Introduction
Protein hydrolysates are reputed in aquafeeds for their antimicrobial, antioxidant and immunomodulatory properties beyond their nutritional value. Although fish protein hydrolysates have been extensively studied (Siddik et al., 2021), the potential use of other sources of animal and plant protein hydrolysates have not extensively evaluated in fish. Among different sources of raw materials to be used for producing protein hydrolysates, rendering by-products have been reported to have relevant nutritional and functional roles in fish nutrition. In particular, the most common evaluated blood by-product in aquafeeds is the porcine spray-dried plasma, although blood protein hydrolysates may be an untapped safe source of animal protein hydrolysates for aquafeeds. The use of rendering blood by-products is an untapped source of highly quality ingredients for aquafeeds, regardless of the change in legislation that allows the use of animal proteins of porcine and avian origin in aquafeeds.
In the present study, authors aimed to evaluate the effects of porcine protein hydrolysates (PPH) on most common key performance indicators like growth and feed efficiency parameters, as well as on its influence on gilthead seabream immunity. Particularly, the immunomodulatory effects of the dietary administration of PPH to a bacterial challenge were tested at two levels: i) by evaluating gene expression level in splenocytes exposed to a short-term ex vivo stimulation with lipopolysaccharide (LPS) and ii) by measuring the antibacterial activity of skin mucus when incubated with different bacterial strains.
Materials and Methods
A control diet was formulated with low levels of fishmeal (FM) (7% FM) to contain 48% crude protein, 17% crude fat, and 22 MJ kg-1 gross energy. Based on this basal formulation, an experimental diet was formulated, in which porcine protein hydrolysate (PPH) (PEPTEIVA®; APC Europe, SA, Spain) was incorporated at 5% at the expense of FM (final FM levels = 2%). The PPH is a hydrolysate of porcine plasma containing 76% protein (>85% of protein in form of peptides smaller than 10KDa), 2.6% crude fat and 14.5% ash. Both diets were isonitrogenous, isolipidic, and isoenergetic.
Fish were fed two experimental diets (4 replicates per diet) for 92 days under the following environmental conditions: water temperature values ranging from 22 to 27 °C, 6.1 ± 0.2 mg L-1 of dissolved oxygen and natural photoperiod. Feeds were distributed four times per day by automatic feeders (feeding rate of 3.3% of the stocked biomass). At the end of the trial, an ex vivo assay with splenocytes from both groups was performed as described in Salomón et al. (2020), and the bactericidal properties of the skin mucus were assayed against Escherichia coli (DSMZ423), and two pathogenic bacteria, Vibrio anguillarum (CECT522T) and Pseudomonas anguilliseptica (CECT899T) that were co-cultured with skin mucus during a 24 h-period (Sanahuja et al., 2019).
Results and Discussion
Fish fed the PPH diet were 4.6% heavier than their congeners fed the control diet with low levels of FM. No differences in length nor condition factor were found. In addition, FCR values were lower in fish fed the PPH diet (1.08 ± 0.06) in comparison to those of the control group (1.23 ± 0.04). Similar results were observed in gilthead seabream fed diets containing high FM levels (46%) (Gisbert et al., 2015). Results from the in vitro assay with splenocytes of fish from both dietary groups revealed that PPH enhanced the systemic immune response of gilthead seabream as gene expression markers indicated. In particular, we found an up-regulation of gene markers involved in the humoral innate response (igM), as well as pro- (il-1β, tnfα) and anti-inflammatory cytokines (il-10, tgfβ1), whereas no significant changes in gene expression were found in terms of mn-sod, cat and lys. Considering that fish skin mucus provides a stable physical, biological, and chemical barrier against invading pathogens, knowledge of its antibacterial capacity when exposed to a pathogenic organism is of relevance. In this sense, the mucus from gilthead seabream fed the PPH showed a higher bactericidal activity than the control group along the 24 h of skin mucus co-culture with the three bacterial species considered. In this sense, skin mucus inhibited the growth of V. anguillarum and P. anguilliseptica; thus, potentially protecting fish from vibriosis and pastelleurosis when fed diets containing low levels of FM. The immunomodulatory properties of the PPH diet may be attributed to the content of plasma in immunoglobulins, albumins, growth factors and biologically active peptides, which may mediate anti-inflammatory and immunomodulatory effects (Pérez-Bosque et al., 2016).
Conclusions
This study showed that porcine protein hydrolysate obtained from blood plasma is a safe and functional ingredient, especially in those formulated with low fishmeal levels. In particular, PPH promotes somatic growth and improves feed performance, whereas at the same time it enhances the immune response of animals. Considering these beneficial properties, this ingredient can be useful for its incorporation in aquafeeds.
Acknowledgments: This work has been financially supported by the project “Nutritional strategies for the improvement of productive performance: the use of functional feeds and health diets in aquaculture (DIETAplus)”, funded by JACUMAR (Ministry of Agriculture, Fisheries and Environment of Spain, MAPAMA) and FEMP (EU), as well as by the ERC (ERC) by the MedAID project (Grant Agreement Nb. 727315).
References
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