This is a collection of cutting-edge mycoplasma methods for the detection, isolation, identification, characterization, and genetic manipulation of the pathogenic mycoplasmas. These step-by-step methods are crafted for successful reproducibility and include biochemical, genetic, and molecular techniques essential to understanding pathogenecity and adhesion to host cells. They also cover the detection of mycoplasmas in cell cultures, an important tool not only in viral diagnosis and research, but also in the production of vaccines and various biological products.
Over the last decade, interest in mycoplasmas has been greatly sti- lated by the spread of contagious bovine pleuropneumonia (CBPP), caused by Mycoplasma mycoides subsp. mycoides SC, in Europe and Africa and the discovery of a possible association between mycoplasmas and AIDS. During this period there has also been a recognition by control authorities and int- national organizations, including the Office International des Epizooties, that mycoplasmas are major causes of economic loss in livestock and continue to be a problem in humans as a result of a range of chronic respiratory and urogenital diseases. However, it is likely that the true significance of my- plasmas is still underestimated because of the difficulties in working with these extremely fastidious microorganisms. Advances in such laboratory technology as DNA amplification has enabled the detection of unrecoverable mycoplasmas in tissue samples and this will provide us with a better understanding of their role in disease. Important developments have also been seen in more conv- tional areas such as serological tests and media formulation enabling improved recovery and identification. Mycoplasma Protocols provides in a single volume up-to-date and easy-- follow methods for the detection, isolation, identification, and characterization of mycoplasmas, with a major emphasis on those of medical and veterinary significance. It also includes biochemical, genetic, and molecular techniques that will form the basis of understanding pathogenicity and, particularly, - hesion to host cells.
Methods in Mycoplasmology, Volume I: Mycoplasma Characterization is the first of a two-volume series that resulted from cumulative efforts to meet the need for standardized techniques in mycoplasmology. The book is organized into five sections. Section A provides an introduction to the mycoplasmas by discussing their characteristics and classification, mycoplasma-animal host interrelationships, and mycoplasma-plant-insect interrelationships. The chapters in Section B examine the morphological and ultrastructural features of mycoplasmas, using techniques such as phase-contrast microscopy, dark-field microscopy, and transmission electron microscopy. Section C discusses the cultivation and nutrition of mycoplasmas. Section D focuses on analytical methods for the characterization of mycoplasmas, including harvest and washing procedures, cell lysis and membrane isolation, and crossed immunoelectrophoresis. Section E describes tests designed to detect specific enzymatic activities or nutritional requirements of mycoplasmas. These include sugar fermentation and arginine hydrolysis tests as well as tests to establish the sterol requirement of mycoplasmas. Section F deals with serological tests. These tests either have specific application to detection of mycoplasma antigens or antibody in plant and animal tissues; are useful in identification of specific groups of mycoplasmas (spiroplasmas); or provide higher sensitivity in measurements of antibodies against mycoplasmas.
Methods in Mycoplasmology, Volume II: Diagnostic Mycoplasmology is the second of a two-volume series that resulted from cumulative efforts to meet the need for standardized techniques in mycoplasmology. The book is organized into five sections. Section A discusses techniques for isolating mycoplasmas from humans. These include the recovery of mycoplasmas from the respiratory and genitourinary tracts, from blood and special tissues, and serological Identification of mycoplasmas from humans. Section B focuses on techniques for isolating mycoplasmas from animals, including the recovery and identification of avian, bovine, caprine, ovine, canine, and feline mycoplasmas. Section C presents techniques for isolating mycoplasmas from cell cultures. These include agar and broth techniques, the use of indicator cell lines, and DNA fluorochrome staining. Section D discusses techniques for isolating mycoplasmas from plants and insects. Section E describes some of the techniques that are useful in evaluating the pathogenicity of mycoplasmas, ureaplasmas, and spiroplasmas. Section F defines some of the techniques useful in virus detection by electron microscopy and presents methods for isolating and characterizing viruses from both mycoplasmas and spiroplasmas.
Mycoplasmas cause some of the most serious and economically significant diseases in livestock and pose major problems for animal health authorities worldwide. Infection has spread in the last five years to new regions and species, but little effective control is available, particularly in developing countries. This work encapsulates the latest research and development on mycoplasmas in sheep, goats and cattle from laboratories all over the world, describing both conventional diagnostic techniques for growth and identification and newly established procedures such as PCR/DGGE. Molecular typing methods are also covered, specifically for use in mycoplasma fingerprinting as well as up to date reviews on the major mycoplasma diseases including contagious bovine and caprine pleuropneumonias, contagious agalactia and many conditions caused by Mycoplasma bovis.
This compendium is the result of the FEMS Workshop on "Rapid Diagnosis of Mycoplasmas" which I organized and which took place in Jerusalem, Israel, August 11-23, 1991. The first week's sessions were held at a resort on the outskirts of Jerusalem and consisted of lectures and discussions. This part was modelled along the lines of the Gordon Conference in the USA, i.e., in an intimate atmo sphere in which everyone could mix and exchange ideas, and was very benefi cial. About 100 scientists from around the world attended the first week. Dur ing the first week, the biology, molecular biology and pathophysiology of myco plasmas, as well as all the main diagnostic methods were covered, including both conventional and the newer technologies. The session on mycoplasmas in the human urogenital tracts was held in conjunction with the Israel Society for the Study and Prevention of Sexually Transmitted Disease. The second week was a laboratory session and was held at the Hebrew University-Hadassah Medical School campus in Ein Karem, Jerusalem. All ex periments were conducted by eminent specialists in their field. The lab session had 36 participants from 19 countries who used the most modern techniques for the diagnosis of mycoplasmas in medicine, veterinary medicine and agri culture. The efficacy of several commercial kits were also tested at this time. I want to again thank everyone who helped and supported this work shop, as well as the authors of the various chapters.
This latest addition to the Methods in Molecular Medicine series, Anti- ral Methods and Protocols, is opportune because there is an increasing int- est in discovering compounds that are effective against both chronic and acute viral infections. A number of the methods described in the volume are unp- lished and their inclusion indicates the speed at which this field is moving. This volume is not a review but each chapter contains methods validated by the experts who have spent time in developing the protocols. The hallmark of this series is the comprehensive way in which the me- ods are described, which includes a list of all the reagents needed for each protocol. Of importance is the section on tips and pitfalls that the authors have discovered while developing their protocols. The manual itself is designed to be used by researchers in universities and industry who are familiar with a range of biological techniques but who want to set up quickly a novel assay system. We encourage a dialog between readers and authors, which may also result in useful collaborations.
This book and its companion, Volume I, concentrate on new procedures--especially those based on the new molecular methodology--developed within the past decade. This volume deals with the new genetic and immunological tools applied to the diagnosis of mycoplasma infections of humans, animals, plants, insects, and all cultures. Volume I outlines the approaches, techniques, and procedures applied to cell and molecular biology studies of mycoplasmas. Diagnostic genetic probes Immunological tools Antibiotic sensitivity testing Diagnosis of specific diseases Experimental infections Diagnosis of mycoplasma infections of cell cultures
