In Vitro Culture of Subanguina picridis in Acroptilon repens Callus,Excised Roots,and Shoot Tissues

The knapweed nematode Subanguina picridis is a foliar parasite that is of interest as a biological weed control agent of Russian knapweed. Attempts were made to culture the nematode in callus, excised roots and in shoots derived from roots of Russian knapweed. In callus tissues, the nematode developed from second-stage juvenile to adult but failed to reproduce; it developed only to the fourth stage in excised roots. However, S. picridis was successfully cultured in vitro in shoots derived from roots. The nematode induced galls on the leaves, petioles, and shoot apices and developed and reproduced inside the galls. Gibberellic acid increased the development rate of the nematode and promoted the formation of males. This is the first gnotobiotic culture of a nematode used for biological weed control.     

Morphological and Biological Parameters of the Knapweed Nematode, Subanguina picridis

Specimens of the knapweed nematode Subanguina picridis (Kirjanova) Brzeski obtained from different host plants were highly variable in measurement and structure. This variability refutes the validity of six Subanguina species attacking plants in the Asteraceae.     

Biology of Subanguina picridis,a Potential Biological Control Agent of Russian Knapweed

The knapweed nematode, Subanguina picridis, forms galls on the leaves, stems, and root collar of Russian knapweed, Acroptilon repens. After being revived from a dormant, cryptobiotic state, second-stage juveniles required at least 1 month in a free-living state before becoming infective. Galls were induced on relatively slow-growing host plants that retained their apical meristems at or near the soil surface for 2-5 weeks. Galls developed extensive areas of nutritive tissue. The nematode was introduced from the Soviet Union and released in Canada for the biological control of Russian knapweed.     

Effects of Temperature,Shoot Age,and Medium on Gall Induction by Subanguina picridis in Vitro

The influence of temperature, shoot age, and medium on gall induction by Subanguina picridis on Russian knapweed (Acroptilon repens) was examined in vitro. The optimal temperature for gall formation was 20 C. Gall induction was delayed as the temperature decreased, and decreased as shoot age increased. Bud primordia (0-day-old shoots and 5-day-old shoots) with an average length of 4.2 mm and 7.9 mm were the most suitable tissues for nematode development and gall formation. Gall formation was more effective on B5G medium than on MSG. Young shoots under slow growth were most suitable for mass rearing of S. picridis.     

Host Range of,and Plant Reaction to,Subanguina picridis

The host range of the knapweed nematode, Subanguina picridis (Kirjanova) Brzeski, under controlled environmental conditions was extended to include, in addition to Russian knapweed, Acroptilon repens (L.) DC., plant species within the Centaureinae, and Carduinae subtribes of the Cynareae tribe of the Asteraceae family. Examination of host response to nematode infection revealed that Russian knapweed was the only highly susceptible host plant. Diffuse knapweed (Centaurea diffusa Lam.) was moderately susceptible, and other plants which formed galls were resistant to S. picridis.     

Interaction between Meloidogyne incognita and Hoplolaimus columbus on Davis Soybean

Greenhouse and laboratory experiments were performed to determine if an interaction exists between Meloidogyne incognita and Hoplolaimus columbus on Davis soybean. Greenhouse tests were performed with three population levels of M. incognita and H. columbus (0, 1,500, 6,000/1.5-liter pot) separately and in all combinations. Dry root weight (DRT) declined nonlinearly and dry shoot weight (DST) declined linearly with respect to increasing initial populations of M. incognita and H. columbus. When the two nematode species were added to the soil together, the amount of DRT and DST suppression by one species was dependent on the initial level of the concomitant species. The final root population of M. incognita or H. columbus declined linearly with increasing initial population density of the concomitant species. H. columbus suppressed M. incognita populations in the soil nonlinearly, but M. incognita had no effect on H. columbus.     

Population Dynamics of Ditylenchus destructor on Peanut

The population development of Ditylenchus destructor in the roots, pegs, hulls, and seeds of eight peanut (Arachis hypogaea) genotypes was studied in the greenhouse. Although all genotypes tested were good hosts for D. destructor, differences in host suitability were observed. Invasion of the plant parts by Ditylenchus destructor proceeded more slowly in genotypes with long growth periods. During the second half of the growth period of these genotypes, D. destructor populations emigrated from the hulls and seeds into the soil but reinfected the pods after a few weeks. The genotypes with the longest growth periods supported the highest number of nematodes when each genotype was harvested at its usual harvest time. The long-season genotypes supported low numbers of nematodes when harvested before crop maturity.     

Survival of Paecilomyces lilacinus in Selected Carriers and Related Effects on Meloidogyne incognita on Tomato

Laboratory and microplot experiments were conducted to determine the influence of carrier and storage of Paecilomyces lilacinus on its survival and related protection of tomato against Meloidogyne incognita. Spores of P. lilacinus were prepared in five formulations: alginate pellets (pellets), diatomaceous earth granules (granules), wheat grain, soil, and soil plus chitin. Fungal viability was high in wheat and granules, intermediate in pellets, and low in soil and chitin-amended soil stored at 25 ± 2 C. In 1985 P. lilacinus in field microplots resulted in about a 25% increase in tomato yield and 25% gall suppression, compared with nematodes alone. Greatest suppression of egg development occurred in plots treated with P. lilacinus in pellets, wheat grain, and granules. In 1986 carryover protection of tomato against M. incognita resulted in about a threefold increase in tomato fruit yield and 25% suppression of gall development, compared with plants treated with nematodes alone. Higher numbers of fungus-infected egg masses occurred in plots treated with pellets (32%) than in those treated with chitin-amended soil (24%), wheat (16%), granules (12%), or soil (7%). Numbers of fungal colony-forming units per gram of soil in plots treated with pellets were 10-fold greater than initial levels estimated at planting time in 1986.     

Impact of Paecilomyces lilacinus Inoculum Level and Application Time on Control of Meloidogyne incognita on Tomato

Microplot experiments were conducted to evaluate the effects of inoculum level and time of application of Paecilomyces lilacinus on the protection of tomato against MeIoidogyne incognita. The best protection against M. incognita was attained with 10 and 20 g of fungus-infested wheat kernels per microplot which resulted in a threefold and fourfold increase in tomato yield, respectively, compared with tomato plants treated with this nematode alone. Greatest protection against this pathogen was attained when P. lilacinus was delivered into soil 10 days before planting and again at planting. Yield was increased twofold compared with yield in nematode-alone plots and plots with M. incognita plus the fungus. Percentages of P. lilacinus-infected egg masses were greatest in plots treated at midseason or at midseason plus an early application, compared with plots treated with the fungus 10 days before planting and (or) at planting time.     

Liquid Culture of the Entomogenous Nematode Steinernema feltiae with Its Bacterial Symbiont

The insect-parasitic nematode, Steinernema feltiae Filipjev strain 42, was reared in liquid culture along with its bacterial symbiont, Xenorhabdus nematophilus Thomas &Poinar. First-stage juveniles developed into reproducing adults in a maintenance salts medium containing resuspended Xenorhabdus cells and the yeast Kluyveromyces marxianus (Hansen) van der Walt or cholesterol. Cultures with media depths greater than 4 mm required aeration. Nematode populations increased as bacterial density increased. An optimal culture system was obtained when the bacteria and nematodes developed in a semidefined medium containing tryptic soy, yeast extract, and cholesterol and were incubated on a rotary shaker at 25 ± 1 C. Under these conditions, up to 86% of the final population were infective juveniles.     

Effects of a Mutant Strain and a Wild Type Strain of Verticillium lecanii on Heterodera glycines Populations in the Greenhouse

A wild type strain ofVerticillium lecanii and a mutant strain with increased tolerance to the fungicide benomyl were evaluated in greenhouse experiments for effects on Heterodera glycines populations. Nematodes were applied at 300 eggs and juveniles per 4,550-cm³ pot (two soybean plants in 4,990 g loamy sand per pot) and at both 300 and 10,000 eggs and juveniles per 1,720-cm³ pot (one soybean plant in 2,060 g sand per pot). With 300 nematodes added per pot, both V. lecanii strains significantly reduced nematode populations in loamy sand (fungus applied at 0.02% dry weight per dry weight loamy sand) and sand (0.006% and 0.06% fungus application rates). The mutant strain applied at 0.002% to sand also significantly reduced cyst numbers. When 10,000 nematodes were added per pot, only the mutant strain at 0.06% significantly decreased population. Various media were tested for isolation of the fungus strains from prills, loamy sand, and sand, but the fungi were recovered from few of the greenhouse pots.     

Histopathology of Ditylenchus destructor on Peanut

The time and mode of entry, and development of Ditylenchus destructor in peanut were studied in field and greenhouse experiments. Few nematodes were present in the cortex of the roots. At 90-120 days after planting, D. destructor was observed in the exocarp at the base of the pod near the point of connection with the peg. The peg was invaded from this primary infection site. The endocarp of the hull was usually penetrated through openings at the base of the mesocarp and sometimes at the pod apex. Numerous D. destructor were present in the testa and the vascular bundles. Nematodes were found in the embryo but not in the cotyledons. The histopathology of D. destructor closely resembles that of the peanut testa nematode, Aphelenchoides arachidis Bos.     

Factors Affecting Population Trends of Plant-Parasitic Nematodes on Rangeland Grasses

The effects of environmental conditions on population trends of plant-parasitic nematodes were studied in experimental plots of five wheatgrasses in the western Utah desert. In a 3-year (1984-86) field study, soil water and temperature affected the population trends of the ectoparasites, Tylenchorhynchus acutoides and Xiphinema americanum, and the migratory endoparasite, Pratylenchus neglectus, on Fairway crested wheatgrass, Agropyron cristatum; ''Hycrest'' crested wheatgrass, A. cristatum X A. desertorura; ''Rosana'' western wheatgrass, Pascopyrum smithii; ''Oahe'' intermediate wheatgrass, Thinopyrum intermedium; and RS-1 hybrid (Elytrigia repens X Pseudoroegneria spicata). The largest soil populations of these nematode species were collected in 1984 under good plant-growth conditions. A reduction in nematode populations occurred in 1985 and 1986, possibly because of low soil-water conditions. There was a positive relationship between high soil water and maximum population densities of T. acutoides in the spring and fall of 1984, and between low soil water and minimum population densities of the nematode in 1985 and 1986. Pratylenchus neglectus populations were affected by soil water, although to a lesser degree than the ectoparasitic nematodes. Population densities of the three nematode species were significantly lower in the drier years of 1985 and 1986 than in 1984. Nematode populations were greater at the lower soil depths in the fall than in the spring or summer.     

Population Development and Influence of Bursaphelenchus xylophilus on Gliocladium virens

Gliocladium virens was isolated from slash pine trees symptomatic and asymptomatic for pine wilt disease with frequencies of 24% and 10%, respectively. Populations of Bursaphelenchus xylophilus, the nematode incitant of this disease, reproduced on this fungus and inhibited its growth. Growth inhibition of the fungus was characterized by an absence of sporulation and by the formation of chains of dark, thick-walled, chlamydospore-like cells. Population increase during a 12-day period following infestation of cultures of the fungus with 10,000 nematodes averaged 3-fold at 16 C, 9-fold at 20 C, and 24-fold at 24 C. In greenhouse studies, nematode recovery from slash pine seedlings coinoculated with both organisms was significantly greater than that obtained from seedlings inoculated with the nematode alone.     

Phytotoxic polyacetylenes from roots of Russian knapweed (Acroptilon repens (L.) DC.)

There are several factors thought to assist invasive weeds in colonization of ecosystems. One of these factors is allelopathy, the negative effect of chemicals produced by one plant on neighboring plants, frequently mediated through root exudates and other plant leachates. Acroptilon repens (Asteraceae) is one of the most invasive and ecologically threatening weed species in western North America. A bioassay-guided fractionation of the root extracts of this plant led to the isolation of five polyacetylenic compounds, of which one [5′-methoxy-1′-(5-prop-1-yn-1-yl-2-thienyl)-hexa-2′,4′-diyin-6′-yl acetate] was hitherto unknown. The structures of these compounds were elucidated on the basis of spectroscopic analysis (IR, ESIMS, ¹H, ¹³C NMR and 2D NMR). All of the compounds obtained, except 1-chloro-4-(5-penta-1,3-diyn-1-yl-2-thienyl)but-3-yn-2-ol, showed phytotoxic activity against Arabidopsis thaliana seedlings. The presence of 4′-chloro-1′-(5-penta-1,3-diyn-1-yl-2-thienyl)-but-2′-yn-3′-ol was detected in the root exudates of aeroponically grown A. repens plants. None of the polyacetylenes isolated in this study were found in Colorado soils collected between September 2006 and July 2007 in an A. repens colonized site. However, polyacetylene 5 in A. repens infested soil from Washington was found in June, 2007. Contrary to our previous report, the compound 7,8-benzoflavone (6) was not detected in root exudates, nor was it encountered in extracts of roots, aerial parts or infested soil. Since we could not repeat this work, the original report has been retracted [Stermitz, F.R., Bais, H.P., Foderaro, T.A., Vivanco, J.M., 2003. 7,8-Benzoflavone: a phytotoxin from root exudates of invasive Russian knapweed [A retraction]. Phytochemistry 64, 493-497.].     

Damage Functions for Three Meloidogyne Species on Arachis hypogaea in Texas

The yield response of Florunner peanut to different initial population (Pi) densities of Meloidogyne arenaria, M. javanica, and an undescribed Meloidogyne species (isolate 93-13a) was determined in microplots in 1995 and 1996. Seven Pi''s (0, 0.5, 1, 5, 10, 50, and 100 eggs and J2/500 cm³ soil) were used for each Meloidogyne species in both years. The three species reproduced abundantly on Florunner in both years. In 1995, mean reproduction differed among the three species; mean Rf values were 10,253 for isolate 93-13, 4,256 for M. arenaria, and 513 for M. javanica. In 1996, the reproduction of M. arenaria (mean Rf = 7,820) and isolate 93-13a (mean Rf = 7,506) were similar, and both had greater reproduction on peanut than did M. javanica (mean Rf = 2,325). All three nematode species caused root and pod galling, and a positive relationship was observed between Pi and the percentage of pods galled. Meloidogyne arenaria caused a higher percentage of pod galling than did M. javanica or isolate 93-13a. A negative linear relationship between log₁₀ (Pi + 1) and pod yield was observed for all three nematode species each year. The yield response slopes were similar except for that of M. javanica, which was less negative than that of isolate 93-13a in 1995, and less negative than that of M. arenaria and isolate 93-13a in 1996.     

Development of Meloidogyne arenaria on Peanut and Soybean under Two Temperature Cycles

Florunner peanut and three soybean cultivars, Centennial, Gasoy 17, and Wright, were inoculated with 48-hour age cohorts of Meloidogyne arenari race 1 second-stage juveniles and placed in a growth chamber set to simulate early season (low temperature) and midseason (high temperature) conditions. Percentages of the initial inoculum penetrating roots 4 and 8 days after inoculation were 2-3 times higher in soybean cultivars than in peanut; 25% on susceptible soybean and 9% on peanut. Penetration and early development of M. arenaria were greater in the higher temperature environment. Penetration percentages were expressed as a function of cumulative degree-days by regression models. Development of M. arenaria 10, 20, and 30 days after inoculation was more rapid on peanut than on soybean. The resistant soybean cultivar Wright had slower development rates than did the other two soybean cultivars. Nematode growth and development were dependent on temperature. In greenhouse experiments, production of eggs by M. arenaria was more than 10 times greater on peanut than on susceptible soybean. The reproductive factor for Wright soybean was less than one, but plant growth parameters indicated that this cultivar was intolerant of M. arenavia.     

Induced Resistance to Meloidogyne hapla by other Meloidogyne species on Tomato and Pyrethrum Plants

Advance inoculation of the tomato cv. Celebrity or the pyrethrum clone 223 with host-incompatible Meloidogyne incognita or M. javanica elicited induced resistance to host-compatible M. hapla in pot and field experiments. Induced resistance increased with the length of the time between inoculations and with the population density of the induction inoculum. Optimum interval before challenge inoculation, or population density of inoculum for inducing resistance, was 10 days, or 5,000 infective nematodes per 500-cm³ pot. The induced resistance suppressed population increase of M. hapla by 84% on potted tomato, 72% on potted pyrethrum, and 55% on field-grown pyrethrum seedlings, relative to unprotected treatments. Pyrethrum seedlings inoculated with M. javanica 10 days before infection with M. hapla were not stunted, whereas those that did not receive the advance inoculum were stunted 33% in pots and 36% in field plots. The results indicated that advance infection of plants with incompatible or mildly virulent nematode species induced resistance to normally compatible nematodes and that the induced resistance response may have potential as a biological control method for plant nematodes.     

Laboratory Culture and Life History of Heleidomermis magnapapula in Its Host, Culicoides variipennis (Diptera: Ceratopogonidae)

The mermithid parasite Heleidomermis magnapapula was maintained in larvae of the midge Culicoides variipennis for 20 months in enamel pans containing nutrient-rich water and polyester pads as a substrate. Inseminated female mermithids were introduced to the pad surface when the host was in the late second or early third-instar. Host larvae were harvested from the pans 9 days after exposure and held in tap water for nematode emergence. Preparasite yield was positively correlated with female nematode size and averaged 1,267 preparasites/female. Male and female nematodes emerged an average of 12.2 and 13.4 days after host exposure, respectively. Supplemental host food (Panagrellus) during the final days of parasitism did not alter time of emergence. Parasites emerging singly were 64% females, whereas superparasitized hosts yielded males (up to nine/host). Nematode carryover into the adult midge normally occurred at a level of 0.5-2.5%. Parasite load (nematodes/ parasitized individual) in midge adults was lower than that of larvae from the same cohort, and adult midges were more likely to harbor female parasites. Exposure of fourth-instar host larvae resulted in higher levels of adult parasitism (up to 17%).