Streptococcus Agalactiae Infection Research Articles

Genome-wide identification of DNA methylation marks associated with resistance to Streptococcus agalactiae in the GIFT strain of Nile Tilapia (Oreochromis niloticus)

Epigenetic mechanism plays a pivotal role in the immune responses of fish. Streptococcus agalactiae (SA) infection is a significant challenge to tilapia aquaculture. DNA methylation involved in immune responses in tilapia against SA is unclear. Here, by using a QTLseq strategy integrated with whole-genome bisulfite sequencing-based DNA methylation analysis (methyl-QTLseq) and RNA sequencing (RNA-seq), we identified 3608 differentially methylated regions (DMRs) and 4355 differentially expressed genes (DEGs) in the spleen of Nile tilapia Oreochromis niloticus. Compared to the susceptible group, the up-regulated DMRs (2449) were approximately twice as many as the down-regulated DMRs (1159). Compared to the susceptible group, lower methylation levels for CpG, CHG and CHH in TEs and for CHG and CHH in genes and significant decreased expression for 8 of the 11 DNA methyltransferase genes in the resistant group were detected. 46 DEGs with differentially methylated promoter regions (DMPs) were significantly enriched in immune processes such as T cell differentiation and adaptive immunity. We detected 4 methyl-QTLs located on LG8:5332501–5,336,400, LG14:2307301–2,353,500, LG16:16889101–16,903,500, and LG22:25238401–2,536,880 across the tilapia genome. The methyl-QTL region located within the chromosome LG22:25238401–25,368,800 contains the most DMRs (91) and DMGs (7). Interestingly, most of the annotated genes that located within or under the peak of the methyl-QTL interval belong to the HOX protein family including hoxa1, hoxa2, hoxa4a and hoxa5a. Bisulfite sequencing PCR and expression analysis indicated that DNA methylation in the promoter regions of hoxa4a, lcp2a, and ccr9a regulated gene expression in spleen in response to SA infection. Overall, our study provides epigenetics evidences involving in immune responses against SA infection in tilapia.

The role of HMGB2 in the immune response of Nile tilapia (Oreochromis niloticus) to streptococcal infection

High mobility group protein B2 (HMGB2) is an abundant chromatin-associated protein with pivotal roles in transcription, cell proliferation, differentiation, inflammation, and tumorigenesis. However, its immune function in Nile tilapia (Oreochromis niloticus) remains unclear. In this study, we identified a homologue of HMGB2 from Nile tilapia (On-HMGB2) and investigated its functions in the immune response against streptococcus infection. The open reading frame (ORF) of On-HMGB2 spans 642 bp, encoding 213 amino acids, and contains two conserved HMG domains. On-HMGB2 shares over 80 % homology with other fish species and 74%–76 % homology with mammals. On-HMGB2 was widely distributed in various tissues, with its highest transcript levels in the liver and the lowest in the intestine. Knockdown of On-HMGB2 promoted the inflammatory response in Nile tilapia, increased the bacterial load in the tissues, and led to elevated mortality in Nile tilapia following Streptococcus agalactiae infection. Taken together, On-HMGB2 significantly influences the immune system of Nile tilapia in response to streptococcus infection.

Effects of hydrogen peroxidase-inactivated vaccine against Streptococcus agalactiae infection in red tilapia (Oreochromis sp.) via different routes of administration

Hydrogen peroxidase-inactivated vaccines have recently developed in controlling infectious diseases in aquaculture. The present study aimed to compare the efficacy of the hydrogen peroxidase-inactivated Streptococcus agalactiae vaccine via different delivery routes (injection, immersion, and oral administration) in red tilapia (Oreochromis sp.) without using adjuvant. Fish were randomly divided into 4 groups including G1: Control treatment (without vaccine), G2: vaccine-based diet (oral administration), G3: vaccinated by immersion and G4: vaccinated by injection. After 6 weeks of the experiment, fish were intraperitoneally injected with S. agalactiae and the mortality was recorded in 14 days. The results showed that lysozyme activity was differentially increased according to the delivery routes of vaccine, organs and time of sampling. However, the specific antibody levels in all vaccinated groups were only increased in week 6 post-vaccination. After the challenge test with S. agalactiae, the serum lysozyme levels in G3 and G4 were significantly higher than the control group (G1), while the total white blood cells and specific antibody levels were significantly increased in G2 and G4 compared to the control (G1). Similarly, the hydrogen peroxidase-inactivated vaccine statistically reduced the cumulative mortality in G2 (35.29%) and G4 (28.95%) compared to G1 (44.12%) after injected with the S. agalactiae. These results showed that the vaccine delivery routes by oral administration or injection may decrease the pathogen and show better protection for red tilapia than the immersion method. Further studies will be investigated to improve the efficacy of hydrogen peroxidaseinactivated vaccine via using different adjuvants.

The HSP70 and IL-1β of Nile tilapia as molecular adjuvants can enhance the immune protection of DNA vaccine against Streptococcus agalactiae infection.

Globally, streptococcal disease caused by Streptococcus agalactiae is known for its high mortality rate, which severely limits the development of the tilapia breeding industry. As a third-generation vaccine, DNA vaccines have shown great application prospects in the prevention and control of aquatic diseases, but their low immunogenicity limits their development. The combination of DNA vaccines and molecular adjuvants proved to be an effective method for inducing protective immunity. This study constructed recombinant plasmids encoding tilapia HSP70 and IL-1β genes (pcHSP70 and pcIL-1β) to verify their effectiveness as molecular adjuvants for S. agalactiae DNA vaccine (pcSIP) in the immunized tilapia model. The results revealed that serum-specific IgM production, enzyme activities, and immune-related gene expression in tilapia immunized with pcSIP plus pcHSP70 or pcIL-1β were significantly higher than those in tilapia immunized with pcSIP alone. It is worth noting that combination with molecular adjuvants improved the immune protection of DNA vaccines, with a relative percentage survival (RPS) of 51.72% (pcSIP plus pcHSP70) and 44.83% (pcSIP plus pcIL-1β), respectively, compared with that of pcSIP alone (24.14%). Thus, our study indicated that HSP70 and IL-1β in tilapia are promising molecular adjuvants of the DNA vaccine in controlling S. agalactiae infection.

Efficacy of the dietary Malva parviflora leaves in modulating immune-antioxidant functions, resistance to Candida albicans and Streptococcus agalactiae infection, digestive/absorptive capacity, and growth of Nile tilapia (Oreochromis niloticus)

Efficacy of the dietary Malva parviflora leaves in modulating immune-antioxidant functions, resistance to Candida albicans and Streptococcus agalactiae infection, digestive/absorptive capacity, and growth of Nile tilapia (Oreochromis niloticus)

Medulla oblongata and NCCs are central defenders against Streptococcus agalactiae infection of the tilapia brain.

Various types of professional immune cells first emerge in fish and likely represent the primordial form and functions. Recent advancements revealed the direct connection between the central nervous system and the immune system in the mammalian brain. However, the specifics of brain-immune networks in the fish and the underlying mechanisms of teleost's brain against pathogen infection have not been fully elucidated. In this study, we investigated the distribution of markers representing cerebral cells associated with protection and professional lymphocytes in the seven major components of the Nile tilapia brain through RNA-Seq assay and observed the most dominant abundance in the medulla oblongata. The subsequent challenge test revealed the non-specific cytotoxic cells (NCCs) exhibited the strongest response against streptococcal infection of the brain. The presence of NCCs in the brain was then confirmed using immunofluorescence and the cytotoxic effects usually induced by NCCs under infection were determined as well. Collectively, these findings contribute significantly to comprehending the mechanism of fish neuroimmune interaction and enhancing our understanding of its evolutionary development.

Vaccines for Streptococcus agalactiae: current status and future perspectives.

A maternal vaccine to protect newborns against invasive Streptococcus agalactiae infection is a developing medical need. The vaccine should be offered during the third trimester of pregnancy and induce strong immune responses and placental transfer of protective antibodies. Polysaccharide vaccines against S. agalactiae conjugated to protein carriers are in advanced stages of development. Additionally, protein-based vaccines are also in development, showing great promise as they can provide protection regardless of serotype. Furthermore, safety concerns regarding a new vaccine are the main barriers identified. Here, we present vaccines in development and identified safety, cost, and efficacy concerns, especially in high-need, low-income countries.

CXCL13/CXCR5 axis mediates IgM+ B cell migration through AKT and STAT3 signaling pathways in Nile tilapia (Oreochromis niloticus)

Chemokine C-X-C motif ligand 13 (CXCL13), originally identified as a B-lymphocyte chemokine, exerts a crucial role in the chemotaxis of B cells through its receptor CXCR5. Although the presence of CXCL13/CXCR5 had been reported in various teleost fish species, its impact on IgM+ B cell still remains unknown. In this study, we investigated the role of chemokine CXCL13 and its receptor CXCR5 in the chemotaxis of IgM+ B cells in Nile tilapia (Oreochromis niloticus). We identified and characterized two isotypes of OnCXCL13 (OnCXCL13a and OnCXCL13b) and CXCR5 (OnCXCR5) in Nile tilapia. The OnCXCL13 is characterized with a typical small cytokine CXC domain and four cysteine, in which the first two cysteines separated by a random amino acid residue, and OnCXCR5 is characterized by seven α-helical transmembrane domains. Both OnCXCL13 and OnCXCR5 were found to be expressed in various tissues and lymphocytes, with the highest expression in the spleen (SPL). Their expression in IgM+ B cells was significantly higher than that in IgM− lymphocytes. Following Streptococcus agalactiae infection, the expressions of OnCXCL13a and OnCXCL13b were up-regulated in peripheral blood (PBL), SPL and head kidney (HK), while OnCXCR5 was significantly up-regulated in SPL and HK. We also found that the recombinant OnCXCL13 proteins had strong chemotactic activity on IgM+ B cells, which could be inhibited by blocking OnCXCR5 with anti-OnCXCR5 antibody. Mechanistically, incubation with OnCXCL13 resulted in the up-regulation of AKT and STAT3 phosphorylation in IgM+ B cells, which could be attenuated by specific inhibitors of AKT and STAT3. Inhibition of AKT and STAT3 also weakened the chemotaxis of OnCXCL13 on IgM+ B cells. Our results suggest that the CXCL13/CXCR5 axis participates in the immune response of tilapia and mediates the chemotaxis of IgM+ B cells, providing insights into the potential role of this axis in bacterial infection and teleost B cell migration.

A Siglec3-like lectin from Nile tilapia (Oreochromis niloticus): Transcriptional profiling upon Streptococcus agalactiae infection and its potential role in immune escape

Siglec-3, also known as CD33, is a type of sialic acid binding immunoglobulin-like lectin that plays a role in regulating cellular activation in the innate immune system and subsequent inflammatory responses. In this study, we cloned the cDNA encoding OnSiglec3-like ORF sequence (1170 bp) in Nile tilapia, contained two Ig-like domains in the extracellular region and one inhibitory ITIM-like motif in the intracellular region. Through qRT-PCR analysis, we found that the mRNA of OnSiglec3-like lectin was highly expressed in the brain and head kidney. When challenged with Streptococcus agalactiae both in vivo and in vitro, we observed a significant up-regulation in the expression of OnSiglec3-like lectin in the head kidney tissues and leukocytes, respectively. Additionally, we investigated the interaction between a recombinant extracellular region protein (referred to as (r)exSiglec3-like protein) and three different phenotypes of S. agalactiae: WT (wild type), ∆cps (cps-deficient mutant), and ∆neuA (sia-deficient mutant) using ELISA. We found that the exOnSiglec3-like protein, when combined with RNA interference and bacterial infection, significantly enhanced the phagocytic ability against S. agalactiae. Furthermore, the expression of four common inflammatory factors (OnIL-6, OnIL-10, OnTNF-α, and OnTGF-β) showed a significant up-regulation. Overall, this study suggests that OnSigelc3-like lectin may play a role in host defence against bacterial infection and mediate immune escape in Nile tilapia.

Serotype distribution, virulence and antibiotic resistance of Streptococcus agalactiae isolated from cultured tilapia Oreochromis niloticus in Lake Volta, Ghana.

Streptococcus agalactiae infection is one of the major factors limiting the expansion of tilapia farming globally. In this study, we investigated the serotype distribution, virulence and antimicrobial resistance of S. agalactiae isolates from tilapia farmed in Lake Volta, Ghana. Isolates from 300 moribund fish were characterised by Gram staining, MALDI-TOF/MS and 16S rRNA sequencing. Serotype identification was based on multiplex polymerase chain reaction (PCR) amplification of the capsular polysaccharide genes. Detection of virulence genes (cfb, fbsA and cspA) and histopathology were used to infer the pathogenicity of the isolates. The susceptibility of isolates to antibiotics was tested using the Kirby-Bauer disk diffusion assay. All 32 isolates identified as S. agalactiae were of serotype Ia. This was notably different from isolates previously collected from the farms in 2017, which belonged to serotype Ib, suggesting a possible serotype replacement. The prevalence of the pathogen was related to the scale of farm operation, with large-scale farms showing higher S. agalactiae positivity. Data from histopathological analysis and PCR amplification of targeted virulence genes confirmed the virulence potential and ability of the isolates to cause systemic infection in tilapia. Except for gentamicin, the majority of the isolates were less resistant to the tested antibiotics. All isolates were fully sensitive to oxytetracycline, erythromycin, florfenicol, enrofloxacin, ampicillin and amoxicillin. This study has improved our understanding of the specific S. agalactiae serotypes circulating in Lake Volta and demonstrates the need for continuous monitoring to guide the use of antimicrobials and vaccines against streptococcal infections in Ghanaian aquaculture systems.

Recombinant lactococcal-based oral vaccine for protection against Streptococcus agalactiae infections in tilapia (Oreochromis niloticus)

Streptococcosis outbreaks caused by Streptococcus agalactiae infection in tilapia aquaculture have been consistently reported and associated with high mortality and morbidity leading to significant economic losses. Existing vaccine candidates against Streptococcus spp. are designed for intraperitoneal injections that are not practical and labor-intensive which have prompted farmers to protect aquatic animals with antibiotics, thus encouraging the emergence of multidrug resistant bacteria. In this study, a live recombinant L. lactis vaccine expressing a 1403 bp surface immunogenic protein (SIP) and a 1100 bp truncated SIP (tSIP) gene was developed and evaluated against S. agalactiae infection in tilapia. Both SIP and tSIP sequences were cloned and transformed into L. lactis. The recombinant L.lactis vaccine was orally administered to juvenile tilapia for a month. Detection of SIP-specific serum IgM in vaccinated groups compared to control groups indicated that recombinant proteins expressed from L. lactis could elicit immunogenic reactions in tilapia. Fish immunized with the tSIP vaccine also showed the highest level of protection compared to other test groups, and the mortality rate was significantly reduced compared to both control groups. The relative percentage of survival (RPS) against S. agalactiae for both SIP and tSIP-vaccinated groups was 50 % and 89 %, respectively, at 14 days post-challenge. Significant up-regulation of IgM, IL-1β, IL-10, TNF-α and IFN-γ were observed at day 34 between the vaccinated and control groups. These results indicated that the recombinant lactococcal tSIP vaccine can elicit both cell-mediated and humoral responses and is recommended as a potential oral vaccine against S. agalactiae infection. Future work will include further in vivo challenge assessments of this vaccine candidate fused with adjuvants to boost immunogenicity levels in tilapia.

Selenium Nanoparticles Ameliorate Adverse Impacts of Aflatoxin in Nile Tilapia with Special Reference to Streptococcus agalactiae Infection

Aflatoxin B1 (AFB1) is a plant-origin toxin that could induce oxidative stress in fish. The micromineral selenium (Se) possesses well-documented antioxidant properties. To assess the ameliorative effects of SeNPs (1 mg/kg fish feed) on oxidative stress induced by AFB1 (500 μg/kg fish feed), Nile tilapia (32.2±1.7 g body weight) were distributed randomly and even in six groups for 8-week feeding trial. Live enzymes, AST, ALT, and ALP levels were increased in the serum of fish fed AFB1-contaminated diet, and the addition of SeNPs could restore normal values compared to the control. The gene expression of antioxidant enzymes, superoxide dismutase (SOD) enzyme and catalase (CAT) enzyme, and DNA fragmentation were significantly increased in response to aflatoxin exposure, while dietary SeNPs could mitigate the generated oxidative stress. The innate immunity, serum antibacterial activity (SAA), oxidative burst activity (OBA), phagocytic activities (PA and PI), and gene expression of cytokines (interleukin (IL)-1β, heat shock protein70 (Hsp), and tumor necrosis factor (TNF)-α) revealed a status of immunosuppression in Nile tilapia fed on AFB1-contaminated diet. These findings showed that fish became more vulnerable to Streptococcus agalactiae infection with a high mortality rate while dietary SeNPs provided a high relative protection level (RPL). From the obtained findings, SeNPs could mitigate the oxidative stress induced by feeding the AFB1 diet and could boost the immunity of stressed Nile tilapia.

Identification, characterization and functional analysis of Rap1 in response to bacterial infection in tilapia

Rap1 (Ras-related protein 1), as a member of the Ras-like small GTP binding protein family, involves in the regulation of cell life activities and mediates a variety of biological events including cell junctions, cell polarity, cell proliferation and differentiation by binding to GDP/GTP. In addition, Rap1 also plays an important role in innate immune response by regulating the adhesion, diffusion and migration of white blood cells. However, it is still unclear how Rap1 affects the innate immunity of teleost fish against bacterial infections. In this study, we amplified the Rap1 from tilapia and conducted the sequence analysis of Rap1 nucleotide and amino acid sequences, then characterized its tissue distribution and expression pattern in response to pathogen infection in tilapia for the first time, and explored the bacteriostatic and bacterial binding effects of Rap1. The results showed that the Rap1 possessed a whole open reading frame (ORF) with 558 bp, encoding 185 amino acids. The Rap1 protein was highly homologous with Rap1 from other species. Conserved domain analysis revealed that Rap1 contained five G box domains that mediated its binding to ornithine. Rap1 mRNA was widely expressed in tilapia tissues, and the highest level was found in spleen. Under Streptococcus agalactiae (S. agalactiae) infection, the expression of Rap1 was significantly up-regulated in head kidney and brain. In addition, the recombinant protein Rap1 hold the ability binding to S. agalactiae and inhibit the proliferation of S. agalactiae. In conclusion, this study not only provided a reference for exploring the function of Rap1 in teleost against pathogen infection, but also enriched the basic theoretical research of Rap1 function and antibacterial immune response in teleost.

A nanocarrier immersion vaccine encoding surface immunogenic protein confers cross-immunoprotection against Streptococcus agalactiae and Streptococcus iniae infection in tilapia

Streptococcosis is a highly contagious aquatic bacterial disease that poses a significant threat to tilapia. Vaccination is a well-known effective measure to prevent and control fish bacterial diseases. Among the various immunization methods, immersion vaccination is simple and can be widely used in aquaculture. Besides, nanocarrier delivery technology has been reported as an effective solution to improve the immune effect of immersion vaccine. In this study, the surface immunogenic protein (Sip) was proved to be conserved and potential to provide cross-immunoprotection for both Streptococcus agalactiae (S. agalactiae) and Streptococcus iniae (S. iniae) by multiple sequences alignment and Western blotting analysis. On this basis, we expressed and obtained the recombinant protein rSip and connected it with functionalized carbon nanotubes (CNT) to construct the nanocarrier vaccine system CNT-rSip. After immersion immunization, the immune effect of CNT-rSip against above two streptococcus infections was evaluated in tilapia based on some aspects including the serum specific antibody level, non-specific enzyme activities, immune-related genes expression and relative percent survival (RPS) after bacteria challenge. The results showed that compared with control group, CNT-rSip significantly (P < 0.05) increased the serum antibody levels, related enzyme activities including acid phosphatase, alkaline phosphatase, lysozyme and total antioxidant capacity activities, as well as the expression levels of immune-related genes from 2 to 4 weeks post immunization (wpi), and all these indexes peaked at 3 wpi. Besides, the above indexes of CNT-rSip were higher than those of rSip group with different extend during the experiment. Furthermore, the challenge test indicated that CNT-rSip provided cross-immunoprotection against S. agalactiae and S. iniae infection with RPS of 75 % and 72.41 %, respectively, which were much higher than those of other groups. Our study indicated that the nanocarrier immersion vaccine CNT-rSip could significantly improve the antibody titer and confer cross-immuneprotection against S. agalactiae and S. iniae infection in tilapia.

Effect of dietary intervention with Capsicum annuum extract on growth performance, physiological status, innate immune response, and related gene expression in Nile tilapia

The red pepper (Capsicum annuum) has gained great attention recently because of its biological and pharmacological characteristics. The present approach aimed to evaluate the effects of C. annuum alcoholic extract (CAE) supplementation on Nile tilapia (Oreochromis niloticus) growth performance, physiological status, some metabolic, immune, and regulatory genes expression, and resistance against Streptococcus agalactiae infection. Fish (22.26 ± 0.19 g) were assigned to four treatments (five replicates, each with 10 fish replicate−1) and fed tested diets for 60 days. The experimental diets were supplemented with CAE at 0, 0.4, 0.8, and 1.6 g kg−1, expressed as CAE0, CAE0.4, CAE0.8, and CAE1.6, respectively. The findings exhibited that CAE dietary supplementation improved growth performance, feed utilization, elevated growth hormone level, and digestive enzyme activities (amylase and protease), and lowered leptin hormone in a level-dependent manner. Boosting the mRNA expression of the transporter proteins (solute carrier family 15 member 2 and solute carrier family 26 member 6) and insulin-like growth factor-1 genes with a decrease in the myostatin gene expression was noticed in the CAE-fed groups. The innate immune (serum bactericidal activity %, complement 3, and phagocytic activity %) and antioxidant (glutathione peroxidase and total antioxidant capacity) parameters were significantly (p < 0.05) improved, and the serum malondialdehyde level was significantly decreased by CAE dietary inclusion. A marked upregulation in the mRNA expression of interleukins (il-1β, il-6, il-8, and il-10), transforming growth factor-β, glutathione peroxidase, and glutathione synthetase genes were observed in CAE-fed groups. Dietary CAE decreased the cumulative mortalities after the challenge with S. agalactiae by 20, 13.33, and 10% in CAE0.4, CAE0.8, and CAE1.6, respectively, compared to the control (40%). Overall, dietary supplementation with CAE could improve growth performance and physiological status, and modulate the expression of several regulatory genes in Nile tilapia. The recommended level of CAE is 1.6 g kg−1 to augment growth and health status.

Molecular characterization and expression analysis of nine toll like receptor (TLR) genes in Scortum barcoo under Streptococcus agalactiae infection

Toll like receptors (TLRs) are important pattern recognition receptors participating in innate immune system. Up to now, no TLR has been identified in Jade perch (Scortum barcoo). In this study, we successfully identified 9 members of TLRs from the Jade perch. Amino acid sequence alignment analysis showed that the whole sequences of these TLRs were highly conserved among different fish species, especially in LRR, TM and TIR domains. Phylogenetic analysis revealed that each SbTLR was successfully grouped into corresponding gene family of teleosts. Expression analysis showed that most SbTLRs mainly expressed in liver, spleen, muscle and skin, while expressed less in brain and stomach. After Streptococcus agalactiae infection, expression of SbTLR2, SbTLR5S and SbTLR22 were significantly upregulated, while SbTLR3, SbTLR5M, SbTLR9, SbTLR13, and SbTLR14 were significantly downregulated. In all, this research first reported molecular characterization and expression profiles of 9 TLRs in Jade perch. These data will make a contribution for better understanding the antibacterial mechanism of TLRs in teleosts.

138 Streptococcus agalactiae infection in a patient with psoriasis under treatment with an IL-17 antagonist

138 Streptococcus agalactiae infection in a patient with psoriasis under treatment with an IL-17 antagonist

Prophylactic effects of dietary caper (Capparis spinosa) extracts on the control of Streptococcus agalactiae infection, growth, immune-antioxidant, and inflammation cytokine responses of Nile tilapia fingerlings

The antibacterial activity of aqueous (AE) or ethanolic extracts (EE) of caper (Capparis spinosa) against Streptococcus agalactiae was evaluated in vitro. Both caper extracts showed antagonistic activity against S. agalactiae and the inhibition zones in case of ethanolic extracts were larger than those of aqueous ones. Additionally, TEM investigations show that S. agalactiae cells treated with both C. spinosa extracts were damaged and degraded and this damage was greater in case of ethanolic extract. Another study was done to assess the promotion effects of dietary caper (C. spinosa) extracts on growth, antioxidant and immune activity, and inflammation cytokine responses of Nile tilapia (Oreochromis niloticus) and its resistance to S. agalactiae infection. However, fish (40 ± 2 g) were fed on diets containing 1.0 and 2.0 g/kg feed of each caper extract as well as the control group (free of caper) for 6 weeks. Fish were intraperitoneally injected (IP) with Streptococcus agalactiae at the end of the feeding trial, and fish mortality was tracked for additional ten days. Compared with other treatments, fish fed on 2.0 g EE/kg feed had higher counts of white and red blood cells as well as higher hemoglobin levels accompanied with lower AST and ALT activities. Antioxidant (superoxide dismutase and catalase activities) and immune, total protein, globulin, lysozyme, and immunoglobulin M) indices were increased along with significant decline in MDA levels in both caper extracts treated fish groups compared to the control group. Significant promotion in fish growth was affected positively with the increase in both caper extracts; particularly, the larger fish growth was observed in the treatment of 2.0 g EE/kg feed. Expressions of IL-1β and IL-8 were declined; meanwhile levels of IL-10, SOD and CAT genes were upregulated in fish fed on 2.0 g EE/kg feed compared to other groups. After being challenged with S. agalactiae infection, fish survival was considerably (P < 0.05) greater in fish groups that fed on diets with caper extracts; particularly 2.0 g EE/kg feed (75%); while all fish fed on the control one were dead. According to these findings, the antioxidant and immune response of Nile tilapia fingerlings is stimulated by ethanolic extract of caper (2.0 g/kg feed), which also enhanced the growth performance and fish resistance to S. agalactiae infection.

Dietary supplementation of Patchouli oil increases the resistance against Streptococcus agalactiae infection in GIFT tilapia as revealed by transcriptome and 16s amplicon profiling

Large-scale disease outbreaks that caused by Streptococcus agalactiae (SA) infection have dramatically affected tilapia production worldwide. Patchouli oil (PO) is a traditional Chinese herbal medicine in treating gastrointestinal symptoms and is considered to be a safe and efficient candidate to treat SA infection in tilapia. In this study, we first investigated the changes of intestinal microbiota and host responses in GIFT tilapia after a 28-day dietary supplementation of different concentrations of PO (basic diet, 5 μL PO/g diet, or 15 μL PO/g diet) and followed by SA infection using an integrative analysis of transcriptome and 16 S amplicon profiling. We observed that dietary supplementation of 5 μL PO/g diet improved food intake and growth in tilapia. Histopathological observation confirmed there were increased surface area of intestinal villi and generally decreased ROS as revealed by the MAD, GSH-PX, and T-SOD activities in the tilapia intestines in response to dietary PO supplementation. The intestinal microbial diversities and community structures were significantly affected, and the colonization of probiotics (e.g., Cetobacterium and clostridium_ sensu_stricto_1) in the mucosa was improved in the dietary PO groups. The relative abundance of Streptococcus (0.033%) in the 15 μL PO/g dietary group was much lower than that in the control group (basic diet; 0.86%). A large number of DEGs were enriched in the immune-related pathways, e.g., lysosomal lumen, MHC protein complex, and toll-like receptor 2 signaling pathway. Our findings suggested that dietary supplementation of PO enhanced intestinal physical barriers, increased the colonization of probiotic bacteria and improved intestinal immunity in tilapia. This provides novel knowledge on PO as a dietary supplement to improve intestinal health against SA invasion in tilapia.

Dietary supplementation of patchouli oil improves resistance to Streptococcus agalactiae infection in genetically improved cultured Tilapia (GIFT, Oreochromis niloticus.) as revealed by RNA-Seq

Pogostemon oil (PO) is a volatile oil extracted from Pogostemon cablin., a traditional Chinese medicinal plants. Clinical studies have confirmed that PO has broad-spectrum antibacterial activity, but the resistance and protective mechanism of PO against Streptococcus agalactiae (SA) infection in Genetically Improved Cultured Tilapia (GIFT, Oreochromis niloticus.) are still unclear. The purpose of our study was to verify the resistance of PO feed to SA infection and reveal the potential immune mechanism. These 252 Tilapias were randomly divided into three groups, each with three replicates. The control group CON was fed with basic diet; These experimental groups PO5 and PO15 were fed with 5 μ L / g, 15 μ L / g PO diet, respectively; After 28 days feeding, Tilapia were randomly selected from CON, PO5, and PO15 groups, respectively: 22 fishes in each group were intraperitoneally injected with PBS to produce NC, NP5, and NP15 groups; other 22 fishes in each group were intraperitoneally injected with SA to produce CK, SP5, and SP15 groups. Twelve hours after SA challenge, spleen tissues were collected to study the effect of PO feed resist SA infection in Tilapia by RNA-Seq. In addition, two independent acute SA challenge experiments were added to verify the protective ability of PO against SA infection. In the transcriptome results, co-expressed genes in MEgreen (divided by WGCNA) were enriched in immune-related pathways by KEGG analysis, including Toll-like receptor signaling pathway, RIG-I-like receptor signaling pathway, MAPK signaling pathway, NOD-like receptor signaling pathway, and C-type lectin receptor signaling pathway. Cluster analysis of 166 shared DEGs responsible to SA infection identified two major clusters with almost opposite expression trends. il-1β (interleukin-1 beta), gpr84 (G protein-coupled receptor 84), and OXER1 (oxoeicosanoid receptor 1) belonged to both share DEGs and key co-expressed genes, were up-regulated by PO after SA infection. In the survival challenge of LD50 concentration of SA, 5μL / g (0.67% vs 53.33%, P < 0.01), 10 μL / g (0.67% vs 53.33%, P < 0.01), and 15 μL / g (33.33% vs 53.33%, P > 0.05) PO feed groups decreased Tilapia mortality compared to basic diet group. In the challenge of SA concentration 10 times higher than LD50, 15 μL / g PO feed (55% vs 95%, P < 0.05) reduced mortality compared to basic diet group, and 5 μL / g PO feed significantly prolonged the median survival time of 1 day. In general, 5–15 μL / g PO feeds improved the resistance of Tilapia to SA infection and reduced the mortality of Tilapia. The results of RNA-Seq provide data support for the molecular mechanism of PO feeds regulating the immune response of Tilapia to SA. Collectively, PO may have the potential to be developed as a dietary supplementation for preventing and treating SA infections.