Decline of Globodera rostochiensis as Influenced by Potato Root Diffusate Movement and Persistence in Soil

Decline of Globodera rostochiensis populations occurring naturally in soil and those added to potato hills and furrow centers in nylon bags was correlated with root weight of Hudson, Rosa, and Katahdin potatoes at two locations in New York. Cysts in bags were added to soil at planting and at 1, 3, 5, 7, and 9 weeks after emergence (AE). Fallow decline required only 2-4 weeks in soil and did not increase with time. Decline due to growing potatoes was greater in hills than in furrow centers, and resistant Hudson potatoes stimulated greater nematode hatch for longer times in both hills and furrows than did resistant Rosa and susceptible Katahdin. Potato root diffusate (PRD) was produced in highest concentration early in the season; decreased egg hatch with time was probably the result of declining PRD production and inactivation of PRD in soil. Decreasing potato row spacing from 92 cm to 46 and 23 cm between rows increased G. rostochiensis decline in furrow centers, with the majority of decline occurring within 1-3 weeks AE. Replanting potatoes after 1 week of trap crop growth failed to favor population reduction over a single full season crop.     

Effects of Initial Nematode Density on Population Dynamics of Globodera rostochiensis on Resistant and Susceptible Potatoes

The influence of resistant and susceptible potato cultivars on Globodera rostochiensis population density changes was studied at different nematode inoculum levels (Pi) in the greenhouse and field. Soil in which one susceptible and two resistant cultivars were grown and fallow soil in pots was infested with cysts to result in densities of 0.04-75 eggs/cm³ soil. A resistant cultivar was grown in an infested field with Pi of 0.7-16.7 eggs/cm³ soil. Pi was positively correlated with decline of soil population densities due to hatch where resistant potatoes were grown in the greenhouse and in the field but not in fallow soil. However, Pi was not correlated with in vitro hatch of G. rostochiensis cysts in water or potato root diffusate. Under continuous culture o f a resistant cultivar, viable eggs per cyst declined 60-90% per plant growth cycle (4 weeks) and the number of cysts containing viable eggs had decreased by 77% after five cycles. The rate of G. rostochiensis reproduction on both resistant and susceptible cultivars was negatively correlated with Pi. These data were used to predict the effect of resistant and susceptible potato cultivars on G. rostochiensis soil population dynamics.     

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.     

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.     

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 of Steinernema scapterisci Nguyen &Smart, 1990

The life cycle of Steinernema scapterisci Nguyen and Smart, 1990 consists of an egg stage, four juvenile stages, and an adult stage (male and female). The cycle from IJ (third stage infective juveniles) to IJ may proceed by one of two routes. If the nutrient supply is sufficient and the population is not overcrowded, the IJ develop to adult males and females of the first generation. Most eggs from these adult females hatch and the juveniles develop through each life stage to become adult males and females of the second generation. Eggs produced by these females develop to IJ. This cycle takes 8-10 days (long cycle) at 24 C. If the nutrient supply is insufficient or if overcrowded, the IJ develop to adult males and females of the first generation, and eggs produced by the females develop directly to IJ. This cycle takes 6-7 days (short cycle). The nematode is less tolerant of lower temperatures and more tolerant of higher temperatures than are other species of the genus. The sex ratio is influenced by temperature. At 15 and 24 C, females constituted 54% and 60% of the population, respectively, but at 30 C females constituted 47% of the population.     

Effect of Initial Nematode Density on Managing Globodera rostochiensis with Resistant Cultivars and Nonhosts

Cropping systems in which resistant potato cultivars were grown at different frequencies in rotation with susceptible cultivars and a nonhost (oats) were evaluated at four initial nematode population densities (Pi) for their ability to maintain Globodera rostochiensis at a target level of <0.2 egg/cm³ of soil. At a Pi of 0.1 to 1 egg/cm³ of soil, cropping systems with 2 successive years of a resistant cultivar every 3 years of potato production reduced and maintained G. rostochiensis at <0.2 egg/cm³ of soil. At a Pi of 1 to 4 eggs/cm³ of soil, 2 successive years of a resistant cultivar followed by 1 year of oats for every 4 years of production were necessary to reduce and maintain G. rostochiensis populations at <0.2 egg/cm³ of soil. At a Pi greater than 4 eggs/cm³ of soil, 2 successive years of a resistant cultivar plus 1 year of oats reduced G. rostochiensis densities to <0.2 egg/cm³ of soil, but the population increased above that density after cropping 1 year to a susceptible cultivar. The numbers of cysts and eggs per cyst in the final population (Pf) of G. rostochiensis were influenced by initial density and the frequency of growing a susceptible cultivar in a cropping system. The lowest number of cysts and eggs per cyst in the final G. rostochiensis population occurred with a cropping system consisting of 2 successive years of a resistant cultivar followed by oats with a susceptible cultivar grown the fourth year of production.     

Comparative Responses of Globodera rostochiensis and G. pallida to Hatching Chemicals

Globodera rostochiensis and G. pallida responded similarly to hatch stimulation by potato root leachate, but proportionally more second-stage juveniles (J2s) of G. rostochiensis hatched than of G. pallida in response to picrolonic acid, sodium thiocyanate, alpha-solanine, and alpha-chaconine. Fractionation of the potato root leachate identified hatching factors with species-selective (active toward both species but stimulating greater hatch of one species than the other), -specific (active toward only one species), and -neutral (equally active toward both species) activities. In a comparison of two populations of each of the two potato cyst nematode (PCN) species, however, greater similarity in response to the individual hatching factors was observed among populations of different species produced under the same conditions than among different populations of the same PCN species. Smaller numbers of species-specific and species-selective hatching factor stimulants and hatching inhibitors than of hatching factors were resolved. In a study to determine whether the different hatching responses of the two species to the same root leachate were associated with different ratios of species-selective and species-specific hatching factors, G. rostochiensis pathotype Ro1 exhibited greater hatch than did G. pallida pathotype Pa2/3 in response to leachate from older plants (more than 38 days old), while G. pallida exhibited greater hatch in response to leachate from younger plants (less than 38 days old); the response of G. pallida pathotype Pal with respect to plant age was intermediate between the other two populations. Combined molecular exclusion-ion exchange chromatography of the root leachates from plants of different ages revealed an increase in the proportion of G. rostochiensis-specific and -selective hatching factors as the plants aged.     

Effects of Host Resistance on Second-stage Juveniles and Adult Males of Globodera rostochiensis

Potato cultivars Katahdin (susceptible) and Rosa (resistant) were exposed to infective second-stage juveniles (J2) of Globodera rostochiensis for varying periods of time, after which root systems were washed and plants were placed in Hoagland''s solution to assess J2 egression and male emergence. After transfer to liquid culture, many J2 egressed from both cultivars, but significantly more egressed from the resistant Rosa than from Katahdin. Juveniles that egressed from Rosa invaded a second host, resistant or susceptible, in significantly fewer numbers than did juveniles that egressed from Katahdin. Also, significantly fewer males developed in and emerged from resistant host roots, relative to susceptible ones. These effects of resistance may be an important component of the tolerance to invasion by G. rostochiensis exhibited by Rosa.     

Major Sperm Protein Genes from Globodera rostochiensis

Three genes in the major sperm protein (MSP) gene family from the potato cyst nematode Globodera rostochiensis were cloned and sequenced. In contrast to the absence of introns in Caenorhabditis elegans MSP genes, these genes in G. rostochiensis contained a 57 nucleotide intron, with normal exon-intron boundaries, in the same relative location as the intron in Onchocerca volvulus. The MSP genes of G. rostochiensis had putative CAAT, TATA, and polyadenylation signals. The predicted G. rostochiensis MSP gene product is 126 amino acids long, one residue shorter than the products in the other species. The comparison of MSP amino acid sequences from four diverse nematode species suggests that O. volvulus, Ascaris suum, and C. elegans may be more closely related to each other than they are to G. rostochiensis.     

Description of Globodera ellingtonae n. sp. (Nematoda: Heteroderidae) from Oregon

A new species of cyst nematode, Globodera ellingtonae, is described from soil collected from a field in Oregon. Second-stage juveniles (J2) of the species are characterized by body length of 365-515 μm, stylet length of 19-22.5 μm, basal knobs rounded posteriorly and pointed anteriorly, tail 39-55 μm, hyaline tail terminus 20-32.5 μm, and tail tapering uniformly but abruptly narrowing and constricted near the posterior third of the hyaline portion, ending with a peg-like, finely rounded to pointed terminus. Cysts are spherical to sub-spherical, dark to light brown and circumfenestrate and cyst wall pattern is ridge-like with heavy punctations. Males have a stylet length of 21-25 μm and spicule length of 30-37 μm with a pointed thorn-like tip. Females have a stylet length of 20-22.5 μm, one head annule and labial disc, heavy punctations on the cuticle, and short vulval slit 7.5-8 μm long. Morphologically this new, round-cyst species differs from the related species G. pallida, G. rostochiensis, G. tabacum complex and G. mexicana by its distinctive J2 tail, and by one or another of the following: shorter mean stylet length in J2, females and males; number of refractive bodies in the hyaline tail terminus of J2; cyst morphology including Granek’s ratio; number of cuticular ridges between the anus and vulva; and in the shape and length of spicules in males. Its relationship to these closely related species are discussed. Based upon analysis of ribosomal internal transcribed spacer (ITS) sequences, G. ellingtonae n. sp. is distinct from G. pallida, G. rostochiensis, G. tabacum and G. mexicana. Bayesian and Maximum Parsimony analysis of cloned ITS rRNA gene sequences indicated three clades, with intraspecific variability as high as 2.8%. In silico analysis revealed ITS restriction fragment length polymorphisms for enzymes Bsh 1236I, Hinf I, and Rsa I that overlap patterns for other Globodera species.     

The Influence of Potato Cultivar on Lipid Content and Fecundity of Bolivian and British Populations of Globodera rostochiensis

The influence of host cultivar on the lipid levels provided by a female to her progeny was investigated with Oil Red O stain and a quantitative image analyzer. A population of Globodera rostochiensis was multiplied at Toralapa Field Station in Bolivia on 25 different potato cultivars grown in that country. The mean neutral lipid content of newly formed second-stage juveniles varied significantly with cultivar over a 200% range. The corresponding range was only 18% and 28% for the same Bolivian and a UK population of G. rostochiensis, respectively, when both completed reproduction concurrently on 10 pot-grown European cultivars in the United Kingdom. Egg numbers per female varied with host for Bolivian cultivars that lack known partial resistance to Globodera spp. There was a 15-fold range between the most and least fecund nematode-host combinations (Kosi and Gendarme). The Bolivian G. rostochiensis population showed only a 2-fold range in mean eggs per cyst when grown on European cultivars in the UK. The fatty acid profiles of lipids from Bolivian G. rostochiensis cysts reared on Bolivian potato cultivars were dominated by C₂₀ (37-64%) and C₁₈ (28-46%) fatty acids and ranged from C₁₄ to C₂₂. The three major fatty acids detected were C_20:4:, C_20:1, and C_18:1. Few differences between cultivars were observed. For a UK population of G. rostochiensis reared on ssp. tuberosum, higher relative percentages of C₁₈ and monounsaturated fatty acids and lower relative percentages of C₂₀ and polyunsaturated fatty acids were found.     

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.     

Relationship between Root Growth of Potato,Root Diffusate Production,and Hatching of Globodera rostochiensis

Hatching response of Globodera rostochiensis in potato root diffusate (PRD) collected by soaking individual potato, Solanum tuberosum, root systems in water for 2 hours was used to assess the relationship between root growth and PRD production. Resistant potato cultivars Hudson and Rosa were used as test plants. Maximum hatch occurred in PRD collected 3 weeks after plant emergence (AE) in the greenhouse, and declined after this time. Hatch was positively correlated with increased root weight only during the first 3 weeks AE. Hudson PRD was consistently more active than Rosa PRD in stimulating hatch, except when adjusted for root weight. Although the results indicated that cells at the root tip produced a more active PRD than cells located elsewhere, PRD appeared to be produced along the entire root. Differences in time length of the vegetative growth phase, extent of root growth, and volume of roots, rather than the production of a more active PRD per se, may explain why Hudson is more effective than Rosa in reducing G. rostochiensis population densities in soil.     

Life Cycle and Reproductive Potential of the Nematode Heterorhabditis bacteriophora Strain HP88

Development of the entomopathogenic nematode Heterorhabditis bacteriophora strain HP88 was studied in vivo with larvae of the greater wax moth, Galleria mellonella, as host and in vitro. At 25 C in vivo, the duration of the life cycle from egg hatch to egg hatch was 96 hours. Juvenile development took 48 hours, with the duration of each juvenile stage ranging from 8 to 12 hours. Under crowded conditions, development proceeded to the infective juvenile (IJ) stage instead of the third juvenile stage (J3). Life-cycle duration and proportion of the various developmental stages in the population were similar in in vitro and in vivo cultures. When in vivo or in vitro development was initiated from the IJ stage, only hermaphrodites developed in the first generation and males appeared only in the second generation. The average (±SD) number of progeny per hermaphrodite was 243 ± 98. The ratio of males to hermaphrodites in the second generation was 1:9.4 ± 6.8.     

Effect of Temperature on Attachment,Development, and Interactions of Pasteuria penetrans on Meloidogyne incognita

The effect of temperature (10, 20, 25, 30, and 35 C) on attachment and development of Pasteuria penetrans on Meloidogyne arenaria race 1 was elevated in growth chambers. The greatest attachment rate of endospores of P. penetrans occurred on second-stage juveniles at 30 C. The bacterium developed more quickly within its host at 30 and 35 C than at 25 C or below. The development of the bacterium within the nematode female was divided into nine recognizable life stages, which ranged from early vegetative thalli to mature sporangia. Mature sporangium was the predominant life stage observed after 35, 40, 81, and 116 days at 35, 30, 25, and 20 C, respectively. The body width and length of M. arenaria females infected with P. penetrans were smaller initially than the same dimensions in uninfected females, but became considerably larger over time at 25, 30, and 35 C. This isolate of P. penetrans also parasitized and completed its life cycle in males of M. arenaria.     

Studies on the Host Range,Biology, and Pathogenicity of Punctodera punctata Infecting Turfgrasses

Punctodera punctata completed its life cycle on Poa annua (annual bluegrass), P. pratensis (Merion Kentucky bluegrass), Lolium perenne (perennial ryegrass), and Festuca rubra rubra (spreading fescue). Minimum time for completion of a life cycle from second-stage juvenile to mature brown cyst was 40 days at 22-28 C. Inoculation by single juveniles indicated that reproduction was most likely by amphimixis. Infestation levels of 50 or 500 juveniles/250 cm³ soil did not affect top dry weight, root dry weight, or total dry weight of Poa annua.     

Effects of Inoculum Density and Egg Age on Establishment of Globodera rostochiensis Populations

The establishment of Globodera rostochiensis Rol populations was examined under greenhouse conditions. The probability of G. rostochiensis population establishment was calculated from the number of plants that produced new cysts with viable eggs following inoculation with various numbers of eggs of different ages. Probability of population establishment was positively correlated with inoculum density but was not affected by the age of eggs used in these experiments. The probability of G. rostochiensis establishment ranged from 5% at densities of 2 eggs/pot to 100% at densities of 25 eggs/pot or greater. At densities of 3 eggs/pot and beyond, there was no correlation between inoculum density and the number of viable eggs/new cyst. Also, the number of plants that produced new cysts was a function of inoculum density and not age of eggs. Juveniles from eggs 1 year old or older were equally as infective as were those from eggs in newly developed cysts (4 months old).     

Development of Heterodera glycines Life Stages as Influenced by Temperature

The effects of temperature on rates of development of Heterodera glycines egg and juvenile stages were examined as a basis for predicting generation times of the nematode on soybean. The relationship of temperature to H. glycines embryonic development between 15 and 30 C was described by a linear model, The calculated basal temperature threshold was 5 C. Thermal optimum for embryogenesis and hatch with low mortality was 24 C. Development proceeded to first-stage juvenile at 10 C and to second-stage juvenile at 15-30 C. Hatch occurred at 20-30 C. At 36 C, development proceeded to the four-cell stage, then the eggs died. The range of diurnal soil temperature fluctuation and accumulated degree-days between 5 and 30 C (DD5/30) had an impact on rate of development of juveniles in soybean roots. From early June to early July, H. glycines required 534 + 24 DD5/30 (4 weeks) to complete a life cycle in the field. During the midseason (July and August), life cycles were completed in 3 weeks and 429 ± 24 DD5/30 were accumulated. Late in the season (September to November), declining soil temperatures were associated with generation times of 4 weeks and slower rates of development.     

Hatching Behavior of Potato Cyst Nematodes from the Canary Islands

The present work investigated early hatching differences in naturally occuring field populations and newly reared populations of potato cyst nematodes from the Canary Islands. Hatching behavior of the two species appears to be distinct, with more juveniles hatched from G. pallida that hatch earlier and over a shorter time than G. rostochiensis. The hatching rate of 3-year-old PCN populations was more than double (mean 44.5% ñ 1) that shown by newly reared populations (mean 19.1% ñ 12.5), and those that could be classified as pathotype Pa 1 (Pa 1 and P 13) were found to hatch particularly poorly. Significant differences were also observed in the juveniles released in tap water between newly reared populations of both species, with mean hatch significantly higher for G. rostochiensis. The results are discussed in relation to the implication that these findings may have for competition between the two species of PCN in the field.