DNA Microarray Analysis
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17 results
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The "Cryptic" Escherichia
- Author: Seth T. Walk
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Citation: Walk S. 2015. The "Cryptic" Escherichia, EcoSal Plus 2015; doi:10.1128/ecosalplus.ESP-0002-2015
- DOI 10.1128/ecosalplus.ESP-0002-2015
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Abstract:
In 2009, five monophyletic Escherichia clades were described and referred to as “cryptic” based on the inability to distinguish them from representative E. coli isolates using diagnostic biochemical reactions. Since this original publication, a number of studies have explored the genomic, transcriptomic, and phenotypic diversity of cryptic clade isolates to better understand their phylogenetic, physiological, and ecological distinctiveness with respect to previously named Escherichia species. This chapter reviews the original discovery of the cryptic clades, discusses available evidence that some are environmentally adapted, and evaluates current support for taxonomic designations of these microorganisms. The importance of these clades to clinical research, epidemiology, population genetics, and microbial speciation is also discussed.
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Biofilm Formation in Candida albicans
- Authors: Jonathan Sewell Finkel, Aaron P. Mitchell
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Source: Candida and Candidiasis, Second Edition , pp 299-315
Publication Date :
January 2012
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Abstract:
This chapter focuses on biofilm formation by the pathogenic fungus Candida albicans. Biofilms are surface-associated microbial communities surrounded by an extracellular matrix. The chapter examines the steps of biofilm formation, from the genes known to function in C. albicans biofilm development, the cell-cell communication within the biofilm, the environmental responses that contribute to biofilm formation, the drug resistance of biofilms, and experimental techniques used to study biofilms. Cell wall genes and adhesins provide mechanisms that promote biofilm initiation, and the transcription factors that regulate their expression couple biofilm initiation with internal and external signals. The study of transcription factors has laid the framework for gene regulatory networks. While much is now known, detailed testing of other known adherence factors, cell wall proteins, transcription factors, kinases, and others is required to identify more genes involved in biofilm formation, and the elucidation of upstream regulation of these factors will allow for greater insights into the signaling events important for biofilm development. Quorum sensing governs functions as diverse as bioluminescence and virulence. It has a vital role in bacterial biofilm dynamics, and its role in C. albicans biofilms is now beginning to be understood. While major challenges lie ahead to further refine the gene regulatory networks and correlate them to in vivo results, it is nonetheless an exciting time for the field of C. albicans biofilm formation.
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Recent Developments in Rapid Detection Methods
- Authors: Lawrence D. Goodridge, Mansel W. Griffiths
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Source: Pathogens and Toxins in Foods , pp 450-459
Publication Date :
January 2010
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Abstract:
The continued presence of pathogenic microorganisms and their toxins in food and drinking water has necessitated the ongoing need for newer, more sensitive and robust analytical systems capable of rapid detection of these contaminants in complex samples. The ideal detection method should be capable of rapidly detecting and confirming the presence of food-borne pathogenic microorganisms directly from complex samples with no false-positive or false-negative results. Rapid detection methods including immunological detection, cell/tissue-based assays and nucleic acid-based assays have been discussed in this chapter. Conventional culture techniques continue to be the gold standard for the isolation, detection, and identification of target pathogens. These methods increase detection times by hours to days, causing preliminary test results to be delayed. These assays are defined as affinity, cell/ tissue, and nucleic acid technologies. Antibody-based detection systems are still considered to be the gold standard of affinity-based testing methods. Aptamers offer several advantages over the use of antibodies in the identification of food-borne microorganisms and toxins. Any microorganism that contains DNA or RNA can be detected using nucleic acid-based assays, but a limitation of these diagnostics is their inability to detect protein-based agents of disease, such as toxins or prions.
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Arcobacter: an Opportunistic Human Food-Borne Pathogen?
- Authors: Irene V. Wesley, William G. Miller
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Source: Emerging Infections 9 , pp 185-212
Publication Date :
January 2010
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Abstract:
Two recent European surveys of patient stool samples ranked Arcobacter as the fourth most frequently recovered Campylobacter-like microbe. Aerotolerant campylobacteria (Arcobacter spp.) were first described as occurring in aborted porcine and bovine fetuses. Handling and consuming raw or contaminated poultry meat are acknowledged potential sources of human Arcobacter infection. Due to the phylogenetic relationship between Campylobacter and Arcobacter, it is logical to assume that animal models, virulence factors (including adherence), invasion, cytotoxicity, and toxin production of the two organisms would be similar. Classification of arcobacters as free-living, environmental organisms should be reflected in the gene content of the Arcobacter genomes. The genomes of some Arcobacter species would be predicted to contain niche-related genes, e.g., osmoprotectant genes in A. halophilus and nitrogen fixation genes in A. nitrofigilis. An multilocus sequence typing (MLST) method that could be used to type five Arcobacter species was recently designed. Future MLST and DNA microarray analyses will provide further insights into Arcobacter divergence. MLST analysis has identified putative horizontal gene transfer (HGT) events in Arcobacter. Members of the genus Arcobacter can be generalized as free-living organisms found predominantly in aqueous environments, occasionally associated with food animals, and infrequently associated with humans. The publication of the complete 2.341-Mb Arcobacter genome sequence has facilitated identification of unique virulence factors and genetic markers to monitor its transmission through the food chain and which underlie its distinctive epidemiology.
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The Nucleoid: an Overview
- Author: Akira Ishihama
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Citation: Ishihama A. 2009. The Nucleoid: an Overview, EcoSal Plus 2009; doi:10.1128/ecosalplus.2.6
- DOI 10.1128/ecosalplus.2.6
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This review provides a brief review of the current understanding of the structure-function relationship of the Escherichia coli nucleoid developed after the overview by Pettijohn focusing on the physical properties of nucleoids. Isolation of nucleoids requires suppression of DNA expansion by various procedures. The ability to control the expansion of nucleoids in vitro has led to purification of nucleoids for chemical and physical analyses and for high-resolution imaging. Isolated E. coli genomes display a number of individually intertwined supercoiled loops emanating from a central core. Metabolic processes of the DNA double helix lead to three types of topological constraints that all cells must resolve to survive: linking number, catenates, and knots. The major species of nucleoid core protein share functional properties with eukaryotic histones forming chromatin; even the structures are different from histones. Eukaryotic histones play dynamic roles in the remodeling of eukaryotic chromatin, thereby controlling the access of RNA polymerase and transcription factors to promoters. The E. coli genome is tightly packed into the nucleoid, but, at each cell division, the genome must be faithfully replicated, divided, and segregated. Nucleoid activities such as transcription, replication, recombination, and repair are all affected by the structural properties and the special conformations of nucleoid. While it is apparent that much has been learned about the nucleoid, it is also evident that the fundamental interactions organizing the structure of DNA in the nucleoid still need to be clearly defined.
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The Cold Shock Response
- Authors: Sangita Phadtare, and Masayori Inouye
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Citation: Phadtare S, Inouye M. 2008. The Cold Shock Response, EcoSal Plus 2008; doi:10.1128/ecosalplus.5.4.2
- DOI 10.1128/ecosalplus.5.4.2
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Abstract:
This review focuses on the cold shock response of Escherichia coli. Change in temperature is one of the most common stresses that an organism encounters in nature. Temperature downshift affects the cell on various levels: (i) decrease in the membrane fluidity; (ii) stabilization of the secondary structures of RNA and DNA; (iii) slow or inefficient protein folding; (iv) reduced ribosome function, affecting translation of non-cold shock proteins; (v) increased negative supercoiling of DNA; and (vi) accumulation of various sugars. Cold shock proteins and certain sugars play a key role in dealing with the initial detrimental effect of cold shock and maintaining the continued growth of the organism at low temperature. CspA is the major cold shock protein of E. coli, and its homologues are found to be widespread among bacteria, including psychrophilic, psychrotrophic, mesophilic, and thermophilic bacteria, but are not found in archaea or cyanobacteria. Significant, albeit transient, stabilization of the cspA mRNA immediately following temperature downshift is mainly responsible for its cold shock induction. Various approaches were used in studies to detect cold shock induction of cspA mRNA. Sugars are shown to confer protection to cells undergoing cold shock. The study of the cold shock response has implications in basic and health-related research as well as in commercial applications. The cold shock response is elicited by all types of bacteria and affects these bacteria at various levels, such as cell membrane, transcription, translation, and metabolism.
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Acetate-Based Methane Production
- Author: James G. Ferry
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Source: Bioenergy , pp 155-170
Publication Date :
January 2008
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Abstract:
This chapter explores the microbiology and biochemistry of acetate conversion to methane, a key component of biomethanation. It provides a fundamental background appropriate for stimulating advances to improve the process that will ensure biomethanation among the competitive alternatives to fossil fuels. Biomethanation of organic matter in nature occurs in diverse habitats such as freshwater sediments, rice paddies, sewage digesters, the rumen, the lower intestinal tract of monogastric animals, landfills, hydrothermal vents, coastal marine sediments, and the subsurface. Methanosarcina species synthesize tetrahydrosarcinapterin (H4SPT) which serves the same function as tetrahydromethanopterin (H4MPT). The aceticlastic and CO2 reduction pathways generate primary sodium and proton gradients that are the only possible driving forces for ATP synthesis. The production of acetate from complex biomass by fermentative and acetogenic anaerobes and the subsequent conversion of acetate to methane by aceticlastic methanogens are of primary importance in the biomethanation process. Aceticlastic methanogenesis is the major factor controlling the rate and reliability of the process; thus, a comprehensive understanding of these methanogens is paramount for developing an efficient process for biomethanation of renewable and waste biomass for use as a biofuel. Although the enzymology of reactions leading from acetate to methane by Methanosarcina species is fairly well understood, there have been only a few investigations reported on the mechanism of energy conservation and regulation of gene expression. Further, global proteomic and microarray analyses have identified a host of proteins and genes in Methanosarcina species, many with unknown functions, that may be important or essential.
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Comparative Genomics of Campylobacter Species Other than Campylobacter jejuni
- Author: William G. Miller
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Source: Campylobacter, Third Edition , pp 73-95
Publication Date :
January 2008
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Abstract:
The best characterized of the campylobacters is Campylobacter jejuni subsp. jejuni, a common commensal of warm-blooded animals, especially birds. The availability of eight new nonjejuni Campylobacter (NJC) genomes permits genomic comparisons between the NJC group and between the NJC genomes and the C. jejuni subsp. jejuni genomes. This chapter compares and contrasts the genomes of Campylobacter species other than C. jejuni subsp. jejuni. The primary phenotypic marker associated with Campylobacter plasmids is antibiotic resistance. Genes encoding resistance to tetracycline, kanamycin, and chloramphenicol have been identified on Campylobacter plasmids. Campylobacter plasmids fall into two basic groups: cryptic plasmids and megaplasmids. Homopolymeric G: C tracts have been identified in all of the NJC genomes. Several genes within the NJC genomes are frameshifted or have other defects and are considered putative pseudogenes. In Escherichia coli, the conjugative F episome cointegrates into the chromosome via homologous recombination between insertion sequences (IS) elements common to both the chromosome and the F plasmid, resulting in an Hfr strain. Helicobacter strains that possess the Entner-Doudoroff (ED) pathway contain a phosphoglucose isomerase (PGI) distinct from those found in the other Epsilonproteobacteria. A complete oxidative tricarboxylic acid (TCA) cycle is present in C. jejuni subsp. jejuni, as evidenced by the presence of all three subunits of succinate dehydrogenase (encoded by sdhABC). The chapter talks about restriction/modification systems, and virulence/pathogenicity loci. Genomic data for the NJC genomes will lead to new typing methods and perhaps culture methods.
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The A Factor Regulatory Cascade That Triggers Secondary Metabolism and Morphological Differentiation in Streptomyces
- Author: Sueharu Horinouchi
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Source: Chemical Communication among Bacteria , pp 363-377
Publication Date :
January 2008
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Abstract:
This chapter deals with the study of the biology and chemistry of γ-butyrolactone-type autoregulators that switch on secondary metabolism and morphological differentiation in Streptomyces. The A factor and receptor system in Streptomyces griseus acts as an all-or-nothing switch for both morphological and physiological differentiation. Escherichia coli carrying afsA produces two new substances that are absent in the broth of E. coli without afsA with their m/z 241 and 213 and the same MS/MS fragmentation pattern as A factor. AfsA is thus the key enzyme for the biosynthesis of γ-butyrolactones. Interestingly, a database search predicts that afsA and its homologs are distributed only in actinomycetes. In S. griseus, A factor production is controlled directly or indirectly by adpA in a two-step regulatory feedback loop. The major streptomycin resistance determinant, aphD, located just downstream of strR, encoding streptomycin-6-phosphotransferase, is also transcribed by read-through from the A factor-dependent strR promoter. The cotranscription of strR and aphD accounts for the prompt induction of streptomycin resistance by A factor and achieves a rapid increase in self-resistance just before induction of streptomycin biosynthesis.
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The Cold-Shock Response
- Authors: Masayori Inouye, Sangita Phadtare
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Source: Physiology and Biochemistry of Extremophiles , pp 180-193
Publication Date :
January 2007
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Abstract:
When a bacterial culture growing exponentially at a temperature optimum for its growth is shifted to low temperature, it exhibits cold-shock response. This is irrespective of the preferred optimum growth temperature; thus all types of bacteria such as psychrotrophic, psychrophilic, mesophilic, and thermophilic bacteria possess cellular machinery to elicit this response. Recent global transcript profiling of Escherichia coli cells undergoing cold shock showed that several genes encoding proteins involved in sugar transport and metabolism were induced by cold shock. Cold-shock response of cold-adapted bacteria is similar to that of mesophiles in aspects such as in many cases a lag phase of growth precedes acclimation to low temperature, specific proteins are induced by temperature downshift, membranes undergo adaptive changes, and enzymes are adapted to function at low temperature. One of the main differences in the cold-shock response of these two types of bacteria is the presence of cold acclimation proteins (Caps) in cold-adapted bacteria. The cold-shock response machinery of cyanobacteria is different from that of E. coli. The two main differences are: (i) the absence of CspA homologs and (ii) the presence of desaturases. Desaturases play an important role in cold-shock response of cyanobacteria. With the advent of DNA microarray technology, several groups have carried out global transcript profiling of cold-shock response of different bacteria. Cellular events occurring during cold-shock response are used in applications such as in food and agricultural industry and in research.
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Escherichia coli at the Intestinal Mucosal Surface
- Authors: Conway Tyrrell, Paul S. Cohen
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Source: Virulence Mechanisms of Bacterial Pathogens, Fourth Edition , pp 175-196
Publication Date :
January 2007
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Abstract:
Escherichia coli is the predominant facultative anaerobe in the gastrointestinal tracts of mammals. A large and growing body of evidence indicates that commensal E. coli strains grow in the intestine on nutrients acquired from mucus. Fluorescence in situ hybridization analysis of mouse intestinal thin slices showed that E. coli BJ4, a rat commensal isolate, is dispersed in the mucus layer but is not associated with the epithelium. Colonization resistance is the ability of a complete intestinal microflora to resist colonization by an invading bacterium. When E. coli and serovar Typhimurium enter the mouse cecum, they initially encounter cecal contents. Since E. coli and serovar Typhimurium fail to grow in cecal contents but grow rapidly in cecal mucus, they must be able to enter the mucus layer and grow by using available nutrients. Genetic screens based on signature-tagged mutagenesis identify individual mutants that are lost from pools of mutants used to infect suitable animals. It is becoming increasingly clear that once E. coli strains reach the large intestine, in order to colonize, they must enter the mucus layer and utilize nutrients there for growth.
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Rapid Methods for the Detection of Foodborne Pathogens: Current and Next-Generation Technologies
- Author: Peter Feng
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Source: Food Microbiology: Fundamentals and Frontiers, Third Edition , pp 911-934
Publication Date :
January 2007
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The history of biotechnology reveals that scientific discoveries lead to technological advances and these are usually followed by a phase of research and development to find applications for the technologies. The authors are currently in another application phase, where new technologies such as real-time PCR (rtPCR), DNA microarrays, and biosensors are providing very sophisticated tools for use in diagnostics. This chapter therefore includes discussions on these "next-generation technologies," which will have an impact on the way one can test for pathogens and toxins in foods. All identification assays require a pure culture of the unknown bacteria, which is then identified most often by its biochemical characteristics. It is a lengthy, labor-intensive, and media-consuming process. The introduction of miniaturized biochemical kits, which provide a quick biochemical profile of bacteria at a great savings in cost, labor, and time, has simplified the process of bacterial identification and continues to be important in regulatory testing of foods. Many of the new methods use next-generation technologies such as rtPCR, biosensors, and DNA chips, which may be more rapid, more sensitive, and capable of multitarget testing and hence are well suited for use in screening large numbers of samples in compliance or food security surveillance programs. Furthermore, comparative evaluation by with standard methods or validation of rapid methods is critical to document their efficacy in detecting foodborne pathogens and toxins.
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Genomics and Proteomics of Foodborne Microorganisms
- Authors: Todd R. Klaenhammer, Erika Pfeiler, Tri Duong
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Source: Food Microbiology: Fundamentals and Frontiers, Third Edition , pp 935-951
Publication Date :
January 2007
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This chapter outlines the basic concepts underlying genomics, proteomics, and microarray technologies of foodborne microorganisms. The heart of all genomics research lies in DNA sequencing. DNA sequencing method utilizes normal DNA replication with a template strand, a primer, DNA polymerase, and a mix of deoxynucleotide triphosphates (dNTPs). DNA microarray analysis is a relatively new technology that allows investigators to take a genome-wide approach to biological systems. While the details of the Gad system were described primarily by experiments with Escherichia coli, genomics and bioinformatics have enabled researchers to identify and study the effects of these genes in other microorganisms. Foods and their microenvironments could potentially be better designed and formulated to minimize the expression of undesirable pathogenic traits (e.g., acid tolerance, virulence, and toxin formation) or to optimize the expression of beneficial properties in desirable microorganisms (e.g., cryoprotection, acidification rates, and adherence to intestinal tissues). The nature of food microbiology has changed dramatically from its historical emphasis on microbial phenotypic properties and behavior to a new perspective dominated by genomic and comparative genomic information. The food microbiologists of the future will become increasingly reliant on genomics and the other omics technologies in their efforts to understand and control microorganisms associated with foods.
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Part III Overview
- Author: James B. Kaper
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Source: Evolution of Microbial Pathogens , pp 245-250
Publication Date :
January 2006
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This chapter presents an overview on pathogenic organisms. Lan and Reeves describe the evolution of three important groups of enteric pathogens, Salmonella enterica, Escherichia coli (including Shigella), and Yersinia spp. At one time, the genus Salmonella was divided into more than 2,000 species based on surface and flagella antigens, but these are now considered to be one species, S. enterica, which is subdivided into seven subspecies and 2,501 serovars. Yersinia pestis is a recently emerged clone that shares 99.7% identity in 16S ribosomal RNA with Y. pseudotuberculosis and might more properly called Y. pseudotuberculosis Pestis. This clone has acquired additional virulence factors such as Yersinia murine toxin (Ymt) and plasminogen activator (Pla) that allow it to colonize the flea gut, be transmitted to a new host via biting, and disseminate hematogenously from the infected site. Clonal associations probably reflect the historical or prehistorical dissemination of M. tuberculosis with human migrations, similar to the associations that have been established for H. pylori. The great diversity of fungi, among which pathogenic species are dispersed across three phyla, plus the lack of information on virulence factors and the pathogenesis of fungal infections make the promulgation of generalized concepts for the evolution of fungal pathogens extremely difficult, if not impossible. In some cases, such as Legionnaires’ disease, archived clinical specimens may reveal that the disease existed decades ago but appropriate diagnostic techniques did not yet exist.
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Immunofluorescent Antinuclear Antibody Tests
- Authors: Arthur R. Bradwell, Richard G. Hughes, Abid R. Karim
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Source: Manual of Molecular and Clinical Laboratory Immunology, 7th Edition , pp 995-1006
Publication Date :
January 2006
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This chapter reviews the history and the method for antinuclear autoantibodies (ANAs) testing on HEp-2 cells and its utility in a laboratory setting. Antinuclear antibody tests have their origin in the “L.E.” cell phenomenon, which was the observation that neutrophils from patients with systemic lupus erythematosus (SLE) ingested other leukocytes. Widespread commercial production of HEp-2 cells and the development of national quality control schemes ensured that the test entered worldwide routine laboratory usage. The nuclear envelope is the membrane that maintains the integrity of the nucleoplasm during interphase. Important autoantibodies are found against most of the cell cycle-related structures that have been mentioned, such as the centromeres, proliferating cell nuclear antigen, mitotic spindle proteins, and centriole proteins. The pattern on HEp-2 cells is usually sufficient to identify most of these autoantibodies. The majority of laboratories use a non-affinity-purified, fluorescein-conjugated second antibody directed against IgG, -A, and -M heavy chains (or anti-IgG heavy and light chains). The HEp-2 cells should be observed by using an epifluorescence microscope fitted with filters appropriate for fluorescein detection. The largest variation in results reported to quality control schemes relates to differences in fluorescence intensity of the samples. The ANA immunofluorescent HEp-2 test is intended to be used for screening and titration of circulating antinuclear antibodies. Autoantibodies occur in both physiological and pathological conditions.
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Molecular Methods for Detection of Antibiotic Resistance
- Authors: Henk J. M. Aarts, Beatriz Guerra, Burkhard Malorny
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Source: Antimicrobial Resistance in Bacteria of Animal Origin , pp 37-48
Publication Date :
January 2006
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The combination of molecular characterization of resistant strains with precise identification of the antibiotic resistance gene(s) or mutation(s) and the genetic elements involved in the dissemination of these genes is an effective approach in the control of the spread of antibiotic resistance. Molecular methods also help to determine the location of the gene and to differentiate between horizontal gene transfer and clonal spread. This chapter describes polymerase chain reaction (PCR) and microarray analysis in detail, and specifies applications of these methods in the investigation of antibiotic-resistant strains. Numerous PCR assays for the detection of antibiotic resistance genes have been developed and the development of microarrays for the simultaneous detection of a large number of these genes and the genetic elements involved in their dissemination is in progress. The implementation of molecular methods in routine analysis can be achieved only when it is supported by the proper validation of the methods and the availability of the necessary controls, reference strains, and educated personnel. The molecular characterization of antibiotic-resistant strains can help to identify atypical resistant strains, describe new outbreak strains at an early stage, elucidate the epidemiology of resistant strains at a genotypic level, and explain the processes leading to the selection of resistant and virulent strains. In addition, molecular methods can allow proper risk assessment with respect to the use of antimicrobial substances. If the transcriptional and translational expression of antibiotic resistance genes can be better understood, molecular methods may replace phenotypic measurements.
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The Paradigm Shift in Microbial Prospecting
- Author: Alan T. Bull
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Source: Microbial Diversity and Bioprospecting , pp 241-249
Publication Date :
January 2004
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This chapter explores the posited paradigm shift from traditional microbiology to bioinformatics in the search for exploitable biology. It also refers to the extraordinary rate of biological data acquisition and deposition in the world's databases. These databases are classified by type (e.g., genomes, gene expression, proteins, RNA sequences, pathology), but the distinction between categories is often arbitrary and individual databases may provide more than one type of information. Integration of the molecular biology databases, although by no means perfect, has attracted serious attention and may be able to deliver useful information on issues such as a protein's structure, function, phylogenetic occurrence, expression, and protein-protein interactions. Similarly, the various bioinformatics research groups and commercial organizations are developing multidatabase query systems that enable interoperability of heterogeneous, formerly incompatible resources. Within the medical sector, bioinformatics objectives have included (i) the detection and identification of pathogens, (ii) the identity of drug targets, and (iii) the search for new drugs and vaccines. The final stratagem to be considered for producing novel exploitable biology is directed evolution, which, as genome sequencing projects continue to grow, promises to become a principal route for search and discovery. Evolution via natural and artificial (sensu traditional animal and plant breeding) selection is dependent on genetic variation, inheritance, and differential reproduction.