Bioecological notes on Xylosandrus compactus (Eichhoff) (Coleoptera Curculionidae Scolytinae), a species recently recorded into Italy

Intense trade flows favour the accidental diffusion of alien plant and animal species harmful to agricultural crops and forest ecosystems in many parts of the world. Particularly prone to such introductions are lignicolous species of woodboring beetles that attack living plants and can develop in lumber or crating wood, such as many species belonging to the subfamilies Scolytinae and Platypodinae (Coleoptera: Curculionidae). Investigations of marked witherings of laurel twigs (Laurus nobilis L.) carried out from autumn 2010 in an area of the Versilia coast (Lucca) revealed that the Black twig borer, Xylosandrus compactus (Eichhoff), was involved in this pathological expression. This species was recently recorded for the first time in Italy on ornamental plants in some localities in the Naples’s province. Here we report its essential external morphological features, with useful characters for its distinction from X. germanus (Blandford), a morphologically similar species, as well as its essential bioecological features, with special reference to what has been observed in the abovementioned area of Tuscany. This should facilitate the rapid identification and verification of new outbreaks and, where possible, the timely implementation of control strategies

BACTERIA ASSOCIATED TO RHYNCHOPHORUS FERRUGINEUS (OLIVIER) (COLEOPTERA DRYOPHTHORIDAE) IN ITALY

Thirty-five bacterial strains were selected on the basis of colony morphology from larvae and pupae of Rhynchophorous (Olivier) ferrugineus collected from infested Phoenix canariensis Chabaud trees growing in different Italian locations. 16S rDNA sequencing showed that bacteria isolated from larvae were mostly Gram positive and belonged to Bacillus, Brevibacillus and Paenibacillus genera. Bacteria isolated from pupae were affiliated to Bacillus and five different Gram negative genera (Alcaligenes, Morganella, Myroides, Providencia and Serratia). Some strains produced lytic enzymes like chitinase, lecithinase, lipase and protease in vitro. The enthomopathogenic strains belonging to Bacillus thuringiensis, Providencia rettgeri and Serratia marcescens species may be considered of interest as potential biocontrol agents of R. ferrugineus

Presence of Destruxin A and Beauvericin in cereals

A LC-MS/MS method for the detection of destruxin A (DTX A) and beauvericin (BEA) in cereals was developed, validated and applied to commercial products collected in Italian markets in the years 2015-2016. Results showed that BEA contaminated 59 % of the samples even if only 15 of them (34%) showed quantifiable residues (comprised between 0.11 and 7.51 ng/g). The sample of red rice contaminated with the highest BEA level was also contaminated with DTX A (0.28 ng/g). Finally, no significant differences were detected between contaminated samples based on the production year and the agronomic technology used (organic or conventional farming)

Metarhizium anisopliae biopesticides and fungus isolates: control efficacy against Rhynchophorus ferrugineus (Olivier) (Coleoptera Dryophthoridae) on different contamination substrata.

The lack of biological insecticides based on Metarhizium anisopliae (Metsch.) Sorokin registered in Italy for the control of the Red Palm Weevil (RPW), Rhynchophorus ferrugineus (Olivier), prompted studies to evaluate the efficacy of two commercial products, Met52® and BioStormTM and of their fungal isolates, M. anisopliae (Man52) and (ManBS) respectively, against the adults. The virulence of the M. anisopliae strains (Man52) and (ManBS) was compared with that of an indigenous M. anisopliae (Man08/I05) strain obtained from R. ferrugineus specimens collected in the wild and that showed to be very virulent against the RPW in previous studies. In both tests the sublethal effects of the treatments on female reproductive potential were examined in relation to the infective substratum. Laboratory results indicated that the commercial formulations mixed directly into the soil were not active in transmitting the infection to RPW adults and in reducing female fecundity and fertility. Diversely the fungal M. anisopliae (ManBS), (Man52) and (Man08/I05) strains inoculated on a rice substratum caused over 80% mortality of the phytophagous. In particular, M. anisopliae (ManBS) and the M. anisopliae (Man08/I05) produced the highest mortality (100%), with LT50 e LT90 reached in 3 and 6 days respectively. M. anisopliae (Met52) strain instead led to 85% mortality of RPW specimens in 28-days but it took longer to reach LT50 (6 days) and LT90 (12 days). The reproductive potential of females infected with the fungal strains was also significantly reduced with respect to the control. This study indicates the possibility of using also M. anisopliae strains in use against other insects of agricultural importance, for the control of R. ferrugineus although their effectiveness is conditioned by the formulations and/or methods employed against the RPW

Autocontamination trap with entomopathogenic fungi: a possible strategy in the control of Rhynchophorus ferrugineus (Olivier) (Coleoptera Curculionidae)

An experimental autocontamination trap was devised to infect Rhynchophorus ferrugineus (Olivier), the Red Palm Weevil, adults with entomopathogenic fungi. The aim was to develop an autocontamination device to support integrated R. ferrugineus control programs. In laboratory bioassays, the delivery system successfully attracted, infected and released weevil adults after they contacted cereal substrata inoculated with indigenous strains of Beauveria bassiana (Balsamo- Crivelli) Vuillemin and Metarhizium anisopliae (Metchnikoff) Sorokin. Tests carried out with the experimental traps showed that M. anisopliae was the more virulent pathogen, causing 75% cumulative mortality in adults, while B. bassiana gave a 45% cumulative mortality. Infectivity of M. anisopliae was not affected by different cereal substrata, i.e. wheat and rice, since curculionid cumulative mortality (95%) and treatment efficiency (95% Abbott) were very high on both of them and Red Palm Weevil LT50 was reached within the same time (15 days). Conidial persistence and germinability of M. anisopliae grown on the rice substratum were examined in field conditions inside traps located in sunny and shady positions in spring, summer and autumn. The results showed that the traps preserved fungal inoculum stability longer in spring and summer than in autumn. No significant difference in M. anisopliae conidial persistence was found between sunny and shady traps during the various seasons

Entomopathogenicity of Beauveria bassiana (Bals.) Vuill. and Metarhizium anisopliae (Metsch.) Sorokin isolated from different sources in the control of Rhynchophorus ferrugineus (Olivier) (Coleoptera Curculionidae)

The entomopathogenicity of an indigenous Beauveria bassiana strain isolated from soil and of B. bassiana and Metarhizium anisopliae strains isolated in Italy from naturally infected Rhynchophorus ferrugineus (RPW) adults, was tested against larvae and adults of RPW in laboratory bioassays. The individuals were infected via direct contact on sporulated mycelia grown on Sabouraud Dextrose Agar or on wheat substrata. M. anisopliae obtained from R. ferrugineus showed the highest efficacy against RPW larvae and adults particularly against individuals contaminated on sporulated wheat, which showed values of cumulative larval mortality of 100% and adult mortality of 90%; LT50 was obtained in 13.1 days in both larvae and adults. B. bassiana strain isolated from soil recorded a lower cumulative mortality on larvae (13%) and adults (13%) treated on inoculated Sabouraud Dextrose Agar. B. bassiana strain isolated from RPW showed cumulative mortality values higher than 50% against larvae treated on inoculated wheat (55%) and Sabouraud Dextrose Agar (53%); LT50 was obtained in 15 days and 21.8.days respectively. Results are discussed with regard to the potential employment of the virulent indigenous strain of M. anisopliae for microbiological control of R. ferrugineus as part of an integrated pest management program

Tenebrio molitor as a Simple and Cheap Preclinical Pharmacokinetic and Toxicity Model

The progression of drugs into clinical phases requires proper toxicity assessment in animals and the correct identification of possible metabolites. Accordingly, different animal models are used to preliminarily evaluate toxicity and biotransformations. Rodents are the most common models used to preliminarily evaluate the safety of drugs; however, their use is subject to ethical consideration and elevated costs, and strictly regulated by national legislations. Herein, we developed a novel, cheap and convenient toxicity model using Tenebrio molitor coleoptera (TMC). A panel of 15 drugs-including antivirals and antibacterials-with different therapeutic applications was administered to TMC and the LD50 was determined. The values are comparable with those already determined in mice and rats. In addition, a TMC model was used to determine the presence of the main metabolites and in vivo pharmacokinetics (PK), and results were compared with those available from in vitro assays and the literature. Taken together, our results demonstrate that TMC can be used as a novel and convenient preliminary toxicity model to preliminarily evaluate the safety of experimental compounds and the formation of main metabolites, and to reduce the costs and number of rodents, according to 3R principles

Effects of cryopreservation and freeze-drying on proteases enzymatic activity of entomopathogenic strains of Beauveria bassiana (Balsamo) Vuillemin and Metarhizium anisopliae (Metchnikoff) Sorokin.

Beauveria bassiana and Metarhizium anisopliae are fungal species known for their entomopathogenic activity; they represent an important alternative to chemical control of many pest insects and appropriate preservation methods are thus necessary to maintain their properties over time. In the present study the effects of cryopreservation and freeze­drying were investigated on virulence determinants such as proteases and specifically on subtilisin-type Pr 1 and Pr 2 that seem to play a predominant role in insect fungal penetration. Three Metharizium anisopliae strains (M 13/ I05, M 13/ I12 and M 13/ I33) and three Beauveria bassiana strains (B 13/ I03, B 13/ I57 and B 13/ I63) were tested. Data obtained from our experiments show that the decrease of Pr 1 and Pr 2 enzymatic activity in B. bassiana and M. anisopliae strains 7 months after storage does not affect their entomopathogenic ability against larvae of Tenebrio molitor compared to the fresh cultures before storage. The role of Pr 1 and Pr 2 as predominant factors in the entomopathogenicity of both fungal species is discussed. No significant differences were found in any strain tested for morphological features using both preservation methods

FIRST INTERCEPTION OF TRICHOFERUS CAMPESTRIS (FALDERMANN, 1835) (COLEOPTERA CERAMBYCIDAE CERAMBYCINAE) IN ITALY

The Velvet Longhorned Beetle Trichoferus campestris (Faldermann, 1835) was intercepted for the first time in Italy, in the Naples harbour, during monitoring activities at entry points carried out in the context of the national project ASPROPI. T. campestris is widely polyphagous and able to colonize several woody species in both agricultural and forest environments. Moreover timber and solid wood items can be equally attractive to this pest. This additional report, in an area where other alien insect species had been identified in the past, emphasises how the national monitoring network represents a key element of the alert system designed to quickly counter the accidental introduction and spreading of exotic insect pests

Assessing the Efficiency of Molecular Markers for the Species Identification of Gregarines Isolated from the Mealworm and Super Worm Midgut

Protozoa, of the taxon Gregarinasina, are a heterogeneous group of Apicomplexa that includes ~1600 species. They are parasites of a large variety of both marine and terrestrial invertebrates, mainly annelids, arthropods and mollusks. Unlike coccidians and heamosporidians, gregarines have not proven to have a negative effect on human welfare; thus, they have been poorly investigated. This study focuses on the molecular identification and phylogeny of the gregarine species found in the midgut of two insect species that are considered as an alternative source of animal proteins for the human diet: the mealworm Tenebriomolitor, and the super-worm Zophobasatratus (Coleoptera: Tenebrionidae). Gregarine specimens were isolated from the gut of both larval and adult stages of T.molitor specimens, as well as from Z.atratus larvae. The morphological analyses were restricted to the identification of the different parasite morpho-types, likely corresponding either to different life-cycle stages or to alternative gregarine species. The samples were also used for the DNA extraction necessary for their genetic characterization. Finally, the efficiency of different molecular markers (i.e., 18S rDNA gene alone or combined with the Internal Transcribed Spacer 1) was assessed when applied either to gregarine species identification and to phylogenetic inference