Lysozyme Gene Research Articles

Constitutively-expressed and induced immune effectors in the house fly (Musca domestica) and the transcription factors that may regulate them.

Insects possess both infection-induced and constitutively expressed innate immune defences. Some effectors, such as lysozymes and antimicrobial peptides (AMPs), are constitutively expressed in flies, but expression patterns vary across tissues and species. The house fly (Musca domestica L.) has an impressive immune repertoire, with more effector genes than any other flies. We used RNA-seq to explore both constitutive and induced expression of immune effectors in flies. House flies were fed either Pseudomonas aeruginosa or Escherichia coli, or sterile control broth, and gene expression in the gut and carcass was analysed 4h post-feeding. Flies fed either bacterium did not induce AMP expression, but some lysozyme and AMP genes were constitutively expressed. Prior transcriptome data from flies injected with bacteria also were analysed, and these constitutively expressed genes differed from those induced by bacterial injection. Binding sites for the transcription factor Myc were enriched upstream of constitutively expressed AMP genes, while upstream regions of induced AMPs were enriched for NF-κB binding sites resembling those of the Imd-responsive transcription factor Relish. Therefore, we identified at least two expression repertoires for AMPs in the house fly: constitutively expressed genes that may be regulated by Myc, and induced AMPs likely regulated by Relish.

Sex and reproductive development impact skin mucosal epithelium immunity, antimicrobial capacity, and up-regulation of immune-related gene of goldfish (Carassius auratus)

This study evaluated the epidermis mucosal capacity of goldfish (Carassius auratus) during different stages of reproductive development in both females and males. In this regard, the activity of mucolytic immune enzymes, i.e., lysozyme, complement and peroxidase, as well as the activity of alkaline phosphatase (ALP) were evaluated. There were five stages for females i.e., immature (f1), cortical alveoli (f2), early and late-vitellogenesis (vtg) (f3 and f4) and ripe (f5); as well as two stages for males spermatogenesis (m1) and spermiation (m2). Some stages were also examined for the mucosal antimicrobial activity against specific pathogens. The results showed that the mucosal lysozyme activity increased significantly during vitellogenesis (P < 0.05), but no lysozyme activity was detected in plasma. On the contrary, the complement activity was only observed in female plasma, and it was significantly higher at f3 compared to the other developmental stages. Both the plasma and mucosal ALP and peroxidase activities showed a significant increase by female reproductive development with the highest amounts at f4. Contrary to the female, no significant changes were observed in plasma and mucosal immune agents and biochemistry of the male. The f5-staged goldfish showed the highest antimicrobial activities against Gram-positive bacteria, i.e., Streptococcus faecium, Staphylococcus aureus and Micrococcus luteus (P < 0.05). Our results also represented the up-regulation of lysozyme (c-lys) gene expression by effects of female maturational development in ovary, liver and skin, while male goldfish showed no significant changes in c-lys expression. Moreover, there were positive correlations between c-lys expression, mucosal lysozyme activity and calcium levels in females (P < 0.01). Overall, our findings revealed that vtg process improves mucosal innate immunity that leads to activate antimicrobial components at spawning season.

Molecular characterization and antibacterial activities of a goose-type lysozyme gene from roughskin sculpin (Trachidermus fasciatus)

Lysozymes, acting as antimicrobial molecules, play a vital role in the host's innate immune response to pathogen infections. In the present study, a g-type lysozyme gene termed Tf-LyzG from roughskin sculpin, Trachidermus fasciatus was firstly reported. The deduced amino acid sequence of Tf-LyzG contained 188 residues and possessed conserved catalytic residues (Glu71, Asp84, and Asp95). Gene expression analysis revealed that Tf-LyzG was widely distributed in the tested eleven tissues with the highest expression in the gill and could be significantly induced post lipopolysaccharide (LPS) challenge. The lysozyme activity of the purified recombinant protein (rTf-LyzG) was found to be most active at pH 5.5 and 37 °C. rTf-LyzG exhibited a wide spectrum of potent bacteriolytic activity against four Gram-positive bacteria and six Gram-negative bacteria. It also displayed a high affinity to polysaccharides on bacteria surfaces including LPS, lipoteichoic acid (LTA), and peptidoglycan (PGN). rTf-LyzG was capable of binding and agglutinating all nine bacteria. Flow cytometry assay further revealed that rTf-LyzG could disrupt the membrane of Micrococcus lysodeikticus which is confirmed by scanning electron microscope (SEM) analysis that reveals blebs around the bacterial cell membrane. In summary, these data indicate that Tf-LyzG is of great importance in the fish immune response against pathogens invasion.

Supplementing dietary selenium nano-particles increased growth, antioxidant capacity and immune-related genes transcription in Pacific whiteleg shrimp (Penaeus vannamei) juveniles

An eight-week nutritional study was carried out to examine the effects of dietary selenium nanoparticles (Se-N) on performance in whiteleg shrimp (Penaeus vannamei) juveniles (1.55 ± 0.04 g, means ± standard deviation). A basal diet (∼ 38 % crude protein, 18.5 kJ/g crude energy) was supplemented with Se-N at five levels, including 0 (control), 0.2, 0.4, 0.8, and 1.2 mg/kg. Three hundred shrimps were distributed randomly into 15 circular fiberglass tanks containing 150-L seawater (20 shrimp/tank). Each dietary treatment was offered to shrimps in three tanks. Shrimps were handfed three times daily based on 7 % of their initial biomass. Water temperature and salinity were 30 ± 0.4 °C and 40 g/L, respectively. The hepatopancreas (HP), hemolymph, and gut were sampled after 56 d of the feeding trial. Shrimps fed Se-N supplemented diets had positive quadratic trends in growth performance (P = 0.011) and feed conversion ratio (P = 0.006). Selenium level increased in the shrimp’s whole body with increasing Se-N in diet and showed both linear and quadratic treands. The activities of plasma phenoloxidase (PO) and, catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx) in the HP were increased by supplementing diet with Se-N and showed positive quadratic treands. The malondialdehyde (MDA) level significantly decreased in the HP of shrimps fed Se-N and had a quadratic treand (P = 0.012). Both linear and quadratic trands were noticed in the relative abundance of prophenoloxidase, lysozyme and penaeidin-3α genes mRNA transcript in the HP of shrimps fed Se-N supplemented diets. However, heat shock protein 70 gene transcript did not show any trend by supplementing Se-N in the experimental diets. In conclusion, based on the second-order polynomial regression analysis the optimum level of Se-N supplementation in a diet for P. vannamei juveniles was estimated to be 0.38 mg/kg diet. Furthermore, extra levels of dietary Se-N decreased growth rate and PO activity that in coincided with the reduction of CAT and GPx activities and elevation of MDA level in HP, indicating that high levels of Se-N in diet can induce oxidative stress in P. vannamei.

Chicken-type lysozyme is a major bacteriolytic enzyme in the blood of the banded houndshark Triakis scyllium

We examined lysozyme activities in the serum and the leukocyte extracts of the banded houndshark Triakis scyllium. The serum exhibited lytic activity, but not the leukocyte extracts. The lytic substance in the serum was of approximately 14 kDa and the N-terminal amino acid sequence was YVYSK. cDNA cloning identified a C-type lysozyme (TsLysC) gene and two G-type lysozyme (TsLysG) cDNA clones of different lengths. The TsLysC gene encodes 149 amino acids residues, and the sequence derived from the N-terminal amino acid sequencing was displayed at position 17–21. TsLysG, on the other hand, contains two ORFs that are homologous to the N- and C-terminal regions of G-type lysozyme of other fish species. TsLysC mRNA levels were high in the liver. TsLysG mRNA level was significantly lower than TsLysC mRNA in the liver.

Lysozyme amyloidosis—a report on a large German cohort and the characterisation of a novel amyloidogenic lysozyme gene variant

Lysozyme-derived (ALys) amyloidosis is a rare type of hereditary amyloidosis. Nine amyloidogenic variants and ∼30 affected families have been described worldwide. The most common manifestations are renal dysfunction, gastrointestinal tract symptoms, and sicca syndrome. We report on the clinical course of ten patients from six families representing one of the largest cohorts published so far. Seven patients carried the W64R variant showing the whole spectrum of ALys-associated symptoms. Two patients—a mother-son pair—carried a novel lysozyme variant, which was associated with nephropathy and peripheral polyneuropathy. In accordance with previous findings, the phenotype resembled within these families but did not correlate with the genotype. To gain insights into the effect of the variants at the molecular level, we analysed the structure of lysozyme and performed comparative computational predictions on aggregation propensity and conformational stability. Our study supports that decreased conformational stability is a key factor for lysozyme variants to be prone to aggregation. In summary, ALys amyloidosis is a very rare, but still heterogeneous disease that can manifest at an early age. Our newly identified lysozyme variant is associated with nephropathy and peripheral polyneuropathy. Further research is needed to understand its pathogenesis and to enable the development of new treatments.

C/N-Dependent Element Bioconversion Efficiency and Antimicrobial Protein Expression in Food Waste Treatment by Black Soldier Fly Larvae.

The black soldier fly (BSF), Hermetia illucens, has emerged as a promising species for waste bioconversion and source of antimicrobial proteins (AMPs). However, there is a scarcity of research on the element transformation efficiency and molecular characterization of AMPs derived from waste management. Here, food waste treatment was performed using BSF larvae (BSFL) in a C/N ratio of 21:1–10:1, with a focus on the C/N-dependent element bioconversion, AMP antimicrobial activity, and transcriptome profiling. The C-larvae transformation rates were found to be similar among C/Ns (27.0–35.5%, p = 0.109), while the N-larvae rates were different (p = 0.001), with C/N 21:1–16:1 (63.5–75.0%) being higher than C/N 14:1–10:1 (35.0–45.7%). The C/N ratio did not alter the antimicrobial spectrum of AMPs, but did affect the activities, with C/N 21:1 being significantly lower than C/N 18:1–10:1. The lysozyme genes were found to be significantly more highly expressed than the cecropin, defensin, and attacin genes in the AMP gene family. Out of 51 lysozyme genes, C/N 18:1 and C/N 16:1 up-regulated (p < 0.05) 14 and 12 genes compared with C/N 21:1, respectively, corresponding to the higher activity of AMPs. Overall, the element bioconversion efficiency and AMP expression can be enhanced through C/N ratio manipulation, and the C/N-dependent transcriptome regulation is the driving force of the AMP difference.

Evaluating the mixture of earthworm meal and wormcast as a protein source for common carp (Cyprinus carpio L.) based on growth performance, antioxidant, immune capacity, lipid metabolism and intestinal health

This study was conducted to evaluate the effects of a dietary earthworm meal mixture (earthworm meal: EM and wormcast: WC) for replacing fish meal (FM) on growth performance, antioxidant capacity, lipid metabolism, and intestinal health of common carp (Cyprinus carpio L.). FM was used as the protein source in the control diet (D1), while 30% FM protein was replaced with an EM and WC mixture in the experimental diets, and the specific mixing ratio was as follows: D2 (15.4% EM), D3 (15.1% EM and 1.8% WC), D4 (14.8% EM and 3.7% WC), and D5 (14.5% EM and 5.8% WC). The growth performance was significantly decreased in the D4 and D5 groups compared with the D1 group, while the condition factor was highest in the D2 group (P < 0.05). In the antioxidant and immune respects, the expression levels of the sod and cat genes in hepatopancreas were remarkably upregulated in the D2, D3, and D4 groups (P < 0.05). Furthermore, serum lysozyme activities and lysozyme gene mRNA levels in head kidney reached the maximum in the D2 group. Additionally, the expression levels of interleukin 10 (il-10) and transforming growth factor β (tgf-β) in head kidney were enhanced in the D2 group (P < 0.05). For lipid metabolism, the expression levels in hepatopancreas of carnitine palmitoyl transferase 1 (cpt-1) were upregulated, while fatty acid synthase (fas) was significantly downregulated in the D4 and D5 groups (P < 0.05). In terms of intestinal health, dietary EM and WC increased the diversity and abundance of intestinal microbiota. The results showed no negative effect of the dietary earthworm meal mixture on growth performance after 30% FM protein was replaced by 15.4% EM or the 15.1% EM and 1.8% WC mixtures, which was beneficial for the health status of treated common carp.

Characterization and functional analysis of a c-type lysozyme gene from obscure puffer Takifugu obscurus

Lysozyme (Lyz) is an alkaline enzyme that hydrolyzes mucopolysaccharides in bacteria and is highly conserved vertebrates and invertebrates. In this study, a c-type lysozyme gene (named ToLyzC) from the obscure puffer Takifugu obscurus was cloned and characterized. The full-length cDNA of ToLyzC was 432 bp, encoding 143 amino acids, with a predicted molecular mass of 16.2 kDa and a theoretical pI of 8.86. The depicted protein sequence contained a LYZ1 domain from 16 to 142 amino acids, seven conserved cysteine residues. Phylogenetic analysis indicated that ToLyzC clustered with Lyzs from other teleost fishes. Quantitative real-time PCR analysis revealed that ToLyzC mRNA was mainly expressed in the liver. The transcript level of ToLyzC gene was significantly upregulated after Staphylococcus aureus and Vibrio harveyi challenge. The optimal pH and temperature of recombinant ToLyzC protein (rToLyzC) lytic activity was detected to be 7.5 and 35 °C, respectively. rToLyzC exhibited significant antibacterial and bacterial binding activities against S. aureus, Aeromonas hydrophila, V. harveyi, and Edwardsiella tarda at different time points. In addition, the morphological changes of V. harveyi cells treated with rToLyzC were observed under scanning electron microscope, which further confirmed the antibacterial and bacteriolytic activity of rToLyzC. Taken together, our current study indicated that ToLyzC is involved in the immune response to bacterial infection in obscure puffers.

Dietary Dracocephalum kotschyi essential oil improved growth, haematology, immunity and resistance to Aeromonas hydrophila in rainbow trout ( Oncorhynchus mykiss )

In the present study, the effect of Dracocephalum kotschyi essential oil (DKE) was investigated on growth, haematology, immune and antioxidant defence system and resistance of rainbow trout juveniles to bacterial infection (Aeromonas hydrophila). For this purpose, the fish were fed a diet containing different concentrations of DKE including 0 (control), 0.2, 0.25 and 0.3 mg/kg diet in three replicates for 60 days. After feeding trial, the fish were challenged with a pathogenic dose of A. hydrophila. Based on the results, immune components in plasma (alternative complement activity [ACH50], IgM content, lysozyme activity, total protein and total albumin) and mucus (protease activity, IgM content and lysozyme activity) significantly elevated in fish fed diet containing 0.2 and 0.25 mg DKE/kg diet compared with other groups (p < 0.01). The plasma bactericidal activity increased in all DKE-supplemented fish. Supplementation of fish with 0.3 mg DKE/kg diet depressed the mucosal lysozyme activity and IgM content. The liver antioxidant enzymes, glutathione peroxidase (GPx), catalase (CAT) and superoxide dismutase (SOD) showed increased activity in response to 0.25 and 0.3 mg DKE/kg diet compared with other experimental diets (p < 0.01). Dietary DKE changed the haematology of the fish. The haematocrit, red blood cell count (RBC), white blood cell (WBC), haemoglobin content and mean erythrocyte cell haemoglobin content (MCHC) increased in response to DKE. In contrast, the mean corpuscular haemoglobin (MCH) decreased in fish fed 0.2 and 0.25 mg DKE/kg diet compared with other diets (p < 0.01). The supplementation of fish with 0.2 and 0.25 mg DKE/kg diet decreased the expression of cytokine-related genes, TNF-α and IL-8. In contrast, the DKE up-regulated the expression of the immune-related genes, C3 and lysozyme. DKE at concentration of 0.3 mg/kg diet depressed the C3 and lysozyme gene expressions. The DKE supplementation decreased the mortality rate of the fish after bacterial challenge compared with non-supplemented ones (p < 0.01). In conclusion, the findings of this study revealed the immune-stimulating effects of DKE at optimized dietary concentrations. In addition, DKE decreased the mortality induced by A. hydrophila infection.

Transformation of food waste to source of antimicrobial proteins by black soldier fly larvae for defense against marine Vibrio parahaemolyticus

Insect biorefinery by black soldier fly larvae (BSFL), Hermetia illucens, has emerged as an innovative technique for the valorization of food waste. However, despite BSFL being an attractive natural source of antimicrobial proteins (AMPs), there is a scarcity of research on the antimicrobial activity and transcriptome expression of AMPs derived from BSFL following waste treatment. In the present study, food waste treatment was performed by BSFL with a substrate C/N ratio ranging from 21:1 to 10:1, marine Vibrio parahaemolyticus (VP) was selected as the model aquaculture pathogen, the antimicrobial activities of AMPs in vitro and zebrafish in vivo were examined, and the molecular mechanism of the C/N-dependent AMP difference was expounded. Findings were made that the AMP extract of C/N16:1 resulted in relatively higher antimicrobial activity in vitro than that of other C/Ns. Further, the AMPs of C/N16:1 exhibited a promising in vivo defense effect for elevating the 96-h survival rate of zebrafish from 0% to 39% after VP infection, comparable to the animal antibiotic sulfamethoxidine. The results of transcriptome analysis reveal that lysozymes were the highest expressed components in the AMP gene family. The C/N16:1 BSFL significantly up-regulated 12 out of 51 lysozyme genes compared with C/N21:1, which likely contributed to the improvement of AMP antimicrobial activity. Further, C/N16:1 significantly up-regulated the expression of lysozyme, glycosyl hydrolase and muscle protein genes compared with C/N21:1, which likely enhanced the defense ability of the immune system, the utilization of the starch-like substrate, and the mobility of the larvae, thereby facilitating the larval transformation and AMP production. Overall, such results indicate that waste C/N ratio interacted with the activity and expression of BSFL AMPs through transcriptome regulation, and the BSFL AMPs derived from food waste could be used for the defense against marine pathogens to support the sustainable development of aquaculture.

Acute ammonia exposure combined with heat stress impaired the histological features of gills and liver tissues and the expression responses of immune and antioxidative related genes in Nile tilapia

Ammonia exposure can be considered more stressful for aquatic animals when it coincides with high temperature. This study was conducted to detect the effects of ammonia exposure and heat stress and their interactions on the histological features of gills and liver tissues and the expression responses of immune and antioxidative related genes in Nile tilapia. Thus, 180 fish were divided into four groups (triplicates), where the first and third groups were kept in clean water without total ammonium nitrogen (TAN) exposure. At the same time, the second and fourth groups were exposed to 5 mg TAN/L. After seven days, the water temperature was raised in the third (without ammonia toxicity) and fourth (exposed with 5 mg TAN/L) groups up to 32 °C and kept under these conditions for 24 h. While the first (without ammonia toxicity) and second (exposed with 5 mg TAN/L) groups were kept under optimum water temperature (27.28 °C) then gills and liver tissues were dissected. Marked upregulation of keap1 was seen in the gills of fish exposed to ammonia/heat stress. The expression of mRNA levels for nrf2, nqo-1, cat, and gpx genes were downregulated in all stressed groups, with the lowest was recorded in the ammonia/heat stress group. The transcription of ho-1 was upregulated in the ammonia and heat stress groups while downregulated in the ammonia/heat stress group. The transcription of the complement C3 gene was downregulated in the livers of heat stress and ammonia/heat stress groups, while the lysozyme gene was downregulated in the ammonia/heat stress group. The mRNA expression levels of nf-κB, il-1β, and tnf-α genes were higher in the ammonia group than in the heat stress group. The highest transcription level of nf-κB, il-1β, tnf-α, il-8, and hsp70 genes and the lowest C3 and lysozyme genes were observed in fish exposed to ammonia/heat stress. The co-exposure to ammonia with heat stress triggered degeneration of primary and secondary gill filaments with telangiectasia and vascular congestion of secondary epithelium while, the liver showed hepatic vascular congestion and visible necrotic changes with nuclear pyknosis. In conclusion, the combined exposure of ammonia and heat stress induced oxidative stress, immunosuppression, and inflammation in Nile tilapia.

Identification of allelic variants, differential expression, and in silico analysis of caprine serum lysozyme gene

Identification of allelic variants, differential expression, and in silico analysis of caprine serum lysozyme gene

Probiotic Bacillus safensis NPUST1 Administration Improves Growth Performance, Gut Microbiota, and Innate Immunity against Streptococcus iniae in Nile tilapia (Oreochromis niloticus).

Probiotics are considered ecofriendly alternatives to antibiotics as immunostimulants against pathogen infections in aquaculture. In the present study, protease-, amylase-, cellulase-, and xylanase-producing Bacillus safensis NPUST1 were isolated from the gut of Nile tilapia, and the beneficial effects of B. safensis NPUST1 on growth, innate immunity, disease resistance and gut microbiota in Nile tilapia were evaluated by feeding tilapia a basal diet or basal diet containing 105 and 106–107 CFU/g for 8 weeks. The results showed that the weight gain, feed efficiency and specific growth rate were significantly increased in tilapia fed a diet containing 106 CFU/g and 107 CFU/g B. safensis NPUST1. Intestinal digestive enzymes, including protease, amylase and lipase, and hepatic mRNA expression of glucose metabolism and growth-related genes, such as GK, G6Pase, GHR and IGF-1, were also significantly increased in the 106 CFU/g and 107 CFU/g B. safensis NPUST1 treated groups. Immune parameters such as phagocytic activity, respiratory burst and superoxide dismutase activity in head kidney leukocytes, serum lysozyme, and the mRNA expression of IL-1β, IL-8, TNF-α and lysozyme genes were significantly induced in the head kidney and spleen of 106 CFU/g and 107 CFU/g B. safensis NPUST1 treated fish. The cumulative survival rate was significantly increased in fish fed a diet containing 106 CFU/g and 107 CFU/g B. safensis NPUST1 after challenge with Streptococcus iniae. Dietary supplementation with B. safensis NPUST1 improves the gut microbiota of Nile tilapia, which increases the abundance of potential probiotics and reduces the abundance of pathogenic pathogens. The present study is the first to report the use of B. safensis as a potential probiotic in aquaculture, and a diet containing 106 CFU/g B. safensis NPUST1 is adequate for providing beneficial effects on growth performance and health status in tilapia.

Evolutionary genomics of APSE: a tailed phage that lysogenically converts the bacterium Hamiltonella defensa into a heritable protective symbiont of aphids

BackgroundMost phages infect free-living bacteria but a few have been identified that infect heritable symbionts of insects or other eukaryotes. Heritable symbionts are usually specialized and isolated from other bacteria with little known about the origins of associated phages. Hamiltonella defensa is a heritable bacterial symbiont of aphids that is usually infected by a tailed, double-stranded DNA phage named APSE.MethodsWe conducted comparative genomic and phylogenetic studies to determine how APSE is related to other phages and prophages.ResultsEach APSE genome was organized into four modules and two predicted functional units. Gene content and order were near-fully conserved in modules 1 and 2, which encode predicted DNA metabolism genes, and module 4, which encodes predicted virion assembly genes. Gene content of module 3, which contains predicted toxin, holin and lysozyme genes differed among haplotypes. Comparisons to other sequenced phages suggested APSE genomes are mosaics with modules 1 and 2 sharing similarities with Bordetella-Bcep-Xylostella fastidiosa-like podoviruses, module 4 sharing similarities with P22-like podoviruses, and module 3 sharing no similarities with known phages. Comparisons to other sequenced bacterial genomes identified APSE-like elements in other heritable insect symbionts (Arsenophonus spp.) and enteric bacteria in the family Morganellaceae.ConclusionsAPSEs are most closely related to phage elements in the genus Arsenophonus and other bacteria in the Morganellaceae.

Effects of salinity on gills' chloride cells, stress indices, and gene expression of Asian seabass (Lates calcarifer, Bloch, 1790).

A 2-week research was carried out to assess water salinity (WS) effects including 0, 15, 35, and 50‰ on osmoregulatory mechanisms and stress indices in Asian sea bass (34.4g) juveniles. Except for fish reared at 50‰, in the other treatments, it gradually decreased to the prescribed WS during a 10-day period (- 5‰ a day). After a 10-day acclimation period, fish were reared at the prescribed WS for 2weeks. Fish reared at 15 and 35‰ had higher chloride cell (CC) counts in the interlamellar region. The number of CC in the interlamellar region elevated with increment of WS up to 35‰, but they were pronouncedly reduced in 50‰ group. The diameter of CC in the interlamellar region was not affected by WS. The smallest nucleus diameter of CC in the interlamellar region was observed in fish reared at 15‰ (P < 0.05). The largest and the smallest amounts of serum aspartate aminotransferase were observed in fish reared at freshwater and 15‰, respectively. Fish reared at 35‰ had the highest serum sodium and potassium contents. Serum chloride content and total osmolality increased with increment of WS (P < 0.05). Serum cortisol and glucose contents gradually increased with elevation of WS up to 35‰; then, their contents remarkably decreased. The relative expression of insulin like growth factor-1 in the liver of fish reared at 35‰ was strikingly higher than that in the other groups. The relative expression of HSP70 gene in fresh water group was pronouncedly elevated compared to other treatments. The relative expression of interleukin-1β in 15 and 35‰ groups was higher than that in the other groups; however, the relative expression of lysozyme gene in the liver of fish reared at fresh water was pronouncedly lower than that in the other treatments. The results of this study suggested rearing L. calcarifer at 15‰ closer to the isosmotic point and better provide its welfare.

Various carbon/nitrogen ratios in a biofloc-based rearing system of common carp (Cyprinus carpio) fingerlings: Effect on growth performance, immune response, and serum biochemistry

In this study, three various carbon to nitrogen ratios including 10:1 (C/N-10), 15:1 (C/N-15), and 20:1 (C/N-20) were designed in a biofloc based system with three replicates. The initial biofloc was formed by adding 200 g common carp commercial diet +1 g urea (46% N) + 50 g bottom soil of carp earthen pond. After 56 days, the highest growth indices and the lowest feed conversion ratio were obtained in C/N-20 treatment (P < 0.05). The raised fish in C/N-10 system revealed significantly (P < 0.05) higher serum glucose, alanine aminotransferase, and aspartate aminotransferase compared to other treatments. However, alkaline phosphatase activity was decreased by increasing C/N ratios (P < 0.05). In addition, no significant influence (P > 0.05) was detected in the content of albumin and total protein among the treatments. The level of immunoglobulin M did not change significantly (P > 0.05) in C/N-15 and C/N-20 groups, while it was significantly higher than C/N-10 group (P < 0.05). The mRNA expression levels of interleukin-1 beta, interleukin-8, tumor necrosis factor-alpha, and lysozyme genes were up-regulated with increasing in the C/N ratios. The anti-inflammatory interleukin-10 gene expression was notably suppressed in C/N-20 compared to C/N10 (P < 0.05). Based on our results, the humoral immune parameters, immune-related gene expression, and zootechnical responses were notably ameliorated in common carp cultured in a biofloc system at a C/N ratio of 20:1.

Acclimatization of transformed seaweed Kappaphycus alvarezii carrying lysozyme gene in culture flask and the cultivation in floating net cage in Pangkep coastal

The in vitro transformation of the lysozyme gene in seaweed K. alvarezii has been successfully executed to increase the viability against ice-ice disease. There were two major stages in this research; (1) transformation of lysozyme gene in seaweed K. alvarezii which was carried out on laboratory scale and the cultivation of gene-transformed explants in the culture flask stored in “culture chamber”; (2) the acclimatization in floating net cages of green nets (mesh size of 1 mm) with cage size of 50 x 50 x 50 cm, the population density of 200 explants and cultivated for two weeks. The explants were then transferred to blue nets (mesh size of 2 mm) with a cage size of 50 x 50 x 50 cm for four weeks of rearing. The plants were then enlarged using a long-line method in the floating net cage, by tying the seaweed using a double line with a gap of 15 cm each. The measurement of weight, bud lengths, and water quality was carried out within 2 weeks. The result shows that the daily growth rate of the transformed seaweed during the regeneration stage in the culture flask was around 0.33-0.4%/day, meanwhile during the acclimatization stage in the green nets the was 0.65-1.6%/day, and even more, increased during the acclimatization stage in the blue nets with DGR of 2.28-2.3%/day. During the enlargement stage in the floating net cages, the lysozyme-transformed seaweed showed an even higher DGR with a value of 3.2-8.2%/day. The results of the integration of the lysozyme gene in seaweed were indicated by the presence of a 670 bp of amplification products, that is the same total length of the 35 S-F promoter fragments and Nos T-R in the expression vector. Based on these results, the lysozyme gene was successfully transformed in K. alvarezii seaweed.

Dietary supplementation with dandelion (Taraxacum officinale) flower extract provides immunostimulation and resistance against Streptococcus iniae infection in rainbow trout (Oncorhynchusmykiss)

Dandelion flower is a well-known phytomedicine due to its therapeutic effects on dyspepsia, bile duct disorders, spleen and liver complaints, and inflammatory diseases. This study aimed to assess the effect of dietary dandelion flower extract (DFE) on skin mucosal and humoral immunity, immune-related gene expression, and disease resistance in rainbow trout. The fingerlings (N = 300, 20.1 ± 0.9 g) were randomly assigned to 15 tanks and fed with different levels of DEF including 0 (control), 1, 2, 3, and 4 g/kg for 56 days. At the end of the trial, the total leukocyte and lymphocyte counts were significantly enhanced in DFE-added groups compared to the control group. Complement components, immunoglobulin M, total protein, and lysozyme showed significantly higher values in the fish treated with dietary DFE at 3 g/kg than others (P < 0.05). The enzymatic activities of the skin mucus samples in the fish fed dietary levels of DFE were significantly higher than the control fish. Skin mucus protein was also improved significantly in 2 and 3 g/kg DFE compared to others. The transcription levels of interleukin-1β and interleukin-6 genes were up-regulated in the fish fed with 3 and 4 g/kg DFE. Interleukin-8 and lysozyme gene expression levels were elevated in 3 g/kg DFE group compared to the control group. Fish mortality after challenging with Streptococcus iniae was significantly reduced from 43.33% in the control group to 73.33% in 3 g/kg DFE group. This study confirmed the beneficial effects of DFE as an immunostimulant in rainbow trout diet and the recommended dose of DFE is 2.49–2.74 g/kg based on the polynomial regression models.

Arms race in a cell: genomic, transcriptomic, and proteomic insights into intracellular phage\u2013bacteria interplay in deep-sea snail holobionts

BackgroundDeep-sea animals in hydrothermal vents often form endosymbioses with chemosynthetic bacteria. Endosymbionts serve essential biochemical and ecological functions, but the prokaryotic viruses (phages) that determine their fate are unknown.ResultsWe conducted metagenomic analysis of a deep-sea vent snail. We assembled four genome bins for Caudovirales phages that had developed dual endosymbiosis with sulphur-oxidising bacteria (SOB) and methane-oxidising bacteria (MOB). Clustered regularly interspaced short palindromic repeat (CRISPR) spacer mapping, genome comparison, and transcriptomic profiling revealed that phages Bin1, Bin2, and Bin4 infected SOB and MOB. The observation of prophages in the snail endosymbionts and expression of the phage integrase gene suggested the presence of lysogenic infection, and the expression of phage structural protein and lysozyme genes indicated active lytic infection. Furthermore, SOB and MOB appear to employ adaptive CRISPR–Cas systems to target phage DNA. Additional expressed defence systems, such as innate restriction–modification systems and dormancy-inducing toxin–antitoxin systems, may co-function and form multiple lines for anti-viral defence. To counter host defence, phages Bin1, Bin2, and Bin3 appear to have evolved anti-restriction mechanisms and expressed methyltransferase genes that potentially counterbalance host restriction activity. In addition, the high-level expression of the auxiliary metabolic genes narGH, which encode nitrate reductase subunits, may promote ATP production, thereby benefiting phage DNA packaging for replication.ConclusionsThis study provides new insights into phage–bacteria interplay in intracellular environments of a deep-sea vent snail.3bKHUh9dEq7Vm5ZK3_XS13Video