The leafhopper Scaphoideus titanus is able to transmit 16SrV phytoplasmas agents of grapevine's flavescence dorée (FD) within 30–45 days, following an acquisition access period (AAP) of a few days ...feeding on infected plants as a nymph, a latency period (LP) of 3–5 weeks becoming meanwhile an adult, and an inoculation access period (IAP) of a few days on healthy plants. However, several aspects of FD epidemiology suggest how the whole transmission process may take less time, and may start directly with adults of the insect vector. Transmission experiments have been set up under lab condition. Phytoplasma‐free S. titanus adults were placed on broad bean (BB) plants (Vicia faba) infected by FD‐C (16SrV‐C) phytoplasmas for an AAP = 7 days. Afterwards, they were immediately moved onto healthy BB for IAP, which were changed every 7 days, obtaining three timings of inoculation: IAP 1, IAP 2 and IAP 3, lasting 7, 14 and 21 days from the end of AAP, respectively. DNA was extracted from plants and insects, and PCR tests were performed to identify FD phytoplasmas. Insects were dissected and fluorescence in situ hybridisation was made to detect the presence of phytoplasmas in midguts and salivary glands. The rate of infection in insects ranged 46–68% without significant differences among IAPs. Inoculation in plants succeeded in all IAPs, at a rate of 16–23% (no significant differences). Phytoplasma load was significantly higher in IAP 3 than IAP 1–2 for both plants and insects. Phytoplasmas were identified both in midgut and salivary glands of S. titanus at all IAP times. The possible implications of these results in the epidemiology of flavescence dorée are discussed.
The phytoplasmal agent of Flavescence dorée (FD) is transmitted to grapevine by the leafhopper Scaphoideus titanus. Even though 30‐45 days are reported to be required from acquisition (by nymphs) to inoculation (by adults), successful transmission could be achieved even after a shorter period, and it could start directly from adults. FD phytoplasma transmission experiments with S. titanus adults revealed that as soon as 14 days from acquisition the leafhoppers were able to transmit the pathogen to healthy broad beans. The results of this work open to new epidemiologic scenarios, which should be taken into account for FD management.
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BFBNIB, FZAB, GIS, IJS, IZUM, KILJ, NLZOH, NUK, OILJ, PILJ, PNG, SAZU, SBCE, SBMB, UL, UM, UPUK
Since a few decades, apiculture is facing important economic losses worldwide with general major consequences in many areas of agriculture. A strong attention has been paid towards the phenomenon ...named Colony Collapse Disorder in which colonies suddenly disappear with no clear explanations. Honeybee colonies can be affected by abiotic factors, such as environmental pollution or insecticide applications for agricultural purposes. Also biotic stresses cause colony losses, including bacterial (e.g. Paenibacillus larvae) and fungal (e.g. Ascosphaera apis) pathogens, microsporidia (e.g. Nosema apis), parasites (i.e. Varroa destructor) and several viruses. In the light of recent research, intestinal dysbiosis, considered as the relative disproportion of the species within the native microbiota, has shown to affect human and animal health. In arthropods, alteration of the gut microbial climax community has been shown to be linked to health and fitness disequilibrium, like in the medfly Ceratitis capitata for which low mate competitiveness is determined by a gut microbial community imbalance. According to these observations, it is possible to hypothesize that dysbiosis may have a role in disease occurrence also in honeybees. Here we aim to discuss the current knowledge on dysbiosis in the honeybee and its relation with honeybee health by reviewing the investigations of the microbial diversity associated to honeybees and the recent experiments performed to control bee diseases by microbial symbionts. We conclude that, despite the importance of a good functionality of the associated microbiota in preserving insect health has been proved, the mechanisms involved in honeybee gut dysbiosis are still unknown. Accurate in vitro, in vivo and in field investigations are required under healthy, diseased and stressed conditions for the host.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Molecular species recognition and identification, based on the mitochondrial cox1 and on the nuclear ITS2, were performed on individuals of Torymus sinensis collected in Italy, on its close relative ...T. beneficus and on native torymids. The automatic-gap-discovery (ABGD) analyses correctly separate almost all morphospecies. On the basis of cox1, individuals of late-spring T. beneficus clustered with T. sinensis, and those identified as early-spring T. beneficus were recognized as a separate entity. Whereas, T. beneficus ecotypes clustered with T. sinensis on the basis of ITS2. Coalescent tree-based methods confirmed these results. The cox1-based recognition of early-spring T. beneficus as a separate phylospecies led us to conclude that this taxon deserves to be treated as a valid species, whereas individuals identified as late-spring T. beneficus might be considered as part of T. sinensis. Morphological identification and BLAST analyses confirmed that no T. beneficus was imported into Italy to control Dryocosmus kuriphilus.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Abstract Scaphoideus titanus is the insect vector of flavescence dorée (FD), a yellow disease of grapevines. Observations on adult females and nymphs of S. titanus showed that this insect is ...associated with a complex microbial community. Ultrastructural analysis showed that the fat body, salivary glands and ovary of the insect harbour microorganisms showing the brush-like structure typically observed in the genus Cardinium . In particular, it has been shown that these symbiotic bacteria are present both in the follicular cells and in the eggs. In addition, cells resembling bacteriocytes, harbouring numerous Cardinium symbionts in the cytoplasm, were observed in the apical portion of the ovary in adult females. These cells are likely responsible for bacterial transmission to the ovary. Optical microscopy showed that the fat body harbours an enormous population of yeast-like symbionts (YLSs). Ultrastructural observations showed that these symbionts are enclosed within specialized cells of the fat body and are also present in the ovary, where they are found in both the follicular cells and the eggs. There is thus evidence that both Cardinium and the YLSs are transovarially transmitted to the offspring. To our knowledge, S. titanus is the sole insect known to transmit two different kinds of symbionts to the eggs, a prokaryote and an eukaryote. Gene sequence analysis and in situ hybridization led to the identification of YLSs as members of the class Sordariomycetes (=Pyrenomycetes). Finally, ultrastructural observation of the midgut content revealed the presence, in both adult females and nymphs, of a complex microbial community, which include a phytoplasma-like microorganism, likely the agent of FD.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
Insects depend on innate immunity only to defend themselves against pathogens and to regulate interactions with many other microorganisms, such as different kinds of symbionts. Recently, it has been ...suggested that immunocytes could play a role in the vectorial capacity of insects leading to an increased interest towards primary immunocyte cultures. We analysed at molecular and cellular level the immune response of the leafhopper Euscelidius variegatus with the aim to provide an in vitro model for studying the insect-microbe interactions. We in vitro cultured and kept alive for more than 3 months E. variegatus immunocytes that showed a mitotic capacity as well as adhesion and phagocytic activities. In situ hybridization revealed that the defensin gene is actively transcribed in cultured immunocytes, while cecropins were not recorded in this species. These promising results obtained with E. variegaus, a leafhopper frequently used as a laboratory experimental model of insect vector of phytoplasmas, will help in developing in vitro tools for the study of the interactions between these pathogens and their vectors.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Insects depend on innate immunity only to defend themselves against pathogens and to regulate interactions with many other microorganisms, such as different kinds of symbionts. Recently, it has been ...suggested that immunocytes could play a role in the vectorial capacity of insects leading to an increased interest towards primary immunocyte cultures. We analysed at molecular and cellular level the immune response of the leafhopper Euscelidius variegatus with the aim to provide an in vitro model for studying the insect-microbe interactions. We in vitro cultured and kept alive for more than 3 months E. variegatus immunocytes that showed a mitotic capacity as well as adhesion and phagocytic activities. In situ hybridization revealed that the defensin gene is actively transcribed in cultured immunocytes, while cecropins were not recorded in this species. These promising results obtained with E. variegaus, a leafhopper frequently used as a laboratory experimental model of insect vector of phytoplasmas, will help in developing in vitro tools for the study of the interactions between these pathogens and their vectors. Key Words: innate immunity; circulating immunocytes; defensin, insect vectors; leafhopper
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
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