Host Tests to Differentiate Meloidogyne chitwoodi Races 1 and 2 and M. hapla

The reproductive factor (R = final egg density at 55 days ÷ 5,000, initial egg density) of Meloidogyne chitwoodi race 2 (alfalfa race) on 46 crop cultivars ranged from 0 to 130. The reproductive efficiency of M. chitwoodi race 1 (non-alfalfa race) and M. chitwoodi race 2 was compared on selected crop cultivars. The basic difference between the two races lay in their differential reproduction on Thor alfalfa and Red Cored Chantenay carrot. M. chitwoodi race 2 reproduced on alfalfa but not on carrot. Conversely, alfalfa was a poor host and carrots were suitable for M. chitwoodi race 1. Based on host responses to M. chitwoodi races and M. hapla, a new differential host test was proposed to distinguish the common root-knot nematode species of the Pacific Northwest.     

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

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

Suppression of Meloidogyne chitwoodi with Sudangrass Cultivars as Green Manure

Meloidogyne chitwoodi race 1 reproduced on Piper sudangrass (Sorghum bicolor (L.) Moench), 332 (sudangrass hybrid), and P855F and P877F (sorghum-sudangrass hybrids), but failed to reproduce efficiently on Trudan 8, Trudex 9 (sudangrass hybrids), and Sordan 79, SS-222, and Bravo II (sorghum-sudangrass hybrids). Meloidogyne chitwoodi race 2 behaved similarly and reproduced more efficiently on Piper, P855F, and P877F than on Trudan 8, Trudex 9, or Sordan 79. The mean reproductive factor for M. chitwoodi races on the poorer hosts ranged from <0.1 to 0.9 under greenhouse and field conditions. Meloidogyne hapla failed to reproduce on any of the cultivars tested. In the laboratory, leaves of each cultivar chopped and incorporated as green manure reduced the M. chitwoodi population in infested soil more than unamended or wheat green manure treatments. Trudan 8, although limited to the zone of incorporation, protected this zone from colonization of upward migrating second stage juveniles (J2) for up to 6 weeks. Leaves of Trudan 8 but not roots were effective against M. chitwoodi, and J2 appeared to be more sensitive than egg masses. Trudan 8 and Sordan 79 as green manure reduced M. chitwoodi in bucket microplots under field conditions.     

Effect of the Mi Gene in Tomato on Reproductive Factors of Meloidogyne chitwoodi and M. hapla

The effect of the Mi gene on the reproductive factor of Meloidogyne chitwoodi and M. hapla, major nematode pests of potato, was measured on nearly isogenic tomato lines differing in presence or absence of the Mi gene. The Mi allele controlled resistance to reproduction of race 1 of M. chitwoodi and to one of two isolates of race 2. No resistance to race 3 of M. chitwoodi or to M. hapla was found. Variability in response to isolates of race 2 may reflect diversity of virulence genotypes heretofore undetected. Resistance to race 1 of M. chitwoodi could be useful in potato if the Mi gene were functional following transferral by gene insertion technology into potato. Since the Mi gene is not superior to RMc₁ derived from Solarium bulbocastanum, the transferral by protoplast fusion appears to offer no advantage.     

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

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

Influence of Meloidogyne chitwoodi and M. hapla on Wheat Growth

Meloidogyne chitwoodi reduced the growth of winter wheat ''Nugaines'' directly in relation to nematode density in the greenhouse, The relationship between top dry weight and initial nematode density suggests a tolerance limit of Nugaines wheat to M. chitwoodi of between 0.03 and 0.18 eggs/cm³ of soil; the value for relative minimum plant top weight was 0.45 g and 0.75 g, respectively. Growth of wheat in field microplots containing four population densities (0.003, 0.05, 0.75 and 9 eggs/cm³ soil) was not affected significantly at any inoculum level compared to controls during September to July, However, suppression of head weights of ''Fielder'' spring wheat grown May-July occurred in microplots initially infested with 0.75 and 9 eggs/cm³ soil. Reproduction (Pf/Pi) was poorer at these two inoculum levels as compared to the lower densities. In another greenhouse experiment, roots of wheat cultivars Fielder, ''Fieldwin,'' ''Gaines,'' ''Hyslop,'' and Nugaines became infected by M. chitwoodi, but not by M. hapla. Reproduction of M. chitwoodi was less on Gaines and Nugaines than on Fielder, Fieldwin, or Hyslop.     

Suppression of Root-knot Nematode Populations with Selected Rapeseed Cultivars as Green Manure

Meloidogyne chitwoodi races 1 and 2 and M. hapla reproduced on 12 cultivars of Brassica napus and two cultivars of B. campestris. The mean reproductive factors (Rf), Rf = Pf at 55 days ÷ 5,000, for the three nematodes were 8.3, 2.2, and 14.3, respectively. All three nematodes reproduced more efficiently (P < 0.05) on B. campestris than on B. napus. Amending M. chitwoodi-infested soil in plastic bags with chopped shoots of Jupiter rapeseed reduced the nematode population more (P < 0.05) than amendment with wheat shoots. Incorporating Jupiter shoots to soil heavily infested with M. chitwoodi in microplots reduced the nematode population more (P < 0.05) than fallow or corn shoot treatments. The greatest reduction in nematode population density was attained by cropping rapeseed for 2 months and incorporating it into the soil as a green manure.     

Effects of Cropping Sequences on Population Densities of Meloidogyne hapla and Carrot Yield in Organic Soil

The influence of various cropping sequences on population densities of Meloidogyne hapla and carrot yield was studied in organic soil under microplot-and field conditions. Spinach, radish, barley, oat, and wheat were poor or nonhosts for M. hapla. Population densities of M. hapla were maintained or increased on cabbage, celery, lettuce, leek, marigold, and potato. Marketable percent-age and root weight of carrots were greater following spinach, oat, radish, and fallow-onion than those following two crops of onion or carrot in microplots. Under field conditions, the carrot-onion-oat-carrot cropping sequence decreased M. hapla population densities and provided a 282% increase in marketable yield of carrot compared to a carrot monoculture. Two consecutive years of onion increased M. hapla population densities causing severe root galling and a 50% yield loss in the following crop of carrot. Based on root-gall indices, carrots could be grown economically for 2 years following radish, spinach, and oat, but not following onion and carrot without the use of nematicides.     

Effect of Soil Temperature on Reproduction of Meloidogyne chitwoodi and M. hapla Alone and in Combination on Potato and M. chitwoodi on Rotation Plants

Meloidogyne chitwoodi developed and reproduced more rapidly than M. hapla in potato roots at 15, 20, or 25 C when both species of nematodes were inoculated simultaneously at 250 or 1,000 juveniles of each. At 30 C significantly more M. hapla than M. chitwoodi females were found at the lower inoculum level after 41 days. More M. chitwoodi than M. hapla juveniles were extracted from soil at 15, 20, and 25 C, but only at the lower inoculum level at 30 C. Potato was considered a more suitable host for M. chitwoodi than M. hapla because of M. chitwoodi''s greater reproduction at 15, 20, and 25 C. Corn and wheat cultivars tested supported M. chitwoodi reproduction at temperatures of 10, 15, 20, and 25 C, but fewest eggs were produced on these plants at 20 C. Temperatures of 10 to 25 C had little influence on the low reproduction of M. chitwoodi on four alfalfa cultivars. M. chitwoodi reproduced on the alfalfa entry Mn PL9HF.     

Incidence and Distinguishing Characteristics of Meloidogyne chitwoodi and M. hapla in Potato from the Northwestern United States

From September 1980 to June 1981, a survey was conducted in the major potato growing regions of northern California, Idaho, Nevada, Oregon. and Washington to determine the distribution of Meloidogyne chitwoodi and other Meloidogyne spp. Meloidogyne chitwoodi and M. hapla were the only root-knot nematode species detected parasitizing potato in all the states surveyed. Meloidogyne chitwoodi occurred alone in 83% of the samples and M. hapla in 11%, with 6% of all samples containing both species. The greater incidence of M. chitwoodi, as compared to M. hapla, may be due to the cool growing season encountered in 1980 (which favored M. chitwoodi but not M. hapla) and to the increased acreage of small grains (which are good hosts for M. chitwoodi but not M. hapla) planted in rotation with potato. Differentiation between these two species can be determined by a differential host test, perineal patterns of mature females, and shape of the tail tip amt of the tail hypodermal terminus of L₂ juveniles.     

Effects of Temperature and Root Leachates on Embryogenic Development and Hatching of Meloidogyne chitwoodi and M. hapla

At 20 C the duration of the embryogenic development of Meloiclogyne chitwoodi and M. hapla was about 20 days. At 10 C the embryogenic development was 82-84 days for M. chitwoodi and 95-97 days for M. hapla. The effect of distilled water and root leachates of potato cv. Russet Burbank, tomato cv. Columbian, and wheat cv. Hyslop on the hatching of eggs of the two root-knot nematode species was investigated at 4, 7, 10, 15, 20, and 25 C (± 1 C). Cumulative egg taatch was no greater in root leachates titan in distilled water, but temperature did significantly affect egg hatch (P = 0.05). Less than 1% of the eggs of both nematode species hatched at 4 C. The percent cumulative hatch at 10 C was significantly less (P = 0.05) than at higher temperatures for both nematodes and significantly more (P = 0.05) M. chitwoodi eggs hatched than did M. hapla eggs. At 15 G the percent cumulative hatch of both species was significantly lower (P = 0.05) than that at 20 and 25 C. The percent cumulative egg hatch of two species did not differ at 25 C, but was higher (P = 0.05) at 25 C than at 20 C. At 7 C the emergence of M. chitwoodi juveniles was about seven times (P = 0.01) greater than that of M. hapla in distilled water.     

Description and SEM Observations of Meloidogyne chitwoodi n. sp. (Meloidogynidae), a Root-knot Nematode on Potato in the Pacific Northwest

Meloidogyne chitwoodi n. sp. is described and illustrated from potato (Solanum tuberosum) originally collected from Quincy, Washington, USA. This new species resembles M. hapla, but its perineal pattern is basically round to oval with distinctive and broken, curled, or twisted striae around and above the anal area. The vulva is in a sunken area devoid of striae. Vesicles or vesicle-like structures are present in the median bulb of females. The larva tail, being short and blunt with a hyaline tail terminal having little or no taper to its rounded terminus, is distinctively different from M. hapla. SEM observations revealed the nature of the perineal pattern and details of the head of larvae and males, and showed the spicules to have dentate tips ventrally. Hosts for M. chitwoodi n. sp. include potato, tomato, corn, and wheat but not strawberry, pepper, or peanut. The latter three crops are excellent hosts for M. hapla. The known distribntion of this new root-knot species presently involves certain areas of Idaho, Washington, and Oregon. The common name "Columbia root-knot nematode" is proposed for M. chitwoodi n. sp.     

Differential Reaction of Alfalfa Cultivars to Meloidogyne hapla and M. chitwoodi Populations

Meloidogyne hapla reproduced and suppressed growth (P < 0.05) of susceptible Lahontan and Moapa alfalfa at 15, 20, and 25 C. At 30 C, resistant Nevada Syn XX lost resistance to M. hapla. M. hapla invaded and reproduced on Rhizobium meliloti nodules of Lahontan and Moapa, inducing giant cell formation and structural disorder of vascular bundles of nodules without disrupting bacteroids. At 15, 20, and 25 C a M. chitwoodi population from Utah reproduced on Lahontan, Moapa, and Nevada Syn XX alfalfa, suppressing growth (P < 0.05). Final densities of the Utah M. chitwoodi population were greater (P < 0.05) than those of Idaho and Washington State populations on Lahontan at 15 and 25 C and on Nevada Syn XX at 15 C, but were less consistent and smaller (P < 0.05) than those of M. hapla on Lahontan and Moapa at 20 and 25 C. Inconsistent reproduction of the Utah M. chitwoodi population on alfalfa suggests the possible existence of nematode strains revealed by variability in alfalfa resistance. No reproduction or inconsistent final nematode population densities with no damage were observed on Lahontan, Moapa, and Nevada Syn XX plants grown in soil infested with Idaho and Washington State M. chitwoodi populations.     

Relationship of Resistance to Meloidogyne chitwoodi (race 2) and M. hapla in Alfalfa

In the Pacific Northwest, alfalfa (Medicago sativa) is host to two species of root-knot nematodes, including race 2 of the Columbia root-knot nematode (Meloidogyne chitwoodi) and the northern root-knot nematode (Meloidogyne hapla). In addition to the damage caused to alfalfa itself by M. hapla, alfalfa’s host status to both species leaves large numbers of nematodes available to damage rotation crops, of which potato is the most important. A nematode-resistant alfalfa germplasm release, W12SR2W1, was challenged with both nematode species, to determine the correlation, if any, of resistance to nematode reproduction. Thirty genotypes were screened in replicated tests with M. chitwoodi race 2 or M. hapla, and the reproductive factor (RF) was calculated. The distribution of natural log-transformed RF values was skewed for both nematode species, but more particularly for M. chitwoodi race 2, where more than half the genotypes screened were non-hosts. Approximately 30 percent of genotypes were non-hosts or very poor hosts of M. hapla, but RF values for M. hapla on susceptible genotypes were generally much higher than RF values for genotypes susceptible to M. chitwoodi race 2. The Spearman rank correlation was positive (0.52) and significant (p-value = 0.003), indicating there is some relationship between resistance to these two species of root-knot nematode in alfalfa. However the relationship is not strong enough to suggest genetic loci for resistance are identical, or closely linked. Breeding for resistance or immunity will require screening with each species separately, or with different DNA markers if marker-assisted breeding is pursued. A number of genotypes were identified which are non-hosts to both species. These plants will be intercrossed to develop a non-host germplasm.     

Vertical Migration of Meloidogyne chitwoodi and M. hapla under Controlled Temperature

Migratory ability of second-stage juveniles (J2) of two Meloidogyne chitwoodi races and a M. hapla population were compared in soil-filled columns at 12, 18, and 24 C. J2 of all populations migrated farthest at 18 C and least at 12 C. Nematode survival was significantly reduced (P = 0.05) at 24 C.M. chitwoodi J2 migrated further and in greater numbers than M. hapla J2 at all temperatures. A comparison with and without a host plant demonstrated no preferential migration toward the plant. Water percolation through the migration columns stimulated upward migration.     

A PCR Assay to Identify and Distinguish Single Juveniles of Meloidogyne hapla and M. chitwoodi

Random amplified polymorphic DNA (RAPD) bands that distinguish Meloidogyne hapla and M. chitwoodi from each other, and from other root-knot nematode species, were identified using a series of random octamer primers. The species-specific amplified DNA fragments were cloned and sequenced, and then the sequences were used to design 20-mer primer pairs that specifically amplified a DNA fragment from each species. Using the primer pairs, successful amplifications from single juveniles were readily attained. A mixture of four primers in a single PCR reaction mixture was shown to identify single juveniles of M. hapla and M. chitwoodi. To confirm specificity, the primers were used to amplify DNA from several isolates of M. hapla that originated from different crops and locations in North America and also from isolates of M. chitwoodi that differed in host range. In characterizing the M. hapla isolates, it was noted that there was a mitochondrial DNA polymorphism among isolates for cleavage by the restriction endonuclease DraI.     

Host Suitability and Response of Asparagus Cultivars to Meloidogyne Species and Races

The host-parasite relationships of asparagus and Meloidogyne spp. were examined under greenhouse and microplot conditions. Meloidogyne species and races differed greatly in their ability to reproduce on asparagus seedlings. Meloidogyne hapla generally failed to reproduce, and M. javanica, M. arenaria race 1, and M. incognita race 3 reproduced poorly, with a reproduction factor (Rf = final population/initial population) usually < 1.0. Only M. arenaria race 2 and M. incognita races 1 and 4 reproduced consistently on all asparagus cultivars tested (Rf typically 1-11). No effect of M. incognita race 4 on host growth was detected. Meloidogyne arenaria race 2 and M. incognita race 1 had slight negative effects (5-10%) on plant and root growth.     

Host Suitability of Twelve Leguminosae Species to Populations of Meloidogyne hapla and M. chitwoodi

Legumes of the genera Astragalus (milkvetch), Coronilla (crownvetch), Lathyrus (pea vine), Lotus (birdsfoot trefoil), Medicago (alfalfa), Melilotus (clover), Trifolium (clover), and Vicia (common vetch) were inoculated with a population of Melaidogyne chitwoodi from Utah or with one of three M. hapla populations from California, Utah, and Wyoming.Thirty-nine percent to 86% of alfalfa (M. scutellata) and 10% to 55% of red clover (T. pratense) plants survived inoculation with the nematode populations at a greenhouse temperature of 24 ± 3°C. All plants of the other legume species survived all nematode populations, except 4% of the white clover (T. repens) plants inoculated with the California M. hapla population. Entries were usually more susceptible to the M. hapla populations than to M. chitwoodi. Galling of host roots differed between nematode populations and species. Root-galling indices (1 = none, 6 = severely galled) ranged from 1 on pea vine inoculated with the California population of M. hapla to 6 on yellow sweet clover inoculated with the Wyoming population of M. hapla. The nematode reproductive factor (Rf = final nematode population/initial nematode population) ranged from 0 for all nematode populations on pea vine to 35 for the Wyoming population of M. hapla on alfalfa (M. sativa).     

Relative Damage Functions and Reproductive Potentials of Meloidogyne arenaria and M. hapla on Peanut

The reproductive potential and damage functions for Meloidogyne hapla and M. arenaria race 1 on Virginia-type peanuts (Arachis hypogaea cv. Florigiant) were determined over 2 years in microplot experiments in North Carolina. Peanut yield suppression and damage to pods as a result of galling were greatest in response to M. arenaria (P = 0.01). Damage functions for the two species were adequately described by the quadratic models: yield (g/plot) = 398 - 17.1 (log₁₀[Pi + 1]) - 17.0(log₁₀[Pi + 1])²; (R² = 0.83, P = 0.0001) for M. arenaria; and yield = 388 - 10.2(log₁₀[Pi + 1]) - 7.5(log₁₀[Pi + 1])², (R² = 0.30, P = 0.0001) for M. hapla. Both species caused galling on pods, but this was more severe in response to M. arenaria. Reproduction of M. arenaria race 1 was greater than M. hapla on peanut, which accounts in part for the more severe pod galling. Peanut was an excellent host for both M. arenaria race 1 and for M. hapla, but reproduction by M. hapla was more variable.     

Identification of Sources of Resistance to Four Species of Root-knot Nematodes in Tobacco

Resistance to the southern root-knot nematode, Meloidogyne incognita races 1 and 3, has been identified, incorporated, and deployed into commercial cultivars of tobacco, Nicotiana tabacum. Cultivars with resistance to other economically important root-knot nematode species attacking tobacco, M. arenaria, M. hapla, M. javanica, and other host-specific races of M. incognita, are not available in the United States. Twenty-eight tobacco genotypes of diverse origin and two standard cultivars, NC 2326 (susceptible) and Speight G 28 (resistant to M. incognita races 1 and 3), were screened for resistance to eight root-knot nematode populations of North Carolina origin. Based on root gall indices at 8 to 12 weeks after inoculation, all genotypes except NC 2326 and Okinawa were resistant to M. arenaria race 1, and races 1 and 3 of M. incognita. Except for slight root galling, genotypes resistant to M. arenaria race 1 responded similarly to races 1 and 3 of M. incognita. All genotypes except NC 2326, Okinawa, and Speight G 28 showed resistance to M. javanica. Okinawa, while supporting lower reproduction of M. javanica than NC 2326, was rated as moderately susceptible. Tobacco breeding lines 81-R-617A, 81-RL- 2K, SA 1213, SA 1214, SA 1223, and SA 1224 were resistant to M. arenaria race 2, and thus may be used as sources of resistance to this pathogen. No resistance to M. hapla and only moderate resistance to races 2 and 4 of M. incognita were found in any of the tobacco genotypes. Under natural field infestations of M. arenaria race 2, nematode development on resistant tobacco breeding lines 81-RL-2K, SA 1214, and SA 1215 was similar to a susceptible cultivar with some nematicide treatments; however, quantity and quality of yield were inferior compared to K 326 plus nematicides.