The Method of Eliminating the Wolbachia Endosymbiont Genomes from Insect Samples Prior to a Long-Read Sequencing.

When extracting DNA of invertebrates for long-read sequencing, not only enough quantity and size of the DNA but, depending on the species, elimination of contamination of endosymbiotic Wolbachia genome also has to be achieved. These requirements become troublesome, especially in small-sized species with a limited number of individuals available for the experiment. In this chapter, using tiny parasitoid wasps (Reclinervellus nielseni) parasitizing spiders as hosts, we developed a method of eliminating the Wolbachia genomes by means of an antibiotic administration to adult wasps via honey solution. Twenty days of rifampicin treatment since their emergence from cocoons resulted in a significant decrease in the Wolbachia genomes while keeping good DNA conditions for nanopore sequencing. An adequate quantity of DNA was then gained by pooling several individuals. The method could be applied to other insects or invertebrates that can be maintained by laboratory feeding with liquid food.

The changing use of the ovipositor in host shifts by ichneumonid ectoparasitoids of spiders (Hymenoptera, Ichneumonidae, Pimplinae).

Accurate egg placement into or onto a living host is an essential ability for many parasitoids, and changes in associated phenotypes, such as ovipositor morphology and behaviour, correlate with significant host shifts. Here, we report that in the ichneumonid group of koinobiont spider-ectoparasitoids ("polysphinctines"), several putatively ancestral taxa (clade I here), parasitic on ground-dwelling RTA-spiders (a group characterised by retrolateral tibial apophysis on male palpal tibiae), lay their eggs in a specific way. They tightly bend their metasoma above the spider's cephalothorax, touching the carapace with the dorsal side of the ovipositor apically ("dorsal-press"). The egg slips out from the middle part of the ventral side of the ovipositor and moves towards its apex with the parted lower valves acting as rails. Deposition occurs as the parasitoid draws the ovipositor backwards from under the egg. Oviposition upon the tough carapace of the cephalothorax, presumably less palatable than the abdomen, is conserved in these taxa, and presumed adaptive through avoiding physical damage to the developing parasitoid. This specific way of oviposition is reversed in the putatively derived clade of polysphinctines (clade II here) parasitic on Araneoidea spiders with aerial webs, which is already known. They bend their metasoma along the spider's abdomen, grasping the abdomen with their fore/mid legs, pressing the ventral tip of the metasoma and the lower valves of the ovipositor against the abdomen ("ventral-press"). The egg is expelled through an expansion of the lower valves, which is developed only in this clade and evident in most species, onto the softer and presumably more nutritious abdomen.

Host manipulation by an ichneumonid spider ectoparasitoid that takes advantage of preprogrammed web-building behaviour for its cocoon protection.

Host manipulation by parasites and parasitoids is a fascinating phenomenon within evolutionary ecology, representing an example of extended phenotypes. To elucidate the mechanism of host manipulation, revealing the origin and function of the invoked actions is essential. Our study focused on the ichneumonid spider ectoparasitoid Reclinervellus nielseni, which turns its host spider (Cyclosa argenteoalba) into a drugged navvy, to modify the web structure into a more persistent cocoon web so that the wasp can pupate safely on this web after the spider's death. We focused on whether the cocoon web originated from the resting web that an unparasitized spider builds before moulting, by comparing web structures, building behaviour and silk spectral/tensile properties. We found that both resting and cocoon webs have reduced numbers of radii decorated by numerous fibrous threads and specific decorating behaviour was identical, suggesting that the cocoon web in this system has roots in the innate resting web and ecdysteroid-related components may be responsible for the manipulation. We also show that these decorations reflect UV light, possibly to prevent damage by flying web-destroyers such as birds or large insects. Furthermore, the tensile test revealed that the spider is induced to repeat certain behavioural steps in addition to resting web construction so that many more threads are laid down for web reinforcement.

Infanticide by a solitary koinobiont ichneumonid ectoparasitoid of spiders.

When encountering an already parasitized host, a parasitoid's optimal choices (superparasitism, host rejection, host feeding or infanticide) seem to depend on the individual species' life history, because the same choice may have different fitness consequences. We demonstrate infanticide under laboratory conditions by a polysphinctine, Zatypota albicoxa, which is a solitary koinobiont ectoparasitoid of spiders. The female always removed any previously attached egg or larva from the body of the host spider, Parasteatoda tepidariorum, with a rubbing behaviour. She rubbed her ovipositor back and forth toward the undersurface of the attached egg or of the saddle under the attached larva to pry it off and laid an egg after removal. When removing a larva, the infanticidal female engaged exclusively in unfastening the 'saddle' which fastens the larva to the body of the spider. All larvae were removed with the 'saddle' attached to the ventral surface of the body. The female invested more time to remove the medium second and the large penultimate instar larvae than to remove eggs and first instar larvae because of the labour involved in unfastening the saddle. Oviposition with infanticide of the medium second and the penultimate instar larvae imposed more time upon the female than that on an unparasitized host. Removal of any previous occupant in spite of the associated labour costs suggests that infanticide will always be adaptive, no matter the time costs to Z. albicoxa, because so much is invested in attacking the host and because the parasitoid cannot detect whether the spider is already parasitized until she achieves subjugation.