Paracoccidioidomycosis (PCM) is a severe disease caused by the dimorphic fungus Paracoccidioides brasiliensis, which is characterized by granulomatous pulmonary and systemic lesions, affecting mainly men between 20 and 60 years of age. Reports of PCM disease in animals are rare, but the disease has been described in armadillos. On the other hand, PCM infection of domestic and wild animals detected by serological or cutaneous tests in the absence of apparent disease has been frequently reported. We present here the case of a female adult Doberman that developed cervical lymphadenomegaly. Histopathological examination of a cervical biopsy specimen revealed active PCM, with an epithelioid, granulomatous inflammation containing numerous yeast-like, multiple budding fungal forms. The diagnosis of PCM was confirmed by immunohistochemistry using a specific antibody anti-gp43 and by nested PCR using primers for the amplification of the gp43 gene region. This is the first report of PCM disease occurring in a dog, an animal that has been shown to play an important role in the natural history of North American blastomycosis.

Paracoccidioidomycosis is a deep mycosis caused by the thermo-dependent dimorphic fungus Paracoccidioides brasiliensis and is prevalent in Latin American countries. We detected the species specific gp43 gene of P. brasiliensis by loop-mediated isothermal amplification (LAMP) in 22 clinical and seven armadillo-derived isolates. The amplified DNA appeared as a ladder with a specific banding pattern. The advantage of the LAMP method is speed; only 3 h were necessary for identification of the organism and diagnosis of the disease. We were also able to obtain positive results from DNA extracted from a paraffin-embedded tissue sample of paracoccidioidomycosis, suggesting that this method may achieve clinical application in the near future.

Paracoccidioidomycosis (PCM) is a life-threatening systemic fungal infection caused by members of the Paracoccidioides brasiliensis complex and P. lutzii. Routine diagnoses of PCM down to the species level using classical mycological approaches are unspecific due to overlapping phenotypes. There is an urgent need for specific, sensitive, and cost-effective molecular tools to diagnose PCM. Variation among the exon-2 of the gp43 gene was exploited to design species-specific primer pairs to discriminate between members of the P. brasiliensis complex and P. lutzii in a duplex PCR assay. Primer-BLAST searches revealed highly species-specific primers, and no significant region of homology was found against DNA databases except for Paracoccidioides species. Primers PbraCx-F and PbraCx-R targeting P. brasiliensis DNA produced an amplicon of 308 bp, while primers Plu-F and Plu-R targeting P. lutzii DNA generated an amplicon of 142 bp. The lower limit of detection for our duplex PCR assay was 1 pg of gDNA. A panel of 62 Paracoccidioides revealed 100% specificity (AUC = 1.000, 95%CI 0.972–1.000, p < 0.0001) without cross-reacting with other medically relevant fungi or human DNA. As a proof of concept, we demonstrated the accurate identification of the P. brasiliensis complex (n = 7) or P. lutzii (n = 6) from a broad range of formalin-fixed, paraffin-embedded (FFPE) tissues of PCM patient’s organs. In four cases, FFPE PCR results confirmed, for the first time, co-infection due to P. brasiliensis (S1) and P. lutzii in the same biopsy. Our duplex PCR assay is useful to detect and differentiate members of the P. brasiliensis complex and P. lutzii, providing clinical laboratories with an important tool to be applied routinely, especially in atypical cases such as those featuring negative serology and positive mycological examination of clinical specimens as well as for the investigation of putative co-infection cases. This will likely benefit thousands of infected patients every year in a wide area of the Americas.

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Pneumocystis jirovecii can cause severe potentially life-threatening pneumonia (PCP) in kidney transplant patients. Prophylaxis of patients against PCP in this setting is usually performed during 6 months after transplantation. The aim of this study is to describe the molecular epidemiology of a cluster of PCP in renal transplant recipients in Brazil. Renal transplant patients who developed PCP between May and December 2011 had their formalin-fixed paraffin-embedded (FFPE) lung biopsy samples analysed. Pneumocystis jirovecii 23S mitochondrial large subunit of ribosomal RNA (23S mtLSU-rRNA), 26S rRNA, and dihydropteroate synthase (DHPS) genes were amplified by polymerase chain reaction (PCR), sequenced, and analysed for genetic variation. During the study period, 17 patients developed PCP (only four infections were documented within the first year after transplantation) and six (35.3%) died. Thirty FFPE samples from 11 patients, including one external control HIV-infected patient, had fungal DNA successfully extracted for further amplification and sequencing for all three genes. A total of five genotypes were identified among the 10 infected patients. Of note, four patients were infected by more than one genotype and seven patients were infected by the same genotype. DNA extracted from FFPE samples can be used for genotyping; this approach allowed us to demonstrate that multiple P. jirovecii strains were responsible for this cluster, and one genotype was found infecting seven patients. The knowledge of the causative agents of PCP may help to develop new initiatives for control and prevention of PCP among patients undergoing renal transplant and improve routine PCP prophylaxis.