Pathogens In Aquaculture Research Articles

Production and characterization of monoclonal antibodies against mandarinfish ranavirus and first identification of pyloric caecum as the major target tissue.

Mandarinfish ranavirus (MRV), also known as a variant of largemouth bass virus (LMBV), is an emerging pathogen in mandarinfish aquaculture. In this study, monoclonal antibodies (mAbs) against MRV were produced and characterized, and 7 mAbs were obtained through Western blotting screening and all 7 mAbs specifically recognized MRV/LMBV but not several piscine iridoviruses as ISKNV, GIV and TFV. By LC MS/MS analysis, the recognized viral proteins by seven mAbs were identified as MRV-pORF47L, MRV-pORF55R, MRV-pORF57L, MRV-pORF77L and MRV-pORF78L, respectively, and all five viral proteins are late expression structural proteins by Western blotting. Based on mAb 1C4, immuno-histochemistry and immuno-histo-fluorescence were performed to re-assess the tissue tropism of MRV. The result showed that abundant reactive signals were observed in infected spleen, kidney as well as intestine and pyloric caecum. Real-time quantitative PCR also demonstrated that spleen as well as pyloric caecum and intestines are the major target tissue upon MRV infection. In infected intestines and pyloric caecum, numerous enlarged, multinucleated cells with intracytoplasmic inclusions were identified as the target cells of MRV, suggesting that MRV serves as a digestive tract pathogen to mandarinfish, which may explain why acute infection of MRV can cause the typical clinicopathology featured by severe ascites.

Detection of Fish Pathogens in Freshwater Aquaculture Using eDNA Methods

Organisms release their nucleic acid in the environment, including the DNA and RNA, which can be used to detect their presence. eDNA/eRNA techniques are being used in different sectors to identify organisms from soil, water, air, and ice. The advancement in technology led to easier detection of different organisms without impacting the environment or the organism itself. These methods are being employed in different areas, including surveillance, history, and conservation. eDNA and eRNA methods are being extensively used in aquaculture and fisheries settings to understand the presence of different fish species and pathogens in water. However, there are some challenges associated with the reliability of results because of the degradation of nucleic acid by several factors. In aquaculture, there are several diseases and parasites detected with these methods. In this review, we discuss different aquaculture diseases and parasites detected with eDNA/eRNA approach and the fate of these nucleic acids when subjected to different water quality and environmental parameters. This review intends to help the researcher with the potential of eDNA/eRNA-based detection of pathogens in aquaculture; this will be useful to predict a potential outbreak before it occurs. Along with that, this paper intends to help people understand several factors that degrade and can hamper the detection of these nucleic acids.

LncRNA MIR144HG-derived miR-144 suppresses antibacterial signaling and facilitates bacteria escape in fish

Growing number studies show that lncRNA is an important regulatory factor and plays an irreplaceable role in innate immunity. However, most studies on lncRNA have focused on mammals and it is unknown in lower vertebrates. In this study, we identify a new lncRNA, named MIR144HG, which can have a negative role in antibacterial immunity of Miichthys miiuy caused by Vibrio anguillarum and V. harveyi. MyD88, TAK1 and p65 are very important pathway genes in antibacterial innate immune responses, whether in mammals or lower vertebrates. Here, we found that miR-144–3p could target MyD88, TAK1 and p65, inhibit host antibacterial response and promote bacterial escape. Further studies found that miR-144–3p can be derived by MIR144HG, and finally enhance the inhibitory effect of miR-144–3p on host antibacterial immunity. Moreover, V. anguillarum and V. harveyi are the two most susceptible Gram-negative pathogens in aquaculture, and the economic losses caused by these two bacteria are immeasurable every year. Our results not only elucidate the mechanism of the lncRNA-miRNA-mRNA axis in antibacterial immune responses, but also provide new insights for understanding the impact of lncRNA on host immunity and bacteria escape.

Screening and identification of lactic acid bacteria with antimicrobial abilities for aquaculture pathogens in vitro

This study aimed to evaluate the antibacterial properties of nine lactic acid bacterial strains. The agar diffusion method (Oxford cup method) was used to assess the antimicrobial activity against pathogenic bacteria in aquaculture. The results showed that all selected strains inhibited the growth of Photobacterium damsel, Vibrio alginolyticus, Listonella anguillarum, Edwardsiella ictaluri, Aeromonas hydrophila, V. arahaemolyticus, Edwardsiella tarda and V. harveyi, but did not inhibit the growth of Metschnikowia bicuspidata. Among these strains of lactic acid bacteria, r1 was selected for its obvious antibacterial activity against eight kinds of pathogenic bacteria by 96-well plate method. Its inhibitory rate reached 96%, and it could inhibit the growth of six pathogenic bacteria at 121℃, 20min and pH 3.5-4.0. It was identified as Lactiplantibacillus plantarum by morphological observation and 16S rRNA sequencing analysis. Because strain r1 was isolated from culture ponds and exerted strong inhibitory effects on pathogenic bacteria, it holds potential as an agent to prevent and control infectious diseases in aquaculture. This study provides a foundation for the development and utilization of probiotics in aquaculture.

Dactylogyrus extensus and Pseudomonas fluorescens dual infection in farmed common carp (Cyprinuscarpio)

Dactylogyrus extensus and Pseudomonas fluorescens are serious pathogens in Cyprinus carpio aquaculture causing severe impacts and substantial economic losses. During the early spring of 2021, abnormal mortalities were reported among farmed C. carpio. Moribund fish showed anorexia, respiratory distress, dermal ulcers, and septicemia. The water analysis revealed low dissolved oxygen (3.4 mg/L), and high un-ionized ammonia levels (0.65 mg/L). Seventy moribund C. carpio specimens were collected and subjected to parasitological and bacteriological examinations. The monogenetic trematode D. extensus was discovered in wet mounts from the gills of all the examined fish samples (100%). The identity of recovered parasites was confirmed by sequencing and alignment of the 28S rDNA gene. P. fluorescens was concurrently identified in the infested fish samples (58.5%) based on phenotypic characteristics using the API20 E. The identity of bacterial isolates was confirmed further by sequencing and alignment of 16S rRNA gene. The IL-1β and MHCII were upregulated in infested fish in tandem with the severity of infections. P. fluorescens isolates displayed high resistance to most of the tested antibiotics. The study is one of the earlier reports on D. extensus and P. fluorescens co-infections in farmed C. carpio and highlights the need of effective control programs to protect fish health and minimize losses.

Dietary supplementation of Pseudoalteromonas piscicida 1UB and fructooligosaccharide enhance growth performance and protect the whiteleg shrimp (Litopenaeus vannamei) against WSSV and Vibrio harveyi coinfection

P. piscicida 1Ub and FOS were evaluated for their potential synbiotic effects on growth, immunological responses, and disease resistance against white spot syndrome virus and V. harveyi coinfection, the major pathogen in whiteleg shrimp aquaculture. Four different supplemented diets were used to feed the experimental shrimp for 40 days: control (control, no probiotic, and prebiotic), probiotic (PRO, P. piscisida 1UB 108 CFU mL−1), prebiotic (PRE, FOS 0.5% w/w), and the synbiotic (SYN, PRO + PRE). Shrimp's body weight, weight gain, specific growth rate, feed conversion ratio, survival, digestive enzyme activity, and metabolism-related gene expression were all evaluated on day 40. After 40 days, shrimp were infected with WSSV as the primary infection and V. harveyi as the secondary infection 24 h later. Shrimp were then grown for seven days and fed with a control diet. Survival, total hemocyte count (THC), differential hemocyte, phenol-oxidase (PO), respiratory burst activity (RB), and immune-gene expression were all analyzed at 0, 3, and 7 days after infection. The results showed that the PRO, PRE, and SYN supplementation improves whiteleg shrimp growth performance, immune responses, and protection against WSSV and V. harveyi coinfection. The increased activity of digestive enzymes and metabolism-related genes correlates with higher growth performance. The increase in THC, PO, RB, and immune-related gene expression after coinfection was associated with a significant reduction in shrimp mortality. Our findings also suggest that supplementing with synbiotics improves the overall performance of whiteleg shrimp significantly more than probiotics or prebiotics only.

Inhibition of microbial pathogens in farmed fish.

Aquaculture, also known as aqua farming, is defined as farming fish, crustaceans, mollusks, aquatic plants, algae, and other marine organisms. It includes cultivating fresh- and saltwater populations under controlled conditions compared to commercial fishing or wild fish harvesting. Worldwide, carp, salmon, tilapia, and catfish are the most common fish species used in fish farming in descending order. Disinfectants prevent and/or treat different infections in aquatic animals. The current review indicates the uses of different disinfectants against some important pathogens in aquaculture, with particular reference to tilapia (Oreochromis niloticus) farming. A single review cannot cover all aspects of disinfection throughout aquaculture, so the procedures and principles of disinfection in tilapia farming/aquaculture have been chosen for illustration purposes.

Valorization of agro-industrial wastes into polyhydroxyalkanoates-rich single-cell proteins to enable a circular waste-to-feed economy

Recovering and converting carbon and nutrients from waste streams into healthy single-cell proteins (SCPs) can be an effective strategy to address costly waste management and support the increasing animal feed demand for the global food supply. Recently, SCPs rich in polyhydroxybutyrate (PHB) have been identified as an effective biocontrol healthy feed to replace conventional antibiotics-supplemented aquaculture feed. PHB, an intercellular polymer of short-chain-length (SCL) hydroxy-fatty acids, is a common type of polyhydroxyalkanoates (PHA) that can be microbially produced from various organics, including agro-industrial wastes. The complex chemical properties of agro-industrial wastes might produce SCPs containing PHA with SCL and/or medium chain-length (MCL) hydroxy-fatty acids. However, the effects of MCL-PHA-containing SCPs on aqua species' health and disease-fighting ability remains poorly understood. This study investigated the feasibility of producing various PHA-containing SCPs from renewable agro-industrial wastes/wastewaters, the effectiveness of SCL- and MCL-PHA as biocontrol agents, and the effects of these PHA-rich SCPs on the growth and disease resistance of an aquaculture animal model, brine shrimp Artemia. Zobellella denitrificans ZD1 and Pseudomonas oleovorans were able to grow on different pure substrates and agro-industrial wastes/wastewaters to produce various SCL- and/or MCL-PHA-rich SCPs. Low doses of MCL-fatty acids (i.e., PHA intermediates) efficiently suppressed the growth of aquaculture pathogens. Moreover, MCL-PHA-rich SCPs served as great food/energy sources for Artemia and improved Artemia's ability to fight pathogens. This study offers a win-win approach to address the challenges of wastes/wastewater management and feed supply faced by the aquaculture industry.

Molecular characterisation of Flavobacterium columnare associated with columnaris disease in freshwater fishes with emphasis on virulence and antibiotic resistance pattern

Flavobacterium columnare is a major aquaculture pathogen that causes columnaris disease. However, only very limited studies have been conducted on its virulence and resistance pattern. Three pathogenic bacterial isolates of F. columnare (CIFRI-RCM8, CFCCSL66 and CFCCO41) were obtained from diseased fish (Labeo rohita, L. catla and Clarias magur) of culture farms in Odisha and West Bengal, India. Clinical signs such as tail rot, skin ulcer and gill discolouration, as well as soft ulceration across the entire body, irrespective of the fish size were noticed. The virulence of the isolates was confirmed by a recursive in vitro experiments. The pathological changes in infected L. rohita were studied to confirm the pathogenicity. Considerable difference in virulence was noticed when fishes were challenged with different F. columnare strains. Among the isolates, CIFRI-RCM8 was the most virulent,whereas CFCCO41 was the least virulent one. All of the isolates were found to be multidrug-resistant showing resistance to cefuroxime, ampicillin, cephalothin, amoxyclav, cefalexin, ceftazidime, carbenicillin, trimethoprim, clindamycin, ticarcillin and penicillin-G. The study suggested that F. columnare is one of the most important etiological agents of disease in freshwater fishes for which appropriate management protocols needs to be developed. Keywords: Antibiotics, Columnaris disease, Flavobacterium columnare, Molecular characterisation, Virulence

Halogenated Indoles Decrease the Virulence of Vibrio campbellii in a Gnotobiotic Brine Shrimp Model.

ABSTRACTIndole signaling is viewed as a potential target for antivirulence therapy against antibiotic-resistant pathogens because of its link with the production of virulence factors. This study examined the antimicrobial and antivirulence properties of 44 indoles toward Vibrio campbellii. Based on the results, 17 halogenated indole analogues were selected, as they significantly improved the survival of brine shrimp larvae challenged with V. campbellii. Specifically, 6-bromoindole, 7-bromoindole, 4-fluoroindole, 5-iodoindole, and 7-iodoindole showed a high protective effect, improving the survival of brine shrimp to over 80% even at a low concentration of 10 μM. To explore the impact of selected indole analogues on bacterial virulence phenotypes, swimming motility, biofilm formation, protease activity, and hemolytic activity of V. campbellii were determined. The results showed that all of the 17 selected indole analogues decreased swimming motility at both 10 μM and 100 μM. Most of the indole analogues decreased biofilm formation at a concentration of 100 μM. In contrast, only a slightly decreased protease activity and no effect on hemolytic activity were observed at both concentrations. To our knowledge, this is the first study of the structure-activity relation of halogenated indole analogues with respect to virulence inhibition of a pathogenic bacterium in an in vivo host model system, and the results demonstrate the potential of these compounds in applications aiming at the protection of shrimp from vibriosis, a major disease in aquaculture.IMPORTANCE Bacterial diseases are a major problem in the aquaculture industry. In order to counter this problem, farmers have been using antibiotics, and this has led to the evolution and spread of antibiotic resistance. In order for the aquaculture industry to further grow in a sustainable way, novel and sustainable methods to control diseases are needed. We previously reported that indole signaling is a valid target for the development of novel therapies to control disease caused by Vibrio campbellii and related bacteria, which are among the major bacterial pathogens in aquaculture. In the present study, we identified indole analogues that are more potent in protecting brine shrimp (a model organism for shrimp) from V. campbellii. To our knowledge, this is the first study of the structure-activity relation of halogenated indole analogues with respect to virulence inhibition of a pathogenic bacterium in an in vivo host model system.

Phytochemical and Antibacterial Effects of Leaf Extract from Mangrove Plant (Avicennia Marina) on Vibrio Parahaemolyticus in Shrimps

Recently, there has been a tremendous increase in the studies addressing the application of bioactive compounds from the natural ecosystem, particularly for medical purposes. Hence, the present study investigated the antibacterial properties of the secondary metabolites possibly contained in the leaves of Avicennia marina (A. marina) for possible prevention of Vibrio parahaemolyticus (V. parahaemolyticus), a devastating bacterial pathogen in shrimp aquaculture. In the current study, secondary metabolites were extracted from the leaves of mangrove plant using ethanol extraction method. The ethanolic extracts were then subjected to phytochemical and antibacterial activity tests. The results from the phytochemical analysis demonstrated that the ethanolic extract from the mangrove plant contained varying amounts of flavonoids, tannins, saponins, polyphenols, alkaloids, steroids, and triterpenoids. However, the number of flavonoids and alkaloids seemed to be higher than the other metabolites. The antibacterial activity analysis through the agar diffusion method has shown that different concentrations (50 ppm, 100 ppm, 200 ppm, and 300 ppm) of the ethanolic extract of A. marina inhibited the V. parahaemolyticus. At 300 ppm, the plant extract exhibited 17.3% antibacterial effectiveness, compared to the antibacterial activity of chloramphenicol. The findings indicated that the secondary metabolites of A. marina have the potential that can be developed as an alternative treatment for aquatic animal diseases in the future.

CONTROLLING AQUACULTURE PATHOGEN BIOFILM BY CRUDE EXTRACT AND SUPERNATANT OF PHYLLOSPHERE BACTERIA

Some infectious-aquaculture pathogens, such as Aeromonas hydrophila, Vibrio harveyi, and Streptococcus agalactiae are able to form biofilm, a protective structure that increases bacterial resistance to some external exposures, for example the antibiotic therapy, making the antibiotic becomes less effective. This ability is supported by the quorum sensing mechanism. Therefore, disruption in quorum sensing mechanism may be a potential way to treat the biofilm formation. The present study was conducted to assess the ability of supernatant and crude extract of some Actinobacteria and phyllosphere bacteria isolates to overcome the biofilms of several aquaculture pathogens. In general, this study consists of several steps, such as primary screening of anti-quorum sensing activity, extraction of bioactive compounds, antibiofilm assay, validation of anti-quorum sensing activity, toxicity assay, and microscopic observation of biofilm. The phyllosphere bacteria isolates showed higher antibiofilm activity compared to the Actinobacteria isolate, where the crude extract had the most ability instead of supernatant. The supernatant and crude extract of all isolates also performed the anti-quorum sensing activities, both qualitatively and quantitatively. Overall, all isolates were potential to be natural sources of antibiofilm agents to control the biofilm of all pathogens used in this study through quorum sensing disruption.

Gut and Gill-Associated Microbiota of the Flatfish European Plaice (Pleuronectes platessa): Diversity, Metabolome and Bioactivity against Human and Aquaculture Pathogens.

Similar to other marine holobionts, fish are colonized by complex microbial communities that promote their health and growth. Fish-associated microbiota is emerging as a promising source of bioactive metabolites. Pleuronectes platessa (European plaice, plaice), a flatfish with commercial importance, is common in the Baltic Sea. Here we used a culture-dependent survey followed by molecular identification to identify microbiota associated with the gills and the gastrointestinal tract (GIT) of P. platessa, then profiled their antimicrobial activity and metabolome. Altogether, 66 strains (59 bacteria and 7 fungi) were isolated, with Proteobacteria being the most abundant phylum. Gill-associated microbiota accounted for higher number of isolates and was dominated by the Proteobacteria (family Moraxellaceae) and Actinobacteria (family Nocardiaceae), whereas Gram-negative bacterial families Vibrionaceae and Shewanellaceae represented the largest group associated with the GIT. The EtOAc extracts of the solid and liquid media cultures of 21 bacteria and 2 fungi representing the diversity of cultivable plaice-associated microbiota was profiled for their antimicrobial activity against three fish pathogens, human bacterial pathogen panel (ESKAPE) and two human fungal pathogens. More than half of all tested microorganisms, particularly those originating from the GIT epithelium, exhibited antagonistic effect against fish pathogens (Lactococcus garvieae, Vibrio ichthyoenteri) and/or human pathogens (Enterococcus faecium, methicillin-resistant Staphylococcus aureus). Proteobacteria represented the most active isolates. Notably, the solid media extracts displayed higher activity against fish pathogens, while liquid culture extracts were more active against human pathogens. Untargeted metabolomics approach using feature-based molecular networking showed the high chemical diversity of the liquid extracts that contained undescribed clusters. This study highlights plaice-associated microbiota as a potential source of antimicrobials for the control of human and the aquaculture-associated infections. This is the first study reporting diversity, bioactivity and chemical profile of culture-dependent microbiota of plaice.

Pathogenicity and transcriptome analysis of a strain of Vibrio owensii in Fenneropenaeus merguiensis

Vibrio is an important conditional pathogen in shrimp aquaculture. This research reported a dominant bacteria strain E1 isolated from a shrimp tank with the method of biofloc culture, which was further identified as Vibrio owensii. To understand the interaction between V. owensii and the host shrimp, we studied the pathogenicity of the V. owensii and the molecular mechanisms of the Fenneropenaeus merguiensis immunity during the Vibrio invasion. Drug susceptibility tests showed that V. owensii was resistant to antibiotics streptomycin oxacillin, tetracycline, minocycline, and aztreonam, but highly sensitive to cefazolin, cefotaxime, and ciprofloxacin, and moderately sensitive to cefotaxime, ampicillin, and piperacillin. Lethal concentration 50 (LC50) test was performed to evaluate the toxicity of V. owensii to F. merguiensis. The LC50 of V. owensii infected F. merguiensis after 24, 48, 72, 96, 120, 144 and 168 h were 1.21 × 107, 1.68 × 106, 6.36 × 105, 2.15 × 105, 7.58 × 104, 5.55 × 104 and 4.33 × 104 CFU/mL. In order to explore the molecular response mechanism of F. merguiensis infected with V. owensii, the hepatopancreas of F. merguiensis were sequenced at 24 hpi and 48 hpi, and a total 40,181 of unigenes were obtained. Through comparative transcriptomic analysis, 86 differentially expressed genes (DEGs) (including 38 up-regulated DEGs, and 48 down-regulated DEGs) and 305 DEGs (including 150 up-regulated DEGs, and 155 down-regulated DEGs) were identified at 24 hpi and 48 hpi, respectively. Annotation and classification analysis of these 391 DEGs showed that most of the DEGs were annotated to metableolic and immune pathways, which indicated that F. merguiensis responded to the invasion through the regulation of material metableolism and immune system genes during V. owensii infection. In the KEGG enrichment analysis, some pathways related to immune response were significantly influenced by V. owensii infection, including phagosome, MAPK signalling pathway and PI3K-Akt signalling pathway. In addition, some pathways related to the warburg effect were also significantly enriched after V. owensii infection, including pyruvate metableolism, glycolysis/gluconeogenesis, and citrate cycle (TAC cycle). Further analysis showed that C-type lectins and ficolin were also play important roles in the immune response of F. merguiensis against V. owensii infection. The current research preliminarily revealed the immune response of F. merguiensis to V. owensii infection at the molecular level, which provided valuable information to further understand the disease control and the interaction between shrimp and Vibrio.

Techno-commercial application of microencapsulated basil oil and thyme oil and their antibacterial activity against aquatic pathogens

A complete study was conducted through manufacturing of Microencapsulation of Basil Oil Powder (MEBOP) and Microencapsulation of Thyme Oil Powder (METOP) to evaluate the anti-bacterial activity against aquatic pathogens. An oil-in-water emulsion were prepared using gum with dextrin and the resultant matrix was spray dried with the average yield of 45% w/w of MEBOP and 50% w/w of METOP. Total oil content in the encapsulated powder in both were found to be 24.39% w/w of MEBOP 25.80% w/w of METOP respectively. Scanning Electron Microscopic (SEM) analysis was also done to confirm the encapsulation of oil and study possible structure of matrix. Microencapsulated powders were further subjected for antibacterial activity against aquaculture pathogens like Vibrio harvey, Vibrio parahaemolyticus, Bacillus cereus, Aeromonas sp, E. coli and Salmonella sp. Results were compared against the respective oils at different concentrations. Phyto- constituents from Basil oil and Thyme oil like Thymol, Linalool, and Methyl chavicol were quantified using HPLC (High-Performance Liquid Chromatography) and GC (Gas Chromatography) which may be responsible for this activity. Furthermore, stress studies were conducted at 60 °C to understand the stability of Encapsulated oil powders to this and its commercial usage in formulations.

Single cell transcriptomics of Atlantic salmon (Salmo salar L.) liver reveals cellular heterogeneity and immunological responses to challenge by Aeromonas salmonicida.

The liver is a multitasking organ with essential functions for vertebrate health spanning metabolism and immunity. In contrast to mammals, our understanding of liver cellular heterogeneity and its role in regulating immunological status remains poorly defined in fishes. Addressing this knowledge gap, we generated a transcriptomic atlas of 47,432 nuclei isolated from the liver of Atlantic salmon (Salmo salar L.) contrasting control fish with those challenged with a pathogenic strain of Aeromonas salmonicida, a problematic bacterial pathogen in global aquaculture. We identified the major liver cell types and their sub-populations, revealing poor conservation of many hepatic cell marker genes utilized in mammals, while identifying novel heterogeneity within the hepatocyte, lymphoid, and myeloid lineages. This included polyploid hepatocytes, multiple T cell populations including γδ T cells, and candidate populations of monocytes/macrophages and dendritic cells. A dominant hepatocyte population radically remodeled its transcriptome following infection to activate the acute phase response and other defense functions, while repressing routine functions such as metabolism. These defense-specialized hepatocytes showed strong activation of genes controlling protein synthesis and secretion, presumably to support the release of acute phase proteins into circulation. The infection response further involved up-regulation of numerous genes in an immune-cell specific manner, reflecting functions in pathogen recognition and killing, antigen presentation, phagocytosis, regulation of inflammation, B cell differentiation and T cell activation. Overall, this study greatly enhances our understanding of the multifaceted role played by liver immune and non-immune cells in host defense and metabolic remodeling following infection and provides many novel cell-specific marker genes to empower future studies of this organ in fishes.

Aeromonas salmonicida aptamer selection and construction for colorimetric and ratiometric fluorescence dual-model aptasensor combined with g-C3N4 and G-quadruplex

Aeromonas salmonicida (A. salmonicida) is an important opportunistic pathogen to aquatic animals that causes severe economic losses to aquaculture, which makes its rapid detection and prevention are critical. In this work, a single-stranded DNA (ssDNA) aptamer (A.s-2) with high specificity to the bacteria was selected by Systematic Evolution of Ligands by Exponential Enrichment (SELEX). The selected aptamer was confirmed with high binding ability and specificity (Kd = 32 ± 8 nM). Furthermore, a novel dual-model colorimetric and ratiometric fluorescent aptasensor was constructed based on the G-quadruplex-modified aptamer and g-C3N4 for sensitive, reliable, and visual detection of the diseased bacteria in fishes. The quantitative detection was achieved in the linear range of 103–107 CFU mL-1 with a detection limit of 1.9 × 102 CFU mL-1. Meanwhile, the semi-quantitative detection can also be performed visually through fluorescence or color changes of the solution, which is suitable for the early diagnosis of pathogen infection in grassroots farms. Moreover, the developed aptasensor was successfully applied to detect A. salmonicida infection in zebrafish samples with satisfactory results. This work provides a framework for the rapid detection of pathogens in aquaculture, indicating its great prospects in food safety.

Phage therapy: An alternate biomedicine against bacterial pathogens in aquaculture

Phage therapy: An alternate biomedicine against bacterial pathogens in aquaculture

Isolation, identification, and characterization of pathogenic Aeromonas hydrophila from critically endangered Acipenser baerii

The Siberian sturgeon (Acipenser baerii) is a long-lived and late-maturing fish; its natural populations are considered endangered and listed in the first class of protected animals in Kazakhstan. To date, attempts have been made to increase the number of sturgeons through artificial reproduction in aquaculture and a release of the resultant juveniles into the Ural River, which flows into the Caspian Sea. This species faces an increased risk of Aeromonas infections. Aeromonas hydrophila is the most important sturgeon pathogen in Kazakhstan, but studies on A. hydrophila infection in Kazakhstan are still inconclusive. In the present work, our purpose was to isolate and characterize dominant bacteria in diseased A. baerii. This isolate, tentatively named AB005, was identified as A. hydrophila in an analysis of its morphological, physiological, and biochemical features and 16S ribosomal-RNA and gyrB gene sequences. A pathogenicity test was carried out for the isolate on healthy Oreochromis niloticus and Acipenser ruthenus via intraperitoneal injection along the caudal peduncle of the fish. Half-lethal doses (LD50) of isolate AB005 for O. niloticus and A. ruthenus were determined: 8.37 × 105 and 2.89 × 106 colony-forming units per milliliter, respectively. Virulence gene profiling revealed the presence of seven virulence genes related to pathogenicity (acyltransferase, phospholipase A, serine protease, heat-stable cytotonic enterotoxin, nuclease, and aerolysins A and B) in this A. hydrophila isolate. Drug sensitivity testing showed that the isolate is sensitive to quinolones, aminoglycosides, nitrofurans, amphenicols, and tetracyclines. The present findings will lay the foundation for future research on this pathogen in Siberian-sturgeon aquaculture.

Study on aptamer based high throughput approach identifies natural ingredients against RGNNV.

Nervous necrosis virus (NNV) is one of the most destructive pathogens in marine fish aquaculture and is capable of infecting more than 50 fish species worldwide, which resulted in great economic losses. Effective drugs for managing NNV infection are urgently required. Medicinal plants have been known for thousands of years and benefit of medicinal plants against pathogens in aquaculture have emerged. Nowadays, the most commonly used method for detecting virus infection and assessing antiviral drugs efficacy is reverse transcription-quantitative real-time PCR. However, the application is limited on account of high reagent costs, complex time-consuming operations and long detection time. Aptamers have been widely applied in application of pathogens or diseases diagnosis and treatments because of high specificity, strong affinity, good stability, easy synthesized and low costs. This study aimed to establish an aptamer (GBN34)-based high-throughput screening (GBN34-AHTS) model for efficient selection and evaluation of natural ingredients against NNV infection. GBN34-AHTS is an expeditious rapid method for selecting natural ingredients against NNV, which is characterized with high-speed, dram, sensitive and accurate. AHTS strategy could reduce work intensity and experimental costs and shorten the whole screening cycle of effective ingredients. AHTS should be suitable for rapid selection of effective ingredients against other viruses, which is important for improving the prevention and controlling of aquatic diseases.