Project description:The bryozoans Bugula neritina and Bugula simplex harbor bacteria in the pallial sinuses of their larvae as seen by electron microscopy. In B. neritina, the bacterial symbiont has been characterized as a gamma-proteobacterium, "Candidatus Endobugula sertula." "Candidatus E. sertula" has been implicated as the source of the bryostatins, polyketides that provide chemical defense to the host and are also being tested for use in human cancer treatments. In this study, the bacterial symbiont in B. simplex larvae was identified by 16S rRNA-targeted PCR and sequencing as a gamma-proteobacterium closely related to and forming a monophyletic group with "Candidatus E. sertula." In a fluorescence in situ hybridization, a 16S ribosomal DNA probe specific to the B. simplex symbiont hybridized to long rod-shaped bacteria in the pallial sinus of a B. simplex larva. The taxonomic status "Candidatus Endobugula glebosa" is proposed for the B. simplex larval symbiont. Degenerate polyketide synthase (PKS) primers amplified a gene fragment from B. simplex that closely matched a PKS gene fragment from the bryostatin PKS cluster. PCR surveys show that the symbiont and this PKS gene fragment are consistently and uniquely associated with B. simplex. Bryostatin activity assays and chemical analyses of B. simplex extracts reveal the presence of compounds similar to bryostatins. Taken together, these findings demonstrate a symbiosis in B. simplex that is similar and evolutionarily related to that in B. neritina.
Project description:Marine sponges are ancient, sessile, filter-feeding metazoans, which represent a significant component of the benthic communities throughout the world. Sponges harbor a remarkable diversity of bacteria, however, little is known about the functional properties of such bacterial symbionts. In this study, we present the genomic and functional characterization of an uncultured ?-proteobacterium associated with the sponge Cymbastela concentrica. We show that this organism represents a novel phylogenetic clade and propose that it lives in association with a cyanobacterium. We also provide an overview of the predicted functional and ecological properties of this ?-proteobacterium, and discuss its complex interactions with surrounding cells and milieu, including traits of cell attachment, nutrient transport and protein-protein interactions.
Project description:Huanglongbing (yellow dragon disease) is a destructive disease of citrus. The etiological agent is a noncultured, phloem-restricted alpha-proteobacterium, "Candidatus Liberibacter africanus" in Africa and "Candidatus Liberibacter asiaticus" in Asia. In this study, we used an omp-based PCR-restriction fragment length polymorphism (RFLP) approach to analyze the genetic variability of "Ca. Liberibacter asiaticus" isolates. By using five different enzymes, each the 10 isolates tested could be associated with a specific combination of restriction profiles. The results indicate that the species "Ca. Liberibacter asiaticus," even within a given region, may comprise several different variants. Thus, omp-based PCR-RFLP analysis is a simple method for detecting and differentiating "Ca. Liberibacter asiaticus" isolates.
Project description:"Candidatus Glomeribacter gigasporarum" is an endocellular beta-proteobacterium present in the arbuscular mycorrhizal (AM) fungus Gigaspora margarita. We established a protocol to isolate "Ca. Glomeribacter gigasporarum" from its host which allowed us to carry out morphological, physiological, and genomic investigations on purified bacteria. They are rod shaped, with a cell wall typical of gram-negative bacteria and a cytoplasm rich in ribosomes, and they present no flagella or pili. Isolated bacteria could not be grown in any of the 19 culture media tested, but they could be kept alive for up to 4 weeks. PCR-based investigations of purified DNA from isolated bacteria did not confirm the presence of all genes previously assigned to "Ca. Glomeribacter gigasporarum." In particular, the presence of nif genes could not be detected. Pulsed-field gel electrophoresis analyses allowed us to estimate the genome size of "Ca. Glomeribacter gigasporarum" to approximately 1.4 Mb with a ca. 750-kb chromosome and a 600- to 650-kb plasmid. This is the smallest genome known for a beta-proteobacterium. Such small genome sizes are typically found in endocellular bacteria living permanently in their host. Altogether, our data suggest that "Ca. Glomeribacter gigasporarum" is an ancient obligate endocellular bacterium of the AM fungus G. margarita.
Project description:Bacteria are generally assumed to be monoploid (haploid). This assumption is mainly based on generalization of the results obtained with the most intensely studied model bacterium, Escherichia coli (a gamma-proteobacterium), which is monoploid during very slow growth. However, several species of proteobacteria are oligo- or polyploid, respectively. To get a better overview of the distribution of ploidy levels, genome copy numbers were quantified in four species of three different groups of proteobacteria. A recently developed Real Time PCR approach, which had been used to determine the ploidy levels of halophilic archaea, was optimized for the quantification of genome copy numbers of bacteria. Slow-growing (doubling time 103 minutes) and fast-growing (doubling time 25 minutes) E. coli cultures were used as a positive control. The copy numbers of the origin and terminus region of the chromosome were determined and the results were in excellent agreement with published data. The approach was also used to determine the ploidy levels of Caulobacter crescentus (an alpha-proteobacterium) and Wolinella succinogenes (an epsilon-proteobacterium), both of which are monoploid. In contrast, Pseudomonas putida (a gamma-proteobacterium) contains 20 genome copies and is thus polyploid. A survey of the proteobacteria with experimentally-determined genome copy numbers revealed that only three to four of 11 species are monoploid and thus monoploidy is not typical for proteobacteria. The ploidy level is not conserved within the groups of proteobacteria, and there are no obvious correlations between the ploidy levels with other parameters like genome size, optimal growth temperature or mode of life.
Project description:HLB is suggested to be caused by the phloem-limited fastidious prokaryotic α-proteobacterium “Candidatus Liberibacter spp.” Previous studies focused on the proteome and transcriptome analyses of citrus 5 to 35-week-after “Ca. L. spp.” inoculation. In this study, gene expression profiles was analyzed using mandarin of Citrus reticulate Blanco cv. jiaogan leaves after 2-year infection with “Ca. L. asiaticus”. The Affymetrix GeneChip® citrus genome were applied to study the molecular pathways mediated by “Ca. L. asiaticus” inoculated 3-year-old jiaogan seedlings. Each of them was graft-inoculated with one sweet orange scions with or without “Ca. L. asiaticus” in Dectember, 2009. RNA samples from three mandarin trees infected with 'Candidatus Liberibacter asiaticus' and three uninfected trees were used for affymatrix genochip
Project description:A range of leaf symptoms, including blotchy mottle, yellowing, and small, upright leaves with a variety of chlorotic patterns resembling those induced by zinc deficiencies, are associated with huanglongbing (HLB, yellow shoot disease), a worldwide destructive citrus disease. HLB is presumably caused by the phloem-limited fastidious prokaryotic ?-proteobacterium 'Candidatus Liberibacter spp.' Previous studies focused on the proteome and transcriptome analyses of citrus 5 to 35 weeks after 'Ca. L. spp.' inoculation. In this study, gene expression profiles were analyzed from mandarin Citrus reticulate Blanco cv. jiaogan leaves after a 2 year infection with 'Ca. L. asiaticus'. The Affymetrix microarray analysis explored 2,017 differentially expressed genes. Of the 1,364 genes had known functions, 938 (46.5%) were up-regulated. Genes related to photosynthesis, carbohydrate metabolic, and structure were mostly down-regulated, with rates of 92.7%, 61.0%, and 80.2%, respectively. Genes associated with oxidation-reduction and transport were mostly up-regulated with the rates of 75.0% and 64.6%, respectively. Our data analyses implied that the infection of 'Ca. L. asiaticus' could alter hormone crosstalk, inducing the jasmine acid pathway and depressing the ethylene and salicylic acid pathways in the citrus host. This study provides an enhanced insight into the host response of citrus to 'Ca. L. asiaticus' infection at a two-years infection stage.