Urease gene‐containing Archaea dominate autotrophic ammonia oxidation in two acid soils

The metabolic traits of ammonia‐oxidizing archaea (AOA) and bacteria (AOB) interacting with their environment determine the nitrogen cycle at the global scale. Ureolytic metabolism has long been proposed as a mechanism for AOB to cope with substrate paucity in acid soil, but it remains unclear whether urea hydrolysis could afford AOA greater ecological advantages. By combining DNA‐based stable isotope probing (SIP) and high‐throughput pyrosequencing, here we show that autotrophic ammonia oxidation in two acid soils was predominately driven by AOA that contain ureC genes encoding the alpha subunit of a putative archaeal urease. In urea‐amended SIP microcosms of forest soil (pH 5.40) and tea orchard soil (pH 3.75), nitrification activity was stimulated significantly by urea fertilization when compared with water‐amended soils in which nitrification resulted solely from the oxidation of ammonia generated through mineralization of soil organic nitrogen. The stimulated activity was paralleled by changes in abundance and composition of archaeal amoA genes. Time‐course incubations indicated that archaeal amoA genes were increasingly labelled by ¹³CO₂ in both microcosms amended with water and urea. Pyrosequencing revealed that archaeal populations were labelled to a much greater extent in soils amended with urea than water. Furthermore, archaeal ureC genes were successfully amplified in the ¹³C‐DNA, and acetylene inhibition suggests that autotrophic growth of urease‐containing AOA depended on energy generation through ammonia oxidation. The sequences of AOB were not detected, and active AOA were affiliated with the marine Group 1.1a‐associated lineage. The results suggest that ureolytic N metabolism could afford AOA greater advantages for autotrophic ammonia oxidation in acid soil, but the mechanism of how urea activates AOA cells remains unclear.     

Coenzyme B12-dependent and independent photoregulation of carotenogenesis across Myxococcales

Light-induced carotenogenesis in Myxococcus xanthus is controlled by the B₁₂-based CarH repressor and photoreceptor, and by a separate intricate pathway involving singlet oxygen, the B₁₂-independent CarH paralogue CarA and various other proteins, some eukaryotic-like. Whether other myxobacteria conserve these pathways and undergo photoregulated carotenogenesis is unknown. Here, comparative analyses across 27 Myxococcales genomes identified carotenogenic genes, albeit arranged differently, with carH often in their genomic vicinity, in all three Myxococcales suborders. However, CarA and its associated factors were found exclusively in suborder Cystobacterineae, with carA-carH invariably in tandem in a syntenic carotenogenic operon, except for Cystobacter/Melittangium, which lack CarA but retain all other factors. We experimentally show B₁₂-mediated photoregulated carotenogenesis in representative myxobacteria, and a remarkably plastic CarH operator design and DNA binding across Myxococcales. Unlike the two characterized CarH from other phyla, which are tetrameric, Cystobacter CarH (the first myxobacterial homologue amenable to analysis in vitro) is a dimer that combines direct CarH-like B₁₂-based photoregulation with CarA-like DNA binding and inhibition by an antirepressor. This study provides new molecular insights into B₁₂-dependent photoreceptors. It further establishes the B₁₂-dependent pathway for photoregulated carotenogenesis as broadly prevalent across myxobacteria and its evolution, exclusively in one suborder, into a parallel complex B₁₂-independent circuit.     

gamma 2-Thymidine kinase chimeras are identically transcribed but regulated a gamma 2 genes in herpes simplex virus genomes and as beta genes in cell genomes.

True gamma or gamma 2 genes, unlike alpha, beta, and gamma 1 (beta gamma) genes of herpes simplex virus 1 (HSV-1), stringently require viral DNA synthesis for their expression. We report that gamma 2 genes resident in cells were induced in trans by infection with HSV-1 but that the induction did not require amplification of either the resident gene or the infecting viral genome. Specifically, to test the hypothesis that expression of these genes is amplification dependent, we constructed two sets of gamma 2-thymidine kinase (TK) chimeric genes. The first (pRB3038) consisted of the promoter-regulatory region and a portion of 5'-transcribed noncoding region of the domain of a gamma 2 gene identified by Hall et al. (J. Virol. 43:594-607) in the HSV-1(F) BamHI fragment D' to the 5'-transcribed noncoding and coding regions of the TK gene. The second (pRB3048) contained, in addition, an origin of HSV-1 DNA replication. Cells transfected with either the first or second construct and selected for the TK+ phenotype were then tested for TK induction after superinfection with HSV-1(F) delta 305, containing a deletion in the coding sequences of the TK gene, and viruses containing, in addition, a ts lesion in the alpha 4 regulatory protein (ts502 delta 305) or in the beta 8 major DNA-binding protein (tsHA1 delta 305). The results were as follows: induction by infection with TK- virus of chimeric TK genes with or without an origin of DNA replication was dependent on functional alpha 4 protein but not on viral DNA synthesis; the resident chimeric gene in cells selected for G418 (neomycin) resistance was regulated in the same fashion; the chimeric gene recombined into the viral DNA was regulated as a gamma 2 gene in that its expression in infected cells was dependent on viral DNA synthesis; the gamma 2-chimeric genes resident in the host and in viral genomes were transcribed from the donor BamHI fragment D' containing the promoter-regulatory domain of the gamma 2 gene. The significance of the differential regulation of gamma 2 genes in the environments of host and viral genomes by viral trans-acting factors is discussed.     

Genomic adaptation to eutrophication of ammonia-oxidizing archaea in the Pearl River estuary

Ammonia-oxidizing archaea (AOA) are ubiquitous in natural ecosystems, and they are responsible for a significant fraction of ammonia oxidation globally. Since the first AOA isolate was established a decade ago, molecular surveys of their environmental distribution [based primarily on amplicon sequencing of the amoA, which codes for the alpha subunit of ammonia monooxygenase (AMO)], show that their habitats are believed to range from marine to terrestrial environments. However, the mechanisms of adaptation underpinning to their habitat expansion remain poorly understood. Here, we report that AOA accounts for almost all of the ammonia oxidizers in the shelf water adjacent to the Pearl River estuary (PRE), with the Nitrosopumilus maritimus SCM1-like (SCM1-like) being the main amoA genotype. Using a metagenomic approach, seven high-quality AOA genomes were reconstructed from the PRE. Phylogenetic analysis indicated that four of these genomes with high completeness were closely affiliated with the Nitrosomatrinus catalina strain SPOT01, which was originally isolated off the coast of California. Genomic comparison revealed that the PRE AOA genomes encoded genes functioning in amino acid synthesis, xenobiotic biodegradation metabolism and transportation of inorganic phosphate and heavy metals. This illustrates the different adaptations of AOA in one of the largest estuaries in China, which is strongly influenced by anthropogenic input. Overall, this study provides additional genomic information about estuarine AOA and highlights the importance of their contribution to nitrification in eutrophic coastal environments.     

Structural variation and rates of genome evolution in the grass family seen through comparison of sequences of genomes greatly differing in size

Homology was searched with genes annotated in the Aegilops tauschii pseudomolecules against genes annotated in the pseudomolecules of tetraploid wild emmer wheat, Brachypodium distachyon, sorghum and rice. Similar searches were performed with genes annotated in the rice pseudomolecules. Matrices of collinear genes and rearrangements in their order were constructed. Optical BioNano genome maps were constructed and used to validate rearrangements unique to the wild emmer and Ae. tauschii genomes. Most common rearrangements were short paracentric inversions and short intrachromosomal translocations. Intrachromosomal translocations outnumbered segmental intrachromosomal duplications. The densities of paracentric inversion lengths were approximated by exponential distributions in all six genomes. Densities of collinear genes along the Ae. tauschii chromosomes were highly correlated with meiotic recombination rates but those of rearrangements were not, suggesting different causes of the erosion of gene collinearity and evolution of major chromosome rearrangements. Frequent rearrangements sharing breakpoints suggested that chromosomes have been rearranged recurrently at some sites. The distal 4 Mb of the short arms of rice chromosomes Os11 and Os12 and corresponding regions in the sorghum, B. distachyon and Triticeae genomes contain clusters of interstitial translocations including from 1 to 7 collinear genes. The rates of acquisition of major rearrangements were greater in the large wild emmer wheat and Ae. tauschii genomes than in the lineage preceding their divergence or in the B. distachyon, rice and sorghum lineages. It is suggested that synergy between large quantities of dynamic transposable elements and annual growth habit have been the primary causes of the fast evolution of the Triticeae genomes.     

基于宏基因组研究植物乳酸菌或博落回提取物对山羊回肠微生物合成维生素B12的影响

【目的】探索研究反刍动物胃肠道微生物合成维生素B₁₂的方法,并评估植物乳酸菌或博落回提取物对断奶山羊回肠食靡微生物合成维生素B₁₂的影响。【方法】选取体重相近年龄相仿的断奶黑山羊20只,随机分为对照组(CON, n=7)、乳酸菌组(LAC, n=7)和博落回组(MAC, n=6)。CON组饲喂普通的日粮,LAC组饲喂基础日粮+10 g/d的植物乳酸菌(Lactobacillus plantarum P-8 strains, 4.0×10⁹ CFU/g),MAC组饲喂基础日粮+0.3 g/d的博落回提取物(Macleaya cordata 3.75%)。试验结束后,采集回肠中段食靡样品。利用宏基因组测序技术,比对最新功能基因数据库VB12Path和公共数据库KEGG,分析植物乳酸菌和博落回提取物对山羊回肠食靡微生物合成维生素B₁₂的影响。【结果】结果显示,比对VB12Path数据库共注释到55个与维生素B₁₂合成相关的基因。与CON组相比,LAC组和MAC组中合成维生素B₁₂基因的丰富度和均匀度降低(P<0.05）。3组间基因的β多样性也有显著的差异(P<0.05);比对KEGG数据库共注释到49个与维生素B₁₂合成相关的基因,LAC组的多样性与CON组没有差异,但MAC组的α多样性显著降低(P<0.05)。值得注意的是,比对VB12Path数据库和KEGG数据库均发现LAC组和MAC组中参与前咕啉2合成途径、参与无氧合成途径、有氧合成途径、参与重排转换途径以及腺苷钴胺素合成途径的部分基因(gltX、cbiT、cobT和btuD等)的丰度均显著地高于CON组(P<0.05)。【结论】2个数据库比对后的相似结果表明博落回提取物在对断奶山羊回肠微生物合成维生素B₁₂相关代谢上与植物乳酸菌的作用相似,均可以通过改变其多样性和提高部分关键基因的丰度,从而影响微生物合成维生素B₁₂的潜能,为后期博落回提取物和植物乳酸菌在畜牧养殖中的运用提供一定的理论支撑。此外,2个数据库比对的差异提示未来研究胃肠道微生物维生素B₁₂相关代谢时,应用多个数据库比对,能更全面精确地进行评价,为后期分析过程奠定研究基础和提供新的思路。     

Survival, mutation and capacity to repair single-strand DNA breaks after gamma irradiation in different Exr? strains of Escherichia coli

Summary Strains of Escherichia coli K-12 and B/r made by transduction of the exrA allele from a B_s-2 derivative have been compared with Exr(W)^– strains derived from B_s-1 and B_s-2 by mutation (E.M. Witkin, 1967). Both transduced exrA and Exr(W)^– strains were almost unmutable by gamma radiation, but the former class were as sensitive to gamma radiation as recA strains and, like them, were unable to repair single-strand DNA breaks as detected by the McGrath-Williams technique. In contrast the Exr(W)^–strains were as resistant to gammaradiation as Exr(W)⁺ strains derived from them and were equally efficient in repairing single-strand breaks. The existence of Exr(W)^–strains suggests that the mutagenicity of single-strand breaks may depend entirely on the way in which they are repaired. The properties of the (Exr(W)^–strains cannot be ascribed solely to the transducable exrA allele.A large effect of diffuse daylight in lowering the molecular weight of DNA on alkaline sucrose gradients is described which, unless prevented, may lead to erroneous results in such experiments.     

Genomes of Thaumarchaeota from deep sea sediments reveal specific adaptations of three independently evolved lineages

Marine sediments represent a vast habitat for complex microbiomes. Among these, ammonia oxidizing archaea (AOA) of the phylum Thaumarchaeota are one of the most common, yet little explored, inhabitants, which seem extraordinarily well adapted to the harsh conditions of the subsurface biosphere. We present 11 metagenome-assembled genomes of the most abundant AOA clades from sediment cores obtained from the Atlantic Mid-Ocean ridge flanks and Pacific abyssal plains. Their phylogenomic placement reveals three independently evolved clades within the order Nitrosopumilales, of which no cultured representative is known yet. In addition to the gene sets for ammonia oxidation and carbon fixation known from other AOA, all genomes encode an extended capacity for the conversion of fermentation products that can be channeled into the central carbon metabolism, as well as uptake of amino acids probably for protein maintenance or as an ammonia source. Two lineages encode an additional (V-type) ATPase and a large repertoire of DNA repair systems that may allow to overcome the challenges of high hydrostatic pressure. We suggest that the adaptive radiation of AOA into marine sediments occurred more than once in evolution and resulted in three distinct lineages with particular adaptations to this extremely energy-limiting and high-pressure environment.Subject terms: Phylogenetics, Metagenomics     

Functional redundancy of seasonal vitamin B12 biosynthesis pathways in coastal marine microbial communities

Vitamin B₁₂ (cobalamin) is a major cofactor required by most marine microbes, but only produced by a few prokaryotes in the ocean, which is globally B₁₂-depleted. Despite the ecological importance of B₁₂, the seasonality of B₁₂ metabolisms and the organisms involved in its synthesis in the ocean remain poorly known. Here we use metagenomics to assess the monthly dynamics of B₁₂-related pathways and the functional diversity of associated microbial communities in the coastal NW Mediterranean Sea over 7 years. We show that genes related to potential B₁₂ metabolisms were characterized by an annual succession of different organisms carrying distinct production pathways. During the most productive winter months, archaea (Nitrosopumilus and Nitrosopelagicus) were the main contributors to B₁₂ synthesis potential through the anaerobic pathway (cbi genes). In turn, Alphaproteobacteria (HIMB11, UBA8309, Puniceispirillum) contributed to B₁₂ synthesis potential in spring and summer through the aerobic pathway (cob genes). Cyanobacteria could produce pseudo-cobalamin from spring to autumn. Finally, we show that during years with environmental perturbations, the organisms usually carrying B₁₂ synthesis genes were replaced by others having the same gene, thus maintaining the potential for B₁₂ production. Such ecological insurance could contribute to the long-term functional resilience of marine microbial communities exposed to contrasting inter-annual environmental conditions.     

Spirochete contributions to the eukaryotic genome

Expanded genome/proteome databases and effective use of sequence alignment tools make it possible to trace the phylogeny of individual eukaryotic proteins and ultimately to identify the prokaryotes that contributed to the last eukaryotic common ancestor (LECA). I developed an application of reciprocal BLASTp that identifies (1) the prokaryotic lineages that have contributed to the nuclear genome and (2) the specific proteins acquired from prokaryotic ancestors. Eight complete eubacterial proteomes were analyzed: two free-living spirochetes, two clostridia, two actinobacteria, and two proteobacteria (one alpha and one gamma). The data reveal a spirochete genetic contribution to the eukaryotic genome including essential proteins involved in DNA binding and repair, cyclic nucleotide metabolism, acyltransferase, and signal transduction. My results, consistent with the sulfur syntrophy hypothesis that posits LECA evolved from a merger of spirochetes (eubacteria) with sulfidogenic eocytes (archaebacteria), confirm the contribution of mitochondrial genes from alpha-proteobacteria. A contribution from clostridia to eukaryote genomes was also detected whereas none was seen from either actinobacterium or Escherichia coli. The complete spirochete and clostridial genetic contributions to eukaryotes and those of other eu-and archaebacteria can be identified by this method.     

Ammonia-oxidizing communities in a highly aerated full-scale activated sludge bioreactor: betaproteobacterial dynamics and low relative abundance of Crenarchaea

Ammonia‐oxidizing bacteria (AOB) have long been considered key to the removal of nitrogen in activated sludge bioreactors. Culture‐independent molecular analyses have established that AOB lineages in bioreactors are dynamic, but the underlying operational or environmental factors are unclear. Furthermore, the contribution of ammonia‐oxidizing archaea (AOA) to nitrogen removal in bioreactors has not been studied. To this end, we investigated the abundance of AOA and AOB as well as correlations between dynamics in AOB lineages and operational parameters at a municipal wastewater treatment plant sampled weekly over a 1 year period. Quantitative PCR measurements of bacterial and archaeal ammonia monooxygenase subunit A (amoA) genes revealed that the bacterial homologue predominated by at least three orders of magnitude in all samples. Archaeal amoA was only detectable in ～15% of these samples. Using terminal restriction fragment length polymorphism analysis, we monitored AOB lineages based on amoA genes. The Nitrosomonas europaea lineage and a novel Nitrosomonas‐like cluster were the dominant AOB signatures, with a Nitrosospira lineage present at lower relative abundance. These lineages exhibited strong temporal oscillations, with one becoming sequentially dominant over the other. Using non‐metric multidimensional scaling and redundancy analyses, we tested correlations between terminal restriction fragment length polymorphism profiles and 20 operational and environmental parameters. The redundancy analyses indicated that the dynamics of AOB lineages correlated most strongly with temperature, dissolved oxygen and influent nitrite and chromium. The Nitrosospira lineage signal had a strong negative correlation to dissolved oxygen and temperature, while the Nitrosomonas‐like (negative correlations) and N. europaea lineages (positive correlations) were inversely linked (relative to one another) to influent nitrite and chromium. Overall, this study suggests that AOA may be minor contributors to ammonia oxidation in highly aerated activated sludge, and provides insight into parameters controlling the diversity and dominance of AOB lineages within bioreactors during periods of stable nitrification.     

Alpha beta lineage-specific expression of the alpha T cell receptor gene by nearby silencers

T cells expressing either the alpha beta or gamma delta antigen receptor (TCR) are distinct cell lineages. The single locus encoding the TCR alpha and delta genes requires special regulation to avoid alpha gene expression in gamma delta T cells. We show here that the minimal alpha enhancer is active in the gamma delta T cell lineage but gains alpha beta lineage specificity through negative cis-acting elements 3' of the C alpha gene that silence the enhancer in gamma delta T cells. The negative elements at the C alpha locus consist of several silencers that work in an orientation- and distance-independent fashion. These silencers also act on a retroviral enhancer that is normally ubiquitously expressed, restricting its activity to alpha beta cells. The alpha silencers are active in non-T cell lines, suggesting that the decision of a cell to differentiate into the alpha beta T cell lineage may involve specific relief from these silencers. Silencers are likely to be as important as enhancers in establishing lineage-specific gene expression in many systems.     

Genomes of Two New Ammonia-Oxidizing Archaea Enriched from Deep Marine Sediments

Ammonia-oxidizing archaea (AOA) are ubiquitous and abundant and contribute significantly to the carbon and nitrogen cycles in the ocean. In this study, we assembled AOA draft genomes from two deep marine sediments from Donghae, South Korea, and Svalbard, Arctic region, by sequencing the enriched metagenomes. Three major microorganism clusters belonging to Thaumarchaeota, Epsilonproteobacteria, and Gammaproteobacteria were deduced from their 16S rRNA genes, GC contents, and oligonucleotide frequencies. Three archaeal genomes were identified, two of which were distinct and were designated Ca. “Nitrosopumilus koreensis” AR1 and “Nitrosopumilus sediminis” AR2. AR1 and AR2 exhibited average nucleotide identities of 85.2% and 79.5% to N. maritimus, respectively. The AR1 and AR2 genomes contained genes pertaining to energy metabolism and carbon fixation as conserved in other AOA, but, conversely, had fewer heme-containing proteins and more copper-containing proteins than other AOA. Most of the distinctive AR1 and AR2 genes were located in genomic islands (GIs) that were not present in other AOA genomes or in a reference water-column metagenome from the Sargasso Sea. A putative gene cluster involved in urea utilization was found in the AR2 genome, but not the AR1 genome, suggesting niche specialization in marine AOA. Co-cultured bacterial genome analysis suggested that bacterial sulfur and nitrogen metabolism could be involved in interactions with AOA. Our results provide fundamental information concerning the metabolic potential of deep marine sedimentary AOA.     

Localization of the α7 integrin gene (ITGA7) on human chromosome 12q13: clustering of integrin and hox genes implies parallel evolution of these gene families

Expression of the α7 integrin gene (ITGA7) is developmentally regulated during the formation of skeletal muscle. Increased levels of expression and production of isoforms containing different cytoplasmic and extracellular domains accompany myogenesis. To determine whether a single or multiple α7 genes underlie the structural diversity in this alpha chain that accompanies development, we have examined the rat and human genomes by Southern blotting and in situ hybridization. Our results demonstrate that there is only one α7 gene in both the rat and the human genomes. In the human, ITGA7 is present on chromosome 12q13. Phylogenetic analysis of the integrin alpha chain sequences suggests that the early integrin genes evolved in two pathways to form the I-integrins and the non-I-integrins. The I-integrin alpha chains contain an additional sequence of approximately 180 amino acids and arose as a result of an early insertion into the non-I-gene. The I-chain subfamily further evolved by duplications within the same chromosome. The non-I-integrin alpha chain genes are localized in clusters on chromosomes 2, 12, and 17, and this closely coincides with the localization of the human homeobox gene clusters. Non-I-integrin alpha chain genes appear to have evolved in parallel and in proximity to the Hox clusters. Thus, the Hox genes that underlie the design of body structure and the Integrin genes that underlie informed cell—cell and cell—matrix interactions appear to have evolved in parallel and coordinate fashions.     

Enhancement of gene detection frequencies by combining DNA-based stable-isotope probing with the construction of metagenomic DNA libraries

Summary DNA-based stable-isotope probing (SIP) using ¹³C-labeled growth substrates as bait is a powerful tool for the selective DNA isolation from microorganisms that are actively involved in consuming these substrates. To enhance the detection frequency of target genes in screens for new natural products, we have combined for the first time DNA-based SIP with the construction of metagenomic libraries. To isolate genes encoding coenzyme B₁₂-dependent glycerol dehydratases an enrichment of glycerol-fermenting microorganisms from a sediment sample of the Wadden Sea was performed by using glycerol–¹³C₃ as sole carbon source. Subsequently, the ¹³C-labeled DNA was separated from the naturally abundant ¹²C-DNA by density centrifugation, and used for library generation. Screening of the constructed libraries for the target genes revealed that the gene detection frequencies employing DNA-based SIP for enrichment of genomes harboring dehydratase genes were 2.1- to 3.8-fold higher than those recorded by using a traditional step with unlabeled glycerol for enrichment.     

Production of 3‐hydroxypropionic acid from glycerol by recombinant Klebsiella pneumoniae ΔdhaTΔyqhD which can produce vitamin B12 naturally

3‐Hydroxypropionic acid (3‐HP) is an important platform chemical that can be used to synthesize a range of chemical compounds. A previous study demonstrated that recombinant Escherichia coli stains can produce 3‐HP from glycerol in the presence of vitamin B₁₂ (coenzyme B₁₂), when overexpressed with a coenzyme B₁₂‐dependent glycerol dehydratase (DhaB) and an aldehyde dehydrogenase. The present study examined the production of 3‐HP in recombinant Klebsiella pneumoniae strains, which naturally synthesizes vitamin B₁₂ and does not require supplementation of the expensive vitamin. The NAD⁺‐dependent gamma‐glutamyl‐gamma‐aminobutyraldehyde dehydrogenase (PuuC) of K. pneumoniae alone or with its DhaB was overexpressed homologously, and two major oxidoreductases, DhaT and YqhD, were disrupted. Without vitamin B₁₂ addition, the recombinant K. pneumoniae ΔdhaTΔyqhD overexpressing PuuC could produce ～3.8 g/L 3‐HP in 12 h of flask culture. However, this was possible only under the appropriate aeration conditions; 1,3‐propanediol (1,3‐PDO) (instead of 3‐HP) was mainly produced when aeration was insufficient, whereas a very small amount of both 3‐HP and 1,3‐PDO were produced when aeration was too high. The production of a small amount of 3‐HP under improper aeration conditions was attributed to either slow NAD⁺ regeneration (under low aeration) or reduced vitamin B₁₂ synthesis (under high aeration). In a glycerol fed‐batch bioreactor experiment under a constant DO of 5%, the strain, K. pneumoniae ΔdhaTΔyqhD, overexpressing both PuuC and DhaB could produce >28 g/L 3‐HP in 48 h with a yield of >40% on glycerol. Only small amount of 3‐HP was produced when cultivation was carried out at a constant aeration of 1 vvm or constant 10% DO. These results show that K. pneumoniae is potentially useful for the production of 3‐HP in an economical culture medium that does not require vitamin B₁₂. The results also suggest that the aeration conditions should be optimized carefully for the efficient production of 3‐HP while using this strain. Biotechnol. Bioeng. 2013; 110: 511–524. © 2012 Wiley Periodicals, Inc.     

Rapid structural evolution of Dendrobium mitogenomes and mito-nuclear phylogeny discordances in Dendrobium (Orchidaceae)

Reconstructing mitochondrial genomes of angiosperms is extremely intricate due to frequent recombinations which give rise to varied sized in Dendrobium mitogenomes and their structural variations, even in most orchid species. In this study, we first sequenced two complete and five draft mitochondrial genomes of Dendrobium using next-generation and third-generation sequencing technologies. The mitochondrial genomes were 420 538–689 048 bp long, showing multipartite (multichromosomal) structures that consisted of variably sized circular or linear-mapping isoforms (chromosomes). The comparison of mitochondrial genomes showed frequent gene losses in Dendrobium species. To explore structure variations of mitochondrial genomes in vivo, we quantified copy numbers of five mitochondrial genes and DNA contents per mitochondrion. The gene copy numbers and the DNA contents showed extreme differences during Dendrobium development, suggesting dynamic structures of mitochondrial genomes. Furthermore, phylogenetic relationships of 97 accessions from 39 Dendrobium species were constructed based on 12 nuclear single-copy genes and 15 mitochondrial genes. We discovered obvious discordance between the nuclear and mitochondrial trees. Reticulate evolution was inferred from the species network analysis in Dendrobium. Our findings revealed the rapid structural evolution of Dendrobium mitochondrial genomes and the existence of hybridization events in Dendrobium species, which provided new insights into in vivo structural variations of plant mitochondrial genomes and the strong potential of mitochondrial genes in deciphering plant evolution history.