[show abstract][hide abstract] ABSTRACT: Background: Molecular methods are essential to define hepatitis C virus (HCV) infection. This study was conduct-ed to evaluate the performance of molecular qualitative and quantitative methods for HCV RNA among chronic patients and individuals during the course of HCV infection. Methods: Single serum samples were obtained from 82 HCV infected individuals where six of them donated serial serum samples (n = 52) during the course of HCV infection. Qualitative (in-house RT-nested PCR and COBAS ® AMPLICOR HCV Test v2.0 and TMA) and quantitative (COBAS ® AMPLICOR HCV Monitor Test v2.0 and bDNA) techniques were employed. Results: TMA presented the highest rate (87.8%) of HCV detection among qualitative tests and it was the most sensitive for HCV RNA detection during the early and late phases of HCV infection. HCV RNA was quantified among 56 samples and significant correlation was observed between the two assays (r 0.92; p < 0.0001). Conclusions: It is concluded that both quantitative methods can be used among chronic and acute HCV cases, but TMA was the most efficient for HCV qualitative detection among chronic cases and in the early and late phases of HCV infection.
[show abstract][hide abstract] ABSTRACT: To evaluate the association between 25-hydroxyvitamin D [25(OH)D] and sustained virological response (SVR) in hepatitis C virus (HCV) infected individuals.
Relevant studies were identified by systematically searching MEDLINE databases up to March 2012 and abstracts of the European and American Congress of Hepatology conducted in 2011. Studies must provide information on SVR and the levels of 25(OH)D3 and/or 25(OH)D2 [henceforth referred to as 25(OH)D] in sera samples from HCV infected individuals. The inclusion criteria were: clinical studies that included HCV infected patients aged older than 18 years regardless of HCV genotype or ethnic group; provided information on SVR rates; and were reported in the English language as full papers. Due to the heterogeneity of studies in categorizing serum vitamin D levels, a cut-off value of 30 ng/mL of serum 25(OH)D was used. Heterogeneity was assessed using I (2) statistics. The summary odds ratios with their corresponding 95%CI were calculated based on a random-effects model.
Overall, 11 studies (8 observational and 3 interventional) involving 1575 individuals were included and 1117 HCV infected individuals (71%) showed low vitamin D levels. Most of the studies included mono-infected HCV individuals with the mean age ranging from 38 to 56 years. Four studies were conducted in human immunodeficiency virus/HCV infected individuals. Regarding vitamin D measurement, most of the studies employed radioimmunoassays (n = 5) followed by chemiluminescence (n = 4) and just one study employed high performance/pressure liquid chromatography (HPLC). Basal vitamin D levels varied from 17 to 43 ng/mL in the studies selected, and most of the HCV infected individuals had genotype 1 (1068/1575) with mean viral load varying from log 4.5-5.9 UI/mL. With regard to HCV treatment, most of the studies (n = 8) included HCV individuals without previous treatment, where the pooled SVR rate was 46.4%. High rates of SVR were observed in HCV individuals with vitamin D levels above 30 ng/mL (OR = 1.57; 95%CI: 1.12-2.2) and those supplemented with vitamin D (OR = 4.59; 95%CI: 1.67-12.63) regardless of genotype.
Our results demonstrated high prevalence of vitamin D deficiency and high SVR in individuals with higher serum vitamin D levels or receiving vitamin D supplementation.
World Journal of Gastroenterology 09/2013; 19(35):5917-5924. · 2.55 Impact Factor
[show abstract][hide abstract] ABSTRACT: BACKGROUND: Crack use constitutes a major problem in cities across Brazil. While existing data suggest that crack use is generally concentrated among disenfranchised young people with extensive health problems and crime involvement, extensive data gaps exist. To address this issue, this study aimed to assess key characteristics of young crack users in two Brazilian cities. METHODS: N=160 regular and young adult (ages 18-24) crack users were recruited by community-based methods in the cities of Rio de Janeiro (Southeast) and Salvador (Northeast). Assessments included an interviewer-administered questionnaire on key social, drug use, health and service use characteristics, as well as serological testing of HBV, HCV and HIV status, and were conducted anonymously between November 2010 and June 2011. Participants provided informed consent and received transportation vouchers following assessment completion. The study was approved by institutional ethics review boards. RESULTS: The majority of participants were: male, with less than high school education, unstably housed (Rio only); gained income from legal or illegal work; arrested by police in past year (Salvador only); had numerous daily crack use episodes and shared paraphernalia (Salvador only); co-used alcohol, tobacco, cannabis and cocaine; had no injection history; rated physical and mental health as 'fair' or lower (Salvador only); had unprotected sex; were never HIV tested; were not HIV, HBV or HCV positive; and did not use existing social or health services, but desired access to crack user specific services. CONCLUSION: Crack users in the two Brazilian sites featured extensive socio-economic marginalization, crack and poly-drug use as well as sexual risk behaviours, and compromised health status. Social and health service utilization are low, yet needs are high. There is an urgent need for further research and for targeted interventions for crack use in Brazil.
The International journal on drug policy 04/2013; · 2.54 Impact Factor
[show abstract][hide abstract] ABSTRACT: BACKGROUND: Enzyme immunoassays (EIA) designed to detect hepatitis C virus (HCV) core antigen and anti-HCV antibodies (HCV AgAb) simultaneously can improve the early detection of HCV infection when molecular diagnostic methods are not widely available. OBJECTIVES: To evaluate the suitability of dried blood spot (DBS) samples for detecting HCV AgAb using commercial EIAs. STUDY DESIGN: Paired serum and DBS samples were assayed using two commercial EIAs for HCV AgAb (Monolisa™ HCV AgAb ULTRA and Murex HCV AgAb). Manufacturer's recommendations were followed for sera while sample volume, incubation time and cut-off (CO) determination were evaluated for the DBS samples. The values of sensitivity, specificity, inter-rater agreement, detection limit, assay precision and stability of DBS samples at different conditions (22-26°C, 2-8°C and -20°C) were determined. RESULTS: It was necessary to increase the DBS sample volume fourfold compared to the sera samples to approximate the DBS Optical Density (OD) values to the sera OD values. Using ROC curve to recalculate CO values for the DBS samples, sensitivity was 97.5% for both EIAs, while the specificity was 99.71% for Monolisa™ HCV AgAb ULTRA and 95.95% for Murex HCV AgAb. Accurate testing results were obtained with DBS samples for 60 days at all conditions evaluated; storage at -20°C resulted in low OD variation. Both EIAs demonstrated the same limit of detection among DBS samples [estimated viral load of 3.1 International Units per millilitre (IU/mL)] and low OD value variability in repetitivity and reproducibility studies. CONCLUSION: DBS samples can be used for the detection of HCV AgAb by EIA as they present comparable performance characteristics and excellent stability among various storage conditions.
Journal of clinical virology: the official publication of the Pan American Society for Clinical Virology 03/2013; · 3.12 Impact Factor
[show abstract][hide abstract] ABSTRACT: BACKGROUND: Oral fluid (OF) sample collection and stability for HBsAg detection are not fully established. This study aims to investigate the applicability of OF collectors and sample stability for Hepatitis B virus surface antigen detection. METHODS: Paired serum and OF samples were obtained from 191 individuals, and Chembio (Chembio Diagnostic System, USA) and Salivette (Sarstedt, Germany) devices were used for OF collection. Two HBsAg enzyme immunoassays (EIAs) were used (HBsAg One kit, Radim, Rome, Italy and ETI-MAK-4, DiaSorin, Vercelli, Italy) to determine the most efficient method according OF collector. Sample volume, incubation time, and cutoff (CO) value were evaluated. The stability of OF samples was determined under different environmental conditions. RESULTS: Chembio samples analyzed using DiaSorin EIA without modification of the manufacturer's instructions, demonstrated a sensitivity of 95.24% and a specificity of 100%. Salivette samples analyzed with Radim EIA with receiver operating characteristic (ROC) curve for calculating the CO showed a sensitivity of 78.26% and a specificity of 89.88%. HBsAg was detected in Chembio and Salivette samples under different environmental conditions, but the Chembio samples were the most stable. CONCLUSIONS: Both collectors can be used for HBsAg detection in OF samples, but some modifications of commercial EIAs should be incorporated for Salivette device. OF samples were reliably stable and could be stored for up to 90 days at 2-8°C.
Journal of Clinical Laboratory Analysis 02/2013; · 1.36 Impact Factor
[show abstract][hide abstract] ABSTRACT: Molecular methods are essential to define hepatitis C virus (HCV) infection. This study was conducted to evaluate the performance of molecular qualitative and quantitative methods for HCV RNA among chronic patients and individuals during the course of HCV infection.
Single serum samples were obtained from 82 HCV infected individuals where six of them donated serial serum samples (n = 52) during the course of HCV infection. Qualitative (in-house RT-nested PCR and COBAS AMPLICOR HCV Test v2.0 and TMA) and quantitative (COBAS AMPLICOR HCV Monitor Test v2.0 and bDNA) techniques were employed.
TMA presented the highest rate (87.8%) of HCV detection among qualitative tests and it was the most sensitive for HCV RNA detection during the early and late phases of HCV infection. HCV RNA was quantified among 56 samples and significant correlation was observed between the two assays (r 0.92; p < 0.0001).
It is concluded that both quantitative methods can be used among chronic and acute HCV cases, but TMA was the most efficient for HCV qualitative detection among chronic cases and in the early and late phases of HCV infection.
[show abstract][hide abstract] ABSTRACT: This study was undertaken to optimize and compare the efficiency of two commercial EIAs for anti-HCV detection (HCV Ab Radim, Pomezzia, Italy and ETI-AB-HCVK-4 DiaSorin, Vercelli, Italy), in dried blood spot (DBS) samples. The long-term stability of anti-HCV on DBS samples stored at three environmental conditions was also evaluated at: 2-8 °C, 20-25 °C, and -20 °C. Paired DBS and serum samples were obtained from individuals with or without anti-HCV. The type of elution buffer, sample and conjugate volume, sample incubation time and cut-off values were evaluated. For both EIAs, a larger sample volume was used, and the cut-off value determined by the manufacturer was employed for Radim EIA; however, ROC curve analysis was used for the DiaSorin EIA. The sensitivity and specificity of Radim EIA on DBS were 97.5% and 99.5%, respectively, and of DiaSorin EIA were 88.9% and 98.9%, respectively. Accurate results were obtained for a period of 117 days using DBS samples stored at all storage conditions, but storage at -20 °C resulted in the lowest variation among the absorbance values. Both EIAs demonstrated the same limit of detection (until dilution of 1:10(4) with estimated viral load of 3.1 × 10(-1) UI/ml), but the Radim EIA was associated with the best performance because a low coefficient of variation was observed in the repetition and reproducibility studies. In conclusion, commercial EIAs can be optimized for anti-HCV detection in DBS samples that are extremely stable at different conditions for more than 100 days.
Journal of Medical Virology 10/2012; 84(10):1600-7. · 2.37 Impact Factor
[show abstract][hide abstract] ABSTRACT: A strategy adopted by different countries to reduce the number of new cases of hepatitis A is the vaccination. However, the mosaic of the epidemiological profile in developing countries has hampered the establishment of a unified nationwide vaccination program. To determinate national vaccination policies, the results of epidemiological studies need to be carefully considered. For this monitoring, the use of oral fluid is very important due to the painless and non invasive collection characteristics. There are few studies investigating which oral fluid collection device is optimal to detect low antibody levels and its use in selecting individuals for vaccination. So, the present study aimed to evaluate different oral fluid collection devices to detect humoral immune response against hepatitis A virus and its application in epidemiological studies. Therefore, 90 matched serum and oral fluid samples were collected from volunteers with different immune status, under ideal conditions of collection (optimization panel); and 224 matched samples in difficult-to-access areas (epidemiological study). Serum was collected by venipuncture and the oral fluid was obtained using three commercial devices: Salivette(®), OraSure(®) and ChemBio(®). Serum and oral fluid were submitted to a commercial immunoblot to detect total anti-HAV antibodies. The optimization panel demonstrated that ChemBio(®) device had the best performance (100% agreement), followed by OraSure(®) (95.4%) and Salivette(®) (90.8%). The optimal collection device (ChemBio(®)), tested in a difficult-to-access area and evaluated under precarious conditions of collection, showed similar prevalence of total anti-HAV between serum and oral fluid, 80.8% and 79%, respectively. A follow-up was performed to evaluate the stability of oral fluid and it was observed that 210 days after the collection it was possible to detect anti-HAV antibodies. Oral fluid can be used to detect low levels of specific-antibody, being important to select age groups to be vaccinated. Therewith, the choice of proper collection device is essential to evaluate HAV antibodies in the epidemiological scenario.
[show abstract][hide abstract] ABSTRACT: Antibody responses to standard regimens of hepatitis B (HBV) vaccination are lower in HIV-infected subjects and the best hepatitis B vaccine schedule in this population is not known.
To assess the immunogenicity and to evaluate predictors of serologic response of a modified regimen of a HBV recombinant vaccine in a cohort of HIV-infected subjects.
HIV-infected subjects received 4 doses (40 μg) of a recombinant HBV vaccine at 0, 1, 2 and 6 months. Demographic information as well as CD4 cell count and plasma viral load were assessed at baseline. Protective and strong responses were defined as an anti-HBs titer ≥10 mIU/mL and ≥100 mIU/mL, respectively and were evaluated one month after the third and the fourth doses.
163 HIV-infected individuals were evaluated 67 (40%) were male and median age was 37 years. Median CD4 cell count was 385 cells/mm(3) and 113 (70%) had undetectable HIV-1 viral load. Protective antibody response was observed in 83 and 91% and a strong antibody response was observed in 62 and 80% of the subjects after 3 and 4 doses, respectively. In a multivariate logistic model undetectable HIV-1 viral load and higher CD4 cell counts were independent predictors of a strong antibody response after 4 doses. Patients with undetectable HIV viral load were almost 3 times more likely to have anti-HBs titers above 100 mIU/mL than those with detectable viral load.
A 4-double-dose regimen of a recombinant HBV vaccine increased response rates and determined higher antibody titers which may translate in prolonged protection agains HBV. Inclusion of a fourth dose of HBV vaccine for HIV-infected subjects should be considered in the public health setting.
[show abstract][hide abstract] ABSTRACT: J Oral Pathol Med (2012) Background: Saliva samples can be used as an alternative fluid for against hepatitis C virus (anti-HCV) detection owing to the ease of collection and excellent acceptability. This study was conducted to optimize a commercial enzyme immunoassay (EIA) to detect anti-HCV in saliva samples. Methods: Ninety-six individuals donated paired serum and saliva samples that were obtained, using a commercial device (Salivette) and spitting into a sterile container. Initially, elution buffer for the Salivette samples, sample volume, incubation time and temperature, and two different anti-HCV EIAs were evaluated. Using the optimized assay, three methods for cut-off calculation were also evaluated. Results: A 20-fold increase in the sample volume for both collection methods was needed. Moreover, the Radim assay was the most appropriate assay for anti-HCV detection in saliva samples, and the quality parameters were increased when a ROC curve was used to determine the cut-off value. Using this optimized assay, the sensitivities, specificities, accuracies, positive and negative predictive values were above 90% for saliva obtained using both the Salivette and spitting methods. Using this assay, discordant false-negative results were obtained for only two Salivette samples and five spitting samples. The concordance kappa was 93% for the Salivette method and 86.1% for the spitting method, demonstrating excellent performance. Conclusions: Saliva samples obtained for both methods can be employed for anti-HCV detection among HCV-infected or HCV-suspected cases, but several modifications must be performed on commercial EIAs to obtain good results. Moreover, samples obtained with commercial devices are more appropriate for anti-HCV detection in saliva samples.
Journal of Oral Pathology and Medicine 06/2012; · 2.06 Impact Factor
[show abstract][hide abstract] ABSTRACT: The objective of the present study was to evaluate four commercial DNA extraction methods and three PCR protocols for hepatitis B virus (HBV) detection in artificially contaminated oral fluid samples. The extraction protocols were selected based on ease of use and cost, and were also compared with respect to sensitivity and cost. Prior PCR optimization was conducted, in which the sample volume for DNA extraction and the concentrations of DNA and Taq DNA polymerase in the PCR were adjusted. One-round PCR, used to amplify the core region of the HBV genome, achieved high levels of sensitivity in comparison with nested and semi-nested PCR experiments that were designed for the amplification of HBV surface protein genes. Of the four extraction protocols evaluated, the RTP DNA/RNA Virus Mini kit and the QIAamp DNA Mini kit gave the highest recovery rates, presenting 20 copies of HBV DNA ml(-1) as the limit of detection. These results suggest that HBV DNA can be detected from oral fluid samples but that the optimization of the PCR assays and the choice of extraction methods must be determined by laboratories before the implementation of this method in routine diagnostics.
Journal of Medical Microbiology 03/2012; 61(Pt 6):844-51. · 2.30 Impact Factor
[show abstract][hide abstract] ABSTRACT: Objective: to describe the knowledge of viral hepatitis among nursing students in two institutions located in two different geographical areas of Brazil. Method: a descriptive, exploratory and quantitative study where 180 students were selected from two nursing higher education institutions (102 in Rio de Janeiro, Southeast and 78 of Mato Grosso do Sul, Midwest), who answered a questionnaire composed of 37 questions about socio-demographic factors and level of knowledge about viral hepatitis after signing a consent form. For data evaluation, a score of knowledge about hepatitis was created based on participants' responses, where "low" (0-21 correct answers), "good" (22-28 correct answers) and "excellent" (29 - 37 correct answers). The variables: age, sex and place of residence were used to assess knowledge of hepatitis and sociodemographic characteristics. This study was approved by the Ethics Committee of the University of Rio Grande (RJ) (protocol number CAAE 0006.0.317.000-08). Results: the mean knowledge about viral hepatitis was 25.95 ± 4.79 showing a good knowledge about viral hepatitis in this population. However, some gaps were analyzed for transmission of viral hepatitis, etiology, and symptoms as well as significant differences in knowledge scores between students from different geographical areas. Conclusions: despite the good general knowledge on the subject, it is necessary to enhance awareness and training strategies on viral hepatitis among nursing students to increase knowledge on this topic. Descriptors: knowledge; hepatitis, viral, human; education; health personnel.
[show abstract][hide abstract] ABSTRACT: Detection of hepatitis B virus (HBV) serological markers in dried blood spot (DBS) samples by enzyme immunoassay (ELISA) has not yet been fully optimized. In this study, the ability to detect three HBV markers (HBsAg, anti-HBc, and anti-HBs) was evaluated in DBS samples using a modified commercial ELISA. Matched serum and DBS samples were obtained from individuals with or without a past history of HBV infection. Sera samples were tested according to the manufacturer's instructions, but for DBS testing, paper diameters, elution buffer, volume of input sample, and cut-off values were evaluated to optimize the assay. Stability studies were done on DBS stored at for up to 180 days at different temperatures. The absorbance values that yielded the maximum sensitivity and specificity were determined based on the area under the ROC curve (AUROC) and chosen as the cut-off value. Using this parameter, sensitivity was 90.5%, 97.6%, and 78% for anti-HBc, HBsAg, anti-HBs assays, respectively. Specificity was 92.6%, 96.7%, and 97.3% for anti-HBc, HBsAg, and anti-HBs assays, respectively. HBV markers could be detected in DBS samples until 63 days after sample collection at most temperatures, but storage at -20°C yielded more consistent results. These results indicate that modified ELISA can be used to detect HBV markers in DBS samples, particularly if the samples are stored appropriately.
Journal of Medical Virology 09/2011; 83(9):1522-9. · 2.37 Impact Factor
[show abstract][hide abstract] ABSTRACT: Matched serum and saliva samples were collected simultaneously from 124 subjects exposed during a hepatitis A virus (HAV) outbreak at a daycare center in Rio de Janeiro, Brazil. All samples were tested for IgM and total anti-HAV antibodies by enzyme immunoassay (EIA). HAV was detected by nested PCR in serum, saliva, and water samples employing primers for the VP1/2A region of the viral RNA; all positive products were then sequenced. The viral load of the matched samples was determined by real-time PCR using the TaqMan system. HAV-RNA was identified by nested PCR in 37.7% of the saliva samples, 29% of the serum samples, and one drinking water sample. The mean HAV viral load was similar in the serum and saliva specimens (10(3) copies/ml). HAV genotypes IA and IB were detected in both specimen types, and the water sample isolate was classified as genotype IB, indicating the existence of more than one source of infection at the daycare center. In six infected patients, a different HAV subgenotype was found in their serum than in their saliva, and this unusual pattern of mixed HAV infection was investigated further by molecular cloning followed by nucleotide sequencing. All clones derived from the saliva samples belonged to subgenotype IB and shared 96.5-100% identity. However, clones derived from their corresponding serum sample belonged to subgenotype IA and shared 90.5-100% identity. This study showed the important role that non-invasive saliva samples can play in the molecular epidemiological analysis of a hepatitis A outbreak.
Journal of Medical Virology 05/2011; 83(5):768-75. · 2.37 Impact Factor
[show abstract][hide abstract] ABSTRACT: Multiple studies have examined the use of oral fluids in modified serum-based assays aiming to replace serum in antibody detection for hepatitis A. However, the reliable detection of HAV immunity in oral fluid requires an extremely sensitive assay; most immunoassays designed for serum antibody determination lack sufficient sensitivity for this purpose. Consequently, an "in-house" competitive enzyme immunoassay (EIA) designed specifically for use with oral samples collected using a ChemBio(®) device was developed to detect total anti-HAV antibodies (IgG and IgM). This system was compared to an in-house competitive EIA and a commercial EIA considered to be the "gold standard" using corresponding serum samples (n=225) to determine the accuracy of the assay and to evaluate the importance of the cutoff ratio for the detection of anti-HAV antibodies in oral fluids. When the median serum cutoff and the optimal oral fluid cutoff (ROC analysis) obtained from the in-house competitive EIA were compared, the oral fluid cutoff was found to be 28.8% higher than the serum cutoff. When different oral fluid cutoff values were compared, a reduction of about 17% was shown to be essential to increase test accuracy. At an oral fluid cutoff value of 0.351, sensitivity and specificity were higher, reaching 91.7% and 86.2% (p<0.001, AUROC=0.915), respectively. The convenience, accuracy and non-invasive nature of the developed method make it a useful alternative to serum-based assays for discriminating between HAV-immune and non-immune individuals.
Journal of virological methods 02/2011; 173(2):169-74. · 2.13 Impact Factor
[show abstract][hide abstract] ABSTRACT: In this study, a modified enzyme immunoassay (EIA) was evaluated for the Hepatitis B surface antigen (HBsAg) among whole saliva and oral fluid samples. Specimens were collected from 115 individuals who gave serum and oral fluid using Salivette (Sarstedt, Nümbrecht, Germany) and whole saliva. Saliva specimens were tested following a modified ELISA, and the results were compared with paired serum specimens that were tested according to the supplier's instructions. Transport buffer for the oral fluids, sample volume for assay, incubation period of sample with conjugate as well as cut-off values were evaluated to optimize the assay. The highest sensitivity and specificity were obtained by increasing the incubation of sample and conjugate to 16 hr and using the area under the receiver operating characteristic curve to calculate cut-off values. HBsAg was detected in 40 oral fluids and 44 whole saliva samples out of 47 paired positive serum specimens and not detected in 64 oral fluids and 63 whole saliva samples out of 68 matched negative sera samples by the ELISA assay. There was excellent agreement between the results for the serum and saliva specimens kappa value (κ): 0.80 for oral fluid and κ: 0.87 for whole saliva and there was excellent reproducibility. Using an optimized protocol, the sensitivities of whole saliva and oral fluid were 93.6 and 85.1%, respectively, whereas specificities of whole saliva and oral fluid were 92.6 and 94.1%, respectively. Our data showed a significant promise for the use of whole saliva and oral fluid together with the modified commercial EIA for Hepatitis B virus infection surveillance.
[show abstract][hide abstract] ABSTRACT: Human papillomavirus (HPV) infection is one of the most common sexually transmitted diseases worldwide. Although two safe and clinically effective vaccines against HPV have been developed, they are not available to the public health network in Brazil. This study was performed to assess knowledge about HPV among seventy-nine professionals who completed a questionnaire about diagnosis, transmission, symptoms, prevention and general information. General knowledge about HPV was high, as most of them recognized that HPV is transmitted sexually (98.7%), the disease can be asymptomatic (82.3%) or warts can be present on the genitals (84.8 ) and the Pap smear is the screening method to identify cellular changes on the cervix (88.6%). However, many professionals did not know that there are now vaccines available for many HPV variants (38.0%) and that not all of them are oncogenic (44.3%). These data show that further educational programs, especially about HPV prevention, are needed in Brazil.
Asian Pacific journal of cancer prevention: APJCP 01/2011; 12(12):3251-6. · 1.27 Impact Factor