Figure 1.
Relationships of nematode species harbouring Wolbachia symbionts.
A phylogenetic cartoon showing the relationships of the nematode species discussed in this work [23]. To the left, the systematic structure of the class Chromadoria is given, and the three major suborders within Rhabditida are highlighted. Lifecycle strategies of the groups are indicated. The fine-scale relationships of species discussed in the text are given to the right. The presence of live Wolbachia infection (+: yes, −: no), evidence of laterally-transferred Wolbachia sequences in the nuclear genome (+: yes, −: no, ?: unknown), and the availability of complete genome sequences (+: yes, −: no, ±: partial genome sequence) for each of the species are indicated.
Figure 2.
Comparison of the Dictyocaulus viviparus proteome to that of other rhabditid nematodes.
Venn diagram illustrating the orthoMCL clustering of the predicted proteome of Dictyocaulus viviparus (DVI) to those of Caenorhabditis elegans (CEL), Heterorhabditis bacteriophora (HBA) and Haemonchus contortus (HCO). The numbers of proteins clustered and the total number of predicted proteins is given below each species' name.
Table 1.
Assembly statistics for the Dictyocaulus viviparus nuclear genome and the Wolbachia-like insertions.
Table 2.
Genome assembly and annotation metrics of D. viviparus and other Rhabditina species.
Figure 3.
Wolbachia sequence in a Dictyocaulus viviparus genome assembly.
A. Taxon-annotated GC%-coverage plot of the primary D. viviparus genome assembly, with contigs that have significant matches to Wolbachia proteins highlighted in red. A total of 193 contigs spanning 1 Mb (out of a total assembly span of 169 Mb) had significant similarity to Wolbachia. B. Circos plot comparing the 25 longest of the D. viviparus genome contigs that contained Wolbachia-like sequence to the genome of the Wolbachia endosymbionts of the filarial nematode Brugia malayi (wBm) [9] and Onchocerca ochengi (wOo). The arcs show BLASTn-derived matches between the contigs and the genome sequences. Transcripts from D. viviparus mapped to the assembly are reported as green lines in the outer circle of the figure. C. Frequency histogram illustrating the different patterns of coverage of the Wolbachia-like scaffolds (black) compared to the nuclear genome scaffolds (green). D. Frequency plot of similarity of D. viviparus Wolbachia-like sequences to wBm (blue) and wOo (the Wolbachia endosymbiont of the filarial nematode Onchocerca ochengi) (red). Each D. viviparus Wolbachia-like segment was split into 500 bp fragments, and the best percentage identity with the reference genomes calculated using BLASTn. E. The Wolbachia-like fragments identified in the D. viviparus genome assembly are co-assembled with nematode genes, and have accumulated multiple inactivating mutations. Two putative Wolbachia insertions in nuclear contigs are shown in views derived from the gBrowse genome viewer. Each panel shows (from top to bottom) the whole scaffold with the zoomed-in region highlighted, the GC% plot for the scaffold, the scale for the zoomed-in region, the read coverage for the zoomed-in region, the genes called by RAST in the zoomed in region and the genes called by AUGUSTUS in the zoomed-in region. The upper plot shows scaffold00357 while the lower plot shows scaffold00506.
Table 3.
Putative Wolbachia-like open reading frames identified in the Dictyocaulus viviparus nuclear genome.
Figure 4.
Comparison of Wolbachia-like insertions from two Dictyocaulus viviparus isolates, and relationships of the Cameroon D. viviparous.
A. 16S rRNA gene fragments from the Cameroon isolate of D. viviparus (obtained through whole genome sequencing) and from the Moredun isolate (from specific amplification) are shown aligned. The genome sequence assembly has three copies of Wolbachia-like 16S genes, two tandemly arranged and truncated in scaffold scaf09320, and one in scaffold scaf01523. B. ftsZ gene fragments from the Cameroon isolate of D. viviparus (obtained through whole genome sequencing) and from the Moredun isolate (from specific amplification) are shown aligned. While we were able to amplify the complete fragment from the Moredun strain, the genome assembly contains only a truncated ftsZ gene (and no consensus is shown for the ∼200 bases of essentially unaligned sequence at the 5′ end of the alignment). C. Bayesian phylogenetic analysis of the complete nuclear small subunit ribosomal RNA (nSSU) genes of the Cameroon D. viviparus and other Dictyocaulus sp., and outgroups (taken from the European Nucleotide Archive). The Cameroon D. viviparus is most similar to the European D. viviparus sequenced previously. RAxML analyses yielded the same topology. The 5′ gene fragment isolated and sequenced from the Moredun strain was identical to the other D. viviparus nSSU sequences.
Table 4.
Possible transcribed genes of Wolbachia origin in the Dictyocaulus viviparus genome.
Figure 5.
Analysis of the phylogenetic relationships of the Wolbachia nuclear insertions in the Dictyocaulus viviparus genome.
Phylogenetic tree inferred from 16S rDNA, groEL, ftsZ, dnaA and coxA loci with maximum likelihood (RAxML) and Bayesian (MrBayes, PhyloBayes) inference. Branch support is reported as (RaxML/MrBayes/PhyloBayes). Strains representing Wolbachia supergroups A, B, C, D, F and H are indicated.
Table 5.
Matches to Wolbachia WO phage in the Dictyocaulus viviparus genome assembly.
Table 6.
Analysis software versions and parameter settings.
Table 7.
PCR test for Wolbachia insertions.