Number Of Archaea Research Articles

EDNA metabarcoding analysis reveals the consequence of creating ecosystem‐scale refugia from deer grazing for the soil microbial communities

AbstractUngulate overbrowsing is a growing problem in forests worldwide due to its prolonged and pervasive impact on plant biodiversity and ecosystem functioning. It has been shown that overbrowsing not only reduces plant species diversity and biomass (i.e., direct effects) but also causes a loss of associated trophic levels that could potentially feedback to influence plant community structure (i.e., indirect effects). One of the primary pathways of such indirect effects that have not been fully examined is the impact of overbrowsing on soil microorganisms. Recent studies have shown that soil microorganisms maintain vegetation diversity and drive succession, so it is of critical importance to understand how soil microbial communities might be affected by or protected from the deer impact. To assess the consequence of creating artificial grazing refugia on the structure and composition of soil microbial communities, we compared the distribution and abundance of soil microbial taxa (bacteria, archaea, fungi) at the fenced versus unfenced control sites in the context of a catchment‐scale field experiment in Japan. The eDNA metabarcoding analysis of soil microbial communities showed that the numbers of archaea and basidiomycetes fungal species were greater in the fenced site than in the control, while no such pattern was found for bacteria and ascomycetes fungi. Despite the lack of significant influence of the fence treatment on taxonomic composition in the soil fungal communities, their functional guild composition was influenced by the fenced treatment, with significant changes in the abundance of animal pathogens. Thus, although the effect of fencing on soil microbial communities is characterized by complex responses that vary from taxon to taxon, our work suggests that creating ecosystem‐scale refugia from deer overgrazing might help sustain certain, if not all, taxa of soil microbial communities.

Effect of cavitated husks on the metabolome of fermentation end products, microbiome and physicochemical parameters of the rumen (i<i>n vitro</i>)

Relevance. Today, a significant place in the feeding of farm animals is given to such inexpensive waste from the oil extraction industry as sunflower husk. In this regard, one of the promising and modern methods for optimizing feed production can be the use of liquid media activated in various ways, in combination with ultrasonic treatment. Materials and methods. The object of research was crushed sunflower husk (1.0 mm) (sample 1), as well as husk sonicated (cavitated) in hydromodules with water (in the ratio “husk: water” 1 : 3 (sample 2)), and with lactic acid (“husk: lactic acid” 1 : 3 (sample 3)). The obtained samples were used in in vitro studies with rumen fluid in the “Daisy D200I” incubator (“Ankom Technology”, USA). The following indicators of the exchange of nitrogenous metabolites in the cicatricial contents were determined – residual and total nitrogen by the Kjeldahl method according to the method of K. K. Akhazhanova (2016); ammonia nitrogen – by microdiffusion method according to Conway. The taxonomic composition of the scar fluid was determined by NGS-sequencing using the “MiSeq device” (“Illumina”, USA). Research results. Data analysis showed that the use of cavitated sunflower husk increases the digestibility of dry matter by 19.0 % (Р ≤ 0.01), and cavitated together with lactic acid husk – by 20.7 % (Р ≤ 0.01) in comparison with crushed husk. When incubating cavitated sunflower husk and cavitated together with lactic acid sunflower husk, there were a decrease in the number of archaea, in particular the class Methanobacteria, by 23.73 % and 20.36 % (Р ≤ 0.05) and an increase in the number of bacteria of the phyla Bacteroidetes by 10.2 % and 11.27 %, Firmicutes – by 4.66 % and 6.86 % relative to the use of crushed husk.

Unique mobile elements and scalable gene flow at the prokaryote\u2013eukaryote boundary revealed by circularized Asgard archaea genomes

Eukaryotic genomes are known to have garnered innovations from both archaeal and bacterial domains but the sequence of events that led to the complex gene repertoire of eukaryotes is largely unresolved. Here, through the enrichment of hydrothermal vent microorganisms, we recovered two circularized genomes of Heimdallarchaeum species that belong to an Asgard archaea clade phylogenetically closest to eukaryotes. These genomes reveal diverse mobile elements, including an integrative viral genome that bidirectionally replicates in a circular form and aloposons, transposons that encode the 5,000 amino acid-sized proteins Otus and Ephialtes. Heimdallaechaeal mobile elements have garnered various genes from bacteria and bacteriophages, likely playing a role in shuffling functions across domains. The number of archaea- and bacteria-related genes follow strikingly different scaling laws in Asgard archaea, exhibiting a genome size-dependent ratio and a functional division resembling the bacteria- and archaea-derived gene repertoire across eukaryotes. Bacterial gene import has thus likely been a continuous process unaltered by eukaryogenesis and scaled up through genome expansion. Our data further highlight the importance of viewing eukaryogenesis in a pan-Asgard context, which led to the proposal of a conceptual framework, that is, the Heimdall nucleation–decentralized innovation–hierarchical import model that accounts for the emergence of eukaryotic complexity.

Archaea in the microbial community of the reindeer rumen in the Russian Arctic

Archaea is the least studied group of the reindeer rumen microbiocenosis. Although the functional load performed by this group of microorganisms in the rumen is large. Methane-forming archaea play a key role in the process of anaerobic decomposition of organic substances, the formation of methane. This study for the first time analyzed the composition of the archaeal part of the microbial community of the reindeer rumen using the T-RFLP method from various regions of the Russian Arctic. As a result, it was found that according to the estimates of the number of archaea by quantitative PCR in the reindeer rumen in the winter-spring period, on average, 108 genomes/g of archaea were observed in individuals of the Yamalo-Nenets Autonomous District, and 109 genomes/in animals from the Nenets Autonomous District Archean. Thus, in the winter-spring period, a lower number of archaea in the rumen was observed in the Yamalo-Nenets Autonomous District. According to the results of the T-RFLP method, 44 to 134 phylotypes were detected in the archaeal community of the reindeer rumen, the Shannon index was 2.02–3.80. The lowest content (up to 11.10 %) of methanogenic archaea of the Methanomicrobia class (including the families Methanosarcinaceae and Methanocorpusculaceae) was revealed in the Nenets Autonomous District, while their presence in individuals of the Yamalo-Nenets Autonomous District reached 36.33 %. Interestingly, in adults of the Yamalo-Nenets and Nenets Autonomous Districts, a significant decrease in the representation of methanogenic archaea of the Methanomicrobia class was noted by 1.38 (P <0.05) and 2.70 times (P <0.01), respectively, compared with young individuals (up to 2 years).

New insight into the biogeochemical cycling of methane, S and Fe above the Sulfate-Methane Transition Zone in methane hydrate-bearing sediments: A case study in the Dongsha area, South China Sea

Microbially-mediated methane, S and Fe cycling in the sediments can largely reduce the flux of methane from the ocean to the atmosphere. Compared to a large number of records regarding these processes in the sulfate-methane transition zone (SMTZ), detailed information of these processes in the zone above the SMTZ is still required. In this study, a short sediment core DS16 (330 cm) collected from the methane hydrate-bearing Dongsha area, Northern of the South China Sea, was examined by geochemical and microbial methods. Pore water dissolved inorganic carbon (DIC) had an increasing trend with the increasing depth of the sediment, while the value of δ13C-DIC showed a decreasing pattern. The high concentrations of DIC (up to 32.88 mM) and relatively negative δ13C-DIC (down to −30.07‰) indicated the occurrence of anaerobic oxidation of methane (AOM) to some extant above the SMTZ in this region. The variation of concentration profiles of H2S (3.69–12.71 mmol/L) and Fe(III) indicated active S and Fe redox reactions occurred in this zone. Notably, both of the decreasing trends of sulfate and Fe(III) correlated well with those of δ13C-DIC, indicating AOM and organic materials coupled with both sulfate and Fe(III) reduction. Moreover, high-throughput sequencing data showed that a number of archaea (ANME-1) and bacteria (Sulfurovum and Shewanella), which are potential methane-, S- and Fe-metabolic related microorganisms, were detected in these sediments. In addition, the functional genes related to sulfate reduction, sulfur oxidation, and Fe uptake were also detected based on metagenomics analysis. Among them, with the depth increase and Fe(III) concentration decrease, the gene related to Fe3+ uptake mechanism decreased, while the gene related to siderophore uptake mechanism remained constant or even slightly increased. Thus, we propose that bacteria acquired Fe(III) via Fe(III) uptake proteins and siderophore uptake proteins in the Fe-deficient environments for reduction, which then might support the occurrence of Fe-AOM. By integrating these data, we suggest the occurrence of active biogeochemical CH4-S-Fe cycling in the zone above the SMTZ under high methane flux.

Impact of inoculum acclimation on energy recovery and investigation of microbial community changes during anaerobic digestion of the chicken manure

ABSTRACTThe aim of this study was to assess the effect of inoculum adaptation on biogas recovery from two identical lab-scale semi-continuous anaerobic digesters (AD) treating chicken waste (i.e. TS and VS contents of ca. 6.2% and 2.9%, respectively) at mesophilic condition (35°C). For the first two months; one of the AD was run with adapted whereas the second AD was run with unadapted granular sludge to chicken manure which was further operated for about 100 more days. In this scope, qPCR analysis and Illumina sequencing were also used to detect microbial community changes inside anaerobic reactors. Molecular analyses revealed that the number of archaea was significantly higher than that of overall archaea compared to the values obtained at the start-up time and methanogens also increased as the operation continued. On the other hand, although average daily biogas production was about 25% higher in adapted AD compared to the unadapted AD (i.e. biogas yields were ca. 0.6 and 0.7 m3/kg VSfeed, respectively), there was not a meaningful change in archaea numbers at the end of the operation. These suggest that changes in the structure of a microbial community lead to changes in biogas production and controlling ultimate methanogenic archaeal community may promote successful methane production in anaerobic reactors.

Hexamerization of Geranylgeranylglyceryl Phosphate Synthase Ensures Structural Integrity and Catalytic Activity at High Temperatures.

The cell membranes of all archaea contain ether lipids, and a number of archaea are hyperthermophilic. Consequently, the enzymes that catalyze the synthesis of membrane ether lipids had to adopt to these rough conditions. Interestingly, the enzyme that establishes the first ether bond in these lipids, the geranylgeranylglyceryl phosphate synthase (GGGPS), forms hexamers in many hyperthermophilic archaea, while also dimeric variants of this enzyme exist in other species. We used Methanothermobacter thermautotrophicus GGGPS (mtGGGPS) as a model to elucidate the benefit of hexamerization. We studied the oligomerization interfaces in detail by introducing disturbing mutations and subsequently compared the stability and activity of generated dimeric and monomeric variants with the wild-type enzyme. Differential scanning calorimetry revealed a biphasic denaturation of mtGGGPS. The temperature of the first transition varies and rises with increasing oligomerization state. This first phase of denaturation leads to catalytic inactivation, but CD spectroscopy indicated only minor changes on the secondary structure level. The residual part of the fold is extremely thermostable and denatures in a second phase at temperatures >120 °C. The analysis of another distant native GGGPS enzyme affirms these observations. Molecular dynamics simulations revealed three structural elements close to the substrate binding sites with elevated flexibility. We assume that hexamerization might stabilize these structures, and kinetic studies support this hypothesis for the binding pocket of the substrate glycerol 1-phosphate. Oligomerization might thus positively affect the thermostability-flexibility trade-off in GGGPS by allowing a higher intrinsic flexibility of the individual protomers.

Archaeal DNA Replication Origins and Recruitment of the MCM Replicative Helicase.

DNA replication is fundamental to the propagation of all life on the planet. Remarkably, given the central importance for this process, two distinct core cellular DNA replication machineries have evolved. One is found in the bacterial domain of life and the other is present in Archaea and Eukarya. The archaeal machinery represents a simplified and presumably ancestral form of the eukaryotic DNA replication apparatus. As such, archaeal replication proteins have been studied extensively as models for their eukaryal counterparts. In addition, a number of archaea have been developed as model organisms. Accordingly, there has been a considerable increase in our knowledge of how archaeal chromosomes are replicated. It has become apparent that the majority of archaeal cells replicate their genomes from multiple origins per chromosome. Thus, at both organizational and mechanistic levels, archaeal DNA replication resembles that of eukarya. In this chapter, we will describe recent advances in our understanding of the basis of archaeal origin definition and how the archaeal initiator proteins recruit the replicative helicase to origins.

Molecular comparative assessment of the microbial ecosystem in rumen and faeces of goats fed alfalfa hay alone or combined with oats

The objective of this work was to compare the biomass and community structure of bacteria, protozoa and archaea communities in samples of rumen and faeces of goats and to what extent the diet (alfalfa hay with or without supplemented oats) offered to them exert an influence. Four cannulated adult goats fistulated in the rumen were used in a cross over design experiment in two experimental periods of 26 days, consisting in 14 days of adaptation, 7 days of sampling rumen contents and 5 days of digestibility measurement. Bacterial, protozoa and archaeal biomass and the communities' structure was assessed by real time PCR (qPCR) and denaturing gradient gel electrophoresis (DGGE), respectively. The numbers of archaea and bacteria in both rumen and faeces were higher and lower, respectively, in animals fed AH diet (P < 0.005). Contrary, protozoal numbers were not affected by the diet but were lower (P < 0.001) in faeces than in rumen. The analysis of the community structure revealed a consistently different population in structure in rumen and faeces for the three studied microbial groups and that supplementing alfalfa hay with oats led to a decrease in the similarity between sites in the rumen and faeces: similarity indexes for bacteria (57 and 27%), archaea (26 and 9%) and protozoa (62 and 22%) in animals fed AH and AHO diets, respectively.

Geochemistry and Microbial Populations in Sediments of the Northern Baffin Bay, Arctic

The Northern Baffin Bay between Greenland and Canada is a remote Arctic area restricted in primary production by seasonal ice cover, with presumably low sedimentation rates, carbon content and microbial activities in its sediments. Our aim was to study the so far unknown subseafloor geochemistry and microbial populations driving seafloor ecosystems. Shelf sediments had the highest organic carbon content, numbers of Bacteria and Archaea, and microcosms inoculated from Shelf sediments showed highest sulfate reduction and methane production rates. Sediments in the central deep area and on the southern slope contained less organic carbon and overall lower microbial numbers. Similar 16S rRNA gene copy numbers of Archaea and Bacteria were found for the majority of the sites investigated. Sulfate in pore water correlated with dsrA copy numbers of sulfate-reducing prokaryotes and differed between sites. No methane was found as free gas in the sediments, and mcrA copy numbers of methanogenic Archaea were low. Methanogenic and sulfate-reducing cultures were enriched on a variety of substrates including hydrocarbons. In summary, the Greenlandic shelf sediments contain vital microbial communities adapted to their specific environmental conditions.

암모니아 산화 고세균의 독립성장에 필요한 결손 유전자 예측

As a number of archaea are ubiquitously found in non-extreme habitats, elucidation of their functional roles becomes currently an emerging issue. However, most of them are unable to grow in pure culture and so it remains to be established. In order to find genes lacking in the genome of an ammonia-oxidizing archaeon (AOA), we here report on the comparative analyses of an AOA genome with those of experimentally or theoretically established minimal genomes for independent growth. We assessed the genes lacking in AOA using logic of clusters of orthologous groups (COG), remote homology, consensus sequence weight matrix, function-based motif or domain, and then further excluded genes encoding hypothetical orarchaea-specific proteins. The results of these combination analyses revealed 19 candidate genes lacking in the genome of an AOA. Thus, our results provide a possibility of inducing independent growth of AOA when supplemented with product (s) of the lacking gene (s), and also give a chance for finding new proteins with novel sequence or structure space even if the predicted lacking-genes will be found using another algorithms or biochemical studies.

Soil microbial community changes as a result of long-term exposure to a natural CO 2 vent

The capture and geological storage of CO 2 can be used to reduce anthropogenic greenhouse gas emissions. To assess the environmental impact of potential CO 2 leakage from deep storage reservoirs on the abundance and functional diversity of microorganisms in near-surface terrestrial environments, a natural CO 2 vent (>90% CO 2 in the soil gas) was studied as an analogue. The microbial communities were investigated using lipid biomarkers combined with compound-specific stable carbon isotope analyses, the determination of microbial activities, and the use of quantitative polymerase chain reactions (Q-PCR). With this complementary set of methods, significant differences between the CO 2-rich vent and a reference site with a normal CO 2 concentration were detected. The δ 13C values of the plant and microbial lipids within the CO 2 vent demonstrate that substantial amounts of geothermal CO 2 were incorporated into the microbial, plant, and soil carbon pools. Moreover, the numbers of Archaea and Bacteria were highest at the reference site and substantially lower at the CO 2 vent. Lipid biomarker analyses, Q-PCR, and the determination of microbial activities showed the presence of CO 2-utilising methanogenic Archaea, Geobacteraceae, and sulphate-reducing Bacteria (SRB) mainly at the CO 2 vent, only minor quantities were found at the reference site. Stable carbon isotopic analyses revealed that the methanogenic Archaea and SRB utilised the vent-derived CO 2 for assimilatory biosynthesis. Our results show a shift in the microbial community towards anaerobic and acidophilic microorganisms as a consequence of the long-term exposure of the soil environment to high CO 2 concentrations.

The microbial diversity of laboratory-scale wetlands appears to be randomly assembled

This study investigated the formation of the microbial communities in two horizontal subsurface-flow laboratory-scale constructed wetlands, one planted and the other one unplanted. The abundance of the predominant functional groups (Archaea, Bacteria and sulphate-reducing bacteria) was determined using fluorescence in situ hybridization and the diversity and community structure of those functional groups were analysed using denaturing gradient gel electrophoresis. The numbers of Archaea, Bacteria and sulphate-reducing bacteria were indistinguishable in both reactors ( P=0.99, 0.80 and 0.55, respectively). The microbial communities in both wetlands were typically no more similar than if they had been randomly assembled from a common source community. Plants did not appear to exert a strong effect on the structure of the microbial communities in the horizontal subsurface-flow constructed wetlands (HSCWs) studied in this investigation.

Community structure of Archaea and Bacteria in a profundal lake sediment Lake Kinneret (Israel)

The microbial community structure of an anoxic profundal lake sediment, i.e., subtropical Lake Kinneret, was analysed with respect to its composition by culture-independent molecular methods including terminal restriction fragment length polymorphism (T-RFLP) analysis, comparative sequence analysis, and quantitative real-time PCR. In particular we were interested in the structure, species composition, and relative abundance of the overall microbial community in the methanogenic sediment layer (0–10 cm depth). Pairwise comparison of archaeal and bacterial 16S rRNA gene T-RFLP profiles obtained from three independent samplings indicated stability of the microbial community. The numbers of Archaea and Bacteria, quantified by real-time PCR, amounted to about 10 8 and 10 10 16S rRNA gene copies cm −3 sediment, respectively, suggesting that Archaea may account for only a minor fraction (approximately 1%) of the total prokaryotic community. Hydrogenotrophic Methanomicrobiales and acetoclastic Methanosaeta spp. dominated T-RFLP profiles of the archaeal community. T-RFLP profiles of the bacterial community were dominated by Deltaproteobacteria, sulphate reducers and syntrophs in particular. The second most abundant group was assigned to the Bacteroidetes–Chlorobi-group. Only one bacterial group, which was affiliated with halorespiring bacteria of subphylum II of the Chloroflexi, showed variation in abundance within the sediment samples investigated. Our study gives a comprehensive insight into the structure of the bacterial and archaeal community of a profundal lake sediment, indicating that sulphate reducers, syntrophs, bacteroidetes, halorespirers and methanogens are of particular importance in Lake Kinneret sediment.

Dense populations of Archaea associated with the demosponge Tentorium semisuberites Schmidt, 1870 from Arctic deep-waters

The associated microbial community in the mesohyl of the Arctic deep-water sponge Tentorium semisuberites Schmidt, 1870 (Hadromerida, Demospongiae) is dominated by Archaea. This is the result of an integral approach applying analyses of microbial lipid biomarkers as well as microscopic investigations using differential fluorescence in situ hybridisation with universal probes and counterstaining with 4′,6′-diamidino-2-phenylindole (DAPI) on sponge sections based on samples collected in the Greenland Sea in 2001, 2002 and 2005. The distribution of isoprenoidal C40 hydrocarbons of the biphytane series suggests that affiliates of both major archaeal kingdoms, the Crenarchaeota and the Euryarchaeota, are present in the choanosome of T. semisuberites. Positive signals using the oligonucleotide probe ARCH915 indicate high numbers of Archaea in the mesohyl of this sponge. Based on optical estimations 70–90% of all microbial DAPI signals accounted for archaeal cells. Archaea in these high proportions have never been described in an Arctic deep-sea hadromerid sponge, nor in any other demosponge species. Similar observations in specimens collected over a time scale of 4 years suggest permanent sponge-Archaea associations.

The origin of the eukaryotic cell: a genomic investigation.

We have collected a set of 347 proteins that are found in eukaryotic cells but have no significant homology to proteins in Archaea and Bacteria. We call these proteins eukaryotic signature proteins (ESPs). The dominant hypothesis for the formation of the eukaryotic cell is that it is a fusion of an archaeon with a bacterium. If this hypothesis is accepted then the three cellular domains, Eukarya, Archaea, and Bacteria, would collapse into two cellular domains. We have used the existence of this set of ESPs to test this hypothesis. The evidence of the ESPs implicates a third cell (chronocyte) in the formation of the eukaryotic cell. The chronocyte had a cytoskeleton that enabled it to engulf prokaryotic cells and a complex internal membrane system where lipids and proteins were synthesized. It also had a complex internal signaling system involving calcium ions, calmodulin, inositol phosphates, ubiquitin, cyclin, and GTP-binding proteins. The nucleus was formed when a number of archaea and bacteria were engulfed by a chronocyte. This formation of the nucleus would restore the three cellular domains as the Chronocyte was not a cell that belonged to the Archaea or to the Bacteria.

A Single Amidotransferase Forms Asparaginyl-tRNA and Glutaminyl-tRNA in Chlamydia trachomatis

Aminoacyl-tRNA is generally formed by aminoacyl-tRNA synthetases, a family of 20 enzymes essential for accurate protein synthesis. However, most bacteria generate one of the two amide aminoacyl-tRNAs, Asn-tRNA or Gln-tRNA, by transamidation of mischarged Asp-tRNA(Asn) or Glu-tRNA(Gln) catalyzed by a heterotrimeric amidotransferase (encoded by the gatA, gatB, and gatC genes). The Chlamydia trachomatis genome sequence reveals genes for 18 synthetases, whereas those for asparaginyl-tRNA synthetase and glutaminyl-tRNA synthetase are absent. Yet the genome harbors three gat genes in an operon-like arrangement (gatCAB). We reasoned that Chlamydia uses the gatCAB-encoded amidotransferase to generate both Asn-tRNA and Gln-tRNA. C. trachomatis aspartyl-tRNA synthetase and glutamyl-tRNA synthetase were shown to be non-discriminating synthetases that form the misacylated tRNA(Asn) and tRNA(Gln) species. A preparation of pure heterotrimeric recombinant C. trachomatis amidotransferase converted Asp-tRNA(Asn) and Glu-tRNA(Gln) into Asn-tRNA and Gln-tRNA, respectively. The enzyme used glutamine, asparagine, or ammonia as amide donors in the presence of either ATP or GTP. These results suggest that C. trachomatis employs the dual specificity gatCAB-encoded amidotransferase and 18 aminoacyl-tRNA synthetases to create the complete set of 20 aminoacyl-tRNAs.

A dual-specificity aminoacyl-tRNA synthetase in the deep-rooted eukaryote Giardia lamblia.

Cysteinyl-tRNA (Cys-tRNA) is essential for protein synthesis. In most organisms the enzyme responsible for the formation of Cys-tRNA is cysteinyl-tRNA synthetase (CysRS). The only known exceptions are the euryarchaea Methanococcus jannaschii and Methanobacterium thermoautotrophicum, which do not encode a CysRS. Deviating from the accepted concept of one aminoacyl-tRNA synthetase per amino acid, these organisms employ prolyl-tRNA synthetase as the enzyme that carries out Cys-tRNA formation. To date this dual-specificity prolyl-cysteinyl-tRNA synthetase (ProCysRS) is only known to exist in archaea. Analysis of the preliminary genomic sequence of the primitive eukaryote Giardia lamblia indicated the presence of an archaeal prolyl-tRNA synthetase (ProRS). Its proS gene was cloned and the gene product overexpressed in Escherichia coli. By using G. lamblia, M. jannaschii, or E. coli tRNA as substrate, this ProRS was able to form Cys-tRNA and Pro-tRNA in vitro. Cys-AMP formation, but not Pro-AMP synthesis, was tRNA-dependent. The in vitro data were confirmed in vivo, as the cloned G. lamblia proS gene was able to complement a temperature-sensitive E. coli cysS strain. Inhibition studies of CysRS activity with proline analogs (thiaproline and 5'-O-[N-(l-prolyl)-sulfamoyl]adenosine) in a Giardia S-100 extract predicted that the organism also contains a canonical CysRS. This prediction was confirmed by cloning and analysis of the corresponding cysS gene. Like a number of archaea, Giardia contains two enzymes, ProCysRS and CysRS, for Cys-tRNA formation. In contrast, the purified Saccharomyces cerevisiae and E. coli ProRS enzymes were unable to form Cys-tRNA under these conditions. Thus, the dual specificity is restricted to the archaeal genre of ProRS. G. lamblia's archaeal-type prolyl- and alanyl-tRNA synthetases refine our understanding of the evolution and interaction of archaeal and eukaryal translation systems.