Influence of dietary Chlorella vulgaris and carbohydrate-active enzymes on growth performance, meat quality and lipid composition of broiler chickens.

Herein, we investigated the effect of Chlorella vulgaris as ingredient (10% of incorporation) in broiler diets, supplemented or not with 2 formulations of Carbohydrate-Active enZymes (CAZymes; Rovabio Excel AP and a mixture of recombinant CAZymes, composed by an exo-ß-glucosaminidase, an alginate lyase, a peptidoglycan N-acetylmuramic acid deacetylase and a lysozyme), on growth performance, meat quality, fatty acid composition, oxidative stability, and sensory traits. One hundred twenty 1-day-old Ross 308 male birds were randomly assigned to one of the 4 experimental diets (n = 30): corn-soybean meal-basal diet (control), basal diet with 10% C. vulgaris (CV), CV supplemented with 0.005% of a commercial CAZyme cocktail (Rovabio Excel AP), (CV + R), and CV supplemented with 0.01% of a 4-CAZyme mixture previously selected (CV + M) during the experimental period lasted from day 21 to day 35. Body weight gain and feed conversion rate of broilers were not affected by C. vulgaris but digesta viscosity increased more than 2-fold (P < 0.001) relative to the control. In addition, neither cooking loss, shear force, juiciness, flavor nor off-flavor was impaired by dietary treatments (P > 0.05). By contrast, the dietary C. vulgaris increased tenderness, yellowness (b∗) and total carotenoids in breast and thigh meats. However, no additional protective effect against lipid oxidation was observed in meat with the inclusion of microalga. Chlorella vulgaris, independently of CAZymes, had a minor impact on meat fatty acid composition but improved the proportion of some beneficial fatty acids. In summary, our data indicate a slight improvement of broiler meat quality and lipid nutritional value, without impairment of broilers' growth performance, thus supporting the usefulness of this microalga in poultry diets, up to this high level of incorporation. By contrast, the selected CAZyme mixtures used do not significantly improve the release of microalga nutrients in poultry diets, through the disruption of microalga cell wall, which warrants further research.

Impact of dietary incorporation of Spirulina (Arthrospira platensis) and exogenous enzymes on broiler performance, carcass traits, and meat quality.

This study assessed the effect of Spirulina (Arthrospira platensis), individually and in combination with exogenous enzymes, on growth performance, carcass traits, and meat quality of broiler chickens. One hundred and twenty Ross 308 male chickens were allocated into 40 battery brooders, with 3 birds per cage, and fed ad libitum a corn-based diet during the first 21 D of the trial. The experimental period lasted from day 21 to 35, during which birds were fed 4 different diets: a corn-soybean basal diet, taken as the control group, a basal diet containing 15% Spirulina (MA), a basal diet containing 15% Spirulina plus 0.005% Rovabio Excel AP (MAR), and a basal diet containing 15% Spirulina plus 0.01% lysozyme (MAL). Body weight gain (P < 0.001) and feed conversion rate (P < 0.001) were improved in control chickens, when compared with those fed with Spirulina. In addition, Spirulina increased the length of duodenum plus jejunum in relation to the other treatment (P < 0.01). Chickens on the MAL diet showed a considerable increase in digesta viscosity (P < 0.05) compared with the control group. Breast and thigh meats from chickens fed with Spirulina, with or without the addition of exogenous enzymes, had higher values of yellowness (b*) (P < 0.001), total carotenoids (P < 0.001), and saturated fatty acids (P < 0.001), whereas n-3 polyunsaturated fatty acid (P < 0.01) and α-tocopherol (P < 0.001) decreased, when compared with the control. In conclusion, the incorporation of 15% Spirulina in broiler diets, individually or combined with exogenous enzymes, reduced birds' performance through a higher digesta viscosity, which is likely associated with the gelation of microalga indigestible proteins. In addition, cell wall of Spirulina was successfully broken by the addition of lysozyme, but not by Rovabio Excel AP. Therefore, we anticipate that the combination of lysozyme with an exogenous specific peptidase could improve the digestibility of proteins from this microalga and avoid their detrimental gelation.

Xylo-oligosaccharides display a prebiotic activity when used to supplement wheat or corn-based diets for broilers.

It is now well established that exogenous ß-1,4-xylanases improve the nutritive value of wheat-based diets for poultry. Among other factors, the mechanism of action of exogenous enzymes may involve a microbial route resulting from the generation of prebiotic xylo-oligosaccharides (XOS) in the birds' gastro-intestinal (GI) tract. In a series of three experiments, the effect of XOS on the performance of broilers fed wheat or corn-based diets was investigated. In experiment 1, birds receiving diets supplemented with XOS displayed an increased weight gain (P = 0.08). The capacity of XOS to improve the performance of animals during a longer trial (42 d) was investigated (Experiment 2). The data revealed that diet supplementation with XOS, tested at two incorporation rates (0.1 and 1 g/kg), or with an exogenous ß-1,4-xylanase resulted in an increased nutritive value of the wheat-based diet. An improvement in animal performance was accompanied by a shift in the microbial populations colonizing the upper portions of the GI tract. XOS were also able to improve the performance of broilers fed a corn-based diet, although the effects were not apparent at incorporation rates of 10 g/kg. Together these studies suggest that in some cases the capacity of ß-1,4-xylanases to improve the nutritive value of wheat-based diets is more related to their ability to produce prebiotic XOS than to their ability to degrade arabinoxylans. The extremely low quantities of XOS used in this study also challenge the depiction of a prebiotic being a quantitatively fermented substrate. These data also bring into question the validity of the "cell wall" mechanism, as XOS elicited an effect with clearly no action on endosperm cell wall integrity and yet the performance effects noted were equivalent or superior to the added enzymes.

Variation in levels of non-starch polysaccharides and endogenous endo-1,4-ß-xylanases affects the nutritive value of wheat for poultry.

1. Endo-1,4-ß-xylanase is known to improve the nutritive value of wheat-based diets for poultry by degrading dietary arabinoxylans. However, broilers' response to supplementation of wheat-based diets with exogenous endo-1,4-ß-xylanase is not always observed. 2. In this study, 108 different wheat lots were analysed for levels of extract viscosity as well as for endogenous endo-1,4-ß-xylanase activity, and the impact of these two variables in animal performance was tested. 3. Results revealed that endogenous endo-1,4-ß-xylanase activity and extract viscosity content varied widely among different wheat lots. Thus, a trial was conducted to evaluate the efficacy of exogenous enzyme supplementation in broiler diets using wheats with different levels of extract viscosity and endogenous endo-1,4-ß-xylanase activity. 4. The data revealed that exogenous enzyme supplementation was only effective when the wheat present in the diet had high levels of extract viscosity (14.8 cP) with low endogenous endo-1,4-ß-xylanase activity (347.0 U/kg). Nevertheless, it is apparent that exogenous microbial xylanases reduce digesta extract viscosity and feed conversion ratio independently of the endogenous properties presented by different wheat lots. 5. The data suggest that extract viscosity and/or endogenous endo-1,4-ß-xylanase activity affect the response to enzyme supplementation by poultry fed on wheat-based diets.

Effect of reduced dietary protein and supplementation with a docosahexaenoic acid product on broiler performance and meat quality.

1. Chicken breast meat is a lean meat due to its low content of intramuscular fat (IMF) resulting in an overall lower acceptability by consumers due to a decrease in juiciness, flavour and increased chewiness. Recently, studies performed in pigs suggested the possibility of increasing IMF by decreasing dietary crude protein (CP) content, an effect possibly mediated through an increased lipogenesis. 2. Dietary supplementation with lipids rich in omega 3 long-chain polyunsaturated fatty acids (n-3 LC-PUFA) may modulate an increase in the content of these fatty acids in meat from monogastric animals and, thus, promote the daily intake of n-3 LC-PUFA by humans. 3. LC-PUFA are very susceptible to oxidation, resulting in off-flavours that affect meat quality and consumers' acceptability. 4. This trial was conducted to assess the effect of reducing dietary CP, from 21% to 17%, on chicken's meat IMF content and, simultaneously, to evaluate if a complementary supplementation with a proprietary n-3 LC-PUFA source (DHA Gold™) could improve meat quality. These effects were assessed by measuring productive performance and meat quality, oxidative stability, sensory traits and fatty acid profile. 5. A reduction in CP content of broiler diets, from 21% to 17%, balanced for lysine, improved performance while it was not sufficient to increase IMF content in chicken meat. In contrast, DHA Gold™ supplementation had a positive impact both in broiler productive parameters and in meat fatty acid profile. 6. In addition, incorporation of 7.4% of DHA Gold™ in the diet promoted carcass yield but negatively affected chicken meat acceptability by consumers, due to a decrease of meat oxidative stability. 7. Overall the data suggest that neither a dietary supplementation with DHA Gold™ nor a reduction in CP have a direct positive effect in the levels of IMF present in broiler meat.

Direct supplementation of diet is the most efficient way of enriching broiler meat with n-3 long-chain polyunsaturated fatty acids.

1. Concentrations of beneficial omega-3 long-chain polyunsaturated fatty acids (n-3 LC-PUFAs) in poultry meat can be improved by increasing the concentration of n-3 PUFA in poultry diets. 2. A decrease in flavour quality is, however, usually associated with the dietary supplementation with n-3 PUFA, which is due to the susceptibility of PUFA to oxidation. 3. This experiment was conducted to study the effects of introducing two different n-3 fatty acid sources (extruded linseed and DHA Gold™, a proprietary algal product rich in docosahexaenoic acid), either separately or together, on broiler productive performance, and meat quality, oxidative stability, sensory traits and LC-PUFA profile. 4. Birds given the algal product displayed better productive performances than animals from other groups. 5. The data revealed an improvement in the fatty acid nutritional value of meat from birds receiving the algal product and an inefficient conversion of α-linolenic acid (LNA) into LC-PUFA. 6. Metabolisation of LNA in vivo is not sufficient to improve meat quality in n-3 LC-PUFA and direct supplementation of the diet with n-3 LC-PUFA is a better alternative to modulate an increase in beneficial fatty acids of broiler meat. 7. The overall acceptability of meat was negatively affected by the dietary supplementation with 7.4% of DHA, in contrast to the supplementation with 3.7% of DHA, which showed to be efficient in improving LC-PUFA meat content without affecting its sensory properties.

The thermostable ß-1,3-1,4-glucanase from Clostridium thermocellum improves the nutritive value of highly viscous barley-based diets for broilers.

1. Microbial ß-1,3-1,4-glucanases improve the nutritive value of barley-based diets for poultry by effectively decreasing the degree of polymerisation of the anti-nutritive soluble ß-glucans. Glycoside hydrolases (GHs) acting on recalcitrant polysaccharides display a modular architecture comprising a catalytic domain linked to one or more non-catalytic Carbohydrate-Binding Modules (CBMs). 2. GHs and CBMs have been classified in different families based on primary structure similarity (see CAZy webpage at http://www.cazy.org ). The role of CBMs is to anchor the appended GHs into their target substrates, therefore eliciting the efficient hydrolysis of structural carbohydrates. 3. Here we describe the biochemical properties of the family 16 GH from Clostridium thermocellum, termed CtGlc16A. CtGlc16A is a thermostable enzyme that specifically acts on ß-1,3-1,4-glucans with a remarkable catalytic activity (38000 U/mg protein). 4. CtGlc16A, individually or fused to the family 11 ß-glucan-binding domain of cellulase CtLic26A-Cel5E of C. thermocellum, was used to supplement a highly viscous barley-based diet for broilers. 5. The data showed that birds fed on diets supplemented with the recombinant enzymes displayed an improved performance when compared with birds given diets not supplemented with exogenous enzymes. However, inclusion of the non-catalytic CBMs had no influence on the capacity of CtGlc16A to reduce the anti-nutritive effects of soluble ß-1,3-1,4-glucans. 6. The data suggest that at elevated dosage rates, CBMs might be unable to potentiate the catalytic activity of appended catalytic domains; this effect may only be revealed when feed enzymes are incorporated at low levels.

Levels of endogenous ß-glucanase activity in barley affect the efficacy of exogenous enzymes used to supplement barley-based diets for poultry.

To improve the nutritive value of barley-based diet for broilers, 2 experiments using 2 different barley lots were performed to evaluate the capacity of a mesophilic cellulase when fused to a ß-glucan specific family 11 carbohydrate-binding module. The data revealed that the recombinant ß-glucanase derivatives were not appropriate for feed supplementation because of a lack of stability at acidic pH levels. However, under the same experimental conditions, a commercial enzyme mixture improved the nutritive value of 1 of the cereal lots used. Analysis of the nutritive value of the 2 barleys revealed intrinsic differences in the levels of endogenous ß-glucanase activity. These differences were extensively evident when the studies were expanded to a range of 64 barley lots. Thus, to clarify the effect of endogenous cellulases on the efficacy of exogenous ß-glucanases used to supplement barley-based diets for poultry, 2 barley lots presenting low and high levels of endogenous plant cell wall-degrading enzymes were selected. These lots were used to prepare 2 barley-based diets, which were supplemented with or without a commercial enzyme product and fed to broiler chicks. The data revealed that the exogenous enzymes were effective when the basal diet presented low levels of endogenous ß-glucanases but were unable to improve the nutritive value of the barley lot displaying higher ß-glucanase activity. Thus, these studies suggest that levels of endogenous ß-glucanases may affect the efficacy of exogenous enzymes used to improve the nutritive value of barley-based diets for broilers. The development of a quick ß-glucanase assay that could be applied for cereal-based feeds may help identify those barley-based diets that are more responsive to the action of feed enzymes.

A family 11 carbohydrate-binding module (CBM) improves the efficacy of a recombinant cellulase used to supplement barley-based diets for broilers at lower dosage rates.

1. Exogenous microbial beta-1,3-1,4-glucanases and hemicellulases contribute to improving the nutritive value of cereals rich in soluble non-starch polysaccharides for poultry. 2. In general, plant cell wall hydrolases display a modular structure comprising a catalytic module linked to one or more non-catalytic carbohydrate-binding modules (CBMs). Based on primary structure similarity, CBMs have been classified in 50 different families. CBMs anchor cellulases and hemicellulases into their target substrates, therefore eliciting efficient hydrolysis of recalcitrant polysaccharides. 3. A study was undertaken to investigate the effects of a family 11 beta-glucan-binding domain in the function of recombinant derivatives of cellulase CtLic26A-Cel5E of Clostridium thermocellum that were used to supplement a barley-based diet at lower dosage rates. 4. The results showed that birds fed on diets supplemented with the recombinant CtLic26A-Cel5E modular derivative containing the family 11 CBM or the commercial enzyme mixture Rovabio Excel AP tended to display improved performance when compared to birds fed diets not supplemented with exogenous enzymes. 5. It is suggested that at lower than previously reported enzyme dosage (10 U/kg vs 30 U/kg of basal diet), the beta-glucan-binding domain also elicits the function of the recombinant CtLic26A-Cel5E derivatives. 6. Finally, the data suggest that exogenous enzymes added to barley-based diets act primarily in the proximal section of the gastrointestinal tract.

Role of a family 11 carbohydrate-binding module in the function of a recombinant cellulase used to supplement a barley-based diet for broiler chickens.

1. Cellulases and xylanases display a modular architecture that comprises a catalytic module linked to one or more non-catalytic carbohydrate-binding modules (CBMs). CBMs have been classified into 52 different families, based on primary structure similarity. These non-catalytic modules mediate a prolonged and intimate contact of the enzyme with the target substrate eliciting efficient hydrolysis of the target polysaccharides. 2. A study was undertaken to investigate the importance of a family 11 CBM, displaying high affinities for barley beta-glucans, in the function of recombinant derivatives of cellulase CtLic26A-Cel5E of Clostridium thermocellum used to supplement a barley-based diet for broiler chicken. 3. The results showed that birds fed on diets containing the recombinant CtLic26A-Cel5E modular derivatives or the commercial enzyme mixture Rovabio Excel AP displayed improved performance when compared with birds fed on diets not supplemented with exogenous enzymes. 4. It is suggested that the enzyme dosage used in this study (30 U/kg of basal diet), was probably too high for the efficacy of the family 11 CBM to be noticed. It remains to be established if the targeting effect resulting from the incorporation of CBMs in plant cell wall hydrolases may be effective at lower exogenous enzyme dosages.

Isoleucine and valine supplementation of a low-protein corn-wheat-soybean meal-based diet for piglets: growth performance and nitrogen balance.

The effects of Ile and Val supplementation of a low-CP, corn-wheat-soybean meal-based piglet diet on growth performance, incidence of diarrhea, and N balance were studied using 60 Landrace x Duroc male piglets in a 4-wk experiment. The 60 individually caged piglets were divided into 5 dietary treatments, each consisting of 12 piglets. Diet 1 was a positive control diet (20% CP); diet 2 was a low-CP negative control diet (17% CP); diets 3, 4, and 5 were low-CP diets to which Ile, Val, or the combination of Ile and Val were added, respectively. All diets were supplemented with Lys, Met, Thr, and Trp to provide the required concentrations of these AA according to the 1998 NRC. Average daily gain and ADFI were similar among pigs fed the positive control, Val-added, and the Val plus Ile-added diets. On wk-2 and wk-4, fecal score was greater (softer feces) in piglets fed the 20% CP level compared with the remaining treatments (P < 0.01). Nitrogen intake was decreased (P < 0.0001) in pigs fed diets containing low levels of CP compared with pigs fed the 20% CP diet. Fecal N excretion (g/d) was decreased (P < 0.05) in piglets fed low-CP diets at wk 1 and wk 4 of feeding, and in urine at wk 4 of feeding. Crude protein levels or AA supplementation had no effect on N retention efficiencies. These results indicate that the supplementation of Val alone, or in combination with Ile, to a low-CP piglet diet with adequate levels of Lys, Met, Thr, and Trp is necessary to achieve maximum performance in pigs consuming corn-wheat-soybean meal-based diets.

Crop beta-glucanase activity limits the effectiveness of a recombinant cellulase used to supplement a barley-based feed for free-range broilers.

1. The supplementation of diets rich in soluble polysaccharides with microbial cellulases and hemicellulases decreases digesta viscosity and promotes broiler performance. 2. In contrast, recent experiments suggest that polysaccharidases are ineffective for improving the nutritive value of pasture biomass used by free-range broilers. However, the feasibility of using cellulases and hemicellulases to improve the utilisation of cereal-based feeds by pastured poultry remains to be established. 3. A study was undertaken to investigate the capacity of a recombinant cellulase from Clostridium thermocellum to improve the nutritive value of a barley-based feed for free-range pastured broilers of the RedBro Cou Nu x RedBro M genotype. 4. The results show that supplementation of a barley-based diet with a recombinant beta-glucanase had no effect on the performance of free-range broilers, foraging in legume-based diets from d 28 to 56. In addition, the results confirm that the lack of effect of the recombinant enzyme in improving the nutritive value of the barley-based feed does not result from enzyme proteolysis or inhibition in the gastrointestinal tract. 5. Significantly, beta-glucanase activity was identified in the crop of non-supplemented animals. The data suggest that endogenous cellulases originated both from the barley-based feed and from the crop microflora. 6. The results presented here suggest that in older birds of slow-growing genotypes associated with free-range production systems, previously unknown sources of beta-glucanases, such as the feed and microbial symbiotic microflora, can affect the effectiveness of exogenous enzymes added to the feed.

Relative toxicity of gossypol enantiomers in laying and broiler breeder hens.

Gossypol, a natural component of cottonseed meal, exists in positive (+) or negative (-) enantiomeric forms, and their levels and ratio could be altered by developing new genetic strains of cotton. Two experiments were conducted to determine the relative toxicity of the individual gossypol enantiomers in laying and broiler breeder hens. In the first experiment, 25 individually caged Hy-Line W-36 forty-three-week-old laying hens were fed a standard corn-soy diet supplemented with either no gossypol or the individual enantiomers at 200 and 400 mg/kg of diet for 20 d (5 hens/treatment). In the second experiment, 15 individually caged Cobb 500 fast-feathering 44-wk-old broiler breeder hens were fed a standard corn-soy-wheat middlings diet supplemented with either no gossypol or the individual enantiomers at 400 mg/kg of diet for 18 d (5 hens/treatment). In both experiments, feed intake, egg production, and egg weight were determined daily. All eggs were individually opened and scored for yolk discoloration. At the end of both experiments, several organ and tissue samples were collected for gossypol analyses. In both experiments, the addition of (+)-gossypol to the diet reduced egg production. Only laying and broiler breeder hens fed (+)-gossypol produced eggs with severe yolk discoloration (score > or = 4). Total feed intake was lower (P < 0.05) in laying hens fed the 400 mg/kg level of (+)-gossypol compared with laying hens fed the other dietary treatments. In contrast, broiler breeder hens consumed less of the diet supplemented with (-)-gossypol. In both experiments, tissue accumulation of (+)-gossypol was higher than (-)-gossypol, with the exception of bile and excreta. The results suggest that in hens the ingestion of (+)-gossypol has a greater effect on egg yolk discoloration than the consumption of (-)-gossypol.

Relative toxicity of gossypol enantiomers in broilers.

Use of cottonseed meal in poultry diets has been avoided in large part because of fear of gossypol toxicity. Gossypol exists naturally as a mixture of 2 enantiomers that exhibit different biological activities. Two experiments were conducted to determine the relative toxicity of gossypol enantiomers on broilers. In the first experiment, 3-d-old broilers were fed a standard diet containing 0, 100, 200, 300, or 400 mg of gossypol from gossypol acetic acid per kilogram of diet from 3 to 42 d of age. This form of gossypol contains both enantiomers in an equimolar ratio. Each dietary treatment consisted of 6 replicate pens of 4 birds. In the second experiment, 3-d-old broilers were divided into 15 pens of 4 birds each and fed a standard diet supplemented with either no gossypol or one of the gossypol enantiomers at 200 or 400 mg/kg of diet from 3 to 21 d of age. In both experiments, feed intake and BW gain were measured. In addition, several organ and tissue samples were collected at 21 d (experiments 1 and 2) and 42 d (experiment 1) of age and analyzed for gossypol. In experiment 1, feed consumption and BW gain were reduced (P < 0.05) at 21 and 42 d for the birds fed the highest level of gossypol. The concentration of gossypol in the heart, kidney, and plasma were equivalent at 21 and 42 d of age. In experiment 2, total feed consumption was reduced only in birds consuming (-)-gossypol, but BW gains were lower for birds fed either enantiomer. However, (-)-gossypol was more detrimental to growth than (+)-gossypol. The liver had the highest tissue concentration of both enantiomers, and accumulation of (+)-gossypol was higher than (-)-gossypol in all tissues examined. No racemization of the enantiomers was apparent in the tissues analyzed. Our results indicated that both gossypol enantiomers were toxic to broilers but that (-)-gossypol was more harmful to efficient broiler production than (+)-gossypol.