This article has been The list of recently completed microbial genome sequencing projects (Table 1) includes genomes of two unicellular eukaryotes, three archaea and a variety of bacteria, including an unusually diverse selection of the Firmicutes. The highlights of these sequencing efforts include complete genome sequences of several important model organisms, including the standard laboratory strain Escherichia coli DH10B, the model halophile Halobacterium salinarum strain R1, the marine cyanobacterium Synechococcus sp. PCC 7002 and the unicellular green alga Chlamydomonas reinhardtii.
 | Table 1 Recently completed microbial genomes (February–March 2008). |
Arguably, the biggest news was sequencing of the genome of E. coli DH10B (Durfee et al., 2008). Among more than a dozen of E. coli strains with completely sequenced genomes, most are pathogenic and only two, MG1655 and W3110, are derivatives of E. coli K-12. Strain DH10B was constructed at Douglas Hanahan's lab at Cold Spring Harbor Laboratory (Grant et al., 1990) as a derivative of E. coli MC1061 designed to serve as a convenient host for cloning and propagation of foreign DNA. Owing to its unusually high transformation efficiency and the ability to maintain large DNA inserts, DH10B became the strain of choice for many genetic engineering tasks and has been extensively used for preparation of mammalian DNA libraries for whole-genome sequencing. Because of this circumstance, the authors were able to replace most of the sequencing with computational analysis of ~4 million sequence reads collected in the course of the bovine genome sequencing project at Baylor College of Medicine. Bovine BAC DNA preparations were found to contain some (< 1%) DNA contamination from the E. coli DH10B host. These DH10B DNA fragments were identified by comparison to the recently updated genomic sequence of E. coli K12 strain MG1655 (Riley et al., 2006), extracted and assembled into contigs. The genomic finishing phase included identification of the DH10B DNA regions that were absent in the strain MG1655 chromosome and closing the gaps between contigs, which still required some sequencing. After the assembly of Wolbachia genomes from Drosophila sequence reads by Salzberg and colleagues (2005), this work is another impressive example of extracting useful information on bacterial genomes from the massive amounts of sequence data accumulated by the eukaryotic genome sequencing projects.
The genome sequence of E. coli DH10B revealed 226 mutations, a 113 kb tandem duplication and an inversion as compared with the genome of E. coli MG1655 (Durfee et al., 2008). Surprisingly, the presence of deoR mutation in DH10B could not be confirmed, which made the causes of the high transformation efficiency of this strain as obscure as ever before.
In addition to DH10B, two other E. coli genomes have been released in March 2008 and will be used for comparative genome analysis. Escherichia coli strain SECEC SMS-3–5 was isolated from a toxic metal-contaminated coastal site at Shipyard Creek in Charleston, South Carolina. Surprisingly, this environmental strain is highly resistant to a number of antibiotics, including ciprofloxacin and moxifloxacin, which is obviously a cause for great concern, see http://msc.jcvi.org/e_coli_and_shigella/. Escherichia coli C str. ATCC 8739 has an altered outer membrane that lacks the outer membrane porin OmpC and contains only OmpF.
Another important model organism with a recently finished genome is the extremely halophilic archaeon Halobacterium salinarum R1. This organism has been first isolated from salted fish in 1920s and has been known under several names, including Halobacterium halobium. Halobacterium salinarum was used in the famous work of Oesterhelt and Stoeckenius (1971) that discovered bacteriorhodopsin, a 26 kDa protein that comprises the simplest membrane proton pump. Bacteriorhodopsin served as a founding member of a vast family of retinal-binding proteins found in a wide variety of organisms and habitats (Beja et al., 2000; Venter et al., 2004). Sequencing of the H. salinarum R1 genome was performed several years ago, although closing the genome proved impossible at that time owing to the abundance of insertion sequences (Pfeiffer et al., 2008). In contrast, Halobacterium sp. NRC-1, whose genome has been successfully sequenced (Ng et al., 2000), remained taxonomically uncharacterized until 2004 when Gruber and colleagues (2004) showed that it also belongs to H. salinarum. Indeed, the recently completed genome sequence of H. salinarum R1 proved nearly identical to that of Halobacterium sp. NRC-1: most of the observed differences were attributable to the presence of insertion sequences (Pfeiffer et al., 2008). Given the significant body of transcriptomic and proteomic data for H. salinarum R1 (Aivaliotis et al., 2007; Klein et al., 2007; Twellmeyer et al., 2007), the availability of the genome sequence should make it an even more useful model organism.
The unicellular green alga Chlamydomonas reinhardtii is used as a model organism to study photosynthesis, cellular division, intracellular signalling and a variety of other topics. At some point it has even been called ‘the photosynthetic yeast’ (Rochaix, 1995). It has distinct advantages in comparison to higher plants because it is unicellular, haploid and amenable to transformation. It can be grown photoautotrophically or heterotrophically and can be genetically manipulated (Grossman, 2000; 2007). In addition, its genome, as well as the recently released genomes of Monosiga brevicollis and Physcomitrella patens, is extremely interesting from the evolutionary point of view.
Monosiga brevicollis is a representative of a small group of Choanoflagellates, unicellular eukaryotes characterized by a single flagellum surrounded by a collar (choane) of microvilli. Choanoflagellates are very similar to the choanocytes, specialized cells that are found in several animal phyla, including sponges, the most primitive group of Metazoa. This makes them particularly interesting objects for studying the origin of metazoans (King et al., 2008). Monosiga brevicollis genes contain numerous introns and might be used to clarify the origin of introns and their role in metazoan evolution.
The genome of the model moss Physcomitrella patens can be viewed as a bridge from aquatic algae, such as Chlamydomonas, to the flowering plants, such as Arabidopsis, which appeared at least 400 million years later (Rensing et al., 2008).
Another interesting genome that may be important for understanding evolution of life is that of Candidatus Korarchaeum cryptofilum, a member of the candidate division Korarchaeota. This group does not include any cultivated organisms but, based on the 16S rRNA phylogeny, was proposed to form a separate archaeal phylum, distinct from Crenarchaeota, Euryarchaeota and Nanoarchaeota (hence ‘cryptofilum’). Extensive sampling of the Obsidian Pool in Yellowstone National Park in Wyoming allowed collection of sufficiently pure DNA samples to perform the whole-genome sequencing. The completed genome reveals a relatively simple metabolism relying on peptide fermentation. It also confirms that K. cryptofilum represents a deep-branching archaeal lineage with limited similarity to Crenarchaeota, Euryarchaeota or Nanoarchaeota, which probably deserves to be considered a separate archaeal phylum.
The three actinobacteria in the current list are all important pathogens: Clavibacter michiganensis ssp. sepedonicus is a phytopathogen causing the wilt and tuber rot in potato, whereas Corynebacterium urealyticum and Mycobacterium abscessus are both human pathogens that cause, respectively, urinary tract infections and infections of skin and lungs (Ripoll et al., 2007; Tauch et al., 2008).
Clavibacter michiganensis ssp. sepedonicus was first described in 1914 as the causative agent of potato ring rot. It is a close relative of the tomato pathogen Clavibacter michiganensis ssp. michiganensis, whose genome was sequenced in 2007 (Gartemann et al., 2008). However, while C. michiganensis ssp. michiganensis can survive both as an endophyte and an epiphyte, C. michiganensis ssp. sepedonicus appears to be limited to the endophytic lifestyle of a potato pathogen (Bentley et al., 2008). Genome comparisons suggest a recent evolution of C. michiganensis ssp. sepedonicus, which resulted in its adaptation to the potato host and included differential gene gain and loss (Bentley et al., 2008).
Mycobacterium abscessus, first described more than 50 years ago, is a rapidly growing mycobacterium, commonly isolated from soil and water. This organism, formerly known as Mycobacterium chelonae ssp. abscessus (Kusunoki and Ezaki, 1992), is an important emerging pathogen that causes a variety of human infections, including skin, ear, soft tissue and lung infections (Brown-Elliott and Wallace, 2002; Petrini, 2006). Although it belongs to the group of so-called non-tuberculous mycobacteria, M. abscessus can cause a chronic lung infection, similar to tuberculosis, particularly in patients with cystic fibrosis and those undergoing immunosuppressive therapy. Mycobacterium abscessus is resistant to many commonly used antibiotics, which makes treatment very difficult.
The marine cyanobacterium Synechococcus sp. PCC 7002 was originally isolated in 1961 in Puerto Rico. Owing to its ability to grow fast, either phototrophically or heterotrophically on glycerol, and natural transformability, Synechococcus sp. PCC 7002 has become a favourite model organism to study oxygenic photosynthesis (see the Donald Bryant's lab web site http://www.bmb.psu.edu/faculty/bryant/lab/Project/Cyano/ for details).
The second cyanobacterium in the list, Cyanothece sp. ATCC 51142, is an aerobic unicellular marine bacterium that is capable of fixing nitrogen and oxygenic photosynthesis (Reddy et al., 1993). As nitrogenase, the enzyme responsible for N2 fixation, is sensitive to oxygen, photosynthesis and N2 fixation cannot occur in the same cell at the same time. Cyanothece overcomes this conundrum by using a diurnal cycle: oxygenic photosynthesis and CO2 assimilation occur during the day time, while N2 fixation occurs during the night (Schneegurt et al., 1994). This turnover is apparently regulated by the circadian clock system, which makes Cyanothece a good model organism to study the mechanisms of circadian rhythm.
The 1.5 Mbp genome of Acholeplasma laidlawii is the largest mollicute genome sequenced to date and the very first one to be sequenced in Russia. Quite appropriately, in Russian street slang, the organism's genus name means something like ‘Why not?’ Like other mycoplasmas, A. laidlawii is a common parasite of animals but has been found also in association with plants, in soil, water and raw sewage. It is one of the most frequently identified contaminants of insect and mammalian cell culture. While lacking a cell wall, A. laidlawii retains the ability of synthesize fatty acids and glycolipids and does not require exogenous cholesterol, which made it a favourite model organism to study the biophysical properties of biological membranes. Acholeplasma laidlawii genome encodes a number of proteins that are not encoded in other mollicutes. These include, among others, components of a signal transduction machinery with two sensory histidine kinases, three response regulators and 14 proteins with diguanylate cyclase (GGDEF) and/or c-di-GMP-specific phosphodiesterase (EAL) domains, which are all missing in previously sequenced mycoplasmal genomes.
Desulforudis audaxviator has not yet been cultivated but appears to be a dominant organism in the deep subsurface environment (hence the species name, which means ‘bold traveller’ and comes from Jules Verne's ‘Journey to the Center of the Earth’). This sulfate-reducing bacterium has been has been described so far only in a single poster at the ASM General Meeting in 2006 (Chivian et al., 2006) and provisionally assigned to a new genus in the clostridial family Peptococcaceae. Desulforudis audaxviator was first identified in South African gold mines and detected in almost all fracture fluids emanating from depths ranging from 1.5 to 3.2 km below the surface (Onstott et al., 2003). Electron microscopy revealed large cells of up to 4 μm in length. Sequencing the D. audaxviator genome was undertaken after analysis of DNA extracted from a borehole water sample collected at 2.8 km depth showed that that more than 93% of that microbial community was Desulforudis- type cells. Preliminary genome analysis indicated the ability of D. audaxviator to utilize CO and fix N2 (Chivian et al., 2006). The authors speculate that D. audaxviator has retained an ancient mode of metabolism that might sustain life on other planets.
The genome of Heliobacterium modesticaldum is the first complete genome sequence from a phototrophic firmicute. This organism is a representative of the family Heliobacteriaceae, which unifies spore-forming Gram-positive bacteria that are capable of anoxygenic photosynthesis. The genome of closely related Heliobacillus mobilis has been reportedly sequenced by Integrated Genomics, but was never publicly released (Mulkidjanian et al., 2006). Heliobacterium modesticaldum is a moderately thermophilic anaerobe that was first isolated from a microbial mat in Yellowstone hot spring and grows best at 50–56°C (Kimble et al., 1995). This organism is capable of fixing nitrogen and can grow either phototrophically or heterotrophically using pyruvate as a carbon source. The availability of the genome sequence will make H. modesticaldum a potential model organism to study the photosynthetic machinery (see the TGRI web site http://genomes.tgen.org/helio.html for more details). It might also help decipher the evolutionary history of anoxygenic photosynthesis, which remains controversial: some authors suggest that heliobacteria possess ancestral photosynthetic machinery (Woese et al., 1985; Gupta et al., 1999), whereas others believe that heliobacteria acquired it through lateral gene transfer (Mulkidjanian et al., 2006). In addition, the ability of H. modesticaldum to grow phototrophically at elevated temperatures using N2 as nitrogen source makes it attractive for use in biotechnology.
Finegoldia magna, formerly known as Peptostreptococcus magnus, is a member of the Gram-positive anaerobic cocci, part of the normal human bacterial flora that colonizes skin and mucous membranes of the mouth and gastrointestinal tract (Goto et al., 2008). Finegoldia magna is an important opportunistic pathogen that is commonly found in clinical samples from infections of soft tissue, bone and joints. The sequenced strain F. magna ATCC 29328 was originally isolated from an abdominal wound.
The lactic acid bacterium Leuconostoc citreum is used in preparation of various processed foods, such as French cheeses, sauerkraut and pickled cucumbers. Over the past several years, L. citreum strains have been isolated from a variety of traditional ethnic foods, including Moroccan soft white cheese; wheat sourdoughs from Southern Italy; pozol, a Mexican traditional fermented corn beverage; traditional fermented milk in South Africa; fermented bamboo tender shoots in North-east India; som-fak, a low-salt fermented fish product from Thailand, and puto, fermented rice cake popular in the Philippines. The sequenced strain L. citreum KM20 has been isolated from kimchi, a traditional Korean dish made of fermented napa cabbage, white radish and other vegetables and seasoned with garlic, ginger and hot red pepper (Cho et al., 2006). Preliminary analysis of L. citreum genome revealed a variety of carbohydrate transporters and glycoside hydrolases, consistent with fermentation of plant material, as well as a mucin-binding protein, consistent with the ability of L. citreum to function as a probiotic (Kim et al., 2008).
Lysinibacillus sphaericus is the recently adopted name of the well-known soil bacterium Bacillus sphaericus, some strains of which are pathogenic for mosquito larvae and widely used for insect control (Ahmed et al., 2007). As noted earlier, two complete genomes of the insect pathogen Bacillus thuringiensis, serovar konkukian and strain Al Hakam, were sequenced primarily because of their pathogenicity to humans (Han et al., 2006; Challacombe et al., 2007). Thus, L. sphaericus strain C3-41 is the first complete bacillar genome sequenced solely because of its mosquitocidal properties. The genome paper (Hu et al., 2008) offers a detailed analysis of L. sphaericus genome and compares it with genomes of six other firmicutes. This comparison reveals a number of significant differences between L. sphaericus and both B. subtilis and B. anthracis, lending further support to the notion that L. sphaericus should be considered a member of a different genus. Remarkably, the closest relative of L. sphaericus was Bacillus sp. strain NRRL B-14905, isolated from surface waters of the Gulf of Mexico (Siefert et al., 2000), whose unfinished whole-genome shotgun sequence (GenBank accession No. AAXV00000000) has been determined at JCVI.
Two more firmicutes with completely sequenced genomes belong to the genus Thermoanaerobacter. Thermoanaerobacter pseudethanolicus strain 39E has been isolated from an algal-bacterial mat in Octopus Spring in Yellowstone National Park in Wyoming and initially described as Clostridium thermohydrosulfuricum (Zeikus et al., 1980). It was later assigned to Thermoanaerobacter ethanolicus and recently renamed T. pseudethanolicus (Onyenwoke et al., 2007). It is a moderately thermophilic (optimal growth at 65°C) anaerobic bacterium that efficiently ferments carbohydrates into ethanol. The ability of T. pseudethanolicus to metabolize xylose makes it attractive for use in bioconversion of lignocellulose to industrial alcohol.
Thermoanaerobacter sp. X514 is a moderately thermophilic bacterium closely related to Thermoanaerobacter ethanolicus. It has been isolated from the deep subsurface environments of Piceance Basin in Colorado (Roh et al., 2002). This organism grew optimally at 60°C using molecular hydrogen as an electron donor for Fe(III) reduction. It could also reduce a variety of metals, including Fe(III), Co(III), Cr(VI), Mn(IV) and U(VI) when using acetate, lactate, pyruvate, succinate, glucose and xylose as electron donors. Metal reduction led to the precipitation of various minerals. Thus, reduction of Fe(III) oxyhydroxide (FeOOH) at temperatures ranging from ~45°C to 70°C led to the production of magnetite Fe3O4 (Roh et al., 2002).
The next two organisms, the α-proteobacterium Methylobacterium radiotolerans and the β-proteobacterium Cupriavidus taiwanensis, are remarkably similar in their ability to form symbiotic associations with legume roots: they both form root nodules and live there, fixing N2 and providing fixed nitrogen to the host plant. At the end of 2007, JGI scientists released the complete genome sequence of the α-proteobacterial methylotroph Methylobacterium extorquens strain PA1, a member of the Rhizobiales (GenBank accession No. CP000908). That genome has now been followed by genomes of two more members of Methylobacterium spp. Methylobacterium radiotolerans strain JCM 2831 is a facultative symbiont of legumes that is capable of nodulation and nitrogen fixation, whereas Methylobacterium sp. 4–46 apparently is not and will be used for comparative genome analysis.
The nitrogen-fixing β-proteobacterium Cupriavidus taiwanensis strain LMG19424 has been isolated from the root nodules of the legumes Mimosa pudica and Mimosa diplotricha in the southern part of Taiwan and originally named Ralstonia taiwanensis (Chen et al. 2001). It was subsequently renamed Wautersia taiwanensis (Vaneechoutte et al. 2004) and, several months later, Cupriavidus taiwanensis (Vandamme and Coenye, 2004). It is one of several β-proteobacteria found to be capable of root nodule formation and nitrogen fixation (Moulin et al., 2001; Chen et al., 2003). The genes responsible for nodule formation and nitrogen fixation were shown to reside on a 0.5 Mbp plasmid. As C. taiwanensis is only distantly related to nodule-forming α-proteobacteria, analysis of its genome could define the set of genes that are required for efficient nodulation of plant roots.
The β-proteobacterium Polynucleobacter necessarius is an obligate intracellular symbiont of the freshwater ciliate Euplotes aediculatus; the organism has not been cultivated outside the host and the host cells cured from P. necessarius die after one or two cell divisions (Heckmann and Schmidt, 1987). However, close relatives of P. necessarius are found in freshwater habitats all over the world and comprise a large fraction of bacteria in the pelagic zone of surface freshwater (Hahn, 2003). This appears to be a case of a relatively minor sequence divergence between a free-living organism and an obligate endosymbiont (Vannini et al., 2007). Complete genome sequence of a free-living Polynucleobacter strain QLW-P1DMWA-1 has been released by the JGI a year ago (GenBank accession No. CP000655). The completion of the P. necessarius genome offers an opportunity to compare the two and gain important clues on the physiology of this important group of bacteria, as well as the genetic determinants of the intracytoplasmic lifestyle.
The first genome of Acinetobacter baumannii, an obligately aerobic bacterium commonly found in soil, water and sewage, as well as in hospital environment, was sequenced in 2007 (Smith et al., 2007b). Genomes of two more strains of A. baumannii have now been sequenced, an antibiotic-sensitive strain A. baumannii SDF, isolated from body lice collected from homeless people living in France (La Scola and Raoult, 2004), and an antibiotic-resistant strain A. baumannii AYE.
Francisella philomiragia, formerly known as Yersinia philomiragia, is a strictly aerobic γ-proteobacterium found in water and fish. It is an emerging pathogen, infecting humans (and fish) with chronic granulomatous disease (Hollis et al., 1989; Mikalsen et al., 2007). The sequenced strain Francisella philomiragia ssp. philomiragia ATCC 25017 was isolated from water in the Bear River Refuge in Utah. Genome comparison of F. philomiragia and Francisella tularensis should help define the pathogenic mechanisms used by these two related bacteria.
The Shewanella genome sequencing project at the JGI has released complete genomes of two more marine bacteria, Shewanella halifaxensis and Shewanella woodyi. Shewanella halifaxensis has been isolated from the Emerald Basin, an unexploded ordnance-contaminated marine sediment site near the Halifax Harbor in Nova Scotia, Canada (Zhao et al., 2006), together with Shewanella sediminis whose complete genome sequence was released by the JGI several months ago (see Galperin, 2007). Like S. sediminis, S. halifaxensis is capable of metabolizing the explosive agent RDX (hexahydro-1,3,5-trinitro-1,3,5-triazine), which is also known as hexogen, hexolite and cyclonite (see http://pubchem.ncbi.nlm.nih.gov/summary/summary.cgi?cid=8490 for the formula). The periplasmic protein fraction of S. halifaxensis transformed RDX almost as well as whole cells, converting it into nitroso derivatives and/or ring cleavage products such as methylenedinitramine (Zhao et al., 2008). Shewanella halifaxensis is not just an attractive organism for bioremediation of unexploded RDX: it is already hard at work, at least in the Halifax Harbor that gave it its name.
Shewanella woodyi is a bioluminescent bacterium that was isolated from seawater and squid ink samples collected from intermediate depth (200–300 m) in the Alboran Sea between Spain and Morocco. These luminous bacteria were unable to ferment sugars but could grow anaerobically using nitrate or nitrite as terminal electron acceptors. The species name was assigned in honour of J. Woodland (‘Woody’) Hastings, a Harvard University professor and a pioneer in studying bacterial luminescence (Makemson et al., 1997).