Intestine Of Fish Research Articles

Prevalence and antimicrobial susceptibility characterization of Escherichia coli isolated from the intestines of freshwater fish from the Mindu dam in Morogoro, Tanzania

Background: The Mindu dam, an open system, serves as a microbial reservoir, housing uropathogenic microorganisms like Escherichia coli. Morogoro municipality relies on the dam for water and fish. This study aimed to assess the antimicrobial susceptibility of Escherichia coli isolated from fish and water in Mindu dam. Methods: In May 2019, a cross-sectional study was conducted in Morogoro municipality. Selected antimicrobials, including ampicillin, tetracycline, cefoxitin, erythromycin, trimethoprim, gentamicin, enrofloxacin, and nalidixic acid, were tested using the disc diffusion method. E. coli ATCC 25922 served as a positive control for quality assurance during bacterial isolation. The potential source of antimicrobial contamination was identified through the multiple antibiotic resistance index. Results: Out of 148 samples, E. coli was found in 24, none in water. E. coli showed high resistance (≥50.0%) to erythromycin (62.5%), nalidixic acid (79.2%), and ampicillin (75%). Additionally, 58.0% of isolates displayed multi-drug resistance across antimicrobial classes, with a multiple antibiotic resistance index ranging from 0.00 to 0.75. Conclusion: Antimicrobial-resistant E. coli in the food chain may heighten the spread of complex urinary tract infections in the municipality. It underscores the necessity for robust municipal surveillance of antimicrobial resistance and effective antimicrobial stewardship for enhanced infection prevention and control.

Nesterenkonia marinintestina sp. nov., isolated from the fish intestine.

A Gram-positive, aerobic, non-motile, irregular short rod, nonspore-forming actinobacterial strain, designated GX14115T, was isolated from fish intestine in Beihai City, Guangxi, China and subjected to a taxonomic polyphasic investigation. Colonies were yellow‒green, circular, smooth, central bulge, convex, opaque and 2.0-3.0mm in diameter after growth on 2216E medium at 30°C for 72h. Growth occurred at 4-45°C (optimum 30°C), at pH 4.5-10.0 (optimum pH 7.5) and in the presence of 0-12% NaCl (w/v) (optimum 3.5%). Chemotaxonomic analysis showed that the main menaquinone of strain GX14115T was MK-7. The major cellular fatty acids were anteiso-C15:0 (44.8%), anteiso-C17:0 (20.5%), and iso-C15:0 (16%). The whole-cell sugars were galactose and xylose. The peptidoglycan type was L-Lys-Gly-D-Asp, and the polar lipids were phosphatidylethanolamine (PE), diphosphatidylglycerol (DPG), one unknown phospholipid (UP), and one unknown glycolipid (UG). The DNA G + C content of the type of strain was 69.5mol%. The 16S rRNA gene sequence analysis revealed that strain GX14115T is affiliated with the genus Nesterenkonia and is closely related to Nesterenkonia sandarakina YIM 70009T (96.5%) and Nesterenkonia lutea YIM 70081T (96.8%). The calculated results indicated that the average nucleotide identity (ANI) values of GX14115T were 74.49-74.78%, to the two aforementioned type strains, and the digital DNA-DNA hybridization (dDDH) values were 20.1-20.7%. Strain GX14115T was proposed as a novel species of the genus Nesterenkonia by the physiological, chemotaxonomic, and phylogenetic data, for whose the name is Nesterenkonia marinintestina sp. nov. The type of strain is GX14115T (= MCCC 1K06658T = KCTC 49495T).

Occurrence of Potentially Pathogenic Bacteria in Commercially Sold Seafood from Misamis Occidental, Philippines

Seafood is a nutritious food that is an enviable component of a healthy diet. Nevertheless, there are health risks associated with the consumption of seafood.
 This study aimed to investigate the occurrence of potentially pathogenic Vibrio spp., Staphylococcus aureus, and Salmonella spp. in 200 seafood samples purchased from several retail outlets in Ozamiz City, Philippines. Different samples analyzed using standard microbiological techniques were clam, shrimp, squid, and fish (anchovy and oil sardines). The Vibrio species identified and isolated were V. parahaemolyticus (47%), V. cholerae (16%) and V. alginolyticus (9%). Thirty percent (30%) of the samples analyzed were positive for S. aureus and 9% of the samples were contaminated with Salmonella sp. The total aerobic plate count mean log10 cfu/g of clams, shrimps, head/gills, intestine of fish (oil sardine, anchovy), and squids were considered as medium to high risk of transmitting pathogenic bacteria. Moreover, the clams and head/gills of oil sardines were considered to be at medium to high risk of transmitting pathogenic V. parahaemolyticus. The findings revealed that seafood samples analyzed microbiologically were loaded with potentially pathogenic bacteria. This result is of public health concern because these organisms are documented as causes of food-borne diseases.

Sensitivity of different organs and tissues as biomarkers of oxidative stress in juvenile tambaqui (Colossoma macropomum) submitted to fasting

The present study was conducted to evaluate the effects of fasting on responses of oxidative biomarkers and antioxidant defenses using different organs and tissues of Colossoma macropomum. The fish were divided into two groups: fed (control) and fasting (7 days). After 7 days, the fish were sampled for assessment of oxidative stress biomarkers (MDA-lipid peroxidation and PCO-protein carbonyl) and antioxidant defenses (SOD-superoxide dismutase; CAT-catalase; GPX-glutathione peroxidase; and GST-glutathione-S -transferase) in the liver, intestine, gills, muscle, brain, and plasma. The results showed an increase in MDA, PCO, SOD, and GPX concentrations in the liver and intestine of fasting fish. In contrast, in the branchial tissue, there was a reduction in the activity of SOD and CAT enzymes in fasting fish. There was also a reduction in CAT activity in the muscle of fasting fish, while in the brain, there were no changes in oxidative stress biomarkers. Plasma showed a relatively low antioxidant response. In conclusion, our results confirm that a 7-day fasting period induced tissue-specific antioxidant responses, but the increase in antioxidant responses was only for the SOD and GPX enzymes of the liver and intestine. Additionally, the liver and intestine were the most responsive tissues, whereas the plasma was the least sensitive to oxidative stress.

Çeşitli Kaynaklardan ve Ticari Gıda Takviyelerinden İzole Edilen Bacillus Türlerinin Bazı Probiyotik Özelliklerinin Karşılaştırmalı Değerlendirilmesi

Bacillus species are gram-positive, aerobic, peritrically flagellated and endospore-forming bacteria. They can be found everywhere in the environment, especially in soil (its common habitat), water, dust or in the air. Probiotics, which have beneficial health effects, constitute an important group of Bacillus species. This study aimed to isolate Bacillus from various sources, identify it molecularly and determine its probiotic properties.For this purpose, eight Bacillus subtilis, Bacillus coagulans and Bacillus clausii strains among 58 isolates from fish intestine, soil, ripened cheese and commercial probiotic supplements were identified and their probiotic properties were characterized. Firstly, Bacillus strains were molecularly identified by 16S rRNA PCR analysis. The growth of Bacillus isolates at various temperatures, salt concentrations, and pH levels, as well as tests for esculin hydrolysis, starch hydrolysis, nitrate reduction, and gas generation from glucose, were all investigated to assess the isolates' physiological and biochemical characteristics. In terms of probiotic potential of Bacillus isolates; tolerance of bile salt, cell surface hydrophobicity, auto aggregation, antibiotic susceptibility tests were conducted. In all analyses, strains obtained from food supplements showed high levels of hydrophobicity and auto-aggregation properties, and the highest values following these strains were observed in Bacillus subtilis strains (F1 and S2) isolated from fish intestines and soil, respectively. All strains showed strong growth features in bile salt conditions. It has been determined that antibiotic sensitivity varies depending on the strain. Overall, high sensitivity to tetracycline has been observed. In summary, this study revealed the potential probiotic properties of Bacillus isolates obtained from different sources. The study also compared these probiotic properties with probiotic Bacillus strains isolated from food supplements.

A novel formula of herbal extracts regulates growth performance, antioxidant capacity, intestinal microbiota and resistance against Aeromonas veronii in largemouth bass (Micropterus salmoides)

In this study, a novel herbal extracts formula (HE) comprising Scutellaria baicalensis, Taraxacum mongolicum, Biancaea sappan and Pulsatilla chinensis extract with a weight ratio of 3.0: 1.8: 3.0: 1.7, was selected based on bacteriostatic test in vitro. Subsequently, a 30-day feeding trial was conducted to evaluate the effect of dietary HE (0, 0.05, 0.10, 0.50 and 1.00%, respectively) inclusion on the performance of largemouth bass (Micropterus salmoides). The results unveiled that dietary HE 0.10% and HE 0.50% inclusion notably increased the specific growth rate (SGR) and antioxidant enzyme (SOD and GPX) activities in the intestine and liver of fish (p < 0.05). Additionally, a significantly greater height of villi and width of villi were observed in the HE 0.50% and HE 0.10% groups, respectively (p < 0.05). Conversely, histological examination indicated that dietary HE1.00% inclusion led to discernible injury to the intestines, with certain villi displaying breakage and shedding. Hematological parameters including white blood cells (WBC) and red blood cells (RBC) were significantly increased, while the erythrocyte indices including average cellular hemoglobin (MCH), and average cellular hemoglobin concentration (MCHC) remarkably decreased in fish fed HE inclusion compared to the basal diet. The 16S rRNA sequence analysis demonstrated that Cetobacterium, Aeromonas and Plesiomonas were the three dominant species in the intestines regardless of diet. Additionally, the LEfSe analysis revealed that Aeromonas was the most affected taxon, which significantly decreased in the HE 0.10%, HE 0.50% and HE 1.00% groups (p < 0.05), while the Cetobacterium content remarkably increased in HE 0.10% and HE 0.50% groups (p < 0.05). Finally, the highest survival rate of fish infected with Aeromonas veronii HZ012 was observed in the group supplemented with 0.50% HE. Collectively, our study revealed that the supplementation of HE at a proportion of 0.10–0.50% could lead to a promotion of growth performance, maintenance of liver and intestinal health, as well as a protective effect against A. veronii infection in largemouth bass.

Effects of selenoprotein extracts from Cardamine hupingshanensis on growth, selenium metabolism, antioxidant capacity, immunity and intestinal health in largemouth bass Micropterus salmoides.

This study aimed to assess the impact of dietary selenoprotein extracts from Cardamine hupingshanensis (SePCH) on the growth, hematological parameters, selenium metabolism, immune responses, antioxidant capacities, inflammatory reactions and intestinal barrier functions in juvenile largemouth bass (Micropterus salmoides). The base diet was supplemented with four different concentrations of SePCH: 0.00, 0.30, 0.60 and 1.20 g/Kg (actual selenium contents: 0.37, 0.59, 0.84 and 1.30 mg/kg). These concentrations were used to formulate four isonitrogenous and isoenergetic diets for juvenile largemouth bass during a 60-day culture period. Adequate dietary SePCH (0.60 and 1.20 g/Kg) significantly increased weight gain and daily growth rate compared to the control groups (0.00 g/Kg). Furthermore, 0.60 and 1.20 g/Kg SePCH significantly enhanced amounts of white blood cells, red blood cells, platelets, lymphocytes and monocytes, and levels of hemoglobin, mean corpuscular volume and mean corpuscular hemoglobin in the hemocytes. In addition, 0.60 and 1.20 g/Kg SePCH increased the mRNA expression levels of selenocysteine lyase, selenophosphate synthase 1, 15 kDa selenoprotein, selenoprotein T2, selenoprotein H, selenoprotein P and selenoprotein K in the fish liver and intestine compared to the controls. Adequate SePCH not only significantly elevated the activities of antioxidant enzymes (Total superoxide dismutase, catalase, glutathione reductase, glutathione peroxidase), the levels of total antioxidant capacity and glutathione, while increased mRNA transcription levels of NF-E2-related factor 2, Cu/Zn-superoxide dismutase, catalase, glutathione reductase and glutathione peroxidase. However, adequate SePCH significantly decreased levels of malondialdehyde and H2O2 and the mRNA expression levels of kelch-like ECH-associated protein 1a and kelch-like ECH-associated protein 1b in the fish liver and intestine compared to the controls. Meanwhile, adequate SePCH markedly enhanced the levels of immune factors (alkaline phosphatase, acid phosphatase, lysozyme, complement component 3, complement component 4 and immunoglobulin M) and innate immune-related genes (lysozyme, hepcidin, liver-expressed antimicrobial peptide 2, complement component 3 and complement component 4) in the fish liver and intestine compared to the controls. Adequate SePCH reduced the levels of pro-inflammatory cytokines (tumour necrosis factor-α, interleukin 8, interleukin 1β and interferon γ), while increasing transforming growth factor β1 levels at both transcriptional and protein levels in the liver and intestine. The mRNA expression levels of mitogen-activated protein kinase 13 (MAPK 13), MAPK14 and nuclear factor kappa B p65 were significantly reduced in the liver and intestine of fish fed with 0.60 and 1.20 g/Kg SePCH compared to the controls. Histological sections also demonstrated that 0.60 and 1.20 g/Kg SePCH significantly increased intestinal villus height and villus width compared to the controls. Furthermore, the mRNA expression levels of tight junction proteins (zonula occludens-1, zonula occludens-3, Claudin-1, Claudin-3, Claudin-5, Claudin-11, Claudin-23 and Claudin-34) and Mucin-17 were significantly upregulated in the intestinal epithelial cells of 0.60 and 1.20 g/Kg SePCH groups compared to the controls. In conclusion, these results found that 0.60 and 1.20 g/Kg dietary SePCH can not only improve growth, hematological parameters, selenium metabolism, antioxidant capacities, enhance immune responses and intestinal functions, but also alleviate inflammatory responses. This information can serve as a useful reference for formulating feeds for largemouth bass.

In vivo and ex vivo studies support the immunostimulant and immunoprotective effect of Damiana (Turnera diffusa Willd) in Almaco Jack (Seriola rivoliana)

Damiana (Turnera diffusa Willd) was evaluated in vitro for antioxidant and antibacterial activities against Staphylococcus aureus and Streptococcus pyogenes (as a preliminary screening assessment) by high‐performance thin‐layer chromatography (HPTLC)-Direct bioautography. A study was performed in vivo to evaluate the effects of Damiana enriched diets at 0.5 % on immune parameters in mucus and serum and gene expression in Almaco Jack (Seriola rivoliana) intestine after two and four weeks; an infection with Aeromonas hydrophila at 1x107 colony forming units (CFU) followed and an ex vivo study was carried out using head-kidney leukocytes. Ferric reducing ability of plasma (FRAP) and 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) assays showed high antioxidant activities in Damiana leaves; even in the ABTS assay, Damiana at 300 μg/mL showed similar activity to ascorbic acid – the standard control. Damiana exhibited strong in vitro antimicrobial activity against S. aureus and S. pyogenes. In vivo studies showed a strong enhancement of myeloperoxidase, nitric oxide, superoxide dismutase, and catalase activities in mucus and serum of S. rivoliana supplemented with Damiana; their immunological response enhanced after infection with A. hydrophila. IL-1β, TNF-α, and IL-10 gene expressions upregulated in the fish intestine challenged with the bacterium. Piscidin and macrophage (MARCO) receptor gene expression up-regulated at week 4 and down-regulated after infection. Intestinal histology results confirm that Damiana not cause inflammation or damage. Finally, the ex vivo study confirmed the immunostimulant and protective effects of Damiana through increased phagocytic, respiratory burst, myeloperoxidase activities and nitric oxide generation before and upon the bacterial encounter. These results support the idea that Damiana has the potential as an immunostimulant additive for diets in aquaculture by enhancing immune parameters and protecting Almaco Jack against A. hydrophila infections upon four weeks of supplementation.

Isolation, identification, and monoclonal antibody development of largemouth bass virus

Largemouth bass virus (LMBV) poses a significant threat to largemouth bass farming, leading to substantial economic losses. In December 2022, massive largemouth bass juveniles died at a fish farm in the city of Xinxiang, China. Through a series of experiments, we conclusively identified LMBV as the causative pathogen. The affected fish displayed anorexia, lethargy, and hemorrhage at the pectoral and caudal fin base. No parasites or pathogenic bacteria were detected on the body surface or gills, or isolated from the diseased fish. Severe hemorrhage, lymphocyte infiltration, and extensive necrosis were observed in the liver, spleen, intestine, and stomach of the moribund fish. The tissue homogenate from the diseased fish induced epithelioma papulosum cyprini cells (EPC) cell death, while no such effects were observed in grouper spleen (GS) cells. Sequence similarity analysis of the major capsid protein (MCP) indicated the virus shared 100% similarity with the LMBV-FS2021 strain, placing it within the Ranavirus genus. Transmission electron microscopy (TEM) observations revealed plenty of hexagonal virions accumulated in the cytoplasm of infected EPC cells. Artificial infection demonstrated that LMBV-XX01 was highly fatal to Micropterus salmoides juveniles, with an LD50 of 103.081 TCID50/fish. RT-qPCR detection confirmed that LMBV appeared in all sampled tissues of the challenged largemouth bass, with significantly higher viral loads detected in the liver and heart compared to other tissues. Additionally, we successfully obtained a highly purified recombinant MCP of LMBV and developed two strains of monoclonal antibodies targeting MCP of LMBV-XX01. Overall, our findings provide valuable materials and insights for the design of prevention strategies and the development of detection methods for LMBV.

Effect of fatty acid-enriched black soldier fly larvae meal combined with chitinase on the metabolic processes of Nile tilapia.

The aim of this study is to determine to what extent the addition of chitinase to black soldier fly (BSF) larval meal enriched or not with long-chain PUFA (LC-PUFA) could improve growth, protein digestion processes and gut microbial composition in Nile tilapia. Two different types of BSF meal were produced, in which larvae were reared on substrates formulated with vegetable culture substrate (VGS) or marine fish offal substrate (FOS). The BSF raised on VGS was enriched in α-linolenic acid (ALA), while that raised on FOS was enriched in ALA + EPA + DHA. Six BSF-based diets, enriched or not with chitinase, were formulated and compared with a control diet based on fishmeal and fish oil (FMFO). Two doses (D) of chitinase from Aspergillus niger (2 g and 5 g/kg feed) were added to the BSF larval diets (VGD0 and FOD0) to obtain four additional diets: VGD2, VGD5, FOD2 and FOD5. After 53 d of feeding, results showed that the BSF/FOS-based diets induced feed utilisation, protein efficiency and digestibility, as well as growth comparable to the FMFO control diet, but better than the BSF/VGS-based diets. The supplementation of chitinase to BSF/FOS increased in fish intestine the relative abundance of beneficial microbiota such as those of the Bacillaceae family. The results showed that LC-PUFA-enriched BSF meal associated with chitinase could be used as an effective alternative to fishmeal in order to improve protein digestion processes, beneficial microbiota and ultimately fish growth rate.

Cell-based homologous expression system for in-vitro characterization of environmental effects on transmembrane peptide transport in fish

All organisms encounter environmental changes that lead to physiological adjustments that could drive evolutionary adaptations. The ability to adjust performance in order to cope with environmental changes depends on the organism's physiological plasticity. These adjustments can be reflected in behavioral, physiological, and molecular changes, which interact and affect each other. Deciphering the role of molecular adjustments in physiological changes will help to understand how multiple levels of biological organization are synchronized during adaptations. Transmembrane transporters, which facilitate a cell's interaction with its surroundings, are prime targets for molecular studies of the environmental effects on an organism's physiology. Fish are subjected to environmental fluctuations and exhibit different coping mechanisms. To study the molecular adjustments of fish transporters to their external surrounding, suitable experimental systems must be established. The Mozambique tilapia (Oreochromis mossambicus) is an excellent model for environmental stress studies, due to its extreme salinity tolerance. We established a homologous cellular-based expression system and uptake assay that allowed us to study the effects of environmental conditions on transmembrane transport. We applied our expression system to investigate the effects of environmental conditions on the activity of PepT2, a transmembrane transporter critical in the absorption of dietary peptides and drugs. We created a stable, modified fish cell-line, in which we exogenously expressed the tilapia PepT2, and tested the effects of water temperature and salinity on the uptake of a fluorescent di-peptide, β-Ala-Lys-AMCA. While temperature affected only Vmax, medium salinity had a bi-directional effect, with significantly reduced Vmax in hyposaline conditions and significantly increased Km in hypersaline conditions. These assays demonstrate the importance of suitable experimental systems for fish ecophysiology studies. Furthermore, our in-vitro results show how the effect of hypersaline conditions on the transporter activity can explain expression shifts seen in the intestine of saltwater-acclimated fish, emphasizing the importance of complimentary studies in better understanding environmental physiology. This research highlights the advantages of using homologous expression systems to study environmental effects encountered by fish, in a relevant cellular context. The presented tools and methods can be adapted to study other transporters in-vitro.

Effect of Moringa olifera leaves on growth and gut microbiota of Nile tilapia (Oreochromis niloticus).

The study was conducted to evaluate the effect of Moringa olifera on the growth and gut health of Tilapia (Oreochromis niloticus). The feed having 30% crude protein was prepared as an experimental diet with 4%, 8% and 10% M. olifera leaf supplementation, respectively. The control diet was devoid of M. olifera leaves. The 10 weeks feeding trial was carried out on 60 fish in aquaria. Fish was fed @ 3% of body weight twice a day. Diet with the high level of inclusion of M. olifera leaves significantly increased the growth rate, Survival Rate (SR), Specific Growth Rate (SGR) and Feed Conversion Efficiency (FCE) in all treatment groups compared to the control group. Similarly, Feed Conversion Ratio (FCR) gradually decreased and found highly-significant. To check the gut health of the Tilapia, random samples were selected and dissected. Nutrient agar was used as culture media to check the growth of bacteria. Pour Plate Method was used for viable colonies count by colony counter. Through staining method, the different bacteria such as Escherichia coli, Salmonella, Shigella and Pseudomonas aeruginosa were identify abundantly in the intestine of control diet fish but less number present in treatment diets groups. These results showed that M. olifera leaves up to 10% of dietary protein can be used for Nile tilapia for significant growth and healthy gut microbiota of fish.

Isolation and Identification of Pathogenic Bacteria from Commercially Marketed Fish in Wukari, North-Eastern Nigeria

Abstract Pathogenic bacteria that are part of the natural microflora can be found in aquaculture products. Microbial pathogens associated with fish can be transmitted to humans who consume or handle the fish. This study aimed to isolate human pathogenic bacteria present in the gills, intestine and skin of apparently healthy and spoil fishes. Tilapia (Oreochromisniloticus) and Cat fish (Clariaslazera) were collected from Wukari old market and Yam market of Wukari Nigeria. Bacteria were isolated from the fish gill, intestine and skin. The growth properties of distinct isolates on specific culture media were used to differentiate and characterize them. Pathogenic bacteria were recovered from various fish species that appeared with different features. The following human pathogenic bacteria were isolated; Escherichia coli and Staphylococcus aureus. The total bacteria count value ranges between 0.12 x 105 to 3.52 x 105 for the tilapia fish and 0.12 x 105 to 3.96 x 105 cfu/ml for cat fish. The identification revealed that Staphylococcus aureus and Escherichia coli was the most abundant pathogenic bacteria and also present in the initial water samples collected. Presence of Staphylococcus aureus and Eshcherichia coli is attributed to contamination of the fish samples by man through handling and processing. These enteric bacteria in fish serve as indicator organisms of faecal contamination and or water pollution. Their presence also represents a potential hazard to humans’ life. The isolates were higher from Yam market samples compared to Wukari old market.

Expansion of Phalacrocorax carbo in reservoirs of the North-Eastern Kazakhstan

Initial data regarding the helminth fauna of the great cormorant (Phalacrocorax carbo) in the Pavlodar region of Kazakhstan have been acquired. The parasite fauna of the great cormorant is influenced by its diet of fish and consists of two fully developed helminth species - the nematode Contracaecum rudolphii Hartwich, 1964 and the cestode Ligula interrupta Rudolphi, 1810. An incidental finding was made when cestodes and their broken pieces were found in the intestines of cormorants. These cestodes were recognised as belonging to the genus Cyathocephalus, which is known to solely parasitize fish in its adult stage. Metacercariae of trematodes, specifically from the family Strigeidae, were found. These parasites are very new to fish intestines and are not commonly found in birds that consume fish. The growing population of great cormorants in the Ertis basin may introduce new kinds of helminths, some of which could have significant implications for the spread of diseases among animals. The prevalence of parasites in cormorants is a significant concern and has the potential to facilitate the transmission of nematodes and cestodes among other species, including fish-eating birds, poultry, fish, and aquatic invertebrates. This creates localised areas where nematodes and ligulidoses can thrive.

Cellulolytic enzymes in Microbulbifer sp. Strain GL-2, a marine fish intestinal bacterium, with emphasis on endo-1,4-β-glucanases Cel5A and Cel8.

Cellulose is an abundant biomass on the planet. Various cellulases from environmental microbes have been explored for industrial use of cellulose. Marine fish intestine is of interest as one source of new enzymes. Here, we report the discovery of genes encoding two β-glucosidases (Bgl3A and Bgl3B) and four endo-1,4-β-glucanases (Cel5A, Cel8, Cel5B, and Cel9) as part of the genome sequence of a cellulolytic marine bacterium, Microbulbifer sp. Strain GL-2. Five of these six enzymes (excepting Cel5B) are presumed to localize to the periplasm or outer membrane. Transcriptional analysis demonstrated that all six genes were highly expressed in stationary phase. The transcription was induced by cello-oligosaccharides rather than by glucose, suggesting that the cellulases are produced primarily for nutrient acquisition following initial growth, facilitating the secondary growth phase. We cloned the genes encoding two of the endo-1,4-β-glucanases, Cel5A and Cel8, and purified the corresponding recombinant enzymes following expression in Escherichia coli. The activity of Cel5A was observed across a wide range of temperatures (10-40 ˚C) and pHs (6-8). This pattern differed from those of Cel8 and the commercial cellulase Enthiron, both of which exhibit decreased activities below 30 ˚C and at alkaline pHs. These characteristics suggest that Cel5A might find use in industrial applications. Overall, our results reinforce the hypothesis that marine bacteria remain a possible source of novel cellulolytic activities.

Intestinal endoparasitism in Corydoras multiradiatus (Siluriformes: Callichthyidae) in Iquitos, Loreto-Peru

Corydoras multiradiatus (Orcés V.1960), commonly called “hog-nosed brochis” and popularly known in Peru as “coridora jumbo” is a species of commercial importance among the Amazonian ornamental fish. It is known that the integrity of the health of aquatic organisms positively increases the production and trade of quality products, however, the opposite negatively affects financial losses and production costs. Among the possible diseases, parasitism stands out, and there is still a lack of information, mainly about metazoans, parasitic species that infect C. multiradiatus. Therefore, this study aimed to identify the species of endoparasite metazoans in C. multiradiatus and evaluate the histological lesions present to provide information to ornamental fish producers about these pathologies. Thirty specimens of C. multiradiatus were collected by fishermen in the municipality of “Belén” in Iquitos, Peru. The results revealed the presence of Tylodelphys sp. and nematode larvae in the intestinal wall and Procamallanus (Spirocamallanus) pintoi (Kohn &amp; Fernandes 1988) present in the intestine. The histopathological analysis of the intestine showed several lesions: necrosis; cell desquamation; infiltration of lymphocytes in the mucosa; transepithelial infiltration of lymphocytes; enterocyte hyperplasia; goblet cell hyperplasia and enterocyte desquamation. The study revealed the presence of endoparasites in C. multiradiatus collected in Iquitos, Loreto-Peru, and a moderate degree of damage caused by histological lesions in the intestine of infected fish. The results of this work will serve the fishermen, collectors and aquarists of the city of Iquitos, who, knowing the endoparasites present in C. multiradiatus and the damage they can cause to the fish, will be able to adopt prophylactic measures in order to guarantee the good condition of the fish for its consequent commercialization at a local, national and international level.

Vibrio cholerae O1 Inhabit Intestines and Spleens of Fish in Aquaculture Ponds.

Vibrio cholerae is the causative agent of cholera, an acute diarrheal disease that spreads locally and globally in epidemics and pandemics. Although it was discovered that fish harbor V. cholerae strains in their intestines, most investigations revealed non-toxic V. cholerae serogroups in fish. Due to the rarity of toxigenic V. cholerae serogroups, it is difficult to cultivate these strains from environmental samples. Hence, here we aimed to uncover evidence of the occurrence of toxigenic V. cholerae in the intestines and spleens of various fish species. By using molecular detection tools, we show that V. cholerae O1 and strains positive for the cholera toxin inhabit both healthy and diseased fish intestines and spleens, suggesting that fish may serve as intermediate vectors of toxigenic V. cholerae. No significant differences were found between the abundance of toxigenic V. cholerae (either O1 or cholera toxin positive strains) in the healthy and the diseased fish intestines or spleens. In conclusion, a variety of fish species may serve as potential vectors and reservoirs of toxigenic V. cholerae as they form a link between the other reservoirs of V. cholerae (chironomids, copepods, and waterbirds). Similarly, they may aid in the spread of this bacterium between water bodies.

Dietary alpha-lipoic acid boosts growth, immune-antioxidant traits, behavior, and transcriptomes of antioxidant, apoptosis, and immune-related genes to combat cold stress in Nile tilapia (Oreochromis niloticus)

Water temperature is a major environmental factor affecting fish activity, physiology, behavior, and growth. Alpha lipoic acid (ALA) has garnered much attention lately as an aqua feed supplement because of its antioxidant effects. Thus, this study aimed to assess the effect of dietary ALA on performance, hematological and immunological indices, behavior and oxidative stress, apoptosis, and genetic inflammation in Oreochromis niloticus following 60 days of cold stress. Two hundred forty fish were divided into four groups (15 fish/replicate, N = 60 fish/group). The control (CNT) and ALA groups were reared at 25 °C and fed a basal diet without any supplementation or a basal diet containing 600 mg ALA/kg diet, respectively. The cold water-stressed group (CWS) and ALA + CWS groups were reared at 18 °C and fed basal and basal diets containing ALA, respectively. The results revealed that dietary ALA supplementation significantly improved feed utilization and growth in fish. ALA also mitigated CWS-induced microcytic hypochromic anemia and hyperlipidemia. Moreover, ALA supplementation considerably enhanced the antioxidant status and boosted intestinal α amylase, lipase, protease, and serum acetylcholinesterase activity of CWS-exposed fish. Besides, ALA supplementation significantly reversed CWS-induced upregulation of proinflammatory genes, pro-apoptotic genes, heat shock protein, and proliferating cell nuclear antigen or downregulation of superoxide dismutase and catalase gene expressions in liver and spleen tissues. Furthermore, ALA supplementation reduced CWS-induced histopathological alterations in fish liver, spleen, and intestine. We can conclude that ALA (600 mg/kg diet) could be recommended as an aqua feed supplement to boost growth, antioxidant capacity, and immunity and attenuate inflammatory and apoptotic reactions of fish reared under CWS-induced.

Emerging role of vitamin D3 in alleviating intestinal structure injury caused by Aeromonas hydrophila in grass carp (Ctenopharyngodon idella)

Bacterial pathogens destroy the structural integrity of functional organs in fish, leading to severe challenges in the aquaculture industry. Vitamin D3 (VD3) prevents bacterial infections and strengthens immune system function via vitamin D receptor (VDR). However, the correlation between VD3/VDR and the structural integrity of functional organs remains unclarified. This study aimed to investigate the influence of VD3 supplementation on histological characteristics, apoptosis, and tight junction characteristics in fish intestine during pathogen infection. A total of 540 healthy grass carp (257.24 ± 0.63 g) were fed different levels of VD3 (15.2, 364.3, 782.5, 1,167.9, 1,573.8, and 1,980.1 IU/kg) for 70 d. Subsequently, fish were challenged with Aeromonas hydrophila, a pathogen that causes intestinal inflammation. Our present study demonstrated that optimal supplementation with VD3 (1) alleviated intestinal structural damage, and inhibited oxidative damage by reducing levels of oxidative stress biomarkers; (2) attenuated excessive apoptosis-related death receptor and mitochondrial pathway processes in relation to p38 mitogen-activated protein kinase signaling (P < 0.05); (3) enhanced tight junction protein expression by inhibiting myosin light chain kinase signaling (P < 0.05); and (4) elevated VDR isoform expression in fish intestine (P < 0.05). Overall, the results demonstrated that VD3 alleviates oxidative injury, apoptosis, and the destruction of tight junction protein under pathogenic infection, thereby strengthening pathogen defenses in the intestine. This finding supports the rationale for VD3 intervention as an essential practice in sustainable aquaculture.

Evaluation of Toxicological Risks and Effects of Microplastics on Nile Tilapia (Oreochromisniloticus) under in Vitro Laboratory Conditions

Microplastics have been reported by many literatures as contaminants of environmental water bodies and are ingested by aquatic organisms due to their small sizes. Knowing the effects of microplastics to fresh water fish which are kept in ponds helps in managing fish keeping practice. The objective of research was to determine the toxicity of microplastics to Nile tilapia (Oreochromis niloticus). The Experiment was done in 80 fish samples. Microplastics which were prepared for the batch experiment were introduced in the aquarium followed with observation for 21 days. The digestion of fish gills and intestines involved 10 mL of 10% (w/v) KOH solution and incubation at 65oC for 24 hours. Engulfed microplastics were determined using stereo microscope and At-IR spectrophotometer for confirmation. Engulfed microplastics were observed to be in mean range of 3.37 x 102±4.04 x 102 to 2.32 x 103 ± 3.57 x 103 particles/kg in gills and 4.68 x 104 ± 3.02 x 104 to4.40 x 104 ± 5.34 x 104 particles/kg in intestines. The observed responses were loss of equilibrium for 35% of fish, abnormal swimming for 49% of fish, abnormal ventilator behavior for 59% of fish, abnormal appearance for 39% of fish and average growth weight increase in control experiment fish was 6.10±2.62 g compared to 1.7 ± 3.62 g in test fish. There was no mortality of Nile tilapia. The responses of fish to the presence of microplastics in aquarium indicated that microplastics had adverse effects to Nile tilapia (Oreochromis niloticus). More researches have to be done on fish physiological changes caused by microplastics.