Comparison of the life cycle of potato cyst nematode (<Emphasis Type="Italic">Globodera rostochiensis</Emphasis>) pathotype Ro1 on selected potato cultivars

The life cycle of Globodera rostochiensis pathotype Ro1 was studied under experimental conditions on selected potato cultivars (Korela, Albina, Vivaldi, Veronika, Vera, Monalisa, Victoria, Maranca, Désirée) in Slovakia during two growing seasons. Two peaks of second stage juveniles (J2) were found in the soil; the first peak three and four weeks after planting in the first and second year, respectively. The last J2 were found on 23 September. The number of J2 found in the second peak was much higher. First J2 associated with roots were observed 18 days, on middle early and seritonous cultivars 34 days after planting, but fourth stage juveniles (J4) were observed 40 days after planting in both cultivar groups. First adult males were found in soil 43 and 46 days after planting, respectively, and the last males two weeks later. White females filled with eggs were observed on roots 61 days after planting. The cycle from hatching of J2 in the soil to the hatching of J2 from brown cysts required 68 days in the first year and 60 days in the second year.     

The Relationship between Temperature and Development in Globodera ellingtonae

A new cyst nematode species, Globodera ellingtonae, was recently described from populations in Oregon and Idaho. This nematode has been shown to reproduce on potato. Because of this nematode’s close relationship to the potato cyst nematodes, G. rostochiensis and G. pallida, an understanding of the risk of its potential spread, including prediction of potential geographical distribution, is required. To determine the development of G. ellingtonae under different temperatures, we conducted growth chamber experiments over a range of temperatures (10.0°C to 26.5°C) and tracked length of time to various developmental stages, including adult females bearing the next generation of eggs. Both the time to peak population densities of G. ellingtonae life stages and their duration in roots generally increased with decreasing temperature. Regression of growth rate to second-stage (J2) and third-stage (J3) juveniles on temperature yielded different base temperatures: 6.3°C and 4.4°C for J2 and J3, respectively. Setting a base temperature of 6°C allowed calculation of the degree-days (DD6) over which different life stages occurred. The largest population densities of J2 were found in roots between 50 and 200 DD6. Population densities of J3 peaked between 200 and 300 DD6. Adult males were detected in soil starting at 300 to 400 DD6 and remained detectable for approximately 500 DD6. By 784 to 884 DD6, half of the eggs in adult females contained vermiform juveniles. Given the similarity in temperature ranges for successful development between G. ellingtonae and G. rostochiensis, G. ellingtonae populations likely could survive in the same geographic range in which G. rostochiensis now occurs.     

Stage-specific Monoclonal Antibodies to the Potato Cyst Nematode Globodera pallida Stone (Behrens)

Using standard hybridoma technology and hierarchical screening, monoclonal antibodies (MAbs) were obtained with specific reactivity against two developmental stages of Globodera pallida. The procedure was based on enzyme-linked immunosorbent assay (ELISA) with homogenates prepared from second-stage juveniles, young adult females, and potato roots. Hybridomas were formed by fusing myelomas with splenocytes derived from mice immunized with either infective juveniles or females of G. pallida. About 600 hybridoma lines were screened from the fusion involving the mouse immunized with juveniles. Two MAbs (LJMAbl &2) were identified with high reactivity toward second-stage juveniles but no reactivity with either potato roots or females of G. pallida. A total of 630 cell lines was screened from the corresponding fusion involving the spleen of a mouse receiving immunogens from adult female nematodes. One MAb (LFMAbl) was obtained with the required specificity against only adult female G. pallida. This work extends the application of monoclonal antibodies in nematology from valuable probes for research and species identification to recognition of developmental stages. These specific MAbs have potential value in plant breeding programs for screening for resistant lines unable to support nematode development.     

Comparison of Compatible and Incompatible Response of Potato to Meloidogyne incognita

One susceptible (D6) and two resistant (E2 and N4) clones of Solanum sparsipilum × (S. phureja × haploid of S. tuberosum) were used to study the responses of potato roots and tubers to race 1 of Meloidogyne incognita (Kofoid &White) Chitwood. The compatible response was characterized by rapid penetration of large numbers of second-stage juveniles (J2) into roots, cessation of root growth, and occasional curving of root tips. The life cycle of M. incognita in the susceptible clone was completed in 25 days at 23-28 C. The incompatible response was characterized by penetration of fewer J2 into roots, necrosis of feeding sites within 2-7 days, and lack of nematode development. There were no differences in response of tubers from resistant and susceptible clones to nematode infection. Small numbers of J2 were detected in tubers, but they did not develop.     

A new system using Solanum tuberosum for the co-cultivation of Glomus intraradices and its potential for mass producing spores of arbuscular mycorrhizal fungi

Freshly harvested potato tubers, Solanum tuberosum var ‘Pukhraj’, were inoculated for transformation with Agrobacterium rhizogenes strain Ri1600. Hairy roots were formed after 8 days of co-cultivation and the transformation efficiency was 40 %. The transformants were transferred from Murashige and Skoog medium (MS) to Modified White’s medium (MW) and finally on a hormone-free minimal medium (M). The putative transformants were confirmed using rolA and rolB gene specific primers for the polymerase chain reaction (PCR) analysis. The root inducing (Ri) T-DNA transformed potato roots were co-cultured with Glomus intraradices (CMCCROC7) to obtain arbuscular mycorrhizal root organ cultures (AM-ROC dual cultures), which were used for studying the symbiosis with Glomus intraradices and the potential for spore production in vitro. Sporulation was comparable with the existing in vitro carrot-dual culture system. Around 60,250 spores/jar could be harvested with around 38,314 extraradical spores/jar and around 21,936 intraradical spores/jar. The new method using potato is certainly promising for the mass production of mycorrhizal biofertilizers. The viability of the spores when tested on potato roots was nearly 100 % and more than half of the roots were colonized 12 weeks after inoculation.     

Enzymatic Digestion of Roots for Recovery of Root-knot Nematode Developmental Stages

Developmental stages of Meloidogyne javanica were successfully released from roots by treatment with commercially available cellulase and pectinase. The average percentage recovery of nematode developmental stages from Dolichos lablab, Elymus glaucus, and Lycopersicon esculentum were as follows: eggs = 526%, J2 = 272%, J3 = 783%, J4 = 549%, adult females = 285%, and total = 425%, expressed as percentages of the counts obtained from stained roots spread on glass plates. Root digestion was more accurate and sensitive in detecting low numbers of nematodes in roots than was the glass plate method. No simple linear, quadratic, or cubic relationship was found between the two methods that would allow a conversion factor to be developed.     

Reproduction of Meloidogyne javanica on Plant Roots Genetically Transformed by Agrobacterium rhizogenes

Reproduction of Meloidogyne javanica was compared on several Agrobacterium rhizogenes-transformed root cultures under monoxenic conditions. M. javanica reproduced on all transformed roots tested; however, more females and eggs were obtained on potato and South Australian Early Dwarf Red tomato than on bindweed, Tropic tomato, lima bean, or carrot. Roots that grew at moderate rates into the agar and produced many secondary roots supported the highest reproduction. Numbers of females produced in cultures of transformed potato roots increased with increasing nematode inoculum levels, whether inoculum was dispersed eggs or juveniles. Females appeared smaller, produced fewer eggs, and were found in coalesced galls at the higher inoculum levels. The ratio between the final and initial population decreased sharply as the juvenile inoculum increased. The second-stage juvenile was preferred to dispersed eggs or egg masses for inoculation of tissue culture systems because quantity and viability of inoculum were easily assessed. Meloidogyne javanica reared on transformed root cultures were able to complete their life cycles on new transformed root cultures or greenhouse tomato plants.     

Development of Meloidogyne chitwoodi on Wheat

Postinfection development of Meloidogyne chitwoodi from second-stage juveniles (J2) to mature females and egg deposition on ''Nugaines'' winter wheat required 105, 51, 36, and 21 days at 10, 15, 20, and 25 C. At 25 C, the J2 induced cavities and hyperplasia in the cortex and apical meristem of root tips with hypertrophy of cortical and apical meristem cell nuclei, 2 and 5 days after inoculation. Giant cells induced by late J2 were observed in the stele 10 days after inoculation. Clusters of egg-laying females were common on wheat root galls 25 days after inoculation. Juveniles penetrated wheat roots at 4 C and above, but not at 2 C, when inoculum was obtained from cultures grown at 20 C, but no penetration occurred at 4 C when inoculum was stored for 12 hours at 4 C before inoculation. In northern Utah, J2 penetrated Nugaines wheat roots in the field in mid-May, about 5 months after seedling emergence. M. chitwoodi eggs were first observed on wheat roots in mid-July when plants were in blossom. Only 40% of overwintered M. chitwoodi eggs hatched at 25 C.     

Differentiation of Two New York Isolates of Pratylenchus penetrans Based on Their Reaction on Potato

The behavior of two isolates of Pratylenchus penetrans on six potato clones was assessed to test the hypothesis that these nematode isolates from New York were different. Four potato cultivars (Superior, Russet Burbank, Butte, and Hudson) and two breeding lines (NY85 and L118-2) were inoculated with nematode isolates designated Cornell (CR) and Long Island (LI). Population increase and egression of nematodes from roots were used to distinguish resistance and susceptibility of the potato clones. Based on numbers of eggs, juveniles, and adults in their roots 30 days after inoculation, potato clones Butte, Hudson, and L118-2 were designated resistant to the CR isolate and susceptible to the LI isolate. More eggs were found in the roots of all plants inoculated with the LI isolate than with the CR isolate. The clones NY85 and L118-2 were inoculated with the CR and LI isolates in a 2 x 2 factorial experiment to assess differences in nematode egression. Egression was measured, beginning 3 days after inoculation, for 12 days. The rates of egression were similar for the four treatments and fit linear regression models, but differences were detected in numbers of egressed nematodes. More nematodes of the CR isolate than the LI isolate egressed from L118-2. Differences in egression of females was particularly significant and can be used as an alternative or supplement to reproduction tests to assess resistance in potato to P. penetrans and to distinguish variation in virulence.     

Differential Response of Thor Alfalfa to Meloidogyne chitwoodi Races and M. hapla

Second-stage juveniles (J2) of races 1 and 2 of Meloidogyne chiiwoodi and M. hapla readily penetrated roots of Thor alfalfa and Columbian tomato seedlings; however, few individuals of M. chitwoodi race 1 were able to establish feeding sites and mature on alfalfa. Histopathological studies indicate that J2 of race 1 either failed to initiate feeding sites or they caused cell enlargement without typical cell wall thickening. The protoplasm of these cells coagulated, and juveniles of race 1 did not develop beyond the swollen J2 stage. A few females of race 1 fed on small giant cells and deposited a few eggs at least 20 and 30 days later than M. chitwoodi race 2 and M. hapla, respectively. Failure of race 1 to establish feeding sites was related to egression of J2 from the roots. The M. chitwoodi race 1 J2 egression from alfalfa roots was higher than egression of race 2 and M. hapla. Egression of J2 of M. chitwoodi races 1 and 2 from tomato roots was similar and higher than that of M. hapla. Thus egression plays an important role in the host-parasite relationship of M. chitwoodi and alfalfa.     

Movement of Potato Root Diffusate Through Soil

Movement of potato root diffusate (PRD) through soil was examined by using the hatch of eggs from Globodera rostochiensis cysts as an indicator. Porous bags containing cysts were placed at increasing distances and depths from potato roots, whose growth was restricted by nylon mesh. Significantly greater hatch was observed up to 50 cm laterally away from potato roots, compared with hatch in fallow soil. Eight weeks after plant emergence, we detected a concentration gradient of PRD, as measured by egg hatch, that decreased with increasing lateral and vertical distance from the root zone. Egg hatch beyond 5 weeks after plant emergence was not attributed to PRD.     

Efficacy of Ethoprop on Meloidogyne hapla and M. chitwoodi and Enhanced Biodegradation in Soil

Responses of egg masses, free eggs, and second-stage juveniles (J2) ofMeloidogyne hapla and M. chitwoodi to ethoprop were evaluated. The results indicated that J2 were the most sensitive, followed by free eggs and egg masses. In general, M. chitwoodi was more susceptible to ethoprop than M. hapla. Ethoprop at 7.2 μg a.i./g soil protected tomato roots from upward migrating M. chitwoodi for 5 weeks. The zone of protection was extended to 10 and 20 cm below the root zone when 3.6 and 7.2 cm water were applied over 8 days. Ethoprop at 1.8, 3.6, and 7.2 μg a.i./g soil degraded faster and killed fewer M. chitwoodi J2 in potato field soil previously exposed to ethoprop than in unexposed soil or sterilized exposed soil. The enhanced biodegradation property of the exposed soil lasted 17 months after the last application of ethoprop. The limited downward movement of ethoprop in the soil, migration of M. chitwoodi J2 into the treated zone, presence of resistant life stage(s) at the time of application, and loss of efficacy due to enhanced biodegradation may have a significant effect on the performance of ethoprop.     

Coating Soybean Seed with Oxamyl for Control of Heterodera glycines

Oxamyl coated on soybean (Glycine max (L.) Merr. cv. Elgin) seeds in solutions of 20, 40, 80, and 160 mg/ml had no serious deleterious effects on seedling emergence and growth when planted in sterile soil. Seedling emergence on day 3 was less than that of the uncoated control, but by day 7 emergence was equal to, or greater than, the control. Shoot and root growth from seed coated with oxamyl in 40 and 80 mg/ml solutions was greater than that of the control. In soil infested with soybean cyst nematode, Heterodera glycines, shoot weight of soybean plants from seeds coated with oxamyl in 80 mg/ml solution was 11 and 9% greater at weeks 3 and 7, respectively, than from uncoated seeds. Numbers of juveniles (J3 and J4) and adults of H. glycines observed on the roots of plants from oxamyl-coated seeds were 83, 42, and 49% less at weeks 3, 5, and 7, respectively, than numbers on the roots of the untreated control. Numbers of J2 extracted from the roots of plants from oxamyl-coated seeds were 75% less at weeks 5 and 7 than those extracted from roots of uncoated seeds. The numbers of J2 extracted from the soil planted to oxamyl-coated seeds were 51 and 33% less at weeks 5 and 7, respectively, than from soil planted to uncoated seed.     

Population Dynamics of Meloidogyne chitwoodi on Russet Burbank Potatoes in Relation to Degree-day Accumulation

Population dynamics of Meloidogyne chitwoodi were studied for 2 years in a commercial potato field and microplots. Annual second-stage juvenile (J2) densities peaked at harvest in mid-fall, declined through the winter, and were lowest in early summer. In the field and in one microplot study, population increase displayed trimodal patterns during the 1984 and 1985 seasons. Overwintering nematodes produced egg masses on roots by 600-800 degree-days base 5 C (DD₅) after planting. Second-generation and third-generation eggs hatched by 950-1,100 DD₅ and 1,500-1,600 DD₅, respectively, and J2 densities rapidly increased in the soil. A fourth generation was observed at 2,150 DD₅ in 1985 microplot studies. Tubers were initiated by 450-500 DD₅, but J2 were not observed in the tubers until after the second generation hatched at 988-1,166 DD₅. A second period of tuber invasion was observed when third generation J2 hatched. The regional variation in M. chitwoodi damage on potato may be explained by degree-day accumulation in different potato production regions of the western United States.     

Nematode Numbers and Crop Yield in a Fenamiphos-Treated Sweet Corn-Sweet Potato-Vetch Cropping System

Nematode population densities and yield of sweet corn and sweet potato as affected by the nematicide fenamiphos, in a sweet corn-sweet potato-vetch cropping system, were determined in a 5-year test (1981-85). Sweet potato was the best host of Meloidogyne incognita of these three crops. Fenamiphos 15G (6.7 kg a.i./ha) incorporated broadcast in the top 15 cm of the soil layer before planting of each crop increased (P ≤ 0.05) yields of sweet corn in 1981 and 1982 and sweet potato number 1 grade in 1982 and 1983. Yield of sweet corn and numbers of M. incognita second-stage juveniles (J2) in the soil each month were negatively correlated from planting (r = - 0.47) to harvest (r = -0.61) in 1982. Yield of number 1 sweet potato was inversely related to numbers of J2 in the soil in July-October 1982 and July-September 1983. Yield of cracked storage roots was positively related to the numbers of J2 in the soil on one or more sampling dates in all years except 1985. Some factor(s), such as microbial degradation, resistant M. incognita development, or environment, reduced the effect of fenamiphos.     

Morphometrics,Illustration, and Histopathology of Sphaeronema rumicis on Cottonwood in Utah

The morphology of a population of Sphaeronema rumicis Kir''yanova found on cottonwood in Utah is illustrated by light and scanning electron micrographs, as well as by drawings. This is the first report of males of S. rumicis, a species also not known previously to occur in North America. S. rumicis females on cottonwood in the United States were smaller than those found by Kir''yanova on sorrel in the USSR. Females and second-stage juveniles (J2) from the United States had slightly shorter stylets than did females and J2 from the USSR. Males were vermiform and had degenerate esophagi. On secondary cottonwood roots S. rumicis induces formation of a syncytium originating from proliferated pericyclic cells. Thick outer walls, wall protuberances, absence of cell wall ingrowths, dense cytoplasm, and hypertrophied nuclei were the main characteristics of syncytia observed in S. rumicis-infected cottonwood roots.     

Measuring variation in potato roots in both field and glasshouse: the search for useful yield predictors and a simple screen for root traits

Aims

Potatoes have an inadequate rooting system for efficient acquisition of water and minerals and use disproportionate amounts of irrigation and fertilizer. This research determines whether significant variation in rooting characteristics of potato exists, which characters correlate with final yield and whether a simple screen for rooting traits could be developed.

Methods

Twenty-eight genotypes of Solanum tuberosum groups Tuberosum and Phureja were grown in the field; eight replicate blocks to final harvest, while entire root systems were excavated from four blocks. Root classes were categorised and measured. The same measurements were made on these genotypes in the glasshouse, 2 weeks post emergence.

Results

In the field, total root length varied from 40 m to 112 m per plant. Final yield was correlated negatively with basal root specific root length and weakly but positively with total root weight. Solanum tuberosum group Phureja genotypes had more numerous roots and proportionally more basal than stolon roots compared with Solanum tuberosum, group Tuberosum genotypes. There were significant correlations between glasshouse and field measurements.

Conclusions

Our data demonstrate that variability in rooting traits amongst commercially available potato genotypes exists and a robust glasshouse screen has been developed. By measuring potato roots as described in this study, it is now possible to assess rooting traits of large populations of potato genotypes.     

Control of Globodera rostochiensis in Relation to Method of Applying Nematodes

Soaking potato tuber pieces for 15 min in 8,000 μg/ml of oxamyl just before planting reduced the number of Globodera rostochiensis cysts that developed on potato roots, but this treatment was phytotoxic. Five foliar applications of 1.12 kg a.i./ha of oxamyl or carbofuran at 10-day intervals beginning when 90% of the plants had emerged suppressed increase in G. rostochiensis densities. Similar foliar applications of phenamiphos were ineffective in controlling G. rostochiensis. Soil applications (in the row at planting) of aldicarb, carbofuran, phenamiphos, ethoprop, and oxamyl at 5.6 kg a.i./ha reduced the numbers of white females that developed on potato roots, but only those treatments involving aldicarb and oxamyl suppressed G. rostochiensis population increase. Combined soil and foliar treatments did not provide any advantage over soil treatment alone, as soil applications of 5.6 kg a.i./ha alone were equal to, or better than, combined soil (3.4 kg a.i./ha) and foliar (2.2 kg a.i./ha) applications in controlling G. rostochiensis.     

Life Cycle and Mating Behavior of Belonolaimus longicaudatus in Gnotobiotic Culture

The life cycle of Belonolaimus longicaudatus was observed in vitro on excised roots of Zea mays. Roots were cultured on Gamborg''s B5 medium in petri dishes with 1.5% agar adjusted to pH 5.8 and incubated at 28 °C in darkness. Second-stage juveniles (J2) fed on the roots and started the second molt (M2) to the third-stage juveniles 2 days after inoculation (DAI). The third molt (M3) to the fourth-stage juveniles occurred 7 DAI, followed by the fourth molt (M4) to males 13 DAI or to females 14 DAI. Nematode gender differences were observed by the end of the fourth molt. The first male appeared 15 DAI and the first female 17 DAI, after which mating occurred. Males were attracted to females, and mating was observed. Mating was required for reproduction. Fertilized females began to lay eggs 19 DAI and continued egg laying without the further presence of males during a 90-day observation. All of the eggs hatched. Unfertilized females rarely laid eggs, and none of the eggs were able to hatch. Feeding took place between each molt and before egg deposition occurred. The first-stage juveniles molted in the eggs 4 days after deposition, and J2 hatched from eggs 5 days after egg deposition. The life cycle from J2 to J2 was completed in 24 days.     

Preferential Colonization of Solanum tuberosum L. Roots by the Fungus Glomus intraradices in Arable Soil of a Potato Farming Area

The symbiosis between plant roots and arbuscular mycorrhizal (AM) fungi has been shown to affect both the diversity and productivity of agricultural communities. In this study, we characterized the AM fungal communities of Solanum tuberosum L. (potato) roots and of the bulk soil in two nearby areas of northern Italy, in order to verify if land use practices had selected any particular AM fungus with specificity to potato plants. The AM fungal large-subunit (LSU) rRNA genes were subjected to nested PCR, cloning, sequencing, and phylogenetic analyses. One hundred eighty-three LSU rRNA sequences were analyzed, and eight monophyletic ribotypes, belonging to Glomus groups A and B, were identified. AM fungal communities differed between bulk soil and potato roots, as one AM fungal ribotype, corresponding to Glomus intraradices, was much more frequent in potato roots than in soils (accounting for more than 90% of sequences from potato samples and less than 10% of sequences from soil samples). A semiquantitative heminested PCR with specific primers was used to confirm and quantify the AM fungal abundance observed by cloning. Overall results concerning the biodiversity of AM fungal communities in roots and in bulk soils from the two studied areas suggested that potato roots were preferentially colonized by one AM fungal species, G. intraradices.