Publications

4476

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.

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 Although metabolic pathways and associated enzymes of anaerobic ammonium oxidation (anammox) of ‘ Ca . Kuenenia stuttgartiensis’ have been studied, those of other anammox bacteria are still poorly understood. reduction to NO is considered to be the first step in the anammox metabolism of ‘ Ca . K. stuttgartiensis’, however, ‘ Ca . Brocadia’ lacks the genes that encode canonical NO‐forming nitrite reductases (NirS or NirK) in its genome, which is different from ‘ Ca . K. stuttgartiensis’. Here, we studied the anammox metabolism of ‘ Ca . Brocadia sinica’. 15 N‐tracer experiments demonstrated that ‘ Ca . B. sinica’ cells could reduce to NH 2 OH, instead of NO, with as yet unidentified nitrite reductase(s). Furthermore, N 2 H 4 synthesis, downstream reaction of reduction, was investigated using a purified ‘ Ca . B. sinica' hydrazine synthase (Hzs) and intact cells. Both the ‘ Ca . B. sinica’ Hzs and cells utilized NH 2 OH and , but not NO and , for N 2 H 4 synthesis and further oxidized N 2 H 4 to N 2 gas. Taken together, the metabolic pathway of ‘ Ca . B. sinica’ is NH 2 OH‐dependent and different from the one of ‘ Ca . K. stuttgartiensis’, indicating metabolic diversity of anammox bacteria.

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.

Abstract Overgrowth of filamentous bacteria in activated sludge wastewater treatment plants (WWTPs) leads to impaired sludge settleability, a condition known as bulking, which is a common operational problem worldwide. Filaments with the Eikelboom 0092 morphotype are commonly associated with such bulking episodes. Members of the uncultured B45 phylotype, which is embraced within the phylum Chloroflexi, were recently shown to exhibit this morphology. Although these organisms are among the most abundant populations recorded in activated sludge processes, nothing is known about their metabolic characteristics. In this study, a genome sequence, representing the B45 phylotype, was retrieved from a metagenome generated from an activated sludge WWTP. The genome consisted of two chromosomes and one plasmid, which were 4.0, 1.0 and 0.04 Mbps in size, respectively. A metabolic model was constructed for this organism, based on annotation of its genome, showing its ability to generate energy by respiration, utilizing oxygen, nitrite or nitrous oxide as electron acceptors, or by fermentation of sugars. The ability of B45 members to ferment sugars under anaerobic conditions was validated in situ with microautoradiography—fluorescence in situ hybridization. The provisional name of ‘Candidatus Promineofilum breve’ is proposed for this species. This study represents the first detailed information on an uncultured genus of filamentous organisms from activated sludge.