# Clinical and Molecular Characterization of Feline Sporotrichosis in the Brazilian Amazon: PCR-Based Identification of Sporothrix brasiliensis

## TL;DR

This study examines feline sporotrichosis in the Brazilian Amazon, identifying Sporothrix brasiliensis as the main cause and optimizing DNA-based diagnostic methods.

## Contribution

The study introduces optimized DNA extraction and PCR methods for identifying Sporothrix brasiliensis in cats, enhancing diagnostic accuracy in the region.

## Key findings

- Most infected cats were young, male, and free-roaming with skin lesions.
- Phenol–chloroform DNA extraction and ITS1–ITS4 primers were most effective for fungal identification.
- All isolates were confirmed as Sporothrix brasiliensis with specific antifungal susceptibility profiles.

## Abstract

Sporotrichosis is a fungal disease that affects both animals and humans. In recent years, cases in domestic cats have increased in the Brazilian Amazon, raising public health concerns. This study analyzed cats diagnosed with sporotrichosis in Manaus, focusing on their clinical signs and behavioral features. In the laboratory, we tested different methods for extracting DNA from the fungus and assessed the efficiency of molecular techniques to identify the fungal species involved. Additionally, we used in silico tools and genetic analysis to confirm the identity of the fungus. Our results showed that most infected cats were young, male, and free-roaming, and had skin lesions. The laboratory tests revealed phenol–chloroform DNA extraction methods and its genetic markers were effective for fungal identification. Molecular analysis demonstrated that the predominant species in the region is Sporothrix brasiliensis, a fungus known for its zoonotic potential. These findings can help to improve diagnostic tools and to provide surveillance information essential for controlling the spread of the disease in the Amazon region.

Sporotrichosis is a zoonotic fungal infection with increasing incidence in the Brazilian Amazon, primarily affecting domestic cats and posing risks to human health. This study characterized the clinical and epidemiological profiles of 29 feline sporotrichosis cases in Manaus and optimized molecular diagnostic methods for Sporothrix species identification. Most affected cats were young (86.2% aged 1–3 years), male (82.7%), and free-roaming or semi-indoor (44.8% each), frequently presenting cutaneous lesions localized at the nasal planum (23.3%), ears (7%), eyes (2.3%) and other facial areas (18.6%). Three DNA extraction methods were compared; the phenol–chloroform protocol yielded the highest DNA concentration and purity, and ITS1–ITS4 primers showed an adequate sensibility for PCR detection. In silico RFLP profiles using common restriction enzymes showed limited discriminatory power among Sporothrix species. ITS sequencing of four high-quality amplicons confirmed all isolates as Sporothrix brasiliensis. Antifungal susceptibility testing of all isolates revealed geometric mean MICs of 0.25 µg/mL for ketoconazole, 0.57 µg/mL for itraconazole, 7.27 µg/mL for amphotericin B, and 64 µg/mL for fluconazole, respectively. These findings provide clinical, molecular, and therapeutic information supporting the diagnosis and surveillance of feline sporotrichosis in the Amazon, reinforcing the need for ongoing veterinary and public health monitoring.