In the past, bacterial phylogeny relied almost exclusively on 16S rRNA gene sequence analysis. More recently, multilocus sequence analysis has been used to infer organismal phylogenies. In this study, the dnaJ chaperone gene was investigated as a marker for phylogeny studies in alphaproteobacteria. Preliminary analysis of G+C contents and G+C3s contents (the G+C content of the synonymous third codon position) showed no clear evidence of horizontal transfer of this gene in proteobacteria. dnaJ-based phylogenies were then analysed at three taxonomic levels: the Proteobacteria, the Alphaproteobacteria and the genus Mesorhizobium. Dendrograms based on DnaJ and 16S rRNA gene sequences revealed the same topology described previously for the Proteobacteria. These results indicate that the DnaJ phylogenetic signal is able to reproduce the accepted relationships among the five classes of the Proteobacteria. At a lower taxonomic level, using 20 alphaproteobacteria, the 16S rRNA gene-based phylogeny is distinct from the one based on DnaJ sequence analysis. Although the same clusters are generated, only the topology of the DnaJ tree is consistent with broader phylogenies from recent studies based on concatenated alignments of multiple core genes. For example, the DnaJ tree shows the two clusters within the Rhizobiales as closely related, as expected, while the 16S rRNA gene-based phylogeny shows them as distantly related. In order to evaluate the phylogenetic performance of dnaJ at the genus level, a multilocus analysis based on five housekeeping genes (atpD, gapA, gyrB, recA and rplB) was performed for ten Mesorhizobium species. In contrast to the 16S rRNA gene, the DnaJ sequence analysis generated a tree similar to the multilocus dendrogram. For identification of chickpea mesorhizobium isolates, a dnaJ nucleotide sequence-based tree was used. Despite different topologies, 16S rRNA gene- and dnaJ-based trees led to the same species identification. This study suggests that the dnaJ gene is a good phylogenetic marker, particularly for the class Alphaproteobacteria, since its phylogeny is consistent with phylogenies based on multilocus approaches.
The extremely diverse genus Lactobacillus is the largest among the lactic acid bacteria, with over 145 recognized species. In this work, which to our knowledge is the largest comparative phylogenomics study of a single genus to date, 12 genomes of Lactobacillus strains were subjected to an array of whole-genome and single-marker phylogenetic approaches, to investigate the case for extracting subgeneric groups and to determine whether a single congruent phylogeny could be identified. We conclude that GroEL is a more robust single-gene phylogenetic marker for the genus Lactobacillus than the 16S rRNA gene, when no whole-genome information is available. Significant incongruence was found, both within a set of trees based on 141 core proteins and within those phylogenies based on numbers of orthologues, concatenated RNA polymerase subunits and single gene/protein markers. This is possibly due to different evolutionary rates, hidden paralogies or horizontal gene transfer. Such phylogenetic ambiguities are efficiently visualized with cluster-networks. Although the genus contains some highly unstable taxa, four subgeneric groups were distinguished. Qualitative and quantitative gene analysis of these groups resulted in three findings: there is a relatively small number of group-specific proteins, the majority of which are poorly characterized; major groupings are functionally better distinguishable by absent genes rather than gained/retained genes; and, finally, a gene cluster possibly involved in purine metabolism is uniquely present in four lactobacilli associated with meat. In conclusion, because of either significantly different branching patterns or the availability of too few members, three of the four identified groups could not serve as the basis for identifying candidate novel genera within the current genus. We therefore suggest targeted sequencing of key taxonomic species identified here, which are likely to add sufficient depth for a future reclassification, followed by phylogenomic analysis involving the core proteins identified here. This will ideally be combined with phenotypic data using a polyphasic approach.
The Prodiscocephalus-like ciliates, or discocephalines, are cephalized organisms that are traditionally considered to be hypotrichs (sensu lato) but whose precise systematic position has long been uncertain. The main reasons for this are that these organisms exhibit several intermediate morphological and morphogenetic features and that hitherto none has been investigated using molecular methods. In the present study, the cortical development of Prodiscocephalus borrori was observed during binary division and this can be summarized as follows: (i) in the parental adoral zone of membranelles, only the posterior end is renewed by dedifferentiation of the old structures; (ii) the oral primordium in the opisthe occurs de novo on the cell surface as seen in other typical stichotrichs; (iii) in both dividers, the undulating membranes anlage does not split longitudinally in the usual way but, instead, divides transversely to form the paroral and endoral membranes; (iv) usually seven frontoventral transverse cirral anlagen are formed in the primary mode which then divide into two sets, one each for the proter and opisthe; (v) both left and right marginal rows divide into two parts, thus giving rise to a post-lateral marginal segment at the posterior end of each; (vi) invariably five caudal cirri are formed at the posterior end of the three rightmost dorsal kinety anlagen. Thus, it was found that, like other related discocephalines, P. borrori exhibits more similarities to stichotrichs than to euplotids. Based on a combination of morphological and morphogenetic data, a phylogenetic tree was constructed which suggests that the discocephalines group within the stichotrichs and separate from the euplotids. In addition, the complete small-subunit rRNA gene (SSU rDNA) of P. borrori was sequenced and analysed. In the resulting SSU rDNA tree, the discocephalines represent an intermediate group between the euplotids and the Stichotrichia–Oligotrichia–Choreotrichia assemblage, albeit with low bootstrap support. From these data, we conclude that the discocephalines might be a divergent, or possibly an ancestral, group within the Stichotrichia. Furthermore, our findings further support the suggestion that these organisms should be considered as a distinct order, i.e. Discocephalida Wicklow, 1982, in the subclass Stichotrichia Small & Lynn, 1985 .