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Chapter: Essential Microbiology: Microbiology: What, Why and How?

How do we know? Microbiology in perspective: to the ‘golden age’ and beyond

We have learnt an astonishing amount about the invisible world of microorganisms, particularly over the last century and a half.

How do we know? Microbiology in perspective: to the ‘golden age’ and beyond

We have learnt an astonishing amount about the invisible world of microorganisms, particularly over the last century and a half. How has this happened? The penetrating insights of brilliant individuals are rightly celebrated, but a great many ‘breakthroughs’ or ‘discoveries’ have only been made possible thanks to some (frequently unsung) development in microbiological methodology. For example, on the basis that ‘seeing is believing’, it was only when we had the means to see microorganisms under a micro-scope that we could prove their existence.

Microorganisms had been on the Earth for some 4000 million years, when Antoni van Leeuwenhoek started out on his pioneering microscope work in 1673. Leeuwen-hoek was an amateur scientist who spent much of his spare time grinding glass lenses

to produce simple microscopes (Figure 1.1). His detailed drawings make it clear that the ‘animalcules’ he observed from a variety of sources included representatives of what later became known as protozoa, bacteria and fungi. Where did these creatures come from? Arguments about the origin of living things revolved around the long held belief in spontaneous generation, the idea that living organisms could arise from non-living matter. In an elegant experiment, the Italian Francesco Redi (1626–1697) showed that the larvae found on putrefying meat arose from eggs deposited by flies, and not spontaneously as a result of the decay process. This can be seen as the beginning of the end for the spontaneous generation theory, but many still clung to the idea, claiming that while it may not have been true for larger organisms, it must surely be so for minute creatures such as those demonstrated by Leeuwenhoek. Despite mounting evidence against the theory, as late as 1859, fresh ‘proof’ was still being brought forward in its support. Enter onto the scene Louis Pasteur (1822–1895), still arguably the most famous figure in the history of microbiology. Pasteur trained as a chemist, and made a lasting contribution to the science of stereochemistry before turning his attention to spoilage problems in the wine industry. He noticed that when lactic acid was produced in wine instead of alcohol, rod-shaped bacteria were always present, as well as the expected yeast cells. This led him to believe that while theyeast produced the alcohol, the bacteria were responsible for the spoilage, and that both types of organism had originated in the en-vironment. Exasperated by continued efforts to substantiate the theory of spontaneous generation, he set out to disprove it once and for all. In response to a call from the French Academy of Science, he carried out a series of experiments that led to the ac-ceptance of biogenesis, the idea that life arises only from already existing life. Using his famous swan-necked flasks (Figure 1.2), he demonstrated in 1861 that as long as dust

particles (and the microorganisms carried on them) were excluded, the contents would remain sterile. This also disproved the idea held by many that there was some element in the air itself that was capable of initiating microbial growth. In Pasteur’s words ‘. . . . the doctrine of spontaneous generation will never recover from this mortal blow. Thereis no known circumstance in which it can be affirmed that microscopic beings came into the world without germs, without parents similar to themselves.’ Pasteur’s findingson wine contamination led inevitably to the idea that microorganisms may be also be responsible for diseases in humans, animals and plants.

The notion that some invisible (and therefore, presumably, extremely small) living creatures were responsible for certain diseases was not a new one. Long before micro-organisms had been shown to exist, the Roman philosopher Lucretius (98–55 BC) and much later the physician Girolamo Fracastoro (1478–1553) had supported the idea. Fracastoro wrote ‘Contagion is an infection that passes from one thing to another’ and recognised three forms of transmission: by direct contact, through inanimate objects and via the air. We still class transmissibility of infectious disease in much the same way today. The prevailing belief at the time, however, was that an infectious disease was due to something called a miasma, a poisonous vapour arising from dead or diseased bodies, or to an imbalance between the four humours of the body (blood, phlegm, yellow bile and black bile). During the 19th century, many diseases were shown, one by one, to be caused by microorganisms. In 1835, Agostino Bassi showed that a disease of silkworms was due to a fungal infection, and 10 years later, Miles Berkeley demonstrated that a fun-gus was also responsible for the great Irish potato blight. Joseph Lister’s pioneering work on antiseptic surgery provided strong, albeit indirect, evidence of the involvement of mi-croorganisms in infections of humans. The use of heat-treated instruments and of phenol both on dressings and actually sprayed in a mist over the surgical area, was found greatly to reduce the number of fatalities following surgery. Around the same time, in the 1860s, the indefatigable Pasteur had shown that a parasitic protozoan was the cause of another disease of silkworms called pebrine´, which had devastated the French silk industry.

The first proof of the involvement of bacteria in disease and the definitive proof of the germ theory of disease came from the German Robert Koch. In 1876 Koch showedthe relationship between the cattle disease anthrax and a bacillus which we now know as Bacillus anthracis. Koch infected healthy mice with blood from diseased cattle and sheep, and noted that the symptoms of the disease appeared in the mice, and that rod shaped bacteria couldbe isolated from their blood. These could be grown in culture, where they multiplied and produced spores. Injection of healthy mice with these spores (or more bacilli) led them too to develop anthrax and once again the bacteria were isolated from their blood. These results led Koch to formalise the criteria necessary to prove a causal relationship between a specific disease condition and a particular microorganism. These criteria became known as Koch’s postulates, and are still in use today.

Despite their value, it is now realised that Koch’s pos-tulates do have certain limitations. It is known for ex-ample that certain agents responsible for causing disease (e.g. viruses, prions:) can’t be grown invitro, but only in host cells. Also, the healthy animalin Postulate 3 is seldom human, so a degree of extrapo-lation is necessary – if agent X does not cause disease in

a laboratory animal, can we be sure it won’t in humans? Furthermore, some diseases are caused by more than one organism, and some organisms are responsible for more than one disease. On the other hand, the value of Koch’s postulates goes beyond just defining the causative agent of a particular disease, and allows us to ascribe a specific effect (of whatever kind) to a given microorganism.

Critical to the development of Koch’s postulates was the advance in culturing techniques, enabling the isola-tion and pure culture of specific microorganisms. The development of pure cultures revolutionised microbiology, and within the next 30 years or so, the pathogens responsible for the majority of common human bacterial diseases had been isolated and identified. Not without just cause is this period known as the ‘golden age’ of microbiology! Table 1.summarises the discovery of some major human pathogens.

Koch’s greatest achievement was in using the ad-vances in methodology and the principles of his own postulates to demonstrate the identity of the causative agent of tuberculosis, which at the time was responsible for around one in every seven human deaths in Europe.

Although it was believed by many to have a microbial cause, the causative agent had never been observed, either in culture or in the affected tissues. We now know that Mycobacterium tuberculosis (the tubercle bacillus) is very difficult to stain by conven-tional methods due to the high lipid content of the cell wall surface. Koch developed a staining technique that enabled it to be seen, but realised that in order to satisfy his own postulates, he must isolate the organism and grow it in culture. Again, there were technical difficulties, since even under favourable conditions, M. tuberculosis grows slowly, but eventually Koch was able to demonstrate the infectivity of the cultured organisms towards guinea pigs. He was then able to isolate them again from the dis-eased animal and use them to cause disease in uninfected animals, thus satisfying the remainder of his postulates.

Although most bacterial diseases of humans and their aetiological agents have now been identified, important variants continue to evolve and emerge. Notable exam-ples in recent times include Legionnaires’ disease, an acute respiratory infection caused by the previously unrecognised genus, Legionella, and Lyme disease, a tickborne infection first described in Connecticut, USA in the mid-1970s. Also, a newly recognised pathogen, Helicobacter pylori, has been shown to play an important (and previously unsuspected) role in the development of peptic ulcers. There still remain a few diseases that some investigators suspect are caused by bacteria, but for which no pathogen has been identified.

Following the discovery of viruses during the last decade of the 19th century, it was soon established that many diseases of plants, animals and humans were caused by these minute, non-cellular agents.

The major achievement of the first half of the 20th century was the development of antibiotics and other antimicrobial agents. Infectious diseases that previously accounted for millions of deaths became treatable by a simple course of therapy, at least in the affluent West, where such medications werereadily available.

If the decades either side of 1900 have become known as the golden age of microbiology, the second half of the twentieth century will surely be remembered as the golden age of molecular genetics. Following on from the achievements of others such as Griffith and Avery, the publication of Watson and Crick’s structure for DNA in 1953 heralded an extraordinary 50 years of achievement in this area, culminating at the turn of the 21st century in the completion of the Human Genome Project.

What, you might ask, has this genetic revolution to do with microbiology? Well, all the early work in molec-ular genetics was carried out on bacteria and viruses and microbial systems have also been absolutely central to the development of genetic engineering over the last three decades. Also, as part of the Human Genome Project, the genomes of several microorganisms have been decoded, and it will become increasingly easy to do the same for others in the future, thanks to methodological advances made during the project. Having this information will help us to understand in greater detail the disease strategies of microorganisms, and to devise ways of countering them.

As we have seen, a recurring theme in the history of microbiology has been the way that advances in knowledge have followed on from methodological or technological developments, and we shall refer to a number of such developments during the course of this book. To conclude this introduction to microbiology, we shall return to the instrument that, in some respects, started it all. In any microbiology course, you are sure to spend some time looking down a microscope, and to get the most out of the instrument it is essential that you understand the principles of how it works. The following pages attempt to explain these principles.

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