Bacillus thuringiensis (Bt) crystal proteins are pore-forming toxins used as insecticides around the world. Previously, the extent to which these proteins might also target the invertebrate phylum Nematoda has been mostly ignored. We have expressed seven different crystal toxin proteins from two largely unstudied Bt crystal protein subfamilies. By assaying their toxicity on diverse free-living nematode species, we demonstrate that four of these crystal proteins are active against multiple nematode species and that each nematode species tested is susceptible to at least one toxin. We also demonstrate that a rat intestinal nematode is susceptible to some of the nematicidal crystal proteins, indicating these may hold promise in controlling vertebrate-parasitic nematodes. Toxicity in nematodes correlates with damage to the intestine, consistent with the mechanism of crystal toxin action in insects. Structure-function analyses indicate that one novel nematicidal crystal protein can be engineered to a small 43-kDa active core. These data demonstrate that at least two Bt crystal protein subfamilies contain nematicidal toxins.

In most free-living eukaryotes studied thus far, heme is synthesized from a series of intermediates through a well defined evolutionarily conserved pathway. We found that free-living worms, including the model genetic organism Caenorhabditis elegans, and parasitic helminths are unable to synthesize heme de novo, even though these animals contain hemoproteins that function in key biological processes. Radioisotope, fluorescence labeling, and heme analog studies suggest that C. elegans acquires heme from exogenous sources. Iron-deprived worms were unable to grow in the presence of adequate heme unless rescued by increasing heme levels in the growth medium. These data indicate that although worms use dietary heme for incorporation into hemoproteins, ingested heme is also used as an iron source when iron is limiting. Our results provide a biochemical basis for the dependence of worm growth and development on heme, and they suggest that pharmacologic targeting of heme transport pathways in worms could be an important control measure for helminthic infections.Caenorhabditis elegans ͉ iron ͉ metals ͉ nematode ͉ prophyrin

1 The species assemblages and abundance of phoretic mites and nematodes associated with the elm bark beetles, Scolytus multistriatus and Scolytus pygmaeus, were studied in Austria. 2 A total of 3922 individual mites were recorded from 144 adults of S. multistriatus and 178 adults of S. pygmaeus. The species spectrum was identical and the relative abundance of mites was very similar for both species of scolytids. Nine mite species, Pyemotes scolyti, Pseudotarsonemoides eccoptogasteri, Trichouropoda bipilis, Tarsonemus crassus, Proctolaelaps eccoptogasteris, Proctolaelaps scolyti, Chelacheles michalskii, nr. Eueremaeus sp. and Elattoma sp. were detected. Two of the nine species, nr. Eueremaeus sp. and Elattoma sp., are documented here as new associates of Scolytus spp. 3 Pyemotes scolyti was the most frequent mite species, and Ps. eccoptogasteri and T. bipilis were relatively common, whereas the other mites occurred occasionally or were rare. 4 The trophic roles of most of the mites associated with S. multistriatus and S. pygmaeus are poorly known, but they may include fungivores, parasitoids of bark beetle broods, predators of bark beetle broods and/or mites and/or nematodes. 5 Besides phoretic mites, two nematode associates were seen on the investigated insects. A species of Cryptaphelenchus occurred under the elytra of both scolytid species, whereas the adults of a Neoparasitylenchus sp. were present inside abdomens of S. multistriatus, but absent from S. pygmaeus.

Symptoms of beech leaf disease (BLD), first reported in Ohio in 2012, include interveinal greening, thickening and often chlorosis in leaves, canopy thinning and mortality. Nematodes from diseased leaves of American beech (Fagus grandifolia) sent by the Ohio Department of Agriculture to the USDA, Beltsville, MD in autumn 2017were identified as the first recorded North American population of Litylenchus crenatae (Nematology, 21, 2019, 5), originally described from Japan. This and other populations from Ohio, Pennsylvania and the neighbouring province of Ontario, Canada showed some differences in morphometric averages among females compared to the Japanese population. Ribosomal DNA marker sequences were nearly identical to the population from Japan. A sequence for the COI marker was also generated, although it was not available from the Japanese population. The nematode was not encountered in Fagus crenata (its host in Japan) living among nematode-infested Fagus grandifolia in the Holden Arboretum, nor has L. crenatae been reported in American beech in Japan. The morphological and host range differences in North American populations are nomenclaturally distinguished as L. crenatae mccannii ssp. n. from the population in Japan. Low-temperature scanning electron microscopy (LT-SEM) demonstrated five lip annules and a highly flexible cuticle. Females, juveniles and eggs were imaged within buds with a Hirox Digital microscope and an LT-SEM. Nematodes swarmed to the tips of freshly cut beech buds, but explants could not be maintained. Inoculation of fresh nematodes from infested leaves or buds to buds or leaves ofF. grandifolia seedlings resulted in BLD leaf symptoms. Injuring dormant buds prior to nematode application, in fall or spring, promoted the most reliable symptom expression. The biogeography and physiology of anguinid nematode leaf galling, and potential co-factors and transmission are discussed. K E Y W O R D S Anguinidae, digital and scanning electron microscopy, foliar nematode, host range, molecular identification, morphometrics, new continent detection, new subspecies, symptom transmission, taxonomy 2 of 15 | CARTA eT Al.

Plant-parasitic nematodes (PPNs) are important pests that cause an estimated ten billion dollars of crop loss each year in the United States and over 100 billion dollars globally. The Animal and Plant Health Inspection Service (APHIS) within the U.S. Department of Agriculture maintains and updates the U.S. Regulated Plant Pest list. Currently, the number of PPNs regulated by APHIS includes more than 60 different species. This review focuses on the top ten most economically important regulated and emerging plant-parasitic nematodes and summarizes the diagnostics of morphological and some molecular features for distinguishing them. These ten major previously described nematode species are associated with various economically important crops from around the world. This review also includes their current distribution in the U.S. and a brief historical background and updated systematic position of these species. The species included in this review include three PPNs considered by the U.S. Department of Agriculture as invasive invertebrates Globodera pallida, Globodera rostochiensis, and Heterodera glycines; four regulated PPNs, namely Bursaphelenchus xylophilus, Meloidogyne fallax, Ditylenchus dipsaci, and Pratylenchus fallax; and the three emerging PPNs Meloidogyne chitwoodi, Meloidogyne enterolobii, and Litylenchus crenatae mccannii.

In 2006, a cyst nematode was discovered in tare dirt at a potato (Solanum tuberosum) processing facility in eastern Idaho. The nematode was found during a routine survey conducted jointly by the Idaho State Department of Agriculture and the USDA Animal and Plant Health Inspection Service through the Cooperative Agricultural Pest Survey program. Extensive additional sampling from two suspect fields led to the identification of the same nematode in a 45-acre (18.2-ha) field located in northern Bingham County. The morphology of cysts and second-stage juveniles and molecular analyses established the identity of the species as the pale cyst nematode Globodera pallida (Stone 1973) Behrens 1975. Morphological characters used for identification included cyst shape, characteristics of cyst terminal cone including nature of fenestration, cyst wall pattern, anal-vulval distance, number of cuticular ridges between anus and vulva, and Granek's ratio (1,4). The second-stage juvenile morphologies critical for identification were the following: body and stylet length, shape of stylet knobs, shape and length of tail and hyaline tail terminus, and number of refractive bodies in the hyaline part of tail (1,4). Diagnosis as G. pallida was clearly confirmed by two molecular tests. First, PCR-RFLP (restriction fragment length polymorphism) profiles of a ribosomal DNA fragment using restriction enzymes RsaI, TaqI, and AluI (2) were consistent with a G. pallida control and not G. rostochiensis. Second, the ribosomal DNA region that extends from the 3′ end of the 18S ribosomal subunit and includes all of ITS1, 5.8S, and ITS2 to the 5′ end of the 28S ribosomal subunit was used to generate sequence for the most accurate species determination. Sequences obtained from three individual juveniles were compared with those from several Globodera species (3), revealing unequivocal similarity to G. pallida. This detection represents a new country record for G. pallida in the United States. Collection of additional information regarding distribution of this nematode within the region is underway. References: (1) J. G. Baldwin and M. Mundo-Ocampo. Heteroderinae, Cyst- and Non-cyst-forming Nematodes. Pages 275-362 in: Manual of Agricultural Nematology. W. R. Nickle, ed. Marcel Dekker, New York, 1991. (2) V. C. Blok et al. J. Nematol. 30:262, 1998. (3) L. A. Pylypenko et al. Eur. J. Plant Pathol. 111:39, 2005. (4) A. R. Stone. Nematologica 18:591, 1973.

Diploscapter, a thermotolerant, free-living soil bacterial-feeding nematode commonly found in compost, sewage, and agricultural soil in the United States, was studied to determine its potential role as a vehicle of Salmonella enterica serotype Poona, enterohemorrhagic Escherichia coli O157:H7, and Listeria monocytogenes in contaminating preharvest fruits and vegetables. The ability of Diploscapter sp. strain LKC25 to survive on agar media, in cow manure, and in composted turkey manure and to be attracted to, ingest, and disperse food-borne pathogens inoculated into soil or a mixture of soil and composted turkey manure was investigated. Diploscapter sp. strain LKC25 survived and reproduced in lawns of S. enterica serotype Poona, E. coli O157:H7, and L. monocytogenes on agar media and in cow manure and composted turkey manure. Attraction of Diploscapter sp. strain LKC25 to colonies of pathogenic bacteria on tryptic soy agar within 10, 20, 30, and 60 min and 24 h was determined. At least 85% of the worms initially placed 0.5 to 1 cm away from bacterial colonies migrated to the colonies within 1 h. Within 24 h, >90% of the worms were embedded in colonies. The potential of Diploscapter sp. strain LKC25 to shed pathogenic bacteria after exposure to bacteria inoculated into soil or a mixture of soil and composted turkey manure was investigated. Results indicate that Diploscapter sp. strain LKC25 can shed pathogenic bacteria after exposure to pathogens in these milieus. They also demonstrate its potential to serve as a vector of food-borne pathogenic bacteria in soil, with or without amendment with compost, to the surface of preharvest fruits and vegetables in contact with soil.

An unusual population of cyst nematode was found in soils collected from a Powell Butte, OR field with a cropping history including potato, wheat, other crops, and significant weed presence. These nematodes could not be placed with certainty into any known species and exhibited some unique morphological features in some specimens. Compared with Globodera pallida, the cyst body length was slightly longer and the second-stage juvenile stylet length was slightly shorter. In some individuals, the J2 stylet knob height was greater and the tail annules were more prominent than in G. pallida, and the tail abruptly narrowed, with a slight constriction near the posterior third of the hyaline terminus. Compared with G. rostochiensis, the hyaline tail terminus had a larger number of refractive bodies, and cysts of this population had a smaller Granek's ratio and fewer cuticular ridges between the anus and vulva. In some individuals, the tail termini of second-stage juveniles were more bluntly pointed, and the stylet knobs were more anteriorly directed with greater height. Unlike G. tabacum, the cyst wall often lacked a network-like pattern and, in some individuals, the juvenile tail terminus distinctly narrowed after a constriction. Molecularly, the population was distinct from G. pallida, G. rostochiensis, and G. tabacum. Multiplex polymerase chain reaction of the internal transcribed spacer (ITS) rDNA region gave results similar to G. tabacum; however, ITS restriction fragment length polymorphism patterns were observed to have individual bands in common with G. rostochiensis and G. pallida. Phylogenetic analysis based on ITS1 and -2 rDNA sequences showed greatest similarity to populations from Argentina and Chile; together, they form a moderately supported clade, distinct from G. rostochiensis, G. tabacum, G. "mexicana," European type G. pallida, and several G. pallida populations from South America.