Professor Alan Wardrop (1921-2003), botanist

Professor Alan Wardrop

Alan Buchanan Wardrop was born in Hobart in 1921. Wardrop was educated at the University of Tasmania, where he obtained a BSc in 1942 and an MSc in 1944 for his work in botany and chemistry. He then spent 1944 and 1945 training RAAF air crews. In 1945 he joined the CSIR (later to become CSIRO) Division of Forest Products and in 1946 was awarded an overseas research scholarship which led to a PhD in botany from the University of Leeds in the UK in 1949. He then returned to CSIRO, where he rose to the level of senior principal research scientist and officer-in-charge of the Section of Wood and Fibre Structure. He was awarded a DSc by the University of Melbourne in 1958.  

Wardrop left the CSIRO in 1964 to return to the University of Tasmania as professor of botany. In 1966 he became the foundation professor of botany at La Trobe University, where he remained until his retirement in 1986, upon which he became emeritus professor in the Botany Department. He played an active role in the academic administration of the university. Professor Wardrop passed away in 2003.

Teachers' notes to accompany this transcript.
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Interviewed by Dr Max Blythe in 1998.

Contents


A well-grounded childhood

Alan, you were born in Hobart, in July 1921. Were there other children in the family?

I had just one elder brother, John.

Your surname is somewhat unusual.

I believe it is quite a common name in Scotland, particularly on the west coast. That is where my father, James Wardrop, was born. He was about three when he migrated to Tasmania with my grandparents in the late 1880s.

Did your parents support you in gaining an education and going into science?

Well, my father was a legal officer working for the Tasmanian government. There wasn't any science in the family, but he was always very supportive of me in anything I wanted to do in relationship to such things. My mother was never opposed to my ambitions but was more protective, – I don't think she wanted me to have to work too hard in life.

Hobart in the 1920s, when you were growing up, must have been relatively quiet.

Yes. It probably had only about 50,000 people. Its geographical setting is very nice, with Mount Wellington just behind, and the beauty of it was that you could walk to the summit and back in one day, through all sorts of vegetation and so on. That was a favourite recreation for me, whether I went alone or with friends.

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Helpful influences at school

What do you remember about beginning your education?

I went to a state primary school – the details are a bit of a blur now – and then to the state high school in Hobart. It was not very large but it had really good teachers and I enjoyed being there.

Did the science teaching in the high school figure well?

Yes, it was very good. In particular, Gordon Brett, who was in charge of teaching physics, made the first chromosome counts of eucalypts and published that in the Royal Society of Tasmania – quite remarkable. I imagine he must have been influenced by the new techniques of seeing chromosomes about that time, but it was unusual for a schoolteacher to have the time to do such a thing. I was impressed by his approach to scientific research: 'If you are going to get into a question, get yourself well prepared, read all you can, and if possible try and see if something doesn't agree with what you think.'

There was a very good chemistry man as well, Victor Crohn. As a matter of fact, the place being so small, you tended to get really good teachers because often they would, particularly in languages, teach part-time at the university and at school.

Would you say you stood out in anything at school – in sports or reading, or academically?

Ahh, in sports I met all requirements! I don't think you could rate it higher than that. I did read a lot, certainly, but I can only say I always did quite well in academic things.

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Building a foundation in biochemistry

I suppose that by the time you got towards your higher school certificate, it was obvious that you would look for a university place. Were your parents supportive of that?

Yes, particularly my father. We were not well off, though, and I think one reason my brother hadn't gone to university might have been a financial constraint. But I was able to get a university scholarship – worth about £50 – which was a great help.

I believe that the University of Tasmania was quite small.

Yes. There were only 15 or 16 students doing honours in the whole of biology.

What did you intend to do at university? I don't suppose you meant to focus on botany right from the start.

For some reason I had a feeling that I would like to do biochemistry, so I aimed at majoring in botany and chemistry. I had done chemistry at school, but biology wasn't taught to males at that time and biochemistry wasn't taught at all in Tasmanian schools.

Did you continue with any physics?

Oh yes. We had a very rigidly structured degree. In first year we did four subjects: physics, chemistry, mathematics and biology. By third year I ended up with botany and chemistry. We had lectures all the way through, complemented by lab work.

So you had a good conventional scientific background. Were there any particular lecturers who fired your mind?

There were a number of them. One quite good physics man, Lester McCauley, had an interest in biophysics, studying in particular (as a physicist) the electric fields around growing roots. He had done his PhD in England – as a student of Rutherford, I think. He stood out for us, however, chiefly for his eccentricity in his personal dress, his manner of talking to you and that sort of thing.

In botany, Professor Gordon was very good. I think he was a student of Bowers, from Edinburgh; he spoke with a strong Scottish accent. And we had just one Chair of chemistry: Edwin Kurth was a chemical engineer, and was engaged in a lot of wartime programs.

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Scientific war efforts

You didn't go straight on to do a PhD, did you?

No. Until about 1946 or '47 there were no PhDs given in Australia and normally you had to go to England or somewhere else overseas for that, but an MSc was the nearest I could do during the war.

Was your MSc course entirely a taught one?

No. We had lectures in physical, organic and inorganic chemistry, but instead of going into the Army to do something towards the war effort we had a project to do, which extended about 18 months. The one that I had was hydrolysing the cellulose of wood to produce monomer sugars which could then be fermented to alcohol, allegedly as a fuel substitute. It didn't seem to me that we were ever going to make enough alcohol to affect the war effort, but it did involve an interesting study of the kinetics of cellulose hydrolysis.

I've been trying to work out which division of science your MSc was actually in.

Well, you would have to say it verged on chemical engineering. We would cook up wood and measure how much had been hydrolysed and then what the sugar yield was, and that sort of thing. Of course, wood hydrolysis had been a common procedure in World War I, when huge kilns were built in Europe to make alcohol. I suppose people had forgotten it all until World War II started.

What other projects were the postgraduate students put onto?

In the Physics Department a lot of work on optical munitions – making lenses and that sort of thing – was going on, but I don't know the details of it.

And as I mentioned, the professor of chemistry was involved in a project. There was a great fuel shortage, so they used to make portable producer-gas units which were carried behind motorcars on little trolleys. Producer gas is mainly carbon monoxide and nitrogen, and you had to blow air over a bed of charcoal, so kilns were built to make very high-carbon content charcoal. One of the 'Kurth kilns' which the professor designed might still be in the hills here in Melbourne.

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Going into the Services

I gather that it was straight after your MSc that you went into the RAAF. Your brother had gone into the Services too, hadn't he?

Yes. He was in the Army for pretty well the whole of the war. He was up in the Pacific Islands.

While you were at university, did the students feel you were missing the war?

I think that was prevalent. As science students we were put into reserved occupations. I don't think I was terribly conscious of that for a long time, but when it came to actually doing work allegedly supporting the war effort, it all seemed pretty useless. I don't want to pose as a great patriot or anything, but going into the Services was the thing to do.

I suppose that by then, late 1943 or so, things were starting to change a bit more favourably. Anyway, I did get in to the Air Force and I started training as a navigator. Our initial training was at Balnarring, on Western Port Bay in Victoria, after which we went to Mount Gambier, in South Australia, for the actual air navigation. And then from there we would do exercises.

Having been relatively isolated in Tasmania for most of your previous life, did you find this period a bit of a revelation?

Well, I found it very interesting. I think it did me a lot of good, in the sense that you got a different perspective on other people and that sort of thing. But after the atomic attack on Japan, it was obvious that the war was coming to an end and we began agitating to get out of the Air Force.

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Starting to look at cell wall structure in wood

Did you get another job fairly quickly?

Yes. I don't recall the exact circumstance. I think I just saw an advertisement for the CSIR, as it was then – the Council for Science and Industrial Research. Later it became the CSIRO.

The Division of Forest Products had about six sections, one of which was on wood structure, with Eric Dadswell – a very eminent wood anatomist – in charge. The particular importance of the section's work was in identifying wood by the arrangement of the cells in it, and also in interpreting the gross properties of wood: its shrinkage, and its strength. But they could see that if you were going to extend this work into wood properties and also into the very important fledgling pulp and paper industry, you needed to know something more than the gross anatomy of the wood. I wanted to look particularly at the very fine structure of the cell walls.

I have always thought of the matrices of specially developed cells which make up wood as massive structures of cell walls, rather like thickened honeycombs of tissue. So these huge trees make me think of great structures of cells that were thickened and then died out, leaving their skeletal structure.

That's right, yes.

The division that you joined was very large, I believe. Wood was quite a priority.

Yes, it was. For example, complementing the wood structure section there was one on wood chemistry, a separate one on physics, and a number of very applied sections on wood preservation, timber mechanics and so on. There would be 20 to 30 people in each section, so it was quite a big laboratory.

They were all in the same building in South Melbourne, so you got good interaction. You could get any information you wanted, particularly with chemistry but also physics. And of course as a lab it had wonderful backup, workshops, if you wanted anything made. (The casino is now on that site, but the division is thriving and well at Clayton.)

That must have been a fascinating time to join the division. There was an interest in the new timbers that had been found during the war in the Pacific area, wasn't there?

Yes. There were many new species coming in all the time, because there was usually a forestry unit with the armies and they would send specimens. But after the war, fortunately for me, there was this suggestion of looking at the structure of the cell walls, which relate to the shrinkage and swelling of the wood as a whole. I didn't particularly want to get into the wood identification kind of thing, so I started to look at what the fine structure was, and the composition, just reading about it.

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How wood is constituted

Then you came across a widely-debated controversy. What was that about?

It was about what the organisation of the cell wall was. I think I should say here what the main wood components are.

In a conifer wood, the main component of the cell walls – about 60 per cent – is cellulose. In a hardwood such as eucalyptus, it is a bit less than 50 per cent. Besides the cellulose, there are non-cellulosic polysaccharides, carbohydrates, making up about 20 per cent. And the remainder is a non-carbohydrate component called lignin, which is basically a polyphenyl.

It is important, when you start looking at the structure of the cell wall, to recognise the molecular nature of cellulose. It is made up of individual molecules, consisting of residues of the sugar glucose, strung together in a very long straight chains of perhaps 10,000. A cellulose molecule would be long enough – but not wide enough – to be seen in an optical microscope. These very long straight molecules of cellulose are aggregated laterally, the lateral aggregates being called microfibrils, and they are partially crystalline. They have perfect arrangement in three dimensions, which means that you can detect them using the techniques for studying crystals, such as polarised light or X-ray diffraction, and so we developed these techniques there at that time.

In explaining the differentiation – the development – I might use the terms 'tracheid', meaning the conducting cells in a conifer, or 'fibres', as the strengthening cells in a hardwood, and occasionally I might interchange the terms.

In the development of the individual tracheid, it is the same as dividing cells just between the bark and the wood in a stem, the cambial layer.

Constantly cutting new cells to the interior?

That's right. In that differentiation from the cambium to the tracheid, there are dimensional changes – the cells expand laterally and longitudinally – during which the cell wall is very thin and is called a primary cell wall. When the dimensional changes have ceased, there are laid down on the inner surface of the primary wall a succession of layers of cellulose and non-cellulosic polysaccharides – the S1, S2 and S3 layers of the secondary wall. When the cell begins to approach maturity, there is the additional deposition of lignin, which begins at the primary wall on the outer side.

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Conflicting views of cellulose arrangement

So where did the controversy come in?

Well, there were two views as to the way the cellulose, in particular, was arranged in the cell wall.

One would think that the existence of these layers would be readily recognised. They can be readily distinguished if you examine them in polarised light, in that because of the crystalline structure of the microfibrils the optical properties of these three layers are different – they appear with different grain structures. One of the main workers in this field at the time was I W Bailey, at Harvard University, and he proposed a model very much like the one I have been talking about.

A contrary conclusion was formed in Leeds, however, by R D Preston. He was a physicist working in the Botany Department because the professor of the time wanted someone trained in physical sciences to be approaching botanical problems. (Preston was working with a very distinguished man, W T Astbury, who did a lot of work on proteins, particularly structural proteins.) Preston built a beautiful little X-ray spectrometer, with which you could get a diffraction diagram of just a single tracheid. And the evidence he obtained from the X-ray diffraction diagram was of only one helix in the whole cell wall.

So while Bailey in America was saying that there were three helical developments, three sheathing collars, Preston was not seeing that?

Oh, he admitted there were three layers, because he too could see them in the polarised light. But because he could only detect one helix from the X-ray data, he postulated that although the mean orientation of the microfibrils was the same in all three layers, the dispersion or the diffraction of orientation in each layer was different. So he proposed that in layer S1, for example, the mean orientation was the same as in S2 but, if you like, the microfibrils wobbled about that orientation. And the same for the S3. On this argument he could explain the optical heterogeneity of the cross-section and the fact that he could only obtain one layer. (I should tell you that I got to know both Preston and Bailey very well in later years.)

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Solving the controversy

Obviously you became interested enough to want to become associated with that problem. Is that why you applied to go to Leeds?

That's right. The CSIRO were offering postwar fellowship scholarships for people who wanted to go away, and I wanted to solve this problem of the different models that had been advanced. Everybody agreed there were three layers, of different optical properties, but was there only one helix or were there three?

Did Preston set you onto this problem for your PhD?

Yes. It was close to his heart. And the solution was really very simple. Imagine a cross-section of the cell, showing the three layers, S1, S2, S3. Now imagine we can cut, with a good microtome, a cross-section. As we alter the plane of section, the optical properties of these three layers will change. The story behind that is too complicated to go into here, but what we did was to section them, starting with a transverse section, and measure the optical properties of the three layers. Then we cut a section steeper, and again steeper, until we got to a longitudinal one.

Now imagine that you plot these optical properties. If there were three layers, in the Bailey model there would come a time, in cutting these sections, when the plane of the section would be in the plane of the microfibril and you would get a maximum of an optical quantity you are measuring. It is called the biorefringent. On this model you would get three curves, one for each layer. But on the Preston model you should have only one. Well, there were three.

It must have been difficult to go and tell that to your PhD supervisor, when he had staked so much of his reputation on there being only one.

It must be said enormously in Preston's favour that he was as hell-bent on getting a solution to this as anyone. At morning tea every day it would be, 'How's it going? How are the measurements?' It was a very congenial sort of thing. And the proof appealed to him because it involved measurement of a physical quantity, with quite complicated interpretation of the optical properties, this biorefringent, at the different planes of section. Although he pulled out every stop in arguing his case – he had a particularly good ability to interpret the optical properties – he acknowledged that we had a clear result. So that was good.

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Highlights and footnotes of the Leeds sojourn

Tell me a bit about your voyage to Leeds.

It was a broadening experience. We went on a ship called the Almeida, which had been torpedoed during the war and then mended by filling up the whole bottom with concrete. So the bowels of the ship were all concrete. From Melbourne to Liverpool took us 7½ weeks – not the best of sea travel, but quite enjoyable. And a number of people who were quite well known in Australia were aboard at the same time, such as George Humphrey, who was Chief of Fisheries at the CSIRO, and Edgar Mercer, who was in the Wool Division.

When you got to Leeds, did that Botany Department strike you as impressive?

Well, the department was in a row of old terrace houses on the Leeds campus; they hadn't started postwar building or anything of that kind. As a matter of fact, it nearly brought me to a sticky end. One of Preston's X-ray diffraction units involved a reciprocating anode, which meant that the target moved backwards and forwards with it, and of course this caused vibration in the operation of the instrument. One day, when I was in the lab – in a house which I think had been plastered for centuries! – trying to line this instrument up, all of a sudden there was a terrible crash and the whole assembly of fluorescent lights came down on top of me and the instrument. It turned out that the lights had only been screwed into the plaster and not into the rafters, and with the vibration of the building they just came loose and fell. So you might say there were some highlights of my time there.

When I arrived, the very well-known Professor Priestley (who had introduced Preston to the department) had just retired. The new professor who came in, Irene Manton, is well known in phycology. She was very keen on electron microscopy as well.

With such distinguished botanists as Priestley, Preston and later Pearsall, that was a remarkable department.

Yes. And they migrated all over the place.

Preston can't have been all that much older than you.

Oh, there is quite a difference. He will be 90 this year and I am 77. But he is young enough to be a very good, lively person to discuss things with.

Would you say he has been undervalued in history?

I think he has, especially in his impact on the timber industry and that sort of thing. He is a very eminent scientist, and was elected quite early to the Royal Society of London. And well, he is a Yorkshireman and I think he had a devilish streak in him – if people tried to be a bit pompous, he'd deal with them by addressing them in dialect. But he was a very nice chap. Even though at times we had quite extended arguments, the way it always ended, late in the afternoon, was, 'Well, let's go up to the pub and have a beer.' There was never any acrimony. Also, he had interesting students around that he was supervising at the time, in different ways. He seemed very popular.

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Persuading annual growth rings to cast light on wood properties

Can you give me an example of Preston's contribution to timber science?

Yes. A cross-section of a stem, we'll say in a conifer, shows growth rings formed annually. It had been observed by German botanists in the 19th century, by measuring the length of the tracheid in successive growth rings, that the length of the cells progressively increased.

Preston made a very important observation about the orientation of the microfibrils in the S2 layer. As the cells increased in length, the angle of the microfibrils decreased. The microfibril orientation in the early cells, say in the first year, would be rather flat, and then in successive years it would get steeper and steeper. Now, to take just one helix at a time, if the orientation is flat, water which is present in the cell walls can't penetrate the crystalline cellulose of the microfibril. So that determines whether the longitudinal shrinkage is great or not. If the helix were flat, you would get a high component of longitudinal shrinkage; if the helix were steep, there would be mainly lateral and very little longitudinal shrinkage. This gave a perfect way to study the effect of microfibril orientation and wall structure on properties, by dissecting wood from successive growth rings.

So Preston was showing the wonderful versatility of change.

Yes, but he was not recognised. He was the first, I think, to establish this relationship. And it is a beautiful experimental model. When I got back, we hammered this very hard, measuring whatever properties we could – shrinkage, inverse swelling, and also mechanical properties like breaking load of the wood. We were able to show a relationship between the orientation of the microfibrils and the breaking load of individual tracheids.

I suppose this work would ultimately enable you to show microscopically the actual properties of a wood – to say quite a lot about its breaking strength – even if you had not seen it before.

Yes, once we knew what the wall organisation was. Remember, this was no one-man band: in that division we were very fortunate in having the chemists and physicists around so that we could always go and get advice on these problems. If you wanted to measure shrinkage, there was somebody there to tell you how to do it in the best possible way.

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Coming into contact with the problem of reaction wood

You spent two years in Leeds, resolving that cell structure debate for your PhD. When you came back to CSIRO did you receive some recognition of the magnitude of what you had done?

Yes, but it didn't make clear the relevance of solving what model was right to the properties. We got on with the microfibril structure work, installing all the instruments to do it and buying our first little electron microscope. It used to sit on a bench, with the high-tension works under the bench. This was in the 1950s, so the lenses were permanent magnets – in the modern ones, they are all electron magnets – and its best resolution was about 10 nanometres. But we had it, and we had the X-ray diffraction unit, good optical and polarising microscopes, and so on.

Alan, would you like to explain to me the interest in reaction wood which you were developing at that time?

There were two things we got interested in about then. One arose from the dimensional changes occurring with the differentiation of the tracheid: the question was what changes in wall organisation, microfibril orientation, occurred during those dimensional changes. And also, through the studies looking at the properties in successive annual rings, we came into contact with the problem of reaction wood – an old problem which the foresters of Europe were looking at in the 19th century.

Consider the axis of a tree and one of its branches. In a conifer, if you cut a cross-section of the branch and look at the annual growth rings, you will find they are eccentric towards the lower side, and the wood there is very abnormal, quite different from the structure I was telling you about. Specifically, the rings are wider; and the tracheids are shorter, the orientation of the microfibrils in the S1 and S2 layers in the tracheids is very much flatter, and the S3 layer is absent. This is popularly known as compression wood, simply because with the weight of the tree that side would be under compression. The interesting thing arises if you suppose that the growing tip of a stem – the leading shoot – is cut off. Normally, the branch will then take over, curving up to become the leading shoot. But this is a massive structure, tremendously strong. In a simple plant like a flower stalk, any movement of the stalk is brought about by a differential growth on the two sides. That can't happen here – this is very thick and very strong, so it has to be bent. The development of reaction wood, or compression wood in this case, involves an actual bending of that massive structure.

Is there a tension in that compression wood, then, that will provide an uplift force?

It pushes the branch up, to put it crudely. But if a thick branch is to bend when the leading shoot is chopped off, a great compressive force must be developed. You can establish that the compression wood is the site of the force, because if you kill the cambium (which generates new tissue) on the side away from it, nothing happens – the compressive force causing the bending is associated with the compression wood. Just how the level of lignification and the different cell wall organisation operate, I don't know. We drew attention to the problem, or added to the evidence, but we never solved it.

We did a lot on that, and also quite a lot of field experimentation, killing the cambium on one part of the stem and not the other. And in the so-called tension wood in the angiosperms, in the hardwoods – a eucalypt, say – you have a similar eccentricity, but the opposite of what you have in the conifer. The structure is again different, abnormal. You can show experimentally that there are enormous growth stresses in the stem.

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Fig tree revelations

Wasn't some work related to tension wood done in the United States on fig trees?

Yes, by Martin Zimmerman and Barry Tomlinson at the Harvard Forest. (Zimmerman was from Switzerland and Tomlinson from Leeds, as a matter of fact.) Their interest in tropical vegetation led them to become very interested in the growth of the fig tree. These trees grow in the normal arborescent form, and from the branches they drop down aerial roots – very thin to start with, but when they hit the ground they go into tension and can be twanged! They are in tension and undergoing contraction.

As part of their field observations, Tomlinson and Zimmerman buried a drum with seed and soil, and marked the level where the drum was buried to. When the roots came down and went into tension, starting to thicken, they were strong enough to pull the drum quite a long way out of the ground.

When the anatomy of these aerial roots is examined, their reaction wood is found to be identical with the tension wood of the fig tree. Although we don't know all the details of the mechanism, this was a good demonstration that the formation of tension wood involved a contractile force. It's a marvellous set-up, really. You have got the stem pulling up and the root pulling down, making a tremendously stable structure. And to complete the story: after this contraction has ceased, then the roots form normal wood and act as a prop. So it is a wonderfully stable structure that develops. That was important to us in establishing the contractile nature of reaction wood.

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Coleoptile contributions to plant growth theory

What part did coleoptiles play in your studies of wood?

They came into the study of the differentiation of, say, the tracheids in a conifer, when cell division takes place in the cambium and the primary wall is formed. This is very thin, and is present during the time of growth or dimensional changes of the differentiating cell, before the secondary wall.

A favourite experimental object for studying the changes in the microfibril arrangement in the primary wall during dimensional change was oat coleoptiles. Imagine a germinating wheat seed of any kind. The first leaf as it grows up is enclosed by a slender sheath made up entirely of a single layer thin-walled parenchyma cells, with no differentiation. This outer sheath is the coleoptile, and the reason it is such a beautiful object for physiological study is that when it is first formed, up to about one centimetre long, it elongates entirely by cell division but after that time – and it can extend through about five centimetres long before the leaf breaks through – the elongation is brought about purely by the extension of the cells already formed. So when it is, say, two or three centimetres long, you can cut a bit out and pull the leaf out of the middle, and then you have got a little cylinder of cells. If you put those in a dish, they will extend, according to what you feed them, or the temperature. The coleoptile was ideal, since this is only a primary wall present, to study the arrangement of the microfibrils.

By this time we had an electron microscope so we could see the microfibril orientation, and we had all the ancillary equipment. So we started to grow the coleoptiles to different lengths, using chemical treatment to separate the individual cells one from another, and then looking at what their structure is. (You can look at this in a polarising light microscope too.)

I think you were about to collapse a hypothesis that had been made.

Well, people were going pretty mad with electron microscopy at this time and you could get all sorts of interpretations. This was prior to the development of good ultramicrotomes to allow you to section things, so you often had to break up the cell walls by very violent methods – by ultrasound or by putting them in a Mixmaster or something of that kind.

There had always been an argument about whether, when the extension period of growth in plants occurred, the growth of differentiating cells was generally uniform or tip. The view that the cells grew at their tips was based mainly on electron microscopy, and it had been advanced by two very eminent people – Frey-Wyssling and Kurt Muhlethaler, who were based at the ETH, in Zurich.

We were able to show, however, in a very simple set of observations, that if you isolate these cells you find that they are interconnected, one with the other, by little pores in the wall by which the cytoplasm communicates with the cell next to it. First of all we observed that these pit-fields, which are easily recognised in the optical microscope, did not increase in number during elongation or in distribution as you would expect if there was tip growth. So, since they did not increase in number, as far as we could see, was there a new cell wall forming? Was it different at the tip than at the middle? No, it wasn't. And then, to clinch it, we fed the coleoptile segments with radioactive glucose, C-14 glucose, and observed that the distribution of the newly formed cellulose was the same all over the cell.

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Under-the-counter research

Is it true that you were actually growing your coleoptiles under benches?

Yes. There was a sort of red-tape situation in which the Department of Forestry in Canberra was in charge of growing trees and the CSIRO Division of Forest Products just looked at the wood. So we had to keep a very low profile on growing anything. These coleoptiles posed no problem, you could grow them in a dish under the bench. But we had to relate this to wood.

Fortunately, at about that time the CSIRO Executive of the day wanted to assess how their labs were going, so James Bonner – from the California Institute of Technology – visited Plant Industry in Canberra and us in Melbourne. I told him about the compression wood story and that we needed glasshouses to be able to grow the plants under different conditions, and also about the coleoptile stuff. He quite liked all this, and after he had left the lab, the chief of the division came round to me and said, 'Well, I've talked to Bonner, and you can have your glasshouse for your coleoptiles.' Of course, we had them already, but it meant we could grow little trees, and we put a glasshouse on the roof of the lab then. That gave us a nice experimental set-up.

So in the very early '60s you could get tissue absolutely as you wanted it, and get right to the bench with it.

Yes, that's right. By the way, things have changed now and there is no problem about growing timber for research. We now have a Division of Forestry and Forest Products, all integrated in every way.

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Itchy feet in an enduring family landscape

By about 1964 you weren't so happy at CSIRO. Why was that?

It's hard to give a specific reason. In the organisation generally, not just in Forest Products, there were changes going on and a lot of discussion of what was pure research and what was applied – a silly discussion, in my opinion. It seemed to me that universities had a complementary approach of doing both research and teaching, and I just wondered whether it wasn't time for a change. I had got my DSc at Melbourne University some years previously, and I began to think that it might be nice to teach.

So you applied for the Chair of Botany at Hobart, and returned to Tasmania?

Yes, to get back into academic work. That was a very good department. My predecessor there was Newton Barber, a distinguished geneticist, so I was a bit of a change. It got off wonderfully well – we got a big grant for a very good Siemens electron microscope – but unfortunately my younger daughter became quite ill in Melbourne and it was apparent that we couldn't move her to Hobart. And also there were a lot of medical facilities in Melbourne. So I was faced with the problem of getting back to Melbourne, either to CSIRO if they would still have me after leaving, or to another job. La Trobe University was just being started, though (that was in 1966, and they took their first students in '67) and I was able to get the Chair of Botany there.

Alan, obviously your family was an important factor in your career, so would you tell us a bit about them? When did you meet your wife?

My wife Beulah is a mathematician, and we met at CSIRO. We were married in 1945, about a year before the opportunity of going to England arose. I had never been out of Australia but she had, just before war broke out. As a matter of fact, her father was a businessman and was travelling in Europe. So she was in Germany at those terrible times of the Nuremberg rallies and so on, and although she was still in her teens that forms an important part of her memories. Anyway, our first child, Martin, was born after we returned, and we have now four children, two girls and two boys.

And they have all done remarkably well.

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Multifaceted academic leadership

You were the first Professor of Botany at La Trobe, weren't you?

Yes. That was a most unusual experience, because very rarely can you plan or even watch something develop from the beginning. It brought with it an awful lot of administration, however.

This being such a new place, you must have had to design laboratories and to think about the kind of department you wanted.

Well, I had a very good lab manager to design the laboratories. I tried to make the department a pretty balanced one, because we had to think of the undergraduate teaching, not just the research.

So you included disciplines like ecology, taxonomy, anatomy, physiology and so on?

Yes. One thing we didn't have was genetics, but we were fortunate that the university created a Foundation Chair in Genetics, with Peter Parsons, and so all the genetics we wanted for our courses was in the same building, on another floor.

Could you continue your microfibril and reaction tissue work there?

Oh, I brought the problems with me. And the links with CSIRO were maintained.

I suspect you rather liked the teaching, but I wonder whether fundamentally you aren't a researcher – you like to base your teaching on having your feet firmly in a laboratory.

I think so. I must admit I do like teaching, but you never get anything the way you really want it. As the administration grew, the teaching became more of a chore, because you dared not take your eye off what money was coming in, and that sort of thing. And an interesting change occurred. When the university was starting, the Vice-Chancellor, David Myers, was working on the assumption that there would be twice as many physical science students as biological ones. Within about five years that was reversed – the swing was occurring all over Australia, I think – and that had tremendous implications on the buildings, of course.

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Fruitful applications of freeze-etching techniques

Quite quickly in the '70s you became involved in freeze-etching technique. Would you like to tell us about that development?

Yes, I'd like to mention it, because I think it was important at the time.

The problem if you want to look at the living contents, the cytoplasm, of a cell, as distinct from the cell walls, is that you have got to kill it. It is usual to use all sorts of violent chemicals such as osmic acid or chromium salts for that purpose, but this naturally causes artefacts in the killing of the cell. So a technique of freeze-etching was developed, particularly by Hans Moor – an associate of Frey-Wyssling and Muhlethaler, whom I mentioned a short while ago.

The technique is based on the fact that if you take, say, a yeast cell, and you freeze it – very fast or very slow – so it is at a temperature of liquid nitrogen, round about minus 190°, and then thaw it out again, if you control the conditions and rate of freezing and rate of thawing you can get about a 95 per cent survival rate. So they built a special machine to enable the whole operation to be done in a high vacuum. You would take frozen cells and put them on a little table, or bench, enclosed in a bell jar which could be evacuated. Built in to this instrument was a microtome knife which had liquid nitrogen pumped through it. You could then move this microtome knife round and shatter the cells sitting on the table – all this being done under high vacuum.

If you had the knife at a temperature a little higher than that of the liquid nitrogen freezing the cells, the water molecules would be picked out of the exposed structure. We would first of all cut the cells with the knife, and then raise the temperature of the knife a little bit and bring it back, whereupon the water molecules would pick out from the exposed surface of the cell. That's the etching part of it: if you had an organelle like a mitochondrion or a plastid in there, you accentuated its substructure by this etching process.

Then, built in to this same bell was a filament, so that having etched it you could evaporate metal onto it.

So that what was on the 'windward' side would stand proud as the metal coated it, like a snowstorm? And what was on the other side would shatter?

Yes. You then strengthened that metal coating with a film of carbon, and at the end of all that you let air into the whole thing and soaked this replica off the surface of a cell which theoretically could have survived the freezing. This was supposed to be a way round this artefact formation, so that when you examined it in the electron microscope, very beautiful images could be got. Of course, with time all this has evolved and people have published papers on the artefacts of freeze-etching! But that happens to everyone.

We got into that technique very heavily, and obtained some nice results in relation particularly to development of the nucleus in the cell. The money for that – £13,000 – came from three pulp and paper companies, of all people, and about a quarter of it from the university.

That was massive money in those days! It would be quite nice to know about some of the structures that you found.

The most spectacular results were achieved by three students in particular. One of them, Brian Fair, from Canada, was working with us. A freeze-etching image of a nucleus which has been shaved off shows that there are two membranes limiting the nucleus, and there are nuclear pores. At this phase of the cell cycle, the membrane breaks down when it divides, but you can see an incredibly regular array of the nuclear pores.

A picture illustrating the work of another student has the outer membrane peeled off and only the inner of the two showing. In the arrangement of the pores and the interaction between the nucleoplasm inside the nucleus and the cytoplasm outside, there is a substructure which changes with the exchange of the materials from the nucleoplasm to the cytoplasm. I won't go too far into that, because we didn't show it. There was also another student who did some very beautiful freeze-etching on collenchyma cells – which reverts a bit to the cell wall story.

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Cell extension: multinet or helicoid?

Let's turn to your collenchyma work, because in the late 1970s you went for a year to Nijmegen, in Holland, to work with somebody rather interesting.

I had corresponded an awful lot with Andre Sassen, at Nijmegen, who was interested in the cell wall particularly. I have mentioned the question of what were the changes in the coleoptile. The primary wall is much simpler; it just appears to be one layer when you look at it. And when you look in the electron microscope, the microfibrils on the inner and outer surfaces are different. On the inner surface they are in a flat helix or transverse, and then, no matter how much the cells extend, on the inner surface they remain that way. But when you look at the outer surface, you see that they become progressively disoriented. In a cell that is elongated a lot, they become perhaps nearly longitudinal on the outside but you have still got new ones forming on the inside, transversed. It can be seen that the new ones are forming transversely all the time, but as the cell elongates they gradually become disoriented.

From that simple observation of the difference on the inner and outer surfaces, a Dutchman, P A Roelofsen – of Preston's vintage, not of Sassen's – proposed a multinet hypothesis. I supported this hypothesis very strongly. I had evidence for it in differentiating cells from cambium, in coleoptiles, and the game seemed to be sewn up there.

But there also occurs in elongating tissues this collenchyma, which has very thick walls.

Yes, strengthening tissue but not as complex as the kind of wood tissues, the tracheids, we have talked about.

That's right. In an optical microscope it has a glaucous appearance. It is partly layered with pectin-rich (non-cellulose-rich) and cellulose-rich lamellae. What so interested us was that when we started to do the cell wall structure and so on, it was not like the multinet even though I tried to interpret it in multinet terms. If you looked at the lamellae in the thick walls of the collenchyma – shadow cast, in a longitudinal section – the interesting thing was that they appeared to have been longitudinally oriented or transversely oriented. The interpretation I put on that was that you had these lamellae and that they were disoriented in the terms of the multinet hypothesis. There seemed to be quite clearly microfibrils, transverse, and so on. This was about the time I went to see Sassen, and we were intrigued by this.

At about that time, a completely different concept was proposed by Bjorn-Paul Rolent, from Paris. We organised a little cell wall symposium in Nijmegen in about 1978, and Rolent came to that. His concept was different from the multinet interpretation and it seemed to be absolutely correct. That is, when the microfibrils are deposited, they are not put down longitudinally and transverse as that would suggest, but they are put down and then gradually shift in orientation as the wall thickens, to form a helicoid. And when you start to use very sophisticated staining techniques for primary walls, even those of the coleoptile, you can detect evidence for this helicoid.

So you'd got it right.

And I think he has got it right! We were trying to do it, but failed dismally. Working with Sassen we did quite a lot of work on the collenchyma as well as the rest, and still our own observations seemed to support the multinet. But at that meeting in Nijmegen I began to have serious doubts, after watching Roelofsen's very sophisticated staining methods. As the evidence has accumulated I think the helicoid concept is pretty generally applicable in extending cells.

That was a good time. Those cell wall meetings have gone on ever since. I think they had 14 or 15 people at the first one, and the last one, in Abyssinia, produced a volume with about 200 papers. That's nice, because you can keep your interest in these things.

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The continuing attraction of unsolved problems

So your strong interest in collenchyma and the microfibrils story continues to some extent in your retirement?

Oh yes. There's been a tremendous development in the techniques of molecular biology, and particularly immunocytochemistry, where you can look at living cells, and also the work on cellulose biosynthesis. In about 1982 I worked for a while with Malcolm Brown at Austin, in Texas. He worked initially with a bacterium, acetobacter, which generates cellulose, and he worked a lot on cellulose biosynthesis. But there are other people in the US, such as Devi Delmar, in California, and a group at the Research School of Biological Sciences, in the Australian National University – including one of our former staff members, Richard Williamson, working on cellulose biosynthesis. Early this year they published a very elegant paper.

Something I didn't get onto is the site of synthesis of cellulose in the cytoplasm, which all relates back into the structures. There are still unsolved problems. We don't really know what determines the orientation of the microfibrils. There is the interesting observation that their orientation parallels that of the microtubules in the cytoplasm, but the exact model of that interrelation I think is not yet clear. They are getting there, though, and there is this beautiful work on biosynthesis. It is a topic that someone like myself can still appreciate, even although I haven't done much about it. La Trobe very kindly provides me with a room, but my own research is becoming less and less.

Alan, it has been great today to go through the various phases of your career. We haven't had time to do justice to it all, but thank you very much.

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© 2017 Australian Academy of Science

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