Plant Science

‘Candidatus Phytoplasma spp.’ are pathogenic bacteria that infect many plant species. ‘Candidatus Phytoplasma pyri’, one of the members of the 16SrX group causes pear decline disease that adversely affects pear crops. To describe the prevalence of ‘Ca. P. pyri’ genotypes in the Czech Republic, 143 pear samples were collected from 41 locations including commercial orchards as well as trees along roads. Phytoplasma was detected by PCR in 115 samples, and it was possible to determine imp gene genotype in 84 samples. The most frequent genotypes were A1, B1, and C, which were identified in 71% of phytoplasma positive samples. ‘Ca. P. pyri’ was present either alone or as a mix of two populations in 88% of genotyped samples, and in another 6% of samples it was found in a mixed infection with ‘Ca. Phytoplasma mali’. A sole infection with ‘Ca. Phytoplasma mali’ was observed in 6% of samples. As for symptoms, 19% of symptomatic samples were found to be phytoplasma negative, and 74% of asymptomatic samples proved to be phytoplasma positive; leaf roll was more often observed in phytoplasma positive samples, while leaf narrowing rather indicated the absence of phytoplasma. The mildest symptoms were observed in samples infected with ‘Ca. P. pyri’ of the A1 imp genotype.

Amycolatopsis coloradensis ATCC 53629 is the producer of the glycopeptide antibiotic avoparcin. While setting up the production of the avoparcin complex, in view of its use as analytical standard, we uncovered the production of a to-date not described ristosamynil-avoparcin. Ristosamynil-avoparcin is produced together with α- and β-avoparcin (overall indicated as the avoparcin complex). Selection of one high producer morphological variant within the A. coloradensis population, together with the use of a new fermentation medium, allowed to increase productivity of the avoparcin complex up to 9 g/L in flask fermentations. The selected high producer displayed a non-spore forming phenotype. All the selected phenotypes, as well as the original unselected population, displayed invariably the ability to produce a complex rich in ristosamynil-avoparcin. This suggested that the original strain deposited was not conforming to the description or that long term storage of the lyovials has selected mutants from the original population.

Huanglongbing (HLB) has turned into a devastating botanical pandemic of citrus crops, caused by Candidatus Liberibacter asiaticus (CLas). However, until now the disease has remained incurable with very limited control strategies available. Restoration of the affected microbiomes in the diseased host through the introduction of an indigenous endophyte Bacillus subtilis L1-21 isolated from healthy citrus may provide an innovative approach for disease management. A novel half-leaf method was developed in vitro to test the efficacy of the endophyte L1-21 against CLas. Application of B. subtilis L1-21 at 104 colony forming unit (cfu ml−1) resulted in a 1,000-fold reduction in the CLas copies per gram of leaf midrib (107 to 104) in 4 days. In HLB-affected citrus orchards over a period of 2 years, the CLas incidence was reduced to < 3%, and CLas copies declined from 109 to 104 g−1 of diseased leaf midribs in the endophyte L1-21 treated trees. Reduction in disease incidence may corroborate a direct or an indirect biocontrol effect of the endophytes as red fluorescent protein-labeled B. subtilis L1-21 colonized and shared niche (phloem) with CLas. This is the first large-scale study for establishing a sustainable HLB control strategy through citrus endophytic microbiome restructuring using an indigenous endophyte.

‘Candidatus Liberibacter asiaticus’ (CLas), the devastating pathogen related to Huanglongbing (HLB), is a phloem-limited, fastidious, insect-borne bacterium. Rapid spread of HLB disease relies on CLas-efficient propagation in the vector, the Asian citrus psyllid Diaphorina citri, in a circulative manner. Understanding the intracellular lifecycle of CLas in psyllid midgut, the major organ for CLas transmission, is fundamental to improving current management strategies. Using a microscopic approach within CLas-infected insect midgut, we observed the entry of CLas into gut cells inside vesicles, termed Liberibacter-containing vacuoles (LCVs), by endocytosis. Endocytosis is followed by the formation of endoplasmic reticulum-related and replication permissive vacuoles (rLCVs). Additionally, we observed the formation of double membrane autophagosome-like structure, termed autophagy-related vacuole (aLCV). Vesicles containing CLas egress from aLCV and fuse with the cell membrane. Immunolocalization studies showed that CLas uses endocytosis- and exocytosis-like mechanisms that mediates bacterial invasion and egress. Upregulation of autophagy-related genes indicated subversion of host autophagy by CLas in psyllid vector to promote infection. These results indicate that CLas interacts with host cellular machineries to undergo a multistage intracellular cycle through endocytic, secretory, autophagic, and exocytic pathways via complex machineries. Potential tactics for HLB control can be made depending on further investigations on the knowledge of the molecular mechanisms of CLas intracellular cycle.

Candidatus Liberibacter spp. are fastidious α-proteobacteria that cause multiple diseases on plant hosts of economic importance, including the most devastating citrus disease: Huanglongbing (HLB). HLB was reported in Asia a century ago but has since spread worldwide. Understanding the pathogenesis of Candidatus Liberibacter spp. remains challenging as they are yet to be cultured in artificial media and infect the phloem, a sophisticated environment that is difficult to manipulate. Despite those challenges, tremendous progress has been made on Ca. Liberibacter pathosystems. Here, we first reviewed recent studies on genetic information of flagellar and type IV pili biosynthesis, their expression profiles, and movement of Ca. Liberibacter spp. inside the plant and insect hosts. Next, we reviewed the transcriptomic, proteomic, and metabolomic studies of susceptible and tolerant plant genotypes to Ca. Liberibacter spp. infection and how Ca. Liberibacter spp. adapt in plants. Analyses of the interactions between plants and Ca. Liberibacter spp. imply the involvement of immune response in the Ca. Liberibacter pathosystems. Lastly, we reviewed how Ca. Liberibacter spp. movement inside and interactions with plants lead to symptom development.