IAWA Journal, Vol. 18 (2),1997: 147-156

FIBRE MATURATION IN THE GIGANTOCHLOA SCORTECHINII by R.J. Murphy & K.L. Alvin

Department of Biology, Imperial College of Science, Technology & Medicine, London SW7 2BB, United Kingdom

SUMMARY

Fibre maturation, which has been shown in a number of to be a process extending over a long period after the culm has reached its full height, has been investigated in comparable internodes (6th above ground level) in culms up to three years old, with special reference to the fibres constituting the free fibre strands immersed in the ground tissue, The possession of such strands is characteristic of this pachymorph species. The fibres of the free strands are notably more heterogeneous in terms of their diameter than those of the fibre caps adjacent to the vascular tissues. It is in some of the larger fibres of the free strands that wall thickening is longest delayed, so that, even after three years, many still remain compa­ ratively thin-walled, especially in the inner region of the culm wall. Fibres retain a living protoplast and appear to undergo progressive septation. Key words: Cell wall, culm, density, septa, vascular bundle.

INTRODUCTION

It has been well established for a number of different bamboos that progressive culm maturation may continue over several years. Thus, Liese (1985) showed that culm strength usually increases with age to reach a maximum in about three years. Fujii (1985), in a study of cell wall fine structure and development in chino, reported that larger fibres continued thickening late into the second year. Alvin & Murphy (1988) found that in cell walls of both fibres and ground tissue parenchyma could go on thickening into the third year. Mohmod et al. (1990) have shown that in three different pachymorph bamboos, including Gigantochloa scorte­ chinii, culm age up to three years has a strong influence on a wide range of mechanical properties. Recently, Liese and Weiner (1996), in a valuable summary of the present state of knowledge of the subject of ageing in bamboo culms, have shown that in viridiglaucescens the culm may undergo ageing processes involving wall thickening and chemical changes for up to twelve years. It is also well known that the rate of fibre wall maturation varies from one part of the culm wall to another. Thus, in the early stages, fibre wall thickening and lignifica­ tion proceeds from the outside of the culm inwards and from the inside of vascular fibre-caps towards the ground tissue. The very early maturing fibres are of relatively

Downloaded from Brill.com10/07/2021 11:08:41AM via free access 148 IAWA Journal, Vol. 18 (2), 1997 small diameter and all reach maximum wall thickness so as to leave a lumen of very narrow diameter well within the first year, It is the larger fibres situated further from the vascular tissues which can continue wall thickening beyond the first year (Alvin & Murphy 1988), Changes in wall thickness with age will inevitably have profound ef­ fects on mechanical properties of the culm. Most anatomical studies on maturation and ageing in bamboos have been carried out on species characteristically lacking free fibre strands, that is, having vascular bun­ dles only of Types I and II according to Grosser & Liese's (1971) and Liese's (1985) system of vascular bundle classification.We have therefore examined aspects of fibre wall thickening in Gigantochloa scortechinii Gamble, one of a number of tropical, pachymorph bamboos characterised by possessing free fibre strands positionally asso­ ciated with the vascular bundles but entirely surrounded by ground tissue. Vascular bundles and their associated fibres are, in this species, very variable in their configura­ tion (Fig. 1). Through most of the culm wall, they are characteristically of either Type III or Type IV. In Type III, the bundle, with its four small fibre-caps (one adjacent to the protoxylem canal, one to the phloem and one to each of the two large metaxylem ves­ sels) has associated a usually large, centripetally placed free fibre strand; in Type IV, free strands are present both centripetally and centrifugally. In the outermost zone, bundles are of Type II, in which there are no associated free strands but the centripetal cap is large. We have paid special attention to aspects of fibre maturation in the cen­ tripetal free fibre strands and the centripetal fibre-cap in the inner part of the outer zone of the culm wall (Fig. I). Fibres in these portions are remarkable for their heterogene­ ity in terms of diameter and wall thickness.

MATERIAL AND METHODS

Three accurately aged culms (one to three years) of comparable diameter (c. 6 cm) and height (c. 7 m) from a stand growing in secondary forest in Peninsular Malaysia were harvested. Anatomical comparison was carried out on the middle section of the sixth internode above soil level. One very immature specimen, estimated to be less that one month old, was also collected. Fresh material was fixed (using a mixture consisting of 5% formalin, 5% acetic acid and 90% of 70% ethanol) and sectioned on a sliding microtome. Sections were stained in safranin and alcian blue and permanently mounted in Euparal. Some difficulty was experienced in sectioning the 3-year-old culm due to its hardness. Culm basic density (oven dry weight! swollen volume) was measured using the water displacement method and expressed as kgm- 3. Three replicates were used for each determination. For observational purposes, the culm wall was divided into three equal zones (Fig. I). By the use of an eye-piece micrometer scale, fibre diameter and wall thickness of individual fibres were measured at ten equidistant intervals along tangential transect lines across the widest part of centripetal free fibre bundles, or, in the case of the outer zone, of centripetal fibre-caps. Three such transects were taken and results averaged for each of the three zones of the culm wall, i. e., the inner part of the outer zone, the middle of each of the middle and inner zones. We chose to measure fibres in the inner

Downloaded from Brill.com10/07/2021 11:08:41AM via free access Murphy & Alvin - Fibre maturation in Bamboo 149 part of the outer zone rather than in the middle of this zone because the centripetal fibre-caps in the inner part resemble more closely in their rather varied diameter and wall thickness those of the free fibre strands in the two inner zones. In the middle and outer parts of the outer zone the fibrous tissue is much more homogeneous and similar to that of the vascular caps throughout (Fig. 1).

RESULTS

Brief examination of the very immature specimen confirmed that vascular bundle fibre­ caps apparently conform in their general pattern of maturation to those in other bam­ boos that have been investigated. The dark staining in Figure 2 indicates wall thicken­ ing limited to fibres adjacent to the vascular tissue. Although so few fibres are yet thick-walled, they are all, by their distribution as well as by their generally smaller diameter, well differentiated from the ground tissue parenchyma. Moreover, the varia­ tion in fibre diameter seen in the free fibre strands in the more mature specimens has already become established at this young stage. Throughout the middle and inner zones of the culm wall, fibres of the vascular caps are consistently smaller in average diameter than those of the free fibre strands (Fig. 1, 3, 4). The narrow fibres (mostly < 15 /lm diameter) characteristic of these caps all thicken to almost their maximum during the first year of culm growth, so that all but a few of the larger, more peripheral fibres have extremely narrow lumina. The same is true of the fibre-caps throughout the middle and outer parts of the outer zone. (All the uniformly dark fibrous tissue seen in Fig. 1 consists of narrow, early maturing fibres.) There is also always much greater variation in fibre diameter within the free strands and, to a lesser extent, in the large centripetal caps in the inner part of the outer zone than in the vascular caps (Fig. 1, 3,4). Measurements show that in the centripetal fibre­ caps in the inner part of the outer zone, about 50% of fibres have a diameter over 20 /lm, of which about 2% are over 30 /lm, and in free fibre strands of the middle and inner zones there are some 65%, of which about 8% are over 30 /lm. Variation in fibre diameter within the free strands is not random; as can be seen in Figures 2-7, there is a tendency for the smaller fibres to be peripheral, with a few groups or isolated smaller fibres further towards the centre. Figure 10, combining data from all three specimens, shows that in both the middle and inner zones of the culm wall, average fibre diameter in the centripetal free strands is significantly greater towards the centre of these strands than at the periphery. This is much less marked in the centripetal fibre caps in the inner part of the outer zone where the average fibre diameter is also smaller. In all three specimens, the walls of most fibres are thickened to some degree, though there is much variation in thickness, especially within the free fibre strands (Fig. 3-7), and even in the 3-year-old culm (Fig. 6, 7) some thin-walled fibres still remain. The proportion of fibres remaining relatively thin-walled is always greater in the inner regions of the culm wall. Observation indicates that it is some of the broader fibres towards the interior of the fibre strands that remain thin-walled the longest and that it is the narrower fibres that thicken earlier (Fig. 4, 5). We have verified from longitudinal sections that it is not the narrow ends of individual fibres that thicken precociously, but that the wall thickens uniformly throughout the length.

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Table 1. Average wall thickness in relation to fibre diameter (flm).

Fibre diameter: <20 flm > 20flm

1 year 5.9 6.4 Outer zone 2 year 7.0 7.5 3 year 7.5 8.6

1 year 3.5 1.9 Middle zone 2 year 5.0 3.3 3 year 7.1 7.9

1 year 3.2 1.3 Inner zone 2 year 3.6 1.8 3 year 6.8 5.6

Table 1, based on the transect data, shows average wall thicknesses of fibres of two size categories (less than and greater than 20 J1m in diameter) in the three zones of each of the specimens. It can be seen that in the outer zone (i.e., centripetal fibre-caps of the inner part of this zone) where thickening is almost fully developed, there is little differ­ ence in wall thickness between fibres of the two categories, the larger fibres in fact having somewhat thicker walls than the smaller. In the free fibre strands of the middle and inner zones, however, larger fibres in the 1- and 2-year-old culms have thinner walls on average than the smaller fibres. In the 3-year-old culm, thickening of the larger fibres is more advanced, especially in the middle zone, though in the inner zone the larger fibres still have on average somewhat thinner walls than the smaller. A small increase in average thickness occurs between the I-year-old and the older culms in the inner part of the outer zone (Fig. lIA). Measurements indicate a 21 % increase in wall thickness from one to two years and a further 10% in the third year. In the middle zone (Fig. 11 B), fibre wall thickness in the centripetal free strands increases by 53% from first year to second and by a further 95% in the third year. In the inner zone (Fig II C), there is no significant increase from first to second year but an increase of about 190% in the third year. Figure 11 also indicates that, in the middle and inner zones of the 1- and 2-year-old culms, the more peripheral fibres have thicker walls than those nearer the centre of the strand, most of which remain relatively thin-walled. In the 3-year-old culm most of the fibres, except still for some towards the centre, especially in the inner zone, are very thick-walled. The dip in the curve for the inner zone of the 3-year-old culm is produced

Fig. 1. Transverse section through the culm wall of the 2-year-old specimen showing character­ istic variation in bundle morphology. Broken lines divide the culm wall into three equal zones. x 20. - Fig. 2. Mid-zone bundle from a very young culm showing fibre wall thickening confined to the caps adjacent to the vascular tissues. The free fibre strand consists entirely of thin-walled cells but is clearly distinguishable from the ground tissue. x 100. - Fig. 3. A typical mid-zone bundle of Type IV from the 2-year-old specimen. x 100.

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Downloaded from Brill.com10/07/2021 11:08:41AM via free access Murphy & Alvin - Fibre maturation in Bamboo 153 by the presence of some fibres which are still thin-walled. There is thus still consider­ able potential for continued wall thickening after the third year. Increase in fibre wall thickness over three years necessitates, of course, the presence of a persistent protoplast. Frequent observation of nuclei provides ample evidence that the fibre protoplasts do indeed persist. Transverse sections of good quality, especially of the 3-year-old culm, were difficult to cut, and perhaps for this reason, cell contents are seldom observed. However, we have prepared longitudinal sections which fre­ quently demonstrate the presence of nuclei (Fig. 8, 9). Longitudinal sections (Fig. 8, 9) also show the presence of septa within many fibres, especially in the 3-year-old specimen. Septa have been reported in a number of bam­ boos (Ghosh & Negi 1959; Parameswaran & Liese 1977) and we have found that in Gigantochloa they are of the same type as have previously been reported. Septa are apparently formed in fibres which are already thick-walled though there is much vari­ ation in wall thickness; they also occur in fibres of different diameters. Septa divide up the fibre into more or less equal uninucleate portions, and presumably originate from cytokyneses. Many septa are thick, with layers of wall thickening continuous longitu­ dinally with the inner layers of fibre wall. We agree with Parameswaran and Liese (1977) in interpreting these as older septa formed initially when the fibre was thinner walled, so that after septa formation wall thickening was laid down on both the septa and on the longitudinal walls. Most commonly septa within an individual fibre appear to be of similar age, as if septation occurred at about the same time. However, occa­ sionally, especially in the 3-year-old culm, fibres are seen containing septa of different thicknesses and presumably therefore of different ages (Fig. 9). Comparison of the three specimens suggests a steady increase in fibre septation with age. Septa are widely distributed within the culm wall but are seldom seen in the small fibres at the periphery or in fibres close to vascular tissues; they occur mainly in the fibres of the free strands and the larger centripetal fibre caps. Mean culm density was 470 (34) kgm- 3 for the l-year-old specimen and 525 (6) and 514 (44) kgm- 3 for the 2- and 3-year old specimens, respectively (figures in paren­ theses are standard deviations). Clearly, culm density increases between year one and later years, but that there is no observed increase from year two to year three seems anomalous as microscopical comparison (Fig. 5, 6) as well as measurements of fibre wall thickness (Table I and Fig. 11) indicate a substantial increase in the third year. The 3-year-old specimen was also noticeably more difficult to section. Mohmod et al. (1990), using fresh Malaysian material of Gigantochloa, reported a progressive in­ crease in culm density with age over three years, though the increase from year one to year two was much greater than that from year two to year three. The reason for the

Fig. 4. A typical Type III bundle from the outer part of the middle zone of the l-year-old speci­ men. x 100. - Fig. 5 & 6. Free fibre strands from bundles comparable to that in Fig. 4 from the 2- and 3-year-old specimens, respectively. x 100. - Fig. 7. Portion of a free fibre strand from the inner zone of the 3-year-old specimen, showing many incompletely thickened fibres, includ­ ing some still thin-walled. x 200. - Fig. 8 & 9. Longitudinal sections through the 3-year-old specimen showing septation of fibres. s = septum, n = nucleus. x 400.

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30

28

26

E::t ~ 24 B'"' v E 22 ;a~ v 20 .n'"' ~ 18

16

14 1 2 3 4 5 6 7 8 9 10 Interval across transect Fig, 10. Fibre diameter. Tangential transects through fibre bundles in the outer (+-+-+), middle (11-.-.) and inner (.A.-A.-A.) zones of the culm wall. Based on combined data from all three culms. apparent anomaly in our density results may lie in variation in the relative amount of ground tissue parenchyma between individual culms. It is perhaps noteworthy that in the three 3-year-old specimen there were relatively fewer free fibre strands.

DISCUSSION

Although the bamboo culm typically attains its full height and develops foliage in its first growing season, some continuing later development of the upper photosynthetic portions and eventual flowering may well demand augmentation of the mechanical sys­ tem within the main axis. Unlike with secondary thickening, bamboos have no potential for any increase in the extent of mechanical tissue once the primary body has been laid down. However, the individual fibres with their retained protoplasts inter­ connected, as they are, by a well developed pit system, retain for a long time the poten­ tial to lay down additional wall material, a process which will of, course, be limited ultimately by fibre diameter. Thus, the possession of fibres of broad diameter will pro­ vide the culm with greater potential for continued development of mechanical support. Our numerical results are admittedly limited but are backed up by microscopic evi­ dence. They confirm the observations of Fujii (1985), who in a study of fibre wall development in Pleioblastus chino reported that larger fibres continued wall thicken­ ing well into the second year. They also agree with the recent investigations of Liese and Weiner (1996) on Phyllostachys viridiglaucescens, who report continued fibre wall thickening over the first two years. These latter authors report also a second phase of fibre wall changes, involving further thickening in much older culms (9-12 years). We have found that in Gigantochloa, even in the 3-year-old culm, there still remains po-

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121 A L EIO -3 C/O ~ 8 c ~ u :s 6

~ 4 ~ ,e 2 ~

0+1----~--~---+--_1----+_--_+--~----+_--~ 2 3 4 5 6 7 8 9 10 12.------. '""' ,B ]'10

C/O ~ 8 .Q u :s 6

~<) 4f ~ • • • .---' .Ep:; 2

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~ 8 c ~ u :s 6

~ 4 ~ ,e 2 ~

0+1--~~--+_--~--~--_1----~--+_--_r--~ 2 3 4 5 6 7 8 9 10 Interval across transect Fig. II. Fibre wall thickness. Tangential transects through fibre bundles in (A) outer, (B) middle and (C) inner zones of the culm wall. Each based on averages of data from three separate transects. +-+ = one year old, .-. = two year old, and .A.-.A. = three year old material. tential for continued wall thickening in the larger fibres of the free fibre strands, most notably in the inner parts of the culm wall. Fibres of the vascular caps, being on aver­ age smaller and undergoing wall thickening earlier, will have little potential for further wall thickening in later years.

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It is noteworthy that the fibres of greatest diameter in Gigantochloa are contained within the free fibre strands and can be clearly recognised even in the very young culm. It is possible that during the early stages of tissue differentiation the 'release' of blocks of potential fibres into the ground tissue may allow some of the elements within them, especially those in the interior of the strand, to expand to a larger final diameter. Work is required on the origin and early development of these strands in bamboos with bundles of Types III and IV. Septate fibres are widespread among vascular plants, having been reported in palms (Tomlinson 1961), in wood fibres and sometimes phloem fibres of many dicotyledons (Metcalfe & Chalk 1950) and in Gnetales (Rodin 1975). According to Parameswaran and Liese (1977), the septa in wood fibres in dicotyledons differ from those in bam­ boos in usually lacking thickening and lignification. The widespread occurrence of septate fibres among vascular plants strongly suggests that they are of functional sig­ nificance, though this is unknown. In bamboos it is possible that they afford some protection to the continuingly important living protoplast.

ACKNOWLEDGEMENTS

We are grateful to the Forest Research Institute, Malaysia for permission to harvest labelled culms and also to Mr Ian Morris of the Biology Department of the Imperial College for photographic assist­ ance. We gratefully acknowledge the receipt of a grant from the Botanical Research Fund in support of this work.

REFERENCES

Alvin, K. L. & R.I. Murphy. 1988 Variation in fibre and parenchyma wall thickness in culms of the bamboo Sinobambusa tootsik. IAWA Bull. n. s. 9: 353-36\. Fujii, T. 1985. Cell-wall structure of the culm of Azumanezasa (Pleioblastus chino Max.). Moku­ zai Gakkaishi 31: 865-872. Ghosh, S.S. & B.S. Negi. 1959. Anatomical features of bamboo used for paper manufacture. Cellulose Res. Symp. CSIRO, New Delhi: 139-148. Grosser, D.& W. Liese. 1971. On the anatomy of Asian bamboos, with special reference to their vascular bundles. Wood Sci. Technol. 5: 290-312. Liese, W. 1985. Bamboos - Biology, sylvics, properties, utilization. Schriftenreihe Gesellschaft Techno!. Zusammenarbeit. No 180. Eschborn. Liese, W. & G. Weiner. 1996. Ageing of bamboo culms. Wood Sci. Techno!. 30: 77-89. Metcalfe, C. R. & L. Chalk. 1950. Anatomy of the Dicotyledons. Clarendon Press, Oxford. Mohmod, A. U., T. Wan, A. Wan & A. Fauzidah. 1990. Anatomical feature and mechanical prop- erties of three Malaysian bamboos. J. Trop. For. Science 2: 227-234. Parameswaran, N. & W. Liese. 1976. On the fine structure of bamboo fibres. Wood Sci. Techno!. 10: 231-246. Parameswaran, N. & W. Liese. 1977. Structure of septate fibres in bamboo. Holzforschung 31: 55-57. Rodin, R. J. 1975. Fiber and laticifer development in Gnetum. Abstr. XII Internat. Bot Congr. I: 231. Tomlinson, P. B. 1961. Palmae. In: C. R. Metcalfe (Ed.), Anatomy of the . Claren­ don Press, Oxford.

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