Device-independent, real-time identification of bacterial pathogens with a metal oxide-based olfactory sensor

A novel olfactory method for bacterial species identification using an electronic nose device called the MonoNose was developed. Differential speciation of micro-organisms present in primary cultures of clinical samples could be performed by real-time identification of volatile organic compounds (VOCs) produced during microbial replication. Kinetic measurements show that the dynamic changes in headspace gas composition are orders of magnitude larger than the static differences at the end of fermentation. Eleven different, clinically relevant bacterial species were included in this study. For each of the species, two to eight different strains were used to take intra-species biodiversity into account. A total of 52 different strains were measured in an incubator at 37°C. The results show that the diagnostic specificities varied from 100% for Clostridium difficile to 67% for Enterobacter cloacae with an overall average of 87%. Pathogen identification with a MonoNose can be achieved within 6–8 h of inoculation of the culture broths. The diagnostic specificity can be improved by broth modification to improve the VOC production of the pathogens involved

Fluconazole resistance in Candida albicans: a review of mechanisms.

Antifungal agents have greatly contributed to the improvement of public health. Nevertheless, antifungal resistant pathogens have increased during the past decade, becoming a serious concern. Candida albicans has been the most extensively studied pathogen in antifungal resistance because of their morbidity and mortality associated with infections in immunocompromised patients. This review describes the antifungal mechanims of the azole fluconazole widely used for the prophylaxis and treatment of candidal infections. The specific molecular pathways occurring in fluconazole-resistance of C. albicans and some issues about new antifungal agents are also discussed

Cytokine pattern secretion by murine spleen cells after inactivated Candida albicans immunization. Effect of cocaine and morphine treatment

In the present work we have analyzed: i) the effect of heat-inactivated Candida albicans immunization on the cytokine production by murine spleen cells; ii) the effect of a subchronic cocaine and morphine treatment on this production. The treatment with a single dose of inactivated Candida blastospores induced interleukin-2 (IL-2), interferon-gamma (IFN gamma) and interleukin-4 (IL-4) production at 24 h after in vitro restimulation of splenocytes. In this model, the exposure to morphine (25 mg/kg, 5 days before, during and 5 days after inoculation with the yeast) significant decreased IL-2 and IL-4 levels, while secretion of IFN-gamma was unaltered. The same cocaine treatment (10 mg/kg) resulted in unchanged levels of the three cytokines tested. The results showed that non-viable Candida cells of this strain induce a predominant Th0 response. This immune effect is in part impaired only by a subchronic administration of morphine

Evaluation of the antifungal effect of EDTA, a metal chelator agent, on Candida albicans biofilm.

OBJECTIVE: Candida albicans biofilm is frequently found on artificial surfaces and the infections related to biofilm are difficult to eliminate, as they require the removal of artificial devices and treatment with antifungal drugs. Nowadays, fungal growth in biofilms is difficult to eradicate with conventional antifungal drugs such as fluconazole. Among chelating agents, disodium salt-Ethylene Diamine Tetraacetic Acid (EDTA) is known to have antifungal activity. In this study, we examined the in vitro activity of the EDTA and the antifungal drug fluconazole against C. albicans mature biofilm. MATERIALS AND METHODS: C. albicans ATCC 20191, fluconazole-susceptible strain, was grown at an inoculum starter of 1 x 106 cells/ml for 72 h in 24-well microtiter plates and was further treated for 24 h with EDTA and/or fluconazole. Antifungal activities in biofilms were expressed as reduction in optical density (OD) determined by a 2,3-bis (2-methoxy-4-nitro-5-sulfophenyl)- 2H-tetrazolium-5-carboxanilide (XTT) colorimetric assay and compared to untreated biofilms. RESULTS: Colorimetric readings revealed that EDTA alone (at 25 and 2.5 mM) significantly reduced fungal metabolic activity in preformed biofilms. Also, EDTA combined with fluconazole significantly reduced the growth of biofilm when compared to biofilm treated with fluconazole alone (at 25 and 2.5 μg/ml). CONCLUSIONS: Our data suggest that the employment of EDTA or other chemicals destabilizers of the biofilm matrix, in combination with antifungal drugs, could lead to the development of new strategies for the management of infections associated to Candida biofilm. Another relevant result of our study suggests that the initial cell concentration, probably through mechanisms of quorum sensing, affects the cellular viability during the process of biofilm formation

Application of electronic noses for disease diagnosis and food spoilage detection

Over the last twenty years, newly developed chemical sensor systems (so-called "electronic noses") have odour analyses made possible. This paper describes the applications of these systems for microbial detection in different fields such as medicine and the food industry, where fast detection methods are essential for appropriate management of health care. Several groups have employed different electronic noses for classification and quantification of bacteria and fungi to obtain accurate medical diagnosis and food quality control. So far, detection and identification of bacterial and fungal volatiles have been achieved by use of e-noses offering different correct classification percentages. The present review includes examples of bacterial and fungal species producing volatile compounds and correlated to infectious diseases or food deterioration. The results suggest the possibility of using this new technology both in medical diagnostics and in food control management. © 2006 by MDPI

INHIBITORY EFFECT OF MORPHINE ON THE CONTROL OF RAT FIBROBLAST CELL-PROLIFERATION

The G(0)/G(1) to S-phase transition in quiescent EL2 rat fibroblast cells stimulated by mitogen (such as the epidermal growth factor) can be blocked by the addition of morphine to the system. The drug must be actively present when quiescent EL2 cells are induced to enter the proliferative slate. Even when morphine is added after mitogenic stimulus, an exposure only within 6 hours is required to inhibit DNA synthesis. These results suggest that morphine can directly influence the proliferation of the nonlymphoid cell system, particularly during the establishment oi a competence state (i.e., G(0)/S phase transition)

Experimental use of a new surface acoustic wave sensor for the rapid identification of bacteria and yeasts.

AIMS:Use of an electronic nose (zNose(TM)) to discriminate between volatile organic molecules delivered during bacterial/fungal growth on agar and in broth media. METHODS AND RESULTS: Cultures of bacteria (Klebsiella pneumoniae, Pseudomonas aeruginosa, Escherichia coli) and yeasts (two Candida albicans strains) were grown on agar and in broth media and incubated for 24 h at 37 degrees C. Headspace samples from microbial cultures were analysed by the zNose(TM), a fast gas chromatography-surface acoustic wave detector. Olfactory images of volatile production patterns were observed to be different for the various species tested after 24 h. Moreover, some strains (two K. pneumoniae, two C. albicans) did not show changes in volatile production patterns within our species. CONCLUSIONS: Our experiments demonstrate that the electronic nose system can recognize volatile production patterns of pathogens at species level. SIGNIFICANCE AND IMPACT OF THE STUDY: Our results, although preliminary, promise exciting challenges for microbial diagnostics

Determination of the in vitro activity of fluconazole against yeast strains using HPLC.

The aim of this study was to evaluate the in vitro activity of fluconazole against two yeast strains by determining the fluconazole content via high-performance liquid chromatography (HPLC). Candida krusei ATCC 6258 and Saccharomyces cerevisiae were tested for fluconazole susceptibility by dilution tests. Fluconazole measurements by HPLC were performed in cultures of the strains at 6, 12, 24 and 48 h. In C. krusei ATCC 6258 cultures, 88.94% of the fluconazole was recovered at 48 h with respect to the level at time 0 h. The drug exhibited major activity at 12 and 24 h. An increase in cell number and elevated concentration of the drug were detected at 48 h. In S. cerevisiae cultures, the measurements revealed a fluconazole content of approximately 51.25% at 12, 24 and 48 h with respect to the level at time 0 h. However, the percentage of fluconazole detected and the minimal inhibitory concentration (MIC) value determined by agar dilution tests suggest a potential for fluconazole resistance in this susceptible strain. The decreases in fluconazole in C. krusei ATCC 6258 and S. cerevisiae cultures at each time point suggest a mechanism of resistance that is not correlated to efflux pumps

(1-3)-β-D-Glucan vs Galactomannan Antigen in Diagnosing Invasive Fungal Infections (IFIs).

Invasive fungal infections (IFIs) are serious and often life-threatening complications in patients with haematological malignancies. Early diagnosis and the initiation of efficacious antifungal treatments could affect the prognosis of these patients. The detection of (1-3)-β-D-Glucan (BDG) could be a promising non-culture-based, noninvasive tool for IFI analyses in haemato-oncological patients, allowing the diagnosis of the two major IFIs, invasive aspergillosis (IA) and invasive candidiasis (IC), with a single test. The aim of this work was to evaluate and compare the use of the BDG in combination with the galactomannan antigen (GAL) assay in order to exclude or confirm suspected IFIs. Sera from 46 haemato-oncological patients (24 with proven/probable IFI and 22 without IFI symptoms) were evaluated retrospectively for the detection of GAL and BDG. In 24 patients, the serum BDG levels facilitated IFI diagnosis: 18 probable IA, 3 proven IA and 3 IC. In the remaining 22 patients, the BDG level helped exclude IFIs. The BDG was positive earlier than GAL in 5/24 cases [three of probable invasive aspergillosis (IA), one of proven IA and one case of proven invasive candidiasis (IC)] and was positive at the same time as GAL in 19/24 cases; in no case was GAL positive before BDG was. The BDG detection is useful, however, the test has a great limitation because it is a completely manual procedure