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Phenotypic and molecular characterisation of Sporothrix globosa of diverse origin from India.

ABSTRACT: Sporotrichosis is one of the neglected tropical diseases causing subcutaneous chronic granulomatous lesion by thermally dimorphic fungi belonging to Sporothrix species. Sporothrix brasiliensis, Sporothrix mexicana and Sporothrix globosa are the common pathogenic species. In Asian countries, S. globosa constitutes nearly 99.3% of all Sporothrix species. We studied 63 cases of sporotrichosis of geographically diverse origin from India and Sporothrix isolates were characterised for its growth in different media, temperatures, ability to assimilate sugars and antifungal susceptibility profile. Molecular characterization was performed by sequencing of the calmodulin (CAL), beta tubulin (BT) and translational elongation factor 1-alpha (TEF-1α) and typing by fluorescent amplified fragment length polymorphism (FAFLP). In patients who presented with fixed (49.2%), lymphocutaneous lesions (23.8%), in 26.9% the details were not known, none had systemic dissemination. All the isolates tested were Sporothrix globosa and that could grow up to 35 °C and unable to grow at and beyond 37 °C. The assimilation of sucrose, ribitol and raffinose helps in identifying S. globosa. Sequences of CAL or BT or TEF-1α can differentiate S. globosa from other species in the complex. FAFLP results exhibited low genetic diversity. No correlation was noted between genotypes and clinical presentation, or geographic distribution. Itraconazole, terbinafine and posaconazole showed good in vitro antifungal activity against S. globosa whereas fluconazole and micafungin had no activity. S. globosa of Indian origin is relatively less pathogenic than other pathogenic Sporothrix species as it does not cause systemic dissemination and in the diagnostic laboratory, incubation of the cultures below 37 °C is essential for effective isolation.

SUBMITTER: Rudramurthy SM

PROVIDER: S-EPMC7966662 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Sporotrichosis is a cutaneous and subcutaneous fungal disease of humans and other mammals, known to be caused by the Sporothrix schenckii species complex, which comprises four species of clinical importance: S. brasiliensis, S. globosa, S. luriei, and S. schenckii sensu stricto. Of them, S. globosa and S. schenckii s. str. show global distribution and differences in global frequency as causal agents of the disease. In the Americas, only three species are present: S. schenckii s. str., S. brasiliensis (so far, only reported in Brazil), and S. globosa. In Venezuela, since the first case of sporotrichosis reported in 1935, S. schenckii have been considered its unique etiological agent. In the present work, the presence of more than one species in the country was evaluated.By phenotypic key features and molecular phylogeny analyses, we re-examined 30 isolates from diverse Venezuelan regions belonging to the fungi collection of Instituto de Biomedicina, Caracas, Venezuela, and national reference center for skin diseases. All isolates were collected between 1973 and 2013, and maintained in distilled water.Sporotrichosis in Venezuela is mainly caused by S. schenckii s. str. (70%). However, a significant proportion (30%) of sporotrichosis cases in the country can be attributable to S. globosa. A correlation between intraspecific genotypes and clinical presentation is proposed.Our data suggest that sporotrichosis various clinical forms might be related to genetic diversity of isolates, and possibly, to diverse virulence profiles previously reported in the S. schenckii species complex. Sporothrix globosa was found to be the causative agent of 30% of sporotrichosis for the Venezuelan cases re-examined, the highest frequency of this species so far reported in the Americas. The high genetic variability presented by S. schenckii s. str. indicates that species distinction based on phenotypic key features could be a challenging and uncertain task; molecular identification should be always employed.

Project description:Abstract Poster session 1, September 21, 2022, 12:30 PM - 1:30 PM Objectives Sporotrichosis is the leading subcutaneous mycosis caused by the Sporothrix (S.) schenckii complex. S. globosa is the causative organism of fixed sporotrichosis in Korea. The preferred regimen of cutaneous sporotrichosis is itraconazole for 3-6 months, however, there were few studies for recalcitrant sporotrichosis. Methods In 2018, we performed a histological examination of a patient who suffered sporotrichosis for 3 years and cultured part of the specimen. Despite various regimens for years, improvement and exacerbation were repeated, so we took another skin biopsy and cultured it in 2021. Isolates from the 2018 and 2021 lesions were identified as S. globosa by ribosomal DNA ITS sequencing (GenBank accession number: MH499862 and MH499863). The in vitro antifungal sensitivity tests were performed by broth microdilution method according to CLSI M38-A2 guidelines or Sensititre YeastOne® manufacturer's instructions. They were incubated at 30°C in a non-CO2 incubator for 7 days. Results In 2018, histologically, we observed chronic inflammatory granuloma comprising lymphocytes, histiocytes, and giant cells, and several spores with periodic acid-Schiff (PAS) staining. Microscopic findings and ITS sequences of rRNA gene were identical with S. globosa. The antifungal susceptibility profile in 2018 revealed sensitive to terbinafine (0.125 μg/ml), and moderate to high MIC values for amphotericin B (2 μg/ml), itraconazole (>16 μg/ml), voriconazole (>16 μg/ml), and echinocandins (>16 μg/ml). Treatment with terbinafine, itraconazole, or amphotericin B, the skin lesions were partially improved, but were not cured. In 2021, we took another skin biopsy and culture specimen. Histopathological and mycological examination results were the same as before. The antifungal susceptibility profile revealed sensitive to itraconazole (0.5/ml), and high MIC for others. Clinically, skin lesions were not improved with the use of itraconazole 200 mg/d. Itraconazole 400 mg/d with local heating induced moderate improvement. There was no evidence of immune deficiency. Conclusion We experienced recalcitrant sporotrichosis that did not respond to itraconazole and terbinafine, and the sensitivity of antifungal was changed. In this case, the combination treatment including local heating, saturated KI may be considered, and frequent antifungal susceptibility tests are needed.

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Phenotypic and molecular characterisation of Sporothrix globosa of diverse origin from India.

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