Steven Sayson. Role of extracellular vesicles (EVs) in host-pathogen interactions. Pneumocystis spp. have an evolutionary loss of many biosynthetic processes, including the ability to synthesize any of the 20 amino acids, co-factors, and the fungal sterol, ergosterol. In place of ergosterol, host cholesterol has been shown to be incorporated into Pneumocystis spp. as the critical sterol. However, the mechanism in which Pneumocystis obtains cholesterol is unclear. In addition to cholesterol, our lab has discovered that mammalian Podoplanin, an abundant alveolar type 1 cell membrane protein, is found within the Pneumocystis membrane. Previous studies have shown that EVs are rich in proteins, lipids, RNAs, and free amino acids; more specifically, EVs are rich in cholesterol and membrane proteins from their host cell. These data suggest that, since Pneumocystis spp. are host-obligate and depend on the host microenvironment for nutrition, EVs are involved in transferring the components required for Pneumocystis metabolism. Current projects in the lab aim to dissect the contents of host- and Pneumocystis- derived EV proteins as well as their role in promoting Pneumocystis survival. These studies will help us in understanding Pneumocystis metabolism and aid in the development of an in vitro culture system.

Alan Ashbaugh. Drug efficacy testing in small animal models of Pneumocystispneumonia (PCP) – Current drugs used to treat PCP are limited by problems of efficacy and toxicity. There is also increasing evidence of resistance to the most commonly used drugs when treating patients with the disease. One focus of our lab is testing investigational compounds and licensed drugs for efficacy against PCP in both a therapy and prophylaxis model. Since Pneumocystis organisms cannot be grown in culture, animals must be used to study these therapies to develop better treatment options for patients. Recent studies have focused on echinocandins and echinocandin analogs due to their inhibition of β-1,3-D glucan synthesis. β-1,3-D glucan is an essential cell wall component of many fungi, including the ascus of Pneumocystis spp. Two promising candidates in these categories are the investigational compounds rezafungin (Cidara Therapeutics) and ibrexafungerp (Scynexis, Inc.). No Pneumocystis organisms were observed microscopically in mice that were allowed to develop PCP and then treated for 8 weeks with rezafungin. Also, mice prophylactically treated with rezafungin for 4 weeks did not allow any residual organisms to activate an infection after cessation of treatment. For ibrexafungerp, mice prophylactically treated for 6 weeks did not allow any active infection to develop after cessation of treatment.

Aaron Albee. Characterizing Pneumocystis spp. reproductive strategies and how they impact Pneumocystis infection will be the focus of my doctoral studies. Pneumocystis spp. is believed to employ both asexual and sexual reproduction during active infection based on previous microscopic observation. Asexual reproduction involves a single trophic cell which undergoes binary fission to produce two identical daughter cells. Previous studies into the life cycle and ultrastructure of Pneumocystis spp. have shown evidence of mating conjugation of two trophic cells and visible signs of sexual reproduction such as chromosomal alignment for recombination. Genomic studies have also identified mating related genes that encode the pheromone receptors Mam2 and Map3, a component of how mating cells recognize other compatible cells. I will undertake a series of experiments to better characterize the structure and biology of these two receptors in Pneumocystis. These experiments will also profile expression of these receptors to identify which of the multiple types of sexual reproduction Pneumocystis spp. uses. The data produced by these studies will help us to better understand both proliferation and infectious spread of Pneumocystis spp.

Sandy Rebholz. Studying the metabolic pathways of the fungal pathogens in Pneumocystis species; the host response to these lung dwelling fungi; in-vitro drug assessments and pre-clinical drug development for the treatment of the pneumonia caused by these fungi; as well as identifying genes of the pathogen in response to excessive extracellular copper found in the hosts’ lungs.  Unfortunately, there is no long-term culture system for Pneumocystis so finding alternate ways in which to grow Pneumocystis such as 3D tissue culture is a priority. Pneumocystis are obligate host-specific fungi that dwell in the lungs of mammals but can cause a lethal pneumonia when the host’s immune system becomes debilitated.  Pneumocystis infects humans receiving therapy for chronic inflammatory diseases like rheumatoid arthritis; in patients after stem cell and solid organ transplants; and in some patients receiving cancer chemotherapy. There are few drugs with which to treat the infection and drug discovery is a major focus of the lab- using molecular approaches such as RNA-seq to discover new drug targets.