Duplication and overexpression of the genes encoding a beta 1,3-glucan synthase confer the intrinsic resistance to echinocandin in Mucor circinelloides

Abstract

ABSTRACTProcedures such as solid organ transplants and cancer treatments can leave many patients in an immunocompromised state resulting in an increased susceptibility to opportunistic diseases including fungal infections. Mucormycosis infections are continually emerging and pose a serious threat to immunocompromised patients. Currently there has been a sharp increase in mucormycosis cases as a secondary infection in patients battling SARS-CoV-2 infections. Mucorales fungi are notorious for presenting resistance to most antifungal drugs. The absence of effective means to treat these infections results in mortality rates approaching 100% in cases of disseminated infection. One of the most effective antifungal drug classes currently available are echinocandins. Echinocandins seem to be efficacious in treatment of many other fungal infections. Unfortunately, susceptibility testing has found that echinocandins have no to little effect on Mucorales. In this study, we found that the model Mucorales Mucor circinelloides genome carries three copies of the genes encoding for the echinocandin β-(1,3)-D-glucan synthase (fksA, fksB, and fksC). Interestingly, we revealed that exposing M. circinelloides to micafungin significantly increased the expression of the fksA and fksB genes when compared to an untreated control. We further uncovered that the serine/threonine phosphatase calcineurin is responsible for the overexpression of fksA and fksB as deletion of calcineurin results in a decrease in expression of all three fks genes and a lower minimal inhibitory concentration (MIC) to micafungin. Taken together, this study demonstrates that the fks gene duplication and overexpression by calcineurin contribute to the intrinsic resistance to echinocandins in Mucor.IMPORTANCEThe recent emergence of mucormycosis cases in immunocompromised patients has become a more prevalent issue. The Mucorales fungi that cause these infections are known to be highly drug resistant, thus treatment options are limited and the mortalities of these types of infections remain unacceptably high. Echinocandins are one of the latest antifungal drugs developed to successfully treat fungal infections, but it remains unclear why Mucorales fungi are resistant to these treatments. In our study, we uncovered three copies of the genes (fks) encoding the drug target for echinocandins. Furthermore, we discovered that the serine-threonine phosphatase calcineurin is regulating the over-expression of these genes, which confers resistance. By inhibiting calcineurin we found that the expression of these drug targets decreases resulting in an increase in susceptibility to echinocandins both in vitro and in vivo