Publications

4729

The amino acid‐binding receptors, a component of ABC transporters, have evolved to cater to different specificities and functions. Of particular interest are cystine‐binding receptors, which have shown broad specificity. In the present study, a putative periplasmic cystine‐binding protein from Candidatus Liberibacter asiaticus (CLasTcyA) was characterized. Analysis of the CLasTcyA sequence and crystal structures in the ligand‐bound state revealed novel features of CLasTcyA in comparison to related proteins. One of the unique features found in CLasTcyA structure was the positioning of the C‐terminal extended loop of one chain very close to the substrate‐binding site of the adjacent monomer in the asymmetric unit. The presence of a disulphide bond, unique to Candidatus Liberibacter family, holds the C‐terminal extended loop in position. Analysis of the substrate‐binding pocket of CLasTcyA suggested a broad specificity and a completely different orientation of the bound substrates in comparison to related protein structures. The open conformation for one of the two chains of the asymmetric unit in the Arg‐bound structure revealed a limited open state (18.4°) for CLasTcyA as compared to open state of other related proteins (~ 60°). The strong interaction between Asp126 on helix‐α5 of small domain and Arg82 (bigger domain) restricts the degree of opening in ligand‐free open state. The dissociation constant of 1.26 μm by SPR and 3.7 μm by MST exhibited low affinity for the cystine. This is the first structural characterization of an l‐cystine ABC transporter from plant pathogen and our results suggest that CLasTcyA may have evolved to cater to its specific needs for its survival in the host.

Grapevine (Vitis vinifera L.) is one of the most important horticultural crops in Iran, with >200,000 ha of cultivated area. Recently, outbreaks of the grapevine yellows Bois noir that is associated with phytoplasma strains related to ‘Candidatus Phytoplasma solani’ were recorded in several Iranian regions. This has resulted in severe economic losses. We carried out a survey in 2015, followed by collection of leaf samples from symptomatic grapevines and weeds. Because no information is available on the molecular epidemiology of ‘Ca. P. solani’ in Iran, multiple gene analyses were carried out here according to molecular characterization of the tuf and vmp1 genes. From the molecular characterization, all of the samples (i.e., grapevines, weeds) were infected with tuf b type. Detailed molecular characterization of the vmp1 gene (i.e., PCR–restriction fragment length polymorphism, sequence analysis) defined five molecular types: V1, V4, V10, V15, and V20. The abundance of Convolvulus arvensis in vineyards and detection of the same ‘Ca. P. solani’ molecular types in grapevines and weeds suggest that C. arvensis has a major role in Bois noir epidemiology of Iranian vineyards. Therefore, control strategies should be developed to manage these host plants to reduce inoculum sources of the phytoplasma in vineyards.

Abstract Rickettsiales are a lineage of obligate intracellular Alphaproteobacteria, encompassing important human pathogens, manipulators of host reproduction, and mutualists. Here we report the discovery of a novel Rickettsiales bacterium associated with Paramecium, displaying a unique extracellular lifestyle, including the ability to replicate outside host cells. Genomic analyses show that the bacterium possesses a higher capability to synthesise amino acids, compared to all investigated Rickettsiales. Considering these observations, phylogenetic and phylogenomic reconstructions, and re-evaluating the different means of interaction of Rickettsiales bacteria with eukaryotic cells, we propose an alternative scenario for the evolution of intracellularity in Rickettsiales. According to our reconstruction, the Rickettsiales ancestor would have been an extracellular and metabolically versatile bacterium, while obligate intracellularity would have evolved later, in parallel and independently, in different sub-lineages. The proposed new scenario could impact on the open debate on the lifestyle of the last common ancestor of mitochondria within Alphaproteobacteria.

AbstractThe size of bacterial genomes is often associated with organismal metabolic capabilities determining ecological breadth and lifestyle. The recently proposed Candidate Phyla Radiation (CPR)/Patescibacteria encompasses mostly unculturable bacterial taxa with relatively small genome sizes with potential for co‐metabolism interdependencies. As yet, little is known about the ecology and evolution of CPR, particularly with respect to how they might interact with other taxa. Here, we reconstructed two novel genomes (namely, Candidatus Saccharibacter sossegus and Candidatus Chaer renensis) of taxa belonging to the class Saccharimonadia within the CPR/Patescibacteria using metagenomes obtained from acid mine drainage (AMD). By testing the hypothesis of genome streamlining or symbiotic lifestyle, our results revealed clear signatures of gene losses in these genomes, such as those associated with de novo biosynthesis of essential amino acids, nucleotides, fatty acids and cofactors. In addition, co‐occurrence analysis provided evidence supporting potential symbioses of these organisms with Hydrotalea sp. in the AMD system. Together, our findings provide a better understanding of the ecology and evolution of CPR/Patescibacteria and highlight the importance of genome reconstruction for studying metabolic interdependencies between unculturable Saccharimonadia representatives.

In recent years, ‘Ca. P. prunorum’, the agent of ESFY was reported from several apricot orchards of Canton of Wallis, the main apricot production region in Switzerland (Genini and Ramel, 2004). The psyllid vector Cacopsylla pruni was also found in several locations in the Lake Geneva area (Ackermann et al., 2006). The presence of the disease and of its proven vector C. pruni at the eastern part of the Lake Geneva area, as well as the existing risk of dissemination of ESFY to other stone fruit orchards along the Lake Geneva are the reasons for which ESFY needs to be studied further. ESFY and its possible dissemination through C. pruni on the territory of Canton of Geneva, bordering France, has never been yet studied. In 2016, visual observations were conducted in several stone fruit orchards near Geneva. Plant material was obtained from peach trees, displaying some of the typical symptoms such premature leaf colouration, leaf-roll, tree decline (Sabaté et al., 2015), in the autumn, when the concentration of the phytoplasma in the upper parts of the trees is the highest. Phloem was prepared from branches and was extracted with a CTAB-based adapted protocol (Lefort and Douglas, 1999). PCR amplification of phytoplasma DNA was achieved with the universal primers: fP1/rP7 (Deng and Hiruki, 1991; Schneider et al., 1995). All positive samples were tested with the ESFY-specific non-ribosomal primers ECA1/ECA2 (Jarausch et al., 1998). Typical symptoms of ESFY, like premature leaf colouration, leaf yellowing with reddish edges,  leaf-roll, severe chlorosis, die-back of top branches and partial or, complete decline of the trees (Figure 1) were found in a peach orchard (GPS coordinates: 46°15'17.4"N 6°12'40.7"E) located in the area of Collonge-Bellerive, close to Geneva city. Ten trees were sampled in this orchard. The presence of ‘Ca. P. prunorum’ was confirmed in two of them (Figure 2 and Figure 3). At the beginning of spring 2017, the monitoring of the ESFY symptoms in the infested orchard was resumed. Early bud break was found on the two infected trees and the infection was confirmed again by PCR (Figure 4 and Figure 5). The same symptoms were observed in many trees of this orchard, which correlated with the previous autumn observations. Trapping confirmed the presence of the insect vector Cacopsylla pruni (Bodnár et al., 2018) in the infected area. In order to understand more about the origin and the diversity of ESFY phytoplasma in this area, additional plant and insect samples will be analyzed. The correlation between symptoms and infected plants will be studied further. We report here for the first time on the occurrence of ‘Candidatus Phytoplasma prunorum’ (‘Ca. P. prunorum’) the agent of ESFY on the territory of Canton of Geneva.

Sponges represent ecologically important models to understand the evolution of symbiotic interactions of metazoans with microbial symbionts. Thaumarchaeota are commonly found in sponges, but their potential adaptations to a host-associated lifestyle are largely unknown. Here, we present three novel sponge-associated thaumarchaeal species and compare their genomic and predicted functional features with those of closely related free-living counterparts. We found different degrees of specialization of these thaumarchaeal species to the sponge environment that is reflected in their host distribution and their predicted molecular and metabolic properties. Our results indicate that Thaumarchaeota may have reached different stages of evolutionary adaptation in their symbiosis with sponges.