Phylogenetic Relationships Among Selected Heteroderoidea Based on 18S and ITS Ribosomal DNA.

In a study of relationships among selected cyst-forming and noncyst-forming species of Heteroderoidea, combined sequences comprised of DNA from part of the conserved 18S ribosomal RNA gene (rDNA) plus the complete ITS rDNA segment were more similar to analyses based on the ITS data alone than to analyses based on the 18S data alone. One of the two noncyst-forming species, Ekphymatodera thomasoni, grouped with cyst-forming species of Heteroderoidea. Bilobodera flexa, also a noncyst-forming species, was separated from all the other taxa by a long branch. Afenestrata koreana, with a weakly sclerotized cyst, grouped closely with H. bifenestra. These observations suggest that phylogenetic analyses using molecular data may aid in our understanding of the evolution of cyst formation in nematodes, including the possibility of secondary loss. The usefulness of molecular phylogenetic analyses in nematodes may depend more on the particular selection of taxa than on mere addition of data from additional genes.

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.

Spatial Patterns of Belonolaimus spp. Among and Within Citrus Orchards on Florida's Central Ridge.

A survey was initiated to determine the incidence of Belonolaimus spp. (sting nematodes) in citrus orchards in the central ridge region of Florida, following widespread damage by these nematodes to young trees replanted after freezing weather in 1989-90. Sting nematodes were detected in 50% of 210 samples and in 64% of 84 orchards surveyed. More orchards in Polk County were infested with sting nematodes (82%) than in counties to the north (36%) or south (48%). Principal component analysis of morphometric data separated six of seven sting nematode populations in northeastern Polk County from six populations in adjacent regions. Stylet:tail ratio for nematodes in northeastern Polk County tend to be > 1.0 and were << 1.0 for all other populations. Patchiness of nematodes within an orchard was associated with stunted trees (23% smaller), reduced root mass density (25% lower), and low fruit yield (57% reduction). Soil texture did not vary among trees of different size in the orchard, but soil water potential between irrigation events was highest beneath small trees with low root mass density. Results of the survey indicate that the incidence of sting nematodes in orchards on the central ridge is much higher than previously estimated and that sting nematodes can cause substantial damage in replanted orchards. Further research is needed to evaluate the significance of sting nematode population variability and its relationship to citrus crop loss in Florida.

Host Response of Ornamental Palms to Rotylenchulus reniformis.

The responses of 20 species of ornamental palms and one cycad (Cycas revoluta) to two populations of the reniform nematode, Rotylenchulus reniformis, from southern Florida were studied in two greenhouse experiments conducted in 1989-1991 and 1991-92. Ornamental palms in pots were exposed to initial population densities of 400 and 1,500 R. reniformis/l00 cm(3) soil for 16 and 15 months, respectively. Nematode reproduction occurred on Acoelorrhaphe wrightii and Washingtonia robusta, but not on the other palms or the cycad. In both experiments, nematode numbers on A. wrightii and W. robusta were significantly smaller than those on cowpea (Vigna unguiculata), a susceptible host of the nematode used as a control in these experiments. Nematodes surviving in pots containing nonhost palms for 16 months retained infectivity and were able to reproduce on susceptible cowpea in a bioassay. Sections from Washingtonia robusta roots infected by R. reniformis females showed the nematode feeding on syncytia formed by endodermal, pericyclic, and vascular parenchyma cells in a manner similar to that reported for other monocot hosts of the reniform nematode.

Pasteuria sp. Parasitizing Trophonema okamotoi in Florida.

Two populations of Trophonema okamotoi parasitized by Pasteuria sp. were found on Liquidambar styraciflua (sweetgum) and on an unidentified tropical grass in north-central Florida. Endospores of this Pasteuria sp. attached to motile vermiform second-stage juveniles (J2) and males of T. okamotoi, but not to other developmental stages. Sporangia and new endospores were produced only inside the bodies of swollen and sedentary third- and fourth-stage juveniles and females that developed in the host roots. No egg masses were produced by infected T. okamotoi females. The endospore diameter from the tropical grass population was 4.93 mum and the central core diameter was 1.97 mum; measurements of endospores from the sweetgum populations were similar. Endospores that were collected from T. okamotoi and added to uninfected T. okamotoi and other plant-parasitic nematodes attached/to J2 of T. okamotoi but did not attach to juveniles and adults of Helicotylenchus pseudorotrustus, Pratylenchus brachyurus, or to J2 of either Meloidogyne arenaria race 1, M. incognita race 1, M. javanica, or Tylenchulus semipenetrans. Pasteuria sp. from T. okamotoi differed from the described Pasteuria species in endospore size, host preference, and rate of attachment.

Mesocriconema ornicauda n. sp. and Ogma floridense n. sp. (Nematoda: Criconematidae) from Two Florida Habitats.

Mesocriconema ornicauda n. sp. and Ogma floridense n. sp. are described from two native habitats of central and northwestern Florida. Mesocriconema ornicauda is closest to M. annulatiforme (De Grisse &Loof, 1967) Loof &De Grisse, 1989, but differs by the shorter stylet of the female (43-50 vs. 54-65 mum) and the moderately conoid tail of the male, which is pointed in M. annulatiforme. Ogma floridense is closest to O. hungaricum (Andrassy, 1972) Siddiqi, 1986. Females differ from those of O. hungaricum by the first of two labial annuli being wider, whereas they are subequal in O. hungaricum. Ogma floridense females differ also by entire or bifid cuticular scales, which are consistently divided into two or four projections in O. hungaricum, the shorter body (360-471 vs. 480-550 mum), the shorter stylet (87-98 vs. 95-100 mum), and the more anteriorly located excretory pore (Rex = 17-19 vs. 21-23).

Tylenchulus palustris Parasitizing Peach Trees in the of nited States.

Most morphological characteristics of three populations of a Tylenchulus sp. from peach roots in Alabama, Arkansas, and Georgia did not differ from those of T. palustris paratypes. However, some mature females differed slightly from those of T. palustris paratypes from Florida. These mature females were more swollen in the posterior portion of their bodies, and they possessed digitate postvulval body sections with round rather than conoid termini. These morphological variants had a wide postvulval section core (PVSC), as do T. palustris paratypes; they did not differ from the paratypes in other characteristics. Second-stage juveniles and males were less morphologically variable and were not different from the paratypes. No males were found in populations from Alabama and Georgia. The Tylenchulus sp. from three peach sites was determined to be T. palustris. This is the first report of T. palustris on an economically important crop.

Distribution, Hosts, and Morphological Characteristics of Tylenchulus palustris in Florida and Bermuda.

Studies on the geographical distribution and hosts of Tylenchulus palustris were conducted over a 3-year period in Florida and Bermuda. Tylenchulus palustris was found on Aster elliottii and Liquidambar styraciflua roots in swamps of northern and central Florida. It was detected also on Borrichia arborescens and B. frutescens roots in tidal marshes of northern Florida and coastal rocklands of southern Florida and Bermuda. Posterior bodies of T. palustris swollen females from Bermuda did not differ from those of the paratypes; however, second-stage juvenile bodies and male tails from Bermuda were longer than those of the paratypes. Greenhouse host tests indicated that Mikania scandens is a host of T. palustris but not of T. semipenetrans.

Host Status of Citrus and Citrus Relatives to Tylenchulus graminis.

Studies were conducted on the host suitability of four citrus rootstocks--rough lemon (Citrus limon), sour orange (C. aurantium), trifoliate orange (Poncirus trifoliata cv. Argentina), and Swingle citrumelo (C. paradisi x P. trifoliata)--to Tylenchulus graminis which was previously considered a "grass" race of T. semipenetrans. In an uncultivated field, sour orange seedlings grown with T. graminis-infected broomsedge (Andropogon virginicus) were not infected with this nematode after 18-month's exposure to T. graminis population densities ranging from < 0.01 to 0.4 second-stage juveniles (J2)/cm(3) soil. In a greenhouse test, two T. graminis populations from two Florida locations did not infect sour orange seedlings grown for 2 years in soil naturally infested with 0.3 and 1.3 J2/cm(3). Rough lemon, trifoliate orange, and Swingle citrumelo seedlings suppressed T. graminis initial population densities of 7 to final values of < 0.1 J2/cm(3) soil. Final values of > 70.0 J2/cm(3) occurred in soil with broomsedge. These findings provide conclusive evidence that T. graminis is a specific parasite of grasses and does not infect citrus.

Tylenchulus graminis n. sp. and T. palustris n. sp. (Tylenchulidae), from Native Flora of Florida, with Notes on T. semipenetrans and T. furcus.

Tylenchulus graminis n. sp. and T. palustris n. sp. are described and illustrated from broomsedge (Andropogon virginicus L.) and pop ash (Fraxinus caroliniana Mill.), respectively. T. graminis resembles T. furcus in having a distinct anus, but T. graminis second-stage juveniles (J2) do not have a bifid tail. T. semipenetrans does not have a perceptible anus. The mature female of T. graminis has a mucronate pointed terminus while T. semipenetrans has a smooth and round terminus. T. graminis males have wider stylet knobs and basal bulb and a longer tail than T. semipenetrans males. T. graminis J2 have a longer posterior body portion (without large fat globules) than T. semipenetrans J2. T. palustris resembles T. semipenetrans in having an undetectable anus but differs by the short and conoid mature female postvulval section. The male of T. palustris has larger stylet knobs and basal bulb than those of T. semipenetrans and a bluntly rounded tail terminus, which is tapered in T. semipenetrans. T. palustris differs from T. furcus and T. graminis in having an undetectable anus, by the conoid postvulval section of mature females, by the shorter and rounded tail of males, and the shorter J2 posterior body section without large fat globules. T. graminis and T. palustris are parasites of indigenous flora of Florida.

Effect of Temperature on Infection and Survival of Rotylenchulus reniformis.

From infestation of lettuce with preinfective females to egg deposition, populations of Rotylenchulus reniformis from Baton Rouge, Louisiana; Lubbock and Weslaco, Texas; and Mayaguez, Puerto Rico, required 41, 13, 7, and 7 days at 15, 20, 25, and 34 C, respectively. No nematode infection occurred at 10 C with any R. reniformis population, and the population from Puerto Rico did not reproduce at 15 C. Nematode survival was not influenced by temperature, since populations from Texas and Louisiana survived for 6 months without a host at - 5 , - 1 , 4, and 25 C. Survival of R. reniformis was substantially influenced by soil moisture. Soil moistures greater than 7% (< 1 bar) aided nematode survival at storage temperature of 25 C, whereas moisture adversely affected nematode survival below freezing. Soil moisture below 4% (> 15 bars) favored nematode survival below freezing but adversely affected nematodes in soils stored at 25 C. Soil moisture effects on nematode survival were less accentuated at 4 and 0 C.

Morphometrics, Illustration, and Histopathology of Sphaeronema rumicis on Cottonwood in Utah.

The morphology of a population of Sphaeronema rumicis Kir'yanova found on cottonwood in Utah is illustrated by light and scanning electron micrographs, as well as by drawings. This is the first report of males of S. rumicis, a species also not known previously to occur in North America. S. rumicis females on cottonwood in the United States were smaller than those found by Kir'yanova on sorrel in the USSR. Females and second-stage juveniles (J2) from the United States had slightly shorter stylets than did females and J2 from the USSR. Males were vermiform and had degenerate esophagi. On secondary cottonwood roots S. rumicis induces formation of a syncytium originating from proliferated pericyclic cells. Thick outer walls, wall protuberances, absence of cell wall ingrowths, dense cytoplasm, and hypertrophied nuclei were the main characteristics of syncytia observed in S. rumicis-infected cottonwood roots.

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.

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.