Evidence That a Laminin-Like Insect Protein Mediates Early Events in the Interaction of a Phytoparasite with Its Vector's Salivary Gland

Abstract

Phytomonas species are plant parasites of the family Trypanosomatidae, which are transmitted by phytophagous insects. Some Phytomonas species cause major agricultural damages. The hemipteran Oncopeltus fasciatus is natural and experimental host for several species of trypanosomatids, including Phytomonas spp. The invasion of the insect vectors' salivary glands is one of the most important events for the life cycle of Phytomonas species. In the present study, we show the binding of Phytomonas serpens at the external face of O. fasciatus salivary glands by means of scanning electron microscopy and the in vitro interaction of living parasites with total proteins from the salivary glands in ligand blotting assays. This binding occurs primarily through an interaction with a 130 kDa salivary gland protein. The mass spectrometry of the trypsin-digest of this protein matched 23% of human laminin-5 β3 chain precursor sequence by 16 digested peptides. A protein sequence search through the transcriptome of O. fasciatus embryo showed a partial sequence with 51% similarity to human laminin β3 subunit. Anti-human laminin-5 β3 chain polyclonal antibodies recognized the 130 kDa protein by immunoblotting. The association of parasites with the salivary glands was strongly inhibited by human laminin-5, by the purified 130 kDa insect protein, and by polyclonal antibodies raised against the human laminin-5 β3 chain. This is the first report demonstrating that a laminin-like molecule from the salivary gland of O. fasciatus acts as a receptor for Phytomonas binding. The results presented in this investigation are important findings that will support further studies that aim at developing new approaches to prevent the transmission of Phytomonas species from insects to plants and vice-versa.

The Immune Response of Hemocytes of the Insect Oncopeltus fasciatus against the Flagellate Phytomonas serpens

“…In the present study, using a model of systemic infection, we have described the interaction between P. serpens and O. fasciatus hemocytes. In agreement with previous observations of both the natural infection of the vector Phthia picta and the experimental infection of O. fasciatus with P. serpens , the parasite survives the invasion of the hemocoel and reaches the salivary gland [4], [25], a condition common to the life cycle of parasites that develop in the hemolymph within their respective vector, such as Plasmodium species and T. rangeli [26], [27]. This mode of infection is widely used for studying the interaction of trypanosomatids that invade the hemocoel of their insect hosts, especially in the model of interaction between T. rangeli with R. prolixus [28], [29].…”

“…These insects founded the colony we maintain in our laboratory in plastic pitchers under a 12 h light/dark cycle at 28°C with 70–80% relative humidity [25].…”

Transovum Transmission of Trypanosomatid Cysts in the Milkweed Bug, Oncopeltus fasciatus

“…A colony of O. fasciatus naturally infected with L. wallacei [7] was established and maintained in our laboratory under a 12 h light/dark cycle at 28°C with 70–80% relative humidity, as previously described [6] , [18] .…”

Natural infection by the protozoan Leptomonas wallacei impacts the morphology, physiology, reproduction, and lifespan of the insect Oncopeltus fasciatus

“…Conversely, no data is available for long-term infections 16 . The transmission of L. wallacei between insects occurs via the ingestion of cystic forms through coprophagy, as well as by transovum transference of cysts that adhere to the eggs during oviposition; i.e., the newly born nymphs are infected by probing the fecal droplets on eggshells 36 . It was common for the O. fasciatus individuals that we used in our laboratory to be naturally infected by L. wallacei 16 .…”