Parasitism of Xiphinema rivesi and X. americanum by Zoosporic Fungi

Living Xiphinema americanum (Xa) and X. rivesi (Xr) extracted from soil samples and stored for 1-5 days at 4 or 20 C contained aseptate fungal hyphae. The fungi directly penetrated the nematode''s cuticle from spores encysted near the head. Penetration through the stoma, vulva, or anus was rare. Catenaria anguillulae (Cat), Lagenidium caudatura (Lag), Aphanomyces sp. (Aph), and Leptolegnia sp. (Lep) were isolated into pure culture from infected nematodes. The pathogenicity of these zoosporic fungi was determined by incubating mixed freshly extracted Xa and Xr in 2% soil extract (pH = 6.7, conductivity = 48 μmhos, 20 ± 2 C) containing zoospores obtained from single-spore isolates. After 4 days, Cat, Lag, Aph, and Lep had infected 78, 18, 13, and 22%, respectively, of the nematodes. Both Xa and Xr were infected by every fungus; however, the relative susceptibility of Xa and Xr to these fungi was not determined. All noninoculated control nematodes remained uninfected and alive. In a second experiment, parasitism of Xa and Xr by Aph and Lep was increased when nematodes were incubated in 2% soil extract for 4 days before exposure to zoospores. In a third experiment, parasitism of Xa and Xr by Cat was greater in diluted saturation soil extract (conductivity = 100-400 μmhos) than in undiluted saturation extract (conductivity = 780 μmhos). Cat produced small zoospores (4-μm-d), bulbous infection hyphae, and assimilative hyphae of varying diameters in nematodes, whereas Lag, Aph, and Lep produced large zoospores (8-μm-d) and tubular, uniform infection and assimilative hyphae in nematodes.     

Catenaria anguillulae in the mermithid nematode Romanomermis culicivorax

Disease epizootics in laboratory cultures of the mermithid nematode Romanomermis culicivorax were caused by a chytridiomycetous fungal parasite, Catenaria anguillulae. Techniques used for mass rearing the nematode provided ideal conditions for dispersal of zoospores and multiplication of the fungus; up to 90% of the postparasites of R. culicivorax were parasitized. The fungus was isolated and grown on yeast extract-soluble starch agar, and the effects of different temperature and pH regimes on the fungus were investigated as possible methods for prevention or control. A temperature suitable for mass rearing the nematodes but unsuitable for the fungus could not be found, as C. anguillulae had a broad temperature range. Adjustment of pH or a short period of chilling (0°C) were more promising control measures. At acid pH (<5), zoospore motility and germination and infection of postparasites were suppressed. The disease was controlled by rearing nematodes in water adjusted to a pH of 4.5.     

The Effects of Temperature on the Infectivity of Romanomermis culicivorax

The survival time (ST) of the preparasitic larvae of Rornanomermis culicivorax was determined by measuring motility at 1, 6, 12, 18, 21, 27, 30, and 37 C; the ST₅₀ at each of these temperatures was 2.3, 2.2, 2.0, 2.0, 1.7, 1.6, 0.9, and 0.7 days, respectively. About one-third of the preparasites infected first-instar larvae of Culex pipiens within 24 h at 27 C. The preparasites were infective at 12 to 33 C with the optimum infectivity at 21-33 C. Lower temperatures decreased the percent infectivity but increased the time that the nematodes remained infective. The time required for host infection increased as the preparasitic larvae aged at 15, 21, and 27 C.     

Species of Phytophthora and Pythium as Nematode-destroying Fungi

Pythium monospermum,, P. aphanidermatum, and Phytophthora palmivora were found to be capable of destroying certain nonstylet-bearing nematodes through endozoic parasitism by hyphae from ingested zoospores. Hyphae of P. monospermum parasitized nematode eggs but could not capture or otherwise prey upon living nematodes. We suggest that endoparasitism of free-living nematodes may be common among Oomycetes in nature.     

Coprinus comatus: A basidiomycete fungus forms novel spiny structures and infects nematode

Nematophagous basidiomycete fungi kill nematodes by trapping, endoparasitizing and producing toxin. In our studies Coprinus comatus (O.F.Müll. : Fr.) Pers. is found to be a nematode-destroying fungus; this fungus immobilizes, kills and uses free-living nematode Panagrellus redivivus Goodey and root-knot nematode Meloidogyne arenaria Neal. C. comatus produces an unusual structure designated spiny ball. Set on a sporophore-like branch, the spiny ball is a burr-like structure assembled with a large number of tiny tubes. Purified spiny balls exhibit moderate nematicidal activity. Experiments show that spiny balls are not chlamydospores because of the absence of nuclei in the structures and quick formation within 3 d in a young colony. Nematodes added to C. comatus cultures on potato-dextrose agar (PDA) and cornmeal agar (CMA) become inactive in hours. Infection of nematodes by the fungus occurs only after the nematodes are immobilized (feeble or dead), probably by a toxin. Electron micrographs illustrate that C. comatus infect P. redivivus by producing penetration pegs with which hyphae colonize nematode bodies. An infected nematode is digested and consumed within days and hyphae grow out of the nematode.     

An ultrastructural study of development and reproduction in the nematode parasite Myzocytiopsis vermicola

An isolate of Myzocytiopsis vermicola, a holocarpic parasite of Rhabditis nematodes, was studied with transmission electron microscopy (TEM) to follow development during infection, asexual and sexual reproduction. Nematodes became infected after attachment of apical cystospore buds to the nematode cuticle. Apical buds were packed with vesicles with dense fibrillar contents, which were absent from the thallus. Some thalli developed into sporangia while others became paired gametangial cells. Zoospore cleavage was often intrasporangial, although during the early stages of an epidemic partially differentiated zoospores usually were released via an exit tube into a fine vesicle. Packets of tripartite tubular hairs (TTH) were not observed in the cytoplasm of either developing or mature sporangia. TEM of sectioned material and whole mounts of zoospores revealed biflagellate zoospores, some without hairs and others with a proximal row of very short hairs on the anterior flagellum. Gametangial contact was via a short, walled fertilization tube and surplus antheridial and oogonial nuclei remained in their respective gametangial cells until disintegration of the periplasm. The mature oospores had a scalloped, electron opaque, epispore wall layer. These observations will be discussed in relation to the likely phylogenetic position of the Myzocytiopsidales within the oomycetes.     

Development and Fecundity of Reesimermis nielseni,a Nematode Parasite of Mosquitoes

Maturation of the mermithid nematode Reesimermis nielseni to the adult stage began by the tenth day after emergence of the nematodes from their hosts at ambient temperatures (24-27 C). Most postparasitic males and females reached the adult stage after 50 and 70 days, respectively. The first females exhibiting egg development and oviposition were observed 25-30 days after emergence, but some oviposition was still taking place 150 days later. Reesimermis nielseni laid an average 2,480 eggs per female over an 18-day oviposition period. A majority of the mature eggs hatched within 7 h after the cultures were flooded. The preparasites are short-lived, and only a few were able to infect exposed hosts after 72 h.     

System optimization for attachment of Pasteuria penetrans to Meloidogyne incognita

For optimal mass production of Pasteuria penetrans in vivo, it is important to develop a system that can ensure 100% nematode attachment of the bacteria and high bacterial infection after inoculation. In this study, effects of endospore concentration and centrifugation parameters on attachment were investigated, followed by evaluation of impacts of centrifugation on endospore dislodgement, Meloidogyne incognita juvenile (J2) mortality, J2 infectivity, and bacterial infectivity. Endospore concentration and percentage of attachment fit well to mass-action and logit models, with the former being superior. Centrifugation had no impact on J2 mortality but had a great impact on endospore dislodgement in sand and water, nematode infectivity and bacterial infectivity. At nematode concentration of 2×10³ J2/mL, the optimal system for endospore attachment was developed which consisted of bacteria at 2×10⁴ endospores/mL, and centrifugation at 9000×g for 3 min three times. This system generated 100% attachment with approximately seven endospores/J2. After inoculation of treated nematodes to tomato plants, the inoculum yielded 47% bacterial infection, superior to 17% infection observed in centrifugation at 6000×g. Endospore dislodgement occurred after placing the centrifuged inoculated nematodes in sand or water for 24 and 48 h, which was more severe in centrifugation at 6000 than at 9000×g. Results also indicated that centrifugation led to lower nematode infectivity, regardless of endospore presence and centrifugation at 9000 or 6000×g, compared with the no centrifugation control.     

Susceptibility of Psorophora columbiae Larvae over Time to Parasitism by Romanomermis culicivorax

The ability of Romanomermis culicivorax preparasites to penetrate and infect Psorophora columbiae decreased substantially after ca. 28 hours. Parasitism at temperatures typical of Louisiana rice fields (i.e., 26, 29, and 32 ± 0.5 C) showed a significant linear decrease (P < 0.01) as the percentage of older larval instars increased at the times of exposure. These data emphasize the need for a synchronous field application of preparasites to challenge the rapid development of early instars of Ps. columbiae. Applications of postparasites rather than insecticide treatments to potential mosquito breeding habitats may offer greater flexibility in larval mosquito control programs.     

A new species of catenaria parasitic on nematodes of sugarcane

Summary A new species,Catenaria vermicola, is named and described. It attacks several plant-parasitic nematodes of economic importance in sugarcane soils of Louisiana. The fungus forms numerous zoospores which encyst on the cuticle, generally near natural openings of the nematode, thereby causing infection. Within the parasitized nematodes hyphae and sporangia quickly form and destroy the victim completely. The following plant-parasitic nematodes were attacked and destroyed:Belonolaimus longicaudatus, Criconemoides rusticum, Hemicycliophora gigas, Hoplolaimus tylenchiformis, Meloidogyne larvae,Pratylenchus brachyurus, P. zeae, Radopholus similis, Tylenchulus semipenetrans larvae,Tylenchorhynchus martini, andXiphinema chambersi.     

Carbohydrate-mediated Recognition of Larval Mosquito Hosts by Romanomermis culicivorax

Proteases, glycosidases, and lectins were tested and the results supported a role in host recognition for glycoproteins containing β-glucose and α-mannose on the cuticular surface of host and parasite. Carbohydrates containing α-glucose, galactose, fucose, or N-acetylglucosamine residues apparently are not involved in nematode attachment. Chitin or a related N-acetylglucosamine polymer was found in R. culicivorax preparasites. Treatment of preparasites with neuraminidase, which hydrolyzes sialic acids, increased nematode attachment to Anopheles freeborni larvae.     

Differential Adhesion and Infection of Nematodes by the Endoparasitic Fungus Meria coniospora (Deuteromycetes)

The conidia of the endoparasitic fungus Meria coniospora (Deuteromycetes) had different patterns of adhesion to the cuticles of the several nematode species tested; adhesion in some species was only to the head and tail regions, on others over the entire cuticle, whereas on others there was a complete lack of adhesion. After adhesion, the fungus usually infected the nematode. However, adhesion to third-stage larvae of five animal parasitic nematodes, all of which carry the cast cuticle from the previous molt, did not result in infection. M. coniospora infected animal parasitic nematodes when this protective sheath was removed. Seven preparations of sialic acid (N-acetylneuraminic acid) gave three types of response in adhesion-infection of nematodes: (i) a significant reduction in conidial adhesions; (ii) no interference with adhesion, but a 10-day delay in infection; and (iii) a delay in infection by 2 to 3 days. The current results support previous findings indicating involvement of sialic acids localized on nematode cuticles in recognition of prey by M. coniospora.     

Life cycle of the endoparasitic nematophagous fungus Meria coniospora: a light and electron microscopic study

The obligate endoparasitic fungus Meria coniospora lives its entire vegetative life within infected nematodes. Conidia of M. coniospora infect the nematode Panagrellus redivivus mainly in the mouth region. The infection, starting with adhesion of conidia to the nematode surface, growth of trophic hyphae, production of conidiophores and conidia, was followed using light, scanning and transmission electron microscopy.This work was supported by grants from the Swedish Natural Science Research Council.     

Anfälligkeit von entomopathogenen Nematoden gegenüber nematodenfangenden und endoparasitären Pilzen

Abstract Susceptibility of entomopathogenic nematodes to nematode-trapping and endoparasitic fungi
Laboratory tests in petri dishes demonstrated that the nematode-trapping fungi Arthrobotrys superba and A. robusta preyed upon the entomopathogenic nematodes Steinernema bibionis, S. feltiae and a species of Heterohabditis. Up to 100% of the nematodes were trapped 48 h after application. Large differences existed in the time required for trapping as well as for mortality after trapping and was dependent on the fungal or nematode species studied. Arthrobotrys robusta , reduced S. bibionis densities in sand-block bioassay chambers. The prior introduction of a microphagous nematode, Panagrellus redivivus , to induce trap formation caused a 52% reduction in S. bibionis levels.
The endoparasitic fungi Harposporium anguillulae and Drechmeria coniospora did not parasitize the entomopathogenic nematodes in petri dish tests. However, Verticillium balanoides , was shown to be a parasite of S. bibionis. Predivivus and a mycophagous species of Ditylenchus were more quickly parasitized than S. bibionis , with parasitism reaching 100, 90 and 30% after 42 days, respectively. In the sand-block chambers V. balanoides did not reduce the, number of recovered S. bibionis juveniles after 14 or 28 days.     

The effect of fungal parasitism and predation on the population dynamics of nematodes in the activated sludge process

Nematophagous fungi, both predators and endoparasites, were found to be common components of activated sludge. Although rotifers and ciliate protozoa (both potential prey) were also abundant, no fungi were parasitic on these organisms. Endoparasitic fungi, which were far more abundant than predators, were able to infect nematodes and complete their life cycles successfully. Neither of the predatory fungi observed were able to produce conidia: an unidentified net-forming species lived saprophytically and failed to capture any prey, although it played a minor role in the formation of microbial flocs, and a single conidium of Dactylella mammillata was observed to capture a nematode by spontaneous trap formation. Several endoparasites were recorded although single species dominated each sludge examined; these were Meria coniospora and Catenaria anguillulae. Both endoparasites were related to the population dynamics of the nematodes with 100% of the nematode population becoming infected at certain times. Clear predator (parasitetprey (host) associations were discernible and these are discussed.     

Glomus fasciculatum,a Weak Pathogen of Heterodera glycines

The occurrence ofchlamydospores of Glomus fasciculatum (Gf) within cysts of the soybean cyst nematode, Heterodera glycines, and the effects of vesicular-arbuscular mycorrhizae on nematode population dynamics and soybean (Glycine max) plant growth were investigated. Chlamydospores occupied 1-24% of cysts recovered from field soil samples. Hyphae of Missouri isolate Gfl penetrated the female nematode cuticle shortly after she ruptured the root epidermis. Convoluted hyphae filled infected eggs, and sporogenesis occurred within infected eggs. G. microcarpum, G. mosseae, and two isolates of Gf were inoculated with H. glycines on plants of ''Essex'' soybeans. Each of the two Gf isolates infected about 1% of the nematode eggs in experimental pot cuhures. The Gfl isolate decreased the number of first-generation adult females 26%, compared with the nonmycorrhizal control. The total numbers of first-generation plus second-generation adult females were similar for both Gf isolates and 29-41% greater than the nonmycorrhizal control. Soybean plants with Gf and H. glycines produced more biomass than did nonmycorrhizal plants with nematodes, but only Gfl delayed leaf senescence.     

An electron-microscopical analysis of capture and initial stages of penetration of nematodes byArthrobotrys oligospora

A detailed analysis was made of the capture and subsequent penetration of nematodes by the nematophagous fungusArthrobotrys oligospora using different electron-microscopical techniques. Capture of nematodes by this fungus occurred on complex hyphal structures (traps) and was effectuated by an adhesive coating, present on these trap cells. The adhesive layer was largely fibrillar in nature and was absent on cells of normal hyphae. Following capture, penetration hyphae were formed at those sites where the trap cell wall was anchored to the nematode cuticle by the adhesive. New walls of these hyphae were formed underneath the original trap cell walls, which were partly hydrolysed to allow growth and development of the penetration tubes through the adhesive coating towards the cuticle. Our observations indicated that the cuticle of the nematode was subsequently penetrated by the penetration tubes by mechanical means. After penetration a large infection bulb was formed from which trophic hyphae arose. Cytochemical experiments indicated that the sites of penetration of the cuticle were intensely stained for acid phosphatase activity. At later stages of infection activity of this enzyme was present throughout the nematode contents; the enzyme was most probably secreted by complex membranous structures associated with the cytoplasmic membrane of the infection bulb and the trophic hyphae.     

Effects of Monoclonal Antibodies,Cationized Ferritin,and Other Organic Molecules on Adhesion of Pasteuria penetrans Endospores to Meloidogyne incognita

The incidence of adhesion of Pasteuria penetrans endospores to Meloidogyne incognita second-stage juveniles (J2) was studied after pretreatment of the latter with monoclonal antibodies (MAb), cationized ferritin, and other organic molecules in replicated trials. Monoclonal antibodies developed to a cuticular epitope of M. incognita second-stage juveniles gave significant reductions in attachment of P. penetrans endospores to treated nematodes. MAb bound to the entire length of J2 except for the area of the lateral field, where binding was restricted to the incisures. Since reductions in attachment with MAb treatment were modest, it is uncertain if these results implicated a specific surface protein as a factor that interacted in binding of the endospore to the nematode cuticle. Endospore attachment was decreased following treatment of the nematode with the detergents sodium dodecyl sulfate (SDS) and cetyltrimethylammonium bromide (CTAB). Endospore attachment to live nematodes was significantly greater than attachment to dead nematodes. Attachment rates of three P. penetrans isolates to M. incognita race 3 varied between isolates. The effects of neuraminidase, pronase, pepsin, trypsin, lipase, and Na periodate on endospore attachment were inconsistent. The cationic dye alcian blue, which binds sulfate and carboxyl groups on acidic glycans, had no consistent effect on endospore attachment. The incidence of endospore attachment was significantly lower but modest, at best, for nematodes that were treated with cationized ferritin alone or cationized ferritin following monoclonal antibody. The lack of consistency or extreme reduction in most experiments suggests that attachment of P. penetrans spores to M. incognita is not specified by only one physico-chemical factor, but may involve a combination of at least two physico-chemical factors (including surface charge and movement of the J2). This points to a need for analysis of combined or factorial treatment effects.     

Signatures of selection in sheep bred for resistance or susceptibility to gastrointestinal nematodes

Background

Gastrointestinal nematodes are one of the most serious causes of disease in domestic ruminants worldwide. There is considerable variation in resistance to gastrointestinal nematodes within and between sheep breeds, which appears to be due to underlying genetic diversity. Through selection of resistant animals, rapid genetic progress has been demonstrated in both research and commercial flocks. Recent advances in genome sequencing and genomic technologies provide new opportunities to understand the ovine host response to gastrointestinal nematodes at the molecular level, and to identify polymorphisms conferring nematode resistance.

Results

Divergent lines of Romney and Perendale sheep, selectively bred for high and low faecal nematode egg count, were genotyped using the Illumina® Ovine SNP50 BeadChip. The resulting genome-wide SNP data were analysed for selective sweeps on loci associated with resistance or susceptibility to gastrointestinal nematode infection. Population differentiation using F_ST and Peddrift revealed sixteen regions, which included candidate genes involved in chitinase activity and the cytokine response. Two of the sixteen regions identified were contained within previously identified QTLs associated with nematode resistance.

Conclusions

In this study we identified fourteen novel regions associated with resistance or susceptibility to gastrointestinal nematodes. Results from this study support the hypothesis that host resistance to internal nematode parasites is likely to be controlled by a number of loci of moderate to small effects.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-637) contains supplementary material, which is available to authorized users.     

Factors influencing the sporulation and cyst formation of Aphanomyces invadans, etiological agent of ulcerative mycosis in Atlantic menhaden, Brevoortia tyrannus

Oomycete infections caused by Aphanomyces invadans occur in freshwater and estuarine fishes around the world. Along the east coast of the USA, skin ulcers caused by A. invadans are prevalent in Atlantic menhaden, Brevoortia tyrannus. From laboratory observations low salinities appear crucial to transmission of the pathogen. To better understand aspects of transmission, we characterized sporulation and cyst formation of secondary zoospores of two isolates of A. invadans at different salinities and temperatures. Sporulation occurred only at low salinities. At room temperature (ca. 20-22 C), using "pond water" augmented with artificial sea salts, the endemic strain WIC and the Thailand strain PA7 of A. invadans produced free-swimming secondary zoospores at salinities of 0, 1 and 2 psu (practical salinity unit = per thousand), but not at 4 psu or higher. Secondary zoospores of another species, ATCC-62427 (Aphanomyces sp.), were observed at 1, 2, 4 and 8 psu but not at 0 and 12 psu. Secondary zoospores of all three isolates, especially WIC, were abundant and motile 1-2 d postsporulation. Sporulation was temperature dependent and occurred over a relatively narrow range. No sporulation occurred at 4, 30 or 35 C for either WIC or PA7. For both strains zoospore production within 1-3 d after the initiation of sporulation was more prolific at 25 C than at 20 and 15 C. At 15 C production of zoospores was sustained over 11 d for WIC and 5 d for PA7. At room temperature single WIC secondary zoospores remained motile 12-18 h. Salinities exceeding 4 psu or vigorous shaking caused immediate cyst formation of WIC secondary zoospores. Exposure to menhaden tissue, but not tissues of other fishes to secondary zoospores (WIC), caused rapid (2 h) cyst formation. Cysts were capable of excysting when transferred to 1 psu water within 2-3 h of cyst formation. Cysts that had remained encysted in 6.5 psu for 24 h did not excyst when transferred to 1 psu water. Salinity and temperature requirements for sporulation indicate that juvenile menhaden must acquire infections during rain or in low salinity oligohaline waters.