Valorization of pretreated biogas digestate with black soldier fly (Hermetia illucens, L; Diptera: Stratiomyidae) larvae.

Increasing concerns related to the negative environmental impacts of food waste havemotivated the development of new solutions to complete the waste cycle of organic residues. One particular "waste" product, the solid digestate from anaerobic digestion, has been identified for further bioprocessing. Black soldier fly (BSF, Hermetia illucens) larvae are known for their great potential in the processing of organic waste. In this study, this potential was investigated to further process the digestate waste stream. Digestate is considered a low potential source of nutrients for larvae due to the presence of different fiber fractions. However, the lignocellulosic matter in this residue could be enzymatically hydrolyzed to release residual carbohydrates. For this study, digestate from a full-scale anaerobic digestion plant in Quebec (Canada) which processes a range of feedstocks (fruits, vegetables, garden wastes, sludge derived from dairy processing and wastewater treatment) was sourced. Digestate was treated with Accelerase® DUET enzyme complex to hydrolyze lignocellulosic matter and compared to a standard diet. For each treatment, 600 four-day old larvae were fed daily with 160 g (70% relative humidity) of diets for 6 days and harvested 3 days later. Although their growth and total biomass were significantly lower than the standard diet, larvae fed on hydrolyzed digestate were almost two times larger than the larvae fed on crude digestate. Furthermore, the content of organic matter, lipids and minerals in the diets and frass were analyzed. Finally, the feasibility of applying BSF treatment for digestate valorization is discussed. According to this study, enzyme-treated digestate does not allow efficient larval growth compared to the standard diet. The development of a more effective method of pretreatment is required for BSF larvae to become an eco-friendly solution for digestate valorization.

Establishment of a comprehensive reference transcriptome for vertebral bone tissue to study the impacts of nutritional phosphorus deficiency in rainbow trout (Oncorhynchus mykiss, Walbaum).

Reducing dietary phosphorus (P) is a common approach to reduce effluent P outputs. The potential resulting P-deficiency is known to negatively impact fish bone condition and might result in vertebral deformities. To date, no large-scale study involving deep sequencing of the bone transcriptome has been conducted in salmonids and vertebral molecular changes remain poorly described. This study aims to provide the first comprehensive vertebral transcriptome for rainbow trout (Oncorhynchus mykiss) to allow functional and quantitative expression studies. Fish weighing 60.8±1.6g, were fed for 27weeks using two practical diets having 0.29% (deficient) and 0.45% (sufficient) available phosphorus (P), respectively. Deep sequencing was conducted using HiSeq2000 Illumina 100 paired-end technology from pooled P-deficient and P-sufficient fish and individuals displaying vertebral deformities. Over 140 million trimmed paired-end reads were assembled de novo with Trinity and resulted in 679,869 transcripts with a mean length of 542.5bp. From these sequences, 340,747 matched with referenced ESTs from rainbow trout. Furthermore, 141,909 and 117,564 sequences were functionally annotated against Nr and Uniprot databases, respectively. Interestingly, we observed putative homologue sequences for most of the key components involved in bone formation and turnover in mammals.

Effect of algal incorporation on growth, survival and carcass composition of rainbow trout (Oncorhynchus mykiss) fry.

A nutritional assay has been conducted with rainbow trout fry (Oncorhynchus mykiss) using a biomass of photosynthetic micro-organisms. The algal biomass was incorporated in the feed at increasing levels (12.5%, 25% and 50%) in order to verify the effects on survival (%), growth (length and mass) and carcass quality of the fish (skin color, water, protein and lipid content). Two commercial feeds (CF, as sold, and 0% A, reprocessed without algae) have been used as controls. After eight weeks at 10 degrees C, the growth has been significantly reduced in fish fed the feeds containing 25% and 50% algae diets (P<0.01). Moreover, an incorporation of algae higher than 12.5% in the feed led to a increase of water content (CF versus other diets; P<0.01) and TL (CF versus 50% A; P<0.01) (wet basis) content of carcasses. Survival was 100% in all treatment groups. Under the experimental conditions used, the results obtained show that a maximum of 12.5% of algal biomass can be incorporated in the feed for rainbow trout fry (O. mykiss) without negative consequences on growth and body content in lipids and energy of fish.

Factors affecting protein release from alginate-chitosan coacervate microcapsules during production and gastric/intestinal simulation.

A series of experiments was performed to evaluate the influence of a number of physico-chemical factors on the diffusion of a model protein, bovine serum albumin (BSA), from dried chitosan-coated alginate microcapsules. Diffusion of BSA was quantified during the microcapsule manufacture processes (gelation, washing, rinsing) and during incubation in conditions simulating the pH encountered during the gastric (0.1 N HCl; pH 1.5) and intestinal (200 mM Tris-HCl; pH 7.5) phases of digestion. Factors tested included alginate and chitosan concentration, calcium chloride (CaCl2) concentration in the gelation medium, loading rate, chitosan molecular mass and pH of the gelation medium. Microcapsule size and gelation time were altered in order to determine their effects on protein retention. Alginate and chitosan concentration significantly influenced BSA retention during microcapsule manufacture and acid incubation, as did calcium chloride concentration in the gelation medium (P<0.05). BSA retention during manufacture was not significantly altered by protein loading rate or pH of the encapsulation medium, however, protein retention during acid incubation decreased significantly with increasing protein loading rate and encapsulation medium pH (P<0.05). Microcapsules that were washed with acetone following manufacture demonstrated significantly increased protein retention during acid incubation (P<0.05). In microcapsules that had been acetone-dried to a point whereby their mass was reduced to 10% of that immediately following encapsulation, protein retention was over 80% following 24-h acid incubation vs. only 20% protein retention from non acetone-dried microcapsules. The presence of calcium in the neutral buffer medium significantly reduced BSA diffusion in a concentration-dependent manner (P<0.05).

Evaluation of protein release from chitosan-alginate microcapsules produced using external or internal gelation.

A series of experiments was undertaken to evaluate the diffusion of a model protein, i.e. bovine serum albumin (BSA), from chitosan-alginate microcapsules produced using either internal or external gelation. Diffusion of BSA was quantified during the microcapsule manufacture processes (gelation, washing, rinsing) and during incubation in conditions simulating the pH encountered during the gastric and intestinal phases of digestion. Encapsulation of an acid phosphmonoesterase permitted in situ protein localization, providing evidence to explain results obtained with BSA. There was significantly greater protein loss from internally versus externally-gelled chitosan-alginate microcapsules during the manufacture process (37.6% versus 4.7%, respectively). Similar trends were observed during 24 h incubation in 0.1 N hydrochloric acid. Increasing alginate concentration from 2-4% (w:v) did not significantly reduce losses from internally-gelled microcapsules. Addition of 0.25 M NaCl to the gelling medium significantly increased protein diffusing during microcapsule manufacture and acid incubation from externally gelled microcapsules. In situ protein localization revealed a higher level of protein near the surface of the microcapsules of externally gelled microcapsules versus internal gelation. The above data indicate that externally-gelled microcapsules are inhomogeneous with a higher concentration of alginate near the microcapsule surface, thus reducing the porosity of the resulting microcapsules. These results suggest that the porous nature of internally-gelled chitosan-alginate microcapsules may result in low encapsulation efficiency, depending on the nature of the product being encapsulated.

Characterization of the inappropriate gonadotropin secretion in polycystic ovary syndrome.

To evaluate gonadotropin release in polycystic ovary syndrome (PCO), one or more of the following hypothalamic-pituitary function tests were performed on 24 patients with the syndrome. These tests included (a) the pulsatile pattern and day-to-day fluctuation of gonadotropin release; (b) effects of exogenous estrogen and antiestrogen (clomiphene) administration on gonadotropin release; and (c) pituitary responsiveness to maximal (150 mug) and submaximal (10 mug) luteinizing hormone-releasing factor (LRF) injections. In 10 of the 14 patients sampled frequently (15 min) for 6 h, luteinizing hormone (LH) levels were elevated above the concentration seen in normal cycling women (except the LH surge). These high LH concentrations appeared to be maintained by and temporally related to the presence of exaggerated pulsatile LH release, either in the form of enhanced amplitude or increased frequency. In all subjects, levels of follicle-stimulating hormone (FSH) were low or low normal, and a pulsatile pattern was not discernible. In four patients, daily sampling revealed marked day-to-day fluctuation of LH but not FSH. That the elevated LH levels were not related to a defect in the negative-feedback effect of estrogen was suggested by the appropriate fall of LH in four patients given an acute intravenous infusion of 17beta-estradiol. This infusion had no effect on FSH levels. In addition, clomiphene elicited rises of both LH and FSH that were comparable to the ones observed in normal women given the same treatment. The clomiphene study also suggested that the positive-feed-back mechanism of estrogen on LH release was intact when the preovulatory rises of 17beta-estradiol induced appropriate LH surges. The elevated LH levels appeared to be related to a heightened pituitary responsiveness to the LRF. This was found in the 11 and 2 patients given maximal (150 mug) and submaximal (10 mug) doses of LRF, respectively. The augmented pituitary sensitivity for LH release correlated with the basal levels of both estrone (P less than 0.025) and 17beta-estradiol (P less than 0.02). The net increase in FSH was significantly greater (P less than 0.001) in the PCO patients than the normal women with maximal doses of LRF. With the smaller dose study none of the injections had a discernible effect on FSH concentrations in either subject. The disparity between LH and FSH secretion could be explained by the preferential inhibitory action of estrogen on FSH release, coupled with a relative insensitivity of FSH release. These data indicate that in these PCO patients the abnormalities of the hypothalamic-pituitary regulation of gonadotropin secretion was not an inherent defect but represented a functional derangement consequent to inappropriate estrogen feedback, which led to a vicious cycle of chronic anovulation and inappropriate gonadotropin secretion.

Effects of exogenous estrogen and progestin on pituitary responsiveness to synthetic luteinizing hormone-releasing factor.

The effect of estrogen and progestin on pituitary responsiveness to 150 mug synthetic luteinizing hormone-releasing factor (LRF) was assessed in premenopausal women receiving sequential (n=12) and combination (n=7) contraceptive steroids. A marked contrast in the time-course and maximal response to LRF was found; a prompt but quantitatively smaller luteinizing hormone (LH) response was seen during cyclic combination therapy, while a delayed (five times) but enhanced (fivefold) LH response was observed during estrogen segments of cyclic sequential therapy. For follicle-stimulating hormone (FSH), the maximum rise was also higher, and the peak response was similarly delayed in the latter group. The quantitative secretion in response to LRF for LH (area under the curve), but not for FSH, was significantly greater (P < 0.01) in subjects receiving sequential, as compared to subjects receiving combination treatment. In both groups, characteristic gonadotropin responses to LRF were reproducible and were independent of the duration of treatment. Since LRF studies were performed during the estrogen segment of treatment cycle in subjects receiving sequential steroids, our data suggest that estrogen exerts a direct feedback action at the pituitary level and that pituitary responsiveness to LRF is augmented by estrogen.

Pituitary gonadotrophin responsiveness to synthetic LRF in subjects with normal and abnormal hypothalamic-pituitary-gonadal axis.

PIP: Pituitary gonadotropin responsiveness to synthetic LRF was studied in normal males, in normal females at different phases of the menstrual cycle, in premenopausal females treated with synthetic estrogen, and in subjects with various abnormalities of the hypothalamic-pituitary-gonadal axis. Iv injection of from 1 to 450 mcg LRF in normal males resulted in an increase of luteinizing hormone (LH) within 2 minutes, with a maximal concentration of LH at a median time of 25 minutes. Maximal follicle stimulating hormone (FSH) levels occurred at a median time of 45 minutes. LH, and to a lesser extent FSH, increased with dose, although a wide variation in quantitative response to the same dose of LRF was seen among patients. In female subjects, the response to LRF varied during different phases of the menstrual cycle. The most sensitive period for pituitary response to LRF for both LH and FSH was just prior to the midcycle surge. Chronic estrogen treatment of premenopausal women enhanced pituitary responsiveness to LRF, with an alteration in both time and magnitude of response. Altered pituitary responsiveness was seen in patients with gonadal dysgenesis, hypogonadotropism, hypogonadotropic hypothalamic amenorrhea, panhypopituitarism, and pituitary tumor.^ieng