Direct Detection Methods of Streptococcus, Enterococcus, and Similar Organisms
المؤلف:
Patricia M. Tille, PhD, MLS(ASCP)
المصدر:
Bailey & Scotts Diagnostic Microbiology
الجزء والصفحة:
13th Edition , p252-253
2025-05-17
607
Antigen Detection
Antigen detection screening methods are available for several streptococcal antigens. Detection of antigens is possible using latex agglutination or enzyme-linked immunosorbent assay (ELISA) technologies. These commercial kits have been reported to be very specific, but false-negative results may occur if specimens contain low numbers of S. pyogenes. Sensitivity has ranged from approximately 60% to greater than 95% depending on the methodology and other variables. Therefore, many microbiologists recommend collecting two throat swabs from each patient. If the first swab yields a positive result by a direct antigen method, the second swab can be discarded. However, for those specimens in which the rapid antigen test yielded a negative result, a blood agar plate or selective streptococcal blood agar plate should be inoculated with the second swab.
Several commercial antigen detection kits are available for diagnosis of neonatal sepsis and meningitis caused by group B streptococci. Developed for use with serum, urine, or cerebrospinal fluid (CSF), the best results have been achieved with CSF; false-positive results have been a problem using urine. Because neonates acquire S. agalactiae infection during passage through the colonized birth canal, direct detection of group B streptococcal antigen from vaginal swabs has also been attempted. However, direct extraction and latex particle agglutination have not been sensitive enough for use alone as a screening test.
Latex agglutination kits to detect the capsular polysaccharide antigen of the pneumococcus have also been developed for use with urine, serum, and CSF, although they are no longer commonly used in clinical microbiology laboratories.
Molecular Diagnostic Testing
Nucleic acid based testing is available and offers a rapid and increased specificity as compared to traditional identification schemes. Polymerase chain reaction is available for the detection of an internal sequence of the CAMP-factor (cfb gene) for group B streptococci. Analyte-specific reagents are available from Roche Applied Science (Indianapolis, Indiana) for the detection of the ptsI gene of group B and group A streptococci. The two groups are differentiated based on specificity of the sequences within the primer pairs for the assay. Gen-Probe Incorporated (San Diego, California) has developed several DNA probe assays for the differentiation of streptococci. The GASDirect test is a DNA probe hybridization assay for the detection of group streptococcal RNA from throat swabs. The ACCUPROBE group B Streptococcus assay is a hybridization protection assay that utilizes a DNA probe for the detection of 16 s ribosomal RNA sequences unique to Streptococcus agalactiae.
A fully integrated automated real-time PCR-based GeneXpert system has been developed by Cepheid (Sunnyvale, California). The system completely auto mates the sample preparation, DNA extraction, amplification, and detection of the target sequence within a closed system. The GeneXpert platform offers a qualitative assay for the detection of group B Streptococcus DNA directly from a swab.
Gram Stain
All the genera described in this chapter are gram-positive cocci. Microscopically, streptococci are round or oval shaped, occasionally forming elongated cells that resemble pleomorphic corynebacteria or lactobacilli. They may appear gram-negative if cultures are old or if the patient has been treated with antibiotics. Gemella haemolysans is easily decolorized. S. pneumoniae is typically lancet shaped and occurs singly, in pairs, or in short chains (Figure 1).
Fig1. S. pneumoniae lancet-shaped diplococci in Gram stain; note the encapsulated organisms as evident by the clear “halo.”
Growth in broth should be used for determination of cellular morphology if there is a question regarding staining characteristics from solid media. In fact, the genera described in this chapter are subdivided based on whether they have a “strep”-like Gram stain or a “staph” like Gram stain. For example, Streptococcus and Abiotrophia growing in broth form long chains of cocci (Figure 2), whereas Aerococcus, Gemella, and Pediococcus grow as large, spherical cocci arranged in tetrads or pairs or as individual cells. Leuconostoc may elongate to form coccobacilli, although cocci are the primary morphology. The cellular arrangements of the genera in this chapter are noted in Tables 1 and 2.
Fig2. Chains of streptococci seen in Gram stain prepared from broth culture.
Table1. Differentiation of Catalase-Negative, Gram-Positive Coccoid Organisms Primarily in Chains
Table2. Differentiation of Catalase-Negative, Gram-Positive, Coccoid Organisms Primarily in Clusters or Tetrads
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