Background: The Src family tyrosine kinases (SFK) are cellular regulatory proteins that influence cell adhesion, proliferation, invasion and survival during tumor development. Elevated activity of Src was associated with increased cell proliferation and invasivity in human papillomavirus (HPV)-associated malignancies; therefore, transduced human foreskin keratinocytes (HFK) were used to investigate whether SFK activation is a downstream effect of papillomaviral oncoproteins. Activation of ubiquitously expressed SFKs, namely Src, Yes and Fyn, was investigated in both proliferating and differentiating keratinocytes.Results: In proliferating keratinocytes, Src, Yes and Fyn mRNA levels were not affected by HPV 16 E6 or E7 oncoproteins, while at the protein level as detected by western blot, the presence of both E6 and E7 resulted in substantial increase in Src and Yes expression, but did not alter the high constitutive level of Fyn. Phospo-kinase array revealed that all ubiquitously expressed SFKs are activated by phosphorylation in the presence of HPV 16 E7 oncoprotein. Keratinocyte differentiation led to increased Yes mRNA and protein levels in all transduced cell lines, while it did not influence the Src transcription but resulted in elevated Src protein level in HPV16 E7 expressing lines.Conclusions: This study revealed that HPV 16 oncoproteins upregulate Src family kinases Src and Yes via posttranscriptional mechanisms. A further effect of HPV 16 E7 oncoprotein is to enhance the activating phosphorylation of SFKs expressed in keratinocytes.

Background: The human papillomavirus (HPV) life cycle is closely linked to keratinocyte differentiation. Oncogenic HPV infection has been shown to hamper the normal differentiation of keratinocytes; however, the underlying mechanisms responsible for this phenomenon are yet to be clarified. Here, we aimed to study the effects of HPV16 E6 and E7 oncogenes on the expression of involucrin (IVL), an established marker of keratinocyte differentiation, in human foreskin keratinocyte (HFK) cells. Results: The differentiation of HFK cells by serum and high calcium significantly increased both the mRNA and the protein levels of IVL. The E6 and E7 oncoproteins of HPV16 together caused strong down-regulation of IVL mRNA and protein both in proliferating and in differentiating HFK cells. To study the effects of HPV oncogenes on the IVL promoter, we made transient transfection assays and luciferase tests and found that HPV 16 E6 but not E7 repressed IVL promoter activity in proliferating HFK cells. The inhibitory effect of HPV 16 E6 on the human IVL promoter could be localised to the proximal regulatory region (PRR) of the gene. Conclusions: These results suggest that the down-regulation of IVL promoter activity by HPV 16 E6 significantly contribute to the inhibition of endogenous IVL expression by the HPV 16 oncoproteins. In contrast, the down-regulation of endogenous IVL expression by HPV16 E7 is probably not caused by a direct and specific effect of E7 on the IVL promoter.

Antimicrobial resistance (AMR) is an increasing public health problem worldwide. We studied some patient-related factors that might influence the antimicrobial resistance. and whether the volume of antibiotic prescribing of the primary care physicians correlate with the antibiotic resistance rates of commensal nasal Staphylococcus aureus and Streptococcus pneumoniae. The socio-demographic questionnaires, the antibiotic prescription and resistance data of commensal nasal S. aureus and S. pneumoniae were collected in the 20 participating Hungarian practices of the APRES study. Multivariate logistic regression analyses were performed on the patient-related data and the antimicrobial resistance of the S. aureus and S. pneumoniae on individual, patient level. Ecological analyses were performed with Spearman’s rank correlations at practice level, the analyses were performed in the whole sample (all practices) and in the cohorts of primary care practices taking care of adults (adult practices) or children (paediatric practices). According to the multivariate model, age of the patients significantly influenced the antimicrobial resistance of the S. aureus (OR = 0.42, p = 0.004) and S. pneumoniae (OR = 0.89, p < 0.001). Living with children significantly increased the AMR of the S. pneumoniae (OR = 1.23, p = 0.019). In the cohorts of adult or paediatric practices, neither the age nor other variables influenced the AMR of the S. aureus and S. pneumoniae. At practice level, the prescribed volume of penicillins significantly correlated with the resistance rates of the S. aureus isolates to penicillin (rho = 0.57, p = 0.008). The volume of prescribed macrolides, lincosamides showed positive significant correlations with the S. pneumoniae resistance rates to clarithromycin and/or clindamycin in all practices (rho = 0.76, p = 0.001) and in the adult practices (rho = 0.63, p = 0.021). The age is an important influencing factor of antimicrobial resistance. The results also suggest that there may be an association between the antibiotic prescribing of the primary care providers and the antibiotic resistance of the commensal S. aureus and S. pneumoniae. The role of the primary care physicians in the appropriate antibiotic prescribing is very important to avoid the antibiotic resistance.

Two Balb/C mouse models of Candida infection were used to detect serum interleukin-6 (IL-6) responses. The first model used systemic infection by Candida albicans ATCC 10231 strain infected through the lateral tail vein of mice without any specific pretreatment. The median Candida burdens of the kidneys were 1.5 x 106 CFU/ml 24 h postinocu lation (p.i.) and 1.2 x 107 CFU/ml 72 h p.i., while median serum IL-6 levels were 479.3 pg/ml and 934.5 pg/ml, respectively. The Candida burden showed significant correlation with serum IL-6 24 h p.i. (R-2 = 0.6358; P = 0.0082) but not 72 h p.i. The second model was a mouse vaginitis model applying intravaginal inoculation of mice pretreated with subcutaneous estradiol-valerate (10 mg/ml) 3 days before infection. Candida cell count in vaginal lavage fluid was 2.8 x 10(6) CFU/ml 24 h p.i. and 1.4 x 10(8) CFU/ml 72 h p.i. Serum IL-6 response was detected in 4 of 15 mice 24 h p.i. and 9 of 15 mice 72 h p.i. Even the responders had low IL-6 serum levels (mean values 29.9 pg/ml and 60.1 pg/ml, respectively) not correlating with Candida cell count in vaginal lavage fluid. In conclusion, serum IL-6 had strong relationship with systemic C. albicans infection while the local C. albicans infection of the vagina led to partial, prolonged and limited serum IL-6 response.

We assessed the diagnostic value of standard clinical methods and combined biomarker testing (galactomannan assay and polymerase chain reaction screening) in a prospective case–control study to detect invasive pulmonary aspergillosis in patients with hematological malignancies and prolonged neutropenia. In this observational study 162 biomarker analyses were performed on samples from 27 febrile neutropenic episodes. Sera were successively screened for galactomannan antigen and for Aspergillus fumigatus specific nucleic acid targets. Furthermore thoracic computed tomography scanning was performed along with bronchoscopy with lavage when clinically indicated. Patients were retrospectively stratified to define a case-group with “proven” or “probable” invasive pulmonary aspergillosis (25.93 %) and a control-group of patients with no evidence for of invasive pulmonary aspergillosis (74.07 %). In 44.44 % of episodes fever ceased in response to antibiotic treatment (group II). Empirical antifungal therapy was administered for episodes with persistent or relapsing fever (group I). 48.15 % of patients died during the study period. Postmortem histology was pursued in 53.85 % of fatalities. Concordant negative galactomannan and computed tomography supported by a polymerase chain reaction assay were shown to have the highest discriminatory power to exclude invasive pulmonary aspergillosis. Bronchoalveolar lavage was performed in 6 cases of invasive pulmonary aspergillosis and in 15 controls. Although bronchoalveolar lavage proved negative in 93 % of controls it did not detect IPA in 86 % of the cases. Remarkably post mortem histology convincingly supported the presence of Aspergillus hyphae in lung tissue from a single case which had consecutive positive polymerase chain reaction assay results but was misdiagnosed by both computed tomography and consistently negative galactomannan assay results. For the galactomannan enzyme-immunoassay the diagnostic odds ratio was 15.33 and for the polymerase chain reaction assay it was 28.67. According to Cohen’s kappa our in-house polymerase chain reaction method showed a fair agreement with the galactomannan immunoassay. Combined analysis of the results from the Aspergillus galactomannan enzyme immunoassay together with those generated by our polymerase chain reaction assay led to no misdiagnoses in the control group. The data from this pilot-study demonstrate that the consideration of standard clinical methods combined with biomarker testing improves the capacity to make early and more accurate diagnostic decisions.