General Immunology and Microbiology

‘Candidatus Phytoplasma solani’ (‘Ca. P. solani’) is a crop pathogen that is a member of the 16SrXII-A ribosomal subgroup. It is also known as stolbur phytoplasma and causes yield losses in several important crops, especially in Solanaceous crops. Different strains of the pathogen are regularly reported all over the world, particularly in the Mediterranean region. In this study, the determination of genetic diversity for the pathogen infecting tomatoes and potatoes was carried out by using multilocus sequence typing analysis for the Tuf, SecY, and Vmp1 genes to gain insight into the epidemiology of ‘Ca. P. solani’ in Turkey. Genetic diversity of the phytoplasmas was investigated by sequence-based phylogenetic analyses and in silico RFLP analysis of related genes. It was determined that all ‘Ca. P. solani’-related strains infecting tomatoes and potatoes were tuf-b, which is linked to field bindweed (Convolvulus arvensis L.). Tomato or potato-infecting ‘Ca. P. solani’-related strains showed similarities with each other; however, the isolates collected from different plants showed genetic differences in terms of the SecY gene. This study indicates that the highest genetic variability of collected samples was found in the Vmp1 gene. RsaI-RFLP analysis of TYPH10F/R amplicons showed that potato-infecting ‘Ca. P. solani’-related strains were found to be similar to some existing V types. However, the V-type of tomato-infecting isolates is not similar to any previously reported V-type. The results indicate that there could be an important genetic diversity of ‘Ca. P. solani’-related phytoplasmas in Turkey. This could indicate various ways in which the pathogen has adapted to the two host plants as a consequence of the various Vmp1 gene rearrangements seen in these two plant hosts. Obtained results also indicate that the epidemiology of ‘Ca. P. solani’-related phytoplasmas in the tomato and potato agroecosystem may be better understood with the use of molecular data on the complex of vmp-types.

The pathogens associated with citrus Huanglongbing symptoms, including yellowing and mottled leaves in Citrus maxima, an important economic crop on Hainan Island of China, were identified and characterized. In the study, detection, genetic variation and phylogenetic relationship analysis of the pathogens were performed based on 16S rRNA and β-operon gene fragments specific to phytoplasma and Candidatus Liberibacter asiaticus. The results indicated that the pathogens—such as phytoplasma strains of CmPII-hn belonging to the 16SrII-V subgroup and CmPXXXII-hn belonging to the 16SrXXXII-D subgroup, as well as Candidatus Liberibacter asiaticus strains CmLas-hn—were identified in the diseased plant samples, with numbers of 12, 2 and 6 out of 54, respectively. Among them, mixed infection with the 16SrII-V subgroup phytoplasma and Candidatus Liberibacter asiaticus was found in the study, accounting for 7.4% (four samples). The phytoplasma strains of CmPII-hn—Tephrosia purpurea witches’ broom, Melochia corchorifolia witches’ broom and Emilia sonchifolia witches’ broom—were clustered into one clade belonging to the 16SrII-V subgroup, with a 99% bootstrap value. The phytoplasma strains of CmPXXXII-hn and Trema tomentosa witches’ broom belonging to 16SrXXXII-D, and the other 16SrXXXII subgroup strains were clustered into one clade belonging to the 16SrXXXII group with a 99% bootstrap value. There were 16 variable loci in the 16S rRNA gene sequences of the tested 16SrXXXII group phytoplasma strains, of which two bases had an insertion/deletion. The strains of Candidatus Liberibacter asiaticus, identified in the study and the strains that had been deposited in GenBank, were in one independent cluster with a 99% bootstrap value. To our knowledge, this is the first report showing that Citrus maxima can be infected by 16SrII-V and16SrXXXII-D subgroup phytoplasmas in China. Moreover, this is also the first report in which the plants are co-infected by 16SrII-V subgroup phytoplasmas and Candidatus Liberibacter asiaticus. More comprehensive and detailed identification and characterization of the pathogens associated with the diseased symptoms in Citrus maxima on the island in China would be beneficial for epidemic monitoring and for the effective prevention and control of related plant diseases.

Grapevine Bois noir (BN) is associated with ‘Candidatus Phytoplasma solani’. It has been recorded in vineyards throughout Europe as well as in different countries in Asia, where it now constitutes a threat to Iranian viticulture. BN is strictly dependent on ‘Ca. P. solani’ strains, wild host plants, and insect vectors. The molecular typing of ‘Ca. P. solani’, based on the nonribosomal gene tuf and the two hypervariable markers vmp1 and stamp, is valuable for the reconstruction and clarification of the pathways of BN spread. In this study, an RFLP analysis was performed on the vmp1 gene, and a single-nucleotide polymorphism analysis confirmed new vmp types in ‘Ca. P. solani’. A stamp gene phylogenetic analysis allowed us to distinguish between the new genotype infections in the grapevines and the ‘weeds’ Convolvulus arvensis and Erigeron bonariensis in Iranian vineyards, highlighting the close genetic relatedness of the strains of ‘Ca. P. solani’ found in Iran and Azerbaijan. The most common genotype in the grapevines was tuf b/V24/stamp III, which was associated with C. arvensis. This information contributes toward the identification of further routes of introduction of ‘Ca. P. solani’ in Iran to sustain the control measures for the management of BN.

The considerable economic losses in citrus associated with ‘Candidatus Liberibacter’ and ‘Candidatus Phytoplasma’ presence have alerted all producing regions of the world. In Chile, none of these bacteria have been reported in citrus species. During the years 2017 and 2019, 258 samples presenting symptoms similar to those associated with the presence of these bacteria were examined. No detection of ‘Ca. Liberibacter’ associated with “huanglongbing” disease was obtained in the tested samples; therefore, this quarantine pest is maintained as absent in Chile. However, 14 plants resulted positive for phytoplasmas enclosed in subgroups 16SrV-A (12 plants) and 16SrXIII-F (2 plants). Although they have been found in other plant species, this is the first report of these phytoplasmas in citrus worldwide.

Florida citrus production has declined 75% due to Huanglongbing (HLB), a disease caused by the pathogenic bacterium Candidatus Liberibacter asiaticus (CLas). Methods to combat CLas are costly and only partially effective. The cross-compatible species Poncirus trifoliata and some of its hybrids are known to be highly tolerant to CLas, and thus can potentially serve as an alternative feedstock for many citrus products. To further investigate the commercial potential of citrus hybrids, three citrus hybrids, US-802, US-897, and US-942, were studied for their potential as feedstocks for citrus co-products using steam explosion (STEX) followed by water extraction. Up to 93% of sugars were recovered. US-897 and US-942 have similar volatile profiles to that of the commercial citrus fruit types and as much as 85% of these volatiles could be recovered. Approximately 80% of the pectic hydrocolloids present in all three hybrids could be obtained in water washes of STEX material. Of the phenolics identified, the flavanone glycosides, i.e., naringin, neohesperidin, and poncirin were the most abundant quantitatively in these hybrids. The ability to extract a large percentage of these compounds, along with their inherent values, make US-802, US-897, and US-942 potentially viable feedstock sources for citrus co-products in the current HLB-blighted environment.

The γ-proteobacterium ‘Candidatus Arsenophonus phytopathogenicus’ is assigned as the major pathogen of “Syndrome des basses richesses”, a sugar beet disease characterised by a reduction in the sugar content of taproots and biomass yield. Despite the economic impact of this bacteriosis, diagnostics for this important pathogen currently rely on end-point PCR detection. Herein, we introduce a TaqMan qPCR for diagnostics of the agent targeting genes encoding a heat shock protein of the Hsp20 family and mannose-6-phosphate isomerase. Quantitation with synthetic oligonucleotides as standard showed that the developed TaqMan qPCR assays enable the detection of up to 100 target copies. A comparison between the TaqMan qPCR and end-point PCR for ‘Ca. A. phytopathogenicus’ detection was carried out on 78 sugar beet samples from different locations in southern Germany. The newly developed assays enable the fast, reliable and sensitive detection of ‘Ca. A. phytopathogenicus’ in sugar beet.