Cell-mediated protection in carp, Cyprinus carpio L., against Aeromonas hydrophila

Abstract. The mechanism of protection against Aeromonas hydrophila infection in carp was studied. Recipient fish, into which pronephric cells from carp previously immunized by dipping in bacterial crude lipopolysaccharide (LPS) at 25°C for 2h were transferred, demonstrated almost the same level of protective ability as an immunized control group. The protective ability was transmitted by non-adherent (to nylon fibre) immune pronephric cells. These non-adherent cells were damaged by anti-carp thymocyte serum and were, thus, considered to be T-like cells. The protective ability was depressed in immunized carp treated with anti-carp thymocyte serum in vivo and was also remarkably reduced in immunized carp whose macrophage function was impaired by Dextran sulphate-500 treatment. These results indicate that the protection shown by carp immunized by dipping in crude LPS is dependent on cellular immunity regulated by a T-like cell-macrophage system.     

Immune protection in carp, Cyprinus carpio L., after immunization with Aeromonas hydrophila crude lipopolysaccharide

Abstract. Vaccination with crude lipopolysaccharide (LPS) induced better protection against infection with Aeromonas hydrophila in carp than vaccination with formalin killed vaccine. Dipping fish in vaccine for 2 h at 25°C was more effective than intraperitoneal injection of the vaccine in procedural simplicity, lower stress loading and the degree of protection acquired. In carp immunized with crude LPS by the dip method, antibodies were not detected by bacteriai agglutination, passive haemagglutination and the agar diffusion tests. The results indicate that the protection against A. hydrophila infection in carp is not dependent on humoral immunity.     

Differences in resistance of carp, Cyprinus carpio L., to atypical Aeromonas salmonicida

Abstract. The degree of resistance to an atypical strain of Aeromonas salmonicida , the causative bacterium of carp erythrodermatitis, was examined in two strains of carp, Cyprinus carpio L.: a Polish line. R3, sixth generation of conventional inbreeding (full-sib matings); and a Hungarian line. R8, fifth generation of conventional inbreeding. Comparisons were made between and within the two strains. Results showed a significant difference ( P < 0·001) in the degree of resistance, with the Hungarian carp showing greater resistance than the Polish carp. Differences within each strain were also observed indicating a maternal influence on resistance. Two transferrin genotypes in three genetic combinations were identified (DD, DG, GG) but were not found to influence resistance.     

Resistance of genetically different common carp, Cyprinus carpio L., families against experimental bacterial challenge with Aeromonas hydrophila

The objective of this study was to determine the differences in disease resistance against artificial infection with Aeromonas hydrophila between genetically different common carp families. Four strains differing in their origin and breeding history were selected from the live gene bank of common carp maintained at the Research Institute for Fisheries, Aquaculture and Irrigation (HAKI, Szarvas, Hungary) to establish families with wide genetic background: Szarvas 15 (15), an inbred mirror line; Tata (T) scaly noble carp; Duna (D), a Hungarian wild carp and Amur (A), an East Asian wild carp. A diallele mating structure was used to allow the assessment of genetic variation within and between the tested 96 families for a variety of traits. The existing technologies of fertilization and incubation of carp eggs, as well as larval and fingerling rearing had been modified because of the large number of baseline populations. Two challenge trials of the 96 families of carp with Aeromonas hydrophila were done. The 10 most resistant and 10 most susceptible families to A. hydrophila were identified from these two challenges. The crosses that produced the most resistant families were mainly those having parents from Tata and Szarvas 15 domesticated strains, while the most susceptible families were from the wild strains Duna and Amur.     

鲤嗜水气单胞菌感染症及其病原生物学特性

对一起养殖鲤(Cyprinus carpio)发生的病害进行了发病情况、临床特征、病理变化等方面的检验.以3尾病(死)鲤进行病变组织中的细菌检查及细菌分离,对分离菌进行了形态特征、理化特性等较系统的表观分类学指征鉴定;同时选择代表菌株进行了16S rRNA基因的分子鉴定,测定了16S rRNA基因序列、分析了相关细菌相应序列的同源性、构建了系统发生树.结果表明,分离菌为气单胞菌属(Aeromonas Kluyver and Van Niel 1936)的嗜水气单胞菌[A.hydrophila(Chester 1901)stanier 1943],代表菌株(HC060718-1株)16S rRNA基因序列长度(不包括引物结合区)为1 454 bp(GenBank登录号:EF669478).择代表菌株做对健康鲤的人工感染试验,表明了分离鉴定的嗜水气单胞菌在被检鲤病例的相应原发病原学意义及较强的致病作用.药敏试验结果显示,对供试37种抗菌药物中的头孢噻肟等23种药物高度敏感,对氨苄青霉素等5种药物敏感,对青霉素G等9种药物耐药.     

鲤嗜水气单胞菌感染症及其病原生物学特性

对一起养殖鲤(Cypr inus carp io )发生的病害进行了发病情况、临床特征、病理变化等方面的检验。以3尾病(死)鲤进行病变组织中的细菌检查及细菌分离, 对分离菌进行了形态特征、理化特性等较系统的表观分类学指征鉴定; 同时选择代表菌株进行了16S rRNA 基因的分子鉴定, 测定了16S rRNA 基因序列、分析了相关细菌相应序列的同源性、构建了系统发生树。结果表明, 分离菌为气单胞菌属(Aeromonas K luyv er and Van N iel 1936) 的嗜水气单[A.hydrophila (Chester 1901) Stan ier 1943] , 代表菌株( HC060718- 1株) 16S rRNA基因序列长度( 不包括引物结合区)为1 454 bp( GenBank登录号: EF669478)。择代表菌株做对健康鲤的人工感染试验, 表明了分离鉴定的嗜水气单胞菌在被检鲤病例的相应原发病原学意义及较强的致病作用。药敏试验结果显示, 对供试37种抗菌药物中的头孢噻肟等23种药物高度敏感, 对氨苄青霉素等5种药物敏感, 对青霉素G等9种药物耐药。     

The histopathology of carp, Cyprinus carpio L., exposed to microcystins by gavage, immersion and intraperitoneal administration

Carp liver, gills, intestine, kidneys, heart and spleen were studied by histology after the fish were exposed to microcystins by gavage, immersion and intraperitoneal administration. Intraperitoneal inoculation with microcystins caused necrosis or dose- dependent degeneration in the liver, gills and kidneys. Gavaging with microcystins caused changes in the histopathology of the liver and gills. Cellular degeneration and necrosis occurred in the liver, gills and kidneys when carp were introduced to a tank containing 1.7 μg ml^?1 of microcystins. Lesions were not observed in the heart, spleen or intestines from any of the treated carp. Microcystins administered by intraperitoneal injection at a concentration of 50 μg kg^?1 were lethal to all fish within 8 h, while gavaging with 250 μg kg^?1 of microcystins caused minimal damage in the tissues studied.     

Responses of fingerling carp, Cyprinus carpio L., to dietary lysine

Abstract. The lysine requirements of fingerling carp, Cyprinus carpio L., for optimum growth and efficiency of food utilization were determined in two experiments by the addition of graded supplements of lysine to a basal diet deficient in this amino acid.
The lysine requirement was estimated to be around 14.5 g/kg dry matter at an ambient temperature of 20°C. At 25°C continuing growth responses occurred up to a dietary lysine concentration of 22.4 g/kg DM. However, when these responses are considered in relation to daily lysine intake, the gross efficiency of utilization of dietary lysine for growth appeared to be unaffected by environmental temperature.
Dietary lysine concentrations exerted no consistent effect upon carcass composition of carp. However, fat deposition tended to increase as food intakes increased in response to lysine additions or to elevated ambient temperature.     

Chemotaxis towards, adhesion to, and growth in carp gut mucus of two Aeromonas hydrophila strains with different pathogenicity for common carp, Cyprinus carpio L

Characteristics that promote bacterial colonization of the intestinal mucosal surface were examined in two strains of the common fish pathogen Aeromonas hydrophila, with different pathogenicity. The characteristics examined were chemotactic activity towards mucus, bacterial adherence to mucus and growth in mucus. Intestinal gut mucus of healthy common carp was used. The results indicate that chemotaxis is not necessary for a bacterium to become pathogenic, but it may be a necessary parameter for a bacterium to be an obligate pathogen. Adhesion also seems to be a factor influencing pathogenicity. The results suggest that higher adhesion to mucus and subsequent growth is associated with differences in pathogenicity.     

Urine properties in carp, Cyprinus carpio L., with sekoke disease

Abstract. Changes in the properties of urine and blood of carp with sekoke disease were studied. With the progress of this disease, the parameters of blood and particularly urine changed significantly. After 120 days, urine flow, osmolarity and concentrations of inorganic ions (K⁺, Ca²⁺, Mg²⁺, Cl⁻) and organic compounds (ammonium, creatine and creatinine, protein, glucose) in the experimental group showed higher values than those of the controls. Even after 60 days, when clinical signs of sekoke disease were not evident, abnormally high concentrations of ammonium, protein and glucose in the urine were found in the experimental group.     

Polysaccharide-induced protection of carp, Cyprinus carpio L., against bacterial infection

Abstract. The efficacy of 10 polysaccharides (curdlan, inulin, krestin, laminaran, lentinan, levan, schizophylian, selerogiucan, yeast glucan and zymosan) to enhance protection of carp, Cyprinus carpio L., against bacterial infection was investigated. Carp were intraperitoneally injected with the polysaceharides (2–l0 mgkg^-1) on days 1 and 4, and challenged with Edwardsiella tarda on day 7. Among the polysaccharides tested, lentinan, schizophyllan and scleroglucan, which are l,6-branchcd-β-l,3-glucans, significantly increased the survival rate. They also induced a protective effect against Aeromonas hydrophila at a dose of 5 mg kg^-1. The ability of the polysaccharides to activate the alternative complement pathway (ACP) was examined by incubating the polysaccharides with carp serum and measuring the residual ACP activity. At a final concentration of 0.l mgml^-1, l,6-branched-β-1,3-glucans greatly reduced (76–77%) the ACP activity. Therefore, it is suggested that the protective effect of the l,6-branched-β-1,3-glucans may be associated with the activation of ACP.     

Ichthyophthiriasis in carp, Cyprinus carpio L.: fate of parasites in immunized fish

Abstract. The host-parasite relationship between O-group carp and the ciliatc Ichthyophthirius multifiliis Fouquet was investigated, with the specific aim of characterizing the fate of parasites encountering immunized fish. Carp were immunized by repeated controlled infections; immunized fish and control fish naive to I. multifiliis were then infected in the caudal fin epidermis by a single controlled exposure to theronts, which were applied in a droplet suspension to the tail surface. The number of parasites present within the caudal fin was monitored over a subsequent 5-day period by means of in situ parasite mapping. Results indicated that, contrary to previous reports, theronts penetrated the skin of immunized fish in numbers comparable to those of fish receiving a primary infection. However, the majority of parasites which penetrated immune skin did not complete normal development; 79% of the parasites which had initially penetrated the immune skin were not relocated within 2h of exposure, and since no parasite material was detected at penetration sites, it was concluded that these parasites had prematurely exited the skin rather than been killed in situ. Subsequently, these sites became populated by leucocytes, predominantly macrophages, and the infiltrations continued for up to 5 days after the initial exposure. In contrast, at sites where mature trophonts had exited the skin of fish following a primary infection, more diffuse leucocytic infiltrations were recorded, and these were predominated by neutrophils. Differences in the response to parasite exit from immunized and previously unexposed control fish skin are discussed, with particular reference to the mode of protection and the fate of parasites encountering immune fish.     

Pharmacokinetics and tissue distribution of oxytetracycline in carp, Cyprinus carpio L., following different routes of administration

Abstract. The objective of this pharmacokinetic study was to investigate absorption, distribution, elimination and bioavailability of oxytetracycline (OTC) in carp, Cyprinus carpio L ., after different routes of administration, OTC was administered intravenously (i.v.), intramuscularly (i.m.) and orally at 60 mg/kg body weight. OTC levels were determined in plasma and several tissues. Analysis of the plasma drug concentration-time curves following i.v. OTC injection revealed three distinct phases. A three-compartment open model was used to derive pharmacokinetic parameters. Compared to mammals, a very extended final elimination half-life was observed (139.8±38.1 h). Following i.m. OTC administration, C_max was 56.8±10.9μg OTC/ml at 14 h post-injection. The Vd area was 2.1 ± 0.66 1/kg. Extreme differences were observed with respect to bioavailability following i.m. and oral administration; approximately 80 and 0.6%, respectively. Following i.m. injection tissue OTC determinations revealed that the drug was accumulating in pronephros, bone tissue and scales. After 21 days the OTC concentrations were 2.9±0.8, 5.2±0.3 and 4.7±3.1 μg/ ml, respectively. In tissue samples from the dorsal region (muscle), including the injection site, OTC could not be demonstrated at that time. The pharmacokinetic data are discussed in relation to the susceptibility of the immune system of fish for modulation.     

Storage of common carp, Cyprinus carpio L., eggs for short durations

In this study, a short-term storage of pre-activated eggs of the Japanese ornamental (koi) carp, Cyprinus carpio L., at three different temperature regimes is reported. Koi eggs were stored at low temperatures (6-9^oC), at variable or high temperatures (12-31C) and at moderate-stable temperatures (20-24.5^oC). Survival of the developing embryos was examined at the first and the second day post-activation, and in hatch-out larvae. Survival was calculated for each treatment by linear regression Y = a-bX (P± 0.01), except for eggs incubated at temperature of about 20 C(P > 0.1).The specific mortality index (b/a 100), as a ratio between the mortality rate (b) and the fertilization rate (a), indicated that the highest mortality is associated with the high and unstable storage temperatures, while the lowest mortality is with the moderate and stable storage temperatures. Eggs can be stored at moderate and stable temperatures for a maximal duration of 6h, yielding survival of hatch-out larvae higher than 50%. In two trials, fertilization potency of sperm stored in a domestic refrigerator (5-9^oC) for 5h, was compared with freshly stripped sperm and no differences in fertilization rates were found between the two treatments.     

Turmeric Powder,Curcuma longa L., in Common Carp,Cyprinus carpio L., Diets: Growth Performance,Innate Immunity,and Challenge against Pathogenic Aeromonas hydrophila Infection

Microbial diseases cause economic loss in aquaculture on a global scale, and the use of commercial antibiotics for disease treatment produces undesirable side effects. As many medicinal herbs have evolved a potent defense against pathogenic bacteria, there is a growing interest in these herbs as sources for natural antibacterial agents. In this context, this study was undertaken to evaluate the effect of turmeric powder (TP), Curcuma longa, on the performance and innate immunity of common carp, Cyprinus carpio L., and its challenge against Aeromonas hydrophila infection. Fish (1.4 ± 0.06 g) were fed 0.0 (control), 1.0, 2.0, or 5.0 g TP/kg diet up to satiation twice daily for 10 wk. After the feeding trial, fish of each treatment were intraperitoneally injected by pathogenic A. hydrophila, and they were kept under observation for 10 d during which the daily mortality was recorded. The growth‐promoting effect of TP was monitored and observed where fish growth and innate immunity increased significantly as dietary TP levels increased. According to the regression curve, the most suitable TP level for optimal fish growth and health was found to be 2.0 g/kg. On the other hand, TP inclusion in the fish diet did not significantly affect fish body contents. The obtained results indicate that TP supplementation is a promising immunostimulant, which could improve fish performance and innate immunity and prevent common carp aermoniosis at a level of 2.0 g/kg diet.     

Regression effect of carp, Cyprinus carpio L., peripheral blood lymphocytes on CHV-induced carp papilloma

Abstract. The effect of peripheral blood lymphocytes (PBL) of carp, Cyprinus carpio L., was investigated on the regression of papilloma which had been induced in carp experimentally infected with Herpesvirus cyprini (CHV). The regression of the papilloma was delayed in carp injected with anti-carp PBL serum. In addition, the spontaneous cytotoxic activity of PBL derived from normal carp was suppressed by injection with anti-carp PBL serum. These results suggest that the spontaneous cytotoxic activity of carp PBL plays an important role in the regression of carp papilloma.     

The influence of antibiotics on the immune system: immuno-pharmokinetic investigations on the primary anti-SRBC response in carp, Cyprinus carpio L., after oxytetracycline injection

Abstract. The present study was performed to obtain further details on the immuno-modulating effects of oxytetracycline (OTC) in carp, Cyprinus carpio L. The immuno-logical study was extended by investigating the pharmacokinetic behaviour of OTC. Oxytetracycline, at a dosage of 60 mg/kg, was administered five times intra-peritoneally at 3-day intervals; the first injection was given 1 day before immunization. The kinetics of the primary anti-sheep red blood cell (SRBC) response, during a short-term OTC treatment revealed that the kinetics of the plaque-forming cell (PFC) response was not significantly affected by OTC nor its solvent. In contrast, the number of plaque-forming cells was significantly decreased. The anti-SRBC antibody production was delayed by 2–4 days in both experimental groups (OTC and solvent). However, within 12–14 days post-immunization, the same antibody levels were detected as in the control group. High OTC plasma levels (>50 μg/ml) were detected about 10 h after a single i. p. injection. A mean plasma elimination half-life (T¹/₂) of 34·5 h was calculated.     

The developmental cycle of epitheliocystis in carp, Cyprinus carpio L.

Abstract. Epitheliocystis infections in the gills of carp from Israel and Portugal occurred in lining epithelial cells as well as mucus and chloride cells. Ultrastructural studies of infected cells revealed several morphological forms of epitheliocystis showing the existence of a pleomorphic developmental cycle. The following stages were observed: chlamydia-like round cells (RC), non-dividing round cells (NRC) and bullet-shaped small cells (SC). All these were identical in shape to RC and SC described from epitheliocystis of other fish, The rickettsia-like cells (PLC and ILC) which occur in epitheliocystis from other fish species and produce the SC were absent in carp epitheliocystis and instead the RC were seen transforming into SC. Transformation to SC, however, occurred at any stage of the developmental process, even before the replication potential of the RC was exhausted, i.e. before reaching the NRC stage.     

Koi herpesvirus epizootic in cultured carp and koi, Cyprinus carpio L., in Taiwan

Koi herpesvirus (KHV) poses a significant threat to cultured koi and common carp, both Cyprinus carpio L. Since the first reported case in Israel in 1998, KHV has rapidly spread worldwide. This study investigates the spread of KHV to Taiwan by collecting 49 cases of suspected common carp and koi infections from 2003 to 2005 for analysis. Clinical signs included lethargy, anorexia, increased respiratory movements and uncoordinated swimming. Hyperaemia, haemorrhage on body surface and necrotic gill filaments were recorded. Gill epithelial hyperplasia, necrosis and eosinophilic intranuclear inclusion bodies were observed by histological examination, while virions were detected using transmission electron microscopy. By detecting the presence of the KHV thymidine kinase (TK) gene and the KHV 9/5 gene using polymerase chain reaction (PCR), 37 cases were identified as KHV-positive, and the cumulative mortality of infected fish was 70-100%. Positive cases showed identical sequences for the genes analysed, implying that they were of the same origin. For the KHV 9/5 gene sequence, these cases exhibited 100% identity with the Japanese strain (TUMST1, accession number AP008984) and 99% identity with the Israeli (KHV-I, DQ177346) and US (KHV-U, DQ657948) strains. Additionally, a loop-mediated isothermal amplification (LAMP) assay was performed and found to be more sensitive than PCR tests, suggesting its potential use as a rapid diagnostic method for KHV. This is the first epidemiological study of KHV infection in cultured common carp and koi in Taiwan.