Adhesion of Candida parapsilosis to Bovine Serum Albumin under Fluid Shear

Candida spp. are common commensal organisms on human skin and mucosae that can lead to invasive diseases among the immunocompromised with substantial associated morbidity, mortality, and cost^1,2,3. Although C. albicans remains an important cause of these infections, non-albicans species such as C. parapsilosis, C. glabrata, C. krusei, C. tropicalis, and C. auris are being increasingly recognized, especially in vulnerable populations and with frequent resistance to available antifungal drugs⁴. Non-albicans species present distinct elements of biology and pathogenesis that are under active investigation.

Adhesion is an important first step in colonization and pathogenesis. Interference with this step may therefore offer an opportunity to stop disease progression at an early stage. Studies of Candida adhesion and invasion have been predominantly focused on static conditions^5,6. These studies have helped define the structure and functions of fungal adhesins in disease^7,8,9. However, adhesion in the bloodstream, gastro-intestinal (GI) tracts, and urinary tracts, and in catheters must occur under conditions of fluid shear flow which places unique constraints upon adhesion. Adhesion under shear requires rapid catch bond formation and the ability to withstand strong pulling forces produced due to the movement of liquids^10,11. The C. albicans adhesin, Als5 has been shown to facilitate shear dependent adhesion^12,13. CpAls7 (CpALS4800) has been previously shown to mediate adhesion of Cp to epithelial cells, and a knockout showed decreased virulence in a urinary tract infection model¹⁴. We demonstrated that CpALS4800 promotes adhesion under physiologically relevant fluid shear conditions¹⁵.

Candida colonization and pathogenesis have been extensively studied in the animal models^16,17,18. The most frequently used models are murine mucosal and bloodstream infections but invertebrate models, such as Galleria larvae, are increasingly being used because of the low cost, rapid throughput, and simplicity. Animal models recapitulate many steps of the human disease process in both the pathogen and host, including the host adaptive and innate immune responses, interactions of yeast with tissues and the microbiota, and yeast responses to the host environment. In contrast, in vitro adhesion assays permit the focus specifically on the adhesion step, and on the experimental manipulation of variables such as shear force, growth conditions of yeast, and adhesion to specific substrates.

Because Cp is capable of growth in both humans and environmental sources, it is likely to be capable of sensing and responding to different environments. In support of this notion, multiple clinical isolates of Cp show low adhesion under fluid shear when grown in the standard yeast growth medium, yeast-peptone-dextrose (YPD), but switch to strong adhesion when grown for a few hours at 37 °C in the tissue-culture medium 199 (M199)^15,19. A detailed protocol is provided here for a medium throughput assay that permits the measurement of adhesion of multiple yeast samples that run in parallel, under defined conditions of growth, fluid shear, temperature, and substrate. The assay has been designed to maximize reproducibility, and to allow for the use of clinical isolates of Cp, as well as strains that have been experimentally manipulated in the lab. The assay as described here, for Cp adhesion to a bovine serum albumin (BSA) substrate, demonstrates that clinical isolates exhibit a range of adhesion, whereas two commonly used lab strains, CDC317 and CLIB214 show poor adhesion.