3.3 Ontogeny of the phloem in the stems of Austrobaileya
scandens
Cross sections of stems at different axial positions revealed
ontogenetic differences in the eustele, which correlated with stem
mechanical properties. In the flexible searcher stems, a large pith was
surrounded by discontinuous primary xylem traces consisting of only a
few tracheids (Figure 5a), but a more continuous phloem layer (Figure
5b). Twining correlated with a sharp increase in the stiffness due to
lignification of the pith and the development of an extra phloem
pericyclic fiber layer (Figure 5c). In wider stems (the ones that
traverse longest distances), the central pith constituted the largest
fraction of the cross-sectional area, and the axial secondary phloem
displayed a lobed morphology (Figure 5d-e), with sieve tubes closer to
the vascular cambium and separated from the pericyclic fibers by
parenchyma (Figure 5f).
In line with the demands for climbing and the need for flexible searcher
branches, a galacturonan-rich pectic epitope, previously related to the
flexibility of sieve tubes (Torode et al., 2018), localized in the sieve
tube walls of the searcher, flexible stems of A. scandens (Figure
6; note that we could not perform immunolocalization in the secondary
phloem due to poor preservation of the twining stems). Different cell
morphologies compose the stems in cross section (Figure 6a), but signal
from this epitope specifically showed the scattered distribution of
sieve tubes (Figure 6b,c). Upon closer inspection, the signal of the
antibody corresponded with the areas of the sieve tubes between the
plasma membrane and the cell walls (Figure 6d-f). In longitudinal
section, the epitope was helpful distinguishing sieve tubes from other
phloem cell types (Figure 6g), which highlighted their longitudinal
silhouette (Figure 6h-i), showing slightly tangential sieve plate
connections (Figure 6j-l).
In the stems supported by twining, and concomitant with leaf expansion,
the morphology of the sieve elements varied, gradually increasing the
number of sieve plate connections at the end of the conduits (Figure
7a,b). In the thickest stems, sieve elements had extremely angled
tangential connections with a great proportion of the tubes covered by
sieve areas (Figure 7b,c). Numerous small pores (0.20µm on average,
n=600), whose radius was similar to those of the leaf petioles,
populated the sieve areas (Figure 7c,d; Behnke, 1986). In addition to
morphology, sieve conduit dimensions increased from thinner to thicker
stems, but their length was shorter at the base of the vine (Figure 7e).