1887

Abstract

The molecular identification and its drug-resistance profile are important to choose the correct therapy.

This work developed a multiplex real-time PCR (mqPCR) for detection of clarithromycin resistance genes for the group.

Isolates received by Adolfo Lutz Institute from 2010 to 2012, identified by PCR restriction enzyme analysis of a fragment of the 65 gene (PRA-65) as type 1 (=135) and 2 (=71) were used. Drug susceptibility test (DST) for CLA were performed with reading on days 3 and 14. Subespecies identification by 65 and B genes sequencing and (41) and genes for mutation detection and primer design were performed. (41) gene deletion was detected by conventional PCR. Primers and probes were designed for five detections: (41) gene full size and with deletion; (41) gene T28 and C28; gene A2058.

In total, 191/206 (92.7 %) isolates were concordant by all methods and 13/206 (6.3 %) were concordant only between molecular methods. Two isolates (1.0 %) were discordant by mqPCR compared to gene sequencing. The mqPCR obtained 204/206 (99.0 %) isolates in agreement with the gold standard, with sensitivity and specificity of 98 and 100 %, respectively, considering the gold standard method and 92 and 93 % regarding DST.

The mqPCR developed by us proved to be an easy-to-apply tool, minimizing time, errors and contamination.

Funding
This study was supported by the:
  • Fundação de Amparo à Pesquisa do Estado de São Paulo (Award Grant #2018/10725-6)
    • Principle Award Recipient: EricaChimara
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2023-03-15
2024-03-28
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