AUTHOR=Villis Paulo C. M. , de Macedo Alessandra T. , Furtado Haryne L. A. , Fontenelle Pedro H. C. , Gonçalves Ingrid S. , Mendes Thayariane L. , Motta Brenda L. A. , Marinho Pedro L. L. , Pinheiro Aruanã J. M. C. R. , Lima-Neto Lídio G. , Monteiro Cristina A. , da Silva Luís C. N. , Ferreira Gabriella F. , Holanda Rodrigo A. , Santos Julliana R. A. 

TITLE=A Study of the Disruptive Effect of the Acetate Fraction of Punica granatum Extract on Cryptococcus Biofilms

JOURNAL=Frontiers in Microbiology

VOLUME=11

YEAR=2021

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2020.568258

DOI=10.3389/fmicb.2020.568258

ISSN=1664-302X

ABSTRACT=<p>Cryptococcosis, caused by yeasts of the genus <italic>Cryptococcus</italic>, is an infectious disease with a worldwide distribution. <italic>Cryptococcus neoformans</italic> and <italic>Cryptococcus gattii</italic> are the species that commonly cause this disease in humans; however, infections caused by <italic>Cryptococcus laurentii</italic>, especially in immunocompromised patients, are increasingly being reported. Owing to the increase in the resistance of fungi to antifungals, and a lack of treatment options, it is important to seek new therapeutic alternatives such as natural products. Among these are plant species such as <italic>Punica granatum</italic>, which is used in folk medicine to treat various diseases. This study aimed to evaluate the activity of the acetate fraction of <italic>P. granatum</italic> leaf extract against environmental and clinical isolates of <italic>Cryptococcus</italic>. Three environmental isolates of <italic>C. laurentii</italic>, PMN, PMA, and PJL II, isolated from soils of different municipalities in the state of Maranhão, a clinical isolate, <italic>C. gattii</italic>, from a patient with neurocryptococcosis, and a standard strain of <italic>C. gattii</italic> (ATCC 32068) were used. The minimum and fractional inhibitory concentrations (MIC and FIC, respectively) and time-kill curve of the extract and fluconazole were determined to assess the susceptibility profile of the fungal isolates. Larvae of <italic>Tenebrio molitor</italic> were infected with <italic>Cryptococcus</italic> strains, and the effects of acetate fraction of <italic>P. granatum</italic> extract and fluconazole on the survival and fungal burden were determined. The extract activity was tested against pre-formed biofilms. The acetate fraction of <italic>P. granatum</italic> extract showed promising antifungal activity against all the species of <italic>Cryptococcus</italic> evaluated in this study, with an MIC value lower than that of fluconazole. The indices obtained in the FIC test indicated that the antimicrobial effect of the combination of the extract and antifungal was indifferent for 80% of the isolates. The <italic>P. granatum</italic> acetate fraction reduced the pre-formed biofilm of some isolates, showing better activity than fluconazole, which is consistent with results from fluorescence microscopy. This is the first study on the use of <italic>P. granatum</italic> and its ability to inhibit <italic>Cryptococcus</italic> biofilms; therefore, further studies and tests are needed to investigate the components and mechanism of action of <italic>P. granatum</italic> against cryptococcosis agents.</p>