Diagnostic performance of an in-house multiplex PCR assay and the retrospective surveillance of bacterial respiratory pathogens at a teaching hospital, Kelantan, Malaysia

Nik Mohd Noor Nik Zuraina, Suharni Mohamad, Habsah Hasan, Mohammed Dauda Goni, Siti Suraiya, Nik Mohd Noor Nik Zuraina, Suharni Mohamad, Habsah Hasan, Mohammed Dauda Goni, Siti Suraiya

Abstract

Respiratory tract infections (RTIs), including pneumonia and pulmonary tuberculosis, are among the leading causes of death worldwide. The use of accurate diagnostic tests is crucial to initiate proper treatment and therapy to reduce the mortality rates for RTIs. A PCR assay for simultaneous detection of six respiratory bacteria: Haemophilus influenzae, Klebsiella pneumoniae, Mycobacterium tuberculosis, Pseudomonas aeruginosa, Staphylococcus aureus and Streptococcus pneumoniae, was developed in our lab. The current study aimed to evaluate the performance of this assay along with the retrospective surveillance of respiratory pathogens at a teaching hospital in Kelantan, Malaysia. Leftover sputa (n = 200) from clinical laboratories were collected and undergone DNA template preparation for PCR analysis. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of the PCR assay were determined in comparison with the gold standard sputum culture. Overall, the accuracy performance of this assay was 94.67% (95% CI: 90.87% to 97.21%) with sensitivity, specificity, PPV and NPV of 100%, 91.67%, 87.1% and 100%, respectively. Based on the organisms detected from sputa, K. pneumoniae ranked as the top isolate (n = 48), followed by P. aeruginosa (n = 13) and H. influenzae (n = 10). Surveillance among the patients showed that the associations of bacterial positive with gender and means of acquisition were found significant (p values = 0.049 and 0.001, respectively). Besides the promising performance of this ready-to-use molecular-based assay for the rapid detection of selected bacteria pathogens, this study also highlighted significant spread of K. pneumoniae RTIs in the community.

Keywords: Haemophilus influenzae; Klebsiella pneumoniae; Multiplex PCR; Pseudomonas aeruginosa; community-acquired pneumonia; respiratory tract infections.

Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1.
Figure 1.
Representative agarose gel image for the diagnostic performance of multiplex PCR assay on the sputum specimens from Hospital USM patients (n = 16). This image shows the presence of target amplicons as single and multiple bands (Lane 7) on positive specimens as well as the absence of amplicon on negative specimens. Lane M, 100 bp DNA ladder; lane N, no template control; lane P, positive amplification control; lane 1–16, presence or absence of PCR amplicons from individual sputa.

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