Publications

4468

AbstractThe epidemiology of ‘Candidatus Phytoplasma prunorum’ was studied in Austria and Hungary from 2014 to 2018. Testing of root samples showed average infections rates of 61 and 40% of the Austrian Prunus spinosa and Prunus domestica spp. insititia samples, respectively. In Hungary, on average 21% of the P. spinosa and 13% of the feral Prunus cerasifera samples were infected. The pathogen was found in 18 out of 19 apricot orchards and PCR positive Cacopsylla pruni were observed at 11 out of 17 sampling locations in both countries. In cage experiments with C. pruni remigrants successful pathogen transmission to Prunus armeniaca, P. domestica and P. spinosa seedlings in budding and foliated developmental stages was recorded, an inoculation access period of 4 hr was sufficient for transmission. A field experiment with ungrafted apricot seedlings planted in 2012 and 2014 indicated a prominent role of the insect vectors for disease spread. In 2017, 40 and 28% of the trees planted in 2012 and 2014, respectively, were infected. Molecular characterisation based on the genes aceF and imp allowed the discrimination between 10 phytoplasma types in apricots. Around 70% of the phytoplasma types in apricots were also common in P. spinosa, in P. domestica spp. insititia and in remigrant C. pruni pointing to a possible pathogen exchange by insects between wild and cultivated Prunus spp. For disease control, vector management over the entire flight period of the remigrants seems necessary; when selecting active compounds, the short inoculation access period of not more than 4 hr should be considered.

Abstract The most abundant aquatic microbes are small in cell and genome size. Genome-streamlining theory predicts gene loss caused by evolutionary selection driven by environmental factors, favouring superior competitors for limiting resources. However, evolutionary histories of such abundant, genome-streamlined microbes remain largely unknown. Here we reconstruct the series of steps in the evolution of some of the most abundant genome-streamlined microbes in freshwaters (“Ca. Methylopumilus”) and oceans (marine lineage OM43). A broad genomic spectrum is visible in the family Methylophilaceae (Betaproteobacteria), from sediment microbes with medium-sized genomes (2–3 Mbp genome size), an occasionally blooming pelagic intermediate (1.7 Mbp), and the most reduced pelagic forms (1.3 Mbp). We show that a habitat transition from freshwater sediment to the relatively oligotrophic pelagial was accompanied by progressive gene loss and adaptive gains. Gene loss has mainly affected functions not necessarily required or advantageous in the pelagial or is encoded by redundant pathways. Likewise, we identified genes providing adaptations to oligotrophic conditions that have been transmitted horizontally from pelagic freshwater microbes. Remarkably, the secondary transition from the pelagial of lakes to the oceans required only slight modifications, i.e., adaptations to higher salinity, gained via horizontal gene transfer from indigenous microbes. Our study provides first genomic evidence of genome reduction taking place during habitat transitions. In this regard, the family Methylophilaceae is an exceptional model for tracing the evolutionary history of genome streamlining as such a collection of evolutionarily related microbes from different habitats is rare in the microbial world.

SUMMARYCandidatus Liberibacter spp. are Alphaproteobacteria associated with plants and psyllid vectors. Most cause plant diseases, including Ca Liberibacter asiaticus (Las) associated with citrus huanglongbing (HLB). Replacing HLB-infected by Las-free citrus trees results in fast re-infection despite psyllid control. To check if HLB could be soil-borne, we performed an insect-free greenhouse-experiment with 130 mandarin seedlings in two citrus-grove soils (A and B), non-autoclaved or autoclaved. Liberibacter-specific 16S-rDNA PCR primers to detect Las were used to search for Ca. Liberibacter spp. in mandarin leaves. Seven plants grown in non-autoclaved soil B showed HLB-like symptoms and tested positive after 2.5 and 8.5 months using three different primer systems: two based on the 16S-rDNA gene (primers HLBas/HLBr and OI2c/OI1) and one based on the rplA/rplJ gene (primers LAA2/LAJ5). DNA segments from these plants amplified by primers OI2c/OI1 were cloned and sequenced; they were 95.9 % similar to Las and 94.8% to Ca. Liberibacter africanus (Laf). The DNA product from Liberibacter-group specific PCR primers for the rplA/rplJ gene was 87.6% similar to that of Las and 78.2% of Laf. As the strain obtained originated from soil and was different from existing Ca. Liberibacter species, this strain may be a new species.

During 2015, samples from 22 apple trees showing proliferation symptoms were collected in southwest Bulgaria and Central and South Poland and tested for phytoplasma presence. ‘Candidatus Phytoplasma mali’ was identified in 18 samples based on results of restriction fragment length polymorphism (RFLP) analysis of the 16S rRNA gene amplified in nested PCR using primer pair P1/P7 followed by R16F2n/R16R2 and F1/B6 primer pairs. The nitroreductase and rhodonase like genes and ribosomal protein genes rpl22 and rps3 were then analyzed using PCR-RFLP technique to study the genetic variability of the  phytoplasma strains. Two restriction profiles, P-I or P-II, were obtained for fragments of 16S rDNA plus 16S-23S spacer region digested with HpaII enzyme. Restriction fragment length polymorphism analysis of nitroreductase and rhodonase like genes using digestion with HincII endonuclease revealed that the all ‘Ca. P. mali’ strains belonged to subtype AP-15. Analysis of rpl22 and rps3 ribosomal protein genes digested with AluI enzyme resulted in classification of detected phytoplasma strains to rpX-A subgroup.