Bacterial colonization of the upper airways of children positive and negative for SARS-CoV-2 during the COVID-19 pandemic.

BACKGROUND: Our understanding of the influence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection on bacterial colonization in the children's upper nasopharyngeal tract during the coronavirus infectious disease (COVID-19) pandemic is limited. This study aimed to determine whether there were any differences in bacterial colonization between asymptomatic children with or without a positive SARS-CoV-2 quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR) results in the community setting. METHODS: A cross-sectional community-based exploratory study was conducted from March to May 2021 in Semarang, Central Java Province, Indonesia. Using stored nasopharyngeal swabs collected from children under 18 years as a contact tracing program, we performed a real-time quantitative (qPCR) for the most important bacterial colonizing pathogens: Streptococcus pneumoniae, Haemophilus influenzae, Staphylococcus aureus, and Klebsiella pneumoniae. RESULTS: Swabs from a total of 440 children were included in this study, of which 228 (51.8%) were RT-qPCR-confirmed SARS-CoV-2 positive. In the 440 children, colonization rates were highest for H. influenzae (61.4%), followed by S. pneumoniae (17.5%), S. aureus (12.0%), and K. pneumoniae (1.8%). The co-occurrence of both S. pneumoniae and H. influenzae in the upper respiratory tract was significantly associated with a SARS-CoV-2 negative RT-qPCR. In contrast, colonization with only S. aureus was more common in SARS-CoV-2-positive children. CONCLUSION: Overall, this exploratory study concludes that there is a significant difference in the bacterial nasopharyngeal colonization pattern between SARS-CoV-2 positive and negative in asymptomatic children in the community in Indonesia.

3' Tth Endonuclease Cleavage Polymerase Chain Reaction (3TEC-PCR) Technology for Single-Base-Specific Multiplex Pathogen Detection using a Two-Oligonucleotide System.

Polymerase chain reaction (PCR) is the standard in nucleic acid amplification technology for infectious disease pathogen detection and has been the primary diagnostic tool employed during the global COVID-19 pandemic. Various PCR technology adaptations, typically using two-oligonucleotide dye-binding methods or three-oligonucleotide hydrolysis probe systems, enable real-time multiplex target detection or single-base specificity for the identification of single-nucleotide polymorphisms (SNPs). A small number of two-oligonucleotide PCR systems facilitating both multiplex detection and SNP identification have been reported; however, these methods often have limitations in terms of target specificity, production of variable or false-positive results, and the requirement for extensive optimisation or post-amplification analysis. This study introduces 3' Tth endonuclease cleavage PCR (3TEC-PCR), a two-oligonucleotide PCR system incorporating a modified primer/probe and a thermostable cleavage enzyme, Tth endonuclease IV, for real-time multiplex detection and SNP identification. Complete analytical specificity, low limits of detection, single-base specificity, and simultaneous multiple target detection have been demonstrated in this study using 3TEC-PCR to identify bacterial meningitis associated pathogens. This is the first report of a two-oligonucleotide, real-time multiplex PCR technology with single-base specificity using Tth endonuclease IV.