Distribution of Transferable Antibiotic Resistance Genes in Laboratory-Reared Edible Mealworms (Tenebrio molitor L.)

In the present study, the distribution of antibiotic resistance genes in laboratory-reared fresh mealworm larvae (Tenebrio molitor L.), their feeding substrates (carrots and wheatmeal), and frass was assessed. Microbial counts on selective media added with antibiotics highlighted the presence of lactic acid bacteria resistant to ampicillin and vancomycin and, more specifically, enterococci resistant to the latter antibiotic. Moreover, staphylococci resistant to gentamicin, erythromycin, tetracycline, and vancomycin were detected. Enterobacteriaceae resistant to ampicillin and gentamicin were also found, together with Pseudomonadaceae resistant to gentamicin. Some of the genes coding for resistance to macrolide-lincosamide-streptogramin B (MLSB) [erm(A), erm(C)], vancomycin [vanA, vanB], tetracycline [tet(O)], and β-lactams [mecA and blaZ] were absent in all of the samples. For the feeding substrates, organic wheatmeal was positive for tet(S) and tet(K), whereas no AR genes were detected in organic carrots. The genes tet(M), tet(K), and tet(S) were detected in both mealworms and frass, whereas gene aac-aph, coding for resistance to amynoglicosides was exclusively detected in frass. No residues for any of the 64 antibiotics belonging to 10 different drug classes were found in either the organic wheatmeal or carrots. Based on the overall results, the contribution of feed to the occurrence of antibiotic resistance (AR) genes and/or antibiotic-resistant microorganisms in mealworm larvae was hypothesized together with vertical transmission via insect egg smearing

Atrasentan and renal events in patients with type 2 diabetes and chronic kidney disease (SONAR): a double-blind, randomised, placebo-controlled trial

Background: Short-term treatment for people with type 2 diabetes using a low dose of the selective endothelin A receptor antagonist atrasentan reduces albuminuria without causing significant sodium retention. We report the long-term effects of treatment with atrasentan on major renal outcomes. Methods: We did this double-blind, randomised, placebo-controlled trial at 689 sites in 41 countries. We enrolled adults aged 18–85 years with type 2 diabetes, estimated glomerular filtration rate (eGFR)25–75 mL/min per 1·73 m 2 of body surface area, and a urine albumin-to-creatinine ratio (UACR)of 300–5000 mg/g who had received maximum labelled or tolerated renin–angiotensin system inhibition for at least 4 weeks. Participants were given atrasentan 0·75 mg orally daily during an enrichment period before random group assignment. Those with a UACR decrease of at least 30% with no substantial fluid retention during the enrichment period (responders)were included in the double-blind treatment period. Responders were randomly assigned to receive either atrasentan 0·75 mg orally daily or placebo. All patients and investigators were masked to treatment assignment. The primary endpoint was a composite of doubling of serum creatinine (sustained for ≥30 days)or end-stage kidney disease (eGFR <15 mL/min per 1·73 m 2 sustained for ≥90 days, chronic dialysis for ≥90 days, kidney transplantation, or death from kidney failure)in the intention-to-treat population of all responders. Safety was assessed in all patients who received at least one dose of their assigned study treatment. The study is registered with ClinicalTrials.gov, number NCT01858532. Findings: Between May 17, 2013, and July 13, 2017, 11 087 patients were screened; 5117 entered the enrichment period, and 4711 completed the enrichment period. Of these, 2648 patients were responders and were randomly assigned to the atrasentan group (n=1325)or placebo group (n=1323). Median follow-up was 2·2 years (IQR 1·4–2·9). 79 (6·0%)of 1325 patients in the atrasentan group and 105 (7·9%)of 1323 in the placebo group had a primary composite renal endpoint event (hazard ratio [HR]0·65 [95% CI 0·49–0·88]; p=0·0047). Fluid retention and anaemia adverse events, which have been previously attributed to endothelin receptor antagonists, were more frequent in the atrasentan group than in the placebo group. Hospital admission for heart failure occurred in 47 (3·5%)of 1325 patients in the atrasentan group and 34 (2·6%)of 1323 patients in the placebo group (HR 1·33 [95% CI 0·85–2·07]; p=0·208). 58 (4·4%)patients in the atrasentan group and 52 (3·9%)in the placebo group died (HR 1·09 [95% CI 0·75–1·59]; p=0·65). Interpretation: Atrasentan reduced the risk of renal events in patients with diabetes and chronic kidney disease who were selected to optimise efficacy and safety. These data support a potential role for selective endothelin receptor antagonists in protecting renal function in patients with type 2 diabetes at high risk of developing end-stage kidney disease. Funding: AbbVie

Larval Performance, Morphological, Behavioural and Elecrophisiological Studies on Tenebrio Molitor L. (Coleoptera: Tenebrionidae)

In questa tesi sono stati condotti studi morfologici, di sviluppo su diverse diete e studi di elettrofisiologia e risposte comportamentali di larve e adulti di Tenebrio molitor. Sono state studiate le strutture sensoriali su palpi mascellari e labiali di larve di T. molitor utilizzando tecniche di microscopia elettronica a scansione. Sulla base della morfologia sono stati classificati nove tipi di sensilli. Un complesso sensoriale è stato osservato sulla punta di entrambi i palpi mascellari e labiali. Le prestazioni di crescita di T. molitor sono state monitorate su quattro diverse diete composte da sottoprodotti dell’industria alimentare: farinaccio e sansa di oliva inclusa in tre diverse percentuali. Le diete con una inclusione fino al 25% di sansa di oliva si sono dimostrate le migliori, con un minor tempo di sviluppo larvale, una riduzione della mortalità e un maggior aumento di peso delle larve. Le risposte comportamentali delle larve di T. molitor a VOCs emessi da diversi substrati alimentari sono state studiate usando biosaggi a quattro e due vie. Il comportamento larvale è stato analizzato utilizzando un software di tracciamento video. I risultati di questo studio hanno dimostrato che le larve di T. molitor sono significativamente attratte dai volatili emessi sia dal farinaccio che da un mix di farinaccio e sansa di oliva (1:1). Le risposte comportamentali degli adulti di T. molitor a VOCs emessi dal farinaccio sono state esaminate in un olfattometro a caduta. Sono state testate cinque dosi di farinaccio, da 1 g a 200 g. Le femmine hanno mostrato un indice di risposta più elevato a dosi più elevate di farinaccio, mentre i maschi erano attratti indipendentemente dalle dosi. Infine, è stato condotto un test preliminare di elettroantennografia (EAG) su 14 diversi VOCs estratti da farinaccio su insetti adulti. In particolare, le maggiori risposte EAG sono state registrate per pentanale, esanale, (E)-2-esanale, (E, E)-2-4-nonadienale, (E, E)-2,4-esadienale.The possibility to use Tenebrio molitor to reduce food wastes were investigated. Different field of the entomological research were used: insect morphology, insect rearing and performances on different diets, characterization of Volatile Organic Compounds (VOCs) emitted from food substrates and electrophysiological and behavioral response of larvae and adult insects to the main VOCs identified. Sensory structures on maxillary and labial palps of T. molitor larvae were investigated using scanning electron microscopy. Based on the morphology nine types of sensilla were classified. A sensory complex were observed on the tip of both maxillary and labial palps. The growth performances of T. molitor larvae were monitored under four different diets composed of organic food by-products: wheatmeal, wheatmeal:olive-pomace 3:1, wheatmeal:olive-pomace 1:1, wheatmeal:olive-pomace 1:3. The first two diets appear favorable, with respect to shortening larval development time, reducing mortality and increasing weight gain. Behavioral responses of T. molitor larvae to VOCs emitted by different food substrates was investigated using four and two-way choice. The larval behavior was analysed using a video tracking software. The results of this study showed that T. molitor larvae were significantly attracted to volatiles emitted both by wheatmeal and by a mixture of wheatmeal and olive pomace (1:1). Behavioral responses of T. molitor adults to VOCs emitted by wheatmeal were also investigated in a two-choice pitfall olfactometer. Five doses, ranging from 1 g to 200 g of wheatmeal were tested. Females showed higher Response Index to higher doses of wheatmeal, meanwhile males were attracted independently to the doses. Lastly, a preliminary electroantennography bioassay (EAG) on 14 different VOCs extracted from wheatmeal on adults is presented. In particular the largest EAG amplitudes were recorded for pentanal, exanal, (E)-2-hexanal, (E,E)-2-4-nonadienal, (E,E)-2,4-hexadienal

Larvae of Tenebrio molitor as a novel food for the future: evaluation of the chemical risk factor.

Due to the rapid increase in world population, the waste of food and resources, and notsustainable food production practices, the use of alternative food sources is currently strongly promoted. In this perspective, insects may represent a valuable alternative to main animal food sources since insects grow and reproduce easily, have a high feed conversion efficiency, and can transform low value of growing substrates into high value food resource [1]. Moreover, insects have a high nutrition potential due to an interesting amino acids composition, an adequate content of minerals, vitamins and polyunsaturated fatty acids. Tenebrio molitor L. (Coleoptera: Tenebrionidae) (yellow mealworm) is particularly efficient in converting the organic substance: it can transform diet substrate with a low nutritional value in a rich protein product, and is characterized by high nutritional value [2]. Different studies highlighted how its larvae are particularly suitable for the human consumption [2], and it is a perfect candidate for rearing with the aim of producing a novel food for the future. Nevertheless, the nutritional value of edible insects is highly variable, and the differences may also depend on their feeding substrates. Moreover, there are some potential safety risks linked with insect consumption: a study on the yellow mealworm larvae, for example, showed that the insects accumulate cadmium and lead in their bodies when they feed on organic matter in soils that contain these metals [3]. In the FoodIN project (Edible insects: new frontiers in food), new feeding substrates were used for rearing T. molitor, in order to investigate the best substrate and rearing conditions and the properties of the insect-based food (i.e. flour and other processed material) so obtained. Five feeding substrates were used for rearing mealworm larvae: 100% organic wheat flour, 100% organic wheatmeal, and organic wheatmeal/organic olive pomace with different percentages: 75/25, 50/50 and 25/75, respectively. In order to assess the chemical risks possibly associated with the insect-based food produced, the determination of Cd, Pb and Hg were carried out on fresh mealworm larvae and their feeding substrates. Cd and Pb were analyzed by Atomic Absorption Spectrophotometry, after microwave digestion of the samples, whereas mercury was determined with a Direct Mercury Analyzer (DMA-1). Preliminary result showed that, in insects, heavy metals concentrations were in the range: Cd, 0.007-0-015 mg/kg ww (wet wheight), Pb, 0.07-0.09 mg/kg ww, Hg, 0.12-0.4 μg/kg ww. Insects reared with 100% organic wheatmeal showed in general higher concentration of heavy metals than insects reared with other feeding substrates. Cd concentrations were lower in insects than in their respective substrates, whereas Pb and Hg concentrations in insects were higher than that of feeding substrates. In agreement with literature data, the results of this study evidenced that heavy metals present in feeding substrates are accumulate in the body of T. molitor. Considering that the legal limit of Cd, Pb for meat, and of Hg for fish food, are 0.05, 0.10 and 0.5 mg/kg (guideline UE n. 1881/2006) respectively, insects showed metals concentrations well below the legal limits. This study demonstrate that mealworm larvae can be considered a safe food from the point of view of Cd, Pb and Hg contamination. This study was financially supported by the Università Politecnica delle Marche, Ancona, Italy, within the project “Edible insects: new frontiers in food-FOODIN”

Addition of Olive Pomace to Feeding Substrate Affects Growth Performance and Nutritional Value of Mealworm (Tenebrio Molitor L.) Larvae

The well-recognized efficiency of Tenebrio molitor larvae to convert low quality organic matter into a nutritionally valuable biomass was exploited to manage solid wastes coming from the olive oil industry, which represent a severe environmental challenge in the Mediterranean area. Three organic pomace-enriched substrates (mixtures middlings/pomace 3:1, 1:1, and 1:3) were assessed, together with 100% organic wheat flour and 100% organic middlings as control feeds. A feeding substrate made up of 25% olive pomace and 75% wheat middlings appeared to be the best compromise between growth performance (larval and pupal weights, survival rate, development time) and nutritional properties of mealworm larvae. In fact, larvae fed the 3:1 feed showed the highest dry matter (DM) yield (38.05%), protein content (47.58% DM), and essential/non-essential amino acids ratio (1.16). Fat content (32.14% DM) and fatty acid composition were not significantly different than those of larvae fed more pomace-enriched feeds

Hermetia illucens in diets for zebrafish (Danio rerio): A study of bacterial diversity by using PCR-DGGE andmetagenomic sequencing

In the present research, bacterial diversity was studied during a 6-month feeding trial utilizing zebrafish (Danio rerio) fed Hermetia illucens reared on different substrates with an emphasis on fish gut bacterial diversity. A polyphasic approach based on viable counting, PCR-DGGE and metagenomic 16S rRNA gene amplicon target sequencing was applied. Two different H. illucens groups were reared on coffee by-products (C) or a mixture of vegetables (S). Viable counts showed a wide variability based on substrate. PCR-DGGE and Illumina sequencing allowed the major and minor bacterial taxa to be detected. Both samples of larvae and their frass reared on the S substrate showed the highest richness and evenness of bacterial communities, whereas zebrafish (ZHC) fed H. illucens reared on substrate C and zebrafish (ZHS) fed H. illucens reared on substrate S had the lowest bacterial richness and evenness. A stimulating effect of bioactive compounds from coffee by-products on the occurrence of Lactobacillaceae and Leuconostoccaceae in H. illucens reared on substrate C has been hypothesized. Zebrafish gut samples originating from the two feeding trials showed complex microbial patterns in which Actinobacteria and Alteromonadales were always detected, irrespective of the diet used. Enterobacteriaceae in fish guts were more abundant in ZHS than in ZHC, thus suggesting an influence of the bioactive compounds (chlorogenic and caffeic acids) in the substrate on Enterobacteriaceae in fish guts. ZHC showed a higher abundance of Clostridia than did ZHS, which was likely explained by stimulating activity on the bacteria in this class by the bioactive compounds contained in H. illucens reared on substrate C. An influence of the microbiota of H. illucens or insect-derived bioactive compounds on the gut microbiota of zebrafish has been suggested. The presence of bacteria consistently associated with zebrafish guts has been found irrespective of the diet, thus attesting to the likely stability of the core fish microbiota