Publications

4858

Abstract Adelgids (Insecta: Hemiptera: Adelgidae) are known as severe pests of various conifers in North America, Canada, Europe and Asia. Here, we present the first molecular identification of bacteriocyte-associated symbionts in these plant sap-sucking insects. Three geographically distant populations of members of the Adelges nordmannianae/piceae complex, identified based on coI and ef1alpha gene sequences, were investigated. Electron and light microscopy revealed two morphologically different endosymbionts, coccoid or polymorphic, which are located in distinct bacteriocytes. Phylogenetic analyses of their 16S and 23S rRNA gene sequences assigned both symbionts to novel lineages within the Gammaproteobacteria sharing <92% 16S rRNA sequence similarity with each other and showing no close relationship with known symbionts of insects. Their identity and intracellular location were confirmed by fluorescence in situ hybridization, and the names ‘Candidatus Steffania adelgidicola’ and ‘Candidatus Ecksteinia adelgidicola’ are proposed for tentative classification. Both symbionts were present in all individuals of all investigated populations and in different adelgid life stages including eggs, suggesting vertical transmission from mother to offspring. An 85 kb genome fragment of ‘Candidatus S. adelgidicola’ was reconstructed based on a metagenomic library created from purified symbionts. Genomic features including the frequency of pseudogenes, the average length of intergenic regions and the presence of several genes which are absent in other long-term obligate symbionts, suggested that ‘Candidatus S. adelgidicola’ is an evolutionarily young bacteriocyte-associated symbiont, which has been acquired after diversification of adelgids from their aphid sister group.

Abstract We investigated seed bugs of the genus Nysius (Insecta: Hemiptera: Lygaeidae) for their symbiotic bacteria. From all the samples representing 4 species, 18 populations and 281 individuals, specific bacterial 16S rRNA gene sequences were consistently identified, which formed a distinct clade in the Gammaproteobacteria. In situ hybridization showed that the bacterium was endocellularly localized in a pair of large bacteriomes that were amorphous in shape, deep red in color, and in association with gonads. In the ovary of adult females, the endosymbiont was also localized in the ‘infection zone’ in the middle of each germarium and in the ‘symbiont ball’ at the anterior pole of each oocyte, indicating vertical transmission of the endosymbiont through the ovarial passage. Phylogenetic analyses based on bacterial 16S rRNA, groEL and gyrB genes consistently supported a coherent monophyly of the Nysius endosymbionts. The possibility of a sister relationship to ‘Candidatus Kleidoceria schneideri’, the bacteriome-associated endosymbiont of a lygaeid bug Kleidocerys resedae, was statistically rejected, indicating independent evolutionary origins of the endosymbionts in the Lygaeidae. The endosymbiont genes consistently exhibited AT-biased nucleotide compositions and accelerated rates of molecular evolution, and the endosymbiont genome was only 0.6 Mb in size. The endosymbiont phylogeny was congruent with the host insect phylogeny, suggesting strict vertical transmission and host–symbiont co-speciation over evolutionary time. Based on these results, we discuss the evolution of bacteriomes and endosymbionts in the Heteroptera, most members of which are associated with gut symbiotic bacteria. The designation ‘Candidatus Schneideria nysicola’ is proposed for the endosymbiont clade.

ABSTRACT “ Candidatus Methylomirabilis oxyfera” is a newly discovered denitrifying methanotroph that is unrelated to previously known methanotrophs. This bacterium is a member of the NC10 phylum and couples methane oxidation to denitrification through a newly discovered intra-aerobic pathway. In the present study, we report the first ultrastructural study of “ Ca . Methylomirabilis oxyfera” using scanning electron microscopy, transmission electron microscopy, and electron tomography in combination with different sample preparation methods. We observed that “ Ca . Methylomirabilis oxyfera” cells possess an atypical polygonal shape that is distinct from other bacterial shapes described so far. Also, an additional layer was observed as the outermost sheath, which might represent a (glyco)protein surface layer. Further, intracytoplasmic membranes, which are a common feature among proteobacterial methanotrophs, were never observed under the current growth conditions. Our results indicate that “ Ca . Methylomirabilis oxyfera” is ultrastructurally distinct from other bacteria by its atypical cell shape and from the classical proteobacterial methanotrophs by its apparent lack of intracytoplasmic membranes.

The potato/tomato psyllid, Bactericera cockerelli transmits the bacterium ‘Candidatus Liberibacter solanacearum’, also known as ‘Ca. L. psyllaurous’, which causes zebra chip disease in solanaceous crops. There have been no studies addressing the effect of the bacterial plant pathogen on the biology of its insect vector. We examined several life-history traits, including 7-day fecundity, hatching percentage, incubation time, nymphal survival percentage, nymphal development time, total development time, and sex-ratio of ‘Ca. L. solanacearum’-positive and -negative psyllid isofemale lines on tomato, as well as adult mortality index of ‘Ca. L. solanacearum’-positive and -negative insects. The only two life-history traits that differed between the ‘Ca. L. solanacearum’-positive and –negative psyllid isofemale lines were 7-day fecundity and nymphal survival percentage, which were significantly lower in ‘Ca. L. solanacearum’- positive lines. The symbiotic bacteria associated with both psyllid isofemale lines were similar, with the exception of ‘Ca. L. solanacearum’, which showed 100% infection in the ‘Ca. L. solanacearum’-positive lines and was not detected in the negative psyllid lines. These results suggest that ‘Ca. L. solanacearum' has a negative effect on population growth rate of its insect vector on tomato.