Publications

4696

Two Gram-negative, facultatively anaerobic, rod-shaped bacteria, strains EF212Tand PS125T, were isolated from the octocoralsEunicea fuscaandPlexaurasp., respectively. EF212Twas isolated from a specimen ofE. fuscacollected off the coast of Florida, USA, and PS125Twas isolated from a specimen ofPlexaurasp. collected off the coast of Bimini, Bahamas. Analysis of the nearly full-length 16S rRNA gene sequences showed that these novel strains were most closely related toEndozoicomonas montiporaeCL-33T,E. elysicolaMKT110TandE. numazuensisHC50T(EF212T, 95.6–97.2 % identity; PS125T, 95.1–96.4 % identity). DNA–DNA hybridization values among EF212T, PS125T,E. montiporaeLMG 24815TandE. elysicolaKCTC 12372Twere far below the 70 % cut-off, with all values for duplicate measurements being less than 35 %. Both EF212Tand PS125Trequired NaCl for growth and showed optimal growth at 2–3 % NaCl, 22-30 °C and pH 8.0. The predominant cellular fatty acids were summed feature 3 (C16 : 1ω6cand/or C16 : 1ω7c), summed feature 8 (C18 : 1ω6cand/or C18 : 1ω7c), C16 : 0and C14 : 0. The DNA G+C content of EF212Twas 48.6 mol% and that of PS125Twas 47.5 mol%. In addition to the genotypic differences observed between the two novel strains and related type strains, phenotypic and chemotaxonomic experiments also revealed differences between strains. Thus, strains EF212Tand PS125Trepresent novel species of the genusEndozoicomonas, for which the namesEndozoicomonas euniceicolasp. nov. andEndozoicomonas gorgoniicolasp. nov., respectively, are proposed. The type strains are EF212T( = NCCB 100458T = DSM 26535T) forEndozoicomonas euniceicolasp. nov. and PS125T( = NCCB 100438T = CECT 8353T) forEndozoicomonas gorgoniicolasp. nov. An emended description of the genusEndozoicomonasis also provided to encompass differences observed in the results of genotypic, chemotaxonomic and phenotypic tests compared from the original and amended genus descriptions.

Three strictly anaerobic, methanogenic strains JH1T, JH4 and JH8 were isolated from rumen of the Korean native cattle (HanWoo; Bos taurus coreanae) in South Korea. The colonies were circular, opaque, and slightly yellowish. Phylogenetic analyses of 16S rRNA gene and mcrA (encoding α subunit of methyl-coenzyme M reductase) sequences confirmed the affiliation of the novel strains with the Methanobacteriales , and Methanobrevibacter wolinii SHT was the most closely related species. The 16S rRNA gene and mcrA sequence similarities between strains JH1T, JH4 and JH8 and M. wolinii SHT were 96.2 and 89.0 % respectively, and DNA–DNA hybridization of the isolates and M. wolinii DSM 11976T showed a 20 % reassociation. Strain JH1T exhibited 92 % DNA–DNA relatedness with strains JH4 and JH8, and their 16S rRNA gene and mcrA sequences were identical. Cells stained Gram-positive and were non-motile rods, 1.5–1.8 µm long and 0.6 µm wide. The strains were able to use H2/CO2 and formate. The optimum temperature and pH ranges for growth were 37–40 °C and pH 6.5–7.0. The DNA G+C content of strain JH1T was 28 mol%. Based on data from this study using a polyphasic approach, the three strains represent a novel species of genus Methanobrevibacter , for which the name Methanobrevibacter boviskoreani sp. nov. is proposed. The type strain is JH1T ( = KCTC 4102T = JCM 18376T).

The objective of this work was to evaluate the reaction of four sweet orange cultivars expressing the attacin A gene to 'Candidatus Liberibacter asiaticus' (Las) infection, a bacterium associated to huanglongbing (HLB) disease. Transgenic sweet orange plants of Hamlin, Natal, Pêra, and Valência cultivars, as well as nontransgenic controls received inocula by grafting budwood sections of HLB-infected branches. Disease progression was evaluated through observations of leaf symptoms and by polymerase chain reaction (PCR) analysis, eight months after inoculation. A completely randomized design was used, with four experiments (one for each cultivar) performed simultaneously. Bacteria title was estimated by quantitative PCR (qPCR). HLB symptoms and Las titers were present in nontransgenic and transgenic plants expressing the attacin A gene of the four sweet orange cultivars, eight months after bacteria inoculation. Five transgenic lines (transformation events) of 'Pêra' sweet orange expressing the attacin A gene have significantly lower Las titers in comparison with nontransgenic plants of this cultivar.

ABSTRACT “ Candidatus Cardinium hertigii” ( Bacteroidetes ) is a maternally inherited endosymbiont known from several arthropods. Its mechanisms for persistence in host populations are mostly reproductive manipulation, though it has been occasionally reported to improve fitness parameters in several hosts. In Culicoides (Diptera: Ceratopogonidae) biting midges, the prevalence of “ Candidatus Cardinium” infection was documented as moderate, with no detectable sex bias. We therefore investigated whether “ Candidatus Cardinium” affects important fitness parameters, such as survival and body size, in Culicoides imicola , a dominant vector species. Field-collected midges were trapped and analyzed for survival under different environmental conditions and antibiotic treatment, taking into account “ Candidatus Cardinium” infection status and parity status (i.e., parous or nulliparous). Additionally, wing lengths were measured as a proxy parameter for body size and analyzed together with “ Candidatus Cardinium” infection data. The findings revealed no difference in survival of Culicoides infected with “ Candidatus Cardinium” and that of uninfected midges in both parity states and under all tested conditions: optimal, starvation, heat, and antibiotic treatment. Beyond survival, no wing length difference was found for “ Candidatus Cardinium”-infected versus uninfected midges. In aggregate, these findings support our conclusion that “ Candidatus Cardinium” does not have an overt effect on the survival and size of adult C. imicola midges. “ Candidatus Cardinium” may affect immature stages or may alter adult reproductive performance.

A new species of Hassallia (Cyanobacteria, Nostocales, Microchaetaceae) from a supralittoral tropical marine biotope is described. Hassallia littoralis is a false-branched nostocalean cyanobacterium with caespitose free filaments or with fasciculated individual filaments not in a common sheath. Filaments are mainly heteropolar, bearing mono- and bi-pored heterocysts and isopolar or heteropolar hormogonia. The sheath is often widening, with pronounced rounded terminals. This new species is defined according to molecular, morphological and ecological criteria, considering data from different stages of its life cycle as well as the 16S rRNA partial gene sequence.

Tomato (Lycopersicum esculentum) crops grown in several departments of Honduras and heavily infested with the psyllid Bactericera cockerelli were observed in April of 2012 with plant symptoms suggestive of “Candidatus Liberibacter solanacearum” infection. B. cockerelli is a serious pest of potato, tomato, and other solanaceous plants and a vector of “Ca. L. solanacearum” (1,2,3,4). The symptoms included overall chlorosis, severe stunting, leaf cupping, excessive branching of axillary shoots, and leaf purpling and scorching (2,3). Disease incidence ranged from 5 to 50% symptomatic plants per field. Tomato (cv. Pony) plant samples were collected from two psyllid-infested commercial fields in the municipalities of Danli and Comayagua in the departments of El-Paraiso and Comayagua, respectively. Total DNA was extracted from leaf tissues of 50 and 20 symptomatic and asymptomatic plants, respectively, with the cetyltrimethylammonium bromide (CTAB) buffer extraction method (1,3). The DNA samples were tested for “Ca. L. solanacearum” by PCR with primer pairs specific for 16S rDNA (OA2 and OI2c) and the outer membrane protein gene (OMB 1482f and 2086r) of the bacterium (1,2). Ten (20%) of the 50 symptomatic tomato samples were positive for “Ca. L. solanacearum” using both primer pairs and the remaining samples were negative for the bacterium with both primer sets. None of the 20 asymptomatic plants tested positive for “Ca. L. solanacearum”. Amplicons from DNA of two plant samples (one plant/municipality) with each primer pair were cloned and four clones of each of the four amplicons were sequenced. BLASTn analysis of the 16S rDNA consensus sequences from the clones (deposited in GenBank as Accession Nos. KC768321 and KC768322) were identical for both locations and showed 99 to 100% identity to several “Ca. L. solanacearum” sequences in GenBank (e.g., JN848753, JN84856, and HM246509). The OMB consensus sequences from the two tomato plants (deposited in GenBank as KC768329 and KC768330) were 100% identical to OMB sequences of Lso in GenBank (CP002371 and JN48754, respectively). To our knowledge, this is the first report of “Ca. Liberibacter solanacearum” associated with tomato crops in Honduras. This bacterium has caused millions of dollars in losses to the tomato industry in the United States, Mexico, and New Zealand (2,3,4). Serious damages to tomato crops due to “Ca. L. solanacearum” could expand throughout Central America, especially in those countries where B. cockerelli occurs. References: (1) J. M. Crosslin. Southwest. Entomol. 36:125, 2011. (2) L. W. Liefting et al. Plant Dis. 93:208, 2009. (3) J. E. Munyaneza et al. Plant Dis. 93:1076, 2009. (4) J. E. Munyaneza. Am. J. Pot. Res. 89:329, 2012.

Cyanobacteria were responsible for the oxygenation of the ancient atmosphere; however, the evolution of this phylum is enigmatic, as relatives have not been characterized. Here we use whole genome reconstruction of human fecal and subsurface aquifer metagenomic samples to obtain complete genomes for members of a new candidate phylum sibling to Cyanobacteria, for which we propose the designation ‘Melainabacteria’. Metabolic analysis suggests that the ancestors to both lineages were non-photosynthetic, anaerobic, motile, and obligately fermentative. Cyanobacterial light sensing may have been facilitated by regulators present in the ancestor of these lineages. The subsurface organism has the capacity for nitrogen fixation using a nitrogenase distinct from that in Cyanobacteria, suggesting nitrogen fixation evolved separately in the two lineages. We hypothesize that Cyanobacteria split from Melainabacteria prior or due to the acquisition of oxygenic photosynthesis. Melainabacteria remained in anoxic zones and differentiated by niche adaptation, including for symbiosis in the mammalian gut.