Isolation and properties of barophilic and barotolerant bacteria from deep-sea mud samples

Several barophilic and barotolerant bacteria were isolated from deep-sea mud samples of Suruga Bay (2485 m depth), the Ryukyu Trench (5110 m depth), and the Japan Trench (land-side 6356 m, and sea-side 6269 m depth, respectivelys. The barophilic bacteria, strains DB5501, DB6101, DB6705 and DB6906, were albe to grow better under high hydrostatic pressures than under atmospheric pressure (0.1 megapascals; MPa). The optimal growth pressures for the barophilic bacteria were approximately 50 MPa at 10°C. The barotolerant strains DSK1 and DSS12 were determined to be psychrophilic, and had optimal growth temperatures of 10°C and 8°C, respectively. The degree of barophily and barotolerance was shown to be very dependent on temperature. For example, at 4°C the barophilic strains were indistinguishable from barotolerant bacteria, whereas at 15°C the barotolerant strains behaved more like the barophilic strains. Based on sequence analysis of 16S ribosomal DNA, all of the strains included in this study belong to the gamma subgroup of the Proteobacteria. Phylogenetic relations between the isolated strains and the known gamma subgroup bacteria suggested that the isolated strains belong to a new sub-branch of this group.     

北极高维度海域海冰嗜冷菌系统发育多样性及其低温水解酶分析

应用样品直接稀释涂布平板、-1℃富集培养和-20℃冷冻24h后富集培养等3种方法,从北极加拿大海盆和格陵兰海的高纬度海域(77°30′N~81°12′N)海冰中分离到37株嗜冷菌。根据其16S rDNA全长序列所进行的系统发育分析表明,分离菌株分属于γ_变形细菌群(γ_Proteobacteria)的Colwellia、Marinobacter、Shewanella、Thalassomonas、Glaciecola、Marinomonas、Pseudoalteromonas和嗜纤维菌_曲挠杆菌_拟杆菌群(Cytophaga_Flexibacter_Bacteroide,CFB)的Flavobacterium、Psychroflexus等9个属。其中有9株菌的16S rDNA序列与已明确鉴定种的相似性在93.4%~96.9%,为潜在的新种。北极加拿大海盆海冰细菌BSi20002与南极威德尔海海冰细菌Marinobactersp.ANT8277的16S rDNA序列相似性为100%,表明在种水平上南、北两极也存在相同的细菌。分离的嗜冷菌在4℃条件下能产生多种大分子物质水解酶类,其中62.6%、51.4%和40.5%的菌株分别能水解Tween_80、明胶和淀粉。     

Growth of Bacteria Degrading Naphthalene and Salicylate at Low Temperatures

A total of 58 bacterial strains degrading naphthalene and salicylate were isolated from soil samples polluted with oil products, collected in different regions of Russia during winter and summer. The isolates were assessed for their ability to grow at low temperatures (4, 8, and 15°C); bacteria growing at 4°C in the presence of naphthalene or salicylate accounted for 65 and 53%, respectively, of the strains isolated. The strains differed in the temperature dependence of their growth rates. It was demonstrated that the type of expression of the Nah⁺ phenotype at low temperatures depended on the combination of host bacterium and plasmid.     

Diversity and association of psychrophilic bacteria in Antarctic sea ice.

The bacterial populations associated with sea ice sampled from Antarctic coastal areas were investigated by use of a phenotypic approach and a phylogenetic approach based on genes encoding 16S rRNA (16S rDNA). The diversity of bacteria associated with sea ice was also compared with the bacterial diversity of seawater underlying sea ice. Psychrophilic (optimal growth temperature, < or = 15 degrees C; no growth occurring at 20 degrees C) bacterial diversity was found to be significantly enriched in sea ice samples possessing platelet and bottom ice diatom assemblages, with 2 to 9 distinct (average, 5.6 +/- 1.8) psychrophilic taxa isolated per sample. Substantially fewer psychrophilic isolates were recovered from ice cores with a low or negligible population of ice diatoms or from under-ice seawater samples (less than one distinct taxon isolated per sample). In addition, psychrophilic taxa that were isolated from under-ice seawater samples were in general phylogenetically distinct from psychrophilic taxa isolated from sea ice cores. The taxonomic distributions of psychrotrophic bacterial isolates (optimal growth temperature, > 20 degrees C; growth can occur at approximately 4 degrees C) isolated from sea ice cores and under-ice seawater were quite similar. Overall, bacterial isolates from Antarctic sea ice were found to belong to four phylogenetic groups, the alpha and gamma subdivisions of the Proteobacteria, the gram-positive branch, and the Flexibacter-Bacteroides-Cytophaga phylum. Most of the sea ice strains examined appeared to be novel taxa based on phylogenetic comparisons, with 45% of the strains being psychrophilic. 16S rDNA sequence analysis revealed that psychrophilic strains belonged to the genera Colwellia, Shewanella, Marinobacter, Planococcus, and novel phylogenetic lineages adjacent to Colwellia and Alteromonas and within the Flexibacter-Bacteroides-Cytophaga phylum. Psychrotrophic strains were found to be members of the genera Pseudoalteromonas, Psychrobacter, Halomonas, Pseudomonas, Hyphomonas, Sphingomonas, Arthrobacter, Planococcus, and Halobacillus. From this survey, it is proposed that ice diatom assemblages provide niches conducive to the proliferation of a diverse array of psychrophilic bacterial species.     

Life cycle of the microbivorous Antarctic Dry Valley nematode Scottnema lindsayae (Timm 1971)

The life cycle of the Antarctic Dry Valley soil nematode, Scottnema lindsayae (Timm 1971) was studied in laboratory culture at two temperatures, 10°C and 15°C. Soil yeast and bacteria isolated with the nematodes were used as the food source. The species reproduced sexually. The higher temperature had a negative effect on the life cycle. The number of eggs per female and the number of juveniles developing per female were greater at 10°C than at 15°C. Juveniles developed faster at 10°C and four juvenile stages were observed outside of the egg at both temperatures. The unusually long life cycle (218 d at 10°C) suggests that more than one austral summer may be required for successful completion. An increase in Dry Valley soil temperatures associated with potential global environmental change may have detrimental effects on soil nematodes.     

Impact of the Antarctic benthic fauna on the enrichment of biopolymer degrading psychrophilic bacteria

Stenothermic cold adaptation was a predominant growth characteristic among biopolymer degrading bacteria from Antarctic shelf sediments. Psychrophilic decomposers of protein (gelatin), chitin, and cellulose accounted for up to 84, 93, and 68%, respectively, of 0°C-isolates from selected compartments of the sediments. Macroinvertebrates were recognized as a selective pressure on these fast-growing (zymogenous) psychrophiles. Psychrophilic properties of growth and biopolymer degradation coincided most in the case of proteolytic isolates. On the other hand, the majority of psychrophilic chitin- and cellulose-decomposers showed less efficient biopolymer degradation at environmental temperatures (0°C). Temperature optima of the activities of pertinent depolymerizing enzymes (e.g., scleroprotease) exceeded by far the temperature optima for growth (between 4 and 12°C). Therefore, it appears likely that enhanced rates of enzyme synthesis at low temperatures play a crucial role for the degradation of detrital organic matter in this permanently cold environment.     

Aspects of the biology of the midge,Eretmoptera murphyi Schaeffer (Diptera: Chironomidae), introduced to Signy Island,maritime Antarctic

Summary The biology of a population of the subantarctic chironomid midge Eretmoptera murphyi Schaeffer, introduced to Signy Island in the maritime Antarctic more than 20 years ago, is described. Investment in reproduction by the parthenogenetic adult females is high, with individuals producing single egg batches containing ca. 85 eggs and having a dry weight of more than twice that of the spent female. In culture, egg development rates to hatching are increased significantly by increasing temperature from 2° to 12°C (a range covering mean summer temperatures found in the species' maritime Antarctic habitat, and natural habitat in the subantarctic). The gelatinous matrix of the egg batch forms a skin on drying, which may reduce further water loss, and allow the eggs or pre-emergence larvae to survive the short periods of desiccating conditions likely to occur in their natural habitat. The biology of E. murphyi is compared with that of the endemic maritime Antarctic species Belgica antarctica, showing much similarity. E. murphyi possesses several preadaptations which allow it to survive the harsher conditions of the maritime Antarctic.     

Effects of temperature on the growth of psychrophilic bacteria from glaciers.

Growth of five strains of psychrophilic bacteria (four Arthrobacter and one Pseudomonas) isolated from glacial deposits was studied at different temperatures. Three strains were facultative psychrophiles, having an optimum temperature for growth at about 25-28 degrees C and a maximum at about 32-34 degrees C. The two Arthrobacter glacialis strains were found to be obligate psychrophiles with an optimum at 13-15 degrees C and a maximum at 18 degrees C. Arrhenius plots showed that A. glacialis could compete with the facultative psychrophilic bacteria only at 0 degrees C, that is, the temperature of its natural environment. The psychrophilic Arthrobacter species studied here are more resistant to thermal stress than are marine psychrophilic bacteria. For Arthrobacter, in contrast to Pseudomonas, temperatures above the optimum induced formation of filaments and abnormal cells. The culture turbidity increased 10 to 30 times, whereas viable count tended to decrease. The thermal block seems to prevent cell wall synthesis and septation, but at a different step for each species.     

Temperature dependence of growth,enzyme secretion and activity of psychrophilic Antarctic bacteria

Five psychrophilic Antarctic bacteria have been selected for their capacity to secrete exoenzymes into culture medium. These strains are able to grow from 0 to about 25° C. However, production of lipase fromMoraxella, -amylase fromAlteromonas haloplanctis, -lactamase fromPsychrobacter immobilis and protease fromBacillus is maximal at temperatures close to that of their environment (—2 to 4° C) and is strongly inhibited at higher temperatures. This thermal effect involves alterations in the secretory pathway in the upper range of temperatures, losses due to the enzyme thermal lability and in some cases to reduction in cell development. The apparent optimal activity temperature of these enzymes is between 30 and 40° C, i.e. about 20° C lower than that of their mesophilic counterparts.     

Production and properties of two types of xylanases from alkalophilic thermophilic Bacillus spp.

Summary Four strains (W1, W2, W3, and W4) of alkalophilic thermophilic bacteria which produced xylanase were isolated from soils. They were aerobic, spore-forming, Gram-positive, and rod-shaped bacteria and hence identified as the genus Bacillus. The optimal temperatures for growth of the four strains were between 45° C and 50° C and pH optima were between 9.0 and 10.0. No growth occurred below pH 7.0 or above 55° C. The four strains produced xylanases in medium containing xylan or xylose under these conditions. The optimal pH and temperature for activities of the four xylanases ranged from 6.0 to 7.0 and from 65° C to 70° C, respectively. The four xylanases were stable in the wide pH range from 4.5 to 10.5 at 45° C for 1 h. All xylanases split xylan to yield xylose and xylobiose.     

A description of seven Antarctic marine gymnamoebae including a new subspecies, two new species and a new genus: Neoparamoeba aestuarina antarctica n. subsp., Platyamoeba oblongata n. sp., Platyamoeba contorta n. sp. and Vermistella antarctica n. gen. n. sp

Seven marine gymnamoebae were isolated from different environments of seawater, slush (pack ice meltwater), and sediment in the Ross Sea area of Antarctica. All amoebae were isolated and maintained at temperatures below 4 degrees C. Growth, rate of locomotion, and general morphology were observed at an environmentally appropriate temperature (1 degrees C) and at room temperature (approximately 25 degrees C). Molecular (srDNA sequences) and microscopical techniques were used to identify the gymnamoebae and establish their phylogenetic affinities. Three isolates (S-131-2, SL-200, and W4-3) were assigned to a psychrophilic subspecies of Neoparamoeba aestuarina, N. aestuarina antarctica n. subsp., one isolate (S-205) was assigned to a new species of Platyamoeba, P. oblongata n. sp., two isolates (W51C#4 & W51C#5) were also assigned to a new species of Platyamoeba, P. contorta n. sp., and one isolate (S-241) was a novel psychrophilic gymnamoeba Vermistella antarctica n. gen. n. sp. Molecular and morphological results revealed that V. antarctica was not related to any described family of gymnamoebae. Strains S-205, W51C#4, and W51C#5 were capable of locomotion at room temperature, while strains SL-200, S-131-2, W4-3, and S-241 exhibited locomotion only below approximately 10 degrees C. Our results imply that the Antarctic environment is host both to cosmopolitan gymnamoebae that have acquired adaptations for existence at low environmental temperature and to apparently novel psychrophilic amoebae described here for the first time.     

Diversity and cold-active hydrolytic enzymes of culturable bacteria associated with Arctic sea ice,Spitzbergen

The diversity of culturable bacteria associated with sea ice from four permanently cold fjords of Spitzbergen, Arctic Ocean, was investigated. A total of 116 psychrophilic and psychrotolerant strains were isolated under aerobic conditions at 4°C. The isolates were grouped using amplified rDNA restriction analysis fingerprinting and identified by partial sequencing of 16S rRNA gene. The bacterial isolates fell in five phylogenetic groups: subclasses and of Proteobacteria, the Bacillus–Clostridium group, the order Actinomycetales, and the Cytophaga–Flexibacter–Bacteroides (CFB) phylum. Over 70% of the isolates were affiliated with the Proteobacteria subclass. Based on phylogenetic analysis (<98% sequence similarity), over 40% of Arctic isolates represent potentially novel species or genera. Most of the isolates were psychrotolerant and grew optimally between 20 and 25°C. Only a few strains were psychrophilic, with an optimal growth at 10–15°C. The majority of the bacterial strains were able to secrete a broad range of cold-active hydrolytic enzymes into the medium at a cultivation temperature of 4°C. The isolates that are able to degrade proteins (skim milk, casein), lipids (olive oil), and polysaccharides (starch, pectin) account for, respectively, 56, 31, and 21% of sea-ice and seawater strains. The temperature dependences for enzyme production during growth and enzymatic activity were determined for two selected enzymes, -amylase and -galactosidase. Interestingly, high levels of enzyme productions were measured at growth temperatures between 4 and 10°C, and almost no production was detected at higher temperatures (20–30°C). Catalytic activity was detected even below the freezing point of water (at –5°C), demonstrating the unique properties of these enzymes.     

Occurrence and some properties of aerobic psychrophilic soil bacteria

Summary The occurrence of aerobic psychrophilic bacteria in different types of soil was studied in regard to all the bacteria present. They constitute a considerable percentage of all the bacteria present in the soils investigated and represent a constant component of the soil microflora, irrespective of the season of the year. They are almost exclusively rod-like, half Gram-negative and half Gram-positive, non-sporulating organisms. It was found, however, that their reaction to Gram-straining varies and depends on the incubation temperature. The temperature of 35°C inhibited the growth of about 40 per cent of the selected psychrophilic strains and caused the development of involutionary forms in 37 per cent of the strains grown. The psychrophilic bacteria isolated from the soils mostly showed ability to decompose urea, and were concerned with ammonification of peptones and reduction of nitrates.Due attention was also given to the problem of adequate incubation temperature in the investigations on soil microorganisms.     

Radiolarian-based transfer functions for estimating paleooceanographic conditions in the south Indian Ocean

A quantitative analysis of 37 radiolarian species in 58 deep-sea surface-sediment samples from the subtropical to the polar regions of the Indian Ocean produced four geographically distinct faunal assemblages (transitional, antarctic, subtropical, subantarctic). Geographic distributions of these assemblages coincide with present-day patterns of sea-surface temperature and water masses. The antarctic factor is almost exclusively found south of today's Antarctic Polar Front. Highest concentrations of the transitional factor are recorded at sites positioned between today's Subtropical Convergence and the Polar Front. The subtropical factor is dominant in sites north of today's Subtropical Convergence.Values of these four faunal assemblages in the surface-sediment samples were regressed onto present-day summer and winter temperatures of the surface waters overlying each of the core-top sites. Resulting transfer functions yield temperature estimates which compare favorably with observed (present-day) summer and winter sea-surface temperatures, with low standard errors of estimate (< ± 1.9°C) and no clear geographic pattern in maps of the residuals (difference between observed and estimated sea-surface temperature).     

Characterization of 15 selected coccal bacteria isolated from antarctic rock and soil samples from the McMurdo-Dry Valleys (South-Victoria Land)

Summary Approximately 1500 cultures of microorganisms were isolated from rocks and soils of the Ross Desert (McMurdo-Dry Valleys). From these, 15 coccoid strains were chosen for more detailed investigation. They were characterized by morphological, physiological and chemotaxonomical properties. All isolates were Grampositive, catalase-positive and nonmotile. Six strains showed red pigmentation and could be identified as members of the genera Micrococcus (M. roseus, M. agilis) or Deinococcus. In spite of their coccoid morphology, the remaining nine strains had to be associated with coryneform bacteria (Arthrobacter, Brevibacterium), because of their cell wall composition and G+C ratios. Most of the strains were psychrotrophic, but one strain was even obligately psychrophilic, with a temperature maximum below 20°C. Red cocci had in vitro pH optima above 9.0 although they generally originated from acid samples. Most isolates showed a preference for sugar alcohols and organic acids, compounds which are commonly known to be released by lichens, molds and algae, the other components of the cryptoendolithic ecosystem. These properties indicate that our strains are autochthonous members of the natural Antarctic microbial population.     

Mesophilic strains of Aeromonas spp. can acquire the multidrug resistance plasmid pRAS1 in horizontal transfer experiments at low temperatures

Both biotic and abiotic characteristics of an ecosystem play an important role in the horizontal transfer of DNA in nature. The abiotic factor temperature has a great impact on such transfers as it controls the metabolic activity of mesophilic microorganisms. Moreover, psychrophilic bacteria, which are not affected by low temperatures, are considered to be potential donors of DNA to mesophilic bacteria under temperature stress conditions. In our study, mesophilic Aeromonas spp. strains isolated from fresh fish were genotypically identified and used as recipients in in vitro conjugal transfer experiments using plasmid pRAS1 from psychrophilic strain Aeromonas salmonicida 718 at three different temperatures (8, 15 and 20 °C). The transfer of the plasmid was confirmed by identifying the elements of the integron in pRAS1. A low temperatures did not prevent the transfer of the pRAS1 plasmid to Aeromonas veronii, A. media, A. hydrophila and A. caviae strains, which showed detectable conjugation frequencies of 10–8 at 8 °C. In other strains of the same species, transconjugants were not detected, which indicated that the genetic background of each strain directly affected the ability to be a recipient of this plasmid at the temperatures tested. Our results demonstrate that mesophilic Aeromonas spp. strains are potential reservoirs of extrachromosomal genetic material. Implications of this plasmid transfer at low temperatures and its possible consequences for human health are discussed.     

Effect of temperature on the growth rates of halotolerant and halophilic bacteria isolated from Antarctic saline lakes

Summary The effect of temperature on the growth rates of Halomonas subglaciescola and a Halobacterium sp., halotolerant and halophilic bacteria isolated from Antarctic saline lakes, was predicted accurately by models which propose a linear relationship between temperature and the square root of the reciprocal of the growth rate. The cardinal temperatures of the strains examined were in general 5–10°C lower than those of taxonomic counterparts isolated from temperate and tropical environments. Temperature dependence data for strains of the Halobacterium sp. and annual temperature profiles of Deep Lake allowed estimation of the in situ potential for growth.     

Influence of temperature on growth rate and competition between two psychrotolerant Antarctic bacteria: low temperature diminishes affinity for substrate uptake.

The growth kinetics of two psychrotolerant Antarctic bacteria, Hydrogenophaga pseudoflava CR3/2/10 (2/10) and Brevibacterium sp. strain CR3/1/15 (1/15), were examined over a range of temperatures in both batch culture and glycerol-limited chemostat cultures. The maximum specific growth rate (mu max) and Ks values for both bacteria were functions of temperature, although the cell yields were relatively constant with respect to temperature. The mu max values of both strains increased up to an optimum temperature, 24 degrees C for 2/10 and 20 degrees C for 1/15. Strain 1/15 might therefore be considered to be more psychrophilic than strain 2/10. For both bacteria, the specific affinity (mu max/Ks) for glycerol uptake was lower at 2 than at 16 degrees C, indicating a greater tendency to substrate limitation at low temperature. As the temperature increased from 2 to 16 degrees C, the specific affinity of 1/15 for glycerol increased more rapidly than it did for 2/10. Thus 1/15, on the basis of this criterion, was less psychrophilic than was 2/10. The steady-state growth kinetics of the two strains at 2 and 16 degrees C imply that 1/15 would be able to outgrow 2/10 only at relatively low substrate concentrations (< 0.32 g of glycerol.liter-1) and high temperatures (> 12 degrees C), which suggests that 1/15 has a less psychrotolerant survival strategy than does 2/10. Our data were compared with other data in the literature for bacteria growing at low temperatures. They also showed an increase of substrate-specific affinity with increasing temperature.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of temperature on growth rates of fungi from subantarctic Macquarie Island and Casey,Antarctica

Summary The linear growth rates of fungal isolates were measured on agar plates at temperatures ranging from 4° to 35°C. Fungi tested included the major fungal colonizers of leaves and litter of the three dominant plant species on subantarctic Macquarie Island, and major fungal species associated with plant and soil communities near Australia's Casey Station on the Antarctic Continent. All fungi grew at 4°C and were classified as psychrotrophs. Maximum growth rates were recorded at temperatures of 10° to 20°C for 13 of the 15 isolates from Macquarie Island and for all six isolates from Casey. Most of the leaf colonizing fungi from Macquarie Island had optimum growth temperatures of 15°C whereas all litter fungi from Macquarie Island and Casey fungi except Thelebolus microsporus had optimum growth temperatures of 20°C or above. Maximum growth of all species was at temperatures above those normally prevailing in their natural environments, with most species growing at 4°C at between 10% and 30% of their maximum rates. However, microclimatic effects may have resulted at times in temperatures near their growth optima. The highest growth rates at 4°C were recorded for Phoma spp. 1 and 2, Phoma exigua and Mortierella gamsii from Macquarie Island and Mortierella sp. 1 from Casey. Thelebolus microsporus and sterile sp. G from Casey also grew relatively fast at 4°C, and these species, and Phoma sp. 3 and Phoma exigua from Macquarie Island had the lowest Q-₁₀ values for the temperature range 4° to 15°C.     

Effects of temperature and day length on the mass balance of Antarctic phytoplankton

Summary Photosynthesis and respiratory carbon losses of freshly collected Antarctic phytoplankton were measured in incubators at 5 temperatures between-2° C and +8°C. The results were used to predict daily growth rates and to define temperature and daylength boundary conditions under which the net balance between photosynthesis and respiration would be positive and allow increases in standing stock. Whereas the Q₁₀ of photosynthesis was 1.4–2.2, the Q₁₀ of respiratory losses exhibited a wide range and higher maxima (2.3–12). Model calculations sugest that under ample light energy supply during long summer days, potential daily growth rates are not severely affected by the low temperatures prevailing in the Southern Ocean. If energy supply is restricted by short days and deep water column mixing, substantially reduced respiration rates at low temperatures may allow the algae to survive.