Microsatellites from the burbot (Lota lota), a freshwater gadoid fish (Teleostei)

We developed 21 polymorphic dinucleotide microsatellite loci, (CA)_n and (CT)_n, for the Holarctic freshwater fish, Lota lota, using an enriched genomic library protocol. The species has an interesting life history because winter‐spawning adults migrate over long distances to form spawning aggregations, a behaviour which should maintain genetic homogeneity across large spatial scales. Availability of the reported microsatellites will facilitate the investigation of population genetic structure with regard to postglacial colonization history and conservation strategies. The primers were screened on 30 individuals from a natural population (Lake Constance, southern Germany), revealing three to 24 alleles per locus with expected heterozygosities ranging from 0.48 to 0.93.     

Food,growth, habitat,and community interactions of young-of-the-year burbot,Lota lota L., in a Precambrian Shield lake

The burbot, Lota lota, is a widely distributed gadid of the northern circumpolar regions of North America and Eurasia. Despite its near ubiquity over much of its range, relatively little is known about its biology during the first year of life.Burbot sac-fry of 3 mm total length, hatched under the ice in early May in Shebandowan Lake. Their first foods following atrophy of the yolk-sac were copepods and cladocerans which they captured pelagically. At first the fry swam in small schools, high in the water column of the near-shore littoral, and fed during the daytime. Upon reaching 30 mm in total length, the burbot fingerlings became solitary and benthic, and fed primarily at night, almost exclusively on the amphipod, Hyalella azteca. Amphipods constituted about 75 per cent by number of all the food consumed by burbot fingerlings in their first year of life. Growth was rapid from May to the end of July, tapered off during August to October, and effectively stopped by November at freeze-up. The principal habitat of burbot fingerlings during the benthic stages of their life was the near-shore littoral, in depths ranging from 0.5 m to 4.0 m. There, they were sheltered by boulder shoals, sunken trees or other debris, or beds of quillworts (Isoetes sp.). The principal ichthyofauna that cohabited with the burbot included the sculpin (Cottus cognatus), the smallmouth bass (Micropterus dolomieui), two etheostomatines (Etheostoma nigrum) and (E. exile) and the yellow perch (Perca flavescens). Burbot fingerlings were subjected to low levels of predation from the nocturnal foraging of walleyes (Stizostedion vitreum).     

The status of the burbot Lota lota (L.) (Gadidae) in Britain

Records of burbot Lota lota (L.) captures from the early nineteenth century to the present have been gathered and arranged watershed by watershed in chronological order. Most are from eastward-flowing river systems from Durham southwards to the Great Ouse, but a few records from westward-flowing systems are considered. In many areas the records imply a decline of burbot numbers and distribution during the present century, though burbot may never have been more than locally abundant. Local over-fishing, pollution and habitat changes are considered the most likely causes of decline. Conservation measures seem desirable.     

Looking for love under the ice: Using passive acoustics to detect burbot (Lota lota: Gadidae) spawning activity

1. Burbot (Lota lota: Gadidae) is a difficult species to manage effectively due to its preference for deep-water habitats and under-ice spawning behaviour, resulting in a poor understanding of its reproductive activity. However, the use of acoustic signalling by burbot as part of their mating system has recently been described and this behaviour may provide a means of investigating questions regarding the spatial and temporal distribution of spawning aggregations using passive acoustic monitoring.
2. We used audio and video recording to confirm that burbot vocalise and that these vocalisations can be detected under field conditions as well as to characterise the relationship between burbot acoustic signalling and spawning behaviour. We also evaluated the feasibility of locating and monitoring burbot spawning aggregations in real time using passive acoustics.
3. Burbot vocalisations were difficult to identify with only about 6% of the recordings containing calls being successfully identified as such in the field. Burbot vocalised more often between sundown and sunrise than during daylight hours. Calls recorded at night tended to be lower frequency, longer duration, and have lower bandwidth than those made during the day.
4. Burbot vocalisations could not be recorded in conjunction with video recordings of spawning activity, indicating that burbot may not call during active spawning, but may use acoustic communication to signal the onset of reproductive readiness and to form pre-spawning aggregations.
5. While burbot calls were readily identifiable, observers had a difficult time identifying burbot calls in real time under field conditions. Passive acoustic monitoring demonstrates considerable potential as a management tool to locate burbot spawning grounds and identify periods of activity, but may not be an appropriate technique for monitoring spawning activity in real time.

     

Comparison of two techniques for the morphometry study on gilthead seabream (Sparus aurata) spermatozoa and evaluation of changes induced by cryopreservation

The development of powerful software has made possible spermatozoa morphology studies. However, some problems have emerged in relation to protocol standardization to compare results from different laboratories. This study was carried out to compare two techniques commonly used (staining vs phase contrast technique) for the morphometry study of gilthead sea bream spermatozoa using an integrated sperm analysis system (ISAS). Spermatozoa morphometry values were significantly affected by the technique used, and phase contrast technique was found to be the more accurate method, showing lower coefficients of variation on spermatozoa morphometry parameters measurements. Moreover, it has been shown that cryopreservation process produces damage in gilthead sea bream spermatozoa, causing negative effects in sperm parameters as spermatozoa morphometry (a decrease in cell volume), motility (from 95 to 68% motile cells) and viability (from 95 to 87% of live cells), being the addition of freezing medium containing cryoprotectant (DMSO) an important factor that caused the morphometry changes.     

Isolation and characterization of new microsatellite markers from the burbot (Lota Lota)

Ten polymorphic microsatellite markers were developed to examine the population structure of the burbot (Lota Lota). The number of alleles ranging from 2 to 14 and the expected heterozygosity ranging from 0.306 to 0.926 were screened in the wild samples from Wusuli River in Heilongjiang Province, China. Eight loci significantly deviated from Hardy–Weinberg equilibrium. No evidence showed significant linkage disequilibrium between pairs of loci. These newly isolated markers would also enrich the available molecular resources of the burbot.     

Diet, prey selection, and growth of larval and juvenile burbot Lota lota (L.)

The diet, prey selection, and growth of larval and juvenile Oneida Lake burbot, Lota lota (L.), was investigated from the time of first hatch in mid-April until their disappearance from the pelagic zone at the beginning of June. Mean length increased exponentially from 3–20 mm on 18 April to 15–00 mm on 29 May. The number and size of prey in burbot stomachs, and the prey size most selected by burbot increased as they grew. Prey width, not prey length, limited the size of prey ingested. The smallest burbot ate and selected highly for the largest prey they could ingest, but older burbot did not. As burbot grew, prey selection was highest first for the rotifer Asplanchna , followed by copepod nauplii, then cyclopoid copepods. Daphnia and calanoid copepods and cyclopoid copepods were selected to similar degrees by 29 May. For a given size, however, cyciopoid copepods were selected more strongly than any other prey type from 18 to 29 May. There were no significant differences in prey selection among offshore, nearshore, and onshore habitats, nor among depths in the offshore habitat.     

Inhibitory effect of osmotic concentration, potassium and pH on motility of the sperm of the North American burbot Lota lota maculosa

Seminal plasma factors maintaining North American (NA) burbot Lota lota maculosa sperm quiescent were examined. Sperm were diluted into buffered saline solutions of various compositions and motility assessed. After 1 h in these solutions at 10° C, aliquots of the suspension were diluted with tap water and motility again assessed. Dilution of sperm in an incubation solution containing Ca²⁺ in the absence of K⁺ initiated sperm motility resulting in low motility when sperm were subsequently diluted in tap water. Incubation solutions with osmolalities >200 mOsm kg⁻¹ and containing 12·5 mM K⁺ prevented the onset of sperm motility and were associated with maximal sperm motility upon dilution in tap water. Sperm maintained at lower osmolalities exhibited limited motility upon dilution in tap water indicating interdependence between K⁺ and osmolality in maintaining sperm quiescent in the presence of Ca²⁺. Sperm kept in incubation solution at pH values < c. 7·5 for 1 h demonstrated reduced motility when subsequently diluted in tap water. That motility of sperm was pH sensitive was further indicated by CO₂ inhibition of motility. Therefore, NA burbot sperm are probably maintained in an immotile state, yet with potential for motility, by combination of high K⁺, osmolality and possibly pH. The results from this study differ from published information on sperm quiescence in the temporally and geographically distinct Eurasian burbot Lota lota lota .     

Influence of cryoprotectants on abnormality and motility of baung (Mystus nemurus) spermatozoa after long-term cryopreservation

Study on the effect of cryoprotectants on abnormality and motility of baung, Mystus nemurus spermatozoa were evaluated using transmission and scanning electron microscopy. Four cryoprotectants, dmso, ethanol, methanol and glycerol at concentration of 10% were tested in triplicates. Three ml of fresh sperm which was diluted with 60 ml of ringer solution was added to each of twelve 5-ml vials containing of 0.50-ml of the cryoprotectants. The vials were placed in an icebox containing dry ice 5 min and then storage into container containing liquid nitrogen for 13 months. The effect of cryoprotectants on the spermatozoa abnormality and motility were significant (P < 0.05). The spermatozoa abnormality was significantly lower in methanol (62.65%) compared with the other cryoprotectants. The spermatozoa motility was higher in methanol, but not significantly different with ethanol (P > 0.05). It is a negative correlation between sperm motility and abnormality. Generally, higher abnormalities of spermatozoa resulted low motility.     

Cryopreservation of the endangered mahseer (Tor khudree) spermatozoa: I. Effect of extender composition, cryoprotectants, dilution ratio, and storage period on post-thaw viability

Several in situ and ex situ conservation strategies have been suggested for the revival of stocks of Tor khudree (Sykes), a threatened species. Cryopreservation of spermatozoa is crucial for the conservation of stocks of endangered species so that sustainable production can be ensured. Among the different extenders, modified fish Ringer (E1) was found to be the best for cryopreservation of T. khudree spermatozoa. Extender E2 appeared the next best. Extenders based on chicken egg yolk and milk powder were found to be unsuitable for the cryopreservation of T. khudree spermatozoa. Among the cryoprotectants, dimethyl sulfoxide provided maximum protection to spermatozoa during freezing and thawing. Propylene glycol and methanol were found to be less effective. Of the four spermatozoa dilutions, 1:10, 1:15, and 1:20 showed better motility rates than 1:5. At the former dilution ratios, the motility rates which were more than 95% prior to freezing were reduced to 80-81 and 43-67%, 10 and 70 days after cryopreservation, respectively. The motility duration did not differ much with increasing storage period at all the dilution ratios. Motility rates generally decreased with an increase in frozen storage. When spermatozoa were thawed and stored at 25 degrees C for varying periods, motility percentage, and duration decreased gradually as the storage period increased; spermatozoa stored up to 40 min after thawing retained 55% motility and were motile up to 77s; these values declined further leading to the complete cessation of motility 70 min after storage. The importance of extender-cryoprotectant mixture, milt dilution, and storage period in developing a protocol for T. khudree spermatozoa cryopreservation is discussed.     

Genetic admixture of burbot (Teleostei: Lota lota) in Lake Constance from two European glacial refugia

The burbot, Lota lota, is the only freshwater species of the codfish family and has a Holarctic distribution. Pleistocene glaciations caused significant geographical differentiation in the past, but its life history characterized by winter spawning migrations over large distances is likely to homogenize populations by contemporary gene flow. We investigated the population genetic structure of 541 burbots from Lake Constance and adjacent Rhine and Danube tributaries in Europe using the entire mitochondrial DNA (mtDNA) control region and 11 microsatellites. Microsatellites revealed considerable population divergence (F(ST) = 0.26) and evidenced recent bottlenecks in two Central European rivers. In accordance to previous evidence two main phylogeographic lineages (Atlantic and Danubian) were found co-occurring at similar frequencies in Lake Constance, where they currently undergo random mating as indicated by microsatellites. The Danubian lineage contributed only a small proportion to the lake's mtDNA diversity, and probably expanded within the lake shortly after its formation approximately 10,000-15,000 BP. The larger Atlantic haplotype diversity suggested a population expansion older than the lake itself. Levels of admixture at microsatellite loci were less obvious due to their high variability, and coalescence methods were used to estimate past admixture proportions. Our results reinforce a model of a two-step colonization of Europe by burbot from an ancestral Danubian refuge, and confirm the persistence of a secondary Atlantic refuge, as proposed to exist for other freshwater fish. We conclude that the present-day burbot population in Lake Constance bears the genetic signature of both contemporary gene flow and historical separation events.     

Seasonal changes in condition indices in adult mature and non-maturing burbot, Lota lota (L.), in the north-eastern Bothnian Bay, northern Finland

The population structure and seasonal changes in condition factors of the burbot in a shallow coastal region of the north-eastern Bothnian Bay are described. The significance of the so-called rest years is examined by comparing condition indices in immature or sterile and mature burbot. The somatic condition index ( K ₂), liver index ( K ₁), intestine index ( K ₁) and gonad index ( K_g ) are determined monthly in terms of organ weight in relation to body length. Approximately 30% of the whole catch of 1052 burbot were non-maturing but were 40 cm or more in length. K ₂, K ₁ and K ₁ were lowest in the autumn, when the first sign of gonad recrudescence was observed in mature burbot. The non-maturing burbot were never in poorer condition than mature ones. As mature and non-maturing burbot dissipated their energy stores during the warmest period of the summer, it is concluded that burbot spending a rest year do not accumulate and store energy reserves over the summer for the next year, and that such rest years, if they exist, do not occur for nutritional reasons.     

Effects of extender composition, cooling rate and freezing on the fertilisation viability of spermatozoa of the Pacific oyster (Crassostrea gigas)

The aim of this research was to optimise protocols for freezing spermatozoa of the Pacific oyster. All the phases of the cryopreservation procedure (choice of cryoprotectant, cooling, freezing, and thawing) were studied in relation to the species of spermatozoa to restore on thawing the morphological and physiological characteristics of fresh semen. The choice of type and concentration of cryoprotectant in which semen is incubated before freezing is fundamental for a successful cryopreservation: the cryoprotectants (dimethylsulfoxide--Me(2)SO, ethylene glycol--EG, propylene glycol-PG, and glycerol in concentrations between 5 and 15%) were tested for their toxicity on the semen exposed up to 30 min at +26 degrees C (room temperature) by evaluating its ability to fertilise and the embryo development to the regular D larval stage. The best cryoprotectants, Me(2)SO, EG, and PG 5, 10, and 15% respectively, were used for the pre-cooling (adaptation/cooling) tests. Two different adaptation/cooling procedures were tested: (A) from +26 degrees C to 0-2 degrees C (2.6 degrees C/min) and (B) at +26 degrees C for 15 min. Lastly, using the cryoprotectants and the adaptation procedure (B) that had given the best results in the preceding stages of the experiment, four cooling rates were tested: 6, 11, 16, and 21 degrees C/min. It was seen that the semen that was incubated with EG 10%, adapted at +26 degrees C for 15 min, and then cooled at a rate of 6 degrees C/min showed a percentage of regular D larvae on thawing comparable to that of fresh semen (p > 0.05).     

Field estimations of temperature-dependent processes: case growth of young burbot

In this study, the growth of young burbot (Lota lota) was temperature-dependent, but varied between age-groups. The growth of age-0 increased with temperatures 4–18°C. Growth of age-groups 1 and 2 was not significantly related to surface water temperatures and varied between the 2 years studied. This study was a re-evaluation of burbot sampled between 1978 and 1980 by plankton net, beach seine, and electric fishing in Lake Kuohijärvi, the Finnish Lake District. In the re-evaluation, growth was studied both as mm day^–1 and as absolute growth (mm), whereas the original study did not include absolute growth. Though the methods should be equivalent, they are not. In our example, time between measurements, temperature, and growth were negatively correlated. When analysing mm day^–1 the estimate variability, therefore, increased with temperature, whereas the variability was homogenously spread on the two axes, growth and degree-day, when analysing absolute growth. Field rate analyses contain an error risk that should be recognised when assessing the temperature-dependency of any processes, e.g. growth or mortality.     

Features of Burbor Lota lota (Gadidae) Biology in the Lower Volga

Journal of Ichthyology - The study presents data on the biology of the marginal population of burbot Lota lota from the lower reaches of the Volga. Individuals aged from 3 to 7 years were recorded...     

Gamete cryobanks for laboratory research: Developing a rapid and easy-to-perform protocol for the cryopreservation of the sea urchin Paracentrotus lividus (Lmk, 1816) spermatozoa

Gamete cryopreservation is a biotechnology that can guarantee a continuous supply of gametes, regardless of the seasonal reproductive cycle. In this study we developed a protocol for the cryopreservation of the sea urchin Paracentrotuslividus spermatozoa, with a view to the creation of cryobanks of semen to be used as a model system in laboratory research and ecotoxicological tests. All the key phases of the procedure were separately considered and the effect on sperm motility was evaluated by means of computer assisted analysis. The best results were obtained using 7% dimethylsulfoxide in 1% NaCl plus 0.04 M trehalose as the extender, at a freezing rate of −20 °C/min. On thawing, in semen samples cryopreserved in accordance with this protocol the velocity parameters of the sub-population of rapid sperm (best performing spermatozoa) did not significantly differ from semen on collection; in addition also the fertilization ability was restored, and about 50% of normal developed plutei larvae were obtained by thawed semen. The developed protocol is rapid and easy-to-perform; moreover, the use of gametes from reared urchins makes it unnecessary to continuously collect specimens from natural populations, making this procedure a promising starting point for the creation of alternative and more sustainable methodologies in laboratory research on sea urchin gametes and embryos.     

Cryopreservation of the Mediterranean mussel (Mytilus galloprovincialis) spermatozoa

The effects of cryoprotectants, cooling rate and freezing on the mussel Mytilus galloprovincialis sperm were evaluated. At the end of each step of the experimental protocol, motility and fertilization ability of sperm were analyzed, compared to fresh semen. Five cryoprotectants were tested in their toxicity level: dimethylsulfoxide, ethylene glycol, 1-2 propylene glycol at 5%, 7%, 10%, 15% and 20% concentration; glycerol and methanol at concentration of 5%, 7% and 10%. The incubation times were 10, 20 and 30 min at 20 ± 1 °C. Only dimethylsulfoxide, ethylene glycol and 1-2 propylene glycol at 5%, 7% and 10% were chosen for the following pre-freezing step. Five adaptation/chilling rates were analyzed: 10 min at 20 ± 1, −2, −1, −0.5 and −0.25 °C/min and the last one was used for testing the best freezing procedure among seven gradients. Particularly, two rapid rates, three slow rates and two double step rates were conducted.Thawing results showed that M. galloprovincialis sperm are very sensitive to rapid pre-freezing and freezing protocols and only a slow procedure assured good motility and fertilization percentages.     

A checklist of protozoan and metazoan parasites of the burbot (Lota lota)

The parasite fauna of the burbot (Lota lota) within its natural range is reviewed. The sent paper summarizes the data on parasites of the burbot from water bodies of Eurasia and North America, based on published monographs, reviewed journals, scientific reports, conference contributions, and PhD theses. The checklist includes all protozoan and metazoan parasites of the burbot. A total of 242 parasite species/taxa were recorded in the burbot (Ki-netoplastomonada--4, Parasitomonada--3, Coccidiomorpha--1. Microsporidea--3, Myxosporidia--35, Pleurostomata--1. Cyrtostomata--3, Peritricha--20. Protozoa incertae sedis--1. Monogenea--8, Cestoda--23, Digenea--50. Nematoda--36, Acanthocephala--28, Hirudinea--11. Bivalvia--5, Crustacea--10). Most parasites belong to digenean trematodes. Most of these species (183 species/taxa) were recorded on Eurasian and only 92--in North America fishes. Several parasite species recorded from the burbot are discussed in relation to host specificity and their geographical distribution.     

Preliminary studies on cryopreservation of snakehead (Channa striata) embryos

This paper reports the findings of the ongoing studies on cryopreservation of the snakehead, Channa striata embryos. The specific objective of this study was to collect data on the sensitivity of C. striata embryo hatching rate to low temperatures at two different developmental stages in the presence of four different cryoprotectants. Embryos at morula and heartbeat stages were selected and incubated in 1 M dimethyl sulfoxide (Me₂SO), 1 M ethylene glycol (EG), 1 M methanol (MeOH) and 0.1 M sucrose solutions at different temperatures for a period of time. Embryos were kept at 24 °C (control), 15 °C, 4 °C and −2 °C for 5 min, 1 h and 3 h. Following these treatments, the embryos were then transferred into a 24 °C water bath until hatch to evaluate the hatching rate. The results showed that there was a significant decrease of hatching rate in both developmental stages following exposure to 4 °C and −2 °C at 1 h and 3 h exposure in each treatment. Heartbeat stage was more tolerant against chilling at −2 °C for 3 h exposure in Me₂SO followed by MeOH, sucrose and EG. Further studies will be conducted to find the best method to preserve embryos for long term storage.     

Protective effects of propolis on cryopreservation of common carp (Cyprinus carpio) sperm

Cryopreservation of sperm is common procedures in aquaculture, particularly used for routine in artificial insemination. However, these application cause damages and adversely affected sperm motility, viability and consequently lower hatching rates. The objective of this study is to determine whether propolis has an effect on cryopreservation and fertilization ability and to investigate the potential protective effect of propolis on spermatozoa of Cyprinus carpio. Many studies have been done in cryopreservation offish spermatozoa, but none of them contain propolis in extender composition. The extenders were prepared by using modified Kurokura Solution to which 10% Me₂SO added with different levels of propolis (0.2, 0.4, 0.6, 0.8 and 1 mg ml⁻¹) and 10% egg yolk (as a control without propolis). The pooled semen samples diluted at the ratio of 1:9 by the extenders were subjected to cryopreservation. The percentage and duration of motility and fertilization tests of cryopreserved sperm samples have been done immediately after thawing and compared with control and fresh semen. The extenders containing propolis exhibited higher percentage motility and motility duration than control group (P < 0.05). Especially the group IV (0.8 mg ml⁻¹ propolis) and the group V (1 mg ml⁻¹ propolis) showed significant positive effects on both post thaw motility and hatching ability. The propolis maintained the integrity of the spermatozoa during the cryopreservation process. Evaluating with its contents, it has been shown that propolis is an appropriate cryoprotective agent in fish semen.