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Borreliose de Lyme em cães e cavalos: caracterização serológica e molecular em animais assintomáticos
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A Borreliose de Lyme (BL), causada por espiroquetas do complexo Borrelia burgdorferi sensu lato (B. burgdorferi s.l.) é, atualmente, a doença mais prevalente da Europa cujo o agente é transmitido por vetores Ixodídeos. É uma doença zoonótica, que afeta diversos hospedeiros vertebrados, entre os quais diversos animais domésticos e silváticos, bem como os humanos.
As espiroquetas responsáveis pela BL encontram-se, normalmente, associadas a pequenos mamíferos, aves e répteis que, enquanto hospedeiros reservatórios, possuem um papel essencial na transmissão e manutenção destes agentes na natureza. Por outro lado, os hospedeiros como o cão, cavalo e Homem têm um papel menos significativo na manutenção e transmissão do agente, sendo considerados hospedeiros acidentais.
A infeção por B. burgdorferi s.l. nos animais é geralmente assintomática, contudo, quando surgem manifestações clínicas, tais como febre e claudicação, estas são comuns a outras doenças, o que dificulta o diagnóstico clínico. Em muitos países, incluindo Portugal, esta doença é subdiagnosticada e pouco reportada, sendo por isso importante recorrer-se a um diagnóstico laboratorial.
O objetivo deste estudo foi caracterizar, por técnicas serológicas e moleculares, um total de 240 amostras séricas de duas populações de animais domésticos assintomáticos para a infeção por B. burgdorferi s.l.. Foram analisadas 114 amostras de cães e 126 amostras de cavalos, originárias de várias regiões de Portugal, caracterizadas em estudos anteriores para a presença de outros agentes transmitidos por vetores, particularmente carraças. A presença de anticorpos IgG anti-B. burgdorferi s.l. foi detetada pelas técnicas de Imunofluorescência Indireta (IFA) e Western Blot (WB), utilizadas como teste de rastreio e teste confirmatório, respetivamente. A presença de DNA borreliano foi pesquisada pela técnica de nested-PCR por amplificação da região intergénica 5S-23S e amplificação parcial do gene flaB. Adicionalmente, foram também submetidas a nested-PCR para amplificação parcial de gene ospC, todas as amostras anteriormente positivas para qualquer um dos alvos genómicos testados.
A ocorrência de amostras com serologia positiva, ou seja, com título superior ou igual a 1:256, foi observada em 29 amostras de cães e em 31 amostras de cavalos, correspondendo a frequências de, aproximadamente, 25% para cada população. Dos 60 soros com imunofluorescência positiva foi confirmada, pela técnica de WB, uma amostra pertencente a um cavalo da região de Lisboa. A presença de DNA de B. burgdorferi s.l. foi detetada, pelas técnicas moleculares, em 7% das amostras de cães e 5,6% das amostras de cavalos. Várias genoespécies foram identificadas, designadamente B. afzelii, em amostras de cães e cavalos, B. burgdorferi s.s., B. valaisiana e B. lusitaniae, apenas em amostras de cavalos. Os distritos de Beja e Lisboa apresentaram o maior número de amostras positivas. Globalmente em 3,5% dos cães e 3,2% dos cavalos foram detetados anticorpos anti-B. burgdorferi s.l. e identificado DNA de B. burgdorferi s.l..
Este estudo evidenciou a exposição ao vetor e o contacto com B. burgdorferi s.l. a que os animais em estudo estão sujeitos, sugerindo a existência de animais assintomaticamente infetados. Os resultados indicam que existem em circulação várias genoespécies em Portugal. No seu conjunto, os resultados obtidos são importantes pois alertam para o risco de BL nos animais e, uma vez que animais domésticos partilham habitats com os humanos, representam um potencial risco de Saúde Pública.
Lyme Borreliosis (LB), caused by spirochetes of Borrelia burgdoferi sensu lato complex (B. burgdorferi s.l.), is presently the most prevalent tick-borne disease in Europe. It is a zoonotic disease that affects several vertebrate hosts, including several domestic and sylvatic animals, as well as humans. The spirochetes responsible for LB are usually associated with small mammals, birds and reptiles, as reservoir hosts, playing an essential role in the transmission and maintenance of these agents in nature. On the other hand, hosts such dog, horse and man are considered “dead-end” hosts and play an insignificant role in the transmission of the agent. Borrelia burgdorferi s.l.-infected animals are usually asymptomatic, however, clinical manifestations such as fever and lameness may appear but they are common to other pathologies, making clinical diagnosis difficult. In many countries, including Portugal, this disease is underdiagnosed and poorly reported, and therefore it is also important to use the laboratory diagnosis. The objective of this study was to characterize by serological and molecular techniques a total of 240 serum samples from two populations of asymptomatic domestic animals for infection for B. burgdorferi s.l.. A total of 114 dog samples and 126 horse samples from several regions of Portugal, which have previously been characterized for the presence of other tick-borne agents, was analysed. The presence of anti- B. burgdorferi s.l. IgG antibodies was detected by both Indirect Immunofluorescence Assay (IFA) and Western Blot (WB) techniques, used as screening and confirmatory tests, respectively. The presence of borrelial DNA was investigated by using nested-PCR targeting the 5S-23S rRNA intergenic spacer region and flaB gene. In addition, positive samples for any of these genomic targets were also submitted to nested-PCR for partial amplification of ospC gene. The occurrence of serologically positive samples, with antibody titre greater than or equal to 1:256, was observed in 29 samples of dogs and 31 samples of horses, corresponding to frequencies of approximately 25% for each population. From the 60 sera with positive immunoreaction, a sample belonging to a horse from Lisbon region was confirmed by the WB technique. The presence of B. burgdorferi s.l. DNA was detected by molecular techniques in 7% of the dogs and 5.6% of the horses. Several genospecies were identified namely B. afzelii, in both dog and horse populations, B. burgdorferi s.s., B. valaisiana and B. lusitaniae, only in the horse population. The regions of Beja and Lisbon showed the highest number of positive samples. Globally, 3.5% of the dogs and 3.2% of the horses showed both antibodies anti- B. burgdorferi s.l. and DNA of B. burgdorferi s.l. This study demonstrated that the populations of dogs and horses have contacted with B. burgdorferi s.l., and suggesting the presence of healthy animals infected with B. burgdorferi s.l.. It also indicating the presence are several genospecies in circulation in Portugal. The results obtained should alert to the risk of LB in domestic animals and also to humans, because they share the same habitats, representing a potential risk to both animal and public health.
Lyme Borreliosis (LB), caused by spirochetes of Borrelia burgdoferi sensu lato complex (B. burgdorferi s.l.), is presently the most prevalent tick-borne disease in Europe. It is a zoonotic disease that affects several vertebrate hosts, including several domestic and sylvatic animals, as well as humans. The spirochetes responsible for LB are usually associated with small mammals, birds and reptiles, as reservoir hosts, playing an essential role in the transmission and maintenance of these agents in nature. On the other hand, hosts such dog, horse and man are considered “dead-end” hosts and play an insignificant role in the transmission of the agent. Borrelia burgdorferi s.l.-infected animals are usually asymptomatic, however, clinical manifestations such as fever and lameness may appear but they are common to other pathologies, making clinical diagnosis difficult. In many countries, including Portugal, this disease is underdiagnosed and poorly reported, and therefore it is also important to use the laboratory diagnosis. The objective of this study was to characterize by serological and molecular techniques a total of 240 serum samples from two populations of asymptomatic domestic animals for infection for B. burgdorferi s.l.. A total of 114 dog samples and 126 horse samples from several regions of Portugal, which have previously been characterized for the presence of other tick-borne agents, was analysed. The presence of anti- B. burgdorferi s.l. IgG antibodies was detected by both Indirect Immunofluorescence Assay (IFA) and Western Blot (WB) techniques, used as screening and confirmatory tests, respectively. The presence of borrelial DNA was investigated by using nested-PCR targeting the 5S-23S rRNA intergenic spacer region and flaB gene. In addition, positive samples for any of these genomic targets were also submitted to nested-PCR for partial amplification of ospC gene. The occurrence of serologically positive samples, with antibody titre greater than or equal to 1:256, was observed in 29 samples of dogs and 31 samples of horses, corresponding to frequencies of approximately 25% for each population. From the 60 sera with positive immunoreaction, a sample belonging to a horse from Lisbon region was confirmed by the WB technique. The presence of B. burgdorferi s.l. DNA was detected by molecular techniques in 7% of the dogs and 5.6% of the horses. Several genospecies were identified namely B. afzelii, in both dog and horse populations, B. burgdorferi s.s., B. valaisiana and B. lusitaniae, only in the horse population. The regions of Beja and Lisbon showed the highest number of positive samples. Globally, 3.5% of the dogs and 3.2% of the horses showed both antibodies anti- B. burgdorferi s.l. and DNA of B. burgdorferi s.l. This study demonstrated that the populations of dogs and horses have contacted with B. burgdorferi s.l., and suggesting the presence of healthy animals infected with B. burgdorferi s.l.. It also indicating the presence are several genospecies in circulation in Portugal. The results obtained should alert to the risk of LB in domestic animals and also to humans, because they share the same habitats, representing a potential risk to both animal and public health.
Descrição
Palavras-chave
Ciências biomédicas Biologia molecular Doenças tropicais Borrelia burgdorferi Borreliose de Lyme Cães Cavalos Técnicas serológicas Técnicas moleculares
Contexto Educativo
Citação
Editora
Instituto de Higiene e Medicina Tropical