Detection of humoral immunity to mycobacteria causing leprosy in Eurasian red squirrels (Sciurus vulgaris) using a quantitative rapid test

Eurasian red squirrels (Sciurus vulgaris, ERS) in the British Isles are a recently discovered natural host for Mycobacterium leprae and Mycobacterium lepromatosis. Infected squirrels can develop skin lesions, or carry the bacteria without showing clinical signs. Until now the clinical diagnosis of leprosy could only be confirmed in squirrels by isolating DNA of leprosy bacilli from carcasses or by establishing the presence of acid-fast bacilli in skin sections of carcasses with clinical signs. In this study, we assessed the performance of a field-friendly diagnostic test for detection of M. leprae/ M. lepromatosis infection in ERS. This up-converting-phosphor lateral flow assay (UCP-LFA) is well established for detection of M. leprae specific anti-phenolic glycolipid-I antibodies (αPGL-I) IgM antibodies in humans and associated with bacterial load. Assesment was performed on serum and blood drops from live squirrels and body cavity fluid samples from dead squirrels. Clinically diseased squirrels showed significantly higher αPGL-I levels than healthy animals or subclinically infected individuals (p<0.0001), both in serum and whole blood drop samples. Subclinically infected animals were identified using molecular methods to detect the presence of leprosy bacilli DNA in punch biopsy tissue samples. In body cavity fluids αPGL-I levels antibody levels were lower than in serum or blood drops. This study shows that the αPGL-I UCP-LFAs presented here allows a field-friendly serological confirmation of M. leprae infection in clinically diseased live ERS. For surveillance purposes, the combination of clinical assessment, αPGL-I UCP-LFAs and molecular methods allow the identification of both diseased animals and subclinically infected animals.