Strategies to enhance production of metabolites in microbial co-culture systems.

Increasing evidence shows that microbial synthesis plays an important role in producing high value-added products. However, microbial monoculture generally hampers metabolites production and limits scalability due to the increased metabolic burden on the host strain. In contrast, co-culture is a more flexible approach to improve the environmental adaptability and reduce the overall metabolic burden. The well-defined co-culturing microbial consortia can tap their metabolic potential to obtain yet-to-be discovered and pre-existing metabolites. This review focuses on the use of a co-culture strategy and its underlying mechanisms to enhance the production of products. Notably, the significance of comprehending the microbial interactions, diverse communication modes, genetic information, and modular co-culture involved in co-culture systems were highlighted. Furthermore, it addresses the current challenges and outlines potential future directions for microbial co-culture. This review provides better understanding the diversity and complexity of the interesting interaction and communication to advance the development of co-culture techniques.

Natural Sunlight-Mediated Emodin Photoinactivation of Aeromonas hydrophila.

Aeromonas hydrophila can be a substantial concern, as it causes various diseases in aquaculture. An effective and green method for inhibiting A. hydrophila is urgently required. Emodin, a naturally occurring anthraquinone compound, was exploited as a photo-antimicrobial agent against A. hydrophila. At the minimum inhibitory concentration of emodin (256 mg/L) to inactivate A. hydrophilia in 30 min, an 11.32% survival rate was observed under 45 W white compact fluorescent light irradiation. In addition, the antibacterial activity under natural sunlight (0.78%) indicated its potential for practical application. Morphological observations demonstrated that the cell walls and membranes of A. hydrophila were susceptible to damage by emodin when exposed to light irradiation. More importantly, the photoinactivation of A. hydrophila was predominantly attributed to the hydroxyl radicals and superoxide radicals produced by emodin, according to the trapping experiment and electron spin resonance spectroscopy. Finally, a light-dependent reactive oxygen species punching mechanism of emodin to photoinactivate A. hydrophila was proposed. This study highlights the potential use of emodin in sunlight-mediated applications for bacterial control, thereby providing new possibilities for the use of Chinese herbal medicine in aquatic diseases prevention.

Alterations of Plasma Biochemical and Immunological Parameters and Spatiotemporal Expression of TLR2 and TLR9 in Gibel Carp (Carassius auratus gibelio) after CyHV-2 Infection.

Cyprinid herpesvirus II (CyHV-2), a highly contagious pathogen of gibel carp (Carassius auratus gibelio), causes herpesviral hematopoietic necrosis disease (HVHND) and enormous financial losses. However, there is limited information available regarding the changes in plasma biochemical and immunological parameters and the response characteristics of Toll-like receptor 2 (TLR2) and Toll-like receptor 9 (TLR9) in gibel carp after CyHV-2 infection. To address this knowledge gap, a sub-lethal CyHV-2 infection was conducted in gibel carp, and the sample was collected daily from 1 to 7 days post infection. The plasma biochemical analyses showed significant decreases in the content of glucose, total cholesterol (TCHO), and total protein (TP), along with marked increases in the level of uric acid, urea, creatinine (CREA), Complement 3 (C3), immunoglobulin D (IgD), and immunoglobulin M (IgM) as well as in the activity of alanine aminotransferase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP), and lactate dehydrogenase (LDH) in the infected group. Compared with the control group, the concentration of cortisol, triglyceride (TG), and Complement 4 (C4) had no noticeable alterations in the infected group. Real-time quantitative PCR analysis showed significant upregulation of TLR2 and TLR9 mRNA expression in the spleen, kidney, brain, liver, intestine, and gill post CyHV-2 infection. Interestingly, a time- and tissue-dependent expression profile has been comparatively observed for TLR2 and TLR9 in the above tissues of gibel carp after CyHV-2 infection, suggesting distinct roles between TLR2 and TLR9 in antiviral response to CyHV-2 infection. Overall, our results demonstrated that CyHV-2 infection led to the disruption of the physiological metabolic process and damage to the liver and kidney, and induced different spatiotemporal expression patterns of TLR2 and TLR9, ultimately stimulating antiviral response via innate and adaptive immune system. These findings may provide a deeper understanding of the host immunity response to CyHV-2 infection and offer novel perspectives for the prevention and treatment and therapeutic drug development against CyHV-2.

Carvacrol Inhibits Quorum Sensing in Opportunistic Bacterium Aeromonas hydrophila.

Bacterial quorum sensing (QS) plays a crucial role in chemical communication between bacteria involving autoinducers and receptors and controls the production of virulence factors in bacteria. Therefore, reducing the concentration of signaling molecules in QS is an effective strategy for mitigating the virulence of pathogenic bacteria. In this study, we demonstrated that carvacrol at 15.625 µg/mL (1/4 MIC), a natural compound found in plants, exhibits potent inhibitory activity against QS in Chromobacterium violaceum, as evidenced by a significant reduction (62.46%) in violacein production. Based on its impressive performance, carvacrol was employed as a natural QS inhibitor to suppress the pathogenicity of Aeromonas hydrophila NJ-35. This study revealed a significant reduction (36.01%) in the concentration of N-acyl-homoserine lactones (AHLs), a QS signal molecular secreted by A. hydrophila NJ-35, after 1/4 MIC carvacrol treatment. Moreover, carvacrol was found to down-regulate the expression of ahyR/I, two key genes in the QS system, which further inhibited the QS system of A. hydrophila NJ-35. Finally, based on the above results and molecular docking, we proposed that carvacrol alleviate the pathogenicity of A. hydrophila NJ-35 through QS inhibition. These results suggest that carvacrol could serve as a potential strategy for reducing the virulence of pathogenic bacteria and minimizing the reliance on antibiotics in aquaculture.

Predation of Cyclopoid Copepods on the Theronts of Ichthyophthirius multifiliis: Shedding Light on Biocontrol of White Spot Disease.

White spot disease, caused by the parasitic ciliate Ichthyophthirius multifiliis, is a significant threat to the freshwater fish farming industry worldwide, resulting in massive mortality and economic losses. Eliminating the free-swimming theronts from the culture environment is considered crucial for the control of I. multifiliis infection. It is well-documented that planktonic ciliates are valuable food resources for macro-zooplankton in aquatic ecosystems. In this study, we developed a fluorescence labeling method for alive theronts and found that cyclopoid copepods Thermocyclops taihokuensis, Mesocyclops spp., Macrocyclops sp., and Paracyclopina sp. present predation on the theronts in co-culture experiments. Laboratory challenge tests further confirmed that the presence of zooplankton in the culture water body significantly reduced the infection of I. multifiliis in goldfish (p < 0.01). Results from this study revealed that cyclopoid copepods have the potential to be used as biological control agents against white spot disease in aquaculture.

In Vitro Antibiofilm Activity of Resveratrol against Aeromonas hydrophila.

Aeromonas hydrophila is a Gram-negative bacterium that widely exists in various aquatic environments and causes septicemia in fish and humans. Resveratrol, a natural polyterpenoid product, has potential chemo-preventive and antibacterial properties. In this study, we investigated the effect of resveratrol on A. hydrophila biofilm formation and motility. The results demonstrated that resveratrol, at sub-MIC levels, can significantly inhibit the biofilm formation of A. hydrophila, and the biofilm was decreased with increasing concentrations. The motility assay showed that resveratrol could diminish the swimming and swarming motility of A. hydrophila. Transcriptome analyses (RNA-seq) showed that A. hydrophila treated with 50 and 100 µg/mL resveratrol, respectively, presented 230 and 308 differentially expressed genes (DEGs), including 90 or 130 upregulated genes and 130 or 178 downregulated genes. Among them, genes related to flagellar, type IV pilus and chemotaxis were significantly repressed. In addition, mRNA of virulence factors OmpA, extracellular proteases, lipases and T6SS were dramatically suppressed. Further analysis revealed that the major DEGs involved in flagellar assembly and bacterial chemotaxis pathways could be regulated by cyclic-di-guanosine monophosphate (c-di-GMP)- and LysR-Type transcriptional regulator (LTTR)-dependent quorum sensing (QS) systems. Overall, our results indicate that resveratrol can inhibit A. hydrophila biofilm formation by disturbing motility and QS systems, and can be used as a promising candidate drug against motile Aeromonad septicemia.

Effects of glycyrrhetinic acid ß on growth and virulence of Aeromonas hydrophila.

Aeromonas hydrophila is a significant pathogen to freshwater farmed animals, and antibiotics are usually used to control the bacterial septicemia caused by A. hydrophila. Due to the severe situation of development and spread of antibiotic resistance, there are stricter restrictions on antibiotics used in aquaculture. To evaluate the feasibility of glycyrrhetinic acid ß (GA) as an alternative therapy against bacterial infection, in this study, an A. hydrophila isolated from diseased fish is used to test the antibacterial, anti-virulence activity and therapeutic effect of GA in vitro and in vivo, respectively. Results showed that GA did not affect the growth of A. hydrophila in vitro, while it could down-regulate (p < 0.05) the mRNA expression of the hemolysis-related genes hly and aerA, and significantly inhibited (p < 0.05) hemolytic activity of A. hydrophila. In addition, in vivo test showed that oral administration of GA was ineffective in controlling acute infections caused by A. hydrophila. In conclusion, these findings suggested that GA was a potential anti-virulence candidate against A. hydrophila, but the application of GA for the prevention and treatment of A. hydrophila-related diseases was still a long way.

QseBC regulates in vitro and in vivo virulence of Aeromonas hydrophila in response to norepinephrine.

The inter-kingdom communication between host and pathogenic bacteria mediated by the host hormones epinephrine (Epi)/norepinephrine (NE)/autoinducer-3 (AI-3) and transduced by the bacterial two-component signal transduction system QseBC has been well demonstrated in mammalian pathogens. Aeromonas hydrophila, a common opportunistic pathogen in freshwater aquaculture, responds to NE by increased bacterial growth and enhanced virulence. However, the underlying mechanisms remain poorly understood. Our study demonstrated that deletion of qseB and qseC significantly inhibited NE-promoted growth, biofilm formation, and hemolytic activity of A. hydrophila. The adhesion ability of ΔqseB and ΔqseC to J774a.1 cells was significantly decreased compared with the wild-type strain in the presence and absence of NE, whereas NE still enhanced the adhesion ability of the mutant and wild-type strains with a similar effect, suggesting that NE-enhanced cell adhesion was independent of QseBC. Moreover, QseBC did not affect the swimming and swarming motility of A. hydrophila with or without NE. Quantitative real-time PCR analyses revealed the down-regulated expression of some virulence-related genes (hly, ast, act, aerA) in each mutant compared with the wild-type strain in the presence of NE. Tilapia infection experiments indicated that deletion of qseB or qseC weakened NE-promoted virulence of A. hydrophila. In conclusion, our study suggests that NE stimulates the growth, biofilm formation, and hemolytic activity of A. hydrophila and enhances the virulence of the pathogen in fish via the QseBC system.

Protective Effects of Glycyrrhiza Total Flavones on Liver Injury Induced by Streptococcus agalactiae in Tilapia (Oreochromis niloticus).

Clinical studies have confirmed that Glycyrrhiza total flavones (GTFs) have good anti-hepatic injury, but whether they have a good protective effect on anti-hepatic injury activity induced by Streptococcus agalactiae in tilapia (Oreochromis niloticus) is unknown. The aims of this study were to investigate the protective effects of Glycyrrhiza total flavones on liver injury induced by S. agalactiae (SA) and its underlying mechanism in fish. A total of 150 tilapia were randomly divided into five groups, each with three replicates containing 10 fish: normal control group, S. agalactiae infection group, and three Glycyrrhiza total flavone treatment groups (addition of 0.1, 0.5, or 1.0 g of GTF to 1 kg of feed). The normal control group was only fed with basic diet, after 60 d of feeding, and intraperitoneal injection of the same volume of normal saline (0.05 mL/10 g body weight); the S. agalactiae infection group was fed with basic diet, and the S. agalactiae solution was intraperitoneally injected after 60 d of feeding (0.05 mL/10 g body weight); the three GTF treatment groups were fed with a diet containing 0.1, 0.5, or 1.0 g/kg of GTF, and the S. agalactiae solution was intraperitoneally injected after 60 d of feeding (0.05 mL/10 g body weight). After 48 h injection, blood and liver tissues were collected to measure biochemical parameters and mRNA levels to evaluate the liver protection of GTFs. Compared with the control group, the serum levels of glutamic oxaloacetic transaminase (GOT), glutamic pyruvic transaminase (GPT), alkaline phosphatase (AKP) and glucose (GLU) in the streptococcal infection group increased significantly, while the levels of total antioxidant capacity (T-AOC), superoxide dismutase (SOD), catalase (CAT) and reduced glutathione (GSH) decreased significantly; observations of pathological sections showed obvious damage to the liver tissue structure in response to streptococcal infection. S. agalactiae can also cause fatty liver injury, affecting the function of fatty acid ß-oxidation and biosynthesis in the liver of tilapia, and also causing damage to function of the immune system. The addition of GTFs to the diet could improve oxidative stress injury caused by S. agalactiae in tilapia liver tissue to different degrees, promote the ß-oxidation of fatty acids in the liver, accelerate the lipid metabolism in the liver, and repair the damaged liver tissue. GTFs have a good protective effect on liver injury caused by streptococcus.

Antimicrobial and Antivirulence Activities of Carvacrol against Pathogenic Aeromonas hydrophila.

Aeromonas hydrophila is a ubiquitous Gram-negative opportunistic pathogen in the freshwater environment and the most common cause of bacterial septicemia in aquaculture. In this study, we investigated the impact of carvacrol, a natural monoterpenoid found in herbs, on the virulence of A. hydrophila in vitro and the antibacterial effect in combination with antibiotics. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of carvacrol against A. hydrophila NJ-35 were 125 µg/mL and 250 µg/mL, respectively. Carvacrol could inhibit the virulence factors (biofilm, protease, exopolysaccharide, and hemolysin) of A. hydrophila, and the antibiofilm potential of carvacrol was further verified by microscopic detection. Transcriptional analyses showed that the gene expression of flaB, ompA, aha, ahp, ela, act, aerA, AhyR, and hly were marked as downregulated. The checkerboard assay results showed that carvacrol did not have an antagonistic effect in combination with antibiotics (florfenicol, enrofloxacin, thiamphenicol, or doxycycline hydrochloride) commonly used in aquaculture but possessed an additive-synergistic effect with neomycin sulfate. In vivo studies demonstrated that carvacrol protected grass carp (Ctenopharyngodon idella) from A. hydrophila infection. Our results indicated that carvacrol possessed significant anti-bacterial and anti-virulence effects on A. hydrophila.

The erection of a new genus, Neotegorhynchus n. g. (Palaeacanthocephala, Illiosentidae), with a redescription of Neotegorhynchus cyprini n. comb. from Cyprinus carpio from the Yangtze River basin, China.

The present paper summarizes acanthocephalan parasites of the family Illiosentidae Golvan, 1960, collected from the intestine of common carp (Cyprinus carpio L.) from Taihu Lake in Wuxi, Jiangsu Province, China. A total of 21 acanthocephalan specimens were found in 6 carps (prevalence of 35%). All the studied specimens were assigned to the family Illiosentidae based on the family-specific morphology of the worms and the presence of 8 cement glands in the males. However, the specimens recently found in common carp differed from all 14 extant genera of the family Illiosentidae in the structure of the reproductive system of both sexes, i.e. i) a vagina lacking a muscular sphincter; ii) the presence of a terminally pointed protruding tail end in the form of a dome with a muscular base; iii) the female genital muscles are fan-shaped cells, each of which has a nucleus and is not attached to the anterior wall of the body; iv) the muscular lip of the bursa of males repeats the curved shape of the posterior end of females, which en copula allows the lip of the bursa to seal to the posterior end of the female. Morphologically Neotegorhynchus n. g. is closest to the genus Tegorhynchus, but differs from it, in addition above features in i) the terminal position of the genital pores of females without a hollow genital vestibule and without a transverse cleft connected to the dorsal terminal genital pore, as in Tegorhynchus brevis; ii) cerebral ganglion at the border of the anterior and middle third of the proboscis vessel; iii) spherical cement reservoir. Molecular studies confirmed Neotegorhynchus n. g. as belonging to the family Illiosentidae, showing less than 98.9% sequence similarity in SSU rDNA and 81.8% in COI with the genus Dentitruncus. Therefore, a new acanthocephalan genus, Neotegorhynchus n. g., is erected, and Neotegorhynchus cyprini n. comb. is designated as the type species and its neotype.

Characterization of the complete mitochondrial genome of Nippotaenia mogurndae Yamaguti and Miyata, 1940 (Cestoda: Nippotaeniidae).

In this study, we report the first complete mitochondrial genome of the tapeworm Nippotaenia mogurndae in the order Nippotaeniidea Yamaguti, 1939. This mitogenome, which is 14,307 base pairs (bp) long with an A + T content of 72.2%, consists of 12 protein-coding genes, 22 transfer RNA (tRNA) genes, two rRNA genes, and two non-coding regions. Most tRNAs have a conventional cloverleaf structure, but trnS1 and trnR lack dihydrouridine arms of tRNA. The two largest non-coding regions, NCR1 (220 bp) and NCR2 (817 bp), are located between trnY and trnS2 and between nad5 and trnG, respectively. Phylogenetic analyses of mitogenomic data indicate that N. mogurndae is closely related to tapeworms in the order Cyclophyllidea.

Protective effects of paeonol against lipopolysaccharide-induced liver oxidative stress and inflammation in gibel carp (Carassius auratus gibelio).

Paeonol (2'-hydroxy-4'-methoxyacetophenone) is a phenol that exhibits antioxidant and anti-inflammatory capabilities. In this study, the underlying mechanism of paeonol against LPS-induced oxidative stress and inflammatory responses in gibel carp was investigated. Three hundred healthy gibel carp were divided into five groups (n = 9), intraperitoneally injected with LPS and thereafter treated with paeonol (16 mg/kg and 64 mg/kg). Fish were anesthetized with MS-222 (100 mg/L), and samples were collected at 72 h to investigate plasma biochemical indexes, liver histopathology, antioxidant enzymatic activity, and TLR receptor-related gene expression. Fish injected with LPS (20 mg/kg) exhibited significantly increased plasma aminotransferase (ALT), aminotransferase (AST), lactate dehydrogenase (LDH), glucose (GLU), diamine oxidase (DAO), and alkaline phosphatase (ALP) levels (P < 0.05). In addition, LPS challenge significantly enhanced myeloperoxidase (MPO) and malondialdehyde (MDA) contents, whereas those of catalase (CAT) and glutathione peroxidase (GSH-Px) decreased (P < 0.05). However, treatment with paeonol attenuated these LPS-induced changes (P < 0.05). The mRNA expression of TLR4, TIRAP, MyD88, TRAF6, NF-κB, TNF-α, IL-1ß, and IL-8, which were activated by LPS challenge (P < 0.05), were downregulated by paeonol. Additionally, histopathological examination demonstrated that paeonol alleviates LPS-induced hepatic tissue lesions in fish. Taken together, the results suggest that paeonol mitigates LPS-induced liver oxidative stress and inflammation in gibel carp.

Paeonol increases the antioxidant and anti-inflammatory capacity of gibel carp (Carassius auratus gibelio) challenged with Aeromonas hydrophila.

Paeonol, a naturally occurring polyphenol isolated from medical plant, has been known to exhibit anti-oxidative and anti-inflammatory effects. In order to evaluate the effect of paeonol on Carassius auratus gibelio infected by pathogenic bacteria Aeromonas hydriphila. 750 fish were randomly divided into 5 groups, which separately treated with 0.85% sterile saline (blank), A. hydriphila (negative control), A. hydriphila with paeonol (4 mg/kg, 64 mg/kg), and A. hydriphila with enrofloxacin (12 mg/kg, positive control). Fish were anaesthetized with MS-222 (100 mg/L), and samples were collected at 6 and 72 h after A. hydriphila challenge. The results showed that compared with the negative group, the survival in paeonol groups marked increased by 14.75% and 18.94%. The plasma immunoglobulin M (IgM) was notably increased, and low density lipoprotein (LDL) was significantly decreased in paeonol groups at 6 h (P < 0.05). The antioxidative enzymes catalase (CAT), total antioxidant capacity (T-AOC) and glutathione peroxidase (GSH-Px) were significantly increased in paeonol groups at 6 h, while malondialdehyde (MDA) and myeloperoxidase (MPO) contents were lower (P < 0.05). The inflammatory related genes MyD88 and TLR-5 were significantly downregulated, and the TLR-3 was significantly increased in paeonol groups at 72 h (P < 0.05). In addition, histopathological analyses showed that the lesion in liver, spleen and caudal kidney were considerably attenuated in paeonol groups. In conclusion, paeonol could increase the survival rate, mitigate oxidative damage, inflammation, tissue lesions, and improve the immunity of gibel carp challenged with A. hydrophila.

Tributyrin Plays an Important Role in Regulating the Growth and Health Status of Juvenile Blunt Snout Bream (Megalobrama amblycephala), as Evidenced by Pathological Examination.

The present study aimed to assess the role of tributyrin (TB) in regulating the growth and health status of juvenile blunt snout bream (Megalobrama amblycephala) through an 8-week feeding experiment. Six groups were fed experimental diets with added TB percentages of 0% (control group), 0.03%, 0.06%, 0.09%, 0.12% and 0.15%. The present results showed that TB supplementation in feed had some positive impacts on FW, WG, FCR and SGR, and the best results were found in the 0.06% TB group (P<0.05). However, TB supplementation in feed had no significant effects on SR, CF, VSI or whole-body composition (P>0.05). TB supplementation in feed increased antioxidant capacity and immunological capacity and attenuated the inflammatory response by increasing the activity of T-SOD, GPx, CAT and the levels of anti-inflammatory cytokines (IL-10 and TGF-ß) and decreasing the levels of MDA and anti-inflammatory cytokines (TNF-α) (P<0.05). Furthermore, TB supplementation improved immunity by increasing the levels of immunoglobulins (IgM and IgG), C3 and IFN-γ (P<0.05). Surprisingly, 0.06%-0.12% TB supplementation significantly increased the content of IL-1ß (P<0.05). However, TB supplementation in feed had no significant effects on the plasma content of GSH, HSP70, IL-8 and the activity of T-AOC (P>0.05). The possible mechanism was that TB activated PI3K/Akt/Nrf2 and inhibits the NF-κB signaling pathway, further regulating the mRNA levels of key genes with antioxidant capacity and the inflammatory response; for example, it increased the mRNA levels of Nrf2, Cu/Zn-SOD, HO-1, CAT, Akt, PI3K, GPx, IL-10, and TGF-ß and decreased the mRNA levels of NF-κB and TNF-α (P<0.05). In addition, 0.06%-0.15% TB supplementation significantly increased the mRNA levels of IL-1ß (P<0.05). TB supplementation in feed had no significant effects on the mRNA levels of HSP70, Mn-SOD and IL-8 (P>0.05). Evidence was presented that TB supplementation decreased the mortality rate caused by Aeromonas hydrophila challenge. In pathological examination, TB supplementation prevented hepatic and intestinal damage. Generally, TB supplementation improved the growth performance of juvenile blunt snout bream. Furthermore, TB supplementation activated PI3K/Akt/Nrf2 and inhibited the NF-κB signaling pathway, regulating health status and preventing hepatic and intestinal damage.

Molecular cloning and functional characterization of the hypoxia-inducible factor-1α in bighead carp (Aristichthys nobilis).

HIF-l is the earliest documented and most widely studied hypoxia-inducible factor (HIF) and plays a key role in the cell hypoxia signal transduction pathway. Particularly, the HIF-1α protein is sensitive to oxygen and plays a critical role in hypoxia regulation. This study is the first to report on the molecular cloning and characterization of HIF-1α in bighead carp (Aristichthys nobilis; anHIF-1α). The full-length cDNA of anHIF-1α was 2361 bp, and encodes an estimated 674 amino acids with a predicted molecular mass of 76.10 kDa and a theoretical isoelectric point of 7.72. Moreover, the conserved basic Helix-Loop-Helix domain along with two Per-ARNT-Sim domains (A/B), and C-TAD were identified in this protein. Interestingly, the tertiary structure of the anHIF-1α protein was found to be extremely similar to that of mice. Multiple comparison and phylogenetic tree results demonstrated that anHIF-1α was highly conserved. Under normoxic conditions, anHIF-1α mRNA transcripts could be detected in all tissues examined with the highest expression level in the heart. With gradually decreasing oxygen concentrations, anHIF-1α mRNA level was upregulated significantly in the gill, liver, kidney, spleen, intestine, brain, and muscle tissues (P < 0.05). Similarly, anHIF-1α was expressed in all examined bighead carp tissues, and the results suggested that the upregulation of anHIF-1α at the transcriptional level may be an important stress response adaptation to hypoxia in bighead carp. Finally, based on the tertiary structure comparative analyses between anHIF-1α with mouse HIF-1α, we think the physiological function, and protein structure of HIF-1α could be compared between fish and mammal in the future.

Transcriptomic analysis of oligochaete immune responses to myxosporeans infection: Branchiura sowerbyi infected with Myxobolus cultus.

The Myxozoa are endoparasites characterized by a two-host life cycle that typically involves invertebrates and vertebrates as definitive and intermediate hosts, respectively. However, little is known about invertebrate-myxosporean interactions, particularly about patterns of host immune defense. We used RNA-sequencing to identify genes that are possibly involved in the immune responses of the oligochaete Branchiura sowerbyi naturally infected with Myxobolus cultus. De novo assembly of the B. sowerbyi transcriptome yielded 119,031 unigenes, with an average length of 896 bp and an N50 length of 1754 bp. Comparative transcriptome analysis revealed 4059 differentially expressed genes (DEGs) between M. cultus-infected and uninfected B. sowerbyi groups, including 3802 upregulated genes and 257 downregulated genes. Among the B. sowerbyi immune factors implicated in the responses to M. cultus infection, DEGs related to lectins, ubiquitin-mediated proteolysis, phagocytosis, oxidative-antioxidative responses, proteases, and protease inhibitors were upregulated. The expression of some immune-related molecules such as calmodulin, heat shock proteins, antimicrobial peptides, lysenin, and serum amyoid A protein were also significantly upregulated. The expression patterns of 14 immune-related DEGs identified by RNA-seq were validated by quantitative real-time polymerase chain reaction. This study is the first attempt to characterize the B. sowerbyi transcriptome and identify immune-related molecules possibly associated with M. cultus infection. It is also the first report of invertebrate host-myxosporean interactions at the transcriptomic level. Our results will facilitate the elucidation of adaptive evolution mechanisms of myxosporean parasites in the definitive host and the genetic basis for differences in resistance of invertebrate hosts of different genotypes to a myxosporean species.

Effects of dietary fenugreek seed extracts on growth performance, plasma biochemical parameters, lipid metabolism, Nrf2 antioxidant capacity and immune response of juvenile blunt snout bream (Megalobrama amblycephala).

Dietary administration of some plant-derived substances have been proved of great economic value in aquaculture. In order to investigate the effects of dietary fenugreek seed extracts (FSE) on juvenile blunt snout bream (Megalobrama amblycephala), a feeding trial was conducted for 8 weeks. The results showed that final weight (FW), weight gain (WG), feed conversion ratio (FCR) and specific growth rate (SGR) were not significantly affected by dietary FSE levels. The whole body lipid contents of fish fed with 0.04%, 0.08% and 0.16% FSE diets were significantly lowered compared to the control group. Dietary FSE diets significantly affected plasma complement component 3 (C3), immunoglobulin M (IgM), albumin (ALB) and total protein (TP). The relative expressions of acetyl CoA carboxylase (ACC), fatty acid synthase (FAS) and sterol regulatory element binding protein-1 (SREBP1) mRNA in the liver of fish decreased significantly with increasing dietary FSE levels from 0% up to 0.04%. FSE supplementation diets lowered the liver pro-inflammatory genes expressions by regulating tumor necrosis factor-α (TNF-α) and interleukin 8 (IL-8) mRNA levels and increased anti-inflammatory genes expression by regulating transforming growth factor (TGF-ß) and interleukin 10 (IL-10). FSE diets increased growth factor-1 (IGF-1) and target of rapamycin (TOR) mRNA levels from 0% up to 0.04%, 0.04% FSE diets significantly increased growth factor-1 (IGF-1) mRNA levels and S6 kinase-polypeptide 1 (S6K1) mRNA levels compared to the control group. 0.04% FSE diets significantly increased superoxide dismutase (SOD) activities and 0.08% FSE diets significantly increased catalase (CAT) and glutathione peroxidase (GPx) activities, 0.16% FSE diets significantly increased total antioxidant capacity (T-AOC) activities compared to the control group. Additionally, compared to the control group, 0.04% dietary FSE significantly up-regulated nuclear factor erythroid 2-related factor 2 (Nrf2) mRNA levels and glutathione peroxidase-1 (GPx1) mRNA levels, at the same time, 0.02%, 0.04%, 0.08%, 0.16% FSE diets significantly down-regulated kelch-like ECH-associated protein 1 (Keap1) mRNA levels. However, no significant effects were observed on copper zinc superoxide dismutase (Cu/Zn-SOD) and manganese superoxide dismutase (Mn-SOD). Our study indicated that dietary FSE could improve plasma biochemical parameters, regulate lipid metabolism related genes, promote Nrf2 antioxidant capacity and enhance immune response of juvenile blunt snout bream.

Effects of dietary tryptophan levels on antioxidant status and immunity for juvenile blunt snout bream (Megalobrama amblycephala) involved in Nrf2 and TOR signaling pathway.

Dietary administration of tryptophan has been proved improving growth performance of fish. An 8-week feeding trial was conducted to investigate the effects of dietary tryptophan level on antioxidant capacity and immune response through Nrf2 and TOR signaling pathway. The results showed that, 0.08% tryptophan level significantly increased plasma aspartate aminotransferase (AST), while immunoglobulin M (IgM) and alkaline phosphatase (ALP) were strikingly increased by 0.40% level. The level of plasma complement component 3 (C3), alanine aminotransferase (ALT) and albumin (ALB) were independent of tryptophan supplementation. Total superoxide dismutase (T-SOD), catalase (CAT), total antioxidant capacity (T-AOC) and glutathione (GSH) activity were increased with increasing dietary tryptophan level until 0.40% and then decreased, while the level of malondialdehyde (MDA) showed a reverse trend. 0.19% and 0.28% tryptophan level significantly improved the glutathione peroxidase 1 (GPx-1) activity. Compared with 0.08% dietary tryptophan level, 0.40% level significantly improved nuclear factor erythroid 2-related factor 2 (Nrf2), GPx, manganese superoxide dismutase (Mn-SOD), CAT and transforming growth factor-ß (TGF-ß) mRNA level, while Kelch-like ECH-associated protein 1 (Keap1) and interleukin 1ß (IL-1ß) mRNA level were significantly decreased. The relative expression of copper zinc superoxide dismutase (Cu/Zn-SOD), heme oxygenase-1 (HO-1), target of rapamycin (TOR), phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K), protein kinase B (Akt) and interleukin 10 (IL-10) were significantly improved by 0.28% diet, while the mRNA level of tumor necrosis factor-α (TNF-α) and nuclear factor-kappa B (NF-κB) were increased by 0.08% diet. Interleukin 8 (IL-8) mRNA level was not significantly affected by dietary tryptophan. Based on MDA and T-SOD value, the optimal dietary tryptophan level of juvenile blunt snout bream was determined to be 0.33% (1.03% of dietary protein) and 0.36% (1.13% of dietary protein), respectively, using quadratic regression analysis.

Morphological variation in Myxobolus drjagini (Akhmerov, 1954) from silver carp and description of Myxobolus paratypicus n. sp. (Cnidaria: Myxozoa).

There is uncertainty in the identification of Myxobolus drjagini, the causative agent of silver carp twist disease, in the literature. An investigation of fish parasites in Lake Taihu, China, revealed several Myxobolus drjagini-like myxosporeans infecting the subcutaneous tissue of the head skin, the olfactory and oculomotor nerves in the cranial cavity, and the intrafilamental epithelium of the gills of silver carp Hypophthalmichthys molitrix (Valenciennes, 1844). Myxospores from the head skin and the nerves were identified as conspecific to M. drjagini based on morphological and molecular data; although the spores from each of the two organs presented morphological variations. SSU rDNA sequence analysis revealed that the sequence of M. drjagini previously deposited in GenBank (AF085179) was invalid. Myxospores from the gills were identified as Myxobolus paratypicus n. sp. The spores were oval, asymmetric in frontal view, 13.8 (12.9-14.9) µm long, 9.9 (9.2-11.1) µm wide, and 7.0 µm thick. Two pyriform polar capsules were unequal in size (ratio above 4:1) with slightly converging anterior ends, and the posterior end of the large polar capsule extended beyond the middle of the spore. The large polar capsule was 7.5 (6.2-8.2) µm long and 5.0 (4.2-5.6) µm wide; the small polar capsule was 2.7 (2.1-3.6) µm long and 1.4 (1.1-1.9) µm wide. Polar filaments were coiled with 7-8 turns in the large polar capsule. The SSU rDNA sequence of M. paratypicus n. sp. was not identical to that of any myxozoan available in GenBank and showed highest similarity with M. drjagini (96%) and Myxobolus pavlovskii (95%) collected from bighead carp and silver carp, respectively.