Dietary inclusion of Clostridium butyricum cultures alleviated impacts of high-carbohydrate diets in largemouth bass (Micropterus salmoides).

A 60-d feeding trial was conducted to explore the potential regulatory effects of dietary Clostridium butyricum cultures (CBC) supplementation in high-carbohydrate diet (HCD) on carbohydrate utilisation, antioxidant capacity and intestinal microbiota of largemouth bass. Triplicate groups of largemouth bass (average weight 35·03 ± 0·04 g), with a destiny of twenty-eight individuals per tank, were fed low-carbohydrate diet and HCD supplemented with different concentration of CBC (0 %, 0·25 %, 0·50 % and 1·00 %). The results showed that dietary CBC inclusion alleviated the hepatic glycogen accumulation induced by HCD intake. Additionally, the expression of hepatic ampkα1 and insulin signaling pathway-related genes (ira, irb, irs, p13kr1 and akt1) increased linearly with dietary CBC inclusion, which might be associated with the activation of glycolysis-related genes (gk, pfkl and pk). Meanwhile, the expression of intestinal SCFA transport-related genes (ffar3 and mct1) was significantly increased with dietary CBC inclusion. In addition, the hepatic antioxidant capacity was improved with dietary CBC supplementation, as evidenced by linear decrease in malondialdehyde concentration and expression of keap1, and linear increase in antioxidant enzyme activities (total antioxidative capacity, total superoxide dismutase and catalase) and expression of antioxidant enzyme-related genes (nrf2, sod1, sod2 and cat). The analysis of bacterial 16S rRNA V3-4 region indicated that dietary CBC inclusion significantly reduced the enrichment of Firmicutes and potential pathogenic bacteria genus Mycoplasma but significantly elevated the relative abundance of Fusobacteria and Cetobacterium. In summary, dietary CBC inclusion improved carbohydrate utilization, antioxidant capacity and intestinal microbiota of largemouth bass fed HCD.

Dietary lipid sources affect growth performance, lipid deposition, antioxidant capacity and inflammatory response of largemouth bass (Micropterus salmoides).

The present study explored the effects of different lipid sources on growth performance, lipid deposition, antioxidant capacity, inflammatory response and disease resistance of largemouth bass (Micropterus salmoides). Four isonitrogenous (crude protein 50.46 %) and isolipidic (crude lipid 11.12 %) diets were formulated to contain 7 % of different oil sources including fish oil (FO) (control), soybean oil (SO), linseed oil (LO) and coconut oil (CO). Largemouth bass with initial body weight of 36.0 ± 0.2 g were randomly distributed into 12 tanks, with 30 fish per tank and 3 tanks per treatment. The fish were fed with the experiment diets twice daily for 8 weeks. The results indicated that the weight gain of largemouth bass fed the FO diet was significantly higher than that of fish fed the LO and CO diets. The liver crude lipid content in FO group was significantly higher than other groups, while the highest liver triglyceride content was showed in SO group and the lowest was detected in LO group. At transcriptional level, expression of lipogenesis related genes (pparγ, srebp1, fas, acc, dgat1 and dgat2) in the SO and CO group were significantly higher than the FO group. However, the expression of lipolysis and fatty acids oxidation related genes (pparα, cpt1, and aco) in vegetable oils groups were significantly higher than the FO group. As to the antioxidant capacity, vegetable oils significantly reduced the malondialdehyde content of largemouth bass. Total antioxidant capacity in the SO and LO groups were significantly increased compared with the FO group. Catalase in the LO group was significantly increased compared with the FO group. Furthermore, the ER stress related genes, such as grp78, atf6α, atf6ß, chop and xbp1 were significantly enhanced in the vegetable oil groups compared with the FO group. The activity of serum lysozyme in vegetable oil groups were significantly higher than in FO group. Additionally, the relative expression of non-specific immune related genes, including tlr2, mapk11, mapk13, mapk14, rela, tgf-ß1, tnfα, 5lox, il-1ß and il10, were all significantly increased in SO and CO groups compared to the other groups. In conclusion, based on the indexes including growth performance, lipid deposition, antioxidant capacity and inflammatory response, SO and LO could be alternative oil sources for largemouth bass.

Effects of Dietary Aflatoxin B1 on Hybrid Grouper (Epinephelus fuscoguttatus ♀ × Epinephelus lanceolatus ♂) Growth, Intestinal Health, and Muscle Quality.

This study investigated the effects of varying doses of dietary aflatoxin B1 (AFB1) on the growth, intestinal health, and muscle quality of hybrid grouper. Four diets with varying AFB1 concentrations (0, 30, 445, and 2,230 µg kg-1) were used. Elevating AFB1 concentrations led to a decline in growth indexes, specifically the weight gain rate and the specific growth rate, although the survival rate remained unchanged. Morphological indicators showed a dose-dependent decline with AFB1 exposure. Intestinal MDA content and hindgut reactive oxygen species (ROS) levels increased, while antioxidant indexes and digestive enzymes decreased with higher AFB1 levels. AFB1 negatively influenced hindgut tight junction protein and antioxidant-related gene expression while promoting inflammation-related gene expression. The presence of AFB1 in the experiment led to a decrease in beneficial intestinal bacteria, such as Prevotella, and an increase in harmful intestinal bacteria, such as Prevotellaceae_NK3B31_group. Muscle lipid and unsaturated fatty acid content significantly decreased, while muscle protein and liver AFB1 content increased dramatically with higher AFB1 concentrations. AFB1 caused myofibrillar cleavage and myofilament damage, leading to increased spaces between muscle fibers. In conclusion, diets with AFB1 levels exceeding 30 µg kg-1 inhibited hybrid grouper growth, while levels surpassing 445 µg kg-1 resulted in hindgut ROS accumulation, inflammation, elevated intestinal permeability, reduced digestive enzyme activity, and compromised muscle quality.

Effects of Clostridium butyricum on Growth Performance, Intestinal Health, and Disease Resistance of Hybrid Grouper (Epinephelus fuscoguttatus♀ × E. lanceolatus♂) Fed with Cottonseed Protein Concentrate (CPC) Replacement of Fishmeal.

An 8-week feeding trial was conducted to investigate the effects of C. butyricum on the growth performance, microbiota, immunity response, and disease resistance in hybrid grouper fed with cottonseed protein concentrate (CPC) replacement of fishmeal. Six groups of isonitrogenous and isolipid diets were formulated including a positive control group (50% fishmeal, PC), a negative control group (CPC replaced 50% of fishmeal protein, NC), and Clostridium butyricum supplemented with 0.05% (C1, 5 × 108 CFU/kg), 0.2% (C2, 2 × 109 CFU/kg), 0.8% (C3, 8 × 109 CFU/kg), and 3.2% (C4, 3.2 × 1010 CFU/kg), respectively, to the NC group. The results showed that weight gain rate and specific growth rate were significantly higher in the C4 group than that in the NC group (P < 0.05). After supplementation with C. butyricum, the amylase, lipase, and trypsin activities were significantly higher than the NC group (P < 0.05; except group C1), and the same results were obtained for intestinal morphometry. The intestinal proinflammatory factors were significantly downregulated, and the anti-inflammatory factors were significantly upregulated in the C3 and C4 groups compared with the NC group after supplementation with 0.8%-3.2% C. butyricum (P < 0.05). At the phylum level, the PC, NC, and C4 groups were dominated by the Firmicutes and the Proteobacteria. At the genus level, the relative abundance of Bacillus in the NC group was lower than that in the PC and C4 groups. After supplementation with C. butyricum, grouper in the C4 group showed significantly higher resistance to V. harveyi than the NC group (P < 0.05). Above all, taking into account the effects of immunity and disease resistance, it was recommended to supplement 3.2% C. butyricum in the diet of grouper fed the replacement of 50% fishmeal protein by CPC.

Modulation of growth performance, antioxidant capacity, non-specific immunity and disease resistance in largemouth bass (Micropterus salmoides) upon compound probiotic cultures inclusion.

An 8-week feeding trial was conducted to evaluate the effects of dietary supplementation of compound probiotic cultures (CPC; Bacillus subtilis, Lactobacillus plantarum and Saccharomyces cerevisiae) on the growth performance, antioxidant capacity, non-specific immunity and disease resistance of juvenile largemouth bass. Triplicate groups of largemouth bass (average weight 42.05 ± 0.02 g), with a destiny of 30 individuals per tank, were fed diets supplemented with different concentration of compound probiotic cultures (CPC) (0%, CPC (0.0); 0.5%, CPC (0.5); 1.0%, CPC (1.0); 2.0%, CPC (2.0)). After the feeding trial, tissue samples of largemouth bass were collected and the challenge test with Aeromonas hydrophila was performed. Results indicated that the CPC supplementation produced no significant difference on the growth performance, feed utilization and body composition of largemouth bass, while significantly increased the cumulative survival rate in the Aeromonas hydrophila challenge test. Meanwhile, the inclusion of CPC elevated the hepatic antioxidant capacity, and the highest activity of antioxidant enzymes, including T-AOC, CAT, GPx and T-SOD, was observed in the CPC (2.0) group. Meanwhile, the transcription of Nrf2/keap1 and antioxidant related genes, including CAT, GPx, GST, SOD1 and SOD2, was significantly elevated with the inclusion of CPC. In addition, the inclusion of CPC improved the non-specific immunity of largemouth bass. The activity of serum lysozyme was significantly elevated in the CPC (2.0) group, while the transcription of RelA and pro-inflammatory factors, including TNF-α and IL-1ß, was inhibited with the inclusion of CPC. Meanwhile, related genes potentially linked to RelA, including TLR2 and p38 MAPK, were detected that their relative expression was significantly inhibited with the inclusion of CPC. The current findings indicated that the inclusion of 2% CPC improved the antioxidant capacity, non-specific immunity and disease resistance of juvenile largemouth bass, and suggested that 2% CPC as a functional additive could be applied to the diet of juvenile largemouth bass in aquaculture practice.

Effects of the Clostridium butyricum on growth performance, antioxidant capacity, immunity and disease resistance of Litopenaeus Vannamei fed with cottonseed protein concentrate (CPC) replacement of fishmeal in diet.

Clostridium butyricum (CB) is a gram-positive bacterium that secretes short-chain fatty acids such as butyric acid and so on. An 8-week feeding trial was conducted to investigate the effects of CB on the growth performance, antioxidant capacity, immunity and resistance to Vibrio parahaemolyticus in Litopenaeus Vannamei fed with cottonseed protein concentrate (CPC) replacement of fishmeal. Six iso-nitrogenous (40%) and iso-lipidic (6%) diets were formulated including a positive control group (PC, 25% fishmeal), a negative control group (NC, CPC replaced 30% of fishmeal protein), and 0.03% (C1, 3 × 108 CFU/kg), 0.12% (C2, 1.2 × 109 CFU/kg), 0.48% (C3, 4.8 × 109 CFU/kg) and 1.92% (C4, 1.92 × 1010 CFU/kg) CB were supplemented on the negative control group (NC). After the feeding trial, the remaining shrimp in each treatment group were subjected to a challenge experiment with Vibrio parahaemolyticus. The results indicated that weight gain rate (WGR), specific growth rate (SGR) in C4 group were significantly lower than those in PC and C2 groups (P < 0.05); the feed conversion ratio (FCR) was significantly higher than that of PC and C2 groups (P < 0.05). There was no significant difference in survival rate (SR) among all groups (P > 0.05). Compared to the PC and NC groups, the total superoxide capacity, superoxide dismutase and lysozyme were significantly higher in the C4 group (P < 0.05); the glutathione peroxidase, acid phosphatase and alkaline phosphatase were significantly higher in the C3 group (P < 0.05); and the malondialdehyde was significantly lower in the C4 group (P < 0.05). The relative mRNA expressions of Toll receptor (TLR), innate immune deficiency gene (IMD), penaiedin3a (Pen3) were significantly down-regulated in the NC group than those in the PC group (P < 0.05). In addition, the relative mRNA expressions of TLR, IMD and Pen3 were significantly up-regulated in all groups supplemented with CB than those in the NC group (P < 0.05). Moreover, the cumulative mortality rate in the NC group was not significantly different from the PC group (P > 0.05) and was significantly higher than those in the C3 and C4 groups (P < 0.05). In conclusion, the CB supplementation on the basis of CPC replacement of 30% fishmeal protein enhanced significantly the antioxidant capacity, immunity and disease resistance of shrimp and improved its growth performance. Therefore, considering the factors of the growth, immunity and disease resistance, the CB supplementation of 0.12%-0.48% (1.2 × 109 CFU/kg-4.8 × 109 CFU/kg) was recommended in the diet of L. vannamei based on the results of this experiment.

Effects of ammonia nitrogen stress on the physiological, biochemical, and metabolic levels of the gill tissue of juvenile four-finger threadfin (Eleutheronema tetradactylum).

In this study, the impact of ammonia nitrogen stress on juvenile four-finger threadfin in pond culture was examined. The 96-hour median lethal concentration (LC50-96h) and safe concentration of ammonia nitrogen were assessed in juveniles with a body weight of 7.4 ± 0.6 g using ecotoxicological methods. The study design included a stress group exposed to LC50-96h levels of ammonia nitrogen and a control group without ammonia nitrogen exposure. To examine the physiological, biochemical, and metabolic effects of ammonia nitrogen on gill tissue, gill tissue samples were collected after 12, 24, 48, and 96 h of stress, with a resumption of treatment after 48 h. Compared to the control group, ammonia nitrogen adversely affected juvenile four-finger threadfin, with LC50-96h and safe concentration values of 20.70 mg/L and 2.07 mg/L, respectively. Exposure to ammonia nitrogen resulted in substantial gill damage, including fusion of lamellae, epithelial cell loss, and proliferation of chlorine-secreting cells. This tissue damage persisted even after a 48-h recovery period. Ammonia nitrogen stress triggered an increase in antioxidant enzyme activity (superoxide dismutase, catalase, and glutathione peroxidase) and malondialdehyde levels in gills, indicating oxidative stress from 12 h onwards. Although enzyme activity decreased over time, oxidative stress persisted even after recovery, suggesting an ongoing need for antioxidant defense. Metabolomics analysis showed significant alterations in 423 metabolites under ammonia nitrogen stress. Key metabolites such as L-arginine, taurine, 20-hydroxyarachidonic acid, 11,12-dihydroxy-5Z, 8Z, and 14Z eicosotrienic acid followed an increasing trend; uridine, adenosine, L-glutathione, and thymidine 5'-triphosphate followed a decreasing trend. These changes reflect metabolic adaptations to stress. In enriched metabolic pathways, the main differential pathways are membrane transport, lipid metabolism, and amino acid metabolism. After 48 h, significant differences were observed in 396 metabolites compared to the control group. Notably, L-arginine, choline, and L-histidine increased, while linoleic acid, adenosine, and glutathione decreased. Amino acid and lipid metabolism pathways were key affected pathways. Under ammonia nitrogen stress, juvenile four-finger threadfin increased the synthesis of unsaturated and saturated fatty acids to cope with low temperatures and bolster immune function by consuming spermidine. This adaptation helps to clear peroxides generated during fatty acid synthesis, thereby protecting cells from oxidative damage. This study provides insights for pond aquaculture and breeding of ammonia nitrogen-tolerant fish strains.

Effects of fermented soybean meal substitution for fish meal on intestinal flora and intestinal health in pearl gentian grouper.

This study explored the role of replacing fish meal protein with fermented soybean meal (FSBM) protein on the growth performance and intestinal morphology, immunity, and microbiota of the pearl gentian grouper (Epinephelus fuscoguttatus♀ × E. lanceolatus♂). Three isonitrogenous and isolipidic diets with increasing levels of FSBM (0%, 20% and 40%; referred to as FM, FSBM20 and FSBM40 diets, respectively) as a replacement for fish meal were selected for this study. The pearl gentian grouper were fed these diets for 10 weeks. The findings revealed that the growth of fish fed the FSBM diets (FSBM20 and FSBM40) were remarkably lower than the fish fed the FM diet. Pathological manifestations of intestinal inflammation, such as shortened intestinal mucosal folds and thickened lamina propria, were observed in the fish fed the FSBM diets. Moreover, the gene expression levels of IL1ß, IL12, IL17, and TNFα were remarkably upregulated in fish fed the FSBM40 diet, in contrast to the gene expression levels of IL4, IL5, IL10, and TGFß1, which were remarkably downregulated (p < 0.05). The FSBM diets significantly affected the stability of the fish gut microbiota. Photobacterium was the dominant phylum in all experimental groups, and the proportion of these bacteria gradually decreased with increasing FSBM substitution. The composition of intestinal flora at the genus level was not the same in the three experimental groups, with a richer composition of intestinal bacteria detected in the FSBM20 and FSBM40 groups (p < 0.05). The correlation between intestinal flora balance and immune gene expression revealed that only Photobacterium was negatively correlated with the above upregulated genes, while other bacteria were positively correlated with these pro-inflammatory factors (p < 0.05). Photobacterium was positively correlated with the above downregulated genes, while other bacteria were negatively correlated with these anti-inflammatory factors (p < 0.05). In conclusion, high levels of substitution of FSBM for fish meal causes intestinal inflammation in pearl gentian grouper. This is likely associated with changes to the intestinal flora. More attention should be paid to the negative role of dietary FSBM on intestinal flora.

Effects of Dietary Alpha-Lipoic Acid on Growth Performance, Serum Biochemical Indexes, Liver Antioxidant Capacity and Transcriptome of Juvenile Hybrid Grouper (Epinephelus fuscoguttatus♀ × Epinephelus polyphekadion♂).

We aimed to investigate the effects of dietary alpha-lipoic acid (α-LA) on the growth performance, serum biochemical indexes, liver morphology, antioxidant capacity, and transcriptome of juvenile hybrid groupers (Epinephelus fuscoguttatus♀ × Epinephelus polyphekadion♂). Four experimental diets supplemented with 0 (SL0), 0.4 (L1), 0.6 (L2), and 1.2 (L3) g/kg α-LA were formulated and fed to three replicates of juvenile hybrid grouper (24.06 ± 0.15 g) for 56 d. The results indicated that dietary 0.4 and 0.6 g/kg α-LA significantly decreased the weight gain rate in juvenile hybrid groupers. Compared with SL0, the content of total protein in the serum of L1, L2, and L3 increased significantly, and alanine aminotransferase decreased significantly. The content of albumin in the serum of L3 increased significantly, and triglyceride, total cholesterol, and aspartate aminotransferase decreased significantly. In addition, the hepatocyte morphology in L1, L2, and L3 all showed varying degrees of improvement, and the activities of glutathione peroxidase and superoxide dismutase in the liver of L2 and L3 were significantly increased. A total of 42 differentially expressed genes were screened in the transcriptome data. KEGG showed that a total of 12 pathways were significantly enriched, including the pathway related to immune function and glucose homeostasis. The expression of genes (ifnk, prl4a1, prl3b1, and ctsl) related to immune were significantly up-regulated, and the expressions of gapdh and eno1 genes related to glucose homeostasis were significantly down-regulated and up-regulated, respectively. In summary, dietary supplementation of 0.4 and 0.6 g/kg α-LA inhibited the growth performance of juvenile hybrid groupers. A total of 1.2 g/kg α-LA could reduce the blood lipid level, improve hepatocyte damage, and increase the hepatic antioxidant enzyme activity. Dietary α-LA significantly affected the pathway related to immune function and glucose homeostasis.

Effects of High-Lipid Dietary Protein Ratio on Growth, Antioxidant Parameters, Histological Structure, and Expression of Antioxidant- and Immune-Related Genes of Hybrid Grouper.

The hybrid grouper (♀ Epinephelus fuscoguttatus × â™‚ E. lanceolatus) is a new species of grouper crossed from giant grouper (E. lanceolatus) as the male parent and brown-marbled grouper (E. fuscoguttatus) as the female parent. We hypothesized that optimal levels of dietary protein may benefit liver function. High-lipid diets are energetic feeds that conserve protein and reduce costs, and are a hot topic in aquaculture today. Therefore, the objective of the research is to investigated the effects of dietary protein level in high-lipid diets on serum and liver biochemistry, liver histology, and liver immune and antioxidant indexes and gene mRNA expression of the juvenile hybrid grouper (♀ Epinephelus fuscoguttatus × â™‚ E. lanceolatus). Six iso-lipidic (161 g/kg) diets were formulated containing graded levels of protein (510 as control, 480,450, 420, 390 and 360 g/kg). Each treatment consisted of three replicates and 30 fish (6.70 ± 0.02 g) in one replicate. After an 8-week feeding experiment, the results indicated the following: (1) With the decreasing of dietary protein level, the specific growth rate (SGR) of groupers increased gradually and then decreased; SGRs of the 390 and 360 g/kg groups were significantly lower than other groups (p < 0.05). (2) In terms of serum and liver, the activity of antioxidant enzymes such as catalase (CAT) and superoxide dismutase (SOD), and the total antioxidant capacity (T-AOC) content, and the activity of immune enzymes such as lysozyme (LYS) and immunoglobulin (IgM) was significantly increased under the appropriate protein level. (3) Based on liver histology, we know that high or low dietary protein levels cause liver damage. (4) Dietary protein levels can significantly affect the mRNA expression levels of an anti-inflammatory factor gene (tgfß), pro-inflammatory factor genes (il6, il8), heat shock proteins, and antioxidant and immune genes (hsp70 and hsp90, gpx, nrf2, keap1). It is concluded that the appropriate protein level can promote the growth performance of groupers, improve antioxidant activity and immune enzyme activity in serum and liver, and enhance the expression of immune genes.

Functional Properties of Protein Hydrolysates on Growth, Digestive Enzyme Activities, Protein Metabolism, and Intestinal Health of Larval Largemouth Bass (Micropterus salmoides).

The present study was conducted to investigate the effects of dietary inclusion of protein hydrolysates on growth performance, digestive enzyme activities, protein metabolism, and intestinal health in larval largemouth bass (Micropterus salmoides). The experimental feeding trial presented in this study was based on five isonitrogenous and isolipidic diets formulated with graded inclusion levels of protein hydrolysates, and it showed that protein hydrolysates improved growth performance, reduced larval deformity rate, and increased the activity of digestive enzymes, including pepsin and trypsin. Gene expression results revealed that the supplementation of protein hydrolysates upregulated the expression of intestinal amino acid transporters LAT2 and peptide transporter 2 (PepT2), as well as the amino acid transporters LAT1 in muscle. Dietary provision of protein hydrolysates activated the target of rapamycin (TOR) pathway including the up-regulation of TOR and AKT1, and down-regulation of 4EBP1. Additionally, the expression of genes involved in the amino acids response (AAR) pathway, ATF4 and REDD1, were inhibited. Protein hydrolysates inhibited the transcription of some pro-inflammatory cytokines, including IL-8 and 5-LOX, but promoted the expression of anti-inflammatory cytokines TGF-ß and IL-10. The 16S rRNA analysis, using V3-V4 region, indicated that dietary protein hydrolysates supplementation reduced the diversity of the intestine microbial community, increased the enrichment of Plesiomonas and reduced the enrichment of Staphylococcus at the genus level. In summary, protein hydrolysates have been shown to be an active and useful supplement to positively complement other protein sources in the diets for largemouth bass larvae, and this study provided novel insights on the beneficial roles and possible mechanisms of action of dietary protein hydrolysates in improving the overall performance of fish larvae.

Tryptophan metabolism and gut flora profile in different soybean protein induced enteritis of pearl gentian groupers.

The substitution of high-level soy meals for fish meal (FM) generally leads to fish enteritis, accompanied by significant variations in gut flora. Relevant studies have pointed out a close relationship between tryptophan metabolism mediated by gut flora and vertebrate inflammatory bowel disease. Present study examines the role of tryptophan metabolism and gut flora profile in fish enteritis caused by different soybean meals. The 960 groupers were randomly assigned into 4 groups (n = 4), which including: (1) FM (the control group, fed with 50% FM feed), (2) SBM40 (replacing 40% FM with soybean meal), (3) SPC40 (replacing 40% FM with soybean protein concentrate), and (4) FSBM40 (replacing 40% FM with fermented soybean meal). Under average temperature and natural light, the groupers were cultivated with feeds of iso-nitrogen and iso-lipid for 10 weeks. The results showed that soybean meal feeds at all experimental levels had negative effects on fish gut physiology and growth performance. Typical enteritis features and fluctuations of immune system occur, which can be observed in the enzyme activities of total superoxide dismutase and lysozyme and in the contents of immunoglobulin M, complement 3 and complement 4. 16SrDNA high-throughput sequencing indicated that it greatly influenced the gut flora with the abundance of maleficent bacteria, like Vibrio, amplified with increasing dietary soybean meals. According to the "3 + 2" full-length transcriptome sequencing, soy meals at the three experimental levels inhibited the key gene expressions of tryptophan metabolic pathway in fish gut, however, there are some differences in the types of key genes that are inhibited. The canonical correlation analysis showed that the changes in key gene expressions in tryptophan metabolic pathway had a positive correlation with the expressions of pro-inflammatory genes (P < 0.05) and negatively correlated with the expression of anti-inflammatory genes (P < 0.05). It is speculated from this study that tryptophan metabolism is closely related to fish soy meal-related enteritis, and the abnormal tryptophan metabolism caused by intestinal flora imbalance may play an important role. In the future research, we can further study the tolerance of fish to soy meals feed from two aspects of tryptophan metabolism and intestinal flora changes.

End-to-End Fovea Localisation in Colour Fundus Images With a Hierarchical Deep Regression Network.

Accurately locating the fovea is a prerequisite for developing computer aided diagnosis (CAD) of retinal diseases. In colour fundus images of the retina, the fovea is a fuzzy region lacking prominent visual features and this makes it difficult to directly locate the fovea. While traditional methods rely on explicitly extracting image features from the surrounding structures such as the optic disc and various vessels to infer the position of the fovea, deep learning based regression technique can implicitly model the relation between the fovea and other nearby anatomical structures to determine the location of the fovea in an end-to-end fashion. Although promising, using deep learning for fovea localisation also has many unsolved challenges. In this paper, we present a new end-to-end fovea localisation method based on a hierarchical coarse-to-fine deep regression neural network. The innovative features of the new method include a multi-scale feature fusion technique and a self-attention technique to exploit location, semantic, and contextual information in an integrated framework, a multi-field-of-view (multi-FOV) feature fusion technique for context-aware feature learning and a Gaussian-shift-cropping method for augmenting effective training data. We present extensive experimental results on two public databases and show that our new method achieved state-of-the-art performances. We also present a comprehensive ablation study and analysis to demonstrate the technical soundness and effectiveness of the overall framework and its various constituent components.

Effects of Dietary Phospholipids on Growth Performance, Digestive Enzymes Activity and Intestinal Health of Largemouth Bass (Micropterus salmoides) Larvae.

While the beneficial roles of dietary phospholipids on health status and overall performances of fish larvae have been well demonstrated, the underlying mechanisms remain unclear. To address this gap, the present study was conducted to investigate the effects of dietary phospholipids on growth performance, intestinal development, immune response and microbiota of larval largemouth bass (Micropterus salmoides). Five isonitrogenous and isolipidic micro-diets were formulated to contain graded inclusion levels of phospholipids (1.69, 3.11, 5.23, 7.43 and 9.29%). Results showed that the supplementation of dietary phospholipids linearly improved the growth performance of largemouth bass larvae. The inclusion of dietary phospholipids increased the activity of digestive enzymes, such as lipase, trypsin and alkaline phosphatase, and promoted the expression of tight junction proteins including ZO-1, claudin-4 and claudin-5. Additionally, dietary phospholipids inclusion alleviated the accumulation of intestinal triacylglycerols, and further elevated the activity of lysozyme. Dietary phospholipids inhibited the transcription of some pro-inflammatory cytokines, including il-1ß, and tnf-α, but promoted the expression of anti-inflammatory cytokines tgf-ß, with these modifications being suggested to be mediated by the p38MAPK/Nf-κB pathway. The analysis of bacterial 16S rRNA V3-4 region indicated that the intestinal microbiota profile was significantly altered at the genus level with dietary phospholipids inclusion, including a decreased richness of pathogenic bacteria genera Klebsiella in larval intestine. In summary, it was showed that largemouth bass larvae have a specific requirement for dietary phospholipids, and this study provided novel insights on how dietary phospholipids supplementation contributes to improving the growth performance, digestive tract development and intestinal health.

AGE challenge: Angle Closure Glaucoma Evaluation in Anterior Segment Optical Coherence Tomography.

Angle closure glaucoma (ACG) is a more aggressive disease than open-angle glaucoma, where the abnormal anatomical structures of the anterior chamber angle (ACA) may cause an elevated intraocular pressure and gradually lead to glaucomatous optic neuropathy and eventually to visual impairment and blindness. Anterior Segment Optical Coherence Tomography (AS-OCT) imaging provides a fast and contactless way to discriminate angle closure from open angle. Although many medical image analysis algorithms have been developed for glaucoma diagnosis, only a few studies have focused on AS-OCT imaging. In particular, there is no public AS-OCT dataset available for evaluating the existing methods in a uniform way, which limits progress in the development of automated techniques for angle closure detection and assessment. To address this, we organized the Angle closure Glaucoma Evaluation challenge (AGE), held in conjunction with MICCAI 2019. The AGE challenge consisted of two tasks: scleral spur localization and angle closure classification. For this challenge, we released a large dataset of 4800 annotated AS-OCT images from 199 patients, and also proposed an evaluation framework to benchmark and compare different models. During the AGE challenge, over 200 teams registered online, and more than 1100 results were submitted for online evaluation. Finally, eight teams participated in the onsite challenge. In this paper, we summarize these eight onsite challenge methods and analyze their corresponding results for the two tasks. We further discuss limitations and future directions. In the AGE challenge, the top-performing approach had an average Euclidean Distance of 10 pixels (10 µm) in scleral spur localization, while in the task of angle closure classification, all the algorithms achieved satisfactory performances, with two best obtaining an accuracy rate of 100%. These artificial intelligence techniques have the potential to promote new developments in AS-OCT image analysis and image-based angle closure glaucoma assessment in particular.

[Effects of Phenol on Activity and Microfauna Community Structure of Activated Sludge].

In order to investigate the effects of phenol on activity and microfauna community structure of activated sludge, phenol with different concentrations were added into the wastewater for activated sludge culturing in a SBR reactor, then the activity parameters of TTC-ETS and INT-ETS of activated sludge were measured, and the dynamics of microfauna community was analyzed. The results indicated that TTC-ETS activity was more applicable to evaluate the effects of phenol on sludge activity than INT-ETS activity. The higher the concentration of phenol was, the more seriously the sludge activity was inhibited. When 50 mg·L-1 of phenol was added into the influent, the inhibition rate of sludge activity was (20.75±10.43)%, while it increased to (39.73±26.92)% with a great fluctuation at the phenol concentration of 100 mg·L-1. The inhibition rate of sludge activity fluctuated at about 40% in the later period of sludge culturing with 300 mg·L-1 of phenol. The inhibitory effects on microfauna community in activated sludge increased with increasing phenol concentration. Phenol had different effects on various groups of microfauna community, i.e., only one group (testate amoebae) was inhibited significantly at a low feeding concentration of phenol, many groups (sessile ciliates, testate amoebae, crawling ciliates, carnivorous ciliates) were inhibited at the phenol concentrations of 100 mg·L-1 and 300 mg·L-1, whereas a few groups (flagellates, nematodes, etc.) were promoted at the phenol concentration of 300 mg·L-1. There was a certain degree of relationship between sludge activity and microfauna under the influence of phenol, which means that Centropyxis aculeata, Plagiocampa mutabilis etc., may be bio-indicators for low sludge activity, and Epistylis lacustris, Bodo lens and Pleuromonas jaculans can be used as bio-indicators for high sludge activity in the activated sludge used to treat phenolic wastewater.