Martinez-Cascales et al.