ABSTRACT Introduction: Hard ticks are hematophagous ectoparasites, vectors of emerging zoonotic pathogens. Until 2011, there was evidence of infection/exposure to Borrelia burgdorferi in humans in Cuba, but the circulation of other agents is unknown. Aim: implement new diagnostic tools, explore infection/exposure of humans to tick-borne pathogens, their presence in vectors and level of knowledge in medical personnel. Methods: Multiplex PCR to detect B. burgdorferi-Anaplasma spp.-Babesia spp., PCR for Borrelia spp., and artificial infection of ticks were implemented. Positivity to infection by B. burgdorferi (2011-2018) was described. Presence of antibodies against borrelias in blood donors from a low-exposure region, antibodies against other zoonotic agents in a population at risk, presence of these agents in ticks, as well as the level of knowledge about Lyme disease in doctors were explored. Results: IPK capabilities for molecular detection of tick-borne pathogens were strengthened. New serological evidence of B. burgdorferi infection was found in patients with suspicion of Lyme disease. Absence of specific antibodies against B. burgdorferi was demonstrated in blood donors. Evidence of antibodies to Anaplasma phagocytophilum, Ehrlichia chaffeensis and Babesia microti were found. The presence of Anaplasma spp., Babesia spp., Rickettsia amblyommii and Coxiella burnetii was demonstrated in ticks. Insufficient knowledge about Lyme disease was found in medical personnel. Conclusions: these results alert the national public health authorities and medical personnel about the risk of infection in individuals exposed to tick bites.

RESUMEN Introducción: Las garrapatas duras son ectoparásitos hematófagos, vectores de patógenos zoonóticos emergentes. Hasta 2011 en Cuba existían evidencias de infección/exposición a Borrelia burgdorferi en seres humanos, pero se desconoce sobre la circulación de otros agentes. Objetivo: implementar nuevas herramientas diagnósticas, explorar infección/exposición de seres humanos a patógenos transmitidos por garrapatas, presencia de estos en vectores y nivel de conocimientos en personal médico. Métodos: Se implementó PCR múltiple para detección de B. burgdorferi sl-Anaplasma spp. -Babesia spp., PCR para Borrelia spp., e infección artificial de garrapatas. Se describió la positividad a infección por B. burgdorferi (2011-2018), se exploró la presencia de anticuerpos contra borrelias en donantes de sangre de una región de baja exposición, anticuerpos contra otros agentes zoonóticos en población de riesgo, presencia de estos agentes en garrapatas, así como el nivel de conocimientos sobre enfermedad de Lyme en médicos. Resultados: Se fortalecieron capacidades del IPK para detección molecular de patógenos transmitidos por garrapatas. Se encontraron nuevas evidencias serológicas de infección por B. burgdorferi en pacientes con sospechas de enfermedad de Lyme. Se demostró ausencia de anticuerpos específicos contra B. burgdorferi en donantes de sangre. Se encontraron evidencias de anticuerpos contra Anaplasma phagocytophilum, Ehrlichia chaffeensis y Babesia microti. Se demostró presencia de Anaplasma spp., Babesia spp., Rickettsia amblyommii y Coxiella burnetii en garrapatas. Se constataron conocimientos insuficientes sobre enfermedad de Lyme en personal médico. Conclusiones: estos resultados alertan a las autoridades nacionales de salud pública y personal médico sobre el riesgo de infección en individuos expuestos a picaduras de garrapatas.

Four variants of the potassium acetate procedure for DNA extraction from ixodid ticks at different stage of their life cycles were evaluated and compared with phenol-chloroform and ammonium hydroxide methods. The most rapid and most efficient variant was validated in the DNA extraction procedure from the engorged ticks collected from bovine, canine as well as from house ticks for the screening of Borrelia burgdorferi sensu lato, Anaplasma spp. and Babesia spp. The ammonium hydroxide procedure was used for non-engorged ticks. All the variants were efficient and allowed obtaining PCR-quality material according to the specific amplification of 16S rRNA gene fragment of the original tick. DNA extracted from the ticks under the study was tested by multiplex PCR for the screening of tick-borne pathogens. Anaplasma spp. and Babesia spp. amplification products were obtained from 29/48 extracts. Ammonium hydroxide protocol was not efficient for two extracts. Detection of amplification products from the PCR indicated that DNA had been successfully extracted. The potassium acetate procedure could be an alternative, rapid, and reliable method for DNA extraction from the ixodid ticks, mainly for poorly-resourced laboratories.