PMC9442803_01
(A) Burn map of the burn injuries sustained by the patient, and timeline of events incurred by the case in Intensive Care Unit (ICU) represented by the green arrow while the red line represents the duration of vasopressors administration. (B) Mucormycosis of the abdominal wall. (B.I.) and (B.II.) Show peristomial ulcerations. (B.III.) Horizontal abdominal scan assessing the intra-abdominal extension of the cellulitis. (C) Evolution and healing of the abdominal lesions in response to IFN-gamma treatment. (C.I.) Lesions before IFN-gamma treatment. (C.II.) and (C.III.) Lesions after 6 days of IFN-gamma and right before surgery. (C.IV.) (C.V.) and (C.VI.) Excision of the abdominal wall and the transposition of the colostomy.
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In parallel, the burn injuries were treated over a course of 11 skin grafts surgeries ( Figure 1 ). After one month in the ICU, as the patient had resolved all organ failures and was slowly improving, two necrotic ulcers were noticed adjacent to the colostomy, and a third one on the left flank ( Figures 1B ). A Computerized Tomography (CT) scan was performed to assess the spread of this abdominal fungal cellulitis which did not show any deep visceral extension of the infection ( Figure 1B ). Macroscopically, IFN-gamma treatment was associated with rapid signs of microbiological cure ( Figure 1C ), showing clear healing of the previously inflammatory and necrotic lesions. The surgery consisted of a large resection of the abdominal wall ( Figure 1C ), transposition of the colostomy, and vacuum therapy with subsequent skin flap graft on the excised zone. Besides common laboratory tests, we collected samples to perform flow cytometry ( Supplementary Figure S1 ), quantification of protein soluble markers ( Supplementary Table S1 ) and transcriptional response ( Supplementary Figures S2, S3  and  Supplementary Tables S2 ,  S3 ), as well as immune functional assays (IFA) ( Supplementary Table S4 ).
Immunomonitoring of cell surface markers using flow cytometry, transcriptomic analysis using the Immune profiling Panel (IPP) prototype and cytokines measured by ELLA platform after being stimulated in Truculture  tubes (Immune functional assays). Markers' measurements are represented on the y-axis, while the x-axis indicates the different time points measured. The highlighted pink area denotes the duration of the IFN-gamma treatment. The vertical blue line denotes the day of the surgery. The boxplots on the left illustrate expression levels in healthy volunteers (green) and ICU patients (red) as reference. (A) Evolution of antigen presentation pathway. (B) Neutrophil-related markers. (C) Lymphocyte-related markers. (D) Cytokine production after immune functional assays.
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Besides common laboratory tests, we collected samples to perform flow cytometry ( Supplementary Figure S1 ), quantification of protein soluble markers ( Supplementary Table S1 ) and transcriptional response ( Supplementary Figures S2, S3  and  Supplementary Tables S2 ,  S3 ), as well as immune functional assays (IFA) ( Supplementary Table S4 ). As expected, treatment with IFN-gamma had a strong effect exhibiting a transient activation of the antigen presentation pathway and monocytes-related markers reflected by the 3-fold increased mHLA-DR expression from 11,447 Ab/C to 36,375 Ab/C (measured on the day of the surgery -  Figure 2A ). CD74 and ARL14EP expression was observed reaching the levels of the healthy reference group ( Figure 2A ). CD74, an HLA-DR antigen-associated invariant chain has a role in MHC class II trafficking to the antigen processing compartments, while, ARL14EP controls the export of MHC class II molecules from endosomal MIIC. Neutrophil cell surface transcriptomic markers: CD177 (neutrophil-specific marker) and S100A9 (a DAMP expressed on both neutrophil and monocyte) ( Figure 2B ) showed a transient downregulation associated with IFN-gamma treatment. Along with a steep increase in transcriptomic markers: CD3D, ZAP70 and IL-7R ( Figure 2C ) was observed at the initiation of IFN-gamma therapy. NK cells (which are a key part of anti-fungal immunity) tended to increase under treatment, and this effect was persistent after the surgery (from 64-120 cells/microL on the day of surgery, and 202 cells/microL a week later) ( Figure 2C ). IL-6, TNF and IL-10 were within the expected range for the septic patients, however, no changes were observed during the IFN-gamma treatment ( Supplementary Table S4 ). Interestingly, the SEB stimulation highlighted a clear immediate increase of IFN-gamma (3-fold) and IL-2 (4.5-fold) cytokines that was maintained until the end of IFN-gamma therapy ( Figure 2D ).