Extended characterization of Corynebacterium pyruviciproducens based on clinical strains from Canada and Switzerland.

The species Corynebacterium pyruviciproducens was described in 2010 based on the features of a single strain. In this report, we describe the chemotaxonomic and phenotypic characteristics of 11 C. pyruviciproducens clinical strains isolated in Switzerland and Canada in comparison to the strain 06-17730(T). Heterogeneities within the type strain were found in the 16S rRNA gene and in biochemical markers. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) identification of this species could not be achieved since currently this bacterial species is not included in the corresponding database. Reliable identification is obtained only with sequence-based identification tools. Results of antimicrobial susceptibility testing of this species with an extended panel of antimicrobials are presented here for the first time.

Identification of a novel 16S rRNA gene variant of Actinomyces funkei from six patients with purulent infections.

Little is known about the clinical significance and laboratory diagnosis of Actinomyces funkei. In this report we describe six clinical cases where A. funkei was isolated from purulent, polymicrobial infections. Conventional identification procedures were compared with molecular methods including matrix-assisted laser desorption/ionization time-of-flight mass spectrometry technique. Analysis of the full 16S rRNA gene sequence of the six investigated strains revealed differences from the A. funkei type strain. DNA-DNA hybridization showed that the clinical strains represent a novel 16S rRNA gene variant within the species of A. funkei.

Comparison of the DiversiLab repetitive element PCR system with spa typing and pulsed-field gel electrophoresis for clonal characterization of methicillin-resistant Staphylococcus aureus.

The emergence of methicillin-resistant Staphylococcus aureus (MRSA) has become an increasing problem worldwide in recent decades. Molecular typing methods have been developed to identify clonality of strains and monitor spread of MRSA. We compared a new commercially available DiversiLab (DL) repetitive element PCR system with spa typing, spa clonal cluster analysis, and pulsed-field gel electrophoresis (PFGE) in terms of discriminatory power and concordance. A collection of 106 well-defined MRSA strains from our hospital was analyzed, isolated between 1994 and 2006. In addition, we analyzed 6 USA300 strains collected in our institution. DL typing separated the 106 MRSA isolates in 10 distinct clusters and 8 singleton patterns. Clustering analysis into spa clonal complexes resulted in 3 clusters: spa-CC 067/548, spa-CC 008, and spa-CC 012. The discriminatory powers (Simpson's index of diversity) were 0.982, 0.950, 0.846, and 0.757 for PFGE, spa typing, DL typing, and spa clonal clustering, respectively. DL typing and spa clonal clustering showed the highest concordance, calculated by adjusted Rand's coefficients. The 6 USA300 isolates grouped homogeneously into distinct PFGE and DL clusters, and all belonged to spa type t008 and spa-CC 008. Among the three methods, DL proved to be rapid and easy to perform. DL typing qualifies for initial screening during outbreak investigation. However, compared to PFGE and spa typing, DL typing has limited discriminatory power and therefore should be complemented by more discriminative methods in isolates that share identical DL patterns.

Prognostic value of genomic alterations in invasive cervical squamous cell carcinoma of clinical stage IB detected by comparative genomic hybridization.

The clinical behavior of invasive cervical carcinoma of clinical stage IB varies considerably in tumors presenting without regional lymph node metastases. The early identification of patients at higher risk for poor outcome may prove useful because these patients would benefit from aggressive adjuvant treatments. In this study, comparative genomic hybridization was applied to evaluate whether genomic aberrations have prognostic significance in cervical carcinoma. Genomic alterations were evaluated in 62 cervical carcinomas of clinical stage IB. DNA sequence losses were most prevalent at chromosomes 4q (53%), 3p (52%), 13q (45%), 4p (44%), Xq (44%), 5q (40%), 18q (37%), and 6q (35%). Several genomic alterations were associated with poor clinical outcome or metastasis. The total number of DNA aberrations/tumor (P < 0.02) and the number of DNA sequence losses/tumor (P < 0.04) were associated with disease-specific survival. 9p deletions were significantly more frequent in carcinomas with lymph node metastasis than in node-negative tumors (P < 0.03). Losses of chromosome 11p (P < 0.0001) and 18q (P < 0.01) were associated with poor prognosis in cervical carcinomas without lymph node metastasis. These data suggest that inactivation of tumor suppressor genes on chromosomes 9p, 11p, and 18q may play a role in the progression of cervical carcinoma.

Expression of p150 in cervical neoplasia and its potential value in predicting survival.

BACKGROUND: A recently cloned novel p150 protein was found to be overexpressed in human breast carcinoma. To the authors' knowledge, no data on p150 expression in any other human tumors have been published previously. METHODS: To investigate whether the expression of p150 correlated with the clinicopathologic stages of cervical neoplasms or the prognoses of patients with these neoplasms, the authors conducted an immunohistochemical study of archival formalin fixed, paraffin embedded specimens. Seventy-two precancerous lesions (CIN), 75 clinical Stage IB invasive squamous carcinomas, and 20 samples of normal squamous epithelia were included. In addition to p150, the Ki-67 labeling index was assessed as a proliferation parameter. The presence of human papillomavirus was analyzed by in situ DNA hybridization. RESULTS: A significant association of p150 with the grade of atypia in cervical neoplasms was demonstrated. The highest expression of p150 was observed in low grade CIN, with subsequently decreasing expression in high grade CIN and invasive carcinoma. For patients with invasive carcinoma, p150 was significantly correlated with clinical outcome. Patients with high expression of p150 had a better prognosis than those with low p150. Those with regional lymph node metastasis and significant p150 expression had longer relapse free survival than those with insignificant p150 expression. Women whose carcinomas demonstrated vascular space invasion or high microvessel density survived longer when p150 was clearly expressed. p150 behaves as a potential tumor marker during early cervical carcinoma development and is later turned off as cells proceed to more advanced stages of their malignant phenotypes. CONCLUSIONS: p150 is a molecular parameter that might become useful in predicting disease progression and determining the prognoses of patients with invasive cervical carcinoma.

Association of p27Kip1, cyclin E and c-myc expression with progression and prognosis in HPV-positive cervical neoplasms.

Recent studies demonstrated that a variety of human cancer cell lines express relatively high levels of p27Kip1 and that this might be associated with increased expression of Cyclin E. There is a feedback inhibitory loop between Cyclin E and p27Kip1, which can be counteracted by elevated c-myc activation. This study analyzed by immunohistochemistry the expression of p27Kip1, Cyclin E and c-myc in a series of HPV-positive cervical tissue samples representing various stages of cervical carcinogenesis, using 13 samples of normal epithelium, 24 low-grade CIN, 63 high-grade CIN, and 69 samples of invasive squamous cell carcinoma. To evaluate the cell proliferation, the Ki-67 Labelling Index (LI) was assessed. The presence of HPV was investigated by in situ DNA hybridization. We did not find any correlation between p27Kip1 expression and Ki-67 LI in normal and tumor tissue samples. There was evidence for an increase of p27Kip1 levels from low-grade to high-grade CIN. Cyclin E, c-myc and the Ki-67 LI were significantly increased during cervical carcinogenesis. Cyclin E and c-myc were positively correlated to cell proliferation in pre-cancerous lesions, but not related to overall survival in invasive carcinomas. Contrary to that, high levels of p27Kip1 are associated with poor overall survival in invasive cervical carcinomas of clinical stage IB. This may reflect the counteracting function of c-myc in blocking p27Kip1, thus representing the worst condition of a disturbed tumor cell cycle in cervical carcinoma, ultimately induced by HPV.

Angiogenesis in cervical neoplasia: microvessel quantitation in precancerous lesions and invasive carcinomas with clinicopathological correlations.

Recently, angiogenic properties have been shown in preinvasive cervical lesions. Our goal was to determine the angiogenesis in cervical intraepithelial neoplasia (CIN) and the relationship between microvessel counts, histopathological parameters, and clinical outcome in invasive cervical carcinoma. One hundred thirty-eight cervical specimens were evaluated; among these 20 were designated normal epithelium, 20 low-grade CIN, 40 high-grade CIN, and 58 invasive carcinoma. Histological sections immunostained for CD31 were quantitatively evaluated for microvessel density. The tumor proliferation rate was determined by the Ki-67 Labeling Index. Comparison of microvessel counts from normal epithelium with those from CIN and invasive carcinoma showed significant increases in precancerous lesions and invasive cancer (P < 0.0001). Microvessel density was found to be associated with the overall survival in women with invasive carcinoma (P < 0.01). There was a significant correlation of microvessel density (P < 0.05) with relapse-free survival in patients with regional lymph node metastasis. A Cox stepwise regression analysis revealed microvessel density, together with depth of invasion, regional lymph node status, and vascular invasion, to be a strong independent prognostic indicator for overall survival in patients with clinical stage IB cervical carcinoma.

Altered expression of mdm-2 and its association with p53 protein status, tumor-cell-proliferation rate and prognosis in cervical neoplasia.

Recent results suggest that p53 inactivation is required for cervical-carcinoma development. The mdm-2 oncogene, which forms an auto-regulatory feedback loop with the normal p53 protein, has been found amplified in human carcinomas, thus abolishing the anti-proliferative function of p53. To investigate whether the mdm-2/p53 interaction plays a role in cervical neoplasms, we performed an immunohistochemical study in archival fixed, embedded specimens that included 178 pre-cancerous lesions (CIN) and invasive squamous-cell carcinomas of clinical stage IB. In addition to p53, we assessed the p53-associated protein, mdm-2, and the Ki-67 labelling index (LI). The presence of HPV was assessed by in situ DNA hybridization. Tumor expression of all nuclear proteins was scored as fraction of positive CIN or cancer nuclei. The analysis demonstrated a significant association of the Ki-67 LI with grade of atypia in cervical neoplasms. p53 accumulation and mdm-2 expression are higher in invasive carcinomas than in pre-cancerous lesions. No correlation was observed with HPV status. An inverse correlation was found between increased tumor-cell proliferation and mdm-2 expression in invasive carcinomas (p < 0.0001). mdm-2 expression was significantly associated with p53 accumulation (p < 0.02). However, the investigated nuclear proteins were not associated with overall survival in patients with invasive carcinomas. Cox stepwise-regression analysis revealed regional lymph node status and depth of invasion to be independent parameters.

Investigation of the Bcl-2 and C-myc expression in relationship to the Ki-67 labelling index in cervical intraepithelial neoplasia.

This is the investigation of the relationship between the neoplastic cell proliferation and the expression of bcl-2 and c-myc in human papillomavirus (HPV)-negative and HPV-positive cervical intraepithelial neoplasms (CIN). The expression of bcl-2 and c-myc was studied using quantitative immunohistochemistry in 20 specimens of normal cervical squamous epithelium and 73 specimens of CIN. To analyze the neoplastic cell proliferation rate, the Ki-67 labelling index was determined; the latter was significantly different between normal epithelium and various grades of CIN (p < 0.0001). The highest proliferation rate was found in high-grade CIN. In precancerous lesions, we found the number of bcl-2 positive cells lower than in normal epithelium, but with a significant difference between low-grade and high-grade CIN (p < 0.0001). The highest percentage of bcl-2 positive neoplastic cells was found in high-grade CIN. C-myc was rarely expressed in normal epithelium. Similar to the Ki-67 labelling index, c-myc immunostaining correlated with the histological grade of CIN, with the highest percentage of c-myc positive nuclei occurring in high-grade CIN (p < 0.0001). In contrast to bcl-2 immunoreactivity, the c-myc significantly was more expressed in high-risk HPV-positive than in HPV-negative lesions. The c-myc expression in CIN is closely related to the neoplastic cell proliferation rate. With progression of intraepithelial neoplasia, bcl-2 production in neoplastic cells increases with high co-expression of c-myc.

Association of tumor induced vascularization with clinicopathological parameters in cervical neoplasm.

Angiogenic properties have been shown in preinvasive cervical lesions. Our goal was to determine the angiogenesis in cervical intraepithelial neoplasms (CIN), the relationship between microvessel counts, histopathological parameters and the clinical outcome in invasive cervical carcinoma. Comparison of microvessel counts from normal epithelium with that from CIN and invasive carcinoma showed significant increases in pre-cancerous lesions and invasive cancer (p < 0.0001). Microvessel density, assessed by CD31 immunostaining, was found to be associated with the overall survival in women with clinical stage IB cervical carcinoma (p < 0.03). There was a significant association of microvessel density (p < 0.05) with relapse-free survival in patients with regional lymph node metastasis.

Analysis of proliferative activity using Ki-67 on cervical precancerous lesions and the relationship to p53 expression.

Neoplastic cell growth rate and the p53 expression have been recently analysed in invasive cervical carcinoma. Samples of 20 specimens with normal cervical epithelium and 73 specimens of dysplasia and carcinoma in situ (CIS) were immunostained with monoclonal antibodies to p53 and Ki-67 to examine the interrelationship between p53, Ki-67 and HPV status in cervical intraepithelial neoplasm. The presence of HPV was assessed by in situ DNA hybridization. Of dysplasias and CIS 79% were HPV positive. The growth rate of neoplastic cells was significantly correlated to the histological grade and the HPV status. The highest proliferation was found in poorly differentiated HPV 16/18 positive precancerous lesions. The analysis of the p53 expression showed no difference between various histological grades. However, the p53 oncoprotein was expressed significantly lower in HPV 16/18 positive neoplasms. The assessment of neoplastic cell growth rate offers a potentially valuable approach to predicting biological behaviour in intraepithelial neoplasms.

Expression of CD44 and variant isoforms in cervical intraepithelial neoplasia.

The cell adhesion molecule CD44 and its variant isoforms have been found to be related to invasive and metastatic character of cancer cells. Their expression in gynecologic precancerous lesions has not yet been reported. Mouse monoclonal antibodies directed against a common epitope (CD44s) and exons 4v, 6v, and 9v were used to study the expression of CD44 and variant isoforms by immunohistochemistry in cervical intraepithelial neoplasia (CIN). Twenty tissue samples with normal cervical epithelium and 57 samples with CIN of different histological grades and different HPV status were included in this study. The standard CD44, CD44-4v, CD44-6v, and CD44-9v were expressed in normal cervical epithelium and in precancerous lesions. In distinct contrast to the normal epithelium, however, the standard CD44, CD44-4v, and 6v showed a reduced expression in precancerous lesions, whereas CD44-9v was significantly overexpressed. Expression of CD44 standard and CD44-4v was correlated with the histological grade but not with the HPV status. Compared with mild and moderate dysplasia, severe dysplasia and carcinoma in situ are associated with low expression of CD44s (P = 0.007) and of CD44-4v (P = 0.03). These observations reveal dynamic changes in CD44 expression during neoplastic cell transformation in cervical intraepithelial neoplasia.

Assessment of EGFR and TGF-alpha expression in relationship to HPV status and Ki-67 distribution in cervical intraepithelial neoplasms.

Expression of epidermal-growth-factor receptor (EGFR), transforming growth factor alpha (TGF-alpha) and Ki-67 proliferation antigen in cervical intra-epithelial neoplasms were analyzed. To examine the interrelationship of TGF-alpha, EGFR, Ki-67 and HPV status in dysplasia and carcinoma in situ, formalin-fixed tissue sections of 92 women were immunostained with monoclonal antibodies to EGFR, TGF-alpha and Ki-67. The presence of HPV was assessed by in situ DNA hybridization. The highest positive TGF-alpha expression was seen in the group of mild dysplasia. The difference was significant between the relatively high expression in mild dysplasia and the low occurrence in severe dysplasia and carcinoma in situ as well. The same relation could be found between TGF-alpha expression in papilloma-virus-negative dysplasia and those with the presence of HPV 16/18. In contrast to these findings, the Ki-67 proliferation marker was intensely detectable in severe dysplasia and carcinoma in situ. Ki-67-stained neoplastic cell nuclei were found in a significantly higher percentage of HPV-positive than in HPV-negative lesions. TGF-alpha over-expression is obviously combined with low proliferating activity and vice versa. Irrespective of the grade of dysplasia or HPV status, EGFR was expressed abnormally as compared with normal squamous epithelium. Over-expression of TGF-alpha in mild dysplasia could be associated with the autocrine pathway of cell-growth regulation. In the presence of HPV 16/18 the EGFR/TGF-alpha pathway for growth stimulation is probably not involved.

Nuclear p53 overexpression is an independent prognostic parameter in node-negative non-small cell lung carcinoma.

The prognosis of operated patients with non-small cell lung cancer (NSCLC) is poor despite thorough pre-operative staging. An improved preselection is needed of patients likely to profit from surgery. This study was undertaken to evaluate the prognostic significance of nuclear p53 overexpression in a cohort of 247 surgically treated patients with NSCLC. It showed that the prevalence of immunohistochemically detectable p53 overexpression varied between different tumour types. p53 overexpression was equally frequent in large cell carcinoma (53 per cent) and in squamous cell carcinoma (54 per cent), but significantly less frequent in adenocarcinoma (34 per cent; P = 0.009). p53 overexpression was particularly rare in bronchioloalveolar carcinoma (positivity in 1 of 17 cases). These variations may reflect aetiological differences between the histological subtypes. p53 overexpression was also associated with high tumour grade (P = 0.0157) and the presence of lymph node metastasis (P = 0.0259), but not with advanced tumour stage. Survival analysis showed no difference in clinical outcome between p53-positive and p53-negative tumours within 101 node-positive tumours. In contrast, survival time was significantly better in p53-negative tumours than in p53-positive tumours within the group of 113 node-negative tumours (P = 0.032). Stepwise regression analysis showed that p53 overexpression is an independent prognostic factor in node-negative NSCLC.

[Hypersensitivity to levobunolol]. / Uberempfindlichkeit gegenüber Levobunolol.

A case is described in which, following local application of glaucoma eye drops containing Levobunolol, a recurring allergic contact eczema with periocular localisation occurred. In the course of the epicutaneous allergy testing a positive reaction (crescendo-type) could be clearly demonstrated experimentally against both the eye drops and their active ingredient, levobunolol-hydrochloride, a beta-blocker (beta-sympatholyticum) out of the class of the aryloxypropylamines.