Lyme disease is the most common tick-borne disease and is caused by a group of bacteria known asBorrelia burgdorferi sensulato (s.l.) complex. Sharing the same genus asB. burgdorferi,Borrelia miyamotoiis a distinct genotype that causes relapsing fever disease. This emerging tick-borne disease is increasingly becoming a concern in public health. To investigate the prevalence ofB. burgdorferi s.l.andB.miyamotoiin ticks first, we developed a PCR (Bmer-qPCR) that targets the phage terminase large subunit (terL) gene carried byB. miyamotoi. A similar approach had been used successfully in developing Ter-qPCR for detectingB. burgdorferi s.l.The terL protein functions as an enzyme in packaging phage DNA. Analytical validation of the Bmer-qPCR confirmed its specificity, efficiency and sensitivity. Second, we designed a citizen science-based approach to detect 838 ticks collected from numerous sites across Great Britain. Finally, we applied Bmer-qPCR and Ter-qPCR to 153 tick pools and revealed that the prevalence ofB. burgdorferis.l. andB. miyamotoiwas dependent on their geographical locations, i.e. Scotland showed a higher rate ofB. burgdorferis.l. and lower rate ofB. miyamotoicarriage as compared to those of the England data. A pattern of diminishing rate ofB. miyamotoicarriage from southern England to northern Scotland was visible. Together, the citizen science-based approach provided an estimation of the carriage rate ofB. burgdorferi s.l.andB. miyamotoiin tick pools and a potential spreading pattern ofB. miyamotoifrom the south to the north of Great Britain. Our findings underscore the power of combining citizen science with the molecular diagnostic method to reveal hidden pattern of pathogen-host-environment interplay. Our approach can provide a powerful tool to elucidate the ecology of tick-borne diseases and may offer guidance for pathogen control initiatives. In an era of limited resources, monitoring pathogens requires both field and laboratory support. Citizen science approaches provide a method to empower the public for sample collection. Coupling citizen science approaches with laboratory diagnostic tests can make real-time monitoring of pathogen distribution and prevalence possible.