Jungeun Park ¹ Sung-Il Cho ¹ Sang-Gu Kang ¹ Jee-Woun Kim ¹ Sunkyung Jung ² Sun Hwa Lee ² Kyou-Sup Han ² Seungsik Hwang ^1*

• ¹ Graduate School of Public Health, Seoul National University, Seoul, Republic of Korea
• ² Seegene Medical Foundation, Seoul, Republic of Korea

Background: With the emergence of COVID-19 cases, governments quickly responded with aggressive testing, contact tracing, isolation and quarantine measures. South Korea's testing strategy primarily relied on real-time reverse-transcriptase polymerase chain reaction (real-time RT-PCR), focusing on cycle threshold (Ct) 1 values, indicative of viral load, to determine COVID-19 positivity. This study examined the long-term time series distribution of Ct values measured in the same laboratory using a nationally standardized testing type and sampling method in South Korea. It aimed to link Ct values, new COVID-19 cases, and the reproduction number (Rt), setting the stage for using Ct values effectively. Methods: This study analyzed nationally collected 296,347 samples Ct values from February 2020 to January 2022 and examined their associations with the number of new cases and Rt trends. The data were categorized into four COVID-19 periods for in-depth analysis. Statistical methods included time series trend analysis, local regression for smoothing, linear regression for association analysis, and calculation of correlation coefficients. Results: The median Ct values across four COVID-19 periods decreased gradually from 31.71 in the initial period to 21.27 in the fourth period, indicating higher viral load. The comparison of trends between Ct values and the number of new cases revealed that the decline in Ct values preceded the surge in new cases, particularly evident during the initial stages when new cases did not undergo a significant increase. Also, during variant 1 Ct, cycle threshold; E, envelope protein; LOWESS, locally weighted scatterplot smoothing; N, nucleocapsid; Rt, reproduction number; RdRp, RNA-dependent RNA polymerase; real-time RT-PCR, real-time reverse-transcriptase polymerase chain reaction; S, spike protein emergence and vaccination rollout, marked shifts in Ct values were observed. Results from linear regression analysis revealed a significant negative relationship between Ct values and new cases (β = -0.33, p < 0.001, R 2 = 0.67). This implies that as Ct values decrease, new case numbers increase. Conclusions: This study demonstrates the potential of Ct values as early indicators for predicting confirmed COVID-19 cases during the initial stages of the epidemic and suggests their relevance in large-scale epidemic monitoring, even when case numbers are similar.