Naive Fish Research Articles

In vivoand in vitroeffects of mercuric chloride and sodium selenite on some non-specific immune responses of blue gourami, Trichogaster trichopterus(Pallus)

The in vivoand in vitroeffects of mercuric chloride and sodium selenite on the non-specific immunity of blue gourami, Trichogaster trichopteruswere studied. Kidney lysozyme activity in fish treated with 0·09 ppm Hg 2+for two weeks was significantly higher than the control, whereas the plasma lysozyme activity remained unchanged. In contrast, all SeO 3 2−-treated fish showed no statistical difference in the kidney and plasma lysozyme activities after two weeks of exposure, except 0·5 ppm SeO 3 2−-treated fish, which had a higher lysozyme activity in plasma. The zymosan-stimulated chemiluminescent (CL) response of head-kidney cells isolated from the naive fish was inhibited in the presence of Hg 2+. This was decreased to about 50% and 10% of the control at 0·045 ppm and 0·09 ppm Hg 2+respectively. On the other hand, the CL response of the head-kidney cells from the naive fish incubated with 4 ppm SeO 3 2−was enhanced significantly. However, the CL response of the head-kidney cells isolated from the fish that were exposed to either Hg 2+or SeO 3 2−for 2 weeks showed no statistical difference to control. The in vivoeffect of Hg 2+and SeO 3 2−on the natural bactericidal activity of plasma was tested by using a fish pathogen, Aeromonas hydrophila(strains L37 and PPD134/91). At 0·09 ppm Hg 2+, the colony forming unit (cfu) counts of the A. hydrophilawere significantly lower than the control. For the 0·1 ppm SeO 3 2−-treated fish, a higher cfu count was encountered for strain L37 but not for strain PPD134/91. On the contrary, 0·5 ppm SeO 3 2−caused no change in the cfu count for strain L37 but decreased significantly the count in strain PPD134/91.

The role of specific antiserum of catfish, Clarias gariepinus, as a defence against Aeromonas hydrophila

Catfish ( Clarias gariepinus) were vaccinated against a virulent (PPD 134/91) and an avirulent strain (L37) of formalin-inactivated Aeromonas hydrophila. The specific antiserum against the avirulent strain, in vitro, showed a significant inhibition in the growth of the avirulent strain as compared to that treated with control serum. In contrast, no significant difference was found in the growth rate of the virulent strain (PPD 134/91) which was treated with either the specific antiserum or control serum. The specific antibodies against PPD 134/91 showed an increased opsonizing effect. However, opsonization did not enhance the bactericidal activity of phagocytic cells against the virulent strain. The specific antiserum did not show any opsonizing effect on the serologically different virulent strain (PPD 122/91). The specific antiserum against PPD 134/91 could inhibit, in vitro, the invasive ability of the same virulent strain of A. hydrophilato epithelial cells. However, no significant protection was encountered in naive fish when they were intramuscularly injected with the virulent strain of A. hydrophilapretreated with the specific antiserum. Hence, the present in vitrofindings suggest that the specific antiserum in the mucus might play an important role in fish disease resistance by preventing serologically similar virulent strains of A. hydrophilafrom attaching and penetrating into the integument at the initial stage of infection.

Serum resistance as a good indicator for virulence in Aeromonas hydrophila strains isolated from diseased fish in South‐East Asia

Abstract. Aeromonas hydrophila strains isolated from diseased fish in South‐East Asia were studied for their virulence in naive tilapia, Oreochrommis aureus (Steindachner), and blue gourami, Trichogaster trichopterus (Pallas). Five of the most virulent strains used in this study shared a common resistance to the killing effect of naive fish serum. Other factors such as lack of autoagglutination in 0.2% acriflavine, instability after boiling, production of an S‐layer, and proteases and haemolysins did not correlate well with virulence. In addition, serotyping could not group all the virulent strains. Therefore, serum resistance is considered as a good indicator for screening virulence of A. hydrophila strains isolated from diseased fish in South‐East Asia.

Differences in relative sensitivity of naive and metals-acclimated brown and rainbow trout exposed to metals representative of the Clark Fork River, Montana

Naive and metals-acclimated juvenile brown (Salmo trutta) and rainbow trout (Oncorhynchus mykiss) were exposed to a metals mixture containing zinc, copper, lead, and cadmium, to compare relative sensitivities to metals concentrations typical of the Clark Fork River, Montana. Differences in the survival responses (tolerance versus resistance) measured for naive fish indicated that the rainbow trout are more tolerant (higher 96-h LC50) of the metals mixture, yet the naive rainbow and brown trout showed similarities in resistance (mean time to death). Differences for metals-acclimated fish indicated that brown trout are more resistant. Thus, the relative sensitivity of brown and rainbow trout varies with both the survival response measured and pre-exposure effects of metals on the physiological condition of the animal. Brown trout continued to acquire metals resistance for up to 5 weeks during acclimation to chronic concentrations of the metals mixture. Hepatic metallothionein and copper residue levels were positively correlated in both naive and metals-acclimated trout, and an energetic cost of metals acclimation was evidenced by reduced weight in brown trout acclimated to the metals. In contrast, rainbow trout demonstrated the least degree of acclimation and no significant growth inhibition was observed.

Effect of Feeding Experience on Growth, Vulnerability to Predation, and Survival of Esocids

Abstract We evaluated direct and indirect effects of feeding experience on growth, food consumption, susceptibility to predation, and survival of esocids. We conducted five experimental stockings of equal numbers and similar sizes (200 mm) of experienced (minnow-fed) and naive (pellet-fed) muskellunge Esox masquinongy (two stockings) and tiger muskellunge (muskellunge × northern pike E. lucius ; three stockings) in reservoirs. Feeding experience had no direct influence on prey consumption and growth. Food consumption was similar in laboratory pool experiments. In contrast, experienced esocids exhibited higher fall survival than naive fish of both taxa. Feeding experience indirectly affected survival, because predation by largemouth bass Micropterus salmoides was higher on naive esocids. Examination of potential mechanisms in field and laboratory experiments suggested predation vulnerability was not affected by differences in habitat selection, foraging behavior, antipredatory behavior, or dispersal. Color...

Experimental evidence of cannibalism and prey specialization in Arctic charr, Salvelinus alpinus

Use of a radiographic technique enabled the study of prey selection and individual specialization in Arctic charr, Salvelinus alpinus, fed with small charr and dry pellets under laboratory conditions. Both naive and experienced fish (mean weight 475 g, mean length 34.9 cm), selected the smallest individuals when offered juvenile charr (6–16 cm) as prey. The selected prey were, on average, 22% of the predator length. Cannibalism appeared to involve individual specialization, since when groups of large charr were given the opportunity to feed on juveniles one day every two weeks, the same individuals were cannibalistic throughout the sampling period of two months. When large charr were presented with alternate cycles of different food types consisting of dry pellets and fish prey, the charr exhibited a strong and consistent feeding specialization with three distinct groups being recognised: cannibals, pellet eaters and non-feeding fish.

Visual and Olfactory Stimuli in Learned Release of Alarm Reactions by Zebra Danio Fish (Brachydanio rerio)

Alarm reactions, innately organized antipredator defensive behavior, are precipitated in zebra danio fish (Brachydanio rerio) by a pheromone, alarm substance, passively exuded from injured skin of conspecific fish. Control over inducement of alarm reactions from fish can be transferred to other stimuli that do not initially provoke alarm. This was first found when an olfactory stimulus (morpholine) presented to fish together with alarm substance elicited alarm behavior from the fish and transformed morpholine into a learned elicitor of alarm from the fish. We observed similar behavior here where fish learned to react to an initially neutral visual stimulus (red light). We also confirmed earlier findings of social transmission of predator recognition by zebra danios. Fish trained to alarm react to light and morpholine were mixed with completely naive fish and tested with light and morpholine alone. All fish in the combined group displayed alarm. The naive fish, separated out and retested with light and morpholine, also exhibited alarm. Thus, the naive fish, never exposed to alarm pheromone, nevertheless learned to recognize predator cues that were socially transmitted by conspecific behavior.

Do anemonefishes use molecular mimicry to avoid being stung by host anemones?

Anemonefishes are known to have a protective mucous coat that allows them to contact the tentacles of their host anemone without being stung. There are two conflicting hypotheses as to the source and biochemical properties of this mucous coating. One hypothesis proposes that anemonefishes acquire anemone substances from their hosts during the behavioral process of acclimation, that protect the fish from being stung. Anemonefishes are considered to use anemone mucus as “chemical camouflage” or “macromolecular mimicry” to avoid recognition as “not-self” by the anemone, and possible subsequent stinging. Another hypothesis is that anemonefishes produce their own protective mucus coat, which lacks substances that elicit cnida (nematocyst and spirocyst) discharge by their hosts. The present study used immunological techniques to test whether the anemonefish Amphiprion clarkii (Bennett) (which was innately protected from two of the anemones used in the present study) has a mucous coat that resembles the external mucus of anemones. Polyclonal antibodies were prepared to the mucus of four species of anemones [ Heteractis crispa, (Ehrenberg) Stichodactyla haddoni (Saville-Kent), Macrodactyla doreenensis (Quoy and Gaimard), and Condylactis gigantea (Weinland)]. Ouchterlony (double immunodiffusion) tests showed that different antigens were present in the mucus of the four anemone species. Anemone antigens were not detected in the mucous coating of either naive (fish that had never before encountered sea anemones) or associated (those living with sea anemones) anemonefishes in Ouchterlony tests. However, more sensitive ELISA (enzyme-linked immunosorbent assay) tests showed that anemone mucus antigens were present in the mucous coating of associated anemonefish, but not naive fish. This showed that an innately protected A. clarkii does not produce a mucus coat that is biochemically similar to that of anemones, but that the same fish does acquire anemone substances in its mucus coat when it associates with anemones in aquaria. It remains to be shown whether these anemone substances actually provide the initial protection for those anemonefishes that must undergo acclimation behavior in order to keep from being stung by their host sea anemones, or additional protection for innately protected fishes.

Evidence for a peripheral olfactory memory in imprinted salmon.

The remarkable homing ability of salmon relies on olfactory cues, but its cellular basis is unknown. To test the role of peripheral olfactory receptors in odorant memory retention, we imprinted coho salmon (Oncorhynchus kisutch) to micromolar concentrations of phenyl ethyl alcohol during parr-smolt transformation. The following year, we measured phenyl ethyl alcohol responses in the peripheral receptor cells using patch clamp. Cells from imprinted fish showed increased sensitivity to phenyl ethyl alcohol compared either to cells from naive fish or to sensitivity to another behaviorally important odorant (L-serine). Field experiments verified an increased behavioral preference for phenyl ethyl alcohol by imprinted salmon as adults. Thus, some component of the imprinted olfactory homestream memory appears to be retained peripherally.

Investigations on the lack of acquired immunity by the Javanese carp, Puntius gonionotus (Bleeker), against the crustacean parasite, Lernaea minuta (Kuang)

During early 1991, an outbreak of lernaeosis in Puntius gonionotus occurred in a manmade lake in the Shah Alam recreational park in Selangor, West Malaysia. The parasite involved was identified as Lernaea minuta. An in vitro investigation was carried out to investigate the possible acquired immunity by P. gonionotus against the infection of L. minuta. The results revealed that fish acquire L. minuta infection regardless of whether they have recovered from previous infection, been injected with plasma from recovered fish or injected with plasma from naive fish. There was no significant difference ( P<0·05) in the intensity of infection between the three groups. Histopathological observations confirmed that L. minuta causes mild inflammation and subsequent pathology in P. gonionotus, compared to that of L. cyprinacea, mainly due to its shallow penetration. The apparent lack of immunocompetence in P. gonionotus against L. minuta was attributed to the less potent pathology caused by the parasite on its host.

Protection against carp erythrodermatitis following bath or subcutaneous exposure to sublethal numbers of virulent Aeromonas salmonicida subsp. nova

Abstract. A bath challenge system was used to infect carp. Cyprinus carpio L., with Aeromonas salmonicida subsp. nova, the causative agent of carp cruthrodermatitis. Bath‐challenged fish became infected with the bacterium exihibitinng typical signs of the disease, Carp that were sublethally bath exposed became infected and exhibited some skin lesions, but after one week, these quickly healed and the animals fully recovered from the infection, Naive fish that had not been previously exposed to the bacterium had mortalities of 100% when infected by the subcutaneous route and 40–60% by the bath route of infection. Carp that received sublethal infections were able to withstand subsequent lethal infection and recover regardless of the route of infection. Sublethally bath‐exposed carp were protected from subsequently lethal challenges of A. salmonicida subsp. nova for at least 5 months.

Cell-mediated Immune Response of Rainbow Trout (Oncorhynchus mykiss) to Larval Pseudoterranova decipiens (Nematoda; Ascaridoidea) following Sensitization to Live Sealworm, Sealworm Extract, and Nonhomologous Extracts

The delaved-type hypersensitivity (DTH) reaction (an in vivo manifestation of cell-mediated immunity) was studied in rainbow trout (Oncorhynchus mykiss) exposed to sealworm Pseudoterranova decipiens infections. Test fish immunized with sealworm extract and live intact worms, were compared with control fish which received either phosphate-buffered saline (negative control) or nonhomologous cestode or nematode antigens (positive control). Test fish immunized with sealworm extract produced a typical DTH reaction to live sealworm, challenge. The DTH reaction was similar to that in mammals and showed lymphoid and mononuclear cell infiltration. The electron microscopical studies revealed the presence of activated macrophages and plasma cells in the reaction zone. The specific response, following immunization with homologous antigens, supports the existence of T-cell function with anamnesis in rainbow trout exposed to sealworm antigens. However, when the test fish were immunized with live sealworms and later challenged with live sealworms, the secondary response was poor and showed no difference from controls and naive fish.

Branchial Morphological and Endocrine Responses of Rainbow Trout (Oncorhynchus mykiss) to a Long-Term Sublethal Acid Exposure In Which Acclimation Did Not Occur

Changes in branchial morphology and in plasma Cortisol, adrenaline, and noradrenaline were quantified throughout an 81 -d exposure of rainbow trout (Oncorhynchus mykiss) to sublethal acidity (pH 4.8) in artificial soft water and after a 5-h acid challenge (pH 4.0) of naive fish and 81-d acid-preexposed fish. Changes in branchial morphology at pH 4.8 were generally very mild and characterized by slight increases in filamental mucous cells and decreases in lamellar mucous cells. Chloride cell numbers and branchial Na+–K+- and total ATPase activities did not change. The filamental epithelium thickened, but the water–blood diffusion distance in the lamellae decreased during chronic exposure. Cortisol was significantly elevated throughout whereas catecholamines exhibited relatively little response. Response to acute pH 4.0 challenge was similar in naive and 81-d acid-exposed fish: epithelial damage, increase in visible mucous cells, loss of chloride cells by necrosis, and high cortisol levels but no changes in lamellar or filamental epithelial thickness, diffusion distance, ATPase activities, or catecholamine levels. Previously reported physiological data from these same trout demonstrated that sensitization rather than acclimation had occurred. Therefore, these observations support the view that acclimation does not occur in the absence of significant branchial damage and repair.

Experience of natural prey and feeding efficiency in three‐spined sticklebacks (Gasterosteus aculeatus L.)

The possibility that efficiency of feeding on different prey in three‐spined sticklebacks (Gasterosteus aculeatus L.) increases by experience of such items was investigated experimentally by comparing sticklebacks reared with only non‐moving prey (frozen brine shrimps) with sticklebacks caught in the wild from localities where a wide range of prey species was available. Fish without experience of natural food scored less well than those from the wild on various components of foraging and so had reduced feeding efficiency. However, feeding efficiency in naive fish improved after just a few days of experience with natural prey. Such experience‐induced changes in foraging efficiency will alter the profitability of prey items for individual fish.

Effects of differential prey mobility on the foraging behaviour of a cyprinid fish,Vimba elongata

The effects of reduced activity in the amphipod Gammarus jossarum on the foraging behaviour of naive and experienced cyprinid fish, Vimba elongata, were tested experimentally. Prey mobility reduced latencies. Generally, latencies declined with increasing experience of the fish. Vimba snapped at prey and inconspicuous particles at a higher frequency when prey were mobile than when they where immobile. The frequency of snaps increased with experience. These results conform to predictions from the search rate hypothesis.

Nature and Time Course of Acclimation to Aluminum in Juvenile Brook Trout (Salvelinus fontinalis). I. Physiology

Chronic exposure (up to 42 d) of juvenile brook trout (Salvelinus fontinalis) to sublethal aluminum at pH 5.2 resulted in a damage/repair acclimation phenomenon. The damage developed rapidly (within 24 h), was centered at the gills, and was characterized by substantial accumulation of Al, a corresponding reduction of gill sialic acid content (a measure of gill mucus), and inhibition of branchial Na+ transport. The corresponding internal effects of this initial damage were losses of electrolytes, hemoconcentration, and impaired tissue O2 delivery (as indicated by elevated lactate). Repair was characterized by progressive reduction of gill Al, restoration of sialic acid content, recovery of Na+ transport, and reduction in hemoconcentration and lactate levels. Accompanying the recovery was progressive development (by day 10 onwards) of increased resistance (i.e. acclimation) to acutely lethal Al. This acclimation was characterized by a reduction in both the rate of mortality and in the magnitude of physiological disturbances relative to control (i.e. Al naive) fish. The increased short-term resistance translated to greatly improved survivorship and correspondingly diminished physiological impact in the face of chronically elevated Al levels (2 wk at &lt; 300 μg Al/L). The acclimation process clearly resulted from specific changes at the gills.

The influence of hunger on feeding behaviour and on the acquisition of learned foraging skills by the fifteen-spined stickleback, Spinachia spinachia L.

Increased hunger and associated feeding motivation increased the foraging efficiency of sticklebacks already experienced with Gammarus or Artemia as prey. Behavioural components additional to the basic foraging sequence were performed less frequently and reaction distance and speed of attack increased as handling time decreased. Foraging efficiency also increased as naive fish gained experience with prey, but the rate of learning was modified by the forager's initial hunger level. Fish deprived of food for 24 h acquired attack and handling skills faster and more thoroughly than those fed to 50 and 95% satiation. Learning increased the profitability of both Gammarus and Artemia. Whether a less profitable prey type can become more profitable than another will, however, depend critically on the prey-specific intrinsic rates of increase and decrease of handling time associated with the independent effects of learning and satiation, and on the forager's initial level of hunger.

The role of learning and memory in the feeding behaviour of the fifteen-spined stickleback, Spinachia spinachia L.

As a result of experience, fifteen-spined sticklebacks began to attack Gammarus and Artemia more efficiently. This was caused by changes in reaction distance, speed of attack, handling time and success in apprehending and handling prey. How quickly foraging efficiency improved depended on the type of prey. Handling times for Artemia decreased more than those for Gammarus; attack and handling efficiency increased correspondingly. Sticklebacks feeding on Artemia also had shorter and less complex foraging sequences. The fish learned about the characteristics of prey items. Gammarus, whose legs lay head-to-tail, were most easily swallowed head-on, and foragers modified their behaviour accordingly. When two types of prey were presented alternately, individuals improved their foraging skills less effectively and handling times decreased less than when on pure diets. Foragers did not retain learned foraging skills indefinitely and became less efficient at dealing with specific prey when these were encountered less frequently. Handling times for prey offered in trials separated by 8 days were similar to those of naive fish. By decreasing handling time, learning increased the profitability (yield of energy/time) of specific prey. Gammarus could thus become more profitable than Artemia, if it predominated in the diet for several feeding bouts, and vice versa.

Lernea cyprinacea L. (Copepoda: Caligidea) in Helostoma temmincki Cuvier &amp; Valenciennes: the dynamics of resistance in recovered and naive fish

Abstract. All recovered and naive Helostoma temmincki lost their infections 30 days after homologous challenge with Lernaea cyprinacea. It is suggested that there is acquired protective immunity in recovered fish. No parasites were detected in five out of 20 recovered fish when the infective challenge dose was low; all naive fish (20) in the same tank were infected. In another experiment, all nine recovered fish were infected when the challenge infective dose was high. Recovered fish lost their infections (i.e. number of fish with no parasites and the number of parasites rejected) much faster than naive fish and this was very evident in the first 2 weeks after challenge. Parasites on recovered fish lost more egg sacs than naive fish during the same period. Eggs from parasites on recovered fish either did not hatch or were not infective or only caused a very low transitory infection in naive fish. It is suggested that, if no naive fish are introduced into a closed system for a period (and this has to be determined under field conditions) after an outbreak, then there will be no infective larvae in the water and the system will now be safe for restocking. This novel approach to control of the parasite L. cyprinacea and the spread of this disease will have to be carefully tested under field conditions.

In vitro Attenuation of Cryptobia salmositica and Its Use as a Live Vaccine against Cryptobiosis in Oncorhynchus mykiss

The present communication reports on the attenuation of a pathogenic hemoflagellate, Cryptobia salmositica Katz (Sarcomastigophora: Kinetoplastida) and its use as a live vaccine against cryptobiosis. The parasite was attenuated by continuous in vitro culture (at 10 C for 55 wk) in minimum essential medium. Attenuated (culture) forms are morphologically similar to virulent (blood) forms. They are however more slender and have a shorter anterior flagellum and a smaller nucleus and kinetoplast. The attenuated form returned to its normal size and multiplied when inoculated into naive Oncorhynchus mykiss. It produced a low parasitemia but did not cause disease (e.g., no exophthalmia or anemia) in fish. At four wk after infection, the vaccinated fish were challenged with the virulent parasite. They were protected from the disease, whereas the control (naive) fish, infected with only the virulent parasite, had the usual clinical signs (e.g., anemia, exophthalmia). No parasite was detected in any of 10 vaccinated fish at 22 wk after challenge with the virulent parasite. However, 5 of 9 fish infected with culture forms and 6 of 9 fish infected with blood forms still had detectable parasites at 26 and 22 wk after infection, respectively.