Photosynthesis: From De Saussure To Liebig.

The dawn of photosynthesis, characterized by the research of Priestley, Ingen- Housz and Senebier, culminated in 1804 with a historical essay of Théodore De Saussure. According to the historians, during the first half of the nineteenth century in which the genesis of the cell theory started off, the research on photosynthesis met a phase of stagnation. Indeed, the literature review of the period does not report particular innovation; however, several scientists (botanists, physiologists, and chemists) supported the thesis of De Saussure with a series of analyses that, in our opinion, deserve to be known. Mirbel, De Candolle, Raspail, Berzelius, Payen, Dutrochet, von Mohl, and other scholars attempted to expand knowledge on photosynthesis but were not able to arrive at a theory that was consistent with a functional mechanism, nor with a suitable chemical model to explain the transformation of the water and carbon dioxide into sugars. A classic case of such inadequacy concerns the discovery of chlorophyll. This compound, isolated in 1818 by Pelletier and Caventou, remained an enigma for many years and was never put in relation with the synthesis of starch. The accurate research of von Mohl led this scientist to believe that the granules of chlorophyll were entirely independent of starch granules, although in many cases these latter were observable inside the granules of chlorophyll. Only in the early forties, Justus von Liebig realized that the assimilation of carbon and hydrogen required a series of chemical reactions that, starting from some organic acids, ended in the formation of sugar. In conclusion, our analysis does not lead to define this period as stagnation but rather as transition, in which the concept of photosynthesis was clear, even though difficult to treat under physiological and chemical views. From the sixties, the researches of Julius von Sachs will open a new road, thanks also to the research carried out in the transition period.

THE CONCEPT OF PLANT LIFE: THE DAWN.

An elementary but correct concept of plant life has come to us in writings of Theophrastus who divided the plant life in its three basic stages: generation, sprouting, growth. This image of plants remained practically unchanged until the seventeenth-century, when the scientific method based on experimentation was introduced by Bacon. The invention of the microscope and the change of the traditional alchemy for an embryonic chemistry allowed some penetrating minds to look upon plants as highly complex living structures, to which had to correspond some specific functions. The observations and deductions of Mariotte, Malpighi, Grew and Ray revealed that the plant was operating as a real factory that, with the contribution of sunrays, changed inert matter in the components of plant structure and that these transformations could give an account for its concept of life. The subsequent work of Hales supported the concept of plant life as a materialistic processes planned by a divine architect. With Hales began a new phase of research, which reached its full development from the nineteenth century.

Elements of plant physiology in theophrastus' botany.

For thousands of years the plants were considered only as a source of food and medicine, and as ornamental objects. Only from the fifth century BC, some philosophers of Ancient Greece realized that the plants were living organisms but, unfortunately, their works have come to us as fragments that we often know from the biological works of Aristotle. This eminent philosopher and man of science, however, did not give us a complete work on the plants, which he often promised to write. From scattered fragments of his conspicuous biological work, it emerges a concept of nutritive soul that, in the presence of heat and moisture, allows plants to grow and reproduce. The task of writing a comprehensive botanical work was delegated to his first pupil, Theophrastus, who left us two treatises over time translated into the various languages up to the current versions (Enquiry into plants, On the causes of plants). The plant life is described and interpreted on the basis of highly accurate observations. The physiological part of his botany is essentially the nutrition: According to Theophrastus, plants get matter and moisture from the soil through root uptake and process the absorbed substances transforming them into food, thanks to the heat. The processing (pepsis, coction) of matter into the food represents an extraordinary physiological intuition because individual organs of a plant appear to perform its specific transformation. Despite that Theophrastus did not do scientific experiments or use special methods other than the sharpness of his observations, he can be considered the forerunner of a plant physiology that would take rebirth only after two millennia.

Humus: latent phase and reality.

The evolution of the concept of humus was in large part dependent on the development of the modern agriculture. This concept derived from the very old practice of manuring and, after a long period of empirical application, was first theorised by Albrecht Thaer, who recognised the organic and inorganic content as the nutritional elements of humus. The role of humus as nutrient was challenged by Carl Sprengel and Justus Liebig, who opposed successfully the mineral theory to the theory of humus. The research in the property of humus, however, continued thanks to agronomists and agricultural chemists during a period in which the mineral theory was consolidated by the first experiments of the hydroponic cultures performed by Julius Sachs and Wilhelm Knop, but several outstanding chemists and microbiologists continued the work initiated by Thaer, giving humus full physicochemical identity and function. In this context shone the figure of Selman Abraham Waksman, whose work was of fundamental importance to the subsequent research in humus, which is still in progress.

Metals essentials for plants: the nickel case.

The long period of research that preceded the discovery of nickel (Ni) essentiality for plants constitutes a paradigmatic case of doubts and uncertainties that often occur in experimental biology. The history of the essentiality of chemical elements that are present as traces in the plant ash (micronutrients) began in the mid-Nineteenth, but it had blurred outlines until Daniel Arnon, towards the mid-twentieth century, fixed the now historic 'criteria of essentiality'. During this rather long time, seven micronutrients were recognised, step by step, as essential for higher plants, (iron, manganese, boron, Zinc, copper, molybdenum, and chloride), at first thanks to meticulous observations of deficiency symptoms and then to the culture of plant on aqueous solutions. The last element to be recognised as essential for plant nutrition was Ni, which was considered a very toxic element for more than a century. Towards the Thirties, Ni became to be regarded as a useful element by some researchers, but the ultimate proof of its essentiality was obtained only in the Eighties, when the American group of Ross M. Welch demonstrated that Ni is a cofactor of the enzyme urease. More recent research shows that Ni improves the nitrogen (N) metabolism and appears to be important for the efficiency of N fixation.

The dawn of photosynthesis.

Photosynthesis may be hold the most important process of plant nutrition, whose essential principles, viz. water, earth, and air, were stated by E. Mariotte and S. Hales between the second half of the seventeenth and the first half of the eighteenth centuries. Subsequently, the pneumatic chemistry demonstrated that the atmospheric air was composed of different kinds of gases. In this context, J. Priestley discovered, in 1772, that the unbreathable air containing high amount of "fixed air" (carbon dioxide) could be made breathable by plants. This English chemist perhaps sensed the importance of this discovery as for the physiology of plant, whilst such importance was clearly perceived by the Dutch physician Jan Ingenhouse. He collected, between 1779 and 1796, a series of experimental results into a reliable hypothesis whose protagonists were air, water and light. Ingenhousz's work was substantiated by the results of the Swiss physiologists Jean Senebier and, in particular, by those of Théodore de Saussure. This latter, in 1804, transformed the hypothesis into a true theory that defined the elaboration of carbon dioxide as nutritional process, and the release of oxygen as a by-product. This theory constituted the ground of photosynthesis for the two successive centuries, distinguished by exciting and splendid research which transformed photosynthesis research into a classic work of scientific genius.

Viruses: are they living entities?

The essence (living or nonliving entities) of viruses has today become an aporia, i.e. a difficulty inherent in reasoning because they shared four fundamental characteristics with livings (multiplication, genetic information, mutation and evolution) without having the capacity to have an independent life. For much time, however, they were considered minuscule pathogenetic micro-organisms in observance of Koch and Pasteur's 'germ theory' albeit no microbiologist could show their existence except their filterability and pathogenetic action. Only some voices based on experimental results raised against this dogmatic view, in particular those of Beijerinck, Baur and Mrowka, without dipping effectively into the dominant theory. The discovery relative to their nucleoprotein nature made between 1934 and 1936 (Schlesinger as for the phage, and Bawden and co-operators as for Tobacco mosaic virus; TMV), together with the first demonstrations of their structures thanks to electron microscopy (from 1939 onwards) started on casting a new light on their true identity, which could be more clearly identified when, from 1955 onwards, phage and TMV proved to be decisive factors to understand the strategies of replication of the genetic material. Following the new knowledge, the theoretical view relative to viruses changed rather radically and the current view looks on these pathogenetic agents as nonliving aggregates of macromolecules provided with biological properties. There is, however, a current of thought, made explicitly by Lwoff that places viruses as compromise between living and non living and, perhaps, as primitive forms of life which have had great importance for the evolution of cellular life. At any rate, viruses are peculiar entities whose importance cannot be unacknowledged.

Recovery. An enigmatic and neglected form of plant resistance to viruses.

This short history relates the main events of a phenomenon called "recovery", characterised by the disappearance of symptoms from leaves of plant affected with an initially severe virus disease and by immunity to reinoculation with the same virus. It is a subject first disclosed in 1916 from a rather strange disease of tomato but that was confirmed for a certainty between the 1920s and 1930s in most suitable virus-host combinations. Several authoritative virologists gave different interpretations of this phenomenon so that, for at least two decades, there was on this subject some confusion. The work of Conway Price and Carlyle Bennett directed the problems towards a right understanding of the phenomenon, that had its final legitimation in the 1960s. The mechanism of recovery was long matter of hypotheses, which, however, did not gave solving responses as for neither the disappearance of symptoms nor the presence of virus in recovered tissues. Only in the 1990s, owing to the discovery of the "gene silencing system" some molecular virologists proved that recovery is a consequence of a mechanism operating at the transcript level due to RNA silencing. Today, recovery appears to be an occasional phenomenon shown by a little number of virus-host combinations, but it is not possible to rule out that in the ancient past it was a rather diffuse phenomenon allowing virus to colonise plant without damage.

Some biological reflections on the concept of life.

Life is the natural phenomenon that has always aroused the largest interest of philosophers, theologians and scientists, on which a new science--biology--was founded two century ago just for throwing light on its mechanisms. As the pre-Hellenic culture was not able to separate distinctly philosophy from science, life was interpreted as a spurious flurry of the activity of Nature, in which religion, magic and science were interlaced in an intricate way. The Hippocratic medicine constituted the first attempt to focus attention on life by collecting some biological knowledge in order to maintain man's health. All the subsequent physiologists (from the Hellenic to the Latin period) benefited from the precepts of the Corpus Hippocraticum as long as the Christian religion imposed its theological rules that favoured the question relative to soul ever more closely interlaced with the physiology of body. The concept of life became therefore subjected to a number of opposite theories with strong prevalence of metaphysical conjectures until the 19th century but, in spite of this imposition, splendid successes were achieved by physiologists and naturalists such as Harvey, Descartes, Haller, Malpighi, Spallanzani, Wolff, and others, who laid the foundation of a biology that has Lamarck as promoter. The importance of Lamarck's biology came from the release from metaphysics with the introduction of physical and structural concepts which permeated the experimental biology to come. Three main events characterised the biology of the 19th century: i) the interplay of the new chemistry with biology, ii) the cell theory, iii) the concept of metabolism. These events led biology to the 20th century, the era of biochemistry and molecular genetics. The discoveries relative to metabolism characterised the first half of this century, while the second half was witness to the internal mechanisms regulating the life of cells, perhaps the most advanced success of the biology of all time. Today, the cell is not only the centre of a metabolism that, on the whole, may be regarded as the material basis of life, but it is also the central control of communication in response to a great number of stimuli. May life be considered on the basis of biochemical and molecular genetical bases? The answer to this question depends on the credo of everybody. In any case, these mechanisms may explain "what" life is, whereas "why" life exists is still matter of philosophy, and then a fully open question.

A history of virus diseases of plants: from the beginning to 1950.

The history of plant virology has given much space to viruses, especially to Tobacco mosaic virus, but very little space to virus diseases. Still, viruses were clearly characterised only more than fifty years after the first observations and descriptions of diseases appearing infected with ineffable agents and, until the 1950s, most of the plant virologists spent a lot of time to study the disease as the preliminary but absolutely necessity in order to identify the virus. The first virus diseases to be investigated were the "tobacco mosaic" in Europe and "rice stunt" in Far East Asia, and both represented useful models for performing a great number of similar researches. The study of virus diseases made necessary the employment of several strategies, and the introduction of new techniques of research. The simple observation of external symptoms, not too selective and requiring broad experience, was followed by histological and cytological analyses which, in the period herein considered, were carried out by light microscopic methodologies. These analyses helped the research of the physiological causes of symptom formation, which, unfortunately, did not always profit from the interest of plant physiologists and biochemists. This schematic series of efforts was not always followed, since research often proceeded in an erratic way, according to the interest or the possibility of single virologists. However, the comprehensive view emerging from the historical analysis of results (for example, from the first textbook of this discipline) allows us to outline that logical sequence of events we have mentioned above. Obviously, the diffusion of viruses in field was one of most investigated line of research, as well as the individuation of the losses produced by virus diseases. From these fields of research (epidemiology and control), it was possible to enter the war to the most pathogenetic viruses by obtaining the first positive successes: this war became more and more pressing and is still current by the use of a very high technology. The work performed during seventy years at first by beginner virologists and afterwards by mature virologists, amounts to a splendid page of history of virology: This page has been written by hundreds and hundreds researchers, most of them quite neglected by the new generation of plant virologists. This history also represents a grateful homage to those researchers.

Alexandr Oparin and the origin of life on Earth.

In the long essay here examined, the Soviet biochemist A. Oparin elaborated and proposed in a coherent and exhaustive way the three main historical phases characterising the origin of life on Earth from an entirely inorganic environment: i) formation of simple organic molecules; ii) appearance of macromolecules interacting with an aqueous substrate so as to form primitively organised microstructures; iii) constitution of metabolically active protocells working as thermodynamically open system. On the whole, Oparin described a biochemical adventure marked by chemico-physics and natural selection, this latter working on microstructure. Oparin worked according to the canons of dialectic materialism applied to Nature by Friedrich Engels and did not give anything to divine and fantastic. He treated all questions biochemically and neglected Morgan's genetics: mitosis, chromosomes and genes were for Oparin conceptual, and then questionable, elements that he set up against the firm bases of biochemico-physics. In such context, proteins and enzymes were the dominant substances of life owing to their manifold activities. Oparin's book was translated into about fifty languages and was a decisive factor for the successive development of detailed experimental research. Even though dated, this book still shows interesting topics on which the specialists are working at the best of the current technology.

Homeostasis: a history of biology.

Homeostasis is a function indispensable for Life that is accomplished through chemical and physical processes. The mechanisms of this function have been disclosed beginning from the 1920s, when Walther Cannon formulated clear rules that transformed an old concept into paradigm. The concept of harmonic equilibrium among bodily humors dates back to the pre-Hellenic thinkers and was converted into medical suggestions by the Hippocratic school. This concept was handed down to posterity by the several schools of Medicine before being resumed by the great naturalist Jean-Baptiste Lamarck in purely physical terms. The experimental Physiology of the second half of the nineteenth-century elaborated a new conception, the so-called milieu intérieur theorised by Claude Bernard. Toward the half of the 20th century, biochemists imposed the functional basis of the concept, whose mechanisms were definitely established as "feedback" and "allostery".

Plant senescence. A complex lesson of biology of development.

Plant senescence has represented a mystery of biology since 1961, when Carl Leopold sensed the main factors that might control it. The discovery of several hormonal classes gave a precious aid to understand the first mechanisms regulating the phenomenon, which was, in particular, studied in leaf. After the discovery of the molecular bases of genetics, several specialists attempted to connect the hormone activity with the molecular genetic work but only during the 1990s, after the discovery of specific senescence-associate genes, the role of promoting-senescence hormones and retarding-senescence hormones was in good part elucidated, with the help of the plant species Arabidopsis thaliana and its mutants. The history of leaf senescence has had its paradigmatic model in the chlorophyll degradation, a process that remained itself a mystery until 1991, when the products of degradation began being isolated and chemically identified. The research into plant senescence is still the objective of specialists, and represents a very complex problem the integration of which in a unique system appears to be yet far.

Praise of biochemistry.

The interplay of chemistry and biology that occurred after Lavoisier's revolution gave the possibility of analysing the first organic molecules present in animals and plants. The will of learning of the first biologists enabled them to gain the biological meaning of many organic molecules and, towards the mid-19th century, to introduce the concept of "metabolism" for explaining the transformation of food into heat and products for both animal and plant tissues. This concept changed the course of both the experimental biology and medicine and made official, almost in an obvious way, the success of biochemistry as the most appropriate discipline for studying cell metabolism. After the discovery of proteins and nucleic acids as the macromolecules of the genetic code, the words "molecular biology" replaced the word biochemistry but this replacement was nominalistic, because the molecular mechanisms involved in growth and development of the organisms are fundamentally of biochemical nature. In this context, life represents a biochemical phenomenon endowed with a specific internal coherence, resting on two fundamental processes: i) a program, expressed by the genetic code, and ii) a modulation of the complex cell metabolism, with the production of chemical work, heat and homeostasis. This conception of life excludes any external planning design, as well as any deterministic and teleological interpretation, and may be considered of validity for the whole evolutionist span, from the formation of the first protocells to the appearance of man.

Photosynthesis: the years of light.

This history relates the research into the light events of photosynthesis performed from the end of the 1930s to the present day. It is an important page of the contemporaneous biochemistry since it deals with the discovery and elucidation of the mechanisms converting light energy into chemical energy available to chloroplast metabolism and, at the same time, releasing oxygen from water. The explanation of these mechanisms has been due to the integrated work of several hundreds of biochemists and biophysicists directed towards specific key points which were perceived by several brilliant scientists. Jan Amesz, Daniel Arnon, Mordhay Avron, Fay Bendall, Lou Duysens, Robert Emerson, Robert Hill, Andre Jagendorf, Bessel Kok, Cornelius van Niel, Sam Ruben, Achim Trebst were among the protagonists that laid the foundation of the biochemical and biophysical processes relative to the energy conversion and water photolysis. In the last twenty years, the precious inheritance left by these scientists has enabled to better understand the structure and function of the apparatus controlling the light events of photosynthesis, Photosystems I and II, an exception singularity in the already complex world of biochemistry.

Plant mitochondria. Fifty years of plant biology.

This history has its conventional opening about twenty years after the statement of the "Cell Theory", when, at the end of the 19th century, several cytologists discovered unusual, minute corpuscles in the protoplasm of both animal and plant cells. These organelles, owing to their polymorphism, were defined "mitochondria" and a pressing debate on their function and origin went on for about half a century. The invention of the electron microscope and the development of a technique of differential fractionation permitting the separation of the cell constituents, made it possible to connect the structure and function of mitochondria and to attribute to them the role of the site of the aerobic respiration. Research on plant mitochondria got a great step forward in 1951, when the American group of James Bonner demonstrated that mitochondria of mung bean accomplished the whole TCA cycle coupled with the oxidative phosphorylation. One of the fundamental phases of that immense cycle of reactions represented by the metabolism of plant cell had thus its completion.

Genetics and virology: two interdisciplinary branches of biology.

Genetics has a tradition that dates back to the Ancient Greeks. It developed, between insight and contradiction, from the post-Renaissance to the mid-1800s, when Mendel and Darwin gave it the first experimental and conceptual bases. From 1910, genetics became a true experimental discipline of Biology thanks to the work of Morgan's group. On the contrary, virology is a relatively young discipline which had origin only after the success of the "germ theory" of Pasteur and Koch, by the hypothesis of the contagium vivum fluidum of Beijerinck, in 1898. In spite of their historical difference, the modern development of the two disciplines had a close connection. In 1922, the geneticist Muller first compared the bacteriophage to the gene and, in 1923, the (phyto)physiologists Benjamin Duggar and Joanne Karrer Armstrong suggested the analogy between gene and Tobacco mosaic virus (TMV). Knowledge on the biochemical nature of gene and virus developed in the early 1940s when the biochemists began to suspect that the nucleic acids might be the genetical determinants for both the bionts. Avery and co-workers discovered in 1944 that DNA was the principle of the transmission of hereditary characters in bacteria and, in 1948, a little group of English (phyto)virologists (Markham, Matthews and Smith) discovered that the RNA of a plant virus (Turnip yellow mosaic virus) was directly involved in virus replication. The fundamental significance of the two discoveries was not gathered by geneticists and virologists, even because the respective groups did not gave the necessary emphasis to their results. Thus, the discovery of the role of the nucleic acids in virus replication is historically attributed to Hershey and Chase for DNA phage, and to Fraenkel-Conrat and the German virologists Gierer and Schramm for plant viruses.

Photosynthesis: 1900-1930.

During the second half of the 19th century Julius von Sachs established the main principles of the photosynthetic production of sugars. From then, a growing number of biochemists and physiologists attended to the process, that appeared like a "black box", in order to detect what came in and what went out of it. The English group of Frederick Blackman gave a remarkable contribution in individuating the close connection between temperature, light and CO2 concentration. Later, the great importance of light was stressed by Otto Warburg, who evaluated the radiant energy necessary to the process in terms of quantum theory. The biochemical mechanism of photosynthesis was interpreted by the main European schools on the basis of Adolf Baeyer's suggestion which posed formaldehyde as the core of the process. Formaldehyde's theory hold engaged the biochemists for about fifty years although some voices rose up against it. However, nobody could put forward more coherent theories until the 1940s, when Sam Ruben and Martin Kamen individuated the cyclic pattern of the process. Ultimately, the first thirty years of the 20th century must be seen as a preliminary stage studded with light and shade even if, in spite of controversial trends, several findings of remarkable interest became to disclose that "black box" as we know today chlorophyll photosynthesis.

The origin of phage virology.

The history of bacteriophage (phage) had its start in 1915, when Twort isolated an unusual filterable and infectious agent from excrete of patients struck by diarrhoea; this discovery was followed by an analogous, and probably independent, finding of d'Hérelle in 1917. For several years phage research made scant progress but great attention was paid to the question of phage nature, which saw the contrast between d'Hérelle and Bordet's views (living against chemical nature, respectively). This situation changed with the independent discovery of lysogeny, in 1925, thanks to Bordet and Bail: this phenomenon was considered of genetical origin, a view that Wollman interpreted by assimilating the properties of phage to those of gene (according to a previous idea of Muller). In the 1930s, Burnet's work opened a new era by demonstrating the occurrence of several species of phages and their antigenic property. In the same period, the physical and chemical characteristics of these viruses were disclosed thanks, in particular, to the work of Schlesinger, who first demonstrated that a virus (phage) was constituted of nucleoproteins. The peculiarity of phage was finally shown after the invention of electron microscope: H. Ruska, in 1940, and Anderson and Luria in the next years, obtained the first images of tailed phages, a finding that strongly helped the investigation on the first steps of the infection process. The decisive impulse to phage virology came from Delbrück, a physicist who entered biology giving it a new arrangement. The so-called "phage group" assembled brilliant minds (Luria, Hershey and Delbrück himself, and later a dozen of other scientists): this group faced three fundamental questions of phage virology, i.e., the mechanisms of attack, multiplication and lysis. In ten years' time, phage virology became an integrant part of molecular biology, also thanks to the discovery of the genetical properties of DNA: in such scientific context, Delbrück, Luria and Hershey's works emerged for the absolute excellence of their results, which led such scientists to Nobel prize. Lysogeny was however neglected by the phage group: this singular property shared by bacteria and phages was instead investigated by Lwoff's group, in Paris, and explained in its fundamental features during the 1950s. The "phage's saga" has gone on being an important division of molecular biology till today, and its history is far from being over.