L everaging on the Cross-Reactivity and recovery of leptospira serovars in stored cultures to mitigate limitations of the Microscopic Agglutination Test

Abstract

Leptospirosis is one of the most widely spread bacterial zoonotic that is endemic in subtropical and tropical countries; accounting for a global annual incidence of 1.03 million human cases and 58,900 deaths. (Alinaitwe et al., 2020). The etiological agents of the disease are spirochetes of the genus Leptospira, comprising over 250 pathogenic serovars(Alinaitwe et al., 2020) Certain serovars are known to be regionally endemic and reserved in certain species of wild mammals and domesticated animals. Cross reactivity was noted in three pairs of serovars when using commercial anti-Leptospira antibodies. The cross-reactive serovar pairs were Hardjo and Sejroe, Icterohaemorrhagiae and Copenhageni, Bratslava and Australis. The same pairs of serovars were cross-reactive (up to 100%) when tested against highly seropositive sera samples (anti-Leptospira antibody titres ≥800 from cattle and dogs. However, cross-reactivity between serovars Icterohaemorrhagiae and Copenhageni was weaker when tested against serum samples (only 40% reactivity). Of the 300 random serum samples tested, matching results between both panels were registered in 293 samples (97.67%). Overall, there was a very strong agreement between the MAT results of the standard MAT panel and the devised shorter one (Cohen's kappa statistic 0.947±0.058; 95% CI 0.909-0.986). The sensitivity and specificity of the shorter panel were 96.97% (95% CI 0.914-0.994) and 98.01% (95% CI 0.950-0.995), while the positive and negative predictive values were 96.0% and 98.5%, respectively. For each serovar, there is less than 50% agreement between the frozen and live cultures, rendering the data from frozen serovars unreliable. The data provided in this study confirms the occurrence of cross-reactivity among Leptospira serovars in the same serogroup in both commercial and non-commercial anti-sera. The shorter panel tested in this study is highly sensitive and specific and compares well with the standard MAT panel, and thus could be used as an alternative panel for diagnosis of leptospirosis in dogs and cattle in Uganda. For each serovar, there is less than 50% agreement between the frozen and live cultures, rendering the data from frozen serovars unreliable.