Publications

4358

Novel halophilic, alkalithermophilic, Gram-type-positive bacterial strains were isolated from sediment of alkaline, hypersaline lakes of the Wadi An Natrun, Egypt. Cells of strain JW/NM-WN-LFTwere rod-shaped, non-spore-forming and non-motile. Strain JW/NM-WN-LFTgrew (at pH55 °C 9.5) between 35 and 56 °C, with an optimum at 53 °C. The pH55 °Crange for growth was 8.3–10.6, with an optimum at pH55 °C 9.5 and no growth at pH55 °C 8.2 or below, or at pH55 °C 10.8 or above. At the optimum pH and temperature, the strain grew in the Na+range of 3.1–4.9 M (1.5–3.3 M added NaCl) and optimally between 3.3 and 3.9 M Na+(1.7–2.3 M added NaCl). Strain JW/NM-WN-LFTutilized fructose, cellobiose, ribose, trehalose, trimethylamine, pyruvate, Casamino acids, acetate, xylose and peptone as carbon and energy sources. Fumarate (20 mM), S2O32−(20 mM), NO3−(20 mM) and iron(III) citrate (20 mM) were utilized as electron acceptors. During growth on sucrose, the isolate produced acetate and formate as major fermentation products. Main cellular fatty acids were iso-branched 15 : 0, i17 : 0 dimethylacetal and 16 : 0 dimethylacetal. The G+C content of genomic DNA was 40.4 mol% (HPLC). On the basis of genotypic and phenotypic characteristics, it is proposed that strain JW/NM-WN-LFTrepresents a novel genus and species,Natranaerobius thermophilusgen. nov., sp. nov. The type strain is JW/NM-WN-LFT(=DSM 18059T=ATCC BAA-1301T). Based on 16S rRNA gene sequence analysis, the strain forms a novel lineage within the class ‘Clostridia’ and clusters with uncultivated bacteria and unidentified strains retrieved from alkaline, hypersaline environments. The phylogenetic data suggest that the lineage represents a novel family,Natranaerobiaceaefam. nov., and order,Natranaerobialesord. nov.

AbstractBackground:  The spectrum of human non‐pylori Helicobacter infections is expanding, with species such as H. heilmannii and H. felis occasionally being associated with gastritis. However, the existence of non‐pylori Helicobacter colonization in asymptomatic subjects has not been evaluated. The aim of this study was to investigate whether Helicobacter species other than pylori are present in the upper digestive tract of asymptomatic human subjects.Materials and methods:  A Helicobacteraceae‐specific semi‐nested polymerase chain reaction (PCR) assay was used to detect Helicobacter‐like organisms in the upper digestive tract of 91 Venezuelan volunteers (aged 18–68 years, 41 females, 50 males). Species were identified by denaturing gradient gel electrophoresis analysis and sequencing of the PCR products.Results:  We detected DNA sharing 99–100% sequence identity in over 300–400 bp with the 16S rRNA genes of H. pylori, H. cetorum, and Candidatus Wolinella africanus in 76%, 16%, and 15% of the subjects, respectively. Multiple colonization was documented in 10% of the subjects: H. cetorum and Candidatus W. africanus (4%), H. pylori and Candidatus W. africanus (4%), and H. pylori and H. cetorum (2%).Conclusions:  Our results suggest that non‐pylori Helicobacteraceae colonization is relatively common in the Venezuelan asymptomatic population. This is the first report documenting the presence of H. cetorum DNA in the human upper digestive tract, and the second report of the recently discovered Candidatus W. africanus.

AbstractA new TaqMan minor groove binding (MGB) probe and new PCR conditions were designed for quick, specific and sensitive detection of ‘Candidatus Phytoplasma mali’. The new probe can distinguish a single mismatch between ‘Ca. P. mali’ and ‘Candidatus Phytoplasma prunorum’, this constituting an improvement over a previously published method. In our study, the relative position of the mismatch in the MGB probe influenced greatly the specificity of detection. Our new real‐time PCR protocol was able to detect one plasmid copy in water and 100 copies in healthy plant DNA extracts. The sensitivity of this new real‐time PCR method, three other real‐time PCR protocols and a conventional PCR with fU5/rU3 primers was compared. Periwinkles and MM106 rootstocks were grafted with infected material and surveyed over time by symptom observation, conventional PCR and real‐time PCR. Phytoplasma infection was detected by symptom observation in all periwinkles within 4 months and in 75% of the MM106 rootstocks by the end of 7 months. Conventional PCR detected phytoplasma infection in all periwinkles within 4 months and in all MM106 rootstocks within 7 months. Best results were obtained by our real‐time PCR, which detected phytoplasma infection in all grafted plants within 3 months after inoculation.

Summary The biodegradation rate of chlorophenols in the environment seems to be limited by a competitive mechanism of O‐methylation which produces chloroanisoles with a high potential of being bioconcentrated in living organisms. In this work we report for the first time the isolation of three soil bacterial strains able to efficiently degrade 2,4,6‐trichloroanisole (2,4,6‐TCA). These strains were identified as Xanthomonas retroflexus INBB4, Pseudomonas putida INBP1 and Acinetobacter radioresistens INBS1. In these isolates 2,4,6‐TCA was efficiently metabolized in a minimal medium containing methanol and 2,4,6‐TCA as the only carbon sources, with a concomitant release of 3 mol of chloride ion from 1 mol of 2,4,6‐TCA, indicating complete dehalogenation of 2,4,6‐TCA. 2,4,6‐trichlorophenol (2,4,6‐TCP) was identified as a degradative intermediate, indicating that 2,4,6‐TCA underwent O‐demethylation as the first step in the biodegradation process. 2,4,6‐TCP was further transformed into 2,6‐dichloro‐ para ‐hydroquinone (2,6‐DCHQ) and subsequently mineralized. The degradation of chloroanisoles could improve the overall biodegradation of chlorophenols in the environment, because those chlorophenols previously biomethylated might also be later biodegraded. Xanthomonas retroflexus INBB4 has two O‐demethylation systems: one is an oxygenase‐type demethylase, and the other is a tetrahydrofolate (THF)‐dependent O ‐demethylase. On the contrary O‐demethylation of 2,4,6‐TCA in P. putida INBP1 is just catalysed by an oxygenase‐type NADH/NADPH‐dependent O ‐demethylase, whereas in A. radioresistens INBS1 a THF‐dependent O ‐demethylase activity was detected.

Summary Recombination is an important process in microbial evolution. Rates of recombination with extracellular DNA matter because models of microbial population structure are profoundly influenced by the degree to which recombination is occurring within the population. Low rates of recombination may be sufficient to ensure the lateral propagation of genes that have a high selective advantage without disrupting the clonal pattern of inheritance for other genes. High rates of recombination potentially can obscure clonal patterns, leading to linkage equilibrium, and give microbial populations a population genetic structure more akin to sexually interbreeding eukaryotic populations. We examined eight loci from nine strains of candidatus Pelagibacter ubique (SAR11), isolated from a single 2L niskin sample of natural seawater, for evidence of genetic recombination between strains. The Shimodaira–Hasegawa test revealed significant phylogenetic incongruence in seven of the genes, indicating that frequent recombination obscures phylogenetic signals from the linear inheritance of genes in this population. Statistical evidence for intragenic recombination was found for six loci. An informative sites matrix showed extensive evidence for a widespread breakdown of linkage disequilibrium. Although the mechanisms of genetic transfer in native SAR11 populations are unknown, we measured recombination rates, ρ , that are much higher than point mutation rates, θ , as a source of genetic diversity in this clade. The eukaryotic model of species sharing a common pool of alleles is more apt for this SAR11 population than a strictly clonal model of inheritance in which allelic diversity is controlled by periodic selection.

Summary Many reports have stated that flagellated protists in termite guts harbour ectosymbiotic spirochetes on their cell surface. In this study, we describe another bristle‐like ectosymbiont affiliated with the order Bacteroidales . The 16S rRNA phylotype Rs‐N74 predominates among Bacteroidales clones obtained from the gut of the termite Reticulitermes speratus . An Rs‐N74 phylotype‐specific probe was designed in this study and used for detection of the corresponding bacteria in the gut by fluorescence in situ hybridization (FISH) analysis. Surprisingly, the signals were detected specifically from the bristle‐like ‘appendages’ of various flagellate species belonging to the genus Dinenympha ; these ‘appendages’ had been believed to be spirochetal ectosymbionts or structures of the protists. The Rs‐N74 bacteria attached to the cell surface of the protists by a tip and coexisted with the spirochetal ectosymbionts. An electron micrograph revealed their morphology to be similar to a typical Bacteroidales bacterium. This bacterium is proposed to represent a novel genus and species, ‘ Candidatus Symbiothrix dinenymphae’, phylogenetically affiliated with a cluster consisting exclusively of uncultured strains from termite guts. A Bacteroidales ‐specific probe for FISH further revealed that this type of symbiosis exists also in various other protists, including parabasalids and oxymonads, and is widespread in termite guts.