Trypsin Activity Research Articles

Development of new mutant alleles and markers for KTI1 and KTI3 via CRISPR/Cas9-mediated mutagenesis to reduce trypsin inhibitor content and activity in soybean seeds.

The digestibility of soybean meal can be severely impacted by trypsin inhibitor (TI), one of the most abundant anti-nutritional factors present in soybean seeds. TI can restrain the function of trypsin, a critical enzyme that breaks down proteins in the digestive tract. Soybean accessions with low TI content have been identified. However, it is challenging to breed the low TI trait into elite cultivars due to a lack of molecular markers associated with low TI traits. We identified Kunitz trypsin inhibitor 1 (KTI1, Gm01g095000) and KTI3 (Gm08g341500) as two seed-specific TI genes. Mutant kti1 and kti3 alleles carrying small deletions or insertions within the gene open reading frames were created in the soybean cultivar Glycine max cv. Williams 82 (WM82) using the CRISPR/Cas9-mediated genome editing approach. The KTI content and TI activity both remarkably reduced in kti1/3 mutants compared to the WM82 seeds. There was no significant difference in terms of plant growth or maturity days of kti1/3 transgenic and WM82 plants in greenhouse condition. We further identified a T1 line, #5-26, that carried double homozygous kti1/3 mutant alleles, but not the Cas9 transgene. Based on the sequences of kti1/3 mutant alleles in #5-26, we developed markers to co-select for these mutant alleles by using a gel-electrophoresis-free method. The kti1/3 mutant soybean line and associated selection markers will assist in accelerating the introduction of low TI trait into elite soybean cultivars in the future.

Effects of Annona muricata Extract on Trypsin, Cathepsin B and Collagenase Activities and Textural Changes in Chilled Macrobrachium rosenbergii.

Texture is an important sensory attribute for overall quality and consumer acceptance of prawns. However, texture is affected during cold storage due to the proteolytic activity of endogenous proteases, resulting in poor quality and a short shelf life. The objective of this study is to determine the inhibitory effects of Annona muricata leaves extract (AMLE) (0, 3, 10 and 20%) on the trypsin, cathepsin B and collagenase activities extracted from the cephalothorax of Macrobrachium rosenbergii. In addition, the textural changes in M. rosenbergii during 20 days of cold storage (4 °C) were also determined. M. rosenbergii were soaked in four different treatments: 0, 3, 10 and 20% AMLE and 1.25% sodium metabisulphate for 10 min at 4 °C. Protease activity was significantly (p < 0.05) reduced at 10 and 20% AMLE. Similarly, cathepsin B showed a significant (p < 0.05) low after treatment at 20% AMLE. The maximum inhibitory activity of trypsin was achieved at 20% AMLE and the standard inhibitor (Tosyl-L-lysyl-chloromethane hydrochloride (TLCK)) compared to the control. Whereas, the lowest collagenase activity was obtained at 20% AMLE compared to the control. These inhibitory effects further maintain the firmness of M. rosenbergii coated with 20% AMLE up to the eighth day of storage when compared to the control. Meanwhile, the highest penetration work was found in the M. rosenbergii coated with 20% AMLE at the twentieth day of storage. In conclusion, treatment at 20% AMLE could be used as a natural preservative to inhibit protease, trypsin and collagenase activity of M. rosenbergii and thus can maintain firmness for up to 8 days of storage.

Hybrids with heterosis possess a more diverse intestinal microbiome than that of carnivorous and herbivorous parental fishes in controlled conditions

The intestinal microbiome plays critical, interactive roles in host nutrition and metabolism and may be involved in the known growth advantage of hybrids. In a previous study, we obtained healthy F1 hybrids (PS) by crossing carnivorous Percocypris pingi (PP, male) and herbivorous Schizothorax wangchiachii (SW, female) and found that the PS hybrids exhibited a growth advantage and that their feeding traits were similar to those of omnivorous S. davidi (SD). In this study, we compared the intestinal histology and anatomy of four fish lines (PP, PS, SW, SD) and, using 16S rRNA gene sequencing, constructed a map of their intestinal microbiome to explore host-influenced intestinal microbiome community composition among them under the same diet. We found that the intestinal anatomical and histological traits of PS and SD were similar, and intermediate between PP and SW, but closer to SW. Although PS and SD have similar feeding traits, their intestinal microbiome composition varied remarkably and, notably, the SD microbiome was closer to that of SW, indicating that phylogenetic effects may play a stronger role in gut microbiome development than the effects of feeding traits. Cetobacterium and Aeromonas were the predominant components of the intestinal microbiome in the four fishes, the former was more abundant in PP, and the latter was more abundant in SW and SD. The relative abundances of Cetobacterium and Aeromonas were lower in PS than in the other three fish lines, but the relative abundances of other microorganisms were generally higher in the PS microbiome, which led to more OTUs and higher alpha diversity. In addition, trypsin activity and amylase activity were strongly positively correlated with the relative abundance of Cetobacterium and Aeromonas, respectively. Although the digestive enzyme activities of PS showed hybrid weakness, its intestinal microbial diversity was higher, which may be related to its growth advantage.

Dietary hydrolyzable tannins reduce growth performance and induce histological damage of Chinese seabass (Lateolabrax maculatus)

Chinese seabass (Lateolabrax maculatus) is a popular carnivorous fish with delicious taste. Although the feed value of condensed tannins has been well documented for L. maculatus, information about the effects of hydrolyzable tannins (HTs) on the growth and health of L. maculatus is limited. This study was conducted to assess the effects of dietary HTs on growth performance, body composition, intestinal digestive enzyme activities, serum metabolites, antioxidant and immune response, and intestine and liver morphology of L. maculatus. A total of 640 fish were randomly divided into four groups with four replicates per group and 40 fish per replicate. Four diets were prepared to contain 0 (G0), 1 (G1), 2 (G2), and 4 (G4) g/kg of HTs. Fish were fed to apparent satiation twice a day during the 56-day feeding trial. Results showed that the final body weight, weight gain rate, specific growth rate, and feed intake were linearly decreased (p&amp;lt; 0.001) as dietary HTs increased. All fish had similar (p &amp;gt; 0.05) whole body compositions. Fish fed G2 and G4 had lower (p&amp;lt; 0.05) intestinal trypsin and lipase activities than those fed G0 and G1, whereas G4 had higher (p&amp;lt; 0.05) aspartate aminotransferase and alanine aminotransferase activities than G0. Serum total antioxidant capacity and lysozyme were linearly decreased (p&amp;lt; 0.01), but the malondialdehyde concentration was linearly increased (p&amp;lt; 0.01) as dietary HTs increased. Intestinal villi in G2 and G4 showed increased deformation, and the vacuolation of liver cells began to appear in G1 and was aggravated as dietary HTs increased. This study showed that HTs should be used with caution due to their growth-inhibiting effect, and the dietary HT level for L. maculatus is recommended to be less than 1 g/kg.

The endoplasmic reticulum acetyl-CoA transporter AT-1 plays a role in the protective unfolded protein response in the exocrine pancreas

Pancreatic acinar cells are responsible for the synthesis and secretion of digestive enzymes. In order to sustain high rates of secretory protein production, acinar cells rely on adaptive mechanisms to mitigate endoplasmic reticulum (ER) stress and maintain secretory homeostasis. The adaptive unfolded protein response (UPR), mediated by the IRE1-XBP1s axis, is essential for acinar function and protects against cellular injury. Recent work has demonstrated that the ER acetyl-CoA transporter, AT-1, is necessary for nascent secretory protein acetylation and export out of the ER and is regulated by IRE1-XBP1s. Furthermore, AT-1 is rapidly downregulated during experimental pancreatitis. To investigate the role of AT-1 in exocrine pancreas function, we generated inducible, acinar-specific AT-1 knockout mice (Ela-Cre AT-1-/-). Loss of AT-1 produced a chronic pancreatitis phenotype characterized by inflammation, immune cell infiltration, tissue fibrosis, and aberrant intracellular activation of the proteolytic digestive enzyme trypsin. Despite the histological and biochemical dysfunction observed, Ela-Cre AT-1-/- mice grow normally, have typical lifespans, and do not lose pancreatic mass. Interestingly, XBP1s expression is significantly upregulated in Ela-Cre AT-1-/- and may confer a protective effect with loss of AT-1. Our data suggests a role for AT-1 in the IRE1-XBP1s axis and highlights the previously undescribed role of protein acetylation in exocrine pancreas physiology. Given the numerous potential downstream effects of dysregulated acetyl-CoA transport, this model will advance our understanding of pancreatic function and disease. NIH This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Influence of artificial diets on biological characteristics and digestive enzymes of Coccinella septempunctata L.

In order to improve the rearing of Coccinella septempunctata L. (Coleoptera: Coccinellidae), nutrients such as shrimp, pollen, honey, and lard were added to the basic artificial diet, and the effects of the artificial diet on biological parameters and digestive enzymes were evaluated. The results show that beetles feeding on the supplemented diet exhibited pupation, emergence, fecundity, and hatching rates that were 102.69%, 125.02%, 162.33%, and 119.90% of those supplied with the basic diet, respectively. The addition of shrimp and pollen to the basal diet improved protease, trypsin, chymotrypsin, and aminopeptidase activity in larvae and female adults. The addition of lard improved lipase activity in female adults, and the addition of honey improved invertase activity in adults of both sexes. This study provides guidance for improving the nutritional benefits of ladybug artificial diets.

Effect of dietary protein, lipid and carbohydrate ratio on growth, digestive and antioxidant enzyme activity of prawn Macrobrachium acanthurus postlarvae

Macrobrachium acanthurus is a freshwater prawn with aquaculture potential. However, little is known about its nutrient requirements, particularly regarding protein and lipid-carbohydrate ratio interactions in the diet. To assess the effect of these macronutrients in growth performance and the activity of digestive and antioxidant enzymes, a 2 × 3 factorial design experiment was conducted using two different levels of protein (P35 and P40) and three lipid-carbohydrate ratios (L:C ratio): 2:1 (20 % L and 10 % C), 1:2 (10 % L and 20 %) and 1:6 (5 % L and 30 % C). Each diet was fed to triplicate groups of juveniles with an initial mean weight of 0.075 ± 0.015 g for 60 days. No significant differences in growth performance were found. Although treatment P35 LC 1:2 showed the highest values of weight gain and specific growth performance, survival rate in this diet was lowest among treatments. Interaction between protein and L:C ratio influenced trypsin, chymotrypsin, leucine aminopeptidase and amylase activities, but protein content alone affected the alkaline proteases. Lipase did not differ among treatments. Catalase, glutathione peroxidase, total antioxidant capacity, phenoloxidase and prophenoloxidase activities were also influenced by protein and lipid-carbohydrate interaction; however, super oxide dismutase activity was only affected by dietary protein level. The highest activities of phenoloxidase and prophenoloxidase were observed in prawns fed diet P40 and L:C ratio 1:6 which suggests oxidative stress. These results suggest that 35 % dietary protein and 10 % lipids and 20 % carbohydrates (L:C ratio 1:2) are optimal for growth performance and activity of digestive and antioxidant enzymes. However, measures should be taken to avoid cannibalism and increase the survival rate.

Improving the activity and thermal stability of trypsin by the rational design

The direct evolution combined with the rational design was performed to improve the catalytic properties of the trypsin. ET mutation trypsin included the mutated sites of S40P, G86P, N192Y and Q194L. The optimum of ET mutation trypsin activity was 80.0 ℃. The half life of wild type trypsin and ET mutant trypsin was 420.1 min or 630.1 min respectively. The activity of ET mutation trypsin increased from 3156.95 U/mg to 3951.68 U/mg. Km of ET mutation trypsin was reduced from 26.2 mM to 25.6 mM. Kcat of ET mutation trypsin increased from 3.309/s to 3.312/s. 150–200 amino acids residues of ET mutation trypsin had the higher hydrophobicity. Based on the molecular docking results, the hydrogen bond between the 48th histidine residue and the substrate became shorter after the ET mutation. The evolution increased remarkably the activity and thermal stability of the trypsin.

Increased trypsin resilience in aqueous-acetonitrile environment when immobilized on glyoxyl-agarose may improve its applicability

Although trypsin is a protease naturally applicable in many processes, it is still possible to increase its efficiency and forms of use. Processes that increase their resilience to different conditions in the reaction medium can expand and/or refine their range of applications. Its performance in the presence of organic solvents, such as acetonitrile (ACN), has been indicated as a promising way to increase the efficiency of digestion processes, such as in sample treatment for MALDI-MS peptide mapping. The maintenance of the activity of trypsin immobilized on glyoxyl-agarose was herein demonstrated in different temperature and ACN ranges. Compared to the soluble, the immobilized enzyme was able to remain active above 50% ACN, where the soluble trypsin showed no activity. Although low, it was still possible to detect about 7% activity at 70% ACN when immobilized form was used. With 7.5% of ACN, the soluble enzyme has already shown loss of activity. The same only occurred with immobilized trypsin at 20% or more. Although the immobilized enzyme may have lower specific activity compared to the soluble one, as confirmed by the determined kinetic parameters, its possibility of reuse was confirmed, with no loss of activity for 5 cycles at 0% and 5% ACN. Added to this is the simplicity that the reaction can be interrupted, through the removal of the enzyme from the medium. What could prevent its own protein chain from contaminating the final product. With these characteristics added, trypsin in its immobilized form presents itself as a promising biocatalyst.

Effects of biomacromolecules on growth, digestibility, digestive enzyme activity, antioxidation, and immunity in broilers

This study aimed to evaluate the effects of two biomacromolecules, fermented wheat bran polysaccharides (FWBPs) and sodium humate (SH), on growth performance, nutrient digestibility, digestive enzyme activity, antioxidant status, and immunity of broilers. A total of 144 male, one-day-old Arbor Acres broilers were randomly divided into three dietary groups: control (CON), 0.4% FWBPs, and 0.1% SH, with six replicates of eight birds. The FWBPs and SH groups had a greater body weight (BW) at 21 and 42 d, average daily weight gain (ADG) in the starter period, average daily feed intake (ADFI) in the overall period, and feed-to-gain ratio (F:G) in the grower period. However, the ADFI was decreased by FWBPs supplementation and increased by SH supplementation in the grower period. The dry matter (DM) and organic matter (OM) digestibility were higher in the SH group at 21 d. At 21 d, the FWBPs group had an increased duodenal trypsin and serum glutathione peroxidase (GSH-Px) activity and immunoglobulin M (IgM) concentration, and a decreased liver malondialdehyde (MDA) concentration. The SH group had increased duodenal lipase activity, serum IgM, and interleukin-2 concentrations but decreased liver MDA concentrations. At 42 d, FWBPs and SH supplementation decreased duodenal trypsin and chymotrypsin and serum superoxide dismutase activity. Jejunum chymotrypsin activity and liver MDA content were decreased in the SH group. In conclusion, dietary FWBPs or SH supplementation during the starter period can improve growth performance and nutrient digestibility and enhance antioxidant capacity and immunity of broilers.

Effects of drinking water temperature in winter on growth performance, water consumption, surface temperature, and intestinal development of geese from 21 to 49 days of age

This research aimed to investigate the effects of drinking water temperatures on growth performance, water consumption, surface temperature, organ indices, blood parameters, and intestinal development of geese, and determine the optimal drinking water temperature for 21 to 49-d geese. A total of 192 twenty-one-day male Yuzhou white geese were allocated randomly to 4 groups with 8 replicate pens per group according to the drinking water temperature (drinking water temperature [7℃-12℃] at ambient temperature [TC], 18℃ [T1], 27℃ [T2], and 36℃ [T3], respectively). The results showed that increasing drinking water temperature did not significantly improve the BW, ADG, and ADFI of geese (P > 0.05), whereas drinking warm water of 36℃ for geese had a trend to increase FCR (0.05 < P < 0.1). Geese in group T2 drank the most water per day on average, whereas geese in group TC drank the least (P < 0.001). Geese in groups T1, T2, and T3 had higher eyes temperatures than group TC (P < 0.001). No significant differences were found on the organ indices and blood biochemical parameters (P > 0.05). Geese from group T1 had higher crypt depth and muscularis thickness of duodenum (P < 0.05), and lower ratio of villus height to crypt depth than other groups (P < 0.001). Geese from group T1 had higher activities of trypsin in duodenum and jejuna and amylase in jejuna at 49 d than other groups (P < 0.01). Overall, these data indicate drinking water at 18℃ can increase water consumption and eyes temperature, and improve the activity of digestive enzymes and promote intestinal development. Under our experimental conditions, we recommend that the optimal drinking water temperature for geese from 21 to 49 d of age is 18°C.

Effects of dietary Humulus scandens extract on growth performance, antioxidant responses and intestinal microflora of Yellow River carp (Cyprinus carpio)

In the present study, the effects of Humulus scandens extract on growth performance, antioxidant responses and intestinal microflora of Yellow River carp (Cyprinus carpio) were investigated. 240 carps were assigned to four treatments received diets supplemented with 0 (control), 0.5, 1 and 2 g/kg Humulus scandens extract for 8 weeks. At end of the trial, the final body weight, specific growth rate (SGR) and intestinal body index of the 1.0 g/kg and 2.0 g/kg group were significantly higher than those of the control group (P < 0.05). No significantly differences were observed for food conversation ratio (FCR) or protein efficacy ratio (PER). The SOD (superoxide dismutase) content in gills of the treatment groups was significantly increased compared to the control group (P < 0.05), GPX (glutathione peroxidase) content in gills of 2.0 g/kg group was significantly higher than that of other groups (P < 0.05). Compared with the control group, the contents of SOD, CAT (catalase) and GPX in the liver of the treatment groups were significantly higher (P < 0.05). The gene expression levels of SOD, CAT and GPX in gill and liver of 1.0 g/kg and 2.0 g/kg group were significantly higher than those of control group (P < 0.05), and the gene expression level of HSP70 in gill and liver was decreased compared to the control group. The trypsin activity, fold height, crypt depth and goblet cell number in 1.0 g/kg and 2.0 g/kg group were significantly higher than those in control group and 0.5 g/kg group (P < 0.05). At the phyla level, the abundance of Actinobacteria in 0.5 g/kg group, Verrucomicrobia in 1.0 g/kg group and Fusobacteria in 2.0 g/kg group were significantly increased compare to the control group. At the genus level, compared with the control group, the abundances of Luteolibacter in 1.0 g/kg group, Gemmobacter and Cetobacterium in 0.5 g/kg and 2.0 g/kg groups were significantly higher. In conclusion, the present study demonstrated that 1.0 g/kg-2.0 g/kg level Humulus scandens extract is beneficial to growth performance, antioxidant activity and intestinal health of Yellow River carp.

Trypsin-Based Chemoenzymatic Assay for Detection of Pollutants and Safety Assessment of Food Additives

Chemoenzymatic assay systems are widely used to detect toxicants in various samples, including food and environment specimens. These methods are based on the ability of various types of toxicant to specifically inhibit/activate the functions of individual enzymes or enzyme systems. The present study examines the possibility of using the proteolytic enzyme trypsin as a specific marker to detect protease inhibitors in different samples. The study shows that trypsin activity is not affected by various heavy metals, pesticides, or quinones at levels considerably greater than their maximum allowable concentrations (MACs) in water bodies. At the same time, the IC50 value for the food preservative potassium sorbate (E202) is 15 mg/L, which is substantially lower than its acceptable daily intake (ADI). The quenching of trypsin fluorescence in the presence of potassium sorbate suggests that inhibition could occur due to the binding of the preservative to the enzyme in the region adjacent to the active center. The trypsin was immobilized in starch gel to ensure its stability in the enzyme inhibition based assay. Single-use reagents were prepared as dry starch disks that could be stored over long periods. Their sensitivity to copper (II) chloride, potassium sorbate, and chromium (III) chloride was similar to the sensitivity of the free trypsin.

Elevated aerial temperature modulates digestive enzyme activities in Mytilus californianus.

The marine intertidal mussel Mytilus californianus aggregates to form beds along the Pacific shores of North America. As a sessile organism it must cope with fluctuations in temperature during low-tide aerial exposure, which elevates maintenance costs and negatively affects its overall energy budget. The function of its digestive gland is to release enzymes that break apart ingested polymers for subsequent nutrient absorption. The effects of elevated aerial warming acclimation on the functioning of digestive gland enzymes are not well studied. In this study we asked whether digestive gland carbohydases and proteases could be overstimulated in warm condition to possibly mitigate the costs related to the heat-shock response. We compared mussels acclimated to a+9°C heat-shock during daily low-tide aerial exposure to mussels acclimated to isothermal tidal conditions in a simulated intertidal system. The results showed fairly consistent activities of cellulase, trypsin, and amino-peptidase across tidal variation and between thermal treatments; however, amylase activity was lower in warmed versus cool mussels across low and high-tide. We also observed the expression of heat-shock genes in gill tissue during warm tidal conditions, suggestive that moderate temperatures during aerial exposure can induce a stress response.

Effects of Replacement of Fish Meal with Poultry By-Product Meal (PBM) on Growth Performance, Digestive Enzyme, and Immunity of Giant River Prawn (Macrobrachium rosenbergii)

A 56-day feeding trial was performed to determine the effect of replacing fishmeal with 0, 25%, 50%, 75%, and 100% PBM (FM) on growth performance, digestive enzyme, and immunity of juvenile giant river prawn (Macrobrachium rosenbergii). Diets were fed in quadruplicate (50 prawns per replicate) to satiation (3–5% of their body weight). The finial weight, weight gain, and specific growth rate of prawns were significantly reduced P &lt; 0.05 when 100% PBM was substituted for fish meal, whereas the remaining test diets showed no variation compared to the control P &gt; 0.05 . There was no significant difference in survival rates. The moisture, crude protein, and crude fat of the prawn were unchanged P &gt; 0.05 . The SOD activity, MDA content, and AKP activity of hemolymph did not change significantly. However, the ACP activity of all replacement groups decreased significantly P &lt; 0.05 . The amylase enzyme activity in the intestine of prawn fed 25% of PBM increased significantly compared with the control group, while the trypsin and lipase activities were unchanged. This study showed that PBM up to 75% replacement group does not affect growth performance, and all replacement groups have no adverse effect on intestinal digestive enzyme activity and immunity in juvenile M. rosenbergii. Therefore, poultry by-product meal could replace fish meal by up to 75%.

Antixenosis of Thodan (AAB), a resistant Musa cultivar to Odoiporus longicollis (Olivier) and characterization of larvicide molecules

Banana pseudostem borer (BPB) Odoiporus longicollis (Olivier) (Coleoptera, Curculionidae) is a serious pest of Musa cultivars. Experimental maintenance of larvae in the live pseudostem of cultivar Thodan, a resistant AAB Musa cultivar has resulted antixenosis followed by death of larvae within a week. Antixenosis was characterized by significant decrease of total hemocytes and sharp changes on the proportionate distribution of different types of hemocytes in larvae. As the number of plasmatocytes, prohemocytes, splenocytes and adipohemocytes decreased the number of granulocytes and oenocytes increased. Antixenosis also caused accumulation of 20-hydroxyecdysone (20E) and significant inhibition on the activities of trypsin like serine protease (TlSP) and phenoloxidase (PO). Phytochemical analysis of Thodan resulted characterization of three larvicides such as Betulinic acid (BA), Stigmasterol-3-O-glucoside (SOG) and Sulfoquinovosyl diacyl glycerol (SQDG), and the content in the pseudostem ranged 0.0027 to 0.007 per cent. All the three larvicides were highly toxic to the larvae with LD50 of 0.38 ppm for SOG, 0.41 ppm for SQDG and 0.83 ppm for BA. Simultaneous action of three larvicides in the live pseudostem resulted resistance in Thodan against infestation by BPB. Susceptible Nendran showed negligibly low content of SOG (0.0011%) and SQDG (0.0013%) but no detectable quantity of BA. Intoxication by all the three larvicides caused significant changes on the proportionate distribution of hemocytes, accumulation of 20-hydroxy ecdysone (20E) and inhibition of TlSP and PO, the enzymes involved in larval metamorphosis and cuticle sclerotisation. This study demonstrated that resistance of Thodan against O. longicollis is due to adverse effect of these larvicides on endocrine system, cuticle development and cytotoxicity of hemocytes. As these larvicide molecules are stable compounds, there is scope for them to be used as substitutes in place of deleterious insecticides for the management BPB.

Effects of nonantibiotic growth promoter combinations on growth performance, nutrient utilization, digestive enzymes, intestinal morphology, and cecal microflora of broilers.

Given the ban on antibiotic growth promoters, the effects of nonantibiotic alternative growth promoter combinations (NAGPCs) on the growth performance, nutrient utilization, digestive enzyme activity, intestinal morphology, and cecal microflora of broilers were evaluated. All birds were fed pellets of two basal diets-starter (0-21 d) and grower (22-42 d)-with either enramycin (ENR) or NAGPC supplemented. 1) control + ENR; 2) control diet (CON, basal diet); 3) control + mannose oligosaccharide (MOS) + mannanase (MAN) + sodium butyrate (SB) (MMS); 4) control + MOS + MAN + Bacillus subtilis (BS) (MMB); 5) control + MOS + fruit oligosaccharide (FOS) + SB (MFS); 6) control + FOS + BS (MFB); 7) control + MOS + FOS + MAN (MFM); 8) control + MOS + BS + phytase (PT) (MBP). ENR, MOS, FOS, SB, MAN, PT, and BS were added at 100, 2,000, 9,000, 1,500, 300, 37, and 500 mg/kg, respectively. The experiment used a completely random block design with six replicates per group: 2400 Ross 308 broilers in the starter phase and 768 in the grower phase. All NAGPCs significantly improved body weight gain (P < 0.01), utilization of dry matter, organic matter, and crude protein (P < 0.05), villus height and villus height/crypt depth in the jejunum and ileum (P < 0.01), and decreased the feed conversion ratio (P < 0.01) at d 21 and 42. MMS, MMB, MFB, and MFM duodenum trypsin, lipase, and amylase activities increased significantly (P < 0.05) at d 21 and 42. On d 21 and 42, MMS, MMB, and MBP increased the abundance of Firmicutes and Bacteroides whereas MMB, MFB, and MBP decreased the abundance of Proteobacteria, compared to ENR and CON. Overall, the NAGPCs were found to have some beneficial effects and may be used as effective antibiotic replacements in broilers.

Enzymatic, skeletal, and histological ontogeny of shi drum (Umbrina cirrosa) larvae under intensive culture conditions

This study was conducted to investigate the structural and ontogenetic development of the skeletal system and digestive tract histologically and enzymatically in larvae of shi drum (U. cirrosa) under intensive rearing conditions until 40days after hatching (DAH). Among digestive enzymes, amylase was detected at 0.89 ± 0.12mUmg protein-1 on the first hatching day. The specific activities of trypsin and lipase were detected simultaneously with mouth opening on 3 DAH with 28.47 ± 3.52 and 2.8 ± 0.32mUmg protein-1, respectively. In addition, pepsin was found for the first time at 0.88 ± 0.21mUmg protein-1 on 15 DAH in association with stomach formation and increased sharply up to 40 DAH. In the structural development of the skeletal system, the development of the caudal fin in larvae was morphologically associated with the flexion of the notochord. It was observed that the shape of the fin and spine, which reached 40 DAH, became similar to the adult shape. Histologically, 3 DAH, the mouth and anus were opened. The formation of the primitive stomach was observed at the end of the seventh day-the pyloric sphincter formed between 13 and 18days. A functional stomach was seen on the 15th DAH. Therefore, U. cirrosa is believed to have critical aquaculture potential that can be cultured under intensive conditions. The developmental profile of skeletal, enzymatic, and histological ontogeny observed in U. cirrosa is similar to that described for other sciaenid species.

The stereoselective toxicity of dinotefuran to Daphnia magna: A systematic assessment from reproduction, behavior, oxidative stress and digestive function

Dinotefuran is a promising neonicotinoid insecticide with chiral structure. In the present study, the stereoselective toxicity of dinotefuran to Daphnia magna (D. magna) was studied. The present result showed that S-dinotefuran inhibited the reproduction of D. magna at 5.0 mg/L. However, both R-dinotefuran and S-dinotefuran had no genotoxicity to D. magna. Additionally, neither R-dinotefuran nor S-dinotefuran had negative influences on the motor behavior of D. magna. However, S-dinotefuran inhibited the feeding behavior of D. magna at 5.0 mg/L. Both R-dinotefuran and S-dinotefuran induced oxidative stress effect in D. magna after exposure. R-dinotefuran significantly activated the activities of superoxide dismutase (SOD) and glutathione S-transferase (GST), while S-dinotefuran showed the opposite effect. S-dinotefuran had more obvious activation effect on the acetylcholinesterase (AchE) activity and trypsin activity compared to R-dinotefuran. The transcriptome sequencing results showed that S-dinotefuran induced more DEGs in D. magna, and affected the normal function of ribosome. The DEGs were mainly related to the synthesis and metabolism of biomacromolecules, indicating the binding mode between dinotefuran enantiomer and biomacromolecules were different. Additionally, the present result indicated that the digestive enzyme activity and digestive gene expression levels in D. magna were greatly enhanced to cope with the inhibition of S-dinotefuran on the feeding.

Small Intestinal Digestive Functions and Feed Efficiency Differ in Different Pig Breeds

Small intestinal growth and health affect its digestion and absorption ability, while little information exists about the small intestinal morphology and function differences among the different pig breeds. Therefore, 90 healthy 35 days of age Taoyuan black (TB), Xiangcun black (XB), and Duroc (DR) pigs (30 pigs per breed) with similar body weight (BW) of the same breed were reared to 185 days of age to evaluate the potential relationship between feed efficiency and small intestinal morphology and function at 80, 125, and 185 days of age. The results show that the TB and XB pigs had lower initial and final BW, ADG, and ADFI and plasma CHO and LDL-C levels, whereas they had higher plasma LIP levels and jejunal trypsin, invertase, lactase, and maltase activities and higher DM, ADF, Tyr, Arg, and His digestibility at 80 days of age compared with the DR pigs. At 125 days of age, TB and XB pigs had lower apparent total tract digestibility and plasma CHO, HDL-C, LDL-C, and NH3 levels; XB pigs had lower DM and NDF digestibility, and TB pigs had higher jejunal lactase and maltase activities. At 185 days of age, TB and XB pigs had lower DM, EE, ADF, and GE digestibility, while having higher plasma ALT and UN levels; TB pigs had higher plasma AST level and jejunal chymase activity. Furthermore, the plasma free amino acid contents, small intestinal VH, and nutrient transporter expression levels differed at different ages. Therefore, the different pig breeds exhibited significantly different growth performance and small intestinal growth, mainly resulting from the differences in digestive enzymes and nutrient transporters in the small intestine.