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Total mercury (THg) analysis of fish tissue requires the samples to be frozen for long-term storage or analyzed immediately after the fish sample has been obtained. Such rigid field sampling protocols do not account for the logistical challenges that can occur if sampling is conducted in remote areas or collaboratively via community-based monitoring initiatives. Here we present a study that examines the loss of THg from Walleye tissue that has thawed for 3 and 5 days, mimicking potential thaw during transport from remote locations. THg concentrations were not significantly influenced (Kruskal-Wallace, p-value > 0.05) by the thaw and refreeze events. The mean THg concentration per day group was 0.67 ug/g, SD = 0.03, 0.65 ug/g, SD = 0.01, and 0.72 ug/g, SD = 0.01 for the control (no thaw), 3-day thaw and 5-day thaw, respectively. Results from this study suggest the analytical integrity of samples may be maintained if freeze-thaw situations arise when conducting community-based mercury monitoring programs., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Purpose: We aimed to evaluate the effect of extending the culture of cleavage-stage embryos to the blastocyst stage in vitrified-warmed cycles on pregnancy outcomes., Methods: This is a retrospectively designed pilot study of a single center. All patients who applied for freeze-all cycle procedures during in vitro fertilization treatment were included in the study. Patients were classified into three subgroups. The embryos obtained were frozen at the cleavage or blastocyst stage. After a warming process, the cleavage-stage embryos were divided into two subgroups: the first group of embryos was transferred (vitrification day 3-embryo transfer (ET) day 3 (D3T3)) on the warming day; for the second group, the embryo culture was extended to the blastocyst stage (vitrification day 3-ET day 5 (after the extension of the embryo culture to the blastocyst stage), (D3T5)). Frozen blastocyst-stage embryos were transferred after warming (vitrification day 5-ET day 5 (D5T5)). Hormone replacement treatment was the only endometrial preparation regimen given during the embryo transfer cycle. The main outcome of the study was live birth rates. The clinical pregnancy rate and positive pregnancy test rate were determined as the secondary outcomes of the study., Results: The study included a total of 194 patients. The positive pregnancy test rates (PPR) and clinical pregnancy rates (CPR) of the D3T3, D3T5, and D5T5 groups were 14.0% and 59.2%; 43.8% and 9.3%; and 56.3% and 39.6%, respectively (p < 0.001 and p < 0.001). The live birth rates (LBR) of patients in the D3T3, D3T5, and D5T5 groups were 7.0%, 44.7%, and 27.1%, respectively (p < 0.001). In subgroup analysis of patients with a poor number of 2PN embryos (defined as having <  = 4 2PN embryos), the D3T5 group had significantly higher PPR (10.7%, 60.6%, 42.4%; p < 0.001), CPR (7.1%, 57.6%, 39.4%; p < 0.001), and LBR (3.6%, 39.4%, 21.2%; p: 0.001)., Conclusion: Extending the culture after warming to the blastocyst stage may be a better alternative than a cleavage-stage embryo transfer., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Shallow thaw (thermokarst) lakes abundant in regions of permafrost-affected peatlands represent important sources of carbon dioxide and methane emission to the atmosphere, however the quantitative parameters of phytoplankton communities which control the C cycle in these lakes remain poorly known. This is especially true considering the roles of permafrost, hydrochemical composition of lakes, lake sizes and season as major governing factors on phytoplankton abundance and biodiversity. In this work, we quantified phytoplankton characteristics of 27 thermokarst lakes (sizes ranging from 115 m2 to 1.24 km 2 ) sampled in spring, summer and autumn across a permafrost gradient (isolated, sporadic, discontinuous and continuous zone) in the Western Siberia Lowland (WSL). The biodiversity indices were highest during all seasons in lakes of the continuous permafrost zone and rather similar in lakes of isolated, sporadic and discontinuous permafrost zone. Considering all seasons and permafrost zones, the biomass and cell number of phytoplankton correlated with Dissolved Organic Carbon (DOC), phosphate, and some metal micro-nutrients (Ni, Zn). The strongest correlations were observed for Cyanophycea during summer, with pH, Ni, Cu, Zn, Sr, Ba (cell number) and Cu, Zn, Ba (biomass), and during autumn, with DOC, K, Cr, Cu, Zn, Ba, Cd, Pb (biomass). Using a substituting space for time approach for climate warming and permafrost thaw and suggesting a shift in permafrost boundaries northward, we foresee an increase in cell number and biomass in continuous permafrost zone in spring and summer, and a decrease in phytoplankton abundance in the discontinuous and sporadic permafrost zones. The biodiversity of phytoplankton in the continuous permafrost zone might decrease whereas in other zones, it may not exhibit any sizably change. However, in case of strong deepening of the active layer down to underlaying mineral horizons, and the release of some limiting nutrients (Si, P) due to enhanced connectivity of the lake with groundwater, the share of cyanobacteria and diatoms may increase., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier B.V. All rights reserved.)

The effects of climate change on water discharge in rivers in cold climates were investigated. To quantify the impacts of air temperature rises on the promotion of snowmelt and associated acceleration of a rise in the level of a river, 10 rivers on Hokkaido, northwestern Japan were chosen. Available data of daily water discharge for more than half a century by the Ministry of Land, Infrastructure, Transport and Tourism (MLIT) were correlated with air temperatures observed at the 8 weather stations operated by the Japan Meteorological Agency (JMA). The analyses show that annual average air temperatures have risen at all 8 sites on Hokkaido for the period from 1954 to 2018. The trends for the 8 sites show a range from 0.119 °C/decade to 0.250 °C/decade with an average of 0.191 °C/decade. Annual snowfall has increased at Sutsu. The trend over the period from 1954 to 2018 was 45.2 cm/decade. Otherwise, annual snowfall has not changed significantly in the period (1954-2018). Accelerations of the rise in the level of the river induced by air temperature rises have been observed in the Teshio River, Ishikari River, Saru River, and Mu River. A delay in the rise in the level of the river caused by an increase in snowfall and the weak warming trend from January to April has been observed in the Shiribetsu River. Although air temperatures have risen year after year, a significant change in annual pattern of daily flow has not been observed in the Syokotsu River, Yubetsu River, Tokoro River, and Abashiri River located in eastern Hokkaido. This can be induced by the weak warming trends in April which may be due to the cold current of the sea of Okhotok., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Several studies have investigated the effect of repeated freeze-thaw (F/T) cycles on RNA detection for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). However, no data are available regarding the effect of repeated F/T cycles on SARS-CoV-2 antibody detection in serum. We investigated the effect of multiple F/T cycles on anti-SARS-CoV-2 IgG detection using an ELISA test targeting the nucleocapsid antibodies. Ten positive and 1 negative SARS-CoV-2 IgG sera from 11 participants, in replicates of 5, were subjected to a total of 16 F/T cycles and stored at 4 °C until tested by ELISA. Statistical analysis was performed to test for F/T cycle effect. None of the 10 positive sera became negative after 16 F/T cycles. There was no significant difference in the OD average reading between the first and last F/T cycles, except for one serum with a minimal decline in the OD. The random effect linear regression of log (OD) on the number of cycles showed no significant trend, with a slope consistent with zero (B=-0.0001; 95 % CI -0.0008; 0.0006; P -value=0.781). These results suggest that multiple F/T cycles had no effect on the ability of the ELISA assay to detect SARS-CoV-2 IgG antibodies.

The physical and chemical consequences of massive ground ice (wedges) melt upon permafrost thaw is one of the central issues of environmental research linked to climate warming in the Arctic. Little is known about the chemical properties of dispersed ground ice abundant throughout permafrost peatlands that can easily melt with increasing active layer thickness (ALT). This is especially pertinent in continental lowlands, that account for sizeable areas of the Arctic, and contain high amount of organic carbon in both solid (peat) and liquid (porewater) phases. Here we studied 8 peat cores (0-130 cm depth)-comprised of porewater from the active layer (0-45 cm) as well as ice dispersed in frozen peat (40-130 cm)-across a latitudinal profile of Western Siberia Lowland (WSL) extending from discontinuous into continuous permafrost zones. Dissolved Organic Carbon (DOC), alkali and alkaline-earth metals (Ca, Mg, Sr, Ba, Li, Rb, Cs), sulfate, phosphorus, some trace elements (Al, Fe, Mn, Zn, Ni, Co, V, As, Y, REE, Zr, Hf, U) were sizably [more than 3 times] enriched in peat ice compared to peat porewaters from the active layer. In most sampled cores, there was a local maximum of strong enrichment (up to factors between 14 and 58) in DOC, P, Ca, Mg, Mn, Fe, Sr, As located 30-50 cm below the active layer. This maximum likely occurred due to solute concentration during full freezing of the soil column during winter. There was a sizable correlation between DOC, Al, Fe and other major and trace element concentrations that suggests strong control of organic complexes and organo-mineral (Al, Fe) colloids on element migration throughout the peat profile. The pool of C, major cations and trace metals in peat ice (40-130 cm) was approximately 3-55 times higher than the pool of these elements in porewaters from the active layer (0-40 cm). A 1-m increase of the ALT over the next 100 years is capable of mobilizing 58 ± 38 Tg of DOC from soil ice into the rivers and lakes of the WSL latitudinal belt (63-67 °N). This fast lateral export of C (3.7 ± 2.7 t C km -2 y -1 ) may double current C yields in WSL rivers (3.4 ± 1.3 t C km -2 y -1 ). A strong increase (150-200%) in riverine export of Zn, P and Cs may also occur while other micronutrients (Fe, Ni, Co, Ba, Mo, Rb) and toxicants (Cd, As, Al) may be affected to a lesser degree (20-30% increase). We propose a global peat ice inventory in permafrost regions is essential for assessing the consequences of permafrost thaw on surface aquatic systems., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Objective: To assess the recovery of thawed blastocysts submitted to quarter laser assisted hatching and examine potential correlations between the procedure and pregnancy rates., Methods: This cross-sectional study included only single-blastocyst transfers performed from July 2017 to December 2018. A total of 765 blastocysts were thawed and immediately submitted to quarter laser assisted hatching in the zona pellucida; they were subsequently incubated for three hours until transfer time, at which time they were examined for collapse or expansion; expanded blastocysts were further evaluated for herniation. The Chi-square test was used in statistical analysis., Results: 627 blastocysts expanded (81.9%) and yielded a pregnancy rate of 40% (251/627). 138 blastocysts collapsed after thawing (18.0%) and yielded a pregnancy rate of 25.4% (35/138) (p=0.001). Additional analysis of the subgroup of expanded blastocysts revealed that the 385 herniated blastocysts (61.4%) yielded a pregnancy rate of 43.9% (169/385). The remaining 242 non-herniated blastocysts (38.6%) yielded a pregnancy rate of 33.9% (82/242) (p=0.013). Statistical significance was attributed to events with a p<0.05., Conclusion: Quarter laser assisted hatching is a safe, valid, and relatively easy-to-use procedure for thawed blastocysts. Blastocysts that expanded and herniated after quarter laser assisted hatching presented statistically superior results.

Arctic permafrost soils contain large amounts of organic carbon and the pollutant mercury (Hg). Arctic warming and associated changes in hydrology, biogeochemistry and ecology risk mobilizing soil Hg to rivers and to the Arctic Ocean, yet little is known about the quantity, timing and mechanisms involved. Here we investigate seasonal particulate Hg (PHg) and organic carbon (POC) export in 32 small and medium rivers across a 1700 km latitudinal permafrost transect of the western Siberian Lowland. The PHg concentrations in suspended matter increased with decreasing watershed size. This underlines the significance of POC-rich small streams and wetlands in PHg export from watersheds. Maximum PHg concentrations and export fluxes were located in rivers at the beginning of permafrost zone (sporadic permafrost). We suggest this reflects enhanced Hg mobilization at the permafrost boundary, due to maximal depth of the thawed peat layer. Both the thickness of the active (unfrozen) peat layer and PHg run-off progressively move to the north during the summer and fall seasons, thus leading to maximal PHg export at the sporadic to discontinuous permafrost zone. The discharge-weighed PHg:POC ratio in western Siberian rivers (2.7 ± 0.5 μg Hg: g C) extrapolated to the whole Ob River basin yields a PHg flux of 1.5 ± 0.3 Mg y -1 , consistent with previous estimates. For current climate warming and permafrost thaw scenarios in western Siberia, we predict that a northward shift of permafrost boundaries and increase of active layer depth may enhance the PHg export by small rivers to the Arctic Ocean by a factor of two over the next 10-50 years., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

This study evaluated the ability of potassium ferrate(VI) and freeze-thaw to stabilise and dewater primary sludge. Potassium ferrate(VI) additions of 0.5 and 5.0 g/L were used as a pre-treatment prior to freeze-thaw. Samples were frozen at -10, -20 and -30 °C, and were kept frozen for 1, 8 and 15 days. The samples were subsequently thawed at room temperature in a setup which allowed meltwater to be separated from the sludge cake via gravity drainage. The meltwater was characterised in terms of fecal coliform, soluble chemical oxygen demand (COD), soluble proteins, soluble carbohydrates, pH and turbidity. The sludge cake was characterised in terms of fecal coliform, total solids (TS) and volatile solids (VS). Freeze-thaw with gravity meltwater drainage reduced the sludge volume by up to 79%. After being frozen for only 1 day, the concentrations of fecal coliform in many of the primary sludge samples were reduced to <1000 MPN/g dry solids (DS), representing >3-log inactivation in some cases. However, pre-treatment of the primary sludge with ≤5.0 g/L potassium ferrate(VI) resulted in significant increases in soluble proteins, soluble carbohydrates, and sCOD, and reduced the effectiveness of stand-alone freeze-thaw. Follow-up experiments using higher doses ranging from 5.1 to 24.9 g/L of potassium ferrate(VI) demonstrated that >5-log inactivation of fecal coliform in raw primary sludge can be achieved within 15 min using 15 g/L of potassium ferrate(VI), and the resulting concentration of fecal coliform in the sludge was 1023 MPN/g DS. Pre-treatment with 22.0 g/L of potassium ferrate(VI), followed by freeze-thaw, with only 3 days frozen, reduced the concentration of fecal coliform to below the detection limit in the meltwater and the sludge cake. This demonstrates that potassium ferrate(VI) and freeze-thaw offers the flexibility to adjust the ferrate(VI) dose to meet treatment requirements for land application, and can be used as a stand-alone sludge treatment technology for primary sludge that achieves both treatment and dewatering., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Objective: This study aimed to investigate and compare the pregnancy and live birth rates in IVF cycles of frozen-thawed embryo transfers and fresh embryo transfers in a group of women with a high risk of Ovarian hyperstimulation syndrome (OHSS)., Material and Methods: The study group consisted of 254 women with a high level of response to controlled ovarian hyperstimulation. The patients who received fresh cycle embryo transfers with calcium infusions are referred to as the Fresh Ca+ group, and those without the calcium therapy are called the Fresh Ca- group; and we used correspondingly similar terminology for the Frozen group., Results: We observed no statistically significant differences between the cycles of fresh and frozen-thawed embryo transfers in patients with a high risk of OHSS in terms of implantation, clinical pregnancy, and live birth rates. Furthermore, these implantation, clinical pregnancy and live birth rates were not different in the cycles with or without calcium treatment. There was no statistical difference in the OHSS rates between the fresh and frozen-thawed cycles; although, the OHSS rates were less in the two calcium infusion groups (Fresh Ca+ and Frozen-thawed Ca+) than in the without-calcium group. There was no OHSS development in the subjects of the Frozen-thawed Ca+ group., Conclusion: Our study results suggest that fresh and frozen-thawed embryo transfers have similar IVF results in patients with a high risk of OHSS. Calcium infusion is beneficial in preventing OHSS without altering pregnancy rates. Both IVF protocols with calcium infusion can safely be applied in high-responder patients without lowering success rates.

Aim: We aim to assess the effect of the state of T cells before cryopreservation on the post-thaw proliferative capacity, phenotype and functional response., Methods: Peripheral blood mononuclear cells (PBMCs) were isolated from a hepatocellular carcinoma (HCC) patient, and the T cells were frozen during cell culture according to our experimental design. After a period of re-culture, the proliferative capacity of the cryopreserved cells, the expression of T cell surface markers and the secretion of IFN-γ and IL-10 were assayed., Results: There was >90% cell viability after thaw in every group. Lymphocytes cryopreserved at day 4, 8 or 12 during the cell culture were allowed to recover for 24 h, whereas lymphocytes cryopreserved while freshly isolated were allowed to recover for 72 h. After the period of re-culture, cryopreservation at day 4, 8 or 12 during T cell culture was not found to alter the T cell subpopulation. The proportions of NKT and Treg cells were unchanged when cells were cryopreserved at day 12 during T cell culture. IFN-γ secretion was not impacted by cryopreservation, and IL-10 secretion was significantly decreased when cells were cryopreserved at day 8 or 12 during T cell culture., Conclusion: The state of T cells before cryopreservation has effects on the post-thaw proliferation capacity, the phenotype and the secretion of IFN-γ and IL-10. Cryopreservation of lymphocytes at day 8 or 12 during the cell culture may be the best choice for T cell immunotherapy., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Rana sylvatica, known as the wood frog, can survive extremely cold temperatures during winter by undergoing full-body freezing, where it tolerates freezing of 65-70% of its total body water. During freezing, cellular dehydration decreases damage to the cell by preventing ice crystallization. Challenged with many stresses, these animals are forced to develop physiological adaptations to osmoregulation and osmoprotection that are necessary to ensure their survival. The purpose of this study was to elucidate a potential mechanism by which the transcription factor, NFAT5, regulates the expression of three osmoregulatory proteins (aldose reductase, SMIT, and BGT-1). These three proteins control cellular concentrations of the organic osmolytes: betaine (BGT-1), myo-inositol (SMIT), and sorbitol (aldose reductase). We studied this mechanism during the freeze-thaw stress in R. sylvatica liver, kidney, and skeletal muscle. Protein expression of BGT-1, SMIT, aldose reductase, and NFAT5 were examined using immunoblotting. We identified that the NFAT5 pathway facilitated osmoregulation in a tissue-specific manner during freezing. In skeletal muscle, we demonstrated that NFAT5 upregulation in thawing led to increases in the protein levels of BGT-1. In liver, NFAT5 was upregulated during freezing, along with aldose reductase. Furthermore, neither of these patterns of expression were observed in kidney as none of these four proteins showed differential expression during freezing or thawing. Therefore, the NFAT5 osmoregulatory pathway appears to be tissue-specific. Our novel findings on a mechanism of osmoregulation in R. sylvatica highlight the importance of studying naturally stress-tolerant animals to identify novel pro-survival pathways., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Rapid Arctic warming is expected to increase global greenhouse gas concentrations as permafrost thaw exposes immense stores of frozen carbon (C) to microbial decomposition. Permafrost thaw also stimulates plant growth, which could offset C loss. Using data from 7 years of experimental Air and Soil warming in moist acidic tundra, we show that Soil warming had a much stronger effect on CO 2 flux than Air warming. Soil warming caused rapid permafrost thaw and increased ecosystem respiration (R eco ), gross primary productivity (GPP), and net summer CO 2 storage (NEE). Over 7 years R eco , GPP, and NEE also increased in Control (i.e., ambient plots), but this change could be explained by slow thaw in Control areas. In the initial stages of thaw, R eco , GPP, and NEE increased linearly with thaw across all treatments, despite different rates of thaw. As thaw in Soil warming continued to increase linearly, ground surface subsidence created saturated microsites and suppressed R eco , GPP, and NEE. However R eco and GPP remained high in areas with large Eriophorum vaginatum biomass. In general NEE increased with thaw, but was more strongly correlated with plant biomass than thaw, indicating that higher R eco in deeply thawed areas during summer months was balanced by GPP. Summer CO 2 flux across treatments fit a single quadratic relationship that captured the functional response of CO 2 flux to thaw, water table depth, and plant biomass. These results demonstrate the importance of indirect thaw effects on CO 2 flux: plant growth and water table dynamics. Nonsummer R eco models estimated that the area was an annual CO 2 source during all years of observation. Nonsummer CO 2 loss in warmer, more deeply thawed soils exceeded the increases in summer GPP, and thawed tundra was a net annual CO 2 source., (© 2017 John Wiley & Sons Ltd.)

Objective: The aim of this study was to perform a comparative investigation of several different thawing protocols and to determine an appropriate protocol for thawing whole bovine frozen ovaries., Study Design: Bovine ovaries were slowly frozen and then thawed by applying different protocols. Ultrastructural change, follicle viability, and the hormone levels of culture supernatant were measured., Results: The percentage of morphologically normal primordial follicles and the hormone levels of culture supernatant in group D (two-step, thawing in water at 39°C) were significantly higher than those in any other group. Moreover, the ultrastructural alteration of oocyte in group D (two-step, thawing in water at 39°C) was slighter than those in any other group., Conclusions: The two-step protocol involving short-term exposure to water at a moderately high temperature (39°C) proved to be a suitable for thawing bovine whole ovaries., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)