Usefulness of Pneumotest-Latex for Direct Serotyping of Streptococcus pneumoniae Isolates in Clinical Samples

Abstract

This study evaluated the usefulness of the Pneumotest-Latex assay for serotyping Streptococcus pneumoniae isolates directly in clinical samples. With an agreement of 88.1% with a PCR-based reference method, this test can be a useful tool for this study purpose, especially in clinical laboratories that do not have access to nucleic acid amplification technologies. The Pneumotest-Latex assay (Statens Serum Institut, Copenhagen, Denmark) is a rapid latex agglutination test intended for serotyping or serogrouping Streptococcus pneumoniae isolates (1). It seems to be promising also for sample-based serotyping, as noted previously by Sanz et al. (2), who used this test for pneumococcal serotyping from incubated blood culture bottles. The aim of this study was to evaluate the usefulness of this test for serotyping S. pneumoniae isolates directly without any previous enrichment step using clinical samples from patients with invasive pneumococcal disease (IPD).(The data included in the manuscript were presented in part at the 21st European Congress of Clinical Microbiology and Infectious Diseases, Milan, Italy, in 2011.) Between January 2007 and May 2013, 4,290 clinical samples were sent to the National Reference Centre for Bacterial Meningitis (NRCBM) (Warsaw, Poland), with pneumococcal DNA detected in 165 samples (3.8%). In daily practice, the NRCBM receives culture-negative samples from normally sterile sites for PCR-based species identification. These samples were sent by local laboratories, with each having Ն250 l volume, as required for molecular diagnostics on specimens from which no growth has been observed after 24 h of incubation. Our study encompassed only those samples with a volume sufficient to perform the Pneumotest-Latex assay. Finally, 64 clinical samples from 59 IPD patients with detected pneumococcal DNA were also studied on the following samples: blood, collected in EDTA or heparin (n ϭ 8), serum (n ϭ 4), cerebrospinal fluid (CSF) (n ϭ 40), pleural fluid (n ϭ 4), and 8 postmortem samples (2 liver, 2 kidney, 1 lung, 1 spleen, and 2 blood samples). Additionally, 18 clinical samples (blood, 7; serum, 1; CSF, 6; postmortem specimens, 4) from 17 patients with invasive meningococcal disease (IMD) were used as negative controls for serotyping.The presence of pneumococcal DNA in the clinical samples was confirmed by PCR amplification of the ply (pneumolysin), lytA (autolysin), and cps (capsular polysaccharide) genes, as previously described (3-5). Clinical samples that were found to be positive for S. pneumoniae were collected for serotyping. Because the Pneumotest-Latex assay is intended for serotyping isolates in pure culture, it was necessary in our procedure to include additional sample preparation steps borrowed from the procedures of commercially available latex agglutination tests that directly detect pneumococcal antigens in body fluids (e.g., the Becton, Dickinson Directigen meningitis combo test and Bio-Rad Pastorex meningitis test). These procedures are based on a reaction with capsular polysacch...

Pneumococcal Capsules and Their Types: Past, Present, and Future

“…However, with the developments of vaccination and the resulting serotype shift, serotyping of pneumococci has returned to the forefront of clinical interest. As long as pneumococcal vaccines are based on PS capsules, there will be a need for methods to determine capsular types (202)(203)(204)(205). A serotyping system should ideally detect all the known pneumococcal serotypes and then distinguish each serotype; however, in view of potentially vast capsular diversity, one needs to consider how much resolution a serotyping system needs and adjust analytical demands accordingly.…”

“…The test is easy to use and can be used by individuals with less training. Consequently, this system has been used extensively since its introduction (204,205) and may be recommended as the most practical and preferred method at present.…”

Identification of Streptococcus pneumoniae and other Mitis streptococci: importance of molecular methods

“…Pneumotest-Latex (SSI) is a fast and easy to perform a test, however, also in this case, cross-reactions with S. mitis and S. oralis were occasionally observed for some reagent pools [ 50 ]. This test can also be applied directly to clinical samples [ 51 ]. Negative results of the Quellung test do not exclude that an isolate represents S. pneumoniae since non-serotypable (rough) pneumococci lacking capsule are observed and in fact, can be very common in some settings [ 52 ].…”

Bacterial Respiratory Infections Complicating Human Immunodeficiency Virus

“…Originally developed for improved serotyping/serogrouping, 100 this procedure has also been found to have diagnostic potential, demonstrating a level of 88.1% agreement with a molecular PCR-based diagnostic method. 101 Quantitative RT-PCR-based detection of pneumococcal nucleic acid, in biological fluids, often combined with simultaneous detection of antibiotic resistance genes, is currently the method of choice for rapid diagnosis of pneumococcal infection. Of several genes investigated, including those encoding autolysin (lytA), pneumolysin, pneumococcal surface adhesin A, Spn9802, and capsular polysaccharides, detection of lytA appears to offer the highest specificity and sensitivity.…”