Navigating cross-reactivity and host species effects in a serological assay: A case study of the microscopic agglutination test for Leptospira serology

BackgroundSerology (the detection of antibodies formed by the host against an infecting pathogen) is frequently used to assess current infections and past exposure to specific pathogens. However, the presence of cross-reactivity among host antibodies in serological data makes it challenging to interpret the patterns and draw reliable conclusions about the infecting pathogen or strain. Methodology/Principal FindingsIn our study, we use microscopic agglutination test (MAT) serological data from three host species [California sea lion (Zalophus californianus), island fox (Urocyon littoralis), and island spotted skunk (Spilogale gracilis)] with confirmed infections to assess differences in cross-reactivity by host species and diagnostic laboratory. All host species are known to be infected with the same serovar of Leptospira interrogans. We find that absolute and relative antibody titer magnitudes vary systematically across host species and diagnostic laboratories. Despite being infected by the same Leptospira serovar, three host species exhibit different cross-reactivity profiles to a 5-serovar diagnostic panel. We also observe that the cross-reactive antibody titer against a non-infecting serovar can remain detectable after the antibody titer against the infecting serovar declines below detectable levels. Conclusions/SignificanceCross-reactivity in serological data makes interpretation difficult and can lead to common pitfalls. Our results show that the highest antibody titer is not a reliable indicator of infecting serovar and highlight an intriguing role of host species in shaping reactivity patterns. On the other side, seronegativity against a given serovar does not rule out that serovar as the cause of infection. We show that titer magnitudes can be influenced by both host species and diagnostic laboratory, indicating that efforts to interpret absolute titer levels (e.g., as indicators of recent infection) must be calibrated to the system under study. Thus, we implore scientists and health officials using serological data for surveillance to interpret the data with caution. AUTHOR SUMMARYSerology is frequently used for disease surveillance, especially in systems that are resource constrained or logistically challenging. Serological testing involves analyzing blood serum samples to detect antibodies with reactivity toward specific pathogens (or more generally, molecular antigens), with the goal of characterizing past exposure to those pathogens. However, these antibodies can be non-specific and may react against other related pathogens or strains - a phenomenon known as cross-reactivity. Interpretation of serological data exhibiting cross-reactivity is difficult and simplifying assumptions are often made (e.g., to interpret the strain that elicits the highest antibody titer level as the infecting pathogen strain). Our work shows that interpreting antibody data requires more nuance and more caution. Both absolute titer levels and relative reactivity against different strains can vary across host species and diagnostic laboratories, so it is essential to interpret these data in the appropriate context. These host species differences in antibody reactivity and cross-reactivity patterns make direct comparisons across species inadvisable.