First published online September 5, 2008
Journal of Experimental Biology 211, 2901-2908 (2008)
Published by The Company of Biologists 2008
doi: 10.1242/jeb.020743
A surface lipid may control the permeability slump associated with entry into anhydrobiosis in the plant parasitic nematode Ditylenchus dipsaci
D. A. Wharton1,*,
L. Petrone2,
A. Duncan3 and
A. J. McQuillan2
1 Department of Zoology, University of Otago, PO Box 56, Dunedin, New
Zealand
2 Department of Chemistry, University of Otago, PO Box 56, Dunedin, New
Zealand
3 Department of Human Nutrition, University of Otago, PO Box 56, Dunedin, New
Zealand
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Fig. 1. Schematic diagram of the three-internal reflection diamond-coated ZnSe
attenuated total reflection infrared (ATR–IR) prism (left). Penetration
depth (dp) and exponential decay of the evanescent wave in
the medium with a lower refractive index
(n1<n2) are shown in detail for one
of the points of reflection (right).
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Fig. 2. Wheat germ agglutinin–fluorescein isothiocyanate
(WGA–FITC)-stained D. dipsaci fourth-stage larvae (L4s)
observed by confocal microscopy, focused on the centre (A) or surface (B) of
the nematodes. Surface labelling of the cuticle can be observed and appears to
be confined to the annulations. Scale bar, 10 µm.
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Fig. 3. Changes in annulation spacing of individual nematodes observed by confocal
microscopy during desiccation of D. dipsaci fourth-stage larvae (L4s)
at 50% RH, 20°C using reflected laser light (open symbols) or on Wheat
germ agglutinin–fluorescein isothiocyanate (WGA–FITC)-stained
specimens (closed symbols).
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Fig. 4. D. dipsaci L4s desiccated at 50% RH, 20°C for 5 min, stained
with Nile Red and observed by confocal microscopy focused on the centre (A) or
surface (B) of the nematodes. Lipid droplets are associated with the surface
of the cuticle, particularly with the cuticular annulations and the grooves of
the lateral alae. Scale bars, 10 µm.
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Fig. 5. Cuticle prints. Samples were prepared as for
Fig. 4 and the material
adhering to the coverslip, after nematode removal, was then observed by
epifluorescence (A) or confocal (B) microscopy. Cuticle prints stained with
Nile Red can be observed, leaving clear impressions of the cuticular
annulations and lateral alae. Scale bars, 10 µm.
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Fig. 6. D. dipsaci fourth-stage larvae (L4s) allowed to dry on a coverslip
and then imaged through the coverslip using DIC optics on a Zeiss
photomicroscope using a x100 objective lens (A) or using confocal
microscopy and reflected laser light (B). Oily material is observed between
the nematodes and the coverslip (arrow in A) or smeared from the surface of
the nematodes, as a result of their movement (arrow in B). Scale bars, 10
µm.
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Fig. 7. Attenuated total reflection infrared (ATR–IR) spectra from the dry
nematodes (with-worms; A), the oily secretion left after the nematodes removal
(without-worms; B), and the spectrum after ether-extraction (C). Absorbance
scale is the same for all spectra. Background was the bare diamond prism.
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Fig. 8. (A) Attenuated total reflection infrared (ATR–IR) spectrum of
purified nematode surface lipid from nematodes exposed to 50% RH and (B)
comparison of part of the spectra from those exposed to 0%, 50% or 98% RH.
Background was the bare diamond prism.
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© The Company of Biologists Ltd 2008
