Genetic and metabolic analyses of Candidatus Liberibacter solanacearum infecting carrot

Insect-vectored plant bacterial pathogens are gaining attention in recent years due to crop threatening outbreaks around the world. Candidatus Liberibacter spp. are infecting crops of different botanical families: Solanaceae, Rutaceae, and Apiaceae and are vectored by psyllids. Five genetic haplotypes (A-E) have been described thus far for the species Ca. Liberibacter solanacearum (Lso). Haplotypes A and B infecting solanaceous plants, haplotypes C-E infecting Apiaceae crops. To better understand the genetic basis that governs host specificity of Lso haplotypes, we sequenced the genome of haplotype D (LsoD). The LsoD genome size is 1.23 Mbp, with a GC content of 34.8% and 1167 predicted genes. Enzyme Commission (EC) numbers were assigned using the JGI software tool and 358 ECs were identified. ECs were mapped to metabolic pathways and compared with other sequenced Liberibacters. Phylogenetic analysis based on ECs and assigned metabolic pathways shows that LsoD groups together with Lso haplotypes (A and B) and is clearly different than Liberibacter species infecting citrus. Differences between LsoD and LsoA/B haplotypes were also found, hinting on host specific enzymes. The LsoD genome was also scanned to identify putatively secreted proteins using the SignalP tool. Thirty-one putative genes were identified, most of them with unknown function. While some genes have homologous in other Lso haplotypes, some were unique to LsoD. By quantitative-PCR we examined the expression of the putatively secreted proteins in the different hosts; the psyllid vector Bactericera trigonica, and carrot. Several genes with significantly higher expression levels in carrot compared with psyllid and vice versa were identified. These genes may have host specific functions. Overall, our analyses reveal genetic and metabolic elements differentiating the carrot-infecting Lso from Lso haplotypes infecting potato/tomato. Research is underway to identify the function of these elements

Genome Analysis of Haplotype D of Candidatus Liberibacter Solanacearum

Candidatus Liberibacter solanacearum (Lso) haplotype D (LsoD) is a suspected bacterial pathogen, spread by the phloem-feeding psyllid Bactericera trigonica Hodkinson and found to infect carrot plants throughout the Mediterranean. Haplotype D is one of six haplotypes of Lso that each have specific and overlapping host preferences, disease symptoms, and psyllid vectors. Genotyping of rRNA genes has allowed for tracking the haplotype diversity of Lso and genome sequencing of several haplotypes has been performed to advance a comprehensive understanding of Lso diseases and of the phylogenetic relationships among the haplotypes. To further pursue that aim we have sequenced the genome of LsoD from its psyllid vector and report here its draft genome. Genome-based single nucleotide polymorphism analysis indicates LsoD is most closely related to the A haplotype. Genomic features and the metabolic potential of LsoD are assessed in relation to Lso haplotypes A, B, and C, as well as the facultative strain Liberibacter crescens. We identify genes unique to haplotype D as well as putative secreted effectors that may play a role in disease characteristics specific to this haplotype of Lso

Modeling trophic dependencies and exchanges among insects’ bacterial symbionts in a host-simulated environment

Abstract Background Individual organisms are linked to their communities and ecosystems via metabolic activities. Metabolic exchanges and co-dependencies have long been suggested to have a pivotal role in determining community structure. In phloem-feeding insects such metabolic interactions with bacteria enable complementation of their deprived nutrition. The phloem-feeding whitefly Bemisia tabaci (Hemiptera: Aleyrodidae) harbors an obligatory symbiotic bacterium, as well as varying combinations of facultative symbionts. This well-defined bacterial community in B. tabaci serves here as a case study for a comprehensive and systematic survey of metabolic interactions within the bacterial community and their associations with documented occurrences of bacterial combinations. We first reconstructed the metabolic networks of five common B. tabaci symbionts genera (Portiera, Rickettsia, Hamiltonella, Cardinium and Wolbachia), and then used network analysis approaches to predict: (1) species-specific metabolic capacities in a simulated bacteriocyte-like environment; (2) metabolic capacities of the corresponding species’ combinations, and (3) dependencies of each species on different media components. Results The predictions for metabolic capacities of the symbionts in the host environment were in general agreement with previously reported genome analyses, each focused on the single-species level. The analysis suggests several previously un-reported routes for complementary interactions and estimated the dependency of each symbiont in specific host metabolites. No clear association was detected between metabolic co-dependencies and co-occurrence patterns. Conclusions The analysis generated predictions for testable hypotheses of metabolic exchanges and co-dependencies in bacterial communities and by crossing them with co-occurrence profiles, contextualized interaction patterns into a wider ecological perspective

Additional file 3:

Annotations for the Cardinium genome from four platforms (IMG/M, Kbase, Rast, MG-Rast) were compared with the manual annotation conducted by Santos-Garcia et al. 2014. The JGI platform had both the absolute highest number of Enzyme Commission (EC) predictions as well as the highest overlap with the manual annotation. Hence, it was selected as the standard annotation tool for all symbionts. (DOCX 113 kb

Additional file 11:

Illustration of the putative complementation at metabolic level detected for the synthesis of Branched Chain Amino Acids and Lysine. The Wolbachia’s lysine biosynthetic pathway lacks its last reaction (argD, EC 4.1.1.20), which is present in Hamiltonella, leading to a complementary potential production of lysine from M-DAP. Synthesis of M-DAP is consistently inferred in most sequenced Wolbachia, providing an intermediate compound for the biosynthesis of peptidoglycan, part of the bacterial membrane. (DOC 108 kb

Additional file 12:

Co-occurrence frequencies of facultative endosymbionts. (DOC 86 kb

Additional file 7:

Illustration of the model used to simulate growth, calculate metabolic overlap, and estimate the effect of specific metabolites on metabolic production. (DOCX 179 kb

Additional file 9:

Orthologous protein clusters of five Wolbachia strains represented as a Euler–Venn Diagram, as described in additional file 1. (DOCX 187 kb

Additional file 1:

Description of symbiont’s DNA extraction and Wolbachia genome assembly. (DOCX 26 kb

Additional file 2:

Comparison of the completeness of the draft genomes using CheckM. (XLS 32 kb