Methanol as cryoprotectant and the suitability of 1.2 ml and 5 ml straws for cryopreservation of semen from salmonid fishes

For salmonid semen, the cryoprotective action of 10% methanol was compared with a 5% dimethyl sulphoxide (DMSO), 1% glycerol mixture, until now one of the most effective cryoprotectants. In Oncorhynchus mykiss (Walbaum), Salmo trutta L. f. fario, Salmo trutta L. f. lacustris and Salvelinus alpinus (L.), semen cryopreserved with both cryoprotectants yielded post-thaw fertilization rates of 90-100% of control with untreated semen at sperm-to-egg ratios of 1.8 × 10⁶-2.4 × 10⁶ spermatozoa per egg. However, at sperm-to-egg ratios of 0.9 × 10⁶-1.2 × 10⁶ spermatozoa per egg, semen cryopreserved with methanol had significantly higher fertilization rates than semen frozen with the DMSO/glycerol mixture. In other studies we obtained similar data for Coregonus sp., Salvelinus fontinalis (Mitchill), Thymallus thymallus (L.) and Hucho hucho (L.), proving that methanol is the most effective and generally applicable cryoprotectant for semen of the studied salmonid species. To facilitate the insemination of large egg batches we investigated the suitability of 1.2 ml and 5 ml straws for deep freezing of semen of Oncorhynchus mykiss, Salmo trutta f. fario, Salmo trutta f. lacustris and Salvelinus alpinus. With 1.2 ml straws the fertilization rates were similar to 0.5 ml straws when using lower freezing and higher thawing temperatures. The 5 ml straws resulted in a fertilization success of only about 40% of fresh semen control.     

Effects of low pH on the reproductive behavior of salmonid fishes

In northern Europe and America, the acidification of lakes and rivers induced by acid precipitation has damaged fish populations. At present, rapidly expanding industrial activities in East Asia have led to continuous increases in the quantity of emission of acidic pollutants, and rain of acidic levels of pH4 has precipitated throughout Japan. In this way, the effects of acid rain on fish populations have also been manifested in Japan. In order to examine the effects of low pH on the reproductive behavior of salmonids which are known to be acid-sensitive species, changes in the frequency of upstream behavior and spawning-related female nest-digging behavior in response to changes in pH were observed in mature hime salmon (land-locked sockeye salmon) Oncorhynchus nerka, brown trout Salmo trutta and Japanese char Salvelinus leucomaenis. Digging and upstream behavior were significantly inhibited in weakly acidic water of pH 5.8–6.4 formulated using sulfuric acid. Land-locked sockeye salmon was the most sensitive to changes in pH among the three species.     

Microsporidian parasites in the gills of salmonid fishes*

Abstract. A microsporidian parasite was found in the gills of the brook trout, Salvelinus fontinalis (Mitchill), which was similar to Loma morhua Morrison and Sprague, a microsporidian parasite in the gills of cod. It was described and designated as Loma sp. Plistophora (=Pleistophora) salmonae Putz, Hoffman and Dunbar in gills of rainbow and steelhead trout, Salmo gairdneri Richardson, was also observed and was transferred to the genus Loma. All microsporidia known in gills of Gadidae and Salmonidae were judged to belong to the genus Loma but the data were not sufficient to provide a basis for a definitive separation of species. Provisionally, Loma morhua, L. branchialis, L. salmonae and Loma sp. were recognized.     

Assessing the consequences of habitat fragmentation for two migratory salmonid fishes

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Fatty acid influence on Prochilodus lineatus (Characiformes,Prochilodontidae) embryo cryopreservation parameters.

This study aimed to evaluate the vitellogenic transference and incorporation of long‐chain polyunsaturated fatty acids (LC‐PUFA) into the membranes of Prochilodus lineatus embryos, aiming to increase the permeability to cryoprotectants and resistance to electric fields. One hundred thirty broodstock of P. lineatus were fed with control (C) or fish oil‐supplemented diets (FO) for 12 months. The fatty acid (FA) profle was determined using gas chromatography. For the neutral fraction, the FO group had a decrease in monounsaturated fatty acids (MUFA) and an increase in n3PUFA and, n6PUFA. To test for cryoprotectant toxicity, embryos were exposed for 20 min to a cryoprotectant solution of 1,2‐Propanediol (Prop) at a concentration of 5 or 6 molar (M). For FO, a reduction in survival of 33.1% was observed in 5 M, and no survival was observed at 6 M. Embryo samples were exposed the six polarized electric fields (3.4–51.6 joules), and with 11.2 J of energy, the control group exhibited reduced survival in 98.3% of the fish, whereas the FO presented superior resistance, exhibiting a survival similar to that of the OJ up to 40.2 J. We conclude that FA were transferred between P. lineatus broodstock to the embryos, with an increase in LC‐PUFA resulting in lower survival rates in the cryoprotectant test in the FO group and a greater physical plasticity of FO embryos to electrical field tests.     

Biophysical properties of salmonid alphaviruses: influence of temperature and pH on virus survival

A series of laboratory studies were undertaken to investigate the survival of salmonid alphaviruses (SAV) under a range of conditions relevant to waste disposal, persistence and spread in the field, and to laboratory studies and testing. SAV was found to be rapidly inactivated in the presence of high levels of organic matter at 60 degrees C at pH 7.2 and at pH 4 and pH 12 at 4 degrees C, suggesting that composting, ensiling and alkaline hydrolysis would all be effective at inactivating virus in fish waste. Testing was conducted under sterile conditions at 4, 10, 15 and 20 degrees C in sea water, half-strength sea water and fresh (hard) water, both in the absence and the presence of added organic matter. Virus survival was shown to be inversely related to temperature, and to be reduced by the presence of organic matter. Calculated half lives (t(1/2)) under these conditions ranged from 61.0 to 1.5 days. Testing in non-sterile sea water resulted in reduced t(1/2) values. The half life of SAV in serum was also found to be inversely related to temperature, emphasizing the need for rapid shipment of samples at 4 degrees C to laboratories for virus isolation studies.     

Evaluation of the influence of different species and cultivars of lupin kernel meal on the extrusion process, pellet properties and viscosity parameters of salmonid feeds

This study examined the influence of different lupin species and cultivars and their inclusion levels on the physical properties of an extruded diet. Lupin kernel meals were included into a fish diet formulation at 0%, 10%, 20% and 30%. Soybean meal was also included in a series of diets on a similar basis as a reference. The diet mashes were extruded using a laboratory-scale APV19:45 twin-screw extruder. The pellets were examined for radial expansion, bulk density, vacuum oil uptakes, sink rate, shear and crush strengths. Inclusion of lupin kernel meal significantly increased bulk density, sink rate, shear and crush strength of the pellets. With the increase in lupin kernel meal inclusion level, a concomitant decline in vacuum oil uptake and radial expansion was also observed. An assessment of the diet mashes using a Rapid Visco Analyser showed that the inclusion of lupin kernel meals increased the rate and degree of gelatinization of the mash starch content compared with that with a similar inclusion level of soybean meal. The inclusion of lupin kernel meals in the diet mash also improved the water-holding capacity of the extrudate, which has important implications for the reduction in extruder wear.     

A uniform method for cryopreservation of semen of the salmonid fishes Oncorhynchus mykiss (Walbaum), Salmo trutta f. fario L., Salmo trutta f. lacustris L., Coregonus sp.

The present study describes a uniform method for cryopreservation of semen of Salmonidae (Oncorhynchus mykiss (Walbaum), Salmo trutta f. fario L., Salmo trutta f. lacustris L., Coregonus sp.). It presents a new type of extender and experiments demonstrating that warming of frozen/thawed semen to 20°C prior to fertilization significantly increases the fertilization rate. Freezing is performed in straws in the vapour of liquid nitrogen and for insemination a diluent technique is used. The consistency of the method was tested by repeating the experiments with different batches of semen and eggs. The following fertilization rates (% of control) were obtained: Oncorhynchus mykiss: 89.6 ± 16.0% (mean ± standard deviation, n= 25, n of control = 20, sperm/egg ratio of 1.6 ± 0.2 × 10⁶ spermatozoa/ egg). Salmo trutta f. fario: 93.8 ± 6.4% (n= 12,9.9 ± 1.2 × 10⁶spermatozoa/egg), Coregonus sp.: 92.8 ± 2.4% (n= 6, 0.5 × 10⁶ spermatozoa/egg), Salmo trutta f. lacustris: 85.0 ± 8.4% (n= 12, 4.8 ± 1.4 × 10⁶ spermatozoa/egg).     

Mercury levels of small fishes: influence of size and catch area

In the present study, we assessed small fishes as potential feed fishes with the lowest mercury levels. The mercury levels of four small pelagic fishes and a benthic fish from the Pacific Ocean and the Sea of Japan, i.e., spotted chub mackerel Scomber australasicus, chub mackerel Scomber japonicus, horse mackerel Trachurus trachurus, round scad Decapterus punctatus, and bastard halibut Paralichthys olivaceus, were determined. Total mercury levels in fish muscle were measured using a reduction vaporizing atomic absorption method. There were no significant correlations between body weight and mercury levels for these species. However, significant differences were observed in the mercury levels of these species between the two catch locations, with fish caught in the Pacific Ocean having higher mercury levels. This difference may be due to the Pacific Ring of Fire, a highly seismically and volcanically active zone, which is located in the Pacific Ocean. Preference for these fishes for use in aquaculture should refer to the area where they are caught in addition to the species.     

Characterization of rainbow trout milt collected with a catheter: semen parameters and cryopreservation success

Collection of fish milt by stripping risks the danger of milt contamination by urine. This may seriously influence milt characteristics and quality, including usefulness for cryopreservation. Urine contamination of milt may be avoided by using a catheter for sperm collection. The objectives of this study were to provide basic characteristics of milt collected with a catheter, to test the usefulness of this milt for cryopreservation, and to correlate characteristics of fresh and cryopreserved semen with sperm fertility rates. Milt from 25 rainbow trout Oncorhynchus mykiss (Walbaum) males were used. All samples were cryopreserved using the pellet method within 1 h of collection, using 0.6 m sucrose and 10% dimethyl sulphoxide (DMSO) as an extender. Catheterization resulted in semen of very good motility (> 90% motile spermatozoa) and high fertilization rates after cryopreservation (mean fertilization rate 81.8 ± 13.3% of control, at a sperm/egg ratio of 2.4 ± 0.3 × 10⁶). Osmolality of seminal plasma and concentrations of sodium, potassium and magnesium ions had low variability, which suggests that they are important for creating a stable environment for sperm storage in the sperm duct. Higher variability of certain seminal plasma characteristics, such as protein concentration and antiproteinase activity, suggests that these characteristics are related to individual semen features of particular males. A strong correlation of seminal plasma zinc concentration with protein concentration may reflect an importance of zinc in semen biology. Cryopreservation caused a significant release of protein and acid phosphatase from spermatozoa. Our results did not reveal any single characteristic of semen collected by catheter that could be used as a powerful predictor of cryopreservation success, presumably because all samples were of high quality.     

低渗溶液浓度对黄颡鱼精子活力和受精率的影响

为提高黄颡鱼(Pelteobagrus fulvidraco)人工授精的稳定性,分别用卵子和精子同步激活法以及预激活精子法,在6个梯度浓度(0%～0.5%)范围的低渗溶液中进行人工授精试验,比较精子活力、受精率和孵化率。结果显示:低渗溶液浓度与精子活力、精子激活率和受精率密切相关,在相同浓度的低渗溶液组中采用预激活精子法的受精率显著高于卵子和精子同步激活法(P<0.05),受精率相对应提高了7.7%～14.5%;低渗溶液的浓度和人工授精方式对孵化率无显著影响。结果表明在浓度0.3%的低渗溶液中采用预激活精子法的效果最好。     

Effects of handling, discrete meals and body weight on the individual variation of gastric emptying parameters

Individual feeding and gastric emptying was estimated for two salmonid species, brown trout (bt), Salmo trutta (L.), and rainbow trout (rt), Oncorhynchus mykiss (Walbaum), fed extruded feed with glass beads, and exposed to repetitive handling, anaesthetizing and X-raying. No significant effect was found to be caused by the handling and experimental procedures. Huge inter-individual (bt: CV_g= 52-104%; rt: CV_g= 19-33%) differences in discrete meal size were caused by interspecific differences in feeding behavior. An exponential model was fitted to the X-ray time series data for each individual for a computation of the initial (Ri) and fractional (R) gastric emptying rates. The individual variations in meal size had no significant effect on the rates. The meal frequency [two meals per day versus postprandial starvation (PS)] significantly affected the fractional gastric emptying rate. The rate decreased significantly for the PS groups while the inter-individual variation increased. Body weight (10-2500 g) significantly affected the fractional gastric emptying rates, which increased by up to 600 g and then decreased. The coefficient of variation for R increased with body weight. An equation for calculation of weight specific food processing rate (FP) is proposed. The FP decreased significantly with body weight. A maximum estimate of FP_i= 2.3 mg g^?1 h^?1 at 12°C for an individual was obtained with a 13.7-g trout. An increased weight-specific consumption (per cent body weight) for the smallest fish demonstrated that FP_g was 5.4 times higher than for the biggest fish, but after normalization with a body weight exponent of 0.75, an estimate of 1.7-2.3 was obtained.     

Determination of the phosphorus fertilization rate for bluegill ponds using regression analysis

Pond fertilization studies usually are restricted by availability of ponds forcing researchers to limit treatments to maintain adequate replicates for statistical analysis. Data from a wide range of fertilizer rates applied over a single season in un‐replicated ponds were treated using regression analysis to establish the optimum phosphorus application rate for bluegill (Lepomis macrochirus) ponds. The response to phosphorus fertilization conformed to a saturation model (R²=0.92). Bluegill production increased with greater fertilization rate up to 3 kg P₂O₅ ha^?1, but production was similar (501–558 kg ha^?1) at rates of 3–7 kg P₂O₅ ha^?1. The observation that 3 kg P₂O₅ ha^?1 per application was adequate phosphorus fertilization for bluegill ponds with sufficient nitrogen agrees closely with the usual recommendation of 4 kg P₂O₅ ha^?1 per application. Results of this research also revealed that nutrient assessment in bluegill ponds can be based on total nitrogen and total phosphorus analyses, and Secchi disk visibility may be used as an index of plankton abundance.     

Contrasting influence of flow regime on freshwater fishes displaying diadromous and nondiadromous life histories

Flow variability structures freshwater fish community traits and life‐history patterns such as migration patterns between fresh and saltwater (diadromy). Few studies, however, have explored relationships between diadromy and flow regime while accounting for other abiotic covariables. The present paper used canonical ordinations to remove the shared variation between groups of explanatory variables that explain variation in fish communities and examine two objectives with New Zealand fishes: (i) to compare the unique contributions of Hydrological Regime, Climate, Habitat and Spatial‐Seasonal data sets to the variation of diadromous and nondiadromous fishes and (ii) to compare the relative contributions of a Hydrological Variability and Low‐Flow data set to community structure of both life‐history patterns. All explanatory variables explained a total of 20.15% and 29.58% of the variation in diadromous and nondiadromous fishes, respectively. Objective 1 analyses showed that the largest unique component of variation was explained by Hydrological Regime for nondiadromous fishes (12.17%), while Climate uniquely explained the most variation in diadromous fishes (4.3%), followed closely by Hydrological Regime (3.08%). Objective 2 analyses showed that Hydrological Variability uniquely explained five and 11 times more variation than the Low‐Flow data set in diadromous and nondiadromous fishes, respectively. Findings illustrate the importance of hydrological regime to New Zealand freshwater fishes. Specifically, aspects of hydrological variability uniquely account for more variation than aspects of low flow. Differing relative influences of hydrology between life‐history patterns suggest that diadromy may mediate the influence of flow regime. Results outline difficulties for environmental flow settings when biota display differing life histories.     

Sperm cryopreservation of silver barb (Barbodes gonionotus): cryoprotectants,cooling rate and storage time on sperm quality

Effectiveness and efficiency of frozen sperm on fertilization and hatching success of eggs from silver barb was examined in relation to cryoprotectants, freezing rate and storage period. Sperm was diluted in calcium‐free Hank's balanced salt solution, equilibrated with dimethylsulphoxide (DMSO), propylene glycol, sucrose or methanol at 5%, 10%, 15% or 20% final concentrations, and frozen in 250‐μL straws using a one‐step freezing procedure (1, 5 and 8°C min^?1 from 25 to ?40°C). Highest post‐thaw sperm motility was found from a treatment using 10% DMSO and 5°C min^?1 (82.2 ± 2.1%), similar to that of 10% DMSO and 8°C min^?1 (87.8 ± 3.2%). Post‐thaw motility of sperm frozen at 5 or 8°C min^?1 was significantly higher than 1°C min^?1. Relative sperm motility declined significantly after 10 months of cryostorage while viability did not change during a 12‐month cryostorage. Average fertilization rates of sperm after 1 and 4 months of storage were 64.5 ± 4.6% and 61.3 ± 3.4%, respectively, similar to those of fresh sperm (69.6–72.3%). Hatching rates of cryopreserved sperm (45.4–51.2%) were similar to those of fresh sperm (51.8–57.8%). This study developed suitable methods for cryopreservation of silver barb sperm that can be used to facilitate hatchery operation.     

Optimization of fertilization rate for maximizing periphyton production on artificial substrates and the implications for periphyton-based aquaculture

The effects of four rates of application of fertilizer, with cow manure (3000 kg ha⁻¹), urea (100 (kg ha⁻¹) and triple super phosphate (TSP) (100 kg ha⁻¹) (treatment F)), treatment F × 0.5 (treatment 0.5F), treatment F × 1.5 (treatment 1.5F) and treatment F × 2 (treatment 2F), on periphyton, plankton and water quality in tropical freshwater ponds were studied. The highest periphyton biomass in terms of dry matter (3.27 mg cm⁻² substrate), ash-free dry matter (2.06 mg cm⁻² substrate) and chlorophyll a (7.49 µg cm⁻² substrate) developed in treatment 1.5F. The ash content of periphyton was lower in treatment 1.5F (38% of dry matter) than in other treatments (57–66% of dry matter). Total ammonia and chlorophyll a of water increased with fertilization rate. Treatment 1.5F (cow manure, urea and TSP at rates of 4500, 150 and 150 kg ha⁻¹ respectively) appears to be the optimum, yielding high quantity and quality periphyton. By supplying a substrate area for periphyton equivalent to the pond surface, it was estimated that this level of fertilization could support a fish production of around 5000 kg ha⁻¹ y⁻¹, without recourse to supplementary food.     

Preservation-induced changes in morphometrics of fishes: influence on prey size-choice in juvenile walleye, Stizostedion vitreum (Mitchill)

Abstract– Juveniles of the Gulf coast population of walleye, Stizostedion vitreum , 22.4–89.9 mm SL ( n =111) shrank significantly in SL and throat diameter over 12 months after being fixed in 10% formalin for 1 week and preserved in 50% isopropyl alcohol thereafter. Significant changes in body depth and total length (TL) were also determined for select larval fish prey that juvenile walleye might encounter in nature. The relationship between fresh SL and throat diameter in juveniles of the Gulf coast population of walleye (22.4–172.4 mm SL) is: throat diameter (mm) =−0.8783 + 0.0786 SL (mm) ( r ²= 0.988, P < 0.0001, n = 135). In laboratory experiments on prey size choice, walleye selected available prey near or less than their mean throat diameter, although several larger prey (7.4% of total) were also consumed. To determine the actual spectrum of prey sizes available to juvenile piscivores, we must adjust the realized size of the predator and prey for changes caused by preservation. Preservation-induced alterations in morphology translated into errors in the perceived pattern of prey size choice in walleye. The preservation-induced changes documented in this study, which are also supported by a number of other empirical data sets, must be included when modeling the relationships between feeding ecology, morphology, growth, and ecologically relevant indices (condition factors, relative weights, etc.).     

The effects of salinity on the fertilization rate and rearing of the Persian sturgeon (Acipenser persicus) larvae

Persian sturgeon eggs were fertilized with different levels of salinities (0.5, 2, 4, 6, 8, 10, 12?ppt), and then each group was incubated in the same salinity until hatch. The fertility (%), hatching rate as well as larvae cumulative mortality rate and abnormality (%) were measured. Our Results revealed that Persian surgeon eggs could be fertilized in the different salinity concentrations but not more than 4?ppt. Moreover, hatching rate decreased with increase in salinities more than 2 and 4?ppt, respectively, and no larvae hatched in 6?ppt salinity. According to these results, the salinity tolerance threshold for Persian sturgeon larvae hatching in brackish water is less than 4?ppt.     

The effects of stream substratum composition on the emergence of salmonid fry

Sternecker K, Geist J. The effects of stream substratum composition on the emergence of salmonid fry.
Ecology of Freshwater Fish 2010: 19: 537–544. © 2010 John Wiley & Sons A/S Abstract – Salmonid fishes are target species for the conservation of freshwater habitats, but their natural reproduction is often insufficient. The emergence of fry is a crucial phase in the life cycle of salmonids and the stream substratum is the key habitat which regulates the emergence success. In this study, brown trout (Salmo trutta) and Danube salmon (Hucho hucho) eggs were exposed to different sediment textures and the emergence and the postemergence survival and growth were observed under constant water chemical conditions in the laboratory. In both species, textural effects on emergence rate, chronology of emergence, survival rate after emergence and growth after emergence were detected. Fine‐textured substratum (5–8 mm) formed a physical barrier to the posthatch migration of salmonids from the interstitial zone to the open water. The time period between the first and the last emerged fish was shorter in treatments with fine texture compared with coarse substratum. The survival rate was higher in treatments of coarser sediment. The effects of different textures on the growth of fry after emergence differed between brown trout and Danube salmon, which can be explained by different life history strategies. These results suggest that physical characteristics of substratum texture can have strong effects on salmonid emergence, and ultimately on the persistence of salmonid populations. They also suggest that biodiversity conservation in stream ecosystems can greatly benefit from an inclusion of the physical characteristics of the stream bed into catchment‐based management plans.     

鱼类应激反应的研究

综述了鱼类的应激反应与监测、养殖生产中应激危害的预防,阐述了应激激素系统下丘脑-垂体-肾间组织轴对鱼类生长、生殖和免疫机能的抑制作用,展望了今后鱼类应激领域的研究方向。