Vana Sypsa

Abstract: A cross-sectional study was carried out in injecting drug users (IDUs) from Greece to assess the seroprevalence of human herpesvirus 8 (HHV-8) and to identify potentially associated risk factors. A total of 288 IDUs were tested for K8.1 antibodies to HHV-8 lytic antigen. Associations between HHV-8 serostatus and potential risk factors were examined using univariate and multivariate logistic regression analysis. Seroprevalence of HHV-8 was 24.3% (95% CI 19.5-29.7), increasing with age from 19.4% in those aged <30 years to 52.9% in those aged 40 years (P for trend=0.003). No statistically significant associations between HHV-8-positive status and gender, educational level, age at first drug injection, needle sharing, number of imprisonments, complications from drug overdose, HIV and HCV were observed. In the multivariate logistic regression analysis, older age (40 vs. <40 years, OR 3.30, 95% CI 1.14-9.56) and report of septicaemia/abscess (yes vs. no, OR 1.80, 95% CI 1.01-3.18) were each independently associated with higher HHV-8 seroprevalence. HHV-8 is highly prevalent in the IDU population in Greece. The independent association between HHV-8 and reported abscess or septicaemia supports the hypothesis that poor hygiene conditions in the setting of drug injection may contribute to HHV-8 transmission.

Abstract: Hepatic progenitor cells (HPC) appear in a variety of liver diseases. Their occurrence in chronic hepatitis C (CHC) remains unclear, and triggering factors have to be elucidated. The presence of HPC in CHC was examined in relation to histological and virological parameters and patient age. Fifty liver biopsies of HCV-infected patients were examined. The presence of HPC was evaluated by immunohistochemical expression of keratin 7 (K7). Double immunostaining with K7 and cell proliferation marker Ki-67 was undertaken. Ductular reaction at the limiting plate, mean number of isolated progenitor cells (IPC) and isolated ductular structures (IDS) were quantified. The predominant distribution pattern of IPC and IDS and the presence of K7(+) hepatocytes were registered. Relationship between ductular reaction, IPC, IDS, presence of K7(+) hepatocytes, and patient age, hepatitis grade and stage, HCV RNA, and HCV genotype was examined. Prominent ductular reaction and increased numbers of IPC and IDS correlated significantly with older age and severe fibrosis/cirrhosis. The above HPC subtypes were not proliferating. Periportal/periseptal distribution pattern of IPC and IDS and presence of K7(+) hepatocytes were significantly more frequent in advanced hepatitis stages and in patients older than 40 years. Intraparenchymal distribution pattern correlated with younger age, lobular activity, and early fibrosis stage. K7(+) hepatocytes were encountered almost exclusively in the periportal pattern and in the presence of interface hepatitis and were more frequent among HCV genotype-1 patients. HPC activation in CHC is a common but diverse phenomenon closely related to patient age and hepatitis stage.

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Abstract: BACKGROUND: Improved sensitivity of HBV-DNA tests is of critical importance for the management of HBV infection. Our aim was to develop and assess a new ultra sensitive in-house real-time PCR assay for HBV-DNA quantification (ultra sensitive RTQ-PCR). RESULTS: Previously used HBV-DNA standards were calibrated against the WHO 1st International Standard for HBV-DNA (OptiQuant(R) HBV-DNA Quantification Panel, Accrometrix Europe B.V.). The 95% and 50% HBV-DNA detection end-point of the assay were 22.2 and 8.4 IU/mL. According to the calibration results, 1 IU/mL equals 2.8 copies/mL. Importantly the clinical performance of the ultra sensitive real-time PCR was tested similar (67%) to the Procleix Ultrio discriminatory HBV test (dHBV) (70%) in low-titer samples from patients with occult Hepatitis B. Finally, in the comparison of ultra sensitive RTQ-PCR with the commercially available COBAS TaqMan HBV Test, the in-house assay identified 94.7% of the 94 specimens as positive versus 90.4% identified by TaqMan, while the quantitative results that were positive by both assay were strongly correlated (r = 0.979). CONCLUSIONS: We report a new ultra sensitive real time PCR molecular beacon based assay with remarkable analytical and clinical sensitivity, calibrated against the WHO 1st International standard.

Abstract: BACKGROUND: The evaluation of academic research performance is nowadays a priority issue. Bibliometric indicators such as the number of publications, total citation counts and h-index are an indispensable tool in this task but their inherent association with the size of the research output may result in rewarding high production when evaluating institutions of disparate sizes. The aim of this study is to propose an indicator that may facilitate the comparison of institutions of disparate sizes. METHODS: The Modified Impact Index (MII) was defined as the ratio of the observed h-index (h) of an institution over the h-index anticipated for that institution on average, given the number of publications (N) it produces i.e. MII = h/10alphaNbeta (alpha and beta denote the intercept and the slope, respectively, of the line describing the dependence of the h-index on the number of publications in log10 scale). MII values higher than 1 indicate that an institution performs better than the average, in terms of its h-index. Data on scientific papers published during 2002-2006 and within 36 medical fields for 219 Academic Medical Institutions from 16 European countries were used to estimate alpha and beta and to calculate the MII of their total and field-specific production. RESULTS: From our biomedical research data, the slope beta governing the dependence of h-index on the number of publications in biomedical research was found to be similar to that estimated in other disciplines ( approximately 0.4). The MII was positively associated with the average number of citations/publication (r = 0.653, p < 0.001), the h-index (r = 0.213, p = 0.002), the number of publications with > or = 100 citations (r = 0.211, p = 0.004) but not with the number of publications (r = -0.020, p = 0.765). It was the most highly associated indicator with the share of country-specific government budget appropriations or outlays for research and development as % of GDP in 2004 (r = 0.229) followed by the average number of citations/publication (r = 0.153) whereas the corresponding correlation coefficient for the h-index was close to 0 (r = 0.029). MII was calculated for first 10 top-ranked European universities in life sciences and biomedicine, as provided by Times Higher Education ranking system, and their total and field-specific performance was compared. CONCLUSION: The MII should complement the use of h-index when comparing the research output of institutions of disparate sizes. It has a conceptual interpretation and, with the data provided here, can be computed for the total research output as well as for field-specific publication sets of institutions in biomedicine.

Abstract: Previous studies showed that high levels of human immunodeficiency virus type 1 (HIV-1) DNA are associated with a faster progression to AIDS, an increased risk of death, and a higher risk of HIV RNA rebound in patients on highly active antiretroviral therapy. Our objective was to develop and assess a highly sensitive real-time multiplex PCR assay for the quantification of HIV-1 DNA (RTMP-HIV) based on molecular beacons. HIV-1 DNA quantification was carried out by RTMP in a LightCycler 2.0 apparatus. HIV-1 DNA was quantified in parallel with CCR5 as a reference gene, and reported values are numbers of HIV-1 DNA copies/10(6) peripheral blood mononuclear cells (PBMCs). The clinical sensitivity of the assay was assessed for 115 newly diagnosed HIV-1-infected individuals. The analytical sensitivity was estimated to be 12.5 copies of HIV-1 DNA per 10(6) PBMCs, while the clinical sensitivity was 100%, with levels ranging from 1.23 to 4.25 log(10) HIV-1 DNA copies/10(6) PBMCs. In conclusion, we developed and assessed a new RTMP-HIV assay based on molecular beacons, using a LightCycler 2.0 instrument. This multiplex assay has comparable sensitivity, reproducibility, and accuracy to single real-time PCR assays.

Abstract: The SLC24A5 gene, the human orthologue of the zebrafish golden gene, has been shown to play a key role in human pigmentation. In this study, we investigate the prevalence of the variant allele rs1426654 in a selected sample of Greek subjects. Allele-specific polymerase chain reaction was performed in peripheral blood samples from 158 attendants of a dermatology outpatient service. The results were correlated with pigmentary traits and MC1R genotype. The vast majority of subjects (99%) were homozygous for the Thr(111) allele. Only two subjects from the control group (1.26%) were heterozygous for the alanine and threonine allele. Both of these Thr(111)/Ala(111) heterozygotes carried a single polymorphism of MC1R (one with the V92M variant and another with the V60L variant). Following reports of the rs1426654 polymorphism reaching fixation in the European population, our study of Greek subjects showed a prevalence of the Thr(111) allele, even among subjects with darker skin pigmentation or phototype.

Abstract: Mutations in the highly conserved tyrosine-methionine-aspartate-aspartate (YMDD) motif are frequently associated with resistance to antivirals and represent a major concern in the treatment of hepatitis B virus (HBV) infection. Conventional methods fail to detect minority populations of drug-resistant viral quasispecies if they represent less than 25% of the total sample virus population. The amplification refractory mutation system real-time PCR (ARMS RT-PCR) was combined with molecular beacon technology using the LightCycler system. The samples from HBV patients selected for assay evaluation included (i) 57 samples from treatment-naïve patients for biological discriminatory ability (cutoff) estimation, (ii) 12 samples from patients with treatment failure that were M204V positive by sequencing, and (iii) 13 samples from patients with treatment failure that were negative for mutation at codon 204 by sequencing. The discriminatory ability of the assay was 0.25% when tested with laboratory-synthesized DNA target sequences. The median mutant-to-wild-type ratio for samples from naive patients tested positive for the wild type and for mutant variants was 0.01% (5th and 95th percentiles = 0.0001 and 0.04%, respectively). A value of 0.04% was selected as the biological cutoff of the assay of clinical samples. In all samples M204V positive by sequencing (12/12), the mutant variant was detected as the predominant population (range, 82.76 to 99.43%). Interestingly, in 5 (38%) of 13 samples negative by sequencing, the M204V variant was detected at a ratio above the biological cutoff (0.05 to 28%). The assay represents an efficient technique for the early detection and quantification of M204V variants before mutant strains emerge to dominate the population.

Abstract: BACKGROUND: The effect of hepatitis B virus (HBV) infection on the natural history of human immunodeficiency virus (HIV) disease remains uncertain. Therefore, a retrospective cohort study was conducted to examine the influence of HIV-HBV coinfection on AIDS development and overall mortality. Moreover, our results were added to those of previous studies in a literature-based meta-analysis. METHODS: Serum samples obtained from HIV-seropositive patients from 1984 through 2003 were retrospectively tested for hepatitis B surface antigen. Multivariable analyses were performed using Poisson and logistic regression models. For meta-analytic purposes, eligible articles were identified and relevant data were abstracted. Pooled estimates of effect were calculated applying fixed and random effects models. RESULTS: The prevalence of chronic HBV infection (documented hepatitis B surface antigen seropositivity for >6 months) among 1729 HIV-positive patients was approximately 6%. The multivariable analyses in our primary study revealed no significant impact of concomitant HIV-HBV infection on progression to AIDS and all-cause mortality. However, a meta-analysis performed on data from 12,382 patients enrolled in 11 studies revealed a significant effect of HIV-HBV coinfection on overall mortality (pooled effect estimate, 1.36; 95% confidence interval, 1.12-1.64). The increased rate of death among coinfected individuals was observed in the meta-analyses of studies conducted both before (pooled effect estimate, 1.60; 95% confidence interval, 1.07-2.39) and after (pooled effect estimate, 1.28; 95% confidence interval, 1.03-1.60) commencement of highly active antiretroviral therapy. CONCLUSIONS: HIV-HBV coinfection seems to affect all-cause mortality, and strategies to reduce liver damage in patients coinfected with HIV and HBV are justified.

Abstract: OBJECTIVE: Skin cancer campaigns have only recently been implemented in a wide scale in Greece. In order to better target those individuals at risk, we aimed to explore sun exposure behaviors in a sample of the Greek population. METHODS: A total of 802 subjects, randomly selected among the residents of the two largest Greek cities, underwent a questionnaire-based telephone interview, assessing the degree of sun exposure patterns and knowledge of skin cancer risk factors. RESULTS: Females more commonly (90%) used sunscreens compared with males (67.5%), and among sunscreen users, only 8% used products with SPF <10. Almost half of the participants (48%) preferred going to the beach during the midday hours. Over exposure to the sun was mostly reported by individuals with phototypes III/IV and of younger age (18-35 years). Most participants were aware of sun exposure as a cause of skin cancer, but among those who did not, 41% were younger than 34 years of age. CONCLUSIONS: In our survey, significant differences in age, sex and phototypes were observed with regard to sun exposure patterns and sunscreen use. Educational campaigns should focus more on younger age groups, encouraging a more complete set of photoprotective measures. However, potential trial limitations inherent in self-reported measures should be taken into account.

Abstract: Mathematical modeling was employed on recent epidemiological data from Mexico in order to assess the impact of intervention strategies on the spread of influenza A(H1N1)v in the setting of the European region. Active surveillance that will ensure timely treatment and home isolation of symptomatic cases in combination with school closure seem to form an efficient strategy to control the spread of influenza A(H1N1)v.

Abstract: The impact of prioritization and of timing of vaccination strategies on reducing transmission of pandemic influenza A(H1N1) was evaluated in a community with the structure of the Greek population using a stochastic simulation model. Prioritization scenarios were based on the recommendations of the United States Centers for Disease Control and Prevention Advisory Committee on Immunization Practices and vaccination was assumed to initiate either before or during the ongoing epidemic. In the absence of intervention, an illness attack rate (AR) of 34.5% is anticipated. Vaccinating the priority groups before the epidemic (pregnant women, people who live with or care for children <6 months of age, healthcare/emergency services personnel, children 6 months-4 years old and high-risk children 5-18 years old) will have a negligible impact on the overall AR. Vaccinating the recommended groups before the epidemic (priority groups as well as all persons 6 months-24 years old and high-risk individuals 25-64 years old) is anticipated to result in overall and age-specific ARs within the range of seasonal influenza (5%-15%). Initiating vaccination early during the epidemic (AR<or=1% of the population) is predicted to result in overall ARs up to 15.2%-19.9% depending on daily vaccination coverage rates. When vaccination is initiated at a later stage (AR: 5%), only coverage of 80% of the whole population at intensive daily vaccination rates would be able to reduce ARs to approximately 15%.

Abstract: The aim of the study was to evaluate the prevalence and risk factors of HPV in a gynecologic population attending outpatient clinics using two new molecular tests. The Amplicor HPV test and the Linear Array (LA) HPV Genotyping test were used for the detection of HPV DNA in 320 women. Multiple logistic regression was used to identify independent prognostic factors of HPV positivity. The agreement between the two methods in terms of their qualitative results was 89.3% (kappa: 0.63). Based on the LA results, the overall prevalence of HPV DNA was 49.1%, 95% confidence interval (95% CI: 43.5%, 54.7%). The prevalence of high-risk HPV types was 30.3%. The predominant types were HPV-6 (24.8%) and HPV-16 (20.4%). Among women with normal cytology, the prevalence of HPV was much higher in those presenting other findings, such as inflammation, than those without other abnormal findings (49.5% vs. 31.5%). On the basis of multivariate analysis, the risk of HPV infection was higher among women with multiple sexual partners [>3 vs. 1: OR = 3.1, 95% CI: (1.5, 7.2)], Pap smear findings [low/high-grade lesions vs. negative: OR = 2.8, 95% CI: (1.2, 6.5)], the presence of warts [yes vs. no: OR = 3.0, 95% CI: (1.5, 6.3)] and no history of child birth [no vs. yes: OR = 2.6, 95% CI: (1.0, 6.7)]. Younger age was an additional risk factor for HPV infection with carcinogenic genotypes [OR for 1 year increase = 0.93, 95% CI: (0.89, 0.98)].

Abstract: A simple, safe and cost-effective treatment protocol in ovarian stimulation is of great importance in IVF practice, especially in the case of previous unsuccessful attempts. hCG has been used as a substitute of LH because of the degree of homology between the two hormones. The main aim of this prospective randomized study was to determine, for the first time, whether low dose hCG added to rFSH for ovarian stimulation could produce better results compared to the addition of rLH in women entering IVF-ET, especially in those women that had previous IVF failures. An additional aim was to find an indicator that would allow us to follow-up ovarian stimulation and, possibly, modify it in order to achieve a better IVF outcome; and that indicator may be the cDNA copies of the LH/hCG receptor. Group A patients (n = 58) were administered hCG and Group B rLH (n = 56) in addition to rFSH in the first days of ovarian stimulation. The number of follicles and oocytes and, most importantly, implantation and pregnancy rates were shown to be statistically significantly higher in the hCG group. This study has also determined, for the first time to our best knowledge, m-RNA for LH/hCG receptors in the lymphocytes of peripheral blood 40 h before ovum pick-up. cDNA levels of the hCG receptor after ovarian stimulation were significantly higher among women receiving hCG compared to those receiving LH. In addition, higher levels were encountered among women with pregnancy compared to those without, although this was not statistically significant due to the small number of pregnancies. It seems that hCG permits a highly effective and more stable occupancy of rLH/hCG receptors and gives more follicles and more oocytes. The determination of cDNA copies could be, in the future, a marker during ovulation induction protocols and of course a predictor for the outcome of ART in the special subgroup of patients with previous failures.

Abstract: A cross-sectional telephone survey on a nationally representative sample of 1,000 Greek households was performed to assess the acceptability of the pandemic influenza A(H1N1)v vaccine, factors associated with intention to decline and stated reasons for declining vaccination. The survey was initiated the last week of August 2009 (week 35) and is still ongoing (analysis up to week 44). The percentage of participants answering they would probably not/definitely not accept the vaccine increased from 47.1% in week 35 to 63.1% in week 44 (test for trend: p<0.001). More than half of the people which chronic illnesses (53.3%) indicated probably not/definitely not. Factors associated with intention to decline vaccination were female sex, age between 30-64 years, perception of low likelihood of getting infected or of low risk associated with influenza, and absence of household members suffering from chronic illnesses. For the majority of the respondents (59.8%), the main reason for intending to decline vaccination was the belief that the vaccine might not be safe. Promotion of vaccination programmes should be designed taking into account the attitudinal barriers to the pandemic vaccine.

Abstract: OBJECTIVES: Limited data exist about the risk factors of melanoma in the Greek population. We investigated the association of melanoma with phenotypic and solar indices in this darker skin population residing in an environment of high ambient ultraviolet radiation. METHODS: Our study included 200 sporadic melanoma cases and 200 age-, sex-matched control subjects. Information on history of sun exposure patterns and cutaneous reaction to sunlight was obtained and a clinical evaluation of pigmentary traits, pigmented lesions, and actinic keratoses was performed. RESULTS: In the multivariate analysis, fair skin (OR: 4.63, for fair skin versus light brown, 95% CI: 1.54-13.92), intermittent sun exposure during childhood (OR: 3.33, >2 weeks/year of sun exposure versus < or =2 weeks/year 95% CI: 1.37-8.09), and outdoor leisure activities (OR: 2.74, 95% CI: 1.28-5.89), but not skin phototype or sunburns, were positively related to the risk of melanoma. In addition to an elevated count of common melanocytic nevi (OR: 6.27, > or =10 nevi versus no nevi, 95% CI: 1.65-23.76) and the presence of clinically atypical nevi (OR: 2.84, 95% CI: 1.16-6.98), solar lentigenes were an independent risk factor of melanoma (OR: 4.33, 95% CI: 1.67-11.22). CONCLUSIONS: Intermittent sun exposure of moderate intensity during childhood/adolescence and outdoor leisural activities, in conjunction with a more resistant skin phenotype to acute sunburns and a strong association with nevi and solar lentigenes was a prominent determinant of melanoma risk in our population.

Abstract: In the recent years, studies of hepatitis B (HBV) and hepatitis C virus (HCV) dynamics have drawn great attention as they provide insight into the process of virus elimination/production and of infected cells decay during antiviral treatment. Estimates of viral dynamic parameters may be used to determine the lifetime of HCV/HBV virions and of infected cells, to estimate how long patients need to be treated and to evaluate antiviral therapies. The implementation of viral dynamics studies is difficult because they involve an intensive blood-sampling schedule and subsequent viral load quantification. In the majority of these studies, a model proposed by Neumann et al. (Science 1998; 282:103-107) is used under various assumptions, such as ignoring the delay in initial viral load decay, assuming time-constant treatment efficacy in reducing virion production and/or complete blocking of new infections, etc. However, only recently the effect of some of these assumptions on the estimated parameters has been evaluated. In this paper we provide a detailed review of the model, its underlying assumptions as well as the assumptions usually made by researchers during the design and analysis of viral dynamics studies. Then, we investigate the effect of these assumptions on the estimated parameters using simulations and draw useful conclusions concerning the analysis of these studies. Real data examples from a clinical trial on hepatitis B are provided as illustrations.

Abstract: A cross-sectional study was carried out in healthy company employees from Greece with the aim of assessing the prevalence of human herpesvirus 8 (HHV-8) and identifying risk factors for this herpesviral infection. Serum samples obtained from 955 subjects were tested for antibodies to HHV-8 by the K8.1 enzyme-linked immunosorbent assay (ELISA). Associations between HHV-8 serostatus and potential risk factors were examined using t-test, chi square test, and multivariate logistic regression analysis. HHV-8 prevalence was 7.6% (95% confidence interval (CI): 6.0%, 9.5%) and it increased with age from 6.5% among <30 years old to 13.8% among > or =50 years old subjects (P = 0.006). HHV-8 seropositivity was independently associated with endoscopic examination (odds ratio (OR): 2.01; 95% CI: 1.09, 3.70; P = 0.026), HBsAg positivity (OR: 5.16; 95% CI: 2.02, 13.20; P = 0.001) and age (OR > or =50 years old vs. <50 years old: 2.09; 95% CI: 1.23, 3.52; P = 0.006). No statistically significant associations between HHV-8 positive status and gender, occupational status, surgery, transfusion, tattoos/body piercing, multiple sex partners, weakness/fatigue, HCV status were observed. HHV-8 is prevalent in Greece. The strong association between HBV infection and HHV-8 positive status supports the hypothesis of an association between these two viral infections. The association between HHV-8 seropositivity and endoscopic examination requires further investigation.

Abstract: BACKGROUND AND OBJECTIVES: The Procleix Ultrio human immunodeficiency virus type 1 (HIV-1)/hepatitis C virus (HCV)/hepatitis B virus (HBV) (Ultrio) assay simultaneously detects HIV-1 RNA, HCV RNA and HBV DNA in individual blood donations. The main objective of the study was to assess the analytical and clinical sensitivity of the multiplex and discriminatory probe assays in samples with a low viral load. MATERIAL AND METHODS: The VQC HIV RNA genotype B, HCV RNA genotype 1 and HBV DNA genotype A standard dilutions were tested in 26 repeats. The probability of detection by Ultrio was compared with previously obtained data of the Procleix Duplex HIV-1/HCV assay on the same reference panels. A selection of 121 anti-HIV-1, 138 anti-HCV and 190 HBsAg positive samples from patients receiving antiviral therapy were tested. The majority of patient samples had a viral load below the detection limit of the diagnostic nucleic acid test assays, which made them suitable to evaluate the performance of the multiplex and discriminatory assays on yield cases with a similar low viral load. RESULTS: The 95% and 50% detection end-points of the Ultrio assay along with the corresponding 95% confidence intervals are 53.7 (32.9-117.2) and 8.6 (6.2-12.1) geq/ml for HIV-1 RNA, 30.3 (19.0-62.4) and 5.2 (3.7-7.2) geq/ml for HCV RNA and 393.7 (147.9-6978) and 54.5 (22.4-143.8) geq/ml for HBV DNA. The analytical sensitivity of Ultrio expressed as a potency factor relative to previously obtained Duplex results on the same HIV-1 RNA and HCV-RNA standard dilutions was 1.09 (0.20-6.10) and 1.11 (0.21-5.89), respectively. The assay detected all 22 HIV-1 infected patients with viral load > 50 copies/ml, and 41 of 99 patients (41%) with viral load < 50 copies/ml, of which 23 (56%) were detected by the discriminatory assay. All 47 patients with HCV RNA load > 521 IU/ml and 10/91 polymerase chain reaction-negative patients with viral load < 50 IU/ml tested positive in Ultrio assay of which five were missed in the discriminatory test. The assay detected 53/55 HBV infected patients (96%) with viral load > 250 copies/ml and 108/135 patients (80%) with viral load < 250 copies/ml of which 17 (16%) were missed by the discriminatory test. CONCLUSIONS: The new Procleix Ultrio assay is as sensitive as the Procleix Duplex assay for HIV-1 and HCV detection meeting the requirements of universal guidelines. The ability of the assay to detect HBV DNA in low viral load samples could be useful for screening blood. Inevitable negative results of discriminatory probe assays caused by stochastic sample variation will reduce the chance of recognizing low viraemic blood donors detected by individual donation nucleic acid test.

Abstract: BACKGROUND: A 9-month course of isoniazid monotherapy is currently recommended for the treatment of latent tuberculosis infection (LTBI) and has been shown to be effective in both children and adults. Reduced compliance with this regimen has forced physicians to explore shorter regimens. The aim of this study was to compare 3- and 4-month combination regimens of isoniazid plus rifampin with a 9-month regimen of isoniazid monotherapy for the treatment of LTBI in children. METHODS: This prospective, randomized, controlled study was conducted over an 11-year period (1995-2005). In period 1 (1995-1998), 232 patients received isoniazid therapy for 9 months (group A), and 238 patients received isoniazid and rifampin for 4 months (group B). In period 2 (1999-2002), 236 patients were treated with isoniazid and rifampin for 4 months (group C), and 220 patients received the same regimen for 3 months (group D). All patients were observed for > or = 3 years. RESULTS: Overall compliance with treatment was good, but patients who received isoniazid monotherapy were less compliant than were those who received short-course combination therapy (P=.011, for group A vs. group B; P=.510, for group C vs. group D). No patient in any group developed clinical disease during the follow-up period. New radiographic findings suggestive of possible active disease were more common in patients who received isoniazid monotherapy (24%) than in those treated with shorter regimens (11.8%, 13.6%, and 11% for groups B, C, and D, respectively; P=.001 for group A vs. group B; P=.418 for group C vs. group D). Serious drug-related adverse effects were not detected. CONCLUSIONS: Short-course treatment with isoniazid and rifampin for 3-4 months is safe and seems to be superior to a 9-month course of isoniazid monotherapy.

Abstract: BACKGROUND: In North America and Europe, human immunodeficiency virus (HIV)-1 infection has typically been dominated by subtype B transmission. More recently, however, non-B subtypes have been increasingly reported in Europe. METHODS: We analyzed 1158 HIV-1-infected individuals in Greece by DNA sequencing and phylogenetic analyses of protease and partial reverse-transcriptase regions. RESULTS: We found that the prevalence of non-B subtypes has increased over time and that this significant trend can be mainly attributed to subtype A, which eventually surpassed subtype B in prevalence in 2004 (42% and 33%, respectively). Multivariate analysis revealed that the year of HIV diagnosis was independently associated with subtype A infection (odds ratio for being infected with subtype A for a 10-year increase in the time period of diagnosis, 2.09 [95% confidence interval, 1.36-3.24]; P<.001). Phylogenetic analysis revealed that the subtype A epidemic in Greece is the result of a single founder event. The date of the most recent common ancestor of the subtype A in Greece was estimated to be 1977.9 (95% highest posterior density interval, 1973.7-1981.9). CONCLUSIONS: Subtype A circulates among the long-term residents of Greece. This is in contrast to the situation in most European countries, in which infection with non-B genetic forms is associated either with being an immigrant or heterosexual or with intravenous drug use.

Abstract: BACKGROUND: Several investigators have described a seasonal variation in the diagnosis of cutaneous melanoma. Limited data exist on the seasonality of melanoma diagnosis in Southern European countries. PATIENTS AND METHODS: The seasonal pattern of diagnosis was analyzed in 404 Greek patients diagnosed with cutaneous melanoma (CM) between 1996 and 2004. A summer-to-winter ratio was determined overall and in relation to gender, age, anatomic site, histopathologic type, and tumor thickness. RESULTS: The summer-to-winter ratio was 1.53 for all patients (95% CI [confidence interval]: 1.15-2.02) with a ratio of 1.83 for women (95% CI: 1.20-2.78) and 1.28 for men (95% CI: 0.87-1.88). A seasonal pattern of melanoma diagnosis was observed for patients younger than 50 years of age (1.70, 95% CI: 1.05-2.74) and between 50 and 69 years (1.64, 95% CI: 1.05-2.56), for melanoma located on the upper or lower extremities (2.50, 95% CI: 1.12-5.56 and 2.23, 95% CI: 1.19-4.18, respectively), for superficial spreading and nodular melanomas (1.73, 95% CI: 1.12-2.69 and 1.52 95% CI: 0.96-2.41) and for melanomas with a tumor thickness of 1-2 mm (1.69, 95% CI: 0.91-3.12) and > 4 mm (2.13, 95% CI: 1.04-4.35). CONCLUSIONS: No major differences were seen in the seasonal distribution of CM diagnosis in a Mediterranean population compared to previously reported results. A better ascertainment of the skin during the summer and an increased awareness due to the melanoma screening campaigns are the more likely reasons for the seasonality of melanoma diagnosis in Greece.

Abstract: This study aimed to estimate the overall HCV genotype distribution and to reconstruct the HCV genotype-specific incidence in Greece during the recent decades. It also focused at the identification of genotype 4 subtype variability in Greek isolates. A total of 1686 chronically infected HCV patients with detectable serum HCV RNA by RT-PCR, belonging to different risk groups were studied. Amplified products from the 5'-noncoding region were typed using a commercially available assay based on the reverse hybridization principle. The HCV genotype-specific incidence was estimated using a previously described back calculation method. HCV genotype 1 was the most prevalent (46.9%) followed by genotype 3 (28.1%), 4 (13.2%), 2 (6.9%) and 5 (0.4%). A high prevalence of genotype 1 (66.3%) in haemophilia patients was recorded whereas HCV genotype 3 was found mainly among patients infected by I.V. drug use (58.2%). Data on the temporal patterns of HCV genotype-specific incidence in Greece revealed a moderate increase (1.3-1.6 times) for genotypes 1 and 4, and a decrease (1.5 times) for genotype 2 from 1970 to 1990, whereas there was a sharp (13-fold) increase for genotype 3. The molecular characterization of 41 genotype 4 HCV isolates belonging to various risk groups revealed that, subtype 4a was the most frequently detected (78%). Phylogenetic comparison of the Greek 4a isolates with all HCV-4a isolates reported worldwide so far revealed a topology which does not discriminate Greek isolates from the others. HCV-4 does not represent a recent introduction in Greece.

Abstract: The COBAS TaqMan HIV-1 test (Roche Diagnostics) was compared with the LCx HIV RNA quantitative assay (Abbott Laboratories), the Versant HIV-1 RNA 3.0 (bDNA) assay (Bayer) and the COBAS Amplicor HIV-1 Monitor v1.5 test (Roche Diagnostics), using plasma samples of various viral load levels from HIV-1-infected individuals. In the comparison of TaqMan with LCx, TaqMan identified as positive 77.5% of the 240 samples versus 72.1% identified by LCx assay, while their overall agreement was 94.6% and the quantitative results of samples that were positive by both methods were strongly correlated (r=0.91). Similarly, in the comparison of TaqMan with bDNA 3.0, both methods identified 76.3% of the 177 samples as positive, while their overall agreement was 95.5% and the quantitative results of samples that were positive by both methods were strongly correlated (r=0.95). Finally, in the comparison of TaqMan with Monitor v1.5, TaqMan identified 79.5% of the 156 samples as positive versus 80.1% identified by Monitor v1.5, while their overall agreement was 95.5% and the quantitative results of samples that were positive by both methods were strongly correlated (r=0.96). In conclusion, the new COBAS TaqMan HIV-1 test showed excellent agreement with other widely used commercially available tests for the quantitation of HIV-1 viral load.

Abstract: BACKGROUND: The aim of the present study was to evaluate human herpesvirus 8 (HHV-8) seroprevalence in Greek hemodialysis patients. Patterns of change in HHV-8 serostatus (seroconversions and seroreversions) over time were also evaluated. METHODS: Serum samples obtained from a cohort of 485 Greek hemodialysis patients were tested for antibodies to HHV-8 by whole virus lysate enzyme-linked immunosorbent assay, and reactive samples were confirmed by means of the orf-73 enzyme-linked immunosorbent assay. HHV-8 seroprevalence at study entry and the incidence of seroreversions and seroconversions per 100 person-years were estimated. RESULTS: The prevalence of HHV-8 antibodies in Greek hemodialysis patients at enrollment was 7.2%. No univariate associations were established between HHV-8 serostatus and patients' characteristics. Incidences of seroreversions and seroconversions were 16.4/100 person-years (95% confidence interval, 7.1 to 32.3) and 0.28/100 person-years (95% confidence interval, 0.03 to 1.02), respectively. Patients 50 years and younger had an increased probability for seroreversion to HHV-8 antibodies than patients older than 50 years (log-rank test, P = 0.018). CONCLUSION: We observed a fair number of seroreversions and a low incidence of seroconversion to HHV-8 infection in hemodialysis patients in Greece. Our data provide indirect evidence that HHV-8 transmission in the hemodialysis setting is uncommon.

Abstract: BACKGROUND: The aim of this study is to evaluate the natural course of hepatitis C virus (HCV) infection in renal transplant recipients infected shortly before or after renal transplantation. METHODS: Seventeen renal transplant recipients with no detectable antibodies to HCV before renal transplantation either seroconverted after transplantation or developed cholestatic syndrome without seroconversion, but with HCV RNA positivity. They were followed up for a mean of 7.2 +/- 4.2 (SD) years after renal transplantation and underwent consecutive liver biopsies. RESULTS: Biochemical abnormalities initially were observed a median of 5.7 months (25th, 75th percentiles, 2.4, 13.9) after transplantation. Initial liver biopsies showed acute hepatitis in 5 patients and chronic hepatitis in 9 patients, whereas 3 patients had histological findings of fibrosing cholestatic hepatitis. During a median follow-up of 2.0 years (25th, 75th percentiles, 1.3, 4.6), the condition of 5 patients, initially with diagnoses of acute hepatitis, deteriorated rapidly, with a median fibrosis progression rate of 0.77 (25th, 75th percentiles, 0.56, 0.86) per year. Six patients with chronic hepatitis progressed with a median fibrosis progression rate of 0.35 (25th, 75th percentiles, 0.15, 0.69) per year in a median of 3.1 years (25th, 75th percentiles, 2.4, 3.5), whereas the other 3 patients with chronic hepatitis with elevated cholestatic liver enzyme levels developed early fibrosing cholestatic hepatitis (1 patient) or vanishing bile duct syndrome (2 patients). Genotype 1 was found in 7 of 9 patients with fibrosing cholestatic hepatitis or vanishing bile duct syndrome (78%; P = 0.049). Six of 17 patients died a median of 6.1 years (25th, 75th percentiles, 1.5, 7.1) posttransplantation; 4 of these 6 patients died of hepatic failure. CONCLUSION: HCV infection acquired shortly before or after renal transplantation frequently is associated with an adverse clinical outcome, characterized by rapid progression of fibrosis, development of cholestatic syndrome, and high mortality rate. Acute hepatitis occurring under maximal immunosuppression is of great prognostic significance, determining a specific high-risk group.

Abstract: Individuals with melanocortin 1 receptor (MC1R) gene variants have been shown to carry an increased risk for the development of melanoma. In this study, we investigated the relationship of MC1R gene variants and the risk of melanoma in 123 melanoma patients and 155 control subjects from Greece. The entire MC1R gene was sequenced for polymorphisms and the results were correlated with host factors and pigmentary characteristics. MC1R polymorphisms were present in 59.4% of melanoma patients compared to 37.5% of controls, yielding an odds ratio (OR) of 2.43 (95% confidence interval (CI) = 1.50-3.96, P < 0.001) for melanoma among MC1R carriers. The risk of melanoma was enhanced in individuals carrying multiple variant alleles (OR = 6.97; 95% CI = 1.86-26.12, P = 0.004). Only the Val60Leu, Arg142His, and Arg151Cys variants were significantly associated with melanoma risk. In stratified analysis, the risk of melanoma among MC1R carriers was not influenced by skin phototype, skin color, or hair color. No association was found between MC1R genotype and the age of onset of melanoma, the tumor location, or the tumor thickness. In conclusion, MC1R polymorphisms are a predisposing factor of melanoma in a southern European population with a relatively low incidence of the disease.

Abstract: AIM: To study the incidence of ulcerative colitis UC in the prefecture of Trikala, Central Greece. METHODS: A prospective and population based epidemiological study of UC from 1990 to the end of 1994 was conducted. Trikala is a semirural prefecture of Central Greece with a population of 138 946 (census 1991). Three gastroenterologists (one hospital based, two private doctors) of the prefecture participated in this study. RESULTS: During the study period, 66 new histologically verified cases of UC were recorded. The mean annual incidence of the disease in 1990-1994 was 11.2 per 10(5) inhabitants (95%CI: 8.7-14.3). There was no difference between men and women (annual incidence: 10.5 and 12.0 per 10(5) inhabitants respectively), either among urban, semirural or rural populations (annual incidence: 11.7, 17.1 and 9.9 per 10(5) inhabitants respectively). The majority (56%) of the patients never smoked and a quarter were ex-smokers. About a half of all cases had proctitis. CONCLUSION: UC is common in Central Greece and its incidence is similar to that in North-Western European countries.

Abstract: We studied viral dynamic parameters in 44 chronic hepatitis B/hepatitis B e antigen (HBeAg)(-) patients treated with pegylated interferon alfa-2b (PEG-IFN) 100 or 200 microg weekly or lamivudine 100 mg daily or the combination of PEG-IFN 100 or 200 microg with lamivudine. Patients receiving PEG-IFN monotherapy exhibited viral load oscillations between weekly injections, which were resolved by the addition of lamivudine. The median pharmacological delay was estimated at 4.1, 5.8, and 1.8 hours in PEG-IFN monotherapy, PEG-IFN 100/200 microg + lamivudine, and lamivudine monotherapy, respectively (P = .44). The median half-life of free virus was 12.7 hours (range, 2.4-69.2 hours). The mean antiviral effectiveness of PEG-IFN 100/200 microg monotherapy was lower than that of lamivudine (82.6% vs. 96.4%; P = .005). The mean effectiveness of PEG-IFN 100 microg + lamivudine and PEG-IFN 200 microg + lamivudine was 92.8% and 94.4%, respectively. The half-life of infected cells ranged from 2.7 to 75 days. The median half-life of infected cells in patients receiving the combination regimens of PEG-IFN and lamivudine was similar to that of lamivudine patients (5.0 days vs. 6.0 days, P = .77). In conclusion, the addition of pegylated interferon alfa-2b in lamivudine treatment was found to neither enhance the potency of blocking HBV production nor the decay rates of infected cells. Supplementary material for this article can be found on the HEPATOLOGY website (http://www.interscience.wiley.com/jpages/0270-9139/suppmat/index.html).

Abstract: BACKGROUND: The aim of this multicenter hemodialysis (HD) cohort study is to prospectively investigate the incidence of hepatitis C virus (HCV) infection in Greece from 1993 to 1995 and delineate early virological and serological events associated with HCV seroconversion in the HD setting. METHODS: Sequential serum samples collected weekly from 562 patients were tested biochemically and serologically by means of a second- (EIA-2) and third-generation enzyme immunoassay (EIA-3). All patients with positive antibody to HCV test results (anti-HCV + ) and sequential samples from seroconverting patients were tested for HCV RNA. RESULTS: Anti-HCV prevalence at study entry was 29% (163 of 562 patients), and viremia was detectable in 110 of 163 anti-HCV + patients (67.5%). HCV incidence was 6.2 cases/100 person-years. Seroconversions could not be attributed to transfusions after study entry (only 1 patient had been administered transfusion), and HD unit was associated with increased hazard for seroconversion ( P = 0.002), even after adjusting for potential differences among their patients. According to Kaplan-Meier estimation, the median interval by which the HCV RNA assay detected HCV infection earlier than anti-HCV testing was 246 and 154 days for EIA-2 and EIA-3, respectively. Detectable HCV RNA and at least 2 consecutive abnormal alanine aminotransferase levels in the preseroconversion period were observed in 29 of 30 (97%) and 14 of 32 patients (44%), respectively. Reductions in HCV RNA levels immediately after seroconversion were transient or did not occur. CONCLUSION: On the grounds of apparent nosocomial transmission, the wide window period of HCV infection in HD patients emphasizes the need for strict adherence to specific infection-control measures in this setting.

Abstract: The prevalence of HIV-1 drug resistance mutations in naïve patients has been previously shown to differ greatly with the geographic origin. The purpose of this study was to prospectively estimate the prevalence of HIV-1 drug resistance in Greece by analyzing a representative sample of newly HIV-1 diagnosed patients, as part of the SPREAD collaborative study. Protease (PR) and partial reverse transcriptase (RT) sequences were determined from 101 newly diagnosed HIV-1 patients, in Greece, during the period September 2002--August 2003, representing one-third of the total newly diagnosed HIV-1 patients in the same time period. The prevalence of HIV-1 drug resistance was estimated according to the IAS-USA mutation table taking into account all mutations in RT and only major mutations in PR region. The overall prevalence of resistance was 9% [95% confidence interval (CI): 4.2--16.2%]. The prevalence of mutations associated with resistance to NRTIs was 5% (95% CI: 1.6--11.2%), for NNRTIs was 4% (95% CI: 1.1--9.8%), while no major resistance mutations were found in PR. No multi-class resistance was detected in the study population. The prevalence of resistant mutations in the recent seroconverters was 22%. For two individuals, there was clear evidence for transmitted resistance based on epidemiological information for a known source of HIV-1 transmission. The prevalence of the HIV-1 non-B subtypes and recombinants was 52%.

Abstract: SUMMARY: The epidemic of hepatitis C virus (HCV) infection is a major public health issue. We conducted a comprehensive analysis to estimate future HCV-related morbidity and mortality, using a model which is the first to take into account currently available treatments. We reconstructed the incident infections per year in the past that progressed to chronic hepatitis C (CHC) in Greece. Then, the natural history of the disease was simulated in subcohorts of newly infected subjects in the presence or absence of treatment using yearly estimates of the number of treated patients obtained from national databases. Annual estimates of the incidence and prevalence of CHC by fibrosis stage, hepatocellular carcinoma (HCC) and mortality were obtained up to 2030. The current proportion of naïve CHC patients receiving treatment in Greece is 1.2% per year. Treatment of 1.2-10% of naïve CHC patients per year would reduce the cumulative number of incident cirrhosis and HCC cases from 2002 to 2030 by 10.8-39.4% and 12.8-39.8%, respectively and decrease the number of prevalent cirrhosis and HCC cases in 2030 by approximately 17-48% compared with the number estimated under the assumption of no treatment. Approximately 17 cirrhosis cases or six HCC cases or 10 premature deaths would be prevented for every 100 treated patients. However, the prevalent cirrhotic/HCC cases because of HCV and HCV-related deaths would not plateau until 2030. Despite the introduction of effective treatment, HCV-related morbidity and mortality will likely increase during the next 20-30 years in Greece. Intensive primary prevention efforts coupled with increased access to the currently available treatments are necessary to control the chronic consequences of HCV epidemic.

Abstract: BACKGROUND: One HIV-1 and HCV assay simultaneously detects HIV-1 and HCV RNA (Procleix, Chiron Corp.). The main intended use of the assay is the testing of blood and blood products in blood banking. STUDY DESIGN AND METHODS: To evaluate the clinical sensitivity of the assay, 164 anti-HIV-1+ and 160 anti-HCV+ patients of different viral load were tested. The assay specificity was determined in 1000 HIV-1- and HCV-seronegative blood donors. The ability of the assay to detect different HCV genotypes was investigated in a total of 40 patients of different genotypes (1-4). Furthermore, to investigate the reduction of the HCV window phase before seroconversion, serial samples of 25 hemodialysis patients who seroconverted to anti-HCV were also tested. RESULTS: The assay detected all 60 HIV-1-infected patients with a viral load of greater than 50 copies per mL and 48 of 104 patients with a viral load of less than 50 copies per mL. Moreover, all 60 patients with an HCV RNA load of greater than 521 IU per mL and 7 of 100 patients with a viral load of less than 50 IU per mL tested positive. The assay specificity was found to be 100 percent. In addition, all 40 patients of different HCV genotypes were successfully detected. Finally, the median time that the assay detected HCV infection before second- and third-generation anti-HCV assay was found to be 183 and 91 days, respectively. CONCLUSION: The assay sensitivity and specificity, its ability to detect different HCV genotypes, and the significant reduction of window period of HCV infection further support its use for improving the safety of blood and blood products.

Abstract: In this study, a comprehensive methodology for modelling the hepatitis C virus (HCV) epidemic is proposed to predict the future disease burden and assess whether the recent decline in the incidence of HCV may affect the future occurrence of cirrhosis and hepatocellular carcinoma (HCC) cases. Using the prevalence of HCV, the distribution of chronic hepatitis C (CHC) patients within the various transmission groups and their infection-onset times, it was possible to reconstruct the incident infections per year in the past that progressed to CHC in Greece. The natural history of the disease was simulated in subcohorts of newly infected subjects using transition probabilities derived either empirically between fibrosis stages 0-4 or from literature review. Annual estimates of the incidence and prevalence of CHC by fibrosis stage, HCC and mortality in Greece were obtained up to 2030. HCV incidence peaked in the late 1980s at five new infections/10,000 person-years. Under the assumption of 20-100% decline in HCV incidence after 1990, the cumulative number of incident cirrhosis and HCC cases from 2002-2030 was projected to be lower by 9.6-48.2% and 5.9-29.5%, respectively, than that estimated under the assumption of no decline. However, the prevalent cirrhotic/HCC cases and HCV-related deaths are predicted to decline in the next 30 years only under the assumption of complete elimination of new HCV infections after 1990. Despite the progress in the reduction of HCV transmission, primary prevention does not seem adequate to reverse the rise in the incidence of cirrhosis and HCC.

Abstract: The LCx HIV RNA quantitative assay (Abbott Laboratories, Delkenheim, Germany) was compared with the Versant HIV-1 RNA 3.0 (bDNA) assay (Bayer, Tarrytown, NY) and the COBAS Amplicor HIV-1 Monitor v1.5 test (Roche Diagnostics, Branchburg, NJ), using plasma samples of various viral load levels from HIV-1-infected patients. Considering the lower limit of the linear range of 50 copies/ml of both assays, the detection range of the LCx was 127/151 (84.1%) versus the 131/151 (86.8%) of the bDNA 3.0 assay, while overall agreement between the two assays was 93.4% (141/151). LCx and bDNA 3.0 results were found to be strongly correlated (r = 0.96). The fitted regression line was described by the equation log10(LCx copies/ml) = 0.05 + 1.06 x log10(bDNA 3.0 copies/ml) with 95% CI for the estimated slope and intercept at 1.01, 1.12 and -0.16, 0.26, respectively. Similarly, the detection range of the LCx was 115/148 (77.7%) versus the 128/148 (86.5%) of the Monitor v1.5 test. A 91.2% concordance (135/148) was observed between these two assays at a cut-off of 50 copies/ml. LCx and Monitor v1.5 results were highly correlated (r = 0.96). The fitted regression line was described by the equation log10(LCx copies/ml) = 0.06 + 1.03 x log(10)(Monitor v1.5 copies/ml) with 95% CI for the estimated slope and intercept at 0.97, 1.09 and -0.16, 0.28, respectively.

Abstract: A randomized trial was conducted to assess the efficacy of daily (QD) or thrice weekly (TIW) administration of interferon-alpha (IFN) in high doses in combination with ribavirin (1.0-1.2 g/day) in patients with chronic hepatitis C (CHC) who were nonresponders to previous IFN monotherapy. Interferon was administered as 10 MU IFN (QD or TIW) for 4 weeks, followed by 5 MU IFN (QD or TIW) for 20 weeks, and then by 3 MU IFN (QD or TIW) for 24 weeks. Sustained virological response (SVR) was evaluated in 142 patients who received at least one dose of medication. One-fourth of the patients achieved SVR, 26% of those treated with IFN QD and 25% of those treated with IFN TIW (P = 0.85). For genotype 1 patients, SVR rates were 32.4 and 15.8% for IFN QD and IFN TIW, respectively, whereas for genotype non-1 patients the corresponding SVR rates were 20.6 and 36.4%, respectively (test of homogeneity: P = 0.031). This finding was further confirmed by multivariate logistic regression analysis where a statistically significant interaction (P = 0.012) was found between treatment and HCV genotype indicating that the IFN QD regimen was superior to IFN TIW among genotype 1 patients whereas, among genotype non-1 patients, the two treatments were similar (odds ratio of SVR in IFN QD vs IFN TIW: 3.33 among genotype 1 patients, 95% CI: 1.00-11.14). In conclusion, re-treatment of patients not responding to previous IFN monotherapy with a combination of high daily dose of IFN with ribavirin may be beneficial for genotype 1 infected patients.

Abstract: The current case study provided an unusual setting to track the evolution of HIV-1 envelope gene over a maximum period of 6 years in two asymptomatic spouses undergoing suppressive highly active antiretroviral therapy. For this purpose, proviral DNA samples taken from uncultured peripheral blood mononuclear cells and spanning the C2-V5 regions of env were analyzed at three sampling points per subject. Two distinct topological patterns were observed in the phylogenetic reconstructions of the genetically linked sequences of the couple: an intermingled pattern and a sequentially shifting pattern in the virus populations of the male index case and his spouse, respectively. Application of three evolutionary models for the amino acid-encoded sites, using the maximum likelihood approach, indicated the operation of positive selection in the region only at the second time point in the woman, before receiving therapy. These findings reinforce the evidence of a crucial role for host-selective constraints on HIV-1 env evolution in vivo.

Abstract: A randomized trial was conducted to assess the efficacy of interferon-alpha (IFN) daily in combination with ribavirin in 301 naïve patients with chronic hepatitis C (CHC). Patients were randomized to receive ribavirin 1.2 g daily (QD) for 48 weeks with either IFN 5 MU (thrice weekly) TIW for 8 weeks followed by IFN 3 MU TIW for 40 weeks (IFN TIW, n = 154) or IFN 5 MU QD for 8 weeks followed by IFN 3 MU QD for 16 weeks followed by IFN 3 MU TIW for 24 weeks (IFN QD, n = 147). Treatment discontinuation rates, because of adverse events, were similar in the two arms (14.9% in IFN TIW and 14.3% in IFN QD, P = 0.87). The proportion of patients with sustained virological response (SVR) was 27.9% for patients treated TIW and 38.8% for those treated QD (P = 0.046). According to logistic regression analysis, patients in the IFN QD arm had 1.7 times higher probability of achieving SVR, than those receiving IFN TIW (P = 0.038). Low baseline viral load (P = 0.017) and genotype non-1 (P = 0.036) were associated with higher SVR rates. Combination of IFN/ribavirin for 48 weeks is more effective when IFN is administered daily for the first 24 weeks in naïve patients with CHC.

Abstract: Recombination plays a pivotal role in the evolutionary process of many different virus species, including retroviruses. Analysis of all human immunodeficiency virus type 1 (HIV-1) intersubtype recombinants revealed that they are more complex than described initially. Recombination frequency is higher within certain genomic regions, such as partial reverse transcriptase (RT), vif/vpr, the first exons of tat/rev, vpu and gp41. A direct correlation was observed between recombination frequency and sequence similarity across the HIV-1 genome, indicating that sufficient sequence similarity is required upstream of the recombination breakpoint. This finding suggests that recombination in vivo may occur preferentially during reverse transcription through the strand displacement-assimilation model rather than the copy-choice model.

Abstract: HBV DNA quantitation is used extensively for the monitoring of treatment of hepatitis B virus (HBV) infection. The aim of this study was to develop a highly sensitive and reproducible real-time PCR (RTD-PCR) assay for the quantitation of HBV DNA using the LightCycler system. The performance of this assay was assessed by analyzing serial dilutions of HBV genomic DNA of known concentration and the lower limit of detection was found to be 1 DNA copy/reaction. By using serial dilutions of plasmid standard, RTD-PCR was determined to quantify HBV DNA in a 10-log10 dynamic range. RTD-PCR was found to be more sensitive than the commercially available tests such as the Quantiplex HBV DNA and the AMPLICOR HBV MONITOR assays. The median coefficient of variation of interexperimental variability was 3.2%. The HBV DNA values obtained with RTD-PCR were highly correlated with assays available commercially. These findings suggest that our RTD-PCR assay combines high sensitivity and reproducibility for HBV DNA quantitation in an incomparable high dynamic range of quantitation.

Abstract: To determine the effect of interferon-alpha2b (IFN-alpha2b) on the long-term suppression of hepatitis C virus (HCV) RNA in patients with persistently normal or near normal alanine aminotransferase (ALT) activity, 76 previously untreated patients with serum HCV RNA and ALT levels <1.5 times the upper limit of normal (ULN) were randomized to receive either interferon-alpha2b (IFN-alpha2b) 5 MU three times a week for 24 weeks (n = 37) or no treatment (n = 39). HCV RNA testing was performed at the end of treatment and after a 6-month follow-up period. Intention-to-treat analysis showed that HCV RNA was detected significantly less frequently in treated than in untreated patients, at the end of both treatment and follow-up (43.2% vs. 7.7%, p < 0.001, and 21.6% vs. 5.1%, p = 0.033, respectively). Among treated patients, sustained virologic response was significantly higher in non-1 than in genotype 1 patients (8 of 26 or 30.8% vs. 0 of 11, p = 0.038). According to multiple logistic regression, untreated patients had a 13.5 times greater risk to be HCV RNA-positive compared with treated patients (p = 0.040). ALT levels flared up in 3 treated and 9 untreated patients (p = 0.07), suggesting that these flare-ups are related to the natural course of chronic HCV infection rather than to IFN-alpha2b. Thus, such patients could benefit from an IFN-alpha2b in combination with ribavirin regimen.

Abstract: To evaluate the safety and antiviral action of interferon-alpha (IFN-alpha) in HIV-1 infection, we undertook a proof of concept study in 27 treatment-naive patients. Eligible patients comprised two groups: the IFN-alphaT group (n = 17), which received 5 MIU IFN-alpha s.c. daily for 32 consecutive days, and the IFN-alphaNT group (n = 10), which did not receive IFN-alpha prior to highly active antiretroviral therapy (HAART), which was commenced on day 28 in both groups. IFN-alphaTreatment was well tolerated in 14 of the 17 patients of the IFN-alphaT group who completed the study. The mean HIV RNA reduction in the IFN-alphaT group on day 14 was 1.1 log(10). Viral load suppression was inversely associated with baseline viral load (p = 0.031). Four weeks after initiation of HAART, IFN-alphaT and IFN-alphaNT group patients had 2.40 and 1.82 log(10) HIV RNA reduction from baseline, respectively (p < 0.001). There was no evidence of cross-resistance with existing antiretrovirals in patients with HIV-RNA rebound after initial plasma viral load decline > or = 1 log(10) during IFN-alpha monotherapy. Thus, low daily IFN-alpha exhibits potent anti-HIV-1 activity in vivo without serious adverse effects. These properties render IFN-alpha an attractive candidate for further assessment as a constituent of HAART.

Abstract: The prevalence of TT virus (TTV) infection in various population groups from Athens, Greece, was assessed by the polymerase chain reaction (PCR) using two primer sets from distinct regions of the genome: the conventional set derived from the open reading frame-1 (ORF-1) and the new, highly sensitive set targeting the region that includes the TATA signal localized upstream of ORF-2. Based on both primer sets, TTV DNA was detected in 42/50 (84.0%) healthy individuals, 42/50 (84.0%) chronic hepatitis C patients, 31/39 (79.5%) acute non-A-E hepatitis patients (group I), 14/16 (87.5%) renal failure patients with acute non-A-E hepatitis (group II), 47/50 (94.0%) intravenous drug users (IVDU), 36/50 (72.0%) hemophiliacs, and 21/31 (67.7%) hemodialysis patients. The presence of TTV was not associated with any particular risk group, and no differences were observed in relation to demographic, biochemical and virological characteristics between TTV DNA-positive and -negative patients. TTV did not seem to have a profound effect on the course of chronic C or acute non-A-E hepatitis either. Phylogenetic analysis revealed that TTV strains circulating in the greater metropolitan area of Athens belong not only to the G1 and G2 genotypes that are encountered worldwide, but also to G3 and to G5 that are found mainly in Europe and Asia, respectively. Further studies will shed light on the role of this highly prevalent virus.

Abstract: Multi-state models defined in terms of CD4 counts are useful for modelling HIV disease progression. A Markov model with six progressive CD4-based states and an absorbing state (AIDS) was used to estimate the cumulative probability of progressing to AIDS in 158 HIV-1 infected haemophiliacs with known seroconversion (SC) dates. A problem arising in such analysis is how to define CD4-based states, since this marker is subject to measurement error and short timescale variability. Four approaches were used: no smoothing, ad hoc smoothing (to move to a later/previous state two consecutive measurements to later/previous states are needed), kernel smoothing and random effects (RE) models. The estimates were compared with the Kaplan-Meier estimate based solely on data concerning time to AIDS. There was an apparent lack of agreement between the Kaplan-Meier and the "no smoothing" estimate. With the exception of the "no smoothing" method, "ad hoc", kernel and RE estimates fell within the range of the 95 per cent CIs of the Kaplan-Meier curve. Simulations demonstrated that the use of raw CD4 counts provides overestimated transition intensities. Compared to the kernel method, ad hoc is easier to implement and overcomes the problem of the choice of bandwidth. The RE approach leads to simple models, since it usually results in very few transitions to previous states, and can handle individuals with sparse data by smoothing their predictions towards the population mean. Ad hoc was the method that performed better, in terms of bias, than the other smoothing approaches.

Abstract: A cross-sectional study was carried out in employees of 17 Greek companies with the aim of assessing the prevalence of hepatitis B (HBV) and hepatitis C (HCV) virus, identifying associated prognostic/risk factors and evaluating the effectiveness of a questionnaire as a pre-screening tool. All participants were asked to complete a questionnaire and a random sample of them was asked to provide a blood sample for hepatitis B core antibody (anti-HBc), hepatitis B surface antigen (HBsAg) and antibodies to hepatitis C (anti-HCV) testing. Individual questions or combinations of them were evaluated in terms of their ability to detect HBV or HCV(+) cases. Of 9085 eligible employees, 6074 (67%) completed the questionnaire. Of 990 samples obtained, 19.9% were anti-HBc(+), 2.6% HBsAg(+) and 0.5% anti-HCV(+). All anti-HCV(+) cases had multiple parenteral risk factors. Multiple logistic regression identified associations between anti-HBc and older age, family members with chronic hepatitis, job category and history of transfusion before 1992. HBsAg(+) was associated with older age and history of transfusion before 1992. None of the risk/prognostic factors had sufficient sensitivity and specificity for HBV but report of at least one risk factor identified all HCV(+) cases. Anti-HCV screening of those with at least two parenteral risk factors not only identified all anti-HCV(+) cases but also resulted in 86% decrease in the screening cost. Under the light of recent treatment advances, targeted questionnaire-based screening of asymptomatic people may prove to be a cost-effective way to face hepatitis C.

Abstract: The relevance of GB virus C/hepatitis G virus (GBV-C/HGV) infections in liver pathology remains unclear. To investigate the epidemiology of GBV-C/HGV in Athens, Greece, sera from 512 subjects were screened for present and past markers of GBV-C/HGV infection using a reverse transcription-polymerase chain reaction (RT-PCR) and a serological assay, respectively. GBV-C/HGV RNA was detected in 18/56 (32.1%), 12/42 (28.6%), and 16/55 (29.1%) patients with acute hepatitis B, C, or non-A-E, and in 5/58 (8.6%) and 18/68 (26.5%) patients with chronic hepatitis B or C, respectively, as well as in 50/133 (37.6%) hemodialysis patients and 10/100 (10%) healthy individuals. The data indicated that GBV-C/HGV seroprevalence is age-dependent; thus, GBV-C/HGV RNA and anti-E2 positivity were shown to be associated with younger age [odds ratio 0.98, 95% confidence interval (CI) 0. 97-1.00, P = 0.017] and older age (odds ratio 1.03, 95% CI 1.01-1.05, P = 0.002), respectively. No significant associations were identified between GBV-C/HGV RNA status and alanine aminotransferase (ALT) levels in either hepatitis or hemodialysis patients. Nevertheless, GBV-C/HGV RNA-positive acute non-A-E hepatitis patients were more likely to manifest a more severe clinical form of acute hepatitis (P = 0.024). Phylogenetic analysis of partial 5'-untranslated region sequences isolated from 18 viremic individuals showed that most GBV-C/HGV strains circulating in the greater metropolitan area of Athens belong to the 2a subgroup. A genetically diverse type 2 sequence that may represent a novel subtype within group 2 was also characterized.

Abstract: AIMS: To determine HIV and hepatitis infection prevalence and correlates with risk behaviour. DESIGN: Cross-sectional study: voluntary, anonymous HIV, hepatitis (HCV, HBV and HDV) surveillance and questionnaire on risk factors. SETTING: Korydallos Prison, Athens and Ag. Stefanos Prison, Patra, Greece. PARTICIPANTS: Of 544 drug users imprisoned for drug related offences, all completed the questionnaire and 533 blood samples were collected. MEASUREMENTS: HIV (by anti-HIV-1), HCV (by anti-HCV), HBV (by anti-HBc, HBsAg) and HDV (by anti-HDV) prevalence. Data on demography, legal status, drug use, sharing of injecting equipment. FINDINGS: Of the 544 drug users, 375 (68.9%) had injected drugs (IDUs) at some time, 35% of whom had injected whilst in that prison. Of the 533 blood samples tested, one was positive for anti-HIV-1 (0.19%), 310 for anti-HCV (58.2%), 306/531 (57.6%) for anti-HBc, 34/527 (6.5%) for HBsAg and 12/527 (2.3%) for anti-HDV. Prevalence rates for IDUs only were 0.27% for HIV-1, 80.6% for hepatitis C, 62.7% for hepatitis B and 3.3% for hepatitis D. Ninety-two per cent of IDUs injecting in prison shared needles, indicating that IDUs inject less but share more during incarceration. Multiple logistic regression revealed needle-sharing as the most important risk factor for HCV infection in IDUs. Prior knowledge of a positive hepatitis result did not appear to inhibit IDUs from practising risky behaviours in prison. CONCLUSIONS: The epidemic of hepatitis B and C among imprisoned IDUs identified by this study constitutes a major public health problem. Prevention programmes, such as counselling, HBV vaccination, community-based methadone maintenance treatment and syringe exchange schemes, are necessary in order to prevent a further spread.

Abstract: An RT-PCR assay using primers from the 5'-UTR of the GBV-C/HGV genome was used to detect viremia, and a serological assay was used to detect past exposure to GBV-C/HGV, in sera from 106 imprisoned Greek intravenous drug users. High seroprevalence rates indicative of the parenteral route of transmission of the virus were found (32.1% for GBV-C RNA and 46.2% for anti-GBV-C E2). These rates were nonetheless lower in comparison to the corresponding rates of HCV infection markers (64.2% for HCV RNA and 77.4% for anti-HCV). Statistically significant univariate associations were observed between GBV-C-RNA positivity and younger age (P=0.006) and HCV-RNA positivity (P=0.024), as well as with higher serum alanine aminotransferase levels (P< 0.001); this latter association was shown to be independent of coinfection with HCV and of age by a multiple logistic regression model. Apparently, GBV-C/HGV had spread readily by needle-sharing in prison, while causing acute subclinical hepatitis in infected inmates. Phylogenetic analysis of the partial 5'-UTR of the GBV-C/HGV genome from 16 seropositive individuals, which delineated their grouping within genotype 2, also revealed a close genetic relationship between two sets of sequences from 4 drug addicts, 3 of whom admitted to sharing needles while imprisoned.

Abstract: BACKGROUND/AIMS: Chronic hepatitis C appears to have a highly variable natural course with 20% of patients developing cirrhosis within 20 years, while the majority of them run a relatively mild course. We studied the relationships of epidemiological, biochemical and virological features with histological severity (grade) and liver disease progression (stage). METHODOLOGY: Liver histology, serum HCV RNA level and HCV genotype were determined in a well-defined cohort of 152 consecutive (100 males, 52 females) patients with chronic hepatitis C. RESULTS: Patients with minimal or mild chronic hepatitis were significantly younger than those with moderate or severe chronic hepatitis (mean age: 41.1 vs 49.5 years respectively, p=0.003). On the other hand, patients with no or mild fibrosis compared to those with moderate or severe fibrosis and to those with cirrhosis were significantly more frequently males (73%, 64% and 43%, p=0.01), parenteral drug users (36%, 11% and 11%, p=0.01) and infected with other than 1b genotype (86%, 52% and 33%, p<0.0001), significantly younger (mean age: 37, 48 and 58 years, p<0.0001) and had significantly lower HCV RNA levels (geometric mean: 6.9, 19.2 and 17.5 x 10(5) eq/ml, p=0.007). Multivariate analysis showed that stage was significantly related only to patient age (p<0.0001), HCV genotype (p=0.0025) and HCV RNA level (p=0.044). CONCLUSIONS: In chronic hepatitis C, histological severity seems to be associated only with patient age, while progression of the disease is mainly associated with patient age, HCV genotype and viremia level.

Abstract: AIM: To investigate the significance of IgM antibody to hepatitis C virus (HCV) core antigen (IgM anti-HCV core) in chronic hepatitis C. METHODS: In a group of 112 patients with histologically proven chronic hepatitis C positive for HCV RNA, IgM anti-HCV core level was studied by a sensitive semi-quantitative enzyme immunoassay. Quantitation of serum HCV RNA was done by a second generation bDNA assay and determination of HCV genotype by RT-PCR and reverse hybridization. RESULTS: IgM anti-HCV core was detected in 72 (64.3%) of the 112 patients. ALT levels were significantly higher in IgM anti-HCV core positive than negative patients. No other significant difference was observed in any of the patients' characteristics between IgM anti-HCV core positive and negative groups. On the contrary, IgM anti-HCV core level was found to be significantly higher in females than in males, in patients with moderate or severe chronic hepatitis, in patients with high HCV RNA levels and in patients infected with HCV genotype 1b. Moreover, IgM anti-HCV core level was significantly correlated with age and ALT level. Multiple regression analysis showed that IgM anti-HCV core level was significantly related only to the HCV genotype (p=0.001), histological grade (p=0.017) and ALT level (p=0.038). CONCLUSIONS: Our data support the hypothesis that IgM anti-HCV core level is associated mainly with HCV genotype and secondly with liver disease necroinflammatory activity. These associations may have implications in the pathogenesis of chronic hepatitis C.