Immune sensing of Candida albicans requires cooperative recognition of mannans and glucans by lectin and Toll-like receptors.

The fungal pathogen Candida albicans has a multilayered cell wall composed of an outer layer of proteins glycosylated with N- or O-linked mannosyl residues and an inner skeletal layer of beta-glucans and chitin. We demonstrate that cytokine production by human mononuclear cells or murine macrophages was markedly reduced when stimulated by C. albicans mutants defective in mannosylation. Recognition of mannosyl residues was mediated by mannose receptor binding to N-linked mannosyl residues and by TLR4 binding to O-linked mannosyl residues. Residual cytokine production was mediated by recognition of beta-glucan by the dectin-1/TLR2 receptor complex. C. albicans mutants with a cell wall defective in mannosyl residues were less virulent in experimental disseminated candidiasis and elicited reduced cytokine production in vivo. We concluded that recognition of C. albicans by monocytes/macrophages is mediated by 3 recognition systems of differing importance, each of which senses specific layers of the C. albicans cell wall.

Serologic diagnosis of allergic bronchopulmonary aspergillosis in patients with cystic fibrosis through the detection of immunoglobulin G to Aspergillus fumigatus.

Allergic bronchopulmonary aspergillosis (ABPA) is seen in approximately 10% of patients with cystic fibrosis (CF) and can be difficult to diagnose. Consensus criteria require the presence of multiple elevated immunologic markers such as total immunoglobulin E (IgE), Aspergillus IgE and Aspergillus IgG, or precipitins for a robust diagnosis. There is some degree of standardization of total IgE and Aspergillus IgE levels, but there is no standardization in the measurement of IgG antibodies or precipitins to Aspergillus. The interpretation of results may, therefore, be confusing. Eighty-seven patients with CF were categorized as having ABPA or as controls, using the consensus criteria and an in-house enzyme immunoassay to measure IgG levels to Aspergillus. All sera from patients were then analyzed by commercial fluorescent immunoassay (FEIA) for the quantitative detection of anti-Aspergillus IgG. FEIA results were analyzed against the consensus conference minimum diagnostic criteria to ascertain a cutoff point, which could predict a diagnosis of ABPA in CF. Eighty patients with CF and with no or incomplete evidence of ABPA had a mean FEIA score of 51.1 mg/L, whereas 7 CF patients with ABPA had a mean FEIA score of 132.5 mg/L. Using receiver operator characteristic curve analysis of the ImmunoCAP (Phadia) IgG score on ABPA versus all other patients gave an area under the curve of 0.933 (estimated SE, 0.027). This analysis provisionally suggested that a score of 90 mg/L may be used as a cutoff point, which would give a sensitivity of 91% and specificity of 88.0% for the diagnosis of ABPA, though this requires further validation. This quantitative approach to Aspergillus IgG measurement in patients with CF along with the results of other tests will hopefully provide a more accurate approach to the diagnosis of ABPA.

A consensus on fungal polymerase chain reaction diagnosis?: a United Kingdom-Ireland evaluation of polymerase chain reaction methods for detection of systemic fungal infections.

The limitations of classical diagnostic methods for invasive fungal infections (IFIs) have led to the development of molecular techniques to aid in the detection of IFIs. Despite good published performance, interlaboratory reproduction of these assays is variable, and no consensus has been reached for an optimal method. This publication describes the first multicenter study of polymerase chain reaction methods, for the detection of Aspergillus and Candida species, currently used in the UK and Ireland by distribution and analysis of multiple specimen control panels. All three Candida methods were comparable, achieving a satisfactory level of detection (10 cfu), and the method of preference was dependent on the requirements of the particular laboratory. The results for the five Aspergillus assays were more variable, but two methods (2Asp and 4Asp) were superior (10(1) conidia). Formally, the overall performances of the two Aspergillus assays were comparable (kappa statistic = 0.77). However, on the Roche LightCycler, there was a clear sample-type effect that greatly reduced the detection limit of the 4Asp method when testing whole blood samples. Therefore, the preferred Aspergillus method relied on the amplification platform available to the user. This study represents the initial process to achieve a consensus method for the diagnosis of IFIs.

Appendicectomy for suspected uncomplicated appendicitis is associated with fewer complications than conservative antibiotic management: a meta-analysis of post-intervention complications.

OBJECTIVES: Recent literature has concluded antibiotic therapy results in fewer complications than appendicectomy for patients with uncomplicated appendicitis. This studies aim was to undertake a meta-analysis of major post-intervention outcomes in patients with suspected uncomplicated appendicitis treated with antibiotics or appendicectomy, and determine which treatment is associated with the lowest rate of major complications. METHODS: We analysed randomised trials of antibiotics vs. appendicectomy in adults with suspected uncomplicated appendicitis. The primary outcome measure was a composite of major complications, peritonitis and intra-abdominal abscess, occurring after appendicectomy or initiation of therapeutic antibiotics. RESULTS: The rate of major post-intervention complications was 0.8% (2/263) in the appendicectomy group and 10.1% (27/268) in the antibiotic group. This difference was statistically significant by the random effects model: Risk Ratio 7.71, 95% C.I. 2.33 to 25.53, Risk Difference 0.09: 95% C.I. 0.05 to 0.13. The Number Needed to Harm (NNH) from antibiotic therapy is 10.7. CONCLUSIONS: Suspected uncomplicated appendicitis has a lower rate of major post-intervention complications when managed with primary appendicectomy compared to antibiotic therapy.

Loss of cell wall mannosylphosphate in Candida albicans does not influence macrophage recognition.

The outer layer of the cell wall of the human pathogenic fungus Candida albicans is enriched with heavily mannosylated glycoproteins that are the immediate point of contact between the fungus and cells of the host, including phagocytes. Previous work had identified components of the acid-labile fraction of N-linked mannan, comprising beta-1,2-linked mannose residues attached via a phosphodiester bond, as potential ligands for macrophage receptors and modulators of macrophage function. We therefore isolated and disrupted the CaMNN4 gene, which is required for mannosyl phosphate transfer and hence the attachment of beta-1,2 mannose oligosaccharides to the acid-labile N-mannan side chains. With the mannosylphosphate eliminated, the mnn4Delta null mutant was unable to bind the charged cationic dye Alcian Blue and was devoid of acid-labile beta-1,2-linked oligomannosaccharides. The mnn4Delta mutant was unaffected in cell growth and morphogenesis in vitro and in virulence in a murine model of systemic C. albicans infection. The null mutant was also not affected in its interaction with macrophages. Mannosylphosphate is therefore not required for macrophage interactions or for virulence of C. albicans.