engleski

The in vitro mycotoxin production of clinical and environmental isolates of Aspergillus fumigatus and A. flavus strains

Moulds from the genus Aspergillus are one of the most common fungi that can infect immunocommpromised patients causing life-threatening invansive aspergillosis (IA) with high mortality rate. IA occurs in patients with hematological malignicity, solid-organ transplantation and AIDS. But why are Aspergillus species predominant fungal isolates in immunocommpromised patients? The mechanism of pathogenicity of opportunistic Aspergillus species is not fully understood and except of host factors various putative virulence factors have been attributed to Aspergillus pathogenicity. These include physical factors (size of conidia and thermotolerance), secretion of enzymes and mycotoxins. Because of that we tested mycotoxin production ability of clinical isolates of Aspergillus strains in comparison with the environmental isolates. 50 strains of A. fumigatus (AFG) and 30 strains of A. flavus (AFL) were collected from immunocompromised patients with IA or with probably IA undergoing hematological malignicity mainly. Environmental isolates (50 of AFG and 30 of AFL) were isolated from soil or air. For in vitro mycotoxin production ability biosynthesis of Aspergillus strains in two media were used: yeast extract (YES) and Czapek-Dox broth (CZA) at 37&plusmn ; 1&ordm ; C and 25&plusmn ; ; 2&ordm ; C during 12 days. Gliotoxin and aflatoxins were identified using TLC, two-dimensional TLC and "bioassay in situ" against Bacillus subtilis NCTC 8236. Concentration of gliotoxin and aflatoxins in biosynthesis were calculated semiquantitatevely on TLC. Gliotoxin was detected only in group of clinical isolates of AFG strains (9 out of 50, 18%). After 3 days of incubation at 37&plusmn ; 1&ordm ; C and 25&plusmn ; 2&ordm ; C, concentration of gliotoxin was in detectable concentration which depend on AFG strains (range from 0.5 to 8 mg/mL in YES at 37&plusmn ; 1&ordm ; C and from 0.1 to 3.5 mg/mL at 25&plusmn ; 2&ordm ; C, range from 0.1 to 2.5 mg/mL in CZA at 37&plusmn ; 1&ordm ; C and from 0.05 to 2 mg/mL at 25&plusmn ; 2&ordm ; C). Prolonged incubation of biosyntheses on both temperatures had effect on higher production of gliotoxin with highest concentration in YES and 37&plusmn ; 1&ordm ; C. Production of aflatoxin B1 was detected after 6 days of incubation in both groups of AFL isolates. Seven strains (23%) of clinical isolates and 11 strains (37%) of environmental isolates of AFL produce aflatoxin B1 in range from 0.024 to 1.2 ug/mL. Production of aflatoxin G1 was also noticed in one clinical isolate of AFL (0.12 ug/mL) and in one environmental isolate (0.024 ug/mL). The role of mycotoxins in etiology of IA and other mycoses are not yet understood. Because of many proved toxic effects of gliotoxin its production in situ probably could contribute pathogenesis of IA. Our research showed that 18% of investigated clinical strains of A. fumigatus produce gliotoxin in vitro. Future research must be performed for detection of mycotoxins in infected tissues with Aspergillus as confirmation of its role in etiology of IA.

Aspergillus fumigatus; Aspergillus flavus; gliotoxin; aflatoxins; aspergilloma

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