Publications

4915

AbstractCandidatus Arsenophonus lipopteni (Enterobacteriaceae, Gammaproteobacteria) is an obligate intracellular symbiont of the blood feeding deer ked, Lipoptena cervi (Diptera: Hippoboscidae). The bacteria reside in specialized cells derived from host gut epithelia (bacteriocytes) forming a compact symbiotic organ (bacteriome). Compared to the closely related complex symbiotic system in the sheep ked, involving four bacterial species, Lipoptena cervi appears to maintain its symbiosis exclusively with Ca. Arsenophonus lipopteni. The genome of 836,724 bp and 24.8 % GC content codes for 667 predicted functional genes and bears the common characteristics of sequence economization coupled with obligate host-dependent lifestyle, e.g. reduced number of RNA genes along with the rRNA operon split, and strongly reduced metabolic capacity. Particularly, biosynthetic capacity for B vitamins possibly supplementing the host diet is highly compromised in Ca. Arsenophonus lipopteni. The gene sets are complete only for riboflavin (B2), pyridoxine (B6) and biotin (B7) implying the content of some B vitamins, e.g. thiamin, in the deer blood might be sufficient for the insect metabolic needs. The phylogenetic position within the spectrum of known Arsenophonus genomes and fundamental genomic features of Ca. Arsenophonus lipopteni indicate the obligate character of this symbiosis and its independent origin within Hippoboscidae.

AbstractCandidatus Liberibacter asiaticus (CLas) is a phloem-limited, gram-negative, fastidious bacterium that is associated with the development of citrus greening disease, also known as Huanglongbing (HLB). CLas is transmitted by the Asian citrus psyllid (ACP) Diaphorina citri, in a circulative manner. Two major barriers to transmission within the insect are the midgut and the salivary glands. We performed a thorough microscopic analysis within the insect midgut following exposure to CLas-infected citrus trees. We observed changes in nuclear architecture, including pyknosis and karyorrhexis as well as changes to the actin cytoskeleton in CLas-exposed midgut cells. Further analyses showed that the changes are likely due to the activation of programmed cell death as assessed by Annexin V staining and DNA fragmentation assays. These results suggest that exposure to CLas-infected trees induces apoptotic responses in the psyllid midgut that should be further investigated. Understanding the adaptive significance of the apoptotic response has the potential to create new approaches for controlling HLB.

AbstractBiocathodes provide a stable electron source to drive reduction reactions in electrotrophic microbial electrochemical systems. Electroautotrophic biocathode communities may be more robust than monocultures in environmentally relevant settings, but some members are not easily cultivated outside of the electrode environment. We previously used metagenomics and metaproteomics to propose a pathway for coupling extracellular electron transfer (EET) to carbon fixation in “Candidatus Tenderia electrophaga”, an uncultivated but dominant member of the Biocathode-MCL electroautotrophic community. Here we validate and refine this proposed pathway using differential metatranscriptomics of replicate MCL reactors poised at the growth potential 310 mV and the suboptimal 470 mV (vs. standard hydrogen electrode). At both potentials, transcripts from “Ca. Tenderia electrophaga” were more abundant than from any other organism and its relative activity was positively correlated with current. Several genes encoding key components of the proposed “Ca. Tenderia electrophaga” EET pathway were more highly expressed at 470 mV, consistent with a need for cells to acquire more electrons to obtain the same amount of energy as at 310 mV. These included cyc2, encoding a homolog of a protein known to be involved in iron oxidation, confirmed to be differentially expressed by droplet digital PCR of independent biological replicates. Average expression of all CO2 fixation related genes is 1.23-fold higher at 310 mV, indicating that reduced energy availability at 470 mV decreased CO2 fixation. Our results substantiate the claim that “Ca. Tenderia electrophaga” is the key MCL electroautotroph, which will help guide further development of this community for microbial electrosynthesis.IMPORTANCEBacteria that directly use electrodes as metabolic electron donors (biocathodes) have been proposed for applications ranging from microbial electrosynthesis to advanced bioelectronics for cellular communication with machines. However, just as we understand very little about oxidation of analogous natural insoluble electron donors, such as iron oxide, the organisms and extracellular electron transfer (EET) pathways underlying the electrode-cell direct electron transfer processes are almost completely unknown. Biocathodes are a stable biofilm cultivation platform to interrogate both the rate and mechanism of EET using electrochemistry and study the electroautotrophic organisms that catalyze these reactions. Here we provide new evidence supporting the hypothesis that the uncultured bacterium “Candidatus Tenderia electrophaga” directly couples extracellular electron transfer to CO2 fixation. Our results provide insight into developing biocathode technology, such as microbial electrosynthesis, as well as advancing our understanding of chemolithoautotrophy.

Significance Mealybugs are plant sap-sucking insects with a nested symbiotic arrangement, where one bacterium lives inside another bacterium, which together live inside insect cells. These two bacteria, along with genes transferred from other bacteria to the insect genome, allow the insect to survive on its nutrient-poor diet. Here, we show that the innermost bacterium in this nested symbiosis was replaced several times over evolutionary history. These results show that highly integrated and interdependent symbiotic systems can experience symbiont replacement and suggest that similar dynamics could have occurred in building the mosaic metabolic pathways seen in mitochondria and plastids.

Summary The anaerobic oxidation of methane (AOM) is mediated by consortia of anaerobic methane‐oxidizing archaea (ANME) and their specific partner bacteria. In thermophilic AOM consortia enriched from Guaymas Basin, members of the ANME‐1 clade are associated with bacteria of the HotSeep‐1 cluster, which likely perform direct electron exchange via nanowires. The partner bacterium was enriched with hydrogen as sole electron donor and sulfate as electron acceptor. Based on phylogenetic, genomic and metabolic characteristics we propose to name this chemolithoautotrophic sulfate reducer Candidatus Desulfofervidus auxilii. Ca . D. auxilii grows on hydrogen at temperatures between 50°C and 70°C with an activity optimum at 60°C and doubling time of 4–6 days. Its genome draft encodes for canonical sulfate reduction, periplasmic and soluble hydrogenases and autotrophic carbon fixation via the reductive tricarboxylic acid cycle. The presence of genes for pili formation and cytochromes, and their similarity to genes of Geobacter spp., indicate a potential for syntrophic growth via direct interspecies electron transfer when the organism grows in consortia with ANME. This first ANME‐free enrichment of an AOM partner bacterium and its characterization opens the perspective for a deeper understanding of syntrophy in anaerobic methane oxidation.

Summary Termite gut flagellates are typically colonized by specific bacterial symbionts. Here we describe the phylogeny, ultrastructure and subcellular location of ‘ Candidatus Adiutrix intracellularis’, an intracellular symbiont of Trichonympha collaris in the termite Zootermopsis nevadensis . It represents a novel, deep‐branching clade of uncultured Deltaproteobacteria widely distributed in intestinal tracts of termites and cockroaches. Fluorescence in situ hybridization and transmission electron microscopy localized the endosymbiont near hydrogenosomes in the posterior part and near the ectosymbiont ‘ Candidatus Desulfovibrio trichonymphae’ in the anterior part of the host cell. The draft genome of ‘ Ca . Adiutrix intracellularis’ obtained from a metagenomic library revealed the presence of a complete gene set encoding the Wood–Ljungdahl pathway, including two homologs of fdhF encoding hydrogenase‐linked formate dehydrogenases (FDH H ) and all other components of the recently described hydrogen‐dependent carbon dioxide reductase (HDCR) complex, which substantiates previous claims that the symbiont is capable of reductive acetogenesis from CO 2 and H 2 . The close phylogenetic relationship between the HDCR components and their homologs in homoacetogenic Firmicutes and Spirochaetes suggests that the deltaproteobacterium acquired the capacity for homoacetogenesis via lateral gene transfer. The presence of genes for nitrogen fixation and the biosynthesis of amino acids and cofactors indicate the nutritional nature of the symbiosis.