As the world continues to fight the COVID-19 pandemic, and hope for an end, questions remain regarding when the SARS-CoV-2 virus first entered the United States. Genomic analysis has put some of the early puzzle pieces together. Now, a new study analyzed blood samples, originally collected through the National Institutes of Health’s All of Us research program, for the presence of antibodies to the virus. The analysis found evidence of SARS-CoV-2 infections in five states earlier than had initially been reported. The results expand on findings from a Centers for Disease Control and Prevention study that suggested SARS-CoV-2, the virus that causes COVID-19, was present in the United States as far back as December 2019.

The research is published in the journal Clinical Infectious Diseases, in the paper, “Antibodies to SARS-CoV-2 in All of Us Research Program Participants, January 2–March 18, 2020.”

Researchers analyzed more than 24,000 stored blood samples contributed by All of Us program participants across all 50 states between January 2 and March 18, 2020. Antibodies against SARS-CoV-2 were detected (using two different serology tests) in nine participants’ samples.

These participants were from outside the major urban hotspots of Seattle and New York City, believed to be key points of entry of the virus in the United States.

The positive samples came as early as January 7 from participants in Illinois, Massachusetts, Mississippi, Pennsylvania, and Wisconsin. Most positive samples were collected prior to the first reported cases in those states, demonstrating the importance of expanding testing as quickly as possible in an epidemic setting.

“This study allows us to uncover more information about the beginning of the U.S. epidemic and highlights the real-world value of longitudinal research in understanding dynamics of emerging diseases like COVID-19,” said Josh Denny, MD, CEO of All of Us and an author of the study. “Our participants come from diverse communities across the United States and give generously of themselves to drive a wide range of biomedical discoveries, which are vital for informing public health strategies and preparedness.”

In studies like these, false positives are a concern, particularly when the prevalence of viral infections is low. Researchers in this study followed CDC guidance to use sequential testing on two separate platforms to minimize false positive results.

All of Us worked with Quest Diagnostics to test samples on the Abbott Architect SARS-CoV-2 IgG ELISA and the EUROIMMUNE SARS-CoV-2 ELISA (IgG) platforms. For a sample to be considered “positive” by the research team, it had to have positive results on both platforms, which target antibodies that bind to different parts of the virus. Both tests have emergency use authorization from the FDA. Sensitivity and specificity of the Abbott and EUROIMMUNE ELISAs and the net sensitivity and specificity of the sequential testing algorithm were estimated with 95% confidence intervals.

“Antibody testing of blood samples helps us better understand the spread of SARS-CoV-2 in the United States in the early days of the U.S. epidemic, when testing was restricted and public health officials could not see that the virus had already spread outside of recognized initial points of entry,” said Keri N. Althoff, PhD, lead author and associate professor of epidemiology at the Johns Hopkins Bloomberg School of Public Health. “This study also demonstrates the importance of using multiple serology platforms, as recommended by the CDC.”

Researchers looked for IgG antibodies in the samples, which do not appear until about two weeks after a person has been infected, indicating that participants with these antibodies were exposed to the virus at least several weeks before their sample was taken. In this study, the first positive samples came from participants in Illinois and Massachusetts on January 7 and 8, 2020, respectively, suggesting that the virus was present in those states in late December.

The study authors noted several limitations to their study. While the study included samples from across the United States, the number of samples from many states was low. In addition, the authors do not know whether the participants with positive samples became infected during travel or while in their own communities. Ideally, this study could be replicated in other populations with samples collected in the initial months of the U.S. epidemic and with multiple testing platforms to compare results.

All of Us expects to release more information following further analysis, and will offer participants whose samples were included in the study an opportunity to receive their individual results. The presence of antibodies in one’s blood sample does not guarantee that a person is protected from the infection or that any such protection will last.