Objectives: The objective of this study was to develop a quantitative 5' nuclease real-time polymerase chain reaction (PCR) assay to diagnose infections caused by Bartonella species. Methods: Between January and April 2013 whole blood samples were collected by convenience from 151 cats (86 domiciled and 65 stray cats). The feline blood samples were subjected to a novel quantitative 5' nuclease real-time PCR (qPCR) for Bartonella species targeting the nictonamide adenine dinucleotide dehydrogenase gamma subunit ( nuoG ) and conventional PCR assays targeting intergenic transcribed spacer, ribC, gltA, pap31 and rpoB , followed by sequencing and basic local alignment search tool analysis. Results: The qPCR assay detected as few as 10 copies of plasmid per reaction. Forty-six (54.4% domiciled and 45.6% stray cats) of 151 sampled cats showed positive results in nuoG qPCR for Bartonella species. The absolute quantification of nuoG Bartonella DNA in sampled cats ranged from 1.1 × 10 ⁴ to 1.3 × 10 ⁴ . Eighteen (39.1%) of 46 positive samples in the qPCR were also positive in conventional PCR assays. The sequencing confirmed that Bartonella henselae and Bartonella clarridgeiae circulate in cats in midwestern Brazil. Conclusions and relevance: The present work provides details of a novel qPCR assay to diagnose infections caused by Bartonella species.

Bartonellaspp. comprise an ecologically successful group of microorganisms that infect erythrocytes and have adapted to different hosts, which include a wide range of mammals, besides humans. Rodents are reservoirs of about two-thirds ofBartonellaspp. described to date; and some of them have been implicated as causative agents of human diseases. In our study, we performed molecular and phylogenetic analyses ofBartonellaspp. infecting wild rodents from five different Brazilian biomes. In order to characterize the genetic diversity ofBartonellaspp., we performed a robust analysis based on three target genes, followed by sequencing, Bayesian inference, and maximum likelihood analysis.Bartonellaspp. were detected in 25.6% (117/457) of rodent spleen samples analyzed, and this occurrence varied among different biomes. The diversity analysis ofgltAsequences showed the presence of 15 different haplotypes. Analysis of the phylogenetic relationship ofgltAsequences performed by Bayesian inference and maximum likelihood showed that theBartonellaspecies detected in rodents from Brazil was closely related to the phylogenetic group A detected in other cricetid rodents from North America, probably constituting only one species. Last, theBartonellaspecies genogroup identified in the present study formed a monophyletic group that includedBartonellasamples from seven different rodent species distributed in three distinct biomes. In conclusion, our study showed that the occurrence ofBartonellabacteria in rodents is much more frequent and widespread than previously recognized.IMPORTANCEIn the present study, we reported the occurrence ofBartonellaspp. in some sites in Brazil. The identification and understanding of the distribution of this important group of bacteria may allow the Brazilian authorities to recognize potential regions with the risk of transmission of these pathogens among wild and domestic animals and humans. In addition, our study accessed important gaps in the biology of this group of bacteria in Brazil, such as its low host specificity, high genetic diversity, and relationship with otherBartonellaspp. detected in rodents trapped in America. Considering the diversity of newly discoveredBartonellaspecies and the great ecological plasticity of these bacteria, new studies with the aim of revealing the biological aspects unknown until now are needed and must be performed around the world. In this context, the impact ofBartonellaspp. associated with rodents in human health should be assessed in future studies.

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.