Veterinary Microbiology, 15 (1987) 71-78 71 Elsevier Science Publishers B.V., Amsterdam -- Printed in The Netherlands Detection of Leptospires in Biological Fluids using DNA Hybridisation B.D. MILLAR Department of Agriculture and Rural Affairs, Bendigo Regional Veterinary Laboratory, Bendigo, Vic. 3550 (Australia) R.J. CHAPPEL Department of Agriculture and Rural Affairs, Attwood Veterinary Research Laboratory, Attwood, Vic. 3047 (Australia) B. ADLER Department of Microbiology, Monash University, Clayton, Vic. 3168 (Australia) ( Accepted for publication 22 January 1987 ) ABSTRACT Millar, B.D., Chappel, R.J. and Adler, B., 1987. Detection of leptospires in biological fluids using DNA hybridisation. Vet. Microbiol., 15: 71-78. DNA extracted from Leptospira interrogans serovar pomona was labelled with phosphorus-32 by nick translation and used as a genomic probe to detect leptospiral DNA. The sensitivity of detection in a 10-/zl spot on nylon membranes was 160 pg of leptospiral DNA or 1.1 X 103 leptos- pires and assays with nylon membranes were somewhat more sensitive than assays with nitrocel- lulose membranes. The probe reacted with the pathogenic hardjo and tarassovi leptospiral serovars, but not with other genera of bacteria. To detect leptospires in body fluids, these were treated to free leptospiral DNA and then concentrated on membranes using a Bio-Dot TM apparatus. Neither serum nor urine interfered with the assay system. The DNA of leptospires added to pig urine was stable for at least 2 h at room temperature and for at least 20 h at -20°C. INTRODUCTION Infection with serovars of Leptospira interrogans is widespread in man and domestic animals and is associated with a range of clinical syndromes. Path- ogenic leptospires colonise the kidney and may be shed in urine for prolonged periods (Faine, 1982 ). Asymptomatic carrier animals are a frequent source of infection and are often difficult to identify. Diagnosis of leptospirosis depends on the demonstration of either serum antibodies or leptospires in blood or urine. However, the presence of serum antibody bears little relationship to current infection or renal shedding ( Burn- 0378-1145/87/$03.50 © 1987 Elsevier Science Publishers B.V.