Development of a point of care (POC) test as an immunobiosensor for okadaic acid detection in mussels

The increase in frequency and intensity of potential risks concerning the exposure of seafood to diarrhetic shellfish poisoning (DSP) toxins, even in areas not traditionally accomplished, raises the number of monitoring analyses for the detection of this class of toxins to guarantee the food safety of the products. In addition to the analysis performed with liquid chromatography-tandem mass spectrometry (LC-MS/MS), reference method in EU, semiquantitative rapid tests for preliminary screening and rapid monitoring plan are applied. Different kits for the detection of DTX family toxins are available on the market, but they require specialised labs and the cost of the analysis per sample is quite high. Based on the current situation, developing an easy-touse and cheap test for measuring okadaic acid, as the main responsible for DSP poisoning, for the implementation of point-of-care (POC) systems is desirable. An immunobiosensor test, based on immunoaffinity reaction utilizing commercially available monoclonal antibodies, has been optimized and validated for the detection of okadaic acid in mussel (Mytilus galloprovincialis) extracts, with a limit of detection of 60 mu g/kg, a sensitivity of 0.68 n degrees of counts/ ug/Kg and a screening test range between of 60-350 ug/kg. The proposed POC immunoassay provides results comparable (r = 0.981) to the ones obtained by other semiquantitative rapide tests, like enzymatic assay applied for routine monitoring plans for the detection of the dinophysistoxins (DTXs) family. The goal of the presented test is the reduction of steps for the toxin extraction and a relevant reduction of the time for the analysis and the provision of a cheap POC analysis system

Chitin and its effects when included in aquafeed

Chitin, the second most abundant polysaccharide in nature after cellulose, has been the sub- ject of increasing interest in recent years, particularly in the aquaculture sector. This review focuses on the effects of chitin in aquafeed on growth, nutrient utilization, gut microbiome modulation, and the immune system of aquatic organisms. Data from the literature showed significant variability in response to chitin by species, with some species benefiting from dietary chitin supplementation in terms of growth and immune health. However, excessive chitin inclusion led to negative effects on the protein and lipid digestibility. These results suggest that optimizing the amount of chitin-containing ingredients in aquafeed requires a deep understanding of each species’ nutritional needs and other studies needed for differ- ent aquaculture species. Furthermore, studies have highlighted the potential of chitin as an immunostimulant and promoter of gut health. In conclusion, chitin appears to be a promis- ing additive for sustainable aquaculture, but further research is needed to define best prac- tices for its use

Effect of dietary chitin on growth performance, nutrient utilization, and metabolic response in rainbow trout (Oncorhynchus mykiss)

The dietary inclusion of ingredients containing chitin has been claimed to be beneficial to fish health. However, controversial results on growth performance and nutrient digestibility have been reported in literature. The present study aimed at assessing the response of rainbow trout (Oncorhynchus mykiss) fed with increasing levels of chitin (0, 1.5, 3.0, and 4.5 %) in a semi-purified basal diet. Protein and lipid digestibility was assessed and after 10 weeks of feeding, 6 h after the last meal, fish were euthanized for the evaluation of growth performance, blood biochemistry, brush border membrane and chitinolytic enzyme activity, gene expression of enzymes and nutrient transporters as well as the major pro-inflammatory intestinal cytokines, and mid intestine microbiota. Dietary chitin inclusion of up to 3 % did not hamper growth performances, nutrient apparent digestibility, and levels of most blood enzymes and metabolites. Diet including 4.5 % chitin negatively affected growth performances and nutrient digestibility. The application of a multidisciplinary approach highlighted that the biomarkers related to gut digestion functionality and inflammation response were altered also in fish fed the diets including 1.5 and 3 % of chitin, while gut microbiota did not differ between dietary treatments, even if some taxa suggested to be important in fish physiology were not recovered. Overall, the results demonstrate a good tolerance of rainbow trout to diets including chitin up to 3 %

Postprandial kinetics of digestive function in rainbow trout (Oncorhynchus mykiss ): genes expression, enzymatic activity and blood biochemistry as a practical tool for nutritional studies

Postprandial kinetics of genes expression of gastric (chitinase, pepsinogen) and intestinal (alkaline phosphatase, maltase) digestive enzymes and nutrient transporters (peptide transporter 1, sodium-glucose transporter 1), Brush Border Membrane (BBM) enzymes activity (alkaline phosphatase, leucine aminopeptidase, maltase, saccharase) and blood biochemistry (triglycerides, cholesterol, protein, albumin, glucose, amino acids) through NMR spectroscopy, were investigated in rainbow trout (Oncorhynchus mykiss) fed a commercial aquafeed. For this purpose, fish were starved 72 h and digestive tract and blood were sampled before the meal and at 1.5, 3, 6, 9, 12, and 24 h after feeding (T0, T1.5, T3, T6, T9, T12 and T24). The postprandial kinetic showed that the expression of the genes involved in digestion and nutrient transport, the activity of BBM enzymes, and the presence of metabolites in blood were stimulated in different ways by the presence of feed in the digestive tract. The expression of most genes peaked 3 h after meal except gastric pepsinogen and maltase in distal intestine that peaked at T9 and T12, respectively. The activity of BBM enzymes were stimulated differently based on the intestine tract. The plasma proteins level increased from T1.5 until T9, while the other blood parameters unvariated during the postprandial period. This study supplied useful information about the physiological effects a single meal as a potential tool for planning nutritional studies involving the digestive functions

Chemical composition and apparent digestibility of a panel of dried microalgae and cyanobacteria biomasses in rainbow trout (Oncorhynchus mykiss)

Despite a growing interest in microalgae and cyanobacteria as potential sources of nutrients in aquafeeds, little information is presently available on their nutritive value for carnivorous fish species. The aim of this study was to evaluate chemical composition and nutrient digestibility of a panel of microalgae and cyanobacteria dried biomasses (MACB), using rainbow trout (Oncorhynchus mykiss W.) as a fish model. Nine test diets were obtained by mixing 80 parts of a reference diet, added with 20 g/kg of acid insoluble ash as an indigestible marker, to 20 parts of each of the following dried whole-cell biomass: Arthrospira platensis, Nostoc sphaeroides, two strains of Chlorella sorokiniana, Nannochloropsis oceanica, Tisochrysis lutea, Phaeodactylum tricornutum, Porphyridium purpureum and Tetraselmis suecica. The digestibility measurements were conducted with rainbow trout (52.4 \ub1 1.5 g) kept in six tank units each including three 60-L vessels singularly stocked with 12 fish and fitted with a settling column for faecal recovery. Per each diet, faeces were collected over three independent 10-day periods. Apparent digestibility coefficients (ADCs) of dry matter, crude protein (CP), organic matter and gross energy (GE) of single MACB were calculated by difference relative to those of the reference diet. The MACBs had heterogeneous chemical composition (CP, from 20 to 69%; Lipid, 5\u201327%; GE, 12.5-\u201322.6 MJ/kg dry matter basis) reflecting their overall biodiversity. Most of them can be considered as virtually good sources of minerals and trace elements and exhibit an essential amino acid profile comparable or even better than that of soybean meal commonly used in fish feeds with P. purpureum showing the best protein profile. The digestibility results obtained with rainbow trout allowed ranking the MACBs into two major groups. A first one, including C. sorokiniana, N. oceanica and T. suecica, resulted in markedly lower (P < 0.05) crude protein and energy ADC (64\u201373%; 51\u201359%, respectively) compared to a second group including P. purpureum, T. lutea and cyanobacteria (CP-ADC, 83\u201388%; GE-ADC, 74\u201390%) while P. tricornutum resulted in intermediate values. Overall, the present study confirms the consistently reported role of cell-wall structure/composition in affecting accessibility of nutrients to digestive enzyme. Based on the overall outcomes, only T. lutea and cyanobacteria actually meet the requirements for being used as protein sources in aquafeeds provided their mass production becomes more feasible and costeffective, hence attractive for the feed-mill industry in the near future

Growth and Welfare Status of Giant Freshwater Prawn (Macrobrachium rosenbergii) Post-Larvae Reared in Aquaponic Systems and Fed Diets including Enriched Black Soldier Fly (Hermetia illucens) Prepupae Meal

Due to the limited application of insect meal in giant freshwater prawn (Macrobrachium rosenbergii) culture, the present study aimed to (i) produce spirulina-enriched full-fat black soldier fly (Hermetia illucens) prepupae meal (HM) and (ii) test, for the first time, two experimental diets characterized by 3% or 20% of fish meal and fish oil replacement with full-fat HM (HM3 and HM20, respectively) on M. rosenbergii post-larvae during a 60-day feeding trial conducted in aquaponic systems. The experimental diets did not negatively affect survival rates or growth. The use of spirulina-enriched HM resulted in a progressive increase in α-tocopherol and carotenoids in HM3 and HM20 diets that possibly played a crucial role in preserving prawn muscle-quality traits. The massive presence of lipid droplets in R cells in all the experimental groups reflected a proper nutrient provision and evidenced the necessity to store energy for molting. The increased number of B cells in the HM3 and HM20 groups could be related to the different compositions of the lipid fraction among the experimental diets instead of a nutrient absorption impairment caused by chitin. Finally, the expression of the immune response and stress markers confirmed that the experimental diets did not affect the welfare status of M. rosenbergii post-larvae

Exploring the biochemical, technical and applicative characteristics of pullulan produced by different strains of Aureobasidium pullulans.

Pullulan is a polysaccharide with multiple beneficial properties that makes it perfect for several applications and for the food industry. The polysaccharide is produced by Aureobasidium pullans, a black yeast also known for its antagonistic properties against plant fungal pathogens. Ten strains, isolated from different environments, were evaluated for their pullulan producing ability. Three strains were selected as main producers (AP1, UOR18, M13). The pgm1 and ugp genes expression was evaluated, showing a significant difference between the strains. Pullulan produced by the strains was biochemically characterized by FT-IR (Fourier Transform Infrared Spec- troscopy), DSC (Differential Scanning Calorimetry) and NMR (Nuclear Magnetic Resonance) analyses. By FT-IR and DSC analysis, the AP1 pullulan displayed to be more capable to entrap moisture in its structure, and by NMR, it showed to be more similar to the commercial pullulan. The biopolymer was formulated as apple coating, used to control Colletotrichum acutatum by in vitro and in vivo assays. The coating was activated by yeast cells that enhanced the antifungal activity of the treatment

Effect of Amaranth addition on the nutritional composition and consumer acceptability of extruded provitamin A-biofortified maize snacks

The objective of this study was to determine the effect of adding Amaranth leaf powder on the nutrient content and consumer acceptability of extruded provitamin A-biofortified (PVA) maize snacks. Flours of four varieties of PVA maize were composited with Amaranth leaf powder at 0, 1 and 3% (w/w) substitution of, respectively, and extruded into snacks. The ash content of the snacks increased from 0.53 g/100 g-0.58 g/100 g to 0.650 g/100g-89 g/100 g and protein content increased from 9.12 g/100 g-10.94 g/100 g when Amaranth was increased from 0% to 3%. Similarly, lysine content increased from 0.10 g/100 g to 0.17 g/100 g, whilst methionine increased from 0.14 g/100 g to 0.19 g/100 g. The provitamin A content of the snacks ranged from 1.29 µg/g to 1.40 µg/g at 0% Amaranth and 1.54 µg/g to 1.78 µg/g at 3% Amaranth. The acceptability of the snacks decreased with increasing Amaranth concentration, only a very small proportion (2-8%) of the panel liked the snacks extremely. PVA maize with added Amaranth leaf powder has a potential for use in nutritious and healthy extruded snacks, but the consumer acceptability of the snacks should be improved

A preliminary study on the degradation of AFB1 by Tenebrio molitor, Rhizopus oryzae and Trichoderma reesei

Recently, genus Aspergillus, a fungus capable of producing aflatoxins, secondary highly toxic metabolites, has spread to new areas. These areas became suitable habitats due to the recent climate changes. The use of aflatoxin-contaminated crops is a cause of great concern in guaranteeing food safety and is responsible for major economic losses along the supply chain. For this reason, several strategies have been investigated to utilize these contaminated products as a possible food or feed resource by reducing or eliminating their aflatoxin content, but with limited relevant success. The presented study was aimed to evaluate a combination of biological processes to use aflatoxin B1 contaminated crops for their reintroduction into the production chain. The high tolerance to AFB1 and the apparent lack of accumulation in yellow mealworm larvae, reared on wheat bran substrates, spiked with increasing AFB1 concentration (0, 125, 250, 500 μg/kg) to obtain proteins of high biological value. Subsequently, the aflatoxin-degrading capacity of Rhizopus oryzae and Trichoderma reesei was applied to insect breeding waste (frass) in a fermentation process to ensure further utilization of biohazardous frass as soil conditioner. Individually, each process proven to be able to reduce the AFB1 present by about 80%, while the combination of the two approaches ensured the total degradation of aflatoxin B1-contaminated substrate and frass, which resulted in the possible production of biomass, that could be used for the feed and agricultural industry