9/18/2020

New paper published in Chemosphere

 A new paper with Ákos' coauthorship was published in the journal Chemosphere. Enjoy!

Novak, M., Baebler, Š., Žegura, B., Rotter, A., Gajski, G., Gerić, M., Garaj-Vrhovac, V., Bakos, K., Csenki, Z., Kovács, R., Horváth, Á., Gazsi, G. & Filipič, M. (2020). Deregulation of whole-transcriptome gene expression in zebrafish (Danio rerio) after chronic exposure to low doses of imatinib mesylate in a complete life cycle study. Chemosphere, 128097.

Abstract

Imatinib mesylate (IM) is an anticancer drug that belongs to tyrosine kinase inhibitors. We report the results of the first investigation of the chronic exposure of zebrafish (Danio rerio) to IM. The exposure to IM (0.01, 1 and 100 μg/L) was initiated in adult fish and continued through hatching and the offspring generation for seven months. In addition to standard toxicological endpoints, induction of genotoxic effects and whole-genome transcriptome of liver samples of offspring generation of zebrafish were analysed. Exposure to IM did not affect the survival and growth of zebrafish, did not cause any histopathological changes, but it induced a marginal increase in the chromosomal damage in blood cells. The whole-genome transcriptome analyses demonstrated dose-dependent increase in the number of differentially expressed genes with a significantly higher number of deregulated genes in female fish compared to male. Differentially expressed genes included genes involved in response to DNA damage, cell cycle control and regulation of circadian rhythm. Based on the low genotoxic activity and the pattern of the changes in DNA damage responsive genes we consider that at current environmental exposure levels, IM represents low risk for genotoxic effects in aquatic organisms. Exposure to IM also induced deregulation of the expression of genes associated with steroidogenesis and hormone metabolism and function, which indicates hormone-disrupting activity of IM that has not been studied so far. The study provide new information on the potential consequences of chronic exposure to the residues of tyrosine kinase inhibitors, which remain to be further explored.

New paper published in Aquaculture Reports

 A new paper with Betti and Ákos as coauthors was published in the journal Aquaculture Reports. Enjoy!

Ljubobratović, U., Péter, G., Demény, F., Kugyela, N., Horváth, Á., Pataki, B., Horváth, Z., Jakabné Sándor, Z. & Rónyai, A. (2020). Reproductive performance in virgin pikeperch (Sander lucioperca L.) females fed different dietary levels of arachidonic acid with respect to the duration of spawning induction. Aquaculture Reports, 18, 100430.

Abstract

Pikeperch (Sander lucioperca) is a species of significant importance for the further development of intensive aquaculture, what makes the total control over species’ reproductive cycle a high priority task. The present study aimed to assess the effect of arachidonic acid (AA) dietary enrichment on the reproductive performance in virgin females (two-years-old; individual weight 1.0 ± 0.2 kg) kept in fully controlled conditions with respect to different time of hormonal application. Two groups of breeders, ARA (fed AA enriched diet) and CTRL (fed control feed), were reared in two separated tanks of the recirculation system over the course of seven months. Five, six, and seven months after the start of trial, batches of five pairs of breeders (in total 15 pairs per each feeding group) from each group were hormonally treated. Ovulations occurred after six and seven months (oocyte diameter range 912−1030 μm), and samples of dry eggs were taken for the analysis of fatty acid profiles. Among the parameters of reproductive success, the embryo survival was significantly affected by the diet, while induction duration affected latency time only. The CTRL group females showed overall higher embryo survival across both reproduction occasions (p = 0.028) than group fed AA enriched diet (35.7 ± 17.1 % and 18.6 ± 13.6 % in CTRL and ARA group, respectively). A significant accumulation of AA was found in ARA group eggs in both reproduction batches. Virgin pikeperch breeders appear to keep at least one month plasticity towards the duration of spawning induction in controlled conditions and optimal oocyte diameter range for artificial reproduction is between 900−1000 μm. Levelling dietary AA with eicosapentaenoic acid during the entire photothermal protocol negatively altered the egg quality.

New paper published in Dose-Response

 A new paper with Ákos' coauthorship was published in the journal Dose-Response. Enjoy!

Kerekes, F., Kollár, T., Gazsi, G., Kása, E., Urbányi, B., Csenki-Bakos, Z., & Horváth, Á. (2020). Investigation of Fertilizing Capacity of Zebrafish (Danio rerio) Sperm Exposed to Heavy Metals. Dose-Response. https://doi.org/10.1177/1559325820919597 

Abstract

The objective of our study was to investigate the effects of heavy metals on the fertilizing capacity of exposed zebrafish sperm, on embryonic survival, and on occurrence of embryonic deformities following fertilization with exposed sperm. It is important to test heavy metals because they are well-known pollutants. Sperm of externally fertilizing species can get in contact with pollutants found in aquatic environment. Zebrafish sperm, despite its advantages, has seldom been used in in vitro toxicological studies and no reports are available regarding the fertilizing capacity of exposed sperm. Zebrafish sperm was stripped and exposed to concentrations of the tested heavy metals (Zn2+, Cd2+, Cr3+, Cu2+, Ni2+, Hg2+, As3+) for 30 or 120 minutes. Calculated half-maximal effective concentration (EC50) values do not differ significantly from those calculated for motility for any of the tested heavy metals, which means fertilization rate can indicate the toxicity of the given substance following exposure of sperm. Thus, its application as in vitro toxicological end point is reasonable. The survival of embryos and embryonic development have not been affected by the exposure of spermatozoa, which means all alterations in spermatozoa caused by heavy metals have been expressed before 24 hours post fertilization.

New paper published in JoVE

 A new paper with Ákos' coauthorship was published in the Journal of Visualized Experiments (JoVE). Enjoy!

Csenki, Z., Horváth, Á., Bock, I., Garai, E., Kerekes, F., Vásárhelyi, E., Kovács, B., Urbányi, B., Mueller, F., Bakos, K. Using Tg(Vtg1:mcherry) Zebrafish Embryos to Test the Estrogenic Effects of Endocrine Disrupting Compounds. J. Vis. Exp. (162), e60462, doi:10.3791/60462 (2020).

Summary

Present here is a detailed protocol for the use of zebrafish embryos Tg(vtg1: mCherry) for the detection of estrogenic effects. The protocol covers the propagation of the fish and treatment of embryos, and emphasizes the detection, documentation, and the evaluation of fluorescent signals induced by endocrine disrupting compounds (EDC).


7/30/2020

New paper published in Cryobiology

A new paper with Ilija's first authorship was published in the journal Cryobiology.

Šćekić, I., Marinović, Z., Lujić, J., Müller, T., Kitanović, N., Urbányi, B., Horváth, Á., 2020. A novel strategy for conservation of European eel (Anguilla anguilla) genetic resources: Cryopreservation of ovarian stem cells. Cryobiology 95, 151–156. https://doi.org/10.1016/j.cryobiol.2020.03.009

Enjoy!

Abstract:

The aim of this study was to develop short- and long-term preservation protocols for European eel ovarian stem cells (OSCs) through hypothermic storage and cryopreservation of ovarian fragments that will assist in current conservation programs of this critically endangered species. Firstly, a freezing procedure was developed by testing different cryomedia and technical aspects of freezing. Utilization of 1.5 M of dimethyl sulfoxide (Me2SO), 0.1 M glucose and 1.5% BSA yielded optimal OSCs survival. Additionally, equilibration of 50-mg ovarian fragments for 30 min and plunging into lN2 at −80 °C displayed the highest OSC viability. Different cooling rates ranging from −1 to −40 °C/min did not significantly affect OSC viability when thawing in a 10 °C water bath. In addition, application of needle-immersed vitrification (NIV), combining ES3 (1.5 M PG and 1.5 M Me2SO) with VS3 (3 M PG and 3 M Me2SO) yielded the highest viability rates. Finally, hypothermic storage (4 °C) of ovarian fragments and ovarian cell suspensions displayed favorable viability of ~90% after 48 h of storage and ~65% after 72 h of storage. The development of OSC preservation methods presents an onset of further development of germline stem cell (GSC) manipulation techniques in this species. Cryopreservation of OSCs can enable a continuous supply of cells for either transplantation or in vitro cell culture thus enabling new and improved management and conservation strategies for this endangered species.

5/12/2020

New book chapter published

A chapter was published with the authorship of Ákos and Dr. Béla Urbányi in the book entitled Reproduction in Aquatic Animals: From Basic Biology to Aquaculture Technology edited by Dr. Manabu Yoshida and Dr. Juan F. Asturiano and published by Springer. The book is the final result of the activities of the COST Action FA1205 AQUAGAMETE as well as the International Symposium on “AQUAGAMETE: Reproduction of Aquatic Animals” held in the Joint Meeting of the 22nd International Congress of Zoology and the 87th meeting of the Zoological Society of Japan, which was held from 14th to 19th November 2016 in Okinawa, Japan. The authors wish to thank Dr. Yoshida and Dr. Asturiano for taking care of the book and making it happen. Enjoy!

Horváth Á., Urbányi B. (2020) Chapter 15 Sperm Cryopreservation of Aquatic Species. In: Yoshida M., Asturiano J. (eds) Reproduction in Aquatic Animals. Springer, Singapore, https://doi.org/10.1007/978-981-15-2290-1_16

Cryopreservation of fish sperm is feasible, and methods have been developed for several hundred species. The procedure includes the collection of sperm and assessment of its quality, dilution in suitable extenders, addition of cryoprotectants, loading into freezing devices, cooling, storage, thawing, and finally post-thaw use of sperm. An alternative to traditional freezing methods is the vitrification of sperm which is promising primarily in smaller model fish species. A wide variety of protocols are available in the literature; however, in spite of the significant progress made by the scientific community, commercial application of fish sperm cryopreservation is still very limited.

New paper published in Fish and Shellfish Immunology

A new paper with Ákos' co-authorship was published in the journal Fish and Shellfish Immunology. The paper was a result of a collaboration among Dr. Daniel Żarski of the Institute of Animal Reproduction and Food Research of the Polish Academy of Sciences, the team of Dr. Patrick Kestemont of the University of Namur in Belgium, Dr. Pascal Fontaine of the University of Lorraine in France and the Department of Aquaculture of Szent István University. It was also financed by the Hungarian-Walloon bilateral project TÉT_14_VL1-2015-0001. Enjoy!

Żarski, Daniel ; Ben Ammar, Imen ; Bernáth, Gergely ; Baekelandt, Sébastien ; Bokor, Zoltán ; Palińska-Żarska, Katarzyna ; Fontaine, Pascal ; Horváth, Ákos ; Kestemont, Patrick ; Mandiki, Syaghalirwa N.M. Repeated hormonal induction of spermiation affects the stress but not the immune response in pikeperch (Sander lucioperca). Fish and Shellfish Immunology 101 pp. 143-151. , 9 p. (2020)

Hormonal induction of spermiation, previously reported to be immunogenic in fishes, is a common hatchery practice in pikeperch, Sander lucioperca. The aim of the present study was to investigate the effects of repeated induction of spermiation in pikeperch, following application of either human chorionic gonadotropin (hCG) or salmon gonadoliberine analogue (sGnRHa) on sperm quality indices as well as on immune and stress response. Mature males of pikeperch (n = 7 per group) were stimulated twice with five days between injections of either hCG (hCG; 500 IU kg−1), sGnRHa (sGnRHa; 50 μg kg−1) or NaCl (control group; 1 ml kg−1) to assess spermatozoa motility with a computer-assisted sperm analysis (CASA) system. During second sampling, blood plasma was sampled for humoral innate immune (peroxidase and lysozyme activities, ACH50), stress (cortisol, glucose) and endocrine (testosterone) markers. In addition, the head kidney was dissected to assay the expression of several immune genes (such as il1, c3, hamp, tnf-α and lys genes). The results indicate that hormonal treatment significantly increased sperm production. Sperm sampled after the hormonal treatment maintained its quality throughout the study, regardless of the sampling time. However, it appears that the application of hCG induced elevated cortisol and glucose plasma levels compared to the control group. Almost all immune markers, except the relative expression of hepcidin (hamp gene), were unaffected by the two hormones applied. The results showed that the induction treatment of spermiation processes in pikeperch resulted in an important physiological stress response for which the intensity varied according to the hormonal agent used. However, this stress response (more profound following application of hCG) was weakly associated with innate immune functions. On the other hand, a significant negative correlation between the expression of several important immune markers (peroxidase activity, relative expression of c3 and il1 genes) and sperm quality indices indicates significant involvement of immune status on sperm quality. The results obtained shed light on immune-system- induced modifications to sperm quality. The data presented here highlight the need for careful revision of broodstock management and selection practices where welfare status as well as individual predispositions of fish to cope with the stress should be taken under the consideration.

5/11/2020

Conference in Croatia

The 55th Croatian & 15th International Symposium on Agriculture was held on February 16-21st, 2020, in Vodice, Croatia. Ákos has given a presentation entitled:
Ákos Horváth, Jelena Lujić, Zoran Marinović, Márton Hoitsy, Béla Urbányi, György Hoitsy:
The use of the hybrid tiger trout as a recipient for interspecific transplantation of spermatogonia

Ákos giving the presentation (photo by R. Balogh)

A trip to Poland

Although the COVID-19 pandemic has anchored us to Hungary without much opportunities to travel, on February 7-9th, thus before the outbreak we (Nevena, Zoran and Ákos) managed to visit our friend, Dr. Daniel Żarski of the Institute of Animal Reproduction and Food Research of the Polish Academy of Sciences. We also had the opportunity to meet Dr. Andrzej Ciereszko, head of Gamete and Embryo Biology group of the institute and members of his team. Our mission was to bring around 100 Atlantic salmon juveniles to Hungary for scientific experiments on their gonads. The fish arrived successfully after a 14-hour journey.

We wish to thank Daniel, Andrzej, Sylwia and all our friends in Poland for their help!

Strengthening the Polish-Hungarian-Serbian friendship at the dinner (photos by D. Żarski)


1/15/2020

Collaboration with Cornell University

In June-August, 2019 we had the pleasure of hosting Logan Goddard, undergraduate student of the College of Agriculture and Life Sciences (CALS) of Cornell University on a 8-week stay in our lab. Logan has participated in the CALS Global Fellows Program and took part in experiments on cryopreservation of eel gonads and carp sperm. He gave an account of his experiences at the 2019 Global Fellows Program Showcase in November 2019. You can read more about this program here: https://cals.cornell.edu/news/global-fellows-program-celebrates-first-year-undergraduate-research-experiences-hungary

10/20/2019

Defense of Patrícia Diogo

On 18th October, Ákos has participated as a member of the examination committee in the defense of the dissertation entitled “Cryopreservation of zebrafish germ cells: technological improvements and methodological standardization for gene banking and management” by Patrícia Diogo at the University of Algarve in Faro, Portugal. Patrícia was a PhD student of our friends Dr. Elsa Cabrita and Dr. Paulo Gavaia and a member of the Aquagroup at the same university and the Centre of Marine Sciences (CCMAR). She gave an excellend presentation of her results and convinced the committee of her profound knowledge in the area of her studies. Congratulations, Dr. Diogo!

Patrícia Diogo (center) with members of the committee (Dr. Paulo Gavaia, Ákos, Dr. Elsa Cabrita, Dr. Catarina Oliveira, Dr. Juan F. Asturiano and Dr. Sérgio Jesus) following the defense

10/03/2019

New paper published in Scientifc Reports

A new paper with the first authorship of Zoran was published in Scientific Reports. Enjoy!

Zoran Marinović, Qian Li, Jelena Lujić, Yoshiko Iwasaki, Zsolt Csenki, Béla Urbányi, Goro Yoshizaki & Ákos Horváth: Preservation of zebrafish genetic resources through testis cryopreservation and spermatogonia transplantation. Scientific Reports volume 9, Article number: 13861 (2019)

Abstract

Zebrafish is one of the most commonly used model organisms in biomedical, developmental and genetic research. The production of several thousands of transgenic lines is leading to difficulties in maintaining valuable genetic resources as cryopreservation protocols for eggs and embryos are not yet developed. In this study, we utilized testis cryopreservation (through both slow-rate freezing and vitrification) and spermatogonia transplantation as effective methods for long-term storage and line reconstitution in zebrafish. During freezing, utilization of 1.3 M of dimethyl sulfoxide (Me2SO) displayed the highest spermatogonia viability (~60%), while sugar and protein supplementation had no effects. Needle-immersed vitrification also yielded high spermatogonia viability rates (~50%). Both optimal slow-rate freezing and vitrification protocols proved to be reproducible in six tested zebrafish lines after displaying viability rates of >50% in all lines. Both fresh and cryopreserved spermatogonia retained their ability to colonize the recipient gonads after intraperitoneal transplantation of vasa::egfp and actb:yfp spermatogonia into wild-type AB recipient larvae. Colonization rate was significantly higher in dnd-morpholino sterilized recipients than in non-sterilized recipients. Lastly, wild-type recipients produced donor-derived sperm and donor-derived offspring through natural spawning. The method demonstrated in this study can be used for long-term storage of valuable zebrafish genetic resources and for reconstitution of whole zebrafish lines which will greatly improve the current preservation practices.

Ákos promoted into professorship

By the Decree 351/2019. (VIII. 30.) of the President of the Republic of Hungary Ákos was appointed as a professor as of 1st September, 2019.

7th International Workshop on the Biology of Fish Gametes

Ákos, Timi, Betti and Nevena have participated at the 7th International Workshop on the Biology of Fish Gametes that was organized between 2-6th September in Rennes, France. As always, the workshop was a wonderful opportunity for scientist working on fish gametes to meet in person and share their newest results. Ákos, Timi and Nevena gave oral presentations durind the sessions while Betti presented a poster. We would like to thank the French colleagues for the wonderful experience and professional organization. Looking forward to seeing everyone in two years in Poland.

Oral presentations by the group:

Tímea Kollár, Bernadett Pataki, Gyöngyi Gazsi, Béla Urbányi, Ákos Horváth: Effect of age on the sensitivity of zebrafish (Danio rerio) sperm
Ákos Horváth, Zoran Marinović, György Hoitsy, Márton Hoitsy, Boglárka Hoitsy, Béla Urbányi, Jelena Lujić: Is the hybrid tiger trout a good recipient for the transplantation of early-stage germ cells?
Ilija Šćekić, Nevena Kitanović, Zoran Marinović, Tamás Müller, Béla Urbányi, Jelena Lujić, Ákos Horváth: Preservation of ovarian tissue in european eel (Anguilla anguilla)

Posters presented or co-authored by the group:

Roman Franěk, Taiju Saito, Tomáš Tichopád, Michaela Fučíková, Zoran Marinović, Jelena Lujić, Ákos Horváth, Vojtěch Kašpar, Martin Pšenička: Germ cell manipulation as a tool for common carp isogenic lines production and management.
Bernadett Pataki, Tímea Kollár, Zoran Marinović, Jelena Lujić, Gyöngyi Gazsi, Roberta Izabella Berta, Béla Urbányi, Ákos Horváth: Inheritance of sperm cryoresistance in zebrafish (Danio rerio)


7/06/2019

Frontires in Reproduction: Molecular and Cellular Concepts and Applications (FIR)

Jelena spent six weeks at the Marine Biological Laboratory (University of Chicago) participating in the course Frontires in Reproduction: Molecular and Cellular Concepts and Applications (FIR). During that time, she had lectures in all relevant methods in reproductive biology such as endocrinology, stem cells, gametogenesis, fertilization, sex determination etc. Most of the time was devoted to laboratory practice and performing experiments regarding signalling, CRISPR Cas9, epigenetics, in situ hybridization, tissue and oocyte culture, germ cell transplantation, oocyte maturation, ICSI, IVF, embryo transfer, ESC microinjection, reproductive immunology, placentation etc.
Beside the educational part, this was the perfect opportunity for networking. The course itself has a very long tradition (since 1998) and the FIR community is very significant in the area of reproductive biology and includes scientists from around the world (circa 400 people).
Beside the support of our University, Jelena`s participation was partly funded by the NICHD and the Burroughs Wellcome Fund.
Jelena can now proudly say that she is a FIRbee 😊

FIRbees 2019
FIRbees 2019 with section directors and organizers
Working mode
Mission accomplished - sperm lab
Science is exciting



Chalk talk about fish surrogate production
Wrap up of the labs from the Section 1
Jelena`s presentation at FIR symposium 2019
FIRbees 2019 at the sunset in the Woods Hole
FIRbees 2019 at the closing reception
FIRbees 2019 at the closing reception
FIRbees during the weekend
It is written in stone, no, in the tissue
Grill party and fun
We are big happy family now
Present from dear colleague Augustin

6/06/2019

Teaching in Pula and Zagreb, Croatia

On the invitation of our colleague Dr. Ana Gavrilović from the University of Zagreb in Croatia, Ákos has spent the period between May 23-27th in Pula and Zagreb. This included teaching the basics of cryobiology, fish sperm and germ cell cryopreservation and their transplantation to undergraduate students of the University of Pula and to graduate students in Zagreb. The trip was also a good opportunity for Ákos to practice his original qualification of a fisheries and aquaculture person as he joined a fishing expedition with Ana and her student Neven Iveša. As you see in the picture below, the catch was quite abundant and a real professional treat for Ákos.

Ákos giving a presentation in Pula

The catch of the day

New review paper published in Theriogenology

A new review paper with the co-authorship of Ákos has been published in Theriogenology. Enjoy!

Juan German Herranz-Jusdado, Victor Gallego, Marina Morini, Christoffer Rozenfeld, Luz Pérez, Tamás Müller, Ákos Horváth, Hiromi Ohta, Juan F. Asturiano: Eel sperm cryopreservation: An overview. Theriogenology, 133, 210-215. https://doi.org/10.1016/j.theriogenology.2019.03.033

The eels are teleost fishes from the order Anguilliformes that includes several species with high commercial value. Due to the high interest for aquaculture production of some eel species and for the need to restore eel species that are endangered, several research groups have directed their research toward developing protocols to cryopreserve the spermatozoa of Japanese eel (Anguilla japonica) and European eel (Anguilla anguilla). In this review, we provide an overview on the different protocols that have been developed so far. The first developed protocols used DMSO as cryoprotectant in both species with good success, obtaining sperm motilities of over 45% in Japanese eel and over 35% in European eel. Moreover, sperm cryopreserved using DMSO was successfully used in fertilization trials, although with low fertilization rates. However, recent studies show that DMSO produce epigenetic changes in eel sperm and therefore, the last developed protocols used methanol as cryoprotectant instead. Cryopreservation protocols using methanol as cryoprotectant, showed improved motility values in both Japanese and European eel. In addition, the latest protocols have been adapted to cryopreserve larger volumes of sperm of up to 5 mL, which is useful for larger scale fertilization trials.
The present study introduces the state of the art and future perspectives of the eel sperm cryopreservation to be applied in aquaculture and biological conservation programs.

4/27/2019

New paper accepted in PlosOne


A new paper in collaboration with our colleagues from University of South Bohemia in Česke Budejovice, Faculty of Fisheries and Protection of Waters (Czech Republic) has been accepted for publication in PlosOne.

Franěk R., Marinović Z., Lujić J., Urbányi B., Fučíková M., Kašpar V., Pšenička M., Horváth Á. 2019. Cryopreservation and transplantation of common carp spermatogonia. PLoS One. 14(4):e0205481. DOI: 10.1371/journal.pone.0205481

Abstract
Common carp (Cyprinus carpio) is one of the most cultured fish species over the world with many different breeds and plenty of published protocols for sperm cryopreservation. However, data regarding preservation of gonadal tissue and surrogate production is still missing. A protocol for freezing common carp spermatogonia was developed through varying different factors along a set of serial subsequent experiments. Among the six cryoprotectants tested, the best survival was achieved with dimethyl sulfoxide (Me2SO). In the next experiment, a wide range of cooling rates (0.5–10°C/min) and different concentrations of Me2SO were tested resulting in the highest survival achieved using 2 M Me2SO and cooling rate of -1°C/min. When testing different tissue sizes and incubation times in the cryomedia, the highest viability was observed when incubating 100 mg tissue fragments for 30 min. Finally, sugar supplementation did not yield significant differences. When testing different equilibration (ES) and vitrification solutions (VS) used for needle-immersed vitrification, no significant differences were observed between the tested groups. Additionally, varied exposure time to VS did not improve the vitrification outcome where the viability was 4-fold lower than that of freezing. The functionality of cryopreserved cells was tested by interspecific transplantation into sterilized goldfish recipients. The exogenous origin of the germ cells in gonads of goldfish recipient was confirmed by molecular markers and incorporation rate was over 40% at 3 months post-transplantation. Results of this study can serve for long-term preservation of germplasm in carp which can be recovered in a surrogate recipient.