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Original Article



Development of a microarray-based method for simultaneous detection and serotyping of Streptococcus pneumoniae from culture-negative serum samples

Avid Hussain Shaik, Vandana Govindan, Geetha Nagaraj, Kadahalli Lingegowda Ravikumar.


Abstract

Streptococcus pneumoniae identification and serotyping are vital for disease management, surveillance, vaccine development and to monitor the emerging serovars. Despite the potential benefits of conventional typing method based on culture isolate, it lacks the ability to type directly from the clinical samples. To address the challenge, a novel microarray method was developed to identify and serotype S. pneumoniae in culture positive and culture negative serum specimens. The custom pneumococcal microarray chip was designed using sure design software and evaluated with 90 reference strains, culture positive and qmPCR (quantitative multiplex Polymerase Chain Reaction) positive (n=8) and culture negative qmPCR positive (n=96) serum samples. To selectively amplify S. pneumoniae DNA from serum, three different methods: a. CPS PCR, b. Whole genome amplification (WGA) and c. Microbial DNA enrichment was assessed. An in-house developed excel based software was used to quantify the signals from each serotype. With a combination of Microbial DNA enrichment, the custom pneumococcal microarray chip identified all reference strain and serum samples serogroup/type information accurately. Of the 104 serum samples, 65 were uniquely identified and 39 were assigned with a combination of their homologous types. 100% concordance was observed with the results of qmPCR and PCRSeqTyping methods with an additional advantage of multiple serotype detection. Our results signify the ability of the microarray technology to identify and detect S. pneumoniae serogroup/type from culture-negative serum specimens. The test is of use even in patients with prior antibiotic treatment and can be used in surveillance and vaccine impact studies.

Key words: Streptococcus pneumoniae, microarray, molecular serotyping, serum






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