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Although studies on snow algae and macroinvertebrates have been frequently conducted on snow patches, only few surveys have been focused on microinvertebrates which reach high biomass and play various trophic roles in other cold habitats. The aims of this study were (1) to search for microinvertebrates in seasonal surface snow patches located on the slope of Mt. Gassan, in northern Japan, and (2) to identify factors determining their distribution associated with snow algal blooms of various colorations (orange, green, and golden-brown) collected from the same sampling site over two seasons (2018, 2019). Microscopic observation revealed presence of two major groups of microinvertebrates: Tardigrada and Rotifera. They were concentrated in green snow colored by blooms of Chloromonas sp. in comparison to orange or golden-brown snow and only a few were found in white snow. Mean body length of tardigrades increased throughout the melt season, their intestine content was green and they laid eggs on colored snow. These results suggest that tardigrades preferentially grew and reproduced on green snow patches. Population densities of tardigrades, rotifers and concentration of chlorophyll a were significantly correlated. Our study indicates that green snow patches in temperate mountainous forests constitute important and unique low-temperature ecosystems for microinvertebrates. Snow covered by algae is an unrecognized novel habitats for tardigrades and rotifers.

Cryoconite is a mixture of mineral and organic material covering glacial ice, playing important roles in biogeochemical cycles and lowering the albedo of a glacier's surface. Understanding the differences in structure of cryoconite across the globe can be important in recognizing past and future changes in supraglacial environments and ice-organisms-dust interactions. Despite the worldwide distribution and over a century of studies, the basic characteristics of cryoconite, including its forms and geochemistry, remain poorly studied. The major purpose of our study is the presentation and description of morphological diversity, chemical and photoautotrophs composition, and organic matter content of cryoconite sampled from 33 polar and mountain glaciers around the globe. Observations revealed that cryoconite included various morphologies including loose and granular forms. Granular cryoconite includes smooth, rounded, or irregularly shaped forms; with some having their surfaces covered by cyanobacteria filaments. The occurrence of granules increased with the organic matter content in cryoconite. Moreover, a major driver of cryoconite colouring was the concentration of organic matter and its interplay with minerals. The structure of cyanobacteria and algae communities in cryoconite differs between glaciers, but representatives of cyanobacteria families Pseudanabaenaceae and Phormidiaceae, and algae families Mesotaeniaceae and Ulotrichaceae were the most common. The most of detected cyanobacterial taxa are known to produce polymeric substances (EPS) that may cementing matter and form granules. Organic matter content in cryoconite varied between glaciers, ranging from 1% to >40%. The geochemistry of all the investigated samples reflected local sediment sources, except of highly concentrated Pb and Hg in cryoconite collected from European glaciers near industrialized regions, corroborating cryoconite as element-specific collectors and potential environmental indicator of anthropogenic activity. Our work supports a notion that cryoconite may be more than just simple sediment and instead exhibits complex structure with relevance for biodiversity and the functioning of glacial ecosystems.

The results show the morphological analyses and spectroscopic studies of snow and glacier algae and their parasitic fungi in Svalbard (High Arctic). Fixed algal cells of two species, Sanguina nivaloides and Ancylonema nordenskioeldii, were imaged using light microscopy, scanning electron microscopy (SEM), and atomic force microscopy (AFM). Fluorescence microscopy using Calcofluor white stain supported the observations of parasitic fungi on the algal cells. Images in brightfield microscopy showed chytrid-like fungi penetrating the cells of both algal species. Parasites were found to colonize the cells of A. nordenskioeldii and hypnozygotes of S. nivaloides, while no fungi infected the cyst stages of S. nivaloides. The autofluorescence analysis revealed the ability of S. nivaloides to glow when excited with different wavelengths, while A. nordenskioeldii did not fluoresce. The hypnozygotes of S. nivaloides emitted brighter fluorescence than the cysts, and the most intense luminosity was observed in the UV range. The Fourier-transform infrared spectroscopy (FTIR) and energy-dispersive X-ray spectroscopy (EDS) spectroscopic analysis showed differences in the chemical composition between samples collected from three different sites. Samples dominated by cyst cells were characterized by the presence of an abundant polysaccharide envelope.

In both natural and built environments, microbes on occasions manifest in spherical aggregates instead of substratum-affixed biofilms. These microbial aggregates are conventionally referred to as granules. Cryoconites are mineral rich granules that appear on glacier surfaces and are linked with expanding surface darkening, thus decreasing albedo, and enhanced melt. The oxygenic photogranules (OPGs) are organic rich granules that grow in wastewater, which enables wastewater treatment with photosynthetically produced oxygen and which presents potential for net autotrophic wastewater treatment in a compact system. Despite obvious differences inherent in the two, cryoconite and OPG pose striking resemblance. In both, the order Oscillatoriales in Cyanobacteria envelope inner materials and develop dense spheroidal aggregates. We explore the mechanism of photogranulation on account of high similarity between cryoconites and OPGs. We contend that there is no universal external cause for photogranulation. However, cryoconites and OPGs, as well as their intravariations, which are all under different stress fields, are the outcome of universal physiological processes of the Oscillatoriales interfacing with goldilocks interactions of stresses. Finding the rules of photogranulation may enhance engineering of glacier and wastewater systems to manipulate their ecosystem impacts.

Snow algae are photosynthetic microbes that inhabit the melting snow surface in alpine and polar regions. We analyzed the pigment and species composition of colored snow collected on Mt. Tateyama in Japan during the melting seasons of 2015 and 2016. High-performance liquid chromatographic analyses of the pigments extracted from the colored snow showed that their composition varied within the study area and were classified into four types: Type A (astaxanthin-monoester dominant), Type B (medium astaxanthin-monoester content), Type C (abundant primary carotenoids and free-astaxanthin), and Type D (abundant primary carotenoids and astaxanthin diesters). Types A and B were most commonly observed in the study area, whereas Types C and D appeared only at specific sites. Analysis of the 18S ribosomal RNA (18S rRNA) gene revealed six major amplicon sequence variants (ASVs) of snow algae, belonging to the Sanguina, Chloromonas, and Chlainomonas groups. The relative abundance of the algal ASVs showed that Sanguina was dominant (>48%) in both Types A and B, suggesting that the difference in astaxanthin abundance between the two types was caused by the production of pigments in the algal cells. The algal community structures of Types C and D differed from those of Types A and B, indicating that the primary carotenoids and astaxanthin diesters were derived from certain algal species in these types. Therefore, astaxanthin-rich Sanguina algae mostly induced the red snow that appeared widely in this alpine area; however, they were partially dominated by Chloromonas or Chlainomonas algae, causing different pigment compositions.

The glacier stonefly Andiperla willinki is the largest metazoan inhabiting the Patagonian glaciers. In this study, we analysed the gut microbiome of the aquatic nymphs by 16S rRNA gene amplicon and metagenomic sequencing. The bacterial gut community was consistently dominated by taxa typical of animal digestive tracts, such as Dysgonomonadaceae and Lachnospiraceae, as well as those generally indigenous to glacier environments, such as Polaromonas. Interestingly, the dominant Polaromonas phylotypes detected in the stonefly gut were almost never detected in the glacier surface habitat. Fluorescence in situ hybridization analysis revealed that the bacterial lineages typical of animal guts colonized the gut wall in a co-aggregated form, while Polaromonas cells were not included in the aggregates. Draft genomes of several dominant bacterial lineages were reconstructed from metagenomic datasets and indicated that the predominant Dysgonomonadaceae bacterium is capable of degrading various polysaccharides derived from host-ingested food, such as algae, and that other dominant bacterial lineages ferment saccharides liberated by the polysaccharide degradation. Our results suggest that the gut bacteria-host association in the glacier stonefly contributes to host nutrition as well as material cycles in the glacier environment.

Snow algae are found from spring to summer on snowfields and glaciers throughout the world. Their blooming darkens snow surfaces, reducing snow surface albedo and accelerating melting. Uncertaintie...

Snow algae are photosynthetic microbes growing in thawing snow. They usually show various morphological cell types. The aim of this study was to carry out microscopic and spectroscopic analysis of different forms of cells of snow algae collected on glaciers in Alaska. Four different shapes of algal cells were observed with the use of bright field LM (Light Microscopy), DIC (Differential Interference Contrast), EDF (Extended Depth Focus), fluorescence microscopy, and SEM (Scanning Electron Microscopy). The cells exhibited the strongest autofluorescence after the exposure to 365-nm excitation light, and the intensity differed among the cell types. Zygotes (cysts) showed the most intense fluorescence. Acridine orange staining revealed the acid nature of the algal cells. The use of Congo red and Calcofluor white fluorochromes indicated differences in the structure of polysaccharides in the cell wall in the individual types of algal cells. FTIR (Fourier-Transform Infrared Spectroscopy) analyses showed the presence of polysaccharides not only in the algal cells but also in the fixative solution. The presence of polysaccharides in the extracellular algal fraction was confirmed by X-ray dispersion spectroscopy (EDS), X-ray photoelectron spectroscopy (XPS), and scanning electron microscopy imaging (SEM). The differences observed in the structure of the cell wall of the different forms of red snow algae prompt further analysis of this structure.

Cryoconite granules are naturally occurring microbial structures on glacier surfaces worldwide. They play a key role in carbon and nitrogen cycling in glacier ecosystems and can accelerate the melting of snow and ice. However, detailed mechanism of nitrogen cycling in cryoconite granules remains unclear. Here, we demonstrate that redox stratification affects the spatial distribution of N cycling processes in cryoconite granules. Based on microsensor measurements for O2, NH4+, NO2– and NO3–, we identified the presence of fine-scale redox stratification within cryoconite granules. Cyanobacteria at the surface layer of the granules created oxic conditions, whereas the inner core of the granules was anoxic. Metatranscriptomic analyses indicated the active occurrences of nitrification in the inner core, whereas denitrification actively occurred both in the inner core and the surface layer of the granules. Cyanobacteria in the inner core of the granules were inactive, and likely dead and being degraded, providing carbon and nitrogen to support nitrifiers and denitrifiers. Quantities of nitrification genes/transcripts were greater in large cryoconite granules than small ones, most likely because nitrogen substrates were more abundantly present in the inner core of large granules due to distinct redox stratification. Our results suggest that the development of a granular structure of cryoconite granules can largely affect carbon and nitrogen cycling on glaciers.

To understand the microbial composition and diversity patterns, cryoconite granules were collected from two geographical areas, i.e., Nepali Himalaya and Greenland, Arctic. 16S rRNA, ITS and the D1/D2 domain sequencing techniques were used for characterization of microbial communities of the four glaciers. The total 13 species of bacteria such as Bacillus aryabhattai, Bacillus simplex, Brevundimonas vesicularis, Cryobacterium luteum, Cryobacterium psychrotolerans, Dermacoccus nishinomiyaensis, Glaciihabitans tibetensis, Leifsonia kafniensis, Paracoccus limosus, Polaromonas glacialis, Sporosarcina globispora, Staphylococcus saprophyticus, Variovorax ginsengisoli, and 4 species of fungi such as Goffeauzyma gilvescens, Mrakia robertii, Dothideomycetes sp., Helotiales sp. were recorded from Nepali Himalaya. Among these, 12 species of bacteria and 4 species of fungi are new contributions to Himalaya. In contrast to this, six species of bacteria such as Bacillus cereus, Cryobacterium psychrotolerans, Dermacoccus nishinomiyaensis, Enhydrobacter aerosaccus, Glaciihabitans tibetensis, Subtercola frigoramans, and nine species of fungi such as Goffeauzyma gilvescens, Mrakia robertii, Naganishia vaughanmartiniae, Piskurozyma fildesensis, Rhodotorula svalbardensis, Alatospora acuminata, Articulospora sp., Phialophora sp., Thelebolus microspores, and Dothideomycetes sp.), were recorded from Qaanaaq, Isunnguata Sermia and Thule glaciers, Greenland. Among these, five species of bacteria and seven species of fungi are new contributions to Greenland cryoconite. Microbial analyses indicate that the Nepali Himalayan cryoconite colonize higher numbers of microbial species compared to the Greenland cryoconite.

Surface albedo of snow and ice is substantially reduced by inorganic impurities, such as aeolian mineral dust (MD) and black carbon (BC), and also by organic impurities, such as microbes that live in the snow. In this paper, we present the temporal changes of surface albedo, snow grain size, MD, BC and snow algal cell concentration observed on a snowpack in northwest Greenland during the ablation season of 2014 and our attempt to reproduce the changes in albedo with a physically based snow albedo model. We also attempt to reproduce the effects of inorganic impurities and the red snow algae (Sanguina nivaloides) on albedo. Concentrations of MD and red snow algae in the surface snow were found to increase in early August, while snow grain size and BC were found to not significantly change throughout the ablation season. Surface albedo was found to have decreased by 0.08 from late July to early August. The albedo simulated by the model agreed with the albedo observed during the study period. However, red snow algae exerted little effect on surface albedo in early August. This is probably owing to the abundance of smaller cells (4.9×104 cells L−1) when compared with the cell abundance of red snow reported by previous studies in the Arctic region (∼108 cells L−1). The simulation of snow albedo until the end of the melting season, with a snow algae model, revealed that the reduction in albedo attributed to red snow algae could equal 0.004, out of a total reduction of 0.102 arising from the three impurities on a snowpack in northwest Greenland. Finally, we conducted scenario simulations using the snow albedo model, coupled with the snow algae model, in order to simulate the possible effects of red snow blooming on snow albedo under warm conditions in northwest Greenland. The result suggests that albedo reduction by red snow algal growth under warm conditions (surface snow temperature of +1.5 ∘C) reached 0.04, equivalent to a radiative forcing of 7.5 W m−2 during the ablation season of 2014. This coupled albedo model has the potential to dynamically simulate snow albedo, including the effect of organic and inorganic impurities, leading to proper estimates of the surface albedo of snow cover in Greenland.

Surface albedo of snow and ice is substantially reduced by inorganic impurities, such as aeolian mineral dust (MD) and black carbon (BC), and also by organic impurities, such as microbes that live in the snow. In this paper, we present the temporal changes of surface albedo, snow grain size, MD, BC, and snow algal cell concentration observed on a snowpack in northwest Greenland during the ablation season of 2014 and our attempt to reproduce the changes in albedo with a physically based snow albedo model. We also attempt to reproduce the effects of inorganic impurities and the red snow algae (Chlamydomonas nivalis) on albedo. Concentrations of MD and red snow algae in the surface snow were found to increase in early August, while snow grain size and BC were found to not significantly change throughout the ablation season. Surface albedo was found to have decreased by 0.08 from late July to early August. The albedo simulated by the model agreed with the albedo observed during the study period. However, red snow algae exerted little effect on surface albedo in early August. This is probably owing to the abundance of smaller cells (4.9 × 104 cells L^-1) when compared with the cell abundance of typical red algal snow (~ 108 cells L−1). The simulation of snow albedo until the end of the melting season, with an algal growth model, revealed that the reduction in albedo attribute to red algae could equal 0.004, out of a total reduction of 0.102 arising from the three impurities on a snowpack in northwest Greenland. Finally, we conducted scenario simulations using the snow albedo model, coupled with the algal growth model, in order to simulate the possible effects of red algal blooming on snow albedo under warm conditions in northwest Greenland. The result suggests that albedo reduction by red snow algal growth under warm conditions (surface snow temperature of +1.5 °C) reached 0.04, equivalent to a radiative forcing of 7.5 W m−2 during the ablation season of 2014. This coupled albedo model has the potential to dynamically simulate snow albedo, including the effect of organic and inorganic impurities, leading to proper estimates of the surface albedo of snow cover in Greenland.

Despite the crucial role of cyanobacteria in various ecosystems, little is known about their evolutionary histories, especially microevolutionary dynamics, because of the lack of knowledge regarding their mutation rates. Here we directly estimated cyanobacterial mutation rates based on ancient DNA analyses of ice core samples collected from Kyrgyz Republic that dates back to ~12,500 cal years before present. We successfully sequenced the 16S rRNA and 16S–23S internal transcribed spacer (ITS) region. Two cyanobacterial operational taxonomic units (OTUs) were detected from the ancient ice core samples, and these OTUs are shared with those from the modern glacier surface. The mutation rate of ITS region was estimated by comparing ancient and modern populations, and were at the magnitude of 10(−7)substitutions/sites/year. By using a model selection framework, we also demonstrated that the ancient sequences from the ice sample were not contaminated from modern samples. Bayesian demographic analysis based on coalescent theory revealed that cyanobacterial population sizes increased over Asia regions during the Holocene. Thus, our results enhance our understanding of the enigmatic timescale of cyanobacterial microevolution, which has the potential to elucidate the environmental responses of cyanobacteria to the drastic climatic change events of the Quaternary.

Cryoconite granules are dark-colored biological aggregates on glaciers. Bacterial community varies with granule size, however, community change in space and their susceptibility to environmental factors has not been described yet. Therefore, we focused on bacterial community from four different granule sizes (30–249 μm, 250–750 μm, 750–1599 μm, more than 1600 μm diameter) in 10 glaciers in northwestern Greenland and their susceptibility to exogenous nutrients in cryoconite hole. A filamentous cyanobacterium Phormidesmis priestleyi, which has been frequently reported from glaciers in Arctic was abundant (10%–26%) across any size of granules on most of glaciers. Bacterial community across glaciers became similar with size increase, and whence smallest size fractions contain more unique genera in each glacier. Multivariate analysis revealed that effect of nutrients to beta diversity is larger in smaller granules (30–249 μm and 250–750 μm diameter), suggesting that bacterial susceptibility to nutrients changes with growth of granule (i.e. P. priestleyi was affected by nitrate in early growth stage).

Cryoconite granules, which are dark-colored biological aggregates on glaciers, effectively accelerate the melting of glacier ice. Bacterial community varies with granule size, however, community change in space and their susceptibility to environmental factors has not been described yet. Therefore, we focused on bacterial community from 4 different granule sizes (30-249 μm, 250- 750 μm, 750-1599 μm, more than 1600 μm diameter) in 10 glaciers in northwestern Greenland and their susceptibility for exogenous nutrients in cryoconite hole. A filamentous cyanobacterium Phormidesmis priestleyi, which has been frequently reported from glaciers in Arctic was abundant (10-26%) across any size of granules on most of glaciers. Bacterial community across glaciers became similar with size increase, and whence smallest size fractions contain more unique genera in each glacier. Multivariate analysis suggests that phosphate, which is significantly higher in one glacier (Scarlet Heart Glacier), is primary associated with bacterial beta diversity. Correlation coefficients between abundance of major genera and nutrients largely changed with granule size, suggesting that nutrients susceptibility to genera changes with growth process of granule (e.g. P. priestleyi was affected by nitrate in early growth stage).

Recent molecular data has strongly suggested that field-collected cysts of snow algae that are morphologically identifiable as the zygotes of Chloromonas nivalis are composed of multiple species. Motile vegetative cells, however, have not been directly obtained from these cysts because of the difficulties involved in inducing their germination. Recently, our comparative molecular analyses, using both field-collected and cultured materials, demonstrated that one Japanese lineage of "C. nivalis zygotes" belongs to C. miwae. Herein, we examined another Japanese lineage of field-collected "C. nivalis zygotes" and a new strain originating from Japan. Our molecular data demonstrated that these two different life cycle stages are conspecific, and that they represent a new species that we herein describe as C. muramotoi sp. nov., based on the vegetative and asexual morphological characteristics of the strain. Multigene phylogenetic analyses showed that this new species was sister to C. miwae. Scanning electron microscopy demonstrated that the cysts of C. muramotoi are different from those of C. miwae, based on the arrangement of the flanges developing on the cell wall.

Snow and ice algal communities were investigated on four glaciers in the Suntar-Khayata Mountain Range in eastern Siberia in Russia over three melting seasons from 2012 to 2014. Two taxa of green algae and five taxa of cyanobacteria were observed on the glaciers. The algal community was dominated by green algae: Ancylonema nordenskioldii in the lower bare ice area and Chloromonas sp. in the upper snow area. The total algal bio-volume showed altitudinal variation, ranging from 0.03 to 4.0 mL m−2, and was greatest in the middle of the glaciers. The altitudinal variations in the algal community were similar on all studied glaciers, suggesting that they are typical in this region. Observations over the three years revealed that there was no significant change in the community structure, but a significant change in the total biomass. Since the mean summer air temperature was significantly higher in 2012 when algal biomass was greater, the difference in algal biomass among the years is probably due to the duration of surface melting. The community structure on the studied glaciers is similar to those on glaciers in Arctic and sub-Arctic regions.

Microbial colonization of glacial ice surfaces incurs feedbacks which affect the melting rate of the ice surface. Ecosystems formed as microbe-mineral aggregates termed cryoconite locally reduce ice surface albedo and represent foci of biodiversity and biogeochemical cycling. Consequently, greater understanding the ecological processes in the formation of functional cryoconite ecosystems upon glacier surfaces is sought. Here, we present the first bacterial biogeography of an ice cap, evaluating the respective roles of dispersal, environmental and biotic filtration occurring at local scales in the assembly of cryoconite microbiota. 16S rRNA gene amplicon semiconductor sequencing of cryoconite colonizing a Svalbard ice cap coupled with digital elevation modelling of physical parameters reveals the bacterial community is dominated by a ubiquitous core of generalist taxa, with evidence for a moderate pairwise distance-decay relationship. While geographic position and melt season duration are prominent among environmental predictors of community structure, the core population of taxa appears highly influential in structuring the bacterial community. Taxon co-occurrence network analysis reveals a highly modular community structured by positive interactions with bottleneck taxa, predominantly Actinobacteria affiliated to isolates from soil humus. In contrast, the filamentous cyanobacterial taxon (assigned to Leptolyngbya/Phormidesmis pristleyi) which dominates the community and binds together granular cryoconite are poorly connected to other taxa. While our study targeted one ice cap, the prominent role of generalist core taxa with close environmental relatives across the global cryosphere indicate discrete roles for cosmopolitan Actinobacteria and Cyanobacteria as respective keystone taxa and ecosystem engineers of cryoconite ecosystems colonizing ice caps.

Glaciers and ice sheets are considered a biome with unique organism assemblages. Tardigrada (water bears) are micrometazoans that play the function of apex consumers on glaciers. Cryoconite samples with the dark-pigmented tardigrade Cryoconicus gen. nov. kaczmareki sp. nov. were collected from four locations on glaciers in China and Kyrgyzstan. The erection of the new genus is based on a unique combination of morphological traits as well as on phylogenetic analyses. The analysis of COI sequences in the new species revealed high genetic differentiation with 9 haplotypes shared among 13 sequenced individuals from three sequenced populations. There was no apparent geographic structure in COI haplotype diversity, which might indicate effective dispersal abilities of the new species. A recovery of numerous live individuals from a sample that was frozen for 11 years suggests high survival rates in the natural environment. The ability to withstand low temperatures, combined with dark pigmentation that is hypothesised to protect from intense UV radiation, could explain how the new taxon is able to dwell in an extreme glacial habitat. We also found that a rare mountain tardigrade Ramazzottius cataphractus (Maucci, 1974) is morphologically similar to the new species, therefore we propose to transfer it to the new genus. Our study indicates that glacier invertebrate fauna is still poorly known and requires intense research.

The community structure of bacteria associated with the glacier ice worm Mesenchytraeus solifugus was analyzed by amplicon sequencing of 16S rRNA genes and their transcripts. Ice worms were collected from two distinct glaciers in Alaska, Harding Icefield and Byron Glacier, and glacier surfaces were also sampled for comparison. Marked differences were observed in bacterial community structures between the ice worm and glacier surface samples. Several bacterial phylotypes were detected almost exclusively in the ice worms, and these bacteria were phylogenetically affiliated with either animal-associated lineages or, interestingly, clades mostly consisting of glacier-indigenous species. The former included bacteria that belong to Mollicutes, Chlamydiae, Rickettsiales, and Lachnospiraceae, while the latter included Arcicella and Herminiimonas phylotypes. Among these bacteria enriched in ice worm samples, Mollicutes, Arcicella, and Herminiimonas phylotypes were abundantly and consistently detected in the ice worm samples; these phylotypes constituted the core microbiota associated with the ice worm. A fluorescence in situ hybridization analysis showed that Arcicella cells specifically colonized the epidermis of the ice worms. Other bacterial phylotypes detected in the ice worm samples were also abundantly recovered from the respective habitat glaciers; these bacteria may be food for ice worms to digest or temporary residents. Nevertheless, some were overrepresented in the ice worm RNA samples; they may also function as facultative gut bacteria. Our results indicate that the community structure of bacteria associated with ice worms is distinct from that in the associated glacier and includes worm-specific and facultative, glacier-indigenous lineages.

Cryoconites are microbial aggregates commonly found on glacier surfaces where they tend to take spherical, granular forms. While it has been postulated that the microbes in cryoconite granules play an important role in glacier ecosystems, information on their community structure is still limited, and their functions remain unclear. Here, we present evidence for the occurrence of nitrogen cycling in cryoconite granules on a glacier in Central Asia. We detected marker genes for nitrogen fixation, nitrification and denitrification in cryoconite granules by digital polymerase chain reaction (PCR), while digital reverse transcription PCR analysis revealed that only marker genes for nitrification and denitrification were abundantly transcribed. Analysis of isotope ratios also indicated the occurrence of nitrification; nitrate in the meltwater on the glacier surface was of biological origin, while nitrate in the snow was of atmospheric origin. The predominant nitrifiers on this glacier belonged to the order Nitrosomonadales, as suggested by amoA sequences and 16S ribosomal RNA pyrosequencing analysis. Our results suggest that the intense carbon and nitrogen cycles by nitrifiers, denitrifiers and cyanobacteria support abundant and active microbes on the Asian glacier.

Antibiotic resistance genes are biologically transmitted from microorganism to microorganism in particular micro-environments where dense microbial communities are often exposed to an intensive use of antibiotics, such as intestinal microflora, and the soil microflora of agricultural fields. However, recent studies have detected antibiotic-resistant bacteria and/or antibiotic resistance genes in the natural environment geographically isolated from such areas. Here we sought to examine the prevalence of antibiotic resistance genes in 54 snow and ice samples collected from the Arctic, Antarctic, Central Asia, North and South America and Africa, to evaluate the level of these genes in environments supposedly not affected by anthropogenic factors. We observed a widespread distribution of antibiotic resistance genes in samples from various glaciers in Central Asia, North and South America, Greenland and Africa. In contrast, Antarctic glaciers were virtually free from these genes. Antibiotic resistance genes, of both clinical (i.e. aac(3), blaIMP) and agricultural (i.e. strA and tetW) origin, were detected. Our results show regional geographical distribution of antibiotic resistance genes, with the most plausible modes of transmission through airborne bacteria and migrating birds.

Cryoconite granules are aggregations of microorganisms with mineral particles that form on glacier surfaces. To understand the processes by which the granules develop, this study focused on the altitudinal distribution of the granules and photosynthetic microorganisms on the glacier, bacterial community variation with granules size and environmental factors affecting the growth of the granules. Size-sorted cryoconite granules collected from five different sites on Qaanaaq Glacier were analyzed. C and N contents were significantly higher in large (diameter greater than 250 μm) granules than in smaller (diameter 30–249 μm) granules. Bacterial community structures, based on 16S rRNA gene amplicon sequencing, were different between the smaller and larger granules. The filamentous cyanobacterium Phormidesmis priestleyi was the dominant bacterial species in larger granules. Multivariate analysis suggests that the abundance of mineral particles on the glacier surface is the main factor controlling growth of these cyanobacteria. These results show that the supply of mineral particles on the glacier enhances granule development, that P. priestleyi is likely the key species for primary production and the formation of the granules, and that the bacterial community in the granules changes over the course of the granule development.

Fourty-two high-rank syntaxa and seven associations of the thallophyte system of syntaxa are either described as new or validated in this paper. Among those, there are the following nine classes: Aspicilietea candidae , Caulerpetea racemosae, Desmococcetea olivacei , Entophysalidetea deustae, Gloeocapsetea sanguineae , Mesotaenietea berggrenii , Naviculetea gregariae , Porpidietea zeoroidis , Roccelletea phycopsis . Eleven orders and ten alliances as well as three associations are described or validated: the Aspicilietalia verruculosae (incl. Aspicilion mashiginensis and Teloschistion contortuplicati ) , the Caulerpetalia racemosae (incl. Caulerpion racemosae ) , the Desmococcetalia olivacei (incl. Desmococcion olivacei ) , the Dirinetalia massiliensis , the Fucetalia vesiculosi (incl. Ascophyllion nodosi ) , the Gloeocapsetalia sanguineae, the Lecideetalia confluescentis (incl. Lecideion confluescentis ) , the Mesotaenietalia berggrenii (incl. Mesotaenion berggrenii, Mesotaenietum berggrenii and Chloromonadetum nivalis ) , the Naviculetalia gregariae (incl. Oscillatorion limosae and Oscillatorietum limosae ) , the Porpidietalia zeoroidis (incl. Porpidion zeoroidis ) , and the Roccelletalia fuciformis (incl. Paralecanographion grumulosae ). Further, five orders, seven alliances and four associations, classified in known classes, were described as well. These include: the Bacidinetalia phacodis, the Agonimion octosporae and the Dendrographetalia decolorantis (all in the Arthonio radiatae-Lecidelletea elaeochromae ), the Staurothelion solventis (in the Aspicilietea lacustris ), the Pediastro duplicis-Scenedesmion quadricaudae and the Pediastro duplicis-Scenedesmetum quadricaudae (both in the Asterionelletea formosae ), the Peccanion coralloidis and the Peltuletalia euplocae (both in the Collematetea cristati ), the Laminarion hyperboreae, the Saccorhizo polyschidi-Laminarietum and the Alario esculenti-Himanthalietum elongatae (all in the Cystoseiretea crinitae ) , the Delesserietalia sanguinei, the Delesserion sanguinei and the Delesserietum sanguineae (all in the Lithophylletea soluti ), as well as the the Rinodino confragosae-Rusavskietalia elegantis and the Rhizocarpo geographici-Rusavskion elegantis (both in the Rhizocarpetea geographici ).

Cryoconite granules are dark-colored spherical aggregates of organic and inorganic material on glacier ice, and are commonly observed on glaciers the world over. The structure of cryoconite granules on Ürümqi glacier No. 1, Tien Shan, China, was analyzed. Granules were distributed over the entire ice surface of the ablation area, and ranged in size from 0.26 to 3.5 mm (mean 1.1 mm). The granule surface was densely covered with filamentous cyanobacteria. Microscopy of a thin section revealed various inner structures. Most granules had concentric layers of dense organic matter, which are probably derived from annual growth of the granules by the activity of cyanobacteria. The number of layers averaged 3.5 and ranged up to 7, which is likely to indicate their mean and maximum growth ages, respectively. Some granules contained two or more subgranules, showing that small granules had combined and enlarged. Such structures suggest that granule formation was mainly due to the activity of filamentous cyanobacteria, and that the granules repeatedly grew and disintegrated over a cycle of several years on the glacier.

Cyanobacterial communities on a glacier in the Qilian Shan, western China, were investigated using microscopic as well as 16S rRNA and internal transcribed spacer gene analyses. Microscopy revealed that there were abundant cyanobacteria on the entire glacier surface and their community consisted mainly of three morphological types. Low-cycle 16S rRNA gene sequences from six clone libraries were grouped into a total of eight cyanobacterial operational taxonomic units (OTUs), defined as 16S rRNA sequences with similarity of 99%. Although the cyanobacterial community based on morphological types displayed no significant differences among the study sites on the glacier, the community based on OTU groups varied among sites. This inconsistency may be due to simple morphology which might hide a large genetic variability. Phylogenetic analysis revealed that the OTU groups included the orders Oscillatoriales, Chroococcales and unclassified, and the majority of OTUs were Oscillatoriales. From the source environments of the cyanobacterial 16S rRNA gene sequences of each OTU on the glacier estimated by BLAST search (>97% similarity), 39.9% were from soil, 38.2% from fresh water and 1.7% from snow and ice environments. Based on geographical records in the database, all cyanobacterial OTUs were matched to those recorded from the Arctic and Antarctica. The results suggest that the cyanobacterial communities on the glacier are common in cold regions of the world and are likely not to be specialized members of the snow and ice biota but also inhabitants of soil and freshwater environments.

Snow algae in a 45.97-m-long ice core from the Tyndall Glacier (50°59′05″S, 73°31′12″W, 1756 m a.s.l.) in the Southern Patagonian Icefield were examined for potential use in ice core dating and estimation of the net accumulation rate. The core was subjected to visual stratigraphic observation and bulk density measurements in the field, and later to analyses of snow algal biomass, water isotopes ( 18 O, D), and major dissolved ions. The ice core contained many algal cells that belonged to two species of snow algae growing in the snow near the surface: Chloromonas sp. and an unknown green algal species. Algal biomass and major dissolved ions (Na + , K + , Mg 2+ , Ca 2+ , Cl − , SO 4 2− ) exhibited rapid decreases in the upper 3 m, probably owing to melt water elution and/or decomposition of algal cells. However, seasonal cycles were still found for the snow algal biomass, 18 O, D-excess, and major ions, although the amplitudes of the cycles decreased with depth. Supposing that the layers with almost no snow algae were the winter layers without the melt water essential to algal growth, we estimated that the net accumulation rate at this location was 12.9 m a − 1 from winter 1998 to winter 1999, and 5.1 m from the beginning of winter to December 1999. These estimates are similar to the values estimated from the peaks of 18 O (17.8 m a − 1 from summer 1998 to summer 1999 and 11.0 m from summer to December 1999) and those of D-excess (14.7 m a − 1 from fall 1998 to fall 1999 and 8.6 m a − 1 from fall to December 1999). These values are much higher than those obtained by past ice core studies in Patagonia, but are of the same order of magnitude as those predicted from various observations at ablation areas of Patagonian glaciers.

The glacier ice worm, Mesenchytraeus solifugus, is a unique annelid, inhabiting only snow and ice in North American glaciers. Here, we analyzed the taxonomic composition of bacteria associated with M. solifugus based on the 16S rRNA gene. We analyzed four fixed-on-site and 10 starved ice worm individuals, along with glacier surface samples. In total, 1341 clones of 16S rRNA genes were analyzed for the ice worm samples, from which 65 bacterial phylotypes (99.0% cut-off) were identified. Of these, 35 phylotypes were closely related to sequences obtained from their habitat glacier and/or other components of cryosphere; whereas three dominant phylotypes were affiliated with animal-associated lineages of the class Mollicutes. Among the three, phylotype Ms-13 shared less than 89% similarity with database sequences and was closest to a gut symbiont of a terrestrial earthworm. Using fluorescence in situ hybridization, Ms-13 was located on the gut wall surface of the ice worms. We propose a novel genus and species, 'Candidatus Vermiplasma glacialis', for this bacterium. Our results raise the possibility that the ice worm has exploited indigenous glacier bacteria, while several symbiotic bacterial lineages have maintained their association with the ice worm during the course of adaptive evolution to the permanently cold environment.

Snow algae in a shallow ice core (6.98 m long) from Yala glacier in the Langtang region of Nepal were examined for potential use as environmental markers in ice-core analysis. The ice core, taken at 5350m a.s.l. in 1994, was estimated to contain 11 annual layers from 1984 to 1994 from the profile of algal biomass. Algal biomass in each annual layer was noted to be correlated with air temperature, and the following two environmental indices which were calculated from air temperature and precipitation at Kyangjing (3920m a.s.l.), the village nearest to Yala glacier: estimated mean snow-cover thickness (MST) and estimated summer mass balance (SMB). Both parameters reflect snow-cover thickness on algal layers, which would be a major determinant of the light available for algal growth on the glacier. Snow algal biomass in the ice core appears to be a good environmental marker for indicating air temperature and accumulation during summer, which is important for understanding the mass balance of summer-accumulation-type glaciers in this region.

Snow algae are cold-tolerant algae growing on snow and ice and have been reported on glaciers in many parts of the world. Blooms of snow algae can reduce the surface albedo of snow and ice and significantly affect their melting. In addition, snow algae found in ice cores can be potential indicators of the paleo-environment, making them of great interest both to the biology and the geophysics of glaciers. A snow algal community was investigated in 2002 and 2003 on Akkem glacier in the Russian Altai mountains, where no information on its biological community has previously been available. Five species of snow algae including green algae and cyanobacteria were observed on the glacier. Red snow due to a bloom of algae (Chloromonas sp.) was visually apparent in the snow area during our study periods. The total algal cell-volume biomass on the glacier ranged from 97 to 1156μL m−2, which is equivalent to that reported previously on glaciers in the Himalaya and Alaska. The community structure showed that Mesotaenium berggrenii and/or Ancylonema nordenskioeldii, which are common species on glaciers in the Northern Hemisphere, were dominant in the ice area, while Chloromonas sp. was dominant in the snow area. Such community structures are similar to those on Alaskan and Arctic glaciers but differ from those on Himalayan and Tibetan glaciers, even though the Altai mountains are geographically closer to the Himalaya and Tibet than to Alaska. The difference in algal communities between the Altaic and other glaciers is discussed together with physical and chemical conditions affecting the algae.

We quantitatively investigated a snow algal community on Tyndall Glacier of the Southern Patagonia Icefield, Chile, at an elevation from 300 to 1500 m a.s.l. We observed 7 species of snow and ice algae (Chlorophyta and cyanobacteria) on the glacier. These species were Mesotaenium (M.) berggrenii, Cylindrocystis (Cyl.) brebissonii, Ancylonema sp., Closterium sp., Chloromonas sp., Oscillatoriaceae cyanobacterium, and an unknown alga. The spatial distribution of these algae differed among the species. M. berggrenii, Cyl. brebissonii, Ancylonema sp., and Closterium sp. appeared mainly in the lower-elevation area (370–770 m a.s.l.), the unknown alga in the higher-elevation area (900–1500 m a.s.l.), and Chloromonas sp. and Oscillatoriaceae cyanobacterium in the middle part of the glacier. The mean cell concentration and total cell volume biomass ranged from 0 to 9.2 × 104 (mean: 1.8 × 104) cells mL−1, from 0 to 327 (mean: 63) μL m−2, respectively. The cell volume biomass generally decreased with altitu...

The altitudinal distribution of a snow algal community was investigated on an Alaska glacier (Gulkana Glacier in the Alaska Range) from 1270 to 1770 m a.s.l.. Seven species of snow and ice algae (Chlorophyta and cyanobacteria) were observed on the glacier surface. These species were Chlamydomonas nivalis, Mesotaenium berggrenii, Ancylonema nordenskioldii, Cylindrocystis brebissonii, Raphidonema sp., and two Oscillatoriaceae cyanobacteria. The altitudinal distribution of snow algae was different among the species: Cd. nivalis was distributed on the middle to upper area, M. berggrenii; A. nordenskioldii, and one Oscillatoriaceae cyanobacterium on the middle to lower area; Raphidonema sp. on the middle area; and Cyl. brebissonii and one Oscillatoriaceae cyanobacterium on the lower area. The total cell concentration and the cell volume biomass of the snow algae ranged from 4·4 × 103 to 9·9 × 105 cells ml−1 and from 33 to 2211 µl m−2 respectively. The cell volume biomass changed with altitude; the biomass increased with altitude below 1600 m a.s.l., and decreased above 1600 m a.s.l. The community structure showed that A. nordenskioldii dominated on the lower part of the glacier, and that Cd. nivalis dominated on the upper part. The species diversity was relatively high at the lowest and middle sites. The pH was 4·7 to 5·3 for snow and 4·9 to 5·7 for ice on the glacier. The altitudinal distribution of snow algae is discussed in terms of the physical and chemical condition of the glacier surface, and is compared with that on a Himalayan glacier. A larger biomass in the snow area on the Alaska glacier than that of the Himalayan glacier is likely due to less frequent snow cover in summer in Alaska. Small amounts of filamentous cyanobacteria on the Alaska glacier may allow washouts of unicellular green algae by running melt water and may cause a different pattern of altitudinal distribution of algal biomass on the ice area from the Himalayan glacier. Copyright © 2001 John Wiley & Sons, Ltd.

N -Acetyl-β- d -glucosaminidase (NAG) is a widely used urinary enzyme for the assessment of renal diseases. We studied the stabilities of NAG isoenzymes in urine at 37°C by enzyme assay and ELISA using a model simulating in vivo conditions. The stabilities were found to be affected by the pH. Under mild acidic condition (about pH 6), there was no significant loss of enzymatic activity of NAG isoenzyme A, enzymatic activity of NAG isoenzyme B and immunological activity of NAG isoenzyme B even after 8 h incubation. In contrast, under alkaline condition (about pH 8), the enzymatic activity of NAG isoenzyme A was rapidly lost, whereas both enzymatic and immunological activities of NAG isoenzyme B were maintained at more than 80% of their initial values. Also, we found that the ratios of endogenous NAG isoenzyme B to total NAG were elevated in alkaline urine samples. These results indicate that NAG isoenzyme A, which is a major isoenzyme in normal urine (pH 5–7), seems to be inactivated in alkaline urine. Our results suggest that for alkaline urine, NAG isoenzyme B should be measured to avoid misinterpretation of total NAG enzymatic activities.

We have developed a new ELISA for quantifying N-acetyl-β-d-glucosaminidase (NAG) isoenzyme B in human urine after raising monoclonal antibodies against the isoenzyme from human placenta. Though the obtained antibodies reacted not only to isoenzyme B but also to A, we could detect isoenzyme B selectively by a two-step sandwich ELISA with a pair of selected antibodies at low pH in the first reaction. The detected limit was 0.5 μg/L for a sample volume of 25 μL. Within-run CVs ranged from 2.5% to 5.4% and between-run CVs ranged from 6.2% to 9.1%. Recoveries of NAG isoenzyme B added to each of three urine samples ranged from 91% to 114%. The dilution curves of urine samples showed good linearity. The cross-reactivity of NAG isoenzyme A was practically negligible (2–3%). The mean value for NAG isoenzyme B in spot urines from healthy adults was 2.9 μg/g creatinine. This ELISA method is rapid and precise enough for routine determination of NAG isoenzyme B in human urine.

Heparin administration to diabetic rats caused no change in VLDL, an increase in IDL and a decrease in LDL on electrophoretic analysis of plasma lipoproteins, while the administration to control rats markedly decreased VLDL and increased IDL and LDL. Both hepatic triglyceride lipase (HTGL) and lipoprotein lipase (LPL) activities in the postheparin plasma were lower in the diabetic rats than in the controls, and the reduction of HTGL activity was greater than that of LPL activity in the diabetic rats. The LPL activity in the adipose tissue was lower in the diabetic rats than in the controls, but the activities in the cardiac and skeletal muscles were similar in the two rats. The HTGL-catalyzed fatty acid (FA) releases from the diabetic VLDL and IDL were lower than those from the normal rat VLDL and IDL, while the LPL-catalyzed FA release in the diabetic rats was not different from those in the controls. The decreases in LPL and HTGL activities and the markedly impaired susceptibility of IDL to HTGL coincide well with the postheparin changes in plasma lipoproteins in diabetic rats, an increase in IDL and a decrease in LDL.

Mouse complement component C7 was purified from serum by a sequential procedure of fractionation precipitation by ammonium sulfate, followed by DE-52 anion exchange chromatography, Protein G affinity column chromatography, Mono S cation exchange chromatography and Superdex 200 gel filtration. The final product contained a highly purified mouse C7 component showing a single band on SDS-PAGE at the apparent Mrs of 90 kDa and 100 kDa under non-reduced and reduced conditions respectively. The yield of C7, which was measured by the biological activity, was 7.0%.

To clarify altitudinal changes in the bacterial community on Gulkana Glacier in Alaska, we analyzed bacterial 16S rRNA gene by low-cycle PCR amplification, denaturing gradient gel electrophoresis (DGGE), and culturing in a snowmelt medium at 4°C. Low-cycle PCR-based cloning revealed the presence of 100 bacterial OTUs; however, 41 OTUs were identified only in a single clone, suggesting that their abundance was limited because of difficulty in predominating on the glacier. In contrast, 17 major OTUs accounted for 57–87% of the clone library at each site, suggesting that they accounted for the major part of the bacteria on the glacier. In addition, five of the 17 OTUs were included in the 21 OTUs cultured in the snowmelt medium. Based on the dominant phylotypes and DGGE results, the bacterial community on the glacier could be divided into three types, corresponding to the snow-covered, snow- and ice-covered, and bare-ice areas of the glacier. Our results suggest that a relatively limited number of bacteria predominate and that each phylotype is adapted to a distinct set of conditions on the glacier.

The expression of a membrane-associated lymphotoxin molecule (mLT) on lymphokine-activated killer (LAK) cells obtained from 18 patients with malignant tumors and its role in the tumor cell killing mechanisms were investigated. LAK cells from tumor-infiltrating lymphocytes (TIL-LAK cells) were mainly composed of CD3-positive cells, whereas LAK cells from peripheral blood lymphocytes (PBL-LAK cells) were mainly composed of CD16- and CD56-positive cells. However, mLT was found to be expressed on TIL-LAK cells as well as PBL-LAK cells. The degree of mLT expression correlated with the killing activity of LAK cells towards L929 cells (r = 0.806, P < 0.01, n = 15), but not with that towards Daudi or K562 cells. Although the degree of mLT expression correlated with the amount of secreted lymphotoxin (LT) in the supernatant of LAK cell culture, the secreted LT itself could not account for the tumor cell killing activity of LAK cells. Polyclonal rabbit anti-LT antibody partially inhibited the killing activities of LAK cells towards L929 cells and this inhibition was found in the combination of autologous tumor cells and PBL-LAK cells. These findings suggest the possibility that the mLT-related cytotoxicity is involved in the tumor cell killing mechanisms of TIL-LAK cells as well as PBL-LAK cells.

In an attempt to clarify the mechanisms of cytotoxicity mediated by CD4+ cytotoxic T lymphocytes (CTL), the expression of perforin and membrane-bound lymphotoxin (LT) (tumor necrosis factor-beta) in herpes simplex virus (HSV)-specific CD4+ human cytotoxic and noncytotoxic T- cell clones was examined. Three HSV-specific CD4+ human CTL clones that showed HLA-DR-restricted cytotoxicity and proliferative response were established. The cytotoxicity of these clones in 5-hour 51Cr release assays was found to be mediated by the directional target cell lysis and not by the release of cytotoxic soluble factors, ie, “innocent bystander” killing. Northern blot analysis showed that messenger RNAs for perforin and LT, which were both considered to be important mediators for cytotoxicity of CD8+ CTL and natural killer cells, were abundantly expressed in HSV-specific CD4+ CTL clones. Expression of perforin in the cytoplasm of CD4+ CTL clones was also detected by immunohistochemical staining using a monoclonal antibody against perforin. In addition, LT bound to the cell surface of CD4+ CTL clones was detected by flow cytometry. In contrast, little or no expression of perforin and LT was detected in three HSV-specific CD4+ noncytotoxic T- cell clones. Although the cytotoxicity mediated by lymphokine-activated killer cells was partly inhibited by addition of anti-LT antibody, it did not show any effect on the cytotoxicity of HSV-specific CD4+ CTL clones. In addition, it was found that cytotoxicity mediated by these CD4+ CTL clones was Ca2(+)-dependent. These data thus suggest that perforin and membrane-bound LT are both expressed in HSV-specific CD4+ CTL, although perforin might be the more important mediator in short- term culture.

Glacier environments harbor cold-tolerant bacteria (Amato et al., 2007; Christner et al., 2003; Segawa et al., 2005). However, a variety of bacteria such as soil and enteric bacteria have been detected from these highly oligotrophic and psychrophilic environments (Segawa et al., 2005; Sheridan et al., 2003). The origin of these bacteria remains unknown. These bacteria may have been supplied at least in part from animals by means such as bird droppings (Sjolund et al., 2008). There are other plausible transmissions, such as local and global atmospheric circulation. In the case of the global atmospheric circulation, a glacier environment may receive bacteria originating from hu-man industrial activities, although most glaciers are located at a distance from such sites of human activi-ties. Emergence of antibiotic resistant bacteria has been considered as a human impact on the environment (Cabello, 2006; Chee-Sanford et al., 2001; Goni-Urriza et al., 2000). The prevalence of antibiotic resistant genes in the glacier environment can be used as an indicator of human impact on the global environment. In this experiment, the prevalence of antibiotic resis-tant genes in glacier ice samples taken from various locations in the northern hemisphere (Alaska, Central Asia) and Antarctica were evaluated by real time PCR array approach using a Taqman probe system. We collected samples of the glacial ice (1 cm in depth) or cryoconite hole samples (Table 1) using a sterilized stainless steel scoop or sterilized syringe, re-spectively. The samples were stored in 50-ml sterile plastic conical tubes (Iwaki, Tokyo). All samples were kept frozen and transported to the Tokyo Institute of Technology, Japan or National Institute of Polar Re-search, Japan. Bacterial DNA was extracted as follows within a Class 100 Clean bench (Sanyo, Tokyo, Model MCV-131BNS) at the National Institute of Polar Research (Tokyo). Ice samples were melted and a portion (1‒

The rat anti-mouse erythrocyte (MRBC) monoclonal antibody (mAb), R13, has been developed. The MRBC membrane protein recognized by R13 (R13-Ag) can be pirified by loading the butanol-extracted MRBC membrane solution on a R13-conjugated Cellulofine column in the presence of 0.1% CHAPS followed by elution with 1% CHAPS. The amino acid sequence of the affinity-purified R13-Ag corresponded to that predicted from the cDNA for the murine heat-stable antigen. It was revealed that the actual heat-stable antigen was composed of 27 amino acids.

Hyperphosphatasemia due to increased intestinal type serum alkaline phosphatase was noted in a 48-year-old male who had asymptomatic liver cirrhosis. The alkaline phosphatase activity in the serum was 828 U/l (our reference range in adults: 57–194 U/l), 94% of which was of the intestinal type as measured by an immunoprecipitation method. The intestinal component of alkaline phosphatase was separated into two major and some minor components using electrophoresis and isoelectrofocusing. One of the major components had similar mobility to that of a standard intestinal enzyme purified from adult intestine. The components were heat-labile and neuraminidase-resistant. Serial lectin affinity chromatography, however, indicated that sugar chain compositions of the alkaline phosphatase were different from those of the standard tissue intestinal enzyme. These results and further enzymological studies suggest that the patient's serum alkaline phophatase basically consisted of several intestine-like isoforms.

In order to clarify the relationship between composition and lipolytic responses to lipoprotein lipase (LPL), very low density lipoproteine (VLDL) from rats or humans were incubated with a commercially available LPL or with a partially purified LPL from postheparin human plasma and fatty acids released from VLDL were determined in vitro. VLDL from rats fed a diet containing 0.25 cholesterol for 6 months were rich in cholesterol and poor in triglycerides, and released less fatty acids from incubation with LPL than those from control rats. VLDL from normo- and hypertriglyceridemic human subjects were incubated with LPL. The fatty acid release poorly correlated with the apoprotein ratios of VLDL, apo C-III/C-II, B/E, and C/E with the exception of apo B/C, but it correlated well with the ratio of triglyceride/either one of the surface components including total apoproteins, free cholesterol and phospholipids in VLDL or the ratio of the triglyceride/total sum of the surface components. The correlation coefficients between fatty acid release and a ratio of triglyceride/total surface components were 0.774 (using the commercially available LPL) and 0.786 (using the partially purified human LPL). The fatty acid release increased after pretreatment of VLDL with phospholipase A2. The phospholipid content of VLDL was reduced without significant changes in other VLDL components. Thus, the responses of VLDL to LPL treatment may depend mainly upon the surface: core relationship of VLDL rather than its apoprotein composition except in rare clinical cases such as apo C-II deficiency.

[1] Red snow caused by algal bloom is common on glaciers and snowfields worldwide. Description of spatial distributions of snow algal blooms is important for understanding snow algae’s unique life in an extremely cold environment and for determining the effect of algae through the reduction of surface albedo. Here we present the spatial distribution of red snow algae on the Harding Icefield, Alaska retrieved from a satellite image. Field observations on the icefield conducted in August 2001 revealed visible red snow, particularly near the snowline. Field measurements of spectral reflectance on the surface revealed the specific spectral absorption of algal pigments. We found a significant correlation between snow algal biomass and a reflectance ratio of SPOT (Satellite Probatoire d’ Observation de la Terre) satellite band of wavelength 610–680 nm to band 500–590 nm. Using this relationship between the reflectance ratio and algal biomass, we estimated the distribution and abundance of red snow across the icefield using a SPOT satellite image. The spatial distribution of red snow on the icefield obtained by mapping the reflectance ratio matched field observations across the icefield with more red algal blooms on the continental than the maritime side of the icefield. Area averaged mean carbon content estimated from the red algal biomass for the icefield on the image was 1.2 kg km 2 .

Biological activity on glaciers has been believed to be extremely limited. However, in Himalayan and Patagonian glaciers, we have found specialized biotic communities, including various cold-tolerant insects and copepods that were living on the glacier by feeding on algae and bacteria growing in the snow and ice. Since these microorganisms growing on the glacier surface are stored in the glacial strata every year, ice-core samples contain many layers with abundant microorganisms. In Himalayan glaciers, we studied snow algae on the glacier surface and in shallow ice cores, and showed that the layers with much snow algae in the ice core could be good boundary markers of annual layers and very useful for ice-core dating. Snow algae can be a new environmental proxy for studies of ice cores from warmer regions such as the Himalayas and Patagonia, where data on chemical and isotopic content data are not reliable because of heavy mixing.

Serum cholesterol, triglyceride and phospholipid levels, liver cholesterol concentration, bile flow, biliary cholesterol, phospholipid and bile acid secretion rates, fecal sterol and bile acid levels and their bile acid compositions were examined in young-old parabiotic rats and compared with those in young and old control rats and young-young parabiotic rats. Bile acid composition was expressed in terms of the cholic acid group/chenodeoxycholic acid group (CA/CDCA) ratio. Body weight (BW) gain decreased after parabiosis especially in old rats, but the liver weight (g/100 g BW), diet-intake, feces dry weight, liver cholesterol concentration and fecal sterol level were almost the same in all the groups. The biliary bile acid secretion rate was higher and the fecal bile acid level was lower in old rats than those in young rats but both the levels became comparable with those in young rats after parabiosis of old rats with young rats. Young rats, however, showed no changes in these levels after parabiosis. The serum cholesterol level and the biliary and fecal CA/CDCA ratios in old rats were higher than those in young rats but decreased after parabiosis with young rats, although they were still higher than those in young rats. The serum cholesterol level in young rats increased after parabiosis with old rats, but not after parabiosis with young rats, and the fecal bile acid level and the CA/CDCA ratio were not changed in either case. It is concluded from these findings that the serum cholesterol level and the CA/CDCA ratio increased with age and that these increases were prevented after parabiosis with young rats, while young rats, although their serum cholesterol level was increased, showed no increase in the CA/CDCA ratio after parabiosis with old rats.

Snow and ice algae are cold tolerant algae growing on the surface of snow and ice, and they play an important role in the carbon cycles for glaciers and snowfields in the world. Seasonal and altitudinal variations in seven major taxa of algae (green algae and cyanobacteria) were investigated on the Gulkana glacier in Alaska at six different elevations from May to September in 2001. The snow algal communities and their biomasses changed over time and elevation. Snow algae were rarely observed on the glacier in May although air temperature had been above 0 ° C since the middle of the month and surface snow had melted. In June, algae appeared in the lower areas of the glacier, where the ablation ice surface was exposed. In August, the distribution of algae was extended to the upper parts of the glacier as the snow line was elevated. In September, the glacier surface was finally covered with new winter snow, which terminated algal growth in the season. Mean algal biomass of the study sites continuously increased and reached 6.3 × 10 μl m−2 in cell volume or 13 mg carbon m−2 in September. The algal community was dominated by Chlamydomonas nivalis on the snow surface, and by Ancylonema nordenskioldii and Mesotaenium berggrenii on the ice surface throughout the melting season. Other algae were less abundant and appeared in only a limited area of the glacier. Results in this study suggest that algae on both snow and ice surfaces significantly contribute to the net production of organic carbon on the glacier and substantially affect surface albedo of the snow and ice during the melting season.

Pollen taxon in sediment samples can be identified by analyzing pollen morphology. Identification of related species based on pollen morphology is difficult and is limited primarily to genus or family. Because pollen grains of various ages are preserved at below 0 C in glaciers and thus are more likely to remain intact or to suffer little DNA fragmentation, genetic information from such pollen grains should enable identification of plant taxa below the genus level. However, no published studies have attempted detailed identification using DNA sequences obtained from pollen found in glaciers. As a preliminary step, this study attempted to analyze the DNA of Pinus pollen grains extracted from surface snow collected from the Belukha glacier in the Altai Mountains of Russia in the summer of 2003. A 150-bp rpoB fragment from the chloroplast genome in each Pinus pollen grain was amplified by polymerase chain reaction, and DNA products were sequenced to identify them at the section level. A total of 105 pollen grains were used for the test, and sequences were obtained from eight grains. From the sequences obtained, the pollen grains were identified as belonging to the section Quinquefoliae. Trees of the extant species Pinus sibirica in the section Quinquefoliae are currently found surrounding the glacier. The consistency of results for this section suggests that the pollen in the glacier originated from the same Pinus trees as those found in the immediate surroundings.

Wistar male rats were treated for six days with broad spectrum beta-lactam antibiotics, latamoxef, and cefotaxime. On the seventh day, the number of fecal anaerobic microbes decreased, total fecal bile acids decreased, and bile acid pools increased. Secondary bile acids such as beta-hyocholic, hyodeoxycholic, lithocholic, and deoxycholic acids decreased in the feces while the primary bile acids, cholic, beta-muricholic, and chenodeoxycholic acids, became predominant. Coprostanol, a microbial metabolite of cholesterol, also disappeared from the feces during the treatment. The cecum enlarged to almost twice the size of that in control rats, whereas the liver weight was not significantly changed. After treatment was stopped, the number of fecal microbes returned to the initial counts within a week, but restoration of bile acid and cholesterol metabolism required at least three weeks.

In some hyperlipidemic patients, low density lipoprotein (LDL) shows several peaks (polydisperse) on polyacrylamide gel disc electrophoreses, though LDL usually shows a single peak (monodisperse). In order to clarify the relationship between the LDL polydispersion and VLDL heterogeneity, LDL and VLDL were prepared from hyperlipidemic patients sera with mono- and polydisperse LDL by sequential ultracentrifugation and fractionated by gradient ultracentrifugation and their compositions were analyzed. Polydisperse LDL was rich in triacylglycerol (TG) and poor in esterified cholesterol (CE) as compared with monodisperse LDL and consisted of the lowest and the medium density subfractions when the LDL was separated into six subfractions. The monodisperse LDL was composed of a single major subfraction of a medium density. VLDL from the patients with polydisperse LDL was relatively rich in the dense and poor in the buoyant subfractions as compared with that from the patients with monodisperse LDL. The subfractions in the former contained more CE and less TG than the corresponding subfractions in the latter. There were no significant differences in the apolipoprotein compositions between those VLDLs. The results suggest that polydisperse LDL might be originated from VLDL that differs in particle sizes, densities and compositions from ordinary VLDL.