The Potential of Thiarubrine C as a Nematicidal Agent against Plant- parasitic Nematodes.

Thiarubrine C, a polyacetylenic 1,2-dithiin isolated from the roots of Rudbeckia hirta (Asteraceae), exhibited strong nematicidal activity in in vitro and growth chamber assays. Thiarubrine C was toxic, in the absence of light, to the plant-parasitic nematodes Meloidogyne incognita and Pratylenchus penetrans at LCs of 12.4 ppm and 23.5 ppm, respectively. A minimum exposure time between 12 and 24 hours was the critical period for nematode mortality due to thiarubrine C. Although thiarubrine C was not totally dependent on light for toxicity, activity was enhanced in the presence of light, especially with the microbivorous nematode, Teratorhabditis dentifera. Upon exposure of M. incognita juveniles to 20 ppm thiarubrine C for 1 hour, infection of tomato plants was greatly reduced compared to untreated checks. Thiarubrine C was also effective in reducing plant infection when mixed with soil 24 hours prior to or at planting, unlike other related compounds such as delta-terthienyl.

First Report of Pratylenchus neglectus in New York.

Soil samples were collected from a field in Wyoming County near Portageville, NY, on 29 October 1992 and 23 November 1993. The field was planted with winter wheat (Triticum aestivum L.) from September 1991 to August 1992, and with potato (Solanum tuberosum L.) cv. Superior from May 1993 to September 1993. Thirty-nine and 45 samples were collected along three transects in 1992 and 1993, respectively. Two species of Pratylenchus were identified, P. neglectus (Rensch) Filipjev & Sch. Stek. (identification confirmed by A. M. Golden) and P. penetrans (Cobb) Filipjev & Sch. Stek. Their combined population densities were 134 ± 18 (mean ± SE per 100 cm3 soil) in 1992 and 195 ± 12 in 1993. P. neglectus was found in all samples, whereas P. penetrans was found in 33 and 78% of the samples in 1992 and 1993, respectively. Where both species occurred together, the density of P. neglectus was 7 to 8 times that of P. penetrans. In both years, spores of a nematode-pathogenic bacterium (probably a Pasteuria sp.) were found attached to the cuticle of P. penetrans (68% with spores in 1993) but were rarely attached to P. neglectus (less than 1% with spores). Although both Pratylenchus spp. are widespread in North America, this is the first report of P. neglectus in New York. Its dominance in the field was unexpected because P. penetrans and P. crenatus Loof are typically the dominant Pratylenchus spp. in potato production regions of northeastern North America (1). The presence of the bacterium, which appeared to be host-specific for P. penetrans, may help explain the dominance of P. neglectus. Reference: (1) R. N. Huettel et al. Am. Potato J. 68:345, 1991.

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/cm(3) of soil. At a Pi of 0.1 to 1 egg/cm(3) 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/cm(3) of soil. At a Pi of 1 to 4 eggs/cm(3) 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/cm(3) of soil. At a Pi greater than 4 eggs/cm(3) of soil, 2 successive years of a resistant cultivar plus 1 year of oats reduced G. rostochiensis densities to <0.2 egg/cm(3) 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.

Characterization of Pratylenchus penetrans from Ten Geographically Isolated Populations Based on Their Reaction on Potato.

Single female cuhures of Pratylenchus penetrans were established from soil and root samples collected from 10 geographically isolated locations in North America. The resultant isolates were used to evaluate nematode egression from and multiplication on roots of potato clones to distinguish intraspecific differences among isolates. The 10 nematode isolates were statistically separated into four groups based on percentage of nematodes that egressed from the P. penetrans-resistant potato done L 118-2. The Cornell (CR), Wisconsin (WI), Long Island (LI), and Adirondack (AD) isolates, selected as representative isolates of each of the four groups, exhibited 53%, 39%, 25%, and 10% egression from L118-2, respectively. Reproduction of these four isolates was measured on three potato cultivars (Russet Burbank, Butte, and Hudson) and two breeding lines (NY85 and L118-2). The LI and AD isolates reproduced well on all five potato clones. The CR isolate reproduced well on Russet Burbank and NY85 but significantly less on Butte, Hudson, and L118-2. Reproduction of the WI isolate was less than the LI and AD isolates but more than the CR isolate on all potato clones tested except Russet Burbank. Reproduction of the WI isolate on Russet Burbank was less than the other three isolates. Based on these results, four distinct intraspecific variants of P. penetrans are proposed: Cornell, Wisconsin, Long Island, and Adirondack.

Lipid Composition of Cyst Stages of Globodera rostochiensis.

Lipid compositional analysis was conducted on the white, yellow, and brown cyst stages of Globodera rostochiensis (golden cyst nematode). Triacylglycerols were the largest lipid fraction in all stages examined, ranging from 55-75% of total lipid. Ethanolamine phosphoglycerides and choline phosphoglycerides were present in high amounts in all cyst fractions, with a total phospholipid content of 20%, 14.7%, and 12.8% in the white, yellow, and brown cyst stages, respectively. Sterols, steryl esters, sphingomyelin, and cardiolipin were found in minor amounts in all three cyst stages and showed greater changes than other classes of lipids relative to cyst stage. The fatty acid compositions of the three cyst stages were similar. Eicosenoic acid (20:1) and arachidonic acid (20:4) were found in higher concentrations than other fatty acids in all cyst preparations; vaccenic acid (18:1) occurred at the third highest concentration. More than 78% of total fatty acids were unsaturated at all cyst stages, and more than 60% were of C20 or longer chain length. The lipid profile of all three cyst stages is consistent with invertebrate adaptation to low-temperature environments.

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.

Effect of Hirsutella rhossiliensis on Infection of Potato by Pratylenchus penetrans.

We evaluated the ability of the nematode-pathogenic fungus Hirsutella rhossiliensis (Deuteromycotina: Hyphomycetes) to reduce root penetration and population increase of Pratylenchus penetrans on potato. Experiments were conducted at 24 C in a growth chamber. When nematodes were placed on the soil surface 8 cm from a 14-day-old potato cutting, the fungus decreased the number entering roots by 25%. To determine the effect of the fungus on population increase after the nematodes entered roots, we transplanted potato cuttings infected with P. penetrans into Hirsutella-infested and uninfested soil. After 60 days, the total number of nematodes (roots and soil) was 20 +/- 4% lower in Hirsutella-infested than in uninfested soil.

Identification of RFLP markers linked to the H1 gene conferring resistance to the potato cyst nematode Globodera rostochiensis.

The H1 gene from Solanum tuberosum ssp. andigena confers high levels of resistance to the potato cyst nematode Globodera rostochiensis and is used extensively in potato breeding. Using a dihaploid segregating population, a search was conducted for linkage between this gene and markers on the potato/tomato RFLP map. A total of 60 RFLP markers covering the entire genome were screened on bulk resistant and susceptible segregants. Linkage was indicated for eight markers on chromosome 5. Individual plant analysis placed the closest marker, CD78, at a maximum map distance of 2.7 cM from H1. A molecular marker for the H1 should be useful both as a correlative screening tool for incorporation of resistance into new cultivars and as starting point for map-based cloning of this important gene.

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).

Probability of Globodera rostochiensis Spread on Equipment and Potato Tubers.

The probability of spreading cysts of Globodera rostochiensis on farming equipment and potato tubers was investigated in naturally infested field plots. The number of cysts recovered from soil that adhered to equipment differed significantly between different pieces of equipment. These differences were related to initial nematode density and, in most cases, to the volume of soil that adhered to the equipment. At an initial density of 0.04 egg/cm(3) of soil, significantly more cysts were recovered from a potato digger than from a potato hiller, cultivator, or plow. At an initial density of 0.90 egg/cm(3) of soil, significantly more cysts were recovered from the plow than from the other equipment. Although the population density was 22 times greater, only 10 times more cysts adhered 3 to equipment used in soil with a density of 0.90 egg/cm(3) of soil than when used in soil infested at 0.04 egg/cm(3). The number of potato tuber samples (4.5 kg) that contained cysts with viable eggs was positively correlated with the initial densities of G. rostochiensis in soil in which they were produced. The percentage of tuber samples with cysts containing viable eggs was 10-12% for tubers harvested from soil with densities less than 1 egg/cm(3) and 30-76% for tubers harvested from soil with densities greater than 4 eggs/cm(3) of soil.

Infection of Pratylenchus penetrans by Nematode-pathogenic fungi.

Eleven fungal isolates were tested in agar dishes for pathogenicity to Pratylenchus penetrans. Of the fungi that produce adhesive conidia, Hirsutella rhossiliensis was a virulent pathogen; Verticillium balanoides, Drechmeria coniospora, and Nematoctonus sp. were weak or nonpathogens. The trapping fungi, Arthrobotrys dactyloides, A. oligospora, Monacrosporium dlipsosporum, and M. cionopagum, killed most of the P. penetrans adults and juveniles added to the fungus cultures. An isolate of Nematoctonus that forms adhesive knobs trapped only a small proportion of the nematodes. In 17-cm(3) vials, soil moisture influenced survival of P. penetrans in the presence of H. rhossiliensis; nematode survival decreased with diminishing soil moisture. Hirsutella rhossiliensis and M. ellipsosporum were equally effective in reducing numbers of P. penetrans by 24-25% after 4 days in sand. After 25 days in soil artificially infested with H. rhossiliensis, numbers of P. penetrans were reduced by 28-53%.

Resistance in Potato to Pratylenchus penetrans.

Potato clones from five different breeding populations were evaluated for their relative resistance and susceptibility to Pratylenchus penetrans. Resistance and susceptibility were distinguished by an index of susceptibility (SI) calculated from the numbers of P. penetrans (including eggs) per g of root of individual clones in relation to that of a susceptible control at 30 or 70 days after inoculation. Evaluations were carried out using 7.5-cm clay pots in a growth chamber at 24 C with 15-hour day length. In the initial evaluation, 70 days after inoculation, the SI of individual clones ranged from 0.01 to 0.75. Clones that supported the least P. penetrans were from a breeding population derived from Solanum tuberosum ssp. andigena that was originally selected for its resistance to the potato cyst nematode, Globodera pallida. In succeeding tests, these clones had a significantly low SI than did susceptible controls or cultivars that were previously reported to possess resistance to P. penetrans, except cv. Hudson. Resistance to P. penetrans from the Pallida-resistant breeding population was incorporated into potato germplasm better adapted to North American growing conditions.

Effects of Host Resistance on the Fecundity of Globodera rostochiensis.

The fecundity of Globodera rostochiensis (RA) females that developed on resistant Rosa and susceptible Katahdin potato cultivars were compared. Cysts collected from each cultivar were bulked, separated into four sizes (> 500 mum, 355-500 mum, 250-355 mum, and < 250 mum), and crushed to determine fecundity as measured by viable egg content (VEC). Fewer and generally smaller cysts developed on Rosa than on Katahdin. Although cyst size significantly (P = 0.01) influenced VEC, cyst age (8 or 13 weeks) had no effect. Regardless of size, cysts produced on Rosa contained significantly fewer viable eggs than did cysts produced on Katahdin. The fecundity of progeny from cysts produced on Rosa was significantly reduced compared with that of progeny from cysts produced on Katahdin. After two generations on Katahdin, the VEC of cysts from a population originating from Rosa was significantly less than that of cysts from a population originating from Katahdin, indicating that in the presence of a pure population of G. rostochiensis RA, the females that develop on the resistant cultivar Rosa represent a diminished rather than a superior selected population.

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.

Relationship Between Morphology and Parasitism in Two Populations of Meloidogyne incognita.

The reliability of morphological characters and host differential plants for distinguishing between two populations of Meloidogyne incognita was studied. Population A (originally from North Carolina) had incognita-type perineal patterns. A single egg mass subpopulation of population A had a mixture of incognita and acrita perineal patterns with 33% of the patterns atypical for either species. Population B (from Georgia) had predominantly acrita-type patterns with only about 5% atypical patterns. The head shapes of males from both populations were mainly M. incognita. On the basis of stylet length, both populations conformed to M. incognita acrita. Both populations were identified as M. incognita race 1 by reaction on the North Carolina differential hosts. Reactions on azalea and pepper gave no clear identification of the populations. We concluded that there is no relation between perineal pattern, male head shape, and parasitism of host differentials with the two populations studied.

Extraction of cyst nematodes from organic soils.

The effects of extraction technique, sample size, soil moisture level, and overflow rate on recovery of Globodera rostochiensis and (or) Heterodera schachtii cysts from organic soils were investigated. A modified Fenwick can (MFC) and an underflow elutriator (UE) described in this paper were evaluated and compared for cyst recovery efficiency and amount of organic flotsam collected. The MFC and UE extracted similar numbers of cysts, but the UE collected 50% less flotsam than the MFC. Sample size was negatively correlated with cyst recovery and positively correlated with amount of flotsam. The amount of flotsam recovered with the MFC was correlated with overflow speed. Presoaking air dried samples for 30 minutes halved the amount of flotsam without affecting cyst recovery. Extracting cysts from wet soil without prior drying resulted in negligible recovery with both extraction techniques. There were no significant differences in cyst recovery of the two genera tested.

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.

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.

Comparison of Compatible and Incompatible Response of Potato to Meloidogyne incognita.

One susceptible (D6) and two resistant (E2 and N4) clones of Solanum sparsipilum x (S. phureja x 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.

Management of Globodera rostochiensis as Influenced by Nematode Population Densities and Soil Type.

The effects of aldicarb, oxamyl, 1,3-D, and plastic mulch (solarization) on soil population densities of the golden nematode (GN) Globodera rostochiensis was assessed in field and microplot experiments with different soil types. Oxamyl was evaluated in both soil and foliar treatments, whereas aldicarb, 1,3-D, and solarization were applied only to soil. Soil applications of aldicarb and oxamyl resulted in reduced nematode populations after GN-susceptible potatoes in plots with initial population densities (Pi) of > 20 and 7.5 eggs/cm(3) soil, respectively, but nematode populations increased in treated soil when Pi were less than 20 and 7.5 eggs/cm(3)soil. In clay loam field plots with Pi of 19-76 eggs/cm(3) soil, nematode densities increased even with repeated foliar applications of oxamyl, whereas nematode populations at Pi greater than 76 eggs/cm(3) soil were reduced by foliar oxamyl. Treatment with 1,3-D or solarization, singly or in combination, reduced GN soil population densities regardless of soil type or Pi. Temperatures lethal to GN were achieved 5 cm deep under clear plastic but not 10 or 15 cm deep.