The ecology and biodemography of Caenorhabditis elegans.

The nematode Caenorhabditis elegans is a well-known model organism for research on aging and life span, but very little is known about its ecology and natural history. The strain N2 is the standard wild-type C. elegans and arose from the progeny of a single hermaphrodite. Since N2 has passed through laboratory culture, the influence of inadvertent selection and genetic drift on C. elegans strains kept in culture is unclear. Because it seems that other wild-type strains have also been subject to lengthy laboratory culture, the life span and biodemography of wild-caught C. elegans is of interest. We recovered C. elegans from snails (Helix aspersa) in ca. 50% of the California locations where we made collections. In experiments with one of the wild-caught isolates, it differed in important demographic properties, mortality, fertility, fitness, and activity patterns, from the standard N2 strain, when both strains were evaluated in a common laboratory environment. The differences were not only statistically significant; they were also large enough to be biologically important. The differences are consistent with the hypothesis that N2 has adapted to laboratory conditions.

Influence of Lysobacter enzymogenes Strain C3 on Nematodes.

Chitinolytic microflora may contribute to biological control of plant-parasitic nematodes by causing decreased egg viability through degradation of egg shells. Here, the influence of Lysobacter enzymogenes strain C3 on Caenorhabditis elegans, Heterodera schachtii, Meloidogyne javanica, Pratylenchus penetrans, and Aphelenchoides fragariae is described. Exposure of C. elegans to L. enzymogenes strain C3 on agar resulted in almost complete elimination of egg production and death of 94% of hatched juveniles after 2 d. Hatch of H. schachtii eggs was about 50% on a lawn of L. enzymogenes strain C3 on agar as compared to 80% on a lawn of E. coli. Juveniles that hatched on a lawn of L. enzymogenes strain C3 on agar died due to disintegration of the cuticle and body contents. Meloidogyne javanica juveniles died after 4 d exposure to a 7-d-old chitin broth culture of L. enzymogenes strain C3. Immersion of A. fragariae, M. javanica, and P. penetrans juveniles and adults in a nutrient broth culture of L. enzymogenes strain C3 led to rapid death and disintegration of the nematodes. Upon exposure to L. enzymogenes strain C3 cultures in nutrient broth, H. schachtii juveniles were rapidly immobilized and then lysed after three days. The death and disintegration of the tested nematodes suggests that toxins and enzymes produced by this strain are active against a range of nematode species.

Revising the standard wisdom of C. elegans natural history: ecology of longevity.

Here, we consider that most of the research concerning Caenorhabditis elegans has been laboratory focused and that only limited research has directly considered the worm's biology relative to its natural history in the wild. We describe that, although the worm has traditionally been considered a soil nematode, we could not find it in soil but frequently recovered it from snails. Finally, we discuss how a better understanding of the natural history of C. elegans may enhance its usefulness as a model organism for studying aging and other phenomena.

Assessment of Host-Induced Selection on Three Geographic Isolates of Heterodera schachtii Using RAPD and AFLP Markers.

ABSTRACT The hypothesis that host plants exert selection pressure on Heterodera schachtii populations was tested. Host selection of genotypes from three genetically distinct isolates of H. schachtii was assessed using cabbage, sugar beet, oilseed radish (Raphanus sativus), and white mustard (Sinapis alba). The plants represent a range of susceptibility to H. schachtii and included R. sativus and S. alba, because cultivars of those species have been used as trap crops for H. schachtii in Europe. Genotypic differences in amplified fragment length polymorphism (AFLP) and random amplified polymorphic DNA (RAPD) markers were detected among the isolates after they reproduced on the different hosts. The poorest host plant, R. sativus, resulted in the greatest number of changes in both AFLP and RAPD markers. Oilseed radish selected nematode genotypes in less than four nematode generations. The nematode population genotypes detected by RAPD analyses after selection on oilseed radish were observed even after nematode populations were transferred back to the other three hosts. The genetic markers that were detected after selection were influenced by the genotypes of the original nematode isolates. The results indicate the utility of RAPDs and AFLPs for identifying and monitoring intraspecific genetic variability in nematodes and for understanding nematode population responses to host plants. Nematode management practices such as using resistant cultivars may alter gene frequencies, thereby reducing the efficacy of the tactic and exacerbating the nematode's potential to damage subsequent crops.

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.

Penetration of Crotalaria juncea, Dolichos lablab, and Sesamum indicum Roots by Meloidogyne javanica.

Penetration of Crotalaria juncea (PI 207657 and cv. Tropic Sun) Dolichos lablab cv. Highworth, and Sesamum indicum by juveniles (J2) of Meloidogyne javanica was assessed to investigate the mechanism by which these plants may reduce nematode numbers in the field. Growth chamber experiments were conducted at 25 C, with vials containing 90 g sand infested with 450 J2; tomato (UC 204 C) was included as a susceptible host. Fifteen days after inoculation, roots were stained and the nematodes within stained roots were counted. Both C. juncea lines were highly resistant to penetration, as they contained significantly fewer nematodes per cm of root and per root system than the other plants. Although containing more nematodes per cm of root than C. juncea, S. indicum and D. lablab had significantly fewer nematodes per root system and per cm of root than tomato. Roots were significantly longer in the plants with the lowest nematode penetration. Although C. juncea, D. lablab, and S. indicum may have potential utility as cover or rotation crops in soil infested with M. javanica, further quantitative information on the reproduction of M. javanica and other nematodes in these plants is needed.

Host Status of Crotalaria juncea, Sesamum indicum, Dolichos lablab, and Elymus glaucus to Meloidogyne javanica.

Reproduction of Meloidogyne javanica on Crotalaria juncea PI 207657 and cv. Tropic Sun, Sesamum indicum, Dolichos lablab, and Elymus glaucus was assessed using a root-gall index, a reproductive index obtained by dividing the final population of juveniles (J2) in soil by the initial J2 population (Pf/Pi), and the number of J2 per gram of root recovered from roots by mist chamber extraction. Lycopersicon esculentum (cv. UC 204 C) was included as a susceptible host. The root-gall index and soil reproductive index were poor indicators of the host status of our test plants as compared with mist chamber extraction of J2 from roots. Lycopersicon esculentum had a mean root-gall index of 7.8. Some plants of S. indicum and E. glaucus had a few galls and other plants had none, with mean root-gall indices of 1.6 and 0.8, respectively. No galls were observed in C. juncea and D. lablab. Lycopersicon esculentum had the highest mean soil Pf/Pi value (mean = 1.93), while in C. juncea and some replicates of S. indicum no soil J2 were found. Even though some replicates had no galls, all replicates supported nematode reproduction. The mean numbers of J2 per gram root after 5 days of mist extraction were 447.7, 223.3, 165.5, 96.9, 42.3, and 41.9 for D. lablab, L. esculentum, E. glaucus, S. indicum, and C. juncea PI 207657 and cv. Tropic Sun, respectively. Accurate assessment of nematode resistance was influenced by sampling time and the nematode extraction technique used. Individual plants of both C. juncea and S. indicum supported nematode reproduction to some extent; however, both C. juncea and S. indicum have potential as cover crops to reduce M. javanica numbers.

Raphanus sativus, Sinapis alba, and Fagopyrum esculentum as Hosts to Meloidogyne incognita, Meloidogyne javanica, and Plasmodiophora brassicae.

Cultivars of oilseed radish (Raphanus sativus var. oleifera cv. Adagio, Nemex, Pegletta, Renova, Siletina, Siletta Nova, and Ultimo), white mustard (Sinapis alba cv. Albatross, Emergo, Maxi, Martigena, Metex, and Serval), buckwheat (Fagopyrum esculentum cv. Prego, Tardo), and phacelia (Phacelia tanacetifolia cv. Angelia) were tested for susceptibility to Meloidogyne incognita race 3 and Meloidogyne javanica. Experiments were conducted in growth chambers at 25 C and 16 hours light for 42 days after inoculation with second-stage juveniles (J2). All cultivars were susceptible to M. incognita and M. javanica. The oilseed radish (cv. Nemex, Pegletta, and Renova) and white mustard (cv. Emergo) were also examined to determine the influence of Heterodera schachtii on susceptibility to Plasmodiophora brassicae as measured by incidence and severity of root galling. All cultivars were susceptible, and neither the severity nor incidence of clubroot galling was affected by H. schachtii.

Enzymatic Digestion of Roots for Recovery of Root-knot Nematode Developmental Stages.

Developmental stages of Meloidogyne javanica were successfully released from roots by treatment with commercially available cellulase and pectinase. The average percentage recovery of nematode developmental stages from Dolichos lablab, Elymus glaucus, and Lycopersicon esculentum were as follows: eggs = 526%, J2 = 272%, J3 = 783%, J4 = 549%, adult females = 285%, and total = 425%, expressed as percentages of the counts obtained from stained roots spread on glass plates. Root digestion was more accurate and sensitive in detecting low numbers of nematodes in roots than was the glass plate method. No simple linear, quadratic, or cubic relationship was found between the two methods that would allow a conversion factor to be developed.

Penetration, Development, and Reproduction of Heterodera schachtii on Fagopyrum esculentum, Phacelia tanacetifolia, Raphanus sativus, Sinapis alba, and Brassica oleracea.

The penetration, development, and reproduction of a California population of the sugarbeet cyst nematode, Heterodera schachtii, was observed on cultivars of cabbage (Brassica oleracea), phacelia (Phacelia tanacetifolia), buckwheat (Fagopyrum esculentum), oilseed radish (Raphanus sativus), and white mustard (Sinapis alba). With the exception of the nonhost, phacelia, all were readily penetrated by second-stage juveniles of H. schachtii. After 38 days at 25 C, no cysts were observed on phacelia cv. Angelia or on the oilseed radish cv. Nemex and Pegletta. Cyst production was low (<2.5 cysts/plant) on the buckwheat cv. Tardo and Prego and most of the oilseed radish cultivars. Cyst production was intermediate (5-14 cysts/plant) on most of the white mustard cultivars, and high on cabbage (20-110 cysts/plant). In microplot studies conducted over 133 days (approx. 450 degree-days, base 8 C), the reproductive index for H. schachtii was greater than 1.0 for cultivars of phacelia, oilseed radish, and white mustard as welt as in fallow treatments, indicating the need for further research on the use of these crops under field conditions.

Applied biotechnology in nematology.

During the past two decades, rapid advances in biotechnology and molecular biology have affected the understanding and treatment of human and plant diseases. The human and Caenorhabditis elegans genome-sequencing projects promise further techniques and results useful to applied nematology. Of course, biotechnology is not a panacea for nematological problems, but it provides many powerful tools that have potential use in applied biology and nematode management. The tools will facilitate research on a range of previously intractable problems in nematology, from identification of species and pathotypes to the development of resistant cultivars that have been inaccessible because of technical limitations. However, to those unfamiliar or not directly involved with the new technologies and their extensive terminology, the benefits of the advances in biotechnology may not be readily discerned. The sustainable agriculture of the future will require ecology-based management, and successful integrated nematode management will depend on combinations of control tactics to reduce nematode numbers. In this review we discuss how biotechnology may influence nematode management, define terminology relative to potential applications, and present current and future avenues of research in applied nematology, including species identification, race and pathotype identification, development of resistant cultivars, definition of nematode-host interactions, nematode population dynamics, establishment of optimal rotations, the ecology of biological control and development of useful biological control agents, and the design of novel nematicides.

Random Amplified Polymorphic DNA Analysis of Heterodera cruciferae and H. schachtii populations.

Heterodera schachtii and H. cruciferae are sympatric in California and frequently occur in the same field upon the same host. We have investigated the use of polymerase chain reaction (PCR) amplification of nematode DNA sequences to differentiate H. schachtii and H. cruciferae and to assess genetic variability within each species. Single, random oligodeoxyribonucleotide primers were used to generate PCR-amplified fragments, termed RAPD (random amplified polymorphic DNA) markers, from genomic DNA of each species. Each of 19 different random primers yielded from 2 to 12 fragments whose size ranged from 200 to 1,500 bp. Reproducible differences in fragment patterns allowed differentiation of the two species with each primer. Similarities and differences among six different geographic populations of H. schachtii were detected. The potential application of RAPD analysis to relationships among nematode populations was assessed through cluster analysis of these six different populations, with 78 scorable markers from 10 different random primers. DNA from single cysts was successfully amplified, and genetic variability was revealed within geographic populations. The use of RAPD markers to assess genetic variability is a simple, reproducible technique that does not require radioisotopes. This powerful new technique can be used as a diagnostic tool and should have broad application in nematology.

Nematicide Efficacy, Root Growth, and Fruit Yield in Drip-irrigated Pineapple Parasitized by Rotylenchulus reniformis.

A 3-year field trial near Kunia, Oahu, Hawaii, was conducted to evaluate four nematicide treatments for efficacy against Rotylenchulus reniformis in drip-irrigated pineapple (Ananas comosus L. (Merr.)). The treatments were (A) preplant fumigation with 1,3-dichloropropene (1,3-D) (336 liter/ ha) and postplant drip application of fenamiphos (3.4 kg/ha) with restricted irrigation, (B) preplant 1,3-D only, weekly irrigation, (C) 1,3-D fenamiphos, weekly irrigation, and (D) postplant fenamiphos only, weekly irrigation. Fenamiphos was applied at 3-month intervals for 1 year after planting in three treatments. Although nematode populations increased in all treatments 1 year after planting, no differences in fruit yield were detected among treatments in the first (plant crop) harvest 19 months after planting. In the second (ratoon) crop (33 months after planting) significant yield differences, larger fruit size, and greater root biomass were obtained in the dual nematicide treatments. Root biomass increased continuously throughout the crop cycle, was greatest near the drip line, and showed a shallow depth distribution (30-40 cm). Rotylenchulus reniformis populations and fenamiphos concentrations were negatively correlated in soil profiles taken 13 months after planting. In the absence of postplant fenamiphos applications, nematode numbers were positively correlated with root biomass.