Amino acid, heme, and sterol requirements of the nematode, Rhabditis maupasi

Rhabditis maupasi, a nematode that occurs in the mantle cavity of Helix aspersa and related North African food snails, requires 5 amino acids (lysine, methionine, phenylalanine, tryptophane and valine) for maintenance of the stage-3 survival larvae, and 5 additional amino acids (arginine, isoleucine, leucine, threonine and, marginally, histidine) for development of these larvae into adults and for reproduction. Tyrosine is beneficial but not absolutely required for reproduction. These results were obtained with axenic R. maupasi grown in a chemically defined medium containing salts, trace metals, purines and pyrimidines, Krebs cycle intermediates, a fatty acid (butyric), vitamins, urea, and a carbohydrate (dextrose). The complete medium contained 18 amino acids; each of 18 test media was deficient in one of the amino acids. In the complete medium and in a medium lacking the "nematode nonessential" amino acids, stage-3 R. maupasi developed into adults and produced one generation of offspring. For continuous cultivation, however, the nematode also required hemin or another iron porphyrin as well as a sterol such as cholesterol.     

Effects of Selected Insecticides and Nematicides on the In Vitro Development of the Entomogenous Nematode Neoaplectana carpocapsae

The effects of organophosphates (mevinphos, phenamiphos, trichlorfon), carbamates (carbofuran, methomyl, oxamyl), a formamidine (chlordimeform), a synthetic pyrethroid (fenvalerate), a chlorinated hydrocarbon (methoxychlor). and an insect growth regulator (diflubenzuron) on in vitro development and reproduction of Neoaplectana carflocapsae were tested by incorporating each chemical into a nematode rearing medium. Organophosphates and carbamates adversely affected development and reproduction at concentrations ≥ 0.1 mg/ml. Phenamiphos was the most toxic, with no nematode reproduction at 0.01 mg/ml. Inoculated infective juveniles developed to adults with some of the organophosphates and carbamates, but limited or no reproduction occurred. Chlordimeform inhibited development at 1.0 mg/ml, while diflubenzuron, fenvalerate, and methoxychlor did not significantly (P > 0.05) reduced reproduction at 1.0 mg/ml. The organophosphate and carbamate nematicides in use for control of plant-parasitic nematodes may be toxic to N. carpocapsae in the soil.     

Photobactin: a Catechol Siderophore Produced by Photorhabdus luminescens, an Entomopathogen Mutually Associated with Heterorhabditis bacteriophora NC1 Nematodes

The nematode Heterorhabditis bacteriophora transmits a monoculture of Photorhabdus luminescens bacteria to insect hosts, where it requires the bacteria for efficient insect pathogenicity and as a substrate for growth and reproduction. Siderophore production was implicated as being involved in the symbiosis because an ngrA mutant inadequate for supporting nematode growth and reproduction was also deficient in producing siderophore activity and ngrA is homologous to a siderophore biosynthetic gene, entD. The role of the siderophore in the symbiosis with the nematode was determined by isolating and characterizing a mini-Tn5-induced mutant, NS414, producing no detectable siderophore activity. This mutant, being defective for growth in iron-depleted medium, was normal in supporting nematode growth and reproduction, in transmission by the dauer juvenile nematode, and in insect pathogenicity. The mini-Tn5 transposon was inserted into phbH; whose protein product is a putative peptidyl carrier protein homologous to the nonribosomal peptide synthetase VibF of Vibrio cholerae. Other putative siderophore biosynthetic and transport genes flanking phbH were characterized. The catecholate siderophore was purified, its structure was determined to be 2-(2,3-dihydroxyphenyl)-5-methyl-4,5-dihydro-oxazole-4-carboxylic acid [4-(2,3-dihydroxybenzoylamino)-butyl]-amide, and it was given the generic name photobactin. Antibiotic activity was detected with purified photobactin, indicating that the siderophore may contribute to antibiosis of the insect cadaver. These results eliminate the lack of siderophore activity as the cause for the inadequacy of the ngrA mutant in supporting nematode growth and reproduction.     

Presence or absence? Primary structure,regioselectivity and evolution of Δ12/ω3 fatty acid desaturases in nematodes

For vertebrates, the adequate supply of polyunsaturated fatty acids (PUFA) by the diet, in particular ω3 long-chain PUFA, is considered essential for neural development, growth and reproduction. In contrast to aquatic ecosystems, ω3 long-chain PUFA apparently are not widely available in the terrestrial food chain. Their de novo synthesis requires the presence of Δ12 and ω3 fatty acid desaturase enzymes, which are absent in vertebrates but present, for example, in the nematode Caenorhabditis elegans (FAT-2 and FAT-1). This raises the question if soil-dwelling nematodes offer substantial supply of these valuable nutritional compounds in terrestrial food webs. BLAST searches in available nematode genomes revealed the existence of fat-2 like genes in almost all clade III–V species, but failed to identify orthologs in clade I–II nematodes. An additional RT-PCR screen across soil-dwelling nematode species identified six novel fat-2 like genes. Hints for the genetic basis of a ω3 (fat-1) desaturase activity was found only in selected clade IV–V species, but not in clades I to III nematodes. Fatty acid pattern analyses following a PUFA-free cultivation and enzymatic characterization of six selected fat-2 or fat-1 like desaturases in yeast confirmed the findings from the genetic approaches. Thus, in similar soil habitats, taxa exist that can synthesize ω3 long-chain PUFA (as Panagrolaimus, Mesorhabditis and Caenorhabditis) whereas others are unable to do so (Acrobeloides, Cephalobus and Oscheius). While these nematodes do not differ in trophic position or major diet, distinction in reproduction mode may have led to the observed variations in desaturase genes.     

Effect and Reproduction of Rotylenchulus reniformis on Sweet Potato Selections

Growth, yield, and quality of 10 sweet potato selections and reproduction of the reniform nematode, Rotylenchulus reniformis, were studied in fumigated and nonfumigated plots in a naturally infested field. Nematode reproduction on the different selections in the field was similar to that reported in the greenhouse but was not related to the effect of the nematode on yield of the different selections. Goldrush supported the least reproduction but was the most severely affected by the nematode, while Centennial supported the most reproduction but was the least affected. Although reniform nematode was not found within enlarged fleshy roots, sweet potatoes were more frequently cracked in nonfumigated plots even when nematode populations were relatively low. One selection, P-104, was resistant to cracking. Yield of all selections tested was significantly reduced when initial populations were moderate to high (1,500-10,000 nemas per 500 cm³). Correlations were made between nematode population parameters and growth, yield, and cracking of the sweet potatoes. The initial populations and the reproduction ratio for the last part of the growing season gave the most significant negative correlations with yield for most selections.     

Interactions Between Calonectria crotalariae and Heterodera glycines on Soybean

The interactions of Heterodera glycines at four egg inoculum levels (0, 100, 1,000, and 10,000 per pot) and three cyst levels (0, 100, and 200 per pot) and Calonectria crotalariae at 500, 5,000, and 50,000 microsclerotia per pot were evaluated on soybean. At the two lowest nematode egg levels, the presence of C. crotalariae did not affect nematode reproduction. At 10,000 eggs per pot, however, nematode reproduction was increased significantly at each microsclerotial level. The increase in nematode reproduction was stepwise at 500 and 5,000 microsclerotia per pot but declined at 50,000 microsclerotia per pot. Similar results were obtained when cysts rather than eggs were used as nematode inoculum. The nematode x fungus interaction significantly affected 60-day plant growth parameters of both Lee 74 and Centennial soybean. The nematode x fungus interaction was antagonistic to plant roots and significantly influenced root injury ratings. The presence of C. crotalariae in tissues of stock plants or plants used as race differentials did not alter the analysis of this population as race 3.     

Hysterothylacium larvae (Nematoda, Anisakidae) in the freshwater mussel Diplodon suavidicus (Lea, 1856) (Mollusca, Unioniformes, Hyriidae) in Aripuanã River, Amazon, Brazil

Larvae of Hysterothylacium use various invertebrates as intermediate hosts. Definite hosts include fish, birds, reptiles or marine mammals. This study describes the occurrence of Hysterothylacium (Nematoda, Anisakidae) larvae parasitizing the pericardic cavity of Diplodon suavidicus (Unioniformes, Hyriidae) specimens collected in the Amazon basin, Brazil. This is the first record of this nematode parasitizing freshwater bivalves in South America. The high prevalence, medium intensity and medium abundance suggest that D. suavidicus acts as intermediate host for Hysterothylacium species in that environment.     

Cultivation of the Pine Wilt Nematode, Bursaphelenchus xylophilus,in Axenic Culture Media

The pine wilt nematode, Bursaphelenchus xylophilus, has been cultured axenically in vitro on soy peptone/yeast extract or modified Caenorhabditis medium supplemented with cholesterol and hemoglobin. Although growth, development and reproduction were best in soy peptone/yeast extract medium, satisfactory population size increases were observed in the chemically defined Caenorhaditis medium.     

Concepts of nematode-fungus associations in plant disease complexes: a review

Plant parasitic nematodes interact with fungi in a variety of ways to cause plant disease complexes. Even some nonplant parasitic nematodes are able to carry fungal spores internally which not only increases their mobility, but also protects them from fungicides. Plant parasitic nematodes frequently wound plants in the process of penetration and feeding. These wounds become subject to infection by fungal pathogens that require aid in penetrating their host. Other nematodes modify plant tissue in such a way that it becomes a better substrate for the fungus and thus increases their growth and reproduction to the detriment of the host. Quantitative and qualitative changes in root exudate which are induced by certain nematodes stimulate the germination, growth, and reproduction of fungal propagules in the rhizosphere. These exudates may also indirectly inhibit components of the rhizosphere microflora (e.g., actinomycetes) which are antagonistic to some plant pathogens. Depending on the species of nematode and fungus, concomitant infections may stimulate nematode reproduction (Pratylenchus-Verticillium) or inhibit reproduction (Heterodera-Fusarium).     

Role of Secondary Metabolites in Establishment of the Mutualistic Partnership between Xenorhabdus nematophila and the Entomopathogenic Nematode Steinernema carpocapsae

Xenorhabdus nematophila engages in a mutualistic partnership with the nematode Steinernema carpocapsae, which invades insects, migrates through the gut, and penetrates into the hemocoel (body cavity). We showed previously that during invasion of Manduca sexta, the gut microbe Staphylococcus saprophyticus appeared transiently in the hemocoel, while Enterococcus faecalis proliferated as X. nematophila became dominant. X. nematophila produces diverse secondary metabolites, including the major water-soluble antimicrobial xenocoumacin. Here, we study the role of X. nematophila antimicrobials in interspecies competition under biologically relevant conditions using strains lacking either xenocoumacin (ΔxcnKL strain), xenocoumacin and the newly discovered antibiotic F (ΔxcnKL:F strain), or all ngrA-derived secondary metabolites (ngrA strain). Competition experiments were performed in Grace''s insect medium, which is based on lepidopteran hemolymph. S. saprophyticus was eliminated when inoculated into growing cultures of either the ΔxcnKL strain or ΔxcnKL:F strain but grew in the presence of the ngrA strain, indicating that ngrA-derived antimicrobials, excluding xenocoumacin or antibiotic F, were required to eliminate the competitor. In contrast, S. saprophyticus was eliminated when coinjected into M. sexta with either the ΔxcnKL or ngrA strain, indicating that ngrA-derived antimicrobials were not required to eliminate the competitor in vivo. E. faecalis growth was facilitated when coinjected with either of the mutant strains. Furthermore, nematode reproduction in M. sexta naturally infected with infective juveniles colonized with the ngrA strain was markedly reduced relative to the level of reproduction when infective juveniles were colonized with the wild-type strain. These findings provide new insights into interspecies competition in a host environment and suggest that ngrA-derived compounds serve as signals for in vivo nematode reproduction.     

Compatibility of Soil Amendments with Entomopathogenic Nematodes

The impact of inorganic and organic fertilizers on the infectivity, reproduction, and population dynamics of entomopathogenic nematodes was investigated. Prolonged (10- to 20-day) laboratory exposure to high inorganic fertilizer concentrations inhibited nematode infectivity and reproduction, whereas short (1-day) exposures increased infectivity. Heterorhabditis bacteriophora was more sensitive to adverse effects than were two species of Steinernema. In field studies, organic manure resulted in increased densities of a native population of Steinernema feltiae, whereas NPK fertilizer suppressed nematode densities regardless of manure applications. Inorganic fertilizers are likely to be compatible with nematodes in tank mixes and should not reduce the effectiveness of nematodes used for short-term control as biological insecticides, but may interfere with attempts to use nematodes as inoculative agents for long-term control. Organic manure used as fertilizer may encourage nematode establishment and recycling.     

In Vitro Embryo Explant Cultures of Peanut to Evaluate Resistance to Ditylenchus destructor

Population densities of D. destructor on embryo explants of 22 peanut genotypes grown in vitro were compared with those in roots and seeds of the same genotypes grown in the greenhouse. During the first 8 weeks after inoculation, the optimum incubation period was 6 weeks for maximum reproduction of Ditylenchus destructor on embryo explants of peanut (Arachis hypogaea L. cv. Sellie) inoculated with 250 nematodes at 25 C. Nematode numbers increased 17-fold. Deletion of MnSO₄ H₂O and a higher KH₂PO₄ concentration in the medium resulted in higher nematode reproduction. Resistance or susceptibility to D. destructor was observed in seeds of several genotypes but was not matched by differences in host suitability in roots. The results indicate that the factor for resistance or susceptibility to D. destructor is synthesized in the seeds of peanut but is not translocated to the roots. Use of embryo explant cultures of peanut as a rapid method to evaluate resistance to D. destructor did not work under the conditions described.     

The Development and Influence of Meloidogyne incognita and M. javanica on Wheat

The effects of soil temperature and initial inoculum density (Pi) of Meloidogyne incognito and M. javanica on growth of wheat (Triticum aestivum cv. Anza) and nematode reproduction were studied in controlled temperature baths in the glasshouse. Nematode reproduction was directly proportional to temperature between 14 and 30 C for M. incognita and between 18 and 26 C for M. javanica. Reproduction rates (Pf/Pi, where Pf = final number of eggs) for Pi''s of 3,000, 9,000, and 30,000 eggs/plant were greatest at each temperature when Pi = 3,000. Maximum M. incognita reproduction rate (Pf/Pi = 51.12) was at 30 C. At 26 C, M. javanica reproduction (Pf/Pi = 14.82, 9.02, and 4.23 for Pi = 3,000, 9,000, and 30,000, respectively) was about half that of M. incognita when Pi = 3,000 or 9,000 but similar when Pi = 30,000. Reproduction of both species was depressed between 14 and 18 C. Shoot and root growth and head numbers were inversely related to soil temperature between 14 and 30 C but were not affected by the Pi of M. incognita when 7 d old seedlings were inoculated. When newly germinated seedlings were inoculated with M. incognita or M. javanica, the Pi did not affect shoot and root fresh weights, shoot/root ratio, and tillering, but it did reduce root dry weight (M. javanica at 26 C) and increase shoot dry weight (M. incognita at 18-22 C). The optimum temperature range is lower for wheat growth than for nematode reproduction. Wheat cv. Anza is a good host for M. incognita and M. javanica, but it is tolerant to both species.     

Modification of Resistance Expression of Phaseolus vulgaris to Meloidogyne incognita by Elevated Soil Temperatures

The effect of temperature on the reaction of susceptible (Canario Divex) and resistant (A 211) bean pure lines to Meloidogyne incognita was studied with soil temperature tanks housed in a growth chamber at 22 or 24 C. Soil temperature remained constant at 16, 22, 24, 26, 30, or 32 C in several trials. Bean line A 211 was resistant at 16 and 22 C but was susceptible at 24 C and above. Resistance to root-knot nematode reproduction was affected by a lower temperature (24 C) than was resistance to root galling (26 C) in A 211. Incubation of A 211 at 30 C for 3 and 16 days after inoculation with M. incognita resulted in a significant increase in nematode reproduction and root galling, respectively. The resistant reactions of A 211 to nematode reproduction and root galling were retained when inoculated plants were incubated at 21 C for a minimum of 16 and 23 days, respectively, prior to high temperature treatment.     

Evolutionary Conservation of Bacterial Essential Metabolic Genes across All Bacterial Culture Media

One of the basic postulates of molecular evolution is that functionally important genes should evolve slower than genes of lesser significance. Essential genes, whose knockout leads to a lethal phenotype are considered of high functional importance, yet whether they are truly more conserved than nonessential genes has been the topic of much debate, fuelled by a host of contradictory findings. Here we conduct the first large-scale study utilizing genome-scale metabolic modeling and spanning many bacterial species, which aims to answer this question. Using the novel Media Variation Analysis, we examine the range of conservation of essential vs. nonessential metabolic genes in a given species across all possible media. We are thus able to obtain for the first time, exact upper and lower bounds on the levels of differential conservation of essential genes for each of the species studied. The results show that bacteria do exhibit an overall tendency for differential conservation of their essential genes vs. their non-essential ones, yet this tendency is highly variable across species. We show that the model bacterium E. coli K12 may or may not exhibit differential conservation of essential genes depending on its growth medium, shedding light on previous experimental studies showing opposite trends.     

Survival and Reproduction of Some Nematodes as Affected by Muck and Organic Acids

Fulvic, humic, acetic, N-bulyric, formic, lactic, and propionic acids were inhibitory to the survival or reproduction of Aphelenchus avenae, Aphelenchoides goodeyi, Helicotylenchus pseudorobustus, Meloidogyne hapla or Xiphinema americanum. Reproduction of H. pseudorobustus and M. hapla significantly increased with increasing amounts of muck added to sand, and with the initial amount of nematode inoculum. All acids except humic and fulvic were lethal, in vitro, to all nematode species tested. When A. goodeyi was treated with fulvic acid, reproduction was reduced significantly when compared with sodium humate or water treatments. Treatment of H. pseudorobustus with fulvic acid (pH 3.5) resulted in a greater reduction in reproduction in soil than did treatment with humic acid (pH 3.5).     

Interrelationship of Heterodera schachtii and Meloidogyne hapla on Tomato

Invasion of tomato (Lycopersicon esculentum L.) roots by combined and sequential inoculations of Meloidogyne hapla and a tomato population of Heterodera schachtii was affected more by soil temperature than by nematode competition. Maximum invasion of tomato roots, by M. hapla and H. schachtii occurred at 30 and 26 C, respectively. Female development and nematode reproduction (eggs per plant) of M. hapla was adversely affected by H. schachtii in combined inoculations of the two nematode species. Inhibition of M. hapla development and reproduction on tomato roots from combined nematode inoculations was more pronounced as soil temperature was increased over a range of 18-30 C and with prior inoculation of tomato with H. schachtii. M. hapla minimally affected H. schachtii female development, but there was significant reduction in the buildup of H. schachtii when M. hapla inoculation preceded that of H. schachtii by 20 days.