Tissue Structure Changes of Aquilaria sinensis Xylem after Fungus Induction

In this study, we analyzed the mechanism and the process of fungal-induced agarwood formation in Aquilaria sinensis and studied the functional changes in the xylem structure after the process. The microscopic structure of the white zone, transition zone, agarwood zone, and decay zone of 12-and 18-months of inoculation A. sinensis xylem was studied. The distribution of nuclei, starch grains, soluble sugars, sesquiterpenes, fungal propagules, and mycelium in xylem tissues was investigated by histochemical analysis. The results show that the process of agarwood formation was accompanied by apoptosis of parenchyma cells such as interxylary phloem, xylem rays, and axial parenchyma. Regular changes in the conversion of starch grains to soluble sugars, the production of sesquiterpenoids, and other characteristic components of agarwood in various types of parenchyma cells were also observed. The material transformation was concentrated in the interxylary phloem, providing a structural and material basis for the formation of agarwood. It is the core part of the production of sesquiterpenoids and other characteristic products of agarwood. Compared with the A. sinensis inoculated for 12 months, the xylem of the A. sinensis inoculated for 18 months was more vigorous. There were no significant differences between the 12 and 18 months of inoculation in terms of sugars and agarwood characteristic products. In production, harvesting after 12 months of inoculation can improve harvesting efficiency.

Linking the Evolution of Development of Stem Vascular System in Nyctaginaceae and Its Correlation to Habit and Species Diversification

Background: The presence of alternative patterns of secondary growth in stems of Nyctaginaceae has been known for a long time. Still, the interpretation of types of cambial variants are controversial. The knowledge on stem anatomical diversity in Nyctaginaceae, which is diverse also in habits, offers the unique opportunity not only to investigate the evolution of complex developments, but also to address how these anatomies shifted within habits and how the acquisition of novel cambial variants and habit transitions impacted the diversification of the family. Methods: We integrated developmental data with a phylogenetic framework to investigate the diversity and evolution of stem anatomy in Nyctaginaceae using phylogenetic comparative methods, reconstructing ancestral states, and examining whether anatomical shifts correspond to species diversification rate shifts in the family. Results: Two types of cambial variants, interxylary phloem and successive cambia, were recorded in Nyctaginaceae, which result from four different ontogenies. These ontogenetic trajectories depart from two distinct primary vascular structures (regular or polycyclic eustele) yet, they contain shared developmental stages which generate stem morphologies with deconstructed boundaries of morphological categories (continuum morphology). Unlike our a priori hypotheses, interxylary phloem is reconstructed as the ancestral character for the family, with three ontogenies characterized as successive cambia evolving in few taxa. Cambial variants are not contingent in habits, and their transitions are independent from species diversification.Conclusions: Our findings suggests that multiple developmental mechanisms, such as heterochrony and heterotopy generate the transitions between interxylary phloem and successive cambia. Intermediate between these two extremes are present in Nyctaginaceae, suggesting a continuum morphology across the family as a generator of anatomical diversity.

Regeneration of vascular tissue through redifferentiation of interxylary phloem after complete girdling in Aquilaria sinensis

We studied the time-course of stem response for six months following complete girdling in branches of Aquilaria sinensis to determine the potential role of interxylary phloem (IP) in this response. It was found that the vascular cambium, as well as its derivative secondary xylem and phloem, regenerated fully through redifferentiation of IP. We confirmed that vascular cambium regenerated within one month after girdling based on observation of new vessels, IP, and secondary phloem fibers. The time-course study showed that IPs made connections with each other, merged, and became larger through the proliferation of IPs parenchyma cells and the cleaving of secondary xylem in a narrow zone 400 to 1000 μm deep inside the girdled edge. This led to the formation of a complete circular sheath of vascular cambium, followed by the regeneration of vascular tissue. It is worth noting that the secondary xylem is regenerated always following the formation of a thick belt of wound xylem.

Development of cambial variant and parenchyma proliferation in Hewittia malabarica (Convolvulaceae) from India and South Africa

Members of the Convolvulaceae are characterized by the climbing habit and occurrence of variant secondary growth. From a histological perspective, the genus Ipomoea L. is the most extensively studied, while other genera have been less studied. Here, stem anatomy of the least studied genus in the family, Hewittia Wight & Arn., represented by Hewittia malabarica (L.) Suresh was investigated using classical histological techniques. In both the samples collected from India and South Africa, stem thickness increased by developing different types of cambial variants such as: neo-formed vascular cylinders, parenchyma proliferation at the phloem wedges, ray-derived cambia from dilating phloem rays, internal cambium, intra- and interxylary phloem. Neo-formed vascular cylinders develop from the parenchyma cells external to the phloem as a meristemoid in thick stems and later in dilating ray cells. With the increase in stem diameter, cells of the phloem wedges showed proliferation by meristematic activity, which form a connection with the cortex by rupturing the primary tissue ring of eustele. Subsequently, development of cambium in phloem wedges and deposition of its derivatives increased the tangential width of rays. Mature thick stems (25–30 mm) give rise to a fissured stem. Intraxylary (internal) phloem development on the pith margin was observed from primary growth onwards and in thick stems secondary intraxylary phloem developed from the internal cambium. Internal cambium is functionally bidirectional and produces secondary xylem internally and secondary phloem externally. In all the samples, patches of unlignified parenchyma embedded within the secondary xylem dedifferentiate and mature into interxylary phloem with the increasing age. Development of cambial variant and structure of the secondary xylem is correlated with the functional significance of the climbing habit.

The occurrence and structure of radial sieve tubes in the secondary xylem of Aquilaria and Gyrinops

New observations of radial sieve tubes in the secondary xylem of two genera and four species of agarwood — Aquilaria sinensis, A. crasna, A. malaccensis and Gyrinops versteeghii (Thymelaeaceae) — are presented in this study. The earliest radial sieve tubes in Gyrinops are formed in the secondary xylem adjacent to the pith. The radial sieve tubes originate from the vascular cambium and develop in both uniseriate and multiseriate ray tissue. In addition to sieve plates in lateral and end walls, scattered or clustered minute sieve pores are localized in the lateral wall of radial sieve tubes. There is a direct connection between radial sieve tubes in ray tissue and axial sieve tubes in interxylary phloem strands (IP), such as (i) connection by bending of radial sieve tube strands, (ii) connection of two IP strands by an oblique bridge, and (iii) connection of two IP strands at a right angle. The average number of radial sieve tubes and interxylary phloem was found to be 1.7 per mm3 and 9.1 per mm2 in the secondary xylem. Considering the higher frequency of radial sieve tubes with the increasing thickness of the secondary xylem, the direct connections between radial and axial sieve tubes could play a significant role in assisting the translocation of metabolites in Aquilaria and Gyrinops.

Formation of medullary phloem in Argyreia nervosa (Burm. f.) Bojer

Histologically, family Convolvulaceae is characterised by the presence of successive cambia, medullary (internal/intraxylary) and interxylary phloem in majority of the species, whereas some of the members are devoid of successive cambia and medullary bundles. The present study on Argyreia nervosa (Burm. f.) Bojer showed the presence of medullary bundles while internal phloem was absent during the primary growth. As the secondary growth progressed, successive cambia initiated from the pericyclic parenchyma. Development of medullary bundles began along with the regular protoxylem and protophloem while formation of intraxylary phloem was observed only after the initiation of secondary growth. Medullary/intraxylary sieve elements began to develop from the marginal pith cells. In thick stems, small segments of internal cambium initiated between the protoxylem and internal phloem. This internal cambium was functionally unidirectional and produced internal phloem centripetally. Developmental particulars are described in details along with its significance.

The occurrence and development of intraxylary phloem in young Aquilaria sinensis shoots

ABSTRACT Agarwoods such as Aquilaria spp. and Gyrinops spp. (Thymelaeaceae) produce interxylary phloem in their secondary xylem and intraxylary phloem at the periphery of the pith, facing the primary xylem. We studied young shoots of Aquilaria sinensis and characterized the development of its intraxylary phloem. It was initiated by the division of parenchyma cells localized in the outer parts of the ground meristem immediately following the maturation of first-formed primary xylem. Its nascent sieve plates bore donut-like structures, the individual pores of which were so small (less than 0.1 μm) that they were hardly visible under FE-SEM. Intraxylary phloem developed into mature tissue by means of the division and proliferation of parenchyma cells. During the shoots’ active growth period, the sieve pore sizes were 0.1–0.5 μm, with tubular elements passing through them. In the maturation stage, large clusters of sieve tubes continued to be differentiated in the intraxylary phloem. In the partial senescence stage observed in a three-centimeter-diameter branch, intraxylary phloem cells in the adaxial part became crushed, and sieve plates had pores over 1–2 μm in diameter without any callose deposition. Before and after the differentiation of interxylary phloem in the first and second internodes, callose staining detected more than twice as many sieve tubes in intraxylary phloem than in external phloem. However, after differentiation of interxylary phloem in the eleventh internode, more sieve tubes were found in interxylary phloem than in intraxylary and external phloem. This suggests that prior to the initiation of interxylary phloem intraxylary phloem acts as the principal phloem. After its differentiation, however, interxylary phloem takes over the role of principal phloem. Interxylary phloem thus acts as the predominant phloem in the translocation of photosynthates in Aquilaria sinensis.

The structure and development of interxylary and external phloem in Aquilaria sinensis

The structure and development of interxylary phloem (IP) and external phloem in Aquilaria sinensis were investigated using light and scanning electron microscopy. Complete IP strands were isolated, measuring 14 ± 4 mm in length and 417 ± 124 μm in width. The outer margin of IP was composed of two to three layers of fusiform parenchyma cells. The development of IP can be divided into five stages: 1) Locally IP starts its differentiation within a small segment of a broad cambial zone, at the cost of xylem differentiation. 2) Inward growth of IP advances, and fibres and sieve tubes differentiate. 3) IP is constricted by the encroachment of immature xylem cells between cambium and immature IP. 4) IP is isolated from the cambium and surrounded by immature, non-lignified xylem tissue. 5) IP is surrounded by lignified xylem tissue, and the fibres within IP become lignified.In all the phloem islands in a ten-year-old stem, sieve elements showed positive staining of callose with aniline blue. However, no staining of callose was observed in the external secondary phloem of agarwood trees collected from two different sites. No sieve tubes or sieve pores were detected by SEM observation of numerous serial cross and radial sections of the external phloem. We therefore conclude that sieve tubes are absent from the external phloem or extremely rare and that the transport of photosynthetic products in the stem of A. sinensis takes place in the interxylary phloem.