The larva of the black soldier fly (Hermetia illucens) has emerged as an efficient system for the bioconversion of organic waste. Although many research efforts are devoted to the optimization of rearing conditions to increase the yield of the bioconversion process, microbiological aspects related to this insect are still neglected. Here we describe the microbiota of the midgut of H. illucens larvae showing the effect of different diets and midgut regions in shaping microbial load and diversity. The bacterial communities residing in the three parts of the midgut, characterized by remarkable changes in luminal pH values, differed in terms of bacterial numbers and microbiota composition. The microbiota of the anterior part of the midgut showed the highest diversity that gradually decreased along the midgut, whereas bacterial load had an opposite trend, being maximal in the posterior region. The results also showed that the influence of the microbial content of ingested food was limited to the anterior part of the midgut and that the feeding activity of H. illucens larvae did not affect significantly the microbiota of the substrate. Moreover, a high protein content compared to other macronutrients in the feeding substrate seems to favor midgut dysbiosis. The overall data indicate the importance of taking into account the presence of different midgut structural and functional domains, as well as substrate microbiota, in any further study that aims at clarifying microbiological aspects concerning H. illucens larval midgut. IMPORTANCE The demand for food of animal origin is expected to increase by 2050. Since traditional protein sources for monogastric diets are failing to meet the increasing demand for additional feed production, there is an urgent need to find alternative protein sources. The larvae of Hermetia illucens emerge as efficient converters of low quality biomass into nutritionally valuable proteins. Many studies have been performed to optimize H. illucens mass rearing on a number of organic substrates and to maximize quantitatively and qualitatively the biomass yield. On the contrary, although insect microbiota can be fundamental for bioconversion processes and its characterization is mandatory also for safety aspects, this topic is largely overlooked. Here we provide an in depth study of the microbiota of H. illucens larval midgut taking into account pivotal aspects such as the midgut spatial and functional regionalization as well as microbiota and nutrient composition of the feeding substrate.

Endophytic fungi (EF) can enhance both plant growth and defense barriers against pests and pathogens, contributing to the reduction of chemical pesticides and fertilizers use in agriculture. Beauveria bassiana is an entomopathogenic fungus showing endophytism in several crops, often associated with a good capacity to limit the development of pests and disease agents. However, the diversity of the protective efficacy and plant response to different strains can be remarkable and needs to be carefully assessed for the successful and predictable use of these beneficial microorganisms. This study aims to select B. bassiana strains able to colonize tomato plants as endophytes as well as to control two important disease agents, Botrytis cinerea and Alternaria alternata, and the pest aphid, Macrosiphum euphorbiae. Nine wild-type isolates and one commercial strain were screened for endophytism, then further characterized for plant-growth promotion plus inhibition of disease development and pest infestation. Four isolates proved to have a good control activity against the biotic stressors tested, but only Bb716 was also able to promote plant growth. This work provides a simple workflow for the selection of beneficial EF, paving the way towards more effective use of B. bassiana in Integrate Pest Management (IPM) of tomato.

Plants generate energy flows through natural food webs, driven by competition for resources among organisms, which are part of a complex network of multitrophic interactions. Here, we demonstrate that the interaction between tomato plants and a phytophagous insect is driven by a hidden interplay between their respective microbiotas. Tomato plants colonized by the soil fungus Trichoderma afroharzianum , a beneficial microorganism widely used in agriculture as a biocontrol agent, negatively affects the development and survival of the lepidopteran pest Spodoptera littoralis by altering the larval gut microbiota and its nutritional support to the host. Indeed, experiments aimed to restore the functional microbial community in the gut allow a complete rescue. Our results shed light on a novel role played by a soil microorganism in the modulation of plant–insect interaction, setting the stage for a more comprehensive analysis of the impact that biocontrol agents may have on ecological sustainability of agricultural systems.

Plant defense peptides represent an important class of compounds active against pathogens and insects. These molecules controlling immune barriers can potentially be used as novel tools for plant protection, which mimic natural defense mechanisms against invaders. The constitutive expression in tomato plants of the precursor of the defense peptide systemin was previously demonstrated to increase tolerance against moth larvae and aphids and to hamper the colonization by phytopathogenic fungi, through the expression of a wealth of defense-related genes. In this work we studied the impact of the exogenous supply of systemin to tomato plants on pests to evaluate the use of the peptide as a tool for crop protection in non-transgenic approaches. By combining gene expression studies and bioassays with different pests we demonstrate that the exogenous supply of systemin to tomato plants enhances both direct and indirect defense barriers. Experimental plants, exposed to this peptide by foliar spotting or root uptake through hydroponic culture, impaired larval growth and development of the noctuid moth Spodoptera littoralis, even across generations, reduced the leaf colonization by the fungal pathogen Botrytis cinerea and were more attractive towards natural herbivore antagonists. The induction of these defense responses was found to be associated with molecular and biochemical changes under control of the systemin signalling cascade. Our results indicate that the direct delivery of systemin, likely characterized by a null effect on non-target organisms, represents an interesting tool for the sustainable protection of tomato plants.

Genome sequencing data have recently demonstrated that eukaryote evolution has been remarkably influenced by the acquisition of a large number of genes by horizontal gene transfer (HGT) across different kingdoms. However, in depth-studies on the physiological traits conferred by these accidental DNA acquisitions are largely lacking. Here we elucidate the functional role of Sl gasmin, a gene of a symbiotic virus of a parasitic wasp that has been transferred to an ancestor of the moth species Spodoptera littoralis and domesticated. This gene is highly expressed in circulating immune cells (haemocytes) of larval stages, where its transcription is rapidly boosted by injection of microorganisms into the body cavity. RNAi silencing of Sl gasmin generates a phenotype characterized by a precocious suppression of phagocytic activity by haemocytes, which is rescued when these immune cells are incubated in plasma samples of control larvae, containing high levels of the encoded protein. Proteomic analysis demonstrates that the protein Sl gasmin is released by haemocytes into the haemolymph, where it opsonizes the invading bacteria to promote their phagocytosis, both in vitro and in vivo. Our results show that important physiological traits do not necessarily originate from evolution of pre-existing genes, but can be acquired by HGT events, through unique pathways of symbiotic evolution. These findings indicate that insects can paradoxically acquire selective advantages with the help of their natural enemies. Parasitic wasps are important insect biocontrol agents. These insects are beneficial for the ecological service they provide, which largely contributes to the control of natural populations of their hosts. Paradoxically, they can be beneficial also for non-host individuals attacked by mistake, if these survive after parasitization. In nature, this can happen to lepidopteran larvae when attacked by a wasp harbouring a symbiotic virus that can mediate horizontal gene transfer. Indeed, this virus, injected along with the egg in the body of the host, can get integrated in the genome of the parasitized larva, carrying along exogenous genes. Because the non-host regularly suppresses the parasitoid egg and/or juveniles, any surviving individual with a stable insertion of new genes in the germ line will represent an evolutionary novelty, with expanded functional capacities, if the resulting gene domestication event confers new physiological traits. The immune function here discovered demonstrates that symbiotic associations can drive unique evolutionary pathways, maximizing the fitness of interacting organisms, which evolve as a complex unit with a shared gene pool.

Acetylcholine (ACh) is one the major neurotransmitters in insects, whose role in mediating synaptic interactions between neurons in the central nervous system is well characterized. It also plays largely unexplored regulatory functions in non-neuronal tissues. Here we demonstrate that ACh signaling is involved in the modulation of the innate immune response of Drosophila melanogaster. Knockdown of ACh synthesis or ACh vesicular transport in neurons reduced the activation of drosomycin (drs), a gene encoding an antimicrobial peptide, in adult flies infected with a Gram-positive bacterium. drs transcription was similarly affected in Drosophila α7 nicotinic acetylcholine receptor, nAChRalpha7 (Dα7) mutants, as well as in flies expressing in the nervous system a dominant negative form (Dα7^DN) of this specific receptor subunit. Interestingly, Dα7^DN elicited a comparable response when it was expressed in non-neuronal tissues and even when it was specifically produced in the hemocytes. Consistently, full activation of the drs gene required Dα7 expression in these cells. Moreover, knockdown of ACh synthesis in non-neuronal cells affected drs expression. Overall, these findings uncover neural and non-neural cholinergic signals that modulate insect immune defenses and shed light on the role of hemocytes in the regulation of the humoral immune response.