Current and past strategies for bacterial culture in clinical microbiology.

A pure bacterial culture remains essential for the study of its virulence, its antibiotic susceptibility, and its genome sequence in order to facilitate the understanding and treatment of caused diseases. The first culture conditions empirically varied incubation time, nutrients, atmosphere, and temperature; culture was then gradually abandoned in favor of molecular methods. The rebirth of culture in clinical microbiology was prompted by microbiologists specializing in intracellular bacteria. The shell vial procedure allowed the culture of new species of Rickettsia. The design of axenic media for growing fastidious bacteria such as Tropheryma whipplei and Coxiella burnetii and the ability of amoebal coculture to discover new bacteria constituted major advances. Strong efforts associating optimized culture media, detection methods, and a microaerophilic atmosphere allowed a dramatic decrease of the time of Mycobacterium tuberculosis culture. The use of a new versatile medium allowed an extension of the repertoire of archaea. Finally, to optimize the culture of anaerobes in routine bacteriology laboratories, the addition of antioxidants in culture media under an aerobic atmosphere allowed the growth of strictly anaerobic species. Nevertheless, among usual bacterial pathogens, the development of axenic media for the culture of Treponema pallidum or Mycobacterium leprae remains an important challenge that the patience and innovations of cultivators will enable them to overcome.

Koch's postulates, carnivorous cows, and tuberculosis today.

With Koch's announcement in 1882 of his work with the tubercle bacillus, his famous postulates launched the rational world of infectious disease and an abrupt social change--strict patient isolation. The postulates, so successful at their inception, soon began to show some problems, particularly with cholera, which clearly violated some of Koch's requirements. Subsequent studies of other diseases and the discovery of entirely new ones have so altered and expanded the original postulates that they now are little but a precious touch of history. The present additions and replacements of the original concepts are skillful changes that several authors have devised to introduce new order into understanding complex viral and prion diseases. In 1988, this knowledge, with the totally rational response of the British population and its cattle industry, was critical in promptly blocking the threatened epidemic of human prion disease. In contrast, the recent upsurge of tuberculosis (TB) in the worldwide AIDS epidemic in developing countries, and the sudden increase in metabolic syndrome in wealthy ones, suggests the need for focused sociobiologic research seeking ways to affect the damaging lifestyle behavior of many less educated populations in both settings. The world awaits an equivalent of Koch's Postulates in sociobiology to explain and possibly avert large self-destructive behaviors.

[A letter of Dr. Matuki Miyazaki to Prof. Masao Ota in 1938].

A letter of Matuki Miyazaki, the director of Kyushu Sanatorium to Masao Ota was found in the materials of Masao Ota stored at the library of University of Tokyo. Miyazaki, who was president of 12th annual meeting of Japanese Leprosy Association in Kumamoto in 1938, thanked Ota for the acceptance of a special lecture. Ota lectured on the title of inoculation of leprosy bacteria to animals.

Tuberculosis, Cholera, Anthrax: Dreadful Culprits.

Dreadful culprits from the minuscule world, indeed, but … what about poverty, war, and terrorism in the macroscopic nowadays world?

Clinical microbiology during the Vietnam War.

During the period of 1965-1968, over two dozen Army microbiologists were deployed to various locations in Southeast Asia in support of the Vietnam War. Their role was to serve both a clinical laboratory mission/function at the mobile Army surgical hospital and mobile laboratory level as well as to perform research roles in all of the facilities. They were essential to the formulation of medical intelligence as well as to the practice of operational medicine in the deployed environment. The results of their laboratory investigations provided commanders and military physicians with critical medical information for patient care, outbreak investigation, and forensic analysis. As with many soldiers in support of the infantry and armor combat forces, most of the work occurs behind the scenes and their contributions are often left out of the historical literature. This article presents a brief overview of microbiology performed by Army microbiologists during the Vietnam War.

[A matter of methods: the historicity of Koch's postulates 1840-2000]. / Alles eine Frage der Methode: Zur Historizität der Kochschen Postulate 1840-2000.

This paper analyses the historical origins and the popularity of 'Koch's Postulates'. In 1884 Friedrich Löffler wrote down the well-known three steps of isolation, cultivation and inoculation as conditions for establishing the existence of a pathogen. These postulates are frequently invoked in textbooks of medical history. Yet they seem to have had little relevance in medical research. Their assumed inventor, Robert Koch, produced numerous variations in his own methodology. However, underlying his work was a sort of trivial ontology of diseases which rendered an experimental reconstruction of human pathologies in animal models practical and meaningful. There were many ways to pursue this end. Koch usually limited his discussion to practical questions related to the course that investigations had to take, while matters of principle were only treated implicitly in his writings. Löffler's achievement was to popularise Koch's views in his postulates. Given that, it is not surprising that the countless references to Koch's postulates which one finds in the 20th century usually refer to the spirit rather than the literal meaning of the postulates. For example, proponents of virology or molecular medicine devise variations of Koch's postulates that serve to relate their own work to Koch's bacteriology. The latter is perceived as the origin of modern experimental medicine. The nature of such references is less historical than anecdotal: referring to a historical object that did not exist as such, these references produce ex traditione credentials for experimental medicine.

[Isolation and identification methods of enterobacteria group and its technological advancement].

In the last half-century, isolation and identification methods of enterobacteria groups have markedly improved by technological advancement. Clinical microbiology tests have changed overtime from tube methods to commercial identification kits and automated identification. Tube methods are the original method for the identification of enterobacteria groups, that is, a basically essential method to recognize bacterial fermentation and biochemical principles. In this paper, traditional tube tests are discussed, such as the utilization of carbohydrates, indole, methyl red, and citrate and urease tests. Commercial identification kits and automated instruments by computer based analysis as current methods are also discussed, and those methods provide rapidity and accuracy. Nonculture techniques of nucleic acid typing methods using PCR analysis, and immunochemical methods using monoclonal antibodies can be further developed.

Acid-Fast Positive and Acid-Fast Negative Mycobacterium tuberculosis: The Koch Paradox.

Acid-fast (AF) staining, also known as Ziehl-Neelsen stain microscopic detection, developed over a century ago, is even today the most widely used diagnostic method for tuberculosis. Herein we present a short historical review of the evolution of AF staining methods and discuss Koch's paradox, in which non-AF tubercle bacilli can be detected in tuberculosis patients or in experimentally infected animals. The conversion of Mycobacterium tuberculosis from an actively growing, AF-positive form to a nonreplicating, AF-negative form during the course of infection is now well documented. The mechanisms of loss of acid-fastness are not fully understood but involve important metabolic processes, such as the accumulation of triacylglycerol-containing intracellular inclusions and changes in the composition and spatial architecture of the cell wall. Although the precise component(s) responsible for the AF staining method remains largely unknown, analysis of a series of genetically defined M. tuberculosis mutants, which are attenuated in mice, pointed to the primary role of mycolic acids and other cell wall-associated (glyco)lipids as molecular markers responsible for the AF property of mycobacteria. Further studies are now required to better describe the cell wall reorganization that occurs during dormancy and to develop new staining procedures that are not affected by such cell wall alterations and that are capable of detecting AF-negative cells.

[Methods of diagnosis and differentiation of plague pathogen: approaches to detection of atypical strains of Yersinia pestis by molecular biology. Part I].

A review of data on the modern methods of detection of typical and atypical strains of the plague agent Y. pestis is given. The history of the development of the molecular-biological tests for the differentiation of Y. pestis from the related microorganisms is presented. The problems facing investigators during the development of these tests are discussed.

[Dyes, indispensable tools for the 19th century's biologic and therapeutic revolution]. / Les colorants, outils indispensables de la révolution biologique et thérapeutique du XIXe siècle.

Born from growing organic chemistry laboratories, dyes were extensively used par textile industry before to be applied in field of biology and therapeutics. Besides their interest for diagnostic techniques due to cell visualization (Virchow, Papanicolaou), dyes allowed scientists to propose scientific hypothesis founding, in conjunction with new microscopy tools, modern basis for biology : tissue constitution, cellular and sub cellular structure, s.o. One of the brightest illustrations of these progresses is the birth of neuronal theory which due to silver print of brain tissue allowed to see intimacy of cerebral structures et propose an operating scheme (Golgi, Cajal). Therapeutic progresses born from dyes chemistry are multiple. First concentrated on the research of antimalarial drugs (Ehrlich) following the use of methylene blue, then generally, anti-infectious drugs, they gave birth to various chimiotherapeutic families: antiseptics, antiparasitic drugs, antibacterial, among which one of the most spectacular illustrations remain sulphonamides preparation.

Flow cytometry of bacteria: glimpses from the past with a view to the future.

Measurement of bacteria and other microorganisms at the level of single cells has progressed enormously over the last couple of decades. Up to the late 1970s, there were no other means than microscopy for observation of single microorganisms, making any type of measurement very cumbersome and tedious, at best. Today, we measure several parameters simultaneously with a precision of a few per cent, and at a rate of 1000 cells per second. The first papers on the use of flow cytometry to measure bacteria appeared only in 1977, although the method had proved highly successful in studies of mammalian cells for almost a decade. There were several reasons for this relatively late introduction, including technical limitations, problems with adequate staining, and, not least, the human factor. Today, flow cytometry has a wide range of microbiological applications, ranging from studies of the bacterial cell cycle and many other cellular characteristics to assessment of antibiotic susceptibility of clinical samples, and monitoring of bacteria and other microorganisms in anything from sewage to sea water. Still, the potential of flow cytometry in microbiology is far from fully utilised. Better instruments and new stains will provide new opportunities to understand, control and exploit this vital part of the biosphere.

From Koch's postulates to biofilm theory. The lesson of Bill Costerton.

The clinical diagnoses of implant infections pose insurmountable difficulties for cultural methods because of their frequent failure when bacteria are growing in biofilms. In 1978 Bill Costerton warned that chronic infections in patients with indwelling medical devices were caused by bacteria growing in well-developed glycocalyx-enclosed biofilms and that bacteria within biofilms resist antibiotic therapies and immune host defenses. Costerton's "biofilm theory" opened two lines of scientific endeavor: the study of the biochemistry and genetics of biofilm formation and function; and, on the other side, the search for new methods for medical diagnosis and treatment of biofilm-centered implant infections. This Editorial and the entire 2012 issue "Focus on Implant Infections" are dedicated to the memory of Bill Costerton, recognized worldwide as the Father of Biofilms for his innovation and body of work on infections caused by sessile bacteria. Bill Costerton was a great scientist, heedful both to the biological aspects of biofilms and to the medical challenges of new diagnostic methods and modern therapeutic approaches to implant infections. But, most of all, he was a charming Maestro for the large number of colleagues and students whose enthusiasm for the science he was able to nourish. Bill passed away on May 12th, 2012 and the entire science community mourns the death of a friend and a leader.