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Abstracts: CryoLetters 36 (4), 2015

 

 

Volume 36, No. 4 July/August 2015

ISSN 0143-2044

 

 


Long-term preconditioning of plantlets: a practical method for enhancing survival of pineapple (Ananas comosus (L.) Merr.) shoot tips cryopreserved using vitrification
Wei-Hsin Hu, Shu-Fen Liu and Song-Iuan Liaw

226-236

 

 


Cold hardiness and range of the myriapod Angarozonium amurense (polyzoniidae, diplopoda, arthropoda) in permafrost environments
Daniil I. Berman, Ekaterina N. Meshcheryakova
and Elena V. Mikhaljova

237-242

 

 


The combined use of honey, garlic (Allium Sativum L.) and skimmed milk as an extender for chilling sheep semen
Rodrigo Jerez-Ebensperger, Lydia Gil, Noelia Gonzalez
and Ignacio De Blas

243-251

 

 


Physiological, cytological and biochemical stability of Medicago sativa L. cell culture after 27 years of cryogenic storage
Liudmila A. Volkova, Valentina V. Urmantseva,
Elena V. Popova and Alexander M. Nosov

252-263

 

 


Effects of orvus es paste on the motility and viability of yak (Bos grunniens) epididymal and ejaculated spermatozoa after freezing and thawing
Megumi Shimazaki, Rentsenkhand Sambuu, Yoko Sato,
Lanh Thi Kim Do, Fuminori Tanihara, Masayasu Taniguchi
and Takeshige Otoi

264-269

 

 


Relationship between supercooling capability and cryoprotectant content in eggs of Pararcyptera microptera meridionalis (orthoptera: acrypteridae)
Xiao-Rong Zhou, Yan-Yan Li, Na Li and Bao-Ping Pang

270-277

 

 


Temporary storage of bovine semen cryopreserved in liquid nitrogen on dry ice and refreezing of frozen-thawed semen
A.M. Abdussamad, M. Gauly and W. Holtz

278-284

 

 


Determination of convective heat transfer coefficient at the outer surface of a cryovial being plunged into liquid nitrogen
Tao Wang, Gang Zhao, Heyu Tang and Zhendong Jiang

285-288

 

 

 

 

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CryoLetters 36 (4), 226-236 (2015)
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LONG-TERM PRECONDITIONING OF PLANTLETS: A PRACTICAL METHOD FOR ENHANCING SURVIVAL OF PINEAPPLE (ANANAS COMOSUS (L.) MERR.) SHOOT TIPS CRYOPRESERVED USING VITRIFICATION

Wei-Hsin Hu1, 3, Shu-Fen Liu2, 3 and Song-Iuan Liaw2*

1Department of Biology, National Museum of Natural Science, Taichung, Taiwan
2Department of Life Sciences, National Chung-Hsing University, Taichung, Taiwan
3Equal contributors
*Corresponding author e-mail:
siliaw@mail.nchu.edu.tw

Abstract

BACKGROUND: The purpose of this study was to develop an efficient cryopreservation protocol for pineapple (Ananas comosus (L.) Merr.) shoot tips. OBJECTIVE: The optimal state of pineapple plantlets was investigated by using sucrose preconditioning to enhance survival after cryostorage. METHODS: To achieve a suitable state of plantlets before cryopreservation, 0.2 M to 0.4 M sucrose concentrations combined with short- (07 days), medium- (1530 days), and long-term (75150 days) preconditioning periods were compared. RESULTS AND CONCLUSION: The highest survival (100%) was achieved using the following procedure: intact plantlets underwent long-term preconditioning with 0.2 M sucrose for 135 days, dissected shoot tips were treated with a loading solution containing 2.0 M glycerol + 0.4 M sucrose for 60 min at 25C and the shoot tips were dehydrated in PVS2 for 2 h at 0C before being plunged in liquid nitrogen. Rewarming was conducted in a water-bath for 30 s at 40C and PVS2 was replaced with a 1.2 M sucrose solution for 30 min at 25C. The shoot tips were transferred on semisolid medium and left in the dark for 1 week, then in dim light for 3 weeks.

Keywords: sucrose preconditioning, pineapple, vitrification

 

 

 

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CryoLetters 36 (4), 237-242 (2015)
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COLD HARDINESS AND RANGE OF THE MYRIAPOD ANGAROZONIUM AMURENSE (POLYZONIIDAE, DIPLOPODA, ARTHROPODA) IN PERMAFROST ENVIRONMENTS

Daniil I. Berman1, Ekaterina N. Meshcheryakova1*
and Elena V. Mikhaljova2

1Laboratory of Biocenology, Institute of Biological Problems (North, Far East Branch), Russian Academy of Sciences, Magadan, Russia;
2Laboratory of Entomology, Institute of Biology and Pedology (Far East Branch), Russian Academy of Sciences, Vladivostok, Russia.
*Correspondence author email:
aborigen@ibpn.ru

Abstract

BACKGROUND: Angarozonium amurense (Gerstfeldt, 1859) is the only one out of more than a hundred diplopod species described in Siberia and the Far East that inhabits regions with solid permafrost. OBJECTIVE: To evaluate the cold hardiness of A. amurense that allows this species to inhabit permafrost regions. METHODS: The survival temperature thresholds and supercooling points (SCP) were measured. RESULTS: The temperature thresholds for adult animals' survival are 8.5C in summer and 27C in winter. Average SCP decreases from 7.70.3С in summer to 16.90.5С in winter. Water content decreases from 55.71.9% in summer to 49.41.6% in winter. CONCLUSION: The cold hardiness of A. amurense sets the record in this class of animals. It allows it to overwinter in the upper 15 centimeters layer of soil in most biotopes of the coldest permafrost regions in North Asia.

Keywords: Diplopoda, Angarozonium amurense, cold hardiness, permafrost.

 

 

 

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CryoLetters 36 (4), 243-251 (2015)
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THE COMBINED USE OF HONEY, GARLIC (ALLIUM SATIVUM L.) AND SKIMMED MILK AS AN EXTENDER FOR CHILLING SHEEP SEMEN

Rodrigo Jerez-Ebensperger1*, Lydia Gil1, Noelia Gonzalez1
 and Ignacio De Blas2

1Department of Animal Pathology, Obstetrics and Reproduction Area, Faculty of Veterinary Medicine;
2Department of Animal Pathology, Infectious Disease Area, Faculty of Veterinary Medicine, University of Zaragoza, Zaragoza, Spain.
*Corresponding author email:
rjerez@hotmail.com

Abstract

BACKGROUND: Sugars are the energetic source for sperm to maintain the metabolic process, and the antibiotics slow down sperm degradation.  OBJECTIVE: To study the effects of rosemary honey as energy source and cryoprotectant in combination with garlic as a natural antibiotic on the quality of ram spermatozoa upon cooling. MATERIALS AND METHODS: The ejaculates from three rams were evaluated at different times during cooling to determine its post-dilution quality. RESULTS: Glycerol and dimethylformamide in conjunction with honey and garlic significantly improve the survival of spermatozoa. CONCLUSION: The addition of honey and garlic reduces sperm deterioration when stored at 4C.

Keywords: Acridine orange; spermatozoa, ram, garlic, honey, OxyDNA, skim milk.

 

 

 

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CryoLetters 36 (4), 252-263 (2015)
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PHYSIOLOGICAL, CYTOLOGICAL AND BIOCHEMICAL STABILITY OF MEDICAGO SATIVA L. CELL CULTURE AFTER 27 YEARS OF CRYOGENIC STORAGE

Liudmila A. Volkova1, Valentina V. Urmantseva1,
Elena V. Popova1,2* and Alexander M. Nosov1,3

1Department of Cell Biology and Biotechnology, Timiryazev Institute of Plant Physiology of Russian Academy of Sciences, Botanicheskaya ul. 35, Moscow 127276, Russia.
2Gosling Research Institute for Plant Preservation, Department of Plant Agriculture, University of Guelph, Ontario N1G 2W1, Canada.
3Faculty of Biology, Lomonosov Moscow State University, Lenin Hills, 1, Moscow 119234, Russia.
*Corresponding author email:
epopova@uoguelph.ca

Abstract

BACKGROUND: The efficiency of long-term cryogenic storage to prevent somaclonal variations in plant cell cultures and retain their major cytogenetic and biochemical traits remains under debate. In particular, it is not clear how stress conditions associated with cryopreservation, such as low temperature, dehydration and toxic action of some cryoprotectants (DMSO in particular), affect post-storage regrowth and genetic integrity of cell samples. OBJECTIVE: We assessed growth, cytogenetic and biochemical characteristics of the peroxidase-producing strain of Medicago sativa L. cell culture recovered after 27 years of cryogenic storage as compared to the same culture before cryopreservation. MATERIALS AND METHODS: In 1984, M. sativa L. cell culture was cryopreserved using programmed freezing and 7% DMSO as a cryoprotectant. In 2011, after rewarming in a water bath at 40С for 90 s, cell culture was recovered and proliferated. Viability, growth profile, mitotic index, ploidy level, peroxidase activity and cell response to hypothermia and osmotic stress were compared between the recovered and the initial cell cultures using the records available from 1984. RESULTS: Viability of alfalfa cell culture after rewarming was below 20% but it increased to 80% by the 27th subculture cycle. Recovered culture showed higher mitotic activity and increased number of haploid and diploid cells compared to the initial cell line. Both peroxidase activity and response to abiotic stress in the recovered cell culture were similar to that of the initial culture. CONCLUSION: Cryopreservation by programmed freezing was effective at retaining the main characteristics of M. sativa undifferentiated cell culture after 27 years of storage. According to available data, this is longest period of successful cryopreservation of plant cell cultures reported so far. After storage, there was no evidence that DMSO had any detrimental effect on cell viability, growth or cytogenetics.

Keywords: alfalfa, cell culture, cryopreservation, peroxidase activity, cytogenetic stability, long-term storage, mitotic index, proliferation.

 

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CryoLetters 36 (4), 264-269 (2015)
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EFFECTS OF ORVUS ES PASTE ON THE MOTILITY AND VIABILITY OF YAK (BOS GRUNNIENS) EPIDIDYMAL AND EJACULATED SPERMATOZOA AFTER FREEZING AND THAWING

Megumi Shimazaki1, Rentsenkhand Sambuu2, Yoko Sato1,
Lanh Thi Kim Do1, Fuminori Tanihara1, Masayasu Taniguchi1
and Takeshige Otoi1*

1Department of Animal Reproduction, Joint Faculty of Veterinary Medicine, Yamaguchi University, Yamaguchi 753-8515, Japan
2Institute for Extension of Agricultural Advanced Technology, Ulaanbaatar, Mongolia
*Corresponding author email:
otoi@yamaguchi-u.ac.jp

Abstract

BACKGROUND: The addition of the detergent Orvus ES Paste (OEP) to semen freezing extenders has been observed to improve the post-thaw survival and longevity of spermatozoa from various species but has never been evaluated for yak spermatozoa. OBJECTIVE: This study evaluated the effects of OEP on the post-thaw motility and viability of epididymal and ejaculated yak spermatozoa.  MATERIALS AND METHODS: Semen samples were frozen and thawed in semen freezing extender supplemented with 0%, 0.375%, 0.75% or 1.5% OEP. The motility and viability of frozen-thawed spermatozoa were evaluated before and after 3 h of incubation. RESULTS: The addition of 0.75% OEP to the freezing extender significantly improved the mean motility and viability values of both the epididymal and ejaculated spermatozoa immediately after thawing, but the beneficial effects on motility disappeared after 3 h of incubation. CONCLUSION: Our findings indicate that the addition of 0.75% OEP is effective for the preservation of yak spermatozoa.

Keywords: Bos grunniens, cryopreservation, detergent, freezing extender, spermatozoa

 

 

 

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CryoLetters 36 (4), 270-277 (2015)
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RELATIONSHIP BETWEEN SUPERCOOLING CAPABILITY AND CRYOPROTECTANT CONTENT IN EGGS OF PARARCYPTERA MICROPTERA MERIDIONALIS (ORTHOPTERA: ACRYPTERIDAE)

Xiao-Rong Zhou#, Yan-Yan Li#, Na Li and Bao-Ping Pang*

Research Center for Grassland Entomology, Inner Mongolia Agricultural University, Hohhot, China.
#These co-authors contributed equally.
*Corresponding author email:
pangbp@imau.edu.cn

Abstract

BACKGROUND: Grasshoppers are major agricultural pests throughout the world. The egg stage is important for the low temperature resistance, and almost all grasshoppers overwinter in the egg stage. OBJECTIVE: To study the relationship between cold hardiness and cryoprotectant content in Pararcyptera microptera meridionalis eggs.  MATERIALS AND METHODS: The supercooling point (SCP) of the eggs was measured, along with the contents of water, fat, amino acids, low molecular sugars and polyols.  RESULTS: SCP, water content and glucose concentration decreased during egg development, whereas the contents of fat, trehalose, glycerol, inositol and sorbitol increased. SCP is negatively correlated with the concentrations of fat, trehalose, glycerol, inositol and sorbitol, but positively with water content and glucose concentration. Among low molecular weight sugars and polyols tested in eggs, trehalose concentration was highest, followed by glycerol. Although total content of free amino acids did not change much, of the tested 17 free amino acids in eggs, proline and glutamine had increased by 46.3% and 13.2%, respectively, and both showed a negative correlation with SCP. Stepwise regression analysis showed that proline, glycerol, trehalose and inositol contribute most to the SCP depression. Cold acclimation at 0C increased the contents of trehalose and glycerol, and decreased SCP. CONCLUSION: The increase of the supercooling capacity in P. microptera meridionalis eggs during development could be attributed mainly to proline, glycerol, trehalose and inositol. Cold acclimation enhances supercooling capacity via glycerol and trehalose.

Keywords: Pararcyptera microptera meridionalis, supercooling capacity, cold acclimation, low molecular weight carbohydrates, amino acids.

 

 

 

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CryoLetters 36 (4), 278-284 (2015)
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TEMPORARY STORAGE OF BOVINE SEMEN CRYOPRESERVED IN LIQUID NITROGEN ON DRY ICE AND REFREEZING OF FROZEN-THAWED SEMEN

A.M. Abdussamad1, M. Gauly2 and W. Holtz*

Department of Animal Science, Georg-August-University Goettingen,Albrecht-Thaer-Weg 3, 37075 Goettingen, Germany
1Current address: Department of Animal Science, Faculty of Agriculture, Bayero University, Kano, P.M.B. 3011, Kano State, Nigeria
2Current address: Faculty of Science and Technology, Free University of Bolzano, Universitaetsplatz 5, 39100 Bozen, Italy
*Corresponding author email:
wholtz@gwdg.de

Abstract

Two experiments were conducted.

The purpose of Experiment 1 was to investigate whether viability of bovine semen stored in liquid nitrogen (-196C) will be adversely affected by temporary exposure to dry ice (-79C). It was convincingly shown that post thaw-motility was not affected, regardless whether semen was thawed immediately or after being returned to liquid nitrogen. Shipping or temporary storage on dry ice, thus, is a viable option.

In Experiment 2, refreezing of frozen-thawed semen was attempted. The proportion of motile spermatozoa was reduced by a factor of ten to between 6.0% and 7.4%, regardless whether thawing occurred directly after removal from liquid nitrogen or after an interim period on dry ice. When semen was refrozen on dry ice before being returned to liquid nitrogen, motility rates were significantly improved (13.0% to 17.0%, P<0.05). In both experiments sperm cells that remained motile displayed vigorous forward movement and normal morphological appearance.

Keywords: Cryopreservation; Semen; Cattle; Liquid nitrogen; Dry ice

 

 

 

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CryoLetters 36 (4), 285-288 (2015)
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DETERMINATION OF CONVECTIVE HEAT TRANSFER COEFFICIENT AT THE OUTER SURFACE OF A CRYOVIAL BEING PLUNGED INTO LIQUID NITROGEN

Tao Wang1, Gang Zhao1,2*, Heyu Tang1 and Zhendong Jiang1

1Centre for Biomedical Engineering, Department of Electronic Science & Technology, University of Science and Technology of China, Hefei;
2Anhui Provincial Engineering Technology Research Center for Biopreservation and Artificial Organs, Hefei, China.
*Corresponding author email:
zhaog@ustc.edu.cn

Abstract

BACKGROUND: Cell survival upon cryopreservation is affected by the cooling rate. However, it is difficult to model the heat transfer process or to predict the cooling curve of a cryoprotective agent (CPA) solution due to the uncertainty of its convective heat transfer coefficient (h)OBJECTIVE: To measure the h and to better understand the heat transfer process of cryovials filled with CPA solution being plunged in liquid nitrogen.  MATERIALS AND METHODS: The temperatures at three locations of the CPA solution in a cryovial were measured. Different h values were selected after the cooling process was modeled as natural convection heat transfer, the film boiling and the nucleate boiling, respectively. And the temperatures of the selected points are simulated based on the selected h values. h was determined when the simulated temperature best fitted the experimental temperature. RESULTS: When the experimental results were best fitted, according to natural convection heat transfer model, h1 = 120 W/(m2K) while due to film boiling and nucleate boiling regimes hf = 5 W/(m2K) followed by hn = 245 W/(m2K). These values were verified by the differential cooling rates at the three locations of a cryovial. CONCLUSION: The heat transfer process during cooling in liquid nitrogen is better modeled as film boiling followed by nucleate boiling.

Keywords: Cryoprotective agent, Cryovial, Convective heat transfer coefficient, Cooling rate, Finite element method

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