Aeromonas Hydrophila Research Articles

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.

Ice nucleation active bacteria metabolites as antibiofilm agent to control Aeromonas hydrophila and Streptococcus agalactiae infections in Aquaculture

ObjectivesThe aim of this study was to quantify and identify metabolites of Ice Nucleation Active (INA) bacteria as an anti-biofilm agent against biofilms of fish pathogens such as Aeromonas hydrophila and Streptococcus agalactiae.ResultsIce nucleation active bacteria, which have the ability to catalyze ice nucleation, isolated from rainwater in previous studies, were used. All INA isolates were tested in several assays, including the antimicrobial test, which uses streptomycin as the positive control and none of the isolates were found positive in the antimicrobial test. As for the quorum quenching assay, it was found that four out of ten isolates were able to disturb the communication system in Chromobacterium violaceum wild type, which was used as the indicator bacteria. On the next assay, all ten isolates were tested for Biofilm Inhibition and Destruction and showed anti-biofilm activity with the highest percentage inhibition of 33.49% by isolate A40 against A. hydrophila and 77.26% by isolate A19 against S. agalactiae. C1 performed the highest destruction against A. hydrophila and S. agalactiae, with percentages of 32.11% and 51.88%, respectively. As for the GC-MS analysis, supernatants of INA bacteria contain bioactive compounds such as sarcosine and fatty acids, which are known to have antibiofilm activity against several biofilm-forming bacteria. Through 16s rRNA sequencing, identified bacteria are from the Pantoea, Enterobacter, and Acinetobacter genera. As for the conclusion, ice nucleation active bacteria metabolites tested showed positive results against pathogenic bacteria Aeromonas hydrophila and Streptococcus agalactiae in destructing and inhibiting biofilm growth.

Exogenous NADH promotes the bactericidal effect of aminoglycoside antibiotics against Edwardsiella tarda

ABSTRACT The global surge in multidrug-resistant bacteria owing to antibiotic misuse and overuse poses considerable risks to human and animal health. With existing antibiotics losing their effectiveness and the protracted process of developing new antibiotics, urgent alternatives are imperative to curb disease spread. Notably, improving the bactericidal effect of antibiotics by using non-antibiotic substances has emerged as a viable strategy. Although reduced nicotinamide adenine dinucleotide (NADH) may play a crucial role in regulating bacterial resistance, studies examining how the change of metabolic profile and bacterial resistance following by exogenous administration are scarce. Therefore, this study aimed to elucidate the metabolic changes that occur in Edwardsiella tarda (E. tarda), which exhibits resistance to various antibiotics, following the exogenous addition of NADH using metabolomics. The effects of these alterations on the bactericidal activity of neomycin were investigated. NADH enhanced the effectiveness of aminoglycoside antibiotics against E. tarda ATCC15947, achieving bacterial eradication at low doses. Metabolomic analysis revealed that NADH reprogrammed the ATCC15947 metabolic profile by promoting purine metabolism and energy metabolism, yielding increased adenosine triphosphate (ATP) levels. Increased ATP levels played a crucial role in enhancing the bactericidal effects of neomycin. Moreover, exogenous NADH promoted the bactericidal efficacy of tetracyclines and chloramphenicols. NADH in combination with neomycin was effective against other clinically resistant bacteria, including Aeromonas hydrophila, Vibrio parahaemolyticus, methicillin-resistant Staphylococcus aureus, and Listeria monocytogenes. These results may facilitate the development of effective approaches for preventing and managing E. tarda–induced infections and multidrug resistance in aquaculture and clinical settings.

Screening of Probiotic Candidates Bacteria as Biocontrol of Aeromonas Hydrophila Pathogen Isolated from Mina Padi Cultivation Area

The agricultural activities in rice-farming system can have an impact on fish farming and the characteristics of bacterial community, especially pathogenic bacteria. Utilization of probiotics as environmentally friendly biotechnology products can be used to improve environmental quality and suppress the presence of pathogenic bacteria. This study aimed to select bacteria as probiotic agents from aquaculture ponds with the Rice-fish farming system. A total of 22 bacterial isolates were isolated from the water and sediment contained in the culture ponds. Based on the screening results, 15 isolates were confirmed as general non-pathogenic bacteria (Aeromonas sp.), 9 isolates had antibacterial activity against Aeromonas hyrophylla and 4 isolates showed high antibiotic sensitivity and were able to synergize. The results showed the Proteus mirabilis, Proteus penneri, Kurthia gibsonii and Bacillus cereus strains. Bacillus cereus strain LB8 has antibacterial activity that can inhibit the pathogen Aeromonas hydrophila with an inhibition zone of 8mm and has a very high sensitivity to antibiotics. These four isolates are able to work together synergistically and can be used as consortium probiotic bacterial agents to suppress the growth of pathogens.

The occurrence of potential pathogenic antibiotic-resistant bacteria in the marine environment in Ha Long Bay, Vietnam

This study aimed to investigate the occurrence of potential pathogenic bacteria from surface water samples collected at 30 stations along Ha Long Bay, Vietnam. Among 116 bacterial isolates identified, the composition analysis indicated 24 distinct bacterial genera, with Vibrio (29.3%), Pseudomonas (23.3%), Staphylococcus (8.6%), and Bacillus (6.0%) being the most dominant. The taxonomic analysis revealed 61/116 (52.6%) isolates as potential pathogens (risk group 2) belonging to 21 bacterial species. Potential pathogens were detected in 28/30 (93.3%) sampling stations, with the most common pathogens including Vibrio alginolyticus (n=25), Vibrioparahaemolyticus (n=5),Enterobacter cloacae (n=3), Pseudomonas mendocina (n=3), Aeromonas hydrophila (n=2), Enterobacter asburiae (n=2), Klebsiella oxytoca (n=2), Bacillus cereus (n=2),and Staphylococcus haemolyticus (n=2). The selected pathogens were highly resistant to Amoxicillin (100%), Piperacillin (100%), Amoxicillin + Clavulanate (85.7%), Ticarcillin (71.4%), Cefoxitin (54.4%), Kanamycin (50%), and Fosfomycin (50%), and less resistant to Trimethoprim + sulfamethoxazole, Fusidic Acid, Tetracycline, Erythromycin, Pristinamycin, Dalacin, and Rifampicin (<25%), and susceptible to the remaining antibiotics. Notably, 12/23 (52.2%) isolates were resistant to at least three antibiotics and were multidrug-resistant bacteria. Our findings underline that the occurrence of pathogenic antibiotic-resistant bacteria in the marine environment of Ha Long Bay presents a potential threat to human health. Further genomic surveillance studies are necessary to manage and control the emergence and spread of pathogenic-resistant bacteria in this area.

Assessment of Conservation Status of Petroleuciscus Borysthenicus Celensis From Gurban River, Romania by Identification of Parasites and Bacteria

The present research was conducted within the framework of a broader investigation with the objective of identifying freshwater parasites and bacteria on national territory, with the aim of gaining a deeper understanding of the relationships between parasites, hosts, and the environment, and to assess the potential detrimental effects of parasitic infestation on the conservation status of fish populations. Within this study, it was documented the first recordings of parasites Vorticella globularia and Epistylis sp. in the Romanian Petro-leuciscus borysthenicus celensis fish species. A total of 42 specimens were collected from the Gurban River to study their infestation status, focusing on the skin, gills, and fins. From the samples analyzed, 16 specimens presented signs of infestation with five distinct parasite species (Ichthyophthirius multifiliis, Dactylogyrus vastator, Trichodina acuta, Vorticella globularia and Epistylis sp) and four bacterial strains (Aeromonas veronii, Shewanella putrefaciens, Aeromona hydrophila, and Citrobacter freundii). The results indicate that the skin is the organ most severely impacted by parasites and bacteria, followed by the gills. The fins, on the other hand, are the least susceptible to infection. Furthermore, the significant amount of parasitic infestation coupled with the high bacterial load of Aeromonas veronii indicates a plausible link between the two.

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

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. 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. 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. 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.

Effect of wogonin, an active ingredient of Scutellariae Radix, on fish infected with Aeromonas hydrophila

Effect of wogonin, an active ingredient of Scutellariae Radix, on fish infected with Aeromonas hydrophila

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-β), autophagy factors (ATG6, ATG8) and immune-related pathway factors (P65, P38), and down-regulating pro-inflammatory factors (IL-1β, 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.

Immune protection of grass carp by oral vaccination with recombinant Bacillus methylotrophicus expressing the heterologous tolC gene

In the field of aquaculture, the enhancement of animal health and disease prevention is progressively being tackled using alternatives to antibiotics, including vaccines and probiotics. This study was designed to evaluate the potential of a recombinant Bacillus methylotrophicus, engineered to express the outer membrane channel protein TolC of Aeromonas hydrophila AH3 and the green fluorescent protein GFP, as an oral vaccine. Initially, the genes encoding tolC and GFP were cloned into a prokaryotic expression system, and anti-TolC mouse antiserum was generated. Subsequently, the tolC gene was subcloned into a modified pMDGFP plasmid, which was transformed into B. methylotrophicus WM-1 for protein expression. The recombinant B. methylotrophicus BmT was then administered to grass carp via co-feeding, and its efficacy as an oral vaccine was assessed. Our findings demonstrated successful expression of the 55 kDa TolC and 28 kDa GFP proteins, and the preparation of polyclonal antibodies with high specificity. The BmT exhibited stable expression of the GFP-TolC fusion protein and excellent genetic stability. Following oral immunization, significant elevations were observed in serum-specific IgM levels and the activities of acid phosphatase (ACP), alkaline phosphatase (AKP), superoxide dismutase (SOD), and lysozyme (LZM) in grass carp. Concurrently, significant upregulation of immune-related genes, including IFN-I, IL-10, IL-1β, TNF-α, and IgT, was noted in the intestines, head kidney, and spleen of the grass carp. Colonization tests further revealed that the BmT persisted in the gut of immunized fish even after a fasting period of 7 days. Notably, oral administration of BmT enhanced the survival rate of grass carp following A. hydrophila infection. These results suggest that the oral BmT vaccine developed in this study holds promise for future applications in aquaculture.

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.

Meningitis caused by Aeromonas hydrophila in Oreochromis niloticus: Proteomics and druggability of virulence factors

Meningitis caused by Gram-negative bacteria is a serious public health problem, causing morbidity and mortality in both children and adults. Here, we propose a novel experimental model using Nile tilapia (Oreochromis niloticus) to study neuroinflammation. The fish were infected with Aeromonas hydrophila, and the course of infection was monitored in the peripheral blood. Septicemia was obvious in the blood, while in the brain tissue, infection of the meninges was present. The histopathological examination showed suppurative meningitis, and the cellular immune response in the brain tissue during infection was mediated by microglia. These cells were morphologically characterized and phenotyped by MHC class II markers and CD68. The increased production of TNF-α, IL-1β and iNOS supported the infiltration of these cells during the neuroinflammatory process. In the proteomic analysis of A. hydrophila isolated from brain tissue, we found chemotactic and transport proteins, proteolytic enzymes and enzymes associated with the dismutation of nitric oxide (NO), as well as motor proteins and those responsible for cell division. After characterizing the most abundant proteins during the course of infection, we investigated the druggability index of these proteins and identified promising peptide sequences as molecular targets that are similar among bacteria. Thus, these findings deepened the understanding of the pathophysiology of meningitis caused by A. hydrophila. Moreover, through the proteomics analysis, important mechanisms and pathways used by the pathogen to subvert the host response were revealed, providing insights for the development of novel antibiotics and vaccines.

Microbiological quality assessment of Clarias gariepinus, Bagrus bajad, and Pangasianodon hypophthalmus fillets

In this study, 80 catfish fillets were randomly collected from Egyptian local markets and retailers. The samples included 20 African catfish (Clarias gariepinus), 20 bayad (Bagrus bajad), and 40 pangasius catfish (Pangasianodon hypophthalmus) fillets. Pangasianodon hypophthalmus fillet samples were divided into 20 white basa and 20 red basa fillets. We conducted a microbiological analysis of catfish fillet samples, evaluating mesophilic aerobic bacteria, psychrophilic aerobic bacteria, H2S-producing bacteria, Staphylococcus spp., Enterobacteriaceae, Coliforms, and fecal Coliform counts. Additionally, we identified the existence of Salmonella spp., Vibrio spp., Yersinia spp., Escherichia spp., Aeromonas spp., and Pseudomonas spp. in the catfish fillet samples. In our study, the psychrophilic bacterial counts in Bagrus bajad (5.21 log CFU/g) were found to be higher compared to the counts in Clarias gariepinus (4.31 log CFU/g) and Pangasianodon hypophthalmus (3.89–4.7 log CFU/g). The fecal Coliform in Clarias gariepinus fillets was significantly higher than in other catfish fillets. We isolated Escherichia coli, Escherichia fergusonii, Aeromonas hydrophila, and Pseudomonas luteola from the catfish fillets, while no Salmonella spp., Vibrio spp., or Yersinia spp. were detected. These isolates were identified using 16S rRNA sequencing and phylogenetic analysis. Furthermore, ten Escherichia spp. were serologically identified, revealing that O26 and O78 were the most commonly occurring serotypes. This study highlights the microbiological analysis conducted on catfish fillets and concludes that the fillet samples from these catfish were of superior quality and deemed acceptable for human consumption.

Target analyses by transcriptome sequencing in peripheral blood cells of grass carp: Potent immune responses in leukocytes and thrombocytes against bacterial and viral infections

Fishes provide abundant protein for humanity, while they are ravaging by bacterial and viral diseases. The defense based on peripheral blood cell immunity is an important means to resist pathogenic infections. However, the immune response of different blood cell communities to bacterial or viral infection remains largely unknown. In this study, grass carp (Ctenopharyngodon idella), the most productive fish in the world, was employed to comprehensively investigate the immune responses of erythrocytes, leukocytes, and thrombocytes after Aeromonas hydrophila or type II grass carp reovirus (GCRV-II) infection by transcriptome sequencing. The data of 45 samples with small intra-group and large inter-group differences were obtained. There were 3921, 15,378, 7019 DEGs in the infected erythrocytes, leukocytes, and thrombocytes, respectively. Through GO and KEGG analysis, they were mainly enriched in the items related to innate and adaptive immunity, including pattern recognition receptor signaling pathways and cytokine signaling pathways. Therefore, we conducted target analyses of vital immune families, including TLRs, RLRs, IFNs, IRFs, immune activators, and antigen presentation genes, systematically clarified the immune responses of blood cells to pathogenic infections. All the three types of blood cells expressed a large number of immune genes involved in anti-infection to varying degrees, among them leukocytes and thrombocytes responded aggressively. This study provides implications for the function of peripheral blood cells and the functional basis of immune genes in pathogenic infections.

Phenotypic, genotypic and virulence traits analysis of aeromonads causing massive mortality in farmed Oreochromis niloticus

Motile aeromonads were identified in earthen-pond-farmed Oreochromis niloticus that suffered massive mortalities in Egypt during the summer of 2020. The fish showed hemorrhagic septicemic signs. Poor management practices and inadequate water quality measures were observed in the affected earthen ponds. Motile aeromonads (n = 31 isolates) were identified from 70 fish specimens. Based on their phenotypic and molecular characteristics, isolates were identified as: Aeromonas hydrophila (n = 12), A. veronii (n = 10), A. caviae (n = 5), and A. sobria (n = 4). Bacteriological examination of farm water samples also revealed aeromonads (n=9) and some fish-nonpathogenic bacteria. The aeromonad isolates recovered from fish exhibited lipase (52.5%) and protease (47.5%) activities and harboured some virulence genes: Ser (62.5%), Aer (55%), ela (37.5%), gcaT (32.5%), Hyl (25%), laf-A (22.5%), and Act (20%). They also harboured numerous antibiotic-resistance genes, including aadA (37.5%), tetC (32.5%), tetA (27.5%), sul 1 (20%), and blaTEM (10%). Virulence and antibiotic resistance genes were also noted in some of the Aeromonas spp. isolates obtained from farm water. Aeromonads were highly resistant to ampicillin, amoxicillin, and gentamicin but highly susceptible to ciprofloxacin and florfenicol antibiotics. Aeromonas spp pathogenicity was confirmed by the experimental infection of Oreochromis niloticus. Our results indicate a positive correlation between excessive tilapia mortalities, motile Aeromonas septicemia and adverse water quality parameters measured during the summer. This study provides data on the virulence, pathogenicity, and antibiotic resistance of motile aeromonads affecting fish and humans, which will be useful for developing efficient therapies.

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.

Structure–activity relationships of the intramolecular disulphide bonds in LEAP2, an antimicrobial peptide from Acrossocheilus fasciatus

BackgroundThe liver-expressed antimicrobial peptide 2 (LEAP2) plays a pivotal role in the host’s immune response against pathogenic microorganisms. Numerous such antimicrobial peptides have recently been shown to mitigate infection risk in fish, and studying those harboured by the economically important fish Acrossocheilus fasciatus is imperative for enhancing its immune responses against pathogenic microorganisms. In this study, we cloned and sequenced LEAP2 cDNA from A. fasciatus to examine its expression in immune tissues and investigate the structure–activity relationships of its intramolecular disulphide bonds.ResultsThe predicted amino acid sequence of A. fasciatus LEAP2 was found to include a signal peptide, pro-domain, and mature peptide. Sequence analysis indicated that A. fasciatus LEAP2 is a member of the fish LEAP2A cluster and is closely related to Cyprinus carpio LEAP2A. A. fasciatus LEAP2 transcripts were expressed in various tissues, with the head kidney exhibiting the highest mRNA levels. Upon exposure to Aeromonas hydrophila infection, LEAP2 expression was significantly upregulated in the liver, head kidney, and spleen. A mature peptide of A. fasciatus LEAP2, consisting of two disulphide bonds (Af-LEAP2-cys), and a linear form of the LEAP2 mature peptide (Af-LEAP2) were chemically synthesised. The circular dichroism spectroscopy result shows differences between the secondary structures of Af-LEAP2 and Af-LEAP2-cys, with a lower proportion of alpha helix and a higher proportion of random coil in Af-LEAP2. Af-LEAP2 exhibited potent antimicrobial activity against most tested bacteria, including Acinetobacter guillouiae, Pseudomonas aeruginosa, Staphylococcus saprophyticus, and Staphylococcus warneri. In contrast, Af-LEAP2-cys demonstrated weak or no antibacterial activity against the tested bacteria. Af-LEAP2 had a disruptive effect on bacterial cell membrane integrity, whereas Af-LEAP2-cys did not exhibit this effect. Additionally, neither Af-LEAP2 nor Af-LEAP2-cys displayed any observable ability to hydrolyse the genomic DNA of P. aeruginosa.ConclusionsOur study provides clear evidence that linear LEAP2 exhibits better antibacterial activity than oxidised LEAP2, thereby confirming, for the first time, this phenomenon in fish.

Inactivation of Aeromonas hydrophila in water by dielectric barrier discharge plasma: Efficiency, mechanism, and application safety

Aeromonas hydrophila (A. hydrophila) is a typical zoonotic pathogen that commonly found in water environments, capable of infecting various aquatic animals and causing different diseases in both animals and humans. In this study, the dielectric barrier discharge plasma (DBDP) is used to inactivate the A. hydrophila in water. The efficiency and mechanism of A. hydrophila inactivation by DBDP were investigated, and the application safety of DBDP also were examined. The results demonstrate that DBDP treatment for 8 min can reduce the concentration of A. hydrophila from 108 CFU/mL to 0 CFU/mL under optimal conditions. Higher discharge voltage and lower concentration is beneficial for the inactivation of A. hydrophila, and O2 as carrier gas and alkaline environment can improve the inactivation efficiency. Reactive oxygen species (ROS) produced in the DBDP system play a significant role in A. hydrophila inactivation. However, molecules like O3 and H2O2 long-lived are likely the main forces responsible for the initial destruction of cell membranes. Subsequently, ROS penetrates the cell membrane, and result in the destruction of main virulence genes of A. hydrophila and the loss of bacterial infectivity. The silver carp challenge experiment indicates that mortality rates and septicemia symptoms in silver carp are not significantly different from the control group when treated with DBDP. Moreover, the total bacterial count in the main organs can be reduced to a safe level. These findings suggest that DBDP is a safe and reliable method for inactivating A. hydrophila in water bodies.

The Effect of Adding Chitosan Nanoparticles Loaded with Sidr Leaf Extract (Ziziphus spina-christi) (CPNs SLE) in Diet on Pangasius sp. Infected Aeromonas hydrophila

One of the bacterial types that affects freshwater fish is Aeromonas hydrophila. These bacteria cause the occurrence of Motile Aeromonas Septicemia (MAS) disease. This study investigates the impact and optimal dosage of chitosan nanoparticles (CNPs) loaded with sidr leaves extract (SLE) on clinical manifestations, growth, hematological parameters, and survival rates of Pangasius sp. infected A. hydrophila. The experimental design employed a Completely Randomized Design with five treatments and three replications, utilizing 150 fish with an average length of 7-9 cm. Treatments included the addition of 10 ppm tetracycline (K+) in the diet and CNPs SLE with concentrations A (0 ppm), B (450 ppm), C (500 ppm), and D (550 ppm). The results demonstrate that incorporating CNPs SLE into the diet yields beneficial effects on infected Pangasius sp., leading to faster recovery from clinical symptoms, enhanced growth rates, and increased survival rates of up to 85%. The optimal dosage of CNPs SLE for Pangasius sp. infected A. hydrophila in the feed is determined to be 500 ppm.

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 β-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.