![]() However, only limited information on the effects of various handling and storage conditions on the stability of bacterial DNA in CSF is available. The stability of target bacterial DNA during CSF storage is an important practical matter in clinical laboratories. Therefore, a negative PCR result can be used with moderate confidence to rule out a diagnosis of infection ( 33). However, the sensitivity and specificity of PCR assay for detection of pathogens may not be better than those of culture assay, which has been standardized and validated for most pathogens, due to the dependability of PCR sensitivity on the assay process. In fact, a number of trials using PCR for detection of a broad range of bacteria in CSF specimens have been reported ( 37, 38, 42, 66). Therefore, performing PCR using a CSF specimen will become the first-line diagnostic test for CNS infections ( 11, 33), due to a sensitivity requiring only a limited amount of CSF, the specificity of the assay, and speed. Thus, the available amount of CSF and numbers of samplings for diagnosis are limited. ![]() The normal adult produces approximately 500 ml of CSF per day, with approximately 150 ml of CSF in the CNS at any given time ( 34). Therefore, detection of even a few microorganisms in CSF by a standardized protocol is a critical matter for diagnosis of such diseases. In particular, recent studies indicate possible involvement of microorganisms in specific diseases of the CNS, including Alzheimer's disease and MS ( 3, 30, 45, 56). Nevertheless, detection, identification, and quantitation of microorganisms in CSF is important in diagnosis of meningitis and other CNS infections. However, it must be noted that detection of microbes in the CSF does not always indicate a CNS infection, since impairment of the blood-brain barrier may permit transit of microbes. Since CSF is considered germfree, detection of microbes in CSF, even in low numbers, provides valuable information about possible infection. Because CSF has important functions, including cushioning the brain, maintaining a constant intracranial pressure, providing nutrients, and removing toxic metabolites from the CNS, an indirect assessment of brain status can be obtained from the CSF. pneumoniae in CSF, will be discussed.ĬSF is widely utilized for diagnosis of diseases of the central nervous system (CNS). ![]() Therefore, in this review general PCR protocols for detection of bacteria in clinical specimens, as well as a specific example of using PCR for detection of C. pneumoniae is difficult to culture in vitro, often low numbers of bacteria may be detected in the CSF of patients with chronic neurological diseases by PCR. In this regard, current studies concerning detection of Chlamydia pneumoniae in CSF obtained from patients with multiple sclerosis (MS) by using PCR provide a good example for discussion of use of the PCR assay in diagnosis. ![]() In particular, a diagnosis based on detection of a few bacteria in clinical specimens by using PCR must be carefully evaluated technically as well as microbiologically. Since a variety of clinical specimens, such as blood, urine, sputum, CSF and others, vary in regard to the nature of the content and amount available, careful design of the PCR assay for each specific specimen before a PCR application is conducted is essential. Even though there are many publications concerning basic protocols of a PCR assay, including DNA extraction and preparation as well as the amplification and detection of amplicons, PCR detection of bacteria in clinical specimens such as cerebrospinal fluid (CSF) has not yet been reviewed. For instance, it is known that the sensitivity and specificity of a PCR assay is dependent on target genes, primer sequences, PCR techniques, DNA extraction procedures, and PCR product detection methods. ![]() However, the application of PCR to clinical specimens has many potential pitfalls due to the susceptibility of PCR to inhibitors, contamination and experimental conditions. In particular, when specific pathogens that are difficult to culture in vitro or require a long cultivation period are expected to be present in specimens, the diagnostic value of PCR is known to be significant. The PCR is the most sensitive of the existing rapid methods to detect microbial pathogens in clinical specimens. ![]()
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