Anaerobic microbial communities in Lake Pavin, a unique meromictic lake in France

The Bacteria and Archaea from the meromictic Lake Pavin were analyzed in samples collected along a vertical profile in the anoxic monimolimnion and were compared to those in samples from the oxic mixolimnion. Nine targeted 16S rRNA oligonucleotide probes were used to assess the distribution of Bacteria and Archaea and to investigate the in situ occurrence of sulfate-reducing bacteria and methane-producing Archaea involved in the terminal steps of the anaerobic degradation of organic material. The diversity of the complex microbial communities was assessed from the 16S rRNA polymorphisms present in terminal restriction fragment (TRF) depth patterns. The densities of the microbial community increased in the anoxic layer, and Archaea detected with probe ARCH915 represented the largest microbial group in the water column, with a mean Archaea/Eubacteria ratio of 1.5. Terminal restriction fragment length polymorphism (TRFLP) analysis revealed an elevated archaeal and bacterial phylotype richness in anoxic bottom-water samples. The structure of the Archaea community remained rather homogeneous, while TRFLP patterns for the eubacterial community revealed a heterogeneous distribution of eubacterial TRFs.     

Phylogenetic Analysis of Methanogenic Enrichment Cultures Obtained from Lonar Lake in India: Isolation of <Emphasis Type="Italic">Methanocalculus</Emphasis> sp. and <Emphasis Type="Italic">Methanoculleus</Emphasis> sp.

The diversity of methanogenic archaea in enrichment cultures established from the sediments of Lonar Lake (India), a soda lake having pH ≈ 10, was investigated using 16S rDNA molecular phylogenetic approach. Methanogenic enrichment cultures were developed in a medium that simulated conditions of soda lake with three different substrates viz., H₂:CO₂, sodium acetate, and trimethylamine (TMA), at alkaline pH. Archaeal 16S rRNA clone libraries were generated from enrichment cultures and 13 RFLP groups were obtained. Representative sequence analysis of each RFLP group indicated that the majority of the 16S rRNA gene sequences were phylogenetically affiliated with uncultured Archaea. Some of the groups may belong to new archaeal genera or families. Three RFLP groups were related to Methanoculleus sp, while two related to Methanocalculus sp. 16S rRNA gene sequences found in Lonar Lake were different from sequences reported from other soda lakes and more similar to those of oil reservoirs, palm oil waste treatment digesters, and paddy fields. In culture-based studies, three isolates were obtained. Two of these were related to Methanoculleus sp. IIE1 and one to Methanocalculus sp. 01F97C. These results clearly show that the Lonar Lake ecosystem harbors unexplored methanogens.     

Comparison of an inactive submarine spring with an active nearshore anchialine spring in Florida

Jewfish Sink is a former anchialine karst feature located in the Gulf of Mexico off the coast of West Central Florida. Freshwater flowed from the feature until 1962 and it is now an anoxic marine basin. The current biodiversity within Jewfish Sink was examined in terms of Bacteria, Archaea, and Eukaryota using a combination of 16S and 18S ribosomal RNA analysis from environmental samples. Analysis of 16S rRNA sequences from microbial mats in the anoxic zones revealed a broad diversity of bacteria (265 clones) and archaea (392 clones), many of which had previously been identified in anoxic environmental samples and are likely to be involved with sulfur, nitrogen, and methane metabolism. Sequence analysis of 785 18S clones revealed that fungi and dinoflagellate sequences dominate the eukaryote sequences. Because Jewfish Sink water is anoxic and high in sulfide, we investigated the effect of Jewfish Sink on the nearby shallow benthic environment. We compared the shallow benthic macrofauna near Jewfish Sink with that near Crystal Beach Spring, an active submarine spring. We found significantly greater species richness, abundance, and diversity of benthic fauna near the Jewfish Sink site than near Crystal Beach Spring. This comparison suggests that greater submarine groundwater discharge in an area with active submarine springs is a significant factor reducing the richness and diversity of the benthic community structure in the nearshore, shallow marine environment.     

Molecular analysis of enrichment cultures of ammonia oxidizers from the Salar de Huasco,a high altitude saline wetland in northern Chile

We analyzed enrichment cultures of ammonia-oxidizing bacteria (AOB) collected from different areas of Salar de Huasco, a high altitude, saline, pH-neutral water body in the Chilean Altiplano. Samples were inoculated into mineral media with 10 mM NH₄ ⁺ at five different salt concentrations (10, 200, 400, 800 and 1,400 mM NaCl). Low diversity (up to three phylotypes per enrichment) of beta-AOB was detected using 16S rDNA and amoA clone libraries. Growth of beta-AOB was only recorded in a few enrichment cultures and varied according to site or media salinity. In total, five 16S rDNA and amoA phylotypes were found which were related to Nitrosomonas europaea/Nitrosococcus mobilis, N. marina and N. communis clusters. Phylotype 1-16S was 97% similar with N. halophila, previously isolated from Mongolian soda lakes, and phylotypes from amoA sequences were similar with yet uncultured beta-AOB from different biofilms. Sequences related to N. halophila were frequently found at all salinities. Neither gamma-AOB nor ammonia-oxidizing Archaea were recorded in these enrichment cultures.     

Analysis and comparison of the microbial community structures of two enrichment cultures capable of reductively dechlorinating TCE and cis-DCE

In order to study the effect of different chloroethenes (electron acceptors) on the bacterial composition of dechlorinating communities, two reductive dechlorinating enrichment cultures were developed that were able to reduce trichloroethene (TCE) and cis-1,2-dichloroethene (cis-DCE) to ethene using hydrogen as electron donor, respectively. The inoculum for the cultures was material from a methanogenic fluidized bed reactor (FBR), which was originally seeded with digester sludge and showed a stable capacity for tetrachloroethene (PCE) reduction to ethene for over six years. Molecular methods were used to determine and compare the microbial communities of these two enrichment cultures. A clone library of bacterial 16S rRNA genes was generated for each enrichment. The clones were screened into different groups by restriction fragment length polymorphism (RFLP) analysis using two different four base pair recognition restriction enzymes. A total of 12 sequence types were identified by phylogenetic analysis of nearly complete 16S rDNA sequences ( approximately 1450 bp). The sequences were affiliated with six recognized phyla of the domain Bacteria: Firmicutes (low G+C Gram-positives), Chloroflexi (green non-sulphur bacteria), Actinobacteria (high G+C Gram-positives), Bacteroidetes (Cytophaga-Flexibacter-Bacteroides), Nitrospira and Spirochaetes. The results led to the identification of an organism closely related to Dehalococcoides ethenogenes to be the presumptive dechlorinator in both enrichments. Different electron acceptors affected the bacterial diversity and the community profiles of the two enrichments. Most of the sequences identified in our dechlorinating enrichments shared high similarities with sequences previously obtained from other enriched dechlorinating cultures and chlorinated-compound-contaminated sediments or aquifers, suggesting these bacteria may have direct or indirect roles in reductive dechlorination.     

High 16S rDNA bacterial diversity in glacial meltwater lake sediment,Bratina Island,Antarctica

The microbial diversity in maritime meltwater pond sediments from Bratina Island, Ross Sea, Antarctica was investigated by 16S rDNA-dependent molecular phylogeny. Investigations of the vertical distribution, phylogenetic composition, and spatial variability of Bacteria and Archaea in the sediment were carried out. Results revealed the presence of a highly diverse bacterial population and a significantly depth-related composition. Assessment of 173 partial 16S rDNA clones analyzed by amplified rDNA restriction analysis (ARDRA) using tetrameric restriction enzymes (HinP1I 5'G/CGC3'and Msp I. 5'C/CGG3', BioLabs) revealed 153 different bacterial OTUs (operational taxonomic units). However, only seven archaeal OTUs were detected, indicating low archaeal diversity. Based on ARDRA results, 30 bacterial clones were selected for sequencing and the sequenced clones fell into seven major lineages of the domain Bacteria; the alpha, gamma, and delta subdivisions of Proteobacteria, the Cytophaga-Flavobacterium-Bacteroides, the Spirochaetaceae, and the Actinobacteria. All of the archaeal clones sequenced belonged to the group Crenarchaeota and phylogenetic analysis revealed close relationships with members of the deep-branching Group 1 Marine Archaea.     

Prokaryotic diversity in Zostera noltii-colonized marine sediments

The diversity of microorganisms present in a sediment colonized by the phanerogam Zostera noltii has been analyzed. Microbial DNA was extracted and used for constructing two 16S rDNA clone libraries for Bacteria and Archaea. Bacterial diversity was very high in these samples, since 57 different sequences were found among the 60 clones analyzed. Eight major lineages of the Domain Bacteria were represented in the library. The most frequently retrieved bacterial group (36% of the clones) was delta-Proteobacteria related to sulfate-reducing bacteria. The second most abundant group (27%) was gamma-Proteobacteria, including five clones closely related to S-oxidizing endosymbionts. The archaeal clone library included members of Crenarchaeota and Euryarchaeota, with nine different sequences among the 15 analyzed clones, indicating less diversity when compared to the Bacteria organisms. None of these sequences was closely related to cultured Archaea organisms.     

T-RFLP技术分析油藏微生物多样性

应用T-RFLP(末端限制性片段长度多态性)技术分析和比较了胜利油田单12区块的一口注水井(S12-zhu)和三口采油井(S12-4、S12-5和S12-19)的油藏微生物多样性。基于T-RFLP图谱的多样性指数表明注入水样品具有更丰富的细菌和古菌多样性。相似性指数表明,样品间细菌群落结构的相似性介于22.4%～30.8%之间,古菌群落结构的相似性介于20.8%～34.5%之间。查询RDP数据库推测这4个油藏样品所共有的优势微生物可能为Pseudomonas属,Marinobacter属和产甲烷微生物。T-RFLP技术能方便快捷的分析微生物多样性,其在油藏微生物多样性研究上的应用可以为MEOR提供有用的信息。     

Archaeal habitats--from the extreme to the ordinary

The domain Archaea represents a third line of evolutionary descent, separate from Bacteria and Eucarya. Initial studies seemed to limit archaea to various extreme environments. These included habitats at the extreme limits that allow life on earth, in terms of temperature, pH, salinity, and anaerobiosis, which were the homes to hyper thermo philes, extreme (thermo)acidophiles, extreme halophiles, and methanogens. Typical environments from which pure cultures of archaeal species have been isolated include hot springs, hydrothermal vents, solfataras, salt lakes, soda lakes, sewage digesters, and the rumen. Within the past two decades, the use of molecular techniques, including PCR-based amplification of 16S rRNA genes, has allowed a culture-independent assessment of microbial diversity. Remarkably, such techniques have indicated a wide distribution of mostly uncultured archaea in normal habitats, such as ocean waters, lake waters, and soil. This review discusses organisms from the domain Archaea in the context of the environments where they have been isolated or detected. For organizational purposes, the domain has been separated into the traditional groups of methanogens, extreme halophiles, thermoacidophiles, and hyperthermophiles, as well as the uncultured archaea detected by molecular means. Where possible, we have correlated known energy-yielding reactions and carbon sources of the archaeal types with available data on potential carbon sources and electron donors and acceptors present in the environments. From the broad distribution, metabolic diversity, and sheer numbers of archaea in environments from the extreme to the ordinary, the roles that the Archaea play in the ecosystems have been grossly underestimated and are worthy of much greater scrutiny.     

Abundance, distribution, and activity of Fe(II)-oxidizing and Fe(III)-reducing microorganisms in hypersaline sediments of Lake Kasin, southern Russia

The extreme osmotic conditions prevailing in hypersaline environments result in decreasing metabolic diversity with increasing salinity. Various microbial metabolisms have been shown to occur even at high salinity, including photosynthesis as well as sulfate and nitrate reduction. However, information about anaerobic microbial iron metabolism in hypersaline environments is scarce. We studied the phylogenetic diversity, distribution, and metabolic activity of iron(II)-oxidizing and iron(III)-reducing Bacteria and Archaea in pH-neutral, iron-rich salt lake sediments (Lake Kasin, southern Russia; salinity, 348.6 g liter(-1)) using a combination of culture-dependent and -independent techniques. 16S rRNA gene clone libraries for Bacteria and Archaea revealed a microbial community composition typical for hypersaline sediments. Most-probable-number counts confirmed the presence of 4.26 × 10(2) to 8.32 × 10(3) iron(II)-oxidizing Bacteria and 4.16 × 10(2) to 2.13 × 10(3) iron(III)-reducing microorganisms per gram dry sediment. Microbial iron(III) reduction was detected in the presence of 5 M NaCl, extending the natural habitat boundaries for this important microbial process. Quantitative real-time PCR showed that 16S rRNA gene copy numbers of total Bacteria, total Archaea, and species dominating the iron(III)-reducing enrichment cultures (relatives of Halobaculum gomorrense, Desulfosporosinus lacus, and members of the Bacilli) were highest in an iron oxide-rich sediment layer. Combined with the presented geochemical and mineralogical data, our findings suggest the presence of an active microbial iron cycle at salt concentrations close to the solubility limit of NaCl.     

Archaeal nucleic acids in picoplankton from great lakes on three continents

Phylogenetic analysis of PCR-amplified 16S rRNA genes revealed the presence of archaea in picoplankton collected from the Laurentian Great Lakes in North America, Africas Lake Victoria, and Lakes Ladoga and Onega in northeastern Eurasia. From 1 to 10% of the rRNA extracted from size-fractionated picoplankton (>0.2 µm but <1.2 µm) collected in the epilimnion and hypolimnion of these lakes was specific to the Archaea, whereas the majority of rRNA was derived from Bacteria. Analysis of the 16S rRNA genes cloned from these samples indicated they were closely related to crenarchaeal sequences that have been widely characterized from marine environments. The presence of nearly identical 16S rDNA clones in several of these geographically disparate lakes suggests a cosmopolitan distribution of specific subgroups of these Archaea in freshwater environments. Despite their abundance in the water column of freshwater lakes, we have no representatives of these crenarchaea in pure culture, and so their physiological characteristics and ecological role remain unknown. Present address (B.P. Keough): Department of Microbiology and Molecular Genetics, Bio-Physical Sciences Building, Michigan State University, East Lansing, MI 48823, Phone: (517) 355-6463; Fax: (517) 353-8957     

Putative ammonia-oxidizing Crenarchaeota in suboxic waters of the Black Sea: a basin-wide ecological study using 16S ribosomal and functional genes and membrane lipids

Within the upper 400 m at western, central and eastern stations in the world's largest stratified basin, the Black Sea, we studied the qualitative and quantitative distribution of putative nitrifying Archaea based on their genetic markers (16S rDNA, amoA encoding for the alpha-subunit of archaeal ammonia monooxygenase), and crenarchaeol, the specific glycerol diphytanyl glycerol tetraether of pelagic Crenarchaeota within the Group I.1a. Marine Crenarchaeota were the most abundant Archaea (up to 98% of the total archaeal 16S rDNA copies) in the suboxic layers with oxygen levels as low as 1 microM including layers where previously anammox bacteria were described. Different marine crenarchaeotal phylotypes (both 16S rDNA and amoA) were found at the upper part of the suboxic zone as compared with the base of the suboxic zone and the upper 15-30 m of the anoxic waters with prevailing sulfide concentrations of up to 30 microM. Crenarchaeol concentrations were higher in the sulfidic chemocline as compared with the suboxic zone. These results indicate an abundance of putative nitrifying Archaea at very low oxygen levels within the Black Sea and might form an important source of nitrite for the anammox reaction.     

An integrated study reveals diverse methanogens, Thaumarchaeota, and yet-uncultivated archaeal lineages in Armenian hot springs

Culture-independent and enrichment techniques, with an emphasis on members of the Archaea, were used to determine the composition and structure of microbial communities inhabiting microbial mats in the source pools of two geothermal springs near the towns of Arzakan and Jermuk in Armenia. Amplification of small-subunit rRNA genes using “universal” primers followed by pyrosequencing (pyrotags) revealed highly diverse microbial communities in both springs, with >99 % of pyrosequences corresponding to members of the domain Bacteria. The spring in Arzakan was colonized by a photosynthetic mat dominated by Cyanobacteria, in addition to Proteobacteria, Bacteroidetes, Chloroflexi, Spirochaeta and a diversity of other Bacteria. The spring in Jermuk was colonized by phylotypes related to sulfur, iron, and hydrogen chemolithotrophs in the Betaproteobacteria and Epsilonproteobacteria, along with a diversity of other Bacteria. Analysis of near full-length small subunit rRNA genes amplified using Archaea-specific primers showed that both springs are inhabited by a diversity of methanogens, including Methanomicrobiales and Methanosarcinales and relatives of Methanomassiliicoccus luminyensis, close relatives of the ammonia-oxidizing archaeon (AOA) “Candidatus Nitrososphaera gargensis”, and the yet-uncultivated Miscellaneous Crenarchaeotal Group and Deep Hydrothermal Vent Crenarchaeota group 1. Methanogenic enrichments confirmed the predicted physiological diversity, revealing methylotrophic, acetoclastic, and hydrogenotrophic methanogenesis at 45 and 55 °C, but not 65 °C. This is one of only a few studies combining cultivation-independent and -dependent approaches to study archaea in moderate-temperature (37–73 °C) terrestrial geothermal environments and suggests important roles for methanogenic archaea and AOA in the carbon and nitrogen biogeochemical cycles in these environments.     

Diversity of bacteria and Archaea in sulphate-reducing enrichment cultures inoculated from serial dilution of Zostera noltii rhizosphere samples

We have analysed the diversity of culturable sulphate-reducing bacteria (SRB) in Zostera noltii colonized sediments from Bassin d'Arcachon (France). Four organic substrates have been tested as well as the combination of H2 and CO2 to select for lithotrophic SRB. All energy sources were supplied in parallel cultures that were amended with yeast extract plus NH4+ and prepared without a source of combined nitrogen, the latter to select for diazotrophic SRB. The 10 different enrichment media were inoculated from serial dilution of rhizosphere samples. The highest dilution cultures yielding positive growth (i.e. 10-7) were studied by molecular techniques (16S rDNA clone libraries, RISA and ARDRA). Lactate as a single organic substrate in combination with a source of combined nitrogen resulted in selection of members of the Desulfovibrionaceae. Surprisingly, when lactate was added without a source of combined nitrogen, Desulfobacteriaceae were selected. A strong influence of the presence or absence of combined nitrogen was also observed for the substrates sucrose and fructose. Whereas the liquid culture growing on sucrose and NH4+ systematically yielded 16S rDNA clones related to an environmental unidentified green sulphur bacterium (OPS185), on plates we were able to isolate a SRB related to Desulfovibrio dechloracetivorans, which likely represents a non-described species. Under diazotrophic conditions, sucrose selected for SRB clones related to the cluster formed by Desulfovibrio zosterae, Desulfovibrio salexigens and Desulfovibrio bastinii. The corresponding isolate obtained on plates showed only low sequence similarity with this closest neighbour (93.8%), and we suggest that it also represents a non-described species. Surprisingly, a 16S rDNA sequence corresponding to an archaeon, i.e. a non-extremophile Crenoarchaeota, was retrieved from several of the SRB enrichment cultures even after subsequent transfers.     

Characterization of the microbial diversity in a permafrost sample from the Canadian high Arctic using culture-dependent and culture-independent methods

A combination of culture-dependent and culture-independent methodologies (Bacteria and Archaea 16S rRNA gene clone library analyses) was used to determine the microbial diversity present within a geographically distinct high Arctic permafrost sample. Culturable Bacteria isolates, identified by 16S rRNA gene sequencing, belonged to the phyla Firmicutes, Actinobacteria and Proteobacteria with spore-forming Firmicutes being the most abundant; the majority of the isolates (19/23) were psychrotolerant, some (11/23) were halotolerant, and three isolates grew at -5 degrees C. A Bacteria 16S rRNA gene library containing 101 clones was composed of 42 phylotypes related to diverse phylogenetic groups including the Actinobacteria, Proteobacteria, Firmicutes, Cytophaga - Flavobacteria - Bacteroides, Planctomyces and Gemmatimonadetes; the bacterial 16S rRNA gene phylotypes were dominated by Actinobacteria- and Proteobacteria-related sequences. An Archaea 16S rRNA gene clone library containing 56 clones was made up of 11 phylotypes and contained sequences related to both of the major Archaea domains (Euryarchaeota and Crenarchaeota); the majority of sequences in the Archaea library were related to halophilic Archaea. Characterization of the microbial diversity existing within permafrost environments is important as it will lead to a better understanding of how microorganisms function and survive in such extreme cryoenvironments.     

Purification,overproduction, and partial characterization of beta-RFAP synthase,a key enzyme in the methanopterin biosynthesis pathway

Methanopterin is a folate analog involved in the C1 metabolism of methanogenic archaea, sulfate-reducing archaea, and methylotrophic bacteria. Although a pathway for methanopterin biosynthesis has been described in methanogens, little is known about the enzymes and genes involved in the biosynthetic pathway. The enzyme beta-ribofuranosylaminobenzene 5'-phosphate synthase (beta-RFAP synthase) catalyzes the first unique step to be identified in the pathway of methanopterin biosynthesis, namely, the condensation of p-aminobenzoic acid with phosphoribosylpyrophosphate to form beta-RFAP, CO2, and inorganic pyrophosphate. The enzyme catalyzing this reaction has not been purified to homogeneity, and the gene encoding beta-RFAP synthase has not yet been identified. In the present work, we report on the purification to homogeneity of beta-RFAP synthase. The enzyme was purified from the methane-producing archaeon Methanosarcina thermophila, and the N-terminal sequence of the protein was used to identify corresponding genes from several archaea, including the methanogen Methanococcus jannaschii and the sulfate-reducing archaeon Archaeoglobus fulgidus. The putative beta-RFAP synthase gene from A. fulgidus was expressed in Escherichia coli, and the enzymatic activity of the recombinant gene product was verified. A BLAST search using the deduced amino acid sequence of the beta-RFAP synthase gene identified homologs in additional archaea and in a gene cluster required for C1 metabolism by the bacterium Methylobacterium extorquens. The identification of a gene encoding a potential beta-RFAP synthase in M. extorquens is the first report of a putative methanopterin biosynthetic gene found in the Bacteria and provides evidence that the pathways of methanopterin biosynthesis in Bacteria and Archaea are similar.     

Phylogenetic analysis of aerobic freshwater and marine enrichment cultures efficient in hydrocarbon degradation: effect of profiling method

Aerobically grown enrichment cultures derived from hydrocarbon-contaminated seawater and freshwater sediments were generated by growth on crude oil as sole carbon source. Both cultures displayed a high rate of degradation for a wide range of hydrocarbon compounds. The bacterial species composition of these cultures was investigated by PCR of the 16S rDNA gene using multiple primer combinations. Near full-length 16S rDNA clone libraries were generated and screened by restriction analysis prior to sequence analysis. Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) was carried out using two other PCR primer sets targeting either the V3 or V6-V8 regions, and sequences derived from prominent DGGE bands were compared to sequences obtained via cloning. All data sets suggested that the seawater culture was dominated by alpha-subgroup proteobacteria, whereas the freshwater culture was dominated by members of the beta- and gamma-proteobacteria. However, the V6-V8 primer pair was deficient in the recovery of Sphingomonas-like 16S rDNA due to a 3' terminal mismatch with the reverse primer. Most 16S rDNA sequences recovered from the marine enrichment were not closely related to genera containing known oil-degrading organisms, although some were detected. All methods suggested that the freshwater enrichment was dominated by genera containing known hydrocarbon-degrading species.     

Adaptations to energy stress dictate the ecology and evolution of the Archaea

The three domains of life on Earth include the two prokaryotic groups, Archaea and Bacteria. The Archaea are distinguished from Bacteriabased on phylogenetic and biochemical differences, but currently there is no unifying ecological principle to differentiate these groups. The ecology of the Archaea is reviewed here in terms of cellular bioenergetics. Adaptation to chronic energy stress is hypothesized to be the crucial factor that distinguishes the Archaea from Bacteria. The biochemical mechanisms that enable archaea to cope with chronic energy stress include low-permeability membranes and specific catabolic pathways. Based on the ecological unity and biochemical adaptations among archaea, I propose the hypothesis that chronic energy stress is the primary selective pressure governing the evolution of the Archaea.     

The unique DNA topology and DNA topoisomerases of hyperthermophilic archaea

Abstract: Hyperthermophilic archaea exhibit a unique pattern of DNA topoisomerase activities. They have a peculiar enzyme, reverse gyrase, which introduces positive superturns into DNA at the expense of ATP. This enzyme has been found in all hyperthermophiles tested so far (including Bacteria) but never in mesophiles. Reverse gyrases are formed by the association of a helicase-like domain and a 5'-type I DNA topoisomerase. These two domains might be located on the same polypeptide. However, in the methanogenic archaeon Methanopyrus kandleri , the topoisomerase domain is divided between two subunits. Besides reverse gyrase, Archaea contain other type I DNA topoisomerases; in particular, M. kandleri harbors the only known procaryotic 3'-type I DNA topoisomerase (Topo V). Hyperthermophilic archaea also exhibit specific type II DNA topoisomerases (Topo II), i.e. whereas mesophilic Bacteria have a Topo II that produces negative supercoiling (DNA gyrase), the Topo II from Sulfolobus and Pyrococcus lack gyrase activity and are the smallest enzymes of this type known so far. This peculiar pattern of DNA topoisomerases in hyperthermophilic archaea is paralleled by a unique DNA topology, i.e. whereas DNA isolated from Bacteria and Eucarya is negatively supercoiled, plasmidic DNA from hyperthermophilic archaea are from relaxed to positively supercoiled. The possible evolutionary implications of these findings are discussed in this review. We speculate that gyrase activity in mesophiles and reverse gyrase activity in hyperthermophiles might have originated in the course of procaryote evolution to balance the effect of temperature changes on DNA structure.