Comparison of Illumigene, Simplexa, and AmpliVue Clostridium difficile molecular assays for diagnosis of C. difficile infection.

We compared the performance of the Simplexa Universal Direct (Focus Diagnostics) and AmpliVue (Quidel Corporation) assays to that of the Illumigene assay (Meridian Bioscience, Inc.) for the diagnosis of Clostridium difficile infection. Two hundred deidentified remnant diarrheal stool specimens were tested by the Simplexa, AmpliVue, and Illumigene methods. Specimens with discrepant results among the three assays and a representative number of concordant specimens were further evaluated by toxigenic culture. The sensitivity and specificity were 98 and 100% and 96 and 100% for the Simplexa Universal Direct and AmpliVue assays, respectively. Both assays are easy to perform, with rapid turn-around-times, supporting their utility in the clinical laboratory as routine diagnostic platforms.

Anti-biofilm forming and anti-quorum sensing activity of selected essential oils and their main components on food-related micro-organisms.

AIMS: The aim of this study was to investigate the effect of clary sage, juniper, lemon and marjoram essential oils (EOs) and their major components on the formation of bacterial and yeast biofilms and on the inhibition of AHL-mediated quorum sensing (QS). METHODS AND RESULTS: Biofilm formation was measured by crystal violet and resazurin staining, and QS inhibition was detected by paper disc diffusion assay. Marjoram EO inhibited Bacillus cereus, Pichia anomala, Pseudomonas putida and mixed-culture biofilm formation of Ps. putida and Escherichia coli and showed the best QS inhibitor effect on Chromobacterium violaceum. For B. cereus, all components showed better antibiofilm capacity than the parent EOs. Lemon EO inhibited E. coli and mixed-culture biofilms, and cinnamon was effective against the mixed forms. Scanning electron microscopy showed the loss of three-dimensional structures of biofilms. CONCLUSIONS: The EOs and components used seem to be good candidates for prevention of biofilm formation and inhibition of the AHL-mediated QS mechanism. SIGNIFICANCE AND IMPACT OF THE STUDY: Biofilm formation on foods and food industrial equipment is a serious problem causing food spoilage and emergence of foodborne diseases. This article highlights the importance of studying EOs as potential disinfectants and food preservatives.

Disparate changes in the expression of transient receptor potential vanilloid type 1 receptor mRNA and protein in dorsal root ganglion neurons following local capsaicin treatment of the sciatic nerve in the rat.

In situ hybridization, quantitative reverse transcription polymerase chain reaction (RT-PCR), immunohistochemistry, and Western blot analysis were applied to study the changes in expression of the major nociceptive ion channel transient receptor potential vanilloid type 1 receptor (TRPV1) after the perineural application of capsaicin or nerve transection. In control rats, quantitative morphometric and statistical analyses of TRPV1 protein and mRNA expression in L5 dorsal root ganglion cells revealed distinct populations of small (type C) and small-to-medium (type B) neurons, which showed very high and moderate levels of TRPV1, whereas larger (type A) neurons mostly did not express this receptor. After either transection or capsaicin treatment of the sciatic nerve, immunohistochemistry and Western blotting demonstrated a massive (up to 80%) decrease in the proportion of TRPV1-immunoreactive neurons and TRPV1 protein at all postoperative survival times. In situ hybridization indicated marked decreases (up to 85%) in the proportion of neurons that expressed TRPV1 mRNA after sciatic nerve transection. In contrast, although perineural treatment with capsaicin resulted in similar substantial decreases in the proportions of type B and C neurons of the L5 dorsal root ganglia 3 days postoperatively, a clear-cut tendency to recovery was observed thereafter. Hence, the proportions of both type B and C neurons expressing TRPV1 mRNA reached up to 70% of the control levels at 30 days postoperatively. In accord with these findings, quantitative RT-PCR revealed a marked and significant recovery in TRPV1 mRNA after perineural capsaicin but not after nerve transection. These observations suggest the involvement of distinct cellular mechanisms in the regulation of the TRPV1 mRNA expression of damaged neurons, specifically triggered by the nature of the injury. The present findings imply that the antinociceptive and anti-inflammatory effects of perineurally applied capsaicin involve distinct changes in neuronal TRPV1 mRNA expression and long-lasting alterations in (post)translational regulation.

Serial quantitative PCR analysis of bone marrow samples from breast cancer patients to monitor systemic micrometastases.

BACKGROUND: We have previously developed a quantitative calibrated PCR assay to measure cytokeratin 19 (CK19) expression in haematopoietic tissue in order to detect systemic micrometastases. PATIENTS AND METHODS: Serial measurements of CK19 expression in bone marrow of patients with primary breast cancer were performed at operation, at 3 weeks and 6 months postoperatively. RESULTS: Reference values for CK19 expression were established by analysing bone marrow samples from 48 healthy female volunteers or patients without epithelial cancer. Samples from breast cancer patients with CK19 values above the upper reference limit were considered positive. Bone marrow samples taken at operation were positive in 29 out of 141 patients (20.6%) and remained positive in 12, turned negative in 4 and were unavailable in 13 at 6 months postoperatively. CONCLUSION: Serial measurements increase the reliability of detecting micrometastases perioperatively. Further studies are in progress to evaluate the relationship between elevated CK19 values and clinical outcome.

A synthetic gamma-lactone group with beta-lactamase inhibitory and sporulation initiation effects.

Previous studies showed that some lactones have beta-lactamase inhibitory or antibacterial effects, others--like A-factor (a gamma-butyrolactone) and its derivatives--stimulate sporulation in Streptomyces griseus strains. Our experiments were aimed at exploring whether synthetic gamma-lactones had such effects. None of the seven gamma-lactones studied showed antibacterial activity, but two of them inhibited beta-lactamases isolated from various bacteria. These two gamma-lactones did not reduce colony formation of murine bone marrow cells in vitro, indicating that they were not toxic to proliferating mammalian cells. Four gamma-lactones, including the two inhibiting beta-lactamase, stimulated sporulation in the non-sporulating S. griseus bald 7 mutant. Further studies of gamma-lactones as potential inhibitors of beta-lactamase seem to be warranted.

Evidence of a role for NAD+-glycohydrolase and ADP-ribosyltransferase in growth and differentiation of Streptomyces griseus NRRL B-2682: inhibition by m-aminophenylboronic acid.

m-Aminophenylboronic acid (APBA) inhibited the germination, growth and sporulation of Streptomyces griseus NRRL B-2682 in an age- and concentration-dependent manner in submerged and solid cultures. When added to cells or cell extracts it irreversibly inhibited NAD+-glycohydrolase and ADP-ribosyltransferase activity. ADP-ribosyltransferase was more sensitive, but inhibition was not complete, even in the presence of 10 mM APBA. The in vivo effects of the inhibitor correlated with its in vitro effect on ADP-ribosylation and on the profile of ADP-ribosylated endogenous proteins. The physiological importance of ADP-ribosyltransferase was supported by the observation that APBA strongly inhibited the growth of a non-sporulating and NAD+- glycohydrolase-negative mutant of the parental strain. The resistance of S. griseus NRRL B-2682 strains able to grow in the presence of APBA was due to permeability factors. A comparison of the ADP-ribosylated protein profiles of S. griseus NRRL B-2682 grown under various conditions showed similarities, but also specific differences. The results suggest that the ADP-ribosyltransferase of S. griseus NRRL B-2682 is an indispensable enzyme for growth and differentiation of the strain. It may regulate the activity of key enzymes or developmental proteins by responding to intra- and extracellular conditions.