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Bacterial recovery from ancient glacial ice.

Ice that forms the bottom 18 m of a 308 m ice core drilled from the Guliya ice cap on the Qinghan-Tibetan plateau in Western China is over 750000 years old and is the oldest glacial ice known to date. Fourteen bacterial isolates have been recovered from samples of this ice from approximately 296 m below the surface (mbs). Based on 16S rDNA sequences, these are members of the alpha- and beta-proteobacterial, actinobacterial and low-G + C Gram-positive bacterial lineages. 16S rDNA molecules have also been amplified directly, cloned and sequenced from the ice-core melt water. These originated from Pseudomonas and Acinetobacter gamma-proteobacterial species. These results demonstrate that bacteria can be recovered from water ice that has frozen for time periods relevant to biological survival through terrestrial ice ages or during interplanetary transport.

Bacteria↗

Antifreeze protein modulates cell survival during cryopreservation: mediation through influence on ice crystal growth.

Antifreeze proteins (AFPs) are extremely efficient at inhibiting ice recrystallization in frozen solutions. Knight and Duman [Knight, C. A. & Duman, J. G. (1986) Cryobiology 23, 256-263] have proposed that this may be an important function of the proteins in freeze-tolerant organisms. We have tested this proposal in vitro by characterizing the influence of AFP on the recovery of cryopreserved cells, which often can survive cooling and yet subsequently be damaged by ice crystal growth during warming. Relatively low concentrations (e.g., 5-150 micrograms/ml) of winter flounder (Pseudopleuronectes americanus) AFP enhance survival of red blood cells cryopreserved in hydroxyethyl starch solutions. This effect is most apparent in samples warmed at suboptimal rates, i.e., where ice recrystallization would be exaggerated. Cryomicroscopy demonstrates that AFP inhibits ice recrystallization in the extracellular regions during the latter stages of the warming cycle. AFP concentrations that enhance survival of red cells confer partial inhibition of recrystallization. Relatively high concentrations of AFP (e.g., 1.54 mg/ml) are much more effective at inhibiting extracellular recrystallization. However, extensive growth of ice around the cell, and concomitant cell damage, is noted. The mechanism for this AFP-induced ice growth is unknown. We propose that there is a delicate balance between AFP-induced enhancement of cell preservation and AFP-induced enhancement of cell preservation and AFP-induced enhancement of cell damage and that this balance hinges on the degrees of inhibition of ice recrystallization and of preferential growth of ice around the cells. We conclude that, under appropriate conditions, one of the proposed functions of AFPs in nature can be emulated, and perhaps have application, in cryopreservation of materials of biomedical interest.

Animals↗

Binding of an oligopeptide to a specific plane of ice.

The alpha-helical antifreeze protein (AFP) from winter flounder inhibits ice growth by binding to a specific set of pyramidal surface planes that are not otherwise macroscopically expressed. The 37-residue AFP contains three 11-amino acid repeats that make a stereo-specific fit to the ice lattice along the <01-12> direction of the (20-21) and equivalent binding planes. When the AFP was shortened to delete two of the three 11-amino acid ice-binding repeats, the resulting 15-residue peptide and its variants were less helical and showed no antifreeze activity. However, when the helicity of the peptide was reinforced by an internal lactam bridge between Glu-7 and Lys-11, the minimized AFP was able to stably express the pyramidal plane (20-21) on the surface of growing ice crystals. This dynamic shaping of the ice surface by a single ice-binding repeat provides evidence that AFP adsorption to the ice lattice is not an "all-or-nothing" interaction. Instead, a partial interaction can help develop the binding site on ice to which the remainder of the AFP (or other AFP molecules) can orient and bind.

Antifreeze Proteins↗

The effect of ice on intra-articular temperature in the knee of the dog.

The effect of surface cooling on intra-articular temperature was examined in dogs' knees. Four treatment protocols were examined: local ice compress application for 5, 15 and 30 min and local ice bath immersion for 15 min. Intra-articular temperatures were recorded using a needle microprobe inserted into the knee and continuous temperature recordings were made before, during and after the treatment. Rectal temperatures were also simultaneously recorded. Intra-articular temperatures rapidly dropped during icing. Five minutes of ice compress application resulted in a 2.2 +/- 1.2 degrees C intra-articular temperature drop with no change in rectal temperature. After 15 min of ice compress application, the joint temperature fell 4.1 +/- 1.3 degrees C with no change in rectal temperature. Thirty minutes of local ice compress application dropped the knee temperature 6.5 +/- 4.0 degrees C with a slight 0.5 +/- 0.3 degrees C drop in rectal temperature. Fifteen minutes of ice water immersion caused a much greater drop in intra-articular temperatures (20.2 +/- 8.4 degrees C) than could be achieved with ice compress. Rectal temperatures fell slightly during immersion (1.6 +/- 0.3 degrees C). After the removal of any type of cryotherapy, intra-articular temperatures continued to drop for several minutes and then a prolonged rewarming period commenced. The mean time required to return to baseline intra-articular temperature varied from 22-60 min, depending on the type and duration of cryotherapy. We conclude that brief periods of topical cold application to a dog's knee can induce significant and long lasting depression of intra-articular temperatures and that this is a local effect not dependent on core temperature cooling.

Animals↗

Frequency, distribution, and management of injuries due to an ice storm in a large metropolitan area.

Patterns of injuries seen in the emergency department (ED) following an ice storm were evaluated and compared to those seen during a control snow period. We conducted a retrospective chart review for an 11-day ice period and an 11-day control snow period using charts from eight metropolitan area EDs. Charts were reviewed for all patients with complaints of falls, motor vehicle crashes (MVCs), or sledding accidents due to ice or snow conditions. The numbers of injuries resulting from falls, MVCs, and sledding accidents for patients of various ages during each of the two periods were compared by chi-square analysis. Of 1,631 patients seen during the 11-day ice period, 1,181 (72%) had injuries due to falls, 192 (12%) were injured in MVCs, and 232 (14%) suffered sledding injuries. Falls on ice resulted in a fracture 39% of the time, whereas MVCs resulted in fracture 14% of the time. Falls accounted for 90% of the fractures during the ice period versus 76% during the snow period. The risk of sustaining a fracture as a result of a fall was significantly higher in older age groups (relative risk [RR] = 2.1), as was the risk of hospitalization (RR = 2.3). There was a significant increase in the number of falls and sledding accidents during the ice period. Thus, ice coverage resulted in a significant increase in fall-related and sledding-related fractures requiring treatment in the ED.

Accidental Falls↗

An immune response to ice crystals in North Atlantic fishes.

In mammals, the presence of crystals composed of small organic molecules, including urate and related compounds, has been shown to trigger an inflammatory response and the subsequent production of specific immunoglobulins (Ig's). Many fishes that are exposed to ice crystals in cold temperate and polar oceans may harbour ice crystals internally. Here, we report evidence for a specific immune response to ice crystals in cold-ocean marine fishes. Using ice nucleation activity as an assay, anti-ice Ig's were detected in the sera of the cold-ocean marine fish species, ocean pout (Macrozoarces americanus) and Atlantic herring (Clupea harengus harengus), but not in the sera of species that are not exposed to ice. Purified Ig's isolated from ocean pout serum using two different protocols showed ice nucleation activity, thus demonstrating the presence of ice binding specificity among these Ig's.

Animals↗

Structural transitions in amorphous water ice and astrophysical implications.

Selected area electron diffraction is used to monitor structural changes of vapor-deposited water ice in vacuum during warm-up from 15 to 188 K. A progression of three amorphous forms of water ice is found with well-defined transitions. The formation of a high-density amorphous form (Iah) at 15 K is confirmed, and the transition to the more familiar low-density form (Ial) occurs gradually over the range 38 to 68 K. At 131 K, the ice transforms into a third amorphous form (Iar), which precedes the crystallization of cubic ice (Ic) and coexists metastably with Ic from 148 K until at least 188 K. These structural transformations of amorphous water ice can be used to explain hitherto anomalous properties of astrophysical ices. The structural transition from Iah to Ial is responsible for the diffusion and recombination of radicals in ultraviolet-photolyzed interstellar ices at low temperatures. The occurrence and persistence of Iar explains anomalous gas retention and gas release from water-rich ices at temperatures above 150 K.

Algorithms↗

Bacterial Activity at -2 to -20 degrees C in Arctic wintertime sea ice.

Arctic wintertime sea-ice cores, characterized by a temperature gradient of -2 to -20 degrees C, were investigated to better understand constraints on bacterial abundance, activity, and diversity at subzero temperatures. With the fluorescent stains 4',6'-diamidino-2-phenylindole 2HCl (DAPI) (for DNA) and 5-cyano-2,3-ditoyl tetrazolium chloride (CTC) (for O(2)-based respiration), the abundances of total, particle-associated (>3- micro m), free-living, and actively respiring bacteria were determined for ice-core samples melted at their in situ temperatures (-2 to -20 degrees C) and at the corresponding salinities of their brine inclusions (38 to 209 ppt). Fluorescence in situ hybridization was applied to determine the proportions of Bacteria, Cytophaga-Flavobacteria-Bacteroides (CFB), and Archaea. Microtome-prepared ice sections also were examined microscopically under in situ conditions to evaluate bacterial abundance (by DAPI staining) and particle associations within the brine-inclusion network of the ice. For both melted and intact ice sections, more than 50% of cells were found to be associated with particles or surfaces (sediment grains, detritus, and ice-crystal boundaries). CTC-active bacteria (0.5 to 4% of the total) and cells detectable by rRNA probes (18 to 86% of the total) were found in all ice samples, including the coldest (-20 degrees C), where virtually all active cells were particle associated. The percentage of active bacteria associated with particles increased with decreasing temperature, as did the percentages of CFB (16 to 82% of Bacteria) and Archaea (0.0 to 3.4% of total cells). These results, combined with correlation analyses between bacterial variables and measures of particulate matter in the ice as well as the increase in CFB at lower temperatures, confirm the importance of particle or surface association to bacterial activity at subzero temperatures. Measuring activity down to -20 degrees C adds to the concept that liquid inclusions in frozen environments provide an adequate habitat for active microbial populations on Earth and possibly elsewhere.

Archaea↗

High-level expression of ice nuclei in a Pseudomonas syringae strain is induced by nutrient limitation and low temperature.

Attempts were made to maximize the expression of ice nuclei in Pseudomonas syringae T1 isolated from a tomato leaf. Nutritional starvation for nitrogen, phosphorous, sulfur, or iron but not carbon at 32 degrees C, coupled to a shift to 14 to 18 degrees C, led to the rapid induction of type 1 ice nuclei (i.e., ice nuclei active at temperatures warmer than -5 degrees C). Induction was most pronounced in stationary-phase cells that were grown with sorbitol as the carbon source and cooled rapidly, and under optimal conditions, the expression of type 1 ice nuclei increased from < 1 per 10(7) cells (i.e., not detectable) to 1 in every cell in 2 to 3 h. The induction was blocked by protein and RNA synthesis inhibitors, indicative of new gene expression. Pulse-labeling of nongrowing cultures with [35S]methionine after a shift to a low temperature demonstrated that the synthesis of a new set of "low-temperature" proteins was induced. Induced ice nuclei were stable at a low temperature, with no loss in activity at 4 degrees C after 8 days, but after a shift back to 32 degrees C, type 1 ice nuclei completely disappeared, with a half-life of approximately 1 h. Repeated cycles of low-temperature induction and high-temperature turnover of these ice nuclei could be demonstrated with the same nongrowing cells. Not all P. syringae strains from tomato or other plants were fully induced under the same culture conditions as strain T1, but all showed increased expression of type 1 ice nuclei after the shift to the low temperature. In support of this view, analysis of the published DNA sequence preceding the translational start site of the inaZ gene (R. L. Green and G. Warren, Nature [London] 317:645-648, 1985) suggests the presence of a gearbox-type promoter (M. Vincente, S. R. Kushner, T. Garrido, and M. Aldea, Mol. Microbiol. 5:2085-2091, 1991).

Bacterial Outer Membrane Proteins↗

Poles apart: biodiversity and biogeography of sea ice bacteria.

This review introduces the subjects of bacterial biodiversity and biogeography. Studies of biogeography are important for understanding biodiversity, the occurrence of threatened species, and the ecological role of free-living and symbiotic prokaryotes. A set of postulates is proposed for biogeography as a guide to determining whether prokaryotes are "cosmopolitan" (found in more than one geographic location on Earth) or candidate endemic species. The term "geovar" is coined to define a geographical variety of prokaryote that is restricted to one area on Earth or one host species. This review discusses sea ice bacteriology as a test case for examining bacterial diversity and biogeography. Approximately 7% of Earth's surface is covered by sea ice, which is colonized principally by psychrophilic microorganisms. This extensive community of microorganisms, referred to as the sea ice microbial community (SIMCO), contains algae (mostly diatoms), protozoa, and bacteria. Recent investigations indicate that the sea ice bacteria fall into four major phylogenetic groups: the proteobacteria, the Cytophaga-Flavobacterium-Bacteroides (CFB) group, and the high and low mol percent gram-positive bacteria. Archaea associated with sea ice communities have also been reported. Several novel bacterial genera and species have been discovered, including Polaromonas, Polaribacter, Psychroflexus, Gelidibacter, and Octadecabacter; many others await study. Some of the gram-negative sea ice bacteria have among the lowest maximum temperatures for growth known, < 10 degrees C for some strains. The polar sea ice environment is an ideal habitat for studying microbial biogeography because of the dispersal issues involved. Dispersal between poles is problematic because of the long distances and the difficulty of transporting psychrophilic bacteria across the equator. Studies to date indicate that members of some genera occur at both poles; however, cosmopolitan species have not yet been discovered. Additional research on polar sea ice bacteria is needed to resolve this issue and extend our understanding of its microbial diversity.

Antarctic Regions↗

Emperor penguins adjust swim speed according to the above-water height of ice holes through which they exit.

Emperor penguins leap from the water onto the sea ice. Their ability to reach above-water height depends critically on initial vertical speed of their leaping, assuming that the kinetic energy is converted to gravitational potential energy. We deliberately changed the above-water heights of ice hole exits, in order to examine whether penguins adjusted swim speed in accordance with the above-water height of the ice. Penguins were maintained in a corral on the fast ice in Antarctica, and voluntarily dived through two artificial ice holes. Data loggers were deployed on the penguins to monitor under water behavior. Nine instrumented penguins performed 386 leaps from the holes during experiments. The maximum swim speeds within 1 s before the exits through the holes correlated significantly with the above-water height of the holes. Penguins adopted higher speed to exit through the higher holes than through the lower holes. Speeds of some failed exits were lower than the theoretical minimum values to reach a given height. Penguins failed to exit onto the sea ice in a total of 37 of the trials. There was no preference to use lower holes after they failed to exit through the higher holes. Rather, swim speed was increased for subsequent attempts after failed leaps. These data demonstrated that penguins apparently recognized the above-water height of holes and adopted speeds greater than the minimal vertical speeds to reach the exit height. It is likely, especially in the case of higher holes (>40 cm), that they chose minimum speeds to exit through the holes to avoid excess energy for swimming before leaping. However, some exceptionally high speeds were recorded when they directly exited onto the ice from lower depths. In those cases, birds could increase swim speed without strokes for the final seconds before exit and they only increased the steepness of their body angles as they surfaced, which indicates that the speed required for leaps by emperor penguins were aided by buoyancy, and that penguins can sometimes exit through the ice holes without any stroking effort before leaping.

Animals↗

Mycosporine-like amino acids in Antarctic sea ice algae, and their response to UVB radiation.

Mycosporine like amino acids (MAAs) were detected in low concentration in sea ice algae growing in situ at Cape Evans, Antarctica. Four areas of sea ice were covered with plastics of different UV absorption exposing the bottom- ice algal community to a range of UV doses for a period of 15 days. Algae were exposed to visible radiation only; visible + UV radiation; and visible + enhanced UV radiation. MAA content per cell at the start of the experiment was low in snow-covered plots but higher in samples from ice with no snow cover. During the study period, the MAA content per cell reduced in all treatments, but the rate of this decline was less under both ambient UV and visible radiation than under snow covered plots. While low doses of UVB radiation may have stimulated some MAA production (or at least slowed its loss), relatively high doses of UVB radiation resulted in almost complete loss of MAAs from ice algal cells. Despite this reduction in MAA content per cell, the diatoms in all samples grew well, and there was no discernible effect on viability. This suggests that MAAs may play a minor role as photoprotectants in sea ice algae. The unique structure of the bottom ice algal community may provide a self-shading effect such that algal cells closest to the surface of the ice contain more MAAs than those below them and confer a degree of protection on the community as a whole.

Amino Acids↗

Effect of the fat globule sizes on the meltdown of ice cream.

The meltdown of ice cream is influenced by its composition and additives and by fat globule size. The objective of this study was to examine the effect of fat globule size and fat agglomerate size on the meltdown stability of ice cream. Therefore, an ice cream mix (10% milk fat) was homogenized at pressures ranging from 0 to 30 MPa in single-stage, double-stage, and selective homogenization processes. The ice cream, produced on a continuous ice cream freezer, was characterized by an optimized meltdown test while, in addition, the fat globule sizes and the free fat content were determined in the mix and the molten ice cream. The meltdown was dependent on the fat agglomerate sizes in the unfrozen serum phase. Agglomerates smaller than a critical diameter led to significantly higher meltdown rates. Homogenization pressures of at least 10 MPa were sufficient to produce a stable ice cream. Furthermore, proof was provided that double-stage homogenization is not necessary for fat contents up to 10% and that selective homogenization is possible to produce stable ice creams. Based on these results a model was deduced describing the stabilizing mechanisms during the meltdown process.

Chemical Phenomena↗

[Microbiological characterization of the accreted ice of subglacial Lake Vostok, Antarctica].

The accreted ice of subglacial Lake Vostok extends upward from the lake water level (a depth of 3750 m) to the bottom surface of the overlying Antarctic ice sheet. All of the accreted ice samples, taken from depths between 3541 and 3611 m, were found to contain pro- and eukaryotic microorganisms, whose number and diversity varied in different ice horizons and correlated, to a certain degree, with the occurrence of organic and inorganic impurities in a given horizon. Some biological objects found in the accreted lake ice, including bacteria, microalgae, and the pollen of higher plants, were morphologically similar to those found earlier in the glacier ice bulk. The others were not. It is suggested that the microorganisms found in the lake ice may come from different locations--the bottom layer of the glacier ice, the bedrock underlying the glacier, and the lake water.

Antarctic Regions↗

The application of the hazard analysis and control points (HACCP) in an ice cream production plant.

The HACCP system is applied in the present study as a preventive food safety approach to control some hazards appearing in one of the ice cream production plants in Egypt. The problem comprised the presence of bacteriological and some chemical contamination in most of the company products. Before applying the HACCP system, the samples examination showed high total mesophilic plat count in 50% of the samples and high coliform count in all samples compared to the level recommended in the Egyptian standards. The highest staphylococcal count (negative for coagulase test) obtained was that of chocolate (1.3 x 10(4) CFU/g) followed by mango ice cream (l.0 x 10(4) CFU/g). Faecal coliform was only positive in mango ice cream. Additionally, the levels of lead, iron and copper have been determined. Copper was higher than the recommended level in all samples and ranged between 0.46 ppm in pistachio ice cream and 2.48 ppm in chocolate ice cream. Most of the CCPs in the ice cream production were mainly due to improper handling and practices throughout the processing steps and also related to the lack of food hygiene knowledge of the workers. The application of HACCP system has successfully reduced the bacteriological hazards in all samples. After applying the corrective actions, the highest total aerobic plate count (8.0 x 10(4)) was that of Swiss chocolate ice cream which corresponds to 50% of the recommended level by the Egyptian standards. Coliform count was less than 10 in all samples except mango ice cream (43 MPN). A significant decrease in staphylococcal count, faecal coliform and trace metal contamination was also recorded in all samples.

Bacterial Infections↗

The structure and complete nucleotide sequence of the murine gene encoding interleukin-1 beta converting enzyme (ICE).

Interleukin-1 beta (IL-1 beta) converting enzyme (ICE) processes the precursor of the cytokine IL-1 beta to a mature, biologically active form in monocytes and macrophages. To further understand the role of ICE in regulating IL-1 beta-mediated biological functions, we have isolated several genomic clones encoding the full-length murine ICE gene. Southern blot comparison of murine genomic DNA and the clones indicates that ICE is a compact, single-copy gene 8616 bp in size. We sequenced the entire gene as well as 1.0-kb segment upstream of the coding region and determined that the gene consists of 10 exons whose organization parallels the functional organization of the ICE proenzyme in that the prodomain and p20 and p10 subunits of ICE are encoded by three clusters of exons. Two initiation sites, 37 and 32 nucleotides upstream of the initiator methionine, were identified by primer extension analysis. The 5' region of the ICE gene lacks a TATA box, a CAAT box, and SP1 sites. However, the presence of a completely conserved 14-bp sequence spanning the transcription initiation site of both the murine and the human ICE genes suggests that this sequence plays a role in transcription.

3T3 Cells↗

Site-specific bone mass differences of the lower extremities in 17-year-Old ice hockey players.

The purpose of the present study was to evaluate bone mass in the pelvis and lower extremities in young ice hockey players, and especially to investigate whether any differences are related to the type and magnitude of weight-bearing loading and muscle stress. The ice hockey group consisted of 22 boys (mean age 16.9 +/- 0.3) from three different ice hockey teams training for about 9 hours/week (mean 9.3 +/- 2.0, range 6-15). The reference group consisted of 22 boys (age 16.8 +/- 0.3) not training for more than 4 hours per week (mean 1.5 +/- 1.5, range 0-4). The groups were matched according to age, pubertal stage, weight, and height. Areal bone mineral density (BMD) was measured in total body, head, pelvis, ala ossis ilii, femoral neck, trochanter, femur diaphysis, and tibia diaphysis using dual energy X-ray absorptiometry. Isokinetic muscle strength of the quadriceps and hamstrings muscles was measured using an isokinetic dynamometer. BMDs of the total body, femoral neck, trochanter, and pelvis, but not of the head, ala ossis ilii, femur diaphysis, and tibia diaphysis, were found to be significantly higher (P < 0.05) in the ice hockey group. Muscle strength of the quadriceps muscles was also found to be significantly higher among the ice hockey players, but this greater muscle strength did not predict any BMD site significantly. However, in the reference group, there was a general strong relationship between muscle strength and BMD. This study has demonstrated significantly higher BMD in adolescent ice hockey players than in referents on a low or moderate level of physical activity. The differences seem to be site-specific and may be related to the type and magnitude of loading acting on each site, during off-season training and preferentially during ice hockey. The nonsignificant differences in BMD of the tibia and femur diaphyses may reflect that the compressive forces acting on these sites during ice hockey are not of sufficient magnitude to influence BMD. High muscle stress in itself, without weight-bearing loading, acting on the ala ossis ilii in adolescent boys does not seem to increase BMD, and an increased muscle strength does not predict BMD in highly trained adolescent boys.

Adolescent↗

The structures and functions of ice crystal-controlling proteins from bacteria.

Many organisms have evolved into unique mechanisms which minimize freezing injury due to extracellular ice formation. Specifically, certain bacteria have produced a few proteins each with different functions. For example, the ice nucleation protein acts as a template for ice formation, which is responsible for imparting ice nucleating activity. The anti-nucleating protein inhibits the fluctuation of ice nucleus formation by a foreign particle in the water drop. Also, the antifreeze proteins depress the freezing temperature, modify or suppress ice crystal growth, inhibit ice recrystallization, and protect the cell membrane from cold-induced damage. In this article, a review on the current knowledge of the structure and the function of these three types of proteins, which are capable of interacting with ice itself or its nuclei from bacteria.

Journal Article↗