Even though lake sediments are globally important organic carbon (OC) sinks, the controls on long-term OC storage in these sediments are unclear. Using a multiproxy approach, we investigate changes in diatom, green algae, and vascular plant biomolecules in sedimentary records from the past centuries across five temperate lakes with different trophic histories. Despite past increases in the input and burial of OC in sediments of eutrophic lakes, biomolecule quantities in sediments of all lakes are primarily controlled by postburial microbial degradation over the time scales studied. We, moreover, observe major differences in biomolecule degradation patterns across diatoms, green algae, and vascular plants. Degradation rates of labile diatom DNA exceed those of chemically more resistant diatom lipids, suggesting that chemical reactivity mainly controls diatom biomolecule degradation rates in the lakes studied. By contrast, degradation rates of green algal and vascular plant DNA are significantly lower than those of diatom DNA, and in a similar range as corresponding, much less reactive lipid biomarkers and structural macromolecules, including lignin. We propose that physical shielding by degradation-resistant cell wall components, such as algaenan in green algae and lignin in vascular plants, contributes to the long-term preservation of labile biomolecules in both groups and significantly influences the long-term burial of OC in lake sediments.

Lactulose is commonly used in pharmacy for constipation and hepatic encephalopathy treatment. The prebiotic effect of lactulose is also often mentioned. However, its cryoprotective effect in combination with lecithin on the main representatives of probiotics has not been tested yet. The 12 taxa of bifidobacteria and Lactobacillaceae members were used for the purpose. These were mixed in a ratio of 1:1 with lactulose + lecithin (finally 5.0% and 1.25%, respectively; LL). The 25% glycerol (G+) solution and cultures themselves were applied as positive and negative controls, respectively. Bacterial suspensions were stored at a mild freezing temperature (-20°C) until the end of the experiment (210th day). The LL solution had a comparable (insignificant difference at the P-value = 0.05) cryoprotective effect as the positive control in five of six bifidobacteria and in three of six representatives of Lactobacillaceae. The better cryoprotective effect was revealed in other Lactobacillaceae. At the end of the experiment, the generally accepted therapeutic minimum (>107 Colony Forming Units/mL) was determined in LL solution in five bifidobacteria and four Lactobacillaceae strains. The presented results improve knowledge about long-term mild cryopreservation of the most commonly used probiotics and could contribute to developing new forms of (nutri)synbiotics.

Long-term preservation of sperm, oocytes, and gonadal tissues at ambient temperatures has the potential to lower the costs and simplify biobanking in human reproductive medicine, as well as for the management of animal populations. Over the past decades, different dehydration protocols and long-term storage solutions at nonfreezing temperatures have been explored, mainly for mammalian sperm cells. Oocytes and gonadal tissues are more challenging to dehydrate so little to no progress have been made. Currently, the detrimental effects of the drying process itself are better characterized than the impact of long-term storage at nonfreezing temperatures. While structural and functional properties of germ cells can be preserved after dehydration, a long list of damages and stresses in nuclei, organelles, and cytoplasmic membranes have been reported and sometimes mitigated. Characterizing those damages and better understanding the response of germ cells and tissues to the stress of dehydration is fundamental. It will contribute to the development of optimal protocols while proving the safety of alternative storage options for fertility preservation. The objective of this review is to (1) document the types of damages and stress responses, as well as their mitigation in cells dried with different techniques, and (2) propose new research directions.

Preservation of evolved biological structure and function in robust engineering materials is of interest for storage of biological samples before diagnosis and development of vaccines, sensors, and enzymatic reactors and has the potential to avoid cryopreservation and its associated cold-chain issues. Here, we demonstrate that \"freezing cells in amorphous silica\" is a powerful technique for long-term preservation of whole mammalian cell proteomic structure and function at room temperature. Biomimetic silicification employs the crowded protein microenvironment of mammalian cells as a catalytic framework to proximally transform monomeric silicic acid into silicates forming a nanoscopic silica shell over all biomolecular interfaces. Silicification followed by dehydration preserves and passivates proteomic information within a nanoscale thin silica coating that exhibits size selective permeability (<3.6 nm), preventing protein leaching and protease degradation of cellular contents, while providing access of small molecular constituents for cellular enzymatic reaction. Exposure of dehydrated silicified cells to mild etchant or prolonged hydrolysis removes the silica, completely rerevealing biomolecular components and restoring their accessibility and functionality.

Understanding the effects of long-term exposure to space environment is paramount to maintaining the safety, health of astronauts. The physical dosimeters currently used on the space station cannot be used to assess the physiological effects of radiation. Moreover, some developed biological methods are time-consuming and passive and cannot be used for active and real-time detection of the physiological effects of radiation in space environment. Here, the SOS promoter: recA-eGFP genetic engineering bacteria was constructed and characterized, and DNA damage effects of some chemical reagents and radiation were evaluated. The results indicated the constructed engineering bacteria can distinguish DNA damage reagents from non-damage reagents and have a good dose-fluorescence effect against Co-60 radiation with the detection limit of 0.64 Gy; in order to overcome the restriction of long-term preservation of bacteria in space environment, the bacteria were freeze-dried, and the protectants were optimized, the storage time of bacteria under dry conditions was explored by accelerated storage experiment. Finally, a microfluidic chip was designed and fabricated for freeze-drying genetic engineering bacteria recovery, culture, and analysis in space environment. This study can provide support for the establishment of on-orbit radiation damage risk monitoring and early warning and can provide basic data for maintaining the health and performance of astronauts on long-term space flight missions. Moreover, the technique developed herein has a great potential to be used as a powerful tool for efficiently screening various radioactive substance, toxic chemicals, drugs, etc. KEY POINTS: • The SOS promoter: recA-eGFP genetic engineering bacteria was successfully constructed, which can distinguish DNA damage reagents from non-damage reagents and possess a good dose-effect relationship against Co-60 radiation. • The bacteria were freeze-dried to overcome the restriction of long-term preservation of bacteria in space environment, and protectants were optimized, and the survival rate of freeze-dried engineering bacteria can be predicted based on the results of accelerated storage experiment. • Microfluidic chip-based culture platform was successfully designed, fabricated, and used for freeze-drying genetic engineering bacteria recovery, culture, and analysis.

Amino acid arrays comprising bioluminescent amino acid auxotrophic Escherichia coli are effective systems to quantitatively determine multiple amino acids. However, there is a need to develop a method for convenient long-term preservation of the array to enable its practical applications. Here, we reported a potential strategy to efficiently maintain cell viability within the portable array. The method involves immobilization of cells within agarose gel supplemented with an appropriate cryoprotectant in individual wells of a 96-well plate, followed by storage under freezing conditions. Six cryoprotectants, namely dimethyl sulfoxide, glycerol, ethylene glycol, polyethylene glycol, sucrose, and trehalose, were tested in the methionine (Met) auxotroph-based array. Carbohydrate-type cryoprotectants (glycerol, sucrose, and trehalose) efficiently preserved the linearity of determination of Met concentration. In particular, the array with 5% trehalose exhibited the best performance. The Met array with 5% trehalose could determine Met concentration with high linearity (R2 value = approximately 0.99) even after storage at -20 °C for up to 3 months. The clinical utilities of the Met and Leu array, preserved at -20 °C for 3 months, were also verified by successfully quantifying Met and Leu in spiked blood serum samples for the diagnosis of the corresponding metabolic diseases. This long-term preservation protocol enables the development of a ready-to-use bioluminescent E. coli-based amino acid array to quantify multiple amino acids and can replace the currently used laborious analytical methods.

In this study, changes in the microbiota of Japanese Black beef carcasses, which are expected to be transported for a long time in chilled temperatures, were investigated. Three Japanese Black beef samples (carcasses A, B, and C) immediately after slaughter were stored at 0 °C for 15 weeks under aerobic and vacuum conditions. The initial bacterial counts were 50 CFU/g for carcass A and less than the reliable quantitative detection limit for carcasses B and C. Under aerobic storage conditions, the bacterial count increased to 8.0 log CFU/g or higher, which is a measure of putrefaction, at 6-9 weeks. Under anaerobic storage conditions, the bacterial counts of carcasses A and C reached 3.5-6.5 log CFU/g, but carcass B showed no bacterial growth during the 15-week storage period. The predominant group was Pseudomonas spp. under aerobic conditions and Serratia spp. under anaerobic conditions. To the best of our knowledge, there is no previous study investigating the transition of microbiota when Japanese Black beef is stored at low temperatures for a long period of time, and the results of this study are considered very important findings for the expansion of international trade of Japanese Beef in the future.

The cryopreservation of dormant buds can be a feasible method for preserving germplasm of cold-tolerant woody plants. In the present study, we evaluated the effects of pre-desiccation, thawing method, and the rehydration of bud sections on the post-cryopreservation recovery of dormant blackcurrant buds in vitro. The estimated recovery of small- and medium-sized buds was 80.1 and 62.7% respectively for desiccated buds and 67.8 and 72.3% respectively for non-desiccated buds. The pre-desiccation of bud sections enhanced the number of the shoots regenerated from vegetative buds (2.3 vs. 4.7). The estimated recovery of fast-thawed buds was better after 14-day than after 7-day rehydration (85 vs. 59%). In slowly thawed buds the difference between 14-day and 7-day rehydration was not significant (73 vs. 62%). The estimated recovery of vegetative and flower buds was 77.7 and 41.1% respectively after 7-day rehydration, and 95.2 and 43.6% respectively after a 14-day rehydration period. The rehydration of bud sections was not necessary for the in vitro recovery of non-desiccated, fast-thawed buds. Of the 23 blackcurrant cultivars cryopreserved using non-desiccated dormant buds collected from a greenhouse, the estimated recovery of 22 cultivars ranged between 42 and 90%.

Thermophilic fungi have several biotechnological and industrial applications such as thermostable enzyme production, biodegradation, and tobacco processing, etc. Thermophilic fungi cannot survive at temperatures below 20 °C. Owing to their inability to grow at low temperatures, they are not stable, so stocking is very difficult. Although a large number of different storage methods are available and described, no method can be universally applied to all fungi. Thermophilic fungi present \"heat-loving\" characteristics, and therefore a new challenge for its preservation and there is no universal protocol for the preservation of thermophilic fungi. The aim of this study was to evaluate the viability, contamination and stability of thermophilic fungi stored under different preservation methods. In this work, 25 thermophilic fungal isolates of species Thermomyces thermophilus, Rhizomucor pusillus, Trichocladium griseum, Melanocarpus albomyces, Malbranchea cinnamomea, Thermothelomyces thermophilus, Thermothelomyces hinnuleus,Thermothielavioidesterrestris, Mycothermus thermophilus, Humicola insolens maintained constant sub-culturing at room temperature, +4 °C and +20 °C, lyophilization at +4 °C, freezing at -20 °C, freezing block at -20 °C and a new technique liquid preservation at room temperature for the periods ranging 5 years. We evaluated the effect of preservation methods by sub-culturing onto either sabouraud dextrose agar (SDA) or yeast extract soluble starch agar (YpSs) on growth, production and viability of spores and macro- and micromorphology. In this study, preservation methods for thermophilic fungi were investigated extensively for the first time and it is clearly shown that freezing block at -20 °C method and lyophilization were better methods for long-term preservation up to 5 years.

Agave tequilana Weber cultivar \'Chato\' represents an important genetic supply of wild severely in decline populations of \'Chato\' for breeding and transformation programs. In this work, the indirect somatic embryogenesis and cryopreservation of Somatic Embryos (SEs) were investigated using the \'Chato\' cultivar as a study case.
METHODS: Embryogenic calli were induced by the cultivation of 1 cm of young leaves from in vitro plants on MS semisolid medium supplemented with 24.84, 33.13, 41.41, 49.69, and 57.98 μM 4-amino-3,5,6-trichloro-2- pyridinecarboxylic acid (picloram) in combination with 2.21, 3.32, and 4.43 μM 6-benzylaminopurine (BAP). The origin and structure of formed SEs were verified by histological analysis. Cryopreservation studies of SEs were performed following the V-cryoplate technique and using for dehydration two vitrification solutions (PVS2 and PVS3).
RESULTS: The highest average (52.43 ± 5.74) of produced SEs and the Embryo Forming Capacity (estimated index 52.43) were obtained using 49.69 µM picloram and 3.32 µM BAP in the culture medium. The highest post-cryopreservation regrowth (83%) and plant conversion rate (around 70%) were achieved with PVS2 at 0 °C for 15 min.
CONCLUSIONS: Our work provides new advances about somatic embryogenesis in Agave and reports the first results on cryopreservation of SEs of this species.