Infectivity of Pratylenchus penetrans on Alfalfa

The infectivity of Pratylenchus penetrans on alfalfa seedlings cv. Du Pulls was studied. The dense root-hair zone was the preferred zone of penetration by females, males, and third-stage larvae. A lesion initially appeared as a water-soaked area at the root surface, becoming yellow and elliptical as the nematode entered the cortex, with dark-brown cells later appearing in the centre as the nematode fed. At 20 C, females penetrated roots earlier, faster, and in greater numbers than either males or third-stage larvae. Females penetrated roots at temperatures from 5 to 35 C, with maximum penetration between 10 and 30 C, while males and third-stage larvae penetrated roots only between 10 and 30 C with maximum penetration a t 20 C. Penetration of roots by females, males, and third-stage larvae increased after storage of 5 C for 35 days, but decreased after storage of 140 days or more. Combinations of the three life stages in pairs neither enhanced nor inhibited penetration of roots by individual life stages; males were not attracted to females. Increasing inoculum density up to 20 nematodes/seedling did not affect penetration.     

Penetration of Celery and Alfalfa Roots by Pratylenchus penetrans as Affected by Temperature

A greater percentage of females than juveniles or males of P. penetrans penetrated celery roots grown in infested soil at 5, 18, or 30 C; the difference was greatest at 5 C. The time of initial penetration of alfalfa seedlings inoculated with single nematodes on water agar varied with temperature. Females penetrated the seedlings earlier and over a wider range of temperatures than did males or juveniles. The rate of penetration was highest for females. After initial penetration, the penetration rate decreased with time. At 13-28 C, approximately 80% of roots were penetrated by females and only 25-30% by males and juveniles by the end of the experiment.     

Nematode Response to Carbofuran

Higher populations of Meloidogyne incognita larvae and Pratylenchus penetrans were recovered from soil treated with carbofuran 10 and 15 days after treatment, respectively, than were recovered from untreated control soil. The number of P. penetrans, however, was lower 50 days after treatment, and symptoms developed only occasionally on the root systems of host plants. Populations of Tylenchorhynchus claytoni inoculated at different distances from the base of corn seedlings growing in carbofuran-treated soil did not move toward the plant, whereas they were attracted in untreated soil from a distance of 12 cm. P. penetrans moved at random in treated agar medium when inoculations occurred 4 cm away from the root tips of tomato seedlings under aseptic conditions. Those nematodes that reached the roots were never observed feeding during a 20-day observation period. Specimens of P. penetrans placed on the developing roots moved at random and never penetrated. In contrast, numerous P. penetrans penetrated roots of seedlings growing in untreated medium.     

Infection of Red Clover Seedlings by Heterodera trifolii Goffart and Pratylenchus penetrans (Cobb)

Penetration of 3-day-old ''Kenland'' red clover seedlings by Heterodera trifolii Goffart and Pratylenchus penetrans (Cobb) was investigated in 50-mm petri dishes on 1% agat or discs of Miracloth® (Chicopee Mills, Inc., New York, N.Y. 10018). Penetration by both nematodes increased arithmetically with increased numbers in the inoculum. H. trifolii larvae slowly penetrated all root tissue but in relatively low numbers, 25-30% of inocula. Swelling of larvae and formation of syncytia within roots was detectable 96 hr after inoculation. Initially, adults of P. penetrans preferentially penetrated the region 3-10 mm behind the root tip, but subsequently they invaded all along roots. P. penetrans penetration efficiency was high, 75-90% of inocula, and penetration was relatively rapid. When introduced simultaneously neither nematode affected the invasiveness of the other. Prior infection by H. trifolii did not affect the invasiveness of P. penetrans.     

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.     

Early Root Response to Meloidogyne incognita in Resistant and Susceptible Alfalfa Cultivars

The early events of Meloidogyne incognita behavior and associated host responses following root penetration were studied in resistant (cv. Moapa 69) and susceptible (cv. Lahontan) alfalfa. Ten-day-old seedlings of alfalfa cultivars were inoculated with second-stage juveniles (J2) and harvested 12, 24, 48, and 72 hours and 7, 14, and 21 days later. Both cultivars supported similar root penetration and initial J2 migration. By 72 hours after inoculation the majority of J2 were amassed inside the vascular cylinder in roots of susceptible Lahontan, while J2 had not entered the vascular cylinder of resistant Moapa 69 and remained clumped at the root apex. Nematode development progressed normally in Lahontan, but J2 were not observed in Moapa 69 after day 7. The greatest differences between RNA translation products isolated from inoculated and uninoculated roots of Lahanton occurred 72 hours after inoculation. Only minor differences in gene expression were observed between inoculated and uninoculated Moapa 69 roots at 72 hours. Comparison of translation products from inoculated versus mechanically wounded Lahontan roots revealed products that were specific to or enhanced in nematode-infected plants. Moapa 69 appears to possess a type of resistance to M. incognita that does not depend on a conventional hypersensitive response.     

Efficacy of Oxamyl Coated on Alfalfa Seed with a Polymer Sticker in Pratylenchus and Meloidogyne Infested Soils

A polymer sticker was used as a coating in which oxamyl was applied to seeds of alfalfa cultivar Saranac for the control of Pratylenchus penetrans and Meloidogyne hapla. The sticker, diluted 1:1 (sticker:water) to 1:5, delayed seedling emergence during the first 4 days after planting. By day 13, however, emergence from all sticker treatments was comparable to the control. Shoot growth of seedlings at day 21 was less than that of the control only from seeds coated with a 1:1 dilution; root growth and nodulation were not affected. Sticker-coated seeds absorbed 30-58% as much water in 3.5 hours as was absorbed by uncoated seeds. Oxamyl concentrations of 40-160 mg/ml in a 1:5 sticker : water mixture had no adverse affect on seedling emergence, growth, and nodulation over 3 weeks. Oxamyl at 160 mg/ml was more effective against P. penetrans than M. hapla. Growth of alfalfa in P. penetrans-infested soil was greater than that of the control in each sampling for 11 weeks. The reduction of number of P. penetrans in soil and roots moderated slowly over 11 weeks from 90% to 60%. Shoot and root growth of alfalfa from oxamyl-coated seed in M. hapla-infested soil were greater than those of the control for 7 and 11 weeks, respectively. The reduction in the number of M. hapla in the soil and roots changed from 80% at 7 weeks to 15% at 11 weeks.     

Histopathology of Pea Roots Axenically Infected by Pratylenchus penetrans

The histological changes in pea roots axenically infected by Pratylenchus penetrans were studied and described. Roots of pea seedlings growing aseptically on the surface of nutrient agar slants were inoculated with axenized nematodes. Six hours after inoculation most of the nematodes introduced were probing the root epidermis, but none had completely entered though a few were observed with their anterior section already in the root. Most of the nematodes penetrated the roots after 12 hr inoculation. From 18 to 24 hr after inoculation the nematodes were mostly in the mid-cortex. Invaded regions of the cortex often showed orange discoloration. As incubation continued, the number of nematodes in these roots increased, and feeding and reproductive activities extended deeper into the cortex. These activities resulted in extensive breakdown of the cortex. No nematodes were observed within the stele of infected roots; however, the endodermis of infected roots stained dark-brown. Gravid female nematodes probed the root endodermis and some endodermal cells appeared to collapse after prolonged probing by the nematode. All stages in the life cycle of the nematode were observed in infected roots; the female to male ratio inside the root was about 5:1.     

Suppression of Alfalfa Growth by Concommitant Populations of Pratylenchus penetrans and two Fusarium species

Growth of alfalfa (Medicago sativa cv. Vernal) seedlings was compared after inoculation with combinations of either Pratylenchus penetrans and Fusarium soloni or P. penetrans and F. oxysporum f. sp. medicaginis. A synergistic disease interaction occurred in alfalfa when F. oxysporum and P. penetrans were added simultaneously to the soil. Alfalfa growth was suppressed at all inoculum levels of P. penetrans and F. oxysporum, but not with F. solani. Seedlings inoculated with the nematode alone gave lower yields than when inoculated with either Fusarium species alone. Fusarium oxysporum, but not F. solani, was pathogenic to alfalfa under similar experimental conditions. Fusarium oxysporum did not alter the populations of P. penetrans in alfalfa roots, whereas the presence of F. solani was associated with a diminished number of P. penetrans in the roots.     

Infection of Seedlings of Alfalfa and Red Clover by Concomitant Populations of Meloidogyne incognita and Pratylenchus penetrans

Invasion of 2-day-old seedlings of ''Buffalo'' alfalfa and ''Kenland'' red clover by larvae of M. incognita and adults of P. penetrans, during 1-3 day periods of incubation at 24 C, was investigated in 50-mm petri dishes on 1% agar. Penetration by both nematodes increased arithmetically with increased numbers in inocula. P. penetrans invaded alfalfa more readily than red cover, but M. incognita invaded red clover more readily than alfalfa. Both nematodes inhibited root-elongation of alfalfa more than that of red clover. In combinations of 10 and 50 of both nematodes, invasion of both plants by both nematodes was the same as for each nematode alone. Penetration by M. incognita into alfalfa, but not into red clover, was significantly reduced when combinations of 50 M. incognita and 200 P. penetrans were inoculated simultaneously. In the presence of large numbers of entrant P. penetrans in both plants, penetration by M. incognita was highly significantly reduced. Penetration by P. penetrans was unaffected in the reciprocal situations.     

Effect of Meloidogyne incognita on Reproduction of Pratylenchus penetrans in Red Clover and Alfalfa

Roots of seedlings of red clover and alfalfa growing on 10⁻¹ Hoagland and Arnon solution agar were inoculated with various combinations of Meloidogyne incognita and Pratylenchus penetrans. Egg-laying by P. penetrans decreased as the number of nematodes, the ratio of entrant M. incognita to entrant P. penetrans, and the priority of invasion of roots by M. incognita increased. Embryogeny and hatching of eggs of P. penetrans, and development of larvae of M. incognita, were not affected. In red clover, the greatest red uction occurred when there were 65 entrant nematodes, the ratio of M. incognita:P. penetrans was 4:1 and M. incognita was inoculated four days prior to P. penetrans. In alfalfa, the less-favorable host for both nematodes, the greatest reduction occurred when there were 45 entrant nematodes, the ratio of M. incognita:P. penetrans was 2:1, and M. incognita was inoculated 4 days prior to P. penetrans.     

Protein Changes in the Vascular Wilt Disease of Lucerne caused by Verticillium albo-atrum R. et B

Field trials confirmed that, whereas the Du Puits variety oflucerne was highly susceptible to Verticillium albo-atrum, ‘Gaetula’hybrids showed slight resistance and certain ‘Swedish’material was moderately resistant. Symptom-free ‘Gaetula’and Swedish plants were used to produce uninfected clonal materialby root propagation. Comparisons of the proteins of such plants with seed-grown DuPuits produced no evidence of a correlation between proteinpattern before inoculation and differences in susceptibilityand resistance. Disc electrophoretic comparisons of infected host material showedthat the pathogen gradually induced a change in the shoot proteinsof Du Puits whereas in the resistant host, after an initialmarked change, there was a development towards normality. Bythe terminal stages of wilting there was a noticeable fall inthe numbers of proteins in both roots and shoots of Du Puits,a finding confirmed both by double diffusion and immunoelectrophoresis.At the same time there was good evidence for a slight increasein protein bands in the resistant material after inoculation. After feeding tritium-labelled leucine to infected Du Puitsplants, a small protein component of fungal origin was detectedin the host. Labelled components from both healthy and infectedDu Puits plants were separated on Sephadex G.25 and compared.     

Effect of Single and Interplantings on Pathogenicity of Pratyenchus penetrans and P. neglectus to Alfalfa and Crested Wheatgrass

Alfalfa is a host of Pratylenchus penetrans and P. neglectus, whereas crested wheatgrass is a host of P. neglectus but not of P. penetrans. In a 120-day greenhouse experiment at 24 ñ 3 C, P. neglectus inhibited the growth of ''Lahontan'' alfalfa and ''Fairway'' crested wheatgrass. There were no differences in persistence and plant growth of alfalfa and crested wheatgrass, or reproduction of P. neglectus, in single plantings of alfalfa (AO) or crested wheatgrass (CWO), or in interplanted alfalfa and crested wheatgrass (ACW) treatments. On alfalfa, P. penetrans inhibited growth and reproduced more than did P. neglectus. Inhibition of plant growth and reproduction of P. penetrans was greater on alfalfa in AO than in ACW treatments. Pratylenchus penetrans did not reproduce on crested wheatgrass, but inhibited growth of crested wheatgrass in interplanted treatments and was avirulent in single planted treatments. Results were similar in a controlled growth chamber experiment at 15, 20, 25, and 30 C. Both nematode species inhibited alfalfa growth at all temperatures, and P. penetrans was more virulent than was P. neglectus to alfalfa at all temperatures and treatments. Plant growth inhibition and reproduction of P. penetrans on alfalfa in single and interplanted treatments were similar at 15-20 C, but were greater in single than in interplanted treatments at 25-30 C. Pratylenchus penetrans was avirulent to crested wheatgrass in the single planted treatments at all temperatures, but inhibited growth of crested wheatgrass in interplanted treatments at 20-30 C. Plant growth and reproduction of P. neglectus on crested wheatgrass was similar in single and interplanted treatments at 20-30 C and 15-30 C, respectively.     

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.     

Penetration and Development of Meloidogyne hapla in Resistant and Susceptible Alfalfa Under Differing Temperatures

Studies were conducted to examine under differing temperatures (12, 16, 20, 24, 28, and 32 C) the penetration anti development of Meloidogyne hapla in resistant lines ''298'' and ''Nev. Syn XX'', and susceptible ''Lahontan'' and ''Ranger'' hardy-type alfalfas. The results indicated that resistance to M. hapla was similar to that previously described for M. incognita in nonhardy alfalfa. Although initial penetration in resistant seedlings was similar to that of susceptible seedlings, nematode larvae failed to establish and develop in root tissues and nematode numbers subsequently declined. In susceptible seedlings, nematode development proceeded rapidly, and egg production began after 5 weeks. Temperature had little influence on the nematode development except to slow the response at the lower temperatures. Other studies were conducted to verify a previously reported immune (no penetration) reaction to M. hapla by the ''Vernal'' selection ''M-4''. When compared to the resistant (penetration without nematode development) Vernal selection ''M-9'' under differing temperatures (20, 24, 28, and 32 C), each selection was equally penetrated by M. hapla but at a lower level than in susceptible Ranger cuttings. Generally, no root galling was observed in either M-4 or M-9; however, very slight galling was found 35 days after inoculation on about 50% of these cuttings when grown at 32 C.     

Control of Pratylenchus penetrans and Meloidogyne hapla and Yield Response of Alfalfa Due to Oxamyl Seed Treatments

Alfalfa (Medicago sativa L. cv. Saranac) seed were soaked for 20 minutes in water, acetone, or methanol containing 10 or 50 mg/ml of oxamyl (Vydate L) or coated with a 2% aqueous cellulose solution containing the same amounts of oxamyl. Seed were analyzed for oxamyl by HPLC immediately after treatment and after 9 and 26 months of storage. Oxamyl content of alfalfa seed did not decline after 26 months of storage. The effects of seed treatment on growth of alfalfa and nematode control were examined using soils infested with Pratylenchus penetrans and Meloidogyne hapla. Germination was not affected by any of the seed treatments. Twenty-one days after sowing, the total growth of alfalfa seedlings grown from seed treated with 50 mg/ml of oxamyl in P. penetrans-infested soils had increased by 62% over controls. Nodulation per pot increased by as much as 267%, and the densities of P. penetrans per gram of root were reduced by as much as 73% compared to control plants. In M. hapla-infested soils, increases in plant growth (32%) and nodulation (71%) also occurred with oxamyl-treated seeds. Root gall reduction (86%) was also substantial due to oxamyl seed treatment.     

Histopathology of Wheat Seedling Roots Infected with Pythium arrhenomanes

Two-day-old wheat seedlings were placed on the edge of a P. arrhenomanes culture for 3 h at 25°C, and transferred into test tubes (18 mm dia.) containing glass beads and 1 ml of sterile water. Roots were sampled every 6 or 12 h for 84 h, and observed with the scanning electron microscope, or serial sectioned for light microscopy. Roots were colonized extensively in the region of root hair formation near the tip within 30 h after inoculation. Extensive penetration occurred 0.1—2 mm behind the root tip, with hyphae breaching the endodermis and gaining entry into the stele. Behind this area, hyphae remained limited to the outer cortical cells, or did not penetrate at all. Hyphae grew intracellularly, and became irregularly inflated inside cells. In most cases, hyphae penetrated the roots directly through the epidermis with appressoria being formed, or through breaks on the surface. However, histological evidence suggested chemical action taking place both outside and inside the cells.     

Inheritance of Resistance to Pratylenchus penetrans in Alfalfa

Fifty-two alfalfa (Medicago sativa L.) clones, randomly selected from the cultivar Baker and the experimental line MNGRN-4, were evaluated for resistance (based on nematode reproduction) to Pratylenchus penetrans in growth chamber tests (25 C). Twenty-five clones, representing the range of nematodes and eggs per plant, were selected and retested. Four moderately resistant and two susceptible alfalfa clones were identified. Inheritance of resistance to P. penetrans was studied in these six clones using a diallel mating design. The S₁, Fl, and reciprocal progenies differed for numbers of nematodes and eggs per g dry root and for shoot and root weights (P < 0.05). Resistance, measured as numbers of nematodes in roots, was correlated between parental clones and their S₁ families (r = 0.94), parental clones and their half-sib families (r = 0.81), and S₁ and half-sib families (r = 0.88). General combining ability (GCA) effects were significant for nematode resistance traits. Both GCA and specific combining ability (SCA) effects were significant for plant size traits, but SCA was more important than GCA in predicting progeny plant size. Reciprocal effects were significant for both nematode resistance and plant size traits, which may slow selection progress in long-term selection programs. However, the GCA effects are large enough that breeding procedures that capitalize on additive effects should be effective in developing alfalfa cultivars with resistance to P. penetrans.     

Response of Peach Seedlings to Infection by the Root Lesion Nematode Pratylenchus penetrans under Controlled Conditions

Twenty-one open pollinated populations of peach rootstock seedlings were evaluated for their response to infection by the root lesion nematode, Pratylenchus penetrans, over a period of 98 days. Nematode-infected peach seedling populations were shorter in plant height and had less shoot weight but more dry root weight than nematode-free controls. Rootstock differences were demonstrated for nematode increase over the 98-day period, and average total numbers of nematodes in soil and roots. Rootstocks were classified into three groups differing in total nematode population levels, ratio of nematode increase, and the number of nematodes per root. The heritable nature of rootstock response to nematodes was evident. Rootstocks showing the lowest response to nematode infection included Tzim Pee Tao, Rutgers Red Leaf, and two progenies of a cross of these two rootstocks.     

Population Dynamics of Pratylenchus penetrans,Paratylenchus sp., and Criconemella xenoplax on Western Oregon Peppermint

Endoparasitic nematode populations are usually measured separately for soil and roots without a determination of the quantitative relation between soil and root population components. In this study, Pratylenchus penetrans populations in peppermint soil, roots, and rhizomes were expressed as the density within a standardized core consisting of 500 g dry soil plus the roots and rhizomes contained therein. Populations of Paratylenchus sp. and Criconemella xenoplax in 500 g dry soil were also determined, thus measuring the total plant-parasitic nematode population associated with the plant. Mean wet root weight per standard core peaked in spring and again in late summer and was lowest early in the growing season and in early fall. Pratylenchus penetrans populations peaked 4 to 6 weeks after root weight peaks. The percentage of the total population in roots reached 70% to 90% in early April, decreased to 20% to 40% in August, and returned to higher percentages during the winter. Rhizomes never contained more than a minor proportion of the population. Mean Paratylenchus sp. populations increased through spring and peaked in late August. Mean C. xenoplax populations fluctuated, peaking in August or September. Populations of all parasitic species were lowest during winter. Evaluation using the standard core method permits assessment of the total P. penetrans population associated with the plant and of changes in root weight as well as the seasonal distribution of P. penetrans.