Publications

4603

‘Candidatus Liberibacter asiaticus’ (CLas) is a devastating pathogen of citrus associated with Huanglongbing (HLB, citrus greening disease). HLB is economically significant in Asia and has destroyed millions of citrus trees worldwide during the last century. Since 2007, when the first molecular evidence for the presence of CLas in North-West Frontier Province of Pakistan (Khyber Pakhtunkhwa) were presented, there have been a limited number of studies reporting the pathogen in different citrus growing districts of the Punjab Province, one of the major citrus producing areas of the country. In this study, a small-scale survey was conducted in citrus groves exhibiting HLB-like symptoms in different districts of Punjab Province. The aim was to obtain current information on the incidence of CLas in the area and complement the previous studies. Conventional and real time quantitative polymerase chain reaction (PCR and qPCR) assays were used for the detection of CLas in the 94 samples tested, followed by sequencing of the PCR amplicons. Overall CLas was detected in 43% (40/94) of the tested samples from Kinnow mandarin (n= 22), sweet orange (n= 12), grapefruit (n= 3) and Feutrell’s early mandarin (n= 3). In agreement with previous reports, this survey confirmed the presence of HLB in the Punjab Province of Pakistan and provided updated molecular evidence for the presence of CLas and disease incidence for four citrus cultivars from five citrus growing districts of the province.

ABSTRACT Citrus huanglongbing (HLB), caused by the phloem‐limited bacterium ‘ Candidatus Liberibacter asiaticus’ ( C Las), is one of the most destructive diseases in global citrus production. Here, we report the functional characterization of a core C Las effector, CLIBASIA_00525 named SDE525, that manipulates host immunity by targeting the citrus nascent polypeptide‐associated complex alpha subunit (CsNACα). We confirmed that SDE525 is a secreted protein that localizes to the nucleus, cytoplasm, and plasma membrane. Notably, the effect of SDE525 exhibits host‐specific context dependence: while its transient expression in the non‐host Nicotiana benthamiana unexpectedly triggered immune responses, in its native host, sweet orange ( Citrus sinensis ), SDE525 interacted with CsNACα to modulate defence, as demonstrated by multiple independent assays. Integrated transcriptomic and metabolomic analyses revealed that this interaction reprograms the host's hormonal landscape, suppressing the jasmonic acid (JA) signalling pathway while simultaneously activating the salicylic acid (SA) pathway. Importantly, under transient overexpression conditions in citrus, this immune reprogramming enhanced resistance against the bacterial canker pathogen Xanthomonas citri subsp. citri —a phenomenon distinct from the susceptibility observed during authentic C Las infection. Our findings uncover a sophisticated mechanism by which a C Las effector rewires host defence signalling, providing fundamental insights into its pathogenesis with potential long‐term implications for future HLB management.

Sweet potato witches’ broom disease caused by phytoplasmas threatens crop productivity in Fujian, China. Here, we report a complete phytoplasma genome consisting of a single circular chromosome (680,179 bp with a G + C content of 29.44%) via the next-generation sequencing of sweet potato witches’ broom-infected samples. A total of 730 genes were predicted, and two 16S rRNAs were revealed to be identical to sequences in ‘ Candidatus Phytoplasma australasiaticum’. Following the latest guidelines for the description of phytoplasma species, we have named the strain ‘ Candidatus Phytoplasma australasiaticum’ SPWBPT2024. The genome contained 33 transporters, 35 putative effectors (including SAP11 and SAP54 with multiple copies), and 2 potential mobile units. Analyses of published peanut and sweet potato phytoplasma genomes revealed similarities between their effectors and the effectors in this phytoplasma genome. A motif analysis was performed to clarify the difference between the two SAP54 copies in the obtained phytoplasma genome, as well as the trends in conserved motifs in other strains. In addition, metabolic pathway-related genes in three genomes were examined, revealing deficiencies in the glycolysis and pyrimidine synthesis pathways. A pangenome analysis revealed that the phytoplasma genome is an open genome. To gain further insights into whole-cell metabolic processes, we reconstructed draft genome-scale metabolic models of the genome. This study provides valuable genomic insights into this phytoplasma, characterizing conserved and strain-specific SAP effectors, as well as distinct metabolic deficiencies that may help advance research to combat phytoplasma infections, including accurate diagnostics, vector transmission control, and antimicrobial interventions.

'Candidatus Phytoplasma solani' is a distinct species of the class Mollicutes (16SrXII group), characterized by phloem-limited localization and strong host dependence. It is widely distributed, mainly in Europe, and affects over 100 plant species, including economically important crops for Moldova. The name reflects its significant impact on plants of the Solanaceae family, causing stolbur disease. It also infects grapevine, pepper, maize, lavender, and other crops. Climate change has favored the spread of the pathogen and its insect vectors into new regions and host plants. Therefore, monitoring and controlling its dissemination in economically important crops is essential. Accurate diagnosis can be achieved by molecular methods, with PCR-based techniques offering a reliable and cost-effective approach. The aim of the study was to determine the presence or absence of infection in some agricultural crops: tomato, maize, potato, pepper, eggplant, wheat, plum and apple growing in experimental fields of Institute of Genetics, Physiology and Plant Protection, MSU. Materials and methods: Various DNA extraction methods were used depending on plant culture: boiling in alkaline solution, DNAzol, SDS, CTAB. Nested PCR analysis was carried out using specific to 'Ca. P. solani' primers designed on the base of chaperonine or 16S rRNA gene sequences. Amplification results were recorded using gel agarose electrophoresis (1.5% agarose, 0.5 x TBE buffer) stained with ethidium bromide and visualized in UV. Results: A comprehensive study of tomato conducted during 2016-2025 showed the continuous presence of the pathogen, with infection rates reaching up to 72% depending on annual conditions, confirming the high susceptibility of tomato to 'Ca. P. solani'. In potato, 14.6% of plants were infected, while sweet pepper showed a much lower incidence of 4%. Overall, the results indicate the predominant distribution of the pathogen within the Solanaceae family, further supported by its detection in the ruderal species Solanum nigrum. The phytoplasma was not detected only in eggplant among the analyzed Solanaceae species. In contrast, molecular diagnostics gave negative results in crops from other families, including maize and wheat (cereals), as well as apple and plum (fruit crops). The pathogen was not detected in orchard trees during either the dormant or fruit-bearing stages. Conclusions: Thus, obtained results demonstrate an ecological and epidemiological specificity of 'Ca. P. solani' for certain host species, characterized by a high incidence in Solanaceae, particularly tomato. Additionally, the study confirms the potential role of ruderal species within the same family as natural reservoirs of the pathogen. These data contribute to a deeper understanding of the spread of the phytoplasma providing a base for effective monitoring and phytosanitary management strategies.