Yaima Arocha-Rosete

Identification and molecular characterization of the phytoplasma associated with peach rosette-like disease at the Canadian Clonal Genebank based on the 16S rRNA gene analysis

Abstract Peach trees exhibiting peach rosette-like disease symptoms and infected by a phytoplasma of group 16SrI ‘Candidatus Phytoplasma asteris’ at the Canadian Clonal Genebank were further tested for the pathogen characterization based on the 16S rRNA gene. Nested PCR with phytoplasma universal primers R16mF2/R1 and R16F2n/R2 resulted in amplification of products of approximately 1.25 kb from all four symptomatic trees tested. Virtual RFLP of the R16F2n/R2 sequenced amplicons with selected restriction endonucleases showed unique RFLP patterns when compared to the described 16SrI phytoplasma subgroups; these data were confirmed by phylogenetic analyses. The phytoplasma was therefore assigned as a member of a new 16SrI subgroup (16SrI-W). Results represent the first report of a new phytoplasma 16SrI subgroup infecting peach in Canada, and provide a valuable tool for further epidemiological studies on this phytoplasma in peach.

Identification and molecular characterization of the phytoplasma associated with a lethal yellowing-type disease of coconut in Côte d’Ivoire

Abstract Coconut palms from the Grand-Lahou region of Côte d’Ivoire exhibiting lethal yellowing-like symptoms and infected by a phytoplasma were further tested for the characterization of the phytoplasma pathogen based on the 16S rRNA gene analysis. Nested PCR with phytoplasma universal and lethal yellowing-specific primers resulted in amplification of products of expected sizes from seven out of 17 symptomatic trees tested, while non-infected symptomless plants yielded no DNA amplification. The 16S rDNA sequence of the phytoplasma detected from Grand-Lahou in coconut trees (KC999037) affected with the Côte d’Ivoire Lethal Yellowing disease showed a 99% sequence identity with that of the Ghanaian lethal yellowing phytoplasma (Cape St. Paul Wilt strain) of group 16SrXXII. The 16S rDNA sequence-based virtual and actual RFLP and phylogenetic analyses revealed that the phytoplasma associated with Côte d’Ivoire Lethal Yellowing in Grand Lahou is as a member of a new subgroup within group 16SrXXII that was designated as 16SrXXII-B. The present results support previous suspicions of disease spread to Côte d’Ivoire from neighbouring Ghana, posing a threat to the survival of the coconut trees in the Grand-Lahou region and for the Ivorian coconut industry.

Identification of Graminella nigrifrons as a potential vector for phytoplasmas affecting Prunus and Pyrus species in Canada

Abstract Prunus and Pyrus species affected with phytoplasma diseases, as well as leafhopper species collected from Prunus and Pyrus fields in Ontario, Canada were tested for presence of phytoplasmas. Preliminary results showed that Graminella nigrifrons is a potential vector for phytoplasma groups 16SrI-W (‘Candidatus Phytoplasma asteris’), and 16SrVII-A (‘Candidatus Phytoplasma fraxini’) to a variety of plant hosts, including peach, apricot, plum and pear. Results showed that G. nigrifrons may be able to transmit both phytoplasma groups simultaneously within the same location and suggest that G. nigrifrons populations appear to have a complex ecology.

Detection and quantification of ‘Candidatus Phytoplasma asterisʼ and ‘Candidatus Liberibacter asiaticusʼ at early and late stages of Huanglongbing disease development

Abstract ʻCandidatus Phytoplasma asterisʼ (CPa) has been identified along with ʻCandidatus Liberibacter asiaticusʼ (CLas) from citrus trees showing Huanglongbing (HLB) symptoms in Mexico. The low titre of CPa and CLas in infected plants is the major constraint to developing an accurate and sensitive method for their detection that would support an effective HLB diagnosis and management programme. This investigation reports the development and validation of a new TaqMan quantitative PCR to detect low titres of CPa in citrus trees with HLB symptoms based on the phytoplasma 16S rRNA gene region. The assay was used to monitor the titre of CPa at early and late stages of disease development in symptomatic and asymptomatic Mexican lime trees in Colima, Mexico. The concentration of CLas was determined by nested-qPCR. Only eight out of 39 sampled branches showed HLB symptoms at the early infection stage. All 39 branches were symptomatic at the late infection stage. The qPCR yielded an increase in the branch incidence and concentration of CLas (49 to 100% and 5 × 103 to 12 × 103 cells/100 ng of total DNA), while CPa showed a decreased incidence and concentration (72to 41% and 199 to 19 cells/100 ng of total DNA) throughout the disease development. Results confirm the occurrence of mixed infection of CLas and CPa in HLB-affected citrus trees, although their epidemiological role in co-infecting remains unknown.

Multilocus DNA sequencing of the whiskey fungus reveals a continental‐scale speciation pattern

Baudoinia was described to accommodate a single species, B. compniacensis. Known as the ‘whiskey fungus’, this species is the predominant member of a ubiquitous microbial community known colloquially as ‘warehouse staining’ that develops on outdoor surfaces subject to periodic exposure to ethanolic vapours near distilleries and bakeries. Here we examine 19 strains recovered from environmental samples near industrial settings in North America, South America, the Caribbean, Europe and the Far East. Molecular phylogenetic analysis of a portion of the nucLSU rRNA gene confirms that Baudoinia is a monophyletic lineage within the Teratosphaeriaceae (Capnodiales). Multilocus phylogenetic analysis of nucITS rRNA (ITS1-5.8S-ITS2) and partial nucLSU rRNA, beta-tubulin (TUB) and elongation factor 1-alpha (TEF1) gene sequences further indicates that Baudoinia consists of five strongly supported, geographically patterned lineages representing four new species (viz. Baudoinia antilliensis, B. caledoniensis, B. orientalis and B. panamericana).

New phytoplasma subgroup identified from Arecaceae palm species in Grand-Lahou, Côte d’Ivoire

Abstract Coconut palms in the south coastal littoral of Grand-Lahou have long been affected by the Côte d’Ivoire lethal yellowing (CILY), associated with subgroup 16SrXXII-B ‘Candidatus Phytoplasma palmicola’-related strains. Recently, foliar discolouration and yellowing symptoms were observed in oil palms (Elaeis guineensis), Rônier palms (Borasus aethiopium), and raffia palms (Raphia vinifera) located in the vicinity of coconut palms (Cocos nucifera) showing CILY-like symptoms in coconut-growing villages of Grand-Lahou. Immature leaf samples were randomly collected from symptom-bearing palms, including coconut palms. Total DNA was extracted and used as a template for a nested PCR assay with universal primers that target the phytoplasma 16S rRNA gene, and the translocation protein (secA) genes. The R16F2n/R2 and SecA amplicons from representative samples of each palm species were purified, cloned, and sequenced. R16F2n/R2 sequences were over 99% identical to phytoplasmas enclosed in group 16SrXXII. Based on iPhyclassifier analyses with 17 restriction enzymes, the phytoplasmas found in C. nucifera, E. guineensis, B. aethiopium and R. vinifera were identical to each other. However, a unique Sau3AI RFLP pattern was observed, and similarity coefficients were lower than 0.97, and so a new 16SrXXII subgroup, 16SrXXII-C, was assigned. Phylogeny based on both R16F2n/R2 and SecA sequences supported this RFLP classification. The presence of the subgroup 16SrXXII-C in the same area where coconut palms are severely infected by the subgroup 16rSXXII-B could lead to different epidemiological constraints that may affect the severity of the coconut disease. Thus, the 16SrXXII-C phytoplasma poses a new threat for other Arecaceae species, which may impact CILY management and control in Grand-Lahou.