Frequency of canine leptospirosis in dog shelters in Veracruz , Mexico

A cross-sectional study was conducted to determine the frequency of canine leptospirosis in 92 dogs from two shelters in the city of Veracruz, Mexico. The microaglutination test (MAT) was used for detecting antibodies against 12 serovars of Leptospira interrogans in dog serum. Of the 92 dogs, eight were positive, resulting in an overall frequency of leptospirosis of 8.6% (CI95% 3.8 to 16.41). With reference to shelter, frequency was 8.3% (4/48; CI95% 2.3 to 19.9) in shelter 1 and 9.0% (4/44; CI95% 2.5 to 21.6) in shelter 2. The most frequent serovar of L. interrogans was Canicola. Frequency by sex of the dogs was 8.8% (4/45; CI95% 2.4 to 21.2) for females and 8.5% (4/47; CI95% 2.3 to 20.3) for males. In conclusion, leptospirosis is present in dogs housed in shelters in the city of Veracruz, Mexico.


INTRODUCTION
Leptospirosis is a zoonotic infectious disease of worldwide distribution that is endemic in tropical and temperate climates, with higher prevalence in tropical countries (Levett, 2001;Adler and de la Peña, 2010;Desvars et al., 2011).Leptospirosis affects humans, domestic and wild animals (Båverud et al., 2009), and the reservoirs are wild or domestic animals such as rodents, cattle or dogs (Kikuti et al., 2012).In urban areas, rodents (mostly rats) are the main carriers of the disease (Oliveira et al., 2012), whereas the dog is considered a dead-end host (Prescott, 2008;Reis et al., 2008).Nonetheless, due to their close contact, dogs pose a risk of infection for humans (Greene, 2006;Adler and de la Peña, 2010).
Leptospirosis is caused by several pathogenic serovars within the species Leptospira interrogans (Båverud et al., 2009;Adler and de la Peña, 2010).Rodents, particularly rats (Rattus norvergicus), are considered natural reservoirs of serovar Icterohaemorrhagiae, and domestic dogs are natural reservoirs of serovar Canicola (André-Fontaine, 2006).However, canine leptospirosis can be caused by other serovars such as Grippotyphosa, Pomona, Bratislava (Birnbaum et al., 1998;Ward et al., 2004;Greene, 2006;Ghneim et al., 2007), Australis, Autumnalis, Ballum, Bataviae and Hardjo (Ward, 2002), which have, among others, mice, pigs and cattle as reservoirs (Bolin, 1996;Birnbaum et al., 1998;Adin and Cowgill, 2000).In dogs, infection usually results from direct contact with urine from an infected animal, or indirectly from contaminated water or moist soil, where the bacteria can survive for several months (Adler and de la Peña, 2010;Raghavan et al., 2011).Acute leptospirosis is characterized by hepatic and mainly renal failure, and common manifestations include anorexia, vomiting, lethargy, jaundice, diarrhoea and bloody urine, and if not treated quickly it can lead to death (André-Fontaine, 2006;Minke et al., 2009).Although serovars Canicola and Icterohaemorrhagiae are usually responsible for acute leptospirosis, serovars Australis, Pyrogenes, Autumnalis and Grippotyphosa have also been found in this form of the disease (Adamus et al., 1997;André-Fontaine, 2006).Chronic *Corresponding author.E-mail: dromero@uv.mx.Tel/Fax: +52 (229) 9342075 or 9344053.leptospirosis is more difficult to diagnose because vaccinated animals can show signs of the disease (Adamus et al., 1997).A vaccinated dog may get infected when exposed to a highly infectious environment or a highly virulent strain (Andre-Fontaine et al., 2003;Klaasen et al., 2003), which could be the case of a dog shelter.Subclinical forms are more common and usually occur in chronically infected animals, which can be carriers for years or for life (OIE, 2004).Clinically recovered dogs can turn into asymptomatic renal carriers and become an important source of infection for humans (CDC, 1972;Trevejo et al., 1998).Seroprevalence of canine leptospirosis has been reported from 1.9% in shelter dogs to 35% in serological surveys (Meeyam et al., 2006;Davis et al., 2008;Zwijnenberg et al., 2008).There are few studies on canine leptospirosis in Mexico, and seroprevalence reported ranged from 4.9 to 35% (Ortega-Pacheco et al., 2008;Jimenez-Coello et al., 2008;2010).In Mexico, vaccination of pet dogs against leptospirosis is practiced by responsible owners, but in the case of shelters, most of the dogs that are taken in at these places are stray, and others have been neglected by their owners, so it is unlikely that these dogs have been vaccinated against leptospirosis.If these dogs are carriers of the disease, they may be a source of infection for their care givers or adopters.Therefore, the aim of this study was to determine the frequency of anti-Leptospira spp.antibodies in dogs from two shelters in the city of Veracruz, Mexico.

MATERIALS AND METHODS
A cross-sectional study was conducted in two dog shelters from the city of Veracruz, Mexico (19°12' North latitude and 96°07' West longitude).Climate is tropical and mean annual rainfall is 1500 mm.These shelters are the only ones in the city and take in stray or unwanted dogs.A total of 92 adult dogs (shelter 1: n = 48; shelter 2: n = 44) were included in the study.The vaccination status of these dogs was unknown because most of them had been picked up as strays, and vaccination is not usually practiced in the shelters due to economic restraints.

Blood samples collection
One blood sample was collected via cephalic or jugular venipuncture from each of the 92 dogs to detect antibodies against Leptospira interrogans.Samples were kept in refrigeration and transported to the laboratory, where they were centrifuged at 3000 rpm for 10 min to separate the serum, which was kept frozen at -20°C until analyzed.

Data analysis
The frequency of seropositive animals was calculated as a percentage of the total number of samples tested.Seropositive animals were examined in relation to sex and shelter.The chi-square test was used to compare both shelters.The STATA® software was used to evaluate the frequency and to calculate the confidence intervals.
According to the sex of the dogs, no difference was found in the present study (Table 2).In Australia, Zwijnenberg et al. (2008) indicate that the female dogs are risk factor associated with the seropositive status, and Miller et al. (2007) mentioned that young and male dogs are more commonly affected by leptospirosis.
In conclusion, leptospirosis was present in the canine population from the two shelters surveyed.Thus, if these dogs or those that tested negative were carriers of the disease, they might spread it to the rest of the population, causing a serious health problem, or they might also transmit it to their human care givers or adopters.Thus, it is important to determine the presence of leptospirosis in dogs from shelters to take preventive sanitary measures to avoid its transmission to the rest of the pack or to the people that are in contact with them.

Table 1 .
Frequency of canine leptospirosis in two shelters from Veracruz, Mexico.

Table 2 .
Frequency of canine leptospirosis with regard to sex in two shelters from Veracruz, Mexico.

Table 3 .
Frequency of serovars of Leptospira spp. in two dog shelters from Veracruz, Mexico.