Healthy Potato Tubers Research Articles

First Report of Bacterial Soft Rot of Potato Caused by Pectobacterium carotovorum subsp. brasiliense in Guangdong Province, China

China is one of the world’s largest potato (Solanum tuberosum L.) producing countries, with a production of 5.68 million ha. Among them, winter-grown potato has been developed rapidly in Guangdong Province in recent years, with an area of ​​nearly 66,000 ha and an output value of about 4 billion yuan. In February 2019, soft rot symptoms were observed occurring on potato (cv. Xisen) in a commercial field (15 ha) in Kaipin, Guangdong, China, with a disease incidence of about 10%. Early symptoms included darkened and water-soaked necrotic lesions at the exterior of the infected tubers. The lesions spread quickly, and the inside of the tubers was slimy and emitted a foul smell. Bacterial exudate from water-soaked areas was also observed. Twenty diseased samples were collected, and the margins between the diseased and healthy areas were cut into pieces (5 × 5 mm), which were surface sterilized using 75% ethanol for 30 s and 2% NaOCl for 1 min, followed by three rinses with sterile water. The sterilized sections were macerated in drops of sterile water, and the extract was streaked onto nutrient agar medium and incubated for 24 h at 30°C. Single colonies were chosen after three times of subculturing (Zhang et al. 2014). Fifteen bacterial isolates were obtained, and a representative isolate KP1 was chosen for further analyses. Its 16S rDNA gene sequence (GenBank accession MK674243) amplified by primer pair 27F/1492R showed 99% identity with Pectobacterium carotovorum subsp. brasiliense (Pcb) strains (nos. CP024780 and CP020350) using BLASTn. Phylogenetic analysis (neighbor-joining method and 1,000 bootstrap values) with strains from different Pectobacterium species showed KP1 was grouped into a clade with Pcb strains, based on the multilocus sequences of gapA (GenBank accession no. MK674244), icdA (MK674245), mdh (MK674246), mtlD (MK674247), proA (MK674248), and pel genes (MK674249) that were amplified using the primer pairs provided in previous studies (Darrasse et al. 1994; Ma et al. 2007). Pathogenicity tests were conducted by injecting 100 µl of bacterial suspensions (10⁸ CFU/ml) into the potato tubers (cv. Xisen). For noninoculated controls, another 10 healthy potato tubers were injected with 100 µl of sterile water. Tubers were placed in an incubator at 30°C in darkness at 95% humidity. After 3 days, inoculated tubers showed diseased symptoms similar to those observed in the field. No symptom was observed on the controls. We reisolated the pathogen from the symptomatic tissues and confirmed it to be Pcb using PCR with 16S rDNA primers. This pathogen was previously reported to infect potato in Inner Mongolia Autonomous Region, Northern China (Zhao et al. 2018). To our knowledge, this is the first report of Pcb causing bacterial soft rot on potato in Guangdong Province, Southern China. This study expanded the geographic location of Pcb as the causal agent on potato in China and helped raise attention on controlling the spread of this pathogen to minimize the losses.

Biological control of potato common scab by Bacillus amyloliquefaciens Ba01.

Potato common scab, which is caused by soil-borne Streptomyces species, is a severe plant disease that results in a significant reduction in the economic value of potatoes worldwide. Due to the lack of efficacious pesticides, crop rotations, and resistant potato cultivars against the disease, we investigated whether biological control can serve as an alternative approach. In this study, multiple Bacillus species were isolated from healthy potato tubers, and Bacillus amyloliquefaciens Ba01 was chosen for further analyses based on its potency against the potato common scab pathogen Streptomyces scabies. Ba01 inhibited the growth and sporulation of S. scabies and secreted secondary metabolites such as surfactin, iturin A, and fengycin with potential activity against S. scabies as determined by imaging mass spectrometry. In pot assays, the disease severity of potato common scab decreased from 55.6 ± 11.1% (inoculated with S. scabies only) to 4.2 ± 1.4% (inoculated with S. scabies and Ba01). In the field trial, the disease severity of potato common scab was reduced from 14.4 ± 2.9% (naturally occurring) to 5.6 ± 1.1% after Ba01 treatment, representing evidence that Bacillus species control potato common scab in nature.

Soft rot of potatoes caused by Bacillus amyloliquefaciens in Guangdong province, China

Potato tubers exhibiting symptoms of soft rot were observed in a field in Baiyun District, Guangzhou, Guangdong Province, China, in March 2016. Bacteria were isolated from the diseased potato tubers. Using morphological and molecular (16S rDNA and gyrB) characterization and pathogenicity tests, a representative isolate of the bacterium was identified as Bacillus amyloliquefaciens. Inoculating the isolate onto healthy potato tubers led to soft rot with the same symptoms as displayed in the field, which confirmed Bacillus amyloliquefaciens as the causal agent. Further testing of the bacterial isolate on other hosts showed that it could also cause soft rot on tomato, onion and pepper. To our knowledge, this is the first report of soft rot of potato tubers caused by Bacillus amyloliquefaciens in the field.

Genetic Diversity of Pathogenic Fusarium semitectum Isolates from Potato Tubers, Using PCR-IGS-RFLP Markers

Fusarium semitectum is one of the important causal agents of dry rot of potato tubers in the world. In order to determine genetic variability among 41 isolates of F. semitectum, morphological and molecular studies were carried out. All F. semitectum isolates were recovered from infected potato tubers with dry rot symptoms collected from four provinces in Iran. According to macroscopic and microscopic characteristics, 41 isolates of F. semitectum were classified in two morphotypes (morphotypes I and II). All 41 isolates were evaluated for their pathogenicity on healthy potato tubers. Tuber rot symptoms were observed on the 21st day after inoculation of Fusarium isolates on the tubers tested. The measurement was done by comparing the depth and width of lesion expansion among the isolates. Molecular characterization through PCR-IGS-RFLP analysis by six restriction enzymes (AluI, BsuRI, Eco88I, MspI, TaqI and PstI) divided the 41 isolates of F. semitectum into two separated clusters that were in accordance with the morphological characterization.

Attraction Behaviors of Entomopathogenic Nematodes (Steinernematidae and Heterorhabditidae) to Synthetic Volatiles Emitted by Insect Damaged Potato Tubers.

Entomopathogenic nematodes (EPNs) play a role in indirect defense of plants under attack by root herbivores. Several investigations have shown that EPNs are attracted or repelled by various volatile compounds (VOCs) released from insect damaged plant roots. We hypothesized that the directional responses of EPNs to the VOCs would be affected by foraging strategy and would vary among species, VOC type, and VOC concentrations. We tested the chemotactic responses of four commercial EPN species (Steinernema feltiae, S. carpocapsae, S. kraussei, and Heterorhabditis bacteriophora) to seven compounds released from insect (Melolontha hippocastani)-damaged (decanal, nonanal, octanal, undecane, 6-methyl-5-hepten-2-one, and 1,2,4-trimethylbenzene) and undamaged (2-ethyl-1-hexanol) potato tubers. Our results suggest that EPNs are able to distinguish herbivore-induced VOCs from those that are typical for healthy potato tubers. In our investigation, nonanal, octanal, and decanal had a greater influence on the movement of EPNs than other tested synthetic volatiles. Decanal was an attractant for H. bacteriophora and S. kraussei at both tested concentrations (as a pure compound and at a concentration of 0.03ppm). The results suggest that the susceptibility to perception of chemical stimuli from the environment is a species-specific characteristic that prevails over the influence of the foraging strategy.

Effect of varieties and tuber sizes on losses of ascorbic acid and phenol content of potato tubers stored under ambient conditions

Effect of Varieties and Tuber Sizes on Losses of Ascorbic Acid and Phenol Content of Potato Tubers Stored Under Ambient Conditions The Investigation was conducted by keeping healthy potato tubers of three different sizes for each genotype in hessian cloth bag at room temperature in four replications. The experiment was carried out to evaluate the effect of varieties and tuber sizes on the ascorbic acid and phenols during the storage period of three months. From the findings, it was concluded that the ascorbic acid content and phenolic content decreased with increasing storage period. The maximum ascorbic acid and phenols were noticed in Kufri Bahar and minimum in Kufri Pushkar. Among the sizes, ascorbic acid had positive, while the phenol content had negative correlation with tuber size. ..

Влияние заражения клубней кольцевой гнилью и обработки монойодацетатом натрия совместно с прогреванием на хранение картофеля

В статье обсуждается влияние на проращивание клубней монойодацетата натрия (МИА) совместно с прогреванием (45 °C, 1 ч) в качестве потенциального агента для борьбы с Clavibacter michiganensis ssp. sepedonicus (Cms) перед закладкой картофеля на хранение. В работе использованы клубни картофеля (Solanum tuberosum L.) сорта Лукьяновский. В эксперименте присутствовало четыре варианта обработки клубней, в каждом варианте было 30 клубней. После проведенных обработок клубни помещали в условия овощехранилища на 8 месяцев. Весной подсчитывали количество ростков, измеряли длину проростков у клубней и проводили клубневой анализ. Полученные результаты статистически обрабатывали с использованием пакета программ Excel. Показано, что заражение клубней Cms снижало способность клубней к прорастанию. Обработка МИА и прогревание здоровых клубней картофеля перед закладкой на хранение подавляло прорастание клубней, что полезно для продовольственного картофеля. Результаты клубневого анализа показали, что прогревание снижало общее количество больных клубней примерно на 39 %, клубни были менее поражены кольцевой и бурой гнилями. Обработка МИА также способствовала снижению количества клубней, пораженных бурой и кольцевой гнилями. Обработка клубней МИА совместно с прогреванием снижала общее количество больных клубней, пораженных проволочником и пустотелых, кольцевой и сухой гнилями, а также паршой. Обработка МИА и прогреванием инфицированных клубней картофеля перед закладкой на хранение снимала подавляющий эффект бактерий на прорастание. Полученный результат актуален для обработки семенного картофеля, так как кольцевая гниль является латентным заболеванием.

Potato virus Y: Contact Transmission, Stability, Inactivation, and Infection Sources.

In glasshouse experiments, two isolates of Potato virus Y 'O' strain (PVYO) were transmitted from infected to healthy potato plants by direct contact when leaves were rubbed against each other, when cut surfaces of infected tubers were rubbed onto leaves, and to a limited extent, when blades contaminated with infective sap were used to cut healthy potato tubers. However, no tuber-to-tuber transmission occurred when blades were used to cut healthy tubers after cutting infected tubers. When leaf sap from potato plants infected with two PVYO isolates was kept at room temperature, it was highly infective for 6 to 7 h and remained infectious for up to 28 h. Also, when sap from infected leaves with one isolate was applied to five surfaces (cotton, hessian, metal, rubber vehicle tire, and wood) and left to dry for up to 24 h before each surface was rubbed onto healthy tobacco plants, PVYO remained infective for 24 h on tire and metal, 6 h on cotton and hessian, and 3 h on wood. The effectiveness of disinfectants at inactivating this isolate was evaluated by adding them to sap from infected leaves which was then rubbed onto healthy tobacco plants. None of the plants became infected when bleach (42 g/liter sodium hypochlorite, diluted 1:4) or Virkon-S (potassium peroxymonosulfate 50% wt/wt, diluted to 1%) was used. A trace of infection remained after using nonfat milk powder (20% wt/vol). PVY infection sources were studied in 2011-2012 in the main potato growing regions of southwest Australia. In tests on >17,000 potato leaf samples, PVY was detected at low levels in seed (4/155) and ware (6/51) crops. It was also detected in volunteer potatoes from a site with a previous history of PVY infection in a seed crop. None of the 15 weed species tested were PVY infected. Plants of Solanum nigrum were symptomlessly infected with PVYO after sap inoculation, and no seed transmission was detected (>2,500 seeds). This study demonstrates PVYO can be transmitted by contact and highlights the need to include removal of volunteer potatoes and other on-farm hygiene practices (decontaminating tools, machinery, clothing, etc.) in integrated disease management strategies for PVY in potato crops.

Molecular phylogenetic and pathogenetic characterization of Fusarium solani species complex (FSSC), the cause of dry rot on potato in Iran

Members of Fusarium solani species complex (FSSC) are common pathogens of potato, causing dry rot in the west of Iran which involved Hamedan, Kermanshah, Eilam and Kurdistan provinces. Therefore, the objectives in this study were to isolate and identify disease-causing FSSC from infected potato tubers based on the morphological and molecular characteristics. Forty-five isolates of Fusarium were obtained from potato tubers collected from the wet market in different regions of the west of Iran and identified as FSSC through morphological characters. All of the isolates were evaluated for their pathogenicity on healthy potato tubers in the planthouse. The tubers rot symptoms were observed on the 21st day after inoculation of Fusarium isolates on the tubers tested. In the tubers inoculation tests, lesion sizes were quite variable; therefore, the measurement was done to compare the depth and width of lesion expansion among the isolates. Based on the sequence data from translation elongation factor (EF-lα) gene and internal transcript spacer (ITS) regions analysis, all of the selected FSSC isolates were divided into two major groups. This is the first report on molecular identification of FSSC strains isolated from potato tubers in Iran and Fusarium falciforme was reported for the first time in Iran.

Scanning electron microscopy and in vitro cultivation of endophytic bacteria from potato tubers afflicted with zebra chip disease

Zebra chip disease (ZCD) of potato (Solanum tuberosum L.) has caused significant economic losses to the potato industry in the USA. ‘Candidatus Liberibacter solanacearum’ Liefting et al. has been associated with ZCD. This non-culturable bacterium dwells within the phloem of plants where other endophytic bacteria may also live. Knowledge concerning phloem colonization by ‘Ca. L. solanacearum’ and other endophytic bacteria may provide a better understanding about disease biology and could facilitate the development of novel disease management strategies. In this study, endophytic bacteria in potato tubers afflicted with ZCD were evaluated by scanning electron microscopy (SEM), in vitro cultivation, and PCR analyses. Bacillus, coccus, and filamentous bacterial cells were observed by SEM in phloem tissue of ZCD-diseased tubers, but were absent in healthy potato tubers. Fifty-one bacterial isolates were obtained in vitro from ZCD-affected tubers and 34 isolates were from non-ZCD tubers. Comparison of 16S rDNA sequences identified bacteria belonging to 16 genera. Seven (Brachybacterium, Enterobacter, Microbacterium, Paenibacillus, Staphylococcus, Stenotrophomonas and Variovorax) were only isolated from ZCD-affected potato tubers, whereas four (Bosea, Nocardia, Sphingomonas and Sphingopyxis) were only isolated from non-ZCD potato tubers. However, a more sensitive PCR analysis diminished the specific association of the bacteria to either ZCD or non-ZCD tubers, suggesting that infection by ‘Ca. L. solanacearum’ influenced the titres of other endophytic bacteria in potato tubers.

Occurrence and Pathogenicity of Fusarium spp. on the potato tubers in Malaysia

Dry rot of potato caused by Fusarium species is a common and commercially important disease of potato tubers found in all production areas of the world, including Malaysia. The objective of this study was to identify Fusarium species associated with wet market potatoes and their pathogenicity on potato tubers in Malaysia. In this survey, 65 Fusarium strains were isolated and identified from diseases decayed potato's segments collected from different regions in Malaysia. All of these 65 isolates, which belong to two morphotypes of Fusarium solani species complex (FSSC) and Fusarium oxysporum, were evaluated to study their pathogenicity on healthy potato tubers. The tubers rot symptoms were observed on the 21st day after inoculation of F. solani and F. oxysporum strains on the tubers tested. In the tubers inoculation tests, lesion sizes were quite variable; therefore, the measurement was done to compare the depth and width of lesion expansion among the strains. The results of the pathogenicity test revealed that F. solani isolates FSO4, FSO12 (morphotype I), FSO18 (morphotype II) and F. oxysporum isolates FOX4 and FOX16 were strongly pathogenic to inoculated potato tubers. The inoculated fungi were re-isolated from the diseased potato tubers to prove the Koch’s postulates. The present study showed that two morphotypes of F. solaniwere associated with tuber rot of potato in Malaysia. This is the first comprehensive report on identity and distribution of major pathogenic fungi causing potato dry rot in Malaysia. Key words: Occurrence, pathogenicity, dry rot, potato, Malaysia.

Cost Considerations in Choosing Group Size for Group Testing in the Seed Potato Certification Program

Viral diseases that can reduce yield and tuber quality are major concerns for potato growers. To produce healthy potato tubers and prevent yield reduction, a certification scheme for foundation class seed potato propagation has been established at the Seed Improvement and Propagation Station in Taiwan. The certification scheme uses an ELISA technique to index specific virus diseases such as Potato virus Y (PVY), Potato leafroll virus (PLRV), Potato virus X (PVX), Potato virus A (PVA), and Potato virus S (PVS). In order to produce foundation class potato tubers, the Dorfman group testing procedure is adopted. The Dorfman testing procedure utilizes a two-stage sampling scheme. The first stage comprises making measurements on all groups. The second stage involves no retesting for negative testing groups and exhaustive testing of all constituent individual samples of positive testing groups. In the seed potato certification scheme in Taiwan, field investigations have usually used groups of 20 plants. However, because of the limitations of available financial resources, the effect of group size on cost is an important consideration. Thus, selection of group size for design considerations is a critical issue. In order to solve the problem, we present the expected cost model as a nonlinear integer programming problem and determine the optimal group size such that the expected cost is minimized. The result demonstrates that the optimal sample size for group testing is 13 plants per group and that the cost, using this optimal group size, is not greatly different from the cost when using 20 plants per group. Thus, 20 plants per group is reasonable for the group testing procedure with regard to cost considerations. The result of this research could be useful in the execution of the seed potato certification program.

First Report of 'Candidatus Liberibacter psyllaurous' in Zebra Chip Symptomatic Potatoes from California.

A disease that severely affects processing potatoes (Solanum tuberosum L.), termed zebra chip (ZC), has been identified in several locations in the United States (Texas, Nebraska, Colorado, Kansas, New Mexico, Arizona, and Nevada), Mexico, and Central America (4). The disease name comes from the rapid oxidative darkening of freshly cut tubers and the dark stripes and blotches that occur in chips processed from infected tubers. Recently, the disorder has been associated with a new 'Candidatus Liberibacter' species in New Zealand (3). Also, a bacterium designated 'Candidatus Liberibacter psyllaurous' has been identified recently in potato plants with "psyllid yellows" symptoms that resemble foliar symptoms of ZC (2). In the fall of 2008, 10 tubers were received at the Prosser laboratory from a commercial potato grower and five had symptoms characteristic of ZC. The tubers, cv. Dakota Pearl, were grown near Lancaster in southern California. The tubers showed rapid oxidation upon slicing and the sunken stolon attachment characteristic of ZC (4). Nucleic acid was extracted from symptomatic tubers (1) and tested by PCR for 'Ca. Liberibacter' species with primer pairs OA2/OI2c (5'-GCGCTTATTTTTAATAGGAGCGGCA-3' and 5'-GCCTCGCGACTTCGCAACCCAT-3') and CL514F/R (5'-CTCTAAGATTTCGGTTGGTT-3' and 5'-TATATCTATCGTTGCACCAG-3'), which amplify from the 16S rDNA and rplJ and rplL ribosomal protein genes, respectively (3). Four of the five tubers with distinct ZC symptoms yielded the expected amplicons with both primer pairs. Two tubers with mild internal discoloration yielded correctly sized amplicons but in lesser amounts than from the severely affected tubers. Nucleic acid from healthy potato tubers yielded no product with these primers. One clone of the 1,168-bp OA2/OI2c amplicon and two clones of the 669-bp CL514F/R amplicon from a strongly positive sample were sequenced in both directions (ACGT, Inc., Wheeling, IL). BLAST alignments of the consensus sequences of the OA2/OI2c and CL514F/R amplicons (GenBank Accessions Nos. FJ498802 and FJ498803, respectively) revealed 100% identity with analogous 'Ca. Liberibacter' sequences reported from ZC-symptomatic potatoes in New Zealand (GenBank Accession Nos. EU849020 and EU919514). The OA2/OI2c amplicon was also identical to a sequence of 'Ca. Liberibacter psyllaurous' (GenBank Accession No. EU812559) from psyllid yellows-affected potatoes in the United States (2) and also showed a 99% identity with sequences from a 'Ca. Liberibacter' species reported in ZC tubers from Kansas (GenBank Accession No. EU921626). Potato crops with symptoms of ZC have been observed previously in California (4), but this is the first identification of 'Ca. Liberibacter psyllaurous' from diseased potatoes grown in California. Since ZC was first reported in the mid- to late-1990s, information from potato growers and processors suggests that ZC is becoming more important. The disease has caused millions of dollars in losses, particularly in south Texas (4). The identification of 'Ca. Liberibacter psyllaurous' in California provides additional evidence that the disease is increasing in importance in other potato-growing regions.

Interceptação de Phoma exigua var. foveata, praga exótica e quarentenária para o Brasil, em germoplasma de batata procedente da França

No Laboratório de Quarentena Vegetal da Embrapa Recursos Genéticos e Biotecnologia, foram realizadas análises fitossanitárias em tubérculos de batata, introduzidos no país para pesquisa, procedentes da França. Alguns desses tubérculos, em exame direto sob microscópio estereoscópio, apresentaram lesões deprimidas de aspecto encharcado, coloração marrom-arroxeada, casca amolecida e picnídios negros sobre as lesões. Em lâminas, sob microscópio de luz, foram observadas características morfológicas do fungo Phoma exigua var. foveata, praga exótica e de quarentena A1 para o Brasil. Para o teste de patogenicidade foram utilizados tubérculos sadios de batata inoculados com cultura pura do fungo isolado dos tubérculos sintomáticos. Sintomas de apodrecimento foram observados após 3 dias da inoculação, evoluindo até o 8º dia, quando o fungo foi re-isolado dessas lesões, em BDA, e confirmado o agente etiológico da doença. Os tubérculos apresentando sintomas foram incinerados e aqueles aparentemente sadios foram tratados com fungicida sistêmico, plantados em quarentenário por duas gerações e re-analisados. Após confirmação da sanidade dos tubérculos, os mesmos foram enviados ao requisitante.

Detection of volatiles released by diseased potato tubers using a biosensor on the basis of intact insect antennae

Comparison of the volatiles emitted by healthy and diseased potato tubers showed that there are volatile patterns specific for infestation by Phytophthora infestans. Some of these marker volatiles can be detected by a biosensor on the basis of intact antennae of the Colorado potato beetle ( Leptinotarsa decemlineata). This biosensor shows responses to diseased potato tubers selectively and is not disturbed by odours emitted by healthy and mechanically damaged potato tubers. One single diseased potato tuber can be detected within up to 100 kg potato tubers. So, this biosensor on the basis of intact insect antennae presents a solution to the problem of damageless screening of large numbers of seed potatoes for fungal infestation.

Effect of Mastoparan on Phospholipase A2 Activity in Potato Tubers Treated with Fungal Elicitor

Participation of phospholipase (PL) A2 in signal transduction has been reported to elicitate a resistance reaction in potato cells by inoculation of an incompatible race of Phytophthora infestans, the late blight fungus, or by treatment with fungal elicitor hyphal wall components (HWC). Mastoparan, a generic G protein activator, has been shown to activate PLA in a G protein-dependent manner in animal cells. We analyzed the effects of mastoparan and the inactive analog Mas-17 on PLA2 activity in potato tubers. In healthy potato tubers, the activation of PLA2 by mastoparan was detected in the soluble fraction, but not microsomal fraction. However, in potato tubers treated with HWC, PLA2 activity was stimulated by mastoparan in both soluble and microsomal fractions. Pretreatment of the microsomal fraction with neomycin, a PLC inhibitor, and staurosporine, a protein kinase inhibitor, inhibited the mastoparan-induced activation of PLA2. This suggested that the PLA2 activation in potato tubers by mastoparan was mediated by the PLC pathway and protein phosphorylation. We also examined the accumulation of potato phytoalexin rishitin. Mastoparan stimulated rishitin accumulation induced by HWC, but did not induce the accumulation. This indicated that mastoparan might activate the signal transduction pathway in the resistance reactions induced in potato tubers.

Sensitivity of different methods for the detection of Ralstonia solanacearum in potato tuber extracts

The sensitivities of various methods for the detection of Ralstonia solanacearum following dilution in healthy potato tuber tissue macerate were compared. Estimated pathogen populations in undiluted macerates, from samples of 200 heel‐end vascular cores each containing a single diseased and 199 healthy tubers, ranged from 1.2 × 106–7.4 × 107 colony‐forming units per ml. Following concentration by high‐speed centrifugation and resuspension in phosphate buffer, the pathogen was detected by all methods studied, including culture on semi‐selective media, enzyme‐linked immunosorbent assay (ELISA), indirect immunofluorescent‐antibody staining (IFAS) of fixed cells, immunofluorescent colony staining (IFCS), detection of specific DNA sequences following amplification by the polymerase chain reaction (PCR) and bioassay in tomato seedlings. Both ELISA and PCR methods were improved by pre‐enrichment of samples in semi‐selective broth prior to testing. A nested PCR method was evaluated which could detect fewer than 10 cells per ml in the potato extracts. Of the other methods only dilution plating on semi‐selective medium and tomato bioassay could detect fewer than 104 cells per ml. In order to combine ease and speed of use with sensitive detection, it was recommended that a series of methods be used for routine screening of potato tuber stocks for infection by R. solanacearum.

The role of endophytic bacteria during seed piece decay and potato tuberization

Healthy potato tubers (Solanum tuberosum L.) cv. Kennebec were found to be internally colonized by non-pathogenic bacterial populations originating from root zone soil. These endophytic bacteria were categorized, on the basis of bioassays, as plant growth promoting (PGP), plant growth retarding (PGR) and plant growth neutral (PGN). Genera isolated from tubers included Pseudomonas, Bacillus, Xanthomonas, Agrobacterium, Actinomyces and Acinetobacter. The PGP and PGR isolates were similarly distributed throughout these genera. Bacterial populations increased in the root zone soil directly adjacent to the seed piece during and immediately following seed piece decay. Bacteria sampled at this time were capable of promoting tuber number and weight. The proportions of PGP, PGR and PGN bacteria in the root zone were altered as endophytic bacteria were released from the decaying seed piece. The study indicates that endophytic bacteria present in the seed tubers may play an important role in seed piece decay, tuberization and plant growth.

Transmission of potato leafroll virus to or from tuber slices by aphids feeding through a membrane

Eye-bearing slices, cut from healthy potato tubers and placed between Parafilm membranes, were inoculated with potato leafroll virus (PLRV) byMyzus persicae. PLRV was detected by ELISA and by transmission tests in tuber slices and in plants grown from the slices of the susceptible cv. Desiree, but not in those of the resistant cv. Arkula. These results suggest that PLRV replication and transport within tuber phloem is controlled by specific mechanisms of resistance.M. persicae was also able to acquire and transmit PLRV toPl floridana from slices cut from tubers of infected plants. The aphids effectively transmitted PLRV from slices cut from the sprouting rose end but they failed to transmit it from slices cut from the heel end of tubers.

Time‐resolved fluoroimmunoassay of potato virus M with monoclonal antibodies

SummaryMouse monoclonal antibodies (MAbs) specific for potato virus M (PVM) were prepared and the properties of three of them were studied. MAb M4C1 is IgG2b, it binds with high affinity to PVM coat protein, to purified virus preparations and recognises PVM in infected potato leaves and tubers. MAb M6D5 is IgG2a and also reacts with PVM coat protein, purified PVM and with PVM in potato leaf and tuber extracts. In double‐antibody sandwich ELISA (DAS ELISA) MAbs M4C1 and M6D5 reacted with all 17 PVM isolates tested. MAb M7 is IgG2b and recognises PVM only in indirect dot ELISA on nitrocellulose filters and viral coat protein on Western blots.MAbs against PVM were used as capture antibodies and europium‐labelled MAbs as conjugates in time‐resolved fluoroimmunoassay (EuTRFIA). The standard EuTRFIA curve of PVM detection is approximately linear over a range of PVM concentrations from 0.5 ng/ml to 1000 ng/ml. The lowest PVM concentration detectable in EuTRFIA was 0.5 ng/ml and correspondingly 6 ng/ml in DAS ELISA. The use of the europium chelate label allows PVM detection in potato leaf and tuber sap at dilutions greater than 10‐‐4 with very low background fluorescence. EuTRFIA with MAbs, with either one or two incubations is about 10–20 times more sensitive for PVM detection than is DAS ELISA. PVM and PVX, mixed with healthy potato tuber sap, were simultaneously tested in a single sample at concentrations lower than 10 ng/ml by double‐label TRFIA using europium‐labelled MAbs to PVM and samarium‐labelled MAbs to PVX.