Susceptibility of Several Common Subtropical Weeds to Meloidogyne arenaria,M. incognita,and M. javanica

Experiments were conducted in the greenhouse to assess root galling and egg production of three root-knot nematode species, Meloidogyne arenaria, M. incognita, and M. javanica, on several weeds common to Florida agricultural land. Weeds evaluated were Amaranthus retroflexus (redroot pigweed), Cyperus esculentus (yellow nutsedge), Eleusine indica (goosegrass), Portulaca oleracea (common purslane), and Solanum americanum (American black nightshade). Additionally, although it is recommended as a cover crop in southern regions of the U.S., Aeschynomene americana (American jointvetch) was evaluated as a weed following the detection of root galling in a heavy volunteer infestation of an experimental field in southeastern Florida. Weeds were propagated from seed and inoculated with 1000 nematode eggs when plants reached the two true-leaf stage. Tomato (Solanum lycopersicum ‘Rutgers’) was included as a positive control. Aeschynomene americana and P. oleracea roots supported the highest number of juveniles (J₂) and had the highest number of eggs/g of root for all three species of Meloidogyne tested. However, though P. oleracea supported very high root levels of the three nematode species tested, its fleshy roots did not exhibit severe gall symptoms. Low levels of apparent galling, combined with high egg production, increase the potential for P. oleracea to support populations of these three species of root-knot nematodes to a degree that may not be appropriately recognized. This research quantifies the impact of P. oleracea as a host for M. arenaria, M. incognita, and M. javanica compared to several other important weeds commonly found in Florida agricultural production, and the potential for A. americana to serve as an important weed host of the three species of root-knot nematode tested in southern regions of Florida.     

Description of a Unique,Complex Feeding Socket Caused by the Putative Primitive Root-Knot Nematode,Meloidogyne kikuyensis

Meloidogyne kikuyensis produces unique galls that form on one side of the root resembling nitrogen-fixing nodules that are produced on legumes in response to infection by Rhizobium and related bacteria. The gall caused by this root-knot nematode is made up of a complex feeding socket composed of several giant cells that are ramified with xylem vessels extending perpendicular from the vascular cylinder. The anterior portion of the second-stage juvenile, which develops into an adult, plugs into this unique feeding socket. The socket and the surrounding parenchyma together form a gall that is very different in morphology from those typically caused by other species of root-knot nematodes. Even though M. kikuyensis was considered to be a primitive species because of its low chromosome count, the complexity of its feeding site and minor plant damage suggests a more derived systematic position.     

Morphological,Molecular, and Differential-Host Characterization of Meloidogyne floridensis n. sp. (Nematoda: Meloidogynidae), a Root-Knot Nematode Parasitizing Peach in Florida

A root-knot nematode, Meloidogyne floridensis n. sp., is described and illustrated from peach originally collected from Gainesville, Florida. This new species resembles M. incognita, M. christiei, M. graminicola, and M. hispanica, but with LM and SEM observations it differs from these species either by the body length, shape of head, tail and tail terminus of second-stage juveniles, body length and shape of spicules in males, and its distinctive female perineal pattern. This pattern has a high to narrowly rounded arch with coarsely broken and network-like striae in and around anal area, faint lateral lines interrupting transverse striae, a sunken vulva and anus, and large distinct phasmids. Molecular data from ribosomal IGS illustrate that M. floridensis n. sp. is different from the mitotic species M. arenaria, M. incognita, and M. javanica. Data from RAPDs confirm it and suggest that this new species lies in an intermediate phylogenetic position between the previous species and the meiotic species M. hapla, M. fallax, and M. chitwoodi. Differential host tests based on annual crops and on Prunus accessions are reported.     

Morphological and Molecular Characterization of Meloidogyne mayaguensis Isolates from Florida

The discovery of Meloidogyne mayaguensis is confirmed in Florida; this is the first report for the continental United States. Meloidogyne mayaguensis is a virulent species that can reproduce on host cultivars bred for nematode resistance. The perineal patterns of M. mayaguensis isolates from Florida show morphological variability and often are similar to M. incognita. Useful morphological characters for the separation of M. mayaguensis from M. incognita from Florida are the male stylet length values (smaller for M. mayaguensis than M. incognita) and J2 tail length values (greater for M. mayaguensis than M. incognita). Meloidogyne mayaguensis values for these characters overlap with those of M. arenaria and M. javanica from Florida. Enzyme analyses of Florida M. mayaguensis isolates show two major bands (VS1-S1 phenotype) of esterase activity, and one strong malate dehydrogenase band (Rm 1.4) plus two additional weak bands that migrated close together. Their detection requires larger amounts of homogenates from several females. Amplification of two separate regions of mitochondrial DNA resulted in products of a unique size. PCR primers embedded in the COII and 16S genes produced a product size of 705 bp, and amplification of the 63-bp repeat region resulted in a single product of 322 bp. Nucleotide sequence comparison of these mitochondrial products together with sequence from 18S rDNA and ITS1 from the nuclear genome were nearly identical with the corresponding regions from a M. mayaguensis isolate from Mayaguez, Puerto Rico, the type locality of the species. Meloidogyne mayaguensis reproduced on cotton, pepper, tobacco, and watermelon but not on peanut. Preliminary results indicate the M. mayaguensis isolates from Florida can reproduce on tomato containing the Mi gene. Molecular techniques for the identification of M. mayaguensis will be particularly useful in cases of M. mayaguensis populations mixed with M. arenaria, M. incognita, and M. javanica, which are the most economically important root-knot nematode species in Florida, and especially when low (<25) numbers of specimens of these species are recovered from the soil.     

Fergusobia/Fergusonina-induced Shoot Bud Gall Development on Melaleuca quinquenervia

Fergusobia nematodes and Fergusonina flies are mutualists that cause a variety of gall types on myrtaceous plant buds and young leaves. The biology of an isolate of the gall complex was studied in its native range in Australia for possible use in southern Florida as a biological control agent against the invasive broad-leaved paperbark tree, Melaleuca quinquenervia. Timed studies with caged Fergusonina flies on young branches of M. quinquenervia revealed that females are synovigenic with lifetime fecundities of 183 ± 42 (standard error; SE) eggs and longevities of 17 ± 2 days. None of the male flies but all dissected female flies contained parasitic female nematodes (range = 3-15), nematode eggs (12-112), and nematode juveniles (78-1,750). Female flies deposited eggs (34 ± 6; 8-77 per bud) and nematode juveniles (114 ± 15; 44-207 per bud) into bud apices within 15 days. Histological sections of shoot buds suggested that nematodes induce the formation of hypertrophied, uninucleate plant cells prior to fly larval eclosion. Enlarged size, granular cytoplasm, and enlarged nucleus and nucleolus characterized these cells, which appeared similar to those of other species galled by nematodes in the Anguinidae. Observations of ovipositional behavior revealed that female Fergusonina sp. create diagnostic oviposition scars. The presence of these scars may facilitate recognition of host use during specificity screening.     

Host Status of 'SeaIsle 1' Seashore Paspalum (Paspalum vaginatum) to Belonolaimus longicaudatus and Hoplolaimus galeatus

Belonolaimus longicaudatus and Hoplolaimus galeatus are considered among the most damaging pathogens of turfgrasses in Florida. However, the host status of seashore paspalum (Paspalum vaginatum) is unknown. Glasshouse experiments were performed in 2002 and 2003 to determine the tolerance of ''SeaIsle 1'' seashore paspalum to a population of B. longicaudatus and a population of H. galeatus, and to compare to ''Tifdwarf'' bermudagrass for differences. Both nematode species reproduced well on either grass, but only B. longicaudatus consistently reduced root growth as measured by root length. Belonolaimus longicaudatus reduced root growth (P ≤ 0.05) by 35% to 45% at 120 days after inoculation on both grasses. In 2003, higher inoculum levels of H. galeatus reduced root growth (P ≤ 0.05) by 19.4% in seashore paspalum and by 14% in bermudagrass after 60 and 120 days of exposure, respectively. Percentage reductions in root length caused by H. galeatus and B. longicaudatus indicated no differences between grass species, although Tifdwarf bermudagrass supported higher soil population densities of both nematodes than SeaIsle 1 seashore paspalum.     

Descriptions of Meloidogyne camelliae n.sp. and M. querciana n.sp (Nematoda:Meloidogynidae), with SEM and Host-Range Observations

Meloidogyne camelliae n.sp. on camellia (Camellia japonica) from Japan and M. querciana n.sp. on pin oak (Quercus palustris) from Virginia, USA, are described and illustrated. M. camelliae n.sp. is distingnishable from other species of the genus especially by its striking perineal pattern having heavy ropelike striae forming a squarish to rectangular outline with shoulders or projections, appearing sometimes ahnost starlike. M. querciana differs from other species by its characteristic perineal pattern round to oval in outline, sometimes with a low arch, and sunken vulva surrounded by a prominent obovate area devoid of striae. M. querciana shows some relationship to M. ovalis, but differs further fxom the latter by longer larvae, absence of annules on head of larvae, and rarity of males. Examination of specimens of M. camelliae n.sp. and M. querciana n.sp. with the scanning electron microscope confirmed observations made by optical microscopy and revealed diagnostic and other structures in greater detail. In greenhouse host tests, M. camelliae infected camellia heavily, showed moderate infection on oxalis, only a trace infection on tomato, and no infection on five other plants tested; and M. querciana attacked pin oak, red oak, and American chestnut heavily, but did not infect nine other test plants. In another test, pin oak seedlings did not become infected when heavily inoculated with and grown in the presence of two populations of M. incognita incognita and one of M. incognita acrita. The common names "camellia root-knot nematode" and "oak root-knot nematode" are respectively proposed for M. camelliae and M. querciana.     

Use of Cucumis metuliferus as a Rootstock for Melon to Manage Meloidogyne incognita

Root-knot nematode-susceptible melons (Cantaloupe) were grown in pots with varying levels of Meloidogyne incognita and were compared to susceptible melons that were grafted onto Cucumis metuliferus or Cucurbita moschata rootstocks. In addition, the effect of using melons as transplants in nematode-infested soil was compared to direct seeding of melons in nematode-infested soil. There were no differences in shoot or root weight, or severity of root galling between transplanted and direct-seeded non-grafted susceptible melon in nematode-infested soil. Susceptible melon grafted on C. moschata rootstocks had lower root gall ratings and, at high nematode densities, higher shoot weights than non-grafted susceptible melons. However, final nematode levels were not lower on the grafted than on the non-grafted plants, and it was therefore concluded that grafting susceptible melon on to C. moschata rootstock made the plants tolerant, but not resistant, to the nematodes. Grafting susceptible melons on C. metuliferus rootstocks also reduced levels of root galling, prevented shoot weight losses, and resulted in significantly lower nematode levels at harvest. Thus, C. metuliferus may be used as a rootstock for melon to prevent both growth reduction and a strong nematode buildup in M. incognita-infested soil.     

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.     

Characterization of a New Burrowing Nematode Population,Radopholus citrophilus,from Hawaii

Karyotype, host preference, isozyzme patterns, morphometrics, and mating behavior of two burrowing nematode populations from Hawaii, one infecting Anthurium sp. and the second infecting Musa sp., were compared with Radopholus similis and R. citrophilus populations from Florida. The population from Anthurium sp. had five chromosomes (n = 5), and that from Musa sp. had four (n = 4). Neither of the Hawaiian nematode populations persisted in roots of Citrus limon or C. aurantium. Anthurium clarinerivum and A. hookeri were hosts of the burrowing nematode population from anthurium in Hawaii and of R. citrophilus from Florida, whereas the two anthurium species were poor hosts of the population from Musa sp. in Hawaii and R. similis from Florida. The isozyme pattern of the population isolated from anthurium was identical to that of R. citrophigus, whereas the pattern of the population from banana in Hawaii was identical to that of R. similis. Mating behavior between the burrowing nematode population isolated from Anthurium sp. and a Florida population of R. citrophilus supports their close taxonomic relationship. Mating was observed between the population from Anthurium sp. and the Florida population of R. citrophilus but not between the Hawaiian burrowing nematode population isolated from Musa sp. and a Florida population of R. citrophilus. These findings indicate that a previously unidentified population of R. citrophilus which does not parasitize citrus occurs in Hawaii.     

Cotton Root Protection from Plant-Parasitic Nematodes by Abamectin-Treated Seed

Abamectin is nematicidal to Meloidogyne incognita and Rotylenchulus reniformis, but the duration and length of cotton taproot protection from nematode infection by abamectin-treated seed is unknown. Based on the position of initial root-gall formation along the developing taproot from 21 to 35 d after planting, infection by M. incognita was reduced by abamectin seed treatment. Penetration of developing taproots by both nematode species was suppressed at taproot length of 5 cm by abamectin-treated seed, but root penetration increased rapidly with taproot development. Based on an assay of nematode mobility to measure abamectin toxicity, the mortality of M. incognita associated with a 2-d-old emerging cotton radicle was lower than mortality associated with the seed coat, indicating that more abamectin was on the seed coat than on the radicle. Thus, the limited protection of early stage root development suggested that only a small portion of abamectin applied to the seed was transferred to the developing root system.     

Aggressiveness and Damage Potential of Central American and Caribbean Populations of Radopholus spp. in Banana

Monoxenic cultures of burrowing nematode populations extracted from banana roots from Belize, Guatemala, Honduras, and Costa Rica were established on carrot discs. Cultures of Radopholus spp. were also obtained from Florida, Puerto Rico, Dominican Republic, and Ivory Coast. The aggressiveness (defined as reproductive fitness and root necrosis) of these populations was evaluated by inoculating banana plants (Musa AAA, cv. Grande Naine) with 200 nematodes/plant. Banana plants produced by tissue culture were grown in 0.4-liter styrofoam cups, containing a 1:1 mix of a coarse and a fine sand, at ca. 27 °C and 80% RH. Banana plants were acclimated and allowed to grow for 4 weeks prior to inoculation. Plant height, fresh shoot and root weights, root necrosis, and nematode population densities were determined 8 weeks after inoculation. Burrowing-nematode populations varied in aggressiveness, and their reproductive fitness was generally related to damage reported in the field. Plant height and fresh shoot and root weight did not reflect damage caused by nematodes under our experimental conditions. Necrosis of primary roots was closely related to the reproductive fitness of the nematode populations. Variation in aggressiveness among nematode populations followed a similar trend in the two susceptible hosts tested, Grande Naine and Pisang mas. All nematode populations had a low reproductive factor (Rf ≤2.5) in the resistant host except for the Ivory Coast population which had a moderate reproductive factor (Rf ≤ 5) on Pisang Jari Buaya. This is the first report of a burrowing nematode population parasitizing this important source of resistance to R. similis.     

Host-Parasite Relationships of Meloidogyne trifoliophila Isolates from New Zealand

Root-infecting nematodes are commonly found on white clover in New Zealand pasture where they reduce yield, nitrogen fixation, and persistence. The dominant root-knot nematode on white clover in New Zealand is confirmed in this study as Meloidogyne trifoliophila by isozyme phenotype comparison with the type population from Tennessee. Results from a host differential test differed in the host ranges of M. trifoliophila and M. hapla from New Zealand locations, with M. trifoliophila failing to reproduce on the standard host plants of the test. The size and character of white clover root galls differ between species as M. trifoliophila galls are large, elongate, and smooth compared to the M. hapla galls, which are small, round, inconspicuous, and generally have adventitious, lateral roots. Culture and identification of root-knot nematode populations from sites in the North Island of New Zealand showed that M. trifoliophila is more widespread and abundant than M. hapla. Similar differential resistant and susceptible galling responses among half-sib families of white clover from a breeding program indicated that all M. trifoliophila populations tested were of the same pathotype. This resistant material was not effective in reducing reproduction of M. hapla. Meloidogyne trifoliophila did not develop to maturity on six grasses tested, but galls were formed on some species.     

Ultrastructure and Development of Pasteuria sp. (S-1 strain), an Obligate Endoparasite of Belonolaimus longicaudatus (Nemata: Tylenchida)

Pasteuria sp., strain S-1, is a gram-positive, obligate endoparasitic bacterium that uses the phytoparasitic sting nematode, Belonolaimus longicaudatus, as its host in Florida. The host attachment of S-1 appears to be specific to the genus Belonolaimus with development occurring only in juveniles and adults of B. longicaudatus. This bacterium is characterized from other described species of Pasteuria using ultrastructure of the mature endospore. Penetration, development, and sporogenesis were elucidated with TEM, LTSEM, and SEM and are similar to other nematode-specific Pasteuria. Recent analysis of 16S rDNA sequence homology confirms its congeneric ranking with other Pasteuria species and strains from nematodes and cladocerans, and corroborates ultrastructural, morphological, morphometric, and host-range evidence suggesting separate species status.     

Resistance of Auto- and Allotetraploid Triticeae Species and Accessions to Meloidogyne chitwoodi based on Genome Composition

The Columbia root-knot nematode Meloidogyne chitwoodi parasitizes several plant species, including grasses that have been developed for semiarid environments, and substantially reduces the productivity of cereals and the longevity of perennial grasses growing under semiarid conditions throughout the intermountain region. Thirty-two auto- and allotetraploid (2n = 28) taxa in the perennial Triticeae were evaluated as possible sources of resistance to M. chitwoodi. Low levels of root galling were observed on roots of all accessions; root-gall indices ranged from 0 (no galls) to 1.95 in the grasses compared to 4.67 for the susceptible ''Ranger'' alfalfa check on a scale of 1 to 6. Even though the gall ratings were low, significant (P < 0.01) differences among accessions of the same species, among species, and among genera with different genomes were observed. Within the reproductive indices, which ranged from 0.01 to 1.20 in the grasses compared to 65.38 for the alfalfa check, there was no difference among genera with different genomes and accessions within the same species and genome; however, there was a significant (P < 0.05) difference among species with the same genomes. This variation can be traced to Thinopyrum nodosum (Jaaska-19), which was the only accession with a reproductive factor greater than 1.00. Based on the data, all auto- and allotetraploids are considered resistant to M. chitwoodi.     

Host Plant Resistance as an Alternative to Methyl Bromide for Managing Meloidogyne incognita in Pepper

Pre-plant soil fumigation with methyl bromide and host resistance were compared for managing the southern root-knot nematode (Meloidogyne incognita) in pepper. Three pepper cultivars (Carolina Cayenne, Keystone Resistant Giant, and California Wonder) that differed in resistance to M. incognita were grown in field plots that had been fumigated with methyl bromide (98% CH₃Br : 2% CCl₃NO₂ [w/w]) before planting or left untreated. Carolina Cayenne is a well-adapted cayenne-type pepper that is highly resistant to M. incognita. The bell-type peppers Keystone Resistant Giant and California Wonder are intermediate to susceptible and susceptible, respectively. None of the cultivars exhibited root galling in the methyl bromide fumigated plots and nematode reproduction was minimal (<250 eggs/g fresh root), indicating that the fumigation treatment was highly effective in controlling M. incognita. Root galling of Carolina Cayenne and nematode reproduction were minimal, and fruit yields were not reduced in the untreated plots. The root-galling reaction for Keystone Resistant Giant was intermediate (gall index = 2.9, on a scale of 1 to 5), and nematode reproduction was moderately high. However, yields of Keystone Resistant Giant were not reduced in untreated plots. Root galling was severe (gall index = 4.3) on susceptible California Wonder, nematode reproduction was high, and fruit yields were reduced (P ≤ 0.05) in untreated plots. The resistance exhibited by Carolina Cayenne and Keystone Resistant Giant provides an alternative to methyl bromide for reducing yield losses by southern root-knot nematodes in pepper. The high level of resistance of Carolina Cayenne also suppresses population densities of M. incognita.     

A Method for Field Infestation with Meloidogyne incognita

A field inoculation method was developed to produce Meloidogyne spp. infestation sites with minimal quantities of nematode inoculum and with a reduced labor requirement compared to previous techniques. In a preseason-methyl bromidefumigated site, nematode egg suspensions were delivered at concentrations of 0 or 10^x eggs/m of row where x = 2.12, 2.82, 3.52, or 4.22 through a drip line attached to the seed firmer of a commercial 2-row planter into the open seed furrow while planting cowpea. These treatments were compared to a hand-inoculated treatment, in which 10^3.1 eggs were delivered every 30 cm in 5 ml of water agar suspension 2 weeks after planting. Ten weeks after planting, infection of cowpea roots was measured by gall rating and gall counts on cowpea roots. A linear relationship between the inoculation levels and nematode-induced galls was found. At this time, the amount of galling per root system in the hand-inoculated treatment was less than in the machine-applied treatments. Advantages of this new technique include application uniformity and low population level requisite for establishing the nematode. This method has potential in field-testing of Meloidogyne spp. management strategies by providing uniform infestation of test sites at planting time.     

Comparison of Methods for Assessing Resistance to Meloidogyne arenaria in Peanut

Use of resistant cultivars is a desirable approach to manage the peanut root-knot nematode (Meloidogyne arenaria). To incorporate resistance into commercially acceptable cultivars requires reliable, efficient screening methods. To optimize the resistance screening protocol, a series of greenhouse tests were done using seven genotypes with three levels of resistance to M. arenaria. The three resistance levels could be separated based on gall indices as early as two weeks after inoculation (WAI) using 8,000 eggs of M. arenaria per plant, while four or more weeks were needed when 1,000–6,000 eggs/plant were used. High inoculum densities (over 8,000 eggs/plant) were needed to separate the three resistance levels based on eggs per gram of root within eight WAI. A gall index based on percentage of galled roots could separate the three resistance levels at lower inoculum levels and earlier harvest dates than other assessment methods. The use of eggs vs. second-stage juveniles (J2) as inoculum provided similar results; however, it took three to five more days to collect J2 than to collect eggs from roots. Plant age affected gall index and nematode reproduction on peanut, especially on the susceptible genotypes AT201 and D098. The genotypes were separated into their correct resistance classes when inoculated 10 to 30 days after planting, but were not separated correctly when inoculated on day 40.     

Meloidogyne lusitanica n. sp. (Nematoda: Meloidogynidae), a Root-knot Nematode Parasitizing Olive Tree (Olea europaea L.)

A root-knot nematode from Portugal, Meloidogyne lusitanica n. sp., is described and illustrated from specimens obtained from olive trees (Olea europaea L.). Females of the new species have a characteristic perineal pattern with medium to high trapezoidal dorsal arch with distinct punctuations in the tail terminus area. The excretory pore is located posterior to the stylet, about 1.5-2.5 stylet lengths from the anterior end. The stylet is 17.1 μm long with pear-shaped knobs. Males have a rounded, posteriorly sloping head cap and head region not annulated. The robust stylet, 24.5 μ long, has large, elongate knobs. Mean length of the second-stage juveniles is 449.5 μm, stylet length 14.2 μm, and tail length 44.1 μm. Scanning electron microscope observations provide further details of perineal patterns and head and stylet morphology of females, males, and second-stage juveniles. Meloidogyne lusitanica n. sp. did not reproduce on any of the differential hosts used to separate the four most common Meloidogyne species. The common name "olive root-knot nematode" is proposed for M. lusitanica n. sp.