Hemotrophic mycoplasmas (hemoplasmas), Bartonella sp., Hepatozoon sp. and Cytauxzoon felis are prominent pathogens that circulate between cats and invertebrate hosts. The present study aimed to detect the presence of DNA from hemoplasmas, Bartonella sp., Hepatozoon sp. and Cytauxzoon felis, and then confirm it by means of sequencing, in blood samples from cats in Cuiabá, MT, Brazil. From February 2009 to February 2011, blood samples with added EDTA were collected from 163 cats that were being housed in four different animal shelters in the city of Cuiabá, state of Mato Grosso, Brazil and from 15 cats that were admitted to the veterinary hospital of the Federal University of Mato Grosso (UFMT). Out of the 178 cats sampled, 15 (8.4%) were positive for hemoplasmas: four (2.2%) for Mycoplasma haemofelis, 12 (6.7%) for 'Candidatus M. haemominutum' and one (0.5%) for 'Candidatus M. turicensis'. One cat (0.5%), a patient that was attended at the veterinary hospital, was coinfected with M. haemofelis, 'Candidatus M. haemominutum' and 'Candidatus M. turicensis', based on sequencing confirmation. Four cats were positive for Bartonella spp.: three (1.7%) for B. henselae and one (0.5%) for B. clarridgeiae. None of the animals showed Cytauxzoon sp. or Hepatozoon sp. DNA in their blood samples. This study showed that cats housed in animal shelters in the city of Cuiabá, state of Mato Grosso, are exposed to hemoplasmas and Bartonella species.