Growth conditions. M. hydrocarbonoclasticus was aerobically cultivated in synthetic medium (SM) containing 1.23% (wtivol) Tris, 0.37% (wtivol) NH,Cl, 0.62% (wtkol)

We sequenced nearly complete small-subunit rRNAs of 54 reference strains belonging to the genera Kbrio, Photobacterium, Aeromonas, and Plesiomonas. We then performed a phylogenetic analysis by comparing the sequences which we obtained with all other known sequences for bacteria belonging to the gamma subgroup of the Proteobacteh (thus providing a data base consisting of 70 sequences for the genera investigated), using methods such as neighbor joining, maximum likelihood, and maximum parsimony, as well as bootstrap, to assess the robustness of each topology. Our results confirmed that the family V?riomceae should include only Photobacterium and Kbrio species (but not Hbrio marinus); that Aeromonas species deserve family rank; and that Plesiomonas shigebides is linked to the family Enterobacterheae. The genera Vdrio, Photobacterium, Aeromonas, and Pleswmonas, together with the family Enterobactkrheae, the family Pasteurekeae, and probably the genus Alteromonas, form a robust monophyletic unit within the gamma 3 subgroup of the Proteokteria.rRNA oligomer cataloging (18, 37, 43), DNA-DNA and RNA-DNA hybridization (3,6,18,32), and 5s rRNA sequencing (15,27,28) have been used previously to study the genera Vibrio, Photobacterium, Aeromonas, and Plesiomonas. Smallsubunit rRNA oligomer catalogs have revealed that these genera, along with the family Enterobacteriaceae, form a branch of the gamma 3 subdivision of the purple bacteria (43). More recently, inter-and intrageneric relationships have been determined by using small-subunit rRNA sequences of Vibrio strains (nearly complete sequences for 10 species [ 141 and partial sequences for Vibrio and Photobacterium species [23]) and Aeromonas strains (10 species), as well as Plesiomonas shigelloides (29). From a taxonomic point of view, broad general agreement has been reached between classification based on a molecular approach and classical classification derived from physiological comparisons and numerical taxonomy. The results of genetic studies suggested, however, that the family Vibrionaceae should include only the genera Vibrio and Photobacterium (9,16,41,42), that the genus Aeromonas should be placed in a different family, the Aeromonadaceae, and that Plesiomonas shigelloides is more closely related to the family Enterobacteriaceae than to the family Vibrionaceae (6, 12, 18, 27, 43).Comparisons of small-subunit rRNA sequences are more informative than either analyses of 5s rRNA sequences or analyses of small-subunit rRNA oligomer catalogs. However, the results of studies in which workers used small-subunit rRNA sequences, which for many species were either not determined or were only partially determined, have left some questions unanswered. Also, the phylogenetic methods used previously to analyze relationships often have not included any estimate of the robustness of the results. Under these conditions, the phylogenetic relationships of all of these species were uncertain. In this study, we sequenced 54 strains and combined our data with previously published seque...

We have analyzed what phylogenetic signal can be derived by small subunit rRNA comparison for bacteria of different but closely related genera (enterobacteria) and for different species or strains within a single genus (Escherichia or Salmonella), and finally how similar are the ribosomal operons within a single organism (Escherichia coli). These sequences have been analyzed by neighbor-joining, maximum likelihood, and parsimony. The robustness of each topology was assessed by bootstrap. Sequences were obtained for the seven rrn operons of E. coli strain PK3. These data demonstrated differences located in three highly variable domains. Their nature and localization suggest that since the divergence of E. coli and Salmonella typhimurium, most point mutations that occurred within each gene have been propagated among the gene family by conversions involving short domains, and that homogenization by conversions may not have affected the entire sequence of each gene. We show that the differences that exist between the different operons are ignored when sequences are obtained either after cloning of a single operon or directly from polymerase chain reaction (PCR) products. Direct sequencing of PCR products produces a mean sequence in which mutations present in the most variable domains become hidden. Cloning a single operon results in a sequence that differs from that of the other operons and of the mean sequence by several point mutations. For identification of unknown bacteria at the species level or below, a mean sequence or the sequence of a single nonidentified operon should therefore be avoided. Taking into account the seven operons and therefore mutations that accumulate in the most variable domains would perhaps increase tree resolution. However, if gene conversions that homogenize the rRNA multigene family are rare events, some nodes in phylogenetic trees will reflect these recombination events and these trees may therefore be gene trees rather than organismal trees.

It is now well established that the clade of simian immunodeficiency viruses (SIVs) infecting west central African chimpanzees (Pan troglodytes troglodytes) and western gorillas (Gorilla gorilla gorilla) comprises the progenitors of human immunodeficiency virus type 1 (HIV-1). In this study, we have greatly expanded our previous molecular epidemiological survey of SIVcpz in wild chimpanzees in Cameroon. The new results confirm a wide but uneven distribution of SIVcpzPtt in P. t. troglodytes throughout southern Cameroon and indicate the absence of SIVcpz infection in Pan troglodytes vellerosus. Analyzing 725 fecal samples from 15 field sites, we obtained partial nucleotide sequences from 16 new SIVcpzPtt strains and determined full-length sequences for two of these. Phylogenetic analyses of these new viruses confirmed the previously reported phylogeographic clustering of SIVcpzPtt lineages, with viruses related to the ancestors of HIV-1 groups M and N circulating exclusively in southeastern and south central P. t. troglodytes communities, respectively. Importantly, the SIVcpzPtt strains from the southeastern corner of Cameroon represent a relatively isolated clade indicating a defined geographic origin of the chimpanzee precursor of HIV-1 group M. Since contacts between humans and apes continue, the possibility of ongoing transmissions of SIV from chimpanzees (or gorillas) to humans has to be considered. In this context, our finding of distinct SIVcpzPtt envelope V3 sequence clades suggests that these peptides may be useful for the serological differentiation of SIVcpzPtt and HIV-1 infections, and thus the diagnosis of new cross-species transmissions if they occurred.

The structure of rhizobial communities nodulating native shrubby legumes in open eucalypt forest of southeastern Australia was investigated by a molecular approach. Twenty-one genomic species were characterized by small-subunit ribosomal DNA PCR-restriction fragment length polymorphism and phylogenetic analyses, among 745 rhizobial strains isolated from nodules sampled on 32 different legume host species at 12 sites. Among these rhizobial genomic species, 16 belonged to the Bradyrhizobium subgroup, 2 to theRhizobium leguminosarum subgroup, and 3 to the Mesorhizobium subgroup. Only one genomic species corresponded to a known species (Rhizobium tropici). The distribution of the various genomic species was highly unbalanced among the 745 isolates, legume hosts, and sites.Bradyrhizobium species were by far the most abundant, andRhizobium tropici dominated among the Rhizobiumand Mesorhizobium isolates in the generally acid soils where nodules were collected. Although a statistically significant association occurred between the eight most common genomic species and the 32 hosts, there was sufficient overlap in distributions that no clear specificity between rhizobial genomic species and legume taxa was observed. However, for three legume species, some preference for particular genomic species was suggested. Similarly, no geographical partitioning was found.

The genomes of the spirochaetes Borrelia burgdorferi and Treponema pallidum show strong strand-specific skews in nucleotide composition, with the leading strand in replication being richer in G and T than the lagging strand in both species. This mutation bias results in codon usage and amino acid composition patterns that are significantly different between genes encoded on the two strands, in both species. There are also substantial differences between the species, with T.pallidum having a much higher G+C content than B. burgdorferi. These changes in amino acid and codon compositions represent neutral sequence change that has been caused by strong strand- and species-specific mutation pressures. Genes that have been relocated between the leading and lagging strands since B. burgdorferi and T.pallidum diverged from a common ancestor now show codon and amino acid compositions typical of their current locations. There is no evidence that translational selection operates on codon usage in highly expressed genes in these species, and the primary influence on codon usage is whether a gene is transcribed in the same direction as replication, or opposite to it. The dnaA gene in both species has codon usage patterns distinctive of a lagging strand gene, indicating that the origin of replication lies downstream of this gene, possibly within dnaN. Our findings strongly suggest that gene-finding algorithms that ignore variability within the genome may be flawed.

We describe a new species on the basis of phenotypic characteristics and the results of an analysis of small-subunit rRNA sequences. Three strains of this organism were isolated from a culture of the toxinproducing dinoflagellate Prorocentrum Zima. These bacteria are gram-negative, strictly aerobic, ovoid organisms that are motile by means of one or two subpolar flagella. They grow at temperatures ranging from 10 to 37°C and in the presence of NaCl concentrations ranging from 0.1 to 2 M and have an absolute requirement for sodium ions. They are strictly aerobic with a nonfermentative type of metabolism and are not able to grow anaerobically in presence or absence of nitrate. They do not denitrify. They exhibit oxidase, catalase, gelatinase, esculinase, P-galactosidase, and (to a lesser extent) amylase activities. The three strains which we examined require thiamine and biotin for growth. They grow only when glucose, trehalose, saccharose, fructose, maltose, pyruvate, malate, citrate, esculin, 2-ketoglutarate, 5-ketogluconate, glutamate, or shikimate is present as a sole carbon source. The three strains have identical small-subunit rRNA sequences. A phylogenetic analysis of these sequences revealed that these bacteria belong to the alpha subdivision of the Proteobacteria and that they form a distinct and robust monophyletic group with Roseobacter denitrifcans and Roseobacter Zitoralis. This result and the general phenotypic characteristics of the organisms place them in the genus Roseobacter, although they do not produce bacteriochlorophyll a, in contrast to previously described Roseobacter species. On the basis of the phenotypic and genetic similarities of these strains, we assigned them to a single species, for which the name Roseobacter algicola is proposed. The type strain is R. algicola FF3 (= ATCC 51440).The genus Roseobacter was created to comprise two bacterial species, Roseobacter denitriJicans and Roseobacter litoralis, which have been isolated from the surfaces of green seaweeds but not from seawater; these aerobic, pink-pigmented bacteria contain bacteriochlorophyll a (16,17). The genus Roseobacter differs phenotypically from the closely related genus Elythrobacter in carotenoid composition, bacteriochlorophyll-protein complex composition, and cell shape (16).Three bacterial strains that were isolated recently from a culture of the toxin-producing dinoflagellate Prorocentrum lima have been shown to produce okadaic acid (14). In this paper we show that on the basis of the results of a phenotypic examination and phylogenetic analyses of small-subunit rRNA these bacteria should be included in the genus Roseobacter, although none of the strains produces bacteriochlorophyll a. All of the data suggest that these three strains belong to the same new species, for which we propose the name Roseobacter algicola; strain FF3 (= ATCC 51440) is the type strain of this species. MATERIALS AND METHODSBacterial strains and growth conditions. Roseobacter algicola ML4, FF2, and FF3T (T = type strain) (14) were isolated f...

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