Experimental design (full details are provided in Methods).

<p>To form <i>Af</i> biofilm, polystyrene disks were placed in tissue culture plates with conidia and media. Disks were incubated to allow the conidia to attach. Following the attachment phase, disks were transferred to new plates containing and incubated for an additional 24 h (line I). Direct interaction of live <i>Pa</i> on <i>Af</i> biofilm formation: Suspensions of conidia and bacteria were combined in tissue culture plates for 16 h. Disks were then rinsed gently, transferred to new plates, and incubated for an additional 24 h (line II). Direct interaction of live <i>Pa</i> on preformed <i>Af</i> biofilm: Fungal biofilms were formed as described. After 16 h the disks were rinsed, transferred to new plates containing <i>Pa</i> suspension and incubated an additional 24 h (lines II and IV). <i>Pa</i> planktonic supernatant assay: To obtain planktonic culture filtrates, <i>Pa</i> suspension was incubated in conical tubes for 24 h. The spent medium was centrifuged to remove suspended cells or debris. The supernatant was filter sterilized and added to wells of a tissue culture plate previously inoculated with <i>Af</i> suspension. Fungal biofilms attached and formed. Disks containing biofilms were washed, transferred to a new plate, and incubated for an additional 24 h (line III). For the preformed biofilm assay, filtered supernatant was added to wells. Disks containing <i>Af</i> preformed biofilms were washed, transferred to the plate containing the bacterial filtrates, and incubated an additional 24 h. <i>Af</i> wells without bacterial supernatant were included as controls (line V). To obtain <i>Pa</i> biofilm filtrates, a suspension of <i>Pa</i> adhered to tissue culture flasks for 2 h (attachment phase). The flasks were rinsed, fresh RPMI-1640 added to the flask and adhered cells formed <i>Pa</i> biofilms for 22 h. The spent medium was then removed. <i>Af</i> conidia forming biofilms or preformed <i>Af</i> biofilms were challenged with the <i>Pa</i> biofilm culture filtrate (lines III and V).