Detection and characterization of Aeromonas salmonicida subsp. salmonicida infection in crucian carp Carassius auratus

Veterinary Research Communications - Aeromonas salmonicida is one of the most important pathogens in salmonids and non-salmonids species. Nevertheless, very little was reported in cyprinids about...     

Pathogenicity of Mycobacterium fortuitum and Mycobacterium smegmatis to goldfish, Carassius auratus

Despite the ubiquitous presence of atypical mycobacteria in the environment and the potential risk of infection in humans and animals, the pathogenesis of diseases caused by infection with atypical mycobacteria has been poorly characterized. In this study, goldfish, Carassius auratus were infected either with the rapidly growing fish pathogen, Mycobacterium fortuitum or with another rapidly growing mycobacteria, Mycobacterium smegmatis. Bacterial persistence and pathological host response to mycobacterial infection in the goldfish are described. Mycobacteria were recovered from a high percentage of inoculated fish that developed a characteristic chronic granulomatous response similar to that associated with natural mycobacterial infection. Both M. fortuitum and M. smegmatis were pathogenic to fish. Fish infected with M. smegmatis ATCC 19420 showed the highest level of giant cell recruitment compared to fish inoculated with M. smegmatis mc(2)155 and M. fortuitum. Of the three strains of mycobacteria examined, M. smegmatis ATCC 19420 was the most virulent strain to goldfish followed by M. fortuitum and M. smegmatis mc(2)155, respectively.     

Structure and function of the melano-macrophage centres of the goldfish Carassius auratus

We have studied the melano-macrophage centres (MMCs) of Carassius auratus as well as the changes they undergo in different experimental conditions. The MMCs of the spleen contain brown and yellow pigments, but dark pigments abound in those of the kidney. They occur in association with splenic ellipsoids and scattered throughout the renal parenchyma. Histochemical tests demonstrated small amounts of lipids and basic proteins, abundant neutral mucopolisaccharides and differences between the spleen MMCs and those of the kidney, specially regarding the presence or absence of haemosiderins and melanins. Electron microscopy shows them to consist of phagocytes containing degenerated debris, sometimes surrounded by a discontinuous layer of flat reticular cells. The MMCs of the spleen and kidney trap carbon material 48 hours after injection of Indian ink. Primary and -more so- secondary immunization with SRBC produces a net increase in number and size of MMCs. Treatment with phenylhydrazine produces a notable and fast increase of the MMCs in both organs. After 5 days the MMCs degenerated and erythroblasts restored the erythroid population. Apparently, MMCs represent non-specific means of phagocytosis related to different physiological processes.     

Thiaminase Activity of Crucian Carp Carassius carassius Injected with a Bacterial Fish Pathogen,Aeromonas salmonicida subsp. salmonicida

Abstract

Dietary thiaminase I is a cause of thiamine deficiency in animals. The physiological significance of thiaminase in the organisms containing this enzyme is not known, nor are the factors causing variation in their thiaminase activity. Tests were performed to evaluate the effect a pathogen might have on thiaminase activity in fish, when analyzed both with a cosubstrate added (CATA tests) and no cosubstrate added (NCATA tests). Pyridine is known as a cosubstrate specific for thiaminase I activity that does not accelerate thiaminase II activity. Crucian carp Carassius carassius known to harbor thiaminase I activity were injected intramuscularly with live Aeromonas salmonicida, a pathogenic bacterium of fish. For comparison, other groups were injected with formalin-killed bacteria and phosphate-buffered saline, respectively; an untreated group of fish was kept as a control. The bacteria did not contain any thiaminase activity. Significantly higher thiaminase activities (CATA and NCATA) were measured in all tissues (whole blood, injected muscle, uninjected muscle, and whole fish homogenates) of fish injected with live bacteria than in the saline-injected and the uninjected groups. The thiaminase activity of blood and that in the injected, inflamed muscle tissue followed different allocation patterns in fish injected with live A. salmonicida. The amount of thiaminase I enzyme appeared to be elevated in the whole blood of injected fish in the absence of natural cosubstrate(s). The thiaminase activity of the injected, inflamed muscle suggested that both the amount of thiaminase enzyme and some yet-unidentified natural cosubstrate(s) were elevated. This suggests that in addition to the enzyme, some cosubstrate(s) of fish or pathogen origin play a regulatory role in the so-far-unknown physiological significance of thiaminase I activity in vivo. It is suggested that the health of fish should be considered when searching for factor(s) affecting its thiaminase activity.     

Comparative pharmacokinetics of enrofloxacin in healthy and Aeromonas hydrophila‐infected crucian carp (Carassius auratus gibelio)

The comparative pharmacokinetics of enrofloxacin (ENR) and its metabolite ciprofloxacin (CIP) were investigated in healthy and Aeromonas hydrophila‐infected crucian carp after a single oral (p.o.) administration at a dose of 10 mg/kg at 25 °C. The plasma concentrations of ENR and of CIP were determined by HPLC. Pharmacokinetic parameters were calculated based on mean ENR concentrations by noncompartmental modeling. In healthy fish, the elimination half‐life (T_1/2λz), maximum plasma concentration (C_max), time to peak (T_max), and area under the concentration–time curve (AUC) values were 64.66 h, 3.55 μg/mL, 0.5 h, and 163.04 μg·h/mL, respectively. In infected carp, by contrast, the corresponding values were 73.70 h, 2.66 μg/mL, 0.75 h, and 137.43 μg·h/mL, and the absorption and elimination of ENR were slower following oral administration. Very low levels of CIP were detected, which indicates a low extent of deethylation of ENR in crucian carp.     

The application of hCG, CPH and Ovopel in successful artificial reproduction of goldfish (Carassius auratus auratus) under controlled conditions

Artificial reproduction of fish is one of the main goals of aquaculture production. The aim of this study is to optimize the method of goldfish reproduction under controlled conditions by comparing the effectiveness of carp pituitary homogenate (CPH), Ovopel and human chorionic gonadotropin (hCG), administered as a one-off dose and inducing two spawns in the same fish within a short time period. Goldfish spawners were stimulated with hCG, CPH and Ovopel, and the results were compared to the fish from the control group, comprised of unstimulated fish. In another experiment, spawn were induced twice within an interval of 21 days with the same group of fish. The best results in the first experiment in terms of the percentage of ovulating females and survival to the eyed-egg stage were achieved after administering hCG (100% and 88.7%, respectively). However, the highest fecundity was observed in fish stimulated with Ovopel (89,960 eggs/kg). It was shown in the second experiment that female goldfish produce higher weight of eggs during the first spawning, but the number of eggs/BW ratio was higher during the next reproduction process. Survival, both that of embryos to the eyed-egg stage and that of spawners, is higher during the first reproduction act.     

Induction of inflammatory cytokines by extracellular products and LPS of the fish pathogen Aeromonas salmonicida ssp. achromogenes in mice and mouse cell cultures

Balb/c mice, injected i.p. with extracellular products (ECP) of Aeromonas salmonicida subsp. achromogenes (Asa), displayed symptoms similar to toxic shock syndrome. The LD(50) observed was between 1.5 and 2.0 microg g(-1) and the mice died within 19 h. Four inflammatory cytokines were measured in mice receiving sublethal ECP doses. TNF-alpha and IL-6 showed a sharp peak in the serum while IL-1 beta and IL-2 were not detected. When peritoneal macrophages were cultivated in the presence of ECP, AsaP1 (a toxic caseinolytic metallo-protease purified from ECP) or LPS, all cultures produced TNF-alpha, IL-6 and IL-1 beta. The same antigens were mitogenic in spleen cell cultures. Furthermore, IL-2 production, which is a normal T-cell response to ConA stimulation, was downregulated in spleen cell cultures from mice injected with ECP.     

Putative immunoglobulin VH genes of the goldfish, Carassius auratus, detected by heterologous cross-hybridization with a murine VH probe

By using a defined cDNA probe for the VH region of a murine phosphocholine-binding myeloma protein (S107) we have defined a family of distinct cross-hybridizing DNA sequences in genomic DNA of the goldfish. The estimated number of the goldfish putative VH family detectable by the S107 probe is about 36. By using two putative goldfish VH probes to analyze, by hybridization, the relationships among seven of the goldfish genomic clones, we have determined that the putative goldfish VH genes recognized by the S107 probe comprise at least several distinct families that are not closely related.     

Quarantine requirements for the importation of black rhinoceros from Zimbabwe into Australia

SUMMARY The proposal by the Zoological Parks Board of New South Wales to import 10 southern black rhinoceros (Diceros bicornis) from Zimbabwe as part of an international project for conservation of the species presented the Australian Quarantine and Inspection Service (AQIS) with a unique challenge. This importation is, at least in the modern era, the first importation of live herbivores from the African continent. Many of the serious animal diseases in the world are endemic in parts of Africa. Knowledge of which of these diseases infect wild species and may be transmitted from the wild species to domesticated species, is limited. This paper describes the strategies adopted by AQIS to facilitate the importation of rhinoceros while maintaining protection of Australian consumers, rural industries, domestic livestock and fauna against the entry and spread of unwanted pests and diseases.     

Surveillance of herpesviruses in koi carp Cyprinus carpio koi and goldfish Carassius auratus cultured in West Bengal,India

Background: Viral diseases create one of the greatest challenges for the rapidly expanding ornamental fish culture sector. In recent years, the host-specific herpesviruses are receiving much attention due to the intensification in aquaculture globally. Cyprinid herpesvirus-2 (CyHV-2, goldfish hematopoietic necrosis virus), and Cyprinid herpesvirus-3 (CyHV-3, koi herpesvirus) are the lethal pathogens of koi carp Cyprinus carpio koi and goldfish Carassius auratus, respectively. In India, West Bengal is the leading state in ornamental fish production and export. Methods: The surveillance of CyHV-2 and CyHV-3 in koi carp and goldfish cultured in West Bengal was carried out between November 2014 and January 2017 as per Office International des Epizooties guidelines. Results: Both fish species were negative for CyHV-3. The CyHV-2 infection was detected in both gill and kidney tissues of apparently healthy and diseased C. auratus from December 2014 to March 2015. Severe necrosis was observed in the gills of infected C. auratus. Coinfection with members of the bacterial genera Aeromonas and Flavobacterium was also observed in the kidney of infected goldfish. The histopathological observations in the kidney of CyHV-2 infected C. auratus demonstrated the pathogenic potential of CyHV-2 and tissue tropism. Conclusions and Clinical relevance: Environmental stressors like low water temperature and the use of wastewater for culture played a vital role in the onset of CyHV-2 infection in the stressed goldfish. The spread of CyHV-2 can likely pose a certain degree of threat to aquaculture especially the unrestricted movement of goldfish. The results of the present study emphasize the necessity of an organized risk assessment to plan for the management strategies on the control and spread of CyHV-2 in Indian aquaculture.     

Host factors associated with the detection of Aeromonas salmonicida and Yersinia ruckeri in Ontario, Canada government fish hatcheries

This paper presents an epidemiological investigation of Ontario Ministry of Natural Resources Fish Health Laboratory data from 1981 to 1997, to determine whether fish species and age were associated with lot-level detection of Aeromonas salmonicida and Yersinia ruckeri in hatchery fish. In stepwise logistic regression, the species brook trout and back-cross (lake trout crossed with the hybrid “splake”) were more likely to test A. salmonicida-positive compared to all other species reared in the hatcheries. Similarly, the species brook trout was significantly more likely to test Y. ruckeri-positive compared to all other species. For both pathogens, the 1–5-month age group was associated significantly with detection. These findings suggest that purposive sampling of higher-risk fish lots could increase the likelihood of detecting both study pathogens.     

Development of a minimum-anesthetic-concentration depression model to study the effects of various analgesics in goldfish (Carassius auratus)

Teleost fish demonstrate the neurophysiologic capacity to experience pain and analgesia. A common model for assessing analgesic effect is the reduction of minimum anesthetic concentration (MAC). The present study adapted the model of MAC depression to evaluate the analgesic effects of morphine, butorphanol, medetomidine, and ketoprofen in goldfish (Carassius auratus). MAC was determined by an up-down method of sequential population sampling, anesthetizing fish with tricaine methanesulfonate (MS-222) in concentration increments of 10 parts per million (ppm), and using intramuscular needle insertion as a supramaximal noxious stimulus. Baseline MAC was determined in triplicate at the beginning (MACi) and conclusion (MACe) of the experiment (approximately 60 days). For drug trials, MAC was redetermined 1 hr after administration of morphine (10, 20, 40 mg/kg i.m.), butorphanol (0.1, 0.2, 0.4 mg/kg i.m.), medetomidine (0.01, 0.015, 0.025 mg/kg i.m.), ketoprofen (0.5, 1.0, 2.0 mg/kg i.m.), or saline control. Each drug/dose was tested in random order with a > 6-day washout period. MACi and MACf were 163 and 182 ppm, respectively, and were significantly different from each other (P = 0.02). All doses of morphine and ketoprofen decreased MAC below MACi. The highest dose of medetomidine decreased MAC below MACi. The lowest dose of butorphanol decreased MAC below MACi, but higher doses increased MAC above MACf. The authors conclude that MAC determination in fish using MS-222 was feasible and reproducible in the short term. The fact that MAC increased over time and/or exposure may limit the usefulness of MS-222 in MAC depression studies. Morphine and ketoprofen decrease anesthetic needs in goldfish and may provide analgesia.     

Virulence, cytotoxic and inflammatory activities of Vibrio anguillarum and Aeromonas salmonicida isolated from cultivated salmonid fish in Sweden

Extracellular products in culture filtrates of Aeromonas salmonicida subsp. achromogenes and Vibrio anguillarum isolated from infected fish have been shown to possess skin inflammatory factor. The extracellular products from Vibrio anguillarum were cytotoxic in HeLa and CHO cells. In addition to the skin lesions, the culture filtrates of V. anguillarum caused necrotic reaction on the rabbit skin. Five of 6 strains of V. anguillarum were lethal to mice after intraperitoneal administration of 3×10⁷ CFU. Only 1 strain of 5 A. salmonicida subsp. achromogenes produced extracellular products which elicited cytotoxic effects in the CHO cells. None of the A. salmonicida subsp. achromogenes strains were lethal to mice. The cytotoxins were inactivated when heated at 65°C for 30 min. The results indicate that the thermolabile exotoxins are non-enterotoxic since they failed to stimulate fluid accumulation in the rabbit ileal loop and did not cause elongation of the CHO cells. The rounding off of CHO cells, as well as of HeLa cells indicate that the exotoxins may play an important role in fish diseases.     

Molecular phylogeny of the kidney-parasitic Sphaerospora renicola from common carp (Cyprinus carpio) and Sphaerospora sp. from goldfish (Carassius auratus auratus)

The genetic relatedness of two kidney-parasitic Sphaerospora species was studied. Although S. renicola, the causative agent of swimbladder inflammation of common carp fingerlings (Cyprinus carpio), and Sphaerospora sp. originating from goldfish (Carassius auratus auratus) were indistinguishable on the basis of spore morphology, they were found to be genetically different as their 18S rDNA sequences shared only 71.9% identical nucleotides. In the phylogenetic trees, Sphaerospora sp. from goldfish grouped with Myxidium truttae (AJ582061) within the clade of the coelozoic freshwater species. Sphaerospora renicola clustered with S. molnari (AF378345) within the group of myxosporeans histozoic in gills. The topology of the six Sphaerospora species on the phylogenetic trees implied that myxospore morphology does not correlate with the genetic relationships, and the genus seems to be polyphyletic.     

The importation of the black rhinoceros (Diceros bicornis) from Zimbabwe into Australia

SUMMARY The paper describes a program to import and breed black rhinoceros ex situ at Western Plains Zoo in Australia. Nine rhinoceros (7 females and 2 males) captured in 1992 in Chete National Park, Zimbabwe, were transported to Australia via Cocos Island. The veterinary treatment of the animals before and during quarantine in Zimbabwe and on Cocos (Keeling) Islands is described. Three animals died; an adult male on Cocos Islands and a juvenile male and an adult female at Western Plains Zoo, Dubbo, New South Wales. The juvenile male died as a result of trauma sustained shortly after arrival and the two adults after developing a severe hepatopathy. The group of 6 females and an additional 4 males imported from the USA in 1994 have adapted well to captivity and to the climate and environment of central west New South Wales.     

In vitro and in vivo studies of the use of some medicinal herbals against the pathogen Aeromonas hydrophila in goldfish

Aeromonas hydrophila is a ubiquitous and opportunistic bacterial pathogen that produces ulcerative dermatitis under stress conditions and inflicts severe losses on global fisheries and fish culture. This study evaluates the antimicrobial potency of aqueous and ethanolic decoction (individual extract) and concoction (mixed extract) of three common medicinal herbs, turmeric Curcuma longa, Tulsi plant Ocimum sanctum, and neem Azadirachta indica, against the in vitro growth of A. hydrophila. Among the decoctions, A. indica exhibited the most potent antibacterial property (P < 0.05) against A. hydrophila. Among the concoctions, both the aqueous and ethanolic triherbal extracts mixed in the ratio of 1:1:1 had higher antibacterial activity (P < 0.05) than the other concoctions and decoctions. Goldfish Carassius auratus (10 +/- 2 g) were challenged with A. hydrophila intramuscularly in the caudal region with two separate doses (days 1 and 3) of 50 microL/fish (1.8 x 10(3) colony-forming units per milliliter). On days 9 (early) and 15 (late) of infection, fish were held in a net and dip treated for 5 min daily in a 1-L solution of 1% aqueous triherbal concoction. Red blood cell (RBC) count, hemoglobin, and hematocrit levels of the infected group were significantly higher (P < 0.05) than those of the control group. In the early treated group, all of the affected profile values returned to near normal, while the late-treated group registered a partial recovery, such as improved RBC count. The derived hematological values, such as mean corpuscular volume, mean corpuscular hemoglobin, and mean corpuscular hemoglobin concentration, of the early and late-treated groups also significantly declined (P < 0.05) but were restored to near normal (P > 0.05) only in the early treated group. The results suggest that dip treatment of A. hydrophila-infected goldfish in an aqueous triherbal concoction had a synergistic restorative effect on the hematological variables.