Anti-Leptospira spp antibodies in cart horses of the city of Guaíba, Rio Grande do Sul State, Brazil

The objective of this study was to evaluate the occurrence of antibodies against Leptospira spp in horses used by communities of informal waste collectors who use horse-drawn cartsin Guaíba, Rio Grande do Sul. Additionally, interviews were conducted with owners to assess the characteristics of the horses, their environment, and work. Serological testing for Leptospira spp. was performed on blood samples collected from 54 horses using the microscopic agglutination test. Results showed that 50.0% of the horses were seropositive for Leptospira spp and the most frequent serogroup was LeptospiraCanicola (88. 8%), followed by Leptospira Pyrogenes (11.1%), Leptospira Bataviae (7.4%), Leptospira Hebdomadis (7.4%), Leptospira Icterohaemorrhagiae (3.7%), Leptospira Australis (3.7%), Leptospira Pomona (3.7%) and Leptospira Copenhageni (3.7%). There were no statistically significant associations between seropositivity for Leptospira spp and age, gender, body score, water source, type of food storage, cohabitation with dogs, rodent infestation, residential flooding, and waste removal services. The prevalence of the Canicola serogroup, as well as a large number of dogs in close relationship with thehorses and the waste collectors, draws attention to the potential of these animals to become a source of Leptospira spp. infection in humans and other animals.


SHORT COMMUNICATIONS
Leptospirosis is a worldwide zoonosis of great sanitary and economic importance that affects human beings and domestic animals (ELLIS, 2015). The disease is caused by pathogenic bacteria of the genus Leptospira, which are classified into serogroups and serovars that vary depending on the diversity of environments and hosts (JARA et al., 2019). In urban areas, the proximity between humans and animals in poor housing conditions increases the risk of infection and the spread of bacteria (MENY et al., 2019). Scenarios like these are found in territories occupied by local informal waste collectors who use horses to transport their cargo in wagons. Horses infected with Leptospira may play a role in the transmission of the bacteria to these workers and other animals. In turn, even asymptomatic human carriers can spread the bacteria through their urine for several weeks (YAN et al., 2010).
Having this context as background, this study aimed to evaluate the occurrence of antibodies against Leptospira spp in horses used by communities of informal waste collectors who use horse-drawn carts, in Guaíba, Rio Grande do Sul.
A cross-sectional study was carried out with serological testing of cart horses and interviews with their owners and caretakers. Guaíba is one of the 34 municipalities that are part of the metropolitan region of Porto Alegre. It is located on the right bank of Guaíba Lake, which is an estuary of the Gravataí, Sinos, Caí, Jacuí, and Delta do Jacuí rivers, passes by Lagoa dos Patos lake and ends at the Atlantic Ocean.
We A total of 54 cart horses were evaluated per convenience. None had been vaccinated against leptospirosis. The owners and caretakers of these horses who agreed to participate in the study were interviewed individually to gather information about the characteristics of the animals, their environment, and work. Horses underwent a physical examination to check for eye lesions and assess body condition, rectal temperature, hydration, and the color of the mucous membranes.
Blood samples were collected for serological tests at the first evaluation and approximately 45 to 60 days after the first tests. Trained veterinary physicians undertook all sampling and the Leptospirosis laboratory at the Instituto de Pesquisas Desidério Finamor in Eldorado do Sul was responsible for serological testing.
Blood serum was used to test for anti-Leptospira antibodies through the microscopic agglutination test (MAT; ELLINGHAUSEN; MCCULLOUGH, 1965). Serum samples were analyzed to detect the presence of 18 serovars of Leptospira pathogens with the following antigens: Autumnalis, Australis, Bataviae, Canicola, Castellonis, Celledoni, Copenhageni, Gripp otyphosa,Hardjo,Hebdomadis,Icterohaemorrhagiae,Javanica,Pa nama,Pomona,Pyrogenes,Serjoe,Tarassovi, and Wolff. Animals that presented 50% or more agglutinated Leptospiras Spp in a 1/100 dilution in the microscopic field, in at least one of the serological evaluations, were considered positive for Leptospira spp. To assess possible associations between a positive Leptospira spp result and animal and environmental characteristics, and type of work, we used the Fisher Exact Test (P <0.05).
This study was carried out with the support of the local government organs overseeing health and environmental issues, the Secretarias de Saúde e do Meio Ambiente. Our research was carried out according to the guidelines of the ethics committee of the IPVDF, Comitê de Ética no Uso de Animais (CEUA), and registered under number 12/2016. It was also approved by the ethics committee of the Ritter dos Reis University Center/RS, Comitê de Ética em Pesquisa com Seres Humanos, and registered under the numbers CAAE 91930918.3.0000.5309 and 2.995.019.
Serological examinations revealed that 50% (27/54) of the animals presented positive reactions to one or more serogroups of Leptospira in at least one of the evaluations, with titers ranging from 1:100 to 1:400. Although the frequency of detection of anti-Leptospira spp antibodies was lower than those reported by other studies involving cart horses from other cities in Brazil (HASHIMOTO et al., 2007;LASTA et al., 2013;FINGER et al., 2014;DIAS et al., 2015), it is still concerning. A large portion of the population from the neighborhoods we tested sustain themselves with load transport and the horses are the main source of traction. These animals maintain close and constant contact with their owners and often share the same home and surrounding environment. Our detection of reactive animals indicates contact with the bacteria and also shows that they circulate in environments where humans and other animals are also exposed. Besides this, if they become chronic hosts and continuously eliminate the bacteria in their urine, the horses can contribute to the transmission of the bacteria to humans and other animals (ELLIS, 2015).
In other cart-horse populations, researchers reported a predominance of the Icterohaemorrhagiae serogroup (FINGER et al., 2014;DIAS et al., 2015). Rodents are the main maintenance hosts of this serogroup (ELLIS, 2015;BOEY et al, 2019). In our study, the only horse which tested positive for Icterohaemorrhagiae serogroup was kept in an environment where rodents were reported.
Antibodies against Bataviae, Australis, Pomona and Copenhageni serogroups have also been previously detected in other Brazilian studies involving cart horses (HASHIMOTO et al., 2007;CASELANI et al., 2012;DIAS et al., 2015). Our findings reinforce the importance of rodent control in homes, since infected mice and rats are reservoirs for these serogroups and also possible sources of infection (FAINE et al., 1999;BENACER et al, 2016). The caretakers of these animals reported the presence of rodents in the environment where the animals were kept, which may explain this result.
The Pyrogenes serogroup has been previously detected in cart horses in Brazil, but at a lower frequency than in our study (HASHIMOTO ET AL, 2007;CASELANI et al., 2012;FINGER et al., 2014;DIAS et al., 2015). In other studies, the detection of this serogroup has been related to cross-reactivity with the serogroup Icterohaemorrhagiae (Jorge et al. 2011;Lelu et al. 2015).
Previous reports have described finding the serogroup L. Hebdomadis in cart horses (CASELANI et al., 2012DIAS et al., 2015. This serogroup has been associated with incidental infections in cattle, dogs and humans (ELLIS, 2015;GUERNIER et al., 2018).
Clinical manifestations related to leptospirosis in seropositive animals were rare and unspecific as previously reported elsewhere (LASTA et al., 2013). This also indicates that there were no cases of active infection during the period of our research.
In urban spaces, environments in which there is waste management and accumulation, flooding, rodents, and contact between different animal species increase the likelihood of leptospiral dispersion and incidental infections in humans and domestic animals (Rafizah et al. 2013;BARRAGAN et al., 2017). We found no significant association between and environmental or management characteristics seropositivity for Leptospira (Table 1). Although no significant association has been demonstrated, these aspects cannot be neglected, as they are known risk factors for Leptospirosis and were present in most of the homes visited. The lack of statistical significance may be related to the reduced number of evaluated animals, interview bias or to observations' homogeneity.
The risk of seropositivity for Leptospira spp has been described as higher for mares and gelding males and increases with the age of horses (Langoni et al., 2001;Sequeira et al., 2020). However, our results did not show this relationship and corroborate similar data about cart horses from other regions of Brazil (Pinho et al., 2014;Dias et al., 2015). This is probably due to the lower number of animals we assessed and the homogeneity of the population.
In conclusion, results show that these cart horses were exposed to Leptospira spp. and also indicate that the bacteria are present in the environments where the horses, people, and other animals interact. The predominance of the Canicola serogroup and the expressive proportion of residences with dogs draws attention to the potential of these animals to become a source of Leptospira spp. infection.