Modeling the human heart ex vivo-current possibilities and strive for future applications.

The high organ specification of the human heart is inversely proportional to its functional recovery after damage. The discovery of induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) has accelerated research in human heart regeneration and physiology. Nevertheless, due to the immaturity of iPSC-CMs, they are far from being an representative model of the adult heart physiology. Therefore, number of laboratories strive to obtain a heart tissues by engineering methods by structuring iPSC-CMs into complex and advanced platforms. By using the iPSC-CMs and arranging them in 3D cultures it is possible to obtain a human heart muscle with physiological capabilities potentially similar to the adult heart, while remaining in vitro. Here, we attempt to describe existing examples of heart muscle either in vitro or ex vivo models and discuss potential options for the further development of such structures. This will be a crucial step for ultimate derivation of complete heart tissue-mimicking organs and their future use in drug development, therapeutic approaches testing, pre-clinical studies, and clinical applications. This review particularly aims to compile available models of advanced human heart tissue for scientists considering which model would best fit their research needs.

RNA-protein interactomes as invaluable resources to study RNA viruses: Insights from SARS CoV-2 studies.

Understanding the molecular mechanisms of severe respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is essential for the successful development of therapeutic strategies against the COVID-19 pandemic. Numerous studies have focused on the identification of host factors and cellular pathways involved in the viral replication cycle. The speed and magnitude of hijacking the translation machinery of host mRNA, and shutting down host transcription are still not well understood. Since SARS-CoV-2 relies on host RNA-binding proteins for the infection progression, several efforts have been made to define the SARS-CoV-2 RNA-bound proteomes (RNA-protein interactomes). Methodologies that enable the systemic capture of protein interactors of given RNA in vivo have been adapted for the identification of the SARS-CoV-2 RNA interactome. The obtained proteomic data aided by genome-wide and targeted CRISPR perturbation screens, revealed host factors with either pro- or anti-viral activity and highlighted cellular processes and factors involved in host response. We focus here on the recent studies on SARS-CoV-2 RNA-protein interactomes, with regard to both the technological aspects of RNA interactome capture methods and the obtained results. We also summarize several related studies, which were used in the interpretation of the SARS-CoV-2 RNA-protein interactomes. These studies provided the selection of host factors that are potentially suitable candidates for antiviral therapy. Finally, we underscore the importance of RNA-protein interactome studies in regard to the effective development of antiviral strategies against current and future threats. This article is categorized under: RNA Interactions with Proteins and Other Molecules > Protein-RNA Interactions: Functional Implications RNA in Disease and Development > RNA in Disease RNA Methods > RNA Analyses in Cells.

Gangliosides are essential endosomal receptors for quasi-enveloped and naked hepatitis A virus.

The Picornaviridae are a diverse family of positive-strand RNA viruses that includes numerous human and veterinary pathogens1. Among these, hepatitis A virus (HAV), a common cause of acute hepatitis in humans, is unique in that it is hepatotropic and is released from hepatocytes without lysis in small vesicles that resemble exosomes2,3. These quasi-enveloped virions are infectious and are the only form of virus that can be detected in the blood during acute infection2. By contrast, non-enveloped naked virions are shed in faeces and stripped of membranes by bile salts during passage through the bile ducts to the gut4. How these two distinct types of infectious hepatoviruses enter cells to initiate infection is unclear. Here, we describe a genome-wide forward screen that shows that glucosylceramide synthase and other components of the ganglioside synthetic pathway are crucial host factors that are required for cellular entry by hepatoviruses. We show that gangliosides-preferentially disialogangliosides-function as essential endolysosome receptors that are required for infection by both naked and quasi-enveloped virions. In the absence of gangliosides, both virion types are efficiently internalized through endocytosis, but capsids fail to uncoat and accumulate within LAMP1+ endolysosomes. Gangliosides relieve this block, binding to the capsid at low pH and facilitating a late step in entry involving uncoating and delivery of the RNA genome to the cytoplasm. These results reveal an atypical cellular entry pathway for hepatoviruses that is unique among picornaviruses.

How many single-nucleotide polymorphisms (SNPs) must be tested in order to prove susceptibility to bacterial meningitis in children? Analysis of 11 SNPs in seven genes involved in the immune response and their effect on the susceptibility to bacterial meningitis in children.

The aim of this study is to describe the prevalence of single single-nucleotide polymorphisms (SNPs) as well as their combinations in genes encoding proteins involved in the immune response in children with bacterial meningitis. The prospective study group consisted of 39 children with bacterial meningitis and 49 family members surveyed between 2012 and 2016. Eleven SNPs in seven genes involved in immune response were analysed. The mean number of minor frequency alleles (MAF) of studied SNPs was lowest in the control group and highest in patients with pneumococcal meningitis. We found that carrying ≥6 MAF of studied SNPs was associated with an increased risk of pneumococcal meningitis. The prevalence of risky variants was noted to be higher in patients with pneumococcal meningitis as compared to the control group. In conclusion, genetic factors are a relevant factor in determining the susceptibility to bacterial meningitis. A statistically significant cumulative effect of mutated variants on increasing the risk of bacterial meningitis was detected. Combining all three SNPs in MBL2 improves the prediction of susceptibility to pneumococcal meningitis. Analysis of risky alleles can help indicate people prone to the disease who are 'gene-immunocompromised'.

Distribution of polymorphisms rs12979860, rs8099917 and rs12980275 IL28B in patients with chronic hepatitis C.

BACKGROUND: The prognosis concerning the treatment of patients with chronic hepatitis C (CHC) is closely related to the genotype of the virus as well as to the factors dependent on the patient. It was proved that polymorphisms of the gene encoding interleukin 28B (IL28B) are associated with sustained viral response, which in the case of profitable variants of IL28B polymorphisms may reach up to 87% of the patients. OBJECTIVES: The aim of the study is to determine the prevalence of alleles and distribution of IL28B polymorphisms genotypes in the examined group of patients with CHC in Wielkopolska Province. MATERIAL AND METHODS: A total of 710 people with diagnosed hepatitis C virus were examined in order to determine the distribution of polymorphisms of gene IL28B rs12979860, rs8099917 and rs12980275. The polymorphisms were evaluated by sequencing of PCR products. RESULTS: The most often noted profitable variant was genotype TT for polymorphism rs8099917 present in 43.5% of the patients, next was AA rs12980275 in 22.5%. The rarest was the profitable variant CC of the polymorphism rs12979860 present in 17.5% of the patients. An occurrence of at least 2 IL28B polymorphisms in the preferred variants (homozygote CC, TT, AA) was found in 239 out of 710 (34%) patients, among which 117 patients had favorable genotypes for all 3 examined polymorphisms. CONCLUSIONS: The SNP distribution of gene IL28 with fixed prognostic value in the population of patients with chronic hepatitis C is different from the general population, and shows the need to evaluate polymorphisms prior to treatment.

Prevalence of IFNL3 rs4803217 single nucleotide polymorphism and clinical course of chronic hepatitis C.

AIM: To evaluate the association of IFNL3 (IL28B) SNP rs4803217 with severity of disease and treatment outcome in chronic hepatitis C (CHC). METHODS: The study enrolled 196 CHC Polish patients (82 women and 114 men in age 20-64) infected with hepatitis C virus (HCV) genotype 1. They were treatment naïve and qualified to pegylated interferon alpha (PEG-IFN-α) and ribavirin (RBV) therapy. The analyzed baseline parameters included: degree of inflammation, stage of fibrosis, viral load as well as alanine aminotransferase (ALT), asparagine aminotransferase (AST) and total bilirubin (TBIL). The analysis of response to therapy included: sustained virological response (SVR), defined as undetectable serum HCV RNA level six month after completion of 48-wk therapy, and relapse, defined as achieving undetectable viral load at the end of treatment but not SVR. HCV genotyping and HCV RNA quantification were performed using commercially available tests. DNA was isolated from peripheral blood mononuclear cells or from buccal cell swabs. In addition to rs4803217, also single nucleotide polymorphisms (SNPs) (rs12979860, rs8099917 and rs12980275) of known significance in predicting of HCV clearance were analyzed. SNPs were determined by high resolution melt analysis and confirmed by sequencing of amplicons. RESULTS: Frequency of rs4803217 genotypes in studied group was as follows: 27.55%; 54.59% and 17.86% for CC, CA and AA, respectively. The rs4803217 SNP, similar to other analyzed SNPs, was not associated with severity of CHC (grade of inflammation, stage of fibrosis, baseline viral load as well as biochemical parameters: ALT, AST, TBIL). It was demonstrated that the rs4803217C allele is associated with SVR (C vs A: P < 0.0001; dose of C allele: P = 0.0002) and non-relapse (C vs A: P = 0.001; dose of C allele: P = 0.002). Moreover, it was found that patients with CC genotype have significantly higher response rates as compared with CA/AA patients (P < 0.0001), whereas patients carrying A allele are significantly predisposed to relapse after treatment (P = 0.0007). Moreover, the association of rs4803217 with SVR was comparable to that of rs12979860 and stronger as observed for rs12980275 and rs8099917. Association of rs4803217 with relapse, was the strongest as compared with the other SNPs. The analysis of combined rs4803217 and rs8099917 genotypes demonstrated that additional genotyping of rs8099917 had no significant impact on the prediction of SVR. Multivariate analysis revealed that among analyzed SNPs only rs4803217 is an independent predictor of SVR (P = 0.016) and relapse (P = 0.024). CONCLUSION: The rs4803217 SNP is a strong, independent and superior predictor of SVR and relapse in HCV genotype 1 infected CHC patients treated with PEG-IFN-α and RBV.

Analysis of TLR2, TLR4, and TLR9 single nucleotide polymorphisms in children with bacterial meningitis and their healthy family members.

BACKGROUND: The aim was to analyse TLR2 rs5743708, TLR2 rs4696480, TLR4 rs4986790, TLR9 rs5743836, and TLR9 rs352140 single nucleotide polymorphisms (SNPs) in children with pneumococcal and meningococcal meningitis and their family members. METHODS: The study group consisted of 39 children with bacterial meningitis (25 with meningococcal meningitis and 14 with pneumococcal meningitis) and 49 family members. Laboratory test results and the course of the diseases were analyzed. Genomic DNA was extracted from 1.2ml of peripheral blood in order to analyze the five SNPs. RESULTS: Patients with pneumococcal and meningococcal meningitis showed a similar male/female ratio, mean age, and duration of symptoms. There were no statistically significant differences in biochemical markers between the two groups. All patients possessed at least one polymorphic variant of the analyzed SNPs. The most common SNP was TLR9 rs352140, detected in 89.7% of patients. No significant differences in SNP frequency were found between patients, family members, and the general population. CONCLUSIONS: The allele frequencies in the population studied are in accordance with the literature data. The study did not find an association between the analyzed SNPs and susceptibility to bacterial meningitis. The role of SNPs in genes coding toll-like receptors and the interactions between them in controlling inflammation in the central nervous system needs further evaluation.

Effect of irradiation on DNA synthesis, NBN gene expression and chromosomal stability in cells with NBN mutations.

INTRODUCTION: The NBN gene product is part of the MRE11/RAD50/NBN complex, which plays an essential role in genomic stability. In the study we try to answer the question what is the effect of irradiation on DNA synthesis, NBN gene expression and chromosomal stability in cells with homozygous c.657-661del, and heterozygous c.657-661del, p.I171V and p.R215W NBN gene mutations. MATERIAL AND METHODS: Immortalized B-lymphocytes with NBN gene mutations were X-ray irradiated at doses of 1, 2, 5 and 8 Gy/min. Radioresistant DNA synthesis rate and the percentage of cells in phase S was analyzed by 3H thymidine and BrdU incorporation assays. NBN gene expression was quantified by real-time PCR with TaqMan fluorescent probe. RESULTS: Increasing the irradiation dose resulted in gradual decrease of 3H thymidine incorporation in all cells, but significantly only in homo- and heterozygous c.657-661del cells (p-values < 0.0001). After irradiation the relative expression of NBN was significantly higher in homozygous c.657-661del and heterozygous p.R215W cells as compared to heterozygous c.657-661del, p.I171V and control cells (p < 0.01). All cells with NBN gene mutations showed significantly higher total number of chromosomal aberrations per metaphase as compared to control cells, with the highest number of aberrations in homozygous c.657-661del cells (p < 0.001). CONCLUSIONS: The results obtained indicate that homozygous c.657-661del mutation affects cell sensitivity to irradiation. Moreover, homozygous variant is associated with disturbance in the activation of cell cycle checkpoints and with defects in DNA repair. In turn, heterozygous c.657-661del, p.R215W and p.I171V mutations do not substantially alter the radiosensitivity.

The methylenetetrahydrofolate reductase 677T-1298C haplotype is a risk factor for acute lymphoblastic leukemia in children.

The etiology of acute lymphoblastic leukemia (ALL) is complex, linked with both environmental exposures and genetic factors. Functional variants of the methylenetetrahydrofolate reductase (MTHFR) gene result in disturbance in folate metabolism and may affect susceptibility to cancer. The study was performed to evaluate whether MTHFR C677T and A1298C polymorphisms, analyzed separately and together, are associated with the development of ALL in a population under 18 years of age of Caucasian ancestry.The study included 117 pediatric patients (59% males, mean age at diagnosis 7.4 ±â€Š5.2 years) with ALL, confirmed by conventional immunophenotyping surface-marker analysis and 404 healthy control subjects (48.5% men, mean age 37.7 ±â€Š11.3 years). The MTHFR C677T and A1298C genotypes were analyzed using allele discrimination tests with Taq-Man fluorescent probes.The MTHFR 677TT genotype was related to a 2-fold increase in risk of ALL (P = .014). The 677T-1298C haplotype was found in ALL patients but not in controls (frequency 0.598%; P <.0001). The observed frequency of carriers of this rare haplotype was 12%, including 677CT/1298CC (1.7%), 677TT/1298AC (6.0%), and 677CT/1298AC (4.3%) genotypes.The MTHFR 677T allele alone or in combination with the MTHFR 1298C allele significantly increases the risk of development of ALL in Polish population under 18 years of age. Further studies of haplotype composition in subjects with the 677CT/1298AC genotype are necessary to assess the risk of childhood ALL.

Interleukin 10 gene single nucleotide polymorphisms in Polish patients with chronic hepatitis C: Analysis of association with severity of disease and treatment outcome.

It is suggested that interleukin 10 (IL-10), as a modulator of immune response, is likely to influence the elimination of hepatitis C virus (HCV), the progression of chronic hepatitis C (CHC) and the response to interferon-based therapy in CHC patients. The aim of the study was to analyze the association of single nucleotide polymorphisms (SNPs) of IL-10 gene with severity of liver disease (degree of inflammation and stage of fibrosis) and outcome of pegylated interferon alpha and ribavirin combined therapy (sustained virological response (SVR) and relapse) in 196 Polish CHC patients infected with HCV genotype 1. The analysis included IL-10 promoter SNPs: -1082(A/G) rs1800896, -819(C/T) rs1800871, -592(C/A) rs1800872 and SNP in the 3' UTR of IL-10 gene: +4529(A/G) rs3024498. Genotyping was performed using PCR-RFLP and HRM analysis. It was demonstrated that the -592C allele is associated with mild hepatic inflammation. Moreover, it was found that the -819C allele might be associated with SVR and that the ACCA haplotype and intermediate IL-10 producer ACC haplotype are associated with SVR and non-relapse. It can be concluded that IL-10 SNPs are associated with severity of disease and response to therapy and may be considered as potential prognostic and predictive markers in CHC.

Does vaccination ensure protection? Assessing diphtheria and tetanus antibody levels in a population of healthy children: A cross-sectional study.

Vaccination effectiveness is proven when the disease does not develop after a patient is exposed to the pathogen. In the case of rare diseases, vaccination effectiveness is assessed by monitoring specific antibody levels in the population. Such recurrent analyses allow the evaluation of vaccination programs. The primary schedule of diphtheria and tetanus vaccinations is similar in various countries, with differences mainly in the number and timing of booster doses. The aim of the study was to assess diphtheria and tetanus antibody concentrations in a population of healthy children.Diphtheria and tetanus antibody levels were analyzed in a group of 324 children aged 18 to 180 months. All children were vaccinated in accordance with the Polish vaccination schedule.Specific antibody concentrations greater than 0.1 IU/mL were considered protective against tetanus or diphtheria. Levels above 1.0 were considered to ensure long-term protection.Protective levels of diphtheria antibodies were found in 229 patients (70.46%), and of tetanus in 306 patients (94.15%). Statistically significant differences were found in tetanus antibody levels in different age groups. Mean concentrations and the percentage of children with high tetanus antibody titers increased with age. No similar correlation was found for diphtheria antibodies. High diphtheria antibody levels co-occurred in 72% of the children with high tetanus antibody levels; 95% of the children with low tetanus antibody levels had low levels of diphtheria antibodies.The percentage of children with protective diphtheria antibody levels is lower than that in the case of tetanus antibodies, both in Poland and abroad, but the high proportion of children without diphtheria protection in Poland is an exception. This is all the more puzzling when taking into account that Polish children are administered a total of 5 doses containing a high concentration of diphtheria toxoid, at intervals shorter than 5 years. The decrease in antibody titers occurring over time is a significant factor in vaccination program planning.Tetanus antibody concentrations were found to be high, but responses to the diphtheria and tetanus components were divergent. The percentage of children protected against diphtheria was significantly lower than protected against tetanus.

Autofluorescence spectroscopy in the differentiation of laryngeal epithelial lesions - preliminary results.

Conclusions Autofluorescence spectroscopy may be a supporting tool for differential diagnosis of changes in laryngeal epithelium. Objectives Early detection and differential diagnosis of proliferative changes in the larynx are still a challenge for laryngologists. The aim of the study was to evaluate the autofluorescence spectroscopy technique to in vitro differential diagnosis of pathological changes in the epithelium of the larynx. Methods Forty-two patients aged 34-79 years were included in the study. The fifty-two tissue specimens, including 10 samples of cancerous lesion, 10 adjacent normal tissue, 10 chronic inflammation, eight cyst, three leukoplakia, four polyp, and seven Reinke's edema, were obtained during laryngological procedures. All tissue samples were independently diagnosed histopathologically. The autofluorescence emission spectra at two excitation wavelengths, 290 nm and 370 nm, were measured for every sample studied. Results The autofluorescence signals of cancerous tissue samples at both excitations exhibited identical emission band shapes of much lower intensities at their maxima as compared to the adjacent healthy tissue samples studied. The autofluorescence spectra intensities of cancerous and normal tissues varied inter-individually. Evident differences in autofluorescence intensities and its band shapes of different pathological laryngeal changes at the 290 nm excitations were demonstrated.

Functional variants of gene encoding folate metabolizing enzyme and methotrexate-related toxicity in children with acute lymphoblastic leukemia.

Methotrexate (MTX) is commonly used agent in therapy of malignancies, including acute lymphoblastic leukemia (ALL). Based on the literature data it is known that MTX elimination and toxicity can be affected by polymorphisms in genes encoding enzymes involved in MTX metabolism. The aim of our study was to investigate the influence of C677T and A1298C polymorphisms in methylenetetrahydrofolate reductase (MTHFR) gene on MTX-induced toxicity during treatment of children with ALL. We also tried to answer the question whether simultaneous occurrence of these two polymorphisms has a clinical significance. MTHFR polymorphisms were assessed in 47 pediatric ALL patients, treated according to intensive chemotherapy for childhood ALL, ALL IC BFM 2009. Prolonged MTX elimination and higher incidence of toxicity were observed for patients with 677T-1298A haplotype. On the other hand, occurrence of 677C-1298A haplotype had protective effect on MTX clearance and toxicity, that was not observed in carriers of 677C-1298C haplotype. In patients with coexistence of studied variants 677CT/1298AC heterozygotes as well as in 677TT/1298AA homozygotes more frequently toxicity incidents were noted. The obtained results suggest that occurrence of 677T allele and coexistence of 677T and 1298C alleles may be associated with lower MTX clearance and elevated risk of adverse effects during MTX-treatment of pediatric ALL patients.

HCV Infection and Interferon-Based Treatment Induce p53 Gene Transcription in Chronic Hepatitis C Patients.

It is suggested that the tumor suppressor p53 gene, classified as an interferon-stimulated gene, is implicated in the interferon (IFN)-mediated innate immunity against viruses. This study aimed to examine the transcriptional response of the p53 gene to hepatitis C virus (HCV) infection and IFN-based therapy in chronic hepatitis C (CHC) patients. The study included 65 CHC patients (HCV genotype 1), treated with pegylated IFN-α and ribavirin, and 51 healthy individuals. p53 gene expression was quantified by real-time polymerase chain reaction in peripheral blood mononuclear cells (PBMCs). Analyses were performed before and at weeks 4 and 12 of treatment. p53 gene expression was significantly upregulated in CHC patients compared with healthy controls and at week 4 of therapy. No significant differences in p53 mRNA expression between rapid virologic responders, complete early virologic responders, and nonresponders were observed. No significant correlation was found between p53 gene expression and viral load. The results obtained indicate that HCV infection and IFN-based treatment induces p53 gene transcription in PBMCs. The p53 gene may therefore play a role in HCV infection but is not directly involved in treatment-induced HCV elimination. Moreover, variations in p53 gene expression do not determine on-treatment response in patients with chronic HCV genotype 1 infection.

Vaccine-Derived Immunity in Children With Cancer-Analysis of Anti-Tetanus and Anti-Diphtheria Antibodies Changes after Completion of Antineoplastic Therapy.

BACKGROUND: Cancer survival rates and longevity of patients after therapy have significantly improved during the last decades. Thus durable protection against infections should be provided. The aim of the study was to compare the levels of vaccine-derived antibodies in children with cancer compared to those of healthy children and to investigate how therapy influences the levels of specific antibodies. PROCEDURE: A group of 40 children, diagnosed with acute lymphoblastic leukemia (ALL) or solid tumor (ST), followed in Poznan University of Medical Sciences Department of Pediatric Hematology, Oncology and Bone Marrow Transplantation, were recruited for evaluation of humoral immunity. Antibody levels were checked before treatment and 3, 6, and 12 months after treatment. RESULTS: In patients with ALL or ST, levels of IgG against tetanus and diphtheria were significantly lower than in the control group. Among ALL patients, 9% remained negative for tetanus and diphtheria antibodies 12 months after therapy. Among patients with ST 3 months after chemotherapy, there were no protective antibodies in 12% against tetanus, and in 18% against diphtheria. All patients reconstituted immunity 6 and 12 months after therapy. CONCLUSIONS: Our data show that a considerable number of cancer patients lose immunity against diphtheria and tetanus after therapy. Compared to ST, patients with ALL lose protective antibody levels more often. Patients with ST reconstituted antibodies after the treatment cessation, while levels in ALL patients remained low.

Heterozygous p.I171V mutation of the NBN gene as a risk factor for lung cancer development.

The NBN gene, also known as NBS1, is located on the chromosome band 8q21.3, and encodes a 754-amino acid-long protein named nibrin. This protein is a member of the MRE1-RAD50-NBN nuclear complex, and is involved in numerous cell processes essential for maintaining genomic stability. Heterozygous variants in the NBN gene, including p.I171V, c.657del5 and p.R215W, have been described as risk factors for the development of several malignancies. However, there is no report regarding the association of these mutations with lung cancer thus far. Therefore, the present study aimed to evaluate whether there is an association between the heterozygous p.I171V, c.657del5 and p.R215W variants of the NBN gene and the risk of developing lung cancer. The frequency of these variants was estimated in a group of 453 adults diagnosed with non-small cell lung cancer (NSCLC) and in healthy controls (2,400 for p.I171V, 2,090 for c.657del5 and 498 for p.R215W). The p.I171V variant was assessed by restriction fragment length polymorphism analysis of polymerase chain reaction (PCR) products, using MunI (MfeI) restriction enzyme, whereas the c.657del5 and p.R215W variants were assessed by the PCR single-strand conformation polymorphism method. A significantly increased risk of developing lung cancer was observed for the p.I171V variant, which was present in 17 (3.75%) of the 453 cases of lung cancer and in 12 (0.5%) of the 2,400 healthy individuals (odds ratio, 7.759; P<0.0001). The results obtained indicated an association between the p.I171V mutation and the development of lung cancer. Therefore, this variant may be considered a risk factor for NSCLC. Prospective studies with larger groups of patients may reveal the potential impact of the p.I171V variant in the occurrence of lung cancer.

Comparison of detection methods for extended-spectrum beta-lactamases in Escherichia coli strains.

INTRODUCTION: Detection of extended-spectrum beta-lactamases (ESBLs) could be a major challenge for microbiologists--the difficulties arise mainly from the phenotypic differences among strains. MATERIALS AND METHODS: Evaluation of ESBLs was performed on 42 strains of E. coli by: 1) DDST on MHA, 2) DDST on MHA with cloxacillin, 3) CT on MHA, according to CLSI, 4) CT on MHA with cloxacillin, 5) Etest ESBL (AB Biodisk), 6) CHROMagarTM ESBL (GRASO), 7) ChromID® ESBL (bioMérieux), and 8) automatic system VITEK2 ESBL test (bioMérieux). RESULT: Positive results were obtained for 20 strains using method 1, for 18 strains using method 2, 17 by method 3, 14 by method 4, 11 by method 5, 39 by method 6, 40 by method 7, and 15 by method 8. Using Etest ESBL 6.0 non-determinable results were obtained. The most consistent results were obtained when comparing the results of method 3 with results of method 2 (97.6%), and comparing the results obtained using methods 3 and 8 (95.2%). CONCLUSIONS: Based on our study we conclude that the chromogenic media can only be used as a screening method for the detection of ESBLs in E. coli rods. Etest is less useful compared to other phenotype methods, due to the impossibility of obtaining results for all the tested strains. Adding cloxacillin to MHA does not increase the frequency of detection of ESBLs in E. coli strains. DDST seems to be the most reliable among phenotypic methods for the detection of ESBLs in E. coli rods.

MicroRNA-155 and microRNA-196b: promising biomarkers in hepatitis C virus infection?

MicroRNAs (miRNAs) are a class of small, evolutionarily conserved, noncoding RNA that regulate several important cellular processes. The versatility of these molecules allowed the accurate predictions that they would also affect the replication and life cycle of HCV. In this review, emphasis has been given to two selected miRNAs: miR-155 and miR-196b. Recent data indicate that miR-155 is overexpressed in HCV-infected patients, inducing an inflammatory state, and promoting virus replication and persistence even after the completion of antiviral treatment. It is also associated with the increased proliferation and inhibited apoptosis of hepatocytes, which promotes the growth of liver tumors. In contrast, miR-196b is reported as a factor inhibiting HCV replication with cytoprotective, anti-inflammatory, and antioxidant properties. Growing evidence suggests that these molecules could be used as potential prognostic and predictive factors and their antagonists or mimics as a promising therapeutic approach in HCV-infected patients.