826 research outputs found
Detection of phytoplasmas in plantlets grown from different batches of seed-potatoes.
No full textDuring 2006 and 2007 eight batches of seed potatoes collected in different locations, and belonging to one cultivar were planted
in spring under greenhouse conditions and tested after 2 months to verify phytoplasma presence. A total of 635 asymptomatic
plantlets were examined. Nucleic acid was extracted from small shoots from either a single plant or from batches of 3 plants each.
Nested PCR on both single and grouped samples with general ribosomal primers, without spacer region allowed specific phytoplasma
detection. Phytoplasmas belonging to diverse ribosomal groups were identified after RFLP analyses according to the batch
tested. Ribosomal subgroups 16SrI-B (related to ‘Candidatus phytoplasma asteris’), 16SrI-C (related to clover phyllody: CPh),
16SrII-D (related to tomato big bud from Australia: TBB), 16SrX-A (related to ‘Ca. P. mali’), and 16SrXII-A (related to stolbur)
were identified in different percentages. After further validation tests, the system can be used to screen high quality seed potatos
for phytoplasmas
Grapevine collections free from pathogens: tools and their application
No full textThe grapevine collections are very important tools to maintain grapevine biodiversity and historical germoplasm as well however in several cases especially grapevine from poor cultivated or non commercial varieties could be infected by several graft transmissible pathogens such as viruses, phytoplasmas and other systemic bacteria. In the majority of the cases these pathogens are not inducing evident symptomatology in short time after grafting therefore the possibly infected material of collection could represent a dangerous pathogen reservoir.
In order to control pathogen presence in already made collections and to prevent the spreading of the above pathogens together with the grapevine germplasm to other collections. Then, it is mandatory to exclude presence of quarantine pathogens such as “flavescece dorée” (FD) phytoplasmas and advisable to exclude relevant pathogens for quality such as viruses and phytoplasmas agent of “bois noir”, by using the most sensitive detection techniques available. It is advisable however to acquire any possible information concerning the phytosanitary status of the circulating grapevine material in order to prevent possible unforeseen outbreak of disease such as those occurred for FD disease when a grapevine insect such as Scaphoideus titanus (previously named Scaphoideus littoralis) was introduced in Europe. It is known in fact that a high number of different phytoplasmas are able to infect grapevine worldwide in the presence of appropriate insect vector or by grafting or micropropagation techniques application and crown gall is an old severely remerging disease at least in the major viticultural areas of EU and US.
First step before transferring germplasm among collection must be the verification of their sanitary status taking into account that tests to verify virus and bacteria presence should be carried out preferably during winter/spring time while those to detect phytoplasmas are more sensitive in Summer and Fall periods and the most sensitive techniques such as ELISA and PCR must be employed.
In the case of germplasm having no clean plants available after the survey it is necessary to clean the material using thermotherapy and or shoot tip culture in order to eliminate the pathogens. These techniques are not eliminating the pathogens from all the produced material therefore molecular tests are again necessary to assess the grapevine health status before the material can be employed for collection and/or field dissemination. In case of virus or phytoplasma infected grapevine germplasm of unique genetic value it must be maintained under insect proof condition while it is infected in order to avoid contamination of other germplasm in the same collection. In the same way the clean germplams should also be protected in insect proof environment in order to avoid its recontamination. It is also very important to keep the collection clean from insect that are virus (mealy bugs and scale insects) or phytoplasma vectors (leafhopper and cixiids) and also the soil must be clean from Agrobacterium tumefaciens and collection should be protected from frost or mechanical damages increasing crown gall dissemination
A project to control “flavescence dorée” outbreaks in hilly areas of the Treviso district (north-eastern Italy).
No full textRicerche sulla identificazione di virus e fitoplasmi fitopatogeni che infettano le piante.
No full textIl progetto proposto si pone come obiettivo la prosecuzione della collaborazione in atto da veri anni in maniera informale fra la prof. Assunta Bertaccini e la dr. Jana Franova in relazione alla presenza di virus e fitoplasmi in piante presenti nella repubblica Ceca. La collaborazione è iniziata nel 1993 con una stage della dr. Franova presso il laboratorio di fitoplasmologia diretto dalla prof. Bertaccini presso l’Università di Bologna. Franova in questa occasione ha appreso alcuni metodi di estrazione degli acidi nucleici da piante sintomatiche ed ha potuto dare inizio a ricerche molecolari per l’individuazione di citoplasmi in materiale vegetale individuato in repubblica Ceca. Nonostante la dr. Franova abbia dovuto assentarsi dal lavoro per maternità due volte nel periodo 1998-2003 la collaborazione è continuata ininterrottamente (vedi risultati della collaborazione). Al momento attuale lo studio di virosi e fitoplasmosi è in espansione presso IPMB-ASCR a C. Budejovice mentre presso il DiSTA di Bologna la metodologia e la ricerca sono a livelli di applicazione di routine. La collaborazione è importante in quanto permetterà alle ricercatrici ceche di diventare autonome nella ricerca e nella identificazione di questi patogeni ed al gruppo italiano di ampliare le conoscenze sulla diffusione geografica di ceppi di patogeni che possono essere seriamente pericolosi per l’agricoltura italiana e della Unione Europea in generale.
Oggetto della collaborazione sarà la risoluzione di alcuni problemi tecnici di laboratorio quali la presenza di falsi positivi o falsi negativi nelle analisi molecolari derivati dall’impiego di PCR o RT-PCR, la scelta dei reagenti (primers) più idonei alle diverse situazioni sperimentali, la identificazione di infezioni miste di citoplasmi e di fitoplasmi e virus.
La collaborazione permetterà inoltre di identificare e caratterizzare fitoplasmi e virus presenti in malattia ad eziologia ancora sconosciuta, l’individuazione di citoplasmi in piante sintomatiche o meno in cui il titolo di tali patogeni e particolarmente ridotto. Le esperienze dei due laboratori potranno essere confrontate e discusse comparativamente onde individuare le migliori modalità di ricerca nel settore
Developing a method for phytoplasma identification in cactus pear samples from California.
No full textCactus pear plants showing proliferation and stunting of cladodes in Californian cultivations were tested in order to define a molecular
methodology for reliable phytoplasma detection. After several unsuccessful trials a simple extraction method was developed
to reduce the mucilage content in nucleic acid preparations that was seriously affecting pathogen detection. Nested PCR on
16S ribosomal gene and RFLP analyses together with sequencing of obtained amplicons allow to verify the presence in symptomatic
plants of 16SrV-A and 16SrI-B phytoplasmas respectively related to ‘Candidatus Phytoplasma ulmi’ and ‘Ca. P. asteris’
TROPICSAFE: WP1: Grapevine yellows Italy
No full textThis dataset contains the underlying data of the following publication:
Y. Zambon, A. Canel, A. Bertaccini, N. Contaldo, 2018. Molecular diversity of phytoplasmas associated with grapevine yellows disease in North-Eastern Italy. Phytopathology, 108:2, 206-214. DOI: 10.1094/PHYTO-07-17-0253-R
In particular, the results obtained from a 3-year survey in different vineyards located in Treviso province (Italy) were presented in tables and figures. The tables 1-3 contain the number of grapevine and insect samples positive to phytoplasma presence and the molecular characterization of these prokaryotes on different genes. Lineages corresponding to different restriction profiles are shown, supporting the findings of a number of variants of different phytoplasmas among both, plants and insects. Table 4 contains the list of GenBank accession number of the sequences used in phylogenetic analyses in comparison with sequences obtained in the survey and listed in table 2
Phytoplasmas associated with grapevine yellows diseases: an overview
No full textPhytoplasmas are obligate intracellular bacterial parasites restricted to the phloem sieve elements of the infected plants and are transmitted by phloem-sucking insects belonging to the families Cicadellidae, Cixidae, Psyllidae, Delphacidae and Derbidae. They are associated with diseases of several hundred plant species, including some economically important crops. Grapevine yellows (GY) is a worldwide disease complex associated with genetically different phytoplasmas. GY-affected Vitis vinifera shows leaf enrollment accompanied by yellowing or reddening, rubbering of the canes and desiccated clusters. Epidemiology of the different GY diseases, undistinguishable based on symptoms observation, strictly depends on the involved phytoplasma because of the insect-vector specificity and their behavior. GY diseases are attributed to infections by at least nine distinct phytoplasmas. In Europe, “flavescence dorée” (FD) and Palatinate grapevine yellows (PGY, present only in Germany), are associated with phytoplasmas classified in the ribosomal group 16SrV, while “bois noir” (BN) is attributed to phytoplasmas classified in stolbur group (ribosomal subgroup 16SrXII-A). In Australia, Australian grapevine yellows is associated to ‘Candidatus Phytoplasma australiense’ (ribosomal subgroup 16SrXII-B), and to ‘Ca. P. aurantifolia’ (ribosomal group 16SrII). Grapevine yellows in Virginia is associated with a ‘Ca. P. asteris’-related strain (ribosomal group 16SrI-A) and X-disease group (ribosomal group 16SrIII) phytoplasmas. In Chile ‘Ca. P. fraxini’ was also associated with GY together with stolbur and 16SrI-B and 16SrI-C phytoplasmas. In Italy and in South Africa ‘Ca. P. asteris’ (16SrI-B) was associated with severe GY epidemics as well. In order to distinguish each GY from the others, an important research topic focuses on developing molecular tools for specific phytoplasmas identification. In Europe, the employment of such methods for the certain exclusion of FD and BN phytoplasmas from grapevine certified propagating material is becoming urgent. PCR-based techniques allowed development of useful tools for the identification of phytoplasmas; standard protocols include nested PCR amplification of phytoplasma 16S rDNA using universal or group specific primers and RFLP analyses in order to determine the taxonomic (ribosomal group/subgroup) affiliation. Further molecular characterization, performed by sequence analyses on genes less conserved than 16S rDNA, found additional markers useful for developing suitable analytical tests for faster and specific detection of FD and BN phytoplasmas. Up to now, innovative molecular approaches developed to this aim are: (i) Real Time PCR and reverse transcription – Real Time PCR for the detection of phytoplasmas associated with FD and BN; (ii) nanobiotransducer for detecting FD phytoplasmas; (iii) multiplex nested PCR for simultaneous identification of FD and BN phytoplasmas; (iv) Ligase Detection Reaction (LDR) DNA microarray to detect and distinguish FD and BN phytoplasmas.
Furthermore, multiple gene sequence analyses (Multi Locus Sequence Typing, MLST) on ribosomal (rplV-rpsC) and non ribosomal (secY, map, uvrB, degV, hlyC, vmp, and tuf) genes highlighted an unexpected genetic heterogeneity among both FD and BN phytoplasma populations, identifying different FD and BN phytoplasma strains that can be associated with specific ecological niches (plant hosts, insect vectors, geographic origin). MLST analyses improved the chance to associate phytoplasma-specific molecular markers with biological features, opening new perspectives for the studies of FD and BN epidemiology
Grapevine crown gall: an old, emerging disease.
No full textCrown gall is considered one of the most important and widespread bacterial diseases of grapevine (Vitis vinifera L.) throughout the world. It is known in Europe for more than 150 years and can be still of great phytopathologic significance in the vineyards and nurseries, especially in cold-climate regions. The disease is predominantly caused by tumorigenic strains of Agrobacterium vitis, more rarely by tumorigenic A. tumefaciens and A. rhizogenes. Unlike A. tumefaciens and A. rhizogenes, that are broad-host-range pathogens, A. vitis is specific to grapevine. Crown gall reduces vigor and yield of grapevines and severe disease may cause partial or complete death of infected plants. High losses occur in nurseries where different graft combinations with visible symptoms are unmarketable and must be discarded. Typical symptoms of crown gall are tissue proliferation (tumors) formed mostly on the lower areas of the trunk and on aerial canes. Tumorigenic and nontumorigenic strains of A. vitis are also able to cause specific root decay and it has been hypothesized that both types may be factors involved in the “replant disease” syndrome. Wounds mainly caused by freezing temperatures or grafting serve as a crucial entry points for the pathogen and its complex infection process. During the infection process DNA fragment from the bacterial tumor inducing (Ti) plasmid is transferred and integrated into the plant genome (interkingdom gene transfer). This leads to the overproduction of the phytohormones auxin and cytokinin, resulting in an uncontrolled proliferation of plant cells and tumor formation. A. vitis is unevenly distributed within systemically infected grapevines and able to survive in vineyard soil, particularly in the vicinity of infected plants and in their debris. Another important aspect is the ability of the pathogen to be latently present within the grapevine, providing an important means of spread over short and long distances by asymptomatic propagation material. Management of grapevine crown gall is not easy considering that no effective chemical control measures are available. However, production of A. vitis-free grapevines is an essential prerequisite for an effective prevention of the disease, and great efforts should be done in this direction. For this reason, shoot tip propagation of grapevine and thermotherapy are available as control measures. Planting of crown gall and cold-resistant cultivars and rootstocks would be a good practice when establishing new vineyards. Biological control of crown gall is another promising approach in the control of the disease and several antagonistic bacterial strains have shown a certain level of efficiency in preventing tumor formation. Indexing of grapevines for the endophytic presence of A. vitis is a very important preventive measure.
Differentiation and identification of tumorigenic strains can be rapidly assessed by PCR using primer combinations specific for bacterial Ti plasmid and chromosomal genes. However, a high level of genetic diversity among Agrobacterium strains may limit the efficiency of PCR. In our studies virC, virD, virF, pehA and 23S rRNA gene-specific primers (Bini et al., Vitis 47:181, 2008; Pulawska et al., Syst. Appl. Microbiol. 29:470, 2006; Suzaki et al., J. Gen. Plant Pathol. 70:342, 2004; Szegedi and Bottka, Vitis 41:37, 2002) were reliable in routine detection and identification of a broad range of Agrobacterium strains occurring in grapevine. However, there is necessity for development and standardization of indexing procedures including protocols of analysis and sampling methods. In the EU and many other European countries, A. vitis is not listed as a quarantine pathogen and is considered as a “quality organism” which significantly reduces the value of propagation material. Therefore, the importance of proper phytosanitary measures in grapevine nurseries and on commercial lots should be emphasized
Presence of phytoplasma infections in tomato plants in Mauritius.
No full textPhytoplasmas were detected and identified in some tomato cultivation areas in Mauritius. Symptoms most frequently observed
were abnormal shoot proliferation, stunting, reduced leaf and fruit size and shortened internodes. In field-grown tomatoes the incidence
of abnormalities rarely exceeded 10%, but under hydroponics up to 100% incidence has been recorded. Two different
phytoplasmas were identified by PCR/RFLP analyses. Field-grown tomatoes were infected with a phytoplasma belonging to ribosomal
subgroup 16SrI-C and the hydroponically-grown tomatoes were infected with a phytoplasma belonging to ribosomal group
16SrV. Further studies need to be done in order to determine their occurrence, incidence, characterization, host range and mode of
transmission, so that eventually the most effective method to control phytoplasma diseases will be determined
Detection of different types of phytoplasmas in stone fruit orchards in Northern Italy.
No full textPhytoplasmas belonging to the ribosomal subgroup 16SrIII-A (X disease and related phytoplasmas) were identified in two plants
of declining cherry during the monitoring of two experimental plum orchards. The same phytoplasmas were identified also in Philaenus
spumarius samples collected in yellow sticky traps located inside the plum orchards. ‘Candidatus Phytoplasma prunorum’
was detected in one declining cherry plant and in samples of Fieberiella florii. Among the insect species tested for phytoplasma
presence Cacopsylla pruni was only captured once, and the sample resulted negative to the molecular tests
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