The Journal of Indian Botany/Volume 1/May 1920/Note on Curvature of Cut Stems of Bryophyllum Calycinum

Layout 4

NOTE ON CURVATURE OF CUT STEMS OF BRYOPHYLLUM CALYCINUM

BY

P. F. Fyson and K. Venkataraman,

Presidency College, Madras.

Loeb in 1917 published (l) the results of a series of experiments on curvature induced in cut stems of Bryophyllum calycinum, sus- pended in moist air by threads, one at each end. He found, and published figures in support, that curvature, apparently due to gravity took place, whether leaves were left attached to the plant or not ; but much more strongly so, when a leaf near the apical end was left, than when all but a basal leaf were removed. He also found that roots were formed adventitiously, on that region only where the curvature was a maximum, and that root-formation was strongest, when an apical rather than a basal leaf was left. From this, he drew the conclusion, that special root-forming and curvature-producing substances (hormones) are manufactured in the leaf, and passed downwards, but not upwards, along with the products of assimilation.

The experiments described here, were made to test these facts and deductions on plants growing in Madras. Healthy stems of Bryophyl- lum calycinum were used and straight pieces 6 to 8 inches long were cut, and in every case the apical bud and the first visible internode, were removed to prevent continued growth, as was done by Loeb.

All the leaves were removed except one or both apical leaves or, one or both basal leaves, and the pieces suspended over water in a glass chamber. No particular difference was observed, which could be correlated with the presence of one or both leaves, whether at the base or apex: But those with one or both apical leaves curved more than those with basal. (Plate I, figs 1, 2 and 3) Loeb in explaining this as due to the formation of geotropic hormones in the leaf and their passage always down the stem, appears to have overlooked the possibility of the curvature being induced by weight, for no attempt apparently, was made to eliminate this. We found however, that if a third thread was used to support the piece at its centre of gravity or if the pieces were laid on a flat board (fig. 11) no curvature took place. (Compare in Plate I, figs. 8, 9, 10 with 5, 6, 7.)

In order to support the piece without introducing other factors, we attached to the piece near its centre of gravity a fine thread, which after passing over a pulley, supported a pan, in which weights were placed to counter-balance, the weight of the piece. Plate II, fig. 27 shows how this was done, and it will be seen that no appreciable curvature took place. In the course of the experiment a lessening of the weight, by loss of evaporated water caused the terminal threads to slacken ; but at the beginning of the experiment, the weight was evenly distributed and the lack of curvature cannot therefore, be due to any pull upwards, at the centre of gravity.

Since the region of curvature is in the weak still unlignified internodes near the apex (vide figs. 1, 2 and 3) it is clear that the bending moment at the point of curvature due to the comparatively heavy, succulent leaves, will be much greater when these are at the apical end, than if they are at the basal (compare figs. 1 and 2 with 3).

In another series of experiments living stems of Coleus still attached to the roots, were laid horizontally, the growing point and the first visible internode, being removed as before. Three pairs of plants were used, and in one of each pair, a pair of apical leaves was left, while in the other one or more near the base. Plate II, fig. 19-26 show the resulting curvature after 24 hours. It will be seen that no definite relationship, can be established between the amount of curvature and the existence or position of leaves Compare Nos. 19 with 20, 23 with 24, 21 and 22 with 25 and 26.

B. The root-forming ' hormones' — Our experiments failed likewise to support Loeb's theory, that root-formation is due to hormones produced in the leaf. Roots were formed freely on the lower side and especially at the region of curvature (figs. 1 and 4) of stems kept damp; but in no apparent relation with the existence or position of leaves. The presence of water, we found to be a much more important factor.

Pieces of stem were placed vertically, in the normal or the reversed position, and one end kept damp, by tying a wet rag round it. In every case roots appeared, whether at the upper or lower end, whether at the apical or the basal. This is exactly in conformity with Klebs results (7).

This occurred also with both isolated and attached leaves of Bryophyllum. Roots appeared always in a day or two, at the parts kept wet. Loeb's results (4) of hanging leaves in different positions, which led him to postulate a flow of root-forming substances towards the base of a leaf appear therefore, to have been accidental. In our experiments, leaves were hung in a vertical plane.

(1) by the petiole

(2) by a hole in the apical end

(3) by a hole at one side and

(4) horizontally by threads through two holes in the lamina. In every case provided the atmosphere was kept saturated, roots appeared with apparently equal facility at any notch.

G. Regeneration. According to Loeb, the growth of roots and shoots', at the marginal notches of a leaf is ordinarily inhibited, by suction of the stem and especially of the growing axillary buds. To verify this : —

(1) A piece of the stem of Bryophyllum had one leaf at the basal portion alone left, and it was left in a moist chamber supported in the horizontal position on a board. On the 7th day (Plate I, fig. 11) roots had grown out from the notches of the single leaf, and the opposite bud showed signs of growth. On the 14th day, the bud opposite the basal leaf had fully grown out and several shoots also had appeared from the notches of the basal leaf. Roots had also appeared from the under-side of the basal node.

Now, if really the opposite bud inhibited the growth in the notches of the basal leaf, one would expect that as the opposite bud began to grow, the roots that had grown from the notches of the basal leaf would not have developed any further. On the contrary, they continued to grow, and shoots also appeared from the notches. In short, the growth in the notches of the leaf, progressed side by side with the growth of the opposite bud. Hence, apparently no inhibi- tion had taken place.

(2) Two pieces of the stem of Bryophyllum, containing each a single node with only one leaf attached were suspended so that part of the lower leaf was sub-merged in water. After 10 days, it was found that the submerged notches of the lower leaf, in each case, had developed roots, while the opposite bud also was developing. (Plate I, figs. 12 and 13).

(3) A piece of the stem of Bryophyllum containing a single node with two leaves attached was arranged so that one of the leaves had a part submerged in water. On the 6th day roots were found to have grown out from the notches of the submerged lower leaf. On the 9th day the bud in the axil of the submerged leaf, as well as the bud in the axil of the opposite leaf had developed. The upper leaf withered and finally fell off, and is therefore not seen in the photo- graph (Plate I, fig. 18). A root had also grown from the piece of stem.

In this case again, if Loeb's inhibition theory were correct, one would expect that since each of the axillary buds inhibited the development of the opposite leaf, none of the two leaves should have grown any roots in their notches. As a matter of fact, the lower submerged leaf produced roots in its notches and its opposite bud also developed, as also the other axillary bud. From this we may conclude that neither the stem itself, nor the opposite bud, nor the axillary bud, exert any absolute inhibition, on the development of the marginal notches, and probably affect it very little.

D. Regeneration in leaves still attached to the living plant. — Several normal, erect, healthy plants of Bryophyllum were found in which roots and shoots had arisen from the notches of several leaves in contact with the parent plant. It should be mentioned that the preceding week had been a very rainy one. Most of the leaves jvhich had grown shoots at their notches were found to be of a slightly paler green colour, but there were also several green leaves among the number. In none of these stems had the terminal buds been injured or destroyed and they were growing quite well. In the case of one plant where the terminal bud, had been accidentally destroyed, the notches in the leaves had not grown out, but the axillary bud lower down, was starting to grow. The stems of all these plants were quite erect and to all appearance normal. There was no indication of their roots having been injured or of any other pathological condition as suggested by Loeb in his objection (5) to Miss Lucy Braun's very similar observation.

Loeb, indeed contended (l) that "when these leaves are attached to the stem of the whole plant, in the natural condition, they never produce roots and shoots in their notches." The chief factors which he mentions, as tending to prevent the formation of roots and shoots on the leaves when in the natural condition, are (l) the growth of buds on the main stem and (2) root-pressure. His explanation is that the formative materials which are required for active growth, are constantly flowing to the terminal bud from all the other parts of the plant and hence the notches in the leaves have no chance of growing. This explanation does not, however, apply to the instances cited above, as in all those cases, the terminal buds on the main erect stems were quite healthy and active.

Turning now, to the second factor that Loeb mentions, he says that the presence or growth of roots on the main stem inhibits the growth of notches in the leaves because. of root-pressure.

It is difficult to conceive how root-pressure can inhibit growth in the notches of the leaf, and Loeb does noc explain how it does. In the concluding passages of his paper (l), he says that attempts to induce growth in the notches of the leaves proved futile. He says " the writer left several leaves of the plant submerged in water for months without any results ", and therefore concludes that root- pressure is the inhibiting factor.

yith a view to find out, whether this was really the case, three leaves of a plant growing in the ground, which had not grown any roots or shoots in the notches of its leaves, were selected and they were left dipping in water contained in a basin, on the 2nd of December, 1919. Three days later on the 5th of December, roots were found to have grown out from the submerged notches of the leaf, to a length of nearly half an inch. On the 8th December, small shoots also had grown out from the notches.

These observations deny the inhibiting influence of root-pressure and suggest the moist external conditions as accounting for the growth. This is quite in consonance with the fact that the notches in the leaves of the plants had grown out just after a week of heavy rain, although root-pressure must then, presumably, have been considerable, because of the very little transpiration that was possible.

The following experiment appears to be conclusive. A pot plant of Bryophyllam was taken and the stem of the plant was cut off leaving only the last basal node with the two leaves attached. The cut end of the stem was sealed firmly with Canada balsam, to prevent water from being forced out and resulting in a diminution of root-pressure. The axillary buds of both the leaves, were destroyed. One of the two leaves was left dipping in a basin of water so that part of it was submerged. On the 6th day, roots were found to have grown out in clusters from the notches jDf the submerged portion of the leaf. Later on small shoots also appeared. Here if, as seems probable, there was considerable root-pressure, the formation of roots and shoots from the leaf- notches was not in any way inhibited.

Summary

Experiments were made to determine whether special root-forming and curvature-producing substances could be recognised as being formed in the leaves, as assumed by Loeb on the results of his observations on Bryophyllum calycinum. They were in the first instance a repetition of his experiments, but with certain modifications to eliminate the effects of weight. We find no evidence in support of the theory of special curvature-producing hormones, nor of root-forming substances. On the other hand our observations confirm those of Klebs on the effect of an 'excess of water on adventitious root-formation.

A further series of experiments on the effect of the growth of an axillary bud on the development of the marginal leaf-notches, failed to confirm Loeb's results. All differences in the rate and amount of growth of roots and shoots from the margins of the leaf, can in our opinion be sufficiently explained by the suction naturally produced, by a developing organ, on the water and sap locally available, without the assumption of any special inhibitory action. Development of the marginal notches we found to be induced readily by an excess o-f water, which appeared to be the sole determining cause. Moreover, our observations entirely negatived Loeb's supposition of an inhibitory effect of root-pressure on this development.

Literature Cited

(1) Loeb ' Rules and mechanism of inhibition and co-relation in the regeneration of Bryophyllum calycinum.' Botanical Gazette, Vol. LJ, p. 249.

(2) Loeb ' Further experiments on co-relation of growth in Bryophyllum calycinum ' Botanical Gazette, Vol. LXII, p. 293.

(3) Loeb ' Influence of the leaf upon root-formation and geotropic curvature in the stem of Bryophyllum calycinum and the possibility of a hormone theory of these processes' Botanical Gazette, Vol. LX1II. p. 25.

(4) Loeb ' On the production of equal masses of shoot by equal masses of sister-leaves in Bryophyllum calycinum' Botanical Gazette, Vol. LXV, p. 150.

(5) Loeb ■ Healthy and sick specimens of Bryophyllum calycinum' Botanical Gazette, Vol. LXV1, p. 69.

(6) Braun ' Regeneration of Bryophyllum calyoinum ' Botanical Gazette, Vol. LXV, p. 191.

(7) Klebs Willkiirliche Entwicklungsanderungenbei Pflanzen. Jena 1903 (Quoted Josfs Plant physiology, Eng., Ed., p. 336.)

Description of Plates

Plate I

Figs. 1 — 7. Pieces of Bryophyllum calycinum hung by two threads.

Figs. 8 — 10. Pieces hung by three threads, the middle thread attached at about the centre of gravity. The positions of the threads, which being invisible against the white back- ground, has been indicated in the original photographs by hand. Apical or basal leaves left as shown. All others removed and terminal bud and first visible internode destroyed.

Fig. 11. Piece of Bryophyllum left on a board. One leaf was left, the others removed as above. The opposite axillary bud has developed but this has not inhibited the growth

at the leaf notches. Note no bending.

Figs. 12 — 15 & 17, 18. Leaves of Bryophyllum- variously isolated and hung with lower part in water showing development at the marginal notches from the submerged parts.

Nos. 12 & 13. The opposite axillary bud allowed to develope.

No. 14. The opposite axillary bud removed.

No. 15. The stem cut in half longitudinally-

No. 16. Leaf hung dry, no;part submerged.

No. 17. No part of stem left attached to leaf.

No. 18. Opposite leaf left attached (but subsequently fell off).

Plate II

Figs. 19 — 24. Rooted plants of.Coleus bicolor placed horizontally. Photographed after *a few chays. One or both leaves near the apex or near the base left attached, all others and the terminal bud removed.

Nos. 25 & 26. No leaves left attached. A lateral bud has deve- loped near the base of No. 26.

Fig. 27. The method of hanging pieces by three threads, adopt- ed for Nos. 8, 9 & 10.