Identification of superficial Candida albicans germ tube antigens in a rabbit model of disseminated candidiasis. A proteomic approach.

The diagnosis of invasive candidiasis remains a clinical challenge. The detection by indirect immunofluorescence of Candida albicans germ-tube-specific antibodies (CAGTA), directed against germ-tube surface antigens, is a useful diagnostic tool that discriminates between colonization and invasion. However, the standardization of this technique is complicated by its reliance on subjective interpretation. In this study, the antigenic recognition pattern of CAGTA throughout experimental invasive candidiasis in a rabbit animal model was determined by means of 2D-PAGE, Western blotting, and tandem mass spectrometry (MS/MS). Seven proteins detected by CAGTA were identified as methionine synthase, inositol-3-phosphate synthase, enolase 1, alcohol dehydrogenase 1,3-phosphoglycerate kinase, 14-3-3 (Bmhl), and Egd2. To our knowledge, this is the first report of antibodies reacting with Bmhl and Egd2 proteins in an animal model of invasive candidiasis. Although all of the antigens were recognized by CAGTA in cell-wall dithiothreitol extracts of both germ tubes and blastospores of C. albicans, immunoelectron microscopy study revealed their differential location, as the antigens were exposed on the germ-tube cell-wall surface but hidden in the inner layers of the blastospore cell wall. These findings will contribute to developing more sensitive diagnostic methods that enable the earlier detection of invasive candidiasis.

Identification of superficial Candida albicans germ tube antigens in a rabbit model of disseminated candidiasis. A proteomic approach

The diagnosis of invasive candidiasis remains a clinical challenge. The detection by indirect immunofluorescence of Candida albicans germ-tube-specific antibodies (CAGTA), directed against germ-tube surface antigens, is a useful diagnostic tool that discriminates between colonization and invasion. However, the standardization of this technique is complicated by its reliance on subjective interpretation. In this study, the antigenic recognition pattern of CAGTA throughout experimental invasive candidiasis in a rabbit animal model was determined by means of 2D-PAGE, Western blotting, and tandem mass spectrometry (MS/MS). Seven proteins detected by CAGTA were identified as methionine synthase, inositol-3-phosphate synthase, enolase 1, alcohol dehydrogenase 1,3-phosphoglycerate kinase, 14-3-3 (Bmh1), and Egd2. To our knowledge, this is the first report of antibodies reacting with Bmh1 and Egd2 proteins in an animal model of invasive candidiasis. Although all of the antigens were recognized by CAGTA in cell-wall dithiothreitol extracts of both germ tubes and blastospores of C. albicans, immunoelectron microscopy study revealed their differential location, as the antigens were exposed on the germ-tube cell-wall surface but hidden in the inner layers of the blastospore cell wall. These findings will contribute to developing more sensitive diagnostic methods that enable the earlier detection of invasive candidiasis (AU)

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Bone microbial decontamination agents in osseous grafting: an in vitro study with fresh human explants.

BACKGROUND: Establishing a safe prophylactic antimicrobial protocol in bone grafting may enhance osseous volume outcomes. The purpose of this in vitro study is to assess human osteoblast response and safety after explant antimicrobial exposure. METHODS: Fresh human bone explants were exposed to three antimicrobials: povidone-iodine (PovI; 0.05%, 1%, and 5%), chlorhexidine (CHX; 0.2% and 1%), and sodium hypochlorite (NaOCl; 2.5%, 4.5%, and 5.25%) at different times (15, 30, 45, and 60 seconds) and concentrations to assess cellular toxicity. Explants were washed three times with saline after exposure. Controls, explants cultured in the absence of antimicrobials, were performed for all experimental situations tested. Trials were conducted in triplicate. Particle size influence on osteoblast growth was determined between bone fragments with a diameter <2 and ≥2 to 5 mm. Test and control groups were monitored by light microscopy to evaluate cellular growth. Osteoblast differentiation and morphology was assessed by alkaline phosphatase activity and scanning electron microscopy (SEM). RESULTS: Osteoblast growth was similar for particles <2 and ≥2 to 5 mm. Alkaline phosphatase control reference values were not significantly different from test groups (0.35 mU/mL ± 0.004 versus 0.34 mU/mL ± 0.009; P >0.05). Light microscopy showed on average 97% osteoblastic growth for bone particles exposed to PovI 5% and CHX 0.2% for all times and CHX 1% up to 30 seconds. The odds ratio of positive osteoblastic growth after a 30-second 2.5% NaOCl exposure was 2.4 times higher than after 5.25%. On average, one of two replicas yielded positive growth with 2.5% NaOCl and one of three with 5.25%. After 60-second explant exposure, positive osteoblastic growth was 7.7 times more likely to occur with 5% PovI or 0.2% CHX than with 5.25% NaOCl (P <0.05). SEM analysis confirmed light microscopy similar cellular adhesion and osteoblast phenotypic features between test and control groups. CONCLUSIONS: Best osteoblastic growth occurred after bone PovI exposure and CHX 0.2%. Cellular toxicity seems to be influenced by the type of antimicrobial, concentration, and exposure time. SEM analysis confirmed absence of osteoblast phenotypic alterations after exposure. Decontamination agents can safely be used in bone transplantation using up to 5% PovI and 0.2% CHX for 1 minute and CHX 1% for 30 seconds.