Detection and quantification of Verticillium dahliae and Verticillium albo-atrum in potato and strawberry plants. T. BORZA, A. GOVINDARAJAN, X. GAO, R. D. PETERS, Z. GANGA, J. RAND, B. BEATON, K. BEST, K. PRUSKI AND G. WANG-PRUSKI. Department of Plant and Animal Sciences, Faculty of Agriculture, Dalhousie University, 50 Pictou Road, Truro, NS B2N 5E3, Canada; (R.D.P.) Crops and Livestock Research Centre, Agriculture and Agri-Food Canada, 440 University Avenue, Charlottetown, PE C1A 4N6, Canada; (Z.G.) Cavendish Farms, New Annan, PE C1N 4J9, Canada; (J.R.) Ivan Curry School of Engineering, Acadia University, Wolfville, NS B4P 2R2, Canada; (B.B.) PEI Department of Agriculture and Forestry, P.O. Box 2000, Charlottetown, PE C1A 7N8, Canada; and (K.B.) Prospect Agri-Services, Cambridge Station, NS B0P 1G0, Canada
Verticillium spp. infect a broad range of plants. In potatoes and strawberries, Verticillium wilt causes decreased tuber and berry yields, respectively. In Atlantic Canada, the disease is caused mainly by two species, V. dahliae Kleb. and V. albo-atrum Reinke & Berthold. Previous work carried out in this geographical area, more than a decade ago, indicated high levels of infestation of potatoes with both V. dahliae and V. albo-atrum, while no such data is available for strawberries. A real-time quantitative PCR (qPCR) method was used for the detection and quantification of V. dahliae and V. albo-atrum in potato stems collected from fields in Prince Edward Island (PEI) and Nova Scotia (NS) and in strawberry plants harvested in fields from NS. For both pathogens, the detection limit using qPCR was 1 to 2 cells/gram of fresh tissue. All potato plants tested from PEI and NS were found positive for V. dahliae (generally > 1000 cells/grams of plant tissue), but not for V. albo-atrum. V. albo-atrum was identified only in an experimental field in which plants were artificially inoculated with V. albo-atrum strain 1856. However, V. dahliae was also identified in these plants, providing additional evidence that presently this pathogen has a ubiquitous distribution in potato plants grown in PEI and NS. The screening of strawberry plants allowed the detection of a few plants infested with V. dahliae, but not with V. albo-atrum. The level of infestation of strawberry plants was found to be very low compared to that observed in potato plants.
Survey of metalaxyl-m resistance in populations of the potato pink rot pathogen (Phytophthora erythroseptica) in Canada and alternative disease management strategies. B. CRANE, R. D. PETERS, L. M. KAWCHUK, A. MACPHAIL, K. A. DRAKE, D. GREGORY AND K. MACDONALD. Crops and Livestock Research Centre, Agriculture and Agri-Food Canada (AAFC), 440 University Avenue, Charlottetown, PE C1A 4N6, Canada; and (L.M.K.) Lethbridge Research Centre, AAFC, 5403–1 Avenue South, Lethbridge, AB T1J 4B1, Canada
Pink rot, caused by the pathogen Phytophthora erythroseptica Pethybr., is a common disease of potatoes in Canada. Pink rot is most prevalent in wet growing conditions in autumn which contribute to pathogen spore release and tuber infections. Traditionally, pink rot has been managed with metalaxyl-m (Ridomil Gold ®) applications either in-furrow at planting or as a foliar spray during the growing season. In recent years, isolates of P. erythroseptica resistant to metalaxyl-m have appeared in potato growing regions of Canada, including the Maritime provinces. In 2013, a two-year national survey was initiated to assess the distribution of resistant strains of P. erythroseptica in Canada. Samples of infected tubers from across Canada were used to obtain isolates of the pathogen for testing for metalaxyl-m resistance using an in vitro agar assay. Resistant strains were recovered from Prince Edward Island, New Brunswick, Nova Scotia, Ontario, and Manitoba in 2013. This indicates an expansion in range and distribution of metalaxyl-m resistant isolates of P. erythroseptica is occurring in Canada. Therefore, alternative management strategies need to be assessed to provide protection against tuber infection by P. erythroseptica. In 2014, a field study was conducted to test the efficacy of in-furrow applications of Serenade SOIL®, Ridomil Gold®, Presidio®, Phostrol®, an experimental treatment, and foliar applications of Phostrol® to suppress tuber infections by metalaxyl-m sensitive or metalaxyl-m resistant isolates of P. erythroseptica during planting and in wet growing conditions in autumn. Ridomil Gold® provided the greatest suppression of seed piece decay by the metalaxyl-m sensitive isolate in the study, but did not suppress infection by the resistant isolate. Plots with other chemical treatments yielded seed piece decay similar to that of the inoculated control for both the metalaxyl-m sensitive and resistant P. erythroseptica isolates. All treatments suppressed tuber infection by the metalaxyl-m sensitive isolate during wet autumn growing conditions compared to the inoculated control. Foliar Phostrol®, Presidio®, and an experimental treatment provided the greatest suppression of tuber infection by the metalaxyl-m resistant isolate during wet autumn growing conditions relative to the inoculated control. In-furrow treatment of Phostrol®, Serenade® or Ridomil Gold® did not provide control of tuber infection by the metalaxyl-m resistant strain. These results suggest that there are some potential new management strategies to control pink rot infections caused by metalaxyl-m resistant strains of P. erythroseptica that continue to spread across Canada.
Characterization of the molecular mechanisms involved in the Pseudomonas fluorescens LBUM223/Streptomyces scabies interaction leading to the biocontrol of potato common scab. M. FILION, T. ARSENEAULT AND C. GOYER. Université de Moncton, Department of Biology, 18 Antonine-Maillet, Moncton, NB E1A 3E9, Canada; and (C.G.) Potato Research Centre, Agriculture and Agri-Food Canada, 850 Lincoln Road, Fredericton, NB E3B 4Z7, Canada
Common scab of potato, caused by Streptomyces scabies, is an economically important disease. There are currently no efficient measures to control common scab. We have previously determined that Pseudomonas fluorescens LBUM223 showed antagonistic properties against S. scabies under in vitro conditions, mainly due to the production of the antimicrobial compound phenazine-1-carboxilic acid (PCA). In this study, we investigated if LBUM223 was able to control common scab under growth chamber and field conditions, and also evaluated the contribution of three mechanisms in controlling common scab development: 1) antibiosis; 2) induced systemic resistance; 3) and transcriptional alteration of the pathogen’s virulence gene expression. The results obtained in growth chamber experiments and field trials were congruent and suggested that PCA production by LBUM223 is required to significantly reduce scab development on potato. Although no reduction in S. scabies population was observed in the rhizosphere or geocaulosphere in the presence of LBUM223, a significant reduction in txtA expression in S. scabies, encoding the production of thaxtomin A, a phytotoxic virulence factor, was observed in the geocaulosphere starting at 10 weeks following planting. Furthermore, LBUM223 did induce long-term overexpression of several defense-related genes in potato, but this systemic response did not seem to significantly contribute to disease control. Taken together, these results suggest that Pseudomonas fluorescens LBUM223 controls common scab development not by antibiosis or by stimulating plant defense responses, but instead reduces S. scabies thaxtomin A production in the geocaulosphere, leading to reduced virulence and symptom development.
What’s in a wart? Insights from the genome of the potato wart pathogen Synchytrium endobioticum. D. L. JOLY, J. CULLIS, C. LEWIS, D. S. SMITH, M. -C. GAGNON, M. P. E. VAN GENT-PELZER, H. VAN DE GEEST, G. J. BILODEAU, T. A. J. VAN DER LEE, X. LI, P. J. M. BONANTS AND C. A. LÉVESQUE. Département de biologie, Université de Moncton, 18 avenue Antonine-Maillet, Moncton, NB E1A 3E9, Canada; (J.C., C.L., C.A.L.) Eastern Cereal and Oilseed Research Centre, Agriculture and Agri-Food Canada (AAFC), 960 Carling Avenue, Ottawa, ON K1A 0C6, Canada; (D.S.S., X.L.) Canadian Food Inspection Agency (CFIA), 93 Mount Edward Road, Charlottetown, PE C1A 5T1, Canada; (M.-C.G., G.J.B.) CFIA, 3851 Fallowfield Road, Ottawa, ON K2H 8P9, Canada; and (M.P.E.v.G.-P., H.v.d.G, T.A.J.v.d.L, P.J.M.B.) Plant Research International, P.O. Box 16, NL-6700 AA, Wageningen, The Netherlands.
Potato wart is a serious disease of cultivated potato (Solanum tuberosum L.) caused by the obligate biotrophic soil-borne chytrid fungus Synchytrium endobioticum (Schilb.) Perc. Even though the disease has been repeatedly reported in numerous locations around the world, its global distribution has been limited by strict quarantine and regulatory measures. In 2000, the US temporarily closed its border to imports of PEI potatoes following the discovery of the fungus, which has then been found subsequently in 2002, 2003, 2007, 2012 and 2014. Genome sequencing of plant pathogens is being increasingly used to improve our understanding of virulence mechanisms, as well as assisting regulatory agencies to develop molecular markers towards more sensitive and accurate detection methods. Isolates of S. endobioticum have been independently sequenced using Next-Generation Sequencing technologies, unravelling a reasonably small genome of ~20 megabases, for which educated annotation revealed under 6,000 genes. Being an intracellular holocarpic pathogen, S. endobioticum secreted proteins are prime candidates to look for virulence factors. Preliminary analyses revealed a relatively small secretome of ~250 putative secreted proteins. Interestingly, one family of secreted proteins represented almost a fifth of the entire secretome. Within this family, high rates of amino acid changes have been identified between two isolates, which suggests diversifying selection is acting on these genes. Functional assays will be developed to screen the function of these putative secreted proteins within plant tissues.
Re-classification of potato isolates of Pectobacterium wasabiae as Pectobacterium kelmani. X. LI, K. YUAN AND S. H. DE BOER. Canadian Food Inspection Agency, Charlottetown Laboratory, 93 Mount Edward Road, Charlottetown, PE C1A 5T1, Canada
With a narrow host range, Pectobacterium wasabiae (Goto & Matsumoto) Gardan et al. (formerly Erwinia carotovora subsp. wasabiae) was originally described as the causal agent of soft rot of horseradish (Eutrema wasabi Maxim.) back in 1987 in Japan. Recently, a group of pectolytic bacteria causing potato tuber decay, aerial stem rot, and/or blackleg-like symptoms in Europe, NA and some other potato production regions were classified as P. wasabiae based mainly on phylogenetic and molecular methodologies. Further investigation on phenotypical characterization indicated that these potato-associated strains differed from the four typical horseradish P. wasabia strains. The potato strains were positive in utilizing or acidification of lactose, melibiose and raffinose, while the four horseradish P. wasabiae strains from Japan were all negative. Revisiting the phylogenetic trees in published studies indicated the presence of phylogenetical differences between these two groups of bacteria. In order to define the classification of the potato strains of the pathogen, we selected a typical isolate CFIA 1002 for decoding its genome sequences using paired-end Illumina HiSeq sequencing technology with TrueSeq V3 chemistry. Sequencing resulted in 8,682,640 reads (insert size of 300 bp) totaled 876,946,640 bp, with approximately 175X genome coverage. Comparative genomic analysis of the draft genome sequences of CFIA 1002 with WPP163 from USA, SCC3193 from Europe, and the original P. wasabiae strain CFBP3394 suggested a need for reclassification of potato isolates, including potato isolates from the US and European, as a new species; we suggest Pectobacterium kelmani. Species-specific real-time PCR assays were developed for diagnostic purposes.
Identification of pathogenic Fusarium spp. in carrots. H. LU, T. BORZA, G. WANG-PRUSKI, R. D. PETERS, J. DRISCOLL AND S. ASIEDU. Department of Plant and Animal Sciences, Faculty of Agriculture, Dalhousie University, 50 Pictou Road, Truro, NS B2N 5E3, Canada; (R.D.P.) Crops and Livestock Research Centre, Agriculture and Agri-Food Canada, 440 University Avenue, Charlottetown, PE C1A 4N6, Canada; and (J.D.) PEI Horticultural Association, Charlottetown, PE C1E 2A1, Canada
In recent years, carrot producers in Prince Edward Island (PEI) have suffered large crop losses due to the development of dry rot caused by Fusarium pathogens. Therefore, Fusarium spp. are considered major pathogens that threaten the PEI carrot industry. Two major pathogenic species, Fusarium avenaceum (Fr.) Sacc. and Fusarium oxysporum Schlecht. emend. Snyder & Hansen, are believed to be the cause of the disease. To distinguish these Fusarium taxa from other Fusarium spp. and from other fungi and to monitor the infestation level of these pathogens in soil, a highly sensitive detection method is required. To achieve this goal, real-time quantitative polymerase chain reaction (qPCR) specific assays have been developed for F. avenaceum, for F. oxysporum as well as for all species in the Fusarium genus. The specificity of theses assays was tested using strains of F. avenaceum, F. oxysporum, F. coeruleum (Lib.) Sacc. and F. solani (Mart.) Sacc. The assays were then used to characterize 68 isolates from carrots grown in different fields in PEI; a large number of these isolates were found to be F. avenaceum. The assays are sensitive enough to allow the detection and the quantification of F. avenaceum and F. oxysporum in soil and plant samples. Data generated using this method will be valuable for developing and testing management strategies to control these Fusarium spp.
Development of a PCR based detection system for five prominent viruses of strawberry. T. D. B. MACKENZIE, A. G. E. GALLAGHER AND M. SINGH. Agricultural Certification Services Inc., 1030 Lincoln Road, Fredericton, NB E3B 8B7, Canada
The cultivated strawberry (Fragaria x ananassa) is a valuable perennial crop, whose productive lifespan and berry yield is often limited by a number of viruses. We have developed a sensitive PCR detection procedure for five of these viruses: Strawberry Mild Yellow Edge (SMYEV), Mottle (SMoV), Vein Banding (SVBV), Crinkle (SCV), and Pallidosis Associated (SPaV) viruses. The five viruses can be transmitted by vegetative propagation of the plants, with varying persistence in different insect vectors, and are usually asymptomatic alone, only causing disease with combined infections, thus virus detection and epidemiology in strawberry is complex and difficult. Our procedure allows for screening large numbers of strawberry leaves harvested at any time in the season, growth stage or level of symptomology of the plant, and can also be used to detect viruses in wild strawberry plants, which can act as a natural virus reservoir. Processing typically takes two days, separated into viral nucleic acid extraction first, followed by a two-step cDNA synthesis/PCR protocol with proprietary primers targeting each virus. The full five-virus screen can be processed simultaneously as two multiplex PCRs for SMYEV, SMoV, SPaV and SVBV, SCV respectively, although simplex or other custom multiplex PCR protocols can also be performed. We routinely process composite samples of several plants mixed together to more cost-effectively screen for uncommon viruses across a large number of plants. This procedure has been commercialized by Agricultural Certification Services, Inc. to service private industry, governmental and research needs beginning in the 2014 growing season.
Updates on potato late blight research in New Brunswick during 2013–2014. K. I. AL-MUGHRABI, K. JAYASURIYA, R. POIRIER, L. M. KAWCHUK, R. D. PETERS, F. DAAYF AND B. PRITHIVIRAJ. Potato Development Centre, New Brunswick Department of Agriculture, Aquaculture & Fisheries, 39 Barker Lane, Wicklow, NB E7L3S4 Canada; (L.M.K.) Lethbridge Research Centre, Agriculture and Agri-Food Canada (AAFC), 5403–1 Avenue South, Lethbridge, AB T1J 4B1, Canada; (R.D.P.) Crops and Livestock Research Centre, AAFC, 440 University Avenue, Charlottetown, PE C1A 4N6, Canada; (F.D.) Department of Plant Science, 222 Agriculture Building, University of Manitoba, Winnipeg, MB R3T 2N2, Canada; and (B.P.) Department of Environmental Biology, Dalhousie University, Truro, NS B2N 5E3, Canada
During 2013–2014, we evaluated the susceptibility of alternative hosts (tomato, pepper, eggplant, petunia, and nightshade) to late blight caused by Phytophthora infestans (Mont.) de Bary US-23 strain. We also assessed the efficacy of fungicides registered in Canada against late blight, both in vitro and in situ. When detached leaves of alternate hosts were inoculated with P. infestans US-23 isolates, nightshade was the most susceptible (Disease Severity ‘DS’ = 9.5 mm) and the severity was comparable to that on potato (DS = 8.9 mm). Late blight severity on petunia (4.7 mm) and tomatoes (4.3–4.7 mm) was significantly (p < 0.05) less than that on nightshade. Eggplant had a DS of 2.1 mm while pepper was the least susceptible (DS = 0.03 mm). Field trials conducted in New Brunswick over 2 seasons examined the following treatments: 1) Control; 2) Confine™; 3) Phostrol™; 4) Zampro™; 5) Bravo®ZN/Dithane™DG; 6) Allegro500 F; 7) Curzate®DF; 8) Acrobat®50WP; 9) Ridomil®GoldMZ68WP; 10) Tattoo®C; 11) Gavel®75DF; 12) Ranman400SC; 13) Headline®EC; 14) Reason®500SC; 15) Revus®; and 16) Cabrio®Plus, alternated with BravoZN/DithaneDG. The effect of seaweed extract alone or alternated with other fungicides was assessed against both early and late blight diseases. Treatments included: 1) Control; 2) seaweed extract (@3.71 L/ha) alternated with Dithane™DG/Bravo®ZN; 3) seaweed extract (@3.71 L/ha, and 4) QuadrisTop™ (@1 L/ha) alternated with Dithane™DG/Bravo®ZN. All fungicides tested in the field suppressed late blight which prevailed during the latter part of the two growing seasons. Seaweed extract alone was ineffective in controlling late blight. Seaweed extract alternated with Dithane™DG and Bravo®ZN or the treatment with QuadrisTop™ alternated with Dithane™DG and Bravo®ZN significantly reduced late blight severity and incidence, as well as early blight severity. In all treatments, marketable yield did not vary significantly.
Occurrence of potato tuber damaging viruses in Canada and evidence of genome reassortment and RNA recombination in Potato mop-top virus. X. NIE, V. DICKISON, X. HU, X. XIONG AND M. SINGH. Potato Research Centre, Agriculture and Agri-Food Canada, 850 Lincoln Road, P.O. Box 20280, Fredericton, NB E3B 4Z7, Canada; (X.H., X.X.) College of Horticulture and Landscape, Hunan Agricultural University, Changsha, Hunan, China; and (M.S.) Agricultural Certification Services, 1030 Lincoln Road, Fredericton, NB E3B 8B7, Canada
In recent years, incidences of potato tubers exhibiting external and/or internal necrosis have been observed in potatoes in Canada as well as many other countries. Several viruses including Potato virus Y tuber necrosis strain (PVYNTN), Potato mop-top virus (PMTV) and Tobacco rattle virus (TRV) have been linked to the tuber necrosis. Various biological, serological and molecular approaches have been used to unveil the tuber necrosis-causing viral pathogens. The complete genome comprising three genomic RNAs of three Canadian and two Chinese isolates of potato mop-top virus were sequenced and analyzed. Two ORFs were found in RNA1 of 6.1 kb, encoding a readthrough RdRp. A CP-readthrough protein was encoded by RNA2 of 3.1 kb. Four ORFs that encoded the triple-gene-block protein (TGBp) and a cysteine-rich protein were found in RNA3 (2.9 kb) of the two Chinese isolates; whereas in the Canadian isolates, only three ORFs encoding TGBps were observed in RNA3. A single nucleotide mutation of A2462 to G2462 abolished the start codon ‘AUG’ for the fourth putative ORF in RNA3 of the Canadian isolates. Based on phylogenetic and sequence similarity analysis at the complete RNA sequence level, each of RNA1, RNA2 and RNA3 could be divided into at least two groups; and interestingly, not all the RNAs in each of the analyzed isolates exhibited the same phylogenetic relationship with that of other isolates. In Canadian isolates Ch9/Ch10/Ch20, all genomic RNAs belonged to group A; and in Chinese isolate Yunnan, all of its RNA belonged to group B; and in Swedish isolate Sw, RNA1 and RNA2 belonged to group A while RNA3 belonged to group B. In Chinese isolate Guangdong, RNA1 and RNA2 belonged to group A, and RNA3 was a hybrid of A and B, possessing a recombinant event at ~nt 1782. Taken together, this research, for the first time, demonstrates that genome reassortment and RNA recombination had taken place during PMTV evolution and diversification processes.
The changing epidemiology of tomato and potato late blight in Canada. R. D. PETERS, L. M. KAWCHUK, F. DAAYF, K. I. AL-MUGHRABI, A. MACPHAIL, D. GREGORY, K. A. DRAKE, M. TRENHOLM AND B. CRANE. Crops and Livestock Research Centre, Agriculture and Agri-Food Canada (AAFC), 440 University Avenue, Charlottetown, PE C1A 4N6, Canada; (L.M.K.) Lethbridge Research Centre, AAFC, 5403–1 Avenue South, Lethbridge, AB T1J 4B1, Canada; (F.D.) Department of Plant Science, 222 Agriculture Building, University of Manitoba, Winnipeg, MB R3T 2N2, Canada; and (K.I.A.-M.) Potato Development Centre, New Brunswick Department of Agriculture, Aquaculture & Fisheries, 39 Barker Lane, Wicklow, NB E7L 3S4, Canada
Late blight of tomato and potato, caused by Phytophthora infestans (Mont.) de Bary, has caused significant economic losses in Canada in recent years. National surveys conducted since 2010 have documented the introduction and spread of novel genotypes of the pathogen, including US-22, US-23 and US-24, which have been responsible for most documented cases of disease. Long-distance transmission of genotypes on infected tomato transplants or potato seed has occurred, in addition to the common local transport of sporangia via wind and rain. In isolated instances, evidence for the generation of novel strains via sexual recombination has also been obtained. These novel pathogen strains have shown differences in host preference as well as sensitivity to fungicides used to control late blight. For example, US-23, which now dominates most pathogen populations in Canada, is highly aggressive on tomato foliage and fruit and potato tubers, but less aggressive on potato foliage. The spectrum of sensitivity to metalaxyl-m found in populations of US-23 is shifting from sensitive to increased resistance to this chemical. Tomatoes, either as infected transplants in garden centres or in home gardens, have become significant sources of infective propagules that can initiate disease in adjacent potato crops. The changing dynamic of late blight in Canada has forced both tomato and potato industries to reconsider their disease management strategies.
Agricultural Certification Services Inc. – a private not-for-profit laboratory playing a major role in disease diagnostics and research. M. SINGH. Agricultural Certification Services Inc., 1030 Lincoln Road, Fredericton, NB E3B 8B7, Canada
Agricultural Certification Services Inc. (ACS) is a not-for-profit laboratory owned by New Brunswick potato growers and located in Fredericton, NB. ACS has been in operation since 1996 providing Canadian Food Inspection Agency (CFIA) approved testing services to potato industries in NB, PEI, NS, ON, QC and AB. ACS is equipped for microbiological, serological and molecular research work and uses novel technologies for testing, such as PCR and real-time PCR along with ELISA. Services offered by ACS include: bacterial ring rot testing; testing for potato viruses A, M, S, X, Y, potato leafroll virus, potato latent virus, potato mop-top virus, potato spindle tuber viroid, late blight, pink rot testing and virus eradication from accessions of potatoes. In addition to potato testing, ACS offers vomitoxin testing in grains, such as wheat, barley, oat and corn. Recently, ACS started new diagnostic services using RT-PCR testing for strawberry viruses, such as Strawberry mild yellow edge virus, Strawberry mottle virus, Strawberry vein banding virus, Strawberry pallidosis virus and Strawberry crinkle virus. ACS has been involved in conducting contract research in collaboration with different organizations, such as the New Brunswick Department of Agriculture, Aquaculture and Fisheries, Agriculture & Agri-Food Canada, McCain’s, Cavendish Farms, National Research Council, Canadian Horticultural Council, DuPont and the CFIA. Currently, ACS is either leading or collaborating on seven different research projects related to PVY strain characterization and management using insecticides and mineral oil.
Detection and quantification of Verticillium dahliae in soil of potato and strawberry fields and its distribution in PEI and Nova Scotia. G. WANG-PRUSKI, T. BORZA, A. GOVINDARAJAN, X. GAO, B. BEATON, K. BEST, Z. GANGA AND K. PRUSKI. Department of Plant and Animal Sciences, Faculty of Agriculture, Dalhousie University, 50 Pictou Road, Truro, NS B2N 5E3, Canada; (Z.G.) Cavendish Farms, New Annan, PE C1N 4J9, Canada; (B.B.) PEI Department of Agriculture and Forestry, Charlottetown, PE C1A 7N8, Canada; and (K.B.) Prospect Agri-Services, Cambridge Station, NS B0P 1G0, Canada
Verticillium dahliae Kleb., in combination with other fungi, is responsible for the early dying syndrome of potatoes which causes important reduction of tuber yields. When present in strawberry fields, V. dahliae can also cause severe economic losses in production. As this fungus has a wide host range and microsclerotia have a long viability, assessing the amount of the inoculum in soil is needed for disease management. The aim of this study was to detect and quantify the levels of V. dahliae propagules in soil samples cultivated with potatoes, strawberries and rotating crops. Molecular methods, such as real-time quantitative PCR (qPCR), provide a faster quantification of V. dahliae in soil samples compared to plating methods. Using a qPCR method, the distribution of V. dahliae in soil was analyzed in four strawberry fields and two potato fields in Nova Scotia, and seven potato fields or fields with rotation crops in Prince Edward Island. The results showed that the qPCR allows the detection of as little as one cell per gram of soil. Higher levels of V. dahliae propagules were found in potato fields and in fields previously cultivated with potatoes. The pathogen was also found in strawberry fields, but lower levels of the pathogen were determined, very likely due to the fumigation practices. Since fumigation is not permitted in Prince Edward Island, the pathogen will continue to represent a significant threat to potato production.
Viruses detected in Canadian potato lots – a brief summary of potato regulatory testing for the past 10 years. H. XU. Canadian Food Inspection Agency, Charlottetown Laboratory, 93 Mount Edward Road, Charlottetown, PE C1A 5T1, Canada
Over fifty viruses have been reported world-wide to infect potato naturally. Most of them have plus-sense ssRNA as their genome that is composed of one or more RNA molecules. Five species in the family of Germiniviridae have ssDNA as their genome and 3 species in the families of Rhabdovirudae and Bunyaviridae have negative-sense ssRNA as their genome. In the seed potato certification program in Canada, seed potato lots are indexed for all viruses and a viroid – Potato spindle tuber viroid. Eleven viruses have been confirmed to infect potato naturally. Four of the 11 viruses, Potato virus S (PVS), Potato virus X, Potato virus Y (PVY, the common strain) and Potato leafroll virus, have a wide distribution across the country and their incidence in seed potato lots is quite low except that of PVS. The diversity of PVY strains appears to be increasing. Potato latent virus, Tobacco rattle virus and Alfalfa mosaic virus have been detected in some potato fields in isolated areas. Potato mop-top virus (PMTV) was confirmed to have a nation-wide distribution 10 years ago. However, the incidence of PMTV is quite low and tuber necrotic symptoms induced by PMTV infection were only detected from tubers of a few susceptible potato cultivars. Potato virus M and Potato virus A have become minor potato pathogens and are hardly detected in seed potato lots for the past 10 years. All regulatory potato testing indicated that Potato aucuba mosaic virus was not present in commercial potato plots. Some viruses stated in the published literature to be present in North America may not be present in Canadian potato lots at all.
Detection and identification of Xylella fastidiosa from maple trees. K. YUAN AND X. LI. Canadian Food Inspection Agency, Charlottetown Laboratory, 93 Mount Edward Road, Charlottetown, PE C1A 5T1, Canada
Xylella fastidiosa Wells et al. causes leaf scorch in more than 100 plant species of agricultural importance. For instance, X. fastidiosa subsp. fastidiosa, the causal agent of Pierce’s Disease (PD) of grapevine, has been considered a regulated pathogen due to the economic losses caused to the grape industry. As a North American indigenous pathogen, X. fastidiosa subsp. multiplex infects many shade trees and other plant hosts, some of which are also the host plants of the PD pathogen, such as almond. Therefore, rapid diagnostics and differentiation of these two closely related pathogens are important for effective management and prevention of PD caused by X. f. fastidiosa. In this study, four samples (M1, M2, U1 and U2) were collected from maple trees showing typical symptoms of bacterial leaf scorch. Isolation of X. fastidiosa was attempted using selective media. X. fastidiosa-like colonies were obtained on isolation plates from one of the samples after more than 15 days incubation. The isolate was identified as X. fastidiosa in a species-specific PCR assay using a common primer set RST31/RST33, resulting in amplification of a 737bp DNA amplicon which was cloned for detailed sequencing analysis. Further analysis of the isolate using a PD-specific TaqMan real-time PCR assay indicated that this bacterium was not the PD pathogen X. fastidiosa subsp. fastidiosa. Based on comparison with published literature, it is likely that the causal agent of the diseased maple trees belongs to X. fastidiosa subsp. multiplex.