Humoral Immunity Links Candida albicans Infection and Celiac Disease

Objective The protein Hwp1, expressed on the pathogenic phase of Candida albicans, presents sequence analogy with the gluten protein gliadin and is also a substrate for transglutaminase. This had led to the suggestion that C. albicans infection (CI) may be a triggering factor for Celiac disease (CeD) onset. We investigated cross-immune reactivity between CeD and CI. Methods Serum IgG levels against recombinant Hwp1 and serological markers of CeD were measured in 87 CeD patients, 41 CI patients, and 98 healthy controls (HC). IgA and IgG were also measured in 20 individuals from each of these groups using microchips sensitized with 38 peptides designed from the N-terminal of Hwp1. Results CI and CeD patients had higher levels of anti-Hwp1 (p= 0.0005 and p= 0.004) and anti-gliadin (p= 0.002 and p= 0.0009) antibodies than HC but there was no significant difference between CeD and CI patients. CeD and CI patients had higher levels of anti-transglutaminase IgA than HC (p= 0.0001 and p= 0.0039). During CI, the increase in anti-Hwp1 paralleled the increase in anti-gliadin antibodies. Microchip analysis showed that CeD patients were more reactive against some Hwp1 peptides than CI patients, and that some deamidated peptides were more reactive than their native analogs. Binding of IgG from CeD patients to Hwp1 peptides was inhibited by gamma III gliadin peptides. Conclusions Humoral cross-reactivity between Hwp1 and gliadin was observed during CeD and CI. Increased reactivity to Hwp1 deamidated peptide suggests that transglutaminase is involved in this interplay. These results support the hypothesis that CI may trigger CeD onset in genetically-susceptible individuals.

Candida albicans Increases Tumor Cell Adhesion to Endothelial Cells In Vitro: Intraspecific Differences and Importance of the Mannose Receptor

The dimorphic fungus Candida albicans is able to trigger a cytokine-mediated pro-inflammatory response that increases tumor cell adhesion to hepatic endothelium and metastasis. To check the intraspecific differences in this effect, we used an in vitro murine model of hepatic response against C. albicans, which made clear that tumor cells adhered more to endothelium incubated with blastoconidia, both live and killed, than germ tubes. This finding was related to the higher carbohydrate/protein ratio found in blastoconidia. In fact, destruction of mannose ligand residues on the cell surface by metaperiodate treatment significantly reduced tumor cell adhesion induced. Moreover, we also noticed that the effect of clinical strains was greater than that of the reference one. This finding could not be explained by the carbohydrate/protein data, but to explain these differences between strains, we analyzed the expression level of ten genes (ADH1, APE3, IDH2, ENO1, FBA1, ILV5, PDI1, PGK1, QCR2 and TUF1) that code for the proteins identified previously in a mannoprotein-enriched pro-metastatic fraction of C. albicans. The results corroborated that their expression was higher in clinical strains than the reference one. To confirm the importance of the mannoprotein fraction, we also demonstrate that blocking the mannose receptor decreases the effect of C. albicans and its mannoproteins, inhibiting IL-18 synthesis and tumor cell adhesion increase by around 60%. These findings could be the first step towards a new treatment for solid organ cancers based on the role of the mannose receptor in C. albicans-induced tumor progression and metastasis.

Clinical factors associated with a Candida albicans Germ Tube Antibody positive test in Intensive Care Unit patients

Background: Poor outcomes of invasive candidiasis (IC) are associated with the difficulty in establishing the microbiological diagnosis at an early stage. New scores and laboratory tests have been developed in order to make an early therapeutic intervention in an attempt to reduce the high mortality associated with invasive fungal infections. Candida albicans IFA IgG has been recently commercialized for germ tube antibody detection (CAGTA). This test provides a rapid and simple diagnosis of IC (84.4% sensitivity and 94.7% specificity). The aim of this study is to identify the patients who could be benefited by the use of CAGTA test in critical care setting. Methods: A prospective, cohort, observational multicentre study was carried out in six medical/surgical Intensive care units (ICU) of tertiary-care Spanish hospitals. Candida albicans Germ Tube Antibody test was performed twice a week if predetermined risk factors were present, and serologically demonstrated candidiasis was considered if the testing serum dilution was >= 1: 160 in at least one sample and no other microbiological evidence of invasive candidiasis was found. Results: Fifty-three critically ill non-neutropenic patients (37.7% post surgery) were included. Twenty-two patients (41.5%) had CAGTA-positive results, none of them with positive blood culture for Candida. Neither corrected colonization index nor antifungal treatment had influence on CAGTA results. This finding could corroborate that the CAGTA may be an important biomarker to distinguish between colonization and infection in these patients. The presence of acute renal failure at the beginning of the study was more frequent in CAGTA-negative patients. Previous surgery was statistically more frequent in CAGTA-positive patients. Conclusions: This study identified previous surgery as the principal clinical factor associated with CAGTA-positive results and emphasises the utility of this promising technique, which was not influenced by high Candida colonization or antifungal treatment. Our results suggest that detection of CAGTA may be important for the diagnosis of invasive candidiasis in surgical patients admitted in ICU.

Diagnosis of invasive candidiasis by enzyme-linked immunosorbent assay using the N-terminal fragment of Candida albicans hyphal wall protein I

Background: The diagnosis of invasive candidiasis is difficult because there are no specific clinical manifestations of the disease and colonization and infection are difficult to distinguish. In the last decade, much effort has been made to develop reliable tests for rapid diagnosis of invasive candidiasis, but none of them have found widespread clinical use. Results: Antibodies against a recombinant N-terminal fragment of the Candida albicans germ tube-specific antigen hyphal wall protein 1 (Hwp1) generated in Escherichia coli were detected by both immunoblotting and ELISA tests in a group of 36 hematological or Intensive Care Unit patients with invasive candidiasis and in a group of 45 control patients at high risk for the mycosis who did not have clinical or microbiological data to document invasive candidiasis. Results were compared with an immunofluorescence test to detect antibodies to C. albicans germ tubes (CAGT). The sensitivity, specificity, positive and negative predictive values of a diagnostic test based on the detection of antibodies against the N-terminal fragment of Hwp1 by immunoblotting were 27.8 %, 95.6 %, 83.3 % and 62.3 %, respectively. Detection of antibodies to the N-terminal fragment of Hwp1 by ELISA increased the sensitivity (88.9 %) and the negative predictive value (90.2 %) but slightly decreased the specificity (82.6 %) and positive predictive values (80 %). The kinetics of antibody response to the N-terminal fragment of Hwp1 by ELISA was very similar to that observed by detecting antibodies to CAGT. Conclusion: An ELISA test to detect antibodies against a recombinant N-terminal fragment of the C. albicans germ tube cell wall antigen Hwp1 allows the diagnosis of invasive candidiasis with similar results to those obtained by detecting antibodies to CAGT but without the need of treating the sera to adsorb the antibodies against the cell wall surface of the blastospore.

Postantifungal Effect of Micafungin against the Species Complexes of Candida albicans and Candida parapsilosis

Micafungin is an effective antifungal agent useful for the therapy of invasive candidiasis. Candida albicans is the most common cause of invasive candidiasis; however, infections due to non-C. albicans species, such as Candida parapsilosis, are rising. Killing and postantifungal effects (PAFE) are important factors in both dose interval choice and infection outcome. The aim of this study was to determinate the micafungin PAFE against 7 C. albicans strains, 5 Candida dubliniensis, 2 Candida Africana, 3 C. parapsilosis, 2 Candida metapsilosis and 2 Candida orthopsilosis. For PAFE studies, cells were exposed to micafungin for 1 h at concentrations ranging from 0.12 to 8 mu g/ml. Time-kill experiments (TK) were conducted at the same concentrations. Samples were removed at each time point (0-48 h) and viable counts determined. Micafungin (2 mu g/ml) was fungicidal (>= 3 log(10) reduction) in TK against 5 out of 14 (36%) strains of C. albicans complex. In PAFE experiments, fungicidal endpoint was achieved against 2 out of 14 strains (14%). In TK against C. parapsilosis, 8 mu g/ml of micafungin turned out to be fungicidal against 4 out 7 (57%) strains. Conversely, fungicidal endpoint was not achieved in PAFE studies. PAFE results for C. albicans complex (41.83 +/- 2.18 h) differed from C. parapsilosis complex (8.07 +/- 4.2 h) at the highest tested concentration of micafungin. In conclusion, micafungin showed significant differences in PAFE against C. albicans and C. parapsilosis complexes, being PAFE for the C. albicans complex longer than for the C. parapsilosis complex.

Producción de la proteína recombinante Kre9 del hongo patógeno Candida albicans con actividad proinflamatoria y protumoral.

Candida albicans es un hongo dimórfico capaz de desencadenar una respuesta proinflamatoria mediada por citocinas que incrementa la adhesión de células tumorales al endotelio hepático, favoreciendo así el proceso de metástasis. La proteína Kre9 de C. albicans está relacionada con la citocina IL-1β, la cual participa en la cascada proinflamatoria. Para estudiar el efecto de la proteína Kre9 en la actividad prometastática de C. albicans, esta se clonó en Escherichia coli mediante el vector comercial pETBlue-2; seguidamente se expresó y purificó mediante cromatografía. Los resultados indican que el procedimiento se llevó a cabo correctamente ya que se identificó la presencia de la proteína mediante SDS-PAGE y Western Blot. Por último, se retiraron las endotoxinas de la muestra mediante tratamiento con agarosa-polimixina y se determinó la concentración final de proteína y endotoxinas.

Carbapenem-resistant and carbapenem-susceptible isogenic isolates of Klebsiella pneumoniae ST101 causing infection in a tertiary hospital

Background: In this study we describe the clinical and molecular characteristics of an outbreak due to carbapenem-resistant Klebsiella pneumoniae (CR-KP) producing CTX-M-15 and OXA-48 carbapenemase. Isogenic strains, carbapenem-susceptible K. pneumoniae (CS-KP) producing CTX-M-15, were also involved in the outbreak. Results: From October 2010 to December 2012 a total of 62 CR-KP and 23 CS-KP were isolated from clinical samples of 42 patients (22 had resistant isolates, 14 had susceptible isolates, and 6 had both CR and CS isolates). All patients had underlying diseases and 17 of them (14 patients with CR-KP and 3 with CS-KP) had received carbapenems previously. The range of carbapenem MICs for total isolates were: imipenem: 2 to >32 mu g/ml vs. <2 mu g/ml; meropenem: 4 to >32 mu g/ml vs. <2 mu g/ml; and ertapenem: 8 to >32 mu g/ml vs. <2 mu g/ml. All the isolates were also resistant to gentamicin, ciprofloxacin, and cotrimoxazole. Both types of isolates shared a common PFGE pattern associated with the multilocus sequence type 101 (ST101). The bla(CTX-M-15) gene was detected in all the isolates, whereas the bla(OXA-48) gene was only detected in CR-KP isolates on a 70 kb plasmid. Conclusions: The clonal spread of K. pneumoniae ST101 expressing the OXA-48 and CTX-M-15 beta-lactamases was the cause of an outbreak of CR-KP infections. CTX-M-15-producing isolates lacking the blaOXA-48 gene coexisted during the outbreak.

HPV infection, a described cause for human cancer: HPV detection and molecular mechanisms of HPV carcinogenesis (Review).

[en]Human papillomavirus (HPV) belongs to the Papillomaviridae virus family and it is one of the most common sexual transmission infections. HPV genome is composed of eight genes, including two early genes and six late genes. Among these late genes, E6 and E7 code for proteins that trigger cell-cycle re-entry in infected cells, which can lead to cervical cancer development. The IARC (International Agency for Research Cancer) proposed a guideline based on Hill’s criteria to determine whether the relation between HPV infection and cervical cancer is causal or not. Epidemiological studies have demonstrated that HPV infection is a necessary but non-sufficient cause for cervical cancer. Furthermore, HPV infection is considered the first necessary cause described of a human cancer, being HPV16 and 18 carcinogenic to humans and the most studied types. Cervical cancer is the second leading cause of cancer death among women worldwide. Different screening programs are carried out with the aim of preventing cervical cancer; such as cytologies and HPV tests. There are two main methods which are equally usable to detect HPV: the real-time PCR assays and the array assays. Regarding the molecular mechanisms of HPV mediated malignancies, E2, E6 and E7 proteins of HPV16 lead to immune response evasion, inducing IL-10 and TGF-β1 gene expression. Besides, E6 and E7 proteins allow cell-cycle reentry, phosphorylating RB and ubiquitinating p53 respectively. HPV genome integration in host genome leads to the alteration of host and viral genes expression, including oncogenes and tumor suppressor genes. However, the differences of E6 and E7 oncoproteins in different HPV types is poorly known due to the fact that almost the most studied HPV type has been HPV16.