Microbe Profile: Buchnera aphidicola: ancient aphid accomplice and endosymbiont exemplar

Buchnera aphidicola is an obligate endosymbiont of aphids that cannot be cultured outside of hosts. It exists as diverse strains in different aphid species, and phylogenetic reconstructions show that it has been maternally transmitted in aphids for >100 million years. B. aphidicola genomes are highly reduced and show conserved gene order and no gene acquisition, but encoded proteins undergo rapid evolution. Aphids depend on B. aphidicola for biosynthesis of essential amino acids and as an integral part of embryonic development. How B. aphidicola populations are regulated within hosts remains little known.

Genome sequence of the English grain aphid, Sitobion avenae and its endosymbiont Buchnera aphidicola

The English grain aphid, Sitobion avenae, is a major agricultural pest of wheat, barley and oats, and one of the principal vectors of Barley Yellow Dwarf Virus (BYDV) leading to significant reductions in grain yield, annually. Emerging resistance to and increasing regulation of insecticides has resulted in limited options for their control. Using PacBio HiFi data, we have produced a high quality draft assembly of the S. avenae genome; generating a primary assembly with a total assembly size of 475.7 Mb, and an alternate assembly with a total assembly size of 430.8 Mb. Our primary assembly was highly contiguous with only 326 contigs and a contig N50 of 15.95 Mb. Assembly completeness was estimated at 97.7% using BUSCO analysis and 31,007 and 29,037 protein coding genes were predicted from the primary and alternate assemblies, respectively. This assembly, which is to our knowledge the first for an insecticide resistant clonal lineage of English grain aphid, will provide novel insight into the molecular and mechanistic determinants of resistance and will facilitate future research into mechanisms of viral transmission and aphid behavior.

Ploidy dynamics in aphid host cells harboring bacterial symbionts

Aphids have evolved bacteriocytes or symbiotic host cells that harbor the obligate mutualistic bacterium Buchnera aphidicola. Because of the large cell size (approximately 100 μm in diameter) of bacteriocytes and their pivotal role in nutritional symbiosis, researchers have considered that these cells are highly polyploid and assumed that bacteriocyte polyploidy may be essential for the symbiotic relationship between the aphid and the bacterium. However, little is known about the ploidy levels and dynamics of aphid bacteriocytes. Here, we quantitatively analyzed the ploidy levels in the bacteriocytes of the pea-aphid Acyrthosiphon pisum. Image-based fluorometry revealed the hyper polyploidy of the bacteriocytes ranging from 16- to 256-ploidy throughout the lifecycle. Bacteriocytes of adult parthenogenetic viviparous females were mainly 64-128C DNA levels, while those of sexual morphs (oviparous females and males) were consisted of 64C, and 32-64C cells, respectively. During post-embryonic development of viviparous females, the ploidy level of bacteriocytes increased substantially, from 16-32C at birth to 128-256C in actively reproducing adults. These results suggest that the ploidy levels are dynamically regulated among phenotypes and during development. Our comprehensive and quantitative data provides a foundation for future studies to understand the functional roles and biological significance of the polyploidy of insect bacteriocytes.

Geography-dependent symbiont communities in two oligophagous aphid species

Aphids and their diverse symbionts have become a good model to study bacteria-arthropod symbiosis. The feeding habits of aphids are usually influenced by a variety of symbionts. Most studies on symbiont diversity have focused on polyphagous aphids, while symbiont community patterns for oligophagous aphids remain unclear. Here, we surveyed the bacterial communities in natural populations of two oligophagous aphids, Melanaphis sacchari and Neophyllaphis podocarpi, in natural populations. Seven common symbionts were detected, among which Buchnera aphidicola and Wolbachia were the most prevalent. In addition, an uncommon Sodalis-like symbiont was also detected in these two aphids, and Gilliamella was found in some samples of M. sacchari. We further assessed the significant variation in symbiont communities within the two aphid species, geographical regions and host specialization using statistical and ordination analyses. Geography was an important factor in shaping the symbiont community structure in these oligophagous aphids. Furthermore, the strong geographical influence may be related to specific environmental factors, especially temperature, among different regions. These findings extend our knowledge of the significance of geography and its associated environmental conditions in the symbiont community structure associated with oligophagous aphids.

Methylation at CpG sites in genomes of aphid Acyrtosiphon pisum and its endosymbiont Buchnera

Pea aphid Acyrtosiphon pisum, a sap-feeding insect, has established a mutualistic relationship with an endosymbiotic bacteria (Buchnera aphidicola) that constitutes an evolutionary successful symbiosis to synthetize complex chemical compounds from a nutrient deprived diet. In this study, led by the presence of DNMT1 and a putative DNMT3 methylase in the aphid genome, we investigated the distribution of the methyl groups on 5'cytosine in CpG motifs on the whole genomes of host and endosymbiont, and looked into their correlation with gene expression. The DNA methylation turned to be present at low level in aphid (around 3% of total genomic cytosine) compared to mammals and plants, but increased to ~9% in genes. Interestingly, the reduced genome of the endosymbiont Buchnera also shows global low level of methyl cytosine despite the fact that its genome does not shelter any DNA methylase. This finding argues for the import of DNA methylase from the host to the endosymbiont. The observed differences in methylation patterns between two clonal variants (host plus endosymbiont) are reported along with the differences in their transcriptome profiles. Our data allowed to decipher a dynamic combinatorial DNA methylation and epigenetic cross talk between host and symbiont in a clonality context that might count for the aphid adaptation to environment.

Insight into the bacterial communities of the subterranean aphid Anoecia corni

Many insect species are associated with bacterial partners that can significantly influence their evolutionary ecology. Compared to other insect groups, aphids harbor a bacterial microbiota that has the reputation of being poorly diversified, generally limited to the presence of the obligate nutritional symbiont Buchnera aphidicola and some facultative symbionts. In this study, we analyzed the bacterial diversity associated with the dogwood-grass aphid Anoecia corni, an aphid species that spends much of its life cycle in a subterranean environment. Little is known about the bacterial diversity associated with aphids displaying such a lifestyle, and one hypothesis is that close contact with the vast microbial community of the rhizosphere could promote the acquisition of a richer bacterial diversity compared to other aphid species. Using 16S rRNA amplicon Illumina sequencing on specimens collected on wheat roots in Morocco, we identified 10 bacterial operational taxonomic units (OTUs) corresponding to five bacterial genera. In addition to the obligate symbiont Buchnera, we identified the facultative symbionts Serratia symbiotica and Wolbachia in certain aphid colonies. The detection of Wolbachia is unexpected as it is considered rare in aphids. Moreover, its biological significance remains unknown in these insects. Besides, we also detected Arsenophonus and Dactylopiibacterium carminicum. These results suggest that, despite its subterranean lifestyle, A. corni shelter a bacterial diversity mainly limited to bacterial endosymbionts.

Triple RNA-Seq characterizes aphid gene expression in response to infection with unequally virulent strains of the endosymbiont Hamiltonella defensa

Background Secondary endosymbionts of aphids provide benefits to their hosts, but also impose costs such as reduced lifespan and reproductive output. The aphid Aphis fabae is host to different strains of the secondary endosymbiont Hamiltonella defensa, which encode different putative toxins. These strains have very different phenotypes: They reach different densities in the host, and the costs and benefits (protection against parasitoid wasps) they confer to the host vary strongly. Results We used RNA-Seq to generate hypotheses on why four of these strains inflict such different costs to A. fabae. We found different H. defensa strains to cause strain-specific changes in aphid gene expression, but little effect of H. defensa on gene expression of the primary endosymbiont, Buchnera aphidicola. The highly costly and over-replicating H. defensa strain H85 was associated with strongly reduced aphid expression of hemocytin, a marker of hemocytes in Drosophila. The closely related strain H15 was associated with downregulation of ubiquitin-related modifier 1, which is related to nutrient-sensing and oxidative stress in other organisms. Strain H402 was associated with strong differential regulation of a set of hypothetical proteins, the majority of which were only differentially regulated in presence of H402. Conclusions Overall, our results suggest that costs of different strains of H. defensa are likely caused by different mechanisms, and that these costs are imposed by interacting with the host rather than the host’s obligatory endosymbiont B. aphidicola.

Isolation of the Buchnera aphidicola flagellum basal body complexes from the Buchnera membrane

Buchnera aphidicola is an intracellular bacterial symbiont of aphids and maintains a small genome of only 600 kbps. Buchnera is thought to maintain only genes relevant to the symbiosis with its aphid host. Curiously, the Buchnera genome contains gene clusters coding for flagellum basal body structural proteins and for flagellum type III export machinery. These structures have been shown to be highly expressed and present in large numbers on Buchnera cells. No recognizable pathogenicity factors or secreted proteins have been identified in the Buchnera genome, and the relevance of this protein complex to the symbiosis is unknown. Here, we show isolation of Buchnera flagellum basal body proteins from the cellular membrane of Buchnera, confirming the enrichment of flagellum basal body proteins relative to other proteins in the Buchnera proteome. This will facilitate studies of the structure and function of the Buchnera flagellum structure, and its role in this model symbiosis.

Some living beings speak up against ‘electron transport chain – chemiosmosis – rotary ATP synthesis’ bioenergetics model

Since 2017, I have argued that affinity/contact-based electron transport chain (ETC) and chemiosmotic rotary ATP synthesis (CRAS) explanation fail to reason the workability or evolution of cellular bioenergetics. Cyanobacteria (Prochlorococcus) can both respire and photosynthesize. In these systems, the aqueous milieu of <0.1 femtoliter (pH 8) cannot afford any free protons to build ‘pmf’’, thereby negating chemiosmosis. The anti-parallel ETCs (NADH→H2O in respiration and H2O→NADH in photosynthesis) would lead to futile cycles, owing to the commonality of cytochromes b6f and c1. Aphids of genus Acyrthosiphon use the inbuilt carotenoid pigments to reduce NAD and synthesize ATP, without elaborate setups like Z-scheme. The classical perceptions deem archaeans to have a unique bioenergetic history, distinct from bacteria/eukarya. Although it is known that archaea lack FoF1ATP(synth)ase, the ‘molecular motor’ is otherwise believed to be conserved in bacterial/eukaryotic systems. However, the genes of Complex V are not clubbed/linked in proteobacterial Rickettsia prowazekii. Several parasitic and a free-living alveolate Tetrahymena thermophila lack essential proteins of Fo module. Complex V is completely absent in the blood-stage malarial parasite (Plasmodium berghei) and Buchnera aphidicola (a proteobacterium, endosymbiont of cedar bark aphid). Monocercomonoides (a flagellate oxymonad rodent-gut symbiont) and human erythrocytes generate ATP without mitochondria! The recently discovered cnidarian parasite Henneguya salminicola does not have respiratory Complexes I, III & IV, the purported classical proton pumps. While the ETC-CRAS model is incompatible with these facts, the newly available murburn alternative accommodates them. The principles of scientific pursuits and Ockham’s razor decide in favor of the murburn model of bioenergetics.