FIGURES AND LEGENDS
FIGURE 1. (A) Adult vines of Austrobaileya scandens in
the greenhouse. (B ) A. scandens vines at the canopy
layer in the tropical rainforest of Queensland (Australia). (C )
Apical part of a ‘searcher’ branch in the greenhouse. (D )
Branches twisted around a stable support without increasing their
diameter. (E ) The average leaf area at each particular node
increased by five fold up to the fully expanded leaves of the seventh
node (p <0.05).
FIGURE 2. Vascular anatomy of the continuum petiole-mid vein ofAustrobaileya scandens leaves. (A ) Mature leaf
of A. scandens . (B ) Vascular tissues of the fiber less
petiole. (C ) Vasculature in the mid part of the major vein
showing a massive layer of pericyclic fibers surrounding the xylem and
the phloem. (D ) Vasculature at the tip of the mid vein.
(E ) Cross sectional areas of the vascular tissues at the three
positions in the midvein: while the cross-sectional areas of xylem (red
squares) and phloem (yellow circles) decreased linearly toward the tip
(p <0.05), the fibers (grey ribbons) occupied a wide
cross-sectional area in the leaf lamina, but not in the petiole.B-D . Cross sections stained with aniline blue to detect callose
of the sieve tube elements. Scale bars: A = 1cm;B -D = 200 µm.
FIGURE 3. Geometrical scaling of the vascular elements in the
leaves of Austrobaileya scandens . (A ) General view of
the leaf veins showing the sampling areas (pink hexagons) and closeup of
the minor veins. (B-C ) Length (squares) and radius (circles) of
the individual vessel elements (red), and sieve tube elements (yellow)
across vein orders; error bars represent standard error at ap <0.05. Inset in panel (A) shows a
fluorescence image section of the leaf stained with the Feulgen
reactive, cleared, and cuticle dissected. Fou, fourth order veins; pet,
petiole; pri, primary vein; sec, secondary vein; ter, tertiary vein.
Scale bar = 500µm.
FIGURE 4. Velocity of the phloem sap in Austrobaileya
scandens leaves. Time-lapse images of a secondary vein showing a 2 min
advancement of esculin hydrate dye from 0min (A ) to 6min
(D ). Insets show the thresholding applied to the sequential
images for the calculation of velocity, with a gradual accumulation of
the dye within the veins. Scale bars = 1000μm.
FIGURE 5. Anatomy of the Austrobaileya scandens stems.
(A ) Cross section of a 2mm diameter ‘searcher’ branch tip with
primary growth, with an extensive central pith. (B ) Same stem
showing callose in the continuous phloem ring (fluorescent green).
(C ) Cross section of a stiff 2mm diameter branch showing a
dramatic increase in lignification of the pith, the xylem, and a
pericyclic fiber cap (fluorescent green). (D ) The phloem forms
a thin layer between the xylem and the fiber cap. (E ) Wider
stems (8mm diameter) with a high degree of lignification in all central
tissues, wide vessels of the xylem, and enlarged areas between the xylem
and the fiber cap. (F ) The fascicular phloem tissue was
separated by axial multilayered phloem parenchyma and the active tubes
were in the vicinity of the vascular cambium. A,C,E: cross
sections of stems stained with acridine orange, which displays
fluorescence of the lignified tissues in fluorescent green and living
tissues in reddish color; B,D,F: cross sections of the stems
stained with aniline blue for callose in the sieve tube elements of the
phloem. F, fibers; p, pith; phl, phloem; xyl, xylem. Scale bars:A-F = 1mm; insets = 100µm.
FIGURE 6. Immunolocalization of LM26 epitope in the sieve tube
elements of searcher branches of Austrobaileya scandens .
(A ) Transverse section showing multicellular arrangement in the
different tissue layers, including three isolated tracheids that compose
the primary xylem (red arrowheads). (B ) Same section displaying
the sieve tube elements of the primary phloem forming a continuous ring
(yellow arrows). (C ) Merged images. (D ) Close up of
the phloem in cross section. (E ) The profile of the sieve tubes
in cross section after immunolocalization (yellow arrows). (F )
Merged images. (G ) Longitudinal section of the branch
displaying he external part in the top and the internal tissues at the
bottom. (H ) Longitudinal profile of the sieve tube element wall
after immunolocalization, yellow arrowheads define the connection
between tubes. (I) Merged images. (J) Detail of the
sieve tube connections. (K) Sieve plate in the primary sieve
tubes are slightly tangential (yellow arrowhead). (L) Merged
images. 4µm thick transverse (A-F ) or longitudinal
(G-L ) sections of the flexible branches displaying
autofluorescence with the 405nm filter (A,D,G,J ),
immunolocalized with the LM26 monoclonal antibody (B,E,H,K ),
and merged images (C,F,I,L ). Ct, cortical tissue; phl, phloem;
sp, sieve plate; vc, vascular cambium; xyl, xylem. Scale bars:A-C = 50 µm; G-I = 25 µm; D-F , J-L =
10 µm.
FIGURE 7 . Morphology of the sieve tube elements in the stems ofAustrobaileya scandens . (A ) Longitudinal view of a sieve
tube element from a flexible branch, 2mm diameter, showing either simple
or compound sieve plate connections (arrows). (B ) Sieve tube
element of an 8mm diameter stem showing numerous sieve plate connections
between tubes (arrows). (C ) Longitudinal view of a sieve tube
element with scanning electron microscopy displaying numerous sieve
plates and sieve areas all along the tube wall (arrows). (D )
Close ups of three sieve plates (analogous to the white dotted square in
figure C), with details of pores. (E ) Sieve tube element length
(squares), and width (circles) across axial stems of different diameter
ranges; bars represent the standard error at a p <0.05.A, B , longitudinal hand sections of the stems stained with
aniline blue to detect callose (bright fluorescence); C,D ,
scanning electron microscopy images showing the sieve plates and the
sieve pores in detail. Scale bars: A-C = 20um; D =
2µm.
FIGURE 8. Sieve tube hydraulic resistance across the aerial
organs of Austrobaileya scandens . The sieve tube resistance drops
from 1.5 to 2 orders of magnitude from the minor leaf veins (squares) to
the base of the stems (circles), even though the distance between them
could be up to 20m. Numbers indicate the relative position along the
transport pathway, noted in the figures, but they do not represent real
distances.