Detection of Pathogenic Leptospira in Dogs
World Small Animal Veterinary Association World Congress Proceedings, 2005
L.O. Castillo Sánchez; M.A. Roa Riol; M.L. Ordoñez Badillo; A. de la Peña-Moctezuma
Grupo de Investigación en Leptospira y Leptospirosis, Dept. de Microbiología e Inmunología, Fac

The role of dogs as carriers of pathogenic Leptospira serovars has been probed elsewhere (Adler and De la Peña-Moctezuma, 2004). Uncontrolled dogs in Mexico city streets become an infection source for leptospiral infection. Personnel at Canine Control Centres (CCC) are in risk of acquiring leptospirosis. Previous serological studies have found leptospiral antibodies in dogs at CCC (Rivera et al., 1999).

To confirm the state as carriers, samples of serum, urine and kidney were collected from 58 dogs without apparent clinical signs at the Louis Pasteur CCC in Aragon, Northern Mexico City. Procedures were followed as in Myers (1985). A Plexiglas working chamber of 65 / 40 / 54 cm was designed to optimize aseptic conditions. 10 ml of blood were collected before euthanasia to obtain serum. After shaving and decontaminating skin with 8% iodine solution, 10 ml of urine were collected directly from the bladder with a sterile syringe through an abdominal incision. One kidney was removed and immersed into 200 ml of a 0.25% benzalkonium chloride solution for decontamination. Three urine drops were inoculated immediately in EMJH broth and two decimal dilutions prepared in the same medium. Another drop was put over a slide for dark field microscopy. After 10 minutes, each decontaminated kidney was cut along its outer edge with a sterile scalpel. A kidney's medulla and cortex macerate was obtained with a sterile scraper made from a 4 cm in diameter punctured metallic bottle cap. The macerate was 1:1 diluted with sterile EMJH medium and 2 or 3 drops inoculated in the same medium and diluted as described. A drop of diluted macerate was observed microscopically. Dilutions were transported to the laboratory and incubated for up to 5 months at 30 C and microscopic observations performed weekly. The microscopic agglutination test (MAT) was performed on 1:50 to 1:6400 diluted sera with each of 12 Leptospira serovars.

36 serum samples (62%) were positive in the MAT with titres from 1:100 up to 1:6400 in at least one serovar. The most common detected serovars were Canicola (23, 39.6%), Bratislava (20, 34.4%), Pyrogenes (19, 32.7%) and Grippotyphosa (11, 18.9%). Serovar Icterohaemorrhagiae was next with 7 positive sera (12%) and then Pomona with 3 positive sera (5.17%). Dark field microscopy from urine samples reported 17 positive samples (29.3%) and 19 positive kidney samples (32.7%). Two leptospira isolates were obtained. One from kidney of dog 46 after 6 weeks and one from urine of dog 28 after 16 weeks of incubation. Dog 46 showed a titre of 1:6400 against serovar Canicola and dog 28 a titre of 1:6400 against Pyrogenes. Noteworthy, samples from both dogs were negative under dark field microscopy. Another 8 dogs showed antibody titres of 1:3200 to 1:6400 against one of more Leptospira serovars but no isolate was obtained from them. Only 2 out of 10 dogs (20%) showing titres higher than 1:3200 were positive in dark field microscopy. Virulence of both isolates was demonstrated by intraperitoneal inoculation in hamsters. Strain 46 was more virulent killing hamsters after 5 days in average and clearer pneumonic gross lesions than strain 28 that killed hamsters after 9 days and showed more discrete lung lesions.

High antibody titres against Leptospira correlate with positive urine of kidney cultures. Dark field microscopy showed no correlation with isolation and a low correlation with titres 1:3200 or higher in MAT. Infected dogs are an important source of pathogenic Leptospira serovars.


1.  Adler B, De la Peña-Moctezuma A. Leptospirosis In: Gyles CL, Thoen CO, Prescott JF: Pathogenesis of Bacterial Infections in Animals, 3rd ed., 2004.

2.  Rivera FA, Roa RMA, Ordoñez BML, De la Peña-Moctezuma A. Seroprevalencia de leptospirosis en perros callejeros del norte de la Ciudad de México. Vet Mex. 1999;30:105-108.

3.  Myers DM. Leptospirosis. Nota Técnica no. 30. Organización Panamericana de la Salud, Oficina Sanitaria Panamericana, Oficina Regional de la Organización Mundial de la Salud. Martínez, Argentina, 1985.

4.  Ultad de Medicina Veterinaria y Zootecnia, Universidad Nacional Autónoma de México

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L.O. Castillo Sánchez

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