Hoss S et al. (2012) Environ Toxicol Chem "Interlaboratory comparison of a standardized toxicity test using the nematode Caenorhabditis elegans (ISO 10872)."

Pluta HJ, Bergtold M, Bluebaum-Gronau E, Menzel R, Hoss S, Mohlenkamp C, Traunspurger W, Brinke M, Ratte HT, Ahlf W, von Danwitz B, Donnevert G

[Environ Toxicol Chem, 2012]

A ring test was carried out within the standardization process of ISO 10872 to evaluate the precision of the toxicity test for the nematode Caenorhabditis elegans. Eight different laboratories tested aqueous solutions of the reference substance benzylcetyldimethylammonium chloride as well as native sediments and soils for toxic effects on the growth and reproduction of C. elegans. Validity criteria were met in all laboratories. Average median- and low-effect concentrations were determined to be 15.1 mg L(-1) (EC50) and 8.7 mg L(-1) (EC10) for growth and 7.5 mg L(-1) (EC50) and 3.8 mg L(-1) (EC10) for reproduction of C. elegans, with ECx values showing a high degree of reproducibility (CV(R) : <21% and <11% for EC10 and EC50, respectively) and repeatability (CV(r) : <20% and <7% for EC10 and EC50, respectively). The toxic effects of the sediments and soils revealed by the different laboratories were well related to each samples' degree of chemical contamination. Moreover, the effects showed an acceptable reproducibility (CV(R) : 5-33% and 0-28% for growth and reproduction, respectively) and repeatability (CV(r) : 3-13% and 0-12% for growth and reproduction, respectively). The present study confirms that the toxicity test with C. elegans according to ISO 10872 is a reliable and precise tool to assess the toxicity of aqueous media, freshwater sediments, and soils.

Crossgrove K et al. (2008) Mol Biochem Parasitol "Orthologues of the Drosophila melanogaster E75 molting control gene in the filarial parasites Brugia malayi and Dirofilaria immitis."

Lochner MC, Maina CV, Robinson-Rechavi M, Crossgrove K

[Mol Biochem Parasitol, 2008]

Filarial parasites cause debilitating diseases in humans and domesticated animals. Brugia malayi and Dirofilaria immitis are transmitted by mosquitoes and infect humans and dogs, respectively. Their life cycle is punctuated by a series of cuticular molts as they move between different hosts and tissues. An understanding of the genetic basis for these developmental transitions may suggest potential targets for vaccines or chemotherapeutics. Nuclear receptor (NR) proteins have been implicated in molting in the free-living nematode Caenorhabditis elegans and have well characterized roles in molting during larval development of Drosophila melanogaster. For example, the D. melanogaster E75 (NR1D3) NR gene is required for molting and metamorphosis, as well as egg chamber development in adult females. We have identified Bm-nhr-11and Di-nhr-6, B. malayi and D. immitis orthologues of E75. Both genes encode canonical nuclear receptor proteins, are developmentally regulated, and are expressed in a sex-specific manner in adults.

Sluder AE et al. (1999) Genome Res "The nuclear receptor superfamily has undergone extensive proliferation and diversification in nematodes."

Mathews SW, Hough DM, Sluder AE, Yin VP, Maina CV

[Genome Res, 1999]

The nuclear receptor (NR) superfamily is the most abundant class of transcriptional regulators encoded in the Caenorhabditis elegans genome, with >200 predicted genes revealed by the screens and analysis of genomic sequence reported here. This is the largest number of NR genes yet described from a single species, although our analysis of available genomic sequence from the related nematode Caenorhabditis briggsae indicates that it also has a large number. Existing data demonstrate expression for 25% of the C. elegans NR sequences. Sequence conservation and statistical arguments suggest that the majority represent functional genes. An analysis of these genes based on the DNA-binding domain motif revealed that several NR classes conserved in both vertebrates and insects are also represented among the nematode genes, consistent with the existence of ancient NR classes shared among most, and perhaps all, metazoans. Most of the nematode NR sequences, however, are distinct from those currently known in other phyla, and reveal a previously unobserved diversity within the NR superfamily. In C. elegans, extensive proliferation and diversification of NR sequences have occurred on chromosome V, accounting for > 50% of the predicted NR genes.

Gissendanner CR et al. (2004) Dev Biol "Expression and function of conserved nuclear receptor genes in Caenorhabditis elegans."

Maina CV, Crossgrove K, Gissendanner CR, Kraus KA, Sluder AE

[Dev Biol, 2004]

The Caenorhabditis elegans genome encodes 284 nuclear receptor (NR) genes. Among these 284 NR genes are 15 genes conserved among the Metazoa. Here, we analyze the, expression and function of eight heretofore uncharacterized conserved C elegans NR genes. Reporter gene analysis demonstrates that these genes have distinct expression patterns and that a majority of the C elegans cell types express a conserved NR gene. RNA interference with NR gene function resulted in visible phenotypes for three of the genes, revealing functions in various processes during postembryonic development. Five of the conserved NR genes are orthologs of NR genes that function during molting and metamorphosis in insects. Functional studies confirm a role for most of these 'ecdysone cascade' NR orthologs during the continuous growth and dauer molts. Transcript levels for these genes fluctuate in a reiterated pattern during the molting cycles, reminiscent of the expression hierarchy observed in the insect ecdysone response. Together, these analyses provide a foundation for further dissecting the role of NRs in nematode development as well as for evaluating conservation of NR functions among the Metazoa.

Robinson-Rechavi M et al. (2005) J Mol Evol "Explosive lineage-specific expansion of the orphan nuclear receptor HNF4 in nematodes."

Robinson-Rechavi M, Maina CV, Gissendanner CR, Laudet V, Sluder A

[J Mol Evol, 2005]

The nuclear receptor superfamily expanded in at least two episodes: one early in metazoan evolution, the second within the vertebrate lineage. An exception to this pattern is the genome of the nematode Caenorhabditis elegans, which encodes more than 270 nuclear receptors, most of them highly divergent. We generated 128 cDNA sequences for 76 C. elegans nuclear receptors, confirming that these are active genes. Among these numerous receptors are 13 orthologues of nuclear receptors found in arthropods and/or vertebrates. We show that the supplementary nuclear receptors (supnrs) originated from an explosive burst of duplications of a unique orphan receptor, HNF4. This origin has specific implications for the role of ligand binding in the function and evolution of the nematode supplementary nuclear receptors. Moreover, the supplementary nuclear receptors include a group of very rapidly evolving genes found primarily on chromosome V. We propose a model of lineage-specific duplications from a chromosome on which duplication and substitution rates are highly increased. Our results provide a framework to study nuclear receptors in nematodes, as well as to consider the functional and evolutionary consequences of lineage-specific duplications.

Gissendanner CR et al. (2008) Dev Biol "The Caenorhabditis elegans NR4A nuclear receptor is required for spermatheca morphogenesis."

Kelley K, Nguyen TQ, Hoener MC, Gissendanner CR, Maina CV, Sluder AE

[Dev Biol, 2008]

The gene nhr-6 encodes the Caenorhabditis elegans ortholog of the NR4A nuclear receptor. We determined the biological functions of NHR-6 through the isolation and characterization of a deletion allele of nhr-6, lg6001. We demonstrate that nhr-6 has an essential role in the development of the C. elegans somatic gonad. Specifically, nhr-6 is required for the development of the hermaphrodite spermatheca, a somatic gonad organ that serves as the site of sperm storage and oocyte fertilization. Using a variety of spermatheca cell markers, we have determined that loss of nhr-6 function causes severe morphological defects in the spermatheca and associated spermathecal valves. This appears to be due to specific requirements for nhr-6 in regulating cell proliferation and cell differentiation during development of these structures. The improper development of these structures in nhr-6(lg6001) mutants leads to defects in ovulation and significantly reduced fecundity of C. elegans hermaphrodites. The phenotypes of nhr-6(lg6001) mutants are consistent with a role for nhr-6 in organogenesis, similar to the functions of its mammalian homologs.

Zhao X et al. (2000) Phytopathology "Species-dependent effects of border cell and root tip exudates on nematode behavior."

Zhao X, Hawes MC, Schmitt M

[Phytopathology, 2000]

ABSTRACT Effects of border cell and root tip exudates on root knot nematode (Meloidogyne incognita) behavior were examined. In whole-plant assays using pea, M. incognita second-stage juveniles (J2) accumulated rapidly around the 1- to 2-mm apical region ensheathed by border cells, but not in the region of elongation. Within 15 to 30 min, J2 which had accumulated within detached clumps of border cells lost motility and entered into a quiescent state. When border cells (and associated root tip exudates) were washed from pea roots prior to challenge with nematodes, no such accumulation and quiescence was induced. Attraction of nematodes by roots was species dependent: no attraction or accumulation occurred in snap bean. Using a quantitative assay, three categories of chemotaxis responses occurred: attraction (pea and alfalfa cv. Thor), repulsion (alfalfa cv. Moapa 69), and no response (snap bean and alfalfa cv. Lahonton). In contrast, total root tip exudates from all three plant species acted as a repellent for M. incognita in the sand assay. An in vitro assay was developed to characterize the induced quiescence response. When total root tip exudate from the tested legumes (as well as corn) was incubated with J2 populations, >80% of the nematodes lost motility. A similar response occurred in Caenorhabditis elegans. Border cell exudates did not induce or contribute to the induction of quiescence. Cocultivation of pea border cells with M. incognita resulted in changes in border cell shape similar to those observed in response to exogenous plant hormones. No such changes occurred in snap bean border cells. Understanding the cell- and host-specific extracellular recognition that occurs between roots and pathogenic nematodes in the early stages before infection occurs could lead to new avenues for disease control.

Tzertzinis G et al. (2010) PLoS Negl Trop Dis "Molecular evidence for a functional ecdysone signaling system in Brugia malayi."

Liu C, Maina CV, Palli SR, Gissendanner CR, Tzertzinis G, Unnasch TR, Egana AL, Robinson-Rechavi M

[PLoS Negl Trop Dis, 2010]

BACKGROUND: Filarial nematodes, including Brugia malayi, the causative agent of lymphatic filariasis, undergo molting in both arthropod and mammalian hosts to complete their life cycles. An understanding of how these parasites cross developmental checkpoints may reveal potential targets for intervention. Pharmacological evidence suggests that ecdysteroids play a role in parasitic nematode molting and fertility although their specific function remains unknown. In insects, ecdysone triggers molting through the activation of the ecdysone receptor: a heterodimer of EcR (ecdysone receptor) and USP (Ultraspiracle). METHODS AND FINDINGS: We report the cloning and characterization of a B. malayi EcR homologue (Bma-EcR). Bma-EcR dimerizes with insect and nematode USP/RXRs and binds to DNA encoding a canonical ecdysone response element (EcRE). In support of the existence of an active ecdysone receptor in Brugia we also cloned a Brugia rxr (retinoid X receptor) homolog (Bma-RXR) and demonstrate that Bma-EcR and Bma-RXR interact to form an active heterodimer using a mammalian two-hybrid activation assay. The Bma-EcR ligand-binding domain (LBD) exhibits ligand-dependent transactivation via a GAL4 fusion protein combined with a chimeric RXR in mammalian cells treated with Ponasterone-A or a synthetic ecdysone agonist. Furthermore, we demonstrate specific up-regulation of reporter gene activity in transgenic B. malayi embryos transfected with a luciferase construct controlled by an EcRE engineered in a B. malayi promoter, in the presence of 20-hydroxy-ecdysone. CONCLUSIONS: Our study identifies and characterizes the two components (Bma-EcR and Bma-RXR) necessary for constituting a functional ecdysteroid receptor in B. malayi. Importantly, the ligand binding domain of BmaEcR is shown to be capable of responding to ecdysteroid ligands, and conversely, ecdysteroids can activate transcription of genes downstream of an EcRE in live B. malayi embryos. These results together confirm that an ecdysone signaling system operates in B. malayi and strongly suggest that Bma-EcR plays a central role in it. Furthermore, our study proposes that existing compounds targeting the insect ecdysone signaling pathway should be considered as potential pharmacological agents against filarial parasites.

Saini AK et al. (2011) BMC Res Notes "Allyl isothiocyanate induced stress response in Caenorhabditis elegans."

Saini AK, Reaney MJ, Tyler RT, Shim YY

[BMC Res Notes, 2011]

UNLABELLED: ABSTRACT: BACKGROUND: Allyl isothiocyanate (AITC) from mustard is cytotoxic; however the mechanism of its toxicity is unknown. We examined the effects of AITC on heat shock protein (HSP) 70 expression in Caenorhabditis elegans. We also examined factors affecting the production of AITC from its precursor, sinigrin, a glucosinolate, in ground Brassica juncea cv. Vulcan seed as mustard has some potential as a biopesticide. FINDINGS: An assay to determine the concentration of AITC in ground mustard seed was improved to allow the measurement of AITC release in the first minutes after exposure of ground mustard seed to water. Using this assay, we determined that temperatures above 67C decreased sinigrin conversion to AITC in hydrated ground B. juncea seed. A pH near 6.0 was found to be necessary for AITC release. RT-qPCR revealed no significant change in HSP70A mRNA expression at low concentrations of AITC (< 0.1 M). However, treatment with higher concentrations (> 1.0 M) resulted in a four- to five-fold increase in expression. A HSP70 ELISA showed that AITC toxicity in C. elegans was ameliorated by the presence of ground seed from low sinigrin B. juncea cv. Arrid. CONCLUSIONS: AITC induced toxicity in C. elegans, as measured by HSP70 expression. Conditions required for the conversion of sinigrin to AITC in ground B. juncea seed were determined. The use of C. elegans as a bioassay to test AITC or mustard biopesticide efficacy is discussed.

Hamilton RG et al. (1984) J Immunol Methods "Limitations of the radioimmunoprecipitation polyethylene glycol assay (RIPEGA) for detection of filarial antigens in serum."

Adkinson NF, Alexander E, Hamilton RG, Hussain R

[J Immunol Methods, 1984]

The performance of the radioimmunoprecipitation polyethylene glycol assay (RIPEGA) was examined for quantitation of filarial antigens (Brugia malayi and Dirofilaria immitis) in serum from infected human and animal hosts and non-infected controls. Multiple PEG concentrations were employed to determine the level of non-specific binding (NSB) in non-exposed human sera (NEHS) containing no filarial antigen. The NSB observed when 3 different 125I-labeled IgG antibodies were added to 26 NEHS varied 3-fold and was correlated significantly with total serum IgM (r = 0.80, P less than 0.005, n = 24) but not with serum IgA (r = 0.37) or IgG (r = 0.45). NSB levels were significantly reduced when a Fab'2 fragment of the 125I-labeled antibody was used, but the correlation of NSB with total serum IgM remained significant (r = 0.57, P less than 0.01). The presence of rheumatoid factor in NEHS sera also significantly increased NSB by an average of 3-fold. These effects eliminated the assay's ability to detect in sera from infected hosts filarial antigen the presence of which could be readily demonstrated by an immunoradiometric assay. The RIPEGA's precision (intra-assay coefficient of variation (CV) = 21% at 35% Bmax) and reproducibility (inter-assay CV = 29% at 35% Bmax) are less satisfactory than many alternative immunoassays. In many cases, positive sera failed to dilute out in parallel with each other or with an antigen-spiked standard reference curve. We conclude that poor performance characteristics currently limit the utility of the RIPEGA for quantitating filarial antigen in human and animal serum.