Active and covert infections of cricket Iridovirus and Acheta domesticus Densovirus in reared Gryllodes sigillatus crickets

Interest in developing food, feed, and other useful products from farmed insects has gained remarkable momentum in the past decade. Crickets are an especially popular group of farmed insects due to their nutritional quality, ease of rearing, and utility. However, production of crickets as an emerging commodity has been severely impacted by entomopathogenic infections, about which we know little. Here, we identified and characterized an unknown entomopathogen causing mass mortality in a lab-reared population of Gryllodes sigillatus crickets, a species used as an alternative to the popular Acheta domesticus due to its claimed tolerance to prevalent entomopathogenic viruses. Microdissection of sick and healthy crickets coupled with metagenomics-based identification and real-time qPCR viral quantification indicated high levels of cricket iridovirus (CrIV) in a symptomatic population, and evidence of covert CrIV infections in a healthy population. Our study also identified covert infections of Acheta domesticus densovirus (AdDNV) in both populations of G. sigillatus. These results add to the foundational research needed to better understand the pathology of mass-reared insects and ultimately develop the prevention, mitigation, and intervention strategies needed for economical production of insects as a commodity

Metarhizium brunneum Blastospore Pathogenesis in Aedes aegypti Larvae: Attack on Several Fronts Accelerates Mortality

Aedes aegypti is the vector of a wide range of diseases (e.g. yellow fever, dengue, Chikungunya and Zika) which impact on over half the world's population. Entomopathogenic fungi such as Metarhizium anisopliae and Beauveria bassiana have been found to be highly efficacious in killing mosquito larvae but only now are the underlying mechanisms for pathogenesis being elucidated. Recently it was shown that conidia of M. anisopliae caused stress induced mortality in Ae. aegypti larvae, a different mode of pathogenicity to that normally seen in terrestrial hosts. Blastospores constitute a different form of inoculum produced by this fungus when cultured in liquid media and although blastospores are generally considered to be more virulent than conidia no evidence has been presented to explain why. In our study, using a range of biochemical, molecular and microscopy methods, the infection process of Metarhizium brunneum (formerly M. anisopliae) ARSEF 4556 blastospores was investigated. It appears that the blastospores, unlike conidia, readily adhere to and penetrate mosquito larval cuticle. The blastospores are readily ingested by the larvae but unlike the conidia are able infect the insect through the gut and rapidly invade the haemocoel. The fact that pathogenicity related genes were upregulated in blastospores exposed to larvae prior to invasion, suggests the fungus was detecting host derived cues. Similarly, immune and defence genes were upregulated in the host prior to infection suggesting mosquitoes were also able to detect pathogen-derived cues. The hydrophilic blastospores produce copious mucilage, which probably facilitates adhesion to the host but do not appear to depend on production of Pr1, a cuticle degrading subtilisin protease, for penetration since protease inhibitors did not significantly alter blastospore virulence. The fact the blastospores have multiple routes of entry (cuticle and gut) may explain why this form of the inoculum killed Ae. aegypti larvae in a relatively short time (12-24hrs), significantly quicker than when larvae were exposed to conidia. This study shows that selecting the appropriate form of inoculum is important for efficacious control of disease vectors such as Ae. aegypti

Doing synthetic biology with photosynthetic microorganisms

The use of photosynthetic microbes as synthetic biology hosts for the sustainable production of commodity chemicals and even fuels has received increasing attention over the last decade. The number of studies published, tools implemented, and resources made available for microalgae have increased beyond expectations during the last few years. However, the tools available for genetic engineering in these organisms still lag those available for the more commonly used heterotrophic host organisms. In this mini-review, we provide an overview of the photosynthetic microbes most commonly used in synthetic biology studies, namely cyanobacteria, chlorophytes, eustigmatophytes and diatoms. We provide basic information on the techniques and tools available for each model group of organisms, we outline the state-of-the-art, and we list the synthetic biology tools that have been successfully used. We specifically focus on the latest CRISPR developments, as we believe that precision editing and advanced genetic engineering tools will be pivotal to the advancement of the field. Finally, we discuss the relative strengths and weaknesses of each group of organisms and examine the challenges that need to be overcome to achieve their synthetic biology potential.Peer reviewe

Repellency of a Wax-Based Catnip-Oil Formulation against Stable Flies

Stable flies, Stomoxys calcitrans (L.), are one of the most serious livestock pests, which cause significant economic loss in the cattle industry. Current practices for managing stable flies are limited to costly sanitation techniques and unsustainable insecticide applications. The present study reports the initial efforts using catnip essential oil as a spatial repellent and the results of field trials using a wax-based formulation to repel stable flies in the cattle feedlot. Electroantennograms showed that catnip oil and its ingredient compounds elicit significant antennal responses from both sexes of stable flies. Catnip oil and ZE- and EZ-nepetalactone showed repellent activity in a single cage olfactormeter study. No behavioral activity was observed from another ingredient compound, caryophyllene. A laboratory dispersal bioassay also showed that stable flies avoided areas treated with catnip oil. Using a solid phase microextraction (SPME) method, the atmospheric concentration of catnip active ingredient compounds (nepetalactones) absorbed by SPME fiber in treated areas was detected at 4 times higher than those in control areas. Application of wax-based catnip pellets in cattle feedlots resulted in \u3e99% repellency of stable flies in treated areas, compared with that in nontreated areas. However, the repellent efficacy of the formulation only lasted 3 h. This is the first study demonstrating the potential application of a plant-based repellent formulation that may be used as an alternative method against stable flies

Increased Phenoloxidase Activity Constitutes the Main Defense Strategy of <i>Trichoplusia ni</i> Larvae against Fungal Entomopathogenic Infections

The cabbage looper Trichoplusia ni is an important agricultural pest worldwide and is frequently used as a model organism for assessing entomopathogenic fungi virulence, though few studies have measured the host response repertoire to fungal biocontrol agents. Here, we quantified the immune response of T. ni larvae following exposure to two entomopathogenic fungal species: Beauveria bassiana and Cordyceps javanica. Results from our study demonstrate that T. ni larvae exposed to fungal entomopathogens had higher total phenoloxidase activity compared to controls, indicating that the melanization cascade is one of the main immune components driving defense against fungal infection and contrasting observations from other insect–fungi interaction studies. We also observed differences in host response depending on the species of entomopathogenic fungi, with significantly higher induction observed during infections with B. bassiana than with C. javanica. Larvae exposed to B. bassiana had an increased expression of genes involved in prophenoloxidase response and the Imd, JNK, and Jak/STAT immune signaling pathways. Our results indicate a notable absence of Toll pathway-related responses, further contrasting results to other insect–fungi pathosystems. Important differences were also observed in the induction of antimicrobial effectors, with B. bassiana infections eliciting three antimicrobial effectors (lysozyme, gloverin, and cecropin), while C. javanica only induced cecropin expression. These results provide insight into the host response strategies employed by T. ni for protection against entomopathogenic fungi and increase our understanding of insect–fungal entomopathogen interactions, aiding in the design of more effective microbial control strategies for this important agricultural pest