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A clinically suspected infection is ultimately confirmed by isolation or detection of the infectious agent. Subsequent identification of the microorganism and antibiotic susceptibility tests further guide effective antimicrobial therapy. Bloodstream infection is the most severe form of infection and is frequently life-threatening, and blood culture to detect circulating microorganisms has been the diagnostic standard. Much of the scientific and technologic advances in blood culture were made from the 1970s to the 1990s; this chapter briefly reviews various aspects of blood culture with emphasis on automated culturing systems.

The association of with peptic ulcer disease and gastric cancer was first proposed by Warren and Marshall in 1983 (Warren and Marshall, 1983). In February 1994, the National Institutes of Health Consensus Development Conference concluded that infection is the major cause of peptic ulcer disease, and all patients with confirmed peptic ulcer disease associated with infection should receive treatment with antimicrobial agents (Yamada et al., 1994). The International Agency for Research on CancerWorking Group of theWorld Health Organization categorized as a group I, or definite, human carcinogen (Versalovic, 2003). Based on the data retrieved during the National Health Interview Survey of 1989, 10% of adult U.S. residents reported physician-diagnosed ulcer disease, among whom one third had an ulcer in the past year (Sonnenberg and Everhart, 1996). In developing countries, the prevalence of carriers can be as high as 70–90%. Most patients acquire the infection at childhood. The prevalence of the infection in developed countries is lower, ranging from 25% to 50% (Dunn et al., 1997). Seroprevalence studies demonstrate an increasing rate in adults of 3–4% per decade (Cullen et al., 1993; Sipponen et al., 1996; Kosunen et al., 1997; Versalovic, 2003).

Immunoassays for the detection of the antigens of microorganisms remain important tools for the diagnosis and management of infectious diseases. Great strides have been made since the introduction of the early precipitation and agglutination assays in increasing the sensitivity, specificity, standardization, and automation of antigen tests (Hage, 1999; Carpenter, 2002; Constantine and Lana, 2003; Peruski and Peruski, 2003). Antigen tests have long been used to detect infectious agents that are difficult, slow, or hazardous to culture. However, antigen detection methods are especially useful for rapid diagnosis, whether in the clinic, emergency department, doctor’s office, or the central laboratory. Recently, simple one-step assays have been introduced that can provide results in 15 min with dramatic benefits to physician decision-making.

In addition to basic microbiological methods, such as microscopy and culture, to detect pathologic organisms, antigen or antibody detection methods by immunoassay and nucleic acid detection by amplification technology have been developed, are commonly used, and will be expanded further for rapid and accurate diagnosis of the common or newly emerging infection-causing agents, such as viruses, in clinical as well as public health laboratories. Since the first competitive radioimmunoassay was developed more than 40 years ago for human insulin detection (Yalowand Berson, 1960), immunoassays have been developed with emphasis on fast and sensitive detection technologies and automated systems. Due to the demand of large-scale screening for epidemiology, blood bank, prenatal care, and diagnosis of HIV and hepatitis, more immunodiagnostic procedures are performed using instruments and reagents similar to traditional immunochemistry platforms, including tests for oncology, toxicology, cardiology, and endocrinology. Immunoassays for detection of host-produced antibodies directed against microorganisms, particularly viruses, is now one of the most widely used analytical techniques in laboratory medicine (Andreotti et al., 2003; Peruski and Peruski, 2003).

Phenotypic testing of bacterial antimicrobial resistance has been widely used in clinical and diagnostic microbiology laboratories. These methods have been well studied and standardized. They have the advantages of being low cost, easy to perform (automated systems), and interpretation criteria readily available for commonly encountered organisms. These assays also are essential for new resistance discovery.

The first step in microbial identification is the phenotypic assessment of the growing colony. In many cases, the colonial morphology such as color, shape, size, hemolytic reaction, and growth characteristics on various selective and differential media can place an organism in a single family, genus, or even species level. In fact, assessing the ability of an organism to grow in various laboratory media and their oxygen requirement coupled with Gram-stain morphology and a few rapid tests such as catalase, oxidase, coagulase, and indole can provide preliminary identification for most of the clinically significant isolates. For example, it is very likely that an organism that grows on MacConkey agar plate and ferments lactose is a member of the family Enterobacteriaceae or that an oxidase-positive non–lactose fermenting Gram-negative rod that has distinct grape odor is .

Bacterial infections account for a large proportion of people admitted to hospitals each year as well as some acquired by patients already in medical care. These can arise from the ingestion of contaminated food or exposure to nonsterile environments through wounds where opportunistic pathogenic bacteria are present. The symptoms and treatment of these illnesses vary and although some clues can be obtained from observing a patient’s symptoms, the causative agent needs to be determined in order for a complete understanding of the nature of the infection, its origin, and the appropriate treatment. It is therefore of immense importance to characterize and identify bacteria wherever they are found in significant quantities, not only to aid clinicians with their diagnosis but also to prevent outbreaks of infections from potential medical, environmental, or terrorist sources. Furthermore, the identification of bacteria needs to be as rapid as possible. In this chapter, the use mass of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is discussed as an emerging technology for the rapid characterization and identification of bacteria.

The most important property of nucleic acid is its nucleotide sequence, which carries the identity of unique organisms. The basic principle of nucleic acid probebased assay is the intrinsic ability of single-strandedDNAorRNAto anneal specifically to a complementary sequence and form a double-stranded hybrid. Most microorganisms encountered in the clinical microbiology laboratory can be identified using conventional methods. However, the conventional/culture-based approach may take a long time to identify slow-growing or fastidious organisms. Nonviable or nonculturable organisms simply can not be identified by conventional/culturebased approach. The nucleic acid probe-based approach provides a rapid, specific way of detecting/identifying microorganisms. Nucleic acid probe-based microbial identification is widely used in clinical laboratories. The probes can be used for identification of microorganism directly from the specimen, from culture, or on formalin-fixed and paraffin-embedded tissue.

The field of molecular diagnostics has rapidly expanded to include technology for accurate and timely determination of clonal relatedness of microorganisms of epidemiological interest as well as the detection of infectious agents in real-time. The predominant technique used for strain characterization has been pulsed-field gel electrophoresis (PFGE), first developed in the early 1980s, to genotype microorganisms by electrophoretic separation of chromosomal DNA by molecular weight (Van der Ploeg et al., 1984). Over the years, this technology has proved to be a powerful tool used alone or in conjunction with restriction endonuclease digestion of the DNA in order to understand the evolution of antimicrobial resistance generated within a single clone and to determine genetic relatedness among microbial strains in industrial and agricultural settings, as well as health care associated epidemiologic investigations. This chapter will discuss the principle characteristics and the clinical applications of PFGE technology, including examples that illustrate its successful application to epidemiology. The strengths and limitations of PFGE are discussed as well as alternative strain-typing methods.