Phytoplasma and phytoplasma diseases: a review of recent research

Numerous yellows-type diseases of plants have been associated with wall-less prokaryote pathogens \u2013 phytoplasma over the last 40 years. These pathogens cannot be grown in axenic culture so that advances in their study are mainly achieved by molecular techniques. Severe disease epidemics associated with a phytoplasma presence have been described worldwide. These include coconut lethal yellowing in Africa and the Caribbean, grapevine yellows in major viticultural areas and various diseases affecting stone and pome fruit plants. Phytoplasma-infected plants exhibit symptoms suggesting a profound disturbance in the normal balance of growth regulators and also yellows symptoms, but very often the symptomatology is not diagnostic. Detection and characterization of phytoplasmas infecting different plant species are now possible with molecular methods, based on the study of 16S rDNA polymorphisms. Molecular diversity of phytoplasmas is also demonstrated by studying genes coding the ribosomal proteins S3, tuf, SecY, amp, imp and other genes. Four phytoplasma genomes have been fully sequenced, including those of two \u2018Candidatus Phytoplasma asteris\u2019 strains, and those of strains of \u2018Ca. P. mali\u2019 and \u2018Ca. P. australiense\u2019. Three of these genomes contain large amounts of repeated DNA sequence, and the fourth carries multiple copies of almost 100 genes. Considering that phytoplasmas have unusually small genomes, these repeats might be related to their transkingdom habitat and to their pathogenic activity. An outlook of recent findings in the field is also repor

Molecular variability and host distribution of ‘candidatus phytoplasma solani’ strains from different geographic origins

The knowledge of phytoplasma genetic variability is a tool to study their epidemiology and to implement an effective monitoring and management of their associated diseases. ‘Candidatus Phytoplasma solani’ is associated with “bois noir” disease in grapevines, and yellowing and decline symptoms in many plant species, causing serious damages during the epidemic outbreaks. The epidemiology of the diseases associated with this phytoplasma is complex and related to numerous factors, such as interactions of the host plant and insect vectors and spreading through infected plant propagation material. The genetic variability of ‘Ca. P. solani’ strains in different host species and in different geographic areas during the last two decades was studied by RFLP analyses coupled with sequencing on vmp1, stamp, and tuf genes. A total of 119 strains were examined, 25 molecular variants were identified, and the variability of the studied genes was linked to both geographic distribution and year of infection. The crucial question in ‘Ca. P. solani’ epidemiology is to trace back the epidemic cycle of the infections. This study presents some relevant features about differential strain distribution useful for disease monitoring and forecasting, illustrating and comparing the phytoplasma molecular variants identified in various regions, host species, and time periods

Anthracnose: A new strawberry disease in Serbia and its control by fungicides

Anthracnose is a destructive disease of strawberry fruits in warm and continental climate. During 2004, in the vicinity of Valjevo, there were severe losses in two strawberry plantations due to fruit anthracnose. Two fungal isolates, GG-6A and GG-JUP were recovered from strawberry stolons and fruits showing severe anthracnose symptoms. Based on morphological and pathological characteristics, and PCR analyses with specific primers of reference species, isolate GG-6A was identified as Colletotrichum gloeosporioides, and GG-JUP isolate as C. acutatum. This is the first identification of C. acutatum in strawberry in Serbia. In order to control strawberry anthracnose, five fungicides and their combinations were applied four times during the flowering. The best fruit protection was achieved by fungicides Metiram + piraclostrobin (Cabrio top), Captan FL and Fludioksinil + ciprodinil (Swich). Less effective were Benomil (Benlate) and Krezoksim-metil (Stroby). Pathogen is transmitted by planting material, so phytosanitary measures are extremely important in preventing the disease

Molecular characterization of ‘Candidatus Phytoplasma mali’ strains in outbreaks of apple proliferation in north eastern Italy, Hungary, and Serbia

During 2005-2008 apple plants of different varieties showing proliferation symptoms were observed in diverse areas of north eastern Italy, Hungary and Serbia. PCR/RFLP analyses showed that all the samples were infected with ‘Candidatus Phytoplasma mali’. In the 16S plus spacer region two phytoplasma profiles (P-I and P-II) were distinguished. P-I profile was detected in reference strains AP, AT1, AT2, in samples from Serbia, and in the majority of samples from Trentino; the P-II profile was prevalent in samples from Veneto; both profiles were identified in samples from Hungary, in some cases together in single samples. The analyses of rpl22-s3 genes allow the identification, in all the samples showing a P-I profile, the presence of phytoplasmas belonging to rpX-A subgroup, while in the samples showing a P-II profile it was possible to distinguish the other three reported rpX subgroups. In the majority of samples from the Veneto region phytoplasmas belonging to rpX-D subgroup were identified, while rpX-B and rpX-C subgroups were identified only in a few samples from Trentino and Veneto regions, respectively. Further RFLP analyses on AP13/AP10 amplicons differentiate among strains belonging to the rpX-A subgroup: the samples from Serbia show AP profiles, while those from Trentino show AT-2 profiles. In the samples from Hungary the presence of AT1, AT2, and AP profiles was identified.Keywords: Apple, ‘Candidatus Phytoplasma mali’, phytoplasma strains, PCR/RFLP analyses, epidemiolog

Complete genome determination and analysis of Acholeplasma oculi strain 19L, highlighting the loss of basic genetic features in the Acholeplasmataceae

BACKGROUND: Acholeplasma oculi belongs to the Acholeplasmataceae family, comprising the genera Acholeplasma and ‘Candidatus Phytoplasma’. Acholeplasmas are ubiquitous saprophytic bacteria. Several isolates are derived from plants or animals, whereas phytoplasmas are characterised as intracellular parasitic pathogens of plant phloem and depend on insect vectors for their spread. The complete genome sequences for eight strains of this family have been resolved so far, all of which were determined depending on clone-based sequencing. RESULTS: The A. oculi strain 19L chromosome was sequenced using two independent approaches. The first approach comprised sequencing by synthesis (Illumina) in combination with Sanger sequencing, while single molecule real time sequencing (PacBio) was used in the second. The genome was determined to be 1,587,120 bp in size. Sequencing by synthesis resulted in six large genome fragments, while the single molecule real time sequencing approach yielded one circular chromosome sequence. High-quality sequences were obtained by both strategies differing in six positions, which are interpreted as reliable variations present in the culture population. Our genome analysis revealed 1,471 protein-coding genes and highlighted the absence of the F(1)F(O)-type Na(+) ATPase system and GroEL/ES chaperone. Comparison of the four available Acholeplasma sequences revealed a core-genome encoding 703 proteins and a pan-genome of 2,867 proteins. CONCLUSIONS: The application of two state-of-the-art sequencing technologies highlights the potential of single molecule real time sequencing for complete genome determination. Comparative genome analyses revealed that the process of losing particular basic genetic features during genome reduction occurs in both genera, as indicated for several phytoplasma strains and at least A. oculi. The loss of the F(1)F(O)-type Na(+) ATPase system may separate Acholeplasmataceae from other Mollicutes, while the loss of those genes encoding the chaperone GroEL/ES is not a rare exception in this bacterial class. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/1471-2164-15-931) contains supplementary material, which is available to authorized users

First Report of 'Candidatus Phytoplasma solani' Infecting Parsnip in Serbia

The parsnip is a biennial plant native to Eurasia. During August and September 2016, typical phytoplasma yellows symptoms were observed on parsnip plants grown in Serbia. To our knowledge, this is the first report of phytoplasma infecting parsnip in Serbia

Identification and ecology of alternative insect vectors of ‘Candidatus Phytoplasma solani’ to grapevine

Bois noir, a disease of the grapevine yellows complex, is associated with 'Candidatus Phytoplasma solani' and transmitted to grapevines in open fields by the cixiids Hyalesthes obsoletus and Reptalus panzeri. In vine-growing areas where the population density of these vectors is low within the vineyard, the occurrence of bois noir implies the existence of alternative vectors. The aim of this study was to identify alternative vectors through screening of the Auchenorrhyncha community, phytoplasma typing by stamp gene sequence analyses, and transmission trials. During field activities, conducted in Northern Italy in a vineyard where the bois noir incidence was extremely high, nine potential alternative insect vectors were identified according to high abundance in the vineyard agro-ecosystem, high infection rate, and harbouring phytoplasma strains characterized by stamp gene sequence variants found also in symptomatic grapevines. Transmission trials coupled with molecular analyses showed that at least eight species (Aphrodes makarovi, Dicranotropis hamata, Dictyophara europaea, Euscelis incisus, Euscelidius variegatus, Laodelphax striatella, Philaenus spumarius, and Psammotettix alienus/confinis) are alternative vectors of 'Candidatus Phytoplasma solani' to grapevines. These novel findings highlight that bois noir epidemiology in vineyard agro-ecosystems is more complex than previously known, opening up new perspectives in the disease management