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E Witze et al. (2002) West Coast Worm Meeting "Biochemical characterization of PLP-1, a regulator of mesendoderm development"

J H Rothman, E Witze, D Hunt, E Field

[

West Coast Worm Meeting,

2002]

The zygotic genes end-1 and end-3 together are required to specify the endoderm progenitor, the E cell. Their expression is restricted to the E blastomere and ectopic expression of either gene is sufficient to convert non-E lineages to an E-like fate. Genetic screens have identified maternal factors, including SKN-1 and POP-1, that are involved in mesendoderm specification and correct end-1,3 expression. To isolate regulators of end-1 that were missed in genetic screens, we used mass spectrometry to identify a transcription factor, PLP-1 (pur alpha like protein) that directly binds to the end-1 promoter and that is similar to the mammalian transcription factor pur alpha. RNAi of plp-1 results in embryos that fail to form a pharynx. Some of the arrested embryos produce extra intestine, suggesting a transformation of non-E lineages to an E-like fate. A very impenetrant gutless phenotype is also observed, which is greatly enhanced in mutants defective in the endoderm-inducing Wnt pathway. However, plp-1 RNAi does not enhance the gutless phenotype observed in mutants in the MAP kinase pathway, which converges with the Wnt pathway to regulate endoderm development, suggesting that plp-1 may function in the latter pathway. To understand how plp-1 might act both positively and negatively on end-1 expression, we have examined the in vitro binding characterisics of PLP-1 and its possible post-translational modifications. PLP-1 binds to a putative Lef-1 binding site, which is required for activation of a minimal end-1 promoter. Surrounding this region, the end-1 DNA apparently adopts a secondary structure in vitro that can be cleaved with an endonuclease specific to cruciform structures. Deletion of the Lef-1 site results in loss of this secondary structure and PLP-1 binding activity. The affinity of PLP-1 for this site appears to be regulated by phosphorylation, as demonstrated by the loss of PLP-1 binding activity when early embryonic nuclear extract is treated with a non-specific phosphatase and by the ability of the extract to phosphorylate recombinant PLP-1 protein. A candidate kinase that might phosphorylate PLP-1 is the nemo-like kinase LIT-1, since it is a component of the MAPK pathway. Consistent with this possibility, we find that in lit-1 mutants, PLP-1 only weakly localizes to the nucleus with a more diffuse staining pattern throughout the cell, unlike in normal embryos in which it is strongly nuclear-localized.

Chrisman, Steven D. et al. (2009) International Worm Meeting "Analysis of Electrotaxis Behavior in C. elegans."

Chrisman, Steven D., Carnell, Lucinda

[

International Worm Meeting,

2009]

C. elegans sense both polarity and amplitude of an electric field by uninterrupted crawling toward the negative pole, a behavior referred to as electrotaxis (Sukul and Croll, 1978). Gabel et al. (2007) have identified specific amphid neurons and the neural circuitry important for this behavioral response by tracking the animal''s movement in a rotating electric field. We are interested in studying the sensory response to an electric field at varying electric field strength in a non-rotating field. We observed that as previously described animal taxis toward the negative pole (cathode). Interestingly, we also observed that decreasing the field strength results in the animals'' responding with an immediate reversal. To examine the sensory mechanism responsible for electrotaxis, we have tested known chemosensory-defective mutants and performed a genetic screen to isolate animals defective in the sensing of the electric field. We have isolated seven candidate mutants. The mutants fall into two general classes: those that taxis to lower field strengths (hypersensitive) and those that respond to higher field strengths (resistant). We are in the process of characterizing these mutants further by recording their response and measuring velocity using the automated tracking software, WormTracker (Ramot et al. 2008), which allows us to observe the behavior of a population of animals simultaneously. Initial experiments indicate the first observable sensory response of animals at low field strengths is to increase velocity, which is followed by a sensing of polarity at increasing field strengths. To determine and quantify the animals'' sensitivity to an electric field, we are measuring velocity at different field strengths over time. Images will be captured at 15 second intervals every 30 seconds as the worms move across the agar plate to the negative pole. The velocities for each group of animals will be measured at field strengths ranging from 0.5 V/cm to 14V/cm. Gabel et al. 2007 Journal of Neuroscience 27(28):7586-7596 Sukul, N.C. and Croll, N.A. 1978 J. Nematology 10, 314-315 Ramot et al. 2008 PLoS ONE 3(5): e2208. Doi:10.1371/journal.pone.0002208.

Chris Gabel et al. (2004) East Coast Worm Meeting "Quantification of electrotaxis"

Chris Gabel, Albert Kao, Aravi Samuel, Dmitri Pavlichin

[

East Coast Worm Meeting,

2004]

In an electric field, the worm has preferred trajectories for forward crawling movement that depend on the direction and magnitude of the electric field. In weak fields (<5 V/cm), worms tend to crawl towards either the positive or negative pole. In moderate fields, ~5 V/cm, worms crawl straight towards the negative pole. In stronger fields, >5 V/cm, worms crawl towards the negative pole in trajectories at an angle to the direction of the field. The angle of approach towards the negative pole increases with field strength, rising from 0 degrees (parallel to the field lines) towards 90 degrees (perpendicular to the field lines). We have quantified these electrotactic movements by tracking wild-type and mutant worms crawling over agar surfaces containing different types and concentrations of ions while responding to electric fields varied in amplitude, frequency, and direction.

Rankin CH et al. (2015) Elife "Finding a worm's internal compass."

Lin CH, Rankin CH

[

Elife,

2015]

A pair of neurons is required for nematodes to be able to navigate using the Earth's magnetic field.

Lei M et al. (2011) J Biomed Opt "Video-rate wide-field coherent anti-Stokes Raman scattering microscopy with collinear nonphase-matching illumination."

Winterhalder M, Zumbusch A, Lei M, Selm R

[

J Biomed Opt,

2011]

A simple scheme for video-rate wide-field coherent anti-Stokes Raman scattering (CARS) microscopy is presented. The method is based on collinear nonphase-matching illumination. The mechanisms leading to CARS signal generation are investigated. We find that refraction-mediated phase-matching is the main effect. Video-rate wide-field CARS microscopy of polystyrene beads and CARS wide-field images of C. elegans embryos are shown, and the capabilities and the limitations of the scheme are discussed.

Fasseas MK et al. (2015) Int J Radiat Biol "Response of Caenorhabditis elegans to wireless devices radiation exposure."

Margaritis LH, Manta AK, Vekrellis K, Skouroliakou A, Fragopoulou AF, Syntichaki P, Fasseas MK

[

Int J Radiat Biol,

2015]

PURPOSE: To examine the impact of electromagnetic radiation, produced by GSM (Global System for Mobile communications) mobile phones, Wi-Fi (Wireless-Fidelity) routers and wireless DECT (Digital Enhanced Cordless Telecommunications) phones, on the nematode Caenorhabditis elegans. MATERIALS AND METHODS: We exposed synchronized populations, of different developmental stages, to these wireless devices at E-field levels below ICNIRP's (International Commission on Non-Ionizing Radiation Protection) guidelines for various lengths of time. WT (wild-type) and aging- or stress-sensitive mutant worms were examined for changes in growth, fertility, lifespan, chemotaxis, short-term memory, increased ROS (Reactive Oxygen Species) production and apoptosis by using fluorescent marker genes or qRT-PCR (quantitative Reverse Transcription-Polymerase Chain Reaction). RESULTS: No statistically significant differences were found between the exposed and the sham/control animals in any of the experiments concerning lifespan, fertility, growth, memory, ROS, apoptosis or gene expression. CONCLUSIONS: The worm appears to be robust to this form of (pulsed) radiation, at least under the exposure conditions used.

Chrisman SD et al. (2016) PLoS One "C. elegans Demonstrates Distinct Behaviors within a Fixed and Uniform Electric Field."

Waite CB, Scoville AG, Chrisman SD, Carnell L

[

PLoS One,

2016]

C. elegans will orient and travel in a straight uninterrupted path directly towards the negative pole of a DC electric field. We have sought to understand the strategy worms use to navigate to the negative pole in a uniform electric field that is fixed in both direction and magnitude. We examined this behavior by quantifying three aspects of electrotaxis behavior in response to different applied field strengths: the mean approach trajectory angles of the animals' tracks, turning behavior (pirouettes) and average population speeds. We determined that C. elegans align directly to the negative pole of an electric field at sub-preferred field strength and alter approach trajectories at higher field strengths to maintain taxis within a preferred range we have calculated to be ~ 5V/cm. We sought to identify the sensory neurons responsible for the animals' tracking to a preferred field strength. eat-4 mutant animals defective in glutamatergic signaling of the amphid sensory neurons are severely electrotaxis defective and ceh-36 mutant animals, which are defective in the terminal differentiation of two types of sensory neurons, AWC and ASE, are partially defective in electrotaxis. To further elucidate the role of the AWC neurons, we examined the role of each of the pair of AWC neurons (AWCOFF and AWCON), which are functionally asymmetric and express different genes. nsy-5/inx-19 mutant animals, which express both neurons as AWCOFF, are severely impaired in electrotaxis behavior while nsy-1 mutants, which express both neurons as AWCON, are able to differentiate field strengths required for navigation to a specific field strength within an electric field. We also tested a strain with targeted genetic ablation of AWC neurons and found that these animals showed only slight disruption of directionality and turning behavior. These results suggest a role for AWC neurons in which complete loss of function is less disruptive than loss of functional asymmetry in electrotaxis behavior within a uniform fixed field.

Chuang HS et al. (2011) Lab Chip "Dielectrophoresis of Caenorhabditis elegans."

Lamb A, Chuang HS, Raizen DM, Dabbish N, Bau HH

[

Lab Chip,

2011]

We demonstrate for the first time the dielectrophoretic trapping and manipulation of a whole animal, the nematode Caenorhabditis elegans. We studied the effect of the electric field on the nematode as a function of field intensity and frequency. We identified a range of electric field intensities and frequencies that trap worms without apparent adverse effect on their viability. Worms tethered by dielectrophoresis (DEP) exhibit behavioral responses to blue light, indicating that at least some of the nervous system functions are unimpaired by the electrical field. DEP is useful to dynamically tether nematodes, sort nematodes according to size, and separate dead worms from live ones.

Pilotte N et al. (2019) Gates Open Res "Laboratory evaluation of molecular xenomonitoring using mosquito and tsetse fly excreta/feces to amplify Plasmodium, Brugia, and Trypanosoma DNA."

Zulch MF, Reimer LJ, Pilotte N, Pryce J, Cook DAN, Williams SA, Minetti C

[

Gates Open Res,

2019]

Background: Results from an increasing number of studies suggest that mosquito excreta/feces (E/F) testing has considerable potential to serve as a supplement for traditional molecular xenomonitoring techniques. However, as the catalogue of possible use-cases for this methodology expands, and the list of amenable pathogens grows, a number of fundamental methods-based questions remain. Answering these questions is critical to maximizing the utility of this approach and to facilitating its successful implementation as an effective tool for molecular xenomonitoring. Methods: Utilizing E/F produced by mosquitoes or tsetse flies experimentally exposed to Brugia malayi, Plasmodium falciparum, or Trypanosoma brucei brucei, factors such as limits of detection, throughput of testing, adaptability to use with competent and incompetent vector species, and effects of additional blood feedings post parasite-exposure were evaluated. Two platforms for the detection of pathogen signal (quantitative real-time PCR and digital PCR (dPCR)) were also compared, with strengths and weaknesses examined for each. Results: Experimental results indicated that high throughput testing is possible when evaluating mosquito E/F for the presence of either B. malayi or P. falciparum from both competent and incompetent vector mosquito species. Furthermore, following exposure to pathogen, providing mosquitoes with a second, uninfected bloodmeal did not expand the temporal window for E/F collection during which pathogen detection was possible. However, this collection window did appear longer in E/F collected from tsetse flies following exposure to T. b. brucei. Testing also suggested that dPCR may facilitate detection through its increased sensitivity. Unfortunately, logistical obstacles will likely make the large-scale use of dPCR impractical for this purpose. Conclusions: By examining many E/F testing variables, expansion of this technology to a field-ready platform has become increasingly feasible. However, translation of this methodology from the lab to the field will first require field-based pilot studies aimed at assessing the efficacy of E/F screening.

Isikman SO et al. (2010) Lab Chip "Color and monochrome lensless on-chip imaging of Caenorhabditis elegans over a wide field-of-view."

Sencan I, Ozcan A, Isikman SO, Oztoprak C, Bishara W, Mudanyali O

[

Lab Chip,

2010]

We demonstrate color and monochrome on-chip imaging of Caenorhabditis elegans samples over a wide field-of-view using incoherent lensless in-line holography. Digital reconstruction of the recorded lensless holograms rapidly creates the C. elegans images within <1 s over a field-of-view of >24 mm2. By digitally combining the reconstructed images at three different wavelengths (red, green and blue), color images of dyed samples are also acquired. This wide field-of-view and compact on-chip imaging modality also permits straightforward integration with microfluidic systems.

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