Asaia, a versatile acetic acid bacterial symbiont, capable of cross-colonizing insects of phylogenetically-distant genera and orders

Bacterial symbionts of insects have been proposed for blocking transmission of vector-borne pathogens. However, in many vector models the ecology of symbionts and their capability of cross-colonizing different hosts, an important feature in the symbiotic control approach, is poorly known. Here we show that the acetic acid bacterium Asaia, previously found in the malaria mosquito vector Anopheles stephensi, is also present in and capable of cross-colonizing other sugar-feeding insects of phylogenetically distant genera and orders. PCR, real-time PCR and in situ-hybridization experiments showed Asaia in the body of the mosquito Aedes aegypti and the leafhopper Scaphoideus titanus, vectors of human viruses and a grapevine phytoplasma, respectively. Cross colonization patterns of the body of Ae. aegypti, An. stephensi and S. titanus have been documented with Asaia strains isolated from An. stephensi or Ae. aegypti, and labelled with plasmid- or chromosome-encoded fluorescent proteins (Gfp and DsRed, respectively). Fluorescence and confocal microscopy showed that Asaia, administered with the sugar meal, efficiently colonized guts, male and female reproductive systems and the salivary glands. The ability in cross-colonizing insects of phylogenetically distant orders indicates that Asaia adopts body invasion mechanisms independent from the host biological characteristics. This versatility is an important property for the development of symbiont-based therapies of different vector-borne diseases

Interactions between Asaia, Plasmodium and Anopheles: new insights into mosquito symbiosis and implications in malaria symbiotic control

Background Malaria represents one of the most devastating infectious diseases. The lack of an effective vaccine and the emergence of drug resistance make necessary the development of new effective control methods. The recent identification of bacteria of the genus Asaia, associated with larvae and adults of malaria vectors, designates them as suitable candidates for malaria paratransgenic control. To better characterize the interactions between Asaia, Plasmodium and the mosquito immune system we performed an integrated experimental approach. Methods Quantitative PCR analysis of the amount of native Asaia was performed on individual Anopheles stephensi specimens. Mosquito infection was carried out with the strain PbGFPCON and the number of parasites in the midgut was counted by fluorescent microscopy. The colonisation of infected mosquitoes was achieved using GFP or DsRed tagged-Asaia strains. Reverse transcriptase-PCR analysis, growth and phagocytosis tests were performed using An. Stephensi and Drosophila melanogaster haemocyte cultures and DsRed tagged-Asaia and Escherichia coli strains. Results Using quantitative PCR we have quantified the relative amount of Asaia in infected and uninfected mosquitoes, showing that the parasite does not interfere with bacterial blooming. The correlation curves have confirmed the active replication of Asaia, while at the same time, the intense decrease of the parasite. The 'in vitro' immunological studies have shown that Asaia induces the expression of antimicrobial peptides, however, the growth curves in conditioned medium as well as a phagocytosis test, indicated that the bacterium is not an immune-target. Using fluorescent strains of Asaia and Plasmodium we defined their co-localisation in the mosquito midgut and salivary glands. Conclusions We have provided important information about the relationship of Asaia with both Plasmodium and Anopheles. First, physiological changes in the midgut following an infected or uninfected blood meal do not negatively affect the residing Asaia population that seems to benefit from this condition. Second, Asaia can act as an immune-modulator activating antimicrobial peptide expression and seems to be adapted to the host immune response. Last, the co-localization of Asaia and Plasmodium highlights the possibility of reducing vectorial competence using bacterial recombinant strains capable of releasing anti-parasite molecules

Characterisation of Refined Marc Distillates with Alternative Oak Products Using Different Analytical Approaches

The use of oak barrel alternatives, including oak chips, oak staves and oak powder, is quite common in the production of spirits obtained from the distillation of vegetal fermented products such as grape pomace. This work explored the use of unconventional wood formats such as peeled and sliced wood. The use of poplar wood was also evaluated to verify its technological uses to produce aged spirits. To this aim, GC-MS analyses were carried out to obtain an aromatic characterisation of experimental distillates treated with these products. Moreover, the same spirits were studied for classification purposes using NMR, NIR and e-nose. A significant change in the original composition of grape pomace distillate due to sorption phenomena was observed; the intensity of this effect was greater for poplar wood. The release of aroma compounds from wood depended both on the toasting level and wood assortment. Higher levels of xylovolatiles, namely, whisky lactone, were measured in samples aged using sliced woods. Both the NIR and NMR analyses highlighted similarities among samples refined with oak tablets, differentiating them from the other wood types. Finally, E-nose seemed to be a promising alternative to spectroscopic methods both for the simplicity of sample preparation and method portability

Paternal transmission of symbiotic bacteria in malaria vectors

Bacteria of the genus Asaia are associated with different species of malaria vectors and are located in the midgut, salivary glands and reproductive organs of female and male mosquitoes. Based on current evidence, the spreading of these bacteria in mosquito populations occurs through different mechanisms: co-feeding, sexual mating, and maternal transmission [1,2]. Even though paternal transmission of insect symbionts to progeny is not commonplace, the presence of Asaia in the male reproductive organs makes this additional transmission route worth being investigated. Here, we show that male-borne Asaia are transferred to females during the mating of Anopheles stephensi mosquitoes. Subsequently, the bacteria acquired by the female are vertically transmitted to the progeny. It would thus be possible to use male mosquitoes, which do not bite, to spread Asaia strains interfering with malaria transmission

Genetic Deletion of the Nociceptin/Orphanin FQ Receptor in the Rat Confers Resilience to the Development of Drug Addiction

The nociceptin (NOP) receptor is a G-protein-coupled receptor whose natural ligand is the nociceptin/orphanin FQ (N/OFQ) peptide. Evidence from pharmacological studies suggests that the N/OFQ system is implicated in the regulation of several addiction-related phenomena, such as drug intake, withdrawal and relapse. Here, to further explore the role of NOP system in addiction, we used NOP (-/-) rats to study the motivation for cocaine, heroin and alcohol self-administration in the absence of N/OFQ function. Conditioned place preference (CPP) and saccharin (0.2% w/v) self-administration were also investigated. Results showed that NOP (-/-) rats self-administer less cocaine (0.25, 0.125 or 0.5 mg/infusion) both under a Fixed Ratio 1 and a Progressive Ratio schedule of reinforcement compared to wild type (Wt) controls. Consistently, cocaine (10 mg/kg, i.p.) was able to induce CPP in Wt but not in NOP (-/-). When NOP (-/-) rats were tested for heroin (20 μg/infusion) and ethanol (10% v/v) self-administration, they showeda significantly lower drug intake compared to Wt. Conversely, saccharin self-administration was not affected by NOP deletion, excluding the possibility of nonspecific learning deficits or generalized disruption of reward mechanisms in NOP (-/-) rats. These findings were confirmed with pharmacological experiments using two selective NOP antagonists, SB-612111 and LY2817412. Both drugs attenuated alcohol self-administration in Wt rats but not in NOP (-/-) rats. In conclusion, our results demonstrate that genetic deletion of NOP receptors confers resilience to drug abuse and support a role for NOP receptor antagonism as a potential treatment option for drug addiction.Neuropsychopharmacology accepted article preview online, 26 August 2016. doi:10.1038/npp.2016.171

Liquid membrane potential in nonisothermal systems.

Electrical membrane potential equations for liquid ion exchange membranes, characterized by the presence of uncharged associated species and by exclusion of co-ions (no electrolyte uptake) have been derived. The irreversible thermodynamic theories already developed for solid membranes with fixed charged site density have been extended to include the different physicochemical aspects of the liquid membranes. To this purpose the dissipation function has been written with reference to the fluxes of all the species present in the membrane. It has been found that the mobile charged site, the counterions, and the uncharged associated species contribute to the electrical membrane potential through their phenomenological coefficients. The electrical membrane potential equations have been integrated in isothermal and nonisothermal conditions for monoionic and biionic systems. The theoretical predictions have been experimentally tested by studying the electrical potential of liquid membranes formed with solutions of tetraheptylammonium salts in omicron-dichlorobenzene

Lyme Disease and Babesiosis: preliminary findings on the transmission risk in highly frequented areas of the Monti Sibillini National Park (Central Italy).

The Monti Sibillini National Park is a recently borne protected area in the central Apennines that has become an important tourist destination. Mountain grasslands and woods have made it also a suitable habitat for sheep and bovine cattle, as well as for wild animals such as the wild bore and the roe deer. Therefore, a preliminary investigation was conducted to assess the risk of transmission of tick-borne zoonoses, such as Lyme disease and Babesiosis, by actively looking for both the arthropod vector and the causative agent. For two consecutive years, ambushing ticks were collected in four distinct geographical areas, comprehensive of many highly frequented tourist places. The tick fauna, tick habitats and the seasonal distribution of the different tick life stages collected with the method of "flagging and dragging" have been reported. Almost all the collected specimens belonged to the species Haemaphysalis punctata (Canestrini and Fanzago) (Acari: Ixodidae), which was found moderately infected with Babesia spp. Only a few Ixodes ricinus (Linnaeus) ticks, the most competent vector of Lyme disease, were found and on PCR examination all of them resulted negative as far as the infectious agent Borrelia burgdorferi sensu lato (Johnson) is concerned

Natural infection of Anopheles maculipennis s.l. with Dirofilaria repens in central Italy.

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Uso della PCR nella diagnosi di Mullus barbatus e M. surmuletus

L’UE con l’adozione del Reg. (CE) 2065/2001, recante applicazione Reg. (CE) 104/2000, ha voluto dare una forte spinta alla “rintracciabilità dei prodotti della pesca” mediante il sistema di etichettatura che obbliga il commerciante a fornire le seguenti informazioni sulle caratteristiche dei prodotti ittici messi in vendita: “il metodo di produzione”, “la zona di cattura” e la “denominazione commerciale della specie”, cioè il nome comune o scientifico del prodotto. Per l’Italia quindi “la denominazione commerciale” deve rispettare l’elenco riportato nell’allegato A del D.M. 27-03-02 e successive modifiche e integrazioni del D.M. 25/07/2005. In questo studio si presentano i risultati per l’identificazione di specie di Mullus barbatus e di Mullus surmuletus, ottenuti mediante un sistema di Polymerase Chain Reaction (PCR) multiplex progettato sul locus 16S rRNA dei 3 generi di Mullidi (Mullus, Upeneus e Pseudopeneus), più diffusi nel mediterraneo

Deciphering microbiota associated to Rhynchophorus ferrugineus in Italian samples: a preliminary study

The Red Palm Weevil, Rhynchophorus ferrugineus (Olivier, 1790) is the most dangerous and deadly pest of date, coconut, oil, sago and other palms. Recently introduced in the Mediterranean basin, it became the most relevant insect pest for ornamental palms in the urban environment. Given the development of an innovative control method based on symbiotic control, we have performed a pilot project to decrypt the microbiota associated to both adults and larval stages of the insect to identify potential tools for biocontrol agents against the Palm Weevil. A number of bacterial species were found associated with the insect. In particular, species of the genera Lactococcus, Proteus, and others were detected