Erythrocytes of the common carp are immune sentinels that sense pathogen molecular patterns, engulf particles and secrete pro-inflammatory cytokines against bacterial infection.

Introduction: Red blood cells (RBCs), also known as erythrocytes, are underestimated in their role in the immune system. In mammals, erythrocytes undergo maturation that involves the loss of nuclei, resulting in limited transcription and protein synthesis capabilities. However, the nucleated nature of non-mammalian RBCs is challenging this conventional understanding of RBCs. Notably, in bony fishes, research indicates that RBCs are not only susceptible to pathogen attacks but express immune receptors and effector molecules. However, given the abundance of RBCs and their interaction with every physiological system, we postulate that they act in surveillance as sentinels, rapid responders, and messengers. Methods: We performed a series of in vitro experiments with Cyprinus carpio RBCs exposed to Aeromonas hydrophila, as well as in vivo laboratory infections using different concentrations of bacteria. Results: qPCR revealed that RBCs express genes of several inflammatory cytokines. Using cyprinid-specific antibodies, we confirmed that RBCs secreted tumor necrosis factor alpha (TNFα) and interferon gamma (IFNγ). In contrast to these indirect immune mechanisms, we observed that RBCs produce reactive oxygen species and, through transmission electron and confocal microscopy, that RBCs can engulf particles. Finally, RBCs expressed and upregulated several putative toll-like receptors, including tlr4 and tlr9, in response to A. hydrophila infection in vivo. Discussion: Overall, the RBC repertoire of pattern recognition receptors, their secretion of effector molecules, and their swift response make them immune sentinels capable of rapidly detecting and signaling the presence of foreign pathogens. By studying the interaction between a bacterium and erythrocytes, we provide novel insights into how the latter may contribute to overall innate and adaptive immune responses of teleost fishes.

Effects of dietary ginger (Zingiber officinale) polysaccharide on the growth, antioxidant, immunity response, intestinal microbiota, and disease resistance to Aeromonas hydrophila in crucian carp (Carassius auratus).

Ginger polysaccharides (GP) promote growth and development in fish. However, the effects of GP on crucian carp remain unclear. The present study investigated the effects of GP on the growth performance, immunity, intestinal microbiota, and disease resistance in crucian carp. Four treatment groups were established with different concentrations of GP (0.1 %, 0.2 %, 0.4 %, and 0.8 %). GP was not added as the control group, and the feeding period lasted for 56 d, followed by a 96-h anti-infection treatment using Aeromonas hydrophila. The results showed that dietary GP significantly improved growth performance, especially in the 0.4 % GP group. Furthermore, GP administration notably increased serum lysozyme (LMZ) activity, digestive enzyme performance, and antioxidant capacity of crucian carp. Moreover, dietary inclusion of GP up-regulated the expression of tumour necrosis factor-α (TNF-α), interleukin-8 (IL-8), interferon-γ (IFN-γ), and nuclear factor kappa-B (NF-κB) genes while down-regulating IL-10 and transforming growth factor-ß (TGF-ß) gene expressions, thus promoting liver health in crucian carp. Additionally, incorporating GP into the diet regulated both the diversity and composition of the intestinal microbiota in crucian carp, explicitly enhancing the relative abundance of beneficial bacteria, such as Fusobacteriota and Firmicutes. Therefore, GP reduces the mortality of crucian carp infected with A. hydrophila. In conclusion, this study provides novel insights into the application of dietary GP in cultured fish and evaluates the value of traditional Chinese medicinal polysaccharides against pathogenic bacteria.

One Step Rapid Sensitive Method for the Diagnosis of Hemolysin Gene of Aeromonas hydrophila by Polymerase Chain Reaction.

Aeromonas hydrophila is a Gram-negative bacterium that has been linked to serious illnesses in both humans and animals. The presence of hemolysin, a virulence factor, is critical in the development of A. hydrophila-related illnesses. As a result, precise and timely detection of the hemolysin gene is critical for efficient diagnosis and prevention of many illnesses. The PCR is used in this study to detect the hemolysin gene of A. hydrophila in a novel, fast, and highly sensitive one-step technique. Specific primers were constructed to amplify a conserved area within the hemolysin gene to achieve both specificity as well as sensitivity. The PCR assay was rigorously optimized, taking temperature, primer concentration, and reaction time into account, in order to maximize the efficiency and reliability of this method. In conclusion, this method's simplicity, sensitivity, and specificity make it highly promising for regular diagnostic applications. Its application would allow for the early detection of A. hydrophila infections, allowing for more effective treatment and control methods.

Growth performance, mucosal immunity and disease resistance in goldfish (Carassius auratus) orally administered with Escherichia coli Strain Nissle 1917.

The current research aimed to shed light on the efficacy of Escherichia coli strain Nissle 1917 (EcN) on goldfish (……) growth, gut immunity, morphology, bacterial nutritional enzyme activity and resistance to Aeromonas hydrophila infection. Fish fed with EcN at 106, 107 and 108 CFU/g feed for 80 days showed an enhancement in growth better than control fish. The gut innate immunity in terms of lysozyme activity, immunoglobulin and total protein levels was increased in the treatment fish with the best result being observed in fish fed EcN at 108 CFU/ g. In addition, an increase was noted in the upregulation of immune-relevant genes, namely lysozyme, interleukin-1ß, inducible nitric oxide synthase and tumor necrosis factor α of fish intestine. A marked surge in the number of proteolytic and heterotrophic bacteria was noted in the gut of fish nourished with the probiotic. Histological studies exhibited an improvement in the intestinal absorption surface area, intraepithelial lymphocyte count and goblet cell density. Significantly higher survival rate was obtained in fish fed EcN at 108 CFU/g compared with the fish fed with the basal diet. These data exhibited the beneficial effect of EcN on goldfish growth, digestive enzymes, intestine heterotrophic bacteria and resistance against Aeromonas hydrophila challenge. This study confirmed the favorable outcomes resulting from the administration of EcN at108 CFU/g.

Joint effects about antibiotics combined using with antibiotics or phytochemicals on Aeromonas hydrophila.

Aeromonas hydrophila is highly prevalent in aquaculture animals and aquaculture environment. Due to the abuse of antibiotics, A. hydrophila can change the antibiotic resistance spectrum directly and affect human health indirectly. The use of combined drugs replacement therapy and the long-term coexistence with drug-resistant bacteria are the reality that human beings have to face in dealing with the problem of antibiotic resistance in the future. This study showed the characteristics and trends through abundant results of combined effects related with the combinations of antibiotic and the combinations of antibiotic and phytochemical on A. hydrophila, and revealed the antagonism probability of combinations of antibiotic and phytochemical is significantly higher than that of the combinations of antibiotic. Meanwhile, the combinations of antibiotic and phytochemical could protect the host cells which also achieved the same effectiveness as combination of antibiotics, and the enrichment pathway was proved to be relatively discrete. In addition, the possible mechanism about the reverse "U" shape of the combined effect curve on wild/antibiotic-resistant bacteria was clarified, and it was confirmed that the antagonism for the combinations of antibiotic and phytochemical might has the significance in inhibiting the evolution of bacterial resistance mutations. This study was aims to provide theoretical basis and some clues for the antibiotic resistance control associated with A. hydrophila.

Rapid discrimination methods for clinical and environmental strains of Aeromonas hydrophila and A. veronii biovar sobria using the N-terminal sequence of the flaA gene and investigation of antimicrobial resistance.

Although the genus Aeromonas inhabits the natural environment, it has also been isolated from hospital patient specimens as a causative agent of Aeromonas infections. However, it is not known whether clinical strains live in the natural environment, and if these strains have acquired antimicrobial resistance. In this study, we performed the typing of flagellin A gene (flaA) of clinical and environmental strains of Aeromonas hydrophila and A. veronii biovar sobria using Polymerase Chain Reaction (PCR) assay with newly designed primers. Detection rates of the clinical and environmental flaA types of A. hydrophila were 66.7% and 88.2%, and the corresponding rates for A. veronii biovar sobria were 66.7% and 90.9%. The PCR assays could significantly discriminate between clinical and environmental strains of both species in approximately 4 h. Also, among the 63 clinical Aeromonas strains used, only one extended-spectrum ß-lactamase-producing bacteria, no plasmid-mediated quinolone resistance bacteria, and only four multidrug-resistant bacteria were detected. Therefore, the PCR assays could be useful for the rapid diagnosis of these Aeromonas infections and the monitoring of clinical strain invasion into water-related facilities and environments. Also, the frequency of drug-resistant Aeromonas in clinical isolates from Okinawa Prefecture, Japan, appeared to be low.

Immunostimulatory effects of dietary verjuice (Vitis vinifera) on immune response and transcription of immune-related genes in juvenile rainbow trout (Oncorhynchusmykiss).

The development of green aquaculture practices has led to the supplementation of fish diets with natural immunostimulants such as organic acids. This study aimed to assess the dietary effects of verjuice (VJ; unfermented unripe grapes; Vitis vinifera) on hematological parameters, skin mucosal immunity, transcriptional immune responses, and antibacterial serum activity against Aeromonas hydrophila in rainbow trout. The fish (51.0 ± 2.4 g) were randomly distributed into 15 tanks and fed ad-libitum thrice daily with diets containing different levels of VJ including 0 (control; VJ-0), 3 (VJ-3), 6 (VJ-6), 9 (VJ-9), and 12 (VJ-12) mL/kg VJ for 56 d. Results showed that immuno-hematological parameters (total white blood cells, neutrophils, and monocytes) were improved in VJ-added groups (P < 0.05). In addition, dietary VJ (9 mL/kg) modulated serum immunological parameters. Skin mucus immunology exhibited a notable increase in alkaline phosphatase, lysozyme activity, alkaline protease, total protein, total immunoglobulin, and esterase levels in VJ-9 group compared with those in the control group (P < 0.05). The mRNA expression of interleukin-1ß, interleukin-6, interleukin-8, and immunoglobulin M were significantly higher in VJ-9 group than in the control (P < 0.05). Furthermore, the results of the antibacterial evaluation showed that A. hydrophila growth was significantly inhibited in the serum samples from VJ-3 to VJ-9 groups after the 56th day and in all VJ-treated groups after the 70th (P < 0.05). In conclusion, dietary VJ is a novel immunostimulant and the optimal dietary supplementation level of 6.65-7.46 mL/kg can effectively improve immune responses in rainbow trout.

Oral administration of recombinant outer membrane protein A-based nanovaccine affords protection against Aeromonas hydrophila in zebrafish.

Aeromonas hydrophila, an opportunistic warm water pathogen, has always been a threat to aquaculture, leading to substantial economic losses. Vaccination of the cultured fish would effectively prevent Aeromoniasis, and recent advancements in nanotechnology show promise for efficacious vaccines. Oral delivery would be the most practical and convenient method of vaccine delivery in a grow-out pond. This study studied the immunogenicity and protective efficacy of a nanoparticle-loaded outer membrane protein A from A. hydrophila in the zebrafish model. The protein was over-expressed, purified, and encapsulated using poly lactic-co-glycolic acid (PLGA) nanoparticles via the double emulsion method. The PLGA nanoparticles loaded with recombinant OmpA (rOmpA) exhibited a size of 295 ± 15.1 nm, an encapsulation efficiency of 72.52%, and a polydispersity index of 0.292 ± 0.07. Scanning electron microscopy confirmed the spherical and isolated nature of the PLGA-rOmpA nanoparticles. The protective efficacy in A. hydrophila-infected zebrafish after oral administration of the nanovaccine resulted in relative percentage survival of 77.7. Gene expression studies showed significant upregulation of immune genes in the vaccinated fish. The results demonstrate the usefulness of oral administration of nanovaccine-loaded rOmpA as a potential vaccine since it induced a robust immune response and conferred adequate protection against A. hydrophila in zebrafish, Danio rerio.

Quercetin disrupts biofilm formation and attenuates virulence of Aeromonas hydrophila.

Aeromonas hydrophila poses significant health and economic challenges in aquaculture owing to its pathogenicity and prevalence. Overuse of antibiotics has led to multidrug resistance and environmental pollution, necessitating alternative strategies. This study investigated the antibacterial and antibiofilm potentials of quercetin against A. hydrophila. Efficacy was assessed using various assays, including antibacterial activity, biofilm inhibition, specific growth time, hemolysis inhibition, autoaggregation, and microscopic evaluation. Additionally, docking analysis was performed to explore potential interactions between quercetin and virulence proteins of A. hydrophila, including proaerolysin, chaperone needle-subunit complex of the type III secretion system, and alpha-pore forming toxin (PDB ID: 1PRE, 2Q1K, 6GRK). Quercetin exhibited potent antibacterial activity with 21.1 ± 1.1 mm zone of inhibition at 1.5 mg mL-1. It also demonstrated significant antibiofilm activity, reducing biofilm formation by 46.3 ± 1.3% at the MIC and attenuating autoaggregation by 55.9 ± 1.5%. Hemolysis was inhibited by 41 ± 1.8%. Microscopic analysis revealed the disintegration of the A. hydrophila biofilm matrix. Docking studies indicated active hydrogen bond interactions between quercetin and the targeted virulence proteins with the binding energy -3.2, -5.6, and -5.1 kcal mol⁻1, respectively. These results suggest that quercetin is an excellent alternative to antibiotics for combating A. hydrophila infection in aquaculture. The multifaceted efficacy of quercetin in inhibiting bacterial growth, biofilm formation, virulence factors, and autoaggregation highlights the potential for aquaculture health and sustainability. Future research should delve into the precise mechanisms of action and explore synergistic combinations with other compounds for enhanced efficacy and targeted interventions.

Comparative Analysis of mRNA, microRNA of Transcriptome, and Proteomics on CIK Cells Responses to GCRV and Aeromonas hydrophila.

Grass Carp Reovirus (GCRV) and Aeromonas hydrophila (Ah) are the causative agents of haemorrhagic disease in grass carp. This study aimed to investigate the molecular mechanisms and immune responses at the miRNA, mRNA, and protein levels in grass carp kidney cells (CIK) infected by Grass Carp Reovirus (GCRV, NV) and Aeromonas hydrophilus (Bacteria, NB) to gain insight into their pathogenesis. Within 48 h of infection with Grass Carp Reovirus (GCRV), 99 differentially expressed microRNA (DEMs), 2132 differentially expressed genes (DEGs), and 627 differentially expressed proteins (DEPs) were identified by sequencing; a total of 92 DEMs, 3162 DEGs, and 712 DEPs were identified within 48 h of infection with Aeromonas hydrophila. It is worth noting that most of the DEGs in the NV group were primarily involved in cellular processes, while most of the DEGs in the NB group were associated with metabolic pathways based on KEGG enrichment analysis. This study revealed that the mechanism of a grass carp haemorrhage caused by GCRV infection differs from that caused by the Aeromonas hydrophila infection. An important miRNA-mRNA-protein regulatory network was established based on comprehensive transcriptome and proteome analysis. Furthermore, 14 DEGs and 6 DEMs were randomly selected for the verification of RNA/small RNA-seq data by RT-qPCR. Our study not only contributes to the understanding of the pathogenesis of grass carp CIK cells infected with GCRV and Aeromonas hydrophila, but also serves as a significant reference value for other aquatic animal haemorrhagic diseases.

A cytokine-like factor 1 homolog acts as a macrophage chemoattractant in grass carp.

Cytokine-like factor 1 (CYTL1) is a small cytokine and has diverse biological functions in mammals. However, whether CYTL1 exists in lower vertebrates is not clear. In this study, we identified cytl homologs in fish and characterized the immune functions in a teleost species, grass carp (Ctenopharyngodon idella). Fish CYTL1 homologs share conserved molecular features with their mammalian counterparts, including 6 cysteine residues in the mature peptide, genomic organization and synteny. Gene expression analysis revealed that cytl1 was constitutively expressed in tissues of grass carp, with the highest expression detected in the heart. Upon infection with Aeromonas hydrophila (A. hydrophila), cytl1 was downregulated in the hindgut, head kidney, skin, and spleen. In the primary head kidney leukocytes (HKLs), stimulation with inactivated A. hydrophila, LPS, poly(I:C), IL-22, IFN-a or IFN-γrel resulted in downregulation of cytl1 expression. Recombinant grass carp CYTL1 protein produced in the HEK293-F cells was potent to induce il-10 expression, but had little effect on the expression of il-1ß and il-6. In vivo experiments revealed that CYTL1 was effective to recruit macrophages to the muscle injected with cytl expression plasmids. Taken together, our results indicate that CYTL1 is a potent chemokine for recruitment of macrophages in fish.

The immune function of heme oxygenase-1 (HO-1) from Nile tilapia (Oreochromis niloticus) in response to bacterial infection.

Heme oxygenase-1 (HO-1), an inducible rate-limiting metabolic enzyme, exerts critical immunomodulatory functions by potential anti-oxidant, anti-inflammatory, and anti-apoptotic activities. Although accumulative studies have focused on the immune functions of HO-1 in mammals, the roles in fish are poorly understood, and the reports on involvement in the defensive and immune response are very limited. In this study, On-HO-1 gene from Oreochromis niloticus was successfully cloned and identified, which contained an open reading frame (ORF) of 816 bp and coded for a protein of 271 amino acids. The On-HO-1 protein phylogenetically shared a high homology with HO-1 in other teleost fish (76.10%-98.89 %) and a lowly homology with HO-1 in mammals (38.98%-41.55 %). The expression levels of On-HO-1 were highest in the liver of healthy tilapias and sharply induced by Streptococcus agalactiae or Aeromonas hydrophila. Besides, On-HO-1 overexpression significantly increased non-specific immunological parameters in serum during bacterial infection, including LZM, SOD, CAT, ACP, and AKP. It also exerted anti-inflammatory and anti-apoptotic effects in response to the immune response of the infection with S. agalactiae or A. hydrophila by upregulating anti-inflammatory factors (IL-10, TGF-ß), autophagy factors (ATG6, ATG8) and immune-related pathway factors (P65, P38), and down-regulating pro-inflammatory factors (IL-1ß, IL-6, TNF-α), apoptotic factors (Caspase3, Caspase9), pyroptosis factor (Caspase1), and inflammasome (NLRP3). These results suggested that On-HO-1 involved in immunomodulatory functions and host defense in Nile tilapia.

Immunoprotective efficacy evaluation of OmpTS subunit vaccine against Aeromonas hydrophila infection in Megalobrama amblycephala.

Bacterial septicemia in freshwater fish is mainly caused by Aeromonas hydrophila infection, which affects the development of aquaculture industry. In the context of sustainable aquaculture, subunit vaccines are of great values because they play positive roles in reducing the overuse of antibiotics and protecting aquatic animals against bacterial infection. In this study, the recombinant outer membrane protein OmpTS of A. hydrophila were used as subunit vaccine to immunize Megalobrama amblycephala, and its immunoprotective effect and host immune responses were evaluated. The survival rates of the vaccinated groups after bacterial infection were significantly higher than that of the control group, especially of the OmpTS high-dose vaccinated group. The better protective effects of vaccinated groups might be attributed to the increased levels of serum IgM-specific antibody titer, the reduced relative abundance of A. hydrophila in various tissues, the increased number of immune-positive cells with different epitopes, the up-regulated expression levels of immune-related genes, and the enhanced activities of antibacterial enzymes. In conclusion, OmpTS subunit vaccine could strongly induce host immune responses in M. amblycephala, thereby enhancing both cellular and humoral immunity, which exhibited excellent and effective immunoprotective efficacy.

Co-infection of Lactococcus garvieae and Aeromonas hydrophila in cultured Nile Tilapia in Kerala, India.

Co-infection of Lactococcus garvieae and Aeromonas hydrophila, has been confirmed from diseased Nile Tilapia (Oreochromis niloticus), Chithralada strain cultured in a freshwater rearing pond of Alappuzha district of Kerala, India. The aetiological agents behind the disease outbreak were bacteriologically proven and confirmed by 16SrRNA sequencing and phylogenetic analysis. PCR detection of the virulent genes, showed existence of adhesin and hemolysin in L. garvieae and aerolysin in A. hydrophila strain obtained. To fulfil Koch's postulates, challenge experiments were conducted and median lethal dose (LD50) of L. garvieae and A. hydrophila was calculated as 1 × 105.91 CFU per mL and 1 × 105.2 CFU per mL respectively. Histopathologically, eyes, spleen, and kidney were the predominantly infected organs by L. garvieae and A. hydrophila. Out of the 13 antibiotics tested to check antibiotic susceptibility, L. garvieae showed resistance to almost 7 antibiotics tested, with a resistance to Ciprofloxacin while A. hydrophila was found resistant to Streptomycin and Erythromycin. Understanding the complex interaction between Gram-positive and Gram-negative bacteria in the disease process and pathogenesis in fish host will contribute to efficient treatment strategies. As a preliminary investigation into this complex interaction, the present study is aimed at phenotypic and genotypic characterization, pathogenicity evaluation, and antibiotic susceptibility of the co-infecting pathogens in a diseased sample of freshwater-farmed Nile tilapia.

Quantitative proteomics investigating the intrinsic adaptation mechanism of Aeromonas hydrophila to streptomycin.

Aeromonas hydrophila, a prevalent pathogen in the aquaculture industry, poses significant challenges due to its drug-resistant strains. Moreover, residues of antibiotics like streptomycin, extensively employed in aquaculture settings, drive selective bacterial evolution, leading to the progressive development of resistance to this agent. However, the underlying mechanism of its intrinsic adaptation to antibiotics remains elusive. Here, we employed a quantitative proteomics approach to investigate the differences in protein expression between A. hydrophila under streptomycin (SM) stress and nonstress conditions. Notably, bioinformatics analysis unveiled the potential involvement of metal pathways, including metal cluster binding, iron-sulfur cluster binding, and transition metal ion binding, in influencing A. hydrophila's resistance to SM. Furthermore, we evaluated the sensitivity of eight gene deletion strains related to streptomycin and observed the potential roles of petA and AHA_4705 in SM resistance. Collectively, our findings enhance the understanding of A. hydrophila's response behavior to streptomycin stress and shed light on its intrinsic adaptation mechanism.

Isolation, characterization and functional analysis of a bacteriophage targeting Culex pipiens pallens resistance-associated Aeromonas hydrophila.

BACKGROUND: Culex pipiens pallens is a well-known mosquito vector for several diseases. Deltamethrin, a commonly used pyrethroid insecticide, has been frequently applied to manage adult Cx. pipiens pallens. However, mosquitoes can develop resistance to these insecticides as a result of insecticide misuse and, therefore, it is crucial to identify novel methods to control insecticide resistance. The relationship between commensal bacteria and vector resistance has been recently recognized. Bacteriophages (= phages) are effective tools by which to control insect commensal bacteria, but there have as yet been no studies using phages on adult mosquitoes. In this study, we isolated an Aeromonas phage vB AhM-LH that specifically targets resistance-associated symbiotic bacteria in mosquitoes. We investigated the impact of Aeromonas phage vB AhM-LH in an abundance of Aeromonas hydrophila in the gut of Cx. pipiens pallens and its effect on the status of deltamethrin resistance. METHODS: Phages were isolated on double-layer agar plates and their biological properties analyzed. Phage morphology was observed by transmission electron microscopy (TEM) after negative staining. The phage was then introduced into the mosquito intestines via oral feeding. The inhibitory effect of Aeromonas phage vB AhM-LH on Aeromonas hydrophila in mosquito intestines was assessed through quantitative real-time PCR analysis. Deltamethrin resistance of mosquitoes was assessed using WHO bottle bioassays. RESULTS: An Aeromonas phage vB AhM-LH was isolated from sewage and identified as belonging to the Myoviridae family in the order Caudovirales using TEM. Based on biological characteristics analysis and in vitro antibacterial experiments, Aeromonas phage vB AhM-LH was observed to exhibit excellent stability and effective bactericidal activity. Sequencing revealed that the Aeromonas phage vB AhM-LH genome comprises 43,663 bp (51.6% CG content) with 81 predicted open reading frames. No integrase-related gene was detected in the vB AH-LH genome, which marked it as a potential biological antibacterial. Finally, we found that Aeromonas phage vB AhM-LH could significantly reduce deltamethrin resistance in Cx. pipiens pallens, in both the laboratory and field settings, by decreasing the abundance of Aeromonas hydrophila in their midgut. CONCLUSIONS: Our findings demonstrate that Aeromonas phage vB AhM-LH could effectively modulate commensal bacteria Aeromonas hydrophila in adult mosquitoes, thus representing a promising strategy to mitigate mosquito vector resistance.

Enhancing immune and antioxidant responses in Nile tilapia through dietary supplementation with Ulva fasciata extract: A study on gene expression and resistance to Aeromonas hydrophila.

BACKGROUND: The study focused on the impact of Ulva fasciata extract (UFE) supplementation in the diets of Nile tilapia (Oreochromis niloticus) on blood and biochemical markers, immune and oxidative responses, and the expression of related genes, with a specific interest in their condition following exposure to Aeromonas hydrophila. METHODS: Four different levels of UFE were tested in the diets: 0% (0 mg kg- 1) for the control group (U0), and incremental additions of 0.05% (50 mg kg-1), 0.1% (100 mg kg-1), and 0.15% (150 mg kg-1) for the experimental groups U50, U100, and U150 respectively. Groups of 45 fish weighing 3.126 ± 0.120 g were fed these diets over 90 days. RESULTS: The study found that groups treated with UFE showed statistically significant enhancements (p < 0.05) compared to the control group. These improvements included increased red and white blood cell counts, higher haemoglobin concentrations, greater packed cell volume, and elevated enzyme activities-specifically, superoxide dismutase, catalase, alanine aminotransferase, and aspartate aminotransferase. Additionally, lysozyme and phagocytic activities were notably higher, especially in the U100 group after exposure. Before exposure to Aeromonas hydrophila, all levels of UFE supplementation led to increased expression of TNF-α and COXII genes and decreased NFκ-B expression. After the challenge, UFE intake resulted in varied expression levels of immune and antioxidant genes (TNF-α, NFκ-B, SOD, and COXII) in the liver, with the most effective responses observed in the U50, U100, and U150 groups. CONCLUSIONS: The findings underscore the potential of dietary UFE as a natural antioxidant and immune booster for Nile tilapia.

Probable Role of Type IV Pili of Aeromonas hydrophila in Human Pathogenicity.

BACKGROUND: Aeromonas hydrophila is a widely recognized broad-spectrum pathogen that primarily targets the gastrointestinal tract. Type IV pili (T4P) are proteinaceous nano-machines located on the bacterial cell surface, playing a crucial role in host colonization and infection. Regrettably, the T4P systems of A. hydrophila remain largely underexplored. METHODS: A. hydrophila genomes with complete genome assembly and annotation reports up to 31 March 2023, were obtained from the NCBI Genome database or KEGG genome database, followed by a global search for T4P secretion system genes. Protein sequences of these manually curetted genes were used as secondary quarry for Synteny analysis. Protein-protein interaction analysis was performed by string analysis and in silico study of genomic islands. RESULTS: We identified 27 orthologs of type IV pili (T4P) nano-machine components in A. hydrophila. These orthologs are primarily distributed across three operons: pilABCD, pilMNOPQ, and pilVWXY. While the first two operons are commonly found in all experimental genomes, the presence of the pilVWXY operon, coding for 11 orthologs, is reported here for the first time in A. hydrophila. Notably, the complete pilVWXY operon is absent in nonvirulent strains. A genomic islands study between a nonvirulent and hypervirulent strain also confirms absence of most of the genes coded by pilVWXY in nonvirulent strain. Interestingly, among the 51 experimental genomes analyzed, the pilVWXY operon was completely absent in 10 strains, most of which are categorized as nonvirulent; Conclusions: The distribution of two major type IV pili (T4P) nano-machines, PilABCDMNOPQ and PilVWXY, is reported here for the first time in A. hydrophila. Additionally, this study suggests a potential role for the PilVWXY nano-machine in establishing human disease.

Probiotic Efficacy in Aquaculture: The Role of Technospore® (Bacillus coagulans) in Improving Nile Tilapia (Oreochromis niloticus) Performance and Disease Resistance: a Study on Gut Health, Immunological Response, and Gene Expression.

This study evaluated the effects of Technospore® (Bacillus coagulans) supplementation on intestinal health, immune response, and Oreochromis niloticus (Nile tilapia) growth performance. The experiment divided fish into four groups: a control group fed an unsupplemented diet and three experimental groups receiving diets supplemented with 0.2 g/kg, 0.4 g/kg, and 0.8 g/kg of Technospore®, respectively. Results indicated that Technospore® supplementation significantly enhanced growth rates and feed efficiency in all treated groups, with the most pronounced improvements observed in the group receiving 0.4 g/kg. Furthermore, the study revealed that B. coagulans supplementation markedly boosted serum immune responses, as evidenced by increased phagocytic activity, phagocytic index, and lysozyme levels, following a challenge with Aeromonas hydrophila. Histological analysis showed improved gut morphology, while gene expression analysis indicated upregulation of immune-related genes, including liver IGF-1, GHR, HSP70, IL-1ß, and TNF-α, as well as spleen TNF-α and IL-1ß and intestinal C-lysozyme and TNF-α, both before and after the bacterial challenge. These findings suggest that dietary inclusion of Technospore® can significantly improve gut health and immune responses in tilapia, potentially serving as an effective prophylactic alternative to antibiotics in aquaculture.

Immunomodulatory, antioxidant, and growth-promoting activities of dietary fermented Moringa oleifera in Nile tilapia (Oreochromus niloticus) with in-vivo protection against Aeromonas hydrophila.

BACKGROUND: Moringa oleifera, a well-known medicinal plant, has been used in aquafeed as a dietary supplement. Based on previous studies, insufficient research is available on the dietary supplementation of Nile tilapia with M. oleifera leaf and seed mixtures, specifically the fermented form. Therefore, this study aimed to investigate the efficacy of fermented (FMO) versus non-fermented M. oleifera (MO) leaf and seed mixtures on immunological parameters, antioxidant activity, growth performance, and resistance to A. hydrophila infection after a 30-day feeding trial on Nile tilapia. METHODS: A total of 180 fingerlings were randomly divided into four groups in addition to the control group (36 fish each, in triplicate). Fish in the tested groups were fed on basal diet supplemented with MO5%, MO10%, FMO5%, and FMO10%, while those in control were fed on basal diet only. After the feeding trial, fish were challenged with A. hydrophila. The immunomodulatory activity of M. oleifera was evaluated in terms of phagocytic and lysozyme activities, immune-related cytokines and IgM gene expression. Antioxidants, and growth-promoting activities were also assessed. RESULTS: The results revealed that fish supplemented FMO markedly in FMO10% group followed by FMO5%, exhibited significant (P < 0.05) improvement in the tested immunological, hepatic antioxidants, and growth performance parameters. Furthermore, the highest survival rate post-challenge with mild clinical symptoms, and the lowest A. hydrophila bacterial count were reported in these groups. Meanwhile, MO10%-supplementation exhibited the opposite trend. CONCLUSIONS: The study' conclusion suggests that fermented M. oleifera leaf and seed mixture is a promising growth-promoting and immunostimulatory feed-additive candidate for Nile tilapia and could reduce the losses caused by A. hydrophila infection.