Hepatitis C in HIV-infected individuals: a systematic review and meta-analysis of estimated prevalence in Africa.

INTRODUCTION Although hepatitis C virus (HCV) screening is recommended for all HIV-infected patients initiating antiretroviral therapy, data on epidemiologic characteristics of HCV infection in resource-limited settings are scarce. METHODS We searched PubMed and EMBASE for studies assessing the prevalence of HCV infection among HIV-infected individuals in Africa and extracted data on laboratory methods used. Prevalence estimates from individual studies were combined for each country using random-effects meta-analysis. The importance of study design, population and setting as well as type of test (anti-HCV antibody tests and polymerase chain reactions) was examined with meta-regression. RESULTS Three randomized controlled trials, 28 cohort studies and 121 cross-sectional analyses with 108,180 HIV-infected individuals from 35 countries were included. The majority of data came from outpatient populations (55%), followed by blood donors (15%) and pregnant women (14%). Based on estimates from 159 study populations, anti-HCV positivity prevalence ranged between 3.3% (95% confidence interval (CI) 1.8-4.7) in Southern Africa and 42.3% (95% CI 4.1-80.5) in North Africa. Study design, type of setting and age distribution did not influence this prevalence significantly. The prevalence of replicating HCV infection, estimated from data of 29 cohorts, was 2.0% (95% CI 1.5-2.6). Ten studies from nine countries reported the HCV genotype of 74 samples, 53% were genotype 1, 24% genotype 2, 14% genotype 4 and 9% genotypes 3, 5 or 6. CONCLUSIONS The prevalence of anti-HCV antibodies is high in HIV-infected patients in Africa, but replicating HCV infection is rare and varies widely across countries.

Poliovirus chimeras replicating under the translational control of genetic elements of hepatitis C virus reveal unusual properties of the internal ribosomal entry site of hepatitis C virus.

Chimeric genomes of poliovirus (PV) have been constructed in which the cognate internal ribosomal entry site (IRES) element was replaced by genetic elements of hepatitis C virus (HCV). Replacement of PV IRES with nt 9-332 of the genotype Ib HCV genome, a sequence comprising all but the first eight residues of the 5' nontranslated region (5'NTR) of HCV, resulted in a lethal phenotype. Addition of 366 nt of the HCV core-encoding sequence downstream of the HCV 5'NTR yielded a viable PV/HCV chimera, which expressed a stable, small-plaque phenotype. This chimeric genome encoded a truncated HCV core protein that was fused to the N terminus of the PV polyprotein via an engineered cleavage site for PV proteinase 3CPpro. Manipulation of the HCV core-encoding sequence of this viable chimera by deletion and frameshift yielded results suggesting that the 5'-proximal sequences of the HCV open reading frame were essential for viability of the chimera and that the N-terminal basic region of the HCV core protein is required for efficient replication of the chimeric virus. These data suggest that the bona fide HCV IRES includes genetic information mapping to the 5'NTR and sequences of the HCV open reading frame. PV chimeras replicating under translational control of genetic elements of HCV can serve to study HCV IRES function in vivo and to search for anti-HCV chemotherapeutic agents.

Hepatitis B and C prevalence in Portugal: disparity between the general population and high-risk groups

Background and aims: The prevalence of anti-HCV and HBsAg in Portugal has been shown to be elevated in high-risk groups, such as intravenous drug-users and incarcerated individuals. However, in the general population, prevalence remains largely unknown. The aims of this study were to estimate the prevalence of anti-HCV and HBsAg in the general Portuguese population and identify associated risk factors. Materials and methods: We carried out a nationwide, population-based cross-sectional study of adults resident in mainland Portugal. Serology for HBsAg, anti-HBc, anti-HBs, and anti-HCV was performed. Anti-HCV-positive individuals were tested for HCV RNA by PCR. Results: Of 1685 participants, 50.6% were men, mean age 50.2±18.3 years. In terms of hepatitis C, the prevalence of anti-HCV was 0.54% [95% confidence interval (CI): 0.2–0.9] and 0.12% (95% CI: 0.0–0.3) were viremic, with peak prevalence among individuals 35–64 years of age (0.8%), men (0.8%), and individuals from Lisbon and Tagus Valley region (1.9%). In terms of hepatitis B, the estimated prevalence of HBsAg was 1.45% (95% CI: 0.9–2.0). A higher prevalence was found in individuals who were 35–64 years old (2.2%), in men (2.5%), and in the Northern region (2.6%). The presence of positive serological markers of hepatitis C virus and hepatitis B virus infection did not correlate with elevated aminotransferases, race, place of birth, and alcohol consumption. Conclusion: These results suggest a low endemicity for both hepatitis B and hepatitis C in the general population, in contrast to a very high prevalence in risk groups, thus suggesting that targeted screening to high-risk groups may be more cost-effective than general population screening.

Antimicrobial and bond strength properties of a dental adhesive containing zinc oxide nanoparticles

Aim: To assess the effect of adding zinc oxide nanoparticles to dental adhesives on their anti-microbial and bond strength properties. Methods: 45 human premolars were cut at the cement enamel junction (CEJ) and the crowns were sliced into buccal and lingual halves. The specimens were classified into three groups, etched with 37% phosphoric acid for 15 s and rinsed for 30 s. Single Bond, Single Bond+5% zinc oxide and Single Bond+10% zinc oxide were used in the first, second and third groups. A cylinder of Z250 composite was bonded and cured for 40 s. For anti-bacterial testing, 10 samples of each group were assessed by direct contact test; 10 μL of bacterial suspension was transferred into tubes containing adhesives and incubated for one hour; 300 μL of brain heart infusion (BHI) broth was added to each tube and after 12 h, 50 μL of bacteria and broth were spread on blood agar plates and incubated for 24 h. Results: The colony count decreased significantly in the second and third groups compared to the first. Conclusions: Incorporation of zinc oxide nanoparticles into dental adhesives increases their anti-microbial properties without affecting their bond strength.

The biological meaning of anti-HBC positive result in blood donors: relation to HBV-DNA and to other serological markers

In order to assess the potential risk of anti-HBc-positive blood donors for post-transfusional hepatitis and to investigate whether other HBV serological markers are capable of identifying the presence of the virus, 1000 first-time blood donors were enrolled between June and July 1997. These donors were screened using routine Brazilian blood center tests (HIV 1 and 2, HTLV 1 and 2, Chagas disease, Syphilis, HCV, HBsAg, anti-HBc and ALT ). The 120 (12%) found to be anti-HBc-positive underwent further tests: HBe, anti-HBe, anti-HBs and HBV-DNA by PCR. Ten cases were HBsAg positive and all were HBV-DNA positive by PCR. Three HBsAg-negative donors were HBV-DNA-positive. Two HBV-DNA-positive donors were also anti-HBs-positive. All the HBV-positive donors had at least one HBV marker other than anti-HBc. Anti-HBc is an important cause of blood rejection. Testing for HBsAg alone is not fully protective and anti-HBc remains necessary as a screening test. The presence of anti-HBs is not always indicative of absence of the virus. The addition of other HBV serological markers could represent an alternative in predicting the presence of the virus when compared with PCR. It is recommended that other studies should be carried out to confirm this finding.

Primary screening of blood donors by nat testing for HCV-RNA: development of an "in-house" method and results

An "in-house" RT-PCR method was developed that allows the simultaneous detection of the RNA of the Hepatitis C Virus (HCV) and an artificial RNA employed as an external control. Samples were analyzed in pools of 6-12 donations, each donation included in two pools, one horizontal and one vertical, permitting the immediate identification of a reactive donation, obviating the need for pool dismembering. The whole process took 6-8 hours per day and results were issued in parallel to serology. The method was shown to detect all six HCV genotypes and a sensitivity of 500 IU/mL was achieved (95% hit rate). Until July 2005, 139,678 donations were tested and 315 (0.23%) were found reactive for HCV-RNA. Except for five false-positives, all 310 presented the corresponding antibody as well, so the yield of NAT-only donations was zero, presenting a specificity of 99.83%. Detection of a window period donation, in the population studied, will probably demand testing of a larger number of donations. International experience is showing a rate of 1:200,000 - 1:500,000 of isolated HCV-RNA reactive donations.

IL28B polymorphism is significantly correlated with ifn anti-viral effectiveness already on first day of pegylated interferon-alpha and ribavirin therapy of chronic HCV infection

Background and Aims: Recently, single nucleotide polymorphisms (SNPs) in IL28B were shown to correlate with response to pegylated interferon-a (IFN) and ribavirin therapy of chronic HCV infection. However, the cause for the SNPs effect on therapy response and its application for direct anti-viral (DAV) treatment are not clear. Here, we analyze early HCV kinetics as function of IL28B SNPs to determine its specific effect on viral dynamics. Methods: IL28B SNPs rs8099917, rs12979860 and rs12980275 were genotyped in 252 chronically HCV infected Caucasian naïve patients (67% HCV genotype 1, 28% genotype 2-3) receiving peginterferonalfa- 2a (180 mg/qw) plus ribavirin (1000-1200 mg/qd) in the DITTO study. HCV-RNA was measured (LD = 50 IU/ml) frequently during first 28 days. Results: RVR was achieved in 33% of genotype 1 patients with genotype CC at rs12979860 versus 12-16% for genotypes TT and CT (P < 0.03). Significant (P < 0.001) difference in viral decline was observed already at day 1 (see Figure). First phase decline was significantly (P < 0.001) larger in patients with genotype CC (2.0 log) than for TT and CT genotypes (0.6 and 0.8), indicating IFN anti-viral effectiveness in blocking virion production of 99% versus 75-84%. There was no significant association between second phase slope and rs12979860 genotype in patients with a first phase decline larger than 1 log. HCV kinetics as function of IL28b SNP. The same trend (not shown) was observed for HCV genotype 2-3 patients with different SNP genotype distribution that may indicate differential selection pressure as function of HCV genotype. Similar results were observed for SNPs rs8099917 and rs12980275, with a strong linkage disequilibrium among the 3 loci allowing to define the composite haplotype best associated with IFN effectiveness. Conclusions: IFN effectiveness in blocking virion production/ release is strongly affected by IL28B SNPs, but not other viral dynamic properties such as infected cell loss rate. Thus, IFN based therapy, as standard-of-care or in combination with DAV, should consider IL28B SNPs for prediction and personalized treatment, while response to pure DAV treatment may be less affected by IL28B SNPs. Additional analyses are undergoing to pinpoint the SNP effect on IFN anti-viral effectiveness.

KRAS mutation testing for predicting response to anti-EGFR therapy for colorectal carcinoma: proposal for an European quality assurance program.

Novel therapeutic agents targeting the epidermal growth factor receptor (EGFR) have improved outcomes for patients with colorectal carcinoma. However, these therapies are effective only in a subset of patients. Activating mutations in the KRAS gene are found in 30-40% of colorectal tumors and are associated with poor response to anti-EGFR therapies. Thus, KRAS mutation status can predict which patient may or may not benefit from anti-EGFR therapy. Although many diagnostic tools have been developed for KRAS mutation analysis, validated methods and standardized testing procedures are lacking. This poses a challenge for the optimal use of anti-EGFR therapies in the management of colorectal carcinoma. Here we review the molecular basis of EGFR-targeted therapies and the resistance to treatment conferred by KRAS mutations. We also present guideline recommendations and a proposal for a European quality assurance program to help ensure accuracy and proficiency in KRAS mutation testing across the European Union.

HBV vaccination of HCV-infected patients with occult HBV infection and anti-HBc-positive blood donors

Anti-HBc positivity is a frequent cause of donation rejection at blood banks. Hepatitis B virus (HBV) infection may also occur in HBsAg-negative patients, a situation denoted occult infection. Similarly, very low levels of HBV-DNA have also been found in the sera of patients with chronic hepatitis C virus (HCV) infection, even in the absence of serum HBsAg. Initially we searched for HBV-DNA in serum of 100 blood donors and 50 HCV-infected patients who were HBsAg negative/anti-HBc positive by nested-PCR and by an HBV monitor commercial test for HBV-DNA. Anti-HBs seroconversion rates were measured in 100 blood donors and in 22 patients with chronic HCV infection after HBV vaccination to determine if the HBV vaccination could eliminate an occult HBV infection in these individuals. Occult HBV infection was detected in proportionally fewer blood donors (6/100 = 6%) than chronic hepatitis C patients (12/50 = 24%) (P < 0.05). We noted seroconversion in 6/6 (100%) HBV-DNA(+) and in 84/94 (89.4%) HBV-DNA(-) blood donors (P > 0.05). All subjects who were HBV-DNA(+) before the first dose of HBV vaccine (D1), became HBV-DNA(-) after D1, D2, and D3. Among 22 HCV-positive patients, 10 HBV-DNA(+) and 12 HBV-DNA(-), seroconversion was observed in 9/10 (90%) HBV-DNA(+) and in 9/12 (75%) HBV-DNA(-) subjects (P > 0.05). The disappearance of HBV-DNA in the majority of vaccinated patients suggests that residual HBV can be eliminated in patients with occult infection.

Variability of anti-PF4/heparin antibody results obtained by the rapid testing system ID-H/PF4-PaGIA

BACKGROUND: Recent studies have shown that a low clinical pretest probability may be adequate for excluding heparin-induced thrombocytopenia. However, for patients with intermediate or high pretest probability, laboratory testing is essential for confirming or refuting the diagnosis. Rapid assessment of anti-PF4/heparin-antibodies may assist clinical decision-making. OBJECTIVES: To evaluate the performance of rapid ID-H/PF4-PaGIA. In particular, we verified reproducibility of results between plasma and serum specimens, between fresh and frozen samples, and between different ID-H/PF4-polymer lots (polystyrene beads coated with heparin/PF4-complexes). PATIENTS/METHODS: The samples studied were 1376 plasma and 914 corresponding serum samples from patients investigated for suspected heparin-induced thrombocytopenia between January 2000 and October 2008. Anti-PF4/heparin-antibodies were assessed by ID-H/PF4-PaGIA, commercially available ELISAs and heparin-induced platelet aggregation test. RESULTS: Among 914 paired plasma/serum samples we noted discordant results (negative vs. low-titre positive) in nine instances (1%; 95%CI, 0.4-1.6%). Overall, agreement between titres assessed in plasma vs. serum was highly significant (Spearman correlation coefficient, 0.975; P < 0.0001). Forty-seven samples tested both fresh and after freezing/thawing showed a good agreement, with one discordant positive/negative result (Spearman correlation coefficient, 0.970; P < 0.0001). Among 1376 plasma samples we noted a strikingly variable incidence of false negative results (none - 82%; 95%CI, 66-98%), depending on the employed ID-H/PF4-polymer lot. Faulty lots can be recognized by titrating commercial positive controls and stored samples of HIT-patients. CONCLUSION: Laboratories performing the assay should implement stringent internal quality controls in order to recognize potentially faulty ID-H/PF4-polymer lots, thus avoiding false negative results.

Inhibitory effect of 9-(2-phosphonylmethoxyethyl)adenine on visna virus infection in lambs: a model for in vivo testing of candidate anti-human immunodeficiency virus drugs.

The acyclic nucleoside phosphonate analog 9-(2-phosphonylmethoxyethyl)adenine (PMEA) was recently found to be effective as an inhibitor of visna virus replication and cytopathic effect in sheep choroid plexus cultures. To study whether PMEA also affects visna virus infection in sheep, two groups of four lambs each were inoculated intracerebrally with 10(6.3) TCID50 of visna virus strain KV1772 and treated subcutaneously three times a week with PMEA at 10 and 25 mg/kg, respectively. The treatment was begun on the day of virus inoculation and continued for 6 weeks. A group of four lambs were infected in the same way but were not treated. The lambs were bled weekly or biweekly and the leukocytes were tested for virus. At 7 weeks after infection, the animals were sacrificed, and cerebrospinal fluid (CSF) and samples of tissue from various areas of the brain and from lungs, spleen, and lymph nodes were collected for isolation of virus and for histopathologic examination. The PMEA treatment had a striking effect on visna virus infection, which was similar for both doses of the drug. Thus, the frequency of virus isolations was much lower in PMEA-treated than in untreated lambs. The difference was particularly pronounced in the blood, CSF, and brain tissue. Furthermore, CSF cell counts were much lower and inflammatory lesions in the brain were much less severe in the treated lambs than in the untreated controls. The results indicate that PMEA inhibits the propagation and spread of visna virus in infected lambs and prevents brain lesions, at least during early infection. The drug caused no noticeable side effects during the 6 weeks of treatment.

Testing the Efficacy of Panax ginseng Extract as a Novel Anti-Quorum Sensing Agent

Quorum sensing (QS) is the phenomenon by which microorganisms regulate gene expression in response to cell-population density. These microorganisms synthesize and secrete small molecules known as autoinducers that increase in concentration as a function of cell density. Once the cell detects the minimal threshold concentration of an autoinducer, the gene expression is altered accordingly. Although the cellular circuitry responding to QS is relatively conserved, the cellular processes regulated by QS, for example are conjugation, motility, sporulation, biofilm formation and production of virulence factors importamt for infection. Since many pathogens have developed resistance to available antibiotics, novel therapies are required to further treat these pathogens. Inhibitors of QS are potential therapeutic candidates since they would inhibit virulence without selecting for antibiotic resistant strains. Previous studies have shown that the Chinese herb Panax ginseng contains compounds that inhibit QS activity. To further characterize this activity, a highly sensitive quantitative liquid assay was developed using a Chromobacterium violaceum AHL mutant, CV026 as a biomonitor strain. After confirmation that P. ginseng aqueous extracts had anti-QS activity, the extract was fractionated on a reverse phase C18 Sep Pak column. Most anti-QS activity was present in the flow through, and compounds eluted with ten percent acetonitrile in water antibacterial activity. We thus conclude that P. ginseng has anti-QS activity and the active compound is water soluble. Compounds from P. ginseng, could be used as a lead structure to design compound with higher anti-QS activity for therapeutic treatments.

Mudanças no âmbito da testagem anti-HIV no Brasil entre 1998 e 2005

OBJETIVO: Analisar mudanças na realização de teste anti-HIV, as razões alegadas entre as pessoas que foram ou não testadas e o recebimento de aconselhamento. MÉTODOS: Estudos transversais conduzidos com homens e mulheres de 16 a 65 anos, com amostras representativas do Brasil urbano em 1998 (n=3.600) e 2005 (n=5.040). Características sociodemográficas, sexuais, reprodutivas e de experiências de vida e saúde foram consideradas na análise. A avaliação das possíveis diferenças nas distribuições das variáveis baseou-se nos testes qui-quadrado de Pearson e F design-based (±<5%). RESULTADOS: Em 1998, 20,2% dos entrevistados haviam realizado o teste e 33,6% em 2005. Foram testadas 60% das mulheres na faixa 25-34 anos, mas as que iniciaram a vida sexual antes dos 16 anos e reportaram quatro ou mais parceiros sexuais nos cinco anos anteriores à entrevista foram menos testadas. Não se observou aumento significativo da testagem entre homens, exceto para os de 55-65 anos, renda per capita entre 1-3 e 5-10 salários mínimos, aposentados, protestantes históricos e adeptos de cultos afro-brasileiros, moradores da região Norte/Nordeste e os que declararam parceria homo/bissexual ou não tiveram relações sexuais nos cinco anos anteriores à entrevista. Não aumentou a freqüência de testagem entre pessoas auto-avaliadas como sob alto risco para o HIV. Entre as mulheres, a freqüência de testagem pré-natal aumentou e a testagem por trabalho diminuiu entre os homens. Em 2005, metade dos testados não recebeu orientação antes ou após o teste. CONCLUSÕES: Houve expansão desigual na testagem, atingindo principalmente mulheres em idade reprodutiva, adultas e pessoas com melhores condições sociais. A testagem parece estar aumentando no País sem a devida atenção à decisão autônoma das pessoas e sem o provimento de maior e melhor oferta de aconselhamento