Publications

4870

Abstract The Miscellaneous Crenarchaeota group (MCG) Archaea is one of the predominant archaeal groups in anoxic environments and may have significant roles in the global biogeochemical cycles. However, no isolate of MCG has been cultivated or characterized to date. In this study, we investigated the genetic organization, ecophysiological properties and evolutionary relationships of MCG archaea with other archaeal members using metagenome information and the result of gene expression experiments. A comparison of the gene organizations and similarities around the 16S rRNA genes from all available MCG fosmid and cosmid clones revealed no significant synteny among genomic fragments, demonstrating that there are large genetic variations within members of the MCG. Phylogenetic analyses of large-subunit+small-subunit rRNA, concatenated ribosomal protein genes and topoisomerases IB gene (TopoIB) all demonstrate that MCG constituted a sister lineage to the newly proposed archaeal phylum Aigarchaeota and Thaumarchaeota. Genes involved in protocatechuate degradation and chemotaxis were found in a MCG fosmid 75G8 genome fragment, suggesting that this MCG member may have a role in the degradation of aromatic compounds. Moreover, the expression of a putative 4-carboxymuconolactone decarboxylase was observed when the sediment was supplemented with protocatechuate, further supporting the hypothesis that this MCG member degrades aromatic compounds.

Sweet cherry (Prunus avium L.) is a deciduous tree originating in the Black Sea/Caspian Sea region where Asia and Europe converge. Being highly valued for its timber and fruit, sweet cherry has been cultivated and naturalized on all continents. Over the past decade, the area of sweet cherry cultivation increased rapidly in China and has reached 140,000 ha. In April 2013, sweet cherry trees (cv. Summit) exhibiting floral virescence symptoms were observed in two orchards located in suburban Taian, Shandong Province, China. The diseased trees developed flowers having white petals with green veins or abnormal floral structures having cupped, green petals. The affected flowers failed to set fruit. A month following the first appearance of the virescence symptoms, the diseased trees became wilted and eventually died. Leaf and stem samples were collected from nine symptomatic and two nearby symptomless trees. Total DNA was extracted from each sample using the Plant Quick DNA Extract Kit (TianGen, Beijing, China). Nested-PCR was carried out using phytoplasma-universal primer pairs P1/P7 and R16F2n/R16R2 (1). All PCR assays with DNA templates from symptomatic samples yielded an amplicon of 1.25 kb, corresponding to the full-length F2nR2 region of phytoplasmal 16S rDNA. No amplicon was generated in PCRs containing DNA templates from symptomless plants. The amplicons were cloned into plasmid vector pMD18-T (TaKaRa, Dalian, China) and sequenced. The obtained sequences were nearly identical, and a representative sequence was deposited into GenBank (Accession No. KF268424). An analysis of the sequence through the iPhyClassifier (4) revealed that the sweet cherry virescence (SCV) disease was associated with infection by a phytoplasma closely related to the reference strain of ‘Candidatus Phytoplasma ziziphi.’ The 16S rDNA F2nR2 region of the SCV phytoplasma shared 99.8% nucleotide sequence identity with that of ‘Candidatus Phytoplasma ziziphi’ reference strain (Accession No. AB052876). A computer-simulated restriction fragment length polymorphism (RFLP) analysis of the SCV phytoplasma 16S rDNA F2nR2 sequence with a set 17 restriction enzymes (3) resulted in a collective RFLP profile identical to the reference pattern of the elm yellows phytoplasma group, subgroup B (16SrV-B). Phytoplasmal diseases of sweet cherry were reported previously in Europe and the etiological agents were phytoplasmas of other groups, including the aster yellows group (16SrI), the X-disease group (16SrIII), and the apple proliferation group (16SrX) (2). To our knowledge, this is the first report of a phytoplasmal disease of sweet cherry in China, and the SCV phytoplasma is a new member of the subgroup 16SrV-B. Presence of 16SrV-B phytoplasmas and their etiological association with various plant diseases in China have been reported previously; affected host plants included jujube, hemp fiber, paper mulberry, Chinese cherry, plum, apricot, red barberry, clover, dianthus, elm, and sunshine tree. Our identification of the SCV phytoplasma expands the known plant host range of the 16SrV-B phytoplasma lineage. The impact of the SCV phytoplasma in the regional ecosystem and in sweet cherry production is being assessed. References: (1) I. M. Lee et al. Int. J. Syst. Bacteriol. 48:1153, 1998. (2) S. Paltrinieri et al. Acta Hort. 550:365, 2001. (3) W. Wei et al. Int. J. Syst. Evol. Microbiol. 57:1855, 2007. (4) Y. Zhao et al. Int. J. Syst. Evol. Microbiol. 59:2582, 2009.