Piero Bianco

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Full professor of Plant Pathology at the University of Milan, after having served as a researcher at the CNR (now IPSP). He deals with virus and phytoplasma diseases of the grapevine and fruit trees. He studies the variability of pathogens in order to develop reliable diagnostic methods and to identify containment strategies based on genetic resistance to pathogens and biocontrol. He has published several articles, both in scientific and technical journals: he is the co-owner of 2 patents and promoter of a startup dealing on phytodiagnostic.

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Resistance to Viruses, Phytoplasmas and Their Vectors in the Grapevine in Europe: A Review

by Etienne Herrbach and Piero Bianco

Control of grapevine virus and phytoplasma diseases is currently based on prophylactic measures a... more Control of grapevine virus and phytoplasma diseases is currently based on prophylactic measures and cultural practices. Certification programs aim to avoid the introduction of diseased grapevines into healthy vineyards, and cultural practices aim to reduce the populations of virus vectors to limit virus spread. These approaches however are of limited effectiveness. Additionally, there is no host resistance to viruses in grapevine identified so far. Genetic engineering provides new approaches to develop pathogen-resistant grapevines. Transgenic grapevines expressing virus-derived genes that can confer resistance have been obtained in several laboratories. The viral coat protein gene has been the most commonly used to engineer resistance. More recently, new approaches based on gene silencing, specifically triggering plant defence mechanisms, have opened new ways to engineer pathogen resistance in grapevines. The possibilities, benefits and advantages, but also the risks involved with the introduction of transgenic grapevines in the field, as well as their acceptability, are discussed in this review.

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Invited Review Grapevine Yellows in Italy: Past, Present and Future

Identification of new 16SrIX subgroups, -F and -G, among 'Candidatus phytoplasma phoenicium' strains infecting almond, peach and nectarine in Lebanon

Abou-Jawdah Y, 2003, CAN J PLANT PATHOL, V25, P28; Abou-Jawdah Y., 2009, Bulletin OEPP, V39, P94,... more Abou-Jawdah Y, 2003, CAN J PLANT PATHOL, V25, P28; Abou-Jawdah Y., 2009, Bulletin OEPP, V39, P94, DOI 10.1111-j.1365-2338.2009.02223.x; Abou-Jawdah Y, 2002, PLANT DIS, V86, P477, DOI 10.1094-PDIS.2002.86.5.477; Cai H, 2008, INT J SYST EVOL MICR, V58, P1448, DOI 10.1099-ijs.0.65615-0; Choueiri E., 2001, Plant Disease, V85, P802, DOI 10.1094-PDIS.2001.85.7.802C; Davis RE, 2010, NEW DIS REPORTS, V20, P35; DENG S, 1991, PHYTOPATHOLOGY, V81, P1475, DOI 10.1094-Phyto-81-1475; Doyle J.J., 1990, FOCUS, V12, P13; Gundersen D. E., 1996, Phytopathologia Mediterranea, V35, P144; Johannesen J, 2008, ENTOMOL EXP APPL, V126, P217, DOI 10.1111-j.1570-7458.2007.00655.x; Kenyon L, 1998, PLANT PATHOL, V47, P671; Khan AJ, 2007, B INSECTOL, V60, P269; Lee IM, 2000, ANNU REV MICROBIOL, V54, P221, DOI 10.1146-annurev.micro.54.1.221; Lee IM, 1998, INT J SYST BACTERIOL, V48, P1153; Lee IM, 2007, B INSECTOL, V60, P239; LOVA MM, 2010, PETRIA, V20, P694; NEI M, 1979, P NATL ACAD SCI USA, V76, P5269, DOI 10.107...

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Candidatus Phytoplasma phoenicium related strains infecting almond, peach and nectarine in Lebanon

Genetic diversity among 'Candidatus Phytoplasma phoenicium'-related strains infecting a... more Genetic diversity among 'Candidatus Phytoplasma phoenicium'-related strains infecting almond, peach and nectarine plants in diverse geographic regions of Lebanon was investigated by virtual restriction fragment length polymorphism (RFLP) analysis of 16S rDNA nucleotide sequences. Calculation of virtual restriction similarity coefficients indicates the presence of two new subgroups, -F and -G, in group 16SrIX. Obtained results open new opportunities for in-depth studies on the distribution of 'Ca. P. phoenicium' strains in plant hosts and insect vector populations from different geographic areas of Lebanon.

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Molecular characterization of 'bois noir' phytoplasma populations from North-Eastern Italy

Genetic diversity of 'bois noir' phytoplasma populations in two vineyards of Verona pro... more Genetic diversity of 'bois noir' phytoplasma populations in two vineyards of Verona province (Veneto region, North-Eastern Italy) was investigated by the use of multilocus analysis of gene sequences amplified in the polymerase chain reaction. Based on results from restriction fragment length polymorphism profiling and nucleotide sequence alignments of 16S rRNA, tuf, hlyC, trxAtruB, cbiQ-glyA, and rplS-csdB genes, 'bois noir' phytoplasma strains were grouped in five SNP lineages. The data suggest that strain composition of BN phytoplasma populations may be modified by ecological relationships in vineyards.

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Impiego del modello EPI-peronospora per un uso sostenibile dei fungicidi

Prevalence of aster yellows (AY) and elm yellows (EY) group phytoplasmas in symptomatic grapevines in three areas of northern Italy

Vitis: Journal of Grapevine Research, 2015

Flavescence doree (FD) and others important grapevine yellows (GY) diseases are known to occur in... more Flavescence doree (FD) and others important grapevine yellows (GY) diseases are known to occur in northern Italy. In this communication we report the results of an extended survey of naturally yellows diseased grapevines growing in three provinces of northern Italy, Vicenza, Brescia and Pavia. The plants were sampled at the end of June and the beginning of August. Direct PCR assay using group-specific primers for universal phytoplasma 16S rDNA amplification or for amplification of 16S rDNA from 16SrI (aster yellows and related phytoplasmas), 16SrIII (X-disease and related phytoplasmas) and 16SrV (elm yellows and related phytoplasmas) phytoplasma groups, and RFLP analysis of amplified DNA, were conducted. Phytoplasmas belonging to group 16SrV were detected only in grapevines in the Vicenza province; whereas, phytoplasmas belonging to group 16SrI subgoup G were found in grapevines in all three provinces.

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Resistance to Viruses, Phytoplasmas and Their Vectors in the Grapevine in Europe: A Review

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Fruit Crop Phytoplasmas

Phytoplasmas: Plant Pathogenic Bacteria - I

Georgian Grapevine Cultivars: Ancient Biodiversity for Future Viticulture

Frontiers in Plant Science

Grapevine (Vitis vinifera) is one of the most widely cultivated plant species of agricultural int... more Grapevine (Vitis vinifera) is one of the most widely cultivated plant species of agricultural interest, and is extensively appreciated for its fruits and the wines made from its fruits. Considering the high socio-economic impact of the wine sector all over the world, in recent years, there has been an increase in work aiming to investigate the biodiversity of grapevine germplasm available for breeding programs. Various studies have shed light on the genetic diversity characterizing the germplasm from the cradle of V. vinifera domestication in Georgia (South Caucasus). Georgian germplasm is placed in a distinct cluster from the European one and possesses a rich diversity for many different traits, including eno-carpological and phenological traits; resistance to pathogens, such as oomycetes and phytoplasmas; resistance to abiotic stresses, such as sunburn. The aim of this review is to assess the potential of Georgian cultivars as a source of useful traits for breeding programs. The u...

Multilocus Genotyping Reveals New Molecular Markers for Differentiating Distinct Genetic Lineages among “Candidatus Phytoplasma Solani” Strains Associated with Grapevine Bois Noir

Pathogens

Grapevine Bois noir (BN) is associated with infection by “Candidatus Phytoplasma solani” (CaPsol)... more Grapevine Bois noir (BN) is associated with infection by “Candidatus Phytoplasma solani” (CaPsol). In this study, an array of CaPsol strains was identified from 142 symptomatic grapevines in vineyards of northern, central, and southern Italy and North Macedonia. Molecular typing of the CaPsol strains was carried out by analysis of genes encoding 16S rRNA and translation elongation factor EF-Tu, as well as eight other previously uncharacterized genomic fragments. Strains of tuf-type a and b were found to be differentially distributed in the examined geographic regions in correlation with the prevalence of nettle and bindweed. Two sequence variants were identified in each of the four genomic segments harboring hlyC, cbiQ-glyA, trxA-truB-rsuA, and rplS-tyrS-csdB, respectively. Fifteen CaPsol lineages were identified based on distinct combinations of sequence variations within these genetic loci. Each CaPsol lineage exhibited a unique collective restriction fragment length polymorphism ...

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Characterization of fungicide sensitivity profiles of Botrytis cinerea populations sampled in Lombardy (Northern Italy) and implications for resistance management

Pest Management Science

Unique resistance traits against downy mildew from the center of origin of grapevine (Vitis vinifera)

Scientific reports, Jan 21, 2018

The Eurasian grapevine (Vitis vinifera), an Old World species now cultivated worldwide for high-q... more The Eurasian grapevine (Vitis vinifera), an Old World species now cultivated worldwide for high-quality wine production, is extremely susceptible to the agent of downy mildew, Plasmopara viticola. The cultivation of resistant V. vinifera varieties would be a sustainable way to reduce the damage caused by the pathogen and the impact of disease management, which involves the economic, health and environmental costs of frequent fungicide application. We report the finding of unique downy mildew resistance traits in a winemaking cultivar from the domestication center of V. vinifera, and characterize the expression of a range of genes associated with the resistance mechanism. Based on comparative experimental inoculations, confocal microscopy and transcriptomics analyses, our study shows that V. vinifera cv. Mgaloblishvili, native to Georgia (South Caucasus), exhibits unique resistance traits against P. viticola. Its defense response, leading to a limitation of P. viticola growth and spo...

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First Report of Grapevine virus A and Grapevine fleck virus in the Former Yugoslav Republic of Macedonia

Plant Disease, 2014

ABSTRACT Grapevine (Vitis vinifera L.) is an economically important crop and can host several dif... more ABSTRACT Grapevine (Vitis vinifera L.) is an economically important crop and can host several different viruses, including those that have to be excluded from certified propagating material in Europe. Among these, the Vitivirus Grapevine virus A (GVA) and the Maculavirus Grapevine fleck virus (GFkV) are phloem-limited viruses that are associated with two different grapevine diseases, Kober stem grooving, belonging to the rugose wood complex, and fleck diseases, respectively. During a survey conducted in 2012 in the former Republic of Macedonia, symptomatic plants with reddening of leaves were collected for laboratory analyses. In this study, grapevine red varieties (Vranec, Francovka, and Pinot noir) from four different localities (Stip, Kavadarci, Valandovo, and Gevgelija) in Macedonia were examined. Thirty-four samples were analyzed by DAS-ELISA using commercially antibodies against Grapevine leafroll associated virus-3 (GLRaV-3). Ten selected samples were processed through DAS-ELISA and molecular assays also for the presence of GVA and GFkV. Total RNA was extracted as previously described (2) and retro-transcribed (RT) using random primers followed by PCR assay with primers GVA-MP (5′-GCCAGAGGTGTTTGAGACAAT-3′) and GVA-CPdt (5′-TTTTGTCTTCGTGTGACAACCT-3′) (1), which amplified a GVA-specific fragment of 986 bp, and with primers GFkV-U279 (5′-TGGTCCTCGGCCCAGTGAAAAAGTA-3′) and GFkV-L630 (5′-GGCCAGGTTGTAGTCGGTGTTGTC-3′) (3), which amplified a GFkV-specific region of 315 bp. Results from DAS-ELISA test showed the presence of GLRaV-3 in 21 tested samples and of GVA and GFkV in six and three out of 10 selected samples, respectively. GVA was found in Vranec and Francovka vines sampled in all the locations mentioned before, while GFkV was detected in Vranec and Pinot noir vines, in Stip, Kavadarci, and Gevgelija. These latter results were confirmed by RT-PCR assays; then, four GVA-specific and three GFkV-specific amplicons were sequenced from both directions to get a 3× coverage. For GVA fragment, a primer pair designed in the internal part of the sequence was also used. BLASTn analyses showed that (i) PCR products amplified with GVA-specific primers shared best nucleotide sequence identities, ranging from 91.7 to 93.7%, with GVA isolate at GenBank Accession No. X75433; (ii) PCR products amplified with GFkV-specific primers shared best nucleotide sequence identities, from 92.5 to 94.7%, with GFkV isolate at AJ309022. These evidences reinforced the serological and PCR results indicating that GVA and GFkV were identified in examined grapevine plants in this study. Nucleotide sequences of GVA (KF594432 to 35) and GFkV (KF594429 to 31) were submitted to GenBank. To our knowledge, this is the first report of GVA and GFkV grapevine viruses in the Former Yugoslav Republic of Macedonia.

Endophytic bacterial community of grapevine leaves influenced by sampling date and phytoplasma infection process

BMC Microbiology, 2014

Background: Endophytic bacteria benefit host plant directly or indirectly, e.g. by biocontrol of ... more Background: Endophytic bacteria benefit host plant directly or indirectly, e.g. by biocontrol of the pathogens. Up to now, their interactions with the host and with other microorganisms are poorly understood. Consequently, a crucial step for improving the knowledge of those relationships is to determine if pathogens or plant growing season influence endophytic bacterial diversity and dynamic. Results: Four healthy, four phytoplasma diseased and four recovered (symptomatic plants that spontaneously regain a healthy condition) grapevine plants were sampled monthly from June to October 2010 in a vineyard in northwestern Italy. Metagenomic DNA was extracted from sterilized leaves and the endophytic bacterial community dynamic and diversity were analyzed by taxon specific real-time PCR, Length-Heterogeneity PCR and genus-specific PCR. These analyses revealed that both sampling date and phytoplasma infection influenced the endophytic bacterial composition. Interestingly, in June, when the plants are symptomless and the pathogen is undetectable (i) the endophytic bacterial community associated with diseased grapevines was different from those in the other sampling dates, when the phytoplasmas are detectable inside samples; (ii) the microbial community associated with recovered plants differs from that living inside healthy and diseased plants. Interestingly, LH-PCR database identified bacteria previously reported as biocontrol agents in the examined grapevines. Of these, Burkholderia, Methylobacterium and Pantoea dynamic was influenced by the phytoplasma infection process and seasonality. Conclusion: Results indicated that endophytic bacterial community composition in grapevine is correlated to both phytoplasma infection and sampling date. For the first time, data underlined that, in diseased plants, the pathogen infection process can decrease the impact of seasonality on community dynamic. Moreover, based on experimental evidences, it was reasonable to hypothesize that after recovery the restructured microbial community could maintain the main structure between seasons.

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Asymmetrasca decedens (Cicadellidae, Typhlocybinae), a natural vector of 'Candidatus Phytoplasma phoenicium

ABSTRACT ‘Candidatus Phytoplasma phoenicium’ is associated with a lethal disease of almond, peach... more ABSTRACT ‘Candidatus Phytoplasma phoenicium’ is associated with a lethal disease of almond, peach and nectarine named almond witches&#39;-broom disease (AlmWB). The disease spread rapidly in Lebanon from coastal areas to elevations exceeding 1200 m, killing over 150,000 trees in a span of two decades. The mode of spread suggested the involvement of efficient vector(s) and Asymmetrasca decedens (Hemiptera, Cicadellidae) was suspected as it is the most abundant leafhopper species present in Lebanese stone fruit orchards. Living A. decedens specimens were collected from fields heavily infested by AlmWB and used in transmission trials on healthy peach almond hybrid GF-677 and peach GF-305 seedlings with an inoculation-access period of 30 days. PCR analysis supported by sequencing showed that A. decedens is a carrier of the phytoplasma, and that the phytoplasma was detected in insect salivary glands and in some inoculated GF-677 and GF-305 seedlings. One year post-inoculation, ‘Ca. P. phoenicium’ was detected in newly emergent leaves of inoculated seedlings. However, the characteristic symptoms of witches&#39;-broom were not observed. PCR amplified fragments from phytoplasma-positive seedlings and from A. decedens samples showed 99.9% nucleotide identity in their 16S RNA region and phylogenetic analysis using a neighbour jointing tree confirmed that the phytoplasmas detected in both insects and inoculated seedlings belonged to 16SrIX-B (D). The present manuscript is the first known report for a leafhopper vector of ‘Ca. P. phoenicium’ and shows that the incubation period of the disease in plants may be longer than 1 year. The importance of phytosanitary control measures, the adoption of a national strategy and regional cooperation in order to contain the further spread of the disease are discussed.

Flavescence dorée phytoplasma affecting grapevine ( Vitis vinifera ) newly reported in Portugal

Plant Pathology, 2010

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Characterization of the microflora associated to Scaphoideus titanus, the insect vector of the" flavescence doree

Bulletin of …, 2008

Several wine production areas in Europe are afflicted by "flavescence dorée" (FD), a grape yellow... more Several wine production areas in Europe are afflicted by "flavescence dorée" (FD), a grape yellows caused by 'Candidatus Phytoplasma vitis', transmitted by the phloem-feeder leafhopper Scaphoideus titanus Ball (Hemiptera Cicadellidae). Symbiotic control of disease transmission, exploiting natural symbionts of the vector, could represent an innovative strategy against FD diffusion. A study was conducted to characterize the microflora associated to S. titanus specimens, collected from FD-contaminated vineyards in the Italian region of Piedmont. By length heterogeneity PCR we identified a major peak associated with almost all the individuals examined. Characterization by denaturing gradient gel electrophoresis confirmed the presence of a major band corresponding to sex-ratio-distorting endosymbionts of the 'Candidatus Cardinium hertigii' group; electron microscopy of tissues of S. titanus showed bacterial cells with the typical morphology of 'Ca. Cardinium hertigii' together with cell morphologies resembling those of 'Ca. Phytoplasma vitis'. Our findings suggest that this endosymbiont, named ST1-C, described for the first time in Cicadellidae, is transovarially transmitted and could have a complex life cycle in the insect body; in addition it colocalizes with 'Ca. Phytoplasma vitis' several host tissues. These results make ST1-C an interesting candidate for the symbiotic control of FD diffusion.

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A Novel Bacteroidetes Symbiont Is Localized in Scaphoideus titanus, the Insect Vector of Flavescence Doree in Vitis vinifera

Applied and Environmental Microbiology, 2006

Flavescence dorée (FD) is a grapevine disease that afflicts several wine production areas in Eur... more Flavescence dorée (FD) is a grapevine disease that afflicts several wine production areas in Europe, from Portugal to Serbia. FD is caused by a bacterium, “Candidatus Phytoplasma vitis,” which is spread throughout the vineyards by a leafhopper, Scaphoideus titanus (Cicadellidae). After collection of S. titanus specimens from FD-contaminated vineyards in three different areas in the Piedmont region of Italy, we performed a survey to characterize the bacterial microflora associated with this insect. Using length heterogeneity PCR with universal primers for bacteria we identified a major peak associated with almost all of the individuals examined (both males and females). Characterization by denaturing gradient gel electrophoresis confirmed the presence of a major band that, after sequencing, showed a 97 to 99% identity with Bacteroidetes symbionts of the “Candidatus Cardinium hertigii” group. In addition, electron microscopy of tissues of S. titanus fed for 3 months on phytoplasma-in...

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Proposal of A New Bois Noir Epidemiological Pattern Related to ‘Candidatus Phytoplasma Solani’ Strains Characterized by A Possible Moderate Virulence in Tuscany

Pathogens

Bois noir (BN), associated with ‘Candidatus Phytoplasma solani’ (CaPsol), is the most widespread ... more Bois noir (BN), associated with ‘Candidatus Phytoplasma solani’ (CaPsol), is the most widespread disease of the grapevine yellows complex worldwide. In this work, BN epidemiology was investigated in a case study vineyard where an unusual CaPsol strain, previously detected only in other host plants, was found to be prevalent in grapevine. Experimental activities included: symptom observation; sampling of symptomatic vines, Auchenorrhyncha specimens, and weeds; molecular detection and typing of CaPsol strains; statistical analyses for determining possible relationships between CaPsol relative concentration, strain type, and symptom severity. Among insects, Reptalus quinquecostatus was the most abundant and was found to be highly infected by CaPsol, while Hyalesthes obsoletus, the main CaPsol vector, was not caught. Moreover, R. quinquecostatus harbored CaPsol strains carrying uniquely the stamp sequence variant St10, also identified as prevalent in vines and in the majority of weeds, ...

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Piero Bianco

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