Ribonucleic　acids　(RNA)　play　active　roles　within　cells　or　viruses　by　catalyzing　biological　reactions,　controlling　gene　expression,　and　communicating　responses　to　cellular　signals.　Rapid　monitoring　RNA　variation　has　become　extremely　important　for　appropriate　clinical　decisions　and　frontier　biological　research.　However,　the　most　widely　used　method　for　RNA　detection,　nucleic　acid　amplification,　is　restricted　by　a　mandatory　temperature　cycling　period　of　≈1　h　required　to　reach　target　detection　criteria.　Herein,　a　direct　detection　approach　via　single-atom　site　integrated　surface-enhanced　Raman　scattering　(SERS)　monitoring　nucleic　acid　pairing　reaction,　can　be　completed　within　3　min　and　reaches　high　sensitivity　and　extreme　reproducibility　for　COVID-19　and　two　other　influenza　viruses’　detection.　The　mechanism　is　that　a　single-atom　site　on　SERS　chip,　enabled　by　positioning　a　single-atom　oxide　coordinated　with　a　specific　complementary　RNA　probe　on　chip　nanostructure　hotspots,　can　effectively　bind　target　RNA　analytes　to　enrich　them　at　designed　sites　so　that　the　binding　reaction　can　be　detected　through　Raman　signal　variation.　This　ultrafast,　sensitive,　and　reproducible　single-atom　site　SERS　chip　approach　paves　the　route　for　an　alternative　technique　of　immediate　RNA　detection.　Moreover,　single-atom　site　SERS　is　a　novel　surface　enrichment　strategy　for　SERS　active　sites　for　other　analytes　at　ultralow　concentrations.　©　2024　Wiley-VCH　GmbH.