Evidence for a global Wolbachia replacement in Drosophila melanogaster

Wolbachia are maternally inherited intracellular α-Proteobacteria found in numerous arthropod and filarial nematode species [1, 2 and 3]. They influence the biology of their hosts in many ways. In some cases, they act as obligate mutualists and are required for the normal development and reproduction of the host [4 and 5]. They are best known, however, for the various reproductive parasitism traits that they can generate in infected hosts. These include cytoplasmic incompatibility (CI) between individuals of different infection status, the parthenogenetic production of females, the selective killing of male embryos, and the feminization of genetic males [1 and 2]. Wolbachia infections of Drosophila melanogaster are extremely common in both wild populations and long-term laboratory stocks [6, 7 and 8]. Utilizing the newly completed genome sequence of Wolbachia pipientis wMel [9], we have identified a number of polymorphic markers that can be used to discriminate among five different Wolbachia variants within what was previously thought to be the single clonal infection of D. melanogaster. Analysis of long-term lab stocks together with wild-caught flies indicates that one of these variants has replaced the others globally within the last century. This is the first report of a global replacement of a Wolbachia strain in an insect host species. The sweep is at odds with current theory that cannot explain how Wolbachia can invade this host species given the observed cytoplasmic incompatibility characteristics of Wolbachia infections in D. melanogaster in the field [6].

Molecular phylogeny of Wolbachia endosymbionts in southeast asian mosquitoes (Diptera: Culicidae) based on wsp gene sequences

Wolbachia are maternally inherited intracellular bacteria that infect a wide range of arthropods and nematodes and are associated with various reproductive abnormalities in their hosts. Insect-associated Wolbachia form a monophyletic clade in the α-Proteobacteria and recently have been separated into two supergroups (A and B) and 19 groups. Our recent polymerase chain reaction (PCR) survey using wsp specific primers indicated that various strains of Wolbachia were present in mosquitoes collected from Southeast Asia. Here, we report the phylogenetic relationship of the Wolbachia strains found in these mosquitoes using wsp gene sequences. Our phylogenetic analysis revealed eight new Wolbachia strains, five in the A supergroup and three in the B supergroup. Most of the Wolbachia strains present in Southeast Asian mosquitoes belong to the established Mors, Con, and Pip groups.

16S rRNA phylogenetic analysis of the bacterial endosymbionts associated with cytoplasmic incompatibility in insects

Bacterial endosymbionts of insects have long been implicated in the phenomenon of cytoplasmic incompatibility, in which certain crosses between symbiont-infected individuals lead to embryonic death or sex ratio distortion. The taxonomic position of these bacteria has, however, not been known with any certainty. Similarly, the relatedness of the bacteria infecting various insect hosts has been unclear. The inability to grow these bacteria on defined cell-free medium has been the major factor underlying these uncertainties. We circumvented this problem by selective PCR amplification and subsequent sequencing of the symbiont 16S rRNA genes directly from infected insect tissue. Maximum parsimony analysis of these sequences indicates that the symbionts belong in the α-subdivision of the Proteobacteria, where they are most closely related to the Rickettsia and their relatives. They are all closely related to each other and are assigned to the type species Wolbachia pipientis. Lack of congruence between the phylogeny of the symbionts and their insect hosts suggests that horizontal transfer of symbionts between insect species may occur. Comparison of the sequences for W. pipientis and for Wolbachia persica, an endosymbiont of ticks, shows that the genus Wolbachia is polyphyletic. A PCR assay based on 16S primers was designed for the detection of W. pipientis in insect tissue, and initial screening of insects indicates that cytoplasmic incompatibility may be a more general phenomenon in insects than is currently recognized.

The phylogenetic relationships of Caulobacter, Asticcacaulis and Brevundimonas species and their taxonomic implications

The phylogenetic relationships among the species of Caulobacter, Asticcacaulis and Brevundimonas were studied by comparison of their 16S rDNA sequences. The analysis of almost complete sequences confirmed the early evolutionary divergence of the freshwater and marine species of Caulobacter reported previously [Stahl, D. A., Key, R,, Flesher, B, & Smit, J. (1992), J Bacteriol 174, 2193-2198]. The freshwater species formed two distinct clusters. One cluster contained the species Caulobacter bacteroides, Caulobacter crescentus, Caulobacter fusiformis and Caulobacter henricii. C, bacteroides and C, fusiformis are very closely related (sequence identity 99.8%). The second cluster was not exclusive and contained the species Caulobacter intermedius, Caulobacter subvibrioides and Caulobacter variabilis, as well as Brevundimonas diminuta and Brevundimonas vesicularis, The marine species Caulobacter halobacteroides and Caulobacter maris were very closely related, with a sequence identity of 99.7%, These two species were most closely but distantly related to the marine hyphal/budding bacteria Hyphomonas jannaschiana and Hirschia baltica, which formed a deep phylogenetic line with Rhodobacter sphaeroides and Rhodobacter capsulatus, Caulobacter leidyia is unrelated to the other species of Caulobacter and belongs to the alpha-4 subclass of the Proteobacteria, forming a distinct cluster with Asticcacaulis excentricus and Asticcacaulis biprosthecium, The taxonomic implications of the polyphyletic nature of the genus Caulobacter and the absence of a type culture for the type species of the genus, Caulobacter vibrioides, are discussed.

Description of Skermanella parooensis gen. nov., sp. nov. to accommodate Conglomeromonas largomobilis subsp. parooensis following the transfer of Conglomeromonas largomobilis subsp. largomobilis to the genus Azospirillum

As a consequence of the transfer of the type species Conglomeromonas largomobilis subsp. largomobilis to the genus Azospirillum, the name of the genus Conglomeromonas must be changed in accordance with Rule 37a(1) of the International Code of Nomenclature of Bacteria. Consequently, it is proposed that the subspecies Conglomeromonas largomobilis subsp, parooensis be transferred to the genus Skermanella gen, nov. as the type species Skermanella parooensis gen, nov., sp, nov. This taxon belongs to an isolated subline of descent in the Azospirillum branch of the alpha-Proteobacteria. The spelling of the specific epithet of Azospirillum largomobile is corrected to Azospirillum largimobile.

Phylogenetic relationships among members of the Comamonadaceae, and description of Delftia acidovorans (den Dooren de Jong 1926 and Tamaoka et al. 1987) gen. nov., comb. nov.

The phylogenetic relationships among members of the family Comamonadaceae and several unclassified strains were studied by direct sequencing of their PCR-amplified 16S rRNA genes. Based on the 16S rRNA gene sequence analysis, members of the family formed a coherent group. The closest relatives are species of the Rubrivivax sub-group: Leptothrix discophora, Ideonella dechloratans and Rubrivivax gelatinosus. The genus Hydrogenophaga formed two subclusters, as did the species of Acidovorax, whereas the five species of the genus [Aquaspirillum] were polyphyletic. Comamonas acidovorans was phylogenetically distant from the type species of Comamonas, Comamonas terrigena. On the basis of this work and previous studies, Comamonas acidovorans is removed from the genus Comamonas and renamed as Delftia acidovorans gen. nov., comb, nov. Descriptions of the new genus Delftia and of the type species Delftia acidovorans, for which the type strain is ATCC 15668(T), are presented.

Description of Gluconacetobacter sacchari sp. nov., a new species of acetic acid bacterium isolated from the leaf sheath of sugar cane and from the pink sugar-cane mealy bug

A new species of the genus Gluconacetobacter, for which the name Gluconacetobacter sacchari sp. nov. is proposed, was isolated from the leaf sheath of sugar cane and from the pink sugar-cane mealy bug, Saccharicoccus sacchari, found on sugar cane growing in Queensland and northern New South Wales, Australia, The nearest phylogenetic relatives in the alpha-subclass of the Proteobacteria are Gluconacetobacter liquefaciens and Gluconacetobacter diazotrophicus, which have 98.8-99.3% and 97.9-98.5% 16S rDNA sequence similarity, respectively, to members of Gluconacetobacter sacchari. On the basis of the phylogenetic positioning of the strains, DNA reassociation studies, phenotypic tests and the presence of the Q10 ubiquinone, this new species was assigned to the genus Gluconacetobacter. No single phenotypic characteristic is unique to the species, but the species can be differentiated phenotypically from closely related members of the acetic acid bacteria by growth in the presence of 0.01% malachite green, growth on 30% glucose, an inability to fix nitrogen and an inability to grow with the L-amino acids asparagine, glycine, glutamine, threonine and tryptophan when D-mannitol was supplied as the sole carbon and energy source. The type strain of this species is strain SRI 1794(T) (= DSM 12717(T)).

A new chemolithoautotrophic arsenite-oxidizing bacterium isolated from a gold mine: Phylogenetic, physiological, and preliminary biochemical studies

A previously unknown chemolithoautotrophic arsenite-oxidizing bacterium has been isolated from a gold mine in the Northern Territory of Australia. The organism, designated NT-26, was found to be a gram-negative motile rod with two subterminal flagella. In a minimal medium containing only arsenite as the electron donor (5 mM), oxygen as the electron acceptor, and carbon dioxide-bicarbonate as the carbon source, the doubling time for chemolithoautotrophic growth was 7.6 h. Arsenite oxidation was found to be catalyzed by a periplasmic arsenite oxidase (optimum pH, 5.5). Based upon 16S rDNA phylogenetic sequence analysis, NT-26 belongs to the Agrobacterium/Rhizbium branch of the alpha-Proteobacteria and may represent a new species. This recently discovered organism is the most rapidly growing chemolithoautotrophic arsenite oxidizer known.

Reassessment of the phylogenetic position of Caulobacter subvibrioides

Determination of the 16S rRNA gene sequence of Caulobacter subvibrioides ATCC 15264(T) (T = type strain) confirmed that this species is a member of the alpha subclass of the Proteobacteria and showed that it is phylogenetically most closely related to the Caulobacter group comprising the species Caulobacter bacteroides, Caulobacter crescentus, and Brevandimonas (Pseudomonas) diminuta, for which 16S rRNA sequences of the type strains are currently available. The closest known relative of strain ATCC 15264(T) among these species is B. diminuta (level of direct pairwise sequence similarity, 95%). On the basis of its previously determined 16S rRNA sequence (accession number M83797), C. subvibrioides is most closely related to Sphingomonas adhaesiva in the alpha-4 subgroup (level of similarity, 97.7%). Analysis of the hydroxy fatty acids of C. subvibrioides ATCC 15264(T) showed that the 2-hydroxymyristic acid which is characteristic of the genus Sphingomonas was absent.

Phylogenetic relationships and uncertain taxonomy of Pedomicrobium species

The phylogenetic relationships among the species of the genus Pedomicrobium were studied by comparing their 16S rRNA sequences, The Pedomicrobium species form a coherent phylogenetic cluster within the genera of the hyphal budding bacteria in the alpha-Proteobacteria. The sequences of two strains of Pedomicrobium australicum were obtained from DNAs extracted from nonviable freeze-dried cells, which are the only source of material available, and were found to be almost identical (level of similarity, 99.9%), Overall, the Pedomicrobium species are closely related, with sequence similarities ranging from 96.2 to 99.9%, Pedomicrobium manganicum is phylogenetically the most distantly related species and exhibits the lowest similarity (96.2%) with Pedomicrobium americanum, Australian isolate Pedomicrobium sp, strain ACM 3067, P, americanum, and P, australicum are all very highly related, with similarities greater than 99%, Pedomicrobium sp, strain ACM 3067 is most closely related to P, australicum (level of similarity, 99.6%) and P, americanum (99.4%), These manganese-oxidizing species are more closely related to the iron-oxidizing species Pedomicrobium ferrugineum than to the other manganese-oxidizing species, P, manganicum. Taxonomic uncertainties resulting from the loss of the type culture of P, australicum are discussed.

Transfer of Blastobacter natatorius (Sly 1985) to the genus Blastomonas gen nov as Blastomonas natatoria comb nov

The budding bacterium Blastobacter natatorius belongs to the alpha-4 group of the Proteobacteria and clusters phylogenetically on a deep branch with Sphingomonas capsulata, with which it shares 93.9% 16S rRNA sequence similarity. On phylogenetic, phenotypic, and chemotaxonomic grounds a proposal is made to transfer B. natatorius to the genus Blastomonas gen, nov. as Blastomonas natatoria comb, nov.

A spongin-boring alpha-proteobacterium is the etiological agent of disease in the Great Barrier Reef sponge Rhopaloeides odorabile

High levels of mortality in the Mediterranean bath sponge industry have raised concerns for the future of sponge farms. Healthy sponges feed predominantly on bacteria, and many harbour a wide diversity of inter- and extra-cellular symbiotic bacteria. Here we describe the first isolation and description of a pathogenic bacterium from an infected marine sponge. Microbiological examination of tissue necrosis in the Great Barrier Reef sponge Rhopaloeides odorabile resulted in isolation of the bacterial strain NW4327. Sponges infected with strain NW4327 exhibited high levels of external tissue necrosis, and the strain was re-isolated from infected sponges. A single morphotype, which had burrowed through the collagenous spongin fibres causing severe necrosis, was observed microscopically. Strain NW4327 was capable of degrading commercial preparations of azo-collagen, providing further evidence of its involvement in spongin fibre necrosis, Strain NW4327 disrupted the microbial community associated with R. odorabile and was able to infect and kill healthy sponge tissue. 16S rRNA sequence analysis revealed that strain NW4327 is a novel member of the alpha-proteobacteria.

Glycogen-accumulating organisms in laboratory-scale and full scale wastewater treatment processes

Laboratory-scale sequencing batch reactors (SBRs) as models for wastewater treatment processes were used to identify glycogen-accumulating organisms (GAOs), which are thought to be responsible for the deterioration of enhanced biological phosphorus removal (EBPR). The SBRs (called Q and T), operated under alternating anaerobic-aerobic conditions typical for EBPR, generated mixed microbial communities (sludges) demonstrating the GAO phenotype. Intracellular glycogen and poly-beta-hydroxyalkanoate (PHA) transformations typical of efficient EBPR occurred but polyphosphate was not bioaccumulated and the sludges contained 1.8% P (sludge Q) and 1.5% P (sludge T). 16S rDNA clone libraries were prepared from DNA extracted from the Q and T sludges. Clone inserts were grouped into operational taxonomic units (OTUs) by restriction fragment length polymorphism banding profiles. OTU representatives were sequenced and phylogenetically analysed. The Q sludge library comprised four OTUs and all six determined sequences were 99.7% identical, forming a cluster in the gamma-Proteobacteria radiation. The T sludge library comprised eight OTUs and the majority of clones were Acidobacteria subphylum 4 (49% of the library) and candidate phylum OPU (39% of the library). One OTU (two clones, of which one was sequenced) was in the gamma-Proteobacteria radiation with 95% sequence identity to the Q sludge clones. Oligonucleotide probes (called GAOQ431 and GAOQ989) were designed from the gamma-Proteobacteria clone sequences for use in fluorescence in situ hybridization (FISH); 92 % of the Q sludge bacteria and 28 % of the T sludge bacteria bound these probes in FISH. FISH and post-FISH chemical staining for PHA were used to determine that bacteria from a novel gamma-Proteobacteria cluster were phenotypically GAOs in one laboratory-scale SBR and two fullscale wastewater treatment plants. It is suggested that the GAOs from the novel cluster in the gamma-Proteobacteria radiation be named 'Candidatus Competibacter phosphatis'.

New arsenite-oxidizing bacteria isolated from Australian gold mining environments - Phylogenetic relationships

Nine novel arsenite-oxidizing bacteria have been isolated from two different gold mine environments in Australia. Four of these organisms grow chemolithoautotrophically with oxygen as the terminal electron acceptor, arsenite as the electron donor, and carbon dioxide-bicarbonate as the sole carbon source. Five heterotrophic arsenite-oxidizing bacteria were also isolated, one of which was found to be both phylogenetically and physiologically identical to the previously described heterotrophic arsenite oxidizer misidentified as Alcaligenes faecalis. The results showed that this strain belongs to the genus Achromobacter. Phylogenetically, the arsenite-oxidizing bacteria fall within two separate subdivisions of the Proteobacteria. Interestingly, the chemolithoautotrophic arsenite oxidizers belong to the alpha-Proteobacteria, whereas the heterotrophic arsenite oxidizers belong to the beta-Proteobacteria.